et h ylfluoroacel amide
00c, 178 -
bp**
77°
178
29
inp
65°
178
7. X-d-4 '11tons’thvlehloroacetamide*
7
mp
--54°
7
8. X-d-Chloroethvll richloroacetamide
7
nip
75°
7
(t. Kthvl X-d-chloroethvloxamale
7
inp
08°
7
10. Methyl X-d-chloroethylcarbamate
7
1.4575
61
29, 134
bpM
100°
7
11. Methyl X-d-chloroel hvl-X-nit rosoca rlaimate
7, 110. 121,
Hu1’
1.4600
no
29,41. no,
102, 55
xy-p-
42
mp
106.5°
42
29, 41
phenvlenediamine
42
mp
102-103
42
29, 41
87, 2-(ft-Chloroethylamino)qninoline hydrochloride
88. «-Benzylamino-0-chlort»-0-pheiiylprop1ophenone
»8k
mp
140 152
48k
29,41
hydrochloride
89. 3-Bcnzylamino-a-bromo-d-phenylpr«piophenone
47
mp
152-156°
58
29
hydrobromide
90. o-Henzylamino-d-bromo-^-phcnylpropiophenone
47
mp
147-149°
58
29
hydrohromidc
C. Dermilives of tertiary amines
• 7
mp
144 147°
58
91. X-Mcthvlcthvleneimine
-..
29
92. d-Hthyleneiminopropionilrile
53j
bp,!
67 69
53j
29
93. Methyl 3-ethylenciminopropionate
53j
bp"’
61 -64°
53j
29
94. j3-Chloroethyldimelhylamine*
134
95. Dimethyl 0-chloropropylamine
If.
no*1
1.4214
16
29, 41
p
123 124°
177e
177c
98. Methyl bis(0-cl11 oroe 1 hy l)a m i nc *
2, 5
Hll18
1.4679
2
29, 41, 121,
134
. ‘V ■ .. - ■ ' • •
#11*
1.4682
115
1.11S
82
—
,r5
1.1203
115
—
bp!u
50.0-50.5°
2
vol*°
2.487
15
99. Methyl 6(’*(/}-ehloroethyl)aminc hydrochloride*
2
mp
107-108°
2
29, 41, 129,
177a
100. Methyl b/s(d-chloroethvl)jimine formate
51a
...
101. Methyl fe/«(d-ehloroetliyliamine picrate
102. Horon fluoride complex of methvl-h(s(d-ehloro-
51a
ethyl)amine
103. Reaction product of methyl 6/x(fPchloroelhyl)-
.50
amine and titanium tetrachloride
104. Methyl 6/s(/3-chloroelhyl)aniinc oxide hydro-
51 e
chloride
29
105. Methyl 6is(d*cyanoethyl)amine
53e
bp1"
177-187°
53c
29
106. Methyl 6/s(tJ-thiocyanoefhyl)ainine
23
Decomposition on distilla-
tion at 1.5 mm
23
29
107. Methyl hydrochloride
23
mp
117-118°
23
29, 41
108, Methyl
109. Methyl 0-chloroethyl-/J-hydro.vycthylaminc hy-
12, 126
...
29, 41
drochloride
12
29
110. Methyl fl-chh.rocthyl-/3-hvdroxyethylainine picrate
12
mp
72 73°
12
... —
111, Methyl /3-acetoxyethyl-d-ehloroethylamine*
32, 126
n ir‘
1.4474
32
43, 106b
bp"14
540
32
bp" 111
46°
32
bp" w
41°
32
SECRET SYNTHESIS VM) PROPERTIES
Table 1 (Conti
n ued).
Compound
Reference
to
synt hesis
Physical properties
Property Reft
‘nmee
Reference to
toxicity
ami
vesica ncy
data
112.
|)imeiliyl ,t, d'-dichlortr-fcri-butvlamine*
113.
Methyl /}-chloroethyI-/3-chloropropylamine*
134
114.
Methyl d-chloroet hyl-d-chloropropylamine*
...
134
115.
Diethvl (4-chlor(x‘l h vlaminc*
116.
Diethyl 3-chloroethvlamine hydrochloride
16
mp
210-211.5°
16
117.
Kthvl bix(#-chloroethyl)Hii)iHe*
2, 10, 120
«n5*
1.4639
2
29, 41, 134,
_■
143
d
1.083
2
hp» ‘
49.0-49.5°
2
TT
vol"
1.59
31
118.
Kthvl bix{ /S-chlomethyl (amine hydrochloride*
2, 10
nip
139-140°
2
137
119.
Kthyl /S-ehloroct hyM-hydroxycthylamine picryl-
sulfonate
17
mp
110 111
17 _
29
120.
0-Methoxvethvl 6f's(j8-chloniethyl)amine
23
«n**
1.4671
23
29, 41
y 1 hix( fi-chloroet hyl)aminc*
16, 120
« n27
1.4029
16
29,41, 134,
—
1 13
,r-«
1.092
16
148
bp*
62-63°
16
vol2"
0.783
31
SECRET N ITROEEX MUSTARDS
Table 1 {Continued).
Reference
to
Physical projicrtics
Heference to
toxicity
and
Compound
synthesis
Property Heference
vesicancy
data
144. Propyl bisl 3-chloroethyl)amine hydrochloride*
16, 120
mp
118-120°
16
137
145.
146. lsopn>pyl-6iJi(d-cMoroetliyl)anune*
2,9
n hM
1.4641
2
29, 41, 143
d
1.053
2
bp2 5
67.0 68.0T
2
' “
mp
13.7
S3
-
V«>l“
0.869
15
147. Isopropyl-5ts(/J-chloroelhyl)amine hydrochloride*
2, 9
mp
210 213 (dec.)
2
It, 137
148. X-/J-Chloruethylpij)cridine
11
bp*
40 11°
11
29. 41
140. X-0-Chloroethylpiperidine hydrochloride
11
mp
230
11
150. Hutyl-6/x(d-chloroethyl)amine
16
no5*
1.4637
16
29, 41
d26
i ,027
16
__ ,
...
bp2-*
89.(V 89.5°
16
‘
vol-®
0.321
31
151. Hut vl-5is(d-chloroethyl)aniine hydrochloride
16
nip
— 96-97*
16
152. 7 -Chlorobut vl-6ix(d-chloroethyl)ainine
29, II
153. y-Oxobutyl 5»«(0-chloroethyl)amine hydrobromide
. . .
29
154. xrC'Hutyl-6i.sO-chlori ir‘
1.4655
16
29, 41
d21
1.028
16
bp*
84-84.5°
16
rr.
v«l,#
0.394
31
155. «er-Rutyl-fc(s(/3-chloroethvl)amine hydrochloride
16
mp
132-138°
16
156, Isob«ilyl-/)ix(/J-chlorocthyllamine
16
«,.2?
1.4597
16
29, 41
- _
.r<
1.0078
16
bp1
81-81.3°
16
- . .
vol20
0.508
31
157, Isobutvl-6/x(d-chloroethvl)ainine hydrochlorides-
16
mp
107-108°
J6
158. 0T/-Hutyl-b/8(#-chloroclhyl)amlne
16
n tf°
1.4710
16
29, 41
• —
if-'
1.032
16
• ,
bp1*
68-69°
16
•
vol2"
0.581
31
159. lcr/-butyl fcr.s-(0-chloroethyl)-
amine hydrochloride
23
mp
112.2-114.2°
23
41
162. /3-Chloroethy!-6i*(d-chluropropyl )aminc*
163. 0-Chloroethyl-fws(0-chloropropyl)amine hydro-
.... .
chloride*
....
164. 1 urfuryl 6/»((?-chloroelhyl)aminc
23
nr.a
1.5033
23
41
1.171
23
bp11-"1 106-107°
23
165. Furfurvl-6is(/3-chloroet hyl )amine hydrochloride
23
mp
88.5-89.5°
23
29, 41, 43
166. Tetrahydrofurfuryl-hisO-chloroethyl )aminc
23
nr.24
1.4877
23
29, 43
d”
1.129
23
“ ‘ .
bp«*
82-84°
23
167. Tetrahydrofurfuryl-6/jt(0-chloroethyl)amiiie hy-
drochloride
168. d/-X-(/J-Chloroet hyl)-2-chloromelliylpiperidinc
23
mp
117 118°
23
hydrochloride*
169. 6(or 7)-Chloroetronccane
30
nn”
1.4913
30
29
170. 6(or 7)-Chloro-l-ehloromothyl-l ,2-dehydropyr-
bp”
111 112°
30
roiizidine hydrochloride
30
mp
122-123°
30
29
171. triaf/J-Chloropropyl)amine
II
bp2
99-103°
11
29
172. X-Kthyl-X-C^-chloroethyDaniline
11
bp°*
102 109
It
29, 41
173, X,X-hid d-( 'hloroet hyl laniline*
11
bp0-7
123°
11
29, 44
mp
43-44°
II
SECRET 65
SYNTHESIS \M) PROPERTIES
Table 1 (Continued).
Reference
to
Physical properties
Reference to
toxicity
and
Compound
synthesis
Pro|>erly Reference
vesica ncy
data *
174. \, N -hig{ (3-C hh in >et hyl t-p-nitrosoaniline
53c
nip
72
53c
20. 41
I7.i, Cychihexyl-fH*(d-chhinx‘t hyl )a mine
16
n n5i
1.1040
16
20, 41
d-'
1.077
16
bp11
105-105.6°
16
vol*°
0.0383
31
17(1. Cvclohc\vl-fci»(itJK,liloroethvl)ainino hydrochloride
16
mp
174 175’
16
177. Benzyl-hisfd-rhlonxOhynnmine
16
Hi)55
1.5334
16
20. 41
(f3
1.112
16
»
b|)-
138-130’
16
178. Itenzvl-b/x(jJ-chlori
<1.0
2
20
1 SO. 1 lop) vl-bix{d-chloroethyl famine hydrochloride
2
41
181. Phenethyl-h»s(d-chloroethyl)ainine
182. X, X. \'-Mrfi/r(«(d-Chliiroelhylfelhylene-
53r
bp*
06°
53k
20 .1.
diamine di hydrochloride
183, l,3-h(»(fei#(d-Ch!orocthyl)amii»o) propane dihydro
40
20, 41
chloride.
184. 1, 3-fu«[ bis( ii-( hloroelhyl )amino]-2-ch!oropropane
32
nip
138 130’
32
20
di hydrochloride
32
nip
141-141 2U
32
20
1 So. hix{ff-(his(d-Chlonx*f hylfaminof-ethyl] sufide*
23
'll.'*
1.5287
23
20
186. 6. Derivatives of quaternary ammonium mils
106. Trinielhyl-d-fluoroct liylaminonium bromide*
mp
244° (dec.)
I77d
177d
107. 1 )imethyl-bi«(d-chlorocth vl)ammoninm chloride
108. d-.\rcloxvcthvl-d-chlon«‘lhvldimclhylammonium
29, 41
iodiile*
126
mp
150° (dec.)
126
100. Mcthvl-/r/.s(d-chloroelhvl)ammonium chloride*
. . .
....
200. MethyHri«(d-chloroethyl)amn!oni»im sulfate*
201. Elhylvinyl-5is(d-chloroelhyl)ammonhim chloride
32
mp
102-106° (dec.)
32
20
202. Triethyl-d-fhionx'thylammonium bromide*
203. d-Carbaimixyel hylet h y 1 -6/*(d-chi oroe t hylfammo-
mp
237° (dec.)
177c
177c
nium chloride
23
20. 41
204. d-KIuoroethylpyriiliniuni bromide*
205. X,X-f»is(d-Ch!orocthvl)pjix'ridinium chloride, mono-
mp
180’
177c
177c
hvdrale
. . .
206. Polymer of mefhyl-fei.’Kd-chloroethyDamine (« = 21
2
20
207. Polymer of met hvl-hi>(d-ehIorcx't hyl (amine
2
20
208. l,4-f»s(d-f ’hlonxdhylf-l ,4-diethylpi|>eraziniuiii di-
chloride
200. Ammonium comiKHind from 1 mole of methvl-fc/«(d-
...
20
chlornethvl)amine ami 2 moles of methyhli-
ethanolamine
20
Detailed studies of the preparation of the four
most important nitrogen mustards on a laboratory,
pilot plant, or manufacturing scale are reported in
the following references.
SECRET 66
NITROGEN MUSTARDS
Agent References
HNl 2, 4, 10, 71. S7, 05, 07, OS, 90, 136. 182d
HN2 2, 4, 5, 82, 120, 158, ICO, 182a, 185a,
185b, e, <1. 102, 104, 196
11X3 2,4,5, 68, 80, 02, i)6, 157, 170. 171, 172
Isopropyl-6is(0-
chlorocthyl)amine 2, 4,83,136
Work has been done on alternate syntheses which
do not employ thionyl chloride. Results have been
discouraging.*4 ** The best alternate method for
HNl uses phosphorous trichloride in place of thionyl
chloride, and gives yields approaching 75 per cent.16
Other reagents used with less success are phosphorous
pentaehlondc,206 phosgene,1'41’-'1 193 and hydrochloric
acid.1'4*’
The alkyl-6is(d-hydroxyethyl)amines and
hydroxyet hy 1) a mine required for synthesis of the
nitrogen mustards have liecn prepared commer-
cially by reaction of primary amines or ammonia
with ethylene oxide. Some work has been reported
in the classified literature on such reactions and on
the purification of technical ethanolamiues for use as
nit rogen mustard intermediates.'s«.iei.i«.i*7,iB».
16»aT2,lS5a,r.l92,194,196
Because of the possibility of a short supply of
ethylene oxide in the event of large-scale nitrogen
mustard manufacture, methods which did not em-
ploy ethylene oxide were investigated for preparing
alkanolamines, particularly RN(riI,(TI2()H)2 (where
R is ethyl, methyl, or isopropyl). The most success-
ful method developed utilizes formaldehyde, hydro-
gen cyanide, and ethyl alcohol as the basic raw
materials and follows these steps:
1. HCN + CHjO CHjOHC’N
2. rn,oiR’N + (C.H:.0)2chs —►
C*Hi(X'H20( 'Il*(’N T CjlUOH
3. 2(',IIiOCTljOCH2CN + 1H2
(f,2H5OCH2OCH2CH,)2NH + Nil,
4. 2(C2H;,0(TI2OCI 12('H2)2NH -(- (C2H;,)2S()4
+ Xa.ro, —> 2(C,HiOCn2OCH2CHI)2Nr,Hi
+ Xa2S04 + h,o + ro2
5. ((‘2H5orH2oriU'ii2)2xr2H, + hci
+ (,11,011 —► (CH2OHCH.)2NC2Hi-H(,l
T- 2rH,(0( ,11.5)2
This “formal” route, in which formaldehyde
cyanohydrin is converted to a less sensitive formal
derivative before hydrogenation, is estimated to Ik*
capable of producing N-ethyl diethanolamine hydro-
chloride at a cost of 25 to 30 cents per pound, at an
annual rate of 10.000,000 pounds.2" A more direct
route, in which formaldehyde cyanohydrin is hydro-
genated directly to diethanolamine (subsequently
alkylated), gave poorer yields and appears to In* a
more expensive process.2" Methyl-6>«(j8-liydroxy-
ethyl)amine has been prepared successfully by hy-
drogenation of diethanolamine in the presence of
formaldehyde.19 Other less advantageous routes to
the alkanolamine intermediates for HXl, HN2, and
HN3 have been explored.1" In addition to the stand-
ard method of preparation from isopropyl amine and
et hylene oxide. isopropyl-Iu’s(/3-hydroxyethyl)amine
has been prepared by hydrogenation of a mixture of
acetone and diethanolamine,19 or by the react ion of
ethylene oxide with isopropyl-d-hydroxycthylamine.
The latter compound is prepared by hydrogenating
a mixture of acetone and-cthanolamine.199 _
6.2.2 Physical Properties
The nitrogen mustards ai*e oils of limited water
solubility. They are miscible with ordinary organic
solvents. Their physical properties have been ex-
tensively studied.
i«7.i«s.i89.jwb.c gome 0f t tie constants having most-
liearing on chemical warfare are presented in Table 2.
6.2.3 Chemical Properties
The nitrogen mustards are basic amines which
form stable salts with strong acids such as hydro-
chloric acid. They are active alkylating agents, and
the physiological reactions responsible for their
toxicity are primarily alkylations. Their reactions
from the biochemical, physicochemical, and physi-
ological mechanism standpoints have been studied
in great detail and are summarized in Chapters 19,
20, and 21. A primary intermediate in their reactions
is a l-(/3-chIoroethyl)ethylenimonium ion, formulated
1 >elow for HN2, which is analogous with (he ethylene-
sulfonium compound1* intermediate in the reactions
of H (see Chapters 19 and 20).33
125.127,130.135,151 .ISltt.b
Self-alkylation is responsible for the dimerization
which occurs slowly when the lower molecular weight
alkyl-6f«(/J-chloroethyl)amines are allowed to stand.
The reaction is rapid in the presence of water. It re-
sults in the deposition of crystalline solids such as the
“dichloroeyclie dimer” which is formed from HX2;
'■ Ktliyloni'-Siilphoninrn Compound ClCH;CIIj
+
SCHjCH.CC
SECRET SYNTHESIS \ND PROPERTIES
67
Tabi.r 2. Physical [»reen exploded without evidence of gross
decomposition except in 75-mm shell under condi-
tions more severe than are encountered in the
field.M 66 89 However, in the most quantitative in-
vestigation that has been made, one of six M47A2
bombs charged HXl flashed on static detonation.66
Although considerable lexicologically effective vapor
was subsequently evolved from the terrain contam-
inated by the explosions in these tests, the dosages
were less by an amount approaching 30 per cent than
would have been predicted from similar tests with H
had no decomposition of HXl occurred during the
explosion or, subsequently, on the contaminated
terrain.66 -—
The available data for I1N2 are of a rather quali-
tative nature. It appears that this agent can be dis-
persed without complete destruction by explosion of
various chemical munitions but that the toxicological
effects of the initial clouds so produced are inferior to
those produced by H or HT.7* 7"7114,47 IH!M*0
Stability on Terrain
Two imjiortant characteristics determining rate of
inactivation on soil and vegetation are solubility in
water and rate of reaction with water. The approxi-
mate data given in Table 3 lead to the prediction of
HXl and IIN2 vapor wore greater than those of If,
as predicted from the relative volatilities, the total
dosage (C7) of evolved vapor was in the eases of HNI
and HN2 only one-half of that anticipated from
similar trials with H. In annulus trials with HXl
and If in Florida, large drops were used on relatively
dry terrain and the 1-hour vapor dosages failed to
reveal significantly greater ground losses for HX3
than for If.6* As stated almve, however, in trials with
single, statically fired homhs the vapor evolution of
HXl was somewhat less than would have been pre-
dicted on the basis of tests with If. Fart of the loss
may have occurred during the explosion of the
bombs, and part subsequently on the terrain. Ter-
rain losses would have been facilitated by the dis-
pei-sion of the agent into small droplets during the
explosion.
6,2.3 Detection and Analysis
Excellent methods for the detection and analysis
of the nitrogen mustards arc available (see Chap-
ters 31 and 37).
For purposes of detection the use of the DH-3 re-
agent (see Chapter 34) is perhaps the most useful.
As used in the United States MO Detector Kit, this
test is approximately as sensitive for the nitrogen
mustards as for H. Collection of 0.1 to 0.2 Mg of HN 1,
HX3, or H from air containing 0.2 Mg 1 or more of
these agents suffices to give a positive reaction.112 A
supplementary test to differentiate nitrogen mustard
from H is somewhat less sensitive.
For purposes of analysis, among the most useful
methods are those utilizing the DB-3 reagent and
those dependent upon the mercurimetric titration of
the chloride obtained by hydrolysis from the nitro-
gen mustard (see Chapter 37).
6.2.6 Decontamination and Protection
The gas mask canister offers complete protection
against the nitrogen mustards.
In general, standard methods of decontamination
effective for H are useful for the nitrogen mustards,
but destruction of the nit rogen mustards by chemical
reaction with bleach or with currently used chlor-
amides is notably less efficient and rapid than in tin*
case of II 117 (see Chapters 21 and 32).
6 3 CHEMICAL STRUCTURE IN It ELA-
TION TO TOXICOLOGICAL POTENCY
Among the nitrogen mustards and related com-
pounds that have been studied (see Table I), the
Tabi.k 3. Characteristics influencing rate of inactivation
of II, fIN' 1, 1I\2, and I1N3 on terrain.
Approximate solubility
Approximate half-
in water at room tom-
life in water at
Agent
peraturc (ppm)
25 C
(minutes)
11
500
8
HXI
1,000 +
1.3
IIN2
13,000 +
4.0
II N3
SO _
2.4
greater losses with HXl and HX2 than with HN3
and TL This prediction is confirmed by the available
field data. There is no evidence in the semiquanti-
tative data of bomb and annulus trials that losses on
moist terrain are greater for HN3 than for II, in
spite of the greater persistence of HN3.*S The results
of British annulus trials with HXl and HN2 on
alkaline sod (Porton downland) indicate that ground
losses are significantly greater than in the ease
of H.1,5 IS* Although the initial dosages of evolved
SECRET TOXICOLOGY
highest toxicological potency is found among the
tertiary 6fs(/3-ehIoroethyl)amines, and, in particular,
in HXl, HX2, HX3, and the propyl and isopropyl
analogs of HXl. The toxicity, vcsicancy, and eye-
injurant action of these compounds is reviewed in the
following sections.
6 1 _ TOXICOLOGY
6.1.1 Detectability by Odor and
Sensory Irritation
HXl, HX2, and 11X3 arc markedly less detectable
by odor or sensory irritation than is H. Testimony
to the insidiousness of the vapors of all three nitrogen
mustards comes from plant accidents in which men.
informed of the potential hazard, were incapacitated
without being aware of having been exposed until
eye and respiratory symptoms developed after a
lapse of several hours.*9174*175
Laboratory (osmoscopic) determinations of the
median detectable concentrations of H, HXl, HX2,
and HX3 are given in Table L Attention is directed
odors. The pilot plant HX3 that was made in Eng-
land has a faint geranium-like odor. Inasmuch as
oral reports indicate that laboratory-prepared sam-
ples do not possess this odor, it may be suspected
that the geranium-like odor was due to an impurity,
possibly associated with the preparation of the ma-
terial in equipment previously used for lewisite. This
pilot plant material was used in the osmoscopic de-
termination cited in the preceding paragraph. Thus,
it is possible that other samples of plant run ILX3
would be even more odorless, and therefore more
insidious.
6. Toxicity
Toxicity data for animals totally exposed to air-
borne HXI, 11X2, and HX3 are set forth in Tables 6,
7, and 8. From the summary presented in Table 5 it
Tabus 5. Summary of toxicides of H, 11X1, I1N2, and
HX3 in the form of vapors.
(See Tables 6, 7, and 8 of this chapter and Table 5 of
Chapter 5 for more detailed data.)
Agent
(mg min nr1)
Mouse* llange for
(/ = 10 min) other species)
Ms! i mated
relative
toxicity
(H_» 100)
H
HX1
TIN2
HX3
1,100 IKK) 2,800
_ - 900 500-3,000
2,000 1,000 6,000
550 500-2,000
100
»100
50
5 100
♦ Fiftocii-diay observation period,
t Approximate.
T vbi.k 4. Median detectable concentrations of 11, 11X1,
HX2, and 11X3 as determined in the laboratory by the
osmoscopic technique.
A Rent
Purity
Median delectable
11
Plant run Ixtvinstein 0.6
00
Vnctiuni-disl ille-
parent only at much higher concentrations.141 Presumably
use of this technique with H and other nitrogen mustards
would give values correspondingly low in relation to the
median delectable concentrations as determined in the labora-
tory.
SECRET 70
MTROGRN ML STARDS
Tabi.k 6. Toxicity of 11 XL The animals
divcn in parentheses.
were totally
exposed, /-K’0,„’s that
are estimated very approximately are
i
HCD*
(mg min in3)
Exposure
time
(min)
Observat ion
licriod
(days)
Analytical
(A)
or
nominal
(X)
cone.
Xiimler
of
animals
Notes
Reference
Mouse
900
10
15
A
2S0
Low-flow chamber
37
900
10
15
A
89
High-flow ehamlier
37
1.300
10
10
X
1 10
Low-flow ehamlier
85
< 1.200
9ti0
30
20-100
15
15
A
A
30
140
Static chamber
Large ehamlier; 90 F; wind
143
1,100
20 100
10 _
A
140
sjieed without effect
I O le
Hat—
(7.50)
10
30
X
10
Low-flow chamber
37, 14a
<1,200
30
15
A
34
Static ehamlier
1 13
800
20-100
10 and 15
A
84
I.argeehamlier; OOF; wind
s|*eed without effect
104e
Guinea pijr
(2,500)
10
30
X
IS
Low-flow chamber
37, 14a
(1 ,.500-3,000)
30
15
A
36
Static ehamlier
143
Rabbit
(1,000 3,000)
10
30
X
5
Low-flow ehamlier
37, 14a
‘MX)
30
15
A
ot>
Low-flow chamber; 90 F
71
(1,000)
30
15
A
IS
1/ow-flow ehamlier; 73+ F
71
( > 4.000)
;io
15
A
- 15
Static ehamlier
143
900
1,100
2a loo
20-100
15
10
A
A
84
84
Ijarge ehamlier; 90 F; wind
speed without effect
104e
910
300
15
A
54
Ijow-flow ehamlier; 90 F
71
Cat
(400)
10
10 30
X
12
Low-flow ehamlier
37, 44a
lh>K
(800)
10
10 30
X
14
Low-flow ehamlier
37, 44a
Goat
(1,500 3,000)
30
15 -
A
9
Static ehamlier
143
Monkey
(1,500)
10
15
X
G
Low-flow ehamlier
37
flow rate, temperature, use of an anesthetic, and
variable delayed deaths due to secondary infections.
Wind speed or flow rate is only of marked im-
portance for toxicity when aerosol is present.37 "*4'*
HNS when present in part as fine drops is much more
toxic at high flow rates than at low flow rates.37 '40
At high flows a greater liquid dose is deposited on the
skin, from which it may be absorbed after exposure
both directly and indirectly as a result of licking and
inhalat ion of vapor.37140
A number of data are available on the toxieities of
the nitrogen mustards to animals exposed totally, by
inhalation only, and by laxly only.37 131 144 In the
ease of mice totally exposed to HNS, the absorption
of the agent from the body surface compares in im-
portance with that directly inhaled.37 It is doubtful,
however, that casualties among troops in the field
would be produced by the systemic effects of nitro-
gen mustard absorbed through the skin except when
the exposures are already more than sufficient to
produce vesicant effects of incapacitating sever-
ity iM.m
Relatively high concentrations of the nitrogen
mustards produce symptoms of irritation during ex-
posure •4.o.7i.7»,*,,>t»..12'.o' hut lower concentrations
arc without immediate effect.7'131 Symptoms then
develop only after a latency of one to several hours
and the most conspicuous pathological changes ap-
pear in the eyes and respiratory tract.84-7'-*3 m
I)ea(h in lethally dosed animals is usually delayed
for one day to two or more weeks, depending on
dosage. Detailed pathological studies have been
made rr.rs.si ,»,m is»,ho ui ,t«
Data on the toxieities and pathological actions of
nitrogen mustards administered percutancously,
orally, and by injection are presented in Chapter 22.
From the practical point of view it may be noted that
production of casualties from (he drinking of con-
taminated water could easily occur. The available
chemical tests are, however, sufficiently sensitive to
reveal potentially dangerous concentrations of the
nitrogen mustards and their toxic products of partial
hydrolysis11 ,1,200)
2
?
A
24
Static chamlier
121
(3,000)
5
10
A
24
Static chamlier
121
(5,500)
10
15
X
12
Low-flow chamlier
44 a
(3,500-7,000)
10
MKT)
X
16
Static chamlier
177b
(3,000- 6,000 r
10
5
A
20
Static chandler
121
(4,000-8,000)
20
10
A
30
Static chamlier
121
(3,000-6,000)
30
7
A
30
Static chandler
121
(>3,800)
60 120
9
A
8
Low-flow chamlier
13!
(2,.500 5,000)
240-450
9
A
14
Low-flow chandler
131
Rabbit
(> 1,200)
2
25
A
24
Static chamlier
121
(1,000-3,500)
5
26
A
24
Static chamlier
121
(4,400)
10
15
X
4
Low-flow chamber
44a
(7,000 14,000)
10
io< ?)
X
12
Static chamlier
177b
(3,000)
10
15
A
10
Stat ic chamlier
121
(2,000-8,000)
20
28
A
30
Static chamber
121
(3,(XX) 6,000)
30
26
A
20
Static chamlier
121
Cat
<1,400
10
10-30
X
8
Low-flow chamlier
4 la-
Don
(2,000)
10
10 30
X
4
Low-flow chamber
44a
Goat
(1,000)
2
13
A
8
Static chamber
121
( < SOO)
5
22 _
A
8
Static chamber
121
(< 1,700)
10
9
A
6
Static chamber
121
(<2,000)
20
9
A
8
Static chamlier
121
(1,000 2,000)
30
9
A
10
Static chamber
121
through clothing is of more practical importance
than action on bare skin because usually most of the
laxly surface, including the areas that are at the
same time the most sensitive and the most critical
for incapacitation, is clothed. In addition to vesi-
cancy through unimpregnated clothing, the degree
of protection offered by chlorarnide-impregnated
clothing and clothing containing activated carbon
mjnires considc• ration.
A further differentiation must l>e made lietween
situat ions in which the skin is relatively cool and dry
and situations in which it is hot and moist. As in the
case of H. high temperatures and humidities and
physical exercise augment the sensitivity of men to
SECRET 72
NITROGEN MUSTARDS
Table 8. Toxicity of 11X3.
Riven in parentheses.
The animals
were totally exposed. /.(Cf)5„’s that
are estimated very approximately are
Analytical'
.
(A)
Ol
KxposiiPe
Observation
nominal
Xumlier
iACtho
time
period
(X)
of
Species
(mjj min/m1)
(min)
(days)
cone.
animals
Xotes
Reference
Mouse
(1,700)
<2
18
A
132
Fine aerosol
144
500-000
10
14-15
A
58
Aerosol-free vapor
101a
.500
10
15
A
230
Low-flow chamber
37
300
10
15
A
60
High-flow chamber; aero-
sol present
37
(10.-,)
10
15
A
20
Vapor; wind tunnel; 05 K
37
1,700
10
10
N
160
lane-flow chamber
78
.-,70
10-100
15
A
130
Vapor, 00 F, 85% humiditv
1 Ole
Hal
(800)
0.25-2
20
A
104
Fine aerosol
144
1,700
10
15
N
28
Low-flow chamber
.54
(SOO 1,500)
10
15
A
IS
Low-flow chamlser
37
(? 1,000)
30
?
A
50
Static chamber; 85 F
139
670
10-100
15
A
60
Vapor, 00 F, 85% humiditv
104 c
Guinea pig >2,300
10
?
\
10
Low-flow chamlicr
37
>1,000
30
?
A
45
Static chamber; 85 F
130
Rabbit
(585)
3-15
15
A
12
Vapor; wind tunnel; 5.5 mph
05 F
—- 37
(1,000-3,000)
10
10
X
11
Low-flow chamber
37
(500)
10-18
15
A
8
Low-flow clmmlicr; vapor
only; 100 F
37
(830)
18 50
15
A
30
Low-flow chamber; vapor
only; 72 F
37
(>1,000)
30
?
A
31
Static chamber; 85 F
130
635
10 100
15
A
70
Vapor; 00 F, 85% humidity
104c
550 ±
Ioiir
15
A
60 ±
Field tests
6.5
Cat
(100 1,000)
10
?
A
32
Low-flow chamber
37
Dor
(400-1,500)
10
?
A
36
Low-flow chamber
37
<1,350
30
?
?
?
I/Ow-flow chandler
187
Goat
(500 1.000)
30
?
A
18
Static chamber; 85 F
130
the vesicant effects of the nitrogen mustards (see
Chapter 23).
So far as is known the time course of development
of injury and incapacitation due to skin injury ap-
pears to be comparable for H and the nitrogen mus-
tards (see Chapters 5 and 23). Some evidence exists
that nitrogen mustard bums are shallower than II
burns and heal more quickly.*9108 n715* On the other
hand nitrogen mustard burns have been referred to
as more tender and painful than H burns.*9191 How-
ever, a sufficiently complete and realistic determina-
tion by means of performance tests of the relative
extent and duration of incapacitation produced by
lesions due to H, HNl, IIN2. and HNS remains to be
made. Thusat present evaluations must l>ebased prin-
cipally on lesion-producing effectiveness rather than
on the more pertinent criterion of casualty-produc-
ing effectiveness. Furthermore there is no informa-
tion as to the effects of large dosages of nitrogen
mustard vapors upon masked troops. In the case of
H it is known that severe exposures under tropical
conditions produce incapacitation within one hour
of exposure tine to temporarily incapacitating nausea
and vomiting followed rapidly by the development
of very severe cutaneous injury."3 No evidences of
systemic injury have been apparent in any of the
man-chamber trials with HNl and HNS at the rela-
tively low dosages that have been utilized.1041’-*-'111
Vesicancy of the LiQrms
1. Cool and temperate conditions. For the produc-
tion of lesions on bare skin when free evaporation is
permitted and decontamination is not practiced,
tin* order of vesicant potency is fl > HNS > HN2
> HNl. The relative weights of the small liquid
drops required to produce blisters at 50 per cent of
the sites of application are;39 44'•e f*h i "91108
II 1
HNS 2-4
HN2 - 4-8
HNl >8
SECRET TOXICOLOGY
73
The agents fall in the same relative order when
evaluated by more realistic tests in which the sizes
of the lesions produced by large drops are com-
parcd.,4VIM-,i“ None of the other nitrogen mustards
and related compounds are as vesicant as HN2 or,
probably, as vesicant as HNl (see Table 1).o.h*u»i
When effective decontamination is practiced I to
5 minutes after contamination, II produces markedly
greater lesions than HN3.1*7 The positions of HNl
and HN2 are not known with certainty; HN2 may
lo only slightly inferior to H m and IlNl somewhat
inferior to HN2.*49 Antivesicant ointments (i.e.,
United States Mo and British A.G. No. 6) available
to Allied troops during World War II do not destroy
the nitrogen mustards as they do H. but. their bases
are good solvents for the nitrogen mustards. The
latter are effectively decontaminated by solvent and
mechanical action when large amounts of ointment
are applied and then wiped off.,w4*n*197 If the oint-
ment is left in place on the skin, the dissolved but
undestroyed nitrogen mustard may slowly exert its
vesicant action and the lesions produced by small
doses of H and 11N3 bi*come comparable in sever-
itv.M.m.e Dilute acids also exert a solvent, action on
the nitrogen mustards, and such oxidizing agents as
permanganate in aqueous solution may l>e utilized
as decontaminants. In addition, some chloramides
not in general use by the Allies during World War II
do destroy nitrogen mustard readily. Notable among
these are 8-43(5,
NCI, NCI,
I I
C.HS—C-=N—C =N—C=N
1 j
and the German Decontaminant 40,
O ('1 O Cl O Cl
111 II I II I
C—N—C—N-C-N
L_ ~~ 1
(see Chapter 24).
Through one or two layers of unimpregnated cloth
(he order of lesion-producing potency is II > HN2
> HNl ~ HNS.5"14*14* The order found for bare
skin is modified because of the importance of vapor
pressure for the transport of the agent through the
cloth and to the underlying skin.
Through cloth impregnated with CC-2 the nitro-
gen mustards gain in effectiveness relative to H lo-
calise of the comparative ineffectiveness of CC-2 as
a decontaminant for nitrogen mustard. Laboratory
data insufficient to permit a conclusive estimate sug-
gest the following order of potency; HN2 > H
> HNl > IIN3.9' Realistic trials under field con-
ditions are lacking.
2. Hot and humid conditions. The scanty avail-
able data do not permit an evaluation of the relative
potencies of the three liquids under tropical condi-
tions. The lesions produced by small doses of the
liquids on resting men and on men exercised to the
|K»inf of sweating under temperate conditions suggest
that the differences which would he observed among
the agents if they were tested under severe tropical
conditions might he less pronounced than those
which have been obtained on relatively cool, dry
skin.44'
Vksu anov of thk Vacuus
1. Laboratory evaluation of potency. Laboratory
data which relate to the production of lesions on
limited areas of the skin of the forearms of men not
acclimated to hot summer weather and exposed un-
der relatively moderate ambient conditions of tem-
perature and humidity demonstrate that on a dosage
basis HNS is equal to or slightly more effective than
II. that IIN2 is definitely inferior to both Tl and
HNS, and that HN1 is greatly inferior to each of the
three other agents (Tables 9 and 10).58M
Table 9. Vapor train tests of the vesicant potencies of
the vapors of H, 11X1, HX2, and 11X3.“
The subjects were at rest. T = 80 F.
Analytical Ct (mg min/m3)
for 50 per cent
resfMinses
Relative
Agent Krytheinas
Hlistcrs
dosage*
H <430
2,300
I
11X1 2,700 +
>21,000
>8
11X2 1,200 +
3,800
2 +
HX3 400 ±
1,800
0 .7 t-
* Reciprocal of vesicant potency.
Tabi.e 10,
Vapor cup tests of the vesicant potencies of
the vapors t
.f 11, UN 1, and HN3.«
The
subjects were at rest. T = 72-
73 F.
Kstimated median vesicating
dosage in mg min/in3
Relative
Agent
(/ = 5-60 min t
dosage*
11
3,500
1
11X1
IS,000
5 +
HX3
3,700
1.1
* Rwiprofftl of vifUfant latency.
These relationships for II, HNl, and HN3 are con-
firmed by arm-chamber studies at high temperatures
SECRET 74
NITROGEN ML'ST ARDS
Tahi.k 11. Basie man-chamber tests with II
and nitrogen mustard vapors:
Cubed States Army data.,""M‘-,l-r
T — 90 F. Relative humidity = 85 per rent. All subjects wore gas masks, shoes, and socks.
Additional
Vapor
exposure
Genital
clothing
Number
dosage
time
Season
protection
and protection
of men
(mg min in1)
(min)
Kffeets
//
Summer
('C-2 impregnated
shorts
None
3
106
10
Moderate erythema of neek.
back, and legs.
Summer
CG-2 impregnated
shorts
None
6
200
20 ±
Severe erythema of neck.
—
thorax, abdomen, and legs;
—
some delayed superficial
vesication.
- ■
uni
SnmmiT
(’('-2 impregnated
—
shorts
None
3
107
11
No effects.
Summer
CC-2 impregnated
shorts
None
3
211
*22
No effects.
Summer
CC-2 impregnated
shorts
None
3
285
30
Questionable erythema of
neck.
Summer
CC-2 impregnated
-
shorts
None
3
520
34
Mild erythema of neck; 1,
mild erythema of back.
Summer
CC-2 impregnated
shorts
None
3
689
41
Mild erythema of nock and
—
body.
Summer
CC-2 impregnated
— ‘ - . ■'
shorts
None*
3
940
44
i moderate and \ moderate
erythema of up|MT trunk.
Summer
CC-2 impregnated
shorts
None
3
1,030
29
| moderate erythema of axil-
— ■
larv folds; J mild cry-
—
thema of upper back and
neck.
■ — -
HN.i*
Winter or
I’n impregnated
early spring
shorts
None
2
90
15
No genital injuries: minimal
Winter or
Carbon-eontain-
erythema over exposed
early spring
ing shorts
None
2
90
15
skin, marked over neck,
- -
back, and anterior axillary
_
folds.
Winter or
Carbon-eontain-
- “
early spring
ing shorts
None
3
150
25
Generalized moderate cry-
thema at 20 hours which
had reached its maximum
and begun to decrease by
96 hours. Krythema most
—
pronounced on neck, hack,
—
and anterior axillary folds.
Winter or
Carhon-eont ain-
early spring
ing shorts
None
4
200
?
3 slight and 5 moderate erv-
thema of trunk and neek;
\ minimal erythema on
legs.
Winter or
Carlxin-rontain-
early spring
ing shorts
None
3
250
?
Slight erythema of trunk,
...
moderate erythema of
neck; § minimal erythema
of legs.
* Wind apecd in the chamber seemed to lx* without effect and has been dforcgi
ided in compiling this table.
SECRET TOXICOLOGY
Table 11 (Continued).
Season
Genital
protection
Additional
clothing
and protection
Vapor Exposure
Number dosage time
of men (mg min in') (min)
Effects
Winter or
Carlton-conlain-
U\J*
—
early spring
ing shorts
None
8 300 ?
* slight, ( moderate, and I
marked erythema of t runk;
more pronounced cry-
Winter or
('arbon-contain-
thema of neck; minimal
erythema of legs.
early spring
ing shorts
None
1 390 ?
Areas of vesication on trunk
and neck; marked ery-
thema with edema and
moist desquamation of
ears and preauricula r
—
areas; slight erythema of
scalp; minimal erythema
of legs.
Winter or
Ca (boils contain*
Xonimpregnated
t 3n0 ?
( slight erythema of neck;
early spring
ing shorts
2-piece herring-
—
J vesication of neck.
twine (will suit,
M5 ointment on
neck.
* Wind speed
in the chamber seemed to be without HTcct and ha** been d»regardf*d in eoinpiliiiK this table.
and humidities with the exception that, when tests
were made with sweating observers acclimated to
hot summer weather, UN I assumed a much more
favorable relative position, requiring only 1.2 to
1.6 times the dosage of H to produce equivalent
lesions.109 The effectiveness of II. HNI, and HNS
vapors in these tests was little affected by the inter-
position of a layer of unimpregnated cloth.109
2, Man-chamber evaluation of potency. The only
available man-chamber tests of the effects of nitro-
gen mustard vapors on observers wearing no cloth-
ing or unimpregnated clothing are summarized to-
gether with representative data for IT in Tables 11
and I2.,tab e ,, r 110 1,1 These data relate to a chamber
temperature of 90 F and relative humidities of 65
and 85 |x*r cent, and to the production of injuries
corresponding only to relatively mild partial dis-
ability.11*
Although the two groups of data show some dis-
crepancies. it seems reasonable to conclude that, in
warm or hot weather and against troops provided
with gas masks but not with protective clothing,
HN3 vapor may approach H vapor in potency as a
casualty-producing agent, particularly when the
genital region is unprotected. HN1 vapor, except
possibly on freely sweating men acclimated to hot
weather, appears to lx- definitely inferior to II and
HN3. The laboratory findings (see the preceding
section) suggest that HNI vapor would be markedly
inferior under cool or temperate conditions.
3. Evaluation of protective. clothing. The merits
and limitations of the ('C-2 impregnated and the
earbon-containing type's of protective clothing are
reviewed in Chapters 26 to 30. In brief, the data 72 73-
■09.111.ns reveal that CC-2 impregnated clothing offers
excellent protection against II, considerable pro-
tection against HN3, and relatively little protection
against TIN I. Thus, in the tropics against troops
protected by this clothing, HNI vapor may be a
more potent casualty-producing agent than H. The
relative positions of ti and HN3 are not known but
may not lx* important because of the high degree of
the protection afforded against both agents.
The best experimental types of carbon clothing
now available offer protection against such large
dosages of II. HNI, and IIN3 (presumably also
HN2) that differences between the dosages of the
agents required to "break” this clothing become of
minor consequence.
1. Protection afforded by ointment. Prophylactic
use of S-330 ointment, and presumably other oint-
ments containing the chloramides available to Allied
troops during World War II, offer little protection
to skin exposed to nitrogen mustard vapor.109
6.1.1 Eye-Injurant Action
Numerous observations on the effect of the nitro-
gen mustards on human and animal eyes demonstrate
that HNI, HN2. and IIN3 are eye-injurants more
insidious than IT and more or less comparable with it
SECRET 76
NITROGEN MUSTARDS
Table 12. Basic man-ehaniber tests with 11 and nitrogen mustard vapors; I'nited States
Naval Research Laboratory data.'1"111
T = 90 F. Relative humidity = 65 per cent. Fxposure time = 60 minutes. All sub-
jects wore gas masks, uniinpregnatcd outer and under clothing, caps, shoes, and socks. The
maximum lesions sustained on various parts of the body over a period of approximately a
week were graded according to the following numerical scale:
0 = No reaction.
1 = Mild erythema.
2 = Moderate erythema.
3 = Intense erythema.
-1 = a. Krylhema with edema.
b. Maceration of axillary skin.
c. Dry sealing of scrotum.
5 = a. Vesicle.
b. Numerous pinpoint vesicles.
e. Crusting or ulceration of scrotum or axilla.
No. of
crusted or
Va|M»r
Severity of injury
ulcerated
Numl>er dosage
Rest of
scrotal
Season of men (mg min m3)
Neck
Scrot um
body
lesions
—
II
March 6
. 50
0.3
0.0
0.1
Julv 5
50
1.2
0 2
0.5
-
March 6
100
1.2
1.2
0,7
July 5
100
1 .!>
0.8
0.9
April 10
150
2.0
0.3
0.7
July 6
150
3.0
2.2
2.1
April 10
200
2.2
2.1
1.2
•JufjT 0
200
4.0
3.2
2.4
April __ 15
250
2 4
3.2
1.0
Julv 6
250
4.2
3.7
2.8
November 0
300
3.3
0,0*
2.4
... - — —
March 5
300
3.4
0.0*
2.9
—
—
JIM
August 10
100
1.3
1.2
0.3
0/10
August 10
200
3.4
1.4
1.2
0/10
August 10
300
3.3
4.6
1.7
7/10
January 8
300
1.0
0.6
0.3
0/10
January 4
450
1.8
1.5
0.6
0/10
January 6
700
2.2
4.0
0.6
4/6
. n . - —
uns
-— .
September 8
50
1.8
0.5
0.2
0/8
Septemlier S
100
3.0
1.9
0.8
1/8
September 8
150
4.0
4.0
I.S
6/8
August 8
150
2.0
0.0*
0.2
OS*
February 6
150
1.5
0.3
0.7
0/6
February 6
250
. 2.5
1.5
0.8
0/6
February 8
350
4.9
3.6
1.7
6/8
* Subjects wore f’C-2 impregnated shorts.
-
in potency .sh.jo.jmi.nub.
c.l86a,112,US,ll!t.l2S.I32,lJS,lM.l3s.M2,141»,U2,lS«.t74.176.1SO.I84
pyl-6fs(d-chloroethyl)amine and isopropyl-6is(0-chlo-
roethypaminc appear to lie somewhat less potent.44'-
81 .134
Effects of the Vapour in* Small Dosages ox
Ik man Eves
The results of observer tests with H, HNl, IIN2,
and HNS as vapors demonstrate that all four agents
are roughly comparable on a potency (dosage) basis
in eye-injurant action. Provisionally it would appear
that HN2 and HNS may be somewhat more potent
than H, and HXl somewhat less potent, but the dif-
ferences cannot be considered to have been estab-
lished with significance (see the following section for
animal data). The importance of the human eye
data merits their more detailed review as follows.
1. II. Critical summary and review 112 m of the
four available sets of data n>r» n».H7«.i9«. suggest that
SECRET TO \ ICO LOG \
77
50 mg min raJ (/ < 8 hours) is the maximum dosage
to which unmasked personnel may lx* exposed with-
out danger of significant eye damage, and that
100 mg min m3 (t = 0 minutes to 7 hours) is the
threshold dosage for production of partial disability.
Extrapolation from the data leads to the estimate
that for offensive purpose's 200 mg min m* (I =
6 minutes to 7 hours) would suffice to produce in-
capacitating conjunctivitis and blepharospasm, with
lacrimation, photophobia, and soreness, and perhaps
with some corneal damage, in the majority of men
for a period of 2 to 7 days, beginning 3 to 12 hours
after exposure. II vapor is somewhat less effective at
very short (i.e., 1 to 2 minute) and very long ex-
posure times.
2. 1IN1.14- A dosage of 90 mg min m* is believed
to represent the beginning of the human casualty
zone, on the basis of tests in which one eye of each of
21 observers was exposed in a respirator facepiece to
5 I min of 11N1 vapor (Cl = 37 to 90. t = 5 to 67
minutes). There was no serious change of vision ex-
cept for three men, exposed to dosages of 41.56, and
90 respectively, who did not think they could shoot
a rifle for 48 hours. Only one of three men exposed to
a dosage of 90 was a “casualty” in this sense. There
was an average delay of 12 hours in (he development
of symptoms, which included gritty feeling, lacrima-
tion, photophobia, blepharospasm, headache, blurred
vision, conjunctival hyperemia, corneal flecks, epi-
thelial* I >edewing, and punctate staining with flu-
orescein. Minor symptoms persisted in one case for
as long as 21 days. The conclusions of (he re[>ort are
transcrilted verbatim as follows:
a. A dosage of I1N1 of 90 mg min ra3 Is prob-
ably the beginning of the human casualty
zone, but with ocular idiosyncrasy casual-
lies can occur at lesser dosages.
b. The average interval between exposure and
onset of symptoms was 13 hours.
c The most common complaint was “gritty”
foreign body sensation in the eye.
d. The most common lesion was flecks of the
corneal epithelial surface which disappeared
spontaneously in 1 to 15 days. Conjunctival
hyperemia occurred almost as frequently.
e. The most annoying symptom was pain in
and behind the eyeball.
f. Other complaints were lacrimation, photo-
phobia, and blurred vision, although there
was never any reduction in visual acuity or
accommodation.
g. Blepharospasm occurred in only two ob-
server and myosis in only one.
h. So far as can be judged from the results ob-
tained, the dosage (Cl) of HNl vapor is a
sufficient index to the degree of damage
anticipated, even though the exposure time
be varied from 5 to 60 minutes (but see
below).
3. HN2.15* Dosages of 40 to 55 mg min m* (t =
(5.5 and It) minutes, respectively) are believed-to
represent the lowest limits of exposure necessary to
produce “disablement” — i.e., certain cases would
call for medical aid and. to an extent depending on
transport and medical facilities, would lx; unable to
take part in operations for a minimal period of 1 to
2 weeks. This conclusion was based on experiments
in which an unstated number of men wearing oro-
nasal masks were exposed in a man-chamber to
dosages of 10 to 55 mg min ra*. The performance of
additional human experiments at higher dosages was
considered to involve an unreasonable risk. There
were no subjective symptoms during exposure. From
8 to 15 minutes after exposure lacrimation and a feel-
ing of grittiness under the lids developed. After 6 to
10 hours the following symptoms had set in: lacri-
mation, photophobia, blepharospasm, and pain in
the eyeball severe enough to prevent sleep. At 24
hours the symptoms were similar but the pain had
become less seven*. There was pupillary constriction,
conjunctival congestion, deep ciliary congestion, and
threshold edema of the corneal epithelium, but no
staining with fluorescein. The condition was resistant
to mydriasis with I percent homatropine but pupil-
lary dilatation and relief of blepharospasm was
achieved by two applications of 1 per cent atropine.
The observers gave their opinion that their efficiency
as soldiers would have been seriously impaired from
6 to 10 hours onward. The duration of the symptoms
was not stated. The report recommends that a dosage
of 70 mg min m3 be aimed at as a minimum for of-
fensive purposes.
4. HN3.""1’'' Of four observers exposed to a dosage
of 20 mg min m3 (I = ?), none experienced any sub-
jective symptoms but all showed moderate conjunc-
tival injection. Their corneas were grossly normal
but examination with the slit lamp revealed moderate
to marked epithelial edema. Of three observers ex-
posed to a dosage of 42 mg min/m* (1 = 7 minutes),
SECRET 78
MTKOGEN Ml STAKUS
Table 13. Eye damage produced in rabbits by the vapors of H, HXt, and
Eight animals were exposed to each agent, at each dosage. The eye damage was graded according to an
arbitrary numerical system1"** which took account of changes in the iris, cornea, conjunctivas, ami lids. The
analytical dosages of the agents were determined by methods adequate to integrate low concentrations
over long times.
K\|>osure time
(min)
I )osagc
(mg min/ni
11 11X3
Eye Dosage
i3) damage (mg min nr1)
Kye
(tamale
MX I
Dosage Eye
(mg min/in*) damage
2
440
20 353
21
485 30
384
19
439 29
10
3:40
29 410—
23
650 15
370
24
389 12
CO
420
23 4 IS
25
4:4,5 12
434
24
400 10
. 200
420
I7
240
347
21 4lt
16
530 11
406 10
360
330
13
all developed lacrirnation, photophobia, and a feeling
of grittiness in the eye. They exhibited marked con-
junctival injection. Their corneas were grossly nor-
mal and did not stain with fluorescein but examina-
tion with the slit lamp revealed epithelial edema and
slight infiltration of the anterior stroma. One de-
veloped moderate edema of the lids. All three were
improving both subjectively and objectively on the
fourth day after exposure. On the basis of the brief
available description it would appear that HN3 pro-
duced effects comparable to those found for 11 in one
investigation l9a b and more severe than those found
for 11 in two other investigations.M9,47“
The clinical reports of plant accidents indicate t hat
the development of eye symptoms due to the vapors
of IINl and HN3 were delayed for several hours.*9174
The same delay was experienced by some workers ex-
posed to HN2 vapor, but others developed eye irri-
tation, lacrirnation, and photophobia immediately
after exposure.17*
Effects of Vapors on Animal Eyes
Although the animal (i.e., rabbit) eye is consider-
ably more resistant to H and nitrogen mustard va-
pors than is the human eye,7S11*,14! it may be assumed
that the relative potencies of the different agents can
lie determined in animal tests.
The most satisfactory available set of comparative
data is summarized in Table 13.,4h l0** The results
suggest that the rabbit is approximately as suscep-
tible to HN3 as to H. IINl is probably more potent
than H and HN3 at very short exposures (i.e., 2 min-
utes) but significantly less potent for exposure times
of 10 to 240 minutes. The results of less rigorously
controlled earlier work witli dogs exposed for 10 min-
utes suggest that HN1, HN2, and HN3 produce
threshold eo meal ilam age at somewhat lower dosages
than H; that at low dosages HNS is the most potent
eye-damaging agent, followed by FIN1, HN2, and H;
and that at Itigher (buTstill moderate) dosages the
differences among the hair eompounds are less con-
spicuous;Mc
In tests with relatively large vapor dosages which
produced severe ocular injury, it was found that the
dosages required to produce equally severe super-
ficial corneal and conjunctival injury were about the
same for each of the t hree nitrogen mustards.67b With
equally severe injury to the superficial corneal tis-
sues, however, the damage to t he deeper tissues (i.e.,
iris and ciliary body) was much the greatest with
1IN2, intermediate with IIN3, and least with I IN 1
and H.s7b The severity of the deep ocular effects pro-
duced by HN2 make it a particularly dangerous
agent from the standpoint of severe and permanent
eye injury.
The results of additional studies on the effects of
nitrogen mustard vapors on animal eyes are to be
found in the following references, some of which con-
tain more or less complete histopathological analy-
S(.S 44(1,43,S7»,.-,S8,70,71.84. H5,123,138.142
The clinical and pathological studies with I1N2
have been reviewed in detail.“
Liquid Contamination of the Eve
Tests on animal eyes with small liquid drops (i.e.,
0.5i mg) of H, IINl, HX2, and HNS demonstrate
that all the agents produce such severe burns, fre-
quently with permanent loss of sight, that any differ-
SECRET KESLLTS OF FIELD TRIALS
79
euees in potency which may exist are relatively un-
important to an evaluat ion of their relative merits as
offensive agents.4**’<'1"1 ,23 , 52 1“ ’4# The observations
tend to emphasize the similarity of the lesions pro-
duced by II and HX3 and the more severe character
of the injury that HXT2 produces in the deeper struc-
tures of the eye.
The effects of small droplets, and in the wind tun-
nel of sprays consisting of fine droplets and vapor,
have also been studied in animal experiments in
order to assess (he relative effectiveness of the agents
in the initial clouds produced by bursting muni-
tions.1-’3 ,SUIK The results indicate that 11X3 may be
slightly less damaging, and HX2 slightly more dam-
aging, than IT. In any event the differences are not
marked.
Decontamination and Treatment
Decontamination can be effected practically only
by prompt lavage of eyes contaminated with (he
agents in the liquid form. There is some evidence that
lavage is of less value with 11X3 than with II.140
Prompt use of dithiocarbamates or of BAL ointment
may be of limited value.46" 57c "»
The subject of treatment has been authoritatively
reviewed.62 The susceptibility to infection of eyes in-
jured by nitrogen mustard and the value of various
types of chemotherapy have recently been investi-
gated, IMo‘l
6.5 RESULTS OF FIELD TRIALS
Field trials with the nitrogen mustards have in-
cluded tests of the vapor return from contaminated
terrain and study of casualties in animals exposed to
clouds of liquid drops and vapor produced by burst-
ing munitions. Xo observer tests have been made to
determine the vesicant effects of evolved vapor in the
field or the hazard to trav ersal and occupation which
is presented by the liquids on soil and vegetation.
The results of (he tests reviewed in Section 6.2.4
attest to the excellent stability of HX3. the probably
adequate but marginal stability of HX1, and the
questionable stability of HX2.
HNl, 11XT2, and 11X3, as well as H. dispersed from
explosive munitions as clouds of liquid drops and
vapor can produce profound eye damage and serious,
often fatal, respiratory injury in unprotected ani-
mals exposed on open terrain (see references cited in
Section 6.2.4). However, such trials may have only
limited bearing on the general utility of the agents
in warfare.
Kvoution of Vapor from Contaminated Terrain
Results of field trials (see Table 14) conducted dur-
ing warm weather at Hushnell, Florida, are avail-
able.64** The tests included both annulus trials and
trials with single, statically exploded M47A2 bombs.
They lead to the following tentative conclusions."2
1. When terrain is similarly contaminated with
HX3 and Levinstein H, the vapor dosage of 11
evolved during the first few minutes is five to eight
times us great as that of HX3, as would be predicted
from the relative volatilities of the agents. With the
passage of time the relative dosage of evobed HNS
vapor becomes progressively greater until, after the
lapse of sufficient time for the completion of the
evaporation process, the total dosages of the two
agents become approximately equivalent. The time
interval after which the evohed dosage of HX3 at-
tainsany specified fraction of the U dosage depends on
the meteorological conditions and the size of the
liquid drops with which the terrain is contaminated.
2. In trials under semitropical meteorological con-
ditions with single, statically fired M47A2 bombs
charged HX3 or Levinstein II, the areas over which
toxieologically significant dosages of HX3 vapor
were obtained within 4 hours amounted to substan-
tial fractions of the areas over which equivalent
dosages of H vapor were obtained (see Table 14).
3. It is estimated 64 that in large-scale attacks un-
der the semitropical conditions prevailing during the
Florida trials the 4-hour vapor dosages obtained
from equal expenditures of M47A2 bombs charged
HX3 or Levinstein H would be:
4-hour vapor dosages,
Meteorological conditions
HX3 as per cent of 11
Woods, clear day
45
Woods, clear night
20
Open, clear day
65
4, At the lower surface temperatures character-
istic of cool or temperate weather, the times after
contamination at which the evolved HX3 vapor
dosages would attain the above percentages of the
H dosages would he greatly prolonged.
5. Under semitropical meteorological conditions
the persistencies of vapor evolution by 11 Nit and
Levinstein H arc not markedly different. Both are,
of course, much less than that of HT (see Chapter 5).
0. 11X3 vapor evolved from contaminated terrain
in the annulus and bomb trials was proved by bio-
assay tests to be toxicologically effective. On the
basis of the respiratory and ocular lesions produced
in rabbits exposed at intervals up to more than 24
SECRET 80
N IT ROC EX MUST A RDS
Table 11.
Results of field trials with IIX1, 11X3, and II; single bomb tests,“MMir
Avg
Avg temp
Area
(artillery
squares)
within the
contours for the
wind
Average
gradient
stated dosages (Cl's in
mg min/nv’) for 0 to 0 +
Agent and
sjx-ed at
ground
r,m - t
4 hour sa
mpling at
a height of 12 inches.
test
Bomb
2m (mph)
temp {(')
in the o|X‘ti
50
100
250
500
1,000
2,500
Meadow, lapse <
•otulilions
11M, lost 4
M47A2
4.43
23.SI
-1.11
0.98
0.01
0.32
0.17
0.09
II, predicted
M47A2
4.43
23.81
-1.11
1.01
0.59
0.29
0.17
0.10
11, observed
M47A2
4,3
17.0
-1.2
0,81
0.51
0.22
0.13
0.07
11X1, tost t>
M47A2
4.0
23.0
+0.7
1.72
1,(3
0.48
0.27
0.15
0.06
II, predicted
M 17A2
4.0
23.0
+0.7
1.77
1.03
0.51
0.29
0.17
0.08
II, observed
M47A2
4.41
21.09
+0.52
2.18
1.22
0.55
0.31
0.17
11X3, test ti
M47A2
3.3
35 2
-1.1
0.76
0.50
0.30
0.18
0.11
0.00
11, predicted
M47A2
33
35,2
1.1
1.34
0.79
0.40
0.23
0.14
0.07
11. observed
M47A2
4.S 32.8
-1.32
0.77
0.48
0.25
0.16
0:10
0.06
II, observed
M70
4.2
36.2
-2.24
0.71
0.45
0.23
0.14
0.09
0,05
Forest, lapse conditions
HX3, test 1
M47A2
1.06
29.3
-1,05
0.57
0.35
0.20
0.13
0.09
0.06
11, predicted
M 47A2
1.06
29.3
- 1.05
0.92
0.59
0.35
0.24
0.10
0.08
11. observed
M47A2
0.1)
27.5
-1.10
1.10
0.72
0.41
0.25
0.10
0.09
11, observed
M70
0.5
26.5
-1.03
0.90
0.67
0.45
0.32
0.22
0.12
11, observed
M70
0.9
28.S
-1.08
0.45
0.30
0.20
0,14
0.10 —
0.07
Forest, inversion
conditions
11X3, test 5
M47A2
0.5
24.5
+0.3
1.18
0.04
0.24
0.12
0.08
0.05
11. predicted
M47A2
0.5
24.5
+0.3
3.83
2.38
1.21
0.71
0.38
0.19~
11, oliserved
M47A2
0.6
21.5
-1.45
1.16
0.00
0.36
0.23
0.13
0.06
II, observed
M 47A2
0.5
19.5
+ 1.70
4. IS
2.78
1.27
0.70
0,20
0.08
II, observed
M70
0.6
25.5
+0.80
2.75
2.02
1.38
0.84
0.47
0.17
hours after exposure, HN3 vapor was significantly
more potent than H vapor.
7. When terrain is similarly contaminated with
HNl and Levinstein II in the form of large drops in
annulus trials conducted in the open in warm
weather, the initial rate of vapor evolution was
greater for HNl than for H, as would be predicted
from the relative volatilities, and the 4-hour dosages
of HNl were nearly twice those of II.
8. In the available single-bomb trials in the open
under semitropical meteorological conditions the
4-hour dosages of HNl vapor were approximately
equal to those obtained with H in similar tests (see
Table II). Approximately 90 per cent of the total
evolved dosage of HNl vapor had been attained
within this time.
9. Analysis of the data indicates (hat the destruc-
tion of HNl during the explosion or, subsequently,
by inactivation on soil and foliage may have been as
much as 30 per cent greater than the loss of II. Tak-
ing these results in connection with those of British
annulus trials155 which indicated 50 per cent de-
struction of HNl on soil, it seems probable that large
variations in per cent destruction may be expected,
depending on the munition utilized and the char-
acter of the terrain upon which the agent is deposited.
Even greater variations might he expected in the
case of HN2.
10. HNl vapor evolved from contaminated ter-
rain in (he annulus and bomb trials was proved by
bioassay tests to be toxicologically effective. In
terms of the respiratory and ocular injuries produced
in exposed rabbits, it was somewhat less effective on
a dosage basis than HNS vapor under similar con-
ditions.
6 6 EVALUATION VS WAR GASES
The instability of HN2 disqualifies it from serious
consideration for use as a war gas. Isopropyl-fus(j8-
chloroe(hyl)aminc is also disqualified because its
somewhat, inferior toxicological potencies are not
counterbalanced by other advantageous properties.
Thus only HNl and HN3 remain as potential sub-
stitute persistent agents for II. In Table 15 are sum-
marized the properties of II, IINl, and HN3 which
bear most directly on an evaluation of their relative
merits and limitations.
The judgment of the present reviewers is in accord
with the principal conclusions of previous assess-
ments:1,2 l,H (I) that IINI and HN3 do not possess
the general utility of H as an offensive agent; and
(2) that in so far as incapacitation of masked enemy
SECRET EVALUATION VS WAR CASES
Tabi.k 15. Properties of If, HN1, and Il\3 hearing on their potential effectiveness
as war gasos.
Pri>|srty
11
UN i
11X3
Storage stability
GimmI
Satisfactory
Kxeellent
Stability On explosion of
Good
Probably sufficient
Good
munitions
Stability on terrain
Good
Good to poor, depending
Good
on the nature and
Density (g ml, 25 (')
1.27
moistness of the ter-
rain
1.09
1.23
Load carried bv M47A2
tilt (pure 11) 71 (Levin-
til
67
Iwnnb ( lb)
stein 11 = 53 lb of
active agent)
Freezing point ((')
11.2 (pure) ca. 8 (I feasible
. ages
layers of CC-2 impreg- _
naled clothing
Injurv-prodneing effective-
Ineffective in reasonably
Ineffective in reasonably
Ineffective in reasonably
ness of vapor against
attainable dosages
attainable dosages
attainable dosages
masked troops equipped
with clothing containing
-
activated carbon
Relal ive injury-producing
1
l_l
3 1
<8
effectiveness of liquid on
bare skin
Relative injury-producing
?
?
?
effectiveness of liquid
-
through (.'( -2 impreg-
—
nated clothing
troops not equipped with chloramide-impregnated
clothing is the primary objective in the use of a per-
sistent- agent, HN1 and IIN3 do not possess the of-
fensive potential of H. At the present time, however,
it is pertinent to arid a discussion of two additional
points.
1, The lack of reactivity of 11X1 and 11X3 with
the chloramides used in the United States and Brit-
ish impregnated clothing of World War II led to
the inference that this ty|ie of clothing would afford
little protection against the vapors of these agents,
and that they would therefore lie more effective
casualty-producing agents than H against troops so
equipped."2118 Recent man-cham1»er tests at 90 F
reveal, however, that subjects exposed in 2 layers of
CC-2 impregnated clothing to 5,000 mg min/m*,
and in 1'2 layers to 1,000 mg min/m3, of HN3 vapor
failed to sustain injuries of incapacitating severity.7*
'Phus CC-2 impregnated clothing affords marked
protection against IIX3 vapor, although not neces-
sarily so much as against II vapor. The explanation
of this unexpected finding is not at hand. On (he
other hand it has been confirmed that CC-2 impreg-
nated clothing affords little protection against UNI.77
However, this lack of protection is at least partially
offset In the additional evidence that HX1 vapor is
relatively ineffective as a vesicant, except possibly
in very hot weather.72
2. It was the intention of the German Army to
use HN3 in high explosive-chemical shells. In the
SECRET 82
NITROGEN MISTAKDS
opinion of the reviewers, tins means of exploiting
HNS merits careful evaluation.When HNS is used
in this way as a harassing and casualty-producing
agent, no other known gases except the Trilons (see
Chapter 9) would be expected to approach if in ef-
fectiveness. It is believed that in high-explosive
bombardments an occasional high explosive-chemical
shell charged TINS and indistinguishable upon de-
tonation from ordinary high-explosive shell would
have l»een used. HNS possesses the stability to with-
stand destruction during the explosion of the shell
and the lack of odor to escape ready detection except
by chemical methods. It is believed that the poten-
tial harassing and casualty-producing effects of the
vapor slowly evolved from the contaminated terrain
might exceed those of the initial cloud. The duration
of danger from the vapor, the time intervals required
for the evolution of casualty-producing dosages, and
the areas over which effects would be produced would
depend on meteorological conditions. As an example
of the order of magnitude of the hazard, however,
reference may be made to the field trial data re-
viewed in Section 0.5 and Table Id. It will be noted
that in warm weather explosion of a single M47A2
bomb (containing 07 pounds of HN8) resulted within
I hours in the attaining of a dosage of 100 mg min m*
of vapor over approximately one-half of an artillery
square, and of 250 mg min nrTover about one-fourth
of an artillery square. A dosage of 250 mg min in*
should more than suffice to produce total disability
of several days’ duration due to eye injuries, and
possibly seven' respiratory injury as well.
SECRET Chapter 7
ARSENICALS
Marshall (lairs, Jonathan IT. Williams, and John .1. Zapp
7.1 INTRODUCTION
IN ji'lv 1917, the Germans not only introduced
mustard gas into World War I, but also employed
for the first time an arsenical chemical warfare agent,
diphenylchlorarsine (DA). Other arsenical agents
were employed by the Germans in rapid succession,
phenyldichlorarsine (PD) in September 1917, ethyl-
dichlorarsine in March 1918, diphenylcyanoarsine in
May 1918, and ethyldibromoarsine in September
1918. Although lewisite ami adamsite were not actu-
ally used in battle, the Allies were preparing at the
end of World War I to use /3-chlorovinyldichIorarsine
(lewisite) and diplienylaminechlorarsine (adamsite),
and were seriously considering the use- of methyl-
dichlorarsine and arsine itself.
There was a distinct feeling on the part of the
Allies that the Germans did not obtain (he maximum
effectiveness from the arsenicals which they used lie-
cause of technical difficulties in methods of disper-
sion, and further that some of the agents which did
not receive battle trial (e.g., lewisite and adamsite)
might become the most effect ive agents of their class.
In view of this, it was natural that attention again
be turned to the arsenical agents at the beginning of
World War II. Accordingly, both the British and the
Americans carried out extensive investigations on
(1) improved methods of preparation of the known
arsenicals, (2) the preparation of small quantities of
new arsenicals, and (3) the physiological action, toxi-
cology, and assessment of military value of these
agents. Although considerable progress was made in
the first two categories, none of the arsenical agents
proved to offer much promise of success in battle for
reasons which are detailed below.
7.2 CHEMICAL SECTION
7.2.1 Lewisite
Lewisite. develo|»ed during World War I, is un-
doubtedly still the best arsenical for gas warfare.
(For a summary of work to 1940, see the bibli-
ography.)'" The preparation of the agent by the
original procedure 104 was complicated and danger-
ous; it involves (he reaction of acetylene with arsenic
trichloride, uring aluminum chloride as a catalyst.
The reaction yields three products:
Asti, + II--C==C—. H —>■ C1CH -CTIAsCl/—►
1^1
(CK’H =CH),Ast'I
I.-2 L3
When aluminnm chloride is used as the catalyst, the
very vigorous reaction leads to a mixture in which
1.-2, 1.-3, tar, and an explosive material are present
with the desired lewisite. The optimum yield of L-l
in this scheme is about 20 per eent.M*"M< 11 was highly
desirable, therefore, to search for other catalysts.
The first work with a catalyst other than AK'U
was carried out in Great Britain in 1938,28Sa where it
was shown that acetylene can be made to react di-
rectly with arsenic trichloride in hydrochloric acid
solution using mercuric chloride as a catalyst. The
yield of L-I was 80-85 per cent based on the arsenic
trichloride and 75 per cent on the acetylene. The
main drawback to this process was the very corrosive
nature of the catalytic solution. A pilot plant oper-
ated by the British at Sutton Oak was found capable
of producing 10 tons per week of “stripped lewisite,”
which analyzed: L-l, 83.7 per cent; L-2, 11.5 per
cent; arsenic trichloride, 2.8 jjer cent; solvent (chlo-
rinated hydrocarbon) 2.0 (>er cent.2**6' W ork in this
country13*'4'14* showed that a batch process for L
using a mercuric chloride catalyst is economically
advantageous.
Work on other catalyst systems proved cuprous
chloride used in conjunction with ethanolamine hy-
drochloride to be one of the best, both for batch
and continuous operations.*-,tt-,*ll88’,®*-200-2',*f*!*0*
Although the reaction rate is somewhat slower
than with IIgCl», the product is cleaner and there is
less of a corrosion problem. It was also shown 2011
that (he cuprous chloride process gives 50 per cent
more production and 5 per cent greater acetylenation
efficiency, and that only one-half the amount of
thionyl chloride or phosgene-hydrochloric acid is
needed in treatment for sludge removal. A plant,
operated by this process at, Sutton (3ak produced
10 tons per week of “stripped lewisite.”28*'
Many workers recognized the desirability of a con-
SECRET 84
ARSENIC \LS
tinuous vapor phase process for the preparation of
L whereby a mixture of arsenic trichloride vapor and
acetylene could be passed continuously over a cata-
lyst. Some degree of success was attained by the use
of mercuric oxide suspended on alumina in an all-
glass reactor.*4 With antimony trichloride as an
activator for the mercuric oxide catalyst, the con-
version was from 30 10 per cent, with yields of 40-
GO jxw cent during the first hour; however, the life of
the catalyst was <|uite short.55
Early in World War II, it became apparent that
there existed a shortage of pure arsenic trioxide used
in the preparation of arsenic trichloride for lewisite
production. Consequently two programs were in-
augurated: (I) the conversion of crude arsenic tri-
oxide to arsenic trichloride; and (2) the use of arsenic
trichloride containing impurities in lewisite produc-
tion by the mercuric chloride process. In a study of
the latter problem it was shown that arsenic trichlo-
ride from crude arsenic t rioxide can lx* used directly
in a lewisite plant. Incidentally it was indicated that
slightly higher absorption rates were obtained when
either 2 per cent antimony trichloride or 1 per cent
ferric chloride had been added to the arsenic tri-
chloride,4-’ This demonstration led to the observation
that, when antimony trichloride is included in the
catalyst layer in the mercuric chloride process, the
output of lewisite is materially increased.55 In pilot
plant operations, it was found that, when the same
volume of SbC'L-containing catalyst (26 per cent
Sb(’L added to the standard HgCI* catalyst) is used
in the standard Hgt’b batch process, the time re-
quired for acetylenation is reduced by about 40 per
cent, whereas the amount of Hg present is 72 per
cent of normal.2"5
The problem of using crude white arsenic in
the production of arsenic trichloride was investi-
gated first on a laboratory scale and then in a pilot
plant.50 57 1% 197 With the use of three different raw-
materials, one of them containing only 51 per cent
arsenic trioxide, for reaction with sulfur monochlo-
ride, yields of 95 per cent baser! on both arsenic anti
chlorine were obtained in pilot plant runs. \\ itli this
experience as a background (he process was trans-
ferred to the Pine Bluff Arsenal,*2 where about 80
tons of specification-grade arsenic trichloride was
produced from two lots of crude arsenic trioxide re-
covered from ore of the Gold Hill, Utah, deposit. A
yield of 95 |H*r cent was obtained based on the
arsenic content of the crude arsenic trioxide. Prac-
tically all of the arsenic trichloride produced in the
experimental runs was consumed in the lewisite
plant with satisfactory results.
It was also demonstrated that arsenic trichloride
of high purity can be prepared from either refined
white arsenic or from low-grade arsenic crudes and
hydrogen chloride in yields of 97 to 99 per cent based
on the arsenic content of the raw' materials.51
In connection with the use of lewisite as a chemical
warfare agent it was necessary to study its corrosive
effect on shell steel. It was shown that plant-grade
lewisite produced by the mercuric chloride process is
practically without action on shell steel (No. 1045)
and may be stored in such steel for long periods of
time at tropical temperatures with insignificant cor-
rosion.-* Under these conditions no pressure is devel-
oped and no deterioration of the lewisite results.
Phosphorus pentoxide may be. used to decrease cor-
rosion slightly, to eliminate the slight rust formation,
and to prevent tin1 increasedn moisture content under
damp storage conditions.
Through other st udies it was found that a 1 1 mix-
ture of lewisite and Levinstein mustard is far more
corrosive than either constituent alone, and that a
1 1 mixture of lewisite and (hiodiglycol mustard is
only one-tenth as corrosive as (he other mixture.29 31
The conclusion reached, therefore; is that pure mus-
tard must be employed if mixtures of it with lew isite
are to be used in chemical warfare.
Several investigations were made in order to dis-
cover agents other than BAL (2,3-dimcrcaptopro-
panol-1) which might serve, to detoxify lewisite or
act as antivesicants for it. In a study of the reaction
products of lewisite and six different dithiols 10 it
was found that the properties of the compounds are
best explained by cyclic formulas of the types:
S—CUR S—CHR
z / \
CICH -ATI As CK’II—CHAs CHIU
“ \ — \ -/
S-CH2 S—CHR
In a study of the reaction of lewisite with thiols, al-
cohols, and amines it was shown that the competitive
rates of formation, or the stability at equilibrium, or
both, of bonds involving arsenic are in the order
As—S > As—O > As -N; hence o-ditIdols appear
to be the most satisfactory reagents for detoxification
of lewisite.10-33
It has been shown that urea peroxide reacts readily
with lewisite to give a nonvesicant product.14 How-
ever, a careful investigation failed to reveal a suitable
SECRET CHEMICAL SECTION
85
method of stabilizing urea peroxide at 60 C for field
use.-’4 Other peroxides were studied and it was found
that 10 g of a 1 1 mixture of sodium perlwwate mono-
hydrate and sodium dihydrogen phosphate mono-
hydrate, either in the form of a tablet or as a powder
dissolved in 50 ini of water, gives a solution equiva-
lent in active oxygen content to a 3 per cent hydro-
gen jK'roxide solution. The conclusion was reached
that HAL if quickly applied is somewhat more ef-
fective- as a preventive for lewisite bums than the
IMuborate-phosphate mixture; however, the latter is
non toxic.M _
T.2.2 Miphatic Arscnicals
Aliphatic arscnicals in wide variety have been pre-
pared for testing as candidate chemical warfare
agents. Major emphasis was placed on alkyldichlor-
arsincs, as it was thought for a while that memliers
of this series might show toxicity equal to that of
lewisite and at the same time exhibit greater chemical
inertness, particularly in reactions with water. How-
ever, it was finally established (hat n-amyl-, isoamyl-,
and /t-hexyldichlorarsine, for example, undergo the
same general reactions as lewisite and react at ap-
proximately (he same rate.36 i here is an opjxirent
difference in the behavior of the alkyldichlorarsines
as compared with lewisite in that the former do not
IiIterate a gas when treated with sodium hydroxide
and the alkylarsine oxides remain in solution longer
than does lewisite oxide.
The alki/Mirhlomrsincs have usually been prepared
by the use of one of the following three schemes;
1. The Meyer reaction.
RX + XajAsO., —> RAsOjNa, + XaX
RAsOjNa* + 211+ RAsOjII, + 2XV
RAsOjH, + SO, + 2HC1 —►
RAsCl, + II,S04+ H20
2. The Kharaseh lead alkyl process.
PbR, + 3AsClj ->- 3RAsCl, + PbCl. + RC1
3. From arsenic trichloride and tertiary arsines.
RjAs-F 2AsClj —3RAsCl,
The Meyer scheme is the one most frequently
used.16 -ij) Incomes less efficient with
the higher alkyl halides, such as heptyl bromide. The
Kharaseh process is particularly good for the prepa-
ration of ethyldichlorarsine in view of the availa-
bility of tetraethyllead.ls The suitability of (he prow-
ess for large-scale production has I wen demonstrated
by pilot plant operations in which the reaction went
readily and smoothly giving a 90 per cent yield.6*
For tho preparation of (Unlkylchhrarstnes, four
principal routes have been followed.
1. The Meyer reaction.
RAsCl, + INaOlJ —>
RAs(ONa)* + 2NaCl + 2114)
RAs(ONa), T R'Br - ■> RR1 As(),Na -)- NaBr
RR'AsO,Na + SO, -f lit I —>
RR'AsCl + XaHSOi
2. The Kharaseh lead alkyl process.
SRAsClj + R.Fb—*. 3RR'AsCl + R'CI + PbCI*
3. The cacodyl process.
4RCOOH -f AsjOj —* 2H.O + ICO, -f (R,.\s)gO
(H,As),0 + 211 Cl -> 2R,.\sCl + 11,0
I. From arsenic trichloride and tertiary arsines.
2R3As + AsClj —► 3R,AsCl
Here, as in the ease of the alkyldichlorarsines, (he
Meyer reaction scheme is the one most commonly
followed.1 5.2X.W.3M.M However, work on the cacodyl
process * o .m has resulted in a marked improvement
in this classical reaction. The improvement is in the
form of a continuous catalytic process wherein va-
pors of the acid and arsenic trioxidc arc passed over
an alkali salt on a pumice support. Although this
process was previously identified only with the pro-
duction of dimethylarsine derivatives, it has been
demonstrated that higher homologs may be pre-
pared in fair yield.17
In the preparation of tertiary arxinrs, three general
reaction schemes have been used:
1. Reaction of Grignard reagents with AsClj,
RAsCU, or RjAsCl.
3RMgCl + AsClj —> RjAs + 3MgCl*
21lMgCI + R'AsCl,—>R,R'As + 2MgCl*
RMgCl -f —> RR,As + MgUl,
2. Reaction of alkylmercuric chloride with arsenic
trichloride.
SRIIgCl + AsClj —> RjAs + 3IIgCl2
3. Disproportionation of RAsCl, or R,AsCl.
2RAsCI2 R,AsCl -f AsClj
2R,.VsC| R3As + RAsClj
RAsCl, + R,AsClRjAs + AsClj
It should be noted that all these methods are labo-
ratory procedures and (hat no large-scale prepara-
tion of an aliphatic tertiary arsine has been at-
tempted.1 6 M:,s
SECRET 86
ARSENIC AI.S
7,2.:$ Aromatic Arscnicals
The standard approach to an aromatic arsenical is
the Bart reaction l>etween an aryldiazonium halide
and sodium arsenite:
ArNsCI + NajAsOj —► Ar.\sO,Na. + NaCl + N,
ArAsO,Na. + SO. + 2HC1 —>
ArAsCl, + Xa,SO, + H.O
Many aromatic arscnicals desired in the toxico-
logical testing program have Ix-en prepared in this
manner.5 5*,m However, the only aromatic arscnicals
produced on any sizable scale during World War II
are diphenylchlorarsine (DA) and diphenylcyano-
arsine (DC). Considerable attention was devoted by
the British to process development studies of those
compounds.280 -90 307 They carried out laboratory and
large-scale tests on two processes for DA prepara-
tion:
1. The Bope-Turner process.
CJFAsCl. + H.0 —► C’ellaAsO + 2HC1
C’sHvVsCh + 3(‘eHiAsO —> 2(CcIU)1AsCl + As,!),
2. The double diazotizat ion process.
CJT.X.C1 + XasAsO, —>
r«H4AsO(ONa)j + N, -f XaC’l
C6H,As()(OXa). + XaHSOj —>
CsIl>As(ONa), + NaHSO,
16H;,As(OXa). T ( sHiX.t 1
(C6H &)j AsOOX a + X. + NaCl
(CsHs).As()OXa + SO. + HC1 —►
(CgllnhAsCl + XallSOi
A considerable improvement in the Pope-Turner
process was effected by the British workers,290 who
worked out the proper conditions for partial hy-
drolysis of phenyldichlorarsine to a stoichiometric
mixture of phenylarsinc oxide and phenyldiehlor-
arsine (3 1 mixed oil), which, when healed to 240-
250 C, was converted to DA in gooil yield. DA is
readily transformed to DC by reaction with 30 per
cent aqueous sodium cyanide at 35-40 C.290
7.2.1 Heterocyclic Arscnicals
From the standpoint of large-scale preparation
work, only one member of this group, adamsite
(DM), was considered important during World
War 11. However, representatives of several other
heterocyclic types were prepared for toxicity testing.
Adamsite is still prepared by the standard pro-
cedure worked out during World War I and involv-
ins tlie reaction of diphenylaminc with arsenic
trichloride:
0 '0 ~(XbO"“
-As
\
Cl
It has been shown 3 that a considerable part of tin1
arsenic trichloride called for in this equation may l>e
replaced by the less expensive arsenic trioxide with-
out a sacrifice in yield.
Furan arscnicals were studied both in this country
and in Great Britain.25101* They were prepared by
reacting a-ehloromerc; ifuran with arsenic trichlo-
ride to give trifurylarsine. From this tertiary arsine
the mono- and di-furyl-chlorarsines were made by
reaction with arsenic trichloride. Similarly thiophene
arscnicals were made from a-thienylmagnesium
bromide and arsenic trichloride ’ and pyridine arsen-
icals were obtained from 3-aminopyridinc by the
Bart reaction.5*
Other miscellaneous heterocyclic arscnicals pre-
pared for toxicity testing include 5, lO-dichloro-5, 10-
dihydroarsanthrene.4 307b i i k m dibenzarsinole chlo-
ride,5’3071’ and 10-chloro-9,10-dihydroarsacridine.58,
307ij k.m.n 'The preferred methods of preparation are
illustrated by the following equations.
1. 5,10-Dichloro-5,10-dihydroarsanthrene.
0 0-N^C1 /\ AsOA'a,
+ Naj r\sOj ► I I
-xo2 _ V-N0*
—AsOjHs
-X..C1
O OH
0—AsOjlli A-AstONa), /\-A»-/\
— +U -*U U
It.Or. Ah
O OH
H2OjAs
cHb - o^b
CljAs q
SECRET PHYSIOLOGIC-\L SECTION
87
2. Dilienzarsinole chloride.
— + .\:ij.\s(), >
o
/
XajO,As IlsOjAs
<0 or-A A
IljOaAs V/\ V
() Oil
o Oil Cl
3. !()-(’hloro-9,10-dihydroarsacridine.
/\ -CH2-/\
+ NajAsOa—►
v-x" V
OrO-Or*t)
AsOaXai AsO.H.
0—ch.-ZN ih*u
i \J mr ~
AsOjHi
COO
— o oh .. a
PHYSIOLOGICAL SECTION
7.3.1 Lewisite
When the United States became actively involved
in chemical warfare during World War I, high hopes
were held for a new agent, /3-chlorovinyldichlor-
arsine, which was prepared and suggested as a candi-
date. agent by (’apt. W. Lee Lewis in 1917. On the
basis of relatively meager laboratory data it was de-
cided to produce this agent, Lewisite (L), in quantity
and to use it in battle. A shipment was on its way to
Europe when the war ended in November 1918.
During 1918, and particularly during the latter
half of the year, the toxicological properties of L were
studied intensively in various laboratories of the
Chemical Warfare Sendee. The .lata obtained dur-
ing this period are well summarized 124 and will not
lx* discussed in detail in this report. However, two
reports'224 225 issued in 1919 are particularly interest-
ing in that they not only summarize the toxicological
data acquired during (he war period but also attempt
to assess the military value of L as a chemical agent.
Since the conclusions of 1919 offer a convenient start-
ing point from which to consider the later develop-
ments which took place in the interval between wars
and during World War II, these conclusions will be
briefly stated.
The effects of liquid L on the skin were studied in
detail on dogs and rabbits.224 It was felt that L was
definitely more damaging to the skin than II and that
the danger of systemic poisoning from I. was con-
siderably greater than with H. It was concluded (hat,
if man were as susceptible as dogs to systemic poison-
ing from L, the minimum lethal dose for man would
lx* 1.4 ml distributed over an area of 5 square inches
for an individual of average size.
No systematic study of the effect of liquid L cm
human skin was carried out. However, it was stated
that ;224
Laboratory workers who have been accidentally burned
with liquid 1. have- given strong evidence for the greater ef-
fectiveness of this substance in man than liquid II. The b
lesions develop with extreme rapidity, are painful and associ-
ated with definite constitutional symptoms. The lesion is not
confined to the skin, fait extends to the deeper tissues. In heal-
ing, dense scar tissue forms, the skin loses its flexibility and
contractures may develop. With liquid II skin burns in man,
pain is less or absent, there are no constitutional symptoms,
the amount of skin destruction is less, and healing occurs with-
out extensive scar formation, formation of contractures, or
permanent disability. .
lu view of the divergence of these views from those
currently accepted, it is well to bear in mind that
these were accidental burns and hence were probably
treated, that the accepted treatment at the time was
application of 5 per cent sodium hydroxide to the
lesion for a period of 80 minutes, and that sodium
hydroxide itself in that strength produces a very
destructive skin effect.
The effects of 1/ vapor on the skin were studied
with dogs, rabbits, and man, and are summarized in
Table 1 «
2. Dibenzursinole chloride.
Table 1. Approximate concentration to produce skin
lesions in 30-minnte exposure.
Rabbit Dog Man
1-ewisite (L) 0.025 mg 1 0.050 mg/I 0.200 mg/I
Mustard (11) 0.200 mg/I 0.050 mg/I 0.025 mg/I
The comparison indicated a lower sensitivity of
man toward L vapor than toward H. The degree of
SECRET 88
arsenic vls
protection afforded by ordinary wet and dry clothing
against I, and H vapor was also studied.
11 was concluded:224
An approximate concentration of .200 mg 1 (of L) is neces-
sary to produce skin lesions in man on exposure of one-half
hour. To lx- effective on parts of the luidv covered with cloth-
ing, il would lie necessary to raise this concentration from
three (3) to one hundred (100) limes, or approximately to a
concentration of .(UK) to 20.0 mg 1 . . . So far as the concentra-
tion required under field conditions to produce cutaneous
lesions in man, 11 should lie regarded as from eight (8) (on
unprotected skin) to a thousand (1000) times (a single layer
of wet wool) more effective than b.
The eye effects of L vapor were studied on rabbits
and dogs. As with skin effects, it was found that rab-
bits were more susceptible to I. vapor than were dogs,
and a comparison with H revealed that ibe relative
susceptibility of the sjteeios toward I, and II vapor
paralleled that Of the skin effects. The data are sum-
marized in Table 2."5 —
from tin* laboratory, tin- cxjieriinental field and the field of
war, our knowledge of the latter is confined entirely to data
front the lalwiratory, 1 he abrupt cessation of experimental
work at the American I niversity following the signing of the
Armistice in November 1018, prevented the carrying out of
field tests with I., preparations for which were already under
way.
In summary of the situation in 1018 it was
stated: 224
\\ e regard t he laboratory data as offering si rong support for
the probability that 1. will prove to have great military value.
Its actual value can only !«• definitely determined, however, by
further ex])erimenl:d data, cs|n-cially those obtainable by field
tests. It would furthermore seem clear that the usefulness of I,
in war would differ quite widely from that of II. kittle effect
should lie expected from the vapor when used against troops
supplied with an efficient mask equipment, because of the low
skin vajair toxicity and the resistance of clothing to penetra-
tion of the vapor. This is the condition, on the other hand, m
which II has liecn found most effective. The usefulness of |,
would lie eon fined to the effect of the substance reaching troops
in the liquid phase (splash or mist) by their coming in contact
with contaminated material, the influence of the hydrolytic
products in contaminating the ground and objects, and the
respiratory effects ami possibly the eye effects of the vapor in
the ease of troops unprotected by mask equipment. In those
respeets L offers many advantages, so far as can be concluded
from the data at hand, over H. Wc feel that L offers sufficient
promise to warrant the most careful further consideration.
Data which are not at present obtainable and which are most,
desirable in ibis connection are as follows;
1. The keeping qualities in steel.
2. The ability of the substance to withstand detonation.
3. The vajsir concentration which it is possible to secure
and maintain in field tests.
4. The vapor concentration necessary to produce eye le-
sions in tests on man.
o. The relative importance of burns by liquid II and vapor
of H in actual warfare.
In conclusion we wish to repeat: We believe that L w ill not
replace II in warfare, and that in any plans for military oper-
ations the production and utilization of II should remain one
of the most important propositions. While very promising, the
military value of L remains to lie established.
Fit the interval between 1919 and 1910 relatively
little research on the toxicology of 1, was carried out
by (lie Chemical Warfare Service, with (he exception
of a dt'tailed study which was published in 1923.1,4
'l itis report has been critically reviewed 124 and will
not l>e discussed in detail, although a few of the re-
sults will be mentioned later in the present report.
It was concluded 1,4 that L is superior to II in that
it gave deeper and more severe bums as well as sys-
temic disturbances leading to death, but the diffi-
culty of setting up effective vapor concentrations
was recognized.
Following (he publication in the open litera-
T\bi.e 2. Approximate concentration' to pro-
duce eye lesions in 30-mimile exposure,
Hahliit Dog Man
D'u-isiic (L) 0.001 me 1 0.020 mg 1
Mustard (II) 0.030jn*/l 0.020 nig/1 0.001 mg 1
The statement was made;224 “If we may lx- allowed
to infer or judge of (he susceptibility iu man without
having an actual determination, the conclusion
would l>e that the eye of man is less susceptible to L
than to II, but such a conclusion can never convex*
the conviction as one based on actual determina-
tion.” No experiments involving the effects of liquid
L on the eye were reported.
I lie respiratory effects of I, vapor were studied on
dogs and compared with the effects of II vapor, it
being found that the dog was approximately twice
as susceptible to L as to H. It was pointed out '225
that the concentration necessary to produce death in
man on respiratory exposure is not known in the ease
of either II or L. hut that in the light of our present
knowledge we can only conclude that on respiratory
exposure, I. is to In- regarded as approximately twice
as effective as II as determined by the concentration
necessary to kill. I his conclusion, as applied to man,
must be made with reservation due to deficiency
of data.”
With resjtecl to the relative military value of L
and II. il was stated;
In attempting a comparison of the relative military value
of the substances II and b, we meet with the fact that while
with the former sulislance we have a very large exjieriencp
SKCRFT physiomm;i<:\l siir.no\
89
ture 335 337 of information that L had been seriously
considered by the Americans as a war gas, the agent
was studied in the laboratories of other nations. The
published German reaction was unfavorable. The
compound was tested in Germany in 1910 338 and
the conclusion reached that it was not reliable as a
war gas because its toxic effects were less lasting than
those of mustard and the irritant effects were so
marked that men would lie warned in time of its
presence. The opinion was offered 340 that the Ameri-
cans were spared a great disappointment by living
unable to use L in World War I. A series of experi-
ments was carried out 339 in which the effects of rela-
tively large doses (one to two drops from an ordinary
eye dropper) of 11 and L on human skin were com-
pared. These exjieriments,- published in 1932, led to
the conclusion that L was inferior to II in producing
skin injury and that its potentialities as a war gas
have l>een greatly overrated. In reference to the cal-
culation 284 331 that 1.4 ml of L applied to the skin of
a man should lie the approximate minimum lethal
dose, it was asserted that this amount was applied
repeatedly to the skin of human livings without giv-
ing evidence of systemic intoxication.339 The Japa-
nese used a 1 1 mixture of II and L against the Chi-
nese at Ichang in 1938, but subsequent information
obtained by (he interrogation of Japanese officers
revealed that the L was added mainly to lower the
freezing point of the H.
The value of L as a chemical warfare agent still re-
mained to be established in 1941. The published Ger-
man opinions were looked upon with distrust, and.
as in World W ar I, the United States undertook the
quantity production of L. The discrepancies in the
literature as to the toxicological effects of L had to lie
resolved and intensive research was carried out both
in the United States and Great Britain.
Properties of Lewisite
Plant run L is usually dark brown in color and pos-
sesses an odor reminiscent of geraniums. Both the
color and odor are due to impurities, which can be
removed if the extra effort involved is considered
worth while. Vis- and leans- isomers exist which
have almost identical toxicides.171 L freezes at
—18.2 C to 0.1C, depending on the purity and
isomers present. The density of liquid L is 1.886 at
20 C, whereas the density of the vapor is 7.1 com-
pared to air. The volatility of L is greater than that
of II and increases somewhat less rapidly than that
of II with increasing temperature. The following data
for I. are calculated from the vapor pressures;;o:
comparative data for 11 are also given.9"
Teinjicratiire
Volatility (mg 1)
I, II
C
I,
II
0
1.06
10
2.23
15
3.2!)
0.41
8.0
20
4.48
0.65
6.0
25
6.14
0.06
6.4
30
8.62
1.30
5.1
35
11.32-
40
15.75
2.82
5.6
L is fairly stable on storage in glass or steel hut is
degraded to a considerable extent on detonation.1,4
The reaction of 1. with UAL and certain related
dithiols30,f 3m,r 3U to form nontoxic complexes has
assumed great importance in the treatment of L
lesions and of arsenical poisoning from L or other
sources. —
The chemical properties which most sharply limit
the usefulness of L as a chemical warfare agent are
the ease with which it reacts with (1) water and
(2) alkalies. In contact with water or moist surfaces,
lewisite is readily hydrolyzed to the oxide which,
although mildly vesicant, is nonvolatile and insolu-
ble in water. Since L “precipitates out” in contact
with moist surfaces it is impossible to maintain high
vapor concentrations in humid atmospheres. Alkalies
decompose L rapidly at ordinary temperatures, and
even alkaline soil315 rapidly destroys the liquid and
imposes a further limitation on its use as a ground
contaminant. 'The maximum efficiency of L is only
attained, therefore, under conditions of low temper-
ature or low humidity, both of which minimize hy-
drolysis, and on dry nonalkalinc terrain.
Physiological Action
Lewisite Vapor. The qualitative effects of L on the
eyes, skin, and respiratory tract have been described
in the open literature21433" and have also been re-
cently summarized.3'® They may be very briefly re-
stated as follows:
1. Eyes. L vapor is extremely irritating to the
eyes, causing pain, lacrunation, and blepharospasm.
The lacrimation and blepharospasm protect in a
large degree from further exposure to the vapor but
if the Cl is sufficiently high the irritation and pain
persist and after a few hours are followed by edema
of the eyelids and conjunctivitis. Permanent damage
is, however, apt to result only from very high con-
centrations difficult to achieve in the field.
Liquid L is capable of causing severe damage to the
SECRET 90
eyes. Pain, lacrimation, and blepharospasm appear
immediately, ami are followed by edema of tin* lids,
iritis, and conjunctivitis. In severe contamination,
ulceration, necrosis, and secondary infection may
lead to blindness or to permanent impairment of
vision.
2. Respiratory tract. L vapor is irritating to the
nasal passages and produces a burning sensation
followed by profuse nasal secretion and violent sneez-
ing. On prolonged exposure coughing results and
large quantities of frothy mucus may Ik* brought up.
The effects of L vapor are so prompt and striking
that men usually mask before enough of tin* com-
pound is inhaled to produce serious injury. However,
in cxjx*rimental animals exposed to vapor in a gas
chamber, injury to the respiratory tract is essentially
similar to that produced by mustard. Edema of the
lung is often more marked and is frequently accom-
panied by pleural fluid.-’18
3. Skin. L vapor usually produces no more than
erythema of the skin, although if the skin is hot and
dry and the vapor concent ration is high, small, shal-
low. turbid blisters may develop and may coalesce to
form large vesicles. Such conditions would seldom
Ik* ivalized in the field.
Eiqnid Lon the skin produces an immediate sting-
ing sensation which fortunately warns of its presence.
If L is allowed to remain on the skin for 5 minutes,
the site of application assumes a cooked appearance,
somewhat resembling that from an acid hum. Ery-
thema develops in a short time around the site of
contamination and is followed by vesication of the
cntiie erythematous area. L can penetrate the skin,
subcutaneous tissue, and muscle, causing extreme
edema and neemsis.
The fluid coinained in vesicles produced by I, tends
to be more opaque than that found in mustard blis-
ters, although it is frequently impossible to distin-
guish L vesicles from mustard vesicles by their
appearance.
The fluid from an I, blister contains 0.8 to 1.3 y of
arsenic per cubic centimeter, equivalent to 2.5 to
4.0 y of original L.'*7
4. Systemic effects. The absorption of a sufficient
amount of L through the skin of dogs may lead to
death within 24 hours and usually within 10 hours.
Table 3. Toxicity of T.
jierioti 10 days, except a
■ vapor. (All figures are lACl).,,, in rng miriTT,
s noted.)
exposure time = 10 min, observation
Total
Inhalation only
Hotly only
Species
exposure
exposure
exposure
Mouse
0.0-1.4 (nom.)71
1.4-1.5 (nom.)75
1.2-1.9 (nom.)77
Mouse
2.8 (nom.)*5
1.6 (nom.)85'
0,3 (nom.)*1'
Mouse
1.5 (anal.)43
1.5 (anal.)85*’
7.0 (nom.)14
Mouse
2.5 2.8 (nom.)17'
Mouse
0.5 (anal.)*6*1*
Rat
1.5 (anal. )**-f _
20.0 (nom.)14
Rat
0.58 (anal.)***-J
,
Guinea piR
1.0 (nnal.)“*‘*
20.0 to 25.0 (nom.) 14-J
Guinea pig
0.47 (anal.)*4’1
Rabbit
1.2 (anal. PHI
15.0 (nom.)14
Rabbit
1.5 (anal.)*4"’*
Goal
1.25 (anal.)*48 **
Cat
—
30.0 (nom.)14-ft
Dor
1,4 (nom.),M §§
30.0 (nom.p- n
40 0 (nom.)*
♦
11- to 14-min exposure. 21-day observation )ieriud.
1
9- to 25-min exjxieurc. 21 -day observation jieriod.
x
tiO- to 180-miii.exposure, 21-day <
observation period.
§
lO- to 10-hub ex|MK-ure. •
1!
7 5- to 13-tnin exposure. 21-day
observation |*riod.
r
60- to 310-min exposure. 21-day ohm* r vat ion period.
♦♦
1 Ott- to 255-min ex|H>turc. 21-day observation period.
'
n
30- to 45-min exposure.
XX
30- to 00-min exposure.
if (<'l = 1.32 for 7 J-mm exposure and I II for 15-nin 9 »-h >ur obiprvatiuii period.
The report .states th it concentrations were determined
both as nominal and analytical but only one set is given ami it
» not charac terized.)
““
-Vufr. N’om. » nominal concent rut ion; i.e., concentration calculat'd from the amount of L volatilized. the flow rate, ami the duration «*f flow.
Amount volatilized (mg)
Nominal coiwentration
—
Flow rale (l/min) X lime (min)
Anal. = analytical concentration: i.c., concentration determined by sampling and chemical
analysis of the atmosphere.
SECRET PJI\ SIOLOGICAL SECTION
91
A few hours after application, the dogs show evidence
of severe intoxication and appear almost moribund.
Death apparently occurs from an intoxication which
interferes with certain vital processes without pro-
ducing sufficient anatomical lesions for complete
characterization of the immediate cause of death. A
frequent accompaniment of systemic intoxication is
a change in capillary permeability which permits loss
of sufficient fluid from the blood to result in homo-
concentration and profound shock. The blood vol-
ume of dogs was observed 224 to fall as low as 3.9 per
cent of laxly weight in burned animals (normal =
9.7 per cent ).
Nonfatal cases may develop a hemolytic anemia,
focal necrosis of the liver, and some injury to the in-
testinal mucosa. —
Toxicity. There is no disagreement over the fact
that L is a highly toxic compound and that it can
produce the physiological effects which have been
described. In order to evaluate the usefulness of I. as
a chemical warfare agent, however, several things
must Lie known. These arc:
1. What dosages of I. are required to kill men or
at least to make them casualties?
2. Can these dosages Lie attained in the field with
a reasonable expenditure of munitions?
3. How easily can the soldier protect himself
against the effects of b?
4. Are the results obtainable through the use of L
in the field likely to be better or worse than those
obtainable with the standard vesicant agent, II?
Toxicity Data
The answer to (1) can only l>e approached experi-
mentally through studies on animals. The toxicity of
I, vapor toward animals of different species is shown
in Table 3. The L{Ct)„0 of L vapor for man is un-
known, but may be estimated (from the data of
Table 3) to lie of the order of 1.2-1.5 mg min/l (ana-
lytical). The L{Ct)bo for body exposure only has been
estimated to be of the order of 100,45 on the basis of
animal experiments and with the assumption that
the absorption of b through the skin is a function of
the ratio of surface exposed to laxly volume.
The toxicity of liquid b applied via the skin for
animals of different species is shown in Table 4. On
the assumption that man would lie as susceptible as
the dog, it was calculated in 1919 224 that the LD-„o
for a 70-kg man would be of the order of 1.4 ml of b
applied over an area of 5 square inches of skin. It is
stated, however, that doses of 1.4 ml can lx* applied
Table 4.
Toxicity of lewisite by
skin application.
Animal
LI)infing kgt
Reference
Mouse
15
87 (Cited bv Smith)
Hat
24
300f
Hat
15
318
Rat
24
240
Rat
20
318
Ralihit
5
318
Rabbit
ti
24 it
Rabbit
«
133
Guinea pig
12
24!t
Dog
38
224
Dog
C!». 70
295a
Goat
24
241
Quit
10
217
repeatedly to men without eliciting any clear-cut
symptoms of arsenical poisoning.”** The I. Dm for
man is probably much greater2*™ than the 40 mg kg
sometimes assumed, A case is reported in which a
worker at Pine Bluff Arsenal suffered accidental
lewisite burns over 20 per cent of his body surface
(mostly on the legs). Tie showed an anemia 10 to 15
days after the burn, but no clear-cut signs of systemic
arsenical poisoning. Tt appears, therefore, that man is
not nearly so susceptible to systemic arsenical poison-
ing from skin contamination with L as was originally
believed.
The toxic dose of L when administered pa rente rally
is much lower than that required by skin absorption.
For example, the LD„n for rabbits is stated in one
British report 227 to be 2 mg kg by either intravenous
or subcutaneous injection, and in another 249 to be
0.5 mg kg by intravenous injection. The intravenous
LD:to for dogs was found to be 2 mg kg as compared
to 38 mg kg by skin absorption.2-' Two mg kg,
injected intraperitoneally, has been given as the
minimum fatal dose for guinea pigs.295"
It is difficult to see, however, how the enhanced
toxicity by parenteral administration can be utilized
in warfare.
Casually production by L may result from the
action of the vapor on the respiratory t ract, or of the
vapor or liquid on the eyes and skin. Assuming that
men will be masked, the probabilities of casualty
production from the inhalation of vapor are small.
Relative to (he eyes, it has been shown that for L to
produce moderate corneal damage in dogs a vapor
Cl of 2.8 mg min 1 (nominal) is required; whereas a
destructive lesion is produced by a Ct of 5.5 (nom-
inal).S2f Analytical concentrations in the above ex-
periments were approximately 50 per cent of the
nominal so that an analytical Ct of the same order as
SKCRKT 92
\RSEMC\LS
the fACt) by inhalation is required to produce mod-
erate eye damage. Since the immediate response of
the eye to b vapor is lacrimation and blepharo-
spasm, both of which protect against further expo-
sure, serious ey e casualties from 1, vapor are not to lx*
expected in conscious men.
biquid b in the eyes is capable of producing de-
structive lesions. It has been estimated176 that a
drop 170 n in diameter in the eye of a man would
make him a casualty for over a week unless immedi-
ately treated. A 0.1-nig drop in the rabbit ey’e caused
IKM'foration of the cornea in approximately 75 per
cent of the cases and permanent disability (as judged
by the1 jx-rsistence of corneal haze) in nearly all
cases.’-’14 In the rabbit eye a 0.1-mg dose of liquid b
produces a maximal lesion. With doses greater than
0-1 mg the severity of the ocular reaction did not
appreciably increase. It has been stated that a dose
of 0.01 to 0.02 mg of liquid b will produce permanent
ocular damage (in rabbits) approximately equal to
that produced by 0.1 to 0,2 mg of liquid H. With
0.05 mg of b most of the eyes are completely de-
stroyed. whereas even 1.4 mg of H does not produce
an equally' severe lesion. Mild, self-limiting injuries
of comparable severity are produced by 0.005 mg
of I, and 0.02 mg of H. It is thus apparent (hat the
severity of the b lesion increases steeply with in-
creasing dosage and rapidly reaches a maximal lesion,
whereas the curve relating severity of the lesion to
dosage of H is much more flat and very large doses
are required to destroy an ey e completely'.
I he threshold Ct for vesication of bare human
skin (forearm) has been estimated as 1,0 mg min 1
(analytical) for a temperature of 55 F and relative
humidity = 70 per cent.24* A Ct of 1.8 at T = 90 F
and relative humidity = 49 per cent caused vesica-
tion of the bare hand in 50 per cent of the men ex-
posed.30* A Ct of 1.5 (analytical) caused vesication of
the neck of six men exposed in the field at T = 60 F
and relative humidity = 41 per cent, but no effect
was obtained on skin covered by ordinary battle
dress.24* A Ct of 1.5 (analytical) at T - 90 F and
relative humidity = 65 per cent caused vesication
on the skin (forearm) of three men (3 3), whereas
a Ct of 1.2 produced vesication in none of three
men (0 3) under the same conditions of temperature
and humidity.21*
biquid b on the bare skin is a very potent vesicant,
the median threshold blistering dose for man Ix'ing
14 ns as compared with 32 ng for II.7* Contrary' to
the opinions held in this country prior to World
War II, recent work has tended to establish the
view*** that in relatively large amounts I, does not
produce as severe skin damage in man as does H.
Although with doses up to about 1 mg of liquid l„
produces skin lesions in men not perceptibly differ-
ent from those resulting from the same amount of
liquid H, the response to larger doses of the two
agents is different. For 2-mg dosages of b and of H,
the lesions produced by I, are less severe and heal
in 2J4 to 4 weeks compared to the 5 to 9 weeks re-
quired for healing of the mustard lesions. One investi-
gation,*** using much larger doses, placed two large
drops (from an ordinary eyedropper) of b on one
forearm and of H on the opposite forearm of a man.
He reported healing of the L lesions in 2(1 days and
of the II lesions in 63 days and stated that these re-
sults were typical of other experiments. It has l>een
pointed out that in rabbits tin- damage produced by
2 mg of liquid L is more severe and slower to heal
t han that produced by 2 mg of liquid 11. The reaction
of rabbit skin toward L is, therefore, not character-
istic of the reaction of human skin. In an investiga-
tion conducted at Port on -,&s it was concluded that b
burns heal more quickly than II burns, are less prone
to infection, and cause less pain during healing. The
question of the comparative severity of lesions pro-
duced by H and b on human skin has recently l»een
reinvestigated,”0 with the result that b Irsrnirs were-
found to be less severe and to heal more quickly than
those caused by the same amount of II (either by
weight or by volume, the dose being 1.0 mg or
0.5 microliters).
It may be noted parenthetically that in 1941 a
statement appeared in United States official chemical
warfare manuals to the effect that the fluid from
lewisite bullae was itself vesicant. However, experi-
ments have been reported *** leading to the conclu-
sion that b blister fluid was neither vesicant nor
irritating and an American investigation in 1943 137
confirmed this conclusion, with the result that state-
ments regarding the vesicancy of b blister fluid have
been withdrawn from recent editions of United
States official manuals.
The toxicity of I, for man is summarized in Table 5.
T he dosages required for b to produce casualties
in men or to kill them appear to have been as well
established as would be possible through the use of
experimental animals in lethal experiments and hu-
man observers in marginal experiments.
As was aptly stated in 1919 255 (he value of b as a
military’ agent depends in large degree on whether
SECRET PHYSIOLOGIC A I, S liCTION
93
Tablk 5. Toxicity of lewisite for
man.
VajKrr
approx. L{Ct)ii,
(analytical)
mg min 1
Liquid
close
mti5"
Death (bv inhalation)
1.2-1.5 (est.)
Death e completely destroyed. The British23*
attempted to assess the danger of systemic intoxica-
tion from liquid I. released in bomb explosions. On
the assumption that the lethal dose for man would
be 1.9 g (a dose which is probably not fatal) it was
concluded that the risk of receiving serious injury
from a bomb charged with L would be no greater
than from a bomb of the same size charged with high
explosive. When unthickened b is released from an
airplane spray tank, the droplets formed are less than
1 mm in diameter.'** Since it has been reported *"*-230
(hat L droplets of less than 1 mm in diameter evap-
orate completely while falling through 2,000 feed, it
is apparent that the employment of unthickened L
from medium altitudes (>2,000 feet) as airplane
spray would be useless. I. may be thickened with
methyl methacrylate and similar materials. The use
of thickened L as airplane spray results in larger
drops (55 per cent of drops >0.5 mg as compared
with 8 per-ccnt of drops >0.5 mg for unthickened
L).1*3 However, when droplets of thickened L strike
a surface, they tend to harden. This effect may be
due to the formation of a skin of L-oxide on the sur-
face of the drop.1*3
A comparison of the casualty-producing effect of
thickened and unthickened L when used as an air-
plane spray from low altitude (100 feet) revealed that
thickened L was less effective in producing casualties
in goats than unthickened L, and that the eye dam-
age caused by the unthickened L was more severe
than that caused by thickened L.163
The tactical value of producing L blisters on hu-
man skin is thrown into very serious doubt by recent
Canadian experiments 3,3 in which observers clad in
battle dress and shirts over long-limbed underwear
and wearing respirators and steel helmets were ex-
posed to airplane spray of L to which had been added
0.55 per cent of thickener. The temperature was
75 F with relative humidity = 89 per cent, and the
contamination density was 0.7 to 5.4 g in2. T he
drops varied between 1.3 and 5.0 mm in diameter.
Of 30 men hit by the spray, 20 developed lesions
which in 7 cases were numerous and prominent but
in other cases were trivial. It was noted that the in-
dividual lesions produced were discrete and circum-
scribed in contrast to the diffuseness of the typical
lesion produced by H spray. After 9 days of compara-
tively strenuous exercise, none of the observers was
the neci -ary dosages can lie set up in the field. Suf-
ficient field exjieriments have now been carried out
to indicate that the requisite dosages are probably
not attainable with any reasonable expenditure of
munitions.
Fiki.d Tkst Data
The concentration of vapor obtained from pouring
50-75 g of I. |s'r square yard on the ground is low
and Cl values obtained are usually not over 4.0 mg
min I.'24 240 The vapor concentration obtained di-
rectly over the contaminated area fell steeply during
the first 30 minutes of the experiments and thereafter
was not dangerous.
In experiments conducted at Edgewood Arsenal
four M70 bombs charged b (total 360 pounds) were
fired statically. Twenty-five yards downwind from
the burst the initial concentration was 0.060 mg 1
but fell to 0.013 mg 1 in 10 minutes. The Ct for 15
minutes was 0.395 mg min I.216 In a further test at
Edgewood an airplane sprayed 610 pounds of un-
thickened L from an altitude of 75 feet over an area
of 76,250 square yards.1*3 Significant vapor concen-
trations directly over the contaminated area were
recorded only for the first 10 minutes and the total
Cl recorded was of the order of 3.
It is apparent from (he above examples that dan-
gerous concentrations of L vapor are difficult to at-
tain in the field, 'Hie reason for this is apparently
the rapid hydrolysis of the vapor and liquid in con-
tact with a moist environment, with possibly the
destruction of some I, by alkaline soil, together with
the fact that the agent may he partially destroyed
by detonation when loaded in munitions. In ex-
tremely hot and dry climates more effective vapor
concentrations may be anticipated.
The effects of liquid L on bare skin might lx*
achieved through ground contamination, bursting
munitions, or airplane spray. However, L is so un-
stable on contact with moisture (hat under ordinary
conditions of humidity it is rapidly hydrolyzed on
the surface of soil or foliage, leaving behind a residue
of L-oxide. The L-oxide. while weakly vesicant, is
SECRET 94
VttSEMCALS
in such condition that he could not carry out military
duties, and in no case had secondary infection de-
veloped. It was concluded that the casually-produc-
ing propensities of II spray are definitely greater
than those of L spray.
Protection* against Lewisite
Lewisite Vapor. The median detectable concen-
tration of L vapor by odor is stated to be 0.014 to
0.023 mg 1. However, the irritating effect of the gas
on the eyes and respiratoiy passages is noticeable at
far lower concentrations, variously estimated as
0.008 mg 1339 and as 0,006 mg I.124 On the basis of
these figures, a concentration of 0.006 mg 1 should
certainly warn troops of the presence of gas and
should lead to masking or to withdrawal from the
toxic atmosphere. The service respirator gives en-
tirely adequate protection to the eyes and respiratory
tract against the effects of L va|K>r. Even in the ab-
sence of the respirator, serious eye effects from L
vapor are unlikely to occur in conscious men since
the immediate response of the eye to L vapor is lacri-
mal ion and blepharospasm, both of which protect
against further exposure.
Ordinary* clothing affords considerable protection
against L vapor. It has been estimated 224 that a
single layer of dry cloth would protect against ap-
proximately three times the concentration of L that
would produce a reaction on bare skin. The British 258
estimated that a Cl of 3.0 4.0 mg min I would be
required to produce an effect under a single layer of
dry serge. In another report, it was found 257 that the
penetration of cloth by L vapor decreases with in-
creasing humidity, and it was suggested that the
reason lies in reaction of L with moisture on the
fibers of the cloth. Complete protection against L
vajKir was afforded by ordinary dungaree shirt ma-
terial, S-330 ointment, and CC-2 impregnated cloth
up to at least Cl 3.3 (analytical) under exposure con-
ditions of 90 F and 65 per cent relative humidity
with 4 hours wear of the clothing after exposure.219
Wet clothing is much more effective in protecting
against L vapor than dry clothing. It has l>een esti-
mated 224 that 100 times the concentration of L that
would produce an effect on bare skin would be re-
quired to | tenet rate a single layer of wet cloth. In
fact the British state that 1. vapor will not burn
through wet clothing.
Liqud Lewisite
Liquid lewisite in the eyes is capable of causing se-
vere damage. However, complete protection against
liquid I- is afforded to (ho eyes hy wearing (lie respi-
rator or (ho eye shield or even by closing the eyes.
Liquid L will penetrate ordinary dry clothing, a
drop of 2.5 mg (1.5 min in diameter) generally caus-
ing vesication through dry service clothing in temper-
ate climates.2*1 Under tropical conditions a 0.4-mg
drop (0.77 mm in diameter) may produce vesication
through light dry clothing.334
Wet clothing protects against liquid L by forming
(he insoluble and nonvolatile L-oxide before the
agent can penetrate to the skin.314 CC-2 impregnated
clothing offers more protection against liquid f. than
does unimpregnated dry clothing, although 5.7 mg
of L produced vesication through a single layer of
CC-2 impregnated cloth,179 indicating that the pro-
tection afforded against L is less than that against II.
( '<*\tI’AH 1SO.V WITH Ml-STAUU
The toxicity of L vapor and II vapor by inhalation
are of the same order of magnitude. However, to pro-
duce systemic effects through the skin, eye damages,
or skin vesica! ion, significantly higher Cl's are re-
quired for I. than for H. Because of the rapid de-
struction of I. liquid and vapor in contact with
moisture or with an alkaline environment the requi-
site Cl's for L would be extremely difficult to attain
in the field. Further, L vapor, unlike II vapor, is not
insidious but gives adequate warning of its presence
by irritation of the eyes and respiratory passages.
Liquid L is more vesicant, than liquid H but the
burns from I. do not. incapacitate men to the same
extent as do burns from H,313 and the L burns heal
more rapidly and are less painful than those from If.
Liquid L on the skin or in the eyes produces an im-
mediate stinging sensation which warns of its pres-
ence. whereas mustard is nonirritating at (he time of
application.
Mustard penetrates ordinary clothing much more
readily than does L, and, since H is more stable than
L, is a Ix-tter choice both for terrain contamination
and vapor return.
Mixtures of H and L have been suggested but have
no advantage over II used alone except with respect
to lower freezing point.
Thkrapv
In 1941, the discovery of a powerful therapeutic
agent against L and other arsenicals was an-
nounced.3"01 This substance. 2,3-dimercaptopropanol-
1, variously known by the code letters HAL and
DTK, will not only destroy arsenicals on contact,
but is capable of minimizing the damage from liquid
SECRET PHYSIOLOGICAL SECTION
arsenicals in the eyes if applied from I to 10 minutes
after exposure, and from liquid arsenicals on the skin
if applied up to 1 hour after contamination.
A discussion of BAL is lieyond the scope of this
report except to say that an ointment containing
BAL was available for issue to United States soldiers.
This ointment was suitable for application to the
skin or eyes and placed in the hands of the soldier a
method of self-help for minimizing the effects of con-
tamination from liquid arsenical agents. Prepara-
tions of BAL were available to physicians for paren-
teral administration and are effective in combatting
systemic intoxication from arsenicals.
Summary
By the end of the World War II, the toxicology of
Lhad been worked out to the point where the dosages
required to produce casualties or death in human liv-
ings were known with a degree of approximation that
is probably sufficient for military purposes.
Field tests, however, showed little promise of at-
taining the requisite dosages of L vapor with any
reasonable expenditure of munitions. The use of
liquid L for gross contamination of personnel seems
feasible only when the agent is dispersed as low-
altitude airplane spray, and the effects produced on
contaminated personnel are so inferior to those pro-
duced by mustard as to create strong prejudice
against the use of L.
Since the powerful antiarsenical agent, BAL. avail-
able to Britain and the Uniter! States in World
War II. will be available to all in the future, there
seems to lie little likelihood that there will ever lie
any incentive for the use of L as a chemical warfare
agent.
7.3.2 Chlorarsine Derivatives Other
Than Lewisite
Lethal Agk.nts
In the Spring of 1918, ethyldichlorarsine (ED) was
us»>d by the Germans as a skin and lung irritant suit-
able for gassing operations to be followed by infantry
assaults.331 *28 There is no mention in Allied official
records of casualties attributed directly to ED, but
the Germans held the compound in high regard. 1 he
United States Chemical Warfare Service investigated
methyldichlorarsinc (MD) during the latter half of
1918 but the compound was not used in battle.
In 1989, the results of a preliminary investigation
by the Chemical Warfare Service 12‘ revealed a lack
of sufficient data for making a definite decision as to
tlu* value of ED as a military agent, hut stated that
“the present available data indicate sufficient poten-
tial value to warrant further study and develop-
ment.” Accordingly, the National Defense Research
Committee [NI >HC] was asked to screen the arseni-
cals for toxicity and stability in order to determine
whether any members of the group were sufficiently
promising to warrant further study or development
as chemical warfare agents. A number of chlorarsine
derivatives were prepared and were studied for toxic-
ity at the University of Chicago Toxicity Labora-
tory [UCTL].
Physiological .1 rtion. The toxic chlorarsine deriva-
tives produce effects which are qualitatively similar
to those produced by I- (q.v.) hut which differ in
degree. Thus, they are all irritant to the respiratory
tract and produce lung injury on sufficient exposure.
The vapors are irritating to the eyes and the liquids
may produce serious eye lesions. The absorption of
either vapor or liquid through the skin in adequate
dosage may lead to systemic intoxication or death.
Local skin damage leading to vesication in man is
usually produced by sufficient exposure to the vapor
or by contact with the liquid.
Vapor Toxicity. The chlorarsines originally
screened for vapor toxicity at the UCTL27 are listed
in Table (>, which shows the results of tests against
Tabik fi. Toxicity of vapor
figures for L(Ct)-,» are in mg
of clilornrsincs for mice. All
min/1 (nominal).
Compound
L(CI) - (Mouse)
lewisite, isomer I
L(Cl)iu = 2.8
lewisite, isomer IT
UCl),u = 2.8
Plant run lewisite, isomer I
Phenyldichlorarsine
lACtU = 3.7
HCI)90 ~ 13.
d-Methoxvcth vldichlorarsine
unstable
5-Ethoxyethyldichlorarsine
unstable
0-C’hloromethoxypropyldichlor-
arsine
(No deaths at Cl = 8.7)
Allyl phenylchlorarsine
Phen y 1 ((3-chh troviny 1 Jclilor-
(No deaths at Cl = 24.44)
arsine
UP On. ~ 1.
Tsoamyldichlorarsine
UCtU ~ 2.
.w-Hul vldichlorarsine
urt),u ~ 12.
6/s(CliloroinclliyI)ehlorarsine
UCtU - 4.5
C 'liloromet hyldichlorarsine
(No deaths at Cl = 43.5)
4-Fentenvldiehlorarsine
L{Cl)u -3.7
A m vldichlorarsine
L(Ct),„ = 2,5
Hut vldichlorarsine
/>(COio ~ 3.5
El hvldichlorarsinc
UCtho - 3.5
0-Kurvldiehlorarsine
(No deaths at Cl = 2,3)
1 leptyldiclilorarsine
UCtU — 13.1
d-Mcthylhutyldichlorarsine
unstable
1 lexyldichlorarsine
UCtho - 3.
Dimethylehlorarsine
L{Cl)lu — 10.
SECRET VKSEMCALS
mice by total exposure for 10 minutes. On the basis
of the information listed in Table fi and information
from the Chemical Warfare Service on ED,129 butyl-
dichlorarsine,1*0 amyldichlorarsine.157 and isoamyl-
dichlorarsine,147 a more detailed investigation was
made of the toxicities toward mice (by total expo-
sure) of the vapors of the alkyldichlorarsinea from
methyl- through hexyl-.49 In order to avoid errors
known to result from different degrees of humidifica-
tion of (he animal’s fur, the mice were exposed for
1 hour to a relative humidity of 20—HO per cent before
exposure to the toxic arsenical. The dichlorarsines
were vaporized with dry nitrogen at 25/30C and
were passed through the 4-1 glass chamber at 11.2
1pm. The relative humidity of (he gases in the cham-
ber did not exceed S |H*r cent.
The vapor toxicities of the alkyldichlorarsines are
given in Table 7, together with the toxicity of phenyl-
dichlorarsine (I’D) and of L for purposes of com-
parison.
The toxicity of several diehlorarsines when applied
to the skin (shaved) of mice is shown in Table 8.
Table 8. Pcrcut
aneons
toxicity of arscnicals
for mice.2*
Compound
Dost*
mjt
No. of
mice
Percent
mortality
10-day
period
Comparison
with
lewisite
I, (plant run)
0.1
10
0
0.3
10
50
0.5
10
UN)
KI)
0.1
4
0
0.5
1
25
<5 L
1.0
4
25
X-Putvldichlor-
0.5
10
10
arsine
1.0
10
30
<5 1,
0-Methylbutvl-
0.5
4
25
dichlorarsine
1.0
4
100
- :■ i.
X-Amyldichlor-
0.1
7
It
arsine
0 3
10
80
= i.
0.5
10
100
Hexvldichlor-
0.1
4
0
arsine
0,5
4
50
1.0
4
50
_ ~i.
Hcplvldichlor-
0.1
4
0
arsine
0.5
4
0
- i »*
1.0
4
50
PD
0.1
10
20
0.3
10
30
= i.
0,5
10
100
Table 7. Toxicity o
if vapor of dielilorarsines for mice.
All figure*
are /i(7)io in mg min/I.
Kxposure time = 10 min; observation period = 10 days.
Agent
Total exposure
Methyldichlorarsine
2,7 (anal.)*3
Kl hvldirhlorarsine
1.555 (anal.)13
Kthvldichlorarsine
3.4 (nom. )«•
Fropvldichlorarsinc
1.4 (anal.)13
Biitvldichlorarsine
1.8 (anal )«
Hutyldichlorarsine
3.7 (nom.)130
Amyldiclilorarsinc
1.4 (anal.)1-1
Anivldiehlorarsine
3.7 (nom.)u7
Isoamyldichlorarsine
3.7 (nom. )’*■
1 lexvldichlorarsine
1.5 (anal. )13
Fhenvldichlorarsinc”
3.4 (nom.)27; 3.3 (noni.)11*
Lewisite
1.5 (anal.)*3
lewisite
2.8 (nom. )27
These data show that none of the dichlorarsines:
tested are more toxie than L and that only amyldi-
chlorarsine and PI) equal L in systemic toxicity.
Thirty-five dihalogenated arsines and thirteen
nmnohalogenated arsines were examined for vesi-
cancy at the LKTL 6S without revealing any vesicant
superior to lewisite.
In general, the dichlorarsines are better vesicants
than the monochlorarsines,79 and the simple alkyl-
dichlorarsines compare favorably with L in respect
to “absolute” vesieaney, i.e., when evaporation of
the liquid from the skin is prevented by covering.
The introduction of a single chlorine atom on the
terminal carbon of a normal aliphatic substituent in
a dichlorarsine or the use of branched chain sub-
stituent groups results in loss of vesicant potency.27
Thus, ED is a more potent vesicant than L 27 when
evaporation from the skin is prevented, and aim 1-
diehlorarsine is a better vesicant than isoarnyldi-
chlorarsine.271”
Eve Effects
The vapors of the chlorarsines are generally irri-
tating to the eyes, leading to lacrimation and bleph-
Examination of the data of Table 7 leads to the
conclusion that all of the dichlorarsines tested, with
the possible exception of Ml), have essentially the
same toxicity toward mice.
Fifty-three dihaloarsiaes were tested at the UC’TL
and the conclusion reached that the members of
the series vary in toxicity up to a maximum in the
group that contains b, ED, and the homologous
straight chain aliphatic dichlorarsines. Data have
also been obtained for a number of monohalogenatcd
arsines,6< but none of these compounds are superior
to L.
References to the vapor toxicities of other halo-
genated arsines will lie found in Table 9.
SECRET PHYSIOLOGICAL SECTION
97
arospasm which protect against further damage.
Liquid Ml) produces a lesion in the rabbit eye which
is less severe than one caused by L.2,9r Liquid KD
produces a lesion in the rabbit eye which is compa-
rable iu severity to that caused by L.2aal* UAL is
effective in the prevention of eye damage from
either MD or KD.2®*bc
Assessment of the Military Value of Chlorarsines
Olhir than I.. Of all the chlorarsines studied, only
MD, KD, PD. hutyldiehlorarsine, and the ainyldi-
ehlorarsines approach L in toxicity and vesicant
potency. Of these, hutyldiehlorarsine is too un-
stable 130 and the amyldiehlorarsines too difficult to
prepare147 to lx considered as chemical warfare
agents.
Thus, after an exhaustive examination of many
compounds, it appears that the l>est of the chlor-
arsines other than L are those which were used (KD
and PD) or considered for use (M D) in World War I.
The status of MI). KD, and PD as military agents
has recently been reviewed2I* with the following
results:
1. MD. The vapor is so irritating that it is easily
detected at low concentrations and would lead to
prompt masking. The vapor is easily hydrolyzed and
the dosage required for skin vesicancy so high that
there is no hope of obtaining vesicant dosages of
vapor in the field. The skin and eye effects of the
liquid are not so damaging as those produced by L.
2. KD. KthyTdiehlorarsine is somewhat superior
to M D but is inferior to L as a casualty agent.
3. PD. The vesicancy, systemic toxicity, and
toxicity by inhalation of PD are equal to those of L,
but PD penetrates clothing less effectively than L
and the volatility of PD is so low that casual! ies from
exposure to the vapor are hardly to be expected in
the field. Like MD and KD, PD is easily hydro-
lyzed.
In view of these facts, it appears that tlie best of
(he chlorarsines are inferior to I, and. since L itself
does not appear to have any future as a chemical war-
fare agent, it can lie assumed that the other chlor-
amines will not be considered further as military
agents for casualty effect. It is interesting to note,
however, that the Allies captured a considerable
number of German artillery shells charged with a
mixture of mustard and PD. Whether this indicates
that (lie Germans held a higher opinion of the effec-
tiveness of PD than the Allies or the mixture was
dictated by other considerations is not clear at the
present time.
7.3..'5 Arsine and Nonhalogenated \rsine
Derivatives
A US IN K
During World War I, the Allies did considerable
exploratory work on the potentialities of arsine as a
chemical warfare agent. In 1919 it was stated; 124
During I lie war many suggestions were made that arsine
should t>e used The popular plan was to use magnesium arse-
nide which would hydrolyze in moist air, setting free arsine.
The experiments made by the Research Division showed that
the hydrolysis does not take place rapidly enough under or-
dinary conditions to give an efficient concentration of arsine.
At the time of the armistice e\|)erimeiits were still under way
to determine whether this material could be used effectively
in the rain. While the use of magnesium arsenide or of any
arsenide was not very promising, there seemed to l>e a distinct
(jossibility of using liquid arsine ... If arsine is to be used in
warfare, it seems probable that it must lie used as liquid.
In 1939. the available data concerning arsine as a
potential chemical warfare agent were summarized 124
with the conclusion that its value would depend on
whether (he canister of the gas mask would afford
sufficient protection against it under all conditions to
which the canister might Ik* exposed.
It was recognized that arsine might be useful as a
casualty agent aside from its lethal effects and ac-
cordingly studies of the toxicity and suitability of the,
compound for chemical warfare use were reinvesti-
gated by both the Americans and the British.
Physiological Action. The physiological action of
arsine has l>een well summarized in the open liter-
ature.**®
In vitro studies have shown that arsine is oxidized
aerobically in aqueous solution, and that this oxida-
tion is catalyzed by hemoglobin.*94* In the presence
of arsine and oxygen, however, the hemoglobin un-
dergoes destruction forming a number of compounds
including met hemoglobin, and a tetrapyrrolie com-
pound whose spectrum resembles that of sulfmel he-
moglobin.2Mb3,M,lh During the reaction of arsine with
hemoglobin about 40 per rent of the arsine taken up
is held in a nondialyzable form, while the remainder
is mostly arsenite with a small amount of arsenate.
There is no reaction 1x9ween arsine and hemoglobin
under strictly anaerobic conditions.2®41*
Arsine is a strong hemolytic agent in vivo; and
in vitro under aerobic conditions only.294b In view of
the known oxidation products of arsine, experiments
were carried out to determine whether the hemolytic
effects of arsine were due to arsenite or arsenate
rather than to arsine per sc, but with negative re-
sults.®3
SECRET 98
VHSEMCA LS
The action of arsine on tissue slices has been stud-
ied and compared with that of arsenide, with the con-
clusion that the effect of arsine in reducing the
oxygen uptake of kidney slices is similar to that of
arsenite.93 3041 HAL protects kidney slices against the
effects of arsine but not of arsenite.93 The action of
arsine on liver slices is not identical with that of
arsenite since the toxicity of arsine increases more
rapidly with increasing concentration, and liver
slices treated with arsine change color, suggesting a
reaction with heme compounds that does not occur
with arsenite-treated liver slices,303'-
Toxicity. Available data on the toxicity of arsine
by inhalation have been summarized.-16 The data
cited are quite variable both for exposures of a given
species and for different species. The LCM for mice
has I>een determiner! as of the order of 0.250 mg 1 for
a 10-minute exposure; 121 2,6 3031 but studies at the
U(TL23 resulted in a figure_of 0.520 ± 0.100 mg I
(analytical), with no apparent explanation of the
discrepancy.
There do not appear to be any satisfactory data
for the LC-m for dogs with 10-minute exposure, but
0.35 mg I for a 30-minute exposure is said to be the
LCsn,,M and lethal concentrations for various ex-
posure periods have been compiled.*'* Rabbits are
apparently less susceptible to arsine than mice, the
LCu> for 10-minute exposure Ixdng estimated to lie
between 0.65 and 0.96 mg 1.176 No satisfactory LC;„,
has lieen reported for cats, but 0.80 mg 1 for 10
minutes caused the death of 3 4 cats within 18 hours
(G-2 Report No. 1322216), whereas cats exposed to
4.1 mg I for 1 minute did not die.23
The LC-3n for rats on 10-minute exposure is of the
same order as that for mice, being between 0.39 and
0.66 mg 1 (G-2 Report No. 1322 *«). The LCht) for
goats on 10-minute exposure is estimated as being
between 1.0 and 2.2 mg 1 (G-2 Report No. 1322216).
Four of fi\ e monkeys died after exposure to 0.45 mg 1
for 15 minutes.3031
No data exist for the AC50 for man, but the mini-
mum disabling concentration has lieen estimated as
2.0 mg I for 2 minutes or 0.2 mg 1 for 30 minutes.'*8
Henderson and Haggard state that exposure to a
concentration of arsine lietween 0.051 and 0.191 mg I
would lie dangerous after 30 minutes, whereas ex-
posure to 0.798 mg I would be fatal after 30 min-
utes.328 British estimates based on the assumption
that 2 mg kg of arsine would lie fatal to man put the
casualty-producing Ct at 14 mg min 1 for a man at
rest and at 4.66 mg min 1 for a man working; and the
fatal Ct at 28 mg min I and 93 mg min I for a resting
man and working man respectively.3" Marly British
results indicated that for the effect of arsine on mice
the product CH rather than Cl was a constant, hut
later investigation showed that for concentrations
greater than 0.5 mg 1, Cl was constant, whereas for
concentrations less than 0.5 mg 1. C'!l was constant.3,Mf
On the grounds that the incapacitation of troops
may he as valuable as their death in most military
situations, and that the incapacitating dose of an
agent may l>e quite different from the lethal dose,
studies were carried out on rabbits to examine the
possibilities.178 The results indicated that exposure of
rabbits to 0.05 mg 1 for 10 minutes caused significant
changes in the oxygen-carrying capacity of their
blood, but that the effect was transient. After 10-
minute exposure to concentrations between 0.13 and
0.20 mg 1 the rabbits were no longer able to maintain
a relatively high red blood cell count, and the de-
crease in oxygen-carrying capacity of the blood was
severe in about half of the animals, whereas with
10-minute exposures to concentrations between
0.234 and 0.40 mg I a marked decrease in hemoglobin
was invariably noted. A similar decrease in the hemo-
globin content of human blood might be expected to
cause severe but sublethal casualties.
Therapy. J idhiul compoimds are effective in the
treatment of arsine poisoning, although BAL-ethyl
ether (2,3-dimereapt opropy I ethyl ether) is more
effective than HAL itself.**8 Since BAL-ethyl ether
is tolerated by human Ixangs in therapeutic dosages
without toxic symptoms, the compound appears to
be suitable for the treatment of arsine poisoning in
man.-’1®
Assessment of Valor as a Chemical Warfare Agent.
The conclusion of (he United States Chemical
Warfare Sendee in 1939 was that the value of arsine
as a chemical warfare agent would depend on the
question of canister protection.'25 The British in 1941
concluded that the only potential method for the
liberation of arsine would lie by high-capacity bombs
and that the only possible advantage over gases of
the phosgene type would be that its detection at low
concentration is more difficult. In order to utilize low
concentrations of arsine, however, exposure must be
prolonged and this is very difficult to obtain short of
excessive effort, so that on the whole arsine should
not merit any particular consideration as an offen-
sive weapon, provided respirator protection is ade-
quate.236
The question of canister protection against arsine
SECRET PHYSIOLOGICA I. SECTION
99
has been summarized as follows; “At one time arsine
was thought to lie a very promising war gas because
it penetrates humidified unimpregnated or copper
oxide impregnated charcoal very readily. With the
introduction of silver impregnation, however, the
protection against arsine was made almost compa-
rable to phosgene. .. .” 100
The weight of arsine that would have to be ex-
pended to produce a lethal concentration is theoreti-
cally about 10 times as great as the weight of phos-
gone required for t lie same purpose.236 Since, in ad-
dition, modem res) lira tors give adequate protection
against it, arsine shows little promise in chemical
warfare.
Noxhalogexatkd Arsixe Derivatives
A numher of tertiary arsine derivatives have been
examined for toxicity. Data for 51 such compounds
were obtained by the UCTL,** and reference to these
and to other tertiary arsines are listed in Table 0.
Tabi.e 9. Arsenical compounds examined as candidate chemical warfare
The com|Miumls in Table It are arranged in the following categories:
1. Derivatives of arsine.
2. Derivatives of primary arsines.
3. Derivatives of secondary arsines.
4. Tertiary arsines. “
5. Quaternary arsenic derivatives.
0. Arsenic analogs of hydrazine.
7. Derivatives of arsenic oxides, sulfides, and amines.
8. Halogen and oxygen derivatives of tertiary arsines,
9. Derivatives of arsenic, arsenic, and arsinic acids. —
10. Arsenic derivatives of uncertain constitution.
British reports describing the examination of com pounds marked with an asterisk are not
Centigrade scale is used throughout the table.
agents,-
all available.
Reference
_ _ to
Compound synthesis
Physical properties- 1{efc™
toxicity
Properly Reference data
Derivatives of arsine
1. Calcium arsenide — — 311
d1*
2.5
311
311
2. Arsine 296b, 311
d”
1.44
296b
23, 311
"
nip
110.1116.0° 296b
*'P
62.8°
296b
3. Arsenic tnflnoride 311,333
d*
2.6659
298a
68, 311
—
nip
8.5°
298a
. . .
i>p
60.4°
29Sa
• . .
vol
152
311
4. Arsenic trichloride* ~ ...
NdIU
1.6009
311
252
,p«
2,163
244
- _ ....
inp
13°
311
bp7*"
129-130
244
...
voP6
84
311
5. Arsenic trichloride - dioxane complex*
227
6, Arsenic trichloride — thioxane complex*
227
7. Arsenic pentafluoride 342
mp
80.4°
298a
Derivatives of primary arsines
bp
52.8°
298a
...
8. Mcthylarsine " 5
bp
T
5
9. Melhyhlifluorarsine* 296c, 298a
d
1 9725
29Sa
298a
nip
30°
296c, 298a
■ ‘ ’ — - - . . .
bp
76°
296c, 298a
10. Mcthyldichlorarsine* 32, 200j, 311
n i/8
1.5588
32
27, 43,68,
311
rp°
1.8358
32
■ ...
mp
42.5°
32
bp7*"
132.5°
32
•
vol1"
74.4
311, 70
* —
68.3
11. Chloromcthyldichlorarsine — 47
bp1®
53°
47
27, 68
...
vol
135
27
SECRET 100
ARSENIC ALS
Compound
Heference
to
synthesis
Physical properl ies
Properly Hefcrenct
Reference In
toxicity
data
12. 2-Chlorovinyldifluorarsine*
113, ISO, 290c
J*
1.97
ISO
121
—
mp
20°
180
bp" 4
bp
vol2*
13.5°
105 110°
31.77
296c
ISO
180
13. 2-Chlorovinyldichlorarsine*
See llibli-
Hu5*
1.6073
43
See llibli-
Lewisite (isomer 1)
ogrnphy
d24
1.879
27
oKraphy
'
mp
2.4°
27
bp10
75°
43
bp16"
1 !KJ°
27
Vol-°
2.3
311
VoI‘°
4.47
70
Lewisite (isomer 2)
See Hibli-
II I.51
1.5900
27
See Hibli-
o^mphy
d2*
1.8681
27
ography
bp10
02.8°
27
. . .
14. 2-( dilorovinvldkhlorarsine-dioxanc complex*
...
bp76”
150.2°
27
227
15. 2-Chlorovinvidibromoarsinc*
231
bp17-1* —
106-107
231
231
in. 2-Hromovinvldibromoarsine*
231
bp”*
132 137°
231
231
17. 2,2-1 Mchlorovinyldichlorarsine*
311
....
311
IS. Elhvlarsine
39
tP*
1.217 “
39
68
bp73*
35 36
39
19. Kthyldifluoroarsine*
112, 296c
d
1.743
296c
_-ii7
‘
... —
nip
38.7°
296c
bp
94.3“
296c
20. Ethvldichlorarsine* (KD)
5, 32, 4S,
n uH s
1.5588
311
27,43, t»S,
58, 311
cP"
1.6595
32
79,311
bp734
153 155°
5
_ .
voli0
21.9
127
—
...
Vol29
30.2
70
21. Et hyldibromoarsiue
58
n if*
1.6405
58
68, 79
(P‘
2.103
58
bp'6
87-88°
58
vol2*
5.72
70
22, 2-Chloroel hyldichlorarsine*
1,111, 114
bp1*
99.8-100°
1
27, 79, 116
23. 2-Hvdroxvelhyldichlorarsine*
227
24. 2-Methoxyethyldichlorarsine
1
(P“
1.693
1
27, 79
bp*
94 95-
1
bp14
102 103°
1
...
25. 2-El box vet hyldichlorarsine
1
iP«
1.605
1
27, 79
bp19
95-97°
1
20. Allvldichlorarsine
1,32,38,
«D79
1.5702
1
105
,p‘
1.6294
32
bp4-*
42°
32
68, 121,
27. 3-Chlorallvldichlorarsine
301c
bp18
104 105°
301c
79, 120
227
28. Fropyldichlorarsinc*
32
no28
1.5297
32
43, 68, 79
,P«
1.5380
32
mp
28.2°
32
—
bp7*
99°
32
;;;
bp76"
vol20
175.3°
12.4
32
70
29. Propyldibromoarsine*
MO. Propyldicyanoarsine *
-rr. .
227
3!)
nip
82-86°
39
68. 79
31. Isopropvldichlorarsine*
227
32. 3-Chloropropyldichlorarsine*
311
311
33. 3-ChloromethoxypropyIdichlorarsinc
5
bp*
130-137°
5
27, 79
Table 9 (Continual).
SECRET PHYSIOLOGIC.*I, SECTION
101
Table 9 (Continued).
Compound
Reference
to
synthesis
Physical properties
Property Roferenco
Reference to
toxicity
data
34. 2-('hlor»>-3-(2-chIorocthyltliio)-l-bulenyldi -hlor-
nrsine*
35. Butyldichlorarsine*
I, 32, 130
d3"
1.4064
32
7~
:N -r
1
N
bp7**
194’
32
121, 130
vol3*
6.3
1.30
36. Bulyldibromoarsine*
37. Hutyldicyamiarsinc
58
inp
61-63°
5S
227
68, 79
38. sw^But-yldichlorarsinc*
32
nua
1.5245
32
27, 68, 79
,p«
1.4128
32
bp7**
182°
32
—
bp11
39“
32
31». 2(or 4)-( 'ldoro-3-methyl-1,3 (or l,2)-bul-adieayl-
dichlorarsine*
227
40. 4-Pentenyldiehiorarsinc
32
»■>**
1.5698
32
27, 68, 79
. *.’
d»
bp5*
1.453
102.5-105.5
32
° - 32
■ ■ - ■
bp!‘
111 114°
32
vol
0.12
27 —
41. 2-Chloro-l-| >oiitonyIdichlorarsinc
38
bp54
130 1330
38
68, 79
42. A my la rsi ne
‘ 58
bp'3"
125-127°
58
68
43. Amyldiclilorarsiuo*
28, 32, 58
ntr*
1.5177
32
27, 43, 157,
—
(P'<
1.4035
32
79
_
bp3*
118°-
32
44. Amlydiliromoarsino
39
bp34"
an11*
213°
1.5760
32
39
68, 79
lP»
1.8804
39
bp18
125.5 127'
39
...
bp734
248°
39
vol
0.399
70
45. Amyldicyanoareinc
58
nip
6(b 69.5°
58
68, 79
46. Isoa my id ichl< >ra rsine
28, 32
an3*
1.5157
32
68, 79, 157
-( ’hloroacetylphcnyldiclilonirsiiie*
227
91, p-Xenyldichlorarsine*
227
92. 2-{4-C'hloroacetylphcnyI)phenyklichlorareine*
227
93. 2-Fluorencdich It >rarsiuc *
227
94. 2-Fhlorenonedichlorarsine*
227
95. o-Benzoylphenyldichlorarsinc*
227-
96. 2,2'-his(Dichloroarsino)still(cne*
... -
227
97. 2- Xa ph 1 hyldichlorareine
73
mp
74.6-75.1°
73
68, 79
98. 2-Furyldichlorarsine*
l,25,301d,
311
no1**
1.6092
25
27, 311
erl ies
Projierty Reference
Reference to
toxicity
data
100. 2-Dichlorarsinophenoxthiin
84K
nip
64 -65°
101 d
68, 79
110. 6-I)ichlorarsino-2-phenvlben/thiazole hydrochloride
84h
111. l»i(t(2-Dichlorarsinoclhvl)Kiilfonc
1
mp
70.5 80.5°
1
68
112. o-Phenvlerie-bbKdichlorarsinc)*
227
113. «(-Phcnvlcne-6/s(dichIorarsine)*
227
114. p-Phenvlcne-fc/s(diehlorarsine)*
58, 311
d
2.15
311
68, 79, 311
-
mp
97 98°
58
bp”
200°
311
115. 1,\-hiM L)ichh)rarsino)-2-nil rolienscne
58
nip
72.6-74.3°
58
68. 79
116. p-Dichh)rostibinophenyldichlorarsine*
...
227
117. 2-I)lchlorarsiuudiphenylchlorarsinc*
227
118. 6is(/i-I)ichlp
160-161°
58
124. Dimel hylthiocyunoarsinc
73
nii”
1.6100
73
68, 79
1.4695
73
bp25
106-107°
73
125, 6i«(Chloromet hyl)chloroarsinc
47
d
ca. 1.85
27
27, 68, 79
bp10
75°
47
-s.-.
126. Reaction product of mercury chloroacctylidc and ar-
senic trichloride*
227
127. 6(’s(2-ChlorovinyI)chlorarsine*(lewisite II)
311
nlt"
1.6096
311
235, 311
d"
1.7047
311
bp'“
112°
311
bp30
136°
311
128. 6i)i(2-('lilorovinyl)cvanoarsine*
.
227
120. 5is(2,2-Dichlorovinyl)chloroarsine*
311
....
311
130. h/s(2-Hromovinyl)hromoarsinc*
231
bp’6
153-158°
231
231
131. bis{ 1,2,2-TrichlorovinyDchloroarsine*
231
bp5
141°
231
231
132. Diet hvlchlorarsinc
58
n o2;
1.5080
58
68, 79
d“
1.368
58
...
bp"
52-54°
58
.
bp™
156°
58
133. Diethylcyanoarsine
58
Bn”
1.4863
58
68, 79
d!h
1.238
58
bp11
80-81 °
58
lip737
190-191°
58
~ . . .
134. Kthylpropylchlorarsine
39
tP”
1.330
39
68, 79
bp-"
82 85°
39
bp7”
176°
39
135. I't hylpropylcyanoarsine
58
n ir7
1.4838
58
68, 79
<7*
1.194
58
bp77
110 113°
58
vol3"
1.45
70
130. Kthylpropylthiocyanoarsine
73
Hi)1*
1.5674
73
68, 79
1.286
73
- ...
bp" 66
102-110°
73
137. Ethylbutylchlorarsine
58
no3'
1.5025
58
68, 79
d36
1,272
58
bp”
89 92°
58
.....
SECRET 104
ARSE MCA I.S
Tabi.K 9 (Continued).
Compound
Ilefcrence
to
synthesis
Pro
Physical proper!
|x‘rty
ics
Hefprrnct*
Reference to
toxicity
data
138. Kthylbutylcyaniarsinc
58
n i,-'_
1.4828
58
68, 79
iP‘
1.152
58
139. h(x(2-Chh>ro-3-(2-chloroethyllhio)-l-butcnvl) chlor-
bp»»
112-112.5°
58
arsine*
227
110. F )ihntvlu«hmrsine
841.
68, 7'.*
141. hidCvclohexyl tehlora rsine*
227
142. Methvlphcnylarsine
73
(P
1.31
73
bp57
108-111°
73
bp"
128-129°
73
'
143. Methvlphcnylehlorareine
58
«n31 ~
1.6022
58
68, 79
■ , .. *
,P<
1.449
58
bp”
127°
58
14 I. Mel hylphenyliodoa rsine*
145. Melhylplienylcyanoarstne
58
n n76
1.5812
58
68, 79
1.372
58
bp="
147-148°
58
14*1. Met hvlphenvlthiocvanoa rsine
58
n n”
1.6577
5S
68, 79
1.433
58
bpI4~'8
176-179°
58
147. m-Chlorophenylmelhylchlorarsine*
...
227
148 Methvl-w-nitrophenvlehlorarsine
73
n,r
1.6272
73
68, 79
roa rsine*
227
163. Diptienylchlorarsine (DA)*
73, 311
n D»
1.6429
73
79. 68, 311,
249
—
1.413
73
' •
mp
38°
244
bp" 7
157 160°
73
vol50
<0.0001
. 311
164. 2-('hlorophenylphenylchlomrsine*
301e
mp
30-35°
301c
227
165. 3-Chlorophenylphcnylchlorareinc*
227
166. 4-ChlorophrnyIphcnylehlora rsine*
.. r.
227
167. 2-Nilrophenylphenylehlorareinc*
227
168. 3-Xitroplienylphenylclilorarsine*
227
169. 4-Nitropheiivlplienylchlorarsine*
227
170, h/>(2-Chlorophenyl)chlorn rsine
30Ie
mp
73 75°
:301c
227
171. 5/>(4-Chlorophenyl K'hlorarsine*
227
172. fc/«(3-Xitri»plienyllehlornrsine*
58
mp
112-113°
58
68, 79
173. 2-Phenvlchlorarsinoaniline hvdroehloride*
....
227
174. 3-Phenvlchlorarsinoaniline hydrochloride*
—r. .
227
175. 4-Plienvlchlorarsinoanilino hydrochloride*
227
176. 67*(»i-Aminophcnyl)chlorarsine dihydrochloride*
227
SECRET PHYSIOLOGICAL SECTION
Compound
Reference
to
synthesis
Physical properl ies
Projierly Hcferi-noe
Reference to
toxicity
data
177.
6(«(/*-Aminiiphcnvl)chlorarsine dihydrochloride*
227
178.
hix{ ;>- Met hoxv phenyl lehlorarsine*
227
179.
1 Jiphenylhromoarsine*
341
. .77
121
iso.
Diphenvlevanoarsine ( DC)*
311
n a50
1.6254
244
311
J*
1.3327
244
inp
31.2°
244
- -
Veil*"
0.0001 0.00015
311
1KI.
2-Chlorophenvlphenvlevanoarsine*
301e
nip
40-42'
:30ic
>27
182.
3-( ’hlorophenylphenylcyanoarsine*
227
183
l-Chlorophenylphenylcyanoarsine*
30 le
mp
102
301c
227
184.
2-Nilroplienylphenylcyanoarslne*
>27
185.
3-X it rophenylphenylcyanoarsine*
227
ISO.
4-Nitrophenylphenylcy»noarsine*
227
187.
bm(‘2-( 'hlorophcnyl )cyanoarsine*
30 le
nip
85-87°
301 e
227
188.
ftj*(4-Chlorophenvl)cyanoar>dne*
227
189.
bi»( 3- Nil ropheiiyl )cyanoarsine*
58
nip
151-152
58
68, 79
190.
2-Phenvlevanoarsi noaniline*
....
227
191.
3-PhenvlevaiUKirsinoaniline*
227
192.
4- Phen vie vain mrsinoaniline *
. . .
227
193.
hixfin-Aminophenyllcyanoarsine*
227
194.
Diphenylt hiocyanoarsiue*
58, 301 j
1.6766
58
68, 79
•
1.379
58
bp*-*
217 219°
58
195.
hix{ m-N i I rophcnyl it hit icy anon mine
58
mp
103-105°
58
68, 79
196.
Phenvl-o-tolylcvnnoarsine*
301e
■227
197.
o-Chlorophenyl-o-tolyfcyanoarsine*
.
227
198.
p-Chlorophenyl-o-tolylcyanoarsine*
227
109.
o-Nitrophenyl-o-lolylcyanoarsine*
227
200.
i/i-Nitrophenyl-o-lolylcyanoarsine*
.
227
201.
;j- N i I rophcny 1-o-toly Icy anoa rs i i in *
—
. T.-
227
202.
Phenyl-w-tolvlcvanoarsine*
227
203.
Phony 1-p-tolylchlorarsinc*
30 le
-...
227
204.
Phenvl-p-tolvleyanoarsine*
30le
227
205.
o-Chldrophenyl-p-tolylcyanoarsine*
....
‘227
206.
«i-Chhirophonyl-/>-tolyleyanoarsine*
227
207.
p-Clilorophenyl-p-tolylcyanoarsine*
. - -
227
208.
p-Nitrophenyl-p-toiylcyanoiirsine*
227
209.
m-(Pheiivlcyanoarsino)bcni!aldehyde*
7. .
227
210.
o-( Phony lohlorarsinolbenzoic acid*
227 -
211.
Methyl o-(phcnylchlorarsino)l)enzoate*
227
212.
w-( Phenvlehlorarsino)bonzoio. acid*
:301b, 301 j
mp
134 136°
3011.
301 h
213.
Methyl m-{phenylchlorarsino)benzoate*
301b, 301 j
,. .
301 h
214.
Methyl »i-(phonylcyanoarsino)l»eiizoate*
301b, 301 j
_
30 Ih
215.
;>-(Phenvlehlorarsiiio)l)enzoic acid*
301b, 301 j
mp
115 117°
3011.
301 h
216.
Mcthvl p-{plieiiylchlorarsino)bcnzoate*
301b, 301 j
• ...
301 h
217.
Methyl p-(phenylcyanoarsino)benzoate*
301b, 301 j
....
301 h
218.
2,4-1 )i methyl phony Ipheny Icy anoarsine*
227
219.
his{ o-Tolvl )cyam mrai ne *
:301c
mp
74°
:30ic
227
220.
bi»(o-i ’a rlxiinel hoxvphcnyl lehlorarsine*
227
221.
o- T ol v 1-m-l ol vlcvanoarsi ne*
227
222.
o-TolvI-p-tolylcyanoarsine*
227
223.
bix( m-T i >lyl )cyanoursine*
. • •
227
224.
t/i-Tol \ 1-p-t«>ly leva noa rsi ne*
227
225.
bi.i( p- T i dyl leyanoarsine
30 le
mp
62“
301 e
227
226.
/«-(Phcnylehlorarsiin»)acetophenone*
301 j
mp
71 72°
30 Ij
227
227.
it) A Phenvlcyalioiirsiiio)«celophenonc*
30 Ij
mp
57-59°
301 j
227
228.
/«-( Phony Ichlorarsinol-uM’hloroa cot ophenone*
301 j
227
229.
;«-( Phony loyanoa rsi no)-t^-chloroacetophenono*
301 j
227
230.
2.1-I)i met hylphenyl-«-loly leyanoarsine*
...-
301f
231.
2,4-l)imothvlphonyl-//-tolylcya noarsine*
301 f
232.
tr-N.aphl hylpheny lehlorarsine*
.,.
„ .
227
233.
d-N aph t hylpheny lehlorarsine*
227
Table 9 (Continued).
SECRET 106
ARSENICA LS
Tvhi.e 9 (Continued).
Compound
Reference
to
synthesis
Physical properties
Pro|.erty Reference
deference to
toxicity
data'
234. a-Xaphthylphenylcyanoarsine*
301e
mp
98 99°
30 le
227
235. 0-Xaphthylphenylcyanoarsine*
227
236. a-Xaphthyl-p-tolylcyanoarsine*
227
237. (8-Xaphlhyltp-lolylcyanoarsine*
227
238. 6is(nr-Xa|ilithyl)ch!orarsiiu-
301 j
mp
163-165°
301 j
239. h.s-or-Xaphthylcyam.arsinc*
301 j
mp
191°
301g
301g
210. 6(s-0-Naphthylchlorarsine*
227
241. b.K-d-Xaphthylcyanoursine*
...—
227
242. Phenyl-(xV-thienylchlorarsine*
301 e
hp0*
150 156°
30 le
227
243. Phony l-fxj-thicnylcyanoarsine*
301c
mp
49-51°
30le
227
bp" *
168-174°
301o
24 1. Phem’l-3-pvridvlchk.rarsine hydrochloride*
227
245. Phenvl-3-pvridvlchh.rarsine melhiodule*
227
246. Phenyl-3-pvridylcyanoarsine hydroohlorido*
...
227
217, 5-Phcnylchlorarsino)-2-chloropyridinc hydrochloride*
248. 5-( Phenvlehlorarsim.)-2-amin. .pyridine dihydrochlo-
•••»-
227
ride*
227 “
249. 6i8(or-Furyl)clilorarsinc* _
1,25, 30Id
nn2S
1.6082
25
68, 79
>"
1.5909
25
:
bp*
122 127°
25
250. 6i*(o-Furyl)cyanoarsine*
39, 30 lo
tivr*
1.5749
39
68, 79
...
1.4857
39
. .5
l.p51
142°
39
251. fcfs(a-Thienyl)chlorarsine*
T..
227
252. ft i»(a-Th io ny 1 )cya noa i> i 110 *
30 le
mp
51 55°
301e
227
bp1’6-
180-182°
301 e
253. his(3-Pvridyl)chlorarsine dihydrochlorido*
58
mp
270-273°
58
68
254. bis( 3- Py ridyl )cyanoarsi no *
227
255. p-Phonylenechlorarsino
1
mp
ca. 145°
I
256. 1-Chlorars indole*
257. 2-(Dielhylaminomethyl)-!,3-dichh.rarsindole hy-
227
drochloride
73
mp
204.2 205.2°
73
68
258. 5-Chlorodihenzarsenole*
5, 3071.
mp
161°
5
68
i.p’-4
230°
5
259, 5-(’lil<.ro-3,7-dinitrodil.enzarsenole*
— 227
260. 5-Chlor.>3-amini slit .enzarsen. tie*
227
261. 5-Chloro-3,7-diaminobenzarsenolc hydrochloride*
227
262. 5-Cvanodil.enzarsenole*
5
mp
178°
5
68
263. 5-l«dodibcnzarsenole*
227
264. 4,1 '-Diphcnylenechlorarsine*
227
265. 5-Chloro-5,10-dihydroacridarsinc*
39, 3071.
nip
113 114°
39
227
266. 2,5-Dichlomd>,10-dihydroaeridflrsine*
307m
mp
116°
291e, 307m
29 ic
267. 5-Cyano-5,10-dihydr. laoridarsino*
307k
mp-
114-115°
307k
285. 291.1
268. 2-Chloro-5-cyano-5,I0-dihvdroacridarsine*
307m
mp
113-114°
307m
285, 291c
269. 5-Chloro-10-oxo-5, lO-dihydroacridarsine*
227
270. 6-C 'hh>r<>-2-mct hyl-5,10-dihydroacridarsinc*
307j
mp
87°
307j
285
271. 5-Cvano-2-melhvl-5,10-dihydroacridarsine
mp
87°
29 If
285, 29If
272. 5-Chtoro-3-met hyl-5,10-dihvdroacridarsine*
mp
65.5-66.5°
307n
29 le
273. IO-AcetyI-5,10-dihydrophenarsazine*
227
274. 10-Acel vl-5,1O-dihvdrophenarsaziiie piorate*
227
275. 10-Trichloroacetyl-5,10-dihydrophenarsazine*
227
276. 10-Chloro-5,10-dihydrophonarsazine (DM )*
38. 73, 311
>r-"
1.648
311
68, 79,311,
104
mp
189-190.4°
73
vol;o
0.00002
31
277. 5-AcetvHO-chloro-5,IO-dihvdrophcnarsazine*
♦ . .
227
278. 5-Propionyl-IO-chloro-5,IO-dihydrophenaraazine*
227
279. 5-Benzoyl-10-clJoro-5,10-dihydrophonarsa2ine*
227
280. 10-Hromo-5,10-dihydrophoua rsazino*
227
281, 10 lodo-5,10-dihydrophenarsazine*
227
282. 10-Cvano-5,10-dihvdrophenarsazine (Cyan DM)*
107, 108
121
283. l(>Thiocj'ams5,10-dihydrophonarsazine*
227
SECRET PHYSlOLCXaCAI, SECTIO>
107
Table 9 (Continued).
Compound
Reference
to
synl hesis
Physical proper! ies
Property Referenc*
Reference to
toxicity
data
2SI.
l(or 3),10-I)ichloro-5,10-dihydropheiiamazine*
227
285.
2,10-Dich!oro-5,10-dihydrophenarsazine*
227
280.
10-Chloro-5,10-dihvdro-4 (?)-nitrophennrsazine*
227
2S7,
l(or 3 ),2,10-Tricliloro-5,1O-dihydrophenarsazine*
227
288.
1,3,10-Triehloro-4j,l O-dihydrophenarsazine*
227
2S!».
l,9(or 3,7),10-Trichloro-5|10-dihvdrophenarsazinc*
227
290.
2,8,10-Trichloro-5, lO-dihydrophenarsazinc*
227
291.
l,2,3,10-Tetnichloro-5,10-dihydropheimrsnzine*
227
292.
2,4,t5,8,l0-Pentahromo-5,10-dihydropl»enarsazine*
■
227
293.
2-Aminu-10-ehloro-5, 10-dihydrophenarsazine hydro-
chloride*
227
291
l(or 3)-Amina-10-chloro-5,10-dihydrophcnarsazine
hydrochloride*
227
295.
1-Amino-lO-ehIoro-5,10-dihydrophenarsazine hydro-
chloride*
—
227
290.
10-( ’hloro-5,10-dihydro-l(or 3)-methyIphenarsazine*
227
297.
IO-(.'hloro-5,10-dihvdro-2-melhylphenarsazine*
.... —
227
298.
5- Ace t y I- 10-cl ik m >-5,10-dihyd ro-2-met hy Iphena rsa-
zinc*
227
299.
lO-Chloro-5, lO-dihvdro-4-mcthylphenarsazine*
227
300.
1 (or 3), 10-Dichh>ro-5,10-dihydro-6-methyIpheimr-
suzine*
. —
227
301.
4-Amino-10-ehloro-5,10-dihydro-7-methylphenar-
sazine hvdn >chh iride *
227
302.
10-Chloro-5,10-dihydropheiiarsazine-l(or 3) car boxy-
—
lie acid*
227
303.
10-('liloro-5, 10-dihydrophenarsazine-4-carhoxylic acid* ...
227
304.
10-Chloro-5,10-dihydro-l(or 3)-0-dimelhylphenar-
sazine*
—
227
305,
10-('hloro-5,10-dihydn>-2,8-dimcthylphenar8a*ine*
227
300.
5-Aeetyl-10-ehloro-5,10*diHydro-2,8~4limet hylphenar-
sazinc*
__
227
307.
1 (or 3 )-Acetyl-1 0-chloro-5,10-dihydrophenarsazine*
~ ~
227
308.
l(or 3)-Acelo-10-hromo-5,l 0-dihydrophenarsazine*
227
309.
10-C'hloro-5,10-dihydro-l ,4,7-t rimet hylphcnarsazine*
227
310.
10-Chloro-5,10-dihydro- 2,4,7-t ri met hylphenarsazine *
. r.
227
311.
10-( ’hloro-5,10-dihydro-l(or 3)-propionylpheuar-
sazine*
—
227
312.
12-Chloro-7,12-dihydrohetiz (a) phenarsazinc*
...
227
313.
7-C'hloro-7,12-dihys
n ir’
1.5665
58
68, 79
d*
1.473
58
bp18
97-103°
58
vol!"
1.72
70
335. 6t«{2-Chioroethvl)methylarsine*
307p
bp**
50-55°
307p
227
336. bis(2-Iodoet hyl )met hvlarsine
307p
bp11
68-69°
307p
337. ft/s{2-F.thoxyethyl)melhylarsine
58
«n“
1.4664
58
68, 79
... —
d*
1.100
58
hp**
124-125°
58
bp“«
230°
58
338. triM.2-Cl.lorovinvl)an«ne*
73
227
339. (riX2,2-T)ichlorovinyl)arsine*
227
340. Triethylarsine
102
68, 79
341. 2-Chlon.ethvUlicthvlarsine*
307p
bp10
53°
307p
227
342. 2-llronmctl.vldiel hvlarsine*
227
313. fcis(2-ChlorocthyIJethylarsine*
307p
bp11
64°
307p
227
344. Ir/sfCvclohexyllarsim*
39
bp!
187-189°
39
68, 79
315. Trioclvlarsine
101a
0.9357
101 a
68, 79
bp‘»
238-240°
101a
'
bp1
181-185°
101a
346. Phcnvlarsenophosijcne
84 i
...
347. 2-(p-Dimethylarsinophenyl)quinoliue
84 j
348. 6/*(2-Chh»r«vinyhphenylarsii.e*
1, 231
tp"
1.384
1
68, 231
bp18
166-171°
231
319. 2-('hloroviiiyldipbcnylarsiiM>*
1
d*
1.327
1
68
350. Triphenylarsine
73
nip
58- 60.5°
73
351. tris( w-X it rophcnyl )arsinc
352. Ins(p-Dimethylaminophenyl)arsine sulfur monochlo-
7
ride addition product*
227
353. Tn-p-lolvlarsine*
...
...
227
354. Dimet hyl-2-pyridylarsinc
84j
68. 79
355. Difurvlmcl hvlarsine*
301e
267
356. 2-Chlorovinvl-5/s(2-furvl)arsii.c
267
»>pj
127°
267
267
357. fri *( 2- Fu ry 1 )arsi ne *
25, .30Id
phena rsazine )*
227
383. 10, I0'-his(5-.\cet vl-5,IO-dihvdroj»henarsazine)*
227
384. I0,10'-6i»(5, IO-l)ihydrophenarsazine) sulfate*
227
385. 2,2',4,4',6,6-ltexanitroarsenobenzene
7
386. 4,4'-Dihydro.xy-3,3'-dinitroarsenolienzene
7
387, 4,4'-1>ihydroxy-3.3',5,5Metranitroarsenol;>enzene
7
Derivatives of arsenic oxides, sulfides, and amines
388. Arsenic oxide*
— _ _
227
389 Kthoxvdichlorarsine
841
68, 79
390. I timet hylaminodifluoroarsine
... -
68
391. Isopropoxydichlorarsine
84 m
bp7*1
155-156°
84m
392. 2-( 'hloro-4,5-dihydro-l ,3,2-oxthiarsenolc
84 in
«n2<
1.6690
84 m
68, 79
,r.i
1.988
84m
bp"-*
72-73°
84m
393. I tiethoxychlorarsine
841
68. 79
394 . 6/«(2-('hloroethoxy)chlorarsinc
47
lip1 *
112 118°
47
68
395. Diisopropoxyfluorarsine
84 j
396 tris( 2- KI uoroet hoxy (arsine
86
68
397. t r/*( 2-C h 1 o roe t h oxy )arsi ne
47
bp10
160-170°
47
68
398. trisi2-( ’hloroel hylt hio(arsine*
84 r
1.5972
S4r
68
399. /ris(Phenylthio)arsine
S4o
nip
92-94°
84 o
68
400. Mel hylarsine disulfide*
227
401. Phenyl met hoxychlorarsine*
227
402; F’henvlet hoxychlorarsine*
227
403. Phenvl-2-chloroelhvllhioehlorarsine*
297a
227
404. o-IIydroxychlorarsinolicnzoic anhydride*
58
mp
146.5-147°
58
OS
hp”
233-230°
58
405. o-Phenylenediarsine oxychloride*
73
nip
150.5-151.5°
73
68
406. Methylarsenic oxide*
311
mp
95°
311
311
bp
ca. 275°
311
407. Methylarsenic sulfide*
227
408. Met hvldimel hoxvarsine*
227
409. Melhyldimethvllhioarsine*
__
227
410. Methyhliethoxvarsine*
227
411. Methyl-5»s(X,X-diethvldithiocarbamvl) arsine*
227
4 12. Met hyl-5/x( N,S-bis(2-hvdroxvethyl (dithioear-
liamyl (arsine*
227
413. Methyld iphenvlt hioarsine*
227
414. Metliyldi-p-loly It hioarsine*
227
415. 2-( ’hlorovinylarsenie oxide (various isomers)*
12, 311
Physical properties vary
with method of prepa-
ration
122, 12
68, 79
416. 2-Chlorovinylarsenic sulfide*
227
417. 2-( 'lilorovinvlarsenie selenide
67
08
418. 2-Chlorovinyldimcthoxyarsine
68, 79
SECRET ARSENIC A LS
Compound
Reference
to
synthesis
Physical proper! ics
Property Reference
Reference t
toxicity
data
419. 2-Cblomvinyldimelhyhhioan.ine
68, 79
420. 2-Chlorovinyldiethoxyarsine*
30
bp17 ,s
84 85°
30
68, 79
421. 2-Cblorovinyl-5is(2-chlor.)cthylthio)arsine*
84p
" ir"
./-"
bp1
1.6100
1.610
SI 86°
Sip
84p
84 p—
68, 79
422. 2-( 3dorovinvl-feis( 2-ethoxyet hoxv )arsiiH»*
227
423. 2-Chlorovinyl-fc.a{2-hydroxycthyIthio)arsine
33
bp*
83.1 .86.7°
33
424. 2-Cldorovinyldintlyloxyarsine*
227
425. 2-('hlorovinyldiisopropoxyarsinc*
227
426. 2-ChIorovinyldi|>entoxyar8inc*
427. 2-( 2-Chl«rovinyl )-5,5-6/s( hydroxy met hyl)-l,5-dihy-
227
dro-l,3,2-dithiarsin
33
mp
127 5-128 5°
3.3
68
428. 2-Chlorovinvldiisooctvloxvars.ne*
227
429. 2-Chlorovinyldiphenylthioarsinc*_
430. 2-Chlorovinyl-5is(N,X-iliethyldithioc»rhamyl)ar-
227
sine*
227
431, Elhylarsenic oxide*
103a, 31)
1.5821
311
68, 79
—
»•*
bp'
1.8019
119°
311
103a
432. Kt hvldimct h vlthioarsine
68
433. Ethyl b)s(2-ch!urocthoxy)arsine
103b
68
434. Kthyldipropoxyarsine —■—
58
«n3" 4
1.4466
58
68, 79
.P7
1.114
58
•
bp'»
86-90°
58
bp™
185 186°
58
435. N-Ethylet hylareeni mide
58
»..**
1.5681
58
68, 79
,P-
1.498
58
-a-
bp’ 4
165 175°
58
436. Propyldiaectoxyarsinc
58
ni>!t
1,4715
58
68, 79
,r-‘
1,335
58
437, .Vrnylarsenic oxide
35
bp13
120 123°
58
438. Isoamylarsenic oxide
35
439. Ilcxvlarsenic oxide
35
440. Phenylarsenic oxide*
39, 311
mp
118-120°
39
68, 291a
441. e-Xit ro phenyl arsenic oxide
7
(Rut varies with method
of preparation)
311
442. w-Xitrophenylarsenie oxide
443. 2,4,6-Trinit rophenylarsenic oxide
444. 2,4,6-Trinilrophenylarscnicdinitrate
58
7
7
mp
184.5-187.5°
58
68, 79
445. PhenvT5ix{2-ehloroelhyllhio)arsine*
297a
227
446. p-IIydroxyplienylarscnic oxide*
227
447. w-Aminophenylarscnie oxide*
448. p-Dimcthylaminophenylarscnic oxide*
449. 3-Ainino-4diydroxvpheny)arsenic oxide hvdroehlo-
7
227
2911.
ride (Mapharsen)
450. o-Arsenosol ten zoic acid*
Commercial
....
334
227
451. m-.\rsenoeohcnzoic acid*
452. 3-Pvridvlarseuic oxide*
—
227
227
453. 2-f’h!oropyridine-5-arsenie oxide*
227
454. 2-Dihenzolhienylarseiiie oxide
84f
455. 4,4 '-biM Arscno8o)biphenyl*
227
456. 5(*(Dimethylarsenie) oxide (cacodyl oxide)
8, 38
bp
149 151°
38
68, 79
457. 2-Chloroethylt hiodimelhylarsine
84 m
bp1*
94 95°
84 m
68, 79
458. bix{ Dimethylarsenic) disulfide*
227
459. his( Diethylarsenic) oxide
17
460. 6i*(hi>-2-Chlorbviiivlarsenic) oxide
235
461. 6is(2-Chlorovinyl)-2-cl.loroelhylihioarsine-
84.,
W I)*6
1.6085
84 q
68
bpni
128-129°
84q
462. 6i«(2-Chloroviiiyl)-l,3-diehloropropyl-2-thioar8ii>c
S4s
bp*-*
148-151°
84s
68
Tabi.k 0 (Continual). PHYSIOLOGICAL SECTION
111
Table 9 (Continued).
('on»|H>und
Reference
to
synthesis
Physical properties
Property Reference
Reference to
toxicity
data
463. Melhylphenylmethoxyarsinc
58
nu"
1.5613
58
68, 79
tP'
1.295
58
bp"-
101 102°
58
464, Mclhylphenylacetoxyarsinc
58
nn"
1.5612
58
68, 79
oxvphenyl) hvdroxyarsinc anhydride*
.
227
472. bi»{o-Carl«)xypln nylpheiiylarsenie) oxide*
227
473, hiM /> (. ‘arboxyphen vlphenvlarsenic) oxide*
•227
474. p-Phenvlene-6(s(phenylarsinc) monoxide*
—
227
475. Acetoxydithienylarsine*
227
476. 1,3-Dihvdroxvars indole*
. .
227
477. bid Dilienzarsenyl-5) oxide*
227
478. b;.<(3,7-D'nitr<>diben*arsenyl-5) oxide*
...
...
227
479. b ts( 3- A tn i in td i be n*a rseny 1-5 ) oxide*
227
480. bis( riienoxarsinvl-10) oxide*
288a
mp
182“
227
481, 10-Chloroe(hvlthio-5,10-dihvdrophenarsazine
84 n
mp
128-153°
84n
482. ?»>(5,10-Dihvdroplienarsatiiiyl-101iixide*
227
483. bis( 5, UP Dihydrophenarsazinyl-10) sulfide*
484. his(b, 10-Dihvdrophenarsazinyl-10) oxide. 10-Chloro-
•••
227
5,10-dihydrophenarsazine (basic DM)*
227
485. 6/«(5-Acctvl-5,10-dihydrophcnars«zinyl-10) oxide*
227
486. b/.s(5-llen/,ovl-5,IO-dihydrophenarsazinyl-IO) oxide*
. .-r-
227
487. 5,10-Dihydroarsa n 1 hrene-5,10 monoxide*
488. 5,10-Dihvdro-5,10-dioxvstibarsanthrene-5,10 mon-
227
oxide* _ _
Halogen and oxygen derivatives of tertiary arsines
227
489. Triphenyldichlorarsine*
227
490. Tri-wi-nitropbenyldibromoarsinc
7
491. Tn-p-tolyldichlorarsine*
227
492. Tri-m-nitrophenvlarsenic oxide
7
493. Tri-m-nitroplienvlarsenie dinit rate
7
mp
147-148°
7
-..
494. o-Carboxypbenylplieiiylmelliylareenic oxide*
495. 7-Plienvlmethvloxidoarsino-2-naj»lithalcne8iilfonie
•227
acid*.
Derivatives of nrsenie, arsonic, anil arsinir arids
- ■”
•227
496 o-Nitroanilinimn arsenate
7
mp
146-147°
7
497. w-Nitroanilinium arsenate
7
mp
114 115°
then
7
—
147-148°
7
498. p-Nilroanilinium arsenate
7
mp
77-78°
7
499. Sodium metbanearsenate hydrate
69
08
500, 2-Cliloroethvleiiearsonic acid*
69
mp
129°
69
68, 79
501. Kthanearsonic acid
fit)
mp
90°
69
68
502. Benzenearsonic acid
58
mp
165°
58
68, 79
503. o-Nitrobenzenearsonic acid
7
504, Cadmium o-nitroliemsenearsonate
34
505 Ijead o-nit rolK-nzenearsonate
7
506. Hi-Nitrolienzenearsonic acid
7 .
...
507. I .ead m-nitrobenzenearsonate
7
508. p-Xilrohenzenearsonic acid
7
509. Sodium 2,4-dinitrobenzenearsonale •
7
510. Magnesium 2,4-dinit rohenzenearsonate
84a
511. Potassium 2,4-dinitrobenzeneareonalc
S4a
...
SECRET 112
ARSENIC\LS
Table 9 (Continued).
Compound
Reference
to
synt hesis
Physical properties
Property Reference
Reference to
toxicity
data
512. Manganous 2,4-dinitrolienzenenrsonate
84a
513. Ferric 2,4-dinitrolienzenearsonate
84a
514, Cohalt 2,4-dinitrolienzenearsonate
84 a
515. Nickel 2,4-dinit rohenzencarsonate
84a
510. Cupric 2,4-dinitrolienzenearsonate
S4a
517. Cadmium 2,4-dinitrolienzenearsonate
34
518. Stannic 2,4-dinitrolienzenoar8onate
84a
510. Barium 2,4-dinit robenzenearsonate
84a
520. Mercuric 2,4-dinitrolienzenearsonate
H4a
521. I-ead 2,4-dinitrobenzenoarsotmte
101 b
522. 2,4,0-Trinitrobenzenearsonie acid
7
523. Sodium 2,4,0-1 rinitrolienaenearsonate
84a
524. Potassium 2,4,6-trinitrobenzeneansoiiate
84a
525. Calcium 2,4,0-trinitrobenzenearsonatc
84 a
520. Cupric 2,4,6-trinitrolienzenearsonate
84a
527. Cadmium 2,4,6-trinitrobenzencarsonate .
- 34
52H, Stannic 2,4,0-1 rinit ml x-nzenoarsonate
84 a
.... TT."
_
529. Barium 2,4,6-trinitrolienzenearsonate
84a
530. Mercuric 2,4,6-trinit rohenzencarsonate
84a
...
531. la-ad 2,4,6-1 rinitmlicii/.enearsonate
7
5142. p-Ilvdroxylx-nzenearsonic acitl*
227
533. 4-Hvdroxv-3-nitrobcntsefiearsonic acid
7
534. la-ad t-hvdroxv-3-nitmlienzenearsonate
7
535. 4-llvdroxv-3,5-diniirolicnzenearsomc acid
7
530. Cadmium 4-hydroxv-3,5-dinitrobenzencarsoiiale
34
537. Lead 4-hvdroxy-3,5-din rtrobenzenearsonate
7
538. ('admium 2,4-dihvdroxy-3,5-dinitrol>enzenearsonale
34
539. o-Arsanilic acid*
227
540. 2- \lnino-5-nitrobenzenearsonic acid*
227
541. p-Arsanilic acid picrale
7
mp
109-170' 7
542. 4-Amino-3,6-dinitmhenzenearsonic acid
7
543. Lead 4-ami no-3,5-dinitmlienzenearsonate
7
544. o-Tolucnearsonic acid*
.... ...
227
545. 2-Hiphenylarsonic acid*
.... . . ._
227
540. 4,4'-14iphonvldiar»onic acid*
227
547. 2-Benzophenonearsoiiic acid*
84f
227
548. S-Metlivl(juinoline-5-arsonic acid
549. 3-Dilienzothiophenearsonic acid
84 f
550. X-Kthvlcarbazole-3-arsonic acid
Sle
551. 1,3-Dihydroxv-l-oxyarsindole*
227
552. 5-llvdroxv-5-oxvdilienzarseiiole*
mp
7250 5
227
553. 5,lli-Dihvdro-10-hydroxy-10-oxyphenarsazine*
227
554. 5,10-1 )ihvdro-10-hvdroxv-l0-oxypl«*narsazine hy-
drochloride*
227
555. 10,10-Dili vdroxv-10-ethylplienoxarsine*
227
550. 5,10-Dihydn>-10,10-diliydroxy-l0-ethylphcnarsazine*
227
Arsenic derivation of uncertain constitution
557. Chlorination product of Propane- 1,3-diarsonic acid*
558. Bv-producl of the preparation of dimethylaminodi-
227
fhioroarsine
(iS
559. Anhydride from o-livdroxvphenylarsenie oxiile*
227
560. Di(fn>(.5,10-dihydrophenarsazine l0)oxalate)-awfate*
227
Xo compounds of unusual toxicity were discovered
in this class, and, although some of the memliers a,>-
pearetl to offer potentialities as irritant smokes, none
showed evidence of lieing significantly Itetter than
the standard irritants DM. DA. and DC-
T.:t.t Irritant Arsenical Smokes
Certain arsenical compounds which arc relatively
nonvolatile and of fairly low toxicity are, neverthe-
less, highly irritating to the respiratory tract when
dispersed as a cloud of very fine particles. Further-
SBCRET PHY SIOLOOICAI, SECTION
113
more. such particles will i»enctrate gas mask can-
isters unless the canisters are fitted with an efficient
particulate filter. Such filters were not available in
World War I. but have since boon developed and are
standard equipment of all nations.
Diphenylchlorarsiue (DA) was introduced by the
Germans in 1917 as a mask-breaking irritant which
was expected to produce temporary casualties and to
cause troops to unmask and expose themselves to the
effects of more lethal agents, usually employed simul-
taneously. In 191S diphenylcyanoarsine (DC’) was
used for the same purpose.
The Allies claimed that these agents were not very
effective but wen1 inclined to attribute their lack of
success in part to the German method of dispersal of
the agents.*20 The Americans produced a new respira-
tory irritant diphenylaminechlorarsine (DM), usu-
ally called adamsite after its discoverer. Dr. Roger
Adams. DM was not used in World War 1. but was
adopted by the United States as their standard irri-
tant smoke and has Iieen found lobe useful in riot con-
trol, since only temporary casualties are produced.
During World War I. the method of dispersing the
irritant smokes was in artillery shell employing a
large burster charge. The particles obtained by this
method are too large to obtain maximal effect from
the agent. The method now used consists of volatil-
ization of the arsenical in a cloud of hot gas produced
in a t hermal generator. The hot arsenical vapor con-
denses to a cloud of very minute particles on contact
with the air.
The Japanese used irritant smoke candles on a
number of occasions against both (’hinese and Ameri-
can troops, although the attacks were always on a
small scale and were apparently undertaken by group
commanders without the sanction of the high com-
mand.
1 hivsiolootcal Action
The physiological action of the irritant smokes has
been summarized in the open literature.320 The effects
of exposure consist of severe irritation to the nose
and throat resembling that from a heavy cold. There
is much sneezing, watering of the eyes, and flow of
mucous from the nose. Headache, pain in the ears
and gums, and nausea are frequently encountered.
A feeling of depression often accompanies exposure
to the arsenical smokes and is thought to lx* largely
psychological in origin.
Toxicity
The /.(fUso’s by inhalation of DA, DC, and DM
have not been determined with very great accuracy
for many species, hut figures quoted*1* lead to the
conclusion that the L{Ct)„0 for man would lx1 greater
than 10 mg min 1. Since the particulate clouds are
nonpersistent. death from inhalation of the arsenical
smokes could be expected only under very unusual
circumstances.
British experiments247 have indicated that, men
exposed to relatively high concentrations of DC
(0.00265 mg I) for periods of 15 to o() seconds and re-
exposnl to the same concentration four times at half-
hourly intervals, do not show any cumulative toxic
effect but rather develop some tolerance to the agent.
A numlxT of compounds were examined by the
British 273without revealing any more
effective than DC, DA, and DM. From the effect of
DC on rabbit eyes it has been concluded that a drop
of <0.1 mm in diameter would probably not cause
permanent damage to the human eye, but that
greater contamination than this might exert a gross
caustic effect which if untreated might lead to total
loss of the eye.*70
Assessment of Military N am e
The particulate filter of modem gas masks affords
adequate protection against the irritant smokes.
Such agents would only be of value, therefore, if they
could lie used for surprise effect before the men could
mask.
Acting on the suspicion (hat the effects of exposure
to the irritant smokes was largely psychological, the
British 251 carried out important experiments in 1942
in which a group of troops exposed to DA and a con-
trol group exposed to a harmless smoke were put,
through an assault course test. The troops were
strongly motivated to turn in a good performance
and were unaware of the fact that there was any dif-
ference in the two types of smoke. The group exposed
to DA remained in a cloud for 2 minutes or until the
generator had burned out, and the mean concentra-
tion of DA was 0.1)2(»6 mg 1. The results showed that
the performance of about two-thirds of the group
exposed to DA was hardly affected at all, that the
performances of the remaining one-third over the
assault course was definitely slower than the control
group, and that about 7 percent of the men exposed
to DA were unable to complete the assault course.
In view of the proximity of the men to the DA gen-
erator and the length of exposure, it was concluded
that the dosage of DA was greater than could lie ex-
SECKET 114
ARSEMCAI.S
pected in the field Further experiment led to the
following conclusions:
1. The effect on fresh troops of DA in concentra-
tions practicable under active service conditions is
almost negligible, apart from the fact that one man
in ten would have great difficulty in keeping on his
gas mask w hen doing heavy work.
2. Experiments on tired troops suggest that con-
centrations of DA practicable under active service
conditions give an average effect which is equivalent
to making fresh troops wear their gas masks.
3. Hence, DA is not an effective weapon even
when used against tired men.
In view of these findings ami the failure to dis-
cover an arsenical smoke significantly more effective
than DA. DC, and DM, little interest remains in the
irritant smokes as chemical warfare agents.
T.t TABULATION OK \USENIC\LS
EXAMINED \S CAN DID \TE
CHEMICAL \\ \RK\RE AGENTS
Table 9 comprises as complete as possible a tab-
ulation of arsenical compounds that have been
examined as candidate chemical warfare agents. Ref-
erences to synthesis, physical properties, and tox-
icological screening data are included.
SECRET Chapter 8
ALIPHATIC MT ROSOC VRBAMATES AND RELATED COMPOUNDS"
Marshall (Inks and liirdscy Ren show
8.1 INTRODUCTION
AsrnvKV conducted by the National Defense
Research Committee [NDRC] revealed that
ethyl X-methvl-X-nitrosoearbamate (“nitrosometh-
ylurethane”) was one of the most disagreeable and
toxic commercially available compounds which had
not received careful study in connection with chemi-
cal warfare.2 Although it proved to I>e insufficiently
toxic to compete with standard agents, synthesis
and assay of related compounds revealed a number
of highly toxic substances. 1'he most promising of
these was methyl N-(0-chloroethyl)-X-nitrosocarba-
matc (KB-Ki).
KB-10 is a persistent agent with a volatility only
slightly less than that of mustard gas (11). Its syn-
thesis, although more involved than that of H. of
lewisite (b), or of the nitrogen mustards, presents no
great difficulty and the required starting materials
are readily available.
KB-10 came under investigation in 1942 at a time
when the nitrogen mustards were being seriously
considered. It was quickly shown that the compound
possesses some of the desirable characteristics of
me thy I -b is (/3-c h 1 oroe (by 1) a n lin e (HX2) — low freez-
ing point, lack of pronounced odor, and effectiveness
as an eye-injurant at low dosages. Interest was also
aroused by the finding that, for mice it is three times
as toxic as H.
Subsequent investigations revealed that: (1) KB-16
possesses inadequate storage stability, and no satis-
factory stabilizer has been found in spite of intensive
search; (2) its eye-injuring potency is not of a differ-
ent order from that of II or b/s(/3-ch loroethy I) amine
(HXd); (3) although more toxic than II and the
nitrogen mustards for mice, it is not so toxic as these
substances for larger species (i.e„ dogs, goats, and
monkeys); and (4) as a vesicant it is markedly in-
ferior to H. Taking these and other findings into ac-
count, assessment of the merits and limitations of
KB-16 led to the conclusion that it does not possess
the.general utility of the standard agent, H, or of the
potentially available nitrogen mustard, IIX3. Ac-
cordingly, KB-1G is not now seriously considered for
use in chemical warfare.
8.2 SYNTHESIS VND PKOPKKT1KS
8.2.1 Synthesis
'1'he aliphatic nitrosocarbamates tested during
World War II (see Table 1) wore prepared by nitro-
sation of the corresponding carbamates, which in
turn were derived from the action of alkyl chloro-
formates on amines. The synthesis of methyl N-(/J-
ch1oroethyI)-X-nitrosocarbamate (KB-10), the only
member of the series that has received detailed study,
involves the following steps.
1. Preparation of N-(0-chloroethyl)earbamate.
Thionyl chloride is allowed to react with ethanol-
amine hydrochloride to produce /3-ehloroethylamino
hydrochloride, which is then treated with methyl
chloroforrnate. Alternatively, methyl chloroformate
can be treated with ethanolamine and the resulting
methyl N-(/8-hydroxyethyl)earbamate converted to
the desired product by the use of thionyl chloride.
The first of these alternatives is preferable (see be-
low). Attempts to prepare methyl N-(0-chloroethy 1 )-
carbamate directly by the action of methyl chloro-
formate on ethyleneimine have not succeeded.4*
H(X,H.CHiNH2HCl + son3—>
arilsCHjNH* HC1
+
ClCOOCHj
I (la)
CnCH,CH,NHCOOCH,
HO('II3CH*NII, + C1COOCII, —>
HOCII2CH*NHCOOCHa
+
sori2 (ib)
I
riCH*CH2NHCOOrHs
Methyl chloroformate may lx* prepared in good
yield either by the addition of methanol to an excess
of liquid phosgene 2 43 or by the reverse addition
of excess gaseous phosgene to methanol.23b The
second alternative gives better yields based on
methanol. An excess of phosgene is required to mini-
• Hascd on information available to NDltC Division 9 as of
November 1, 19-15,
SECRET 116
ALIPHATIC MTKOSOCARBAMATES AND RELATED COMPOUNDS
Table 1. Aliphatic nitrosocarbamates and related compounds examined a.s candidate chemical warfare agents.
The compounds are arranged in four major categories in the following sequence: (1) nitrosocarbamates, (2) nitrosu-
amides, (3) nitrosoamines, and (4) miscellaneous carbamates.
The following abbreviations are used: no', refractive index at / C; earbamate
2
bp,,!
76-78°
2
10
a. Kthvl X-ethyl-X-nilrosoearbamatc
Commercial
lip35
80- 84 ’
10
6. Methyl X-(£t-chhiroet hyl )-X-mtrosoearbamate
2,21a, 23c, 13
flu1'
1.4666
41
10, 41, 14
d,24
1 .3053
II
bp*-*
72 76
2
-- .
vol5"
0.600
11
7. Kthvl X-(d-cliliiroethvl)-X• nitrose>carbainate
2, 21a, 43
bp1"
02-03°
2
10
•
vol111
0.426
11
8. /3-Fluoroethyl X-((Kchloroethyl i-X-nitrosocar-
bamale
21b
bp-
118-121“
2le
10
9. Isopropyl X-(d-chlorocthyl)-X-nitrosocarbamate
2
bp* *
80°
2
10
10. Butyl X-O-chloroethyD-X-nil rosoearbamate
2
bp" 6
95*
2
10
11. Methyl X -(3-1 iromoe I hvl )-X-n it rosocarl m male
2Id, 54c
bp1
110-115“
21 d
10. 14
12. Mcthvl X-(d-hvdroxvetliv 1 )-X-nitrosocarbamate
2
bp" •
00 95“
2
10
13. Methyl X-(3-chloropropyl)-X-nitriKsocarbamate
2
bp* 3 * •*
75-80°
2
10
4 4. Methv l-N-bwtvl* X-n i t rosoca rba mate
2
l'l>-
70-72°
2
10
15. Met hyl X-8-{fl'-ehloroet hyl t hio)-et hyl-X -nil roso-
ca rba male
44
.16. Methyl X-phenethyl-X-nitrosocarbamate
21d
Cannot lie distilled
2 Id
10
AT ilrosoaniides
~ .
17. X-(3-(’hIoroelhvl)-X-nitrosoformamide
2
bp* *
78-80“
2
18. X-(3-Chloroethyl )-X-nitrosoacetamide
2
bp" s
70 72°
2
10
10. X-Met hyl-X-nit rosofluoroacet amide
51
bp11
84°
51
50
.V it rnsoa mines
20. X-Xit rosopiperidine
2
bpu
94-06°
2
10
21. X-Xit rosomorpholine
2
mp
28°
2
10
bp14
105 107°
2
22. X,X '-Dinitrosopiperazine
2
mp
155 157“
2
23. d-Chloroethvlmelhvimtrosoamine
54b
♦ . .
44
24. hist3-Chloroelhvl(nitrosoamine
2
Cannot lie dist died
2
10, 44
25. 4-Methvl-4(methylnitrosoaminoVpcnlanone-2
10
26. X,X '-Dime!hyl-X,X '-dihitroso-p-phenylene-
diaminc
15
nip
149-150°
15
It)
27. X,X '-6/s(3-(bloroethyl)-X,X '-dinitroso-/>-phenyl
enediaminc
15
mp
106.5°
15
10
MisreUancous carbamates
28. Methyl X-fd-cliloroethvlVX-nilrocarbamate
2
bp"*
95-100°
2
10
vol20
0.138
11
20. Methvl X-d-ehloroelhvIca rba mate
2
no50
1.4575
27
10, 4 4
.
bp14
100°
2
30. Kthvl X-isobutvlearbamate
15
bp“
04°
15
10
31. Kthvl X-isoamvlcarbamate
15
no20
1.4333
15
10
•
bp**
109°
15
32. Kthvl X-methoxycarhamate
10
SECRET 117
Tabi.e 1 (Conti mint).
Compound
Reference
to
synthesis
Physical properties
Pro|ierty Reference
Reference to
toxicity
data
33. Mcthv! N-ethylthioIcarbamale
12
«.r’s
1.4078
12
10
50
12
10
dIJ
1.067
12
—
bp-4
109.5-110.5°
12
bp37
110-121.5°
12 —
35. Mi'llivl X-et li vldil liioearbamate
12
«irr
1.6130
12
10
-r-4
1.151
12
:
bp* *
121-122°
12
SYNTHESIS AND PROPERTIES
mize formation of methyl carbonate. Distillation of
the crude methyl ehloroformate is not necessary.49
2. Pivparation of HR-16 hy nitrosat ion of methyl
X-{0-ehloroe(hyl)earbamate. This step may l»e ef-
fected hy nitrous acid in solution or hy nitrous gases
either with or without a solvent. The action of ni-
trous gases on methyl X-(/3-ehloroethvl)carhamate
in the absence of a solvent is the most rapid and
convenient.
— UNO. or
(It IU II XH('(XX H,
nitrous gases
(’1CH,( I I2X (XO)COOCH* (2)
Reaction (la) was employed in the original lal>o-
ratory preparation of KB-16.!*u/9-Chloroethylamine
hydrochloride prepared essentially according to
Ward fil from solid ethanolamine hydrochloride and
thionyl chloride in chloroform was treated as a solid
suspended in ether or benzene with aqueous alkali
and methyl ehloroformate. The resulting methyl
N-(0-el11oroethy 1)carl>amate was purified hy distilla-
tion, diluted with ether or benzene, mixed with a so-
lution of sodium nitrate, and nitrosated by the addi-
tion of nitric or sulfuric acid. Overall yields of 62 per
cent were obtained. Alternatively, N-(/J-chloro-
el hyljcarbamate was nitrosated under anhydrous
conditions by the use of nitrous gases. Flash distilla-
tion was used to purify the final product and appears
to be the only feasible method. The above procedures
were used with little modification for the synthesis of
the first samples investigated in Great Britain.54*
The method is well suited for large-scale runs.
Preparation of KR-16 by the alternative procedure
utilizing reaction (lb) is also convenient for labora-
tory scale work and can be carried out in overall
yields of 65 per cent.4143 It is less readily modified
for use on a larger scale because the hydroxycar-
bamatc must be distilled and the conversion of this
intermediate to the eh lorn compound has not been
achieved in yields greater than To per cent.
The first met hod,-as modified for production on a
larger scale, has been simplified by: (1) elimination
of the isolation of ethanolamine hydrochloride and of
d-chloroethylamine hydrochloride, both of which are
hygroscopic; (2) combination of the hist three steps
into one; (3) reduction of the large excess of thionyl
chloride and sodium nitrite; (4) the use of a single
solvent (chloroform or benzene) in reduced quantity
throughout the reaction steps; and (5) elimination
of all distillations except that of the methyl N-(0-
chloroethyl)carbamate.9 233 49 A brief description of
the modified process follows. Ethanolamine in chloro-
form,49 in benzene,23® or in the absence of a solvent 9
is treated with dry hydrogen chloride to produce
ethanolamine hydrochloride. Thionyl chloride is then
added directly (if no solvent was used in the first
step, benzene is added at this point), and the mixture
is heated to convert the ethanolamine hydrochloride
to 0-chloroethvlamine hydrochloride. The mixture is
then diluted with water, and caustic alkali and
methyl ehloroformate are added. After the acylation
is complete, the organic layer is separated, washed,
dried, and stripped of solvent. The crude methyl
X-(d-ch 1 oroethyI)call>amate thus obtained is then
distilled under diminished pressure. It has been ob-
tained in yields of 77.5 per cent in runs utilizing
24-5 lb of ethanolamine.23®
If nitrosat ion is carried out by slowly acidifying a
mixture of the carbamate in benzene or ether with an
aqueous nitrite solution, the reaction is slow and a
large excess of sodium nitrite is necessary.*-23® When
the reverse addition is used and (he reaction mixture
is strongly acid, a slight excess of sodium nitrite is
sufficient and the reaction proceeds rapidly.4*
SECRET 118
\LIPIIATIC MTROSOCAUB AMAXES VM) RELATED COMPOUNDS
Aqueous nitrosations were used in all investiga-
tions where scaling up the synthesis of KB-16 was
tried, but it was subsequently shown that the re-
action of nitrous gases with methyl 0-chloroethyl-
carbamate in the absence of a solvent is quantitative
and almost instantaneous.21' 2*r This variation pos-
sesses a number of practical advantages.
1. Solvent is eliminated and effective reactor ca-
pacity is thereby increased.
2. The purity of the final product is sufficient to
obviate the need for flash distillation.
3. Equipment is simplifier! and the total time
cycle is reduced.
4. The method should allow preparation of the
agent in inlu shortly before use, or in shell subsequent
to firing. This would eliminate the problem of storage
stability ami greatly lessen the hazards involved in
synthesis.
Preliminary design data and cost estimates for a
plant to produce KB-16 at the rate of 500 tons i>er
month have been submitted. The calculations were
based on the use of aqueous nitrosation in the final
step.*
N-(0-Chloroethy!)-N-nitrosoacetamide, a highly
toxic analog of KB-16, has been preparer! on a labora-
tory scale by nitrosation of an ethereal solution of
N-(/3-ch!oroethyl)acetamide with oxides of nitrogen.
Yields of 60 per cent of material purified by flash
distillation were obtained.*
8.2.2 Physical Properties
KB-16 is usually obtained as an orange-red limpid
oil of limited thermal stability. It is soluble in water
to the extent of 0.7 g 100 g, and is completely misci-
ble with ordinary organic solvents. Although it has
not been obtained in crystalline form, it assumes a
semisolid state at —65 C; at —25 (.' it is a viscous oil.
The density of KB-16 is 1.3053 g ml at 25 C,41 the
refractive index 1.4085 at 25 and the boiling
point 100 (’ at 15 mm, 89 (’ at 6.5 mm, 86 C at
5.5 mm, 82 at 1 mm, and 75 C at 2 mm.43
The volatility of KB-16 is 0.87 mg/I at 25 C,
slightly less than the corresponding value of 0,96
mg 1 for f«s(0-chloroethy 1) sulfide (H). Several de-
terminations of the volatility (or vapor pressure) as
a function of temperature have been made.2" 53 The
vapor pressure at temperatures in the range of inter-
est for chemical warfare is given by the following
equation:11
log p (mm Hg) = 8.91282 — —
The standard free energy of formation of KIM6
and several thermodynamic constants of the inter-
mediate carbamate have been calculated from the
results of a series of calorimetric and equilibrium
studies.7
Chemical Properties
KB-16 decomposes within 48 hours in water or in
aqueous bicarbonate solutions.’ In the former case
about 40 per cent of the nitrogen appears as nitric
acid, the remainder disappearing from the reaction
mixture. Only 5 per cent of the chlorine appears as
chloride ion. In bicarbonate solution more than 00
per cent of the nitrogen is lost, presumably .as nitro-
gen gas, and carbon dioxide and methanol are pro-
duced. About 80 (x-r cent of the chlorine appears as
chloride ion; the remainder is bound to carbon, pre-
sumably in the form of ethylene chlorohydrin. The
production of chloride ion at pH 8 is not significantly
altered in the presence of substances which react
with the 0-chloroethyl groups of (he sulfur and nitro-
gen mustards. The decomposition of ethyl N-(/J-chIo-
roethyl)-N-nitmsocarbamatc in aqueous solutions is
similar to that of KB-16.*
One of the most characteristic reactions of KB-16
is its rapid and complete decomposition with evolu-
tion of nitrogen when treated with alcoholic ammonia
or primary aliphatic amines.* Solutions of ammonia
or ethanolamine in ethylene glycol have therefore
been recommended as personal or laboratory decon-
taminants.2 However, the distinct possibility that
substances of the nitrogen mustard type may be
formed by this reaction should lx* considered in the
choice of a decontaminant (see below). Secondary
amines and primary aromatic amines react relatively
slowly with KB-16.
In aqueous solutions, the reaction with ammonia
and with primary amines is slower, perhaps because
of the low solubility of the nitrosocarbamate. At
pH 8, the main reaction with primary amino groups
is carboalkoxylation, as has been demonstrated by
the isolation of methyl and ethyl N-benzylcarba-
mates as products of the reaction of bcnzylamine
with methyl and ethyl N-(0-chIoroethyl)-N-nitroso-
carbamates, respectively.* Secondary amines (di-
ethanolamine) are also carboalkoxylated.
In ethereal solution, reaction with bcnzylamine
leads to methyl N-benzylcarbamate and N,N'-di-
benzylethylenediamine, (he latter probably arising
through bcnzyb/S-chloroethylamine its an inter-
mediate.’
SECRET SYNTHESIS VM) PROPERTIES
1 10
The amino groups of a-amino acids also react with
KB-16, but the reaction is more sluggish than those
of primary aliphatic amines and does not appear to
go to completion. With cysteine, the reaction pro-
ceeds along several lines; both amino and sulfhydryl
groups disappear. 6/s-S-(Cys(einyl)ethane, probably
arising through the intermediate S-(d-chloroethyl)-
cysteine, has been isolated as a product of this re-
action.* Nitrosomethylurethane also carbethoxylates
the amino group of cysteine, but is far less active
toward the sulfhydryl group than is KB-16.*
In solutions containing egg albumin KB-16 reacts
/3-chloroethyl group into amines and into the sulf-
liydryl compounds. The intermediate products are of
the sulfur and nitrogen mustard type, and undergo
further reactions characteristic of these substances
(see Chapter 19).
With regard to loci of action in tissues, it may be
noted that reactions of the sulfur and nitrogen mus-
tards involve a process of thermal solvolytic activa-
tion in water (see Chapter 20). On the other hand,
the alkylation of benzylamine by KB-16 in ether so-
lution demonstrates that this agent need not lie so
activated. As a result, it is possible that KB-16 can
ru' h. ■('H;- \( n o) • co • or n>
i ~~ ' n
+ II:0 at /(II 8 + RNH, i + HS■ CH2 -('II(NI!;)• COOII
\ i i i r ]
CK H: CHj\H + IK)CO (M il, ClCIIrCIIrMI + R-NHCOOCHa + HSCIUCIUCOOH
I ! II-
NO ~ NO NO NH-COtX'Hj
- H.O - H.O ~ - H-0
t T 11
CICHjCHN, CX), + IKK'Il, CICH-CIIN. ClCHjCHN,
' - ” k
+ 11,0 + R-NH- | + TIS.CIl,.CH(NH,) COOII
ClCH, CH,.OII + N, C1CH,.CH. NH-H + N, CICI1,.CH2 S CII2 -f N,
+ R-NH, 4- HS C'H, CH(NII,) COOII
RNH CH, CH, NH R + HC1 COOH ('OOII
CH-CH. S-CHj-CIIj-S.CH., i'll + MCI
xh, kn,
slowly with the liberation of 1 mole of nitrogen per
mole of nitrosocarbamate but no decrease in the
amino nitrogen content of the protein occurs.*2S The
reaction with hemoglobin is rapid 25 and in this case
some amino groups disappear, possibly by carbo-
methoxylation, although it has not been possible to
ascertain the mode of reaction.*
The following scheme of reaction, reminiscent
of those of Klobbie &s and V. Pechmatin M for the
breakdown of nitrosomethyl- and nitrosoethylur-
ethanes. has been proposed to explain many of the
observed reactions of KB-16.* Thus KB-16 can be-
have as a chloroalkylating agent, introducing the
react in fatly phases of tissues, whereas reactions of
the nitrogen mustards in these loci are not equally
probable.*
i!.2.V Detection and Analysis
KB-16 reacts with the 1)11-3 reagent to produce
the characteristic blue color. Samples as small as
25 //g may be detected by use of the DB-3 tube of tin*
United States Army M-9 Detector Kit according to
the standard procedure. By heating to 200 C, the
sensitivity of the tube can be increased sufficiently
to permit the recognition of I Mg.'*’* in the absence of
a guard tube, the spotted dick test of the British
SECRET ALIPHATIC MTROSOC \RB A \1 AXES AM) RELATED COMPOLNDS
Vapor Detector Kit gives an overall blue color.41
Acidified iodoplatinate paper is bleached by the
vapor of the agent. Other procedures for detection
involve the use of the diethylamine and diphenyl-
benzidine reagents or the Liebermann reaction.
Positive reactions given by the decomposition
products of KB-16 limit the usefulness ot these
methods.
The most useful method for the analysis of KH-lti
depends upon the quantitative evolution of nitrogen
which occurs when the compound is treated with
primary amines ,6i **r or with alcoholic alkali.41 This
method is suitable for use as an assay method or for
analysis of samples collected in chamber or field
tests, anil has the advantage of specificity to the ex-
tent that it measures only the nitrosated material.
The Griess reagent as used for nitrites can be em-
ployed for the field or chandler analysis of this
agent.4*
A more detailed discussion of the detection and
analysis of the nitrosocarbamates will be found in
Chapter 34.
8.2.5 Stability
KB-16 and its homologous esters are thermally un-
stable. Decomposition with gas evolution occurs at
rates which make storage impractical.2-41 In steel
containers with 25 per cent void, the pressure in-
crease per day is appreciable at temperatures as low
as 4 C and amounts to about 2 psi at room tempera-
ture.3 At high temperatures the decomposition be-
comes even more rapid, the pressure increase in glass
with 50 per cent void amounting to about 4.5 psi per
day at (50 C.41 No significant difference between the
rates of pressure development in glass and steel con-
tainers has lieen observed (unpublished data), even
though steel appears to be attacked.41 The purity of
(he sample has a considerable effect on the rate of
thermal deeomposition, carefully purified material
decomposing at a lower rate than crude material.2*11
The stability of preparations made by nit-rogation
with nitrous gases is as good as or better than that of
flash-distilled samples. Decomposition is accelerated
by acidic and phenolic substances and by zinc and
magnesium oxides.2 41 There is disagreement as to
the effect of traces of water, weak bases, or contact
with metals other than steel.2-41
The gas produced during !Ik- decomposition of
KB-16 consists principally of nitrogen; oxides of
nitrogen, carbon dioxide, and hydrogen chloride
have also been identified.I*h 2*c-41
In spite of intensive searches for a stabilizer to
prevent or minimize the spontaneous decomposition
of KH-16, little success has been achieved. Few of the
tested substances were of any value and none pro-
duced a pronounced increase in storage stability.
The tested compounds include organic and inorganic
bases, acids and derivatives of acids, hydroxy and
mercaptan derivatives, oxidizing agents, inert liquids
and solids, salts and complexes of heavy metals, and
numerous miscellaneous compounds.2*"41
Few reliable data are available on the stability of
KH-16 to detonation in munitions.18"1 30 31 11 The re-
sults of a field trial with 105-mm shell supply no
definitive information.3" In a small chamber, detona-
tion of a 75-mm shell charged KH-lti resulted in
more or less complete destruction of the agent;w in
similar tests ethyl-6fx(/3-ehlonx‘thyl)amine (HNl)
was also destroyed but /n«{0-chloroc(hyl)amine
(HNS) was not. It may lx* noted that (he conditions
of these tests were more severe than would 1m> en-
countered in the field, and that HNl can effectively
lx* dispersed from M47A2 bombs.-' KH-lti is not de-
stroyed to any great extent by the milder explosions
that occur when it is dispersed from glass bottles
either in the field by means of a standard detonator 31
or in a 2-cu m chamber by means of a blasting cap or
detonator.18"1 31
K.2.6 Decontamination
Rapid surveys of the reactions of KH-lti, with
emphasis on reactions of possible use in decontamina-
tion. have been carried out both in this country and
in Great Britain.20 47 The reagents examined included
bleaching powder, chloramides, a number of inor-
ganic salts in solution, mineral and organic bases, at
least one strong oxidizing agent, and reducing agents.
Solid bleach or lime slurry would appear to be suit-
able for field use. Caustic soda or alcoholic ammonia
has been recommended for laboratory use, and aque-
ous ethanolamine for personal use. In line with these
recommendations, groups concerned with the synthe-
sis of the agent have used solutions of ammonia or
ethanolamine in alcohol or ethylene glycol for per-
sonal, laboratory, and pilot plant decontamina-
tion.32,11 As stated above, ammonia or primary
amines should be used with caution because of the
possibility of producing toxic intermediates.*
For treatment of eyes contaminated with KH-lti,
mild alkalies and reducing agents (e.g., HAL) should
be more effecthe than in the case of nitrogen mus-
tards.2*-47
SECRET CHEMICAL STRICTURE IN RELATION TO TOXICITY
121
8.2.7 Protection
The canisters of modern gas masks afford ade-
quate protection against the vapor KB-16.13 41 For
details the reader is also referred to the Summary
Technical Report of NDRC Division 10.
Tlie chloramidc impregnation of clothing would
appear to offer little resistance to KB-16. because
(his agent fails to react with chloramine-T or with
various impregnites.2" It may lx4 assumed that cloth-
ing containing activated carl ion would effectively
exclude the vapor of the agent.
a..i CHEMICAL STRICTURE IN RE
LATION TO TOXICITY
In Table 2 are presented data on the toxicity for
mice of compounds in which the N-nitroso, N-(/J-
Table 2. Toxicities of X-subst it uted aliphatic carbamates for mice.
Most of the data arc taken from reference If). The mice were observed for 10-15 days after exposure for 10
the stated nominal concentration. In the case of one compound, methyl N-methyl-N-nitrosoearbamatc, the
obtained from reference 48 ami relate to rats exposed for 30 minutes.
minutes to
data were
Mortality
Structural
Nominal cone.
for 10-min
('ompound
formula
(mg 1)
The prototype Compound
Methyl N-(3-chloroethy 1 )-X-nitrosocarbamate (KB-16)
(’ICIT/H, o
\ !!
—- -
-
x—r—och,
0.03(5
/^’in
Effect of replacement of the \~nilroxo group
—~
Methyl X-(d-chloroethvl)carbamate
cich.ch, o
_
—
\ 1!
X—C -OCH,
1.0
0 20
11
Methyl X-(d-chloroethyl)-X-nit rosocarbamate
nciijCH, o
\ 1!
-
N—C—OCH,
/
1.3
0 20
OtN
Ethvl X-{0-chloroethyl)-X-nitro8ocarbumate
C1CH,CH, O
V 11 . .
N—C—OTjHi
0.075
—
Ethyl X,X-f>/s(/t-chloroethyl)carlwmate
ricHjCii, o
\ 1!
'
X C—OC,H,
/
0.4
0 40
CICHiCHi
X-(3-chli iri«-thvl)-X-n it rosoaeet amide
CICHjCH, O
—
\J--nu
on/
O.Olfi
Li ’fcO
— -.
X, X -bix( /3-ch loroet hy 1 facet a midc
CICHjCH* O
\ !! -
X—6—CH,
0.5
0/10
/
CICHjt'H,
Effect of replacement of the \ -{ff-chloroelh yl) group
—
Methyl X-met by 1-X-nil rosocarbamate
(II, O
\ . !!
N—C OCH,
0.20 (30 min)
!4 (rats)
1
X
\
0.13 (30 min )
34 (rats)
Methyl X-(d-bromoethyl)-X-nit rosocarbamate
BK'HjCH, O
\ li
X C -OCH,
0.2
0/10
'
/
ON
0.82
10/20
SECRET 122
ALIPHATIC MTHOSOCAKBAMATKS AM) BELATED COMPOUNDS
Table 2 (Continued).
Mortality
Structural Xominal cone.
for lO-iniu
Compound
formula
(niR 1)
exposure
Mcthvl X-(/3-hvdroxvcthvl)-X-nitn>socarbamatc
HOCHCH.
o
\
II
X C OCH,
0.3
0/20
OX
Methyl X-(d-chloropropyl)-X-nitTOsocarbamatc
CIIjCHCICHi
()
— \
II
X—C—OCH,
0.3
0/10
OX
Mi thy 1 X-butyl-X-uitroeocarbamnte
CIliCIIjCH-CH*
O
■'
”
y
NT—C OCH,
0.3
1/20
/
OX
Methyl X-phcnethyl-X-nitrosooarbamatc
(VIC CH,CII,
o
-\
It
/
OX
X ('—OCH,
0.97
0 20
Kffrrl of rtplocrmrnl of the mi lhuxy group
Ethyl X-d-chloroethy 1-X-nil rosoearbanmtc
CK’TIjCHi
0
II
X
-C—OCH2C-Hi
0.07a
IJ J}
-
/
OX
-
(3-Fluorocthvl X-(^-chlorcx'thvl)-X-nitrosoc!irbaniate
CK'HjCHj
O
0.1
11/15
\
11
0.2
11 20
X -C OCHjCIIjF
0.5
10 20
—■/
OX
Isopropyl X-(P-ehloroethvl)-X-nit rosoearbanmtc
CK'ITjCHj
o
0,1
0 20
\
\
II
0.12
20/20
X
- C—OCH(CHj),
0.2
16/18
■
/
OX
Butyl X-(/3-ehloroctbyl)-X-nit rosocarbamate
cicH.cn,
O
\
11
X
-C OC.1E
0.10
U'm
’ '• - ■_
/
ON
0.41-
LC'*
X-0-<-hloroethyl)-X-nit rosoai-ct amide
CICHjCHj
0
\
II
X-€ -CHj
0,040
—
/
OX
chloroethyl), and methoxy groups of KB-16 are re-
placed by other substituents. On (he basis of these
data, and subject to their limitations, the following
conclusions can be "drawn.
1. The N-nitroso group is essential for high toxic-
ity. Its replacement by another group has always re-
sulted in at least a 30-fold reduction in toxic potency.
2. The N-(/3-chloroethyl) group is essential for
highest toxicity, but moderate toxicity is possessed
by some compounds in which it is replaced by an
alkyl group (e.g., ethyl N-methyl-K-nitrosocarba-
mate apparently possesses one-tenth the potency of
ethyl N-(jJ-chloroethyl)-N-nitrosocarbamate)..
3. The methoxy group, although optimal, is not
essential for high toxicity. Its replacement by a
methyl group (to form N-(/3-chIoroethyl)-N-nitro-
soacetamide) results in an insignificant decrease in
potency, and the ethoxy analog is about one-half as
toxic as KB-16.
Toxicity data for other species, vesicancy tests,
and determinations of eye-injuring potency arc not
sufficiently complete to permit analyses of the rela-
tive potencies of members of this series. Such data
as are available indicate the relative superiority of
KB-16 and are not inconsistent with the other con-
clusions drawn from (he toxicitv tests with mice.
SECRET 123
TOXICOLOGY
Compounds which possess a fluoroacetate-Hke toxic-
()
H
ity by virtue of the presence of an FCH-C— group
are an exception to this generalization (see Chap-
ter 10).
8.4 TOXICOLOGY
Of the following toxicological sections, those on
detectability by odor anti sensory irritation, vesi-
ca ncy, and eye-injuring action bear most directly on
the evaluation of KB-16 and related compounds as
chemical warfare agents.
8.4.1 Detectability by Odor and
Sensory Irritation
The vapor of KB-16 has a pleasant odor, some-
times described'as sweet or fruity, which can be de-
fected by smell only at concentrations several times
greater than those required for H. Men exposed to
relat ively high concent rations (i.e., 70 jug I nominal)
for 30 seconds detect the odor but experience no
sensory irritation,41 and concentrations as high as
0.2 41.4 mg 1 elicit no signs of irritation in animals.1*1’
Those properties of KB-16 vapor, considered in rela-
tion to its eye-injuring potency and the delayed on-
set of the injuries caused by casualty-producing
dosages (see the following section), confer upon it
some insidiousness. However, in this regard it is not
notably superior to some of the nitrogen mustards
(e.g., HNS), which are probably less easily detected
by odor and not notably inferior in eye-injuring
potency (see Chapter 6).
Laboratory determinations of the median de-
tectable concentrations in jug/1 of KB-16, II,
ethyl-6w03-chlorocthyl)amine (HNl), and HNS arc
tabulated below. Attention is directed primarily to
the relative values, inasmuch as the absolute values
arc not necessarily of significance for field conditions.
Agent Mg/1 Reference
II Plant run Levinstein 0.6 34
Pure thiodiglycol 1.8 35
KB-16 _ 7 ± 16 1,41
HNl Plant run 13 34
Pure _ 17 33
1IN3 Plant run 15± 37
The vapor of N-(/3-ehloroethyl)-N-nitrosoaeeta-
mide does not possess the insidiousness of KB-16.16f l?i
8.4.2 Toxicity
Inhalation- Toxicity
In Table 3 data on the toxicity of KB-16 vapor
Tabi.e 3. Inhalation toxicity of Kli-16 in comparison
with mustard gas (H) and /mO-chlorocthyl)aminc (11X3).
Approximate nominal L('.„, in mg 1 for 10-tnin
exposure and 15-day ohservalion [icriod.*
S|iecics
KB-16 11 MX3
Mouse
0.(130 (259)t41« 0.12"*1,1 0.12§ls‘
Hal
Guinea pig
Rabbit
Gat
Dog
Goal
Monkey
0.1-0.2 (19)"“> 0.1“'.•» o.2
0.035 (60)i
0.2 ± (17),c,‘ 0.21*1 >0.216*
>0.2 (7)*0.281® 0.14'“
0.1-0.2 (11 )'«'■•* 0.07'*' 0.08,4k o
0.1 (H)i«'*.i.ud 0.07161 o.l">k‘*“
0.3 (6)'T1 0.19t«
0.2-0.5 (6),Tt* 0.08"4’ .... ■
* In the ease of KB*16 some deaths occurred among the larger «|M*eit's
after observation periods as long as 15 30 days and were includcii in inti-
mating the Lf.’se’a.
t The figures in parenthesis give the nutnlier of animals U|»oii whieh the
estimated LfW« are based.
X Analytically determined concentration.
§ The analytical LCh* is about 0.055 mg 1.
for various animal species arc set forth in comparison
with corresponding data for 11 and HN3. One of the
early observations arousing interest in KB-16 was
the discovery that for mice it is several times more
toxic than H. When tests were made with other se-
cies, however, no such differential in favor of KB-Hi
was found, except possibly in the ease of the rat. In-
deed, it may be questioned whether KB-16 is as toxic
as H for (he larger mammalian species which have
been studied.
The only evidence bearing on the relation of tox-
icity of KB-16. to exposure time is the demonstration
that the L{Ct)io for mice is approximately the same
for 30-minute exposures as for 10-minute expo-
sures, IBhand the result of a single experiment in which
a dog succumbed 23 days after exposure to a total
vapor dosage of approximately 1,100 mg min/m*
administered during three 8-hour periods on suc-
cessive days.17,1
During exposure to KB-16 vapor at concentra-
tions as high as 0.2 0.4 rng/1, animals exhibit no
signs of discomfort or irritation.161’ Concentrations
considerably in excess of the 10-minnte LC„o s occa-
sionally caused closing of the eves, but the irritation
was mild and not accompanied by profuse lam-
ination.
The development of symptoms after gassing with
KB-16 is usually delayed for 12-24 hours and follows
the same general pattern in different speeies.41*,’-nd
Respiratory distress becomes prominent. The ani-
mals appear depressed and stop taking food and
water; as a consequence weight loss may be precipi-
SECRET 121
ALIPHATIC MTKOSOCARUAAtATKS AM) RELATED COMPOUNDS
(ous. Severe eye injuries also develop (see Sec-
tion 8.4.4). In nonfalal eases the symptoms slowly
subside. In fatal cases respiration may become la-
bored and terminate in death after 3-10 days, or the
animals may slowly waste away and die as late as
3-4 weeks after exposure.
The principal pathological changes occurring in
animals gassed with KB-16 are found in the eyes
(see Section 8.4.4) and the respiratory tract.4
In mice the most severe changes are confined to the
nasal and nasolaryngeal mucosa, and an exudate,
first fluid and later mucopurulent, is often produced
in sufficient amount to block the air passages. The
trachea and bronchi show much less damage. The
lungs may become hyperemic but pulmonary edema
is minimal and pneumonia does not ordinarily de-
velop. In larger species, perhaps liecause the larger
size of the air passages permits further penetration
of the vapor, nasal injury is accompanied by severe
pathological changes in the deeper parts of the respir-
atory system. The larynx, trachea, and bronchi are
severely involved, and pulmonary injury with con-
solidation occurs. The pneumonia often appears in
focal patches around the bronchi. Degenerative ma-
terial cast off from the larger respiratory tubes often
blocks some of the larger and smaller bronchial pas-
sages.
In general the bone marrow is little affected.4 6
Atrophy of the lymphoid organs with rhoxis of the
lymphocytes of the thymus gland and the splenic
follicles has been reported in some species ,7g but was
not found to he conspicuous in another investiga-
tion.4 Hematological studies fail to reveal the con-
spicuous changes in numbers of circulating white
blood cells which characterize severe intoxication
with the sulfur and nitrogen mustards, although a
rise followed by a fall in lymphocyte count has been
reported in mice exposed to about eight L(Cl)M
dosages of KB-16 vapor.17* In some instances limited
degenerative changes, possibly secondary to respira-
tory embarrassment, have been observed in the liver
and kidney. In mice and rats the digestive tract from
upper esophagus to anus is often markedly distended
with gas but no hemorrhages or perforations have
been observed;4 the distention is probably caused
by swallowing of air, which occurs because of mouth
breathing and difficulty in respiration.
The available data suggest that the principal
pathological effects of ethyl N-(/J-chloroethyl)-N-
nitrosocarbamate and of N-(0-ehloroe(hyI)-N-nitro-
soaeetamide are similar to those of KB-16.®,®1*1"
Toxicity through the Skin
In its actions on and through the skin, KB-16 is
relatively ineffective as a lethal agent when com-
pared with H, HNl, or HNS. In spite of the high
sensitivity of mice to the inhaled vapor, exposure of
only the bodies of animals of this species to KB-16
vapor at a nominal dosage of 6,900 mg min m3 (/ =
10 min) caused no deaths within 15 days; 11,300 mg
min m* killed 1 5 unshaved mice; and 13,000 mg
min m3 killed 6 0 mice with shaved backs.16,1 For
other agents -approximate nominal L(Cl)b0’s {I =
10.min) for mice upon body only exposuiv are: H,
1.000 mg min nv1; HNl, 5,000 mg min m3; HNS,
2.000 mg min m* (analytical value — 1,000); and
b. 2.100 mg min m*.u The toxicity of liquid KB-16
applied to the shaved skin of mice is also low when
inhalation of vapor is minimized. Necrosis, of the
skin and ulcer formation occur at the site of appli-
cation but a minimum value for the /,/>j0 is believed
to be 62 mg kg.4 Corresponding values for other
agents are: II, 92 mg kg; HNl, 13 mg kg; HNS, 7
20 mg kg (see Chapter 22). It may be noted that all
of these values are high in comparison with the per-
cutaneous /.Dio’s for some of the compounds con-
sidered in Chapter 9.
Toxicity by Injection
Parenteral injections, although they have no direct
bearing on chemical warfare, supply useful informa-
tion concerning the toxicological properties of KB-16.
//tin's upon intravenous injection are: mouse, 0.45
mg kg; rat, 1.1 mg kg; and rabbit, approximately
2.0 mg kg.4 The subcutaneous LD,n for the mouse is
9.0 mg/kg;4 and those for the rat and rabbit ap-
proximately 8 and 20 mg kg, respectively.41 Even
large doses are without immediate pharmacological
effects, and subsequent developments reveal no neu-
rological injury, central nervous or gastrointestinal
action, pronounced leueopenic action, or significant
changes in the total number of circulating white
blood cells.4 The conspicuous pathological changes
occur in the lungs, which become distended, moist,
and hyperemic. They sink in water and on cutting a
pinkish, foamy fluid runs from the lungs and trachea.
A small amount of pleural fluid accumulates. The
heart is often dilated but gross pathological changes
elsewhere are conspicuously absent. Venous con-
gestion of the liver and myocardial injury with focal
necroses are occasionally but not constantly ob-
served. The thymus gland and spleen are usually but
not markedly decreased in size — probably (he re-
8 EGRET TOXICOLOGY
125
suit of a nonspecific lymphoid involution. In some
instances (e.g., in rabbits receiving large doses) there
is evidence of lymphocytic fragmentation in the
spleen, lymph nodes, and thymus. The bone marrow
usually appears normal, although evidence of leueo-
blastic stimulation apjiears in some rabbits.4 It has
been reported that one of two dogs receiving 18
mg kg intravenously died in 4 days with aplasia of
the bone marrow and drastic leucopenia, involving
both lymphocytes and granulocytes,* It levs been
concluded that intravenously injected KB-16 causes
death by producing fatal pulmonary edema, which
develops slowly over a jieriod of 2 8 days.4 5
Injections by various routes demonstrate that
KB-16 reacts with lilx-ration of gas (presumably X2)
in the first capillary bed it reaches.4 Circulatory
stasis may occur, in some eases possibly because of
-vessel spasm or thrombosis, so (hat contact w ith the
tissue may lie. prolonged. These observations give an
explanation for the finding that the principal patho-
logical changes following inhalation or intravenous
injection occur in the lungs. The liberation of gas,
which occurs in the tissues to which injected KB-16
is first carried and which also occurs when KB-16 is
added to tissue suspensions, undoubtedly contributes
ischemic injury to the chemical injury produced by
the direct reactions of KB-16 with tissue compo-
nents. The liberation of gas following inhalation of
KB-16 vapor is presumably not sufficient to be sig-
nificant.
Toxicity by Mouth
KB-16 is moderately toxic when administered by
stomach tulie. The hi)-,o’s are in the order of 20
mg kg for the rat and 15 mg kg for the rabbit.41 The
substance is immediately irritating, as evidenced by
vomiting in dogs, and it produces severe esophageal
and gastric damage.17® In the rat, vesicles similar to
those produced by lewisite oxide have been found in
the stomach at autopsy.41 Lung pathology has been
observed in some cases,7* and the absorption of
KB-16 from the gastrointestinal tract has been
demonstrated by the appearance of gas bubbles in
the hepatic portal vein.4 Death occurs after from
one-half day to many days and is preceded by pro-
nounced weight loss when survival is prolonged.17e 41
That KB-16 presents some hazards as a water con-
taminant is demonstrated by the virtually 100 per
cent mortality of mice, rats, and dogs whose supply
of drinking water was contaminated with 0.5 1.0
mg ml.17' Only few deaths occurred when the water
was contaminated with 0,1 mg ml. In most of the
experiments the contaminated water was freshly
prepared each day. In spite of the fact that aqueous
solutions of KB-16 decompose within 48 hours (see
Section 8.2.3), mice whose drinking water was con-
taminated with 1.0 mg ml of KB-16 at the beginning
of one experiment died almost as quickly as those
whose contaminated water supply was freshly pre-
pared at daily intervals.
K.t.3 Vesicant Action
In comparison with H, the vesieancy of KB-16 is
of a low order 11 41 46 and screening tests indicate that
none of the related compounder is more potent.14 A
direct comparison of “absolute vesieancies,” de-
termined by application of agents diluted with ben-
zene and covered to prevent evaporation, reveals
that KB-16 is about 1 ;1 to 1 4 as potent as II.41 As
shown in Table 4, small doses of liquid KB-16 applied
For
I. arc
made
of the
of 63
Table 4. Vcsicancy of K B-16.14
the sake of comparison, data obtained with H and
included. All applications of the vesicants were
during winter weather (January 1943) to the skin
forearms of human subjects at room temperatures
72 F and relative humidities of 14-37 per cent.
Days after
Dose
applica-
Agent
(mb)
tion Erythemas Blisters
KB-If)
200
2 7/28 (4 mm) 0/28
7 7/9 ... 1/9 (2 mm)
II
65
2 112/119 (7 mini 70/119 (5 mm)
I-
95
2 285/290 (8*mm) 279/290 (6 mm)
to tlie skin in the usual way (i.e., undiluted and with
evaporation permitted) produce far less injury than
do II or L. 'I'ested more realistically in relatively
large doses (drops t.l mm in diameter), it produces
injuries which after 3 days are no more severe than
those elicited by HN3 or IIN2.46 It is known from
other data (see Chapter 6) that, under the moderate
conditions of temperature and humidity prevailing
in the above test, these nitrogen mustards are no
more than one-fourth as vesicant as H. All observa-
tions14414652" indicate that skin injuries due to
KB-16 require considerably longer (i.e., 5 6 days)
to attain maximum severity than do those usually
produced under similar conditions by II, L, or the
nitrogen mustards.
It should be noted that all of the above observa-
t ions were confined to applications of the liquid agent
to the not visibly sweating skin of physically inactive
subjects at moderate temperatures and humidities.
SECRET 126
ALIPHATIC NITROSOCARBAM AXES A ND REL ATED COMPOUNDS
No determinations have been made of the vesicant
potency of liquid KB-16 on hot, sweating skin, of the
vapor under any conditions, or of the effectiveness of
either the liquid or the vapor through ordinary or
protective clothing.
B i t Eye-Injuring Action
Numerous observations on the effect of KB-16
vapor on human and animal eyes demonstrate that
the agent is an insidious and potent eye-inju-
rant.16* h r g,h i k l7*,d *‘ f i i 24* “6 S2'44’5'* h As has been
mentioned, no exposure symptoms are produced in
animals by even high concentrations (i.e., 0.2-
0.4 mg 1), and exposures entirely undetected have
sufficed to produce moderately severe injuries in
laboratory workers.The onset of injury and ac-
companying symptoms is more delayed than in the
ease of H. and much more delayed than in the east*
of the arsenieals. There is an asymptomatic latent
pericxl of many hours. Maximal damage develops
after from two to several days, and recovery is pro-
tracted. Corneal edema, opacity, and delayed but
extensive vascularization are the most prominent
symptoms. The conjunctivas are also injured, al-
though less extensively than in the ease of H. Iritis
occurs but is not so conspicuous as in eyes exposed
to IIN2 or H. Delayed relapses such as occur in the
case of H have not been observed.
An interesting preliminary report lfi,‘ indicates that,
in addition to the injuries just described, severe
retinal damage can be produced in animals by ex-
posures to relatively small dosages of KB-16 vapor
which produce only moderate conjunctivitis and
slight and transient superficial keratitis. Changes in
the retinas of eats examined 3 14 days after exposure
of the animals to O.Oii mg I for 10 minutes consisted
of: (1) slight increases in glial cells and perivascular
macrophages, with hyperchromatieity of ganglion
cells; (2) restricted zones of perivascular cuffing with
leucocytes, resembling a periarteritis; and (3) intense
chorioretinitis with subhyaloid hemorrhages, migra-
tion and phagocytoses of pigment, and extensive
chromatolysis and destruction of ganglion cells. ( om-
parable exposures to II Nl produced no morphologi-
cal changes in the retinal ganglion cells, although
clusters of leucocytes adhering to the endothelium of
the blood vessels represented a difference from the
normal retina. Exposures to H vapor (0.04 mg 1 for
10 minutes) likewise produced the clustering of leuco-
cytes, and itt addition isolated small patches of cho-
rioretinitis; however, changes in the neural elements
wore absent or at most mild compared with those
produced by KB-16 at the slightly higher dosage.
Data to be summarized in the following paragraphs
lead to the conclusion that KB-16 vapor is a dis-
tinctly more insidious eye-injuring agent than II
vapor but not necessarily a more potent injurant
when assessed on a dosage {Cl) basis. In this respect
KB-16 is similar to HN3 (see Chapter 6).
Exposuke of 11 rsiAS Eves to Small Dosages ok
KB-16 Vapok
The eye injuries produced by the vapor of KB-16
at small and minimal dosages may l>est be illustrated
by citation of the ease histories of accidentally ex-
posed laboratory workers.
In one case 38,1 some KB-16 was splashed on the left side of
the face. It was immediately decontaminated and the liquid
presumably did not enter the eye, as the worker was wearing
glasses. Nevertheless as a precaution the eyes were quickly
washed wit h water. There were no ocular symptoms out he day
of the accident. On the following day both eyes were slightly
sore but normal duties could be carried out. Ophthalmic ex-
aminations 3-22 days after the accident revealed the following
effects. J days. The conjunctivas were hypercmic, the injection
being more marked in the palpebral aperture than elsewhere.
The cornea did not slain with fluorescein but scattered epi-
thelial cells showed hydropic degeneration. Tin-pupils were
normal and there was no iritis. 4 day*. The eyes were more un-
comfortable and the patient experienced slight difficulty in
keeping them open. The conjunctivas were more hyperemic
and there was epithelial bedewing all across the palpebral
ajierture. All the limbal blood vessels were congested. The
substantia propria of the cornea was normal. The lids were
slightly swollen. There was no eheinosis and no iritis. 5 days.
The symptoms were worse and an attack of blepharospasm
and photophobia occurred. Suhepithelial cellular infiltrates
could be seen in the left eye. V days. The congestion was worse
and the left cornea still 1 axle wed. 7 days. The patient felt that
his sight was worse. Visual acuity was reduced from 6/12 (on
the fourth day) to6/24. The congestion was more marked and
the superficial layers of the substantia propria were densely
infiltrated with celts but not edematous. 9 days. The lids were
slightly sticky and puffy, the conjunctivas very injected. The
interior of the eyes was normal. 11 days. The limbal loops
showed great activity and appeared to be advancing on to (he
corneas from all meridians. 16 days. The right eye was slight ly
lx*tter, the left showed further roughening of the epithelium
and slight edema of the substantia propria. The limbal loops
were advancing. 20 days. Photophobia persisted and the eyes
appeared worse. Conjunctival injection was marked. The
margins of both corneas were invaded with a rich superficial
vascular net. The corneas were full of cells at all levels. 22 days.
The limbal ls were still extending. The symptoms were
somewhat alleviated but definite objective improvement had
not started. Comment. The main points of interest are the
absence of immediate symptoms, the long latent period, and
the delayed recovery, even though the dose was insufficient to
produce pupillary contraction or iritis. The prognosis was
SECRET TOXICOLOGY
127
considered good in view of the course of the case next to he
descril>cd.
In a second case a chemist hud been working with KB-lti
for 3 days during which time tie smelled nothing and experi-
enced no sensation to suggest that he was being exposed to
the vapor. On the second day his eyes were slightly bloodshot
but not painful. On the evening of the third day of work he
had a severe headache and pain in the eyes, and on awakening
during the night found himself unable to keep his eyes open.
He was examined at 3-22 days after tic commenced Ids work.
3 days. The lids were only slightly swollen but the patient was
unable to keep his eyes open. There was lacrimafioii but no
discharge. The conjunctivas were not very congested and there
were no hemorrhages. In the paljiebral aperture there was a
band of epithelial edema and punctate staining. The pupils
were small and their reaction to light poor. The patient had
nasal discharge. .{ days. The eyes were still closed and the
pain, now a gritty feeling, was relieved by phenacetin. The
corneas appeared improved. The lids were slightly reddened.
■> days. The gritty feeling jiersisted. There were no signs of
iritis but the limbal loops were beginning to encroach on the
corneas, d days. The eyes were much (letter and could be kept
ojien for periods of an hour or more, with attacks of blepharo-
spasm lietween. The conjunctival injection was almost limited
to the paljiebral apertures. The epithelium was bedewed but
an ocular infiltration was lieginning under Bowman’s mem-
brane. The deep structures were normal. 7 days. The eyes
could lie kept open much better but lacriniation jiersisted.
There were infiltrates throughout the substantin propria.
days. Photophobia persisted and there was a slight whitish
discharge, the epithelial bedewing in the jialjjehral ajiertures
was less pronounced. The conjunctivas were still slightly in-
jected. 11 days. The eyes were about as above except that new
vessels wore extending in ojien loops onto the corneas. Photo-
phobia jiersisted. 15 days. The newly formed sujierficial vessels
on the cornea were beginning to empty. 20 days. The photo-
phobia had jiraclieally disapjieared and the eyes were nearly
normal on macroscopic examination. 2 i days. The eyes wore
practically symjitomlcss. The limbal loojis had extended onto
the cornea all around in both eyes but wore mostly empty and
disappearing. Suliepithelial infiltrating cells were fewer. A
few" hydropic cells remained in a line on the jialjicbral fissure
of one eye. Comment, the main points are that the exjmsure
was unsusjiecled and symptoms delayed for 2 days. They
then became severe. Virtually comjjlcte recovery had occurred
within 22 days.
A number of investigators working with KB-16
and presumably receiving minimal vapor dosages
have developed mild ocular changes.16* The conjunc-
tivas showed at most only mild congestion. Exami-
nation of the corneas revealed superficial punctate
nebulae largely peripheral in location and almost al-
ways confined to the interpalpehral area. The nebulae
sometimes escaped detection on slit lamp examina-
tion but were seen after fluorescein staining. Clini-
cal notes also mention a Stahli’s line, unduly
prominent corneal nerves but no alteration in comeal
sensibility, and fine punctate hyaline areas best seen
with lateral illumination or rctroillumination and
disappearing within 2-3 weeks. The observed changes
were of a type commonly observed in various non-
specific irritations of (he eye and are difficult to
evaluate. ’1 hey were insufficient to produce signifi-
cant subjective symptoms or loss of visual acuity.
Nevertheless, in view of the slow development, of the
pathological changes caused by KB-16 vapor, it has
been recommended that individuals showing such
lesions avoid any possibility of further exposures for
1 2 weeks.-6
Comparison- of KB-16 with Other Agents ox
Basts of Effects of Relatively Large Vapor
Dosages on Animal Eyes
Quantitative comparisons of the potencies of dif-
ferent agents in terms of the dosages necessary to
produce eye injuries of casualty severity are difficult
at l>est and for KB-16 there exist no detailed quanti-
tative studies such as have l>een made with H.-9 Of
the numerous interim studies with animals, two m
permit a more or less direct semiquantitative com-
parison between (he potencies of KB-16 and II. Both
studies (Tables 5, 6, and 7) indicate (hat the poten-
Tabus 5. Effects of the saturated vapors of KB-16 and
H on the eyes of rabbits: effect of exposure time on sever-
ity of injury.3,“
The eyes were protopsed and exposed for the stated
limes at 22-24 C to vapor cups containing KB-16 or H.
At 23 ( ' the volatility of KB-16 is approximately 0.8 ing/1,
that of It approximately 0.0 mg/l.” The severity of the
resulting lesions is tabulated.
Exposure
time Agent
(sec) KB-16
II
15 Mild conjunctival le-
Minor conjunctival Ic-
sion, slight corneal lc-
sion. Rapid recovery.
sion. Rapid recovery.
30 Injury variable. Rare
Injury variable. Com-
perforation, occa-
plcte recovery in
sional complete recov-
most cases.
cry.
60 Mild permanent dam-
Moderate permanent
age. Perforation in
damage. No cases of
some cases.
perforation.
120 About same as 1-min-
Severe damage. Many
ute exposure.
cases of perforation.
cies of the two agents are of (he same order of mag-
nitude, the principal difference being that the onset
of damage and possibly the rate of recovery are more
delayed in the case of KB-16. In so far as this con-
SECRET 128
ALIPHATIC MTKOSOCAHBAAIATES AM) RELATED COMPOI ADS
Table (i. Effects of the saturated vapors of KB-16 and
H on the eyes of rabbits: tabulation of types and relative
severities of injuries.51*
The eves were protopsed and exposed for 1 minute at
22-24 C to vapor cups containing KB-16 or II. At 23 C
the volatility of KB-16 is approximately 0.8 tng/1, that
of 11 approximately 0.9 mg 1."
Characteristic of
Agent
injury
KB-10
II
Latent jieriod for severe injury
18-36 hr
6 16 hr
Conjunctival reaction:
4-
4-4-4- -
Redness
4-4-4-
4- 4-
Cheinosis
4-
4-4-4*
1 lemorrliagic necrosis
0
4-4-
Ischemic necrosis
0
— 4-4-4-
Corneal reaction;
4-4-4-
4-4-
Edema
4-4-4-
4-4-
Vascularization
4- 4- 4- 4"
4- 4-
I'lceration
4-
4- 4-
Residual opacity
4-
4- 4-
Purulent discharge
+
4- 4-
Iritis
4-
4-4-
Relapse
0?
- -4-4-
Table 7. KfTeets rties which presumably under-
lie the necrotizing action of KB-16 and the less toxic
related compounds were reviewed above (Sec-
tion 8.2.3). In resume two types of react ion have been
demonstrated to occur with substances of biological
interest in aqueous solutions at pH 8. First, a gen-
eral pro)MMlv of N-alkyl-N-nit rosocarbamic acid
esters is the capacity to transform HNH2 groups
into H• XII CO ()Br groups (carbomethoxylation,
carbethoxylation, etc.). This reaction characterizes
not only KB-16 and the corresponding ethyl ester,
but also u'trosocarbamates (i.e., ethyl N-methyl-N-
nitrosocarbamate) which do not contain a /3-chloro-
ethyl group attached to nit rogeh. Second, interaction
of KB-16 or the homologous ethyl ester with o-amino
acids results in the disappearanee of amino groups,
and, in the ease of cysteine, of the sulfhydryl group
as well. Substances analogous to the “one-armed”
sulfur and nitrogen mustards are presumed to lie
intermediates in these reactions, and conceivably
may be toxic by virtue of the alkylating power of
their 0-chloroethyl groups. The /S-chloroethyl group
of KB-16 itself is relatively unreact ive and neither of
the above-described reactions corresponds to the
principal mode of interaction of the sulfur and nitro-
gen mustards with amino, sulfhydryl, and other
physiologically important groups (Chapter 19). The
difference in mechanism is further emphasized by the
fact that KB-16 reacts in nonaqueous media with
the amino group of benzylamine, whereas reactions
of the sulfur and nitrogen mustards dejxmd upon a
preliminary solvolytic activation in water.
The reaction of KB-16 with hemoglobin in vitro
supplies a model for possible react ions of t oxicological
significance, and the absence of a comparably vigor-
ous reaction with egg albumin suggests that the ef-
fects of the agent in the cell may lie confined to only
some of the biologically important molecules and re-
active groups. Biochemical studies do, in fact, reveal
that some enzyme systems are readily poisoned by
KB-16, whereas others are not.
In one study with enzyme systems in vitro, the
effects of KB-16 were compared with those of H.'*
The three tested systems were inhibited by KB-16,
but not so effectively as by II; previously hydrolyzed
KIM6 was without effort. Purified yeast hoxokina.se
was inhibited 60 per cent by 0.006 M KR-16 and
at) per rent by 0.003 M H. 1’hosphoereatinc phos-
phokinaso was not significantly inhibited by 0.002 ,1/
and was inhibited 20 jxt rent by 0.006 M KB-16,
whereas H at 0.001 .1/ produced an inhibition of
90 per cent. Inorganic pyrophosphatase was in-
hibited 70 per cent by 0.001 M 11 and only 35 per
cent by 0,002 .1/ KB-16.
The respiration (oxygen consumption) of slices of
tissue from a variety of organs was inhibited by
treatment with 0.001 M KB-16.24 In general the in-
hibition was greater (even complete) in the absence of
added oxidizable substrates than in the presence of
glucose, lactate, pyruvate, or other carbohydrate
intermediates. The degree of inhibition increased
with time in some instances. In contrast with its
effect on “oxygen consumption, KB-16 had but a
slight effect on glycolysis as measured by carbon
dioxide output or lactic acid production. Some but
not all aspects of the metabolism of pyruvic acid by
tissue slices were markedly affected by KB-16. Oxi-
dation of pyruvate (utilization in presence of oxygen)
was inhibited, but considerable species and organ
variation occurred. The disrnutation of pyruvate as
measured by its utilization by chopped brain in the
absence of oxygen was inhibited to a smaller extent,
and its decarboxylation by dried yeast was unaf-
fected. The synthesis of carbohydrate from pyruvate*
by kidney slices (rat) was almost completely in-
hibited by 0.001 M KB-16, but another condensation
reaction, the synthesis of acetoacetate from pyruvate
by chopped pigeon liver, was almost unaffected. Ex-
perimenls with rat kidney indicated that the oxi-
dative deamination of natural amino acids (i.e.,
glutamic) is greatly inhibited by KB-16 but that
d-amino acid oxidase is unaffected. KB-16 had little
effeet on the oxidation of citrate and fatty acids by
various preparations. Cholinesterase (Stodman) was
inhibited by KB-16 but the agent had no significant
effect on a number of other enzymes including the
following: carboxylase, succinic dehydrogenase, cyto-
chrome oxidase, choline oxidase, pepsin, and urease.
In summary, the primary effects of KB-16 seem to
be flue to the inactivation of certain essential pro-
teins. Prominent among the sensitive substances
appear to be the activating proteins of pyruvic oxi-
dase and /-amino acid oxidase. Inasmuch as the re-
actions appear to be irreversible, the combatting of
injury by KB-16 should be based primarily on pre-
vention of the reactions.
SECRET 130
ALIPHATIC NITKOSOC ARB A M ATES AM) RELATED COMPOUNDS
Tabi.e 8. Properties of KIMfi, mustard gas (H), and fnsO-chlorocthyl)aniine
(11X3) bearing upon
their utility as
chemical warfare agents.
•
Agent
Property
KIM 6
11
HN3
Storage stability
poor
good
excellent
Explosion stability
questionable
good
good
Factors influencing stability on moist terrain:
Solubility in water (ppm at room temperature)
7,000
500
so
Half life in water (min at 25 (')
8 ±
2.4 +
Volatility (mg 1 at 25 (’)
0.87
0.00
0.12
Freezing point (C)
<—50
14.3*
- 3
5-9t
Density (g/ml at 25 C)
1.21
1.27
I 24
Median detectable cone, (ugl)
7 ±
o.tif
15±
1.8*
Relative eve-injuring potency
l±
1
1±
Relal ive vesicant potency of liquid on not visibly sweating skin <0.25
1
0.25-0.5
Relative vesicant potency of vapor on sweating skin
?
1
O f. 0.9
* Parc It.
t Ij-vinstrin H.
_
As is the case with II and the nitrogen mustards,
instillation of very small amounts of KB-16 into the
eye results in an inhibition of mitosis in the corneal
epithelium.341* This effect is exerted by less than one-
thousandth of the minimal dose causing clinically
visible lesions.
R.5 EVALUATION AS WAR GASES
KB-16 and the most toxic related compounds (i.e.,
ethyl X-(/3-chloroet 1 iy 1 )-X-ni trosoca rbamate and X-
(0-chloroethyl)-N-nitrosoacetamide possess insuffi-
cient storage stability to be seriously considered for
large-scale manufacture for purposes of chemical
warfare. It has been suggested that this difficulty
might he overcome by nitrosating the stable inter-
mediate, methyl X-(j8-ehloroethyl)carbamate, with
nitrous gases just before use, or by development of a
munition designer! to effect the nitrosation shortly
before firing or even thereafter However, comparison
of the other properties of KB-16 with those of such
persistent agents as II and HX3 (see Table 8) leads
to the conclusion that KB-16 does not possess suffi-
cient general utility to merit such special treatment.
Moreover, in the opinion of the authors, it would not
deserve serious consideration even if a method for its
stabilization should Ive forthcoming.
KB-16 does possess certain desirable features —
low freezing point, lack of pronounced odor, and
effectiveness as an eye-injurant at low dosages. The
available data do not permit the conclusion that the
vapor dosages necessary to produce casualties among
unmasked troops by eye or respiratory injuries would
l>e of a different order than the dosages required in
the cases of II and HN3. Given equivalent low vapor
dosages, however, KB-16 because of its less pro-
nounced odor would be a more insidious and there-
fore more effective agent than H. On the other hand,
it would not have this advantage over HN3, which
is less odorous.
Because of the necessity of assuming that enemy
troops will be equipped with gas masks, current doc-
trine giv es greater weight to the vesicant effects than
to the eye-injuring potency or inhalation toxicity of
a persistent agent not having either much less odor
or much greater potency (or both) than II, KB-16,
or 11X3. Thus, the relatively low vesicant potency of
KB-16 places it at a great disadvantage in compari-
son with H.
SECRET Chapter 9
FLUOROPHOSPHATES AND OTHER PHOSPHORUS-CONTAINING
COMPOUNDS
By Marshall (inks and Binhey Renshaw
9.1 INTRODUCTION
Approximately 200 phosphorus-containing com-
pounds of widely varying structures were ex-
amined as candidate chemical warfare agents during
World War II, Only the few represented by the
dialkyl fluorophosphates, the diamidophosphoryl
fluorides, the alkyl cyanamidophosphates, and the
alkyl duorophosphonates have merited detailed ex-
amination. The individual compounds that have re-
ceived most attention are:
2. Diary.idophosphoryl Jluoridcs.
CH,
\
N
) / \
/ CH, \/
/ P
CH, /\
\/ F
N
/
cn,
hid I hmethylamido)phosphoryl fluoride (TL 792, T-2002)
1. Diailkyl Jluorophosphates.
CH,—O O
\ /
P
/ \
— ch,—o f
Dimethyl fluorophosphate
(PF-1, TL 311, T-1035)
CH, CH2—O 0
\ /
1*
/ \
CH, CH,—() F
Diethyl fluorophosphate
(TL 345, T-1036)
3. Alkyl cyanamiduphosphates.
CH,
\ “
N
/ \ O
CH, \ /
P
/ An
ch,ch2—o
Kthyl dimethylamidocyanophosphate
(MCE, Tahiti), Lc 100, TL 1578, T-2104)
CH, CH,CH,
\ \
CH—O CH—O
/ \ o / \ o
CH, \ / CH, \ /
P - p
CH, /\ CH5CH2 - /\
\ / F \ / F
CH—0 CH—O
/ /
CH, CH,
4. Alkyljluorophosphonales.
CH,
\
, CH—0 O
J / \ /
CH, P
/ \
/ F
H,C
Isopropyl methanefluorophosphonate
(MF1, Sarin, T-I44, TL 1618, T-2106)
Diisopropvl fluorophosphate
(PF-3, DPF, TL 466, T-1703,
1152)
Di -scc-hutyl fluorophosphate
(TL 1206, T-1835)
H,C—ch2
/ \ - -
H,C CH—o
\ / \ o
11*0 CH, \/
HjC CH* 1*
/ \ / \
H2C CH—O f
\ /
II2C CH,
CH,
_ \
CH—O O
/ \
CH, P
CH,CH2
Isopropyl ct hanefluorophosphonate
(TL 1620, T-2109)
Dicyclohexyl fluorophosphate (TL94!, T-1840)
SECRET 132
FUUOROPHOSPH VTES \ M) IMIOSPIIORUS-COXT VI \l \C COMPOUNDS
The dialkyl fluorophosphates were described in the
open literature in 1932. The British undertook their
examination as war gases in 1911 and much work on
them was subsequently carried out in the United
Kingdom and United States*
They are parasympathetic stimulants and cholin-
esterase poisons of high potency. For some sjx'cies
(e.g., the monkey), PF-3 and di-scr-butyl fluoro-
phosphate are more toxic than any of the standard
United States or British chemical warfare agents. At
lethal concentrations they are” “quick-kill” agents,
their action being only slightly less rapid than that
of hydrogen cyanide (AG). However, their relatively
low volatility, at 25 G 30 mg 1 for PF-l, S mg 1 for
PF-3, and 1,8 mg 1 for di-xcr-butyl fluorophosphate.
puts them in a class with the persistent agents and
would render difficult the rapid administration of a
lethal dose under field conditions. Chief interest in
them has arisen from their action on the eye. They
produce extreme constriction of the pupil, interfer-
ence with the muscles of accommodation, potentially
dangerous congestive iritis, and severe pain behind
the eyes. PF-3 and di-scc-butyl fluorophosphate at a
dosage of 50 mg min nr* produce pupillary constric-
tion, and PF-3 at about 300 mg min nr* produces the
other harassing symptoms just mentioned. However,
by 1943 and 1911 careful assessments led to t he con-
clusion that in practice these effects would be harass-
ing rather than casualty producing. It is believed
that troops supplied with gas masks would not he-
come casualties from attack with the fluorophos-
phates except under circumstances where standard
non persistent agents would have equally or more
severe consequences. A useful interim summary of
work on the fluorophosphates was prepared by Di-
vision 9 in 1944.37
The diamidophosphoryl fluorides proved to be
about as toxic as the fluorophosphates but to be less
potent in their action on the eye. Their chief point of
interest is that they are extremely stable in water and
upon oral administration are among the most toxic
of the known synthetic compounds.
The dialkyl fluorophosphates appear to be eclipsed
in toxicological potency and potential value as
chemical warfare agents by the alkyl cyanamido-
phosphates and alkyl fluorophosphonates. These
compounds, known collectively as Trilons (a name
assigned to them by the Germans), first came to the
attention of United States and British workers after
the termination of hostilities in Europe in the spring
of 1945. It was then discovered that the Germans had
manufactured largo quantities of MCE for use in
bombs and high explosive-chemical shell. They had
been attempting also to prepare MFI on a large scale
tint had I>een unable to overcome difficulties in its
synthesis.
The Trilons an* similar in mode of action to the
fluorophosphates but are considerably more potent
both in terms of inhalation toxicity and in the pro-
duction of eye effects. For the monkey the L{Ct)„o’s
of MCE and MFI are in the order of 250 and 150 mg
min m3, respectively. In man MCE at the extraor-
dinarily low dosage of 3.2 mg min m3 produces
pupillary constriction. Dosages in the order of 15 to
20 proved to Im> definitely harassing because of ocular
and systemic effects, and it would seem that 30 mg
min m* might suffice to produce significant partial
disability. Quantitative eye data on MFI are not
available to the reviewers. Although MCE is some-
what less volatile than mustard gas (H) and is sus-
ceptible to hydrolysis, MFI has (he rather high
saturation concentration of 16 mg 1 at 25 C and is
quite stable. Moreover, it is virtually odorless.
it would seem that the Trilons are the one new
group of chemical agents discover'd during World
War II which merit serious consideration for adop-
tion as standard agents. Their use in high explosive-
chemical shell, indistinguishable on detonation from
ordinary high-explosive munitions, should 1k> care-
fully evaluated and assessment made of the relative
casualty-producing effects of (1) the initial cloud of
droplets and vapor and (2) the subsequent vapor
evolution from the contaminated terrain.
Division 9 has participated in work on the Trilons
only to (he extent of performing limited studies on
synthesis, detection, and analysis. Most of the re-
ports on work done by other agencies have become
available during the period when the division was
terminating its activities. Some of these reports may
not have come to the attention of the reviewers. It
has not been possible to render the review of the
Trilons as complete as the discussion of the other
agents of major importance. A summary of the field
trials conducted at Raubkammer after the defeat of
Germany has not been included, and a complete
assessment of t he value of t heTrilons as chemical war-
fare agents has not been undertaken in this chapter.
9.2 SYNTHESIS \ND PROPERTIES
9.2.t Synthesis
Many methods have been used in the synthesis of
the compounds listed in Table 1. The following dis-
SECRET SYNTHESIS AND PROPERTIES
133
Table 1. Fluorophosphatcs, amidocyanuphosphates, fluoropbosphonates, and other phosphorus compounds examined as
candidate chemical warfare agents.
The compounds are arranged in the following general classes; (1) derivatives of phosphine, (2) derivatives of primary
phosphines, (3) tertiary phosphines, (4) oxygen, sulfur, and nitrogen derivatives of trieovalenl phosphorus, (5) phosphorus
pentahalides and related com|xmnds, (6) phosphoric' and phosphonic acid derivatives and their sulfur analogs, (7) quarter-
nary phosphonium salts, and (8) miscellaneous compounds.
The following abbreviations arc used; n'B, refractive index at / C’; ', density in g ml at I , specific gravity at
/, C in reference to water at fj C; mp, melting point in C; bp'1, boiling |M>int in V at /» mm Hg; vp', vapor pressure in
mm Ilg at 1 C; and vol', saturation concentration (volatility) in mg l at 1 C.
Centigrade scale is used throughout the table.
Reference
Reference to
(’onqsnind
to
synthesis
Physical pro|H*rlies
Property Reference
toxicity
(hit a
1, Phosphorus trifluoride
10, 10tH>,
bp
101.1°
I06d
11, 106a
2. Phosphorus monochlorodifluoride
106d
lll{>
151-5°
10< id
11
3. Phosphorus dichloromoiiofluoridc
11
4. Phosphorus trievanide
30d
bp° 5
150'
30d
■>. Phenylphosphine
27g
(sublime
11, 18
6. Kt hyldichlorophosphine
2
bpT«o
9-1-97°
2
11
7, Kt hyhlicyanophosphine
27r
bp'"
94- 9(5°
27r
11, 60d
S, Phenyldiehlorophosphine
—
!). Phenvldicvanophosphinc
27d
tP;
1.1660
27d
II. IS
—
— . .
bp*
100
27d
...
—
bp20
145°
27d
*
mp
35
27d
10. Phenyhlithiocyanophosphine
27f
11, IS
11. p-Chloro|)henyldichIorophosphine
12. p-ToIyldichlorophosphine
13. a-Xaphl hyldichlorophosphine
. T __
■ —...
14. 2-Dibenzofuryldicyanopbosphine
27h
11
15. 3-( X-Kl hylcarbtizole) dichlorophosphine
16. 2-Phenoxthiindicyanophospltine
27h
27h
... —
• • • •
H
1)
17. Triehloromethylphosphinc —_
27f, 77
11, IS
18. Triethylphosphine
27d
bpj«
127.5°
27d
11
1ft. Tributylphosphine
27b
20. Trioclylphosphine
27.1
bp‘
234-237°
27d
11. 18
-
mp
30°
27d
21. Tridecylplaisphine
27h
... ■
11, 18
22. Diethylphenylphosphine
27f
II. 18
23. 1 tiallylphenylphosphine
27f
24. Dibittylphenylphosphine
27e
p5"
185°
27e
- —
bp"*
116°
27e
fp
25°
27e
25. difluoropb(>sphine
27n
II
39.
Dietbvlaminodichlorophospbinc
27d
1.196
27d
11
bp'*
72 75°
27d
. . ,
'
—
189°/atmos.
27d
10
N,X-his((3-('blon>etliyl)aminodicliloropb<>.sphine
1. 27m
II
41.
Klhyl-(d-cbloroethylthin)ehlomphosphinc
27p
bp" *
89-92"
' 27P
11, 18
42.
DiplKMiyI-/3-chlorsphin®
82
bp'
74 75°
82
82
48.
5i>(T)imelhylaniino)fluorophosphine
10
iP*
0.975
10
11
—
bp
120
10
40.
KlbvW»is(£-fluoroethoxy)phosphinc
35a
bp*-*
40 4'.)°
35a
II
50.
Pbcnyldiet hoxyphosphine
27g
11, 18
51.
Phcnyl-Wsfo-ehlorophenoxy Iphosphiue
27h
11, 18
52.
K(bvl-6)*(ifFchloroelliyllhio)phosphine -—
27p
WO5*
1.5600
27p
11, 18
bp
115-120°
27p
— ...
53.
Phcnyl-5is(methylthio)phosphine
271.
It. 18
54.
55.
Phenyl-fnsfd-chloroethyllbiojphosphine
p-Dimethylaminophenyl-bisfd-ehloruethyllhio)-
271
II, 18
phosphine rnonoelhylate
27n -
....
11. 18
50.
Phenyl-fci*(d,p'-dichloroisopropylthio)phosphinc
27q
II
57.
Phenyl-5iK(hu1y!lhio)phosphinf
27h
....
—
11, 18
58.
Dimethyl hydrogen phosphite
27h
....
• •
11, 18
59.
fws(d-Fluoroelhyl) hydrogen phosphite
bp"
109 110“
104p
I04p
00.
1 )iisopropyl hydrogen phosphite
bp17
82.5°
I04f
104f
fil.
Tri met hoxyphosphine
27h
11, 18
02.
T riel hoxyphosphine
27c
iP”
0.968
27c
11, 18
_ . . .
l.p7'"
I55-156“
27e
bp"
36-38°
27e
63.
/rr.s(f)-Flnorriet hoxv )))hosphine
30(1
bp" 4
100-103°
30d
II
04.
tris{ d-Chloroethoxy Jphosphinc
27f
II, 18
65.
trfs( d-Rromocthoxy)phosphine
27m
II, 18
66.
Trihutoxyphosphinc
34h
1';"
98-100°
105e
104l>
129. Diisopropyl fluorophosphate (PK-3)
Sec text
1.3780
105e
Sec text
hp-4
84-85°
10.5e
hp:su
183“ (est)
105c
fl>
- 93°
Vol*°
5.84
12
)30. btx(fi,ii1held,ireiis.ipn>|ivl) fluorophosphate
10.5k
hp"-3 *
163 165”
10.5k
131. Dihulvl fluorophosphate
28h
105a
132". Di-sn’-hut vl fluorophosphate
I05f, 10.51
hp- -
02-64°
105f
Sea1 text
voli0
1.12
12
133. Diamyl fluorophosphate
... ■
tOlh
13-1. Diisoamyl fluorophosphate
I05f
l»l»"
145 118°
I05f
1041.
135. W«(o-Klhylpn>pyl) fluorophosphate
105k
hps »
97-98°
105k
10-lj
13(5. Dieyelohe.xyl fluorophosphate
105i
«l>
1.4558
281
See text.
hp0-*
116°
I05i
voP»
0 0044
12
V • ,
137. bix( 1,3-DimelhyllnilyI) fluorophosphate
105k
1»PIT
102 103°
10.5k
UMj
138. hix(‘2- Methyleyclohexyl) fl\iorophosptuite
I05p
hp1"
120°
105p
I05p
139. hlx(a-Ca els'! hoxvet hyl) fluorophosphate
hp" 14
137°
105p
ia5k
hp" ‘
126 128°
105k
10-lj
140. Di|dienyl fluorophosphate
2sj, ia*if
bp"
106-108“ _
I0.5f
lOlR
141. fus(Triethvllead) fluorophosphate
105c
nip
>200°
105e
105e
142. Diethyl chlorophosphate
11, 18, 104h
143. fcfsO-Fluor, tethyl) ehlorophosphale
30d
i.p"4
108-112°
30d
-
144. Diethvl evanophosphate
J05r
hp"
95 97“
105r
105r
145. Diethyl lhioevanophosphate
105e
hp14
115-125°
105e
I04d
14R. Kthvl X-plienvIamidofluorophospliale
105n
rap
.50° —
105n
105n
117. Methyl N.N-diethvlamidoehlorophosphatc
25
«i>
1.4443
25
69d
!4H. Kthvl X, X-di met hvlamidocya n. .phosphate
hpn,*-*-i*
19 49.2°
25
(MCK)
21,25, 105r
no
1.4243
25
See text
iPn
1.077
25
hp"-*
56-58°
25
f|>
.50.0°
60
vol*4
0.567°
69c
vol“
0.612
60
—
149. Methyl X, X-diet hylam idocyan. .phosphate
35b
bp"4
65 66°
35b
69el
150. Kthvl X, X-diet by la midocy an..phosphate
151. Methyl X,X-f»/x(0-chloroethyl)amidocyanophos-
30c
tide
phatc
69el
152. hix(Dimelhvlami«lo)phosphoryl fluoride
10, 105m, 105o
ip1
1.110
10
See text
l.p14
86°
105m
. .
, _ ... .
hp!
50°
10
vol.**
2.16
12
153. hix( ButvlnmiphosphoiyI fluoride
105m
rap
59.5“
105m
I04ei
154. 6)«(Diethylamido)phosphoryl fluoride
105m
m,"
1.4321
28m
lO-lo
hp5 4
83 87°
28m
hp5"
124.5-125.5
105m
155. bis(Morpholido)phosphoryl fluoride
105und
synthesis
Pr
»|ierty
Reference
data
1(12. bix{ Diinethylainidoiphosph.iryl chloride
104r
1 03. Diethvl fluorothiophosphate
2Sk
bp"'
55°
28k
11, 18
HU. Isopropyl inethaneflnorophosphonate (MFI)
Sr-e text
wna
1,37‘K)
09.1
.See text
d1-
1.0941°
09.1
. ; , , .
bp15
50.5 57
25
Vol2i
10.4
69e
105. Isopropyl ethanefluorophosphonate
22
»nsi
1.3872
22
See text
«n55
1.3817
09e
. v ■ £.*
d»
1 .0552
09c
bp“
07-68°
22
.7.
/
vol;s
11.0
09e
1 (VO. 2-C’hlor.ihexenc-l-phosphonie achy
27k
107. Dimethyl met hanephosphonate
271
11,18, 09a
10S. bi«(d-(’hloroethyl) met hanephosphonate
271
—. . . .
II, 18. 09a
109. Di-.w-bnl vl fluoromethanephosphonale
ia5k
lip3
90 100“
10.5k
KUj
170. Diethyl 2-flnomet hanephosphonate
105s
bpH
200202
105s
405s
171. Diethyl 2-chloroothanephosphonate
27k
II
172. Dimethyl propane-2-phosphonate
271
. . -
11, 18, 09a
173. hi.-i(J < 'hi. .methyl) pmpane-2-phosphonate
271
11, IS, 09a
171, Diethyl o-toluenephosphonate
105s
hp"
155°
105s
175. Diethvl earbelhoxvmel hanephosphonate
27e
d'
1.139
27e
11, IS
bp
259’(a(inos)
- 27c
'•p'5
149-150°
27e
170. (l-C'liloroethyl diethyl phosphate
bpw
144-145°
104f
lOlf
177. frts(d-C'hloroethyl) phosphate
77
178. /ri«(d,/3'-Diehloroisopropy'l) phosphate
...
77
179. Irjs(o-Crcsyl) pliosphale
77
1 SO fns{2-MellivlJO-propylphenyl) phosphate
77
181. IriM. Ethvlt llio) phosphate
bp"
172-174°
104f
104f
182. Diethyl amidophosphate
bp" *
131 138°
104f
lOlf
mp
45.5°
104f
.
183. Diethyl X-ethvlamkh»phosphate
30b
bp° 1
90°
;ioi.
184. Diethyl N,X-diethylamidophosphatc
30b
bp*
90°
30b
11, 18
185. Dime!hvl X,N-bisi/J-chlorocthyljamidophosphate
69d
180. Diethvl X, X-bix(jj-chh>roethylJamidophosphale
30c
bp"
104-165.5°
30c
11
187. hi>(/J-Ch 1 oroethy 11 hin) X , X-h/.sfd-ehloroei hyDamido-
phosphate
27o
n,i*
1.5525
27o
II, 18
d4
1.472°
27«
bp""1
155 100°
27o
188. /m(Dimethylamido)phosphate
10
bp*
81°
10
11
189. Trimethyl thiophosphate
27h
II, 18
190. Tricthyl thiophosphate
bp19
100°
104k
101K
191. Phosphoniuin iodhle
27d
sublimes
02.5°
27d
192. I(ir(ikis{( ’hloromet hyl )phosphonium chloride
27e
nip
192-193°
27e
1 1, IS
193. ft-Chloroel hyltriet hvl phosphoniuin iodide
‘27k
11
194. fOHrmnocthyltricthylphosphoniuni bromide
27c
mp
235°(d)
27c
11, 18
195. Triethyl phenyl phosphoniuin iodide
27h
—...
11, 18
190. Triethyl-p-tolylphosphonium iodide
27h
II, 18
I!I7. Triallylphenylpliosphonium bromide
27k
11, 18
198. Triphenylphospholietaine
27h
11, IS
199. jS-Chloroelhyltriphcnylphosphoninm iodide
27h
11
200. d-Ilromoethvltriphcnvlpliosph. mium bromide
27d
mp
208°
27.1
11, 18
201. 3-Chloroacetoiiyltriphenylpliosphonium chloride
27h
11, 18
202. d-('hloroetliyl-een obtained using hydrogen fluoride as the
fluoridating agent .m
A well-eoolcd solution of phosphoryl dichlorofluoride in dry
ether is treated with cyclohexanol. After complete removal of
hydrogen chloride, the resulting dicyclohexyl fluorophosphale
is isolated in 50 |>er cent yield by fractional distillation under
diminished pressure,1061 Attempts to prepare this compound
by the phosphite or the phosphoryl chloride methods have
l)een unsuccessful.
The following compounds have Ifcen prepared by this
method:
diethyl flnorophosphate ,“h
dipropyl flnorophosphate lu6-
diphenyl flnorophosphate l#5(
Wsfethylthio) fluorophosphale ,osr
W*(/>-fluoroethyl) fluorophosphale ,oy
his{0-Chlorocthyl) flnorophosphate lu6k
hidd-methyleyclohexyl) flnorophosphate ,05P
dicyelohexyl fluorophosphale 581 >"*■
2. Phosphoryl chloride method — synthesis of dimethyl flu-
orophosphale (PF-!). Phosphoryl chloride is treated with
methanol at — 78 C and the mixture is allowed to come to
room temperature slowly. Hydrogen chloride is evolved rap-
idly for a period of 2 hours. The mixture is transferred to a
copper vessel and treated with hydrogen fluoride. Fractiona-
tion following a crude distillation gives PF-1 in 34.5 to 38.8
per cent yield.2"
The following compounds have been prepared by this
method;
dimethyl fluorophosphale !5r
diethyl fluorophosphale ,8e
diisopropyl fluorophosphale
h/s(d-chloroethyl) fluorophosphale 281
A slight modification of the method has been used to pre-
pare ethyl /8-ehloroethyl flnorophosphateISl and methyl
ethyl fluorophosphate.” By using thiophosphoryl chloride as
a starting material, derivatives of fluorothiophosphoric acid
have been prepared, and, by the use of one mole of alcohol
j»er mole of phosphoryl or thiophosphoryl chloride in the first
step, derivatives of difluorophosphoric or diflnorothiophos-
phoric acid.J8o *>,0i[
3. Dialkyl hydrogen phosphite method — synthesis of diiso-
propyl fluorophosphale (PF-3), A cooled solution of isopropyl
alcohol in either carbon tetrachloride or ether is treated with
a solution of phosphorus trichloride in the same solvent and
then blown with air and treated with ammonia to remove hy-
drogen chloride. Filtration and fractionation give diisopropyl
hydrogen phosphite in 82.5-89 per cent yield. This material
is then chlorinated in 71-80 per cent yield to give diisopropyl
* Improvements on Lange’s preparation of ammonium
fluorophosphate from ammonium bifluoride and phosphorus
jjentoxide have Is-en reported by Marquina.118
SECRET SYNTHESIS AND PROPERTIES
139
cblorophosphatc, which is purified first hy blowing to remove
hydrogen chloride and then by fractional distillation. Fluori-
nation is accomplished by gentle heating of diisopropyl chloro-
phosphatc in benzene with jiowdcred sodium fluoride. PF-3
is obtained in 84 |ht cent yield; the overall yield is thus in the
order of GO per cent. Hydrogen fluoride can also l>e used as a
fluorinating agent.58* Distillation of the intermediates can lie
eliminated and the whole process carried out in the original
solvent (carbon tetrachloride) without greatly decreasing the
yield.1"’1'
The following comjiounds have been prepared by this
method:
dimethyl fluorophosphate 4
diethyl fluorophosphate Ksa
diisopropyl fluorophosphate 4-“.,IB*
di-we-bulyl fluorophosphate ll4“
diisoamyl fluorophosphate ,"i(
his( 1,3-dimet hylbutyl) fluorophosphate lu6k
botfor-cartiet boxyct hyl) fluorophosphate ,l£‘k
5i«(a-ethylpropyl) fluorophosphate ll6k
6js(d,d'-dichloroisopropyl) fluorophosphate U6k
fws(/*-fluordethyl) fluorophosphate ,®t
The last step of the reaction can lie modified so as to replace
chlorine by groups other than fluorine. Diethylthiocyano-
phosphate and diethyl amidophusphate have been prepared
in this way.1115*
Semitechnieal .syntheses of PF-1 and PF-3 have
been carried out by the dialkyl hydrogen phosphite
method, but only with the latter compound have
enough runs lieen made to standardize conditions. A
description of the procedures used follows.
I. Semi technical preparation of diisopropyl fluorophosphate
(PF-3).* The main reaction was carried out in a 130-gallon
I.astiglas-lincd jacketed vessel equipjied with a lined and
coated gas inlet pipe, a propeller-type stirrer, a charging pipe,
sight glasses, manometer connections, and a bottom outlet.
Steam or refrigerating brine could be circulated through the
jacket. The reactor was connected to a 10-foot
steel tower, 6 inches in diameter, which was fitted with a lead
coil condenser from which distillate could lie passed to either
of two receivers. The bottom outlet of the reactor was con-
nected to the top of a 40-gallon filter tank equipped for vac-
uum filtration.of the slurry and return of the filtrate either to
the reactor or one of the receivers. Since plugs developed at
the Ixittom outlet in many runs, an additional connection lie-
tween the gas inlet and the top of the filler tank was provided
to allow transfer of the slurry by this route. All vacuum lines
led to a 35-gallon separator tank which was connected to a
three-stage steam ejector. Drain lines leading to a 40-gallon
lead decontaminating tank were provided. A separate still
was provided for benzene distillation.
In a typical run, 212 lb (3.54 pound-moles plus 1 per cent
excess) of isopropyl alcohol ( <0.2 per cent water) was cooled
with brine to — 5C in the jacketed reactor. Phosphorus tri-
chloride (100 lb, 1.16 pound-moles) was added gradually with
cooling and stirring over the course of 4 hours, during which
the temperature was not allowed to exceed 12 C. The system
was kept under slightly diminished pressure (about 700 mm).
The mixture was then stirred for \2 hour before applying
the full vacuum of the steam jet. Chlorine was passed into the
reaction mixture at a rate of 12 pounds per hour with contin-
ued cooling. The end of the reaction {10 hours) was indicated
by a drop in temperature, even though the rate of How of
chlorine was increased. A total of 122 lb of chlorine (1.72
pound-moles, 48 per cent excess) was used.
To remove excess chlorine, hydrogen chloride, and iso-
propyl chloride, the stirred mixture was kept under vacuum
for 2 hours, during which time the leni|ierature was gradually
raised to 20 C by passing steam into the jacket of the reactor.
Ten gallons of benzene was then added and distilled off under
reduced pressure at a maximum temperature of 30 C. The
last t races of hydrogen chloride were removed by adding an
additional 10 gallons of benzene and distilling under reduced
pressure at reactor temperatures not exceeding 50 C.
After cooling to 20 C, 19 gallons of benzene recovered from
a previous run was added. Dry sodium fluoride (95 |ier cent
pure, 123.4 lb,-2.8 pound-moles, 142 [>er cent excess) was in-
troduced into the reactor through an inlet line by means of a
funnel. The stirred slurry was heated to reflux during I hour
and held at reflux for 4 hours; it was then cooled and filtered.
After washing the filter cake with three 5-gallon portions of
benzene, the filtrate and washings were combined, collected in
the cleaned reactor, and distilled under minced pressure. The
lienzene forerun containing about 2 per cent of product was
collected to lie used in the following run. One hundred and
fifty-eight pounds (74 per cent of theory based on phosphorus
trichloride) of FF-3 was obtained. The entire run required
44 hours. An additional 20 hours was necessary to decontam-
inate and dry the system in preparation for the next run.
Preliminary design and round cost estimates for a full-scale
plant to produce 500,000 lb jit*r month of PF-3 by a batch
process have been "drawn up using data obtained during oper-
ation of this pilot plant. It is estimated that the capital cost of
the complete plant would tie $700,000. Estimated manufact ur-
ing costs are $0.37 per pound of product, $2,222,000 per man-
ufacturing year.5
Hound cost estimates for a plant producing PF-3 by a con-
tinuous process have also been-prepared.' Although fewer ex-
perimental data arc available (the estimates are based on
laboratory scale work only),4 a smaller capital outlay and
lower operat ing costs seem possible.
A total of 13 kg of PF-3 has been prepared at the British
Research Establishment at Sutton Oak by a batch process
resembling that just descrilicd.1"2
2. Pilot plant preparation of dimethyl fluorophosphate
(PF-1). Three pilot plant runs on a process similar to that al-
ready described for PK-3 have been carried out to produce a
total of 35 lb of PF-1. This experience was not sufficient to
allow standardization of conditions, but it was found that the
temjieraturcs required, which are somewhat lower than those
in the PK-3 process, could be maintained without difficulty.
Because of mechanical difficulties, yields approaching those
obtained in the laboratory (72 per cent) were not realized in
these three runs.1
DIA M IDO PHOSPHOR VL F UOK IDES
A number of compounds in this series have been
prepared by Ibe application of the following more or
less straight forward methods.
SECRET 140
FU OROPHOSPHAXES AM) I’llOSPUOUUS-CONT VIM M; COMfOlADS
1. The action of amines on phosphoryl dichloro-
fluoride.
2. The controlled action of amines on phosphoryl
chloride followed by fiuorination.
3. The action of amines on phosphoryl fluoride.
The first of these methods appears to be general.
It is carried out by adding a solution of phosphoryl
dichlorofluoride in ether, benzene, or toluene to four
moles of the amine in the same solvent. After filtra-
tion from the precipitated amine hydrochloride, the
product is isolated by distillation or crystallization.
The following compounds have been prepared in this
way;
dianilidophosphoryl fluoride ,nSf
6j«(dimethylamido)phosphoryl fluoride 1 "
fe/s(diethylamido)phosphoryl fluoride
fcf.v(butylamido)phosphoryl fluoride iu:,m
bbv(cyclohcxylamido) phosphoryl fluoride ,95m
6/»(met hy lanilido)phosphoryl 11 uoride 'li5,u
6/*(benzylamklo)phosphoryl fluoride l05m
A modification of this method involving prior
treatment of phosphoryl dichlorofluoride with one
mole of alcohol has yielded ethyl N-phenylamido-
fluorophosphatc.H*“
The second method also appears to be general, and
avoids the use of the difficultly available phosphoryl
dichlorofluoride. The fiuorination of fns(alkvlamido)-
phosphoryl chlorides proceeds somewhat less readily
than that of dialkyl chlorophosphates. The method
has been used successfully with 6/«(anilido)phos-
phoryl fluoride and with 6 rs(di me thy lamido) phos-
phoryl fluoride.1050
The third method suffers from the disadvantage
that a large part of the fluorine is wasted. It has Wen
used to prepare 6/s(dimethy!arnido) phosphoryl flu-
oride.19
Alkyl Cyanoamidophosphates
Although vague but persistent rumors of a new
German gas, Trilon, reached Allied hands from time
to time during World War II through intelligence
channels, no reliable information as to the nature of
this gas or gases became available to the Allies until
the spring of the German surrender, when German
munitions charged with a new agent were captured.
The agent was very quickly identified as ethyl di-
mefhvlamidocyanophosphale (AICE) and an in-
tensive study of it covering all phases of interest to
chemical warfare was started. About the same time
an intelligence team interviewing members of the
staff of the I. G. Werke, Elberfcld, reported that this
compound had been discovered in 1937 by 1. G.
Elberfcld du ring a search for new insecticides, and
that in the following year an even more toxic and in-
sidious substance, isopropyl methanefluorophospho-
nate, had been discovered.72 Both compounds had
lx*en imported to the War Ministry under its standing
order to the German chemical industry regarding the
reporting of toxic substances.
The laboratory method of synthesis of ethyl di-
methylamidocyanophosphate (MCE), disclosed in
detail by (he 1. G. representative's, made use of the
following steps.72
1. The interaction of t wo moles of dimethylamine
and one of phosphorus oxychloride, first at 30 (' and
finally at 120 C, to produce dimethylamidophos-
phoryl chloride in 93 per cent yield.
2. The action of sodium cyanide and ethanol on
dimethylamidophosphoryl chloride to give MCE in
90 per cent yield.
This procedure has lieen checked in at least two
laboratories in this country 24 25 and the German
claims substantially confirmed, although the yields
obtained were not so high. No detailed study of the
reactions was carried out. A novel alternative method
for laboratory preparation of the agent has Ix-en
used in Great Britain.195 In this procedure, diethoxy-
phosphorus chloride is allowed to react with
dimethylamine and the resulting diethoxydimethyl-
aminophosphorus is treated with cyanogen iodide to
give MCE directly.
In 1939-40 the Germans began pilot plant produc-
tion of MCE at Munsterlager, near Bremen, and ex-
perienced no difficulty in the manufacture of 50 tons
of the material. Construction of a large plant at
Dyhemfurth near Breslau was begun in January
1940, but production did not begin until April
1942.‘2‘3 In the plant process, chlorobenzene was
used as a reaction medium in the final step. Initially
the product was stripped to a content of approxi-
mately 5 per cent of chlorol>enzene. Eater a product
containing 20 per cent chloroltenzene was standard-
ized. Both 105-rnm shells and 250-kg bombs were
charged with the agent.7* A total of 10,000 to 12,000
tons ,3of MCE was produced. It is worth noting
that the figure 12,000 tons represents 18 jkt cent of
the, total German production of war gases of all
kinds,113 which gives some indication of how largely
this agent figured in the plans of the Germans.
MCE is a high-boiling, fairly stable liquid pos-
sessing a faint fruity odor. The pure material is color-
less, but as technically produced MCE is dark brown.
SECRET SYNTHESIS VM> PROPERTIES
It boils at 85 C under 1.5-mm pressure, at 120 C
under iO-mm pressure, and at 250 C with some de-
composition at atmospheric pressure."-’ Its density
at 20 C is 1.077 and its refractive index (//*„) is
1.4240.60 Its vapor pressure appears to be about one-
half that of II "* and can 1m* represented as a function
of temperature by tin* following equation:*
5,750
logio /'(mm) = 11.545 - _ ~ •
Its volatility at 25 C is 0.567 mg I.**1 The tactical
use of the agent as an aerosol produced by heavy-
walled shell equipped with large bursting charges
apjH*ars to have Ihmmi envisaged by the Germans.
MCE is claimed by the Germans to bo the opti-
mum compound of this scries as regards toxicological
properties,7* but this assertion has not been verified
in this country since no - comprehensive synthetic
program was established to explore the field.
A related compound of relatively high toxicity was
encountered during an attempt to prepare the iso-
propyl analog of MCE by the simultaneous action
of sodium cyanide and isopropyl'"alcohol on dimethyl-
amidophosphoryl chloride. In this case the cyano
group alone was introduced, and dimethylamido-
cyanophosphoryl chloride was obtained in 68 per
cent yield. Its toxicity is approximately one-half
that of MCE®
Alkyl FLroKOPUoscnoNATKs
Mention has been made of the discovery of this
class in 1938 by members of the staff of I. G. Elber-
feld. The optimum compound of the series, isopropyl
methanefluorophosphonate (MFI), Ls several times
as toxic for most species as is MCE, is more volatile,
and is also more difficult to detect by odor. It aroused
great interest among the Germans, but in spite of
intensive efforts to develop manufacturing methods,
production on a plant scale was never realized.
As first reported to intelligence teams, the labora-
tory preparation of MFI proceeded as follows.72
Dimethyl hydrogen phosphite is prepared in 90 per
cent yield by the action of methanol on phosphorus
trichloride, and is converted into dimethyl methane-
phosphonate in 85 per cent yield by the action of
metallic sodium followed by methyl chloride. Finally,
methanephosphoryl chloride, produced by the action
of phosphorus pentachloride on dimethyl methane-
phosphonate, is converted to MFI by the simultane-
ous action of sodium fluoride and isopropyl alcohol.
The yields in these steps are 90 and 82 per cent re-
spectively.
Attempts by both American and British groups to
use this scheme without modification were not en-
tirely successful. In tins country yields greater than
14 per cent w ere not obtained in the nv*t Inflation step
even w hen methyl iodide was substituted for methyl
chloride or when the reaction was carried out in an
autoclave at 125 C. By using dimethyl sulfate, how-
ever, yields of 77 per cent were obtained in this step.21
British workers were able to carry out the methyla-
tion step in 59 per cent yield by using a modification
of the original German procedure in which dimethyl-
hydrogen phosphite was alkylated by treatment with
sodium sand in dry ether followed by methyl chlo-
ride.10* Neither group obtained greater than 42 per
cent in the final fhioriuation and esterification.
After this work was well under way, additional in-
formation became, available from intelligence sources
to the effect that the Germans had used sodium
methoxide in methanol instead of metallic sodium in
the methylation step 72 71 and that two alternative
methods for the final fiuorination. one using sodium
fluoride and the other hydrogen fluoride, were pos-
sible, The use of hydrogen fluoride made possible
operation at lower temperatures but introduced cor-
rosion problems. Few details on the actual operation
of the final step are available.7*
The substitution of higher alcohols for methanol
in the first step of this process appears to be advan-
tageous. Dimethyl hydrogen phosphite is rather
unstable, is water-soluble, and its"sodium salt is in-
soluble in organic solvents. Diethyl hydrogen phos-
phite has given better results in the hands of British
workers, particularly in the methylation step,251"4*
whereas the use of butanol to give dibutyl hydrogen
phosphite followed by methylation with dimethyl
sulfate and sodium methoxide was adopted as opti-
mum for a simplified process suitable for pilot plant
use by NDRC workers.21 In the latter example, the
solubility of sodium dibutyl phosphite in organic
solvents appears to be distinctly advantageous. By
this method methanephosphoryl chloride can be
obtained in 79 per cent overall yield.24 Other improve-
ments made during this study were substitution of a
water-wash for filtration to remove sulfate salts after
the methylation. and combination of the first three
steps to eliminate all distillations except that of
methanephosphonyl chloride.
The isomerization process of Arbusow ns has also
been used to prepare dialkyl methanephosphonates.
Dimethyl methanephosphonate is obtained in 95 per
cent yield by heating trimethyl phosphite with
SECRET " FLLOROPHOSPHATES AND PIlOSPHORtS-CONT VIM NO COMPOl' \DS
methyl iodide,105* whereas a similar reaction using
tributyl phosphite yields 89 per cent of dibutyl
methanephosphonate.24
A novel process well suited for conversion to plant
scale operations has been developed on a laboratory
scale for the synthesis of the ethyl analog of MFI.
In this process tetraethyllead is allowed to react un-
der nitrogen with phosphorus trichloride to give 89
to 96 per cent of the theoretical yield of ethylphos-
phorus dichloride, which is then converted in 85 to
95 per cent yield to ethanephosphoryl chloride by the
action of sulfuryl chloride. Treatment with sodium
fluoride and isopropyl alcohol converts this substance
into isopropyl ethanefluorophosphonate in 72 to
85 per cent yield. The fust two steps can be carried
out in the same vessel.” The resulting ethyl analog
of MFI has about three-fourths the toxicity of MFI
itself. No attempt has been made to synthesize MFI
by a similar process using tetramethyllead, w hich is
reputed to lx* much less easily handled than tetra-
ethyllead.
Pilot plant production of MFI has not Ireen under-
taken in this country. The efforts of the Germans to
produce this substance on a plant scale were not suc-
cessful. Although intermediates for the material were
made in substantial quantity (300 tons of dimethyl
hydrogen phosphite, 5 to 10 tons of dimethyl meth-
anephosphonate, and 1 to 2 tons of methanephos-
phoryl chloride were produced), not more than }/'i ton
of MFI itself was produced.73-111 Corrosion ap(>eared
to have been the principal source of difficulty. Equip-
ment shortages necessitated the use of resin-coated
equipment where stainless-steel or glass-lined equip-
ment would ordinarily have been used. Silver-lined
equipment was resorted to in some cases.72 73
MFI is a colorless, almost odorless liquid broiling
at 59 C at 8 mm of mercury. Its volatility at 25 C is
16.4 mg/1.*9" It is less stable than MCE, but can be
stabilized by the addition of 0.5 per cent of diethyl-
amine.
9.2.2 Chemical Reactions, Detection,
and Analysis
Studies on the chemistry, detection, and analysis
of phosphorus compounds as candidate chemical war-
fare agents have Ireen limited almost exclusively to
PF-3, certain of its close relatives, and MCE.
Dialkyl Fluorophosphates
Solutions of PF-1 in 0.9 per cent saline lose virtu-
ally all toxicity in 3 hours. This deterioration is re-
tanled by buffering the solutions near neutrality but
is markedly no (‘derated by buffering at /d 1 9.7.'®*'
PF-3 is hydrolyzed slowly at room temperature by
water to give fluoride ion and diisopropyl phosphoric
acid. This hydrolysis is less than 50 i>er cent com-
plete in 15 hours and is still incomplete after 25
hours.* In neutral aqueous solutions at body temper-
ature the half-hydrolysis time is about 9 hours.46 In
2 per cent aqueous alkali PF-3 is rapidly hydrolyzed
at room temperatures, although more concentrated
alkalies appear to retard this hydrolysis.IOSe his( I )i-
methylamido)phosphoryl fluoride appears to be con-
siderably more stable to hydrolysis than PF-3.**
In contrast to the ease1 with which fluoride ion is
freed by aqueous alkalies, the isopropyl groups of
PF-3 are very resistant to alkaline hydrolysis. For
example, no isopropyl alcohol can be detected after
refluxing with 10 per cent sodium hydroxide for
72 hours.*'* Advantage is taken of this resistance to
hydrolysis in several of the analytical procedures for
PF-3 based oir determination of fluoride ion, the
titration of which is interfered with by phosphate ion
but not by alkyl phosphates.7,1*-1*51
The kinetics of hydrolysis of PF-3 have been stud-
ied in several laboratories.In addition to the
marked catalysis by alkali already noted, the reaction
is also acid-catalyzed, and thus m pure water is auto-
catalytic. In buffered solutions the hydrolysis is
pseudomonoraoleeular. The observation of a pro-
nounces! acceleration by phosphate ion suggests that
the decomposition may be subject to general base
catalysis as well as acid catalysis, although acetate
ion is the only other anion which has been observed
to have an accelerating effect.20
When hydrolysis of PF-3 is allowed to proceed in
acid solutions, the course of the react ion may become
complex. For example, in some experiments, acetone
and isopropyl phosphorous acid were formed in addi-
tion to fluoride ion, and no phosphate ion could Ik*
detected. Other dibasic acids were likewise absent.
Acetone is also formed when acid solutions of diiso-
propyl phosphoric acid are treated with sodium flu-
oride. It has not always been possible to reproduce
these experiments, however, and the mechanism by
which acetone and isopropylphosphorous acid are
formed is not yet clearly understood.20
PF-3 does not react with sodium hypoiodite to give
iodoform and does not react with thiosulfate ion.s,b c
Methods for the detection and analysis of com-
pounds of the fluorophosphates series are summa-
rized in Chapters 34 and 37. The following general
SECRET SYNTHESIS AND PROPERTIES
143
remarks may be supplemented by reference to these
chapters.
1 he fluorine atom of PF-3 and related compounds
is readily converted to fluoride ion on hydrolysis and
any detection methods depending upon (he recog-
nition of fluorine ion are thus applicable to these com-
pounds. The ability of fluoride ion to bleach metallic
lakes of certain dyes or its etching effect on glass has
l>een utilized for recognition.14 41 •42-5s 04 M10*c
A device making use of the etching effect has I>een
examined by (he British.95-9®07
The decomposition of volatile fluorine compounds
by hot platinum filaments or hot platinized silica gel
to produce hydrogen fluoride is applicable to mem-
Imts of the fluorophosphate series.14-M
Detection of PF-3 collected upon plain silica gel
tulies can lie accomplished by testing either for
fluoride ion or for phosphate ion after suitable treat-
ment. The DB-3 reagent may also be used.13
Chemical methods for the detection of fluorine
compounds, including PF-3, in water have been de-
veloped.4142 Use of the miosis produced by PF-3 as a
method for detection of this agent in water has also
been proposed. It is claimed that 25 to 50 ppm can
bn detected in 3 minutes by this method without
injury to the eye.40
The analysis of PF-3 has been accomplished by
volumetric, colorimetric, or gravimetric determina-
tion of the fluoride ion produced by alkaline hydroly-
sis, Alternately, phosphate ion can l>e determined
colorimetrically after vigorous acid hydrolysis with
hydrobrornic, hydriodic, or sulfuric acids.7-*’50-80-81 10"
10ie,t,10«f
Methods suitable for use in field and chamber
analyses of PF-3 have been descril>ed.3 5,161
Ethyl 1)imkthvlamidocvanuhhosi*hate (MCE)
MCE is readily dest royed in either acidic or basic
solutions.*1-"2 In alkaline solutions, cyanide ion is
liberated rapidly even in the cold, the half life at 25 C
twang 5 minutes at pi I 8.5 and 30 minutes at pH
7.5.61 In acid solutions rapid liberation of dimethyl-
amine occurs, the half life in solut ions of pi I 1 being
2 minutes, that in solutions of pH 3, 90 minutes. The
substance has maximum stability at pH 4.5, where
its half life is 7 hours with respect to both cyanide
ion formation and dirnethylamine liberation. Solu-
tions of maximum stability result from hydrolysis in
unbuffered solutions, since the hydrolysis products
are acidic and self-buffering in the range pH 1 to 5.*1
In solutions of high acidity (i.e., 3 normal), hydro-
gen cyanide as well as dimethylamine is liberated
rapidly but complete degradation to phosphoric acid
results only from boiling the substance with mineral
acids.1 “
Bleach and chlorinating agents react readily with
MCE to yield CK.70112
MCE is extremely hygroscopic, and moist solu-
tions of it slowly liberate AC."1-6*1! Its faint fruity
odor cannot be relied on for detection.91 1,4 Its median
detectable concentration as determined with the
osmoscope is 2.2 pg l.Mh
The standard liquid vesicant detectors, l»oth Brit-
ish and American, give positive reactions with MCE.
This is true of the II papers of the kit, food testing,
and of the M-6 paper, M-7 crayon, and M-5 detector
paint of the United States Chemical Warfare Service,
and of the British Detector, Gas, Ground. The Brit-
ish differential detector powder gives a yellow color
with the agent.***-70-112 The black dot (AC) tube of
the M-9 detector kit has about the same sensitivity
for MCE vapor as it has for AC itself (20 pg) but is
considerably less sensitive than the German AC tula*
(sensitivity 2-3 gg). The red dot (nitrogen mustard)
tube gives a nonspecific test.70 The British pocket
vapor detector gives no reaction with the agent.112
The ready production of cyanide ion and a volatile
amine on alkaline and acid hydrolysis, respectively,
together with the production of phosphate ion on
ultimate hydrolysis, can be taken as confirmatory
identification."2
For field or chamber analysis, MCE can be col-
lected in 1.25 normal sodium hydroxide and titrated
with silver nitrate,23 62 *** or (for small amounts) esti-
mated colon metrically with sodium pic rat e.23-6*k -6*1
Phosphorus colorimetry using molybdivanadophos-
phate is also suitable if the sample, collected in al-
kali. is fumed with perchloric acid or otherwise
completely decomposed. The sensitivity of this
method is several times as great as that of those
already described.23 62 Attempts to adapt the DB-3
method to the analysis of MCE have not been en-
tirely successful.6*11
9.2.3 Stability
1 )IALKVL FH OKOI’HOSPHATKS
PF-3 is stable when stored in glass at 25 C. When
stored in steel at 65 C, slight decomposition lakes
place as indicated by sludge formation. This decom-
position continues at an increased rate when the
sample is removed and stored in glass at 25 C. This
SECRET 144
FLUOROPHOSPHATES AND PHOSPHOIU S-CO\T\[M\C COMPOUNDS
effect may be due to the action of light and dissolved
iron salts.*
In the presence of steel at 58 Gl) C, diethyl fluoro-
phosphate appears to stable for several months.'-8®
Both PE-1 and PE-3 are resistant to flashing. No
temperature has been found at which dimethyl Hu-
orophosphate flashes; PE-3 can l>c made to Hash
feebly over a narrow temperature range.-60
Ethyl Dimkthylamidocyanophosphate (5ICE)
Technical MCE containing 20 per cent mono-
chlorobenzene is reported by the Germans to be
stable even on prolonged storage.”
It is also claimed by them that MFI, when stabi-
lized with < 1 percent of diethylamino, can be stored
in iron anti that it is stable in methanol solution. It
was supposed to have been used in such solutions.71
9.2, t Decontamination
Dialkyl Elcohophosphates
Bleach suspensions and dry bleach react vigor-
ously with PE-3 and presumably with other Huoro-
phosphates, and normal field decontamination, pro-
cedures as used for vesicants should be effective. The
chloramides S-4GJ ami S-32S do not react with
PE-3 or diethyl Huorophosphate, nor do dilute solu-
tions of calcium hypochlorite.3 -" 04
The ease of hydrolysis of the fluorine atom of the
dialkyl fluorophosphates by water alone varies con-
siderably with structure. PE-1 is 72 per cent hy-
drolyzed after standing I hour in water at 21 C; the
diethyl compound, 21 percent; and the PE-3, 1 per
cent. However, dilute alkalies at room temperature
produce rapid hydrolysis of all three esters.3 Lime
slurry should thus be an effective decontaminant.
Dilute solutions (approximately 0.4 per cent) of
sodium hydroxide have been proposed for skin de-
contamination.
Mere hosing of contaminated areas with water
should mitigate the vapor hazard produced by PE-3,
since it is soluble to the extent of 1.5 per cent in
water.04
Ethyl Dimethylamidocyanophosphate (MCE)
In the Dyhernfurth plant of the Germans, equip-
ment used for the synthesis of MCE was decontam-
inated by steam and ammonia. Surface decon-
tamination, in the absence of steam, was done by
solutions of ammonia or of amines.72
Alkalies or bleach and water have been recom-
mended by the Chemical Warfare Service for decon-
tamination, but it is recognized that the production
of CK by flip action of bleach on MCE might prove
hazardous under some conditions.70
1.2.5 Protection
Dialkyl Fliorophospiiatks
Adequate protection against dialkyl Huorophos-
phates appears to be provided by United States.
British, German, and Japanese canisters, and it is
doubtful whether canister [>enetration by these
agents will over be a significant problem. Repre-
sentative United States, German, and Japanese
canisters have been tested against PE-1, PE-3, and
methyl ethyl Huorophosphate; ail afforded good pro-
tection.57 The standard United States Navy can-
ister provides complete protection against PE-3 as
does the British l.t. Mk. II canister.75111
Ethyl D im kth v l a m i di>cy a nophi»e11ate (MCE)
Completely adequate protection against the vapor
of MCE isafforded by American, British, and German
canisters 62 70 72 90 "2 and it is implied in intelligence
reports that the German canister gives adequate
protection against MEL72 American canisters
(M-ll and M-lOA-1) give adequate protection
against ethyl dimethylamidocyanophosphate as an
aerosol (particle size 2 w, concentration 100 Mg 1,
How rate 32 1pm), but the Canadian canister, which
has a resin wool pad-type filter, allows serious }>ene-
t rat ion after 5 minutes.70 It is to be noted that the
tactical use of the agent contemplated by the Ger-
mans was as an aerosol.
Combined activated carbon-aeration treatment of
water contaminated with MCE gives excellent re-
moval of cyanide ion, odor, and color but does not
remove organic phosphorus if the water has been
standing more than 15 hours after contamination.6®' k
9.3 TOXICOLOGY
0.3.1 Detectability by Odor and Other
Physiological Signs
The Triions and fluorophosphates may be detected
by (1) odor, (2) a feeling of tightness in the chest
and or throat, and (3) pupillary constriction.
MCE and PE-3 have faint, sweetish odors. The
available osmoscopic data for these and other agents
are presented in Table 2. It is apparent that the
Huorophosphates are relatively . odorless. Crude
MCE (German shell filling) is more readily detected
but does not possess so pronounced an odor as II.
MFI is said to be odorless, or practically so.10"
SECRET TO\IGOLOCV
145
Table 2. Detectability by odor of MCE,
phates, and other representative agents as (let:
the osmoscopic leeliniqiie.
flnorophos-
ermined by
Median
detectable
Agent
cone, (gg 1)
Reference
MCE (German shell filling)
2.2
66h
PE-3
36
lit
Dimethyl fbiorophosphate
IS
49
1 bet hyl fluorophospbatc
la
49
II (plant-run Ix'vinstein)
Of.
51
11 (pure thiodiglycol)
1.8
65
HN3 (plant run)
to
66m
AC
34
39
CG
1.4
38
mask donned before a large dosage had reached the
eyes or lungs. Sufficiently low concentrations to
escape these means of detection would he revealed
after some minutes or an hour by pupillary constric-
tion, and the mask applied if more prolonged ex-
posure were unavoidable. Thus, except upon very
sudden exposure to high concentrations of vapor and
aerosol, dosages sufficient to produce systemic ef-
fects would seem to be theoretically avoidable. It is
much more difficult to detect exposures to small
dosages sufficient to produce miosis and the other
harassing but not disabling symptoms descrilted in
Section 9.3.2, and it is in this sense that MCE and
PF-3 may be considered insidious. Accidental ex-
posures to undetected dosages that resulted in these
symptoms are reviewed in the next section. It has
lx*en emphasized that PF-3 is readily absorbed by
lacquer, rublxT, clothing, and hair. The gradual de-
sorption of vapor can result in obtaining, within con-
fined spaces, concentrations which suffice to produce
eye effects but which may remain undetected until
these effects appear.*7
The lack of odor of MFI may not prove to be so
great an advantage as would appear at first sight if
throat irritation and feeling of chest constriction
should prove to be definite indications of the inhala-
tion of very low concentrations.
9.3.2 Eye Effects
The vapors of the fluorophosphates and Trilous
are absorbed directly by the eye and produce con-
traction of the pupil (miosis) and interference with
the muscles of accommodation. As a consequence
harassment due to poor dim light vision in dim light
and to pain and difficulty of focusing is experienced. A
potentially dangerous congestive iritis can develop
and pain behind the eyeball frequently becomes very
severe. These ocular symptoms can l>e relieved by
(repeated) instillations of a mydriat ic (e.g., atropine)
but the subject is left with a dilated pupil and para-
lyzed accommodation. The concomitant systemic
effects often include a feeling of tightness in the chest,
nausea, and vomiting. No data are available concern-
ing the exposure of human subjects to dosages suffi-
ciently large to produce more severe disability.
Studies ox Aximals
Tests of the effects of dialkyl fluorophosphates on
the eyes of animals originally served (I) to demon-
strate beyond reasonable doubt that cautious trials
with human volunteers (see the next section) could
In man-charnber experiments a German shell fill-
ing containing MCE with 20 per cent monochloro-
benzene was detected at a concentration of 1.6 Mg 1
by 2 of 10 subjects.M The pure agent seemed to be
more odorous and was detected at a concentration
of 0.35 ag 1 br each of 4 subjects.'18 It is possible that
the Germans considered decreased detectability by
odor to be one advantage of the addition of mono-
chlorobenzene to MCE. In one of the man-chamber
experiments with PF-3, concentrations of 37 to 70
Mg 1 remained undetected by odor.42 In a field (an-
nulus) test PF-3 could lie detected at an average
concentration of 0.5 Mg 1 but the odor waanorisUffl-
ciently characteristic to t>e easily identifiable.
Man-chamber experiments indicate that throat
irritation and a feeling of tightness in the chest are
apparently more sensitive indicators of exposure to
MCE and PF-3 than are the odors. In the case of
MCE, 6 of 10 observers exposed to the German shell
filling at a concentration of 1.6 gg I experienced the
feeling of chest constriction, as did each of, the 4 who
were exposed to 0.35 Mg I of pure MCE.** In the case
of PF-3 each of 18 subjects exposed to 8.2 gg 1 —
not detected by odor — experienced throat irritation
and a feeling of chest constriction within 60 to 90
seconds.7*
Pupillary constriction to pin-point size1 develops
within a matter of minutes upon exposure to mod-
erate dosages of MCE and PF-3, although it is longer
delayed at the minimal effective concentrations (see
the following paragraph).
The foregoing suggests that troops having masks
available could protect themselves against danger-
ous dosages of MCE and PF-3 if they could take
note of odor, feeling of chest constriction, and pupil-
lary size. High concentrations could lx* detected
quickly by odor or chest and throat signs, and the
SECRET 146
FLl OROFHOSFH VTF.S AM) FHOSFHORIS-COXTAIMAG COMFOl NDS
be carried out without risk of causing permanent eye
damage, and (2) to determine the relative miotic
potencies of some of the compounds.
Various observations have demonstrated that
pupillary constrict ion is produced in rabbits and mon-
keys at dosages considerably smaller than those re-
quired to cause permanent ocular injury or marked
systemic cffects.2*r *®* Ma'49 79 10u,h The factor of
safety in the case of PF-3 is most strikingly illus-
trated by experiments with rabbits. Instillation into
the conjunctival sac of a nearly lethal dose of the
liquid (i.e., 1.15 mg kg) and repeated instillations of
smaller doses, while eliciting intense miosis, lacrima-
tion, and a transient increase of intraocular pressure,
caused no permanent ocular injury.*** Similarly, al-
though vapor dosages of less than 1.000 mg min m*
sufficed to induce marked pupillary constriction,
dosages of 15,000 mg min m* caused no permanent
damage.*®*10411 In the case of both PF-1 and PF-3
the vapor dosages necessary to produce miosis in the
rabbit are considerably smaller than those required
to kill.104* With the monkey, a species that is excep-
tionally sensitive to the lethal actions of PF-3 and
di-sec-butyl fluorophosphate, the difference between
dosages producing miosis and serious systemic poi-
soning may be smaller,-’8"1 p q -r■M *4 as may also he the
ease with man (see next section).
Tests with rabbits have demonstrated that PF-3
is a markedly more potent pupillary constrictor than
are (he dimethyl, diethyl, dipropyl, or diallyl es-
ters.49'0411 It not only produces constriction at lower
dosages, but also for longer times.49 7M04b Illustrative
data are presented in Table 3.
ating the relative potencies of this compound and of
PF-3 ait* not available. Dicyclohexyl fluorophosphate
also appears to be an effective miotic that produces
pupillary constriction after a somewhat greater la-
tency than characterizes the compounds just men-
tioned.'041 Its potency relative to that of PF-3 is not
known.
The high miotic potency of MCE is illustrated by
observations on animals 69,-*s but quantitative com-
parisons with the fluorophosphates are not available.
At high doses MCE can produce conjunctival hemor-
rhages. **
Observations ox Human Subjects
MCE appears to l>e considerably more potent in
producing eye effects than any of the_fluorophos-
phates.** Although a dosage of 0.7 mg min m* (t =
2 was without effect on the eyes. 3.2 mg
min m* it — 2 minutes) produced slight but definite
miosis. Dosages of I t to 21 mg min m5 produced a
seven* harassing effect of several days’ duration. The
action of these* dosages was characterized by the fol-
lowing symptoms, not all of which were observed in
all the subjects: pin-point constriction of the pupils,
lasting for several days; severe frontal headache;
retrobulbar pain, tightness in (he chest, and cough-
ing; pain on focusing on near objects; slight blurring
of l>oth distant and near objects; slight blurring of
peripheral visual fields; nausea and vomiting; en-
gorgement of the bulbar conjunctival, anterior cili-
ary, and radial iris vessels, and of (he vessels at the
base of the iris; acute ciliary tenderness; and fall in
intraocular tension. This symptomatology was usu-
ally almost completely relieved within an hour after
the instillation of either atropine or hyoscine solu-
tion, but the effects of the treatment did not persist.
In the absence of treatment the symptoms became
most harassing 24 to 4S hours after exposure and
persisted in gradually decreasing intensity for several
tiays thereafter. In the case of one observer exposed
to 30 mg inin/m* (/ = 10 minutes), the harassment
was very severe and, in addition to the effects men-
tioned above, visual acuity was markedly reduced
and had not returned to normal 17 days after ex-
posure.
Data on the eye effects of MFl are not available.
PF-3 produces symptoms similar to those caused
by MCE but is definitely less potent. From the data
presented below it would appear that exposure to
10 mg min up of PF-3 vapor produces about the
same effects as exposure to 3 mg min/m* of MCE
Table 3. Relative miotic effects of several dialkyl fluoro-
phosphates in rabbits."’41.
The animals were exposed for 3 minutes to nominal con-
centrations of 1/50,000 (0.11 to 0.16 nig/1).
Dialkyl ester of
fl iiorophospln >ric
acid
Average prr cent of initial
pupil diameter
After After After
10 min 100 min 300 min
Dimethyl (PF-1)
32
82
100
Diethyl
27
58
85
Dipropyl
45
68
96
Diisopropyl (PF-3)
16
31
52
Diallyl
27
46
67
That di-scc-butyl fluorophosphate is a very potent
miotic is revealed by the production of marked pupil-
lary constriction within 10 minutes after the exposure
of monkeys to 50 mg min m* (I = 2 minutes).*5
Animal data adequate to provide a basis for evalu-
SECRET TOXICOLOGY
147
vapor. At larger dosages, 200 100 mg min/m* of
PF-3 may correspond roughly to 14 to 20 mg min/m*
of MCE,
PF-1 and diethyl fhiorophosphato are definitely
less potent miolies than PF-3. 6/.s( Dime thy lamido)-
phosphoryl fluoride is also less potent, probably
much less so. Di-sec-butyl fluorophosphatc appears
to he somewhat more potent than PF-3 but defi-
nitely less potent than MCE.
The observations on which (he above statements
tire based may lx* abstracted as follows.
1. Ethyl dimelhylamidocynriophosphate (MCE). The results
of one controlled laboratory study are available.*8 In addition
there have been accidents which demonstrate that exposures
to undetected concentrations of (he vapor can produce ex-
treme pupillary contraction and in addition congestion of the
eyes.**!'51 In other instances a feeling of tightness in the chest
has accompanied and given warning of the exposure.
Four subjects exposed in a man-chamber to 0.7 mg min nf1
(I = 2 minutes) detected the odor of the agent and cxjierienced
a brief feeling of tightness in the chest. They developed no
miosis.
Ten additional subjects were exposed lo a dosage of 3.2 mg
min /m5 (I = 2 minutes). Only two noticed any smell. Six ex-
jiericnced a very slight feeling of constriction in the chest.
Slight miosis develop'd in all after 140 to GO minutes.
Ten subjects, some of whom had liccn exposed 4 hours
previously in the preceding group, were exposed to 14 mg
min/m* (I = 2 minifies). The gas was detected faintly by
smell and those not previously exposed felt a slight tightness
in the ehest. Soon after exposure all subjects had contraction
of the pupils which persisted for 48 hours. Severe headache
and pain in the eyes followed unless atropine was administered.
Vascular injection of the eyeballs was present. Difficulties of
focusing were experienced. Vomiting on the day after exposure
occurred in four of the subjects.
Three additional subjects were exposed to 14 mg min/m*
(t = 10 minutes). The odor and a feeling of tightness in the
chest were detected. Pupillary constriction, headache, rhinor-
rhea, nasal congestion, and other symptoms developed rapidly
and jiersisted for several days in the aliscnce of treatment.
Visual acuity at moderate illuminations was not markedly
affected.
Five additional subjects were exposed to 21 mg min/m*
(I = 10 minutes). They became severely harassed by (he
symptoms (hat developed. The symptoms and their limes of
onset (minutes, in parenthesis) were tightness in the chest
(1.5 to 8), coughing (1.5 to 0), pin-jsiint pupils (10), lamina-
tion (2 lo 10), retrobulbar pain (8 to 19), conjunctival con-
gestion (2 to 10), “tingling” of the eyelids (6 to 10), rhinor-
rhea (fi to 120), frontal headache (13 lo 18), difficulty of seeing
distant objects (11,14 — two cases), difficulty in seeing near
objects (15 — one case), and constriction of the |ieripheral
visual fields (15 — one case).
One subject with one eye protected was exposed to 30 mg
min in5 (I = 10 minutes). The protected eye was unaffccted-
The pupil of the exposed eye began lo contract within 4 min-
utes and had become fully contracted within 12 minutes.
Visual acuity in dim light had markedly deteriorated within
an hour and had not fully recovered 17 days later. Moderate
conjunctival and severe ciliary congestion had develop'd
within 3 hours. The subject was unable to sleep for two nights
because of severe pain above and behind the exposed eye.
2. Dimethyl fluoraphosphate (PF-t). At low dosages PF-1
is not so potent a harassing agent as PF-3, nor does the pupil-
lary constriction which it induces persist as long. Although
this ester is considerably less readily detected by odor than
PF-3, it is more irritating to the throat and chest. In subjects
expost“d to nominal concentrations as low as 5.7 jig/1 (1/10*)
it producer! a tightening sensation in the throat.111** No eye
effects were noted when subjects were exposed, presumably
for short times, to this concentration or to one four times as
great.
At a considerably higher concentration, li t 1 (1 50000),
the throat, sensation was not more marked but eye effects were
produced: an exposure of 30 seconds’ duration (Cl - 57 nig
min ni5) produced in five of seven subjects some pupillary
constriction and discomfort but no spasm of the muscles of
accommodation; exjmsures of 1 to 5 minutes’ duration
(Cl = -111 to 570) produced within 5 to 10 minutes pupillary
constriction lasting for an hour or more, and, in 60 jier cent
of the subjects, a marked spasm of the muscles of accommo-
dation.
3. Diethyl flnorophnsphale.'3 This ester also apjiears to be
considerably less potent than PF-3. In twelve subjects 2-min-
utc exposure to a nominal concentration of 139 eg/l (Cl —
278 mg min m*) produced throat irritation within 10 to
30 seconds, then a painless tightening sensation in the chest,
and finally coughing toward the end of the exposure. Within
30 to fiO minutes the pupils had partially contracted and their
reflexes to light and accommodation were absent. There was
no significant alteration in visual acuity in daylight or in sim-
ulated twilight, although the sensitive Rangefinder Test
revealed harassment. The size and reflexes of the pupil had
returned to normal within 18 hours. At no time was there more
than minimal congestion of the iris in any of the subjects.
4. Diisopropyl fluorophmphate. (PFS).**'’ *•• ,8- *’• s:-:*-
71, M. 10)t, lute
a. Find (preliminary) British examination.'"* Ten minutes
after exposure of two subjects to a nominal dosage of
24fi mg min m* (0082 mg 1 for 3 minutes), the pupils lie-
gan to constrict and subsequently were reduced to pin-
point size, with the result that the lalwratory appeared
dim. The olwervers exjicrienccd difficulty and pain in focus-
ing, eye ache, and headache. A I wok could lie read only if
held within a few inches of the eye. The miosis and diffi-
culty of accommodation jiersisted for 2 to 3 days in the
case of the older volunteer (over 60 years of age) and for
almost a week in the younger (28 years). The report does
not mention extraocular symptoms.
Upon exposure of two additional subjects to a nominal
dosage of 82 mg min 'nr1 (0.0082 mg I for 10 minutes) the
effects did not develop for about 30 minifies but then ap-
peared as descrllied above and persisted for 3 days. The
subjects could read only with pain and difficulty. Vision in
dim light was poor. Distant vision was. impaired but re-
covered sometime liefore near vision had returned to nor-
S ECU FT F LUO R OPII OSH 11 AXES AM) PHOSPHORUS-COATAIMING COM POL M>S
mal. The eyes of one observer were congested for about a
day beginning I day after exposure.
b. Preliminary American observations.**-33 1 pon exposure of
only the eyes of several subjects to a nominal dosage of
approximately 300 mg min m* (0.1 mg 1 for 3 minutes)
only one subject reported slight subjective eye irritation
during exposure. Miosis became maximal within 20 to 30
minutes and persisted for about a week. The subjects ex-
|s‘iienced difficulty in accommodation and found reading
painful during the first 2 days; they had less difficulty after
1 to 2 days in spite of the maintenance of pupillary con-
traction and the development of irritation (congestion),
eye ache, and headache. There was only a slight decrease in
far vision; Snellen charts could lie read ulxiut as well as be-
fore exposure. The subjects reported that their night vision
was poor. Atropine and adrenaline instillations gave relief
but had to lie repeated daily. In two men accidentally ex-
cised for several hours to low and undetected concentra-
tions. a viewed object first appeared clearly but then
rapidly liecame blurred, accommodation was slightly re-
duced, and nearsightedness apparently increased.4’
Four men whose eyes only were exposed to approximate
dosages of 111 to 210 mg min m3 (0.037 0.070 mg 1 for
3 minutes) detected no odor and experienced no discom-
fort during exposure. They subsequently developed miosis,
difficulty of focusing and blurred vision, eye ache and head-
ache, conjunctivitis and a gritty sensation in the eye, and
twitching of the eyelids. Visual acuity as tested by Snellen
charts was not reduced. Four men accidentally exfwsed to
low, undetected concentrations experienced eye effects as
just described and, in two cases, nausea. There is some evi-
dence, that in two cases visual acuity in dim light was
reduced.53
c. Scrawl British examination.7* Subjects were exposed to
nominal concentrations of vapor in a large man-chamber
and subsequently examined for pupil size, pupillary re-
flexes to light and accommodation, and acuity of near and
distant vision iis tested with Jaeger and Snellen Test Type
indices both in daylight and in simulated twilight (approxi-
mately 0.4 footcandle).h The general condition of their
eyes was also examined and their performance on the
Rangefinder Testc in ordinary daylight and simulated
twilight b determined.
All six subjects exposed to 41 mg min/m3 (0.0082 mg 1
for 5 minutes) complained of throat, irritation about 1 min-
ute after the start of the exposure and of “tightness in the
chest” after about 1.5 minutes. Three hours later (he pupil
was only slightly contracted and pupillary reflexes to light
and accommodation were present and normal. The tests
for visual acuity revealed no significant deterioration
either in ordinary light or in simulated twilight. The aver-
age degree of harassment for the group as measured by the
Rangefinder technique was 36 |>er cent in ordinary light
and 50 per cent in simulated twilight. At no time was there
congestion of (he iris and no subject experienced headache
or other discomfort.
I'pon exposure to 00 mg min in3 (0.033 mg 1 for 3 min-
utes) all of 18 subjects experienced throat irritation after
about 50 seconds of exposure and complained of “tighten-
ing of the chest” within about 1.5 minutes. It took 4 to
6 hours for maximal miosis to develop, at which time pu-
pillary reflexes were absent. The tests for visual acuity (as
described) revealed no significant change although the
Rangefinder test indicated It |«*r cent harassment with
ordinary lighting and 100 jw-r cent in simulated twilight.
Congestive iritis with accompanying headache and photo-
phobia developed within IS to 21 hours. Atropine sulfate
(1 per cent) proved more effective than homatropine (I jier
cent) in dilating the pupils and relieving the iritis.
Of 12 subjects excised to 328 mg min m* (0.164 mg I
for 2 minutes) all experienced throat irritation and a feel-
ing of tightness in the chest within 30 to 105 seconds. Them
was also some coughing, but no eye irritation, Incrimalion,
or blepharospasm occurred. The pupils were only partially
contracted 30 minutes after exposure but bad contracted
nearly to pin-point size within 3 hours. Pupillary reflexes
were then absent. A definite deterioration in acuity for dis-
tant vision bad develo]>ed within 30 to 60 minutes after
exposure and was not notably more marked when tested
in simulated twilight (see preceding paragraph) than when
tested at higher levels of illumination. The individual alter-
ation in near vision was variable til'both tested levels of
illumination but for the group as a whole there was definite
deterioration. Twenty-four hours after exposure distant
vision had improved but near vision had deteriorated fur-
ther. The subjects without exception complained of head-
ache, the pain being referred to above or behind the eyes
and being sufficiently intense to interfere with sleep. There
was well-marked congestive iritis and conjunctival con-
gestion but no edema of (lie lids, conjunctiva, or cornea.
The average degree of harassment for the group as meas-
ured by the Rangefinder technique was 63 |ior cent in or-
dinary light and 100 jier cent in simulated twilight.
Granted that the symptoms caused some discomfort and,
at night, visual harassment, they were not considered to lie
of a disabling nature in the recorded opinion of the British
Ophthalmic Panel and Medical Subcommittee.""
d. Third British examination,84 The eves only of sixteen sub-
jects were exposed for 5 minutes in a const ant-flow device
to analytically determined dosages of 40 to 250 mg min/m3.
Subsequent clinical examination included observations on
pupil size, visual acuity at high and relatively dim illumi-
nations, the near point of accommodation, the threshold of
scotopic vision, and the condition of the conjunctiva,
cornea, and iris.
In summary, with dosages up to 101 mg min/m3 the
effects produced by the vapor on pupil size and (he accom-
modative mechanism were not considered of serious con-
sequence. However the three subjects exposed to 250 mg
min m3 developed a congestive iritis associated with pain-
ful symptoms and consequently were considered to 1k>
partially disabled for 3 to 6 days.
The pupil had contracted to a minimum diameter of 1 to
2 mm within less than 1 day at all dosages and within I
hour at dosages above 116 mg min/m3. The miosis l«‘gan
'• This level of illumination was far greater than would l>c
encountered at night and the tests therefore give no adequate
measure of the handicaps which the subjects would have
experienced in night fighting.
r This technique has been dcscrilHMp* and critically dis-
cussed."1’
SECRET TOXICOLOGY
to abate after 2 days at the lower dosages and after 3 days
at the higher ones, but was not completely relieved for 5 to
K days. During the first days prolonged dark adaptation
resultixl in no pupillary dilatation.
Visual acuity tested with Snellen charts in bright light
(17 footcandles) showed practically no deterioration. In-
deed, the uncorrccted vision of myopes was improved, as
a result of the small pupil size. Visual acuity in relatively
dim light was determined by lowering the illumination of
tho Snellen chart until the smallest type which the subject
could read at high illumination was no longer legible. Be-
fore exposure the average illumination recorded for the
twelve eyes of six observers was 2.8 footcandles (range
0.32-10.5). Twenty-four hours after exposure to 40 or
< 80 mg min/in 5 it was 0.4 footcandles (range 0.32-17).
This change was not considered consistent or marked It
must Ik- emphasized that the tested range of illuminations
was sufficiently high that cone (not rial) vision was living
measured, and that the results throw no light on the im-
pairment which may have been produced in night vision.
Among subjects exposed to 40 to 191 mg min m* the
near |x»int of clear vision was brought in, indicating in-
creased ciliary tension. However the absence of serious im-
pairment of distant vision indicates that any existing
spasm uf the muscles of accommodation was not a serious
handicap at the light intensities employed in the tests.
From the changes in pupil size caused by the PF-3 vapor
one might have expected as much as a 16-fold rise in sco-
topic visual threshold. Actually the change in threshold
brightness level, as measured with the Craik Adaptometer
1 to 2 hours after exposure to 116-191 mg min/m*, was only
2- to 10-fold (average 5+ fold in six subjects).
Subjects exposed to 100-191 mg min m3 generally de-
veloped conjunctival hyperemia 2 to 3 days after exposure.
At 250 mg min/m* the hyjiercmia was much more severe
and developed within 24 hours. In addition a marked and
potent ially dangerous congestive iritis, accompanied by
painful symptoms, made its appearance.
Examination with the slit lamp revealed no corneal
changes, nor were changes noted upon opt halmoseopic ex-
amination in instances where miosis had been abolished
with a mydriatic.
Among the subjective symptoms reported by the sub-
jects were mistiness before the eyes, eye ache, and diffi-
culty of seeing in the dark.
Instillation of homatropinc (0.43 minim) had to lie re-
peated three times at hourly intervals in order to obtain
significant pupillary dilatation in two observers who de-
veloped congestive iritis following exposure to 250 mg
min m*. After the third instillation the congestive symp-
toms were relieved but paralysis of accommodation oc-
curred and the observers became partially disabled
because of blurry vision,
c. American assessment.^• ” -*71’.>■ One subject was exposed
in a man-chamlier to a dosage of 181 mg min m* (I = 0.7
minutes), eight subjects to 165 mg min in* (/ = 8.7 min-
utes), one subject to 2140 mg min in* (I — 10.7 minutes),
and six subjects to 244 mg min m* (1 = 9 minutes).
All the men exposed to 165 mg min m* experienced a
slight feeling of tightness and constriction in the chest,
apparent 1 2 hour after exposure and particularly noticcable
several hours later. Instances, of rhinorrhea, diarrhea,
nausea, and vomiting (one case) occurred but, except for
the rhinorrhea, may not have been due to the effects of the
PF-3. Xo muscle tremors —a sign which might be ex-
pected to herald serious systemic poisoning were ob-
served.
Of the men exposed to 211 mg min m* (0.027 mg 1 for
minutes), five of six experienced a fleeting feeling of chest
constriction while in the chandler. This returned and per-
sisted for 2 days, being accompanied by coughing in two
cases. All the subjects developed rhinorrhea within an hour
after the exposure. Only one developed nausea, and he
vomited twice. There were no abdominal eramps or muscle
tremors. One volunteer exposed to 290 mg min in* de-
velops! constant nausea for a day following exjsisure, ex-
perienced abdominal cramps, and exhibited increased nasal
secretion, lie had no muscular tremors and felt no chest
constriction.
The majority of men in both groups had diminished dis-
tant vision, which was caused by a spasm of the muscles of
accommodation. Although the resultant false myopia
measured between 1.75 and 0.5 diopters, because of the
small pupil sine the visual acuity was not diminished
greatly. The greater part of the diminution of distant vi-
sion had developed within 45 minutes after exposure. Fur-
ther deterioration occurred at 3 hours in some cases. Re-”
covery occurred at 2 to 7 days, being slightly more rapid
in the subjects exposed to the smaller dosage than in those
exposed to the larger.
Maximal miosis developed within 10 to 15 minutes after
exposure. Among the men exposed to 105 mg min in3, the
pupils liegau to relax after 1 to 3 days and attained normal
size and activity after 3 to 9 days. Among lhose exposed to
244 mg min m* relaxation did not begin until after the”
third day and complete recovery required 5 to 11 days.
All the volunteers showed a diffuse conjunctival injec-
tion which required 5 days to clear up.
Concurrently with the development of pupillary con-
striction and spasm of accommodation, the near point of
accommodat ion moved to within 3 to 6 cm from the cornea,
and it became increasingly difficult for the men to focus
after gazing into infinity. Small type could be read but
several seconds were required before it could lie seen
clearly. Without exception the men complained of intense pain
when they attempted to perform visual tasks within IS inches.
Recovery of the untreated eyes gradually occurred over an
average of 3.5 days after exposure to the lower dosage and
4.5 days after the larger.
Except for one man who exhibited a transient rise in in-
traocular tension, all displayed a subnormal tension for
several days.
A performance test (in daylight) showed no decrease in
efficiency of marksmanship and all of the men felt that
they could competently discharge such military tasks as
guard duty, vehicle driving, and rifle firing.
The reports state that prolonged questioning failed to elieit
any symptoms of defective night vision, all the volunteers
feeling that their visual acuity at night was proportional
to (hat in the day. This statement is at variance with the
results obtained in other tests and observations. It would
not be anticipated that the vision of men with maximally
SECRET 150
FLUOKOPHOSPH \TES VXD PHOSPHORUS-CONTAINING COMPOUNDS
Compound
Mouse
HCl)ia in mg min/m3
(10 min nominal)
Uange for all
Monkey tested s|M‘eies
Increase of
/dm-with
increase of
exposure
time
LDiV
Rabbit,
intra-
venous
(mg/kg)
Mouse,
percu-
taneons
Kthyl dimethylamidocyanophosphate (MCE)
380
250
200 1,000
Slight
0.1 ±
1 f-
Isopropyl methanefluorophosphonate (MFI)
250
150
100-300
Definite
0.02
1 ±
Isopropyl elhanefluorophosphonate
330
200
150 700 ±
Definite
1.7
Dimethyl fluorophosphate (PK-1)
2,000
2,500- > 12,000
Marked
3
30
Diethyl fluorophosphate
8,200
7,000- > 11,000
?
35
Diisopropyl fluorophosphate (PF-3)
5,900
000
000 >8,000
Slight
0.4
72
Di-wr-hutyl fluorophosphate
5,200
250 ± 150
250->18,000
?
Dievelohewl fluorophosphate
1,100
i ,ooa 8,ooo
9
6(s(Dimethyiamiilo)phosphoryI fluoride
950
950- > 4,000
?
3 +
Table 4. Summary of toxicifics.
contracted pupils would l>e normal at illuminations suffi-
ciently low to confine visual function to the rods.
The serum cholinesterase concentration of all the sub-
jects was reduced by the exposure to PF-3 to 1 to.') per cent
of the normal value.
f. Additional accidental exposurtWorkers acciden-
tally exposed to undetected concentrations at the American
pilot plant developed extreme miosis of 1 week’s duration.
Difficulty of night vision was stressed.*
In a report on a similar incident at the liritish pilot plant
emphasis was placed on pupillary contraction, blurring of
vision, especially in artificial light, headache, and light-
ness in the chest.100 In another incident 7* undetected cx-
posures to vapor produced miosis, pi sir vision in dim light,
difficulty in focusing, twitching of the eyelids, nasal dis-
charge, and (in some cases) conjunctivitis. There was no
mention of headache or chest symptoms.
5. Di-scc-bulyl fluorophosphate.'^1 This ester has been
given only preliminary tests with four human subjects. L'pon
exposure to a nominal concentration of I/10* (Cl = approxi-
mately 15 mg min m3) all noticed a tightness across the chest
but three of the four fell that it. was not sufficient to call for a
respirator. About 5 minutes after the subjects left the cham-
ber, miosis set in, became intense, and persisted for 5 days.
Comparison with the results obtained with PF-3 at compa-
rable and somewhat greater dosages T,-,ov would indicate that
the di-scc-butyl ester may be the more potent miotic agent.
(5. hint Dimdhytnmido) phonphoryl fluoride.,IMo Exposure of
four volunteer subjects to about 45 mg min m1 (7.3 Mg 1 or
1/10*, for 5 to 7 minutes) produced no observable ocular or
systemic effects. This compound is therefore less effective,
perhaps very much less effective, than PF-3.
9.3.3 Toxicity
Inhalation and Injection Toxicitiks
Toxicity data for the more intensively studied Tri-
Ions, fUiorophosphales, and dialkylamidophosphoryl
fluorides are given in Tables 5 through 17 and are
summarized in Table 4. It is apparent that the Tri-
bal.- arc the most toxic volatile agents considered in
Table 5. Toxicity of ethyl dimcthylnmidoeyanophos-
phale (M('K) by inhalation.
The animals were totally exposed to the vapor of the
agent. Concentrations were nominal except when other-
wise designated.
Exposure
N umber
time
UOtho
of
Species
(min)
(mg min/nr1)
animals
Reference
Mouse
2
100
100
C9d
5
385
100
69d
5
500-750
22
104s
10
380
400
69,1,e
10
220*
140
87
10
500-750
IS
104s
20
420
100
69d
30
420
100
69d
00
670*
120
87
120
840*
60
87
Hat
5
750-1,000
14
104s
10
750-1,000
10
104s
10
500-1,500
14
69c,e
10
304*
2:40
87
20
385*
54
66k
60
620*
60
87
120
1,200*
120
87
Guinea pig
5
1,000 ±
6
104s
10
1,000-2,000
6
104 s
10
393*
81
87
10
500 1,500
12
69e,c
00
740*
50
87
120
1,500*
48
87
llahhit
10
1,000*
15
87
10
>4,000
6
104s
10
>2,000
4
69c, e
10- 62
840*
55
G6h
Cat
7.5
300 800
3
104s
10
2.50
8
69e,e
I tog
10
400
4
69c,e
Goat
10
700*
9
87
14-23
400-700*
10
66h
13-120
765*
30
66k
20
1,400* t
38
66k
Monkey
5-10
400 ±
5
104s
10
250
4
69e,c
10
180*
3
87
* Analytically determined concent ration
t Angora goal*.
SECRET TOXICOLOGY
Table 0. /./>„o’s of ethyl d imcthy la midi >eya qo| >hns-
phate (MCK).
The figures in parentheses arc the ininiber of animals
used.
Route of
Approximate
administration
S|tecies
/.Din (mg kg)
Reference
Intravenous
Mouse
0.15
(15)
69c
_
Rat
0.006
(35)
66h
Rabbit
0.0025
(40)
6011
0.18
(14)
87
0.125
(15)
104s
DoK
0.084
(20)
06i
0.116
(0)
69c
Subcutaneous
Mouse
0.4
(25)
87
Rat
0.3 0.4
(42)
87
Guinea pig
0.2
(20)
87
-Rabbit
0.5
(30)
87
Percutaneous
Mouse
1.0
(70)
—69c
>4
(20)
OOg
Rat
18 35
(47)
87
Guinea pig
35
(43)
87
Rabbit
2.5-3.0
(5)
OOg
3.3
(00)
09h
35
(19)
87
Dog
30-50
(4)
69c
Goat
>5
(2)
OOg
1.1
(21)
OOh
-
3
(17)
87
Monkey
0.3
(0)
69c
I’er oh
Rat
3.7
(107)
66ii
8
(20)
87
Rabbit
10.3
(51)
OOh
Dug
5-11
(12)
OOh
Table 8. LI)
(MFI):
Tin* figures
used.
,,,’s of isopropyl
in parentheses
methanefluorophosphonate
are the numlier of animals
Rout*? of
administration
S| levies
Approximate
1.1I.J (nig kg)
Reference
Intravenous
Hat
0.0-15
(50)
OOi
Rabbit
0,0 Hi
(14)
OOi
Percutaneous
Mouse
1.08
(40)
OOd
Rabbit
0.025
(10)
(ilii
Per ok
Rat
0.55
(00)
OOi
Table 0. Toxicity of isopropyl elhanefluoruphosphonate
by inhalation.
The animals were totally exposed to the vapor of the
agent. All concentrations were nominal.
Kxposure
Number
_
time
/.(Oh,
of
Sjreeies
jiniii)
(rnjj min in')
animals
Reference
Mouse*
5
245
80
title
10
330
120
60d,c
10
350 1,000
8
104t
30
570
60
title
Rat
10
260
6
69e
10
<350
4
Kilt
Guinea pig
10
>210
6
60e
10
350 1,000
4
1041
Rabbit
10
230
4
60c
10
350-1,000
4
104t
Cat
10
170
6
title
Dog
10
230
4
60c
Monkey
10
210
3
60e
* SuhoutaiM’tms LDw (8 mice) “ approx. 0.4
L/)w (40 slaved mire) =1.7 mg/kg.1^
mu, kg. '***
Percutaneous
Table 7. Toxicity of isopropyl imThahcfluoroptios-
phonate (MFI) by inhalation.
The animals were totally exposed to the vapor of the
agent. All concentrations were nominal.
Species
Exposure
time
(min)
urt),a
(mg min/in’)
Number
of
animals
Reference
Mouse
5
230
100
GOe
10
250
lOf)
title
10
1.50-230
22
104t
15
34.5
120
69e
20
360
100
Otte
30
420
100
title
Hat
10
300
IS
6tld,c
10
150-250
12
1041
Guinea pin
10
ISO
18
69d,e
10
150-2.50
12
104t
Rabbit
10
120
6
Gtld.c
10
1.50 2.50
5
104t
Cat
10
100
10
69d,e
I)oK
10
100-150
8
09d,e
Monkey
10
150
5
G9d.i-
cate Unit the species variation in susceptibility is not
prononneed. On the other hand the animal species
exhibit considerable variation when MCE, anti par-
ticularly tiie fiuorophosphates, come into considera-
tion.1* This variation makes estimates of I lie human
lethal dosage precarious. In the case of PF-3 com-
parison of the systemic effects produced upon ex-
posure of human subjects (see preceding section) and
of monkeys2SI|,r'63 64 83 to dosages in the order of
300 to 400 mg min m* indicate clearly that man is
the more resistant species. Man was affected but not
prostrated,-whereas the monkeys were severely pros-
trated and some were killed. How much more re-
sistant man is to PF-3 than the monkey is not known,
nor does the same relationship necessarily hold for
the Trilons.
this volume. The fiuorophosphates are considerably
less toxic, although the potency of PF-3 and di-scc-
butyl fluorophosphate for the monkey approaches
that of tho Trilons. The limited data for MFI indi-
'* The high L{Ct)j,of PF-1 and PF-3 (possibly also
MCE) for the rabbit are due largely to inhibition of respira-
tion. \\ hen inhibitory respiratory reflexes are suppressed, or
when t lit* agents are injected, the rabbit is not found to lie
excessively resistant.**•'4.“
SECRET 152
FLI'OUOPIIOSPII \TKS V ND IMIOSPIlORLS-CONT VIM NO COMPOUNDS
Tarle in.
Toxicity of dimethyl fluorophosphate (PK-1)
by inhalation.
The animals were totally
exposed to the vapor of the
a ({cut. All
concentrations were nominal. Injection and
tiercHlaneous toxicity figures are Riven in the footnotes.
Hxeept for 1
the mouse /,((V )..„’s, the figures are very rough
approximations based on only a few animals of each
sjH'cies.
Kx|>osure
time
unu
S|ierics
(min)
(mg min m*) Heferenee
Mouse*
1
1,200 201
2
1,740 20h
10
2,550 11
10
3,000 — 40
30
5,000 ± 26f
120
>5,000 20f
Hat
1
1,800 ± 26f,l
10
3,000-0,000 20a, I04b
Guinea pig
1
7,000 + 20f,l
0.5 4
8,(XK) ± 20f
Habbitf
1
> 12,000 20f, 104b
('at t
1
0,000 + 201
r>o*s
1
6,000 + 201
Goat
3 5
20,000 + 26f
House fly
10
<30 201
Mosquito '
10
<30 201
» Mouse ii.tr:
sve non* LI) w » 0.15 mg kg/** Mouac infra peritoneal
me
: kg ** Mouse jx-r
cutaneous LI)ao (70 mice) » 0.72
mg animal, or approx. 3tt mg kg.4*
t Habbit infra
ivcnous LDu = 2 I
mg kg >
t ('at int ravel
hour LD*o = 1.5 mg
kg."1
§ I>-« intravc
nous /,Os. = 12 mg kg.5*
|| Antra urr/yjJi.
Table 12. Toxicity of diisopropyl fluorophosplmto
(PF-3) by inhalation.
The animals were totally exposed to the vapor of the
agent. All concentrations were nominal. Kxeept for the
mouse and rat s, the figures are rough approxi-
mations based on relatively few animals per species. A
total of 39 monkeys were exposed.
_
Exposure
time
««)».
Species
(min)
(mg nun nr')
Reference
Mouse
1
4,000
I04e
2
3,800
104e
5
2,700
I04e
10
3,500
I04e
It)
5,500
49
10
5,900
26c,p
—
30
4,500
1 Ole
100
>0,400 —
26p
Ral
1
4,200
104h
2
3,000
I04h
5
2,850
10 th
10
2,800
104h
30
4,500
I04h
Guinea pig
10
>8,200
104b
Rabbit
10
(8,000 ±)
104b
Dog
10
5,000 +
8.3
Goat
10
6,000 7.000
S3
Monkey
2
500-800
261
2-15
500 ±
63, 64
10
800 +
83
100
1,000-2,000
26p,r
Table 13. LD.
•u’s of diisopropyl fluorophosphate (PF-3).
Route of
Approximate
administration
Species
L/)jo(mg/kg)
Reference
Intravenous
Rabbit
0.3-0.4
— 07b
0.4 +
53
0.5-0.75
32a
Cat
<3
33a
Goat
0.8 +
53
Monkey
0.1-0.2
48, 0«a, 07f
Intramuscular
Rat
2-
321.
Rabbit
0.75-1.0
67b
Sulieutancous
Mouse
4 ±
80, 104e
Rat
3 +
80
Rabbit
I +
86
•
Dog
3 ±
80
Goal
1 +
SO
Percutaneous
Mouse
72 +
40
(1.45 mg mouse)
Per os
Mouse
30.8
08a
2 +
80
Rat
5-10
20r
6 +
80
Rabbit
0.8
08b
Ry eye
Rabbit
1.4
32a
Table 11. Toxicity of diethyl fluorophosphate by
inhalation.
The animals were totally exposed to (he vapor of the
a Kent. All exposure times were for 10 minutes and all
concentrations were nominal. Kxcept for the mouse
/.(COjo’s, the figures are very rough approximations
based on 6 to IS animals per speeies.
UCt)„
S|ieries
(mg min/m3)
Reference
Mouse*
8,200
II
■ ‘ - ' -
4,100
- 40
4,000-6,000
104b
Kat
7,000-14,000
20b, 104 b
Guinea pig
7,000 14,000
26b, 104b
Rabbit
>14,000
104b
* Mourn* pcrrutapwHis /-/>** (00 mice) ■** 0.70 i
35 me k^.**
mg,animat, or approx.
As indicated by (he data of (he toxicity tables, the
“rate of detoxification” as measured by the increase
of the lethal dose with increase of time of its admin-
istration. is marked for PF-1, moderate for MFI
and isopropyl ethanefluorophosphonate, and slight
but definite for MCE and PF-3. More detailed data
hearing on the rate of detoxification as determined
by inhalation and injection experiments will Is1 found
in the references cited in the tables. Other references
are also pertinent.l#f * w “ “
Both the Triions and the fluorophosphates are TOXICOLOGY
153
Tabu 14. Toxicity of di-scc-hutyl fluorophosphale by
inhalation.
The animals were totally exposed to the vapor of the
agent. All concentrations were nominal. Except for the
mouse L{Ct)i,’s, the figures are very rough approxima-
tions based on 2 to 23 animals of each sj)ecies.
Exposure
time
UCtu
Species
(min)
(mg min/in’)
Reference
Mouse
10
5,140
20r
10
5,400
104i
Rat
10
4,000 10,000
20r, 104 i
Guinea pig
10
>18,000
20r, 104i
Rabbit
10
5,000-10,000
20r, I04i
Gat
10
0,000 ±
2Gr
Dog
10
4,er sixties.
UClU
SjH'ries
(nig min/nv1)
Keference
Mouse
800
1011
I,I00
26n
If at
1,200 ±
26n, Kill
(Iniitea pig
6,000-10,000
ion
Hal.bit
1,2(Xh 2,800
2fin, 1011
Dog
1,000 1.100
201, 20u
Table 10.
Toxicity of 6i«(diinethylami4,000
20j, 104o
2
4 f
80
Rabbit
> 2,000
104o
3±_
0 +
3 +
80, lOlo
Cat
2 t-
80
Goat
2 +
80
Monkey
>1 or 2
SO
“quick-kill” agents. Although occasional deaths are
delayed for 1 or 2 days, most lolhally poisoned ani-
mals die within 2 hours after exposure, and the ma-
jority during or within a few minutes after exposure.
Detailed statements may be found in the references
cited in the toxicity tables. A special study has shown
that PF-1 and PF-3 are only slightly slower in speed
of action than hydrogen cyanide (AC), although the
lower volatilities of the Huorophosphates would make
high concentrations relatively difficult to attain in
the field.44
Symptoms and Pathology
The symptoms produced by exposure to the Tri-
Ions and fluorophosphates are those which character-
ize the nicotinic and muscarinic actions of parasym-
pathomimetic agents in general. There are also
evidences of central nervous stimulation. Although
there are variations according to species, agent, and
dosage, frequent mention has been made of the fol-
lowing: lacrimation and salivation; apprehension;
coughing, dyspnea, and gasping; hyperexeitability,
incoordination, and ataxia; tremor, muscular twitch-
ings, and convulsions; sometimes bronchospasm,
pilomotor stimulation, urination, and defecation;
general weakness ami depression; and finally cessa-
tion of respiration. Detailed descriptions for the
various agents and species may be found in the refer-
ences cited in the toxicity tables (see also the refer-
ences given under the section “Protection and Treat-
ment”).
Respiratory failure is probably the usual primary
cause of death.*3 However, the action of the agents
as revealed, for instance, by a study of PK-3,33*
clearly involves most of the important systems in the
body and the weakest link in the chain of events
leading to death is questionable. One point of view
is that bronchospasm may be important in some
species, including man. This is not t rue in the cat.33
Because of the early time of death, pathologi-
cal changes frequently are not conspicuous at au-
topsy.261** '£1X7-,(M Dicyclohexyl fluorophosphale,
which seems to act somewhat more slowly than the
other Huorophosphates, has berm found to produce
in rabbits a marked pulmonary edema and edema of
the perivascular connective tissue, marked pulmo-
nary hyperemia, large areas of ataleetasis, hepatic
congestion and incipient central atrophy, and slight
lymphorhexis.1*'" The action of 6fs(dimethylamido)-
phosphoryl fluoride is slower still ami the pathologi-
SECRET 154
Fl.l OHOPHOSnr ATES V\'D PHOSPHOR17S-CONT M M NO COMPOl M)S
Table 17. Toxicity
of vapors through the ski
n (body only exposures)
F,X|sisiire
Dot
age (Cl in mg min in’)
I ime
Nominal
Analytical
Agent
Species
(min)
cone.
cone.
Mortality
Reference
Dimethyl fluornphosphate (l’F-1)
Mouse
10
124,000
122,000
0/6
20*1
10
51,600
48,200
3/6
20*1
15
13,000
12,000
0/6
26*1
Kthvl dimelhylamidoevannphosphate
Mouse
10
3,850
2,500
UCt)u
00c, d
(Mt'K)
10
1,000
750*
Uri)M
00c
no
0,000
3,000
UCl)m
00,1
Guinea pig
10
11,200
0,100
0 6
69c
Dog
10
11,200
0, UK)
0/1
69c
210
45,400
20,000
0/1
69*1
227 360
80,000 ±
urnM
00k
(8 animals)
Hablut
77-282
10.000 +
UCDu,
Oflli
(30 animals)
Isopropyl methaneflnorophnsphonate MFI
Mouse
10
8,720
1 20
69e
6/s(jM-'hloroelhvl) sulfide (II)
Mouse
10
3,500
UCI),a
15
Habhit
13.5
2,000
0/1
15
18
4,000
0/1
32
5,800
i/i
35
8,(*)0
i/i
- —
00
13,400
0/1
80
20,500
—1/1
"
rh>g
00
0,550
0/1
15
-
CO
0,000
1/1
-
30
15,400
1/1
00
17,000
1/1
00
24,000
1/1
Iris(,%{ ’hloroet hvDamine (H X3)
Mouse
10
800
UCl)M
15
Rabbit
47-140
>5.500
urt)M
20t,u
Dog
30
13,300
0/1
15
45
14,500
0/1
75
21,400
l/l
100
51,000
1/1
* Skin of miee shaved.
cal changes are somewhat different. Attention has
been directed to marked pleural effusion, pulmonary
edema and hyperemia, and inflammation of the sub-
mucosal layer of the tracheal and broncheal epi-
thelium.36'ksB The references cited in the toxicity
tables shoijld lx* consulted for more detailed patho-
logical information.
Effects on and Timorr.H the Skin
Neither the Tritons nor the fluorophosphat.es exert
a vesicant action.ll !*d s', *6h k e S|7,9° With regard
to absorption of the agents through the skin, the
• lata given in Table 17 suggest that vapor dosages
reasonably attainable in the field would not produce
significantly severe systemic effects percutaneously
in the cases of the larger animals nor, presumably, in
man. Moreover, ordinary clothing can be expected
to afford some protection against the vapors; C(-2
impregnated clothing, considerable protection; and
carbon clothing, virtually complete protect ion.6#Jr
On the other hand, liquid contamination of the skin
with MCE or MFL is potentially very dangerous
(see Tables (i and 8).**h' In the ease of MCE rapid
removal of the liquid by blotting is effective treat-
ment. Apparently chloramides do not react readily
with MCE. Consequently antigas ointments are of
limited value except in so far as their application can
facilitate the removal of the agent by solvent or me-
chanical action. In experiments with rabbits it was
found that interposition of a single layer of plain
herringbone twill increased by 6- to 8-fold the dose
of MCE that must lie applied to the skin to cause
death. A single layer of CC-2 impregnated cloth in-
creased the dose 10- to 12-fold; two layers of this
cloth, 20-fold; and one layer of carbon cloth, 15-
fold.M,, i Thus clothing, particularly protective cloth-
ing, is of considerable value in preventing the
absorption of lethal doses of this agent.
Protection and Treatment
Numerous substances and procedures for prophy-
laxis and therapy of the systemic effects of fluoro-
SECRET 155
TOXICOLOGY
phosphate and Trilon poisoning have been inves-
tigated.4*4 * b *-32 33*-44 *8* b.*.h. i,k,«7».b.r.M,*!to.h,t04h.n.« Qf
these the injection of atropine and magnesium sulfate
seems to offer the most promise and has been recom-
mended for use in the event of human poisoning.**3®7*
These therapeutic agents suppress the autonomic
symptoms. Injection of Nembutal in addition will
control the convulsions which occur in MCE poi-
soning,*"1 •S7 However, in severe poisoning the action
of these drugs will merely delay but not prevent
death. In any event it is essential that therapy be
instituted promptly. Adequacy of protection by the
gas mask has been mentioned, as have Ix'en methods
of I renting eye effects and of preventing percutaneous
absorption of liquid contamination.
Physiological Mechanism
In 15)41 and 1912 British workers reported that
the dialkyl fluorophosphales are very potent inhibi-
tors of cholinesterase.1"3 1043 h Since that time exten-
sive studies have been made on the clinical pathology
and biochemistry of action of these compounds,-*' *•
m a,uj moro recently of the Trilons,
which have also proved to be potent antieholines-
terases.M« '' ' «7.6oe,f h i 'i')ie results have already begun
to appear in the open literature and need not be re-
viewed hem. It is obvious that the agents will In*
of great value as tools in physiological and biochemi-
cal research. Their possible use in the treatment of
myasthenia gravis has also been under investi-
gation.
SECRET Chapter 10
METHYL FLU01U)ACETATE AND RELATED COMPOUNDS*
Brnhey Ren show and Marshall dates
lo.i INTRODUCTION
Rkhokts that methyl fluoroacetate is highly
toxic were received from Polish investigators
by the British in 1912 and prompted extensive stud-
ies in the United Kingdom and United States. Flu-
oroaeetie acid and many simple derivatives including
salts and esters, 0-fluoroethanoI and its esters, anti
salts and esters of y-fluorobutyric acid, y-fluoro-0-
hydroxybutyric acid, and y-fluorocrotonie acid,
proved to l>e highly toxic by inhalation, injection,
and ingestion. These compounds produce death, usu-
ally after a latency of one-half to several hours, by
action on the heart or central nervous system.
Compounds of this group are not seriously con-
sidered for large-scale use in chemical warfare at the
present time because: (1) although very toxic for
some species, (he human lethal and incapacitating
doses are believed to be comparable to, or consider-
ably greater than, those of the current ly standardized
persistent and nonpersistent agents; (2) the stable
derivatives do not possess sufficiently high vapor
pressures to be dispersed from available munitions
in high concentrations as nonpersistent agents; and
(.3) the gas mask affords adequate protection.
The silts of fluoroacetic and related toxic acids
are nonvolatile, stable in aqueous solution, and ap-
proximately as toxic when administered orally as
when injected. They are, therefore, potential water
poisons in warfare and are proving to lx1 highly ef-
fective bait poisons for rodents.
The compounds of this group selectively poison
enzyme systems and as inhibitors will 1h» of value in
the study of intermediary metabolism.
10.2 Synthesis and Properties
Approximately 160 aliphatic fluorine compounds
have U’en prepared by NDRC and British investi-
gators for evaluation as eliemieal warfare agents.
The compounds, their physical properties, and refer-
ences to their synthesis and toxicity are listed in
Table I.
10.2.1 Synthesis
In general, the syntheses have Ixx-n effected by the
fluorination of corresponding chlorine and bromine
compounds by treatment with metallic fluorides,
usually anhydrous potassium fluoride, less frequently
silver fluoride, mercuric fluoride, or antimony fluo-
ride. according to known procedures.19-'22■“•w.ioi.im.im
Application to the fluorinated compounds of
standard synthetic methods has resulted in a variety
of derivatives including representatives of most of
the common aliphatic types.
The methods may lx- illustrated by the following
examples.
1. Methyl fluoroacfiate '».**•>»*.!«» has been pre-
pares! on a large laboratory scale (50 lb) by heating
methyl chloroacetate under pressure with anhydrous
potassium fluoride at 220 (' for 5 hours. The product
is distilled directly from the pressure vessel and is
purified by fractionation. Yields in the neighborhood
of 75-77 per cent are obtained.
The compound is a colorless mobile liquid with a
faint ester-like odor. Unlike the other haloacetates,
it has no lacrimatory properties. It boils at 104.5 C,
freezes at —35 C, and is soluble in water to the extent
of about 15 per cent. Its physical properties have
been thoroughly investigated.u-*7-*Stt
2. Sodium fluoroacrlate, l9-22-92<‘ because of its prom-
ise as a rodent icide, has been prepared on a much
larger scale than has any other member of the fluoro-
acelate series. Complete pilot plant conditions were
worked out during the course of preparing 1,000 lb 22
for use in experimental rodent-control projects. It is
prepared by saponification of ethyl fluoroacetate
" Rased on information available to Division 9 of the Na-
tional Defense Research Committee [NDRC] as of October
1. 1945.
Attention is directed to a recent paper hy ,J. S. C. Marais,
entitled Monofluoroaretir Arid, The Toxic Principle nf "Gif-
bhmr" Dirhapetnlnm rymosnm (Hook) Emjl., Ondcrstepoorl
Journal of Veterinary Science and Animal Industry 20, 67 73
(1944). The early Dutch settlers in South Africa gave the
name “Gifhlaar” to a plant the leaves of which are poisonous
to livestock. A numlxT of toxicological and chemical studies
have Ix-cn made in South Africa since about 1900, and it is
apparently a remarkable coincidence that the active principle
was lx-ing identified there at the same time that fluoroacetic
acid derivatives were being actively studied as potential
chemical warfare agents in the United Kingdom and United
States. Although the South African literature corroborates
many of the chemical and toxicological findings summarized
in this chapter, a cursory survey fails to reveal data permitting
an independent estimation of the human lethal dose.
SECRET 157
INTRODUCTION
Table 1. Aliphatic fluorine compounds examined as candidate chemical warfare agents.
The compounds are arranged in three major categories in the following sequence: (1) compounds containing not
more than one fluorine atom attached to any carbon atom; (2) compounds containing two fluorine atoms attached to any
one carbon atom; and (3) compounds containing three fluorine atoms attached to the same carbon atom. Within each major-
category compounds are arranged in sequence according to the following types: hydrocarbons, alcohol derivatives, amines,
carbonyl derivatives, and acid derivatives.
The following abbreviations are used; n’n, refractive index at 1 C;
1.708
52
52
mp
12 18°
52
bp“
55-5G°
52
vol54
70.1
_ 52
- ...
4. Icd-Butvl fluoride
3, 19
bp™
14.5-16°
3
13
5. /J-K!uoroethanol
19, 92c
nn!“
1.3618
14
13, 91c, 92c
-- — _ _
iP*
1.0913
14
bp7**
99 100°
14
r~.
vol5*
74.8
14
, . .
6. Methyl j3-fluoroethvl ether
91 f
bp
ea. 60°
91f _
91 f
7. Chloromethyl 0-fInoroethyl ether
19
bp5i
35-40
19
13
8. 0-Chloroethvl 0-fluoroelhyl ether
19
bp”
55 58°
19
13
9. /3-Fluorocthvl d-hydroxycthyl ether
19
n u!S
1.1050
19
13
bp"
01-62°
19
10. Methvl £-fluoroethoxyacetate
19
bp1*
8.5-88“
19
11. d-Fluoroethyl phenyl ether
92f
mp
41
92f
01 h
_x_ ■
“ ...
bP>7
92.5°
92f
12. 2 '-Flin>ro-2,4-dinitrophenetolc
19
mp
89-91°
19
13
13. 0-Fluoroelhvl d-naphlhvl ether
92g
mp
49.5 50°
92g
91 h
14. b/»(d-Fluoroeethvl X-nit roso-X-(d-chloroethyl)carlmmate
19. 30
bp5
118-121°
19
13
27. d-F’luoroethyl glycine hydrochloride
92j
mp
150.5°
911
911
28. (J-Fhioroethyl betaine hydrochloride
92j
mp
122°
911
911
29. 0-Fluoroethyl nitrite
19
"o50
1.3589
14
13
p‘*°
128°
14
vol5*
47.0
14
SECRET 158
METHYL FLL’ORO ACETATE WO RELATED COMPOUNDS
Compound
Reference
to
synthesis
Physical pro|ierti*
Projierty R
'ferenee
Reference to
toxicity
data
33. bi.s(/3-Fhioroethyl) carltonale
19
>p‘
71-72°
14
Vol*4
1.15
14
34. U-Fluoroethvl chlorosulfonate
19. 92g
•>p!"
85 86°
10
13, 91f
33. b/«(d-Fluoroethvl) sulfate
19, »r2g
«ir*
1.4177
14
13. 91 f
—
d?4
1.3678
14
bp?
SO-81°
14
vol54
0.425
It
36. fris(tf-Fluoroethyl) arsenite
19
bp! 4
132 134°
19
13
37. Mrakis(P-Fluoroethvl) silicate
19
bp"4
102 104.5°
19
13
38. trix(H-Fluoroethvl) borate
bp
192°
91 o
91o
39. d-Fluoroethoxydichlorophosphine
19, 31 e
•»P”
50
31e
13
. * . -
bp™
140-145°
31 e
40. fcjsfd-FluoroethyD hydrogen phosphite
92n
bp15
109 110°
92n
91 h
41. his( Diethvlamino)-/S-flii(»roelhoxyphosphine
19
b|'a
108-111°
19
13
42. Kthvld>f*(f3-fluoroethoxy) pTiosphine
19
bp1'-4
4(F49
19
13
43. trisfp-Fluoroethyl) phosphite
19
bp11-4
100 105°
19
13
44. his(d-Flu•
1.3759
14, 24q
13, 91c
—
dP
1.0826
14, 24q
bp™
114-118“
14, 24q
'
vol5*
68.57
14, 24q
79.
Klhvl dichlorofluoroacctate
47h
bp7J0
131.5-132°
47b
13
SO.
d-Fluoroethvl fluoroaeelate
19, 92d
1.3802
14, 19
13, Old
bp«
85°—
14, 19
—
vol54
7.81
14, 19
81.
0-Clilorocthyl fluoroaeelate
19, 92d
it'-"
1.3160
14
13, 9lc
_
bp51
86“
14
Vol=*
3.55
14
82.
Allvl fluoroaeelate
19, 921
n ir"
1.4063
14
13, 911
...
iP"
l.OOtil
14
—
bp50
64.5-65°
14
-
vol5*
31.69
14
. . .
83.
Propyl fluoroaeelate
92e
bp
135 137°
92e
91c
84.
Isopropvl fliloroacetate
19. 92e
n n50
1.3804
14
13, 91c
(Pa
1.033
14
bp™
121-123°
14
-=-
vol5*
62.31
14
85.
/J-Ethylhexyl fluoroaeelate
59a, 59c
86.
Plicnvl fluoroaeelate
19
mp
01.5-63°
19
13
87.
/)-('hlorophenyl fluoroaeelate
19
mp
52-54°
19
13
88.
Cliolestervl fluoroaeelate
92f
mp
144 144.5°
92f
91 h
89.
Met hvlenc-4«',>t(monofluoroaeel ate)
921
mp
57“
921
t>21
90.
Gl ve< >1 bts( m< >n< (fluoroaeetate)
92f
bp17
110-141°
92f
13, 91 h
91.
Fluoroacetvlcholine chloride
7T.
92.
Fluoroacet vlsalicylic acid
19, 92j
mp
141-144°
19, 92f
13, 91b
—
mp
131.6°
93.
S-(J-chlorocthvl fluorothiolaeetate
19, 92j
bp10
80 81°
19
13, 911
94.
Phenyl fluorothiolaeetate
92f
mp
36.5-37.5°
92f
91h
bp'*
132°
92f
...
95.
Methyl fluoroselenolacetate
11
nu-“
1.4879
14
13
iP"
1.573
14
—
bp7”
130-132°
14
Vol55
69.95
- 14
96.
Fluoroacet vl fluoride
19, 92e
bp70"
35-40°
19, 92e
13, 91f
97.
Flui >r< meet vl ehloriile
19, 92b
»i.5“
1.3831
14
13, 91c
iP"
1.3530
14
...
bp75*
69 71°
14
. . .
—
vol5*
C>07
14
98.
Fluoroaeetonitrile
19, 92«
m.50
1.3324
14
13
bp™
78°
14
vol5*
260
14
99.
Fluoroacet vl isot hioevanate
19
nu5“
1.5327
14
13
,P«
1.3527
It
—
bp*°
76°
14
vol5*
15.51
14
...
100.
Fluoroacet ic anhydride
92e
bp15
88-89.5°
92c
91c
101.
Fluoroacet amide
19, 92h
mp
108°
19, 92b
9lc
102.
X- M et hy Ifliu iroaeet a mide
92b
nip
64°
92b
103.
X - Xi t roso-X-mclhy lflu< >roaeel a mide
92e
bp1*
84°
92e
91c
SECRET 100
METHYL FLL’ORO ACETATE AND RELATED COMPOUNDS
Tabus 1 (Continued).
Compound
Hcference
to
synthesis
Physical properties
Property Hcference
Reference to
toxicity
data
104. X-/J-Chlor«)cthylfluoroacet amide
19, 92e
mp
fi5°
19, 92e
13
bp" 3
77°
19, 92c
105. X-0-Tlydroxyethylfluoroacetamide
92e
mp
ca. 21°
92c
bp"1
114°
92e
106. X.X-Diethvlfluoroacctamide
19
bp"
86°
19
13
107. X,X-6fs(d-Chlor(>ethyl)fluoroacctamide
92e
nip
64.5°
31a, 92e
bp“-°*
102°
31a, 92c
108. a-Flnoroacetanilide
23
mp
73 74°
23
20
109. Fluoroacetvlglvcinc ethvl ester
92f
mp
50 50.53
92f
9lh J
110. Fluoromethylfluoroacetvlurea
921
mp
84°
921
921 /
111. 2-Fluorocthane-l-sulfonyl chloride
92g
bp13
81.5-84.5°
92g
91 h
112. Methyl o-fh i«.ropropiona t c
92c
bp
106.5-108.5°
92c
91 f
113. F.thvl jl-fluoropropionatc
19
bp‘*
65-68°
19
20
li t. Diethvl fluommalonatc
9lc
bp"
84 86°
91c
91c
115. S«>dium y-fluorobutyrale
19
13
110. Methyl y-fluorobulyrate
10. 19
«ir°
1.3887
10
13
1.0662
10 —
—
, .....
lip1"0
79°
10
—
...
vol3*
39,6
10
...
117. Methvl nr-fluoroisobutyrate
92c
bp
108-1014°
92c
91c
1 IS. Ethvl a-flu■»*.»?<■ has been prepared on
a large laboratory scale (50 lb) by heating anhydrous
potassium fluoride with ethylene ehlorohydrin at
ISO (’ for 4 to 5 hours. Yields of 53 per cent are ob-
tained. Except for the difference in temperature the
reaction is carried out as described for methyl fluoro-
acetate. During the heating ethylene oxide is formed
almost quantitatively from the ethylene ehlorohydrin
but appears to he the product of a reversible side re-
action. Attempts to produce 0-fluoroelHanoi from
ethylene oxide and hydrogen fluoride have been un-
successful.
/3-FIuoroethanol is a colorless liquid with a pleasant
alcohol-like odor. It boils at 102.5 C at atmospheric
pressure and is completely miscible with water.
Several of its physical properties have been deter-
mined.14-*Se
4. Mrlhyl y-fluorobulyrate,019 is prepared by
treating trimethyleneehlorobromide with sodium
cyanide to produce a mixture of -y-chloro and -y-bromo-
butyron it riles. The mixture is then heated under
pressure with anhydrous potassium fluoride at 200 C
to give -y-fluorobutyronit rile, which is converted to
SECRET 162
METHYL KLCOROACETATE WD BELATED COMI’Ol ,M)S
the corresponding methyl ester by treatment with
methanol and acid. The overall yield of methyl
y-fluorobutyrate obtained in preparations on a large
laboratory scale has been about 25 per cent for the
three-step process.
5. Methyl y-Jlnoro-0-hydroxybutyrnte, methyl fi-
chloro-y-fluorobutyriile, and methyl y-fluororrotu~
nnte 1019 are prepared as follows. Epichlorohydrin
when heated under pressure at 225 C with potassium
fluoride is converted into epifluorohydrin. Treatment
of the latter with anhydrous hydrogen cyanide and a
small amount of sodium cyanide gives y-fluoro-/3-
hydroxybutyronitrile in excellent yield. This inter-
mediate is converted to methyl y-fluoro-jS-hydroxy-
butyrate by treatment with methanol and acid.
Methyl /J-chlorb-y-fluorobutyrate is produced by the
action of thionyl chloride and pyridine on the hy-
droxy compound. Methyl y-fluorocrotonate may
then he formed by dehydrohalogenation of the /it-
ch loro compound with triethylamine. Yields are good
except in the case of the first step involving the eon-
version of epichlorohydrin to epifluorohydrin. It has
not yet been possible to raise the yield of this step
above 40 per cent, although 70 to 74 per cent of the
unconverted epichlorohydrin is recovered in a form
suitable for re-use. The overall yield based on the
amount of epichlorohydrin utilized is 46 per cent for
methyl y-fluoro-/3-hydroxybu(yrate, 39 per cent for
methyl /3-chloro-y-fluorobutyrate, and 33 per cent
for methyl y-fluorocrotonate. All three esters are
stable colorless liquids.
An alternative method not requiring high-pressure
equipment has been developed for the preparation of
methyl y-fluorocrotonate on a laboratory scale.19
Methyl y-bromocrotonate, prepared by bromination
of methyl croton ate with N-bromosuccinimide, is re-
fluxed at atmospheric pressure with anhydrous po-
tassium fluoride. The product is slowly dist illed from
the mixture as the reaction proceeds. Yields of ap-
proximately 10 per cent are obtained.
In addition to the synthetic procedures already
described, a variety of methods has been used to pre-
pare other fluorinated aliphatic compounds. .Men-
tion may be made of (he preparation of difluoro- and
trifluoroaeetic acids by the oxidation of 1,1-dichloro-
3,3-difluoropropene and l,l,2-trichloro-3,3,3-triflu-
oropropene, respectively 15 (see Chapter 40); the
preparation of several ethers of /3-tetraHiioro-
ethanol by the addition of alcohol to tetrafluoro-
cthylene; the synthesis of /3-fluoroethyl thiolacetate,
from which /3-fluoroethylmercaptan may be obtained
by hydrolysis, by the peroxide-catalyzed addition
of thiolacetic acid to vinyl fluoride;33 the synthesis
of tetrafluoro-1,2-dinit methane and chlorotrifluoro-
1.2-dinitroethane by the addition of nitrogen tetrox-
ide to (he eorresponding halogeiuited olefines; and
the synthesis of derivatives of t-fluorocaproic acid
from eyelohexanone through t-hydroxycaproic acid
and the corresponding bromo-eompound, which is
treated with silver fluoride.wk
l».2.2 Chemical Properties
Methyl fluoroacetate has l>een the subject of most
of the work on the chemistry of fhoaliphatic fluorine
compounds considered in this chapter. It is readily
hydrolyzed to fluoroaeetic acid and methyl alcohol,
the half life of the ester in water buffered at pH 7
being less than 1 hour.21 On the other hand, the flu-
orine atom can be removed from the molecule only
by relatively drastic treatment. No reagent has been
found which will bring about rapid replacement at
room temperatures. As an example, no fluoride ion
is produced by heating methyl fluoroacetate for
5 minutes with 20 per rent alcoholic potassium hy-
droxide, although more prolonged heating (18 hours
on steam bath) does result in the incomplete libera-
tion of fluoride ion.95d Concentrated acids at steam
bath temperatures hydrolyze the fluorine atom at
unspecified rates.1*0 Under physiological conditions of
pH and temperature, no fluoride ion is liberated in
72 96 horn's in the presence of any of a variety of ni-
trogen bases, sulfur compounds, and inorganic salts.2"
A further example of the chemical inertness of the
fluorine atom in fluoroacetates is given by the follow-
ing comparison of the rates of replacement of halogen
by sulfite in the ethyl esters of fluoroaeetic, cliloro-
acetic, and bromoacetic acids; 99
Temp
Bi molecular
Compound
C _
velocity constant
Kthyl bromoacctatc
25
18.3
Ethyl chlonmoetate
25
0.13
Ethyl fluoroacetate
45
4,5 X 10-»
Limited data on the storage .stabilities1’ of both
h A recon) report from the Chemical Warfare Service
{TCI H 345, Surveillance of Fluorine Compounds, September 5,
1015) testifies to the stability of methyl fluoroacetate and re-
lated compounds with respect to pressure development as
follows; (I) methyl fluoroacetate and 0-fluoroelhanol arc
stable in 75-mm steel shell with respect to pressure for at
least 1 year at tin C; (2) methyl 7-flnorobutyrate docs not de-
velop pressure in fi months at fi5 C when in contact with a
steel strip in glass apparatus; and (3) methyl -y-fluoro-
d-hydroxybntyratc develops a pressure of about 120 psi at
35 per cent void in glass apparatus at 65 C in either the pres-
ence or absence of a steel strip. CHEMICAL STKICTIHE IN RELATION TO TOXICITY
163
methyl fluoroacetate and sodium fluoroacetate also
illustrate the high stability of these com]K>unds.
Methyl fluoroacetate undergoes no visible change on
storage for S months at 00 (’ in glass containers in the
presence of varnished steel, but a slight deposit of
silica forms in the presence of bare steel.* Sodium
fluoroacetate undergoes no visible change, loses no
weight, and does not alter in fluoride content on
storage for 30 days at 65 (’ in tin-plated cans; the tin
surfaces show no change.47'-
Methyl fluoroacetate resists oxidation by aqueous
permanganate or chromate. In the presence of
chromic acid plus concentrated sulfuric acid, pro-
duction of hydrogen fluoride occurs, slowly in the
cold and rapidly on heating.so
Methyl fluoroacetate exhibits a thiosulfate de-
mand on heating at 100 C.sl
There is no evidence that other members of this
scries differ strikingly from methyl fluoroacetate in
the stability of (he fluorine atom, or that- they exhibit
peculiarities in the reactions of the more common
functional groups.
10.2.3 Detection and Analysis
The fluorine atom in compounds of the fluoro-
acetate type is too stable toward hydrolysis to make
practical the use of this reaction for purposes of
identification. Therefore, recourse is had to oxidative
or thermal decomposition producing hydrogen flu-
oride. which is then detected by its etching effect on
glass or by its ability to bleach metallic lakes of ap-
propriate dyes.,7i5i so.s'.ssi, y device making use of
the etching effect to produce a nonwet table surface
in small glass tubes has been examined by the Hrit-
ishd” v-■!W Hot platinum filaments and hot platinized
silica gel both decompose volatile fluorine compounds
and both have been utilized in experimental appar-
atus designed for field use.17 "
Satisfactory tests for fluoroacetate ion in water
have been developed.4,,M
All quantitative methods for determination of
fluorine in compounds of the fluoroacetate type have
involved the conversion of the organically bound
fluorine to fluoride ion, which is then determined by
one of the standard methods.
The detection and analysis of aliphatic fluorine
compounds are reviewed in more detail in Chap-
ters 34 and 37.
10.3 CHEMICAL STRUCTURE IN
RELATION TO TOXICITY 24i 44 , i ,
In Table 2 are listed representative compounds
which do and do not possess to a marked degree the
Table 2. Aliphatic fluorine
compounds illustrating Ihc relationship between molecular structure and toxicity. -
Compounds exhibiting definite fluoro-
Compounds exhibiting no or only
acetate*- or 7-fhiorobut yra t e-like
slight fluoroacetate- or 7-fluoro-
■
toxicity*
licfcrence
bulyrate-Iikc toxicity*
Reference
’ - — „
.4 rids
Fluoroaeetic acid
91 e, 92c
Difluoroacelic acid
13
Salts
Sodium fluoroacetato
13,20,34a
Sodium chloroacetate
13, 38c
Sodium y-fluorobutyratc
13
Sodium bromoaectatc
13, 3Sc
Sodi 11 in 7-fliiet hy 1 7-fluon >bnt yra te
Sec Table 4
Ethyl i-fluoro valerate
91 r
d-Fhioroethyl 7-fluombutyrate
See Table 4
Ethyl oj-fl uorohendcca noa te
9lr
Methyl 7-fluorolhiolbutyratc
Sec Table 4
Methyl
Sec Table 4
Methyl 7-fliioro-/J-hydroxybutyrate
See Table 4
*
SECRET 164
METHYL FLUOROAEETATE AM) RELATED COMPOUNDS
Table 2 (Continued).
Compounds exhibiting definite fluoro-
(omiKiunds exhibiting no or only slight
acetate- or ■y-fluorolmtyrate-like
fluoroacetate- or >-fluorobutyrate-
toxicity*
Reference
like toxicity*
Reference
Methyl y-fluoro-d-hydroxyt hiolbutyratc
See Table 4
Methyl -y-fluorocrotonate
See Table 4
Diet hyl fluoromalonatc
91c
Ethyl t-fluorooaproate
!Mo
(3-Fluoroethyl t-fluorocaproate
91r
Ethyl uf-fluorocaprate
!)lr
_
Anhydrides
Fluoroacetic anhydride
91c, 95c
Nitriles
*
-
Fluorojicctonit rilef
13. 91c
—
y- Fl u orobu tyron i t rile f
13
y-Fluorocrotoni t rile t
13
Trifluoroacctonit rile
13
Aldehydes
Fluoroacetaldchvde
91 o
Amides
Fluonwcetamidc
91b, 92b
X-d-chlon*et hvl fluoroacet amide
13
X -ni t r< iso-X -met hvl flm >n meet amide
91c
;
Arid halides
—
Fluoroacetyl chloride
13, 91c
Acetyl fluoride
13
Eluoroaeetvl fluoride
13, 91f
('hloroacetyl fluoride
13, 91c
— - ‘
Hutvryl fluoride
13
Crotonyl fluoride
13
—
,4 Icohols
3-Fluoroethanol
See Table 4
•y-FluoroppopanoI
59b
Esters of Fluoroethanol
mnnoUi-FluorOflhyl) derivatives
0-Fluorocthyl chloroformate
13
d-Fluoroethyl acetate
91 f
0-Fluoniel hvl fluoroacetate
See Table 4
d-Fluoroethvl ehloroaeetate
91c, 92g
- -
nitrite
13
.
Dichloro(/S-fluorocthoxy)phosphine
13
3-Fluorocthyl sulphury! chloride
9lf
—
hid 0-FIuoroel hyl) derivedives
his( 3-FI uoroel h yl) carbonat e
13
bis(3-F1 nonx>thy 1) fluorophosphate
91 f
bisfd-Kluoroethvl) ehloromaleate
13
6/s(3-Fluorocthvl) sulfate
13, 9lf,92g
—
Di-3-fluoroelhyl hydrogen phosphite
91 h
Ethyl his(3-fluoroethoxy )phosphinc
13 —
'
lris{ d-Fl uoroel hyl) derived ives
_ -
tn'M3-Flu-fluorobulyrate-likc toxicity would, for any specie* at doses equal to or slightly greater than
those listed
for methyl fluoroacetate or methyl >-fluorob«tyn»tr in
Tables 3 and 1, produce the characteristic symptoms after the usual latent period (see below), and
at least some deaths within 2 days.
—
f May posse** .slight activity, hut markedly less than corresponding esters.
t Produce methyl fluoroacetate symptoms but only
at somewhat higher concentrations.
SECRET TOXICOLOGY
165
Table 3. Toxicity of methyl fluoroacetate.
With the exception of the entries market! with an asterisk, the figures are approximations based on
limited numliers of observations.
Species
LC\„ (mg/1)
f = 10 min
Reference
Intra-
venous
Reference
LD,„ (mg kg)
Subcutaneous
Reference
Oral
Refer-
ence
Dog
0.025*
24t
0.08
241
0.1 0.2
73
0.1-0.2
73
Cat
(0.025-0.05)t
24 i
0.2*
40b
0.3
73
0.3
73
Rabbit
0.065*
241
0.33*
241
0.3 0.5
24 i, 73
0.5
73
Guinea pig
0.15
24d, Die
0.2
73
0.5
73
Goal
0.2
76
<2.0
51
1.0
73
1.0
73
Hat
0.3
24d, 73. 76
2.5
73
3.5
73
Mouse
3,2*
24f,t
17*
2 it
5-20
24i, 73
(5-0)
24 j, 73
Rhesus monkey
0.8 2.0
73, 76
5-15
51
10 12
73
10 12
73
Ccrcopitheeus monkey
* , .
>50j
01 q
Krog
100 200
40a
t Hstimatr based on susceptibility h> £pftuort*‘lhanol.
X Intraperilotiral injection.
'
characteristic toxicological properties of methyl
fluoroacotate and methyl y-fluorobutyrate, Com-
])oiiuds which produce the characteristic toxicological
actions fall into the following categories;
1. The following acids, in some cases tested only
as salts and esters: fluoroacetic, y-fluorobutyric,
y-fluoro-d-hydroxybutyric, /1-ehloro-y-fluorobutyric,
y-fluorocrotonic, t-fluoroeaproic, and w-fluoroeaprie.
2. Other simple derivatives of the above acids and
their thiol analogs, including anhydrides, amides,
aldehydes, and acid halides, but not the nitriles.
3. /J-Fluoroethanol, its esters, and certain other
derivatives.
The following compounds do not evoke the char-
acteristic toxic effects;
1. Di- and poly-fluoro derivatives of the toxic
mono-fluoro compounds.
2. Chlorine, bromine, and iodine analogs of the
toxic fluorinated derivatives.
3. Fluoride-liberating compounds such as acid
fluorides.
4. Derivatives of aliphatic acids in which the
fluorine atom is not in the terminal position (i.e.,
methyl a-fluoropropionate, methyl o-Huoroisobutyr-
ate, ethyl a-fluorobutyrate, and diethyl fluoromalon-
ate).
5. w-Fluoro derivatives of aliphatic acids with an
odd number of carbon atoms (e.g., ethyl 0-fluoro-
propionate, ethyl 5-fluorovalerate. and ethyl «-fluoro-
hendecanoate).
Thus, the F-CH»-group appears to be essential.
Its presence is not sufficient, however, and presum-
ably it must form the end of a chain of an even num-
ber of carbon atoms. It is also necessary that the
proper group, usually an oxygenated one, form the
other end of the chain (e.g., methyl fluoroacotate is
highly toxic, l-ehloro-2-fluoroethane is less so, and
fluoroacetonitrile Ls relatively nontoxic). That other
features of the molecules play a role in determining
the degree of toxicity by inhalation is also revealed
by the large differences which exist between the pre-
cisely determined LCi0'* for a number of related de-
rivatives containing one and twoF ■ CHi-groups,241 •"44
and by the large differences in toxicity which are
associated with various in methyl
y-fluorobutyrate (see Table 4).
tot _ TOXICOLOGY
id.1.1 Toxicity for Animals
The toxicity of methyl fluoroacetate for animals is
set forth in Table 3 and may be evaluated in com-
parison with hydrogen cyanide, the LDi0 of which
is in the order of 1 mg 'kg for most species, including
man (see Chapter 2). It is noteworthy that: (1) the
species variation is unusually large — the dog is ap-
proximately 100 times more susceptible than the
mouse or monkey and two tested species of monkeys
show considerably different susceptibilities; and
(2) the compound is approximately as toxic when
administered by mouth as when injected intrave-
nously or subcutaneously.
The toxicity of /3-fluoroethanol for various species
is comparable to that of methyl fluoroacetate; d-flu-
oroethyl fluoroacetate, the most toxic member of the
fluoroacetate group, is somewhat more potent
(Table 1). Methyl y-fluorobutyrate and related com-
pounds (Tables 2 and 4) produce toxic effects similar
in a general way to those of methyl fluoroacetate but
exhibit less pronounced species variation, principally
SECRET 166
MKTim. KLUOUOACET \TE \M> RELATED COMPOUNDS
Table 4. Inhalation toxicides of fluorinaled aliphatic compounds.
With the exception of the mouse LCsn’s, the figures are approximations based on limited data.
LCao (mg/1, nominal, for I —
10 min)
Monkey
Guinea
Compound
(Rhesus)
Mouse
Rat
P'g
Rabbit
Cat
Dug
Methyl fluoroacetate
0,8-2.O'*-18
3.2“**
03>*i,n.7«
0.15**1
0.065***
0.025s
0-Chloroelhyl fluoroacetate
0.7Me
0.2 ± ’*<’
0.15+**•'•*“
o.r*-
(i-Fhiornet h vl fluon meet a t e
0.63s**
0.2 ±,,J
0.07s** »w
Q.0591,1
fJ-Fluoroelhanol
1.5»M-«
1.2s-"
0.2+5M.76
0.15s'-'-*"-
0.025s*1*
0.03.VXI
(0.007)
Methvl y-fluorobutyratc
0.5S4b
0.12s*'
0.35+slh
0.07s1«•
0.035s**1
0.0351***
0.05s!">
Met liyl -,-fluorol hiolbut yr-
ate
0.004-11
d-Chloroet hvl y-fluorobut yr-
j
ate
>0.3-0
0.051s1)
0.1 + s'i
....
d-Kluorocthvl y-fluorobulyr-
9
ate
o.5+s*b
0.077s4'
0.2 s lh
0.035s*1*
<0.075=*
0.025s**1
0.025s*
Methyl d-ehloro-y-fluorobu-
tvratc
0.16s**
Methyl y-fluoro-^-hydroxy*
-
butyrate
0.2-|B
0.023s**"
. .T.
<0.063s*1'
0.P
<0.0(43-*
Mel hvl y-fluoro-jj-met hoxy-
butyrate
....
>0.1“"
>0. !«'■
Met hvl y-fluoro-0-hydroxy-
-r ;
thiolhutyrate
0.2-1P
<0.03S4P
...»
<0.063s*1,
0.063s “
/S-Chloroethyl y-flumo-d-hy-
—
—
droxybulyrate
0.048s*’
Met h vl y-fluoroerotonatc
<0.5Mh
0.080s*1
0.15s**1
....
Ix'cause of a much greater toxicity for mice. When
tested on monkeys, the members of this group are
more toxic than methyl fluoroacetate but not so
toxic as either mustard gas or phosgene.
Methyl fluoroacetate, and presumably also (he
•y-fluorobutyrate derivatives, are detoxified in the
laxly, but only at a slow rate.24'1* '-403-73-76'911’'4
Changes in with changes in exposure time
over the range I to 100 minutes have been observed
in experiments with methyl fluoroacetate, 0-Huoro-
ethyl fluoroacetate, and /3-fluoroethanol, but the
effects are not large.24'1*'73 76 The L{Ct)„o of methyl
■y-fluorobutyrate for mice is the same for exposures
of 1, 10, and 100 minutes,24* and that of methyl y-flu-
orocrotonate may not be significantly different for
exposures of 10 or 100 minutes or for two fractional
exposures at a 24-hour interval.241 Summation of the
effects of multiple sublethal doses of methyl fluoro-
acetate administered at daily intervals by mouth,
injection, or gassing has been observed but some de-
toxification occurs and, with sufficiently small incre-
ments, the equivalent of several lethal doses can l*e
tolerated.40m’73'il,f However, species differences appear
to exist; successive small doses produce a more pro-
nounced cumulative effect in guinea pigs than rats,
the latter species probably developing an increased
resistance to the poison.9,1 Indeed, recent data dem-
onstrate that a small dose (approximately 0.1 JJ)„o I
administered orally or subcutaneously confers a sta
tistieally significant, degree of resistance upon nib\
tested 24 hours later with an LD:,n administered;
orally or intramuscularly.470 A similar phenomenon:
has berm reported for orally administered sodium j
fluoroacetate, but it would not appear that the ele-j
vat ion of resistance is sufficient to affect the value of
the salt as a rodentieide.
A characteristic latency is associated with the
visible effects of poisoning by methyl fluoroacetate
and related compounds. Even 10 to 20 times the
lethal dose produces sympt oms only after a minimum
delay of 15 minutes.24'1 Survival times of animals dy-
ing as a result of inhalation of median lethal dosages
are almost always at least 1 hour, usually 2 to 12
hours, less frequently 12 to 21 hours, and rarely
longer.24'1'91' The derivatives of y-fluorobutyric acid
act similarly to the fluoroacetates but the latent
period may be somewhat briefer and the recov-
ery of sublethally poisoned animals more pro-
tracted.24*11’4
There are two immediate causes of death in methyl
fluoroacetate poisoning: action on the heart, culmi-
nating in ventricular, fibrillation and circulatory
failure; and stimulation of the central nervous sys-
tem, producing convulsions, apnea, and death with-
SECRET TOXICOLOGY
107
out severe cardiacr abnormalities.51 The relative
severity of the two effects is not the same in different
species: the cardiac action is the primary cause of
death in monkeys, goats, and rabbits; effects on the
central nervous system predominate in rats, cats,
ami dogs.4"515sk 73 Transient but sublethal central
nervous effects occur in some species (e.g., Rhesus)
which eventually die with ventricular fibrillation,
and cardiac effects in other species (e.g., the eat)
which die of respiratory failure following severe con-
vulsions.51 The poisoned heart has a decreased ex-
citability and the effects are not due to diminished
coronary blood How.51
In experiments on three monkeys methyl *y-fluoro-
butyrate produced cardiac depression and arrhyth-
mias, as well as marked parasympathetic symptoms,
but ventricular fibrillation has not lieen observed.58,1
/3-Fluoroethyl y-fluorobut vrate, likewise tested on
only three monkeys, produced effects similar to those
of methyl fluoroaeetate but was more toxic.58,1 Both
compounds produced effects similar to methyl fluoro-
acetate in the cat and rabbit.681
The symptoms associated with poisoning by
methyl fluoroaeetate and related compounds have
been deserilied in detail for various species ?4', E h1'1,
5i.58k,73,stia and can lx* interpreted as resulting from
the actions of the poisons on the heart or nervous
system, or both.
Pathological studies «».J h.7j »ik.., jn animals dying
acutely from single doses reveal no significant
changes other than signs referable to venous con-
gestion. In animals exposed repeatedly to sublethal
doses until death ensues,911 ,q there are found the
sequelae of protracted venous congestion att ributablc
to heart failure, definite abnormalities in the myo-
cardium, changes in the kidney which may or may
not be secondary to disturbances in the metabolism
of other organs, and changes of doubtful significance
in some other organs; no unequivocal pathological
changes have been observed in the nervous system.
in.t,2 Physiological Mechanism
A number of clinical pathological and biochemical
studies have been made to throw light on the cellular
mechanism of action of methyl fluoroaeetate and re-
lated con Ipou lids. l8"4,11' 2H,34,38.40,51,58l,.f .j.k.l, 73. 9U.li.g.j,
m.n.q.n.ss xiipir heterogeneous character precludes a
review of all the isolated facts whieli eventually may
prove to he of significance.
The evidence is strong that methyl fluoroaeetate
does not owe its toxicity to the liberation of fluoride
ion at critical loci in the body. In accord with chemi-
cal studies on the stability of (lie fluorine atom (sec*
Section 10.2.3), none of a large manlier of biochemi-
cally important substances,*1 including some with a
high reactivity toward organic halogens, liIterates
fluoride from methyl fluoroaeetate at physiological
conditions of yd I and temperature; -8 383 nor is fluoride
ion liberated when the ester is incubated with rat
tissues.18 Moreover, methyl fluoroaeetate does not
show a marked tendency to inactivate enzymes
which arc highly susceptible to fluoride.38"
It has been proposed as a working hypothesis that
all the lexicologically active compounds under con-
sideration may be the precursors of some common
toxic material, possibly the fluoroaeetate ion, which
could l»e. produced, for example, by hydrolysis of
esters, oxidation of /3-fluoroethanol, and /3-oxidation
of the 7-fluorobutyrates.91' Although this hypothesis,
which conceivably could explain the facts set forth
in Section 10.3, “Chemical Structure in Relation to
Toxicity,” has not as yet been submitted to sys-
tematic test, (lie following findings may be cited as
bearing upon it.
1. Sodium fluoroaeetate, fluoroacetic acid, and
fluoroaeetamide possess approximately the same
toxicity as methyl fluoroaeetate and produce symp-
toms after a comparable latency.51 01,1 The latency
in poisoning by the ester is not, therefore, determined
by time for hydrolysis. However, this does not imply
that hydrolysis of the ester may not be a necessary
prelude to the initiation of toxic action. Tissues and
blood contain a methyl fluoroaeetate esterase,18 38*
which in the rat is sufficiently active to afford the
ester a half life of not more than a few minutes — a
fraction of (he usual latent period for symptoms.18
2. That the characteristic effects of methyl fluoro-
acetate and sodium fluoroaeetate on the myocardium
do not require in vivo chemical changes in other organs
is suggested by experiments on eviscerated rabbits 61
« Tt may !«■ noted that the conclusions concerning cardiac
effects have been based on detailed, continuous electro-
cardiographic observations,** and that species which exhibit
central nervous stimulation concomitantly develop abnormal
electroencephalograms in the absence of notable cardiac
irregularities.578
d Arginine, serine, histidine, tyrosine, proline, asparagine,
glutamic acid, lysine, tryptophane, alanine, glycyl glycine,
imidazole, guanidine, cysteine, glutathione, S-allyl thiourea,
d-mercapt oet Hanoi, 2,3-dimerca ptopropani >1, carbobcnzoxy
methionine, Ihiodiglycol, benzylamine, triethanolamine, tetra-
cthanolarnnionium chloride, hexamethylenetetramine, and
d-aminolienzoic acid; or sodium thiosulfate, sodium sulfide,
sodium bisulfite, or sodium iodide.
SECRET 168
MKTin L ELI OROACETATE AND RELATED COM 1*0 U NDS
and proved by tests with isolated, perfused hearts of
the cat.51 rabbit,91 ,n and guinea pig,31" and with the
isolated papillary muscle of the cat.51 A similar con-
clusion with respect to effects on the central nervous
system is suggested by the finding that local appli-
cation of methyl fhioroacetate to one cerebral hemi-
sphere produced convulsive discharges after the usual
latency for symptoms; although the convulsions were
generalized, the effect of the poison on the treated
hemisphere ajqx’ared to be greater than on the con-
tralateral areas.31*
3. On the contrary, it may be necessary for /3-flu-
oroethanol to undergo chemical change, possibly by
oxidation to fhioroacetate in the liver. This is sug-
gested by the finding that the alcohol exerted no
effect on the isolated, perfused heart when tested at
concentrations at which methyl fhioroacetate pro-
duced marked decreases in rate and survival time.31*"
4. If the toxicity of the y-fluoro butyrates and
other longer-chained nominated aliphatic acids de-
pends on the production of fhioroacetate by 0-oxida-
tion, their relatively high toxicity for some species
(Table 2) would require that they l>e concentrated
to a greater degree than the fiuoroacetates at critical
loci in the body. That the /3-oxidation of 7-Huoro-
bot y rate is not prerequisite for all its actions upon
biological systems is indicated by evidence that,
methyl 7-fhiorobutyrate is not converted to Huoro-
acetate by rabbit kidney cortex in vitro, in spite of
the fact (hat both compounds markedly inhibit the
oxidation of acetate by this preparation.38* Substitu-
tions on the /3-carbon atom are, however, important
determinants of inhalation toxicity, as is revealed by
the widely differing toxicities of a number of the
butyric acid derivatives listed in Table 4.
Changes indicative of a derangement in carbo-
hydrate metabolism in methyl fhioroacetate poison-
ing in various mammalian species are increases in
blood sugar,58,173 non protein nitrogen,40*-75 inorganic
phosphate,58j lactic acid,40*-5801 pyruvic acid,5Kb and
lactate-pyruvate ratio.586 In rabbits there is a marked
reduction in liver glycogen 58i and, in the heart,
marked decreases in total acid-soluble phosphorus
and organic soluble phosphorus.5** Serum potassium
and calcium show only minor increases.588 75
Negative results have been obtained in many but
not all the studies on the effects of fhioroacetate on
enzyme systems in vitro and on the metabolism of
tissues obtained from poisoned animals or treated
with the poison after isolation.18-34 3*-40-58b f-,-3!* 38
Illuminating experiments have been performed
with an isolated skeletal muscle, the sartorius of the
frog, hut have not lx*en extended as yet to cardiac
muscle.18-401’The resting oxygen consumption and
the contractility of the* unfatigued sartorius are not,
affected by the poison at a concentration of 0.005.1/,
but the extra oxygen consumption following activity
is strongly inhibited.18A01’*1 The inhibition is associ-
ated with a greatly decreased resynthesis of phos-
phocreatine1X and abolition of the delayed heat
production normally associated with aerobic re-
covery of stimulated muscle.401’r Similarly, the extra
oxygen consumption produced by pretreatment of
the isolated muscle with the stimulants caffeine and
dinitrophenol is essentially abolished by fluoro-
acetate.1*-40' Similar changes are produced by sodium
azide but the mechanism of action is different: azide
inhibits cytochrome oxidase and adenyl pyrophos-
phatase, whereas methyl fhioroacetate has no in-
hibitory action either upon these enzymes or upon
cytochrome reductase.18
The possibility jhat fhioroacetate inhibits lactic
acid dehydrogenase is suggested by the findings that
the isolated frog sartorius utilizes pyruvate (also
acetate) but does not oxidize added lactate,40'' and
that the effects of fhioroacetate upon the isolated
guinea pig heart are counteracted by pyruvic acid
derivatives but not by sodium lactate.31'" An iiuiitm
study on lactic acid dehydrogenase likewise revealed
an inhibition of the enzyme prepared from yeast,40,1
although in another experiment the enzyme pre-
pared from heart muscle was not reported to lx* in-
hibited by methyl fhioroacetate.3*' Data relating to
the production of lactate by the stimulated poisoned
muscle under anaerobic conditions are not con-
sistent.18 401’ In the case of rabbit kidney cortex prepa-
rations in vitro, however, methyl fhioroacetate in-
hibits the oxidation of glucose and certain intermedi-
ates of carbohydrate metabolism but it has no effect
on the anaerobic phase of carbohydrate degradation
resulting in the formation of lactic acid. The latter
findings have led to the suggestion that a locus of
action may be at one of the steps in the dehydro-
genation of pyruvate via the citric acid cycle.18
The effect of fhioroacetate on the oxygen con-
sumption of stimulated skeletal muscle has been
found to be reversible,1* offering some hope that
methyl fhioroacetate poisoning may eventually be
subject to treatment. Moreover, if the oxygen con-
sumption of heart muscle should prove to lx; as
easily inhibited by methyl fhioroacetate as that of
stimulated skeletal muscle, the possibility would
SECRET TOXICOLOGY
1G9
exist that a therapeutic agent could be found in a
carbohydrate intermediate the oxidation of which is
not strongly inhibited.1*
The slight therapeutic value of procaine and p-
amiuobenzoic acid in fluoroacetate-poisoned monkeys
and the absence of a corresponding effect with other
antifibrillatory drugs (see Inflow) suggested the al-
ternative possibilities that p-aminolx*nzoic acid
might l>e fluoroacctylated, thereby detoxifying the
poison, or that the toxicity of the fluoroacetate might
be associated with an inhibitory action on normally
occurring acetylations. However, experiments reveal
that (he monkey does not acetylate, and therefore
probably does not fluoroacetylate, p-aminobenzoic
acid,5*1 that the acetylation of p-aminohippuric acid
by rabbits is not markedly affected in fluoroacetate
poisoning,341, and that fluoroacetate does not inhibit
the acetylation by liver slices of sulfanilamide, p-
aminoljenzoic acid, or choline.3** h On the other hand,
fluoroacetate does produce some inhibition of the
utilization of acetate in vitro by rabbit heart and
kidney 18 preparations and by rat kidney, liver, and
heart slices.3*1’ In (he case of Corynebaeterium rrca-
tinovorar and of yeast, the inhibition is almost com-
plete.3** These and other findings have led to the sug-
gestion that fluoroacetate may produce a profound
alteration in the metabolism of carbohydrate by vir-
tue* of a specific inhibitory effect on the oxidation of
acetate.3*0 However, poisoned caffeine-stimulated
muscle does utilize acetate.4110
The resting potential of frog peripheral nerve is
sensitive to concentrations of methyl fluoroacetate
as low as 0.001.1/.340The potential was reduced in
poisoned nerves by a period of anoxia, and the oxi-
dative recovery was little affected by addition of
acetate or acetyl phosphate, but was counteracted
by addition of pyruvate.
to.t.:t Therapy
* No satisfactory procedures for the treatment of
fluoroacetate poisoning have been discovered. Tests
have been made of substances and procedures de-
signed to prevent convulsions, to stimulate respira-
tion, to stimulate diuresis and excretion of the poison,
to prevent ventricular fibrillation and otherwise* re-
store the failing heart, to promote detoxification by
fluoroacetylation, to compete for enzyme systems
with fluoroacetate, and to supply necessary metabo-
lites the formation of which may lx* cut off by the
action of the poison on enzyme systems.1* •*4r 151 ■
4fte,e,i,i,k,l,73.»l«.i.in
Intracardiac inject ions of procaine accompanied by
artificial respiration and cardiac massage through the
thoracic wall temporarily restore an organized beat
to the monkey heart fibrillating as a result of methyl
fluoroacetate poisoning; but fibrillation recurs and
eventually proves fatal in spite of continued treat-
ments and tlie presence of subcutaneous deposits
of procaine.51 ssd
Administration of large doses of sodium p-amino-
ben zoa t e t o a n esl h e t i zed monkeys{Rhesus and Afrits)
poisoned with one LDMOdose of methyl fluoroacetate
corrects the cardiac disturbances and saves the ma-
jority of animals.5** * However, (he value of this
treatment is limited inasmuch as it does not save
monkeys poisoned with larger doses5*' or rabbits
poisoned with four /,/>i0’s;-*1"' nor does it combat
the lethal action of methyl fluoroacetate in the rat,
a species which dies of central nervous lather than
cardiac effects.581
Large concentrations of sodium acetate (0.1 per
cent) prolong the survival of the isolated rabbit
heart perfused with depressant concentrations of
methyl fluoroacetate, but the acetate ion has exerted
little or no protection when administered to the
poisoned animal.4*1 Similarly, the sodium salt and
other derivatives of pyruvic acid protect the isolated
guinea pig heart poisoned with methyl fluoroacetate
but have little or no therapeutic value in vivo A""
Various anesthetics have been shown to l>e effec-
tive in controlling the convulsions associated with
methyl fluoroacetate poisoning,-’41 51 -91*-' but even
when combined with respiratory stimulants they do
not decrease the mortality.*1* Sodium pentabarbital
is contraindicated because it increases the mor-
tality.81'
The following additional substances and proce-
dures have been without significant value under the
tested conditions in saving the lives of animals poi-
soned with methyl fluoroacetate or /3-fluoroethanol:
artificial respiration;91* artificial respiration plus
sodium phenobarbital;-4f 73 oxygen plus carbon di-
oxide;73 urethane, paraldehyde, or chloral hydrate,
with or without theophylline or coramine;9lE the-
ophylline; Mr,91i bromide;91* Dilantin (sodium di-
phenyl hydantoin); 73 91* morphine hydrochloride;91* '
quinidine, digitalis, quinidine-digitalis, or caffeine;5’9'
papaverine;5*' yohimbine;581 anticholinesterase drugs
or aconite;5** atropine;’40 5*0 ephedrine;iHF0 thia-
mine;5*' glucose;54* potassium salts;5,e*,m calcium
salts; 240 •Mc barium salts;5** 2,3-dimercaptopropanol
(HAL), acetophenone, or cobalt acetate.91'"
SECRET 170
METHYL FLL’OIIO ACETATE AMI RELATED COMPOUNDS
10.4.4 Toxicity for Man
Alan is among the species which are relatively re-
sistant to methyl fluoroacetale. Direct evidence
comes from the results of ingestion of the compound
by a British volunteer.91' Upon taking an oral dose
of 0.1 mg kg in water he experienced no symptoms
other than a slight, possibly psychogenic, feeling of
unsteadiness upon standing up l}4 hours after the
dose was taken. Similar ingestion of 0.65 mg kg pro-
ducer! no symptoms other than a feeling of unsteadi-
ness for a few minutes 1 hour after the dose and a
slight malaise 5 hours later; however, the subject
continued work in the lalxiralory with no obvious
loss of efficiency and his electrocardiogram and elec-
troencephalogram, recorded at frequent intervals,
showed no deviation from the normal, ft is to lx>
noted that the dose ingested was greater than (he
LDhp for guinea pigs, rabbits, cats, and dogs.
Various lines of evidence 73 80 suggested that the
lethal dose per os is in the order of 6 8 mg kg. Ex-
posure of workers for prolonged periods to low con-
centrations of the vapor9* produced marked weak-
ness, reluctance for any physical effort, and strong
mental depression with periods of nervous irritation
difficult to control, followed by physical and mental
exhaustion, drowsiness, and giddiness; a few days’
rest resulted in marked improvement.e
Assuming (1) that the above estimate of the lethal
dose per os for man is correct, (2) that the toxicity of
methyl fluoroacetatc is more or less independent of
the route or rate of administration, and (3) that
100 per cent absorption of inhaled vapor occurs, one
may calculate that the lethal vapor dosage for a
70-kg man breathing 10 1pm (relative inactivity)
would lx* 50,000 mg min m3, corresponding to a
10-minute LCm of 5 mg 1; for a man breathing 40
1pm, corresponding to exercise intermediate bet ween
a walk at 5 mph and a slow run,- the figure would be
12,500 mg rnin m3, the equivalent of 1.25 mg I for
10 minutes. Although the validity of this method of
calculation has been questioned,7* it has been shown
to yield good approximations when applied to the
flog and rabbit, the only larger species for which
both the /.('.mi’s and LI),n’s have lx*en determined
with precision.*4* For defensive purposes the British
have estimated the L(Ct)-M at 4,000 and 7,000 mg
min in3.76-79 For most species the margin between
the convulsive and lethal doses is small.
The mild, indistinctive odors of methyl fluoro-
aeetato and 0-fluoroethanol make it possible that
large vapor dosages could lx* inhaled undetected. It
has liven reported that methyl fluoroacetale at
0.05 mg 1 is just detectable, that at 0.2 0.3 mg 1 it
would easily lx* overlooked, ami that at 0.1 mg 1 it
possesses a fruity smell and may produce a slight
feeling of tightness in the chest.79 a,<- Most of the
7-fluorobutyrates are probably somewhat more odor-
ous than methyl fluoroacetafe but it has Ijoen empha-
sized that methyl y-fluoro-d-hydroxy butyrate pos-
sesses only a very slight odor, similar to but much
fainter than that of ethyl lactate.-'1’
A prominent symptom in severe poisoning is the
occurrence of repeated and severe convulsions indis-
tinguishable from status epileptieiis;73 less dramatic
symptoms may include nausea, vomiting, dizziness,
and fall in body temperature.*1" In addition to the
symptoms, tests with urine mar aid in the recog-
nition of fluoroacetatc poisoning, for a toxic, lluorinc-
containing substance not present in normal urine is
excreted.91* ' n q The fluorine may be converted to
fluoride and detected by chemical test 9"’ or the
urine given to rats by stomach tube, the character-
istic symptoms of fluoroacetale poisoning73 then
Ix-ing produced.*1" q
In the absence of specific therapeutic procedures
for fluoroacetale poisoning, cases can at present only
be treated symptomatically. Morphine has been
recommended to allay distress, anxiety, and con-
vulsions, but barbiturates (i.e., pentobarbital so-
dium) are contraindicated. 9,*_
to.3 EVALUATION AS W AR GASES
Evaluation of the potentialities of methyl fluoro-
acetafe and related compounds in terms of available
data and present concepts of chemical warfare indi-
cates that none of the derivatives possesses the gen-
eral utility of currently standardized gases. They
remain a subject of some military concern, however,
in view of (heir potential use as food and water
poisons (see Section 10.6) or for other -penal pur-
poses.
For man methyl fluoroacetatc is not appreciably
more toxic, and in all probability is considerably less
toxic, than currently standardized gases. The lethal
vapor dosage, calculated above on the basis of the,
demonstrated low oral toxicity to be in (he order of
12,000 mg min m3 for ventilation rates correspond-
ing to moderate physical activity and several times
* These symptoms, experienced by Polish chemists,
prompted the initial toxicological examination of the effects
of methyl fluoroacetate on animals.
SECRET EFFECTIVENESS VS FOOD AND WATER POISONS
171
this value for men at rest, may he compared with the
minimum dosages of standard agents currently
recommended as adequate for the following tasks:71
comparable to methyl tluoroacetate range from that
of methyl tluoroacetate (i.e., 119 mg 1 at 25 (.’) down
to very low values. Thus, agents of any desired de-
gree of persistence are potentially available. Al-
though the indistinctive odor and relative difficulty
of detection by chemical means would confer upon
these compounds a certain insidiousness, their lack
of effectiveness on the eyes and skin renders them
inferior in general utility as persistent agents to such
vesicants as mustard gas and fiv.s(/3-chloroethyl)-
amine (HN3). Except in drinking water, (heir decon-
tamination offers no special problems.*0
Methyl tluoroacetate possesses excellent storage
stability (see Section 10.2.2) and its explosion sta-
bility is believed to be sufficient to permit its dis-
persal from chemical munitions now in use.2,i It is
potentially available in quantity.
There are no data bearing upon the toxicity for
man of the derivatives of y-fluorobu lyric acid. On
the basis of the comparative toxicides of these de-
rivatives and of methyl tluoroacetate for the monkey
(Table 1), they would be suspected of being some-
what more toxic for man than is methyl tluoroacetate;
They are, however, less volatile (Table !) and nota-
bly more difficult to manufacture.
10.6 POTENTIAL EFFECTIVENESS VS
FOOD AND \\ \TEU POISONS IN
\\ VRFARE112* 0-7*-7* *
The chemical and toxicological properties of
methyl tluoroacetate and related compounds make
them potential water and food poisons. They are
approximately as toxic when administered orally as
when injected or inhaled; to a degree they may act
as cumulative poisons; and they are not readily de-
tected by smell or taste. Although methyl fluoro-
acetate itself undergoes hydrolysis, the resulting
fluoroaeetic acid is stable and toxic.
At concentrations of 0.1 per cent or less in water,
methyl tluoroacetate has no smell or taste; *6 ]/% 1 of
a 0.1 per cent solution would probably be lethal for
man (see Section 10,4.1). This concentration in milk
is readily accepted by rats and dogs,1*2 although it is
not freely accepted in otherwise pure drinking water
by rats, nor are /3-fluoroethanol and sodium fluoro-
acetate at 0.01 percent (100 parts per million) freely
accepted by mice.21' - The effectiveness of sodium
fluoroaeetate as a rodent bait poison is discussed
below.
Filtration of contaminated water with charcoal
Dosage
Task
Agent
(mg min nr1)
To produce a large propor-
Phosgene
3,2tK»
tion of deaths or severe
Hydrogen
casualties in surprise at-
cyanide
5,000
tacks with nonpersistent
Cyanogen
gases (dosages to lx- ob-
tained within 2 minutes)
chloride
11,000
To produce skin burns of
Mustard
sufficient severity to to-
vapor
1,000(T > 80 F)
tally disable 50 |ht cent
2,000 4,000
of masked troops not
equipped with protective
clothing (dosages to—he
obtained within 4 hours)
—
(T = (10-80 F)
To produce eye damage of
Mustard
sufficient severity to
cause temporary blind-
ness among troops not
wearing gas masks
vapor
200
In view of the relatively low toxicity for man, it is
apparent from a consideration of (he physical proper-
ties of methyl fluoroaeetate (Table 5), the most
volatile stable compound of the group,1 that it would
Tabus 5. Physical properties of methyl fluoroacetate and
of currently standardized nonpersistent agents.
Property
Methyl
fluoro-
acetate
Ilydro-
gen
cyanide
Cyano-
gen
chloride
Phosgene
Liquid density
(g/ml at 25 C)
1.17
0.68
1.2
1.36
Boiling point, C
104
26
12.6
8.3
Freezing |>oint, G
— 35
-13.4
-7
-104
Latent heat of evap-
oration, e;d, g
100
210
135
GO
Vajxjr pressure.
nun Ilg
at 25 C
20
740
1,200
1,400
at -20 C
ss
180
230
Volatility, mg/1
at 25 C
119
1,060
at -20 G
145
680
1,400
be more difficult than in the case of the standard
nonpersistent gases to achieve in the field vapor
dosages sufficiently large to be lethal in surprise at-
tacks; and, in view of the effectiveness of the can-
ister,*0 the breaking of the gas mask cannot be con-
sidered a feasible task.
The volatilities of /3-fluoroethanol and of various
stable fluoroaeetate derivatives having toxicities
1 The more volatile p-ffiioroethyl nitrite, fluoroacetyl fluo-
ride, and fluoroacetyl chloride arc chemically unstable. 172
METHYL FLUOROACETATE AM) RELATED COMPOUNDS
removes methyl fluoroacetate but not the hydrolytic
product, fluoroacetic acid.45 Filtration with charcoal
plus pyridine is said to remove not only methyl flu-
oroacetate but also, from neutral solution, sodium
Huoroacetate as well.44 The detection of fluorine com-
pounds in contaminated water is discussed in
Chapter 54.
10.7 USE AS RODENTICIDES
Sodium fluoroacetate was one of several substances
studied in the chemical warfare program in the
United Kingdom and the United States which were
recommended by Division 9 of NDRC to/he Fish
and Wildlife Service of the Department of Interior
for test as rodenticides.4' Preliminary tests with
small samples (200 lb) submitted by Division 9 were
so successful that the division subsequently pre-
pared an additional 1,000 lb - for large-scale field
trials. Field campaigns in a number of states and in
military establishments in this country and abroad
were conducted by the Fish and Wildlife Service, by
the Typhus Control Unit of the Public Health
Service, and by the medical departments of the
Army and Navy. The results demonstrate that
sodium fluoroacetate, coded 1080, is one of the most
promising available roden t icides.46d *0‘744
Sodium fluoroacetate possesses the following re-
quirements of a good rodent icific: high toxicity and
acceptability, stability, lack of volatility, lack of irri-
tation and toxic properties for human skin, lack of
inflammability, and potential availability in quan-
tity at reasonable cost.464’ The oral lethal doses for
various species of rats and other rodents of concern
in public health and agriculture range between 0.1
and 5 mg kg.41*100 The substance is effective in baits
at much lower concentrations than in the case of
other rodenticides. Excellent results have been ob-
tained in field trials utilizing 0 oz of sodium fluoro-
acetate per 250 lb of cereal or ground meat bait.4"*
Water solutions are also highly effective. A concen-
tration of I 2670 has been recommended for general
use,70 although concentrations ten times greater
(i.e., 5 oz gal) are sufficiently acceptable to rats and
have been used with good results.4"'
As in the ease of other rodenticides, the possibility
of accidental human poisoning cannot lie ignored,
particularly in the absence of effective methods for
treatment of fluoroacetate poisoning. However, it
would appear that concentrations sufficiently low to
make accidental human poisoning improbable may
still be effective in rodent control. The human lethal
dose is believed to be of the order of 5 10 mg kg
(see Section 10.4.4). On the other hand, the oral
lethal doses for eats and dogs are very low (0.1 to
0.5 mg kg) and, therefore, the likelihood of acci-
dental poisoning of these species confers a certain
disadvantage upon sodium Huoroacetate.
The sodium salts of y-fluorobutyric, y-fluoroqS-hy-
droxybutyric, and y-fluorocrot onic acids are several
times as toxic for rats as is sodium fluoroacetate.
However, in view of the already high toxicity of the
lat ter, this apparent disadvantage is more than offset
by the greater difficulty and expense of their prepa-
ration.
SECRET Chapter 11
CADMIUM, SELENIUM, AND THE CARBONYLS OF IRON
AND NICKEL
By John .1. Zapp
ill INTRODUCTION
IN tiik skakch for new chemical warfare agents,
the toxic properties of certain metals were not
neglected. The increasing use of cadmium in indus-
try, for example, had revealed that the inhalation of
finely divided cadmium metal, the oxide, or salts was
capable of producing severe lung edema comparable
with that produced by phosgene.**-*8 Selenium com-
pounds showed similar properties,80 and, although
somewhat less toxic than cadmium on an absolute
basis, they produced physiological effects much more,
promptly. Being inorganic, these agents offered
promise for inclusion in burning-type munitions or
incendiaries as well as for dispersion by high-ex-
plosive shell. The carbonyls of iron and nickel
aroused considerable interest not only liecause of
their inherent toxicity, but also because they break
down catalytically in contact with gas mask carbon,
yielding carbon monoxide which is not absorbed in
the canister but passes into the mask.9*-1’ Thus the
carbonyls might be valuable in attacking either
masked or unmasked troops, (’om pounds of mercury,
thallium, tin, antimony, lead, chromium, and ger-
manium were screened by the University of Chicago
Toxicity Laboratory [LTCTL],7 but without reveal-
ing any of special interest for chemical warfare
purposes.
The part of Division 9 of the National Defense
Research Committee [NDHCj in the field of heavy
metals was largely one of screening the toxicity of a
great number of compounds, many of which were
prepared under Office of Scientific Research and
Development [DSHD] contracts. The detailed in-
vestigation of the promising compounds, including
investigations of cadmium and selenium and the
carlHHivls, was carried out by the Chemical Warfare
Service and by the Directorate of Chemical Warfare
in Canada.
11.2 CADMIUM
n.2.1 Physiological Action
Cadmium, its oxide, and salts are toxic by any
route of administration, but their particular signifi-
ounce in chemical warfare lies in the fact that finely
divided dusts can l>e set up either by thermal com-
bustion of incendiary mixtures containing cadmium
or by the explosive dispersal of cadmium compounds.
These- dusts are quite toxic by inhalation,*4-*8 pro-
ducing lung edema comparable with that observed
in phosgene poisoning.
Exposure to high concentrations of cadmium
causes some early respiratory irritation, which pro-
gresses to marked dyspnea within a few hours. Two
cats exposed to a high concentration of cadmium
oxide fume for 30 minutes 66 showed on autopsy ex-
tensive acute pulmonary injury with edema, injury
to the bronchioles and alveolar ducts, and acute
alveolar emphysema. Liver and kidney damage was
also found. Exposure to lower concentrations of
cadmium fumes or dust results in a temporary irri-
tation of the respiratory tract which disappears
shortly after cessation of exposure only to reappear
within about 12 hours with increasing severity ac-
companied by general malaise. Within about 24 to
48 hours dyspnea is marked and cyanosis occurs
prior to death.10-1™•“ On autopsy the lungs are
found to lie firm, but with interstitial and perivascu-
lar edema and extensive hemorrhage. Liver and kid-
ney show evidence of fatty infiltration.
Several cases of human poisoning by inhalation of
cadmium have been reported. In one of these, re-
porter! in 1858,60 three men were exposed to cadmium
carbonate dust. Symptoms did not occur until sev-
eral hours after exposure and then consisted of
dyspnea, dizziness, vomiting, and diarrhea. One
patient apparently cont racted pneumonia by second-
ary infection, but all three recovered eventually.
Fifteen cases of human cadmium poisoning from in-
halation of cadmium oxide fumes, two of which were
fatal, have been reported from Canada.67 In all these
cases, dyspnea, which did not become severe until
several hours after exposure, was the most prominent
symptom, although the majority of cases also ex-
hibited gastrointestinal symptoms. The two men
who died showed congestion of the lungs, pulmonary
edema, hemorrhage into the lungs, atelectatic areas,
proliferative interstitial pneumonitis, and catarrhal
SECRET
173 174
CVDMII M, SEIXML'M, AM) CARBONYLS OF IKON AM) NICKEL
bronchitis. Liver and kidney damage was also
present.
11.2.2 Toxicology
The toxicity of cadmium oxide by inhalation is
summarized in Table 1 for various species and ex-
ditions where particles do not agglomerate,49 It is
worth noting that prior to the Canadian experi-
ments4'* there was a tendency to assume that the
toxicity of cadmium oxide for man would closely re-
semble that for the monkey, making the human
L(Ct)„o of the order of 15 mg min 1. This estimate
would seem to bo entirely too high. The true toxicity
of cadmium oxide for man may be as great or greater
than that of phosgene (see Chapter 3).
The toxicity of cadmium metal itself ami of cad-
mium compounds other than (he oxide by inhalation
was tested at the UCTI,. The results are shown in
Table 2. So far as mice are concerned, there is con-
Tabi.k
1. Toxicity of cadmium oxide by
inhalation.
Kxposure
T4Ct)H,
time
S|X'cies
(mg inin/1)
(min)
Reference
Mouse
0.5
15-30
60
0.87
10
17
0.58
10
15
0.34
10
7
Rat
2.0—
2
40
l.l
5
40
0.78
10
to
0.0
30
40
1.3-1.8
5-10
46
Guinea pig
3.0
15 30
60
Rabbit
3.0
15-30
60
>1.8
5 10
46
<5.2
10
IS
Goat
<1.6
5-10
46
Dog
3.0
1(T
45
Monkey
15-20
-40
45
-
15 ±
15
45
21 +
;«)
45
Man
1.5-2.0
75-90
49
Table 2, Inhalation toxicities of
for mice.7
A = analytical concent ration;
concentration.
cadmium compounds
X = nominal
Compound
Ct (us
compound)
(mg min 1)
Ayr.
Ct particle
(as Cd) diameter
(mg min/1) (m) Mortality
Cadmium
0.38 A
0.38
<0.2
18,20
('admium
0.17 A
0.17
<0.2
18/19
Cd oxide
0.31 A
0.30
<0.2
uctu
Cd chloride
2.3 A
1.4
<0.5
lACtU.i,
CM fluoride
1.8 N
1.2
?
0/20
CM fluoborate
0.5 N
1.9
?
8/20
Cd fluosdieale
0.7 X
2.1
?
9/20
CM sulfide
1.35 A
1.05
<0.3
5/20
CMsclcnate
2 27 -X
0.03
?
0/20
CM nitrate
3.85 A
1.4
<0.5
UCtu,,
CM phosphate
6.5 X
3.07
?
2/20
posure times. Some of the variability in results is un-
doubtedly due to variation in particle size of the
cadmium oxide. When disperser! from most incendi-
ary munitions the median particle size is usually less
than 1.0 n in diameter, but agglomeration of particles
frequently takes place. This point has been particu-
larly emphasized in the estimate of the L(Ct)i0 for
man,49 in which the elementary particles were less
than 0.3 n in diameter, but in which the cloud actu-
ally consisted of large numbers of small agglomerates
of 1.0 to 2.0 n in diameter, with a small number of
agglomerates 40 n or greater in diameter. The L{Cl)i0
of 2,0 mg min 1 was based on analytical CV’s ob-
tained in an experiment set up to duplicate condi-
tions which resulted in two cases of fatal cadmium
oxide poisoning in an industrial plant. There ap-
peared to be every reason to believe that the concen-
tration of cadmium oxide in the original accident
was not greater than that obtained in the duplicate
experiment, but the L(r/)50’s for rats and rabbits
exposed in the duplicate experiment were about
twice those previously obtained with arc-produced
cadmium oxide fumes. The difference was attributed
to the greater median particle size in the duplicate
experiment and led to the hypothesis that the L{Ct)h0
for man might be as low as 1.5 mg min 1 under eon-
siderable variability in I ho t oxicity of the difTercnt
cadmium compounds even when dosages are calcu-
lated in terms of the cadmium content of the com-
pound. It is also of interest that cadmium metal
itself is more toxic than any of its salts. In this in-
stance, the combination of cadmium with anions
which are themselves toxic resulted in decreased
rather than enhanced toxicity. Unfortunately it is
not possible from the data to assess the effect of
part icle size on the different est imates of the o’s
but taking the results at their face value it would
appear that cadmium oxide is (he most toxic of the
cadmium compounds. This fact is fortunate since
the oxide is easily prepared in the field by the com-
bustion of incendiary munitions containing cadmium
metal.15'-4"*’4®
11.2.3 Assessment of Value as a
Chemical Warfare Agent
Cadmium appears to be a promising material for
addition to incendiaries if toxicity as well as fire is
SECRET SELENIUM
175
Tabi.k
3. Toxicity of selenium dioxide by inhalation.
Kx) insure
Time to
Cl
t ime
death
S|>ecics
(ms min/I)
(miu)
Mortality
(hr)
Reference
Mouse
2.30*
10
0 20
7
Hat
2:30t
- 10
0/6
7
Rabbit
o.89t
20
4/6
6, 6.5, 40, 132
27
6.59t
10
4 /6
3.8, 11, 13,32
27
13.1Ht
20
6/6
2.8, 3, 3, 5, 5.5, 8
27
Goat
5.89t
20
0.2
27
6.59f
10
2/2
5.5, 84
27*
—
S.83f
30
3/4
6, IS. 130
27
13.i8t
20
2/2
4,4.5
27
* Set i* di?»| termed by
atomization of aqueous solution.
-
f SeO? formed and disperst-d by detoiuiti
ion of Sr;liigh-explowive mixture, IVak range of particle size 0.0 to 1.0 p in
diameter.
desired. The cadmium oxide which results from the
combustion of cadmium metal in incendiary mixes
is odorless and probably not irritating enough in the
presence of smoke and burning materiel to be de-
tected by odor.
Cadmium oxide smoke is brown in color, however,
and may he detected by appearance after it has been
used a few times. Cadmium chloride may also be dis-
I>ersed from burning munitions IS:’V” and lethal con-
centrations may lx* obtained in mixtures which are
indistinguishable in appearance from harmless
screening smokes. Attempts have been made to dis-
perse cadmium compounds by the explosion of mu-
nitions containing cadmium or its compounds,
5» 55,64 ]ulj this method of dispersal is relatively in-
efficient because of the rapid .agglomeration and
settling of the cadmium part icles.
One drawback to the use of cadmium in offensive
warfare is the delay in appearance of toxic effects,
since, as has I teen pointed out, dyspnea does not
usually become severe until at least 12 hours after
exposure. If. however, incendiary at lacks are planned
against industrial installations, large stores of ma-
teriel, cities, and the like, such targets are usually
well beyond the front lines and a delay in the appear-
ance of toxic effects can lx* readily accepted. From
the available data it would appear that cadmium
might play a very important role if a military re-
quirement for toxic incendiaries should arise.
11.3 SELENIUM
A review of the toxicity of selenium as a potential
indust rial hazard appeared in 1938.** At that time it
was known that selenium compounds were toxic
when ingested and that hydrogen selenide was toxic
on inhalation. On this basis it was predicted that
soluble dusts such as selenium oxides (SeO>, Se03,
i CScOrt, I LSeO,) and certain halogen compounds
might lx- toxic Ix'eanse of the ease by which they
could lie absoii>ed from the lungs and gastrointestinal
tract. These toxic dusts might he set up by the com-
bustion of incendiary mixtures containing selenium
or its compounds or by the detonation of explosives
containing selenium. Hence selenium, like cadmium,
was investigated as a possible chemical warfare
agent.
tl.3.1 Physiological Action
The action of selenium appears to be similar to
that of cadmium, with the exception that the onset
of toxic effects is rnoiv rapid after exposure to sele-
nium, Goats and rabbits exposed for periods of 10 to
HO minutes to selenium oxide smoke showed dyspnea
and tachycardia on removal from the exposure
chamber. Animals receiving a fatal dose usually died
within 21 hours and sometimes within 3 hours. On
autopsy, pronounced pleural effusion and pulmonary
edema were found, plus hemorrhages in the lungs,
heart, and kidneys, and marked congestion of the
glomeruli and spleen.27
H.3.2 Toxicology
All workers agree that in absolute terms selenium
is less toxic than cadmium. The toxicity of selenium
oxide toward various species is shown in Table 3.
Other selenium compounds were screened for toxicity
at the UCTL without revealing any of greater in-
terest or effectiveness than the oxide.7
11.3.3 Assessment of Value as a
Chemical Warfare Agent
Selenium oxide smoke differs from cadmium oxide
smoke in the following respects; (I) it is less toxic
than cadmium oxide; (2) it is acrid and irritating to
SECRET 176
CAIIMIIM, SELEMIM, AND CARBONYLS OK IRON \M> MCKEL
the respiratory tract, whereas cadmium oxide is
odorless and relatively nonirritating; (3) it is white,
whereas cadmium oxide is brown; (4) it kills or dis-
ables more quickly than cadmium oxide. In contrast
to cadmium, which was found to l>e most effective in
incendiary munitions, selenium has been studied
mainly in explosive-type munitions.57'*1-"-**
Since selenium oxide is white, it would not lie de-
tected by appearance alone if used in conjunction
with ordinary screening smokes. On the other hand,
its irritant properties might well load exposed troops
to mask promptly. Whereas its relatively rapid
action as compared with that of cadmium is a
desirable feature, it is doubtful whether this out-
weighs its lower absolute toxicity. The potential use-
fulness of selenium oxide as a chemical warfare agent
cannot be accurately assessed on the basis of avail-
able information. If t here is a future requirement for
this type of agent, further experimentation, and par-
ticularly field trials, are in order.
li t NICKEL CARBONYL \NU IRON
CARBONYL
Nickel carbonyl, Ni(CO)(, was discovered in 1890,
and iron pentacarbonyl, Fe(CO);„ in 1891. Both eom-
|mumls can 1m* made to dissociate into carbon mon-
oxide and the pure metal under controlled condi-
tions;-and this reaction forms (he basis for the com-
mercial preparation of pure nickel (Aloud process),
a method which is in use to the present day. Iron
carbonyl was more difficult to prepare than nickel
carbonyl, the yield l»eing only about 1 per cent of
theoretical, so that the Mond process .was not eco-
nomical for the preparation of iron on a large scale.
In the 1920's, however, iron carbonyl found some
use in Europe as an “antiknock” for gasoline, and
more recently it has lieen an important source of the
pure, finely powdered iron which is used in powder
metallurgy.
The toxicity of the metal carbonyls was recog-
nized as early as 1891 7,1 and was extensively investi-
gated and reported in 1907 1908.70 Chemical war-
fare interest in the compounds arose primarily from
two facts; (1) they are toxic enough to merit con-
sideration as agents for use under certain circum-
stances where they might not be readily detected,
and (2) they dissociate readily in contact with the
active carbon of the gas mask, releasing four or five
volumes of carbon monoxide per mole of carbonyl.
The carbon monoxide is not absorbed by the canister
of the service gas mask. Therefore, the carbonyls
provide an indirect way of bringing carbon monoxide
into offensive chemical warfare.
11.1.1 Physiological Action
When iron or nickel carbonyl comes into contact
with moist air. dissociation into carbon monoxide
and a finely divided metallic salt takes place. Tins
salt appeal's to be a hydrated basic carbonate of
somewhat uncertain composition. Thus, when a [Ma-
son is exposed to an atmosphere into which iron or
nickel carbonyl has been released, lie breathes a
mixture of varying proportions of the metallic car-
bonyl, carbon monoxide, and a dust of finely divided
metallic salt. What part each of these components
may play in the toxicological picture will be discussed,
but for the moment the discussion will be limited to
the overall effects of inhalation of an atmosphere
known to contain originally iron or nickel carbonyl.
Since the physiological action of the two compounds
is essentially the same, they will lie discussed to-
gether. -
Armit in 1907 70 described the sequence of events
in human cases of nickel carbonyl poisoning as fol-
lows:
... immediately after having Iwcn exposed to air contain-
ing plant-gas there was giddiness, and at times dyspnea and
vomiting. These symptoms passed off rapidly ns soon as the
patients were brought into the fresh air. After 12 to 30 hours
the dyspnea returned, cyanosis appeared, and the temperature
liegan to l?e raised. Coughing with more or less blood-stained
sputum occurred on the second day or later. The pulse rate
became increased hut not in proportion to the respiratory
rate. Delirium of varying types frequently occurred, and a
variety of other signs of disturbance of the central nervous
system was noted. Death took plaee in the fatal cases lictween
the 4lh and 111li days. The chief changes found post mortem
were hemorrhages in the lungs, edema of the lungs, hemor-
rhages in the white matter of the braifTfin one case this was
very extensive), while some doubt exists as to whether any
blood changes were present ,
'Phis sequence of events parallels closely that of a
fatal case of human nickel carbonyl poisoning de-
scribed in 1934,72 in which death occurred on the
seventh day following exposure. The reaction of
mice, rabbits, cals, dogs, guinea pigs, and goats is
similar to that of man.’h,,b-c-,,>I0-7u
If masked troops are exposed to air containing
iron or nickel carbonyl, (ho carbonyl is catalytically
decomposed in contact with the active carbon of the
mask, leaving the finely divided metallic salt and
carbon monoxide. The metallic dust is efficiently
retained by the particulate filter of the service mask, NICKEL CARBONYL AND IRON CARBONYL
177
but the carbon monoxide passes through. When
mice were exposed to atmospheres which had origi-
nally contained iron or nickel carbonyl but which
had then I>een passed through active carbon, deaths
which occurred were entirely due to carbon monoxide
ami bore no similarity to those resulting from ex-
posure to (he carbonyls peans and the investigation of its proper-
ties from the standpoint of assessment as a possible
chemical warfare agent were studied under Divi-
sion 9 of the National Defense Research Committee
[NDRC] during the |»eriod 1942-1915. Earlier work
by British investigators had shown that ricin (com-
monly coded as “W”) could l>e dispersed as a par-
ticulate, nonpersistent, toxic cloud by explosion of
bombs containing a susjiension of ricin in carbon
tetrachloride. Notable progress was made by NDRC
investigators in all phases of the work with ricin.
Processes for the extraction of ricin from castor
beans and cold-pressed castor bean pomace were the
subject of laboratory and pilot plant studies.Timing
the laboratory inv estigations the protein was crystal-
lized; the crystals were not completely homogeneous
but mpresent the purest ricin so far obtained. The
pilot plant development culminated in a process of
extraction of castor Itean pomace with water and
purification of the toxin by two precipitations with
sodium sulfate. A water solution of the purified toxin
was spray-dried to give a dry product with a mass
median diameter of 0 8 g. This was air-ground to
give “disjiersible ricin” with a mass median diameter
of 2.5-3.5 m. which was approximately half as toxic
(by injection) as crystalline ricin.
Early work on the physiological action of ricin
resulted in the development of a bioassay procedure
in mice which was used to determine the toxicity
of various ricin preparations. Later studies investi-
gated the inhalation toxicities of various ricin prepa-
rations, which are a function of the intrinsic toxicity
of the material and the particle size distribution in
the inhaled particulate cloud.
Toxoids have l>een prepared from ricin by various
means, most successfully by treatment with formalin.
The toxoid has been used to produce in horses and
rabbits antiriein serums. These have been purified
and concentrated as antiriein globulin fractions that
were made available for therapy in case of accidental
exposure. Immunization against ricin appears im-
practical at present because of the short duration of
passive immunity in animals, and the toxicity and
local necrotizing action of toxoid preparations avail-
able for use in inducing active immunity.
The detection and assay of ricin in the field is a
difficult problem. Sensitized guinea pigs afford the
most sensitive, rapid, and specific means of detection
through their anaphylactic response. Hemagglutina-
tion and precipitin tests have been used; chemical
tests are less specific. Determination of particle size
distribution forms an important part of the assess-
ment of ricin and all other particulate clouds-(see
Chapter 15). Field trials employing these analytical
means and animals exposed to (he clouds to deter-
mine toxicity have been conducted to evaluate ricin
as a war gas and determine the efficiency of various
munitions for its dispersal.
Hiein is most efficiently dispersed from small high
explosive-chemical bombs as a suspension in carbon
tetrachloride of the most finely divided material
available. On the basis of airplane stowage such
bombs are estimated to be seven times as effective
as bombs charged with phosgene.
Processing all of the castor beans used in this coun-
try (based on 1911-1944 consumption) by the opti-
mum procedure bast'd on pilot plant experience
would yield approximately 1,000 tons of dispersible
ricin annually at a cost of about $13 per pound. This
is a significant quantity of a material which might
be used as a unique nonpersistent agent in gas war-
fare, difficult to detect and disturbing to morale be-
cause of its delayed toxic action. Ricin has served as
a model substance, presenting problems in prepara-
tion, protection of personnel, detection, assay, and
dispersal similar to those presented by other materi-
als investigated in the field of bacteriological warfare.
Some minor duplications appear in the subsections
of this chapter, which were written by different
authors.
12.2 PREPARATION OF RICIN b
The isolation from castor beans of products con-
taining the toxic principle known as ricin has been
recorded many times in the open literature within
the past 60 years.64 During World War I ricin was
• By Arthur C. Cope.
b By Joseph Dec.
SECRET
179 ISO
K1C1N
examined as a candidate chemical warfare agent and
its preparation was studied.21 The investigation of
the preparation and properties of ricin pertinent to
its use as a chemical warfare agent was renewed in
Great Britain 53 early during World War II and in
this country under NDRC’ Division 9 during the
fall of 1942.’ 4
The objective of developing a process for the large-
scale production of ricin in a form suitable for dis-
persion from munitions was attained.” During the
course of this development about 3,800 lb of ma-
terial was products I on a pilot plant scale.1 1133 Also
of considerable importance was the preparation of
ricin in a crystalline form.2 for the first time.
A complete review of the great number of products
containing ricin whose preparation has Iwen recorded
both in the open and classified literature is beyond
the scope of this chapter. Emphasis is placed herein
on the products studied most extensively during
World War II and on the studies leading to their
preparation. These include crystalline ricin; two
products used in field trials with munitions, 470 BM
199 and 1,703; and the material used for the prepa-
ration of toxoid, HI. A process for the large-scale
production of ricin is outlined.
12.2.1 Crystalline Ricin
The isolation during the late suminor-of-4643-of
the material responsible for the toxicity of crude
ricin preparations in crystalline form was a signal
achievement.2 Neither the first crystals isolated nor
any of the crystalline materials subsequently pre-
pared 1216 could be shown to be single substances.21*
Since the crystalline material was the most toxic
fraction ever isolated from crude ricin, studies were
initiated to determine its physical and chemical
properties, composition, and physiological behavior.
Properties
The crystalline material is a protein of the globulin
type,215 although the crude toxin shows albumin-like
solubility behavior. Repeated crystallizations fail to
increase its toxicity,212 which has been assayed to
be 500 9 and 750 12 TU (depending on the method of
evaluating TU; for definition of the toxicity unit
known as TU see Section 12,5). The protein is soluble
in acid or alkaline solution, is least soluble in the
range of pH 5.0 to 8.0,2 12•’* and is more soluble in the
presence of other proteins,21* Its ultraviolet light
absorption spectrum is similar to that of a typical
protein,21* and it has a specific optical rotation of
— 26.3 UUraccntrifuge and electrophorcsis measure-
ments showed tlie material to l>e fairly homogene-
ous.2 3 However, solubility measurements indicated
the crystalline material to consist of a solid solution
of more than one component.21* On the basis of
sedimentation and diffusion studies the molecular
weight has been estimated at 36,000* and 77,000.*
The rate of denaturation of the crystalline material
in aqueous solution to a product insoluble at pH 5.1
has Iwen determined at 65.3, 71.5, 7S.1, and 86.5 (\
and from pH 2 to pH 11.14
The chemical composition of the crystalline ma-
terial has been investigated, but not exhaustively.
Evidence was obtained that the rf-amino acid content
of an acid hydrolyzate of the toxin cannot be more
than 3 per cent.16 On a moisture- and ash-free basis
a sample of three times crystallized ricin was found
to contain 16.23 + 0.4 per cent nitrogen.’* From the
titration curve of crystalline ricin in water and in
the presence of 8 per cent neutral formaldehyde the
numbers of basic, amino, imidazole, and carboxyl
groups were deduced.1* The amide nitrogen, alkali
labile ammonia, hydroxvamino acid, arginine, his-
tidine, aspartic acid, and glutamic acid 15 contents
have been determined by chemical analysis. The
amino acid analyses referred to account for 50 per
cent of the weight of the protein and 60 per cent of
the nitrogen. The protein was found to contain 1.34
per cent sulfur and less than 0.1 per cent phos-
phorus.
Preliminary to the first successful crystallization,
ricin-sodium sulfate cake, an amorphous product
(describes! in Section 12.2.5), was fractionated with
ammonium sulfate at pH 6.8 to concentrate the
toxin.2 The moist solid was dissolved in a minimum
of water and allowed to stand at 5 C. A granular pre-
cipitate formed, which gradually became crystalline
on standing for several weeks. The crystals were
isolated, suspended in water, and dissolved by the
addition of a little hydrochloric- acid. The solution
was adjusted to pH 6.8 and allowed to stand at 5 C.
Recrystallization was complete in 2 or 3 days.
Crystallization procedures more rapid and pro-
ductive than the original method were developed.’*•’*
Two extractions of the ricin-sodium sulfate cake
with 10 parts of sodium sulfate solution (19 g Nar
SOi TOO ml H20) were found to leach away many
of the gummy low molecular weight impurities with-
out appreciable loss of the toxin.14 One useful pro-
cedure 15 involved extracting the residue with water
and allowing the solution to stand overnight in a 181
PREPARATION OF RICIN
refrigerator. The precipitate which formed was re-
moved and dissolved in water with the aid of a little
acid. The solution was neutralized, seeded with a few
crystals, ami stores! in the cold for several days to
yield a crystalline precipitate, which was separated
and recrystallized. A modification of this procedure
was performed starting with 1 kg of ricin-sodium
sulfate cake.1*The yield was about 70 g of crystalline
material, which is 7 per cent by weight or about
35 per cent of the toxin content of the starting ma-
terial. Reerystallization was complete in 12 36 hours
with an 85 90 per cent recovery.
A 2- to 21 (-hour dialysis of a 20 per cent aqueous
solution of ricin-sodium sulfate cake also served to
remove the low molecular weight impurities.1'" The
dialyzed solution after filtration and standing in a
refrigerator yielded a crystalline precipitate. The
percentage yields were comparable with those ob-
tained in the procedure involving preliminary puri-
fication with sodium sulfate solution.
In an attempt to obtain a pure sample of ricin for
an absolute standard, 60 g of 4 times crystallized
material was extracted 25 times with 0.1 per cent
sodium sulfate solution at /dl 7.0 and 10 C.1S The
residue of al>out 6 g was recrystallized. Solubility
studies on this product have not yet been made.
Although the product is probably the purest sample
of ricin obtained thus far, its allergen content has
linen estimated at about 0.1 per cent on the basis of
animal assay.12 -
Numerous experiments were performed in the
study of the crystallization of ricin which led to the
procedures just described.21215 Flotation-purified
ricin and the ball-milled and hammer-milled products
(described in Section 12.2.5) were less satisfactory
than ricin-sodium sulfate cake as the starting ma-
terial;12 however, crystalline material has been ob-
tained from flotation-purified ricin.15 Although a
short dialysis of a solution of ricin-sodium sulfate
cake is satisfactory for the preliminary purification,
exhaustive dialysis is not.12 Some impurities can also
be removed by adsorption on ('elite or floridin.12
12.2.2 Amorphous Klein
Studies on the preparation of amorphous ricin have
been extensive and a great number of products of
varying properties and content of toxin, non toxic
protein, proteose, and salt have been obtained.4-9,15 34
Crude ricin is soluble in water ami dilute salt solu-
tions. In (he dry state the products are normally
stable at room temperature and denatured at ele-
vated temperatures.4-34**-33 The stability decreases
with increasing moisture content.4 3#b Aqueous solu-
tions are less stable than the dry product at both
room and higher temperatures.415 33
Starting Material fob Preparation ok Hicin
Samples of ricin prepared from castor beans of dif-
ferent sizes and colors seem to be identical in physi-
cal, chemical, and immunological properties.13 4 The
maximum variation in toxin content of the different
beans which were examined in one laboratory was
34 per cent.4
The beans contain about 50 per cent oil and the
toxin is best isolated after removal of a substantial
portion of this oil. The castor bean pomace which is
obtained in the laboratory using a Carver press 4 or
in industry using a hydraulic press 11 contains about
15 per cent oil and is satisfactory for the aqueous ex-
traction of the toxin. A pomace containing 1-2 per
cent oil can lie prepared by extraction of either
ground castor beans or cold-pressed pomace with
suitable organic solvents.4 34 If desired, the bean hulls
can lie removed from (he pomace by flotation in or-
ganic solvents.4
Hydraulir-pressed castor bean pomace is prepared
commercially by castor oil producers. In one of the
commercial processes the castor beans are ground,
heated to about 60 C, and presses:!.11 This cold-
pressed pomace is recommended as the starting ma-
terial for the large-scale production of ricin.411 Com-
mercially, this product is extracted four times with
heptane at 82-87 C to obtain the remaining castor
oil and then blown with steam to recover the residual
heptane.” The latter step also serves to detoxify the
pomace, which is sold as fertilizer. Tests on a labora-
tory and pilot plant scale showed that no appreciable
detoxification occurs during the extraction with hot
heptane.4 11 Efforts to find an economical procedure
for recovery of the residual heptane without detoxi-
fication of the pomace were unsuccessful.11 Extrac-
tion of the cold-pressed pomace with water at pll 3.8
to remove the toxin and subsequent solvent extrac-
tion yielded castor oil containing free fatty acid.11
Extraction of Toxin from Bean Meal
Among the solvents which have been used to ex-
tract t he toxin from castor l>eans or the pomace arc
water, dilute salt solutions, glycerol, ethylene glycol
containing a little water, and diethylene glycol con-
taining a little water. Water and dilute salt solutions
are the most efficient and economical extractants for 182
RiCIN
the toxin.4 Ten per cent saline is slightly more effec-
tive than water; however, it also dissolves more non-
toxic material, most of which is coagulable protein.4 15
About 3' i 4 parts of water at pH 3.S to 1 part of
pomace seems to be most satisfactory. Less nontoxic
protein is dissolved at pH 3.8 than at pH 7.0 and
filtration is accomplished more easily.4 Extraction at
temperatures approaching 70 C proceeds more
rapidly than at room temperature but is accom-
panied by denaturation of the toxin.4
Isolation of Toxin fhom Aqueous Extract
of Pomace
The toxin may !>e precipitated from the aqueous
extract of pomace by nonaqueous solvents, by picric
acid and similar precipitants, and by inorganic salts.
Organic solvents such as alcohols and ketones pre-
cipitate the toxin from aqueous solution but rapidly
denature it at room temperature. At temperatures
below 0 C acetone has been used to precipitate and
wash the toxin.4 31 The use of ammonium sulfate,4 34
sodium chloride,4 "13 and sodium sulfate 3 4 "-'5 for
precipitating and fractionating the toxin has re-
ceived considerable study. Ammonium sulfate has
l>een used for precipitating the toxin on a pilot plant
scale.35 Sodium sulfate is now regarded as the best
precipitant. It is sujrerior to sodium chloride because
it gives better fractionation, is less sensitive to
changes in pH, and precipitates the toxin more com-
pletely.411 15 The importance of temperature control
during precipitation, filtration, and drying when
sodium sulfate is used have been st udied.4 Many data
on the salting out of the toxin with different amounts
of sodium sulfate and at different pH have been ob-
tained.3 4 " 15 These data were useful in the develop-
ment of the process for the large-scale production of
amorphous ricin (Section 12.2.3)."
Better yields of the toxin have been obtained in
the laboratory than in the pilot plant.4 " 15 In the
methods preferred by some investigators 415 slightly
less sodium sulfate is used than in the proposed large-
scale process and the first precipitation is performed
at pH 3.8. In one laboratory run,15 during which the
isolation of the toxin was followed by chemical anal-
yses and toxicity determinations, the product
amounted to 2.3 per cent of the pomace weight. It
contained 10.4 jier cent nitrogen and 32.5 per cent
inorganic material and had a TU value of 190. Of
the toxicity present in the extract, 92 per cent was
recovered. The product obtained at a similar stage
in the pilot plant process amounted to 1.4 per cent
of (lip pomace weight. Procedures involving a .single
precipitation of the toxin with sodium sulfate yielded
in (he laboratory products with TU values above
200,415 but these methods were not satisfactory on a
pilot plant scale because of operational difficulties.41'
Removal of water by lyophilizat ion of solutions of
partially purified ricin yields products of good ap-
pearance and stability.4 " Dialysis can serve to re-
move much organic and inorganic impurity and in
neutral solution leads to a precipitate of amorphous
ricin.4
Comminution of Amorphous Uiriv
Since the toxicity by inhalation of ricin aerosols
increases with decreasing particle size,* considerable
effort was directed toward developing a method to
produce finely divided, readily dispersible material
without concomitant denaturation of the toxin, '['he
process involving spray drying and air grinding of
partially purified ricin was the best solution found to
the problem." Prior to this solution an appreciable
number of other methods were considered and ex-
plored.*11 - —
The particle mass median diameter of freshly pre-
cipitated crude ricin is 1 2 g, but as the moist filter
cake is dried the particles agglomerate. The final
precipitation was performed under various condi-
tions with the objective of obtaining a product that
could 1k» ground readily to fine particles." Among the
conditions investigated were temperature of precipi-
tation, agitation during precipitation, addition of
sodium sulfate as a dry powder or from saturated
solution, variation in amounts of sodium sulfate used,
addition of colloids, addition of seeding agents, addi-
tion of nonionic wetting agents, and transfer of the
freshly precipitated product to a volatile liquid. A
2-hour ball-milling test was used for comparing all of
the samples obtained in this series of tests. None of
the experimental products showed significantly su-
perior grinding properties. Lyophilizatipn of solu-
tions of partially purified ricin proceeds without de-
toxification to give a friable mat-like solid.4 " Ball-
milling the solid reduced the particle size to a mass
median diameter of 6 g in 33 per cent less time than
that required with precipitated air-dried material."
The detoxification which accompanied the ball-
milling was 20 |M»r cent less than with precipitated
air-dried material." Lyophilizat ion of a pomace ex-
tract yielded a gummy product."
Flotation-purified riein-sodium sulfate cake (de-
scribed in Section 12.2.1) was used in ball-milling, PREPARATION OF RICIN
183
colloid-milling, and hammer-milling experiments.11
Hammer-milling gave products with particle mass
median diameters no smaller than 20 g. Colloid-
milling was even less effective. For about a year ball-
milling appeared to be the most promising method
for obtaining a finely divided material, and this
method was investigated intensively.1148 *' The opti-
mum conditions using an Abbe 4-jar mill fitted with
It4 gallon “specimen” type porcelain jars, which
were found to give a 4- to 6-ju product, involved
(4) steel balls for the milling. (2) low milling temper-
ature ( — 20 C), (3) low moist ure content ricin, and
(4) milling a susjxmsion of ricin in carbon tetrachlo-
ride." Factors affecting the ball-milling that were
studied included the vehicle, grinding media, temper-
ature, time, and moisture content of the amorphous
ricin." The ball-milling time necessary to give a
4- to 6-n product, was proportional to the load of
ricin in the jar, 1 lb of ricin requiring 8 hours. Ball-
milling a high moisture content material at room
temperature or in the dry state resulted in more de-
naturation of the protein than otherwise. Even under
the above optimum conditions at least 50 per cent
detoxification accompanied ball-milling the material
to a mass median diameter of 4-6 m " The toxicity
loss was reduced somewhat by drawing off the fine
particles as they were formed.51
A combination of spray drying and air grinding
was found to give a product with a mass median di-
ameter of 2.5-3.5 m with little denaturation of the
starting material." A spray dryer was constructed
and conditions for its operation investigated." Fac-
tors such as type of nozzle, solution concent ration,
atomizing air pressure, drying rate, drying temper-
ature, and amount of drying air were studied. Under
optimum conditions at an operating rate of 1 lb of
product per hour the product has a particle mass
median diameter of 6-8 m and is 95 per cent soluble
in water. The spray-drying process is superior to the
ball-milling method from the standpoints of low
toxicity loss, processing time required, safety, and
cost.
Several types of air-grinding equipment were in-
vestigated for the comminution of spray-dried ricin."
A grinder previously developed by the Eagle Pencil
Company was found to be the Ixst of the types ex-
amined. Optimum conditions for its operation in a
low humidity room were determined. Under optimum
conditions the product with a particle mass median
diameter of 2.5-3.5 n and a TU value of 225 is ob-
tained at a rate of 1 lb per hour. A reduction in
toxicity of about 5 per cent accompanies the air-
grinding operation.
I2.2.;i A Process for the Production
of Finely Divided Klein 11
On the basis of considerable laboratory and pilot
plant data a process for the production of finely
divided ricin at the rate of 20 lb per day has been
outlined. The equipment and manpower necessary
for this scale of o|x* rat ions have been determined.
The process involves extraction of the toxin with
water from castor bean pomace, two precipitations
of the toxin by addition of sodium sulfate, spray
drying of a solution of the partially purified toxin,
and air grinding of the spray-dried material. It was
estimated that the cost of such a pilot plant would be
approximately $125,000 and that the cost of pro-
duction at the 20 lb per day rate would be about
$10 per pound. The cost of operating a plant to pro-
duce 2,000 lb of “dispersible ricin” daily was esti-
mated to lx> approximately $13 per pound of product.
The product has a particle mass median diameter of
2.5-3.5 ju and a toxicity value of 225 TU. The yield
is 0,05 per cent basts! on (he pomace and would have
amounted to about 1,050 tons annually during the
years 1941-1944 if the castor beans crushed in this
country during those years had been processed by
this method." Reworking of the by-products from
the spray-drying anti air-grinding operations and
reuse of the nitrogen-containing sodium sulfate sepa-
rated in the flotation step should increase the yield
to about 0.85 jx*r cent.
Starting Material
The starting material for (his process is commer-
cially available hydraulic-pressed castor bean pom-
ace which has not been solvent-extracted to remove
the residual oil and subsequently steamed. The pom-
ace produced by one company averages 8.0-8.5 per
cent moisture, 14.0-16.0 per cent oil, and 4.6-5.0
per cent nitrogen. The pomace is ground in a ham-
mer mill prior to extraction.
Extraction of Pomace
The recommended conditions for extraction of the
toxin from pomace are as follows:
\\ ater for ext faction 350 per cent of pomace
weight
pi I 3.8 ± 0.1
Acid to adjust pH 5 per cent H2S04
Agitation time 60 minutes (not critical)
SECRET 184
RICIN
Temperature of extraction 25 C
Filtration Continuous vacuum fil-
ter
Filter aid 7 per cent of pomace
weight
Water for washing 50 per cent of pomace
weight
Under these conditions at least 97 j>er cent of the
extractable toxin is recovered. The amount of water
used is the minimum necessary to produce a slurry
that can be handled satisfactorily in plant scale
equipment. Sulfuric acid is preferred over hydro-
chloric acid because of lower cost and lower corrosion
/rate. Continuous vacuum filtration at a higher pH
y is not possible because of the changed physical char-
acter of the slurry. The filter aid is necessary to in-
sure a satisfactory filtration rate. Filtration with the
vacuum filter proceeds about 30 times faster than
with a recessed plate type filter.
First Precipitation and Filtration
The optimum conditions for precipitation of the
toxin from the extract and subsequent filtration were
determined to lie as follows:
Salt usage 20 per cent Na2SO,,based
on filtrate weight
pH 7.0
Alkali to adjust pH 12 per cent NajCO*
Temperature 25 C
Time of precipitation 20 minutes
Filtration Continuous vacuum filter
Filter aid 4 per cent of slurry weight
Wash solution 20 per cent of 16.7 per
cent NaoSOi, based on
weight of extract
Under these conditions 50 per cent of the total
nitrogen in the extract remains in solution and Is
eliminated in the filtrate, whereas less than 2 per cent
of the toxin is lost. Precipitation at pH 7-8 was found
to remove 6-10 per cent more nontoxic nitrogen than
at pH 3.8. Increasing the temperature from 25 to 35
C and varying the precipitation time from 15 to 60
minutes showed no appreciable effects. The rate of
filtration with a vacuum filter was 3±2 times that with
a plate and frame filter press, filter aid being neces-
sary to obtain a satisfactory filtration rate in both
cases.
A full-scale pilot plant run was made to determine
whether a single precipitation process would give a
product suitable for spray drying.11 Filter aid was
not used, because previously it had been found not
possible* to reduce the sodium sulfate content of a
product containing filter aid by a process involving
flotation in carbon tetrachloride. Despite the absence
of filter aid, which made the filtration very slow, the
dried product separated very poorly in carbon tetra-
chloride, The product, which amounted to 1.0 per
cent of the original pomace, contained 11.0 per cent
nitrogen and had a toxicity value of 200-250 ITT.
The operational difficulties encountered indicated
this one-step process to be unsatisfactory on a pilot
plant scale.
Second Extraction and Filtration
The optimum conditions for extraction of the toxin
from the ricin-sodium sulfatc-guhr moist filter cake
were found to be as follows:
Water for extraction 300 per cent of wet cake
weight
pH 3.8 ±0.1
Acid to adjust pH 5 per cent T USth
Filtration Continuous vacuum filter
Water for washing 25 per cent of slurry
weight
An additional 10 per cent (based on the pomace
extract) of nontoxic nitrogen is removed during this
operation. The pH was varied from 3.8 to 9.0, and it
was found that 5 per cent (based on pH 3.8 extract)
more nontoxic nitrogen is removed at pH 3.8 than
at pH 9.0. The filtration is very rapid because of the
large amount of filter aid present.
Second Precipitation and Filtration
The recommended conditions for the second pre-
cipitation of the toxin and subsequent filtration are
as follows: ~
Salt usage 20 per cent allow-
ance being made for the
sodium sulfate in the
filt rate
pH 7.0
Alkali to adjust pH 12 per cent Na*COg, or
more dilute
Temperature 25 C
Time of precipitation 45 minutes
1* iltration Plate and frame filter press
Filter aid None
Washing None
Drying of the filter cake can Ik* accomplished in
6-10 hours using a three-section hot-air dryer oper-
ated at successively increasing temperatures from
55 (' to 75 C. The dried product is given a slight
SECRET PREPARATION OF RICIN
185
grind, passed through a five- to ten-mesh screen, and
slurried in five parts of carbon tetrachloride. The
toxin is removed from the surface of the mixture and
dried. The sodium sulfate which settles to the bottom
is used in the precipitation steps.
A quantity of partially purified ricin was produced
by the process outlined except that the product was
dried at 50 C. The product was obtained in 0.85 jx*r
cent yield based on the pomace, contained 13.0 per
cent nitrogen, and had a TU value of 250 3(H).
Pilot plant tests indicated that a minimum of 20 lb
of sodium sulfate is necessary to prevent loss of toxin.
Approximately 3 (>er cent more non toxic nitrogen is
removed at pH 7.0 than at pH 3.8. Operation at 35 C
instead of 25 C removes 2 per cent more nontoxie
nitrogen, but about 2 per cent more toxin is lost.
Since filter aid cannot be employed in this step, the
use of a vacuum filter, which requires filter aid, is not
possible. However, the physical character of this
second precipitate |iermits a satisfactory filtration
rate with a plate and frame filter press. Washing the
filter cake with sodium sulfate solution (19.5 lb
NajSOj 100 lb H«0) does not result in .sufficient puri-
fication to warrant a washing operation.
The utility of a third precipitation of the toxin
with sodium sulfate was invest igated. No appreciable
purification was obtained without concomitant loss
of toxin.
Spray Drying and Air Grinding
A 20 per cent aqueous solution of the above flota-
tion-purified product is spray-dried under certain
prescrilied conditions at the rate of I lb per hour to
give solid particles, which are 95 per cent soluble in
water and have a mass median diameter of 6-8 m-
The solution for the second precipitation step can be
spray-dried but it would contain about 50 per cent
sodium sulfate. It was not found possible to separate
the sodium sulfate from a spray-dried product by
flotation in carbon tetrachloride.
Air grinding of the spray-dried material is carried
out under certain defined conditions in an air grinder,
previously developed by the Eagle Pencil Company,
at a rate giving about 1 lb of product per hour. This
operation reduces the toxicity of the material about
5 per cent. The product has a particle mass median
diameter of 2.5-3.5 m and a TU value of 225.
12.2.1 Four Vmorplious Ricin Products
The four amorphous ricin products described in
this section a re of particular interest because of the
considerable extent of studies performed with them.
The preparations known as (!) ricin-sodium sulfate
cake, (2) 470 H.M 199. and (3) 1,703 represent suc-
cessive stages in the development of an amorphous
ricin product in a form suitable for dispersion from
munitions, and (4) HI was used for the preparation
of a toxoid.
Ricin-Sodiim SriiKATE I’akk 1
A total of 1,550 lb of the product known as ricin-
sodium sulfate cake was prepared on a pilot plant,
scale at the request of NDRC Division 9,' and an
additional 2,000 lb was prepared for the Canadian
government.11 The method used in these operations,
which was based on a procedure previously developed
in another laboratory,4 utilized the facts that crude
ricin is soluble in water and insoluble in saturated
aqueous solutions of sodium chloride and sodium
sulfate. Subsequent studies resulted in a marked
improvement in the method of preparation (Sec-
tion 12 2.3)."
Castor beans were the starting material and an
Anderson expellee was used for expressing the oil
from the beans. From each ton of beans was obtained
810 lb of #3 grade castor oil. The expellee cake, which
contained 13.1 [>er cent oil, 11.2 per cent moisture,
and 4.6 per cent nitrogen, was ground in a hammer
mill. Three parts of water at 15 20 C were mixed with
the ground cake, the mixture agitated for 1 hour, the
pH adjusted to3.8 + 0.1 with 5 percent hydrochloric
acid, and the slurry filtered in a plate filter press.
The filtered extract at pH 3.8 + 0.1 and 17 C was
saturated with sodium chloride to precipitate the
toxin. The precipitate was separated by filtration,
sufficient guhr being used to insure a satisfactory
filtration rate. V sample of dried filtered cake was
found to contain 33 per cent guhr and 33 per cent
sodium chloride. The wet precipitate was mixed with
five parts of water and the mixture adjusted to pH
8.0 with 5 per cent sodium hydroxide solution. The
mixture was agitated for 1 hour and then filtered to
remove guhr and other impurities. The filtrate was
saturated with sodium sulfate, allowance being made
for the sodium chloride present. The mixture was
adjusted to pH 7.0 and then filtered at 35 10 C. The
filter cake, about I inch thick, was dried in trays for
6(U72 hours at a maximum temperature of 60 C and
then packaged.
About 55 per cent of the toxicity available in the
starting material was present in the ricin-sodium
sulfate, cake. The TU value '■* of the cake was 100-
SECRET 186
RICIN
125. Analysis of the product showed 4.4 per cent
moisture, 46.6 per cent ash, and 8.6 per cent nitrogen,
of which 97 per cent was soluble and 45 per cent co-
agulable.9 Electrophoretic and ultracentrifugal stud-
ies indicated the cake to consist of several compo-
nents with toxicity and hemagglulinating power
associated with only the Bl fraction.3 Other studies
indicated it to Ire composed of (1) the toxin, (2) a
nontoxic protein otherwise very similar in properties
to the toxin,3 16 (3) a dye derived from the bean
shells, (1) an allergen, (5) an unidentified substance
which tends to keep the toxin in solution at pH 7.0,
(6) proteoses,14 mid (7) inorganic salts.12
Preparation 470 B.M 199 —
About 100 lb of the product designated as 470 BM
199 was prepared 11 for field trials at Dugway Prov-
ing Ground22 and Suffield Experimental Station,
Canada.43 44 This ball-milled material was (he best
available in sizable quantities from the standpoint
of high toxicity and small particle size for the field
tests held during the spring and summer of 1944.
Ricin-sodium sulfate cake was the starting ma-
terial for (he preparation of 470 BM 199. The cake
was ball-milled for 15 minutes in an Abbe porcelain
jar mill to yield a product that would pass through
a 40-mesh screen and (hen slurried with 5 parts of its
weight of carbon tetrachloride. The sodium sulfate
tended to settle to the bottom of the mixture and the
ricin concentrated at the Turface where it was re-
moved by scooping with a wire screen. This flotation
step reduced the salt content of the cake from about
45 per cent to 15-18 jrer cent. The flotation-purified
ricin was suspended in carbon tetrachloride and the
slurry ball-milled for 8 hours at room temperature
in I1 4 gallon capacity Abb£ porcelain jar mill using
5,s-inch steel balls. The product was tray dried at
60 C for 2 hours and then at 82 C for 1 Yi hours,
which gave a white friable cake readily disintegrated
by ball-milling for 5 minutes.
Considerable denaturation of prot ein accompanied
the ball-milling operation. The TU values found for
different samples of this material ranged from 00 to
100.1-9 Examination of a representative sample
showed a particle mass median diameter of 6.3 n, 4.4
per cent moisture, 15.4 per cent ash, and 13.35 |>er
cent nitrogen, of which 04 per cent was soluble and 14
per cent was coagulable.9
Product L703 11
A total of about 60 lb of spray-dried air-ground
ricin was prepared.11 Eot 1,703 was examined in the
laboratory fur toxicity by inhalation after dispersion
as a dust 9 and similar lots L701 and 1,82(3 were tested
in the field at the Suffield Experimental Station,
Canada.46 The small mass median diameters, 3.1 n
for 1,703 and 3.3 g for I.S'it),2'1'' are particularly note-
worthy.
The starting material for (he preparation of spray-
dried air-ground ricin was (1) riein-sodium sulfate
cake partially purified by flotation in carbon tetra-
chloride, and included some (2) ball-milled and
(3) hammer-milled products. Preliminary to spray
drying, these materials were partially purified by
another precipitation with sodium sulfate. The stal l-
ing material was stirred with I parts of water, (he
pH of the mixture adjusted to 7.0 ± 0.1, guhr added,
and the mixture filtered at 30 C. Sodium sulfate
(10.2 per cent of filtrate weight) was added to the
filtrate. The resulting slurry was adjusted to a pir
of 7.0 ± 0.1 and filtered at 30 35 C. The filter cake
was dried at 00 C for 10 hours, ball-milled for 5 min-
utes to pass a 40-mesh screen, and the sodium sulfate
content was reduced by flotation in carbon tetra-
chloride, Spray-drying 20 per cent aqueous solutions
of this flotation-purified ricin gave materials with
particle mass median diameters of 0-8 p.
The spray-dried materials were processed in an air
grinder to yield products with TU values averaging
200 and mass median diameters of 2.5-3.5 p." Analy-
sis of lot L703 showed 2.0 per cent moisture, 19.7 per
cent ash, 13.2 per cent nitrogen, of which 91 percent
was soluble and 45 per cent coagulable, a TU value
of 100, and a mass median diameter of 3.1
Preparation BU
Preparation B1 is of interest because of its use for
the preparation of toxoid. It was prepared as follows:
7* 2 K of ricin-sodium sulfate cake, which contained
71 mg of insoluble nitrogen and 050 mg of soluble
nitrogen, was suspended in water and centrifuged.
The precipitate was washed twice with 30 ml of
water, to which was.added for the second washing
about 0.1 g of sodium sulfate. To the solution and
washings (300 ml) was added 175 ml of warm satu-
rated sodium sulfate solution to precipitate the toxin,
and the mixture was allowed to stand overnight. The
precipitate was centrifuged and reprecipitated twice
from a volume of 150 ml with 87.5 ml of warm (37 C)
saturated sodium sulfate solution. Additional toxin
can be recovered from the filtrates.
B1 is about two-thirds as toxic as the crystalline
material. The molecular weight of B1 was determined
SECRET PHYSIOLOGICAL ACTION
187
to be 85,000 and the isoelectric point to Ik* 5.2. Crys-
talline ricin and Bl seemed to differ only in toxicity,
since by immunochemical, ultracentrifugal, and
electrophoretic criteria they appeared to be identical.
12.5 PHYSIOLOGICAL ACTION'
Systematic work on the use of ricin as a chemical
warfare agent was begun in the United States during
the fall of 1942. Its immediate objective was the pro-
duction on a pilot plant scale of a sufficient quantity
of an active product to make possible field trials of
methods of dispersal of this novel type of agent.
Such toxicological work as was done at this time was
directed toward assisting in the control of (he plant
process and toward (he accumulation of basic data
on the inhalation toxicities of the product in various
species of animal.
When it Ix'came evident that the bulk production
of a satisfactory material was feasible,1 1 u the ques-
tion arose of the form in which it should be prepared
for dispersal in the field. On the basis of experience in
England,it was decided that it should be re-
duced to a finely divided dry powder which could be
introduced into munitions either in the dry state or
in suspension in an inert volatile liquid. This decision
made urgent the need for an extensive investigation
of the relation between the particle size distribution
in a toxic dust cloud and the inhalation toxicity of
the cloud. Thereafter the chief emphasis of all as-
pects of the program was on this complex problem.
It was recognized that the significance of the pro-
gram did not rest solely upon the potentialities of
ricin as an agent for chemical warfare. Ricin was con-
sidered, rather, as a readily available prototype of
other unstable nonvolatile toxic agents of biological
origin which might be exploited as offensive agents
by one or other of the warring nations.
The following subsection contains a summary of
the available information on the parenteral and in-
halation toxicities of standard preparations of ricin.
This is followed by a review of the symptoms and
pathology of ricin poisoning and a brief discussion of
the mechanism of its action.
12,3.1 The Parenteral Toxicity of Ricin
Details of methods of bioassay, of methods of field
detection and assessment, and of the relation of parti-
cle size to inhalation toxicity will be found in Sec-
tions 12.5 and 12.0 and in Chapter 15, respectively.
The summary which follows is concerned only with
the toxicities for various species of standard prepa-
rations of ricin under laboratory conditions.
Ricin has been stated to be toxic for all verte-
brates.5 Frogs are sensitive only if kept in a warm
environment." Few invertebrates ap|x>ar to have
been tested. The motility of a ciliate has been found
to be arrested by low concentrations of ricin,5 but the
relation between this effect of a preparation anti its
toxicity for higher animals has not been established.
The results of the few laboratories that have made
comparative assays of a single preparation on a range
of animal species are summarized in Tables 1 and 2.
In Table I they are given as toxicities relative to the
toxicity for the rabbit. The high sensitivity of the
rabbit is well attested, but there is not full agreement
on the order of sensitivities of other species. The ma-
jority of the toxicities recorded in the literature have
been based upon very few animals and are scarcely
more than orders of magnitude. The most extensive
series of observations are those made at the Uni-
versity of Chicago Toxicity Laboratory [UCTL],*
but even (best* can be accepted as precise only for the
mouse and for the rabbit.
Table 1. Relative LD-
o’a (approximate) of
ricin for dif-
ferent sjx'cies
*
Author
Osborne
Field
Hunt
OSRD 5525’
Date
1905
HI 10
1918
1945
Reference
til
56
2d
9
Route
Sultou-
lutrn-
Subcu-
Subcu-
tancous
muscular tancous
tancous
Rabbit
I
1
1
1
Rat
1
1.5
Guinea pig
7
8
5
3
Mouse
8
S
Sheep
2
D‘>g
7
16
2
Cat
2
16
10
Goal
30
♦ An entry of 10
in this tabh
hidicatea
that for the species in question.
ricin wan found to be one-tenth
an toxic as for the rabbit.
etc.
In Table 2 some of the data on which Table 1 was
based are given in absolute units. The preparation
to which they refer exhibited about 28 per cent of
the toxicity of crystalline ricin based on comparative
assays on mice. Although the crystalline material is
not believed to be molecularly homogeneous, it is
definitely the most toxic material which has been
prepared in contemporary work. It is suggested,
therefore, that the best estimate of the attainable
toxicity of ricin is obtained by dividing the LD:,,, for
r By R. Keith Caiman.
SECRET 188
R ICIN'
Table 2. Kstimated LD
... (Mg /kg)
Author
Osborne
Field
OSHD 5525s (10-day
observation)
Crystalline
Standard riein
ricin* (computed)
Rabbit
0.5 (7-day)
100 (2-day)
0.1
10
3
Hat
15
4
Guinea pig
3.2 (7-day)
100 (2-day)
0.8
30
9
Mouse
80
24
Sheep
20
7
Dog
500 (2-day)
0.6
20
7
Cat
100 (2-dav)
0.2
100
30
Goat
• ... 3
• (3-day)
♦ The OSRD obwtvatiflns wort*
made on the
1 pilot plant product (stand-
ard i irtu). 1
ririn.
tits had 28 per rent
of the toxicity for mic
c of crystalline
tanoons and intravenous toxic! lies for I ho rabbit*
are in the ratio of 1/5.
• 2.3-2 Toxicity by Inhalation
1 he importance of the partic le size* distribution of
the airborne toxin has been emphasized in the intro-
duction. In one extensive investigation of this prob-
lem,* two methods of varying the particle size wore
user!. In one, animals were exposed to atomized aque-
ous solutions of ricin containing varying amounts of
glycerol. The mean particle size in the aerosol varied
with the amount of nonvolatile solvent in the solu-
tion. The other type of experiment was the exposure
of animals to dust clouds generated from powdered
standard ricin which had been reduced to varying
degrees of fineness by milling or spray drying. The
results are summarized in Table 3;
standard ricin by 3.5. The figures given in the last
column of Table 2 have l>een derived in this manner.
The very high toxicities recorded by Field M for
his preparation find no explanation. It is highly im-
probable that they represent a product many times
more toxic than crystalline ricin. On the other hand,
his figures and those of Osborne do suggest that
some of the early investigators of ricin succeeded in
purifying the toxin to a degree approaching the
purity of the crystalline material.
The Relation of the Survival Time to the Dose
The early investigators recognized that the time
of survival of animals injected with ricin varied from
a few hours to sc*vend weeks depending on the dose
administered. This relation has been investigated
for mice and, less extensively, for rats in several
laboratories ».».■».«.« ;ind has formed the basis of the
accepted method of bioassay (Section 12.5). The
dose-survival time curves for mice obtained in one
laboratory *‘l& have been found to approximate rec-
tangular hyperbolas which may be represented by
the-equations
D{t — 11) = 430 (intravenous)
D{t — 13) = 1,150 (intraperitoneal)
D(t - 16) = 2,500 (subcutaneous)
where I is the survival time in hours and D is the dose
in micrograms of crystalline ricin per kilo body weight.
Route of Injection
The above results indicate that the relative toxic-
ities for the mouse by subcutaneous, intraperitoneal,
and intravenous injection, respectively, are (for the
smaller doses) approximately 1 2.2 6. The subcu-
A. Inhalation toxicitii
Table 3 —
s of atomized solutions of standard
ricin.
MMD (M)
L(Cl )i„ (mg, min /in5)
1.4 4.0
fi.fl
Rabbit
4
s
10
Guinea pig
7
15
Mouse—
9
40
45
Dog
24
45
Cat
24
50
Rat
50
120
Monkey
100
Preparation
MMD(m)
Rail-milled
10 0.3
Spray-dried
5.9 3.1
Atomized
solution
1.4
~
Relative lone Hies
—
Mice
3.5
2.8
0.5
0.5
100
Rabbits
5.7
5.3
30
100
% mass below
3 fi
7.5
10.0
3.0
15
loo?
% mass below
2 fl
3.0
3.2
0.0
H
100?
B. Inhalation toxieilies of dry dusts of standard ricin.
lt would appear that the toxicity increased as the
mass median diameter [.MMD] of the cloud dimin-
ished. Indeed, then- is some justification for the eon-
elusion. in the cases of mice and rabbits, that the
toxicity was roughly proportional to the fraction of
the airborne mass which was present in particle sizes
smaller than 2 3 n in diameter. The reader is re-
minded that the MM I) is an inadequate description
of the characteristics of a dust cloud in which the
particles differ in shape and density as well as in size
and is referred to Chapter 15 for a discussion of the
SECRET PfIVSIOLOGIC V1. ACTIO\
189
relation of these factors to the probability that an
inhaled particle will penetrate the nasal barrier.
Although the most toxic aerosol was (hat with the
MM I.) of 1.4 n it is improbable that this represents
the maximum attainable inhalation toxicity. Some
allowance for nasal retention and for incomplete re-
tention in the lungs should probably he made. Even
so, the inhaled doses of the 14-g aerosol for mice and
rabbits, which may he computet! from the minute
volumes of respiration and the L{Cl)50’s, are approxi-
mately equal to the /./>.,o’s by intravenous injection.9
That is to say, Hein is at least as toxic hy inhalation
as by vein. That it is probably more toxic in the
lungs is indicated by the fact that the approximate
when solutions were injected directly into the
trachea of rabbits, was 0.5 Mg kilo. In cats, dogs, and
rats it was about 5 Mg kilo.9 In contrast with these
results were the very low toxicities resulting from the
nasal instillation of ricin.9
When solutions of ricin are instilled in the eyes of
animals in sufficient amount, enough may he al>-
soribed to l>e lethal.-4 Only small amounts are neces-
sary to produce serious local injury. The instillation
of 1.5 Mg of crystalline ricin produced corneal damage
in a rabbit’s eye which disappeared in 10 11 days.9
A particle of 100 m in diameter (0.5 Mg) implanted in
the eye resulted in a conjunctival reaction persisting
for a week. Corresponding lesions in the eyes of rats
and guinea pigs required five to ten times this dost*.
It must be remembered that only large particles will
impinge in the eye from a cloud and that such parti-
cles will tend to precipitate rapidly under wind con-
ditions favorable for the persistence of a fine particu-
late cloud. Clouds of fine dusts such as are highly
toxic by inhalation would therefore be unlikely to
contain a concentration of coarse particles which
would present a serious hazard to the eyes.
12.3.3 The Toxicity of Ricin for Man
The ingestion of two castor beans lias been fatal in
man.24 54 6 8 It has l>een estimated that this corre-
sponds with a lethal dose of about 0.3 mg of purified
ricin per kilo. It has been suggested that ricin is
about 100 times as effective by vein as by mouth.*"
On this basis the intravenous lethal dose for man
would lx* as small as that for the rabbit. Such com-
putations arc highly precarious, hut other evidence
has been advanced to indicate that man is quite
susceptible to ricin poisoning.33 41
Elsewhere in this section are descrilied symptoms
of mild poisoning in a number of individuals who had
probably lioen exposed to low concentrations of air-
borne ricin. It is significant however that no serious
casualty has occurred in the pilot plant, in the ex-
plosion pit at Dngway Proving Ground, or in labora-
tories studying the dispersal of ricin. The atmos-
pheres in all these places must have l>ecn contam-
inated with ricin dust.
It has also been suggested that the handling of
solutions of ricin presents a skin hazard,4 hut the
opinion of most investigators who have long worked
with such solutions is that the hazard is small if ele-
mentary cleansing precautions are taken.
12.3.1 Symptoms of Intoxication
Laboratory animals show no evidence of intoxica-
tion for several horn's after the injection of a dose
which will kill them in 24 hours. Thereafter their fur
liecomes ruffled, they grow restless, and refuse food.
As the time of death approaches, diarrhea is frequent,
breathing becomes dyspneic, (heir bodies feel cold to
the touch, and their eyes may become sealed with
exudate. Finally, the animals become moribund and
die in coma or, more frequently, after a series of
violent convulsions. With smaller doses the sequence
of events is similar, hut their time course as well as
the initial latent period arc more protracted.
Some 150 cases of poisoning in man have been re-
viewed.24 •64 M Most of these have been the result of
the accidental eating of castor beans. In some cases
weakness and prostration were the only symptoms.
In more serious attacks, there was nausea and vomit-
ing, epigastric pain, cramps in the limbs, a weak
pulse, and a rapid respiration with a rise in body
temperature. Fatal cases passed into collapse fol-
lowed by convulsions. Symptoms might be delayed
for 2 to 14 days, or, surprisingly, might be evinced
within 1 hour after ingesting the t leans.
Among the personnel working with ricin in the
United States throughout 1943-45, there were no
serious cases of poisoning, although there were a
number of minor illnesses attributable to exposure.
These were probably the result of inhaling airborne
toxic dust. Two types of reactions among laboratory
workers have !>een distinguished.9 One — the im-
mediate reaction — resembles that of an individual
sensitized to a foreign protein. The symptoms have
varied from a protracted bout of sneezing to a severe
asthmatic attack with violent coughing and retching.
The symptoms disappeared within an hour. The
second type of reaction probably corresponds to the
toxic effect in animals. Symptoms were delayed for
SECRET 190
HIC1N
4 to 8 hours. There was then a sharp febrile response,
tight ness of the chest, tracheitis, aching joints,
nausea, dyspnea, and coughing. Some hours later
the onset of profuse sweating was commonly the
signal of the alleviation of most of the symptoms.
Somewhat similar observations have been made
by the British, who have obtained local and general
reactions by the intradermal injection of very small
doses of ricin preparations 33 (see Section 12.4.3).
I2.a .3 Pathology
Accompanying the outward signs of intoxication
in animals has been noted an early fall in body tem-
perature,52 which may be preceded by a rise,57 In
rabbits, it has been reported that the blood pressure
falls from 100 to 65 mm of mercury at an early stage
and remains at this level until death.7 There appear
to l>e no notable changes in the blood picture.27 It is
generally agreed 94 7 52 that about 20 hours after
the injection of an dose there is a leucocytosis,
with a simultaneous increase in both lymphocytes
and polymorphonuclear leucocytes,10 A transient fall
in red cell count has been recorded,9 10 but others
report no change in red cell count, in red cell volume,
or in sedimentation rate.52 Within 20 hours after an
LD n dose the clotting time was found to increase to
three times its normal value and remain at this level
till death several days later.7 An extreme terminal
hypoglycemia 7 and acidemia 62 have been observed
in rabbits and in rats and a rise in blood phosphatase
has been reported.52
Careful reports of the gross and microscopic pa-
thology of animals dying after the parenteral admin-
istration of ricin are found in the early literat ure.55-57
This information is reviewed and extended in Chem-
ical Warfare Service Monograph 37, writ ten in 1918.21
Between this time and 1940, students of ricin became
preoccupied with the chemical and immunological
characterization of the toxin and with the hemag-
glutmating activities associated with it. Little was
added to our knowledge of the physiological action
of the* toxin. During World War II, extensive patho-
logical examinations of animals poisoned with ricin
were made in England,*9 in the United States,91"-22
and in Canada.47 Some of these were confined to
post-mortem examination of animals killed by the
injection or inhalation of the toxin,9 22 39 whereas
others relate to animals sacrificed at chosen times
after the parenteral administration of lethal or
sublethal doses.10 59 Bearing these differences in pro-
cedure in mind, it may lx* said that there is substan-
tint agreement between the laboratories referred to
and the early reports in the open literature.45 57-6i It
is possible, therefore, to summarize the situation in
the following general conclusions.
RahENTER AL A DM IXISTKAT Io\
1. There is mild to moderate congestion and
edema of the lungs.
2. There is mild degeneration of the intestinal
epithelium at suprulethal doses only.
3. There is necrosis of the liver at and below
IJ)Mi doses.
t. There is hyixrphisia of the spleen at .sublethal
doses and involution at higher dosage.
5. There is fragmentation and involution of the
thymus at all doses.
6. There is congestion and delayed necrosis of the
adrenal in rats but not in rabbits.
The occurrence of pin-point hemorrhages through-
out the body lias been emphasized by some 17 43 but
minimized by others. Less consistent'findings have
been necrobiosis of reticuloendothelial cells and hone
marrow, cloudy swelling of the kidneys, and fatty
degeneration of heart muscle. No differences be-
tween the effects of crystalline ricin and of amorphous
preparations have been observed 10 nor have any
striking differences in the responses of different
species been observed.*
Exhalation
The pathology is almost entirely confined to the
thorax.9 '" The lungs are dark and greatly increased
in weight and are filled with edema fluid. The ab-
dominal organs are normal except, for some fatty de-
generation and, occasionally, hyaline infiltration and
necrosis of the liver.
Ingestion
The effects of ingesting the toxin have been in-
vestigated in fatal cases of poisoning in man.24,54-**
The chief post-mortem findings have been extreme
congestion of the stomach and intestines.
12.it.a The Mechanism of Action of Ricin
Such pathological work as was carried out in the
United States in 1943-45 was incidental to the pro-
gram outlined in the introduction. No systematic
investigation of the mechanism of action of ricin
was undertaken and our knowledge of this subject
remains fragmentary. We are, indeed, as ignorant
of the nat ure of the action of ricin as we are of the
actions of those bacterial toxins which exhibit a sirni-
SECRET IMMlNOLOGY
191
lar delayed effect and ill-defined pathology. Apart
from revealing local effects depending upon the route
of administration, pathological reports lie tray no
characteristic lesions which would indicate the in-
trinsic nature of the toxic action.
The death of animals in convulsions is probably
the result of hypoglycemia. It has been found (hat
the blood sugar of rabbits and rats remains normal
until a few hours before death, when it falls precipi-
tously to convulsive levels.7 62 The toxic action, how-
ever, is not primarily a reversible disturbance in
carbohydrate metabolism. The liver glycogen is
found to Ik- very low at death, but it has not been
possible to induce glycogen storage in poisoned ani-
mals by injecting glucose to maintain a normal blood
sugar level. Nor has life been prolonged by this
means.7
One of the earliest theories of the action of riein
was that it was an enzyme. This was thought to ex-
plain its great potency. It was also thought that its
delayed action might plausibly lx attributed to the
time required for the enzyme to build up a lethal con-
centration of the hypothetical product of its activity.
In this connection it should be borne in mind that
several enzyme activities — phosphatase, lipase,
esterase — are exhibited by extracts of castor beans.
Purification of the toxin is not, however, accom-
panied by enhanced enzyme activity. Indeed, it has
been stated that crystalline riein is free from phos-
phatase and lipase action.1213 Recently a Canadian
laboratory has reported that riein preparations hy-
drolyze adenosine triphosphate (ATP).43 60 They
further observed that riein inhibited the I mat of the
isolated frog’s heart and that the Ixat was restored
by the addition of ATP. This would suggest that
riein may act by interfering in those basic metabolic
reactions whereby the energy of metabolism is con-
veyed to the functioning structures of tissues. Data
are, however, not yet available to indicate whether
the concentration of crystalline riein which is re-
quired for effective adenosine triphosphatase action
is such as to make plausible the hypothesis that its
lethal action is dependent on this property. More-
over, in one investigation 7 the action on the frog’s
heart was not confirmed. No increase in nucleotide
in the blood of animals poisoner! with riein was ob-
served. Riein did not cause a hydrolysis of ATP in
the blood of dosed rats.62
It may lw submitted that it is just as plausible to
attribute a disturbance in metabolism to the blocking
or distortion by the toxin of the action of an enzyme
native to the rolls of the animal as to consider it to
be the result of the invasion of those rolls by a foreign
enzyme in the form of the toxin. —
An early theory of the action of rieiu was based on
the hemagglutinating properties of riein prepara-
tion.24*8 If (his action were manifested in vivo pro-
found disturbances in circulation might be respon-
sible for the toxic effect. Unfortunately the concen-
trations of riein required to agglutinate red cells in
vitro are greater than those established in body fluids
by lethal doses of riein. Moreover, the agglutination
of red cells is inhibited by serum 1 '* and crystalline
riein is very much less potent as an agglutinin than
are cruder preparations.216 Finally, the absence of
thrombotic lesions would seem to deny the theory.
Although the hypothesis has little to support it, it
should lx* recorded that t issue cells as well as erythro-
cytes have iKaai shown to be agglutinated by crude
riein and, in the case of the tissue, cells, the action is
accentuated rather than inhibited by addition of
serum.**
One investigator52 has drawn attention to (he
similarity lietween intoxication by riein and circula-
tory shock. He has found some evidence of dimin-
ished blood volume in poisoned rats and of reduced
peripheral circulation in the rabbit. The latter effect
he was inclined to attribute to pooling of blood in the
splanchnic area. He considered, but dismissed, (he
thought that this condition might be due to capillary
blockage resulting from agglutination in vivo. An in-
cidental observation bearing on this question was
that the rate of absorption of iron from the gut and
the amounts deposited in tissues were increased in
poisoned animals. He draws attention to a similar
observation on animals in peptone shock.69
In conclusion it is worthy of remark that no effect
of riein on unicellular organisms or isolated tissues
has been clearly established. Much more work in this
field is desirable as are more detailed studies of the
time course of metabolic disturbances in poisoned
animals and the level of differentiation of tissue or-
ganization and function at which susceptibility to
poisoning first becomes manifested.
12.t IMMUNOLOGY d
In (he United States active research on the im-
munology of riein was initiated in February 1943 by
NDUC Division 9 (Section 9.4.2, Immunochemical
d By Birdscy Uenshaw.
SECRET 192
HK.1N
Studies).3 ,8 Related work was subsequently taken
up by other NDRC investigators,*141417 by the
Chemical Warfare Service,2- 23 2,1 27 28 31 and by the
Commit tec* for Medical Research.19 At the time the
NDRC research began there were available, in addi-
tion to the open literature* on riein, an account of
studies carried out for the Chemical Warfare Service,
during World War 124 and reports on preliminary
work conducted in the United Kingdom during 1040,
1941, and 1042.33-3®'40'41 More recently Canadian in-
vestigators have made a significant contribution.43
The principal objective was to provide and eval-
uate immunological procedures for protection against
and treatment of riein poisoning. With respect to
protection, the aim — not yet attained — was the
production of a toxoid which could practically lx*
used to immunize troops. With respect to treatment,
the problem — now satisfactorily solved— was the
production and evaluation of potent antiricin serums
and antibody globulin preparations. A secondary ob-
jective was the study and evaluation of immunologi-
cal methods for detection and estimation. By-
products of the immunochemical work have been
significant contributions to the purification ami
physicochemical characterization of riein.31*
For purposes of orientat ion it may be stated at the
outset that immunological, ultracentrifugal, and
electrophoretic studies on riein preparations from
castor beans of different source and color have failed
to reveal the existence of more than one heat-labile,
toxic antigenic protein.318 On the other hand, a non-
crystalline fraction (HI) from castor beans, which
by these criteria is identical with crystalline riein, is
not so toxic as the latter.319' Furthermore, solubility
studies do not reveal the crystals to be homogeneous,2
It is also known that castor beans contain, in addi-
tion to heat-labile toxic protein, one or more heat-
stable antigenic substances of low molecular weight
(allergen); small amounts of allergen appear to be
present even in crystalline riein.14 43
12.4.1 Preparation of Riein Toxoids
Incomplete success has attended efforts to produce
from riein a toxoid possessing high antigenic potency
coupled with negligible toxicity and skin-necrotizing
properties. The available toxoids are satisfactory for
eliciting vigorous antiricin production in animals.
The best has been recommended for the active immu-
nization of volunteers on an experimental scale but is
not considered suitable for practical use in the routine
immunization of troops.
The most satisfactory toxoid has boon prepared by
formalinization of the toxin as follows: 3 l8m riein at a
concent rat ion of 0.5 nig riein nitrogen per milliliter in
0.15.1/ sodium chloride plus 0,02.1/ phosphate buffer
at /di 7.4 is treated with 5 per cent formalin for
5 days at 37 C. Originally, partially purified pilot,
plant riein (Hi fraction |Si-Vvas used.
Recently crystalline riein has been utilized with sim-
ilar results 1Ja and will undoubtedly be employes! in
all future work. For best results the toxoid is pre-
cipitated with alum or protamine. The resulting
toxoid is about one-thousandth as toxic for mice as
native riein.3-,8ml However, subcutaneous injection
of as little as 0.1 eg of the toxoid nitrogen produces
skin necrosis in some rabbits,19* and in the form of an
aerosol the toxoid is only about 15 times less potent
than native riein as a lung injurant.19*
Some observers Ixdieve that formalinization in a
more alkaline medium yields a better toxoid.14 Un-
doubtedly a greater diminution of toxicity is effected
under these conditions, but the indications are
(hat antigenicity is more than correspondingly re-
duced.3ISrtu Precise evaluations of toxoids prepared
at pH values differing by only 0.1-0.2 unit are not
available.21 The concentration of formalin is not
critical; even high concentrations do not effect com-
plete detoxification, and 0.5 per cent suffices to pro-
duce a toxoid suitable for many purposes.3 Some
consideration has been given to the chemical re-
actions that occur during toxoid formation.1414
No success has attended numerous attempts to
produce a toxoid more effective than that just de-
scribed. Among the procedures to which riein has
been subjected are the following: oxidation with
chlorine or permanganate;40 ultraviolet irradiation
at low intensities31St‘-f and for short times at high
intensities; *IXu acetylation;3I8ik tryptic diges-
tion;19a<' peptic digestion; 19b,‘ treatment with nin-
hydrin;14 and heating.14A toxoid prepared by shaking
riein with toluene showed some promise in pre-
liminary tests but remains to be completely evalu-
ated.191’ Injections of formalinized toxoid treated
with normal serum and of specific precipitates of
formalinized toxoid with antiricin rabbit serum
proved unsatisfactory for active immunization.318'
A few additional procedures have been suggested 21
but were not evaluated before the work terminated.
A finding of significance is that a purified but non-
crystalline fraction (HI) prepared from pilot plant
riein is immunologically identical with crystalline
riein but possesses only 00 per cent of the toxicity of
SECRET 193
IMMUNOLOGY
the latter.* 17b r l,“ This observation suggested (1) that
some form of detoxified riein either exists in castor
beans or is produced in the process of extraction and
purification, and (2) that crystallization effects at
least a partial separation of the toxic from the de-
toxified material. That detoxified material immuno-
logically indistinguishable from riein is indeed pres-
ent in castor beans is suggested by the further finding
that crude aqueous extracts from the beans also pos-
sess considerably more immunologically active ma-
terial per unit amount of toxic material than does
crystalline riein.*asu.i**,!- ptp p, nmv p has been possi-
ble to effect only a very incomplete separation of the
toxic and nontoxic fractions.1** * However, further
study of the conditions and factors responsible for
the origin of detoxified riein in castor beans might
lead to a solution of the toxoid problem.'19r In such
work the changes which may take place in develop-
ing and germinating beans should be examined.
There is evidence that castor bean allergen *7 is
not completely removed from the heat-labile riein
by crystallization M or even by repeater! recrystal-
lizations.14 Injections of a toxoid containing even
small amounts of allergen conceivably might render
men hypersensitive to the allergen contained in sub-
sequently injected toxoid, and to sublethal dosages
of airborne riein containing allergen. Some workers
are inclined to minimize* the practical importance of
this possibility; to others14 it has been a source of
great concern. Animal experiments bearing on the
point are reviewed in Section 12.4.4, and limited
human data are presented in Section 12.4.3.
12.4.2 Antiricin
Potent antiricin rabbit, horse, and goat serums
have been obtained by a series of injections, first
subcutaneous and subsequently intravenous, of riein
toxoids.*1,28 Immunization can be continued with
alum-precipitated but otherwise untreated riein. For
therapeutic purposes the hyperimmune serums may
be used as such, but the antibodies are preferably
purified to lessen the likelihood of immediate reac-
tions and scrum sickness.
Standardization of Antiserums
During World War 11 antiricin has been estimated
with reference to an American Standard Antiserum
arbitrarily assigned a potency of 100 units ml.!# 3,1
Each milliliter of this serum* contains antibody
equivalent to about 7,500 mouse LDh«doses of riein;
that is, by the toxicity test described below it neu-
tralizes 200 ng of crystalline riein nitrogen1*1 or
500 ng of nitrogen of the relatively impure pilot plant
preparation against which it was first tested.*11
Two tests have been developed for the quantitative
assay of anti riein titer: M*®
1. Toxicity-neutralization. Solutions of known
amounts of riein and of antiserum are mixed in 0.9
per cent saline, incubated at 57 C for x/i hour, and in-
jected int rapedtoneally into mice. The minimum
volume of scrum in the mixture for which mice sur-
vive for 10 days is considered to he equivalent to the
amount of riein used. The toxicity-neutralization
test may be used to defect as little as 0.2 unit of anti-
body and is the method of choice if time, permits.
2. Inhibition of hemagglutination. Portions of
riein (e.g., 2 ,ug in saline) are mixed with decreasing
volumes of scrum and saline is added to a volume of
0.8 ml. After incubation at 37 (' for hour, 0.2 ml
of a 4 per cent suspension of washed human erythro-
cytes of blood group () are added. The extent of
agglutination is refolded after shaking and incubat-
ing at 37 C for 1 hour. The minimum amount of
serum which completely inhibits hemagglutination
is considered equivalent to the amount of riein used.
Because of nonspecific inhibition by normal serum,IM
this test cannot be used to measure less than 5 units
of antibody per milliliter.18*'1 In the choice of riein
for use in this test consideration must be given to the
fact that the hernagglut mating properties can be re-
versibly masked under some circumstances.31518111’31*
Potency op Antiserums
The use of graded series of injections of riein t oxoid
and/or native riein has yielded in rabbits antiserums
having potencies as high as 250 imits/ml .* In horses
serums possessing 150 units of antibody per milliliter
have been obtained.* However, few animals have
been observed with circulating antibody titers greater
than that of the standard, and most animals in any
series will attain titers more or less below it. Never-
theless, the pooled scrum from a group of adequately
immunized rabbits possesses what can be considered
for therapeutic purposes a high and effective titer.
Purification of Antiricin
To reduce the possibility of reactions from the
therapeutic use of antiricin scrum, methods which
had been used for the partial purification of other
antibodies were applied.* Sufficient experience has
* Available at the Medical Research Lalwatory, Kdgewood
Arsenal.
SECRET 194
R1C1N
been gained to make possible the production of con-
centrated, partially purified horse or rabbit antiricin
rapidly and on as large a scale as any program might
require.
Antiricin globulin from immunized rabbits was
obtained in almost quantitative yield by 45 per ecut
saturation of the diluted serums with sodium sulfate
at 37 C. The precipitate was dissolved in water,
merthiolate added as a preservative, and the solution
sterilized by passage through a Chamberland fil-
ter.18* 1 The ampouled material possessed an anti-
ricin potency of 50 to 125 units ml and was pre-
pared in sufficient quantity for distribution to the
Chemical Warfare Service and NDRC laboratories
engaged in work on ricin. Prompt intravenous in-
jection of 25 ml was recommended in the event of
accidental inhalation of ricin aerosols.
Horse antiricin was partially purified by isolation
of the pseudoglobulin fraction '*"' *,* h or by peptic
digestion by the Parfentiev method.1*5 3lh The latter
method was used to process Ifi 1 of horse plasma
assaying 50 units of antiricin per milliliter. The yield
was 1,090 ml of purified, modified globulin solution
assaying 500 units/ml.1 * —
Therapy with Anthucin
Antiserum or purified antibody globulin is of con-
siderable therapeutic value if promptly adminis-
tered. Its effectiveness rapidly decreases with in-
crease in the time between poisoning and therapeusis,
and no benefit is obtained after the delayed symp-
toms of poisoning have appeared.’18 31A0
Serotherapy is effective against injeetions of at
least several lethal doses of ricin if sufficient antiricin
is administered promptly.318 31 Typical data for mice
are presented in Table 4.181
After exposure to airborne ricin the pathological
effects occur mainly in (he lungs31' and the animals
are not completely protected against pulmonary in-
jury even by immediate therapy with injected anti-
serum.31'1 11 However, the use of antiserum has defi-
nite life-saving value up to 6 hours after gassing and
is perhaps of limited lienefit even as late as 10
hours.3181"31'1 Illustrative data are presented in
Table 5.18n The data reveal the desirability of utiliz-
ing a large amount of antiriein. Equivalent amounts
of rabbit antiserum, purified rabbit antibody globu-
lin. and purified horse antibody pseudoglobulin ap-
Table 4. Therapeutic use of antiricin in mice poisoned
with riein by intraperitoneal injection.*"i
Mice were injected intraperitoneally with about 20
lethal doses (2 of BI ricin nitrogen) and subsequently
injected intraperitoneally with ten times the neutralizing
equivalent of antiricin (rabbit antiserum).
Mortality
Treatment 0-24 hr 24-48 hr 2 10 da
ys Total
Xo serum 36 0 0
36/36
Serum 0.5 hr
after ricin 0 0 0
0 37
Serum 2 hr
after ricin 10 5
6/38
Serum 5 hr
after ricin 9 3 9
21/37
Tabus 5. Therapeutic use
of antiriein in mice poisoned
with ricin by inhalation.”"
—
Mice
were exposed for 10 minutes to an aqueous aerosol
of ricin at a nominal concentration (about 8 neritoneally at various times after theexposure.
Anliricin
Time of
admin-
treat-
Mortality
istered
ment
0 4 day!
* 4-8 days 8-10 days Total
100 units
None
20
0 0 - 20/20
1 hr
0
0 I 1/17
4 hr
2
3 0 5/19
10 hr
5
9 2 16/18
2-4 hr
17
3 0 20/20
10 units
None
10
0 0 16/16
1 hr
2
4 1 7 20
4 hr
1
10 0 11/18
10 hr
11
7 2 20/20
pear to possess approximately t lie same therapeutic
efficacy.*1*1' "
Passive Immcnitv
Passive immunity results from the injection of
high-titer antiserum or partially purified antiri-
cin.31s 3u b c *‘*f On the basis of these animal experi-
ments, however, the protection cannot be expected
to persist for more than 1 or 2 weeks at most. Thus
passive immunization would have limited usefulness
as a practical method for protecting troops.
12. t.3 Active Immunization against Ricin
Injection or inhalation of native ricin in small doses
evokes antiricin formation31*' and immunity2740
in surviving animals. The response is sometimes
striking, particularly after repeated administration
of the toxin. Practically speaking, however, active
immunization must be attained the use of a
toxoid.
( This material, put, up in amjxniles each containing 12 ml,
is available at the Medical Research Laboratory, Eclgcwood
Arsenal.
SECRET IMMUNOLOGY
Table fi. Resistance to airborne riein of rabbits immunized with six injections of formalinizcd ricin.19*'
Tin; exposures in the gassing chamber were of 10 minutes’ duration. The 10-minute IAfor nonimrmmized rabbits was
about 0.5 gg ricin nitrogen per liter. Thus the exposures were to approximately 4 and 20 times the A,(C/)5,i dosage.
Interval between Serum antibody
Cone, of ricin in
Lung pathology
Serum antibody
titer in survivors
last toxoid injee- tiler before eX|>o-
gassing chamber
Deat hs in 10
in survivors 14 days
14 days after
tion and exjxjsurc sure (units)
(#ig nitrogen /I)
days
after exposure.
exposure
12 da vs 0.8 3.3
2.3
0 10
- to + + +
1-8
1.0-3.3
11.8
3/9
+ to -f- T +
6.5-12
3 months <0.2 0.8
2.3
0/7
- to 4-4-4-
8->30
<0.2 0.2
0.8
4/7
+ + to 4- 4~ +
12->30
5j months <0.2 0.6
2.1
3/8
— to 4- 4-
2 4
<0.2-11.2
11.0
5/7
-(- to 4-4-4-
S- >30
Sn diks with Animals
Several injection schedules have been used for
studies with rabbits on the development of active
immunity to inhaled ricin.3!9' The first schedule con-
sisted of three subcutaneous injections at 5-day in-
tervals of formalinized toxoid in the amounts of
2.5, 5, and 10 /ig of ricin nitrogen per animal, respec-
tively. This schedule resulted in circulating antibody
levels of 1-3 units per milliliter and many animals
survived exposure to about 20 L{Ct)„0 dosages of air-
borne ricin 10 days after the last injection. However,
(he injections of toxoid produced severe skin re-
actions with necrosis. A schedule of six injections
conferred equal or greater immunity and the severity
of the local reactions at the sites of injection was
greatly reduced, although necrosis was not absent in
all instances.19ac A dosage sequence of 0.1, 0.2,
0.5, 2, 10, and 20 of toxoid nitrogen is believed
preferable to a schedule composed of doses of 0.1,
0.5, 2, 5, 10, and 20 gg.
Some of the characteristics of the antibody re-
sponse and protection effected in rabbits by six
toxoid injections are illustrated by (he data of
Table 6.,9r' Maximum antibody response and pro-
tection is attained 10 to 20 days after the last injec-
tion. At this time the circulating antibody levels are
1 to 3 units per milliliter of serum. The animals are
immune to the lethal effects of at least several
dosages of airborne ricin, but the development of
lung lesions is not prevented. Circulating antibody
titer then falls progressively to reach levels of the
order of 0.2 unit per ml after 2 to 1 months.3 '■*' 3,‘-
In spite of the low level of circulating antiricin, some
resistance to airborne ricin porsis(s.,’,(i‘, t ,!k:
After circulating antiricin has reached a low level,
an injection of toxoid produces only a moderate in-
crease in circulating antiricin.3 |Sr u In contrast, a
very striking increase in circulating antiricin, to
5-30 Units per ml of serum, is produced by a single
exposure to a sublet 1ml dosage of airborne ricin.,1lH" u-
lih The effect is to be seen when the exposure follows
by only 12 days the last of a series of toxoid injec-
tions. If appears to be mow marked after longer
times, when the circulating antiricin evoked by the
toxoid injections has fallen to low values.1'1' A second
exposure to airborne ricin does not elicit a pro-
nounced further increase in the circulating anti-
bodies.l8u These findings suggested that, for pur-
1 roses of active immunization, controlled exposure to
aerosols of ricin toxoid might effectively reinforce the
effects of toxoid injections. No studies on immuniza-
tion by inhalation of toxoid have as yet been made,
however, except for one experiment in which previ-
ously unt reated rabbits were given a single exposure
to airborne formalinized toxoid.,9a Twelve days later
none of the survivors had develojred a circulating
antiricin titer as great as 0.2 unit per ml. Challenge
exposures to airborne ricin were not made.
Actively sensitized guinea pigs possess consider-
able immunity against the toxic effects of ricin.S18,, t
Subsequent to exposure to 15 40 L(Ct)5n dosages of
airborne ricin, a high proportion of the animals (i.e.,
31 of 33) that recovered from the initial anaphylactic
reaction survived indefinitely. This degree of re-
sistance was present in the three animals tested as
late as 116 to 173 days after sensitization.
Although the results show that considerable re-
sistance to inhaled ricin can be achieved by immu-
nization with formalinized toxoid, it has been empha-
sized that the resistance has been measured by a
statistical increase in the number of animals surviv-
ing challenge exposures and that the surviving ex-
SECRET RICIN
posed animals usually develop lung lesions.19*- Some
of these lesions are severe and apparently predispose
the animals to bronchopneumonia.
Human Immunization
With regard to immunization of large numl>ers of
troops, representatives of the Surgeon General
(Army) have indicated that practical considerations
make it highly desirable to limit the numlier of in-
jections of toxoid to one or at most two or three
spaced over a period of 1 to (5 weeks.20 The animal
experiments give no reason to believe that effective
immunization can be obtained with so few injections
of currently available toxoids at doses sufficiently
small to preclude very severe local reactions. There
is, moreover, no evidence that effective immunity,
if once produced) would persist at high levels for
more than a few months.
No experiments on human immunization have
been made. However, the use of formalinized toxoid
at a dosage schedule similar to that employed for
immunizing rabbits has been recommended as safe
for test with volunteers. It was felt that data on the
local effects produced and on the levels of circulating
antiricin attained would help to orient the further
course of the work.
Numerous scrum samples from men working with
ricin have been assayed for circulating antiricin by
the toxicity-neutralization lest.3,,8dNo signifi-
cant amounts of circulating antibody were found be-
fore exposure to ricin. Considerable antiricin (i.e., up
to 2.5 units ml) was found in the serums of men who
had handled ricin at the pilot plant for several
months or more. The highest levels were found in
men having histories of either cuts and abrasions or
symptoms traceable to ricin. There is no direct evi-
dence as to the degree of immunity possessed by
these individuals.
In the experience of the University of Chicago
Toxicity Laboratory two types of reaction to acci-
dental exposure to ricin have been observed.9 17,1
One, a delayed reaction, sets in after a latent period
of 5 to S houre. A febrile response then occurs, ac-
companied by tightness of the chest, tracheitis, aeh
ing joints, nausea, dyspnea, and coughing. In e viewed with suspicion in the
case of detection and analysis of unknown ricin
samples because of the possibility that hemaggluti-
nating properties can la: masked.3-1518,1131*
PnKOI CITIN' RKACTION
Although the addition of ricin to normal serum
produces a precipitate under certain conditions,
much smaller amounts suffice to produce a specific
precipitate with antiricin serum. Thus (he precipitin
reaction is highly specific and sensitive; it is capable
of delecting I pg of ricin nitrogen within 5 minutes,
and much smaller amounts in longer times.3-18'1-1
Although less subject than hemagglutination to ex-
traneous conditions, it must, be borne In mind that
serum protein is precipitated by the ions of heavy
metals which are present in smokes of various kinds.3
With the limitation that different ricin preparations
possess different ratios of toxic potency to immuno-
logical activity (Section 12.4.1), the quantitative
precipitin test affords a very accurate method for
the estimation of ricin.318p r■* Under optimal condi-
tions it is accurate to about 1 per cent.
Anaphylactic Responses of Sensitized Animals
The anaphylactic response of actively sensitized
guinea pigs appears to provide the most rapid, spe-
cific, and sensitive method for the detection of air-
borne ricin or ricin dusts that have settled on sur-
faces.322 The animals must Ire watched, however,
ami the possibility of desensitization guarded
against.318,1 Passive sensitization was earlier em-
ployer! by British investigators 33 40-41 but active im-
munization has the advantages of inducing much
more prolonged sensitization and. probably, greater
sensitivity.3-,sik’n p-' The rapidity and sensitivity of
the reactions was demonstrated by tests in which
characteristic responses were evoked within 1-3 min-
utes after exposure to ricin dust at the lowest nominal
concentrations tested, 0.03 mK ricin nitrogen per
liter.3Tin’s concentration was in the order of
one-thousandth the 10-minute LCi0 for mice.
Anaphylactic reactions of guinea pigs are known
to 1m> highly specific. In the case of the studies with
ricin, however, them has been debate, not yet re-
solved. as to whether the reactions are due to sensi-
tivity to the toxin itself or to contaminatings castor
bean allergen. The evidence that guinea pigs can be
sensitized to ricin itself, irrespective of the possibility
that hj-persensitivily to allergen may also occur, may
be summarized as follows:
J. Guinea pigs passively sens!(ized by intravenous
injections of rabbit antiserum to a fraction (Bl),
which contained in relatively purified form most of
the toxin in pilot plant ricin, were subsequently in-
jected with fraction Bl and with fraction B3, a
gummy fraction presumably containing much castor
bean allergen but virtually free of ricin itself. Injec-
tion of fraction Bl uniformly produced fatal ana-
phylactic shock, whereas injection of fraction B3
produced much less severe reactions. The animals
which received fraction’ B3 were fatally or severely
shocked by subsequent injections of Bl.318k.
2. All guinea pigs that had been immunized by
injections of ten times recrystallized ricin, of pilot
plant ricin, or of alum-precipitated ricin toxoid
showed anaphylactic responses when injected intra-
venously with crystalline ricin or when exposed to
airborne pilot plant or crystalline ricin at relatively
low coticent rations.3 •17d 181
3. Guinea pigs injected with ricin develop con-
siderable immunity to the toxic effects of ricin (Sec-
tion 12.4.3). In general, immunity in guinea pigs goes
hand in hand with hypersensitivity.
Since the completion of this work, Canadian in-
vestigators 43 have reported failure of attempts to
elicit anaphylactic responses in guinea pigs sensitized
with crystalline ricin and exposed to airborne pilot
plant or crystalline ricin. Animals immunized with
pilot plant ricin showed weak responses. On the
other hand, animals given a single injection of castor
bean allergen M-R7 reacted vigorously to crystalline
ricin as well as to pilot plant ricin in low concentra-
tions. These data, considered in conjunction with
supplementary results obtained by the use of the
Schultz-Dale technique, led to the conclusions that
* The sensitivity chiimcd for the hemagglutination reac-
tion in reference 27 appears to be in error.
SECRET 198
iiir.iv
the sensitization was to allergen rather than to toxin,
and that allergen is present in crystalline ricin. Addi-
tional evidence that a small amount of allergen is
present even in many times recrystallized ricin has
since l>een presented.14
Further work is required to clarify the apparent
discrepancies. In any event, it is evident that guinea
pigs can be prepared in such a way as to render thorn
highly susceptible to anaphylactic shock upon ex-
posure to very low concentrations of all known riciir
preparations.
The impract icability of employing (he reactions of
animals other than dogs as routine methods of de-
tection in warfare has often been emphasized. How-
ever, general considerations as well as the results ob-
tained during field trials with ricin 3 22 would indicate
that sensitized guinea pigs could be of great value in
the hands of special officers assigned the duty of
checking upon the possible use of protein toxins by
an enemy.
12.5 \SSA Vh
It was early recognized that the toxicity of castor
beans was associates! with the water-soluble, heat-
coagulable protein of the beans.*4 55 The presumption
was that the toxicity was the unique property of a
single protein component, and this hypothetical com-
ponent was designated ricin. In 1943, a crystalline
protein was isolated from extracts of castor beans.5-15
Its toxicity was reproducible and was about twice as
great as that of the most active amorphous product
available.9-15 This result greatly strengthened the
presumption that there is present in castor beans a
single toxic protein component. However, although
the crystalline product has met some, it has not met
all, of the criteria of molecular homogeneity which
are required of a single protein.2-315 The possibility
cannot yet be rigorously excluded that the toxin is a
complex whose components may. some day, be sepa-
rated from one another and may then be found to be
active only in association with one another. As long
as this possibility remains, the only assay of the toxin
of castor bean preparations to which no objection can
l>e raised is an estimation of the toxicity under ap-
proved experimental conditions. Theoretically, the
measurement of any physical, chemical, or biological
property which has !>een shown to l>e quantitatively
related to the toxicity should serve as an assay. Un-
fortunately, the demonstration of the existence of
such a relation cannot he complete until the pure
toxin has I>een isolated and fully characterized. A
variety of properties of extracts of castor beans have
been proposed as bases for assay and these will lie
reviewed briefly. The problem of bioassay will, how-
ever, lie considered first and in fuller detail.
12.3.1 Bioassay
All vertebrates that have been tested are suscep-
tible to ricin (see Section 12.3). Few observations
have lieen made on invertebrates. It has been re-
ported 5 that the motility of certain sfieeies of pro-
tozoa is arrested by the addition of ricin to the
medium, but it has not been established that the po-
tencies of a series of preparations of ricin are propor-
tional to their toxicities. Difficulties in controlling
the action on the protozoa led to abandonment of
the attempt to use these organisms as a means of
assay.
Ricin acts slowly on vertebrates. With minimum
lethal doses, animals seldom die in less than 5 or <>
days, and may survive for weeks. Assays based upon
the estimation of median lethal doses are, therefore,
protracted and require an arbitrary choice of obser-
vation period. They also require large numbers of
animals for statistical validity because of incidental
variables such as casual infections. Where many
assays must be carried out, there are obvious advan-
tages in the adoption of a method which gives quick
results and is economical of animals.
The value of establishing a relation between the
dose of ricin and the survival time in a given species
as a basis of a method of assay was urged in 1918.24
In this country the problem has been investigated
in three different laboratories.5 *15 ®1 Work was also
done in Canada41 and in England.®-37 The mouse
has been the favored animal, being preferred to the
rat.12 Several homozygous strains have been used,4 9
the most popular one in the United States being the
CFl strain of white mice developed by Carworth
Farms, New City, Rockland County, New York.5 *-14
A method of assay based upon 24-hour mortalities
has l>ecn adopted. Groups of five or preferably ten
mice weighing 20-25 g are injected with a series of
graded doses by the intraperitoneal route. The indi-
vidual survival times of the animals are recorded and
the mean survival time for each dose is derived. By
interpolation in an accepted dose-survival time rela-
tionship, a value for the dose corresponding to a
mean survival time of 21 hours is, then, obtained for
each experimental dose. This 24-hour lethal dose has
1 By R. Keith Canaan.
SECRET ASSAY
199
been designated the toxicity unit [TU], Because of
the skewness of the distribution of survival times,
one investigator 4 prefers to convert each individual
death time in a group to a TU and to average these
to give the TU for the group.
It has been customary to express the TU in micro-
grams of the preparation per 20-g mouse though it is
sometimes more useful to express it in terms of the
total nitrogen or the eoagulable nitrogen present in
the preparation. The desirability of making a simul-
taneous assay of a standard product has l>een empha-
sized by all laboratories concerned with the evalu-
ation of ricin preparations.5 915 When this is done,
one may then readily express the toxicity of an un-
known preparation in terms of the per cent of the
standard which it contains. Crystalline ricin is the
logical reference standard.
No difference in susceptibility to ricin of the two
sexes of the CF1 strain of mice has been detected.9
In the cases of two other strains, small differences
have lieen recorded.5-4- Fean mice seem to lie more
resistant than fat mice of the same weight.4 42 This
is probably because a greater proportion of the body
weight of the lean animals is active tissue. Studies of
the effects of diet 5 also have led to the conclusion
that the toxicity is a function of the ratio of active
tissue to body weight. Mice to be used for assay
should he fiee from parasites. The susceptibility of
young mice is found to be greater when the environ-
mental temperature is elevated and is reduced at low
temperatures.4-42 This is presumably due to corre-
sponding changes in body temperature. Frogs are
also sensitive to ricin only at elevated temperatures.*5
In the conduct of assays with mice, the temperature
at which the mice are kept should be controlled. The
preferred temperature has been close to 25
In one laboratory,5 automatic devices for the con-
trol of temperature, the injection of the animals, and
the recording of individual death times have been
employed with the object of rendering the conditions
of assay as uniform as possible.
If the preparation of ricin is very active, it must be
diluted to a concentration of 10 50 mg per liter be-
fore injection. The danger has been emphasized 5 of
losses by adsorption on the walls of the vessels in
which the solution has Ix-en prepared. Adsorption is
said to I>e significant even when paraffin-coated
vessels are used. To reduce this error, a solution of
0.3 percent egg albumin has Ixren used as the diluent
on the principle that the excess of inert protein will
inhibit the adsorption of the toxin.5 Shorter sur-
vival times are observed when this procedure is
adopter!,5 * IS but there has Ihh-ii debate as to whether
this was due to the suppression of adsorption or was
the result of a synergistic action of the egg albumin.
The modified technique has not found general ac-
ceptance.9 14 When a simultaneous assay of crystal-
line Hein is made and the result is expressed as the
per cent of crystalline ricin in the product, it is prob-
ably immaterial whether water or 0.3 per cent egg
albumin is used as the solvent. On the other'hand,
toxicity unit values derived from assays of egg al-
bumin solutions of ricin arc consistently smaller
than those of aqueous solutions and are not directly
comparable with them.
The Dose-Sihvival Time CTkve koh Mice
For a given route of injection, this curve approxi-
mates a rectangular hyperbola of the type repre-
sented by the relation:5 9 15
if) - /)-) __
it ~ tm)
where I) is the observed dose, I is the observed sur-
vival time, and k is a constant characteristic of the
preparation of ricin. Dm and tm are constants having
the qualities of an extrapolated minimum lethal dose
and an extrapolated minimum survival time respec-
tively. Over a wide range of lethal doses a single pair
of values of 1)m and f,„ fils the experimental observa-
tions only very roughly. Over restricted ranges of
survival times, however, values of the constants can
l>e so chosen as to fit the data quite satisfactorily.*
For short survival times, /)„. is small relative to D
and may be ignored. Then, since the toxicity unit is
the value of D when I = 24 hours, the above equa-
tion can be rewritten in the form:
TO-B?
(< -
This assumes that tm is independent of the nature of
the material which is being assayed. If this assump-
tion is not correct (and it has been implicitly ques-
tioned) 5 the assay of an unknown product by in-
terpolation in a standard curve would lie subject to
error. However, extensive observations 915 have indi-
cated that the use of tm = 13 hours satisfies the in-
traperitoneal data for both crystalline ricin and
standard ricin over a range of survival times of about
18 to 30 hours. These laboratories have, accordingly,
adopted the following equation for general use:
SECRET 200
RICIN
TU = H
D (f - 13)’
In order that the uncertainty of interpolation should
l>e minimized, it is recommended that values of TU
should lx* computed only from mean survival times
falling within the limits of 21 and 28 hours.
A summary of the data of one laboratory 9 on
crystalline ricin and standard ricin is given in Table 7.
In Table 8 will lx- found a comparison of the results
M'iik Quantitative Precipitin Method3
Antiserums prepared by the injection of crystalline
ricin have been found to precipitate from extracts of
castor beans, and amorphous preparations generally,
not only the toxin, but a noutoxie protein.* '6 Since
this material is antigenically indistinguishable from
the toxin, it may l>e called a natural toxoid. In the
preparations that have been tested, the ratio of toxin
to toxoid has varied from 1 1 to 2 I. Only by the
process of crystallization has a separation of the two
antigenic components been accomplished. If this
limitation of the "method is borne in mind, the pre-
cipitin technique (see Section 12.4) is a valuable
method of assay. It requires only a few micrograms
oLpurified material and gives a positive result within
a short time.
H KM AGGLU T l N ATI O N
There is a wealth of evidence that the hemagplu-
t mating activities of ricin preparations do not parallel
their toxicities.*5-16 24 :t4 Crystalline ricin has, for ex-
ample, only about 20 per-cent of the agglutinating
power of amorphous preparations which are consid-
erably less toxic.*16 The method can, therefore, have
only limited use.
A method of assay based on hemagglutination
(sex* Section 12.1) has been proposed for use in the
field.2-’ 23 26 Agglutination tests are rapid and require
only small samples of material. However, the cus-
tomary method of evaluating the agglutinating
potency of a sample is only coarsely quantitative
and depends on subjective discrimination by the ob-
server. An attempt to increase the objectiveness and
precision of the method has been described.16
Enzymic Activity
The observation that crystalline ricin does not ex-
hibit the esterase, phosphatase, and lipase activities
of crude preparations eliminates these properties as
means of assay.5 It has recently been reported 43,50
that ricin preparations hydrolyze adenosine triphos-
phate. If it should be established that this activity is
proportional to the toxicity in a representative series
of preparations, a valuable alternative to bioassay
may become available.
Chemical Methods
No chemical property of the toxin is known which
distinguishes it from other heat-eoagulable water-
soluble proteins of the l>ean. However, water ex-
tracts from castor bean contain little coagulable pro-
Tabu-; 7. Summary of a series of assays of crystalline
ririn and of standard ricin carried out at intervals over a
period of 13 months.’ The CFl st rain of mice was used.
Solutions were prepared and diluted with water.
Crystalline ricin Standard ricin
Male Female
Male Female
Total number of assays 23 22
13 12
Mean toxicity unit
pig of material
—
I>er 20-g mouse 2.00 2.04
0.06 7.01
Standard deviation 0.21 0.17»
0.7!t 0.72
Table S. Dose-survival time relationsJor different routes
of injection. The CF1 strain of mice was used. Solu-
tions were prepared in water and injected in a volume
equal to I |xt cent of the body weight.
Standard equation: 0(1 — l„) = k.
Intravenous Intraperitoncal Subcutaneous
k (rig-hours)
8.C 23 - 50
l„ (hours)
11 13 1C
TU ng 20 k
0.0C “ 2.1 0.3
LD:o egkg
2.2 10.4 22.1
of intravenous, intrapeiitoneal, and subcutaneous
injections.* The method of intravenous injection is
technically too difficult for routine assays. All in-
vestigators have agreed that intraperitoneal injec-
tions yield more precise and reproducible results than
do those by the subcutaneous route. Subcutaneous
toxicities have been found to vary to a remarkable
degree with the concentration of the solution which
is injected.*
12.5.2 Alternative Methods of Assay
A variety of properties of ricin preparations have
l»eon proposed as bases for assay. These include the
antigenic properties, the hemagglutinating potency,
and various enzymic activities which have luxm
found in crude preparations of ricin. The weight of
evidence is that none of these are specific for the
active toxin. Some of them are valuable for the com-
parison of limited types of preparation, but must be
supplemented by bioassays in critical situations.
SECRET evaluation as a war <;as
201
tein other than the toxin and the toxoid.915 In such
extracts, an estimation of the heat-denaturuble pro-
tein gives a result only slightly greater than the esti-
mation of the protein precipitated by antiserum.915
In extracts prepared with salt solutions, on the other
hand, a large additional amount of coagulable pro-
tein is present.15 54 Fortunately this is denatured at
or below /dl 4. If such extracts are acidified and
filtered, the soluble coagulable protein which remains
corresponds closely to the sum of the toxin and tox-
oid. The heat-coagulable protein has usually been
estimated as the difference between the soluble pro-
tein before and after boiling for 15 minutes to 1 hour
at 100 C. Any acceptable met lux 1 of protein determi-
nation may be used which is adapted to the amount
of protein present.'5
I2.r>.;t Field Detection and Assay
For the rapid detection of airborne ricin, the sensi-
tized guinea pig is undoubtedly the most sensitive
and specific*-" (see Section 12.4). The maintenance
and care of sensitized animals in the field, however,
present many difficulties. Moreover, there is some
question whether the anaphylactic reaction is elicited
by the toxin or by a so-called allergen which may be
separated from it.5 53
Any other method of detection or assay requires
the collection of samples adequate in amount for the
test which is to be performed. Certain color tests have
.been proposed and are both sensitive and rapid.5 In
so far as they are simply tests for protein or for the
carbohydrate commonly associated with protein,
they are entirely nonspecific and are of value only in
indicating the possible presence of ricin. More spe-
cific, but less suited to field work, are the hemag-
glutination and precipitin tests. Some limitations of
the former have been mentioned. The precipitin re-
action is decidedly more specific and more accurate.
It has, however, been pointed out that the heavy
metals present in some smokes will give nonspecific
precipitates with serum proteins. Neither test gives
an immediate response. An assay of toxicity is, of
course, the most dilatory of all.
It may lx* appropriate, in conclusion, to remind
the reader that the hazard of exposure to a non-vola-
tile airborne toxin cannot be evaluated simply from
the time of exposure and the concentration of toxic
material in the cloud. The inhalation toxicity is de-
termined in large measure by the particle size distri-
bution in the cloud. The relation of particle size to
toxicity is discussed in Section 12.3 and in Chap-
ter 15, w here methods of evaluating the particle size
distribution in a cloud are reviewed. In contemporary
field trials22 with standard ricin, it was found profit-
able to assay the cloud not only for toxicity and for
particle size but also for total protein and for heat-
coagulable protein. The two latter estimations pro-
vided useful information on the extent to which the
method of dispersal resulted in detoxification and
denaturation of the material with which the muni-
tions had l>een charged. The results of field trials are
reviewed in Section 12.6. ,
/
12.6 EVALUATION VS V WAR GAS1
The performance of field trials on munitions
charges! ricin and the interpretation of (he results of
these trials in terms of evaluation of the agent as a
war gas rest in large part on the laboratory researches
in the United States, Canada, and Great Britain on
particulate sampling and bioassay (Chapter 15b
The most significant criterion for effectiveness of
ricin in the field was bioassay by animals exposed to
the particulate cloud. Physical measurements were
essential to an understanding of the reasons for poor
or good results in the several trials and as a guide for
design of subsequent trials. Low toxicity in the field
could be associated with many variables, including
large particle size arising from compaction and aggre-
gation, thermal inactivation of the sensitive protein
agent, inefficient munition functioning, and meteoro-
logical conditions. The field trials also rested on the
prior development of pilot plant methods for the
preparation of finely divided ricin (Section 12.2).
12.6.1 Kelative Efficiency of Dispersion
by Different Munitions
The principal types of munitions and chargings
which have been studied for the dispersion of ricin
are the following:
1. High explosive-chemical bombs charged with a
suspension of ricin in carbon tetrachloride. Bombs of
this type, with steel casings and axial bursters, were
employed in the British experiments carried out in
1941 ■i3 M and in the recent Canadian trials.4* Muni-
tions of this type retain to a significant degree the
effectiveness of ordinary HE fragmentation bombs.
2. Light-case metal bombs charged with dry ricin.
The Canadian 4 lb L.C. bomb was a metal can hold-
ing about 550 g of ricin and fitted with a small burster
• By Stanford Moore.
SECRET 202
IUC1N
(e.g., 20 g of nitroguanidine and 70 g of sodium bi-
carbonate).'”43 44
3. Base ejection bombs charged with dry riein.
The U.S. M-71 10-lb tail ejection incendiary bomb
was modified for use with about 385 g of dry riein."-24
4. Gas ejection bombs charged with dry riein.
N DRC Division 10 carried out developmental work
on a two-compartment bomb holding liquid carbon
dioxide or compressed air in one compartment, which
on functioning ejected the particulate charging from
the second compartment."
5. Plastic and glass bombs charged with a susjkmi-
sion of riein in carbon tetrachloride. Experimental
munitions of this type developed by NDRC Divi-
sion 10 were similar to (1) above but with plastic or
glass casings instead of steel.44
The lines of investigation on dispersion of riein
from the various types of bombs at the several field
experimental stations have led to the same general
conclusions. The results indicate that high explosive-
chemical bombs charged with a 35 per cent suspen-
sion of riein in carbon tetrachloride ate superior to
the dry powder munitions in their ability to put up
a cloud in which the volume mass median diameter
is sufficiently small to pass the nasal barrier.®*-33-3*-
37-44 46 This conclusion confirms the earlier analysis
of the problem made by British investigators in
19413,-36 on the basis of a less complete series of
experiments.
In the field trials plastic bombs have given results
comparable with those obtained with steel bombs
but the British 4-lb HE Chem Type F Mk I steel
bomb, as used in the later Suffield trials, possessed
the advantages of availability in standard design and
durability in transport.
The lower dispersion efficiency of the munitions
charged dry powdered riein was largely the result of
the formation of aggregates in the particulate
clouds.22 Comparisons were based on parallel tests
employing a given sample of powdered riein set up
both in the dry and suspension forms.44 In trials with
dry samples, aggregation of the initial particles of
the riein charging to yield a cloud of larger mass
median diameter was increased by increase in the
moisture content of the charging or in the relative
humidity of the atmosphere."
In the 1941 British trials 33 36 it was concluded that
bombs filled with riein suspended in carbon tetra-
chloride were at least three times as effective as simi-
lar bombs filled with a solution of riein in water. In
more recent tests with plastic bombs the solutions in
water were also found to lie loss stable to detonation
than suspensions in carbon tetrachloride.17c-,i-f The
munitions were functioned in a stainless-steel ex-
plosion chamber at the NDRC University of Chicago
Toxicity Laboratory and a material balance deter-
mined. No measurable denaturation was observed in
the ease of the suspensions in carbon tetrachloride,
whereas a 40 per cent loss in toxicity occurred with
the aqueous Hein solutions. Chamber trials on the
plastic munitions were also carried out at the Divi-
sion 10 NDRC Munitions Development Labora-
tory.'
12.6,2 Comparison \silb Bombs Charged
Phosgene
On the basis of the early trials with suspensions of
riein in carbon tetrachloride the British investigators
concluded in 19413336 that bombs filled with riein
were about as effective as phosgene bombs of the
same size. With improvements made in the pilot
plant manufacture of dispersible riein since that
date, and progress in the testing of munitions, the
relative effectiveness of riein has been increased to a
position well above that of phosgene. The compara-
tive data have been analyzed by the Suffield Experi-
mental Station.46 From calculations of (he dosage
contours from the field trial data the munition ex-
penditures required for 80 per cent coverage of a
target area with a riein dosage of at least 100 nig/
min m3 have l»een calculated. The L(Ct)ui of riein
for man is not known. The results of the field experi-
ments indicate that for goats in the field the L(Ct)w
of the present pilot plant samples of riein dispersed
by the 4-lb UK Chem Type F bomb is about 100
mg min nr*. For the present calculations it is assumed
that this value holds for man. Employing (he meth-
ods of calculation applied to the test data on phos-
gene 33 it is estimated that for 500-lb clusters of
Type F bombs an expenditure of 1.2 clusters (43 lb
of riein) per 100x100 yard square would cover
about 80 per cent of the target area with an L(Cl)t,o
dosage on open terrain (neutral temperature gradi-
ent; wind speed less than 12 rnph). For 500-lb bombs
charged phosgene under the same conditions the
estimated expenditure is 8 bombs (1,000 lb of phos-
gene) pel- 100x100 yard square for coverage by a
dosage of 3,200 rng/min/m* within 30 seconds or
4 Ixjmbs for coverage within 2 minutes. The com-
parison is based on tests with a batch of spray-dried
1 These are reviewed in the Summary Technical Report of
Division 10.
SECRET EVALUATION AS A WAR GAS
203
air-ground ricin with a volume median diameter of
3.3 /i which yielded clouds of volume mass median
diameter of about 15 g.
From this it is concluded that ricin appears to !>e
at least seven times as effective as phosgene on the
basis of aircraft stowage when the comparison is
based on a 30-second dosage of phosgene. If the
L{Ct)so for phosgene is considered to be high by even
a factor of two there would still l»e a margin in favor
of ricin. Since ricin in carbon tetrachloride gives no
detectable odor, the comparison on the basis of a
30-second dosage is suggested as the fairest compari-
son. On the basis of weight of active agent employed,
rather than the weight of munit ion, ricin has a superi-
ority over phosgene of 40 to I from these data.4*
12.6.3 Ricin as a ar Gas
Ricin is an odorless powder capable of hieing dis-
persed as a particulate or dust cloud. The absence of
odor and the complexity of the consequent detection
problem in the field w ould render ricin more insidious
than any standard U. S. or British chemical war-
fare agent. Comparison with the German Trilons
(Chapter 9) would present a closer differentiation
problem. The physiological effects of ricin are de-
layed. Lung injury, similar in character to that pro-
duced by phosgene, can lead to deaths at from one
to several days after exposure. Ricin can be dispersed
in munitions not readily distinguishable from stand-
ard HE bombs.
For detection in the field attention has been given
to hemagglutination tests and to the use of ricin-
sensitized guinea pigs (Section 12.4). These methods
are intrinsically more difficult in practice than the
simple means for detecting such agents as mustard
gas or phosgene by odor or chemical tests. The IT. S.
and British gas masks, when well adjusted, give
complete protection against any dosage of ricin likely
to be produced in the field.37 The immunization of
troops against ricin and serum therapy present diffi-
culties, as outlined in Section 12.4.
From the. few tests on (he persistence of ricin in
(he field it has been concluded that the major part
of the agent is rapidly dissipated in the particulate
cloud. Only in the area immediately around the point
of burst was ground contamination sufficient to be
measurable. In tests in which sensitized guinea pigs
were allowed to run through the brush in this area a
possible hazard was detectable for about 3 days in
dry weather.32
As a result of the progress made during-World
War II on (he preparation and dispersion of ricin it
must be considered that hrall-out chemical warfare
it is possible that ricin could lx? employed in a prac-
tical role in chemical Supply and manu-
facture would place a ceiling on (he scale of use but,
would not prevent the accumulation of significant
quantities of this agent. It has been estimated that
the cost of production of dispersible ricin on a large
scale would be approximately $13 per pound (Sec-
tion 12.2).
In the course of the research during World War II
the work on ricin has served to advance the knowl-
edge on the general problem of particulate disper-
sion. In some respects ricin has served as a model
substance for work on the dispersion of agents of
similar chemical and physical properties in the re-
lated research in the field of bacteriological warfare.
SECRET Chapter 13
AROMATIC CARBAMATES
Arthur C. Cope
13.1 INTRODUCTION
Beginning in 1913 under the auspices of the Na-
tional Defense Research Committee [NDRC],
search for a superior nonvolatile toxic agent was un-
dertaken by several cooperating laboratories. Cri-
teria for the agent sought were extreme toxicity on
subcutaneous injection, rapid lethal action, ready
availability through practical synthesis or otherwise,
and sufficient stability for military use and storage.
A survey of the open literature 17 and information
currently available concerning the toxicity of chem-
ical warfare agents guided the search. Among the
more toxic classes of substances known, botulihus
toxin, other bacterial toxins, and potent plant toxal-
bumins (particularly ricin) were considered unsuit-
able because of slowness of their toxic action,
immunological characteristics, and in some cases in-
adequate sources of material for possible use on a
considerable scale.
The alkaloids physostigmine and aconitine were
high on the list of toxic substances.
I I
CHa CH,
Physostigmine
Aconitine is the more toxic of the two, but its com-
plete structure is unknown, and search for a toxic
agent among simpler related compounds is unprom-
ising because minor structural modification of the
toxic aconite alkaloids often destroys their toxicity.
On the contrary, many synthetic N-alkylcarbamates
related to physostigmine are highly toxic, and search
for a superior agent in this class appeared more
promising. For this reason, and after failure to ob-
tain highly toxic compounds in several other classes,
the investigation soon turned to a thoiough explora-
tion of the carbamates. Similar studies were con-
ducted at an earlier date in England by R. D. Ha-
worth and his associates, and in Canada by Leo
Marion and others. The following investigations of
carbamates reported in the open literature preceded
all the classified work.
After establishment of the structure of physostig-
mine. by Stedman and Barger,42 a number of syn-
thetic analogs were prepared by Stedman. Several
of his papers 13 47 describe the synthesis and mi-
otic properties of such analogs, but contain no toxi-
cological information. White and Stedman 4 ■ report
a detailed pharmacological study of miotine, the syn-
thetic miotic of choice from the group, including
toxicity data for this substance and three related
carbamates. Aeschlimann and Reinert49 report
toxicity and other pharmacological data for physo-
stigmine and 44 related synthetic carbamates, many
of which had been prepared earlier by Stedman.
Stevens and Bell tel ** also have investigated physo-
stigmine substitutes, and report chemical and toxic-
ity data for 27 related carbamates. The toxicity data
for such compounds are recorded in the open liter-
ature.484*’5’
As a result of the classified British and Canadian
work, TL 1071 (British code T-1708) was the lead-
ing candidate in the carbamate group. The com-
pounds TL 1217 and TL 1299 proved to be the
agents of choice on the basis of the NDRC work.
(If,
(iV)COXHCH, ([ V>COXIICH3 AoOONHCH,
u u u
NfCdLhCILI NfCdbhCIbCl X(CH3)jI
Tt, 1217 TL 1299 11, 1071
(T-1708)
Additional compounds that received more or less
detailed study were:
CII,
/ytcoxiiCHi if Ycoxiich,
NfCjHilsC'HjX X(CHj)jX
TL 1217; X - I TI. 1071; X -1
TL 1299; X = Cl TI, 1236; X = Cl
TL 1317; X = CILSO, TL 1185; X = ClIiSO.
TL 1186; X = USO,
(I \)('OXHCTb
chv
X(CH,).X
TL 1210; X - I
TI. 11.53, X = Cl
TL 1188; X = CUiSO,
SECRET SYNTHESIS
205
0OCON(CH,)j /Vk’OXHCH,
X(CHj)jN[I J
CII(CHi)t CH(CH,)t
TL r,90; X - t TL 1327; X = I
TL 1443; X » Cl TL 134.>; X - Cl
In the following pages work on the carbamates
which might be of some practical importance as toxic
agents is summarized. Investigations which led to
selection of leading candidates are mentioned briefly.
13.2 SYNTHESIS
13.2.1 m-Dielhylaininophenyl-N-methylear-
hamate metbiodide ('LL 1217)
The most practical preparation of TL 1217 is the
following sequence:
0)11 CH.NCO- jj Vx'ONHC.'lb ri4 l> i[ \kqxhch,
V - v
NtC.IL), N(C.ITi). MtMLf.CH.I
This process was operated successfully on a pilot-
plant scale.15 Approximately 360 lb of methyl isocya-
nate were prepared by reaction of methylamine and
phosgene in the vapor phase to give methyl ear-
Oamyl chloride, which was converted to methyl
isocyanate by treatment with pyridine in toluene.
The average yield was 81 per cent, w-Diethylamino-
phenol (a commercial dye intermediate) dissolved
in dry benzene was refluxed w ith an excess of methyl
isocyanate for several hours, m - Die thy lam i nophenyl-
N-methylcarbamate was isolated in yields of over
80 per cent by evaporating the solvent under reduced
pressure, filtering, washing, and drying. TL 1217 was
prepared by reaction of m-diethylaminophenyl-N-
methylcarbamatc with methyl iodide in acetone un-
der reflux. After addition of ethyl acetate, the prod-
uct was recovered by filtration in yields of 79 to
86 per cent. The product so obtained was of high
purity, as verified by elementary analyses, use of a
special analytical procedure involving hydrolysis and
determination of carbon dioxide and methylamine,13
and toxicity tests.
Essentially this same procedure had been used
earlier for the preparat ion of TL 1217 on a laboratory
scale.1-25 This compound is among the group de-
scrihed by Aeschlimann and Reinert49 and by R. D.
Haworth.23 Prior to development of a practical syn-
thesis of methyl isocyanate, m-diethylaminophonyl-
N-methylcarbamate was prepared on a large labora-
tory scale in yields of 71-86 per cent by reaction of
m-diet In land nophenol with phosgene in the presence
of diethylaniliuo, followed by reaction with methyl-
amine.23 u This procedure was developed from a
similar method used by Marion.29 32
is.2.2 m-Dielliylaininopbenvl-N-mcthyl-
carbaiuatc inethochloridc (TL 1299)
The most practical preparation of TL 1299 is the,
following;
CH.xro l^jOCONHt'H,
N(C,IL), X(CiH»)jCHjC1 N(C|H»)jCHjC1
This process was operated successfully or a pilot
plant scale.15 Redistilled w-dielhylaminophenol was
treated w ith an excess of methyl chloride in an auto-
clave at 100 C. After cooling and evaporation of the
excess methyl chloride, the product was purified and
isolated in 77 per cent yield by grinding, washing,
and drying. Yields in this step on a laboratory scale
were 95 per cent.,sm-DiethyIaminophenol methochlo-
ride was converted to TL 1299 by reaction with
methyl isocyanate in dimothylforraamidc as a solvent
and a mixture of triethylamine and glacial acetic acid
as a catalyst. Yields on a pilot plant scale were 91 per
cent .15 This process was developed to a high state of
perfection in an intensive laboratory investigation,18
in which yields of 94-97 per cent and 90-92 per cent
in the two steps were obtained, or 86-5)0 per cent
overall. A useful laboratory synthesis of methyl Iso-
cyanate from methylamine and phosgene also was
developed in this work,13 and was used until it was
superseded by the pilot plant process 15 for this essen-
tial intermediate.
Prior to development of the above process, TL
1299 was prepared by a different procedure. m-Di-
ethvlaminophcnyl-N-met hylcarbamate was | > repared
first from w-diethylaminophenol by the phosgene-
methylamine procedure, with diethylaniline as the
acid acceptor (yield 78 per cent).2 314 This product
was converted to (he methosulfate salt (TL 1317)
by reaction with methyl sulfate (yield 75-79 per
cent), and TL 1317 was converted to TL 1299
through reaction with anhydrous calcium chloride in
methanol containing hydrogen chloride (yield 74 per
cent).2 3 Earlier TL 1299 was prepared in high yields
from TL 1217 and silver chloride.2 3 5 14
13.2.3 (2-Metliyl-5-diinethvlaminopbenyl)-
N-metbvIcarbamatc methiodide (TL 1071)
TL 1071 (British code T-1708) was commonly
called the “Haworth compound” during the NDHC
SECRET 206
AROMATIC CARR \MATES
investigations, since it was the leading candidate
from the British work. Details of Haworth’s method
of preparation could not be obtained, but Canadian
reports on the synthesis ->6-34 were available and served
as a basis for further developments.
The intermediate 2-methyl-5-dimcthy land nophe-
nol was obtained from the National Aniline Division
of the Allied Chemical and Dye Corporation, where
it was prepared from p-toluidine by methylation,
sulfonation, and alkaline fusion:
(ML (ML C1L (ML
6-6-6"" - 6"
XHi X(ClLh X((MLh X((MLh
2-Methyl-5-dimethylaminophenol was converted into
the N-methylcarbamate by treatment with phosgene
in the presence of diethylaniline, followed by methyl-
amine (yield 75 80 per cent).14 With methyl isocya-
nate available,14 this intermediate could lx? used in
preparation of the N-methylcarbamate, which has
Ix'en prepared in that manner in 85 per cent yield on
a small scale.4 TL 1071 was prepared from the N-
methylcarbamate and methyl iodide in acetone in
95 per cent yield,14 In a Canadian pilot plant opera-
tion, 39 lb of TL 1071 were pro pans! from 2-methyI-
5-dimethylaminophenol by this process, with an
overall yield of 39 per cent.33
Other quaternary salts differing from TL 1071 only
in the anion were prepared. Among these were the
methosulfate, TL 1185,14 which was hydrolyzed
slowly with aqueous hydrochloric acid or water to
the acid sulfate, TL 1186.14 The latter on treatment
with calcium chloride yielded the methochloride,
TL 1236. The preferred procedure for preparing this
compound was to heat the crude methosulfate with
an alcoholic solution of calcium chloride for 20 hours.
Overall yields from the N-methylcarbamate were
77-87 per cent." Treatment of the N-methylcarba-
male with methyl chloride also yielded TL I236.5
13 2.1 (1-Met In 1-3-diiuethy land nophenyl )-
N-me th vicar ha male methiodide (TL 1216)
TL 1216 was prepared during the Canadian work,27
and became known during the NDRC investigations
as the Haworth isomer. It was synthesized in Divi-
sion 9. NDRC. from 4-methyl-3-dimethylandnophe-
nol, which was prepared by the National Aniline
Division of the Allied Chemical and Dye Corpora-
tion from o-toluidine:
xil N((’iit)> n(cil)* x(cn,)s
Bolh the phosgenc-mct hylaminc procedure "27 ami
methyl isocyanate 4 were used in preparing (he
N-mothylearbamate. The methiodide, TL 1216, was
preparer! from the N-mothylcarhamateA 14 Other
salts prepared were the methoehlorido * ('IT, 1453)
and the methosulfate (TL 1 188). Synthetic metliods
employer! paralleled those described for TL 1071.
13.2.5 (3*lsoj>ropvl-l-dimethylaminophenvl)«
\, N-dimelhvlearhamate inetliir>t)irie
(TL 599)
TL 599 is the most toxic of the carbamates de-
scribed by Stevens and Beutel.53 Its preparation on
any scale is hindered by lack of a practical source1 or
synthesis for w-isopropylphenol, the essential start-
ing material. An investigation of.eight routes to (Ids
compound was made,'* of which the most satisfactory
started with benzoic acid and continued through
methyl m-hydroxyl>enzoate and m-hydroxyphenyl-
dimethylcarbinol, by way of the Grignard reagent.
The remaining steps in the synthesis of TL 599 were
the following:'*
A„„ Aon _ Aon _
U °AJ«V
CUfClbh CUCCILh (t l(Cli,)2
AOH (Cl.hNCOC, A<)fox«;i..).
—-—>
CIKClbh CtKClLh
t| \)c;on((ti3)5
I(CH:Axily
Cll(CILh
Compounds in the corresponding N-monomothyl-
carbamate series also were prepared (TL 1327, TL
1345).91*
13.2.6 Synthesis of Other Aromatic
Carbamates for Toxicity Tests
In addition to the carbamates described in the
preceding sections which were the subject of rela-
tively intensive laboratory or pilot plant investiga-
tions, many similar compounds were prepared on a
small scale for toxicity tests, in the search for the
most toxic and readily synthesized agent in the
SECRET TOXICOLOGY
207
group. The following inferences contain the results
of such investigations, and an indication of the
classes of compounds studied where that information
can l>e stated concisely; otherwise they are classified
as miscellaneous.
Ref.
Classes of Carbamates Described No.
M iseellaneous; sulfur analogs 1
Derivatives of polyhydric plienols -1
Derivatives of 3-dielhylaminophenot, 3-dimethvl-
amiiiophenol, 2-methyl-5-diinethylaminophcnol and
2-methyl-a-diethylaniinophcnol 5
Derivatives of 4-dimethylaminotbymol and 4-dimethyl-
ammoeurvacrol 6
Hoinologs and analogs of Doryl (aliphatic carbamates) 7
Derivatives of p-aminophenol, 4-methyI-3-aminophe-
nol, 3-meihyl-l-anunophcnol, 2-tnethyl-5-aminophe-
nol . . S
Derivatives of 5,6,7,8-tetrahydronaphthol-l; 3-iso-.
pfopyl-4-aminophenol; miscellaneous ...... 9
Derivatives of 3,.5-dimethyl-t-aminophenol 10
Derivatives of 3-alkyl-4-aminophenols II
Tb 1299, the corresponding X,X-dimelhylcarbamate
methiodide (TL 1238) and methochloride (TL 1422) 13
Derivatives of 2-melhyl-5-diiuelhylaminophenol, 4-
metbyl-3-dimethylaminophenol, 2-tnelhyl-5-die(hyl-
aminophenol, m-dielhylaminopbenol 14
Derivatives of 3-isopropyl-4-aminophem>l, 2,6-diiso-
propyl-4-aminophenol, 2-isopropyl-5-aminophenol,
4-lsopn»pyl-3-aminophenol, 4-isopropyl-2-aminophe-
nol; arsenic analogs — . 19
Miscellaneous; toxicity, data only on compounds pre-
pared by R. D. Haworth .... . . ~. ..... 23
Miscellaneous 24
Derivatives of m-dimelhylaminoplienol 25
Derivatives of 2-melhyl-5-dimethylaminophenol . . . 26
Derivatives of 4-metKyl-3-dimcthylaminophenol , . 27
Denvativc-s of m-diethylaminophenol ....... 2tl
Derivatives of 2,4-dimet hyl-5-dimethytaminophcnol 31
Miscellaneous 34
Miscellaneous 43
Miscellaneous : 44
Miscellaneous . . . .~ 45
Miscellaneous 46
Miscellaneous 47
Miscellaneous 48
Miscellaneous 49
Miscellaneous 50
Miscellaneous 51
Miscellaneous . 52
Miscellaneous 53
13.3 STABILITY
The toxic aromatic carbamates of possible practi-
cal importance are reasonably stable at 65 C, show-
ing little decomposition after 2 months storage.16
The two labile groups in such compounds are the
carbamate and quaternary salt linkages. The carbam-
ate group is subject to thermal decomposition to
methyl isocyanate and the corresponding phenol,
and to hydrolysis to the phenol, mothylamine, and
carbon dioxide, or related products. The quaternary
salt groups are subject to decom positiou at elevated
temix’ratmcs to an alkyl halide and the correspond-
ing tertiary amine. If the carbamates are kept dry,
they have good thermal stability. The same pre-
eaution protects them from hydrolysis. Hydrolysis is
very rapid in alkaline solutions, and slow at an acid
pH. As a precaution to insure stability, the carbam-
ates may be crystallized from solvents containing
hydrogen chloride. Alternatively, acidic stabilizers
such as sodium acid sulfate or hydrazine dihydro-
chloride may he added.
A number pf the more toxic carbamates were ex-
amined for relative stability 16 at a time when it ap-
peared that stability might lie a decisive factor in
choice of a superior agent. The following conclusion
was reached concerning thermal stability: variation
in the anion of the quaternary ammonium salt re-
sults in (he following order of decreasing stability:
methosulfate > methiodide > methochloride. Com-
paring stabilities toward hydrolysis, two N,N-di-
methylcarbamates were much more stable than two
N-mcthylcarbamates (TL 1071 and Tb 1217), which
in turn were more stable than two N-arylcarbam-
ates. Ultimately the two agents chosen as superior on
the basis of toxicity and ease of manufacture (TL
1217 and TL 1299) were determined to be sufficiently
stable for any anticipated use.
One factor with an important bearing on stability
is the hygroscopic character of some of the carbam-
ates. TL 1299 is quite hygroscopic in humid weather.*
TL 1217 is not, and largely for this reason became
the agent of choice. TL 1299 could be handled satis-
factorily if it were needed on a large scale by con-
trolling the humidity of (he rooms in which it would
be crystallized, dried, and packaged. Whereas this
could be done readily on a full manufacturing scale,
on (he large laboratory and pilot plant scale it was
much simpler to employ the nonhygroscopic methi-
odide, TL 1217.
13.1 TOXICOLOGY
A report has been prepared 12 which summarizes
much of the toxicological work done in this country,
in Britain, and in Canada on the aromatic carbam-
ates. Tables from this report, reprinted as Table 2
of this chapter give toxicity data for the 319 aro-
matic carbamates and closely related compounds
known to have been tested.
Aromatic carbamates prepared as part of the
NDRC program were submitted to the University
of Chicago Toxicity Laboratory for testing. There
SECRET 208
AROMATIC C ARU VACATES
they received a TL (Toxicity Laboratory) number,
and were tested for subcutaneous toxicity to mice.
Two to five mice were injected subcutaneously with
doses of 80, 10. 20, 10, 5, and 1 mg kg of body
weight, at dilutions such that each mouse received
approximately I per cent of its body weight in a
suitable nontoxic solvent (usually water). Any com-
pound that killed at 1.0 mg kg was screened further
and LDio determinations were made for all those
killing at less than 0.5 mg kg. The data obtained are
listed in Table 2, together with similar toxicity data
obtained elsewhere for other aromatic carbamates.
A number of factors influencing toxicity determina-
tions made by injection were studied carefully for
the more important aromatic carbamates. One of the
most important was the animal species used in test-
ing. The leading candidates were tested in several
animal species, since the object of the search was to
select an agent toxic for all species. TL 1217 proved
to be very toxic for all species in which it was tested.
TL 1345 is the most toxic compound tested in mice,
but as Table 1 shows,12 it presents no marked superi-
intraperitoneal (In the single comparison available
for rats the in! ra peritoneal route was the more
effective.)
3. Carbamates are relatively ineffective when ad-
ministered by stomach 1ul>e. 25 to 500 times as much
material being required to kill as by injection.
The carbamates are toxic when administered by
inhalation as aerosols,'- but do not show the ex-
traordinary toxicity in comparison with standard
chemical warfare agents which characterizes them
when toxicitics determined by injection are com-
pared.
The aromatic carbamates are “quick-kill” agents
capable of producing severe parasympathomimetic
effects terminating in death. Death occurs rapidly,
for example, in 5 to 20 minutes after subcutaneous
injection in dogs. The symptoms produced are similar
in all species which have been examined. They con-
sist of salivation, evacuation of bowels and bladder,
restlessness and incoordination, and fibrillary muscu-
lar movements. Respiratory movements are quick-
ened and labored. Coma is accompanied or preceded
by convulsive movements. Respiration appears to
cease first, the heart lieating, usually irregularly, for
some moments after respiration has failed. Muscular
twitching persists for some time after failure of res-
piration and cardiac activity.
The aromatic carbamates are powerful cholin-
esterase poisons, and produce marked changes in
the blood. Because of medical and toxicological in-
terest in them, their physiological mechanism of
action has received considerable study. Most of this
work may be located through certain leading infer-
ences,20 ■2U hd e Atropine or atropine and pento-
barbital administered intravenously have been rec-
ommended as antidotes for the carbamates.36 41 Anti-
dotes can lx* demonstrated to be useful in animals,
but must be administered quickly (at the onset of
symptoms) because of the very rapid toxic action of
the carbamates.
For references to toxicity assays on the carbamates
in addition to the summary previously mentioned 12
see the Bibliography.2211 *’<’ ‘12J 2S-3II S5 S7
I3,r. RELATIONSHIP BETWEEN CHEMI-
CAL STRUCTURE AND TOXICITY
Relationships existing between chemical structure
of the aromatic carbamates and their toxicity have
been pointed out in some detail.12 The following prin-
cipal conclusions can be drawn from the available
Table 1.
1345 for v
Subcutaneous toxicitics of TL 1217 and TL
a rious species.
Sjiecies
LT)a„ (mg kg)
(| \)COXHCHj (|^OCOXIICH,
V a(C",wv
CIKCII.),
TL 1217 TL 1345
Mouse
0.129 0.047
Rat
ea. 0.400 0.103
Guinea pig
0.097 ca. 0.050
Rabbit
ca, 0.150 ca. 0.075
Cat
ca. 0.075 ca. 0.100
Dog
ca. 0.075 ca. 0.100
Monkey
ca. 0.200 ca. 0.150
ority over TL 1217 when other species are con-
sidered.
Other factors considered in precise toxicity de-
terminations were the concentration of the solution
injected; the strain, sex, body weight, and age of the
mice used in LDh0 determinations; and the effect of
the temperature of the environment of the assay
animals. A number of compounds were tested by
various routes of administration, and the following
conclusions were reached.12
1. The carbamates tested were about twice as
toxic intravenously as by any other route.
2. Subcutaneous injection was more effective t han
SECRET CHEMICAL STRUCTURE V ND TOXICITY
209
toxicity data (figures cited refer to subcutaneous
toxicity in mice).
1. The most toxic compounds contain both a
carbamate and a quaternary salt group.
2. The carbamate group is more intimately con-
nected with toxicity than is the quaternary salt
group. This conclusion follows from several lines of
evidence:
a. The quaternary ammonium group can lie re-
placed with sulfonium or arsonium without change
in order of magnitude of toxicity. For example:
-
X(C2Hi)jCH,I \s(C4I).CHl
Tl, 1217 TL 1306 TL 1501
- 0.120 mg kg LIh* =* 0.37 mg kg LIhu =» 0,5 mg/kg
h. The 6/s-N,N-dimethylcarhaniate of catechol is
highly toxic even though it contains no basic group;
int roduction of a quaternary salt group in this com-
pound results in diminished toxicity.
OXX)X(CHi)j jj \x'ON(CHj),
X’ON(CH,), I(CH,),nI!Jo( 0N(CH,)j
TL 1015 TL 1165
Llha = 1.4 mg/kg Toxic dose > 10 mg/kg
c. Quaternary salts derived from amiuophcriols
are not very toxic, but the N-methylcarbamates de-
rived from some of them am highly toxic.
jj^jOCONIICH,
NfC-HshCHjCl N(Cyis),CII,Cl
TL 1300 TL 1200
Toxic dose about 40 mg/kg Lhut =• 0.09 mg/kg
d. Structural changes in the carbamate group in
related series of compounds may produce enormous
changes in their toxicity.
0 OCONHCTI, /\sCOXHCI1j
c”V
N(CIIj),I X(CII,)jI
TL 1216 TL 1230
LOfco = 0.17 mg, kg Toxic dose > 80 mg/kg
0DCOX HCITj (| \xX)N(CH,)»
V
X(CiHj)jCIIjI X(CjHi)jCH,T
TI- 1217 TL 1238
/-/)» * 0.120 mg/kg LDu> » 0.175 mg/kg
CH,CH,
0OCON CHj HC,Hj
\hjCH, {J
N(C,II,fcCHJ N(C.II4)jCH,,I
TL 13(6 TI. 1-181
Toxic liosc about 1 nut k(t Toxic dose about 5 mg kg
IK'(,11 HCdbOCH,-,,
N(C’:Ilj)iCUjl X(C.H.U’UI
TI- 1433 TI- 1412
Toxic dose > 8(1 mg kg Toxic dose > SO mg kg
o. Changes in the quaternary salt group in a series
iu whieh the N-methylcarbamate group is kept con-
stant produce smaller changes in toxicity.
OCOXHCIIj OCOXIICH,
(S o
N^xtdhhi
c*n.
TI, 1178 TI, 1323
LDw = 0.270 mg/kg /,/>» = 0.135 mg/kg
OCOXIICH, OCOXHCIIj
f) (S
(C,Hi)j CIJjCHfzEIIi
tl. 1217 TL 1435
LD& — 0.129 mg/kg LDm =» 0.102 mg/kg
OCOXIICH, OCOXIICH,
- (QLxCjHOjI
(CHjIjCH,
TI- 1434 TL 1259
LDw - 0.10 mg/kg LDx = 0.23 mg/kg
OCOXHCIIj
A -
VfIl!l
(CjHg),
TI, 1324
/.Dm = 0.48 mg/kg
II. In general, the N-methylearhamates are mom
toxic than corresponding N,N-dimethyIcarhamates.
Of 20 such pairs of compounds tested, (he mono-
met hy lea rhamates were more toxic in I t eases (for
some pairs they were 10 to 40 times as toxic); in (ho
other G cases they were approximately equal. No
other substitution on the carbamate nitrogen which
was investigated led to compounds as toxic as the
hi-methyl and N.N-dimcfhyl derivatives.
SECRET 210
VROM \TIC CAR HAM AXES
4. With few exceptions, the most toxic compounds
were those in which the X-methylearltamato and
quaternary salt groups were in the meta orienta-
tion.
0OCOX H( ’ll, /VxX)N IICH,
X(CH»)jI {J
\(CH,M
Toxic dose 130 mu kg . TL 1178
Llho ■** 0.27 mg/kg
OK'OXHCTb
TL 1007
Toxic dose about 20 me kg
5. Methyl substitution in the nucleus ortho or
para to the carbamate produces no great change in
the toxicity of m-quatemary compounds, and may
result in slight ly more toxic substances. Similar sub-
stitution by bigber alkyl groups leads to less toxic
compounds.
Oil,
OGC’DNHCHj a Xk'OXHCH, fi %HX)XncH,
V CI,V
N(C1I,U XfCHhl N(CIE)J
TL 1178 TL 107! TL 1216
LDw = 0.27 mg kg £/>» «■ 0.111 mg kg LDm = 0.17 mg kg
ti. The series with an alkyl substituent meta ami
tlie quaternary salt para to the carbamate group
contains some extremely toxic compounds. In the
most toxic homologs of this type 11 the alkyl group
is isopropyl.
0OCONIICIh j| \ K OX(CIb),
UClb)A\J
CH(('I!,), 011(011,),
TI, 1327 Tl. 309
/.Dm = 0.0H7 mg/kc LOao =• 0.083 mg kg
7. The toxicity of aromatic carbamates'substi-
tuted by quaternary salt groups resides in the cation.
Of the various stilts, the chlorides have l>een found to
Ik? somewhat more toxic than would be calculated on
a molecular weight basis. Other salts with the same
cation have toxieifies proportional to their molecular
weights.
Table 2. Toxicities of aromatic carbamates and related compounds.
The following tables” contain the toxicity data available as of March 1945 for aromatic carbamates and closely related
sulstances (319 in all). The tables are subdivided into IS structural classes, as follows;
I Benzene compounds with one carbamate group, and
no quaternary ammonium group.
IT Benzene compounds with two carbamate groups and
no other groups.
III Benzene compounds with two carbamate groujis and
other groups.
IV Benzene compounds wit h 1 hree carbamate groups and
no other group.
V Benzene compounds with one carbamate group and
one quaternary ammonium group in the ortho
position.
VI Benzene compounds with one carbamate group and
one quaternary ammonium group in the ortho
position and alkyl groups.
VII Benzene compounds with one carbamate group and
one quaternary ammonium group in the meta
position.
VIII Benzene compounds with one carbamate group and
one quaternary ammonium group in the meta
position and other substituents.
IX Benzene compounds with one carbamate group amt
one quaternary ammonium group in the para posi-
tion (including thiocarbamalcs).
X Benzene comjmunds with one carbamate group and
one quaternary ammonium group in the para posi-
tion and other substituents.
XI Benzene compounds with one carbamate group and
an alkyl side chain having a quaternary ammo-
nium group.
XII Benzene compounds with one carbamate group and
two quaternary ammonium groups. -
XIII Benzene compounds with one carbamate group and
one sulfonium or arsonium group.
XIV Carbamates of naphthalene derivatives.
XV Carbamates of quinoline and isoquinolinc deriva-
tives.
XVT Carbamates of aliphatic alcohol derivatives,
XVII Miscellaneous carbamates.
XVIII Carbamides and carbazates.
The tables represent a revision of a similar review issued on
June 15, 1914,19 and follow the system of classification used in
the earlier summary. Entries in the column headed “Code”
have the significance noted in the Glossary.
In the column headed “Route and Solvent” the following
abbreviations are used:
Sc.W. = subcutaneous injections in water.
Se.P. = subcutaneous injections in propylene glycol.
Sc.O. = subcutaneous injections in olive oil.
Sc.M. - subcutaneous injections in mineral oil.
Sc.Imp. — subcutaneous implantation of dry solid.
Iv.W. = intravenous injection in water.
Iin.Iinp. = intramuscular implantation of dry solid.
Ip.W. = intraperitoncal injection in water.
Oral W. = administered by stomach tul>e, in water.
pll-t indicates that this acidity was achieved with Mcll-
vaine’s buffer.
Whenever the room temperature during the determination
was recorded, it was listed immediately following the /.//,„
figure.
SECRET
1. Benzene compounds with
one carbamate group, and no
quaternary
ammonium group.
Route
—
and
Dose
Code
Xamc
Structure
solvent
Species
mg kg
Kffect
All-1
Carhamie acid, X-methvl-
OCOXI1C1L
Iv.
Mice
>50
U\«
phenyl ester
0
TL-1113
Carhamie acid, X,X-dimcthyl-
V
OCOX(CHi)
Sc.M.
Mice
80
0/2
phenyl ester
/\
40
0/2
0 -
20
0/2
TL 1218
('arhamthiolicacid, X-methyl-
X/
SCOXTICHj
Sc. 1’.
Mice
SO
2/2
p-lolyl ester
/\
40
0 2
o
20
0 2
V
Cl I,
TL-997
Carhamie acid, X-methyl-2-
OCOXHCI1,
Sc.Tmp.
Mice
80
0/2
All 2
nitrophenyl ester
Qxo,
Tv.
Mice
:«
/./),*
TI, 948
Carhamie !acid, X-methyl-3-
V
OCOXHC1L
Se.lmp.
Mice
80
0 2
nitrophenyl ester
/\
to
0/2
20
0/2
VNO*
TI.-947
Carhamie acid, X-methvl-4-
OCOXHCIL
Se.Tmp.
Mice
80
0/2
nitrophenyl ester
/\
40
0/2
()
20
0/2
\/
xo*
-
T 1,-980
Carhamie acid, X-methvl-2-
OCOXHCIL
Sc.Tmp.
Mice
80
0/2
hydroxyphcnyl ester
A
40
0/2
_
O’-
20
0/2
TL 1010
Carhamie acid, X,X-di methyl-
\/
OC'OX(ClL)i
Sc.W.
Mice
80
0/2
2-hydmxyphenyl ester
A
40
0/2
0m
20
0/2
TL-979
Carhamie acid, X,X-diethvl-
\/
(X 'OX(CdL),
Sc.Tmp.
Mice
80
0/2
2-hydroxy phenyl ester
A
-
40
0/2
O’"
20
0/2
TL 1101
Carhamie acid, X,X-dimelhyl-
V
OC()X(CH3).
Sc.O.
Mice
80
0/2
2-allyloxvphenvl ester
A
40
0/2
r jOCHjCl I CIT.
20
0/2
TL-1110
Carhamie acid, X,X*-dimethyl-
V
OCOX(CH.h
Sc.M.
Mice
SO
— 0/2
1-all vl-2-met la ixyphenyl
A
40
0/2
ester
o
—
20
0/2
\/
C1I>CH—CUj
CHEMICAL STRUCT THE VM> TOXICITY
211
SECRET 212
AROMVTIC CARBAMATES
Tabus 2, Section I (Continual)
Code
Name
Structure
Route
and
solvent
Species
Dose
mg/kg
KfTeet
TL-1111
Carbamic acid, X.X-dimethvl-
OCOX(CHj)j
Sc.M.
Mice
80
0/2
2-mcthoxy-4-propylphenyl
A
40
0/2
ester
[ jOCIb
20
0/2
V
CHjCIljCHi
TL-lllG
Carbamic acid, X.X-dimethvl-
OCOX(CIL).
Sc.M
Mice
SO
0 2
4-« llyl-2-mc t hoxy-5-n i fro-
A
40
0 2
phenyl ester
f jOCH,
20
0/2
o,x
V
ClijCH—CIIj
IT. Benzene compounds with two carbamate groups
and no other groups.
TL-lOlo
Benzene, 1,2-hia{ methyl-
OCOXHC1L
Sc. W.
Mice
40
2/2
carbamyloxy)-
20
2/2
—
[ jOCOXTICII,
10
0/2
5
0/2
TIi-978
Benzene, 1,2-hts(dimcthyl-
OCON(CHj)j
Sc.P.
Mice
1.4
LD*
«.
carbamyloxy)-
A
-- 1
f jOCOXCCIbh
TL-1118
Benzene, 1,2d>is{diel hylear-
V
OCOX(CdI.,b
Sc.P
Mice
SO
0/2
bamyloxy)-
A
_ 40
0/2
1
r JocoxicdCb
20
0/2
TL-1119
Benzene, l,2-h/.s{X-penta-
V
OCOXCdl,,,
Sc.M
Mice
SO
0/2
met h vlenecarbam vloxv )-
A
40
0/2
J
r pcoxciii.o
20
0/2
TL-1112
Benzene, 1,3-6is(dimct hylcar-
X/
OCOXTCIIj),
Sc.W.
Mice
so
0/2
bamyloxy)-
A
—
40
0/2
1
20
0/2
1
1 l0C0X(CHj)j
TL-1114
Benzene, 1,4-6£s(dimet hylcar-
OCON(CHi)j
Sc.P
Mice
SO
0/2
bamyloxy)-
A
40
0/2
o
20
0/2
V
OCOX(CTL)i
TL 1348
Benzene, 1,4-6/s(met hylcar-
OCOXHCIlj
Sc.P.
Mice
SO
0/2
bamyloxy)-
A
40
0/2
o
20
0/2
v
OCONHCH,
—
III. Benzene compiunds
with two carbamate groups and other groups.
TL 1117
Benzaldehydc, 3,4-Wa(dimethyl-
OCOX(CHs)j
Sc.P.
Mice -
so
0/2
carbamyloxy)-
A
40
0/2
r |ocox(cHi)i
20
0/2
V
CHO
SECRET 213
CHEMICAL STRICTURE AND TOXICITY
Code
Xame
Structure
Route
and
solvent
Species
Dose
mg
Kffcct
TL-1157
Benzyl alcohol, 3,4-6»s(di-
OCOX(CIIa)I
Sc. W.
Mice
10
1/2
met 11 ylcarl >a myloxy)-
/\
5
0/2
[ jocoxccn.h
i
0/2
X/
CHjOH
TL 1160
Dimcthvlamine, X-[3,4-5is(di-
OCOX(CHj)i
Sc.W.
Mice
10
2/2
me(hylcarbamyloxy)benzyl]
/\
5
2/2
—
hydrochloride
f jOCOX(CIIi),
i
0/2
A J
0.5
0/2
—
9 CII2X(CHj)j-HC1
TL-981
Benzene, 1,2-bis( dimethyl-
OCOX(CTI3)t
Sc.lmp.
Mice
80
0 2
carbarn yloxv)-4-ni tro-
A
Sc.O.
SO
0/2
r pcoxxciuh
40
0/2
—
NO,
—-
TL-I017
Benzene, 1,2-hi«( d i me t hy 1 ca r-
OCOX(CIlj);
Sc. N/10
M ice
to
2/2
1 mmy loxy )-1-amino-
/\
Ac.
20 _ -
2/2
—
f pcox(cn5)s
10
2/2
I J
5
0/2
XH,
TL 1155
Benzene, 4-(dirne(hylamino)-
OCOX{CH,)i
Sc.W.
Mice
10
0/2
1,2-Ws{diincthylcarbamyl-
A
5
0/2
oxy)-, methiodide
f ]()COX(CHj)j
1
0/2
{J
0.5
0/2
-
X(CH,),I
TL-1150
Benzene, 1,2-W«(dimet hylcar-
OCOX(CIIj)i
Sc.O.
Mice
10
0/2
bamyloxy)-3-allyl-
/\OCOX(CH,),
5
1
0/2
0/2
X^JciIjCH—CHj
0.5
0/2
TL-1158
Benzene, 1,2-6/.<.{ dimethyl car-
OCOX(CIL)j
Sc.O.
Mice
10
0/2
1 >a my 1< »xy )-3-pn tpy 1-
|/\oc 'OX (CII a) j
5
1
0/2
0/2
I JtTLCTLCH,
0.5
0/2
TL-1102
Benzene, 1,2-/>w(dimetliylcar-
OCOX(GH,)i
Sc.O.
Mice
10
0/2
bamyloxy)-4-allyl-
A
5
0/2
i
0/2
-
Kj
0.5
0/2
CHjCH=CH, __
TL-1156
Benzene, 1,2-his(dimel hylcar-
OCON(CH,),
Sc.M.
Mice
10
0/2
bamyloxy)-4-propyl-
A
5
0/2
f |OCOX(CH,)t
i
0/2
f.
0.5
0/2
CHtCHiCHi
TL-1086
Benzene, 1,3-b/»(X-met hylcar-
OCOXIICH,
Sc.P.
Mice
40
0/2
bamyloxy )-2-ni t ro-
Ano,
20
0/2
I Jocoxiich,
—
Table 2, Section III {Continued)
SECRET 214
VROM A TIC CAR RAM AXES
Tahle 2, Section III (Continued)
Route
and
Dose
Code
Name
Structure
solvent
Species
mg kg
Effect
TL-1120
Benzene, 1,3-b/x( X-met hylear-
OCOXIICIl,
Sc.W.
M ice
SO
0/2
bainylo\y)-2-amino hydro-
chloride
/\xh2iici
10
20
0,2
0/2
1
sJdcoxiicii,
ti, 1310
Renzene, 1,4 -bis( met hylearba-
OCOXIICIl,
Sc.P.
Mice
. SO
0 2
mvloxy )-2,G-dimet hyl-
A
40
0 2
CHI
0".
OCOXIICIl,
- '
20
0 2
TI, 1350
Ben zone, 1,4-his( met hylcarba-
myloxy )-2-isopropy 1-5-
OCOXIICIl,
A
Sc.P.
Mice
so
10
2 2
12
rnctlivl-
( VlKCH.I,
20
1 2
nij
I J
10
0 2
v
5
0/2
• .
Ot ONIRIC
IV. Benzene compounds with three carbamate
groups and no other group.
TI 1115
Renzene, 1,2,3-//ix( dimethyl-
OCOXfCHOs
Sc.W.
Mice
40
2/2
carbamvloxv)-
A
20
2/2
■ - 1
f KX'OX(CTI,)s
1 k)cox(cii3),
10
it
0/2
0/2
V.
Benzene compounds with one carbamate group and one quaternary ammonium group in the ortho position.
TL-903
Carbamie acid, N-mcthyl-2-
aminophcnyl ester hydro-
OCOXIICIl,
A
Sc.W.
Mice
so
40
0/2
0/2
chloride
[ jXTIj-IlCl
--
20
0/2
T-{ ?)
Carbamie acid, N-methyl-2-
dimethylaminoplicnyl ester
\/
OCOXIICIl,
A
Sc.
Mice
430
ldm
methiodidc
r |X(cn,)ji
VI. Benzene coin|)oimds with one carbamate group
i ami one quaternary
ammonium
group in the ortho position ami alkyl groujis.
TL-1488
Carbamie acid, X-methyl-2-
di met hyla mi no-4-isopropyl-
OCOXIICIl,
A
Sc.W.
Mice
SO
40
0/2
0/2
phenyl ester methiodidc
r ]x(cu,),i
20
0/2
V
CIKC1I,),
SR-13
Carbamie acid, X.X-dimethyl-
2-dimethvlamino-4-methvl-
OCOX(CHj)-
A
Sc.
Mice
Approx. 200
LD,„
phenyl ester hydrochloride
f
CH,
SECRET CIIE.MKUL STRUCTURE AND TOXICITY
215
Tabi.k 2, Section VI (Coulin
iied)
Route
and
Dose
Code
Name
Structure
solvent
Species
n>R kf;
K fleet
SB-14
Carbamic acid, N,N-dimethyl-
OCOX(CH,)j
Sc.
Mice
2.0
I.D,„
2-dimet hvlamino-4-methyl-
/\
phenyl osier mclhiodidc
| |N(CHj)jl
V
CH,
SB-15
.Carbamicacid, \,\-dimethyl-
OC()N(CIIi)t
8c.
Mice
27
2-dimet hvlamino-4-ethyl-
A
_
phenyl ester hydrochloride
r jN(CHj)jll(T
"'“I
V
c3h.
SB-10
('arhamic acid, X,X-dimethyl-
OCON(CHj)j
Sc.
Mice
1.25
LDia
2-di met hvlamino-4-ethyl-
A
phenyl ester mclhiodidc
r |N(cn»)»i
V
c,h5
-
SB-17
Carhalnic acid, N,N-dimethyl-
OCX)N(CIIi)*
Sc,
Mice
>400
UK,
_
2-dimet h vlamino-4-isopro-
A
■ _ - ”
-
pylphenyl ester hydrochlo-
1 |N(Cii;)-nci
“ ride
IJ
—
•
CH(CH»)*
SB-18
Carhamic, acid, N,N-dimcthyl-
()CON(CIIi)j
8c.
Mice
4.8
LDU
2-dimet hylamino-4-isopro-
A
pylphenyl ester nicthiodide
r ]N(cn,).i
-—
—
—
■- — ■
V
ciuciia).
SB-10
Carhamic acid, X,X-dimcthyl-
OCOX(CIIj),
8c.
Mice
> 500
UK„
2-dimethylamino4-/frl hutyl-
A
phenyl ester hydrochloride
f jN(CHi)jliCI
--
V
C(CHj)i
SB 20
Carbamic acid, N,\-diinethyl-
OCOX(CH,h
8c.
Mice
13.5
LDn,
2-dimet hylamino-4-fcr< butyl-
A
ptienyl ester methiodide
f |X(CHj)jI
V
C(CHj)j
SB-21
Carhamic acid, N.N-dimethyl-
OCOX(CHj)j
Sc.
Mice
> 500
LDM
2-dimet hylnmino-4-fcrl
A
• .
amylphenyl fitter hydro-
f ]Nr(cir3h-nci
chloride
v
—
CVCCXCII.h
SB-22
Carbamic acid, N,\-diinrthyl-
OCOX((dlj)j
Sc.
Mice
12
ldm
2-dimct hylamino-4-/crf
A
amylphenyl ester met hi-
(slide
V1
CtlUC(CHa)k
SECRET 216
UlOMATIC CARBAMATES
Table 2 (Continued)
VII. Benzene compounds with one carbamate group and one quaternary ammonium group in the meta position.
Code
Name
Structure
Route
and
solvent
Dose
Species mg/kg
Kffect
TL 1309
Phenol, 3-(dielhylamino)-
nicthochloride
Oil
A
Sc.W.
Mice 80
40
20
22
2/2
0/2
-
1 1X(CoH:,)2CH.C1
10
0/2
T-1122
Carhamie acid, 3-dimethyl*
aminophenyl ester methio-
dide —
OCOXIIj
Sc.
Mice 37
U). o
AH 11
Carhamie acid, 3-dimethyI-
aminophenyl ester met ho*
sulfate
OCOXII*
(^X(CH,),Sp4CH,
lv.
Mice 0.7
/.a.
TL 94ti
Carhamie acid, X-methyl-3-
aminophenyl ester hydro-
chloride
OCOXIICH,
1 lxH,HCI
Sc.W.
Mice — SO
40
20 —
0/2
0/2
0/2
ATC12
Carhamie acid, X-methyl-3-
dimet hylaminophcnyl ester
hydrocliloride
OCOXIICH,
(^X(CII,),HC1
Iv.
Mice 15
T-1152_
Carhamie acid, X-incthyl-3-
dimethylaminophcnyl ester
mcthiodhle
OCOXIICH,
(^)nt(cii,),i
! I 1 |
■i-i-i
Mice 0.44
Rabbit 0,26
Mice 30
LDi o
TL 1178
Carhamie acid, X-methyl-3-
dimethylaminophenyl ester
methiodide.
OCOXIICH,
[^)x{CH,),l
Sc.W.
Iv.W.
Mice 0.27
Mice 0.115
(Sec p. 219)
UK,
Llh ,
TL-1226
Carhamie acid, X-inctliyl-3-
OCOXIICH,
Sc.W.
Mice 0.140
LDtt
dimet hylaminophcnyl ester
methochloridc
,)3('i
Iv.W.
Mice 0.070
LDu
T-1090
Carhamie acid, X-met hyl-3-
dimclhylaininophenyl ester
OCOXIICH,
A
Sc.
Mice 0.37
Uho
met liochloride
Mx(CII,),CI
All 13
Carhamie acid, X-met hyl-3-
dimet hylaminophcnyl ester
met hosul fate
OCOXIICH,
A
l\(CH,),SOtCH,
Iv.
Mice 0.1
UK..
TL 1323
Carhamie acid, X-melhvl-3-
OCOXIICH,
Sc.W.
Mice 0.135
Uho
T-T194
dimethylaminophenyl ester
cthiodide (
A
Sc.
Sc.
Mice 0.38
Rabbit 0.13
Uho
ldm
1
]X(CH,),C,H5I
SECRET CHEMICAL STUUCTLRE AND TOXICITY
217
Code
Name
Structure
Route
and
solvent
Species
Dose
Effect
AH 11
Carbamic arid, X-methyl-3-
rlhylmcthylaininophetiyl
ester met hobromide
OCOXIIGlij
s Jx(CIIi)jC-jIIiHr
Iv.
M ice
0.15
UK*
TI. 1134
Carbamic arid, X-methyl-3-
diniel hylaminophenyl ester
propyl bromide (
OCOXHCII, Sc.W.
IX(CIFj)iCMjCHjCIIaBr
Mice
(Sec p. 210)
0.100
(78 K)
UK*
Tbi i:r.
Carbamic acid, X-methyl-3-
dimel hylaminophenyl ester
ally! bromide
OCOXHCII, Sc.W.
*1
IX(CH,bCH,CH—Cll-llr
Mice
(See p. 210)
0.102
(82 K)
UK,
TL 1324
Carbamic acid, X-methyl-3-
dibulylaminoplienyl ester
met h iodide
OCOXHCII,
Ix(C4H,WHJ
Sc.W.
Mice
0.48
UK*
AR-15
Carbamic acid, X-mefhyl-3-
diel by la minophenyl est er
hydrochloride
OCOXIICH,
I.X(CjHt),HCI
Iv.
Mice
5.0
UK*
Tb-1217
Carbamic arid, X-methyl-3-
diet hylaminophenyl ester
met h iodide
OCOXHCH,
lx(C.H„).CHd
Sc.W.
Sc.W.
Sc.W. pH4
Sc W.
Mice
Mice
Mice
0. pi>?
(See p. 210)
0.122*
0.120
0.135
0.007
LDiu
U),„
LD_,
LDW
T-1123
All-If.
TE-1200
Prep. 1
Prep. 2
Prep. 3
Prep. 3
Carbamic acid, X-methyl-3-
diet hylaminophenyl ester
methochloride
OCOXHCII,
IX(C,Hi),CH,CI
Sc.
Tv.
Sc.
Sc.W.
Sc.W.
Sc.W.
Sc.W.
Mice
Mice
Mice —
Mice
Mice
M ice
Mice
(See p. 220)
0.20
0.1
0.13
O.OOOf
0.007
0.105$
0.005§
UK*
UK,
UKo
UK,,
ldm
UK*
UK*
TI. 1317
Carbamic acid, X-methyl-3-
diethylaminophenyl ester
methosnlfate
OCOXHCII,
Qjxccai.MCH^SO,
Sc.W. pi 14
Sc.W.
Sc.W.
Sc.W.
Mice
Mice
Mice
Mice
(Sec p. 220)
0,100
0.114
0.107
0.102
IjDs t
U)M
UK,
UK*
TI-1250
Carbamic arid, X-methyl-3-
diethylaminophenyl ester
eth iodide
ocoxncii,
In(CiIU).1
Sc.W.
Mice
0.23
UK,
AR-31
Carbamic acid, X,X-dimcthyl-
3-dimet hylaminophenyl
ester acid tartrate
OC’ON(C’Hj), Iv.
A
|X(CH,>X—CHOHCOOH),
Mice
00
UK*
* At 8.'. F. t At 75 F. J At 77 F.
s At 76 F.
Table 2, Section VII (Continual)
SFX’RET \ R OM AT 1C C A KB A M A TES
Code
Name
Structure
Route
and
solvent
SjICCieS
Dose
mg/kg
KITcct
TL-1321
SH-24
Carbamic acid, X,N-dimethyl-
3-dimethylaminophenyl
ester methiodidc
OCOXfClb,).
(^)x(CII,),I
Sc.W.
Sc.
Mice
Mice
0.475
0.55
Uho
U),,
AR-32
SB- 23
TL-1304
Carbamic acid, X.X-dimothyl-
3-dimethyIamitiopliciiyl
ester methosulfatc
(Prosl igmine)
OCOX(CH3),
(^X(CII,)1SO,CH3
Tv.
Sc.
Mice
Mice
(See p.
0.5
0.45
220)
UK,
LI),,
TL-1238
Prep. 1
Prep. 2
Prep. 3
Prep. 3
Carbamic acid, X,X-dimethyl-
3-diethylaminophenyI ester
melhiodide
ocox(cir3h.
(^)x(c.iii).cn3r
Sc.W.
Sc.W.
Sc.W.
Iv.W.
Mice
Mice
Mice
M ice
(See p.
80
40
20
10
0.125*
O.ITot
0.089f
220)
2/2
2 2
0 2
0 2
LDia
Uho
/.D,„
TL-1422
Carbamic acid, X,N-dimethyl-
3-diethylaminophenyl ester
met bf (Chloride
OCOX(CII3h
[^)x(CsIi5)iCII3Cl
Sc.W.
Sc.W.
Sc.W.
Mice _
Mice
Mice
(See p.
0.058J
0.108$
0 1 (HI
220)
Uho
W>».
UK,
TL-1481
Carbamic acid, X-elhyl-3-di-
ethylaminophcnyl ester
melhiodide
OCOXHCjHs
(^N(C;1W’IU
Sc.W.
Mice
10.0
5.0
2.5
1.0
2/2
2/2
0/2
0/2
AR-21
Carbamic acid, X-elhyl-3-di-
met hylaminopheny! ester
methosulfatc
OCOX HCjHs
1 J\'(CHj)3SO,CHj
Iv.
Mice
1.0
LDoo
A11-36
Carbamic acid, X-ethyl-X-
incthyl-3-dimethylamino-
phenyl ester methosulfatc
cii,
/
OCOXCsHs
A
Iv.
Mice
3.5
LJ)*o
;■ ' - !
1 lx(CITJ)JS01CH1
AR-33
Carbamic acid, X,X-diethyl-
3-dimethylaminophenyl
ester methosulfatc
OCOX(CiHs)i
(^N(CH.)3SO.CH3
Iv.
Mice
8
/v/2%0
AH--19
Carbamic acid, X-allyl-3-di-
methylaminophenyl ester
hydrochloritle
OCOXIICI!jCH—('Hj
[^Jx(CII3):-HCI
iv.
Mice
150
ID,,
AR-34
Carbamic acid, X,X-diallyl-
3-dime(hylaminophenyl
ester methiodidc
OC<)X(CH.CII-CH,)l
Mx(CH3)3I
Iv.
Mice
10
LD,o
• At S3 F
t At 75 F. t At 80 K.
5 At 71 F. 1) At 7
3 F.
Table 2, Section VII (Continued)
SECRET CHEMICAL STRIXTLKE AND TOXICITY
219
Code
Name
St met ure
Route
and
solvent
SjM-cies
Dose
111)!; kg
Effect
All-20
Carhamic acid, X-allyl-3-di-
methylaminophenyl ester
met la >sulfatc
OCOX HCHjCll—Cll-
1 Ix(C1I,)jSG4CIIi
Iv.
Mice
0.75
L/Xo
All 24
Carhamic acid, X-phenyl-3-
dimethylaminophcnyl
ester hydrochloride
‘ /
OCOX lie, 11=,
|^\x(CH,VIICl
Iv.
Mice
25
/■/),.
AH 25
Carhamic acid, X-phenyl-3-
dimethylaminophenyl ester
met hosulfatc
OCOXIlCslli
1 lx(CHj)jSOiCIIj
Iv.
Mice
2
/-/), 0
All 22
Carhamic acid, X-lx»iizy 1-3-
dime thy la mi noplwnyl ester
hydrochloride
OCOXIlCHjtMIs
3),-IIC1
Iv.
Mice
50
ADS„
AH-23
Carhamic acid, X-benzy)-3-
dimcthylaminophenyl ester
mcthosulfate
OCOXTICHjC.Hs
(^)x(CHJ)iSOtCH3
Iv.
Mice
0.1
LDsa
T-1125
Carhamic acid, N-benayl-3-
(li met hylaminophenyl ester
methiodide
OCOXHCH.CdL
(^)n(CIL),I
Sc.
Sc.
Mice
Rabbit
0.35
0,20
I.D.,,
Lfho
TL 1308
Carhamic acid, N,X-penta-
melhylenc-3-dimethyl-
aminophenyl ester
methiodide
OCOXCJI.o
1 Ix(CIIj)jI
Sc.W.
M ice
10
5
2.5
2/2
2/2
0/2
All-35
Carhamic acid, X,X-penta-
met hylenc-3-di methyl-
aminophenyl ester
mcthosulfate
OCONCsHi#
(^)x(CII,)JSO,CHi
Iv.
Mice
0
TL 1340
Carhamic acid, X,X-|>enta-
met hylene-3-diel hyl-
aminophenyl ester
methiodide
OCOXCJI.o
1 jx(CJU,
Sc.W.
Mice
5
1
0.2
0.1
5/5
3/5
0/5
0/5
CH,I
T 1207
Carhamic acid, X-(4-melhoxy-
pheuyl)-3-dimcthylamino-
phenyl ester methiixlide
ocoxnocit3
A
1 IX(CHa) J
Sc.
Mice
0.24
LD*
TL 1442
Carhamic acid, X-(4-methoxy-
phenyl)-3-dielhylamino-
phenyl ester methiodide
N>OCII,
A
1 IX(CjlTi)jCH,I
Sc.F.
Mice
SO
40
20
0/2
0/2
0/2
Table 2, Section VII {Continued)
SECRET 220
AROMATIC CARBAMATES
Route and
Code solvent
Species
(at
Effects
various doses)
TE 1178 Sc.W.
Rat
Rabbit
G. pig
L>og
Cat
0.10
0.125
0.25
0
.5
1.0
0/2
0/1
0/2
0/2
0/1
0/2
2/2
2/2
0/1
0/2
1/2
2/2
2/2
0/2
2 2
2 2
2/2
2/2
1/2
TL 1434 Sc.W.
Rat
Rabbit
G. pig
Cat
Dog
0.025
0.0,50
0.100
0.200
0/2
0/2
0/2
0 2
2/2
1/2
0/2
1/2
1/2
2/2
1/2
2 2
2/2
2/2
2/2
—
TL-1435 Sc.W.
Rat
Rabbit
G. pig
Dog
0.050
0.100
0,200
0 2
0/2
0/2
2/2
0/2
2 2
2/2
2/2
2/2
TL 1217 _ Sc.W.
0.0.5
0.1
0.2
0.3
0.4
Rat
0/2
0 2
0/7
6/7
Rabbit —
0/2
2/2
G. pig
0/2
2/2
Dog
1/2
22
2/2
Cat
0/2
2/2
2/2
-r— *
Sheep
0/2
3/3
Goat
0/2
2
-'5
2/3
Monkey
0/2
2/3
1"
Route and
Effects
Code solvent
Species
(at various doses)
TL-1299 1in.Imp.
0.025
0.05
0.1
0.2
0.3
(2ml sample)
Goat
0/1
0/1
l/l
Monkey
0/2
0/4
1/1
1/1
Sc.W.
I)o(J
0/3
1/3
8/10
TL-1317 Sc.WT.
0.05
0.1
0.2
0.3
Rat
0/2
3/5
2/2
J- P'K
0/2
1 /5
5/5
Rabbit
0/2
1/2
2/2
(*at
0/2
2/2
TL 1394 Sc.W.
0.2
0.5
1.0
1.5
- .
Rat
0/2
1/2
2/2
Rabbit
0/2
2/2
2/2
> pig
0/2
2/2
2/2
2/2
Dog
0/2
1/2
1/2
—
Cat
0/2
1/2
2/2
TL-1238 Sc.W.
0.05
0.1
0.15
0.2
0.3
0.4
1.0
Rat
0/2
0/2
0/2
1/2
2/2
Rabbit
0/2
1/2
2/2
L l>ig
0/2
3/0
2/2
Cat
0/2
2/2
Dog
0/2
/2
2/2
T LI 422 Sc.W.
0.05
0.10
Rabbit
0/2
2/2
Dog
0/2
2/2
Table 2, Section VII (Continued)
SECRET CHEMICAL STKCCTl HE VXD TOXICITY
221
Table 2 (Continued)
VIII. Benzene compound* with one carbamate group and one quaternary ammonium group in the
met a position anti other .substituents.
Code
Name
Structure
Route
and
solvent
Species
Dose
mg/kg
Kffect
TL-12r.fi
Carbnmic acid, 2-me I hy 1-5-
OCONH,
Sc.
Mice
10
2/2
dimethylainiiu.phenyl ester
/\
5
2/2
methiodide
cn/ ]
1
0/2
0.5
0/2
TL-1184
Carbnmic acid, N-melhyl-2-
OCONHCHj
Sc.P.
Mice
80
1/2
mcthyl-5-dimelhyltimiiio-
A
40
0/2
phenyl ester
C11/ ]
20
0/2
I jN(CIfj)i
Sc.W. pH3
Mice
SO
5/5
V
40
5/5
20
1/5
T 1708
Carbamie acid, N-methyl-2-
OCONHCHj
Sc.
Mice
0.1-0.12
LDw
TL-1071
me! lrvl-5-dimet h vlamino-
/\
Sc.W.
Mice
0.115
/.Djo
phenyl ester methuxlide
CIl/ ]
Sc.W. pH4
Mice
0.1 as
LD„ 0
1 tN'(CU,)J
Sc.W. pH!
Mice
0.107
Uh 0
V
Sc.W. pH4
Mice
0.102
/.Dju
TL-1236
Carbnmic acid, N-methyl-2-
OCONHCHj
Prep. 1
methvl-5-dimethylamino-
/\
Sc.W.
Mice
0,075
ID, 0
Prep. 2
phenyl ester methochloridc
ch/ ]
Sc.W.
Mice
0.064
LD,a
Prep. 2
—
1 In(CH,).C1
Ip.W.
Mice
0.088
Uh.
Prep. 2
V
Sc.W.
Rats
0.100
LDi0
Prep. 2
-
Ip.W.
Rats
0.078
LD,.
— Prep. 2
Oral W.
Rats
2.5
LD, 0
Prep. 2
-
Sc.W.
Dogs
2.0
4/10
1.0
0/3
Prep. 2
:
Im.Imp.
Monkeys
0,0.50
1/4
Prep. 3
: &.w.
Mice
0.070
Uh0
Prep. 3
-
—-
Iv.W.
Mice
0.035
TL-1185
Carbnmic acid, N-methvl-2-
OCONHCHj
Sc.W.
Mice
0.110
LDia
met hyl-5-dirnet hylarni no-
/\
(See p. 224)
phenyl ester methosulfate
ch/ ]
1 InCCIDjSOjCH,
TI,-1186
Carbamie acid, N-methyl-2-
OCONHCHj
Sc.W.
Mice
0.103
metliyl-5-diinethylamino-
/\
(See p. 224)
phenyl ester methosulfuric
CH/ ]
acid
1 iN(CH,),HSO,
TL-1340
Carbamie acid, N-methvl-2-
OCONHCH,
Sc.W.
Mice
0.090
ID-,.
methyl-5-dimet hyhimino-
/\
—
phenyl ester ethiodide
CH/ ]
'
t Jn(ch,),
1
-
CjHjI
TL-1339
Carbamie acid, N-methyl-2-
OCONHCH,
Sc.W.
Mice
~ 0.075*
LDw
methyl-5-dimcthylamino-
/\
phenvl ester elhochloridc
ch/ ]
TIn(CHj)»
V I
c2h*ci
TL-1257
Carbnmic acid, N-methyl-2-
OCONHCH,
Sc.W.
Mice
0.125
T-1739
met hvl-5-dietli vlamino-
/\
1
Mice
0.2
phenyl ester methiodide
ch/ ]
I ]N(C..Hs)jCH,I
• At 8i F.
SECRET 222
ARC) M A TIC C \ H B A M AXES
Route
and
Dose
(‘(Hie
Xante
Structure
solvent
Sjteeies
ntR/kg
Effect
TL -1202
Carbamic acid, X-methyI-2-
met hyl-5-( X-Ih'mzvI-X -
OCOXHCllj
/\
Sc.W.
Mice
10
5
2 2
2 2
methylamino)phenyl
CH/ ]
2.5
0 2
ester methochloride
I Jx(CIIi)*
2
0, 2
X/ 1
CH.C,H»Cl
T I,-1201
Carbamic acid, X-methyl-2-
melhyl-5-(X-allyl-X-meth-
vlamino)phcnyl ester me lb-
OCOXHCH,
C\l/ | (Cll.hd
Se.W._
Mice
0.077
UK,,
—
ochioride
\/ X C1 IjC 11 —CH,
TL-1511
Carbamic acid, X-methyl-2-
met hyl-5-dimel bylamino-
phenyl ester 0-hydroxy-
ethiodide
OCOXHCH,
CM A
1 Jx(CHi)iI
Sc.W.
Mice
o.ns*
v' 1
CHjCHiOIl __
TL 1512
Carbamic acid, X-methyl-2-
OCOXHCH,
Sc.W.
Mice
0.050 f
/./Co
met hvl'5-dimethylamino-
/\
■ = , —
phenyl ester aeetonylchlo-
CH/ ]
ride ...
1 IX(CHj)iCI
CH-COCII,
—
TL 1513
Carbamic acid, X-methyl-2-
ocoxiich.
Sc.W.
Mice
0.1051
U),a
one t by 1-5-d i me t by 1 a mi no-
/\
phenyl ester carbethuxy-
CH/ ]
methochloride
1 1x(CH,)2C1
CHCOGCjIIj
T-1722
Carbamic acid, X-methyl-3-
OCOXHCH,
(In saline)
Mice
0.75
dimethylamino-O-elhyl-
phcnyl ester melhiodide
C.«A
1 IX(CH,),I
(In buffer
solution)
Mice
1.30
LDka
T-1700
Carbamic acid, X-methyl-5-
OCOXHCH,
9
Mice
250-300
TL 1501
dimethylainino-2-Lsopropyl-
/\
(In buffer
phenyl ester melhiodide (CII,)jCH[ |
solution)
Mice
125
LDu
1 IX(CH,),I
Sc.W.
Mice
80
0/2
-
10
20
0/2
0/2
T-T778
Carbamic acid, X-methyl-2-
OCOXHCH,
?
Mice
175
Uho
cyclohexyl-o-dimet hylamino-
/\
phenyl ester melhiodide
CeHnf ]
1 JX(CH,),I
•
T-1842
Carbamic acid, X-methyl-2-
OCOXHCH,
?
Mice
45
LDi o
cbloro-5-dimcthvlamino-
A
phenyl ester hydrochloride -
1 lx(CH,),-HC1
-
T-1S00
Carbamic acid, X-methyl-2-
chloro-5-diinethylaminO'
phenyl ester melhiodide
OCOXHCH,
cA
?
Mice
4
LDm
1 lx(CH,),I
TT-1523
Carbamic acid, X-methyl-3-
OCOXHCH,
Sc.W.
Mice
0.120
1st ipropy 1-5-d i me t hy huni no-
A
(78 F)
phenyl ester melhiodide
f 1
~
(CHj).HCl JX(CIIj)jI
* At:
•e F. t At 73 F. t At 74 F.
Tabi.k 2, Section \111 (Continued)
SECRET CHEMICAL STRUCTURE VM) TOXICITY
223
Tab IE 2, Sectum VIII (Continued)
Code
Name
Structure
Houle
and
solvent
Species
Dose
»"K kg
Effect
T ITtiS
Carhamic acid, X-mclhyl-3-
OCOXHCHj
?
Mice
10-15
LD, o
dimethvlamino-4-inethvl-
/\
phcnvl ester hydrochloride
I IX(CIIi)*- LIC1
\r
CH,
T I.-1187
Carhamic acid, X-methyl-l-
OCOXHCHj
Sc.P.
Mice
80
2 /2
mcthyl-3-dimcthylamino-
/\
40
2/2
phenyl ester
20
2/2
—
I JX(CII,),
10
0/2
V
5
0/2
cir.
Sc.W. pl!3
Mice
so
5/5
40
5/5
20
5/3
10
0/3
5
0/5
TIM216
Carhamic acid, X-methvl-4-
OCOXHCHj
Sc.W.
Mice
0.170
LD,„
iuethvl-3-dimcthvlamino-
/\
phenyl ester methiodidc
fi
1 lX(CIIj)aI
“ -
V
CHS
—
Tl. I t53
Carhamic acid, X-mcthyl-4-
OCOXHCH,
Sc.W.
Mice
0.130
LDi o
mcthyl-3-dimctliylamino-
A
—
(75 F)
phenyl ester mcthochloride
1 lx(cn,)jCi
\r
CH,
TL U20
Carhamic acid, X-methyl-4-
OCOXHCH,
Sc.W.
Mice
0.155
hO.»
met h vl-3-dimel hylami no-
/\
(72 F)
phenyl ester ethiodide
( ]
• . —'*
I IxXCHjJjCjHJ
—
CU9
TIM 188
Carhamic acid, X-methyl-4-
OCOXHCH,
Sc.W.
Mice
0.200
U) ss
methyl-3-dimethylamino-
/\
—
(See p. 224)
phenyl ester methosulfatc
1 Ix(CHj)jSOjCH,
CH,
TL-1354
Carhamic acid, X-methyl-l-
OCOXHCH,
Sc.W.
Mice
0.005
/>D«
met hvl-3-dimet hylamino-
/\
—
]>henvl ester allvl bromide
( IX(CH,),CHjCH
—CHJIr
' -
V
CH,
TIM 338
Carhamic acid, X-methyl-4-
OCOXHCH,
Sc.W.
Mitre
10
3/3
met hyl-3-met hylbenzyl-
/\
5
2/3
aminophcnyl ester met ho-
| I
i
0/3
bromide
1 JXCHjCfHs
0.5
0/3
\/ \
CH. (CH,),Br
T-17(19
Carhamic acid, X-mcthvl-3-
OCOX HCH,
(In buffer
Mice
70
LD„a
dimethylamino-4-isopro-
/\
solution)
pylphenyl ester hydrochlo-
f 1
ride
I Jx(CH.hHCI
CTI(CH,)»
SECRET 224
AROMATIC CARB \MATES
Code
Name
Structure
Route
and
solvent
S[H'oies
Dose
ntR/kK
Kffect
TL-1502
Carbamic acid, X-methvl-3-
OCOXHCH,
Sc.W
Mice
0.51
CDi0
dimelhylamino-4-isoprt)-
A
(75 F)
T-1721
pyipbcnyl ester methiodide
(
Mice
1
1
lx(CH,),l
——==■"
CII(CH,),
T-1770
Carbamic acid, X-methvl-2,4-
OCOXHCH,
9
Mice
10
LD,,
diincthyi-3-dimelhylainino-
A
phenyl ester hydrochloride
CHJ
1
JX(CHj)j*HC1
CH,
T-1707
Carbamic acid, X-methvl-2,4-
OCOXHCH,
9
Mice
0.1
U>.n
dimcthvl-.Vdimethvlamino-
A
phenyl ester methiodide
CHJ
—
1
|X(CH,),I
V
CH|_
TM710
Carbamic acid, X-inethyl-3-
OCOXHCH,
9
Mice
0.17
LD*,
dimethylamino-5- methyl-
A
plienyl ester methiodide
1
1
-
CHJ
lX(CH,),I
T 1741
Carbamic acid, X-methvl-3-
OCOXHCH,
?
Mice
0.4
dimethylamino-4-ethyl-
A
phenyl ester methiodide
(
—
—
1
Jx(CH,),I
C,H,
—
TL 1237
Carbamic acid, X,\-dimethyl-
OCOX(CH,V
Sc.W.
Mice
10
2/2
2-met hy 1-5-di met hylamino-
A
5
2/2
phenyl ester methiodide
CHJ
2.5
2/2
1
2.0
0/5
V
1.0
0/5
TL-1423
Carbamic acid, X, X-dimcthyl-
OCOX(CHj),
Sc.W.
Mice
10
2/2
2-met hvl-5-diethvlami me
A
5
2/4
phenyl ester methiodide
CHJ
1
__
0.5
0/5
1
lX(CjHi),CH,l
0.25
0/5
TL-1325
Carbamic acid, X,X-dimcthyl-
OCOX(CH,h
Sc.W.
Mice
10
2/2
4-met hyl-3-dimct hylamino-
A
5
2/2
phenyl ester methiodide
1
f 1
i
1/2
1
[ Jn(CH,),I
_
0.5
0/2
V
0.2
0/2
CH,
—
TI,-1487
Carbamic acid, X-methyl-X-
OCOXCH;,OCH,
Sc.W.
Mice
2.5
5/7
mot hoxy-2-met hyI-5-di meth-
A
1.0
0/7
ylaminophcnyl ester meth-
CHJ
]
0.5
1/10
iodide
1
0.25
0/10
TL-1300
Carbamic acid, X.X-penta-
OCOXCJI.o
Sc.W.
Mice
SO
1/2
methvlene-2-methyl-5-dimeth-
A
40
0/2
ylaminophcnyl ester meth-
CHJ
1 i
20
0/2
iodide
1
1 IX(CII,),1
TL-1355
Carbamic acid, X,X-penta-
OCOXCjH.o
Sc.W.
Mice
20
2/2
methvlene-4-metbvl-3-di-
A
10
2/2
met hylaminophenyl ester
1
f I
_ 5
0/2
met hiodide
1
1 IX(CH,),I
1
0/2
\/
CH,
Table 2, Section VIII (Continued)
SECRET CHEMICAL STRUCTURE AND TOXICITV
225
Table 2, Section VIII (Continued)
Route
and
Dose
Code Name
Structure
solvent Species mg. kg Effect
TL-1239 Carbarn! biolic acid, N-metby
-
SCOXIICHj
Sc.W. Mice 80
0/2
3-diinctliylamino-4-mcth-
/
\
40
0/2
ylphenyl ester mcthiodidc
f
— -
20
0/2
I
|X(C11,),I
/
\
CM,
Route ami
Effects
Code solvent
Species
(at various doses) _
;TL 1185 Sc.W.
0.1
0.2
0.3
0.4
1.0
/
Rat
0/2
1 /2
2/2
Rabbit
0 2
1/2
1 2
• 2/2
G. pig
0/2
2/2
—
Dor
0 2
1/2
1/2
Cat
0/2
2,2
2/2
Sbeep
0/1
0/2
0/2
0 2
i/r
_
Goat
0/2
2/5
0/2
0 2
Monkey
0/2
TE-11 Sti Sc.W.
0.05
0.1
0.2
0.3
Rat
0/2
2/2
Rabbit
0/2
2/2
—
G. piR
0/2
2/2
2/2
Dor
0/2
1/2
0/2
TI.-1188 Sc.W.
0.1
0.2
0.3
0.4
Rat
0/2
1/2
0/2
0/2
— -
Rabbit
0/2
1/2
1/2
G. piR
0/2
2/2
Dor
0/2
2/2
1/2
...
IX. Benzene compounds with one carbamate group and one quaternary ammonium group in the
para [>osition (including thiocarbamates).
Code
Name
Structure
Route
and
solvent
Species
Dose
mg/kg
Effect
TL-913
Carhamic acid, N-methyl-1-
OCONIICHj
Sc.W.
Mice
80
2/2
aniinophenyl ester hydro-
A
40
0/2
chloride
o
__
20
0/2
■
NHj-IICl
T 1088
Carhamic acid, X-methvl-1-
OCONIICHj
Sc.
Mice
50
I-tDt, o
AH-17
dimethylaminophcnyl ester
A
Iv.
Mice
2
LDa o
TL-101)7
methiodidc
( i
Sc.W.
M ice
80
2/2
I J
10
2/2
V
20
2/2
N(CIIj)jl
10
0/2
5
0/2
TL-M69
Carhamic acid, X-methyl-4-
OCONHCH,
Sc.W.
Mice
10
2/2
dimethylaminophenyl ester
A
20
2/2
ethiodide
f ]
10
0/2
.
v
5
0/2
N(CH,)jC}H J
secret 226
AROM ATIC C ARR A MATES
Code
Name
Structure
Route
and
solvent
Species
Dose
mg kg
Effect.
TL 1150
Carhamie acid, X-methyl-4-
OCOXTICHj
Se.W.
Mice
10
2 2
di met hy la minopheny 1 ester
/\
5
2 2
allyliodide
()
2.5
0/2
\/
(ClLi'ijXCTLCU—CTljI
TL-1431
Carhamie acid, X-methyl-4-
OCOXI1CH j
Sc.W.
Mice
40
9/9
diethylaminophcnyl ester
A
20
1 2
methiodide
[ ]
10
0/2
v
5
0/2
X(C;lL’,):CHc4
.
-
.
TL 1132
Carhamie acid, X-methyl-4-
ocox iic Hi
Sc.W.
Mice
10
2/2
--
diethylaminophcnyl ester
A
20
1/2
allyliodide
f 1
10
0/2
v
I)
0/2
X(CdL)iI
CH;CH—CH*
TL 1430
Carhamie acid, X-methyl-4-
OCOXIIC1L
Sc.W.
Mice
SO
2 2
-
diethylaminophenyl ester
A
40
1 /2_
ethiodide
f i
20
0/2
v
10
0/2
'
X(C-lL).d
TL-1437
Carhamie acid, X,X-diinetliyl-
OCOX(C!L>.
Sc.W.
Mice
SO
1/2
4-dimet hylaminophenyl
A
40
0/2
ester ethiodide
o
20
0/2 _
V
XCCHahCsHJ
TL-1486
Carhamie acid, X,X-dimethyl-
OCOX(CH3)2
Sc.W,
Mice
SO
2/2
4-dimethylarninophenyl ester
A
40
1/2
/3-hydroxyet hobromidc
f 1
20
0/2
'\J
10
0/2
X(CIIj)2Br
-
—
|
CILCILOII
TL-1470
Carhamie acid, X,X-dimethyl-
OCOX(CIL).
Sc.W.
Mice
80
2/2
4-dimet hylaminophenyl
A
40
1/2
ester allyliodide
f I
20
0/2
v
10
0/2
N(CHi)jI
• — '■
CH*CH-=CH,
TL 1438
Carhamie acid, X,X-dime(hyI-
OCOX(CTL),
Sc.W.
Mice
80
0/2
4-diet hylaminophenyl ester
A
—: ■ _
40
0/2
met hiodide
o
20
0/2
V
X(CjHs)iCHjI
TL-1472
Carhamie acid, X,X-dimethyl-
OCOX{(TL,)j
Sc.W.
Mice
SO
0/2
4-diel hylaminophenyl ester
A
40
0/2
allyliodide
()
20
0/2
V
N(CjH»)»l
-
(’IIjCII—CHj
Table 2, Section IX {Continued)
SECRET CHEMICAL STRUCT I.'RE AND TOXICITY
227
Code
Name
Structure
Route
and
solvent
S|iecies
Dose
mg kg
Kffect
TL-1471
Carbamic acid, N,X-dinielhyl-
OCOXtCH.h
Sc.VV.
Mice
SO
0/2
4-diet hylaminophenyl ester
/\
40
0/2
ethaxlidc
||
20
0/2
.
v
—'
X(C.IU)3I
TL 1220
Carbamlhiolic acid, X-methyl-
SCOXHCH,
Sc. I*.
Mice
80
1/2
4-nit rophenyl ester
A
40 —
1/2
■—
I 1
20
0/2
u
10
0/2
-
V
NO.
TL-1258
Carbamthiolicacid, X-methyl-
SCOXHCH,
Sc.W.
Mice
SO
0/2
4-dimet hylaminophenyl
A -
10
0/2
ester methiodide
0
20
0/2
V
X(CII3),1
—
TL-1054
Carbamthiolthionic acid, X,X-
SCSX(CH,),
Se.P.
Mice
40
0/2
dimethyl-4-nit rophenyl
A
20
0/2
ester
o
T
V
xo.
—
TL-1128
Carbamthiolthionic acid. X,X-
SCSX(CII3)s
Sc.W.
Mice
SO
0/2
dimethyl-l-aminoj)henyl
A
10
0/2
ester hydrochloride
()
20
0/2
V
NHj-HCl
TT-1179
Cnrhamtbiolthionie acid, X,X-
SCSX(CIIj),
Sc.W.
Mice
SO
1/2
dimethyl-4-dimet hylamino-
A
(suspension)
40
0/2
phenyl ester metliiodidc
()
—
20
0/2
v
X(CH,),I
—
X. Benzene compounds with one
carbamate group and one quaternary ammonium group
in the
—
para position and other substituents.
TL-1478
Phenol, 3-isopropyl-i-dimetli-
OH
Sc.W.
Mice
80
2/2
ylamino-, methiodide
A
40
2/2
—
20
0/2
McHfCIh),
•
10
0/2
X(CH,),I
TL-1322
Carbamic acid, X-methyl-2-
OCOXHCII,
Sc.W.
Mice
0.51
LDia
isopropyl-4-dimet hylamino-
A
phenyl ester methiodide
r jcii(cii,)i
—
v
X(CII,),T
TL 1446
Carbamic acid, X-methvl-3-
OCOXHCH,
Sc.W.
Mice
10
2/2
methvl-4-ecics
Dose
mg kg
Effect
TL1147
Carbamic aeid, X-niclliyl-3-
OCONHCH,
Sc.W.
Mice
It)
2 2
incthyl-1-diinct hylamino-
A
5
2 2
phenyl ester cthiodide
1
2/2
1 Jen,
0.5
2/2
V
0.25
0/5
N(CHi)jCj!Isl
TL-1448
Carbamic acid, N-methvl-3-
OCONHCHj
Sc.W.
Mice
0.24
/,/>.„
met hyl-4-dimet hylamino-
— A
(73 F)
phenyl ester allyliodide
( |
vrH‘
(CTOjNCHjCII—CHS1
TL-1434
Carbamic acid, N-methyl-3-
OCONHCHj
Sc.W.
Mice
JO
2/2
mcthyl-4-diel hylamino-
A
5
2 2
phenyl ester methiodide
I I
i
1 2
I Jell.
0.5
0/2
V
0.25
0/2
N(C*Hi)iCIIiI
TL 1407
Carbamic aeid, N-methyl-3-
OCONHCHj
Sc.W.
Mice
0.145
LD> o
ethyl-l-dimethylamino-
A
(74 F)
phenyl ester methiodide
| 1
_ -
—
\ A'11'
N(CH,),I ~
TI/-1468
Carbamic aeid, N-methvl-3-
OCONHCHj
Sc.W.
Mice
0.39
LD, „
propyl-4-di met hytami no-
A
(78 F)
phenyl ester methiodide
( |
—
I ICHjCH-CHj
N(CH,),I
TL-1381
Carbamic acid, N-methyl-3-
OCONHCH,
Sc.W.
Mice
10
2/2
isopropyl-4-dimet hylamino-
A
5
2/2
phenyl ester hydrochloride
2.5
2/2
1 JCH(CHj)j
1.0
1/2
—
V
0.5
0/2
—
N(CH,VHC1
_ _
TL-1327
Carbamic acid, N-mcthvl-3-
OCONHCH,
Prep. I
isopropyl-4-dimet hylamino-
A
Sc.W.
Mire
0.067
Prep, 2
phenyl ester methiodide—
Mice
0.070
LD„
1 JCH{CH,)j
(79 F)
Prep. 2
V
pH 4
Mice
0.064
LD, „
N(CH,),I
. —
TL 1345
Carbamic acid, N-methyl-3-
OCONHCH,
Prep. 1
isopropyl-4-dimcthylamino-
A
Sc.W.
Mice
0.045*
LD„n
Prep. 2
phcnvl ester methochloridc
I |
Sc.W.
Mice
0.0474
LD:,n
Prep. 2
1 JCH(CHj),*
Sc.W. pH 4
Mice
0.050}
Prep. 2
V
Sc.W.
Rats
0.1035
U)M
N(CH,),C1
(See p. 234)
TL-1522
Carbamic acid, N-methyl-3-
OCONHCH,
Sc.W.
Mice
0.057}
LDia
isopropyl-4-dimethylamino-
A
—
phenvl ester alls ! bromide
1 k'lKCH,),
(CH,hNCH,CII -CH,Br
TL-1475
Carbamic aeid, N-methyl-3-
OCONHCH,
Sc.W.
Mice
10
2/2
but vM-dimet hvlamino-
A
5
2/2
phenyl ester methiodide
i
0/2
1 ICILCH-CTLCH,
0.5
0/2
N(CH,),I
«
At 00 F. t At 87 F.
4 At 78 F. i At 81 F.
SECRET CHEMICAL STRl’CTUKE AND TOXICITY
229
Route
—
anil
Dose
Code
\ ante
Structure
solvent
Species
niK/kK
Effect
TL 1476
CarLamic arid, X-mcthyl-3-
OCOXHCII,
Sc.W.
Mice
10
2/2
am vl- 1-dimet hylamino-
/\
5
2/2
phenyl ester methiodide
1
i
0/2
1 ICILCILCILCII.CIL
0.5
0/2
X(CH,),I
TL-1416
Carhamic arid, X-mclhvI-3-
OCOXIICH,
Sc.W.
Mice
10
2/2
cyclopentyl-4-dimethyl-
/\ CIL -CM,
5
2/2
aininophcnyl ester meth-
i
0/2
iodide
Jen
0.5
0 2
V V
ciL-cn,
X(CH,),I
TL 141K)
Carhamic acid, X-melhyl-3-
ocoxncit,
Sc.W.
Mice
10
2/2
hexyl-4-dimethylamino-
/\
5
2/2
phenyl ester methiodide
1
2.5
2/2
-
v JcJL,
1.0
0/2
V
0.5
0/5
X(CH,),I
Tl, 14 SO
Carhamic acid, X-metlivl-2,5-
OCOXIICH,
Sc.W.
Mice
0.325*
hDi o
dimelhvl-4-dimethvlamino-
/\
phenyl ester elhiodide
chj
I
\JVlh
- : ■
XiCFLWdU
-
TL-1254
Carhamic acid, X-methvl-3,5-
OCOXHCII,
Sc.W.
Mice
80
2/2
dime) hyl-4-dimcthyla mino-
A
40
2/2
phenvl ester hvdroiodidc
1
20
0/2
chJ
ijc".
10
0/2
—
X(CH,)jHI
—
TL 1482
Carhamic acid, X-methvl-4-
ocoxiich.
Sc.W.
Mice
0.145f
LD.0
dimet h vhuninocarvacrvl
A
ester ethiodide
CHJ
1
-
1 lcH(CH,),
X(CH^C,HSI
SB-26
Carhamic acid, X-mcthvl-4-
OCOXHCII,
Sc.
Mice
23
LD*
dimet hylamiiiothymyl
A
ester hydrochloride
{Clique
1
_
nJch,
X(CH,),-HO
SB-27
Carhamic acid, X-melhvI-4-
OCOXHCII,
Sc.
Mice
0.22
LDi0
dimet hylamiiiothymyl
A
ester methiodide
(Client
1
—
iyH'
-
X(CH,),I
TL 1451
Carhamic acid, X-methvl-2,6-
OCOXIICH,
Sc.W.
Mice
80
2/2
dils< ipropvl- 4-dimet hyl-
A
40
0/2
aininophcnyl ester met h-
(Clique
( |CH(CII,),
20
0/2
iodide
KJ
X(CH,),I
SB-2
Carhamic acid, X,X-dimethvl-
OCOX(CH,h
Sc.
Mice
>400
LDtri
4-dimolhvlamino-2-mcthyl-
A
phenyl ester hydrochloride
l)"‘
V
X(CIL), TIC1
* At 75 F. t At 73 F.
Table 2, Section X (Continued)
SECRET AROMATIC CARBAMATES
Table 2, Section X (Continued)
Code
Name
St met tire
Route
and
solvent
Species
Dose
mg ku
KlTcct
SB 3
Carhamic acid, X,X-diniethvl-
OCOX(CHi).
»Sc.
Mice
6.5
4-dimelhvlamino-2-methyl-
/\
phenyl ester methiodide
()"■
— .
v
N(C1I,),I
TL-I313
Carhamic acid, X,X-dimethvl-
OCOX(CHj)j
Sc.W.
Mice
0.17
4-dimethylamino-2-isopropyl-
A
phenyl ester methiodide
[ jcii(cih)j
V
SB 1
Carlmmicacid, X,X-dimethvl-
OCX)X(CII,)*
Sc.
Mice
105
t-diinet hvlainino-3-rnel hvl-
/\
--
phenvl ester hydrochloride
f 1
U1"'
-- —
X(CHj)i-IICl
SB 5
Carhamic acid, X,X-dimethvl-
OCOX(CHi)j
Sc.
Mice
13.0
LD,„
4-diiiiclhvlamino-3-methvl-
/\
phenvl ester methiodide —
I ]
-
X(CH,)»1
TL 11 10
Carhamic acid, X,X-dimethvl-
cuj
SB-6
('arbainic acid, X,X-dime(hy 1-
OCOX(CH,)»
8c.
Mice
45
4-dimethylamino-3-ethyl-
/\
phenyl ester hydrochloride
( ]
—
1 ICjII*
_* .
X(CHj)»-IK1
SB 7
Carhamic acid, X.X-dimethyl-
OCOX(CH.h
8c.
Mice
1.15
LD it,
TL-1412
4-dirnet hvlamino-3-elhyl-
/\
Sc.W.
Mice
10
2/2
j>henvl ester met hiodide
f 1
5
2/2
I kyi,
i
1/2
V
0.5
0/2
X(CHj)iI
SB-8
Carhamic acid, X,X-dimcthyl-
OCOX(CHi)j
Sc.
Mice
0.075
LDio
TIXiOy
l-dimcthylainim>-5-isopro-
A
Sc.W.
Mice
0.080
Uh 0
pvlphenvl ester methiodide
l i
Sc.W.
Mice
0.080
l JcM(CHj)j
Ip.W.
Mice
0.108
LDi 0
-
V
Ip.w.
Mice
0.220
U)i*
X{CIIj)jI
Ip.w.
Mice
0.265
I'D it.
(See p.
234)
TL-1460
(. 'arhamic acid, X, X-dimet hyi-
OCOX(CH3)j
Sc.W.
Mice
10
2/2
3-propvl-4-dimet hylamino-
A
5
2/2
phenyl ester methiodide
1 1
i
2/2
1 JCIIiCHjCHi
0.50
0/2
V
0.25
0/2
X(CHj)iI
SECRET CHEMICAL STRCCTl RE VM) TOXICITY
231
Table 2, Section X (Continued)
('ode
Name
Structure
Route
and
solvent
S|>eeies
I lose
niK k*
Kffect,
TL 1143
Carlmmic acid, X,X-dime!hyl-
OCOX(CHah
Sc.W.
Mice
0.065
ldm
3-isi >i >n >py l-4-di met hyl-
/\
(73 F)
arninophenyl osier met ho-
I I
chloride
1 IcirtcH,),
-
X(CHj)jCl
-
TL-1521
Carbamic acid, X,N-dimcthyl-
OCOX(CIL)-
Se.W.
Mice
0.182
3-isopropyI-4-dimethyl-
/\
(71 F)
aminoplienyl os tor elhio-
f 1
dide
1 lcH(CHj)«
XXClLbCdU
TL-1461
Carbamic acid, X,\-dimethyl-
OC()X(CHj)i
Sc.W.
Mice
10
2 2
3-hut vl-4-dimet hvlamino-
/\
5
2 2
phenyl ester met hiodide
\ |
i
0 2
-
1 kcn,),cH,
0.5
0 2
x(ciis)a
TI/-14152
('arbamie acid, X,X-dimclhyl-
ocoxxcn,).
Se.W.
Mice
10
2 2
3-umyl-4-dimelhvlamino-
/\
5
22
phenvl ester incthiodidc
( 1
i
0 2
•
1 I(CHj)iC11j
0.5
0/2
Tl. 1163
Carbamic acid, X,X-dimel hyl-
()COX(Clh):
Sc.W.
Mice
10
2/2
3-hexyl-l-dimet hvlamino-
A
5
2/2
phenvl ester mcthia mic acid, X, X -diuicthyl-
OCOX(ClUh
Sc.W.
Mice
10
2/2
3-cyclopentvl-4-diinethvl-
/\ CHj —CH;
5
2/2
arninophenyl ester
1
2/2
methiodide
1 Jem
0.5
0/2
—
V \
CH,—t
H,
-
TL-1166
Carbamic acid, X,X-dimethyl-
OCOX(CH3)i
Sc.W.
M ice
80
0/2
3-phenvl-l-dimethylamino-
/\
10
0/2
phenyl ester methiodide
1 1
20
0/2
VCcH'
X(CIT,)al
SB-28
Carbamic acid, X-methyl-4-
OCOXIICHj
Sc.
Mice
2.1
LD„
dimethylarainocarvacryl
/\
ester hydrochloride
1 P1*
(CHahlld \
X(('lf»)** 1IC1
'
SR-29
Carbamic acid, X-methvl-4-
OCOXIICHj
Sc.
Mice
0.09
LDm
dime! hylaminocarvacryl
/\
ester methiodide
I FH»
{cii.wid 1
X(CHj)jl
SIX’HIT 232
AROMATIC CARBAMATES
Taiu-E 2, Section X (Continued)
Code
Name
Structure
Route
and
solvent
Species
Dose
mg kg
K fleet
SHI 1
Carbamicaeid, X.X-dimcthvl-
OC()X(CHj)j
Sc.
Mice
20
LD> o
4-dimct hvlaminocarvacrvl
/\
ester hydrochloride
1 Flh
(C1I3).1IC1 1
x(cii»)*-nca
SIM 2
Carbamic aeid, X,X-dimethyI-
0(ox(cir,)-
Sc.
Mice
0.24
U)»
4-dimct hvlaminocarvacrvl
/\
ester methiodide
(cn,)*nd 1
X(Clh),I
—
SIM)
Carbamic acid. X.X-dimcthvl-
OCOX(CHj)-
Sc.
Mice
ICO
Uh 0
l-dimet hylaminot hymyl
(CH,yiic/\
ester hydrochloride
It
ijcu.
—
X(CHj)i- IIC1
■
-
SU-10
Carbamic acid, X,X-dimethyl-
OCOX(Clf3)j
Sc.
Mice
0.72
LDt o
•1 -dime! hylaminot hymyl
/\
ester methiodide
(CH3)2U('| ]
[JVH.
_ .
X(CHa)»I
TL Hilo
Carbamic acid, X,X-dimethvl-
OCOX(CIL)i
Sc.P.
Mice
SO —
0/2
3,o-di me t hy 1-4-nit r< rphenyl
/\
40
0/2
ester
f 1
20
0/2
chJ Jciu
XOj
- —
TL-1233
Carbamic acid, X,X-dimetliyl-
OCOX(CHj)i
Sc.W.
Mice
SO
0/2
3,5-dimcthyl-4-di methyl-
/\
40
0/2
aminophenyl ester hydro-
20
0/2
iodide
cnJ leu,
X(CHj)ilII
TL-1377
Carbamic acid, X-cthvl-3,5-
OCOXIICjH,
Sc.W.
Mice
SO
0/2
diisopropyl-4-di methyl-
/\
—
40
0/2
aminophenyl ester
[ I
20
0/2
(CH.hCllI IcH(CII»)j
-
X((’H,)j
TL 1077
Carbamic acid, X,X-dicthyl-
OCOX(CjH5),
Sc.O.
Mice
80
0/2
3,5-dimethyl-4-nil roso-
/\
40
0/2
phenyl ester
I I
20
0/2
CllJ IcH,
- ■—
xo
TL-1197
Carbamic acid, X,X-diethvl-
OC()X(Cdl„)5
Sc.P.
Mice
80
0/2
3,.5-dimethyl-t-nit rophenyl
/\
40
0/2
ester
f 1
20
0/2
CllJ ICH,
s/
NO,
TL -907
Carbamic acid, X',X-diethvl-
OCOX(CjHi)j
Sc.W.
Mice
80
2/2
4-dimet hylaminot Itymyl
/\
40
1/2
ester methiodide
]
20
0/2
K/"'
10
0/2
X(CHj)J
SECRET CHEMICAL STRUCTURE CM) TOXICITY
233
Code
Name
Structure
Route
and
solvent
Species
Dose
tng kj?
Effect
TL 778
Carbamie acid, X.X-dielhyl-
OCOXCJLO
Sc.W.
Mice
SO
0 2
eneoxy-4-dimelhylamino-
/\
40
0/2
carvacrvl ester met ho-
20
0 2
cldoride
\
X(CH,),C1
TL 770
Carbamie acid, N,\-dielliyl-
OCOXC.IbO
Se.W.
Mice
80
1/2
eneoxy-4-dimet hyiamino-
A
40
0/2
carvacrvl ester methiodide
—
10
0/2
(CH.JjIIcI 1
—
X(GHa).r
TL 1073
Carbamie acid, X,X-6(S(2-
ocoxcciun.cib
Sc.W.
Mice
SO
1/2
chloroethyl )-4-dimet hyl-
A
'
40
1/2
luninothymyl ester metho-
(CHiVt ’1I| ]
*
20
0 2
cldoride .
V"'
10
0/2
X(CIIj)>CI
TL-1079
Carbamie acid, X-(2-chloro-
C’jHs
Sc.O.
Mice
SO
0/2
ct h vl )-X -et h vl-4-uit roso-
40
0/2_
thvmyl ester
ocox
20
0/2
A x
—
(CIIihCHj 1 CILCILC1
u™-
—
X/
xo
TL-1080
Carbamie acid, N, X - fciX 2-
OCOX(C 11 jC 1 IjCl ),
Sc.O.
M ice
80
0/2
chloroethyl )-4-»itro-
A
40
0/2
thvmyl ester
20
0/2
— -
l/H-
X/
xo,
TL 900
Carbamie acid, X-(2-chloro-
CjHs
Sc.W.
Mice
80
2/2
e I hyl )-X-ethyl-4-dimet h-
/
40
0/2
ylaminothymyl ester
ocox
20
0/2
—
methiodidc
A x
(ClL)JICf 1 CILCH..C1
1 Jch8
v
X(CIIi)jI
TL-1074
Carbamie acid, X-(2-chloro-
C.H,
Sc.W.
Mice
SO
2/2
ct hy 1)-X -ct hy)-4-dimet h-
/
40
2/2
vlaminothymvl ester
ocox
20
0/2
methochloride
A \
10
0/2
(cir,)2cnf ] cilcilci
A-
-
X(CHj)jCl
— -
TL-104S
Carbamie acid, X,X-6m(2-
ocox(c,n4ci).
Sc.W.
Mice
so
0/2
chloroethyl M-dimelhyl-
A
40
0/2
amiiiolhymyl ester meth-
(C1I,)2IIC| 1
20
0/2
iodide
V"‘
X(C!la)jI
TL-119S
Carbamie acid, X-(2-ehloro-
CjHi
Sc.P.
Mice
80
0/2
—
ethyl)-X-ethyl-3,5-di-
/
40
0/2
inethyl-4-nil rophcnyl
OCOX—C H,C I I, Cl
-
20
0/2
ester
A
—
chJ Ich,
—
X/
xo.
Tabi.k 2, Section X (Continued)
SECRET 234
A ROMATIC CARRAMAXES
Code Name
Structure
Houle
and Dose
solvent Species mg kg F fleet
TL 1075 Carhamic aeid, X,X-6/s(2-
ehloroet hyl )-3,5-dimet hyl-
4-nilrosophenyl ester
t’llj
DC'O\(C1 l.-C 11..Cl)j Set). Mice 80 I 2
/\ 40 0 2
| 20 0 2
K/'"‘
xo
TL-1255 Carhamic acid, N',N-l»i*(2-
chlorocthyl V-3,5-dimethy 1-4- _
nitrophcnyl ester trihy-
drate CH;
OCOXTCIM'IIiCDj 3H;0 Sc.P Mice 80 0 2
/\ — 40 0,2
20 0 2
A
xo.
TL-1413 Carhainie acid, X,X-pcnta-
niet h vlene-3-ot hyl-4-di-
ocoxcdf,.
A
Sc.W. Mice 80 2,2
40 2 2
methylaminoplienyl ester
metliiodide
A"'
X(CHj)jI
. 20 2 2
It) 0 2
5 0.2
TL 1414 Carhainie acid, N,X-penta-
rne t hylcne-3-isopn ipyl-4 -
dimcthylnminophenyl ester
methiodide
OCOXCill,.
X(C».)3l
Sc.W. Mice 0.51 /jDju
(78 F)
TL-1418 Carhamic acid, X.X-penla-
methyleno-3-cyclo|>entyl-
4-dimethylnminophenyl
ester methiodide
OCOXCJLo
or
CUi-t
X(CH»)J
Sc.W. Mice 80 2/2
’ll. 40 2/2
20 1/2
10.J 0/2
•II,
TL 1049 Carhamic acid, X.X-penla-
inethyleiic-4-diinelhyl-
aminothymyl ester mctho- (CI1,)2C11
chloride
OCOXCill,,.
Qr".
x(cn,)3a
Sc.P, Mice 0.36 LDio
TL-968 Carhamic acid, X,K-penta-
met hylene -4-di met hyl -
aminothyniyl ester meth- (CIIs)»HC
iodide
(K'OXCiH,,,
0™-
X(CHj)il
Sc.W. Mice 0.44 LI)ia
TL-777 Carhamic acid, X,X-|ienta-
methylcne-4-dimet hyl-
aminocarvacryl ester
methiodide (CH»)*IIC
OCOXCill,..
0~
X(C1I3),I
Sc.W. Mice 3 2/3
2 0/3
1 0/3
Iv.W. Mice 3 3/5
2 0/5
TL-1106 Carhamic acid, X,X-pen(a-
mct hy lene-3,5-d i rne 1 hyl-
4-nitrophenyI ester
CII,
OCOXCill, „
6"-
xo.
Sc.P. Mice 80 1/2
40 1/2
20 0/2
10 0/2
Tahi.k 2, Section \ (Continual)
SECRET CHEMICAL STRUCTURE \ X l> TOXICITY
235
Table 2, Section X (Continued)
Route
and
Dose
Code
Name
Structure
solvent
Species
nig kg
Kffect
TL I2G0
Carlmmic acid, X,X-penta-
OCOXC»H,.
Sc.W.
Mice
80
0 2
mcthvlene-3,5-din»clhvl-
\
40
0/2
4-dimcthylaniinophcnyl
f
]
20
.02
ester hydroiodulc
CIlJ
X(ciij)j m
TL 1465
Carlmmic acid, X-phenvl-
OCOX HC.Hi
Sc.W.
Mice
.40
2/2
3-i-sopropyI-4-dimethyl-
/
\
20
2/2
atliinophcnyl ester meth-
(
1
10
•> o
iodide
I
JcH{CIIi).
—
5
1 /2
\
/
2,
5
0 2
X(C1I.)J
Route and
Kffccls
( ’ode
solvent
Species
(at
various doses)
TL-114S
Sc.W.
0.2
0.3
0.5
1.0
Hal
0/2
2/2
Rabbit
0/2
2/2
-
0. pig
0/2
1/2
1/2
2/2
T I.-1345
Sc.W.
0.025
0.05
0.1
0.15
0.2
tisi sample;
G- pig
0/2
1/2
2/2
Rabbit
0/2
2/2
1/2
4/4
Dog
0/2
1/2
1/2
2/2
Cat
0/2
1 2
2,2
2/2
Monkey
. . . “
0/2
3/3
SB-8
Sc.W.
0.1
0.2
0.3
TL-500
Rat
3/6
6/6
„ .
Rabbit
1/3
2 3
G. pig
1/4
4/5
5/5
Dog
0 2
2/3
3/5
Cat
0/3
2/2
XT. Benzene compounds with one carbamate group and an alkyl side chain having a quaternary ammonium group.
Code
Name
Struct tire
lloutc
and
solvent
Sjiecies
Dose
mg/kK
Kffcct
Til HO
Carliamic acid, X-methyl-2-
dimctbylaniinomethyl-
phenyl ester met hiodidc
OCONHCH,
[ |CII2X(CH,)»I
Sc.
Sc.
Mice
Rabbit
7.2
3.5
ID, o
AK-3!»
Carbamic acid, X,N-dimet hyl-
2-diethylaininomctbyl-
phenyl ester hydrochlo-
ride
\/
OCOX(CH>)t
Iv.
Mice
1.5
/,D,.
All-40
Carbamic acid, X,X-diiueth\T
2-diet hylaminomct hyl-
phenyl ester inethiodide
0(‘0N(CHj)s
[ VlI,X(C,II»W
KJ \n.
Iv.
Mice
0.5
UKu
SECRET 236
AROMATIC CARBAMATES
Table 2, Section XI (Continued)
Code
Name
Structure
Route
and
solvent
Sjieeies
Hose
mg /kg
Effect
T-(?)
Carbainic acid, X-methyl-N-
( X'-me thylcarba myl )-2-di-
methylaminomcthylphenyl
ester methiodide
CH,
OCOX
\
/\ COXHCHj
r jCH*N(CHiM
Sc.
Mice
343
1.D, o
T-20G5
Carbainic acid, X-mcthyl-2-
(1-dimelhylamino-n-pro- —
pyllphenyl ester methiodide
v
OCOXHCH,
/XcilCIIiCH,
\/X(CH,).I
Sc.
Mice
10
t
T-2068
Cart Mimic acid, X-methyl-2-
(1 -diliiethylamilio-n-pro-
pyOphenyl ester hydro-
chloride
OCOXHCH,
/\ciich3cii,
• HC1
Sc.
Mice
40
/.D„
T-1890
Carbainic acid, X-methyI-2-
di met hytaminomet hyl-C-
methylphenyl ester hydro-
cliloride
OCOXHCHi
CH/ |CH,N(CH,),HCl
?
Mice
350
I'Diu
T-1S0I
Carbainic acid, X-methyl-2-
dimethylaminomcthyl-5-
methylpheny! ester hydro-
chloride
_ OCOXHCH,
( ]CIliX(CH,),HCl
c,v -
?
Mice
150
LDt,o
T-1802
Carbamic acid, X-melhyl-2-
dimeltiylaminomet hyl-4-
methylphenyl ester hydro*
- chloride
OCOXHCH,
j^CHjXtCH.VHCI
CH,
?
Mice
140
LDio
T-1893
Carbainic acid, X-methyl-2-
dimcthylaminomethyI-4-
methylphenyl ester melh-
iodide
OCOXHCH.
J^jC'H.X(CH,),T
CH,
?
Mice
75
LDi (,
T-1847
Carbamic acid, X-methyl-2-
(o-dimethylaminoethyl)-4-
methylphenyl ester hydro-
ctiloride
OCOXHCH,
/\ciIN(CH,)i • HCI
O.
CH,
?
Mice
70
IjD$ 0
T-184C
Carbamic acid, X-mcthyl-2-
(o-dimet hylaminoel hyl )-4-
methylpheayl ester meth-
iodide
OCOXHCH,
/\CHX(C11,),I
Oh.
CH,
?
Mice
12
LD.u
T-1821
Carbamic acid, X-methyI-3-
(dimet hylaminomet hyl)-
phenyl ester hydrochloride
OCOXHCH,
(^)t'HIX(CH.)lIICI
Mice
10
LDi0
SECRET CHEMICAL STRUCTURE AND TOXICITY
237
Table 2, Section XI (Continued)
Code
Xante
Structure
Route
and
solvent
Species
Dose
mg/kg
Kffecl
T-1825
Carbamic acid, X-methyl-3-
(dimethylaminomethyl)-
phenyl ester methiodide
OCOXHCHi
(^)cll,X(CUJ)J
?
M ice
7
IDi0
T 1887 ?
T-1939 ?
Carbamic acid, X-methyl-3-
(/3-dimet by laminoet hylV
pbeuyl ester methiodide
1
OCOXHCHi
QcH,CH,X(CH.),I
9
?
Mice -
Mice ca.
100
7.5-10
LD, o
I.D,„
A R 28
Carbamic acid, X-methvl-3-
OCOXHCHi
Iv,
Mice
1.0
T 18-13
(o-diinet hy laminoet hyl)>
phenyl ester hydrochloride
(miotine)
1
V CH,
Iv.
Mice
Rabbit
G. pig
Rat
Mice
0.5
1.0 ± 0.5
1.0 ± 0.5
1.0 ± 0.5
1.0 ± 0.5
Lf}$v
LD* o
T-1894
Carbumic acid, X-methyl-3-
(o-dimethylaminopropyl)-
phenyl ester hydrochloride
1
OCOXHCHi
?
Mice
3.0
LDu
T 1895
Carbamic acid, X-methyl-3-
(o-dimethylaininopropyl)-
phenyl ester methiodide
1
OCOXHCHi
c,h6
?
Mice
5.0
LDm
AR 29
Carbamic acid, X-methyl-3-
(a-dimethylaininoethyl)-
6-meihoxyphcnyl ester
hydrochloride
CHaOj
OCOXHCH,
[^CilXiCHihHCl
CH,
Iv.
Mice
0
h Dsn
AR30
Carbamic acid, X-melhyl-3-
(o-dimethyl:iiiiinoethyl)-6-
mcthoxyphenyl ester
methiodide
CII30|
OCOXHCHi
CH,
Iv.
Mice
5
Uhi
T-1886 ?
T 1938 ?
Carbamic acid, X-methyl-3-
(d-dimet hylaminoel hyl )-
phenyl ester hydrochloride
1
OCOXHCHi ?
f)
1 tcH,CHiX(CHi)j • HCl
Mice
Mice
35
Approx. 3.0
LD$q
LD$u
T-2040
Carbamic acid, X-methyl-3-
(2-dimcthylamino-n-]»ropyl>-
phenyl ester hydrochloride
1
OCOXITCH,
[ | X(CH,),-HCI
N/CHiCHCH,
Sc.
Mice
Approx. 16
Uhl
T-2064
Carbamic acid, X-inethyl-3-
(2-dimclhylamino-n-prcpyl)-
phenyl ester methiodide
i
OCOXHCH,
2CHX(CH.),l
CH,
Sc.
Mice
0.6
LDjri
SECRET 238
\KOM VTIC CARBAM VTES
Route
and
Hose
Code
Name
Structure
solvent
Sjiecies
mg kn
KfTect
T 2038
Carbamic add, X-methyl-3-
(3-dimet hy la mino-n-but yl y
plienvl ester hydrochloride
OCOXHCH,
■A, X(CH3)j - IIC1
So.
Mice
9
Uhu
1
1 JcHiCll.CU
CH,
- -
T '2039
Carbamic acid, N-methyl-3-
OCOXHCH,
Sc.
Mice
1(1
U):, o
(3-dimet hylamino-n-butyl)-
/\ X(CH,),1
phenyl ester methiodide
1 JcHjCHjCH
\/
CH,
—
T-1845
Carbamic acid, N-methyl-4-
OCOXHCH,
7
Mice
00
diniel hvlrminomelhyl-
/\
phenyl ester hydrochloride
o
V
CHiX(CHa)j. HC1
T-1844
Carbamic acid,X-methyl-4-
(a-dimethylaminoct hyl )-
phenyl ester hydrochloride 1
OCOXHCH,
o
f
Mice
25
LD„ o
CH,CHX(CH,), HC1
AR-28a
Carbamic acid, X-me thy 1-4-
(rt-dimethylaminoethyl )-2-
methoxypheuyl ester hydro-
OCOXHCH,
[ pCH,
Iv. “
Mice
1-1.5
Uho
chloride
1/
CH,CHX(CH,V HC1 .
T-1S96
Carbamic acid, X-methyl-4-
(«-dimct hyl ami nopropyl)-
OCOXHCH,
A
?
Mice
300
LDta
phenyl ester methiodide
I 1
—
CH,CH3CHX(CH,),I
T 1834
Carbamic acid, X-methyl-4-
(d-riimet hylaminoct hyl )-
plienyl ester hydrochloride
OCOXHCH,
0
CHiCHiX(CH,)2-HCl
7
Mice
10
LDbn
AR 41
Carbamic acid, X,X-dimethyl-
4- (/3-dimct hylaminoet hyl)-
OCOX(CH,)i
A
Iv.
Mice
15
TJKo
phenyl ester hydrochloride
f 1
C11.CH2X(CH,)jHC1
AR 42
Carbamic acid, X,X-dimethyl-
4-(/J-dimct hylaminoct hyl>
phenyl ester methiodide
OCOX(CH,)i
n
Iv.
Mice
55
UK*
CH3CH3X(CH,),I
T-1935
Carbamic acid, X-methyl-4-
(y-dimet hylaminopropyl y
phenyl ester hydrochloride
OCOXHCH,
0
7
Mice
5-7.5
Uh0
V
CHiCH,CH,X(C H,), • IIC1
Tabi.k 2, Section \I (Continued)
SECRET CHEMICAL STRUCTURE AM) TOXICITY
239
Code
Name
Structure
Route
and
solvent
Species
Dose
mg kg
KtTeet
T-1936
Carbamic and, X-meihyl-4-
(y-dimet hylaminopropyl )-
phenyl ester methiodide
OCOXHCHj
0
9
Mice
Approx. 50
Llh o
T-15)81
Carbamic acid, X-methyl-4-
(7-dirnethylamino-n-butyl)-
phcnyl ester hydriK'hloride
CH,CHiCHsX(CHj),I
OCOXHCHj
n
Sc.
Mice
1(H)
LDu
CHiCIIjCHN(CHi)j- HCl
T-1U82
t 'arbamic acid, X-met by 1-4-
(■y-diinetliylamino-n-lHityl)-
phenyl ester methiodide
CTIi
OCOXHCHj
o
Sc.
Mice
40
LDt0
\/
CI I..CH.(' H X (C H ijal
-
TT.-H15
Carbamicacid, X,X-dimethyl-
3d/3-2-pyndy lothyl Iphcnyl
ester methiodide
CHj
OCOX(CIIj)i
0-0 -
Sc.W.
Mice
0.33
(78 F)_
LD, o
—
i
CHd
T 1827
XII. Benzene compounds
Carbamic acid, X-mcthyl-2,4-
&i«(dimethylamino)phenyl
ester dihydrochloride
with one carbamate group ami two
OCOXHCHj
| |X(CHj)»-HCl
quaternary ammonium groups.
? M ice tit)
LD„
T-1826
Carbamic acid, X-methyl-2,4-
6i«(dimethylamino)phenyl
ester dimethiodide
V
x(cn,VHa
OCOXIICII,
9
Mice
7
LDuo
T 1800
T-1S11
X(CITj)31
Carbamic acid, X-methyl-2,5- OCOXHCIIj
bis( ilimet hylamino)phenyl /\
ester dihydrochloride | |X(CHj),-HCI
(CIIi)>X • IICll 1
9
Mice
50-75
/.Dio
T-1810
Carbamic acid, X-methyl-2,5-
his{ di mcthylam ino)phcnyl
ester dimethiodide
OCOXHCH,
( ]X(CH,).I
(CHj)jXll \
9
Mice
500-1,000
/./Co
A11-27
Carbamic acid, X-methyl-3-
£methyl-(d-dielhylam!iio-
ct hyl )-aminojphenyl ester
hydrochloride
OCOXIICII, Iv.
! In cHiCH;x(c,h,), • iici
Mice
0.1
/.Dso
CH,
-
. “ .*
T\bi.e 2, .Section XI (Continued)
SECRET 240
AROMATIC C VR BA MATES
Code
Name
Structure
Route
and
solvent
Species
Dose
mg kg
IcfTeet
T 1780
Carbamic acid, X-methvl-4-
OCOXHCH,
?
Mice
16
LLK,
[met hyl-{ d-diel hylamino-
/\
ethyl)-amino1phenyl ester
( ]
dihydrobromide
U
CHi—X—CHiCIIjX(CiIIi)j-llllr
X-1779
Carbamic acid, X-incthyl-4-
OCOXHCH,
9
Mice
100
LDm
[methyKd-dicthylammo-
/\
*
et hyl)-amino]phenyl ester
f ]
monomet h iodide
v rij
cifj—x circifxtc.ihh
T-1S33
Carbamic acid, X-methvl-5-
OCOXHCH,
9
Mice
500 2,500
/XL..
dimet hylamino-2-di met hvl-
/\
aminomethylphenyl ester
( VlLXCClhh 1IC1
dihydrochloride - \
XITI. Benzene compound
with one carbamate group and one sidfonium
or arsonium group.
XL-1306
Carbamic acid, X-methyl-3-
OCOXHCH,
Sc.W.
Mice
0.370
LDia
methylthiophenyl ester
A —
—
met humiliate
( i
1 kCH.^SO.CH,
.
XL 1452
Carbamic acid, X-rnethvl-2-
OCOXHCH,
Sc.W.
Mice
so
1/2
dime! hylarsinopheny! ester
A
40
0/2
methiodide
r Wh,),i
20
0/2
XL 1479
Carbamic acid, X,X-dimeth\l-
V
OCOX(CH,)>
Se.W.
Mice
10
2/2
3-diinelhylarsinophenyl
A
5
2/2
ester methiodide
l 1
1
2/2
1 lAs(CH,),I
—
0.5
0/2
XL-1504
Carbamic acid, X-methvl-3-
ocoxiich j
Sc.W.
Mice
1.0
2/2
diet hylarsinopheny 1 ester
A
0.5
2/2
methiodide
\ 1
0.25
0/2
I lAs(CjH5),CH,T
0.125
0/2
XL-1459
Carbamic acid, X,X-dimethyl-
OCX)X(CH,),
Sc.W.
Mice
80
0/2
4-dimct hylarsinophenyl
A
40
0/2
ester methiodide
o
20
0/2
\/
As(CH,),I
XTV
’. Carbamates of naphthalene derivatives.
XT-1096
Carbamic acid, X-methyl-2,4-
OCOXHCH,
Se.B.
Mice
SO
2/2
dinitro-l-naphthyl ester
AA
40
0/2
(Tr
20
0/2
W
xo.
XL-1053
Carbamic acid, X-methvl-1,6-
xo.
Se.P.
Mice
40
1/2
dinit ro-2-naphthvl ester
AA
20
0/2
( Y pcoxcn,
A
10
0/2
HA/
Table 2, Section XII (Continued)
SECRET CHEMICAL STRUCTURE VXD TOXICITY
Code
Route
and
Name Structure solvent
Species
Dost*
mg kg
Effect
T-1889
Carhamic acid, X-melhyl- ?
5,6,7,8-tctrahydro-5-di- /\/\
methylamino2-naphthyl f s f jOCOXIICHj
ester methiodide I I 1
Mice
20
LDbo
X
(CH.).l
T-iaS8
Carhamic acid, N-methyl- /\/\ 7
5,B,7,8-tetrahydro5-di- \ * \ pCOXHClh
melhyhunino2-naphthyl 1 ' J 1
ester hvdrohromide \/ \y
X
(CHjVHUr—
Mice
4.0
/.Dm
TL 110*'.
Carhamic acid, X-melhyl- OCOXIlCIIj Sc.W.
o,fi,7,S-tetrahydro4-di-
methylamino-l-naphthylester I si
methiodide 1 J I
Mice
0.31
(74 F)
ID. o
X(CHS)3I
XV. Carbamates of quinoline and isoquinoline derivatives.
T-1934
Carhamic acid, X-methyl-8- /X/\ 7
quinolinyl ester hydro 1 T 1
chloride III
CIIjXIlCO -HC1
II
0
Mice
Approx. .*>00
LD. o
All-37
Carhamic acid, X,X-dimelhyl- /\/\ lv.
8-quinolinyl ester hydro
chloride 1 1 J
(CIIi)-XCO 1ICI
II
()
Mice
150
LD,o
A H 1S
Carhamic acid, X-rhelhyl-8- /\/\ Tv.
Mice
0.1
LDu
T-(7)
quinolinvl ester methiodide ( T 1 lv.
Mice
10
LDt o
1 1 J Sc.
Mice
90
\y^s/ sc.
CHjXHCO dial (In buffer
|| solution)
O
Mice
31
uh„
AH 3S
Carhamic acid, X,X-dimcthyl- /X /\ lv.
8-quinolinyl ester metho
sulfate
Mice
0.5
ID SO
(CHahXCO ‘|
11 CHaSO.CITa
— 0
—
T 1072
Carhamic acid, X-methyl-1- /X/\ ?
met hyl-1,2,3,4-tet ra hydro
7-quinolinyl ester hydro CHjXHCOl 1 SJ
cldoride || \/'X
o I
CHj-UCl
Mice
30
LD,»
T-11)73
Carhamic acid, X-mclhyl-1- /\/\ 7
methyl-1 ,2,3,4-tet rahydro
7-quinolinyI ester meth- CllaXIlCOl Is!
iodide 11 X/'X
o I
(CH.hl
M ice
0.33
Table 2, Section XIV (Continued)
secret 242
AROMATIC C ARB AM AXES
Houle
ami
Dose
Code
Name
Structure
solvent
Species
mi! kg
Kffect
T 1937
Carbamic acid, \-methvl-l-
?
M ice
Approx. 45
/./> „
met hyl-1,2,3,4-tel rahydro-
1 S I
8-quinulinyl ester meth-
iodide
CII, XI ICO
11
o
Ax^
1
(CHS)2I
T-1970
Isoquinolinc, 2-methvi-
CILXIICO,
V\
s XCHj llCl
V
•}
Mice
20
1,2,3,4-tetiahvdro-5,tt-5(.s-
A
(X-methvlearbamyloxy)
(11, XI ICC)/
J
hydrochloride
I
- /
T-1071
Isoquinoline, 2-met hyl-
1,2,3,4-tel ra h v d ro-5, G-6is-
ClliXHCXL
/\
9
M ice
GO
/,/),„
f X-met hvlearba in vloxy)
CM XHCo/
J" s X(CIL)j 1
melhiodide
T 1968
Isoquinolinc, 2-methyl-
Cl LX 1 ICO- /\
Mice
Approx. 400—
/,/).,0
1,2,3,4-tct ra h vdro-tLZ-
800
bis( X-met hvlearhamyl-
ClLXHCoJ
1 s XCII, I1C1
oxy) hydrochloride
\/
—
T 1909
Isoquinolinc, 2-niethyl-
1,2,3,4-tel rahydro-6,7-
C1LXHCO, /X
9
Mice
>800
/./),„
j * X(CILM
bis( X - met h v Icarha my 1-
ClLXHCoJ
oxy) methiodidc
'v
XVI. Carbamates of aliphatic alcohol derivatives.
TL 1251
Carbatnic acid, 2-(dibutyl-
I(C4IL)5XCILC1LOCOXIL
Sc.W.
Mice
80
0/2
_ amino)-cthyl ester bulo-
40
0/2
iodide
20
0/2
TL 1224
Carbamic acid, X-met hvl-2-
KC.lDACHjCI LOCOX IIC ’1L
Sc.W.
Mice
80
0/2
(dibutylamino)-cthyl ester
40
0/2
butoiodide
20
0/2
TL-1234
Carbaraic acid, 2-(dictfiyl-
1(C-I LLXCH .C1LOCOX1L
Sc.W,
Mice
SO
0/2
amino)-ethyl ester ethio-
40
0/2
dide
20
0/2
TL-1152
Carbamic acid, X,X-di-
1( Cjl DjXCIIjC 1 LOCO X (CH 3 >2
Sc.W.
Mice
80
0/2
met hvl-2-diet hvlamino-
40
0/2
ethyl ester ethiodide
20
0/2
TL 1151
Carbamic acid, X-mct hvl-2-
I(Cj 1LLXCILC1LOCOX I1CH,
Sc.W,
Mice
80
0/2
diethylaminocthyl ester
40
0/2
ethiodide
20
0/2
CIIjC I LOCOX HC11 j
Sc.W.
Mice
80
0/2
TL 1154
Carbamic acid, X-methy 1-2-
xy )-, met hiodide
I Sc W.
CIL(C2IU=XCI1.(IICJLOCOXHCI1j
1
ocoxiicn.
Mice
SO
40
20
0/2
0 2
. 0 2
TL 1514
I lexync, 2,5-6i«(X-me( hyl-
carbamyloxy)-
high melting form
('—CII(Cllj)0( OXIIClIi Sc. P.
I
C (IRCIIdOCOXHCH,
Mice
SO
~ 10
20
0/2
0/2
0 2
TL 1515
Ilexyne, 2,5-bis( X-methyl-
ea rba n ty luxy)-lo\v melt-
ing form
C—CIKCliriOCOXllClU Sc.P.
i -
<' -cii(cn5)Ocoxiicn,
Mice
SO
40
20
0/2
0/2
0, 2
T-{?)
Carlmmic add, X-benzyl-
2-dimetliylaminoetliyl
ester met hiodide
I(CII1)1XCil.( 1 Sc.
Mice
6.25
/-/)» 0
T-(?)
Carbamie aeid, X,X-di-
1 ienzy 1-2-dimet hyl-
aminoethyl ester
met hiodide
>"<0
RCIIjIjXCILCILOCOX
Mice
75
Lt>i„
T (?)
Carbamie add, 3-di-
methylami nopropyl
ester methiodide
l(CHi),XClI;( 'IIjCITjOCOX Hi Se.
M ice
37.5
LDa
T-(?)
Carbamie aeid, 4-di-
met hy laminobutyl
ester methochloride
CKCTDiXTCIDjCHjOCOXTIi Sc.
Mice
12.5
LD&a
T-(?)
Carbamie aeid, 10-di-
mcthylaminodecyl
ester methoehloride
C ’l(CH3)JX(CH,)9CH,0( OX 1 r2 Se.
Mice
75
LD„
T-1090
Carbamie acid, 5-di-
methylaminoamyl
ester methoehloride
C1(CI I,i)5X(CI DiCH/KOX 11, Se.
Mice
20
LDiB
T 1124
Carbamie acid, X-methyl-
4-dimetliylaminol)eiizyl
ester methoehloride
('1( CI I,),X ■ I I.OCOX HCHj Sc.
Mice
79
LDsa
T (?)
Carbamie acid, X-methyl-
2-dimef hylaminoel hyl
ester methoehloride
C1(CH,)3XCH2CHi(XOXIICHj Sc.
M icc
15
T (?)
Carbamie aeid, X,X-di-
met hy 1-2-dimet hyl-
aminoethyl ester
methiodide
KCID.XClIiCHiOCOXtCIU), Sc.
Mice
20
LDin
T-(?)
Carbamie acid, X-ethyl-
2-di met hylaminoel hyl
ester methoehloride
Cl(CH,)jXCI l.CHOCOXIICdL Sc.
Mice
60
LDb o
T- (?)
Carbamie add, X,X-di-
elhyl-2-rlimelhyl-
aminoethyl ester
methiodide
I(('II,),NCI12CHiOCOX(CjI lib Sc.
Mice
42.5
LDi o
SECRET 244
AROMATIC CARBAMATES
Code
Xame
Structure
Route
and
solvent
Sjiecies
Dose
mg kg
Effect
H» H,
I I
T (?)
Carbnmic acid, X,X-penta-
mcthylene-2-dimet hyl-
aminoethyl ester
mcthiodide
- ■ — 1 1
C—C
/ \
I(CHj),XCH;CIIiOCOX CH,
\ /
C—C
1 1 —
Sc. _
Mice
1
LUW
T (?)
T-(?)
T 1003
Carbamic acid, X-allyl-
2-dimethylaminoethyl
ester mcthochloride
Carbamic acid, X-phenyl-
2-dimcthylaminoethyl
ester mcthiodide
Morpholine, X-(d-earba-
myloxyet hyl)-,
methcx'hloride
H, H,
C1(CHi)»XCIIjCHsOCOXIICIIjC1I—■ CH,
Sc.
l(CH,)iXCH,CIW)COXIIC»Hi Se,
Cl
CHr X—CH,CH,OCOXH, Sc.
/ \
CH, CH,
i i
Mice
Mice
Mice
37.5
450
175
LDJV
Uhn
* ‘ .
it
CH, CH,
\ / -
_ O
—
TL-1380
AR-II
All 45
TI.-HOO
XVII. Miscellaneous Carbamates,
Physost igmine salicylate CHa Sc.W.
CHjXIICOOp \ | j|
V'n V
ch, ch,
c,h,o,
Physostigmine salicylate Iv.
Physost igmine mcthiodide Tv.
Ammonium comjmund, snbsti- CHj Sc.P.
tuted di mcthyl-[/J-(X- /\
methylcarbamyloxy)-y-(3,4- 0 j
inethylenedioxyphenyl)pro- / 0
pyl] (3,4-methylcnedioxy- /\S
benzyl) iodide
M i«!C
Hats
G. pigs
Rabbits
Cats
Dogs
M ice
M ice
Mice
0.370
1.500
1.500
1.500
1.2Q0_
1.000
0.800
1.400
1.200
1.000
0.5
0.75-1.0
80
40
20
__ J.l)W
0/2
0/2
0/2
2/2
2/2
2/2
1/2
1/2
0/2
I. IK,
/,/),„
0/2
0/2
0/2
- •—
CII,
----
1
CHjNHCOCH
ii i
■
it i
0 CH,
1
X(CH,)J
CH,
■
0.
\ O
0
\l
CH,
Table 2, Section XVI {Continual)
SECRET CHEMICAL STKI CTI RR AMD TOXICiTV
245
Code
Name
Structure
Route
and
solvent
S|>ccies
Dose
mR/kg
Kffect
TL-1411
Carhamic acid, X-methyl-
II
Sc.W.
Mice
80
0/2
3-di met h via in ino-d-born vl
40
0 2
ester mcthiodide
/VOCOXHCH,
20
0/2
—
1 lx{CIIj)jl
N/
11
- —
AR-43
Carhamic acid, X-melhyl ester
A A
Tv.
Mice
GO
Uh„
of Harmol hydrochlorkle
( i (i
-
1 L 1 Jocoxiicii
cn.
SB 2o
Carhamic acid, X,X-dimcthvl-
0(’()X(CHj)i
Sc.
Mice
120
LD
/t-pyridyl ester hydrochlo-
/\
ride
(1
V
VXHCI
XVIII. Carbamide* and carbazates.
TL-1517
Carbazic acid, 2,2-dimethvl-o-
OC*OXHX(CIIj)i
Sc.W.
Mice
80
0 2
dimethylamino-2-methyl-
A 1
40
0/2
phenyl ester dimethiodide
cu/ ]
20
0/2
1 Jn(CH,)3
Vi
TL-1516
Carbazic acid, 2,2-dimethyl-
OCOXHXfCIIsb
Sc.W.
Mice
80
0/2
5-dimethvlamino-2-methvl-
40
0/2
phenyl ester dihydrochloride
ch/ |
20
0/2
VyxfCH,),
AR 2fi
Carbazic acid, 2-phenyl-3-di-
OCONHXHCtlU
lv.
Mice
0.25
LD.„
methylaminophenyl ester
/\
met hiodidc
[ 1
_ ' -
TT.-1402
U rea, 1 -(4-hy drox v-2,3,o-t ri-
11 X—COXIICH,
Sc.P.
M ice
80
0/2
me thylphenyl)-;4-methyl-
Acn.
- ■
40
0/2
1 1
-
20
0/2
CII\ X'H,
-
on
TL-1401
Benzene, 1,4-bis
CITj—X —COXIICHj
Sc.W.
Mice
SO
0/2
(1,3-di met hyhireido)-
A
40
0/2
()
20
0/2
V
CHj—N—CONHCH*
Tabus 2, Section XVII (Continued)
SECRET Chapter 1I
MISCELLANEOUS COMPOUNDS PREPARED OK EXAMINED AS
CANDIDATE CHEMICAL WARFARE AGENTS
Marshall dates
ill INTKODUCTION
1.\ table 1 of this chapter m grouped all those
compounds which for one reason or another have
not. l«>en subjectod to detailed toxicological examina-
tion. Wit.li the average example, these substances
showed insufficient toxicity to he seriously considered
as chemical warfare agents, although other consider-
ations, such as limited availability, or
lack of means for tactical employment have influ-
enced decisions to abandon exploration of some com-
pounds or the classes to_\vhieh they belong.
Several of the compounds included or classes cov-
ered have I>een treated in other chapters of this vol-
ume. For example, cadmium, cadmium oxide, other
cadmium compounds, some selenium derivatives, and
several metallic carbonyls form the subject of Chap-
ter 11. The tabulation of this chapter is intended to
supplement such chapters by including references to
the preparation and screening of the less promising
members of such classes for the sake of completeness.
Although a number of compounds examined by the
British have been included in the tabulation, no at-
tempt has been made to give comprehensive coverage
to British screening tests, since such systematic lists
are provided elsewhere.**
Perhaps worthy of mention in passing is the .sub-
stance diehloroformoximo (“phosgene oxime”). It
was examined in this country ljecau.se intelligence
reports and published literature indicated that some
attention had been paid it, by the Germans and per-
haps by the Russians. Diehloroformoxime possesses
marked irritating action against skin which is mani-
fested by an immediate burning sensation and the
production of blisters. For this reason, the substance
has Ik'Cii proposed as a “nettle" gas, but its limited
stability, relatively low toxicity, and difficult prepa-
ration preclude serious consideration of it as a chem-
ical warfare agent.
Diehloroformoxime exists w hen pure as a colorless
solid of nip 39 40 C. It boils at 129 C w ithout decom-
position at atmospheric pressure*3 and at 47 49 C
at 23 nun 42:1 and is soluble in water and in organic
solvents. It is rapidly destroyed by alkalies and is
slowly hydrolyzed by water.®* It possesses a pene-
trating and unpleasant odor and attacks (be mucous
membranes and the eves severely.** The substance
appears to lie reasonably stable, when pure and kept
from contact with moisture ■’*'■** or when stored in
anhydrous ether solution 60 but crude material rap-
idly decomposes on standing.44*
Three distinct methods of preparation are de-
scribed in the open literature:
1. The reduction of triebloionitmsomethane by
hydrogen sulfide or aluminum amalgam,64
2. The action of chlorine on fulmintc acid *' or on
mercury fulminate,*2
3. The chlorination of chloroisonitrosoaeetone.**
The first anti third of these methods have been
briefly examined by investigators under Division 9 of
(lie National Defense Research Committee £NDRC]
with disappointing results.42l 44b The first gave rise to
unspecified yields of material of poor quality which
decomposed in less than a day; the
second gave only 30-40 per cent yields of crude ma-
terial. The material which was examined physiologi-
cally by the University of Chicago Toxicity Labora-
tory melted below 35 C, and it is doubtful whether
a pure sample of diehloroformoxime has been pre-
pared or exanrned in this country.
The chlorination of fulminic acid salts has been in-
vestigated briefly in England.*** The yields obtained
(24 45 per cent) did not approach those claimed by
Birckonbach and Sennewald.61 It was found that
twice reerystallized material is considerably more
stable than distilled material and can l»e stored for
several weeks without undergoing appreciable de-
composit ion.
The related dibromoformoxirne has also Ijcen pre-
pared and screened for toxicity.511’ It is less toxic than
the prototype.
SECRET MISCELLANEOUS COM POL NDS AS CHEMICAL AVAR FAKE AGENTS
Tahlk I. Miscellaneous compounds prepared or examined as candidate chemical warfare agents
The compounds in Table 1 arc arranged in two large groups;
(1) derivatives of heavy metals; and (2) miscellaneous
organic compounds Within the heavy metals group, the compounds are classified according to the periodic
group of the
mi
tal, and, among each group of the periodic tahlc, according to
increasing atomic number.
Tite miscellaneous organic
compounds have liccn arranged according to the Heilstein system.
The following abbreviations are used: refractive index at1 C; sjiccific gravity at t,
C in reference
to water at
hi
nip, melting point in (4; hpJ', boiling jxdnt in C at p mm lig;
vp', vapor pressure in mm
Ilg at 1 C; vol', saturation
concentration (volatility) in mg 1 at I C; and dec. p., dcconi|M)sition point.
llrilish reports concerned with those compounds marked hv at
asterisk are not all available in this country.
Centigrade scale is used throughout the table.
Reference
Refer, to
to
Physical projicrties
toxicity
Compound synthesis
Property
Reference
data
1.
Cupric (luoroaeelate 52
51
2.
Cupric 2,4-dinitrols‘n/eiiearsonale 40a
3.
Cupric 2,4,fi-(riiiitrobenzenearsonatc 40a
... ....
—. . ,
t;
Silver nitRite Commercial
24
5.
Zinc fluoborate 40r
24
fi.
Zinc fluosilicate 40i|
24
7.
Strontium fluolroratc 40r
24
8.
Strontium fluosilicate 40r
24
it.
Cadmiumt Commercial
... - ....
See ('hap. 11
10.
Cadmium fluoridet 40o
—
21
11.
Cadmium chloridet Commercial
24
12:
Cadmium uitratef Commercial
24
13.
Cadmium osidef
24
14.
Cadmitiih sulfidef Conimercial
24
15.
Cadmium sclenide 22
24
Hi.
Cadmium selenite 40q
24
17.
Cadmium selenatet 22
... ....
... *
24
18.
Cadmium phosphite
24
19.
Cadmium phosphate!
24
20.
Cadmium fhiol>ora(et lOp
24
21.
Cadmium fluosilieatcf 40q
dcc.p. Approx. 100“
40q
24
22.
Cadmium lactate 6
23.
Cadmium butyrate .5.
24.
Cadmium caproatc 6
25. Cadmium palmitate fi
. . _
20.
Cadmium oleale fi
27.
Cadmium stearate (j
...
28.
Cadmium naphthenate 6
29.
Cadmium oxalate 6
30.
Cadmium malonate fi
31.
Cadmium maleate fi
. ; .
32.
Cadmium fumaratc 6
33.
Cadmium succinate 6
34.
Cadmium malate fi
35.
('admium tartrate - - - 6
30.
Cadmium glularale 6
... ....
37.
Cadmium adipate 6
...
38.
Cadmium mucatc fi
39.
Cadmium citrate 6
40.
Cadmium chelate of acetylacctonc 0
dcc.p. 280-285°
6
41.
Cadmium enolate of ethvl nitromalonate 0
42.
Cadmium salt of nitrated oxidized starch Commercial
24
13.
Cadmium salt of 2,4-din it rophenol — 6
... ....
44.
Cadmium picrule 0
dec.p. 250°
fi
45.
Cadmium chelate of dinitroresorcinol fi
40.
Cadmium styphnate fi
47.
Cadmium m-nilrobonzcncsiilfonate fi
....
48.
Cadmium 2,4-diiiitrohenzencsuifonatc 0
Cadmium p-nitrolicnzoate fi
49.
50.
Cadmium 2,4-dinitrolienzoate 6
...
51.
Cadmium 3,5-dinitrobenzoate 6
t These compounds are discussed more fully in Chapter 11.
SECRET 248
MlSGEULANEOl S COMPOUNDS \S CHEMICAL WARFARE AGENTS
Compound
Before ncc
to
synlliesis
Physical projH'rties
Property Hcferencc
Refer, to
toxicity
data
52. Cadmium 2,4,6-trinitrolicnzoatc
6
53. Cadmium chelate of salicvlaldchvdc
6
mp
>300
0
54. Cadmium chelate of salicylaldoxime
6
mp
>300°
6
55. Cailinium salicylate
6
dcc.p.
280° 290°
6
56. Cadmium 3-nil rosalicylate
6
57. Cadmium 5-nil msalicvlate
6
58. Cadmium pbthalate
40k
50. Cadmium o-nitrocinnamate
6
(iO. Cadmium /«-nitrocinnamalc
j «
61. Cadmium p-nitrocinnatnale
/ 8
62. Cadmium s;ilt of hcxanitrodiphcnylamine
f ti
(13. Cadmium o-nitrolieiueneareunate
6
64. Cadmium 2,4-dmitrol)cnzencarsonatc
0
65. Cadmium 2,4,6-lrinitrobeii7.cnearsonate
6
66. ('admium 3,5-dinitro-4-bvdroxybcnzcncarsonatc
6
1)7. Cadmium 3,5-dinil ro-2,4-325°
6
73. Dimethvleadmium
6
i>p -
08-99°
6
74. Diclhvlcadmium
6
bp5"
62 64°
6
75. Dipropylcadmium
6
I'P"
67°
ti
76. Barium fluoboratc
4 Op
mp
>200°
40p
24
77- Barium fluoeilicate
40p
mp
>200°
lOp
78, Barium succinate
....
70. Barium salt of 2,4-dinilrophenol
40c
<80. Barium 3,5-dmitrobenzoate
40e
81. Barium 2,4,6-trinil robenzoate
40c
82. Barium salt of dipicrvlamine
lOc
83. Barium 2,4-dinitrobenzcnearsonate
40a
84, Barium 2,4,6-trinilrobcnzcncarsonatc
40a
85. Barium 5-nitro-2-furoate
10c
24
86. Barium 5-nit ro-2-furvlacrvlale
40e
87. Mercuric chloride
Commercial
24
88. Mercuric fluoroacelatc
52
51
80. Mercury salt of nitrated oxidized starch
Commercial
24
00. Mercuric 2,4-dinitrohenzcnearsonate
40a
01. Mercuric 2,4,6-trinil robcnzcncarsonatc
40a
02. Chlorovinylmercuric chloride
40j
24, 33
03. Butvlmercuric iodide
40e
04. Butvlmercuric hydroxide
40c
05. 2-( ’hloromercurifuran
28
mp
151-152.5°
28
24, 33
06. 2,5-( IJicliloromercuri)furan
28
24
07. 2-Chloromcrcurithiophenc*
28
mp
183-184°
28
24, 33
08. 2,5-f>is(Chloromercuri)thiophene*
00. Difurylmercury
40c
IO
CO
CO
100. Thallous fluoride
15
bp
298°
15
24
■ _ —
mp
288“
15
101. Thallous fluoborate
15
i»p8
300“
15
24
102. Thallous selenite
4 Or
103. Thallous fluosilicate
15
bp*
340°
15
24
104. Thallous ethoxide
15
, , .
105. Thallous /3-chIoroethylmcrcaplide
40p
mp
>300°
4 Op
106, Thallous formate
15
107. Thallous acetate
15
24
108. Thallous fluoroacelatc
52
51
100. Thallous trifluoroacetale
40p
mp
116-110°
lOp
24
110. Thallous salt of ethyl nitromalonale
40c
Table 1 (Continued)
SECRET MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
249
Table 1 (Continued)
Compound
Hefercnee
to
synthesis
Physical pro|H‘rlies
Property Reference
Refer, to
toxicity
data'
111. Thallium sail of nitrated oxidized starch Commercial
24
112. Thullous lienzoate
15
113. Thallous /j-nilrolx'iizoate
15
114. Thallous 2, 1-dinit robenzoate
15
115. Thallous 3,5-dinitrobenzoate
15
110. Thallous /H-trifluorometbvlbenzoatc
40p
24
117. Thallous salt of 2f4,6,2',4',6'-hexaaitrotiiphenyl-
amine
15
118. Thallous furoate
15
1 111, Thallous o-nitro-2-furoate
15
120. Thallous 5-nil ro-2-fnrylacry late
15
...
. r.
121. Thallous N - me t h v Id i t h iocarba mate
15
122, Thallous X,X-dimelhvldi(hiocarbamale
15
mp
121-125°
15
123. Thallous X-ethvldithiocarbamatc
15
121, Thallous X-isopropyldit hiocarhama te
15
125. Thallous X,X-diethy!di(hiiK'arl>amate
15
bp""1
190°
15
120. Thallous X-butvldithiocarbamate
15
'HP
110 111°
15
137, Thallous X,X'-diisoj>ropyldilhioearl>amate
15
....
128. Thallous X-cvcIohewlilittiioearliamate
15
-v . . •
129. Thallous X, X-dibit tyldilhiocarba male
15
bpo.oi-ae
230-235°
15
130. Thallous X,X-diisobutvldithiocarbamato
15
nip
mp
75 77°
165-105.5*
15
15
131. Dimcthylthallium fluoride
15
....
132. Dimethylthallium iodide
15 —
....
133. Dimethylthallium hydroxide
15
131. Dimethylthallium fluoborate
15
mp
303°
15
135. Dimethyllhalliuni fluosiheate
15
nip
>300°
15
130. Dimethylthallium cthoxidc
15
24
137. Dimcthylthallium ethylmercaptide
24, 33
138. X-Dimethyllhallium dimethylaminc
40i
...
139. X-Dimelhylthallium diethylamine
40i
110. X-Dimethylthallium dibutylamine
40!
141. Dimelhvlthallium acetvlacetone
15
mp
214 215°
15
142. Dimethylthallium cthvl acctoacetate
15
nip
128-130°
15
143. Dimcthylthallium trifluorohexoylacetone
2 4, 33
I It, X-Dirnethvlthallium mellivlaniline
4 Of
....
145. Diniethyllh.allium salieylaldchyde
15
mp
200°d
15
140. Dimethylthallium X,X-dicthyldithiocarbamate
15
bp1
130°
15
24, 33
117. Dimethylthallium X,X-diLsopropyIdithioear-
15
bp4
bp*
138°
130°
15
15
24, 33
bamate
bp55
115°
15
.. . ’ •
mp
150°
15
148. Dimcthylthallium X.X-dihutyldilhiocarhanmte
15
bp"-*
147 148°
15
149. Dimelhvlthallium X,X-diisobutyldithioearbamatc
15
bp»»
104-105°
15
24
150. Dicthylthallium bromide
15
mp
73-74
15
151. Diethylthallium ethoxide
15
152. Dietlivltballiuin Irifluoroaeetate
15
mp
233-235°
15
153. Diethylthallium aectvlacctone
15
dcc.p.
240°
15
154. Diethylthallium benzoylacetone
40*1
155. Diethylthallium thioacetale
15
mp
isi-i83°
15
150. Dipropvllhalliuin ethoxide
15
157- Dipropyllhallium-d-camphor-l 0-sulfonate
15
15
158. Diisopropylthallium chloride
15
mp
150°d ■
15
15
159. Dibutvlthallium fluoride
15
mp
22t>-230°
15
15
100. Dibulvllhallium chloride
15
mp
240-245°
15
15
161. Dibutvlthallium bromide
15
mp
245-250°
15
15
102. Dibutvlthallium iodide
15
mp
220-225°
15
15
103. Diisoamylthallium acet vlacetone
40d
104, Diphenylthalliuin chloride
40(1
— , . .
SECRET 250
MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
Table 1 (Continued)
Reference
Refer, to
to
Physical projwri ics
toxicity
Conqmuml
synthesis
Property
Reference
data
165. Diphenylthalliuin iodide
15
mp
>300°
15
166. Diphcnylthallium hydroxide
40.1
167. Difuryllballium fluoride
15
mp
235 210.1
15
. , .
168. Difurvlthallium iodide
15 —
mp
238-240°
15
166. TctrainethylKermaniimi
24
170. Stannic 2,4-dinit rolicnzenearsoiinte
40a
171. Stannic 2,4,6-trinitrobenzcncarsonate
40a
172. Rutvltin triiodide
2
bp‘
154°
2
2
173. Dipropyltin dibromide
2
bp"1
112°
2
2
«
mp
49-50°
2
2 ~
174. Dibutyltin diiodide
2
bp55
1572
2
2
175. Di-terl-butvltin dibromiile
2
bpu
128° —
2
2
176. 6is(2-Pyridyl)tin bromide
10f
... -
177. Trimethyltin bromide
2
bp*
46-47
2
— 2
17S. Trimethyltin hydroxide
2
sublim.p. 105-108°
2
179. Triethvllin hydride
2
bp*-
36°
2
180. Triethyltin bromide
2
1'P
216-217°
2
2 24
181, Tripropyltin hydride
2
bP*
t>5‘
2
21
dt*
1.1452
2
182. Tripropyltin bromide
_ 2
bp*
123°
2
2, 24
183. Triisopropyltin bromide
2
bp1
79°
2
184. Triisopropyltin iodide
2
bp4
108 no
2
2
185. Tributyltin hydride
2
bp*
115°
2
24
d,n—
1,108
2
186. Tributyltin chloride
2
bp1 5
119°
2
2
200°
40q
24
209. Ix'ad salt of nitromethane
1
210. Ixad salt of nitroaminogiianidinc
1
211. la-ad salt of dinitrotartaric acid
1
212. Ix-ad-w-nilrolxuizenesulfonate
1
213. Ix>ad 2,4-dinitrolx-nzenesnlfonate
1
214. Lead lienzoate
I
215. Leaad m-nitrolienzoatc
1
217. Ixad p-ni(robenzoate
1
218. I>ead 2,4-dinitrobenzoate
1
219. Ijead 3,5-dinitrolienzoate
1
....
...
SECRET MISCEI.I. VN KOI S COMPOUNDS VS CHEMICAL WARFARE AGENTS
251
Tabi.E I (Continued)
Hefereiex1
Refer, to
tft
Physical properties
toxicity
Compound
synthesis
Property
Reference
data
220. I/oad 2,4,6-trinitrohenzoate
1
221. I*'ad sail of p-nitrophcnylhydroxamic acid
1
222. Load salt of m-phcnylenedinitroamine
1
• ...
223. I.ead «-nit rolrenzenearsoimle
1
224. Ijcad m-nit rolienzenearsonatc
1
225, Ix-ad 2,4-dinit rolrcnzenearsonatc
. . *
220. Iz>ad 2,4,6-trinitrohenzcncarsonnte
I.
227. Ix'ad 3-nit ro-4-hvdroxvbenzcnoarsoimlc
1
228. Iz*ad 3,5-dinitro-l-hydro.\vhen2encarsonate
1
....
220. 1 .cad 3,5-dmil ro-1-aminobcnzenearsonate
1
..:—
230. Iz\id 5-nitrofuroate
I
231. Iz-ad 5-nit rofurvlacrvlate
1
....
232. Diethvllead dinitnUe
1
233. Diethvllead scionito
16
mp
>286"
16
234. Dicthylh'ad fcis(p-chlorolx'nzoale)
16
mi)
18.551
16
235. Dicthvllcad hix(»«-hromohenzoate)
16
nip
178 179 d
16
—.,.
230. Dietlivllcad b/s(m-nilrolxMizoate)
16
mp
179 18051
16
237. 1 )ict livllcad b/s(/>-tohiate)
16
mp
18651
16
238. Diet livllcad hiM N -lint vlant hranilatc)
16
mp
169 109.551
16
230. Dicthvllcad dinicotinatc
16
mp
14351
16
240. Dicthvllcad dithioacetatc*
56a
mp
84.5-85°
56a
241. Dihutvllcad dinitratc
1
242. Diphcnyllead dinitratc
1
243. b/«(»»-Nitropheiiyl)lead dichloridc
1
211. bis{ m-N ilrophcnyl)lcad dibromidc
1
215. hix( i/i-Xil rophenvl )lcad diiodide
1
246. 6i*(m-Nitrophenvl)lead dinitratc
1
....
217. his(m-Nitrophcnyl)lcad oxide
1
....
248. 'Primetlivllcad p-tolucncsulfonalc*
— .
249. Trictlivllcad thiocyanate*
16
mp
26.5 27°
16
24, 33
250. Triethvllead sclcnocyanate*
16
mp
33-34°
16
251. Trict livllcad nitrate
I
252. 6/s(Tricthv!lcud) fluosilicate
*
24
253. Triet hyl-p-chlorot hiocthoxylead
40m
24
254. Triethvllead fluoroacetate
56e
mp
180.5°
56e
55c
255. Triethvllead a-chlorocrotonale
16
mp
153 155°
16
256. Triethvllead acid oxalate
16
mp
>300°
16
257. bi,s(Trietlivllcad) oxalate
16
mp
>300°
16
258. hiM Trict livllcad) fumarate
16
dcc.p.
165°
10
259. b(s(Trictlivllcad) adipate
16
mp
>360°
16
260, f>is(Ti iothyllcad) d-eamphoratc
16
mp
>310°
16
16
261. (rt.s(Triclhyllcad) citrate
16
nip
>350°
16
262. Triethvllead m-chlorol>cnzoatc
40d
263. Triethyllcad p-chlorobenzoate
16
mp
123 124°
16
264. Triethvllead o-hromolienzoatc
16
mp
134-135°
16
265. Triethvllead w-bromol>enzoate
16
mp
113-11-1°
16
16
266. Triethvllead p-bromobenzoate
16
mp
127-128°-
16
267. Triethvllead o-iodolrenzoatc
16
mp
138.5 139"
10
268. Triethvllead m-iodolienzoate
16
mp
135-1.36°
16
269. Triethvllead /Hodolienzoatc
16
nip
129.5-130.5°
16
270. Triethvllead <>-nitrot>enzoate
16
mp
142-14351
16
16
271. Triethyllcad w-nitrobenzoate
16
mp
1 72 I73°d
16
272. Triethvllead p-nitrolienzoate
16
mp
1 67-168.551
16
273. Triethvlleadsalicvlatc
16
mp
75-76°
16
274. Triethvllead p-anisate
16
mp
97 98°
16
. . .
275. Triet livllcad /eaniinobenzoatc
16
dcc.p.
265°
16
276. Triethvllead p-aminolienzoate monohydralc
16
mp
84-86°
16
277. Triethvllead X-inethvlanlhranilatc
16
mp
132.751
10
278. Triethvllead N’-phcnvlanfhranilatc
16
mp
124.5-125°
16 .
279. Triethvllead phcnylaectatc
16
nip
96-97°
16
280. Triethyllcad /r-aminophcnylaeetatc
40d
281. Triethyllcad phcnylpropiolate
16
mp
149-15051
16
16
SECRET 252
MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
Compound
Reference
to
synthesis
Physical properl
Projierty
ics
Reference
Refer, to
toxicity
data
282. Tricthvllead cinnamate
10
mp
122 123°d
16
283. Triethvllcad /4-lie nzoylacry late
16
mp
139 110(1
16
284. Triethylload 9-fluorcnccarl>o.\ylate
10
dcc.p.
208°
16
285. Trie thy Head /3{3-naj»hthoyl (propionate
16
mp
134 135°
16
16
280. Tricthvllead diphcnylacetale
16
mp
164-165“
16
287, Trielhyllead triphcnylacetatc
16
mp
I34-136°d
16
288. Triethvllcadsulfanilamidc
56b
mp
171°
56b
56b
289. Triethvllcad furcate
16
mp
156-157°d
16
290. Tnethylleail furylacrylate
16.
nip
132133 d
16
291. Tricthyllcad lepidino2-eai lioxylale
16
mp
153 155“
16
16
dcc.p.
197-199'
16
292, Triethvllcad X -c t h vlcarba zi)le-3-earlx>.\yla 1 e
16
mp
19551
16
16
293. Triethvllcad thioacctatc*
56a
mp
44°
56a
294. Tricthyllcad cvclohexvlsulfinatc
16
mp
132-134’
16
295. Triethvllcad p-tcluencsultiimlc
16
mp
86 88°
16
296. Triethvllcad o-tohienesulfonate*
50a
mp
87“
56a
297. Tricthvllead p-toluencsulfonate*
298. Triethvllcad 2-amino-5-tohicncsuJfonate
16
mp
21051
16
299. Triethvllcad imphthalciie-2-sulfonate*
300. Tricthyllcad d-camphor-l 0-sulfonate
16
mp
- 172°
16
16
301. Tricthvllead p-tolylthiosulfonatc
16
mp
109°
16
302. Tricthvllead methanesulfoliamidc*
66b
mp
97“
56b
56b
303. Tricthvllead mcthanesulfonanilidc*
56h
mp
115.5°
56b
56b
304. hi.s(Triethvllead) niethaliedisulfonate
50b
56b —
305. hits(Triethvllcad) methanedisnlfonanilidc
561)
56b
300. Tricthvllead cthancsulfonanilidc
56b
mp
110°
56b
56b
307. Triethvllcad benxenesulfonaraide
55a
308. Triethvllcad p-aminolxaizenesulfonamidc
16, 56h
mp
173 174°
10
56b
309. Triethvllcad o-tolucncsulfonamidc*
56b
mp
133°
56b
310. Tricthyllcad p-toluenesulfonamidc*
. . . ~
56a
311. Tricthvllead p-toluciicsulfoiianilide*
56b
mp
134°
56b
312. Tricthvllead p-tolucncsmlfon-p-ehlonuiilhle
56b
mp
111.5°
56b
56b
313. Triethvllcad p-toluenesulfon-p-bromanilidc
56b
mp
117°
56b
50b
314. Trielhvllead (7carl»oxvl)enzenesulfoiimiide*
561)
mp
135°
56b
315. Tripropyllcad o-tohicnesu donate
56a
mp
87°
56a
310. Tripropyllcad p-tolucncsulfonatc
50a
mp
7:4-74.5“
56a
317. Tricthvllead l-amino-d-naphthalcncsulfonatc
16
mp
238 240°
16
318. Tripropyllcad mclhanesulfonamide*
56b
mp
67°
56b
56b
319. Tripropyllcad hcnzcncsulfonamidc
55a
320. TripropyHead p-aminobenzcncsulfonainide
50b
mp
101°
56b
56b
321. Tripropyllcad p-lohiciicstilfotiainlide
56b
mp
104°
56h
55a
322. Tripropyllead /Mohicncsulfon-p-chloranilide*
5Hh
mp
123°
56b
56b
323. Tripropyllcad o-carboxyhcnzcncsulfoninikle
56b
mp
130°
56b
56b
324. Tributvllcad p-tolucncsulfonate
56a
mp
81-82°
56a
325. Trihutvllead naphthaleiie-2-sulfonatc
56a
mp
08°
56a
320. Triphcnyllead nitrate
1
mp
(sinter)
220-225°
1
327 Tri(»«-nitrophcnyl)lcad chloride
i
.77”
328. Tri(m-nilroplicnyl)lcad nitrate
i
.... —
329. Tel ramet hyllcad"
24
330. Triethvlallvllcad dimer
10h
331. Antimony trifluoridc
Commercial
24
332. Ethyldichlornsl ihine
13
bp'
62-83°
13
24
—
A
2.182
13
333. p-Thiocyanophcnyldichlorost ihine*
334. p-r.lhvlthiophenyldicliloroKt ihine*
335. p-(3-Chlorocthvllhio>phcnvl dichlorostibine*
330. p-Phcnylcncarsincst ihine tetrachloride*
337. l>is(«i-Aminophcnyl)chloroMtihine dihydrochloride
24
338. his( m-Aminophcnvl thydroxystihine
24
339. 5,10-1 lichloro-5,10-dihydrost ibarsan t hrcnc*
340. Diphenyl-o-l hknylst ihine *
Table 1 (Continued)
SECRET MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
253
TabI-K J (Continued)
Compound
Reference
to
synthesis
Physical projjerties
Property Reference
Refer, to
toxicity
data
341.
Phenyhlit hienylst ihim*
342.
Trifury (antimony
lOe
• .V. —
24, 33
343.
/r/*( 5-iert-B u ly 1-2-f ury 1 )anl i mon y
40f
24, 33
344.
(m(2-Pyridyl)antimony
40g
24, 33
345.
Trimelhvlstihine sulfide*
346.
fci«(Trimelhylstil>o)t risulfide*
...
347.
bis( Diphenylstibine)sulfide*
34.8.
Sulfate of 6i.s(m-aminophenyl)hydroxystibine*
349.
5,10-Dihydro-5,10-di< >xys t il >a rsanl hrene-5,10-
monoxide
330.
1 )iphenylbismuth thiocyanate*
56d
331.
//■/*( 2-lurvl )bismuth
4 Of
352.
Chromyl chloride
28
bp
114°
28
24
353.
('hromium hoxacarbonyl
40g
Jc5
1.912
28
354,
('hromium 5-nilro-2-furoate
40c
355.
Tungsten carlxmyl
36
mp
125°
36
35
yp6T
1.2
36
350.
Manganous 2, l-dinitrol)enzenearsonatc
40a
357.
Iron jxmtacarbonyl*
Commercial
See Chap. 11
35.8.
Ferric 2,4-dinitrobenzenearsonatc
40a
.77
359.
Cohaltous fluohorate
24
300.
Salcominc
Commercial
35
361.
Cobalt 2,4-dinitrobcnzcncarsonate
40a
362.
Nickel carbonyl* ~
. . —
See Chap. 11
303.
Nickel fluoborate
40q
24
364.
Nickel fluosilicate
4 Or
mp
>275°
4 Or
24
305.
Nickel 2,4-dinitrolienzenearsonate
40a
300.
Chlorine
Commercial
24, 59
307.
Bromine
Commercial
24, 59
368.
Nitrogen fluoride
— ...
24
369.
Ammonium fluoride
Commercial
24
370.
Lithium hypochlorite
24
371.
Hydrogen sulfide
. . .
mp
-85.5°
59
59
—
bp
-60.3°
59
372.
Sulfur monofluoride*
54, 57
bp
-35°
54
54, 57
_
-99°
mp .
-105.5°
54
...
,u"
1.5
54
373.
Sulfur tetrafluoride*
54, 57
bp
-40°
54
54, 57
374.
Sulfur hexafluoride
...
mp
-124°
54
24
375.
1 hsulfurdeeafluoride
See (’hap. 4
See Chap. 4
376.
Thionvl fluoride*
54, 57
bp
-43.8°
54
24, 57
-
mp
-129.5°
54
377.
Sulfuryl fluoride*
54, 57
bp
-52°
54
24, 57
mp
-120°
54
378.
Salfuryl cblorofluoride
. . .
24
379.
Pvrosulfurvl chloride
...
24
380.
Hydrogen selenjde
bP
-41.5°
59
59
381.
Sodium selenitic
Commercial
24
382.
Selenium monochloridef
59
bp;“
127°
59
59
2.7741
59
nun
1.5962
59
383.
Selenium monobromidet
-
384.
Selenium hexafluoridet
57
sublim.p.
-46.6°
54
57
385.
Carbon snlfidcsclenidcf
. . .
386.
Carlm>ii diselenidet
22
bp
117 118°
22
24
387.
Selenium oxychloridef
23
bp21
84 85°
23
24, 33,59
nip
10.9°
59
nil5”
1.6516
59
t These compound* and other selenium compounds are discussed more fully in Chapter 11.
SECRET 254
MISGEI.I.ANEOl S COMPOUNDS AS CHEMICAL WARFARE AGENTS
t'oni|>ound
Reference
to
synthesis
Physical properties
Property Reference
Refer, to
toxicity
data
388.
Selenium ox v bromide!
38!).
Selenium oxide
Commercial
24
390.
Selenium dioxide!
391.
Chloroselenious acidf
392.
Sodium selenite __
24
393.
Hydrazine hydrate
Commercial
24
391.
Ammonium fluosilicate
Commercial
24
393.
T riehloronit rosomethanc
7
24
3s< ipn >pa ne
49a
bpioo
40
49a
405.
2-Nitrobulene-I
vol3"
32.36
26
24
40(4.
1,4-Dil>romo-2-l mtene
7
nip
54°
7
24, 33
407.
/n‘s( Chloromel hyl )uit romel hane
47a
24, 33
408.
3-Chlon>-3-nit rosopentane
7
bp1'
bo35
44°
1.4190
7
7
24
.
d,3i
1.016
7
409.
Methyl sulfite
...
24
410.
Methyl silicate
7
hpiw
75“
7
24
411.
Dimethyl sclenide
22
bp
56-58°
22
412.
Trirncthylselenonium fluoride
20
rf®
1.378
20
24
«ir"
1.4600
20
413.
Dimethyl tclluride*
414.
Monoehloromet hyl sulfate*
415.
f>/.t(Chloromet hvl) sulfate*
416.
his( Chloromel by I) et her*
417.
6/s(Bromomcthyl) etlier
59, 401
mp
bp
d3"
-34°
154 155°
2.2013
59
59
59
24, 59
418.
fl-ChlorovinylsoIenium chloride*
23
nip
86°
23
24
419.
Ethyl sulfite
24
420.
Ethyl fluorosulfoiiate
24
421.
Ethyl chloroselenite*
422.
Ethyl selenomcrcaptan*
... .
423.
Elhoxysclenyi chloride*
i>p,s
81.5-82.5°
55e
55c
424.
Diethyl sclenide*
22
bpls
79 82°
22
24
425.
Diethyl diselenide*
426.
Diethyl tclluride
427.
(r/jf(/J-ChIoroethyn liorate
40s
lip1
97-99°
40s
24
428.
ltlrnkis(J}-( 'hloroelhyl) orl hosilicatc
21
bp1
142 113°
21
24, 33
429.
/3-Chloroelhvl nitrite
7
l.p’"
33°
7
24. 33
Till30
,P"
1.4115
1.212
7
7
430.
Methyl 5-ebloroethvl sulfite*
... —
24
431.
hi${(}-{'hloroelhyl) sulfite*
...
24, 33
432.
6(x(d-Chlomethvl) selenite*
nip
44-45°
55e
55c
433.
d-Chloroet hylsulfnryl chloride*
12
bp° s
60-64°
12
24
t These compound* and other selenium compounds are dis
■usseti more fully in Chapter 11*
Tabi.k t (Conlinutd)
SECRET MISCELLANEOUS COMPOUNDS AS CHEMICAL AY A K FAKE AGENTS
255
Tabus 1 (Continued)
Reference
Refer, to
to
Physical properties
toxicity
Compound
synthesis
Property
Reference
data
134. f»(s($-Chloroelhyl) sulfate
12
bpos
117-133°
12
24
435. W.P
113-117°
27
24
n ir"
1.4330
27
430. 3-Rrorao-2-propyn-l-ol
27
bp=
49-53°
27
24
MU*"
1.5140
27
440. 3 Iodo-2-pmpvn-l-ol
27
bp5
82-85°
27
24, 33
mp
40-43°
27
44 U Methyl 2-propynyI ether
27
bp
01 65°
27
24
/(u'4
1.4052
27
4 12. Methyl 3-bromopropynyl-2 ether
27
bp1*
34-38°
27
21
air*
1.4703
27
443. 3-Chloroallvl alcohol
24, 33
114. Allyl methyl ether
. . .
24
445. si/m-Dichloroisopropyl eblorosnlfinatc —
32
l>l>15
108 110°
32
24, 33
«w*°
1.5130
32
-
—. . .
J--a
1.432
32
446. 2-Xilro-l-butanol silicate
40b
....
4 17. Ethinyldimcthylvinyl carbinol
Commercial
24, 33
448. 2-Butyne-l ,4-diol
Commercial
24
440. bi.x-0-ChloructhyI formal
12
bp1-
02-04°
12
24
450. Mcthylformylchloride oxime
19
bp
63-66°
19
24
mp
-64 to —60°
19
»Da
1.4193
19
— —
—
d*
1.135
19
451. Acetaldehyde a zinc
7
bp
96 08°
7
3, 24
«ua
1.4370
7
452. i lemiaoetal of chloral and chlorctonc
471.
mp
68-69°
47b
453. Chloral oxime
7
bp*9
69 70°
7
24, 33
7lu“
1.4905
7
•
d,a
1.571
7
454, Acrolein
Commercial
24
455. Propionaldehyde azine
7
bp
139-141°
7
3, 24
n ir5
1.4497
7
456. Acetone azine
7
bp
120-133°
7
3, 24
nu4
1.4511
7
457. 6jx(Selenoacelone)*
....
458. Chloroacetone oxime
7
bp*
70-71°
7
24
. , .
n
1.4777
7
1.221
7
459. Bromoaectonc
21
bp13
35.5-36.5°
21
24
400, Butyraldehyde azine
7
bp17
77-78°
7
3. 24
«na
1.4504
7
461. Methylethylketone azine
7
bp-»
71 72°
7
3, 24
n u’5
1.4517
7
462. l-Bromobutanonc-2
21
bp30
62-66°
21
24
«n15
1.4700
21
. . .
463. 3-Bromobutanone-2
21
bp-
49 53°
21
24
H,,11
1.4595
21
464. Selenovaleraldehyde *
. . ,
465. Diethylkelone azine
7
bp“
94-96°
7
3, 24
Bn3
1.4539
7
406. 1 - LI romopen t a n one-2
21
467. 3-Bromopent anonc-2
21, 58
bp15
75-76°
21
24
«u!1
1.4576
21
vol-" -
21.59
26
468. a-Chloromesitvl oxide
30
bp31
66-69°
39
24
400. 1-Hy droxy-2-pcntync-4-one
27
bp3
79-83°
27
21
«ir°
1.4587
27
vol”
0.111
26
SECRET 256
MISCELLANEOUS COMPOUNDS AS CHEMICAL W Alt FA HE AGENTS
C oin pound
Reference
to
S) nthesis
Physical pr< >i>ert ies
Property Hi
■ferenec
Refer, to
toxicity
data
470. l-Mcthoxy-2-pcntync-4-one
27
bp3
47-50°
27
24. 33
471. Carbon sulioxide
38
n n!H
1.4462
27
24
472. 1 ,l,4,4-Tetraethoxy-2-butyne
27
»>P:
97-102°
27
24
«iru
1.4346
27
473. Diketone
*»pm
43°
38
24
474. Hydrocyanic acid
See Chap. 2
Sec Chap. 2
47.5. Sodium cyanide
Commercial
...
24
476. Triallvl orlhoformate
24
477. 2-Propynyl formate
27
bP
105-109°
27
24, 33
478. Allvl formate
a,,1'1
1.4203
27
24
479. a, d-Dicblorovinyl acetate
49c
bp11
41-13°
49c
24
480. (3-Triazoethyl acetate
21
bp'-“
74°
21
21
«ie"
1.4345
21
1.123
21
481. Acetyl fluoride
12
bp
20-22°
12
24
482. Acetyl azide
2L
...
483. Acetonitrile-boron trifluoride addition product
39
mp
118-120°
39
24
484. Methyl selenolaeetale
22
iip”
29 31°
22
485. Sodium chloroacetatc
Commercial
bP
112-114°
22
21
486. Chloroacetyl fluoride
49n
bp™
366.2
26
24
-
vol-°
74-76°
49u
487. Sodium bromoacctatc
24
4SS. Itromoa cetyl bromide
Commercial
24. 33
489. Sodium iodoacetate
24
490. Ethyl iodoacetate
40k —
24, 33
491. Propiolic acid
27
bpa
73-77°
27
24, 33
492. Methyl propiolate - —
27
bp
100-102°
27
24, 33
493. Ethyl propiolate
27
»>P
119-120°
27
2 4, 33
494. p-Chloroethyl propiolate
27
bp‘3
n ir“
—79-82°
1.4588
27
27
24, 33
49.5. Allyl propiolate
27
hpeo
70-73°
27
24
— .
«i.lss
1.4378
27
496. Hromopropiolic acid
27
mp
85.5-87°
27
24, 33
497. Methyl bromopropiolate
27
bp“
40 45°
27
24
.
tiua
1.48S4
27
498. Acrylonitrile
Commercial
bp
75.5-76°
24
499. Methyl a-cliloroacrylatc
7
bpin
51-55*
7
500. 0-Chloroacrylonitrilc
»u!0
dt'°
1.4400
1.201
7
7
24
501. o,0-Dichloroacrylonitrile
Commercial
bp®"
58-59°
24, 33
.502. o,d,/3-Trichloniacrylonit rile
48
mp
17 19°
48
24
—
bp710
141-142°
48
503. Ethyl /3-chloropropioniminocstcr hydrochloride
12
rnp
96°d
12
504. o,o,d-Trichloropropionitrile
505. Mcthoxytetrolic acid
24
27
bp’
114-118°
27
24, 33
506. Methyl methoxytetrolatc
27
nij!t
lip’
1.4669
56-58°
27
27
21, 33
507. Crotonyl fluoride
19e
n n5"
bp
1.4438
88°
27
49e
24
508. Ethyl vinylacctifhinoester hydrochloride
12
nip
90 100°d
12
509. Allyl cyanide
24
510. y-Chlorocrotononitrile
49m
bp10
60-62°
49m
24
.511. /8-Chlorocrolononitrile
48
bp7*
125.5 126.5°
48
24
512. Butyryl fluoride
49c
bp
6.5-67"
49c
24
513. Methyl a-chloroisobutyratc
. . .
24
514. a-Triazobutyric acid
21
bp"7
80°
21
24, 33
515. Methyl o-nitro-0-methylerotonatc
49p
nir4
bp5<
1.4536
129-125°
21
49p
Table I (Continued)
SECRET MISCELLANEOUS COM POINDS AS CHEMICAL WARFARE AGENTS
257
Tahi.e 1 (Continued)
Compound
Reference
to
synthesis
Physical proper!
Property
ies
Reference
Refer, to
toxicity
dat a
516. Methyl melhoxyacetate
24
517. Dimethyl diglycolatc
Commercial
...—
24
518. hi*{(}-{ 'hloroelhyl) diglycolatc
7
1>PS
195-199°
7
51ft. Formaldehyde cyanohydrin
24
520. Methyl 2,2,2-trieblorolaetate
Iftq
i.p*
92-94°
4ttq
24
521. Chloralcyanoliydrin
4!th
mp
59-60°
49h
522, d-Cvanoelhyl nitrite
Iftd
cannot be distilled
49.1
523. l-Chloni-l-isoiiitrosoacctone
1ft
mp
108-109°
19
24, 33
524. l-Cbloro-2-methylglyoximc
1!)
mp
183-IS4°d
19
2 4, 33
525. Triehloronectylcyanide.
49g
bp
118-121°
49g
24
526. Vinyl mncocblurate
Commercial
vol20
0.289
26
2 4. 33
527. Hexaehlorodimethyl oxalate
4fth
mp
79-80°
4ftb
24
528. Oxalvl fluoride
10
i>p
approx. 2 3°
10
24
52ft. Oxalvl chloride
12
bP
64-65°
12
24. 33
530. Methyl cyanoformate
42c
bp
98 9ft
42e
24
531. Chlorocyanofomuddosimc
42.1
bp5
53-549
42d
24, 33
mp
54 56
4 2d
532. Cyanogen
....
7T.
24
533. Diethyl dichloromalonale
17
i»pu
115-116°
17
24
no54
1.4386
17
534. 1 Wet hv! ethoxvmethylenem.-donate
47c
1.4600-1.4620
47c
24
535. Ktlioxymethyleneinjilononitrile
4ftj
mp
60-63°
49j
24
536. Diethyl diethoxymcthvlmalonatc
47d
bp1*
133°
47d
24
n u2«
1.4220
47d
537. Dimethyl acetylene* 1 icarlsixylate
4ftc
bp2"
1.17
26
24. 33
vol29
98-99°
49o
538. Diethyl acetylenedicarboxylate
27
bp3
84-88°
27
24, 33
-- — . . .
nr.2"
1.4435
27
53ft. h/xf $-(' h loroe t hy 1) acetylenedicarboxylate
27
bp*
175 21551
27
24
nu1'5
I..5004
27
540. Diallyl acetylenedicarboxylate
27
bp*
112-118°
27
24
■ -
tin" ‘
1.4718
27
541. Diisopropyl acetylenedicarboxylate
27
bp*
103-107°
27
24
*
...
nDlil
1.4408
27
542. f)/s(2-Ethylhexyl) acetylenedicarboxylate
.... ""
24
543. Dimelliyl maleate
27
bp
199-204°
27
24, 33
544. Dimethyl fumaratc
bp
189-192°
27
24, 33
545. Diallyl fumaratc
27
bp2"
137-140°
27
24
546. Fumaryl chloride
48
bp'3
62 63°
48
24
517. Dimethyl chloroinaleale
24, 33
548. Diethyl bromomalcate
49g
i.P°‘
85-86°
49g
24
54ft. Diethyl chlorofnmaratc
• 27
i>p5u
137 139°
27
24. 33
550. Chlorofumaronitrile
Commercial
24. 33
551. Diethyl bromofnmarale
27
bp8
120-123°
27
24
552. Chlorofnmaryl chloride
27
bp21"
140-143°
27
24
553. Dimethyl dibromomaleate
17
bp"
134-136°
17
24
554. Dicthvl «,a'-diehlorosuccinate
17
bp*
106 108°
17
24
555. Dimethyl «,o'-dibroniosiiccinate
. 17
mp
00-61°
17
24
556. Dimethyl tctrachlorosuccinatc
24
557. Dimethyl «,«'-dicbloroglntarate
17
bp1
95-96°
17
24
558. Dimethyl or.o'-dichlorondipate
17
bp2
126 128°
17
24
«rr*
1.4660
17
55ft. fusCTriehloromethyl) carbonate
24
560. Methyl ft-chloroelliyl carlxmate
24, 33
561. 6i«(£-CbIor.s‘thyl) carbonate
12
bp11
119 122°
12
24
562. Methyl fluorocarlsinale
4ftg
bp
43-15°
49g
24
503. Methyl chlorocarbonatc
24
564. Trichloromethyl cblorocarlsmate (diphosgene)
See Chap. 3
See Chap. 3
.565. Ethyl chlorocarbonatc
...
24, 33
566. 0-Chloroethyl cblorocarlsmate
7
bp™
152.5°
7
24
»i>t0
1.4465
7
d.20
1.3825
7
SECRET 258
MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
Table I (Continued)
Compound
Reference
to
synthesis
Physical pro|>erl ies
Property Reference
Refer, to
toxicity
data
Allvl cbloroearbonate
40f
bp
107-111°
40f
24
.'>68. Methyl triazoformate
4!li
bp
97-101°
40i
\ 24
560. Carlsmvl chlon iflnoride
See Chap. 3
See Chap, 3
570. Carbonv] chloride (phosgene)
8ee Chap. 3
See Chap. 3
571. X.X-Dichlorourelhanc
40d
24, 33
572. Dimethyl azoformate
7
bp“
08°
7
24
bpe
104“
7
«u:*
1.4180
7
t/.,rn
1.222
7
573. 6is(/3-Chlorocthyl) azoformate
7
bp'
140-143°
.-z- 7
33
(Id"
1.4752
7
•IF
1.300
7
574. Cyanogen chloride
See ('hap. 2
See Chap. 2
575, Diehloroformoxime (phosgene oxime)
42a, 56c, 62,
bp-1
47MO°
42a
21, 55b
64, 66
1V1|)
30 10
63
—
- -
bp
129°
03
576. Cyanogen bromide
Commercial
24
.577. 1 >ibromoformoxime
55b
....
55b
578. Methyl chlorothiolformatc
■12c
b|>
Ill 112°
42c
21
' -
—
mr"
1.4001
42c
. . . -
if-'"
1.200
42c
570. Tridden .met hyl chlorothjolformate-
42c
lip-4
153 162°
42c
24
> 1.52
42e
,po
1.654
42c
580. Thiophosgene
42c
bp
73-76°
42c
24
581. Thiocarlsmyl chloride polymer
42c
7-.—
24
582. Acetyl thiocyanate
40h
bp"
60.5 61.0°
40h
24
583. CarlMWiielhoxy isothiocvanate
40h
bp14
58-61°
49h
24
584. Methyl thiocyanate
28
bpJ«
128-129°
28
24
«ua
1.4081
28
•hr'
1.0732
28
585. 2-Chloroethvl thiocyanate
5
— ...
24, 33
586. Hexyl thiocyanate
28
bp1 5
84 87°
28
24
ttu
1.5650
28
.
•hr*
0.041
28
587. Dodecyl thiocyanate
28
bp14
177 170°
28
24
«i>“
1.460
28
—
mct hyl) et her*
. ,T~
24
501. Aceloliyl thiocyanate
40e
....
24
502. Cyanogen sulfide*
, . .
503. Methvl chlon>dithkiformafe
42c
bp13
47 10
42c
24
504. Allvl sclcnourea*
505. 1,3- Diseleiiocyanopropane*
506. Cyanogen diselenide*
5!>7. (3-Chloroethencselcnjnvl chloride*
28
24
508. Elhaneseleninie acid
23
500. Elhaneseleniiivl chloride hydrate
23
mp
72-75°
23
24
GOO. N, \-Dimef hvlformamule
Commercial
24
601. 2,5-6/s(N-Methylearbarnyloxy)-3-he.vyne (two
forms)
35
602. X,X-DimethvIcarl>.imvl fluoride
49k
bp**
05"
19k
24
603. Ihmcthvlc.irbamyl chloride
11
bp
166 168°
II
24
604. Methvl isocyanate
37
bp
37-30°
37
35
605. Dimethylsulfamyl fluoride*
4 On
bp14
48.5°
4 On
24, 55d
606. Dimethvlsulfamvl chloride
11
bp* *
34
II
24, 33, 55d
607. X, X-T )ict h vlchloroacet amide
Commercial
24, 33
608. X,X'-Dicthyloxamidc
n
nip
175°
11
24
600. Ethyl X,X-6/s(j3-chlorocthyl) carbamate
39
bp"
131-132°
39
24
dif5
1.4688
39
,if
1.214
39
SECRET MISCELL VXEOCS COMPOl M)S AS CHEMICAL WARFARE AGENTS
Compound
Reference
to
synthesis
Physical pro [Kit
Pro(>erty
It'S
Reference
Refer, to
toxicity
data
010. \-But vlmaleimide
24
Oil. Dibutvlearbamyl chloride
11
l',p*
108-109°
ii
24,33
012. 4-Chlon>|ieii( vldielhylamine hydrochloride
34
mp
99.5 100°
34
24
013. Kthvlenediamine thiosulfate
24
014. 1,0-llexanedianiine
Commercial
24
015, 1,0-Hexanediol diisocvanate
Commercial
21, 33
010, /1-DimethvlaminoelbvI formate
14
bp7*
125-130'
14
24, 33
n if0
1.4262
14
“ ■
0.905
II
017. d-Dimethvlaniinoellivl acetate
14
bp1*'
148-151°
14
24, 33 —
a u10
1.4178
14
d„r"
0.928
11
618. fJ-MetKvIelhvlaminoethyl formate
14
bp711
117-150°
14
24, 33
.
_
«..»*
1.4287
14
0.919
11
610. /J-Methvletlivlamiinielhvl acetate
14
bp711
162 163°
14
24. 33
«!!*"
1.4226
14
dio50
0.918
14
020. Formylcholine chloride
14
mp
144 146°
14
24
021. Acetylcholine chloride
Commercial
24
022. ('arhaminoylcholine chloride (Doryl)
35
023. ti-( N-Met bvlcarbamvloxv )ei hvlt rimet hylain-
mcmium chloride
49b
24.33
024. 0-( X-Pro|*vlcarl*iimyl)ch(>line iodide
24
025. (i-( N-HutvlcarhamyDcholine iodide
24
026. /3-Diet hylaminoethyl formate
14
bp7*
157-160°
14
2 4, 33
n i>so
1.4358
14
0.900
14
027. (J-DiethylaminoethvI acetate
14
bp61
101-103°
14
24, 33
W|.5“
1.4259
14
djo™
0.911
14
028. /S-Diethvlaminocthvl carbamate elhiodidc
29
mp
150-150.5°
29
24
629. 0-Diethylaminoelbyl N-methylcarbamate eth-
iodide
29
nip
90-92°
29
24
030. /l-l)ihul vlarniniK-thyl carbamate but oh slide
29
nip
99.5-100.5°
29
24
631. /J-Dibutylaminoethyl N-mcthylearbamate bulo-
iodide
29
mp
100 101.5°
29
24
032. vdroxycthvl )methylamine
('ommercial
24
033. hylaminoethyl formate
14
i,p7
126-127°
14
24, 33
—
n if"
1.4698
14
1.045
14
034. Mcthyl-l(i*{d-fornio.\yethylfamine
14
bp7
110-111°
14
24, 33
«u20
1.4501
14
-l ti.amy la mi nopropyl X-methylcarbamate
amvliodide
29
nip
78-83°
29
24
611. I-Diethylamino-2,3-6«f(X-mcthylcarbaiiiyloxy)
melhiodide
29
mp
122-123.3°
29
24
642. Ethyl diazoacelatc
21
bp1-
42 43°
21
21
nip"
1.1592
21
043. 1 )imel h vlaminoacet onit rile
49h
bp11
55-56°
49h
24
644. /r/xfd-Thioevanoel hyl famine
...
51 —
<>45. Trifluoromel hvlsilieane
24
610. T rich Ion >mel hvlsilieane
24
647. Dichlorodiraethylsilicane
24
Tabi.k I (Cimtinueil)
SECRET 260
MISCELLANEOUS COMPOUNDS VS CHEMICAL WARFARE AGENTS
Compound
Reference
to
synthesis
Physical projicrties
Pro|*crty Reference
Refer, to
toxicity
data
HIS. Chlorotrimcthylsilicane
24
649. Ethvltrifluorosilicane
. . .
24
650. Kthyltrichloroeilicuno
4 On
bp7*"
96-98°
40n
24
651. Tetracthylsilicanc
40f
24, 33
652. Trifluoropropylsilicanc
24
653. Trichloropropylsilicane
24
654. Trichloroisopropylsilicane
24
655. ButyK rifluorosilicane
...
....
...
24
656. Buty 11rieldorosil icane
. _♦ . .
21
657. Tributvlboron
40a
65S. 1 l/xachlorncvc!ohexane (impure)
Commercial
24
659. rt-Broino-2-chloro-6-uiirotoluenc
Commercial
24,33
600. 2-N'ilro-l-phcnylpropene
18
bp'°
64.5 65.5“
18
mp
139°
IS
24. 33
66!. 1,2-fus(fl-Chlorocthyl)bcnzenc
32
i>p
120-122°
32
24
602, Phenvl chlorocaHamate
24, 33
663. 2,4,6-TrichIorophcnyl chlorocarbonatc
24, 33
664. Picrvl silicate
40n
065. Cvclohoxvl dithiocyanate*
666. o-C’hlorophcnyl thiocyanate*
~ . .
667. m-Chlorophenyl thiocyanate*
668, p-('lilorophcnyl thiocyanate*
669. roacel<»|>lu-ii<>nc
24
702. tt.o-Dichloruacetophcnone
24
703. «-ChIi>n>-f>-nitrosoaci‘(ophcnone
47e
mp
138,1
17c
* - .
704. a-('hh.r(..p-phcnylacctophen(.nc
24
705. Phcnylpropargyl aldehyde
27
i»p”
114 117°
27
24
—
n i>'5
1.0029
27
700. Phcnylpropargyl acetal
27
l.p*3
153 150°
27
24
n [i3
1.5100
27
707. n-Bromopropiophem>nc __
24
70S. Selemicyanoacetophenone*
709. S-Isonitrosocamphor (d)
44e
mp
154-155°
14c
24
— -
bp1"
179°
44e
710. 1,3,5-/ r /.t( C h 1 oroa ce t y 1) lie ll ze nc
32
mp
148 1.50°
32
24, 33
711. 2-MethvH,4-naph(li1 K'nzoylazide
21
mp
07 08°
21
24
713. rt-Hromolien/oylcyanide
59
mp
29°
59
24, 69
in."
132-134°
59
714. Methyl phcnylpropiolate
27
i»p!
04 99°
27
24
715. n-Amyl-X-(-/>-phonylencdi-
lla
...
24
amine
34
mp
178 180°
34
24, 33
SECRET MISCELLANEOUS COMPOUNDS AS CHEMICAL AVARFARE AGENTS
Table 1 {Continued)
Compound
Reference
to
synthesis
Physical proper!
Property
ies
Reference
Hefer. to
toxicity
data
743. X, N '-bi*{r}-i ’hloroc.trbethoxy V/i-phenvlenecii-
aniine
34
mp
201°
31
24
74!. l,4-5(s(X,X'-Oimcthylureido’ibenzene
30
nip
229.5-230.5°
30
24
74">. '-Disulfinvl-p-phcuylenediarnine
40a
nip
115-110°
40a
35
740. p-l)imcthvlaminoanilinc
Commercial
24, 33
74 7, p-Di md hylaminophenyl isothioeyanate
34
bp5
148-150°
34
24
mp
09-70°
34
748, p-Dimcthylaminophenyl isothioeyanate hydro-
chloride
34
mp
144-145°
34
24
741*. p-Dimct hylaminophenyl isothioeyanate moth-
iodide
34
mp
171°
34
24
750. X-\lelhyl-X-(p-dimethylaminophenyl)thiourea
hvdrochloride
24
751. X,S-I)iincthyl-N'-(p-dimethylaminophenyl)-
thiuu rea 11 y droit slide
24
752. X,X '-Di met hvl-p-phenylenedian due
34
bp”
157 100’
34
24, 33
■ _ —
mp
53-54°
34
753. X,X‘-Dinudhyl-p-phenylonediamilie dihydro-
chloride
34
nip
224 (1
34
24
754. X,X'-Dirarbethoxy-X,.X'-dimethyl-p-phenyl-
enediamine
34
nip
106-107°
34
24
755. X,X,X'.X '-Telrameth\T«-pheii vlenediamine
34
bp"
92 93°
34
24. 33
750. X, X,X ',X '-Tetramethyl-o-phenylenediamine
—meihiodide
34
mp
19451
34
24
757 X,X, X ',X '-Telramethvl-m-phenvlenediamine
34
bp10
121-124°
34
24, 33
758. X,X,X',X'-Tctramrthyl-m-phenylencdiamine
meihiodide
34
mp
18751
31
24
759. X,X,X',X '-Tetramethyl-p-pheiiylenediaminc
Commercial
24, 33
700. X,X,X',X'-Tetramethyl-p-phenylenediamine di-
hvdrochloride
34
mp
22251
34
24
701. X,X,X',X'-Tetrame4hyl-/)-phenyleiiediamine
meihiodide
34
mp
20051
34
24
702. i>-Phenylene-f)ts(oxasMdidone-3) —
34
mp
253 254°
34
24
703. X ',X '-Diet hvl-X,X-di met hvl-u-phen vlenediamine 34
bp,u
137°
34
24, 33
mp
203-205°
34
764. X,X'-Dielhyi-X,X'-diincthyl-p-|i!icnylenedi-
ainine
34
bp”
150-151°
34
24, 33
705. X,X',X '-Trielhyl-X'-methyl-p-phenylenediamine
34
bp"1
144°
34
21, 33
mp
22°
34
766. X,X,X'-Triethyl-X'-mcthyl-p-phenylencdiamine
dihvdrochloridc
34
mp
22051
34
24
707. X,X,X'-Triel hyl-X'-methyl-p-phenylenediaminc
meihiodide
34
mp
177°
34
24
70S. X,X,X ',X '-Tetradhvl-p-phenvlenediamine
34
bp”
155-156°
34
24, 33
mp
51-52°
34
709. X,X,X',X'-Tet raelhyl-p-phony lenediaminc
meihiodide
34
mp
185°
34
24
770. X, X’ '-his(d-Hydroxyet hylVp-phenvIenediainine
34
mp
123
34
24
771. X,X'-6is(l-Methyl-l-diethylarninohiityl)-/>-
phonylenediamine
34
bpio*
145°
34
24
772. X-Melhyl-X'-(p-dimei hylaminomet hvlphenyl)-
ihiourea hvdrochloride
-
24
773. Polymer of X,X '-decamet hylenc-X,X '-dime! hy 1-
l,4'-diamiiiodiphenylmethanc ftis-metho-
bromide
21
774. Phenylhydrazine
Commercial
24
775. X-Carbomet hoxv-X '-phenvlhvdrazine
49f
mp
114.5-110°
49f
24
770. p-Phciivlenedihydrazine dihvdrochloride
- 34
dec.p.
200°
31
24
777. Tetra-m-nil rophenvlsilicon
40a
24
77S. Tel rahvdrofurfuryl alcohol
Commercial
24
779. Tetrahvdrofurfurvl fluorocarbonate
49r
bp-15
92-94°
49r
24
780. Tetrahydrofurfuryl chlorocarbonate
49o
bp1"
81-83°
49o
SECRET MISCELLANEOUS COMPOUNDS AS CHEMICAL WARFARE AGENTS
263
Tabi.k 1 (Continued)
Compound
Reference
to
synt hesi.s
Physical propel
ProjxTty
1 ies
Reference
Refer, to
toxicity
data
78!.
3,6-K|h rxvcvel, tbcxcnc
24, 33
782,
Adduct of fumn and maleic anhydride
7
nip
110111°
7
24, 33
783.
Furan
Commercial
24
784.
Met hylfuran
Conunercial
24
7 So,
l-(2-Furvl)-2-nilroethyIene
IS
bp"'
74 75°
18
24, 33
786.
I 42- FuryI)-2-nit roprojiene
IS
inp
hp10
110°
48.5-49.5°
18
IS
24, 33
787.
Furfurvl alcohol
Commercial
mp
125°
IS
24
788.
2-Furaldchvde
Commercial
24
780.
Furoic acid
Commercial
24
790,
5-Hvdmxy-2-cbl«roinethyl-y-pyroBC
12
nip
162163°
12
791.
5-Meth<>xv-2-ehloromethvl-> -pvrone
12
mp
119 120°
12
24
7! >2.
5-llydroxy-2-henylethyl)-pyrro-
colinium bromide
25
24
812.
Oct ahydn>4-(d-hy dr, ixj'ct hyl)-2-met hylpy rm-
colinium bromide
25
24
813,
4-( d-Aco t, ixyet hyl)-oc( ahy dro-2-met hylpyrro-
colinium chloride
25
_
3
24
814.
2-TriacetvlnoreholyIoetahvdropyrrocoline
25
mp
75-95°
25
24
815.
N-ChlorocarhamyIpi|ieridine
24
816.
3-{ Pi)S‘ndvl-X-carliMiml (choline iodide
24
817.
1-Pipcridylsulfamyl chloride
. ..v
24
818.
X -<•)-11 vdroxyet hylpiireridine
12
bp™
95-96°
12
819.
2-Pijieridyloethyl X-mel hylcarbamale
methiodide
29
mp
103-105°
29
24
820.
X-Cyanoinethylpiperidine
49h
i.p"
83 84°
49h
24
821.
2-Vinvlpvridine
12
822.
Coniine (o-propvlpi|)eridine)
Commercial
24
823.
2-0-1 lydroxyel hyl (pyridine
12
bp"4
84 90°
12
824.
3-Ilromoacetylpyridine hvdrobromidc
24
825.
Xicotfne
Commercial
. .
24
826.
2-( X-CarlK,mcthoxvamino)pyridine
49j
mp
• 122°
49j
24
827.
4-(/S-I)imcthvlaniinoethyl)pyriiline
30
bps"
135 145°
30
35
828.
4-(tf-l)imethviaininoot hyl)pvridine dime) hiodide
30
rnp
207-208°
30
35
829.
2-(/3-Hydroxyethylamino)pyridjne
12
bP”
180-185°
12
mp
109 110°
12
X These substances were obtained from natural sources.
SECRET MISCELLANEOUS COMPOUNDS VS CHEMICAL WARFARE AGENTS
Tabi.E 1 (Continued)
Reference
*
Refer, to
_
to
Physical properties
toxicity
Compound
synthesis
Property
Reference
dat a
830. 2-Carbclh»xvoxv-4-carbcthoxyaininopyrindane
34
inp
139-141°
34
24
831. 4-Carbet hoxvamin<>-2-p-tosyloxy-4-pyriiidane
34
nip
132.5°
34
24
852. 4-Amim>-2-liydroxypyrindine
nip
309“
34
24
833. 4-Acetylamim>-2-liydroxypyrindinc
24
834, 2-Aectoxv-4-acelvlaminopyrindine
X . .
24
835. 2-Methylpyrrocoline hydrochloride
25
nip
01 02°
25
24
836. 2-Plie n vlpyrn icolinc
25
mp
214-215°
25
24
837. 3-Acet vl-2-rnct hvlpyrroeolinc
25
nip
83-85°
25
24
838. 2-Triacol vlnorcholvlpyrrocolinc
25
nip
109-170.5°
25
24
839. 1,2,3,4-Tctracarbomethoxyquinolizinc
25
nip
180-188°
23
24
840. 4,7-Dichloroquinoline
47c
mp
84-85°
17c
24
841. 2-p-N itrophenylqninolinc
401
842. 8-Methoxy-5-methylquinoline
....
24
843. 2-{»i-DimelhylamiiiophenyI)quinoline
40k
. . X
844. 2-4 />-! )iincthylaminophenyl )qnin-1,3-isoquino-
lincdione —
Commercial
— ■
- 24
847. 9-Vinvlcarbaxole
24. 33
SIS. Kthvhme-N-nilrofcourea
19
mp
102 I04d
19
24
849. 1,4-1 >itlhyl-l ,4J»?s{3-hyilnixvi'thyl)pi|MTazinium
diehloride
24
850. 2-PhenyIimidazo-[l ,2-o] pyridine hydrobroniide
25
mp
122-124°
25
24
851. N-Morpholinoacetonitrile
49i
nip
60’
49i
852. 1,4-Selenoxan-4-dichloridc*
. . . *
853. Phenoxtellurinc*
—
854. 10,10-Dichlorophenoxtellurine*
855. Bisajiometliylbrucinc hydrochloride
24
850. Bisapomethylhrucine diacetate
24
857. Dimcthvlfuraxane
■ 12
bp760
153°
12
24
858. Dimethylfurazane oxide
12
bp5*
170-171°
12
859. Dicarbellmxyfurazane oxide
12
bp!s
173°
12
24
860. Methvl N-{5-totrazalvDcarbainate
49g
nip
>300°
49g
801. Product of thermal destruction of cyanogen
chloride
—
3.5
862. Veratrinc
Commercial
...
24
803. Ricin*
Sec Chap. 12
See Chap. 12
804. Ficin
24
865. Lubricating oil, S.A.E. No. 10
24 -
860. Fog oil, SGF No. 1
24
SECRET PART II
SPECIAL PHYSIOLOGICAL AM) TOXICOLOGICAL STUDIES
SECRET Chapter 15
THE ASSESSMENT OF PARTICULATES AS CHEMICAL
WARFARE AGENTS
William L. Doyle and l\. Keith Cannon
I5.i INTRODUCTION
During tiik vkars 1941 1915, more than 1,500
compounds were examined in the United States
as potential chemical warfare agents. The volatilities
of the majority were so low that they could have
little offensive value if used in the form of vapors.*11
On the other hand, a few were intrinsically so much
more toxic or more vesicant than were the standard
chemical warfare agents61012 that the question of
disjiersing them in particulate form commanded con-
sideration. Ricin (\V), for example, was several score
times as toxic as phosgene, whereas l,2-6ts(/3-chloro-
ethylthio)ethane (Q), when applied in a solvent to
the skin, was vesicant at one-tenth of the minimal
blistering dose of mustard (II).
Apart from observations incidental to the study of
the screening power of smokes, little attention was
pair! to the toxicological properties of particulate dis-
persions until the decision was made to submit finely
powdered ricin to field tests. As a result of this de-
cision, an expanding program of work was under-
taken on the physical and toxicological assessment of
dispersions of this material. As a result of the experi-
ence gained, the investigations were later extended to
a study of the vesicant effects of aerosols of f».s(/3-chlo-
roethylthioethyl) ether (T), Q, and fm(/3-chloro-
ethyl)amine (HX3).
The point of departure of all the work was an ap-
preciation of the paramount importance of particle
size in determining the effectiveness of a particulate
cloud (see Table 1). In the first place, the particle
size determines the stability of the cloud under given
meteorological conditions. Secondly, it controls the
fraction of the area dose which will impact upon an
obstacle in the path of the cloud, and therefore de-
termines the hazard to the unprotected skin and
eyes of an individual in the cloud. Finally, the im-
pacting characteristics of the particles also control
the inhalation toxicity of the cloud, since they affect
the fraction of the inhaled material that will pene-
trate to and lie retained in the lungs.
The problem of the assessment of particle size in
fine liquid particulates had been greatly advanced by
Table 1
warfare t
. Relation of pari
•haracteristics.
tide diameter to chemical
Pari icle
diameter
(microns)
Type of cloud
Characteristics of clouds
10 3 - 10
- Vapor
(molecular)
Airborne, non|»ersistent, sub-
ject to laws of diffusion.
Invades lungs, eyes, cloth-
ing, and skin.
10 '
Aerosol
Airborne and non|)ersistent.
Invades lungs. Din’s not
impact out of streamlines.
I to 5
Fine particu-
lates
As for aerosol except that it
is more readily filtered and
the lung retention is more
complete. Five a is close
to up[»er limit of nasal
penetration.
20 to 100
Particulates
('loud persists in mild lapse
conditions. Does not reach
lungs. Impacts on surfaces
and should invade eyes
and skin.
200
Sprays
Sediment rapidly. Impact
efficiently. Not dealt with
in this chapter.
the British in (lie invention of (he cascade impaetor.33
The attempt was made to adapt (his instrument to
the assessment of clouds of solid particles. However,
the irregular size, shape, and density of the particles
raised a number of difficulties which have not yet
boon satisfactorily resolved. Much fundamental
work has, however, been carried out on the calibra-
tion and use of the cascade impaetor with dusts.1318
The relation of particle size to the inhalation
toxicity of toxic particulates was investigated by
direct assays in animals of various species. Ricin
aerosols of controlled ranges of particle size were
utilized.lsi At the same time the filtering character-
istics of the human nose were measured by observa-
tions of the extent of penetration of a variety of
nontoxic particulates.13'1 •,8ch The effectiveness of dis-
persions of nonvolatile and of slightly volatile vesi-
cants on human skin and on the eyes of animals were
investigated as a function of particle size, wind
speed, etc.,3k
SF.CRFT ASSESSMENT OF PARTICULATES AS CHEMICAL WARFARE AGENTS
The results obtained in this work have shown
clearly that the significance of the size of the air-
borne particles cannot be reduced to any simple
formula. However, the following broad conclusions
would appear to be justified and may serve to indi-
cate the status of the problem.
1. The size, shape, and density of the particles, as
well as the wind speed and other meteorological con-
ditions, all contribute to the aerial behavior of a
particulate cloud. Particles with effective diameters
greater than alxmt. 100 n sediment rapidly in a stable
atmosphere. They will remain airborne for significant
periods only under conditions of considerable turbu-
lence. On the other hand, clouds containing effective
concentrations of particles smaller than 0.1 n in di-
ameter are subject to rapid aggregation. For example,
if a cloud with a concentration of I mg 1 were com-
posed initially of very small particles, it would attain
relative size stability only when the average particle
diameter had grown to about 0.7 m-
—These considerations lead to the conclusion that
the problem of toxicological effectiveness may be
restricted to a consideration of clouds whose parti-
cles (if of unit density) fall within the size range of
0.1 to 100 n.
2. In general, toxic agents are much more effective
if they enter the lungs than if they are retained in the
nose. The probability that a particle will penetrate
the nasal barrier increases as the size of the particle
diminishes. Available evidence indicates that the
optimum size for penetration to and retention in
human lungs probably lies within the limits of
0.5 3 ft in diameter.l8* h The optimum size for lalx>ra-
tory animals is appreciably lower.18* These values are
for resting animals and are further lowered at the
high ventilation rates associated with exercise.
3. The probability of the impaction of a particle
on a surface in the path of a cloud increases with the
size of the particle. At moderate wind sjieeds, the
fraction of the area dose which may lie expected to
impinge on the surface becomes significant if the
particle size is above 10 m and becomes an important,
fraction of the area dose above a size of about 70 a*-46 4r
It would api>ear from items 2 and 3 that no single
dispersion can exploit to the full the potentialities of
an agent which, like Q, is both vesicant by contact
and toxic by inhalation. This is a fundamental di-
lemma which imposes serious limitations on the
offensive potentialities of aerosols of this type.
4. If munitions were available which would dis-
perse particulate material in either of the optimum
size ranges indicated above, new orders of inhalation
and of vesicant effectiveness in the field should he
obtainable. Such munitions have not yet l>een ade-
«iuately «leveloj)ed.3'J
15.2 TYPKS OF I’AKTTCUIATKS
15.2.1 Sternutators
Classical sternutators such as diphenylamine-
chlorarsine (adamsite), diphenylaminecyanoarsine
(cyan DA), and toxic sternutators such as aconitine
and nitrophenyldichlorarsine are primarily harassing
agents. These agents act at concentration time prod-
ucts (Ct's) considerably lxdow (hi mg min in*. At
present there is little interest in these agents because
they are stopped by available masks and because
trained troops carry on effectively despite their pres-
ence. It is possible, however, that the utility of
sternutators has not been adequately considered.
Larger particles than those that have been utilized
may be more harassing. German interest in mixtures
of sternutators with mustard 16 may indicate at-
tempts to hide the presence of more toxic agents.
15.2.2 Toxic Particulates
1. Inorganic substances, e.g., cadmium selenium.
2. Synthetic organic compounds, e.g., aromatic
carbamates.
T Naturally occurring substances, e.g., ricin (W).
The metals are thermally stable and may be in-
corporated in standard smoke incendiary S!> or high-
explosive weapons. As indicated in Chapter 11, these
substances are not more toxic than standard chem-
ical warfare agents, but they may be used without
ready detection in various types of munitions.
Although the aromatic carbamates are consider-
ably more toxic than standard agents,10 they are un-
stable in aqueous solution and to heat. For these
reasons, little serious consideration has been given
to their use as particulate clouds.
Ricin (W) is intrinsically somewhat more toxic
than the best of the carbamates. It is also thermo-
labile. Its toxicity when dispersed as a cloud has been
studied extensively in the laboratory and prelim-
inary field trials, using special munitions, have been
carried out. This interest in ricin was not entirely
dependent on its own merits as a toxic agent. It was
recognized as a prototype of toxic protein materials
of bacterial origin which wore known to have even
greater toxicity but which were less conveniently
prepared and handled.
SF.GRKT EFFECTIVENESS OF PAIIT1CULATE CLOL DS
15.2.3 Vesicants
1. Volatile vesicants, e.g., mustard (II), tris(0-
chloroethyljamine (HN3).
2. Nonvolatile vesicants, e.g.,12 b is (/3-c Ido root hy I-
thio)ethane (Q).
All the members of this group are toxic, but not so
toxic as (hose in Section 15.2.2, (1) and (2). They
are, however, vesicant. The best nonvolatile vesi-
cants are intrinsically more toxic and more vesicant
than the volatile ones. Q is inherently 10 to 20 times
as vesicant as II and at least 5 times as toxic.7,91*a l8h
They should lx* more difficult to detect than the
volatile agents. In the field they will not be expected
to create a vapor hazard, but, by contamination of
equipment, should establish a contact hazard for
bare skin which it would be difficult to eliminate by
decontamination. It is doubtful if nonvolatile vesi-
cants can be effective through clothing.1*1 I*1’ The
volatile members of this group can lid dispersed
thermally, by means of airplane spray, or by high
explosive-chemical shells. The particle size achieved
will markedly influence the action of the agent. Thus,
very small pat t ides (0.2 1.0 g in diameter) may Ik*
non vesicant because of streamlining, but will be
more toxic by inhalation and will also yield (he great-
est and most rapid vapor return. Larger particles
(5 to 25 g) will have greater vesicancy, but probably
at the expense of toxicity. The largest particles (200 -
2,000 g) may lx* most effective for the penetration of
clothing, particularly of the permeable protective
type. The largest particles may also create a contact
and traversal hazard.
These statements are broad and tentative general-
izations based upon contemporary views of the char-
acteristic behavior of particles of different diameter.
Many of these generalizations require further experi-
mental investigation. 'Hie results of contemporary
work are reviewed in Section 15.5.1.
15.3 TilK EFFECTIVENESS OF PAR-
TICULATE CLOUDS
15.3.1 Stability
Srdimcntation. The rate of sedimentation of
particles in a static atmosphere increases with in-
crease in particle size. Computations based upon
kinetic considerations indicate that precipitation lx*-
comes rather rapid when the effective diameters of
the particles exceed about 100 g. Clouds of such large
particles could Ik* maintained in the air for significant
periods only under strongly turbulent conditions.
They are sprays rather than clouds and find their
natural use for contact and ground contamination
— e.g., as airplane sprays. The opt imum part ide size
for such sprays depends on a variety of factors —
the speed of the plane, the turbulence, the vola-
tility of the agent, etc. — which it is not the
province of this chapter to discuss.|J Suffice it to
say that for direct assault upon exposed personnel
there is general agreement that the optimum range
of particle size to obtain massive and diffuse con-
tamination with a vesicant is about 0.3-2 mm in
diameter. If ground contamination is the objective,
the upper limit of size may be unimportant.
Coagulation. Although the rate of sedimentation
sets the upper limit of size in a jiersistent aerosol, the
tendency of particles to coalesce upon collision estab-
lishes a lower limit of particle size stability. On simple
considerations of collision frequency, the half life of
a particle should l>e roughly proportional to the con-
centration of the aerosol. With increase in particle
size, the concentration required to give a fixed half
life increases with the mass, and therefore with the
cube of the radius of the particles. It has been esti-
mated47 that the half life of an aerosol containing
5 X 10* particles jw-r milliliter is 6 minutes at room
temperature. For particles of 0.1 g in diameter and
unit density, this corresponds with a concentration
of 2.5 nff I. For particles of 1 g in diameter, the cor-
responding concentration is 2.5 gg 1. If these clouds
were initially established in higher concentrations,
they would aggregate until the numbers of particles
in unit volume had fallen to a relatively stable level.
It should be noted that the tendency to coagulate
does not lead directly to a reduction in mass concen-
tration, but rather to an increase in average particle
size. The phenomenon is important, therefore, only
if clouds of small particle size are required. There is
no purpose in attempting to disperse a particulate in
a smaller particle size than can be sustained by the
concentration that is to Ik* established. If a concen-
tration of I yg I is accepted as the lowest which the
toxicity of the material would justify, then the small-
est particle size which it is worth while attempting to
disperse is of the order of 1.0 g.
In summary, therefore, considerations of rates of
sedimentation and of coagulation suggest that we
should concern ourselves with the behavior of air-
borne particles in the size range of 0.1-100 g in diam-
eter, corresponding to a HP-fold range of particle
mass.
SECRET 270
ASSESSMENT OF PARTICULATES AS CHEMICAL WARFARE AGENTS
15.3.2 The Significance of Particle Size
The tactical use of a chemical warfare agent in the
form of a cloud is, in general, to lie justified only
when the conditions of the operation will l>e such
that personnel exposed to the cloud will absorb
casualty-producing doses through the lungs or
through the skin. When the cloud is a true vapor,
the actual dost* that is inhaled under standard con-
ditions of respiration can be predicted from the prod-
uct of the concentration and the time of exposure
(Cl). The amount of vapor absorbed from the skin
under standard conditions of temperature and hu-
midity can, likewise, be predicted from the Cl, since
the rate of diffusion of the vapor to, and (he rate of
penetration of, the skin may generally lx* taken to lx*
proportional lo the concentration. It follows that an
effective dosage on the target can lx* assured if muni-
tions expenditure is properly adjusted to the meteor-
ological condit ions.
The tactical requirements cannot lie formulated so
simply when the cloud is composed of particles with
colloidal or larger dimensions. In this situation, the
concentration of the agent may actually be less im-
portant than the sizes of the airborne particles. It
has been noted that this is a factor in determining
the stability of a particulate cloud. In the case of an
agent which is toxic by inhalation the particle size
also controls the proportion of the inhaled material
which is filtered out of the inspired air in the respira-
tory passages. Likewise, in the case of a vesicant, the
amount of material deposited upon an exposed sur-
face at a given Cl and wind speed is a function of the
particle size. In brief, the effect ive dose of an inhalant
and of a vesicant depends on the impinging char-
acteristics of the particles which, in turn, depend
upon the size and density of the particles.
15.3.3 The Impingement of Particles
The amount of an airborne particulate which will
deposit on an object in the path of a cloud w ill ho the
sum of the amount which impinges upon the object
and the amount which is deposited under gravity.15
Since this discussion has ix*en limited to clouds in
which the rate of sedimentation is small, considera-
tion may l« confined to the amoimt which impinges
on the object.
The probability that a spherical particle w ill im-
pinge upon a cylindrical surface in its path is given 45
by
/* = «( 1 + ■-) (1 - e~^/D) (1)
In this equation, n is the velocity of the particle, <1 its
diameter, and p its density. D is the diameter of the
target, while a and n are constants. It will lie seen
that the tendency to impinge increases with the size
and with the density of the particle and also with the
wind speed. It depends also on the size and shape of
the target. According to Sell,15 when D » , a =
0.75 and « = 050 egs units. More recent experi-
mental data give values of a = 0.75 to L and a =
70 egs units.
It may Ik* assumed that all liquid particles which
impinge upon a surface will remain adherent to it,
but. it is not expected that this will Ik* true for a solid
particulate. In this ease the probability of adherence
to the surface may be much lower than the probabil-
ity of impingement. The magnitude of the losses will
depend upon the, nature of the surfaces of (he target
and of the particulate.
15.3.1 The Effective Vesicant Dose
The to\icologica!Iy effective dose of a nonvolatile
vesicant may lie taken to be proportional to the
amount of the agent which is deposited on unit area
of a surface exposed to the cloud in question. This
amount is given by the product of Cl, u, and/, where
n is the wind speed and / is an impaction factor. The
latt er corresponds "with the fraction of t he area dose,
which is deposited on the target. When the cloud is
homogeneous with respect to size and density and
the adhesiveness of the surface for the particles is
high, / is given by P in equation (1). When the cloud
is heterogeneous, it is necessary, in principle, to
measure the distribution of the total concentration
over the particle sizes which are present and to de-
rive an overall impaction factor for the cloud.
P is an exponential function of the size of the parti-
cle. When its value is considerably less than unity, it
increases rapidly with small increases in particle size.
For this reason, the impacted dose of a heterogene-
ous cloud may l>e largely determined by the rela-
tively small number of the larger particles which are
present. A heterogeneous cloud may, therefore, Ik* a
much more effective vesicant than a homogeneous
cloud having the same mass median diameter.
Further complications are introduced if, as may
occur with solid particulates, the particles vary in
shape and density as well as in volume. A very im-
portant example of such variations is (he formation
SECRET PRODUCTION VND CONTKOT, OF PARTICl LATE CLOUDS
271
of loose irregular aggregates of low density from
smaller primary particles of uniform density. This
type of aggregation is prone to occur during the dis-
persion of powdered materials, particularly if they
are somewhat hygroscopic. The effects of these com-
plicating factors on the impingement of solid parti-
cles are elalxjrated in Section 15.4.3.
When one is dealing with a particulate cloud of a
slightly volatile agent such as HNS, consideration
must lx? given to the toxic effectiveness of the vapor
as well as to that of the dispersed phase. It must bo
rememliered, also, that the characteristics of the
cloud continually change with time. The particulate
phase suffers progressive loss in concentration and
size as volatilization pnx-eeds until a pure vapor
cloud results. An analysis has been made of the fac-
tors which determine the rate of evaporation of air-
borne particles.”
It is of interest to note that the wind speed has two
opposed effects on the tact ical efficiency of a vesicant
particulate cloud. The greater the velocity of the
wind, the lower Is the concentration of an agent
which is being generated at a fixed rate. On the other
hand, the greater the wind speed, the gieater is (he
impaction efficiency of a given concentration of the
particles.
Experimental studies of the relation of particle
size to the vesieancy of aerosols of Q, T, and TINS
arc reviewed in Section 15.5.1.
15.3.5 Effective Inhaled Dose
It is generally acknowledged that toxic particles
are less effectively absorbed from the nasal and
respiratory passages than from the alveoli of the
lungs. Considering the pulmonary toxicity alone, the
effective dose of an inhalant may be given as the
product of Cl, v, and (1-/) where v is the minute vol-
ume of respiration, and/is the fraction of the inhaled
material which is trapped in the respiratory passages.
It will lx1 agreed that this fraction is determined in
large measure l»y the amount of impaction in the
nose. It may lx* expected to vary with the species of
animal, and, to some extent, from animal to animal
of the same species. It will also vary with the physi-
ological state of a single animal. To the extent that
impingement in the nose determines/, an increase in
rate of respiration will, by increasing the velocity of
the particles in the nasal passages, result in a greater
nasal retention and a reduced effective dose.
The question of (he extent to which particulate
material which enters the alveoli is retained and al>
sorlxxl has been investigated in a preliminary way.
The results are summarized in Section 15.5.2.
Experimental studies of the effects of particle size
on the toxicity of ricin for animals and on the reten-
tion of nontoxic particulates in the human nose are
reviewed in Section 15,5 and in Chapter 12.
15.1 LABORATORY PRODUCTION AND
CONTROL OF PARTICULATE CLOUDS
15.1.1 Dispersal
Liquid* and Solutions. In a few speeial studios the
Sinelair-LaMer homogeneous smoke generator has
Ik'cii used.1 3 Thermogenerators may lx> employed
for the dispersal of stable, slightly volatile agents.
In most cases, however, various types of atomizer
have-been used under conditions of operation which
have been empirically determined to give clouds of
the desired characteristics. Preliminary studies of
the fundamental proi>erties of atomizers have been
reported.Mb
A useful method of producing clouds of varying
particle size from a standard atomizer has been the
following. A nonvolatile cosolvent is mixed in vary-
ing proportions with a dilute solution of the agent in
a volatile solvent. When these mixtures am atom-
ized, the mass median diameters of the particles in
the cloud vary with the proportion of nonvolatile
solvent in t he original mixture. For example, glycerol
has been found to bo a satisfactory cosolvent for
aqueous solutions of ricin and dibutyl phthalate for
solutions oLnonvolatile vesicants.13*-,8b
Solids. Electric arcs employing the toxic agent as
one component of the electrodes provide useful
sources of finely divided metals and their oxides.
Thermal generation of toxic clouds by the incorpora-
tion of the agent in incendiary or fuel block mixes
may also lx? used when the agent is thermostable.
Sucli thermal generators tend, however, to give dis-
persions which coagulate rapidly.**-40-41
The obvious alternatives in the ease of a thermo-
labile solid such as ricin are to disperse by atomiza-
tion of a solution or to generate a dust cloud from a
finely comminuted powder. Most devices which have,
been desenix'd for the dispersal of powders lead to a
fractionation of the sample. In some it is the smaller
particles, in others, the larger particles which tend to
disperse the more rapidly. In most there occurs a
considerable formation of loose aggregates in the
cloud. Although some attempts have been fairly suc-
cessful,nblsh no really satisfactory method for the
SECRET 272
ASSESSMENT OF PAKTICl I VIES AS CHEMICAL WAKFAKE AGENTS
uniform dispersal of a powder at a rate of a few milli-
grams a minute has been described. The devices
which lead to least aggregation in the cloud have the
disadvantage of a variable rate of delivery. (See
Section 15.(1 for dispersal in (he field.)
15.1.2 Measurement of Size
The assessment of particle size in a cloud requires
not only the observation of the range of sizes in the
cloud, but also the amounts of material in the differ-
ent size categories. The results are comprehensively
expressed as curves in which the cumulative amount
of material is shown as a function of the diameter of
the particle. Three types of curves may lx- distin-
guished, according to whether the particle diameter
is plotted against (1) the number, (2) the volume, or
(3) the mass of airborne particles. From these curves
may be derived respectively a number median di-
ameter [XM D], a volume median diameter [A MD],
and a mass median diameter [MMD], The number
distribution is appropriate if one is interested in ef-
fects dependent on the numlter rather than on the
mass of airborne particles — as, for example, in the
knockdown of mosquitoes. The volume distribution
has no particular practical significance, but is the
form in which results must be cast if the densities of
the particles are not known and the amount of ma-
terial must be evaluated from microscopic observa-
tions of the numbers and diameters of the particles
in the sample. The mass distribution is the descrip-
tion of particle size which is most significant to the
problem of the vesicant and toxic effects of the cloud.
The clouds from atomized liquids have fairly typi-
cal distributions and the densities of the particles are
uniform. In such conditions the MMD is sufficient
to characterize the cloud satisfactorily. In dust
clouds generated from powders, on the other hand,
the distribution of sizes may be quite abnormal, the
unitary particles may lx- far from spherical, and
many aggregates of low density may I>e present. The
MMD of such a cloud may be a quite misleading in-
dex of the impaction efficiency of the cloud. The
complete mass distribution is required for the char-
acterization of such a cloud.
Methods. When dealing with dusts it is desirable
to make counts of the undispersed material for com-
parison with the airborne cloud. The MM I) and the
range of sizes in a given preparat ion are best deter-
mined by direct microscopic examination It the M MD
is below 10 g. The work is tedious and various meth-
ods have been discussed to save labor but critical in-
vestigatora agree with Fairs on the procedure to lx*
followed. Hard and fast rules for (lie numlxr of parti-
cles to bo counted cannot be stated.130 The statistical
features of the problem are well presented by Dalla-
valle.4- The suitability of a laboratory or field proce-
dure for the measurement of the particle size in a
cloud depends upon the size of the particles, whether
they are liquid or solid, upon the time available for
sampling, and upon the concentration. Optical meth-
ods suited to the analysis of homogeneous smokes
have been developed.* These methods are not readily
applicable to heterogeneous clouds, but some at-
tempts in this direction have been made.13 In general,
the optical methods result in neglect of the relatively
small numbers of coarse particles winch may carry
an appreciable fraction of the mass. An instrument
capable of photoelectric measurement of the surface
area of individual particles is not theoretically impos-
sible. In view of the labor required in available pro-
cedures, some such device is highly desirable.
Ultra microscopic and dark field observations of
falling particles have frequently been employed.18
Such methods are limited to particles small enough
to remain airborne prior to observation and to con-
cent rations so low that coagulation is avoided. There
is great danger that large particles will be lost in the
sampling procedures^prior to observation.
'The thermal precipitator47 is very useful for parti-
cles below 5 to 10 n in diameter, provided that the
cloud is available for a sufficient period of time so
that the necessarily slow sampling rate provides an
adequate sample.
For clouds ranging from 2.0 to 50 g in diameter,
there is one instrument at present which avoids many
of the difficulties inherent in other methods. This is
the cascade impact or.2* •:w It merits more detailed
consideration than those already referred to.
]5.t.3 'The Cascade Impactor
This instrument consists of a series of four jets ar-
ranged in series so that the sampled cloud impinges
at four increasing velocities on to suitably prepared
microscope slides (A,B,C, and D). In this way the
particles are separated into four impacted groups.
'The size ranges trapped on successive slides overlap
to some extent, but the MM 1> of the material on a
" The British workers employed the effective drop size
[EDS] m place of the MMD to characterize the slides. The
EDS is approximately the size lielow which !)8 jjer cent of the
number of particles on each slide is found and for most clouds
is about 1 times the diameter of the mass median.ui
SECRET PRODUCTION AND CONTROL OF PARTICULATE CLOUDS
273
particular slide is, under favorable conditions, char-
acteiistic of that slide. Under such conditions, there-
fore. it is necessary only to measure the amount of
material on each slide in order to obtain a rather
satisfactory assessment of the mass distribution.
The amount of material on a slide may lie computed
from microscopic counts or by chemical analysis.
The cascade impactor has a number of obvious ad-
vantages over single jet instruments such as koni-
ometers, the Owen’s jet, etc. [t was originally de-
vised and calibrated 23 30 for the assessment of
liquid particulates. For nonvolatile liquids quite pre-
cise data can lie obtained if proper consideration is
given to the following variables.
In the first place the MMD of the material im-
pacted on any one slide depends to some, extent, on
the MMD of the cloud as a whole. It deqiends also on
the shape of the distribution curve of the cloud.
Values of the MMD’s on the four slides have been
determined experimentally for clouds of MMD 5,
10. 10. and 100 g.13' Impingement of a particle on a
given slide is a question of statistical probability.
The particles of a homogeneous cloud are distributed
over more than one slider Calculations have been
made of the mass distribution on the slides which
should be obtained with strictly homogeneous
clouds.Secondly, the MMD of the particles
on a slide depends on the velocity of operation of the
impactor. Experimental results indicate that the
MMD is proportional to the reciprocal of the square
root of the flow rate.iai lHh
As the result of the analysis of the counts of a
large number of slides a characteristic mass distri-
bution curve has been constructed.181* By means of
this curve it is possible, by chemical analysis of the
amounts of material on the slides in a given experi-
ment, to arrive at a fair estimate of the MMD’s on
the four slides. When an instrument lias been cali-
brated in this way, the use of chemical methods of
analysis eliminates the very tedious process of micro-
scopic assessment of the slides.
Dust Clouds. The use of the, cascade impactor for
the assessment of clouds of solid particles was first
investigated in this country.1*"-' -,,h It will be evident
from what lias already lieen said that its use for this
purpose is complicated by a number of factors which
arise from the diversity of the characteristics of solid
particles. Solid particles may be highly irregular in
both shape and density. It has lieen found, for ex-
ample, that samples of ricin prepared by the spray
drying of aqueous solutions consisted largely of hoi-
low spheres. When this material was further de-
graded by air grinding the product was chiefly in the
form of thin disks. Again, the particle size distribu-
tions in dusts may lie quite different from those char-
acteristic of atomized sprays. The spray-dried ma-
terial referred to was remarkably uniform in size,
whereas ball-milled preparations of Hein contained a
wide range of particle sizes with a large number of
extreme fines. Finally, the adhesion of impinging
solid particles may lx* incomplete and the degree of
slippage may change progressively as the slide be-
comes coated with the agent.
These factors combine to give a wider distribution
of particle sizes on a single slide than is obtained
when an atomized liquid is assessed. When the parti-
cles are not. spherical, the problem arises of the
proper method of computing their volumes from
observations of their dimensions under the micro-
scope. Serious errors may arise if they arc1 treated as
if they were spheres. The volume of a sphere is
0.52 UP. Hey wood 44 has listed some of the factors by
which the cube of the observed “diameter” should be.
multiplied when the particles depart from the spheri-
cal. The factor fora rounded particle is given as 0.54,
for a prismoidal object it is 0.47, and for a tetra-
hedral particle it is 0.38. A mean value of 0.5 is sug-
gested for a heterogeneous assembly of nonspherical
part ides.
Recent work has confirmed the validity of this
factor for slides (’ and D, but it has not always been
possible to apply it to slides A and B because it is on
these slides that the large highly irregular and often
disk-like particles are found. To measure the mean
lateral dimensions of such particles and compute
their volume as though they were spheres leads to an
MM1) for the slide which is much greater than the
true value. Some attempt should be made to measure
the thickness of plate-like objects and to calculate
their rectangular volumes.
The frequent occurrence of aggregates in a solid
particulate has proved to Ik1 particularly trouble-
some.41 The MM I) of t he particles impacted on a
particular slide varies with the square root of the
density. Since the density of a loose aggregate may
1)0 less than one-tenth of that of the unitary particles
of which it is composed, it is evident that the presence
of many aggregates on a slide may profoundly change
the MMD of that slide. The problem of the density
to be assigned to an aggregate in order to compute
its mass is also a difficult one. Microscopically the
l)est that can be done is to take a few representative
SECRET ASSESSMENT OF PARTICULATES VS CHEMICAL WARFARE AGENTS
aggregates, count the numlier of unitary particles in
them, and sum their volumes. The density may then
be taken to be the ratio of this volume to the volume
of the whole aggregate treated as a sphere.
Many aggregates disintegrate when they impact
on a slide. They will be assessed as though they corre-
sponded in impinging properties with the unitary
particles of which they were composed, although the
latter would probably not have appeared on that
slide had they not been aggregated. The result will
be artificially to reduce the MMD below its real
value.
When unusually large (Kirticles are present in a
cloud, losses may occur by impingement on the walls
of the orifice of the instrument, particularly when
the impactor is operated in a-static cloud. Under
conditions of isokinetic sampling of clouds moving
with average wind velocities it has boon calculated
for liquid, droplets that orifice losses become appar-
ent with droplets about 50 g in diameter and in-
crease as the size further increases. Similar calcula-
tions for aggregates with a density of 0.1 indicate
that the upper limit for reliable sampling is about
100 g. The upjjer limits for static clouds are probably
appreciably below these figures because of increased
turbulence around the leading edge of the orifice.
Summary. The emphasis which has been laid
upon the evaluation of the sizes of particles on the
impactor slides has tended to distract attention from
the fact (hat the cascade impactor does not measure
the size of a particle but rather its impingement tend-
ency. Most of the difficulties in applying the instru-
ment to the assessment of dusts have been in express-
ing the distribution of impacted material in terms of
volumes and masses computed from microscopic ob-
servations of the dimensions of the particles. This
has been a necessary preliminary to the calibration
of more direct methods of interpreting the results
obtained. In so far as the toxicity of particulate ma-
terial is a function of the amount of material which
will impact in the nose or on exposed surfaces the
efficiency of a cloud should, most logically, be de-
scribed in terms of its impaction factor under stand-
ard conditions. The use of particle size to characterize
the cloud is a convention which may, perhaps, be
discarded when instruments which measure impinge-
ment have been properly calibrated.
The MMD of slide B of the cascade impactor is
close to or slightly greater than the maximum size of
particles which have been found to penetrate the
nasal barrier in most animals. The fraction of air-
home material which collects on slides B, (', and D
under standard conditions of operation should, there-
fore, be somewhat greater than the effective inhala-
tion dose. Calibration of the instrument in such a
way as to establish a relation between these two
fractions should make possible an estimation of the
inhalation toxicity of a cloud from a chemical analy-
sis of the impact or slides alone.
The effective dose of a vesicant is dependent on
the fraction of airborne material which is large
enough to impact efficiently. Most of this fraction
should be captured by slide A. The analysis of im-
pactor slides operated in clouds of nonvolatile liquid
vesicants should lead without much difficulty to
satisfactory estimates of the effective vesicant dose.
15.5 PARTICLE SIZE \ND TOXICITY
15.5.1 Vesicant Effects
The dose of a particulate which is deposited on_an
object, depends upon the amount settling out plus the
amount impacting.45 The amount impacting will vary
with the wind speed, density of the particle, area of
the particle, diameter of the target, and nature of the
surface of the target. A heterogeneous cloud of
MM I) 2.0 g may have the same impactibility for a
given surface as a homogeneous cloud of M MD t.O g.
The impingement pattern on the object will vary
with particle size from a diffuse pattern with vapors
atid stpokes to a localized (upstream surface) mosaic
with coarse sprays. The volatility of the agent and
the rate of absorption of the material by the target
will affect the physiologically effective dose.
Preliminary indications of the order of magnitude
of effect of particle size on vesicancy of a nitrogen
mustard (HN3) and a nonvolatile vesicant (T) were
obtained by exposures of forearms in a wind tun-
nel.13k~m,l8“ At 5 mph wind speed and under condi-
tions of temperature (about 80 F), relative humidity,
and skin resistance (sweating index) such that a
vapor of HN3 at a Ct of 1,200 mg min nr produces
an erythema, the following tentative conclusions
were reached. Smokes of MMT) below 2.0 g are less
effective than vapor. A heterogeneous (atomized)
cloud of MM D 2.0 g was equally as effective as vapor.
A heterogeneous cloud of MMD 8.0 g was twice as
effective but the erythema was more localized. HN3
is less volatile than mustard. T is practically non-
volatile. The relation of volatility and particle size
to vesicancy is illustrated by the following relation-
ships. By topical application of single drops to fore-
SECRET PARTICLE SIZE VND TOXICITY’
275
arms it lakes 10 limes as much IIN3 to produce the
same skin reactions as a given amount of T.7
As a 2. 0-/i (heterogeneous) particulate, a Cl of to
of T (area dose = Cl X 5 mph) is the equivalent of
a Cl of 1,200 mg min m* of 11N3; T is thus 27 1 hues as
vesicant as TINS. When the particle size is raised to
8.0 g, a Cl of 0 to 10 of T is as effective as a Cl of 600
of HN3, thus demonstrating a factor of 100 or more
in the vesieaney of these agents.18* These findings
demonstrate the importance of designing munitions
which will dis|>erse the chosen particulate in an opti-
mum size range.
Owing to the great effect of temperature and
humidity 4 817 on skin reactions to given exposures,
it is difficult to generalize from these data to other
agents and conditions. By employing the appropriate
factors for comparison of-vesicant power,71xh com-
parisons may l>e made with the values given in
Chapters 5 and 0. Under the conditions obtained in
the exjKM'iments described in the preceding para-
graphs, I] vapor is about one-half as effective as
HN3 vapor. The effect of evaporation of the agent
after deposition on the skin has hern found to bo
approximately (he same for H and ITN3,,8a b despite
differences in volatility. It will be indicated in the
next section that of a heterogeneous cloud of T of
MM I) 8.0 n. only 10 15 per cent will penetrate the
human nose.
The results on vesieaney in relation to particle
size apply to exposed skin areas. The presence of
clothing profoundly modifies the situation. A droplet
of nonvolatile agent on the surface of clothing can
under some circumstances !>e considered innocuous,
whereas a volatile agent will generate vapor which
may lie drawn over the underlying skin by the bel-
lows effect of clothing. Numerous tests have been
carried out on the droplet diameter required to pene-
trate clothing by wetting the cloth. The sizes in-
volved are well above the particulate range consid-
ered here. Relatively few data on the penetration of
clothing by small particles are available for chemical
warfare agents. The amount of a particulate found
on clothing is a function of filtration and impaction.
A given expenditure of agent will with increasing
wind speed deposit decreasing amounts on (and
through) the cloth by filtration (bellows effect) but
will deposit increasing amounts by impaction, espe-
cially for coarser particulates. For nonvolatile sub-
stances the amount penetrating cloth by impaction
forces appears to l>e a small fraction of the amount
penetrating by filtration.I8b
For nonvolatile materials there is definite disad-
vantage to the use of particulate clouds coarser than
1 to 2 n in diameter if penetration of clothing is to he
achieved. Increased turbulence at higher wind speeds
appears to reduce the percentage that penetrates by
filtration.18'*
15.5.2 Penetration of the Nose ,
Initial experiments were designed to determine the
particle diameter at which 50 per cent of the mass of
a given cloud passes the nose.1*-'-*-2* Values obtained
on four human subjects are given in Table 2.
Tmilk 2. Penetration of the human nose by particulates.
Agent
Diameter for
50 per cent
Density Flow rate penetration
(g-ml) (1pm) (n)
Corn oil
Dry NalK'Oj
0.93 17 5.0
00 IS
2.2* 17 2.1
00 0.8
* Actual density in
tides.
i nose somewhat lower Ijecause of hydration of par-
These experiments wen* extended in an attempt to
determine the percentage penetration of the nose at.
various sizes for materials of differing physical char-
acteristics, e.g., liquid corn oil of density 0.9 and dry
NaHCOs of density 2.0, and at various rates of
breathing. The results are presented in Figures 1-
and 2. It is of interest that there is little difference in
rir.rRE 1. Nasal retention of particulates in man.
nasal penetration Ik*tween flow rates of 17 and 29 1pm
(unpublished data). A change from 17 1pm to 60 1pm
changes the value for 50 per cent penetration from
2.1 to 0.8 n. Regardless of flow rate or density, parti-
SECRET 276
ASSESSMENT OF PARTICULATES \S CHE.MIL VL WARFARE AGENTS
eles 10 g in diameter have approximately a 10 per
cent chance of penetrating the nose.
Over the size range shown in Figures 1 and 2 it
would ap|icar that for a given particle diameter the
MASS MEDIAN DIAMETER IN MlC»0NS
FiiiCRE 2. T.ung retention of particulates in man.
lung retention is about 20 per cent more efficient
than the nose. From another viewpoint, the same
efficiency in retention obtains for particles in the
nose which are 2.5 times the diameter of those in the
lung.
When molecular dimensions arc reached the nasal
and lung retentions increase al>ove those found at
0.2 g.18'-29
15.5.3 Inhalation Toxicity in Animals
Mice, rats, and rabbits were exposed, while at rest,
to particulate clouds of W in glycerol at controlled
HMD’s. 'Phe relation between particle size and
L{Ct)ho is shown in Figure 3. For the size range 0.5 to
7.0 g the effect is much more pronounced in rats and
mice than in rabbits. These data, which are reviewed
in more detail in Chapter 12, should be compared
with those of British authors29 using other tech-
niques.
13.6 DISPERSIBILITY OF PARTICULATES
In the laboratory it is relatively simple to prepare
clouds of unitary particles by atomization, thermal
generation, or in electric ares. Previously comminu-
ted powders may also Ik- disperses! largely as unitary
particles in special apparatus. Munitions capable of
dispersing previously comminuted powders in the
unitary state have yet to be developed. Powders
differ in ease of dispersibility, as shown in various
Fioitre 3. -Inhalation toxicity of ricin in relation to
particle size.
laboratory tests. Sueli tests tire, however, generally
meaningless in terms of dispersibility by field muni-
tions. The factors involved in field munitions which
are difficult to scale up from laboratory tests include
aggregation phenomena occurring prior to, at. the
time of, and immediately subsequent to dispersal.
These aggregation phenomena are influenced by ge-
ometry, strength of materials, brisance of explosives,
and scale of munition.** •**
To date field experiments 39 have, however, almost
universally confirmed the finding 15 that suspensions
in organic nonsolvent media result in much higher
dispersion efficiencies than can l>e obtained by use of
gas ejection munitions or standard munitions with
dry fillings. Owing to low bulk density of dry fillings,
suspensions permit a higher ratio of active filling to
munition weight.
15.7 FIELD ASPECTS OF PARTICULATE
ASSESS M ENT
The outstanding observation resulting from fit !d
experiments on dispersion of previously comminuted
powders by field munitions is the fact that the frac-
tion of material airborne in the size range of the
original filling is generally insignificant. Most of the
mass of the filling appears in a highly aggregated
SECRET FIELD ASPECTS OF PARTICULATE ASSESSMENT
277
state. Field sampling must not only evaluate the
gross clumping and spillage but also account for the
low toxicity (in terms of chemical Cl or area dose) of
the more lastingly airlxxrne clouds. The frequently
occurring light fluffy (snowflake) aggregates are gen-
erally not encountered in laboratory investigations.
They are particularly deceptive since they may be
readily disrupted in the sample and thus appear as a
group of component unitary particles.
The orifice velocity of field sampling equipment
should not deviate markedly from the wind speed if
particles above 50 g in diameter are to lx* readily
sampled. Where power-operated devices ait* em-
ployed, the requirements of pump capacity for ap-
propriate sampling of particulates, which are higher
than for vapors, may cause some embarrassment.
In early exjx*riments fine wires were tested as
sampling devices but discarded because of (he differ-
ing impaction efficiencies for small and large parti-
cles.31’ By employing wires or tubes of three different
diameters, however, the relation of collection effi-
ciency to wire diameter can be utilized to calculate
particle size and area dose from the mass of material
collected on each size of wire."**r d 44 This device
dispenses with power requirements when sampling
in wind speeds above 3 mph. At I mph corrections
for settling are required.
There are marked difficulties in the assessment of
initial clouds containing vapor and particulate con-
centrations. The chemical drop trap 24 and the chem-
ical selector*4 indicate possible methods to be de-
veloped. The use of impingers M or filters is recom-
mended when numerical values of (he MMD or
impactibility are not required, 'fixe total chemical Ct,
without regard to size, can be measured for dry par-
ticulates with filters. Hayon-aslx’.stos, esparto-as-
bestos, and gas mask filter papers may lx- used. For
use with \V those papers are unsuitable owing to the
strong adsorption of the protein on the paper. Cellu-
lose acetate filter batts 12 do not absorb proteins ami
in addition are soluble in appropriate organic liquids.
Another qualitative device for evaluation of aggre-
gates is the “sticky finger.” Ha
Methods have been developed and results obtained
during the period 1911 to 1945 which indicate the
desired particle size for various purposes. In the same
jxeriod, however, no adequate munition capable
either of producing such sizes or dispersing materials
already prepared at those sizes has been developed.
Thus, at the date of writing, W (which has in the
laboratory several score times the toxicity of phos-
gene) has (in the field) been found to be only seven
times as toxic as phosgene in the best munitions
available.**
SECRET Chapter 16
APPARATUS AND TECHNIQUES UTILIZED IN TOXICOLOGICAL
STUDIES ON CHEMICAL WARFARE AGENTS
By //. .1. Wooster and W. L. Doyle
16.1 INTRODUCTION
In this chaptku are summarized methods de-
veloped and utilized for toxicological studies at
the University of Chicago Toxicity Laboratory
[UCTL]. Pertinent contributions of other NDRC
Division 9 contractors arc included, but no attempt
is made to review systematically developments made
by other agencies.
The apparatus and methods are described under
the following major headings: (1) gassing chandlers,
(2) methods of dispersing agents into chambers,
(3) sampling equipment, (4) precision methods of
testing inhalation toxicity, and (5) methods of test-
ing vesicants. Each section starts with a discussion
of the relevant principles and is followed by a brief
description of specific items of equipment and pro-
cedure, together with an evaluation of the merits and
limitations of each. Descriptions and construction
details for the more important items of equipment
will l)e found in the reports listed in the Bibliography
and referred to in the text.
The work leading to the development of appara-
tus included in this report was initiated prior to
March 15, 1945, at which time the contract with the
University of Chicago was assumed by the Chemical
Warfare Service. Subsequent work has Den reported
where it was in logical extension of apparatus initi-
ated under the prior contract.
J6.2 GASSING CHAMBERS
16.2.1 General Description of Design
The earliest form of gassing chamlier was a closed
container in which the animals were placed and the
agent dispersed. Despite the simplicity of such an
apparatus, its use introduces many complexities.
The actual concentration of agent in it at any one
time is a result of the action of at least two variables
— the rate at which the agent is sprayed into (he
chamber and the decrease of the concentration. The
latter is influenced in several ways — absorption on
the chamber walls, chemical changes of the agent
(the hydrolysis of dichlordialkyl arsines, for exam-
ple), and, in the ease of particulates, aggregation of
the smaller particles. Animals kept, in a closed eham-
Ikm for any period of time may /hange the carl ton
dioxide content of the air sufficiently to distort their
respiratory patterns. Nominal concentrations in such
chambers are almost meaningless, and analytical
concentrations are difficult to interpret.
Lehmann, in Germany, in a long series of investi-
gations (1884 1918) st udied the effects on animals of
various toxic vapors used in industry, llis method
was to expose animals in a modified Pettenkofer
respiration apparatus to a continuous flow of air con-
taining a constant and known concentration of the
agent being studied. Almost all the gassing chambers
used in this country since 1918 are based on this
constant Row, or “dynamic” principle. (It should lie
noted that English workers, in many of their screen-
ing runs, employed “static” chambers during World
War II.)
The ratio of chamber volume to air flow is critical
in the design of such chandlers. Silver23 derives the
basic equation covering chamber equilibration t imes:
tv = 4.(5 X I
0
where = time for the chamber concentration to
attain 99 per cent of the theoretical
nominal concen trat ion.
a = volume of the chamber in liters.
h = (he rate of air flow in 1pm.
It will Ixi seen from this that a chamber having an
air flow of 1 chamber volume per minute will come to
equilibrium in about 5 minutes; with 10 chamber vol-
umes per minute equilibrium is attained in 0.5 min-
ute. A (puck equilibration time has several advan-
tages— momentary changes in concentration, such
as ( hose produced by the introduction of animals, arc
quickly rectified, and unstable materials have less time
in which to decompose. The saving in material by the
use of a shorter equilibration time is overbalanced
by the larger amount of material necessary to set up
SECRET GASSING CHAMBERS
279
a given concentration, but a 5-g sample is generally
adequate for a single test of a substance toxic at
0.3 mg 1. When slowly volatile materials, which exist
as both vapors and aerosols, are dispersed in cham-
bers of very high How rates, such as the auxiliary
chamlier for the 200-1 medium How chamber (see
below),-effects of the flow rate on toxicity may lx1
encountered.
The flow of air through the chamlier may Iic pro-
duced by either positive or negative pressure. In
most chamliers negative pressure is used, to mini-
mize the tendency for toxic materials to escape into
the laboratory. Standard equipment for this is a
gear-type (Roots) blower V-belted to an electric
motor. An air ejector is used on the chamlier for the
large Benesh atomizer, and water aspirators have
been used on some small smoke chamliers. Positive
pressure has been used on three chambers — a large
screening smoke chamlier, a small chamber used for
testing the toxicity of gasoline, and the microline.
In these chamliers the air How is controlled by the
volume of air blown into the chamlier.
Standard equipment for measuring air flow
through the larger chamlx'rs has lieen an orifice in a
Monel plate in the effluent, line between (he chamber
and the filters. A differential manometer, filled with
butyl phthalate, is connected to each side of the
orifice. To calibrate such a flowmeter a large dry-
type gas meter is connected to the chamber and all
other openings are sealed. A working calibration
chart is prepared from these readings. The dry meter
is calibrated by positive displacement of a measured
volume of air.
The standard orifices are about 0.8 inch in diam-
eter. Because of their location they are subject to
contamination and corrosion. When aerosols are used
in the chamber they tend to clog up the hole and
make it smaller. It is advisable to recalibrate such
orifice flowmeters at least once a year. A much larger
orifice (about 2.5 inch) has been used in the chamber
for the large Benesh atomizer, which was designed
specifically for use with smokes. An inclined differ-
ential manometer is necessary to read accurately the
small pressure gradient resulting from the use of such
a large orifice.
The effluent from these chambers contains a large
proportion of the original toxic material. It is passed
through replaceable charcoal filters. When nonvola-
tile vesicants have lx*en used, the filters become
heavily contaminated, and their removal and re-
charging is a hazardous procedure. The effluent from
all chambers plus (he effluent from all rooms is drawn
olT by ml ary bh>wers and discharged into a large
incinerator stack. The dilution afforded by the stack
provides a larger margin of safety and in some cases
permits dispensing with charcoal filters. The UCTL
stack has an average inside diameter of I6J2 feet
and is 100 feet high. Under normal conditions the
stack discharges 750,000 cfm.
The chamlx'rs are of metal and or glass construc-
tion. The all-metal chaml>ers are constructed of
welded sj'fi-inch mild steel plate, which is protected
on the inside with a baked-on vitreous or bakelite
resin (l.ithcole) enamel. Connections to these cham-
lx'rs are made with standard plumbing pipe fittings.
Ten-liter wide-month glass bottles with holes drilled
in them have been used for several small chambers.
The chamber on the small Benesh machine is made
entirely of triplex safety glass, cemented together.
Composite structure is represented by the 400-1
chamlier. made from a length of Pyrex industrial
pipe 12 inches in diameter, fitted with brass ends,
and the 488-1 chamber, made of metal lined with
plate glass.
One of the more important procedures in gassing
animals is the method of introduction of animals into
the chamber. The simplest method, which is entirely
feasible with mice, is to open a port and insert the
caged animals. This is routinely done with the small
smoke chambers and with some of the larger cham-
bers which have small auxiliary ports on their larger
doors. With larger animals, some sort of sliding car-
riage for the cages must lie provided. This is, in
essence, a three-sided box, the ends of which are
plates, and the bottom an open structure. The side
view can be represented by | J. Either end may
serve as a closure for the opening in the chamber side.
When such a carriage is rapidly pushed into a cham-
ber, a certain piston action is exerted. The carriage
on the big Benesh machine was designed to avoid
(his. When the carriage is out of the chamber, closure
is provided by a vertically sliding glass panel. Thus
(he carriage needs only the form ). In high-flow
chambers the animals may be placed in the chamber
before the agent is put in. This is practicable because
of the short equilibration time of such chambers.
However, it should not be used for short exposures to
substances which deviate markedly from Haber’s
law.
One difficult problem in the design of chambers is
the position of the port through which the agent is
to lie introduced. This may be at either the top or
SECRET 280
VI’P.AHATL'S \M> TKCHMQLES IN TOXICOLOGICAL STI'DIES
the side of the chambers. The top is a somewhat more
convenient, location, inasmuch as all the parapher-
nalia connected with dispersal may be placed on top
of the chamber out of the way. When dealing with
gases or with aerosols set up by a baffled atomizer,
the position is not so important as with other de-
grees of dispersion because the materials enter the
chamlier at a low velocity and loss by impaction on
inlet tubes is negligible. With concentric atomizers
dispersing semi volatile materials, introduction from
the top means that the spray must undergo a right-
angle bend to get. into the chamlier, with consequent
loss on the mixing bowl. A jet fed in from the side of
the chamber must lie aimed with care to clear the
animal cages. It would seem advisable to design
future chandlers with provision for the optional use
of either route of entry.
Little attempt has been made so far to control the
temperature and humidity of air entering the cham-
lier. In most cases the chambers withdraw air from
the lalioratorv and operate at the ambient temper-
ature and humidity. In the microline provisions were
made to humidify the entering air. Some small cham-
bers have lieen operated in a thermos fated water
bath. The most elaborate regulation is in the man-
chamlier, which has automatically controlled equip-
ment for heating or cooling, and varying the water
content of the entering air, as well as temperature
control of the room surrounding the chamlier.
16.2.2 Description of Specific Chambers
Rectangular Chambers Larger than 200 Liters
.QUO-Liter Standard Chamber.g -3 The first large
chamber used at UCTL was built from a design
standard at Ed go wood Arsenal. This chamlier is
fitted with a sliding carriage 8 inches high and 15
inches wide. This, at most, can hold 4 cats or rabbits,
or 20 guinea pigs or rats. The chamber air flow can
be regulated between 50 to 90 per cent of the cham-
ber volume per minute. A wooden sliding carriage
with stocks for surrounding the necks of exposed
animals was made to study body and head exposures.
In use, this chamber was found to have several
limitations. Animals larger than cats could not be
exposed routinely (singledogs were used in body ex-
posures). Appreciable difficulty was encountered in
working with lewisite, owing to wall loss at the low
How rates --e.g., the nominal LC:,o of lewisite for
mice was approximately three times as high with the
standard chamber as with the Benesh machine.
SSO-Liter Standard Chamber.* This chamlier is
identical in principle and operation with the 100-1
chain her. The sliding carriage is somewhat higher in
relation to the height of the chamlier. Its cross-
sectional dimensions are 23x55 Inches. This gives
it a maximum animal capacity of I small dogs, or
2 large dogs, or 1 or 2 goats, or 6 monkeys. A mixed
group of 1 small dog, 1 rabbits, 1 cats, 10 guinea pigs,
10 rats, and 20 mice can Ik? exposed at the same t ime.
At a later period a small door was built into the
outside plate of the sliding carriage, making it possi-
ble to put small animals into the chamber without
pulling out the carriage.
This chamber has been calibrated with mustard
gas (11), using (hi1 Northrop 1 itrimeter.3*1 The air
flow was 700 1pm. and the nominal concentration
may be expected to be in error by about + 5 percent-
The concentration built up is the same in all parts of
the chamber within 1 per cent and the drop in con-
centration on moving the carriage in or out is prob-
ably not more than 5 per cent.
This chamlier has, perhaps, Ik'om the most con-
sistently useful for general work.
300-Liter Medium Flow Chamber,u This chamlier
was designed to provide a chamber in which dogs and
other large animals could be exposed to agents at
rates of chamber exchange comparable to those at
which mice had been exposed in smaller chambers.
By interchanging a glass door on (he side of the
chamber for one which is provided with platforms
and head stocks, mice, rats, or guinea pigs may I>e
exposed to gases either by inhalation or by body ex-
posure alone. A similar arrangement can be attached
to the front carriage for similar exposures of cats,
rabbits, or dogs.
Some time after the chamber was built an auxiliary
high-flow chamlier was added.14 The new chamber
was built onto a removable side plate which could be
substituted for the side door. The cross-sectional
diameter of the high-flow chamber is about one-ninth
that of the main chamber. Air is drawn from the
main chamber into the auxiliary chamlier and thence
into the exhaust line. When the chamber is operated
at 500 1pm, the velocity is increased to 3 mph just
before the toxic agent reaches the animals, with a
minimum velocity of 0.5 mph in the center of the
compartment in which the animals are exposed.
These velocities may be increased or decreased by
varying the air flow. The incident velocity may lx;
changed by changing the size of the slits through
which (he air stream enters.
The high-flow chamlier is 18x7x3 inches. It.
SECRET GASSING CHAMBERS
is divided into three compartments by two longi-
tudinal walls, each of which contains 10 slits, 2x
Yi incl u*s. the slit size may be varied. The compart-
ment in which the animals are exposed is 18x3x3
inches, and is located lx-tween the other two com-
partments. the long, slender compartments on each
side have openings in the floor through which ana-
lytical samples may lx- drawn. The inner of the small
compartments is o|x*n on the side communicating
with the main chamber, and on the distal side has the
slotted wall openings. It serves as a mixing chamber
to insure that all the animals are exposed to the same
concentration. The outer of the two compartments
has a slotted inner wall through which the air stream
leaves. The effluent is carried away through an open-
ing in the end of this chamber. Analytical samples
can lx* drawn Ix-fore and after the agent passes the
animals.
Animals may be exposed by total exposure, or by
body or head exposure atone. A special manifold is
provided for the last two types. The lower portion of
the side plate to which the high-flow chamber is
attached can also be used for either Ixx.lv or inhala-
tion exposures at low flow rates. Total exposures for
low flow rates can be carried out by placing animals
in the main chamber. Animals may, therefore, lx-
exposed simultaneously to high and low flow rates
either by total exposure, body exposure, or inhalation
exposure.
The 200-1 chamber differs in several design details
from the standard chambers. The carriage is provided
with castors, making it more convenient to slide it in
and out. In the standard chambers the bare metal of
the door seats against the bare metal of the chain) tor.
In this chamber sponge rubber gaskets are pro-
vider!. The toxic agent is usually admitted at the top
of this chamber instead of at the side.
In the use of this chamber a good agreement has
lx‘on obtained between analytical and nominal con-
centrations. L{CI):M’s obtained by this chamber cor-
respond with those* obtained in the small high-flow
chamlxws. This is not the case with values obtained
from the 100-1 standard chamber.
429-Lttrr (Ham-Lined Chamber. This chamlx*r was
designed specifically for use with aerosols. It is lined
with plate glass. The sliding stainless-steel animal
carriage is attached to a glass panel which forms the
front wall of the chamber when the carriage is in
place. When this is not used a counterweighted glass
panel drawn down from the top seals the chamber.
Interlocks are provided so that the carriage cannot be
pushed in until the sliding panel is fully raised. This
scheme is a trifle complicated and requires two oper-
ators for rapid action, but has the advantage that it
does not exert the plunger effect of the usual cham-
ber carriage. A small circular auxiliary port in the
panel on the carriage permits caged mice to be placed
in the chamber without opening the main door.
A large air injector is used as a pump to exhaust
air from the chamber. This gives a maximum air flow
of 900 1pm at 35 lb air pressure. An inclined differen-
tial manometer, reading across a large orifice, gives
t he chamber air flow. Such a large orifice is less sensi-
tive to fouling than those commonly used, flic air
injector is also less subject to fouling with aerosols
than gear-tyjx' blowers. Variations in the pressure of
the air running the Venturi are corrected by a
diaphragm-actuated regulator.
The glass lining makes this chandler particularly
easy to clean. It is much.quieter in operation than
the mechanically driven chambers.
This chamber has lieen calibrated with 11 by means
of the Northrup titrimeter, at a nominal concentra-
tion of 38.8 gg 1 and an air flow of 1.5 chamber vol-
umes per minute.21® The following conclusions were
drawn:
1. The Ct calculated from the nominal concentra-
tion will be in error by about -flG |x*r cent.
2. The 10-minute Ct calculated from an analytical
concentration measured at about the mid-point of a
10-minute exposure will be in error by about +3 per
cent.
3. The Ct calculated from an analytical concen-
tration based on a sample drawn over the entire
period should be in error by less than 3 per cent.
4. In general, errors caused by the fall in concen-
tration that occurs upon pushing in the animal cages
may Ik; neglected for 10-minute exposures and can lx*
corrected by analytical samples drawn at intervals
during the entire exposure period.
Screening Smoke Chamber.316 This chamber was
designed for the repeated exposures of animals to low
concentrations of agents employed as screening
smokes. It was made large enough for monkeys to
live in and was fitted with automatic controls. The
chamber is 4 feet square and 7 feet high. Its volume
is 3,078 I. The base is a concrete block fitted with a
drain and lined with sheet metal. The top is wooden,
as are the comer posts. The sides are of glass. A com-
mon wooden door with a glass panel is let into one
side, this door is weafherstripped. The whole struc-
ture is lined with a very heavy wire mesh.
SECRET 282
APPARATUS AM) TECHNIQUES IN TOXICOLOGICAL STUDIES
Two fans are used with this ehamlier. A continu-
ously running exhaust fan provides ventilation. An
intermittently operating centrifugal blower giving
2,180 1pm is mounted on the top of the ehamlier. Just
Inflow the ceiling outlet is a suspended baffle plate.
A six-jet atomizer (DeVilbiss experimental model
No. 7030 1), connected to the compressed air line,
discharges into the inlet of the centrifugal blower. A
General Electric time switch controls the solenoid
valve, feeding compressed air to the atomizer and the
relay actuating the inlet fan. These go on and off to-
gether, in a cycle of 30 minutes on and 30 minutes
off.
The chamber was found to come to equilibrium in
10 (±2) minutes. 'This is somewhat longer than the
theoretical time. Forty-five |>er cent of the equi-
librium concentration is reached after I minute,
and eighty per cent at 5 minutes.
Lnrrimotor Chamber"'* This chamber is essentially
a 400-1 standard ehamlier with a maximum air flow
of 1,0001pm. The adaptation for use with lacrimators
consists of three ports projecting from the center of
the ehamlier walls on three sides. Eye pieces, which
fit the ports snugly, consist of rubber diaphragms
edged with rubber tubing. Swimming goggle frames
are cemented around holes cut in the diaphragm.
The sternutator provision consists of industrial-type,
nose and mouth respirator masks connected to the
ehamlier with lengths of gas mask host*. There are
six of these.
The subjects signal their response by tapping keys,
located under the ports, which cause signal magnets
to mark an automatically timed rotating kymo-
graph. The subject taps the key when irritation is
first experienced, and again when he feels tears start-
ing to form. Thereafter he depresses the key each
time he is forced to close his eyelids and releases it
when the lids are once again open. At the end of the
run there is a graphic record of the onset of irritation
and of lamination, as well as of (he periods during
which the eyes were open or closed.
Owing to the low priority assigned to lacrimators
and sternutators, this chamber was never extensively
used or completely calibrated.
Great Lukes Man-Chamber’""*"r" * This cham-
ber was designed for the exposure of human subjects
under conditions of temperature and humidity con-
trollable by the investigator and independent of
ambient conditions.
The chamber is made of :,s-inch boiler plate, lined
with i6-inch sheet lead. Its volume, exclusive of
Ilie air lock, is about 17.300 I. Tlic maximum How
rate through the ehamlier is about 5,100 1pm. All
control of concentration (H has l>eon (he only agent
used) is dune with the Northrup titrimeter, so that
exact values for this flow are not so necessary as when
an attempt is made to estimate the nominal con-
centration.
This chamber is equipped with automatic pneu-
matic controls for temperature, relative humidity,
rate of flow, and pressure. They function as follows;
1. All air coming into the chamber passes through
a eommeicial air-conditioning unit. It emerges from
this into the chandler at 26 F, saturated with water
vapor. When warmed to 70 F, this air is at about
35 per cent relative humidity. The temperature and
relative humidity of this air represent the lowest
levels at which the ehainber can be operated,
2. The desired wet bulb and dry bulb tempera-
tures are set on the controlling-recording apparatus
and the steam lines are opened. Heating is cont rolled
by a steam coil controlled by the dry bulb tempera-
tures. Lowered wet bulb temperatures cause the
automatic humidity valve to open, injecting steam
into the ehamlier. When the wet and dry bulb tem-
peratures reach the desired values, the humidity
valve closes and the by-pass dampers open; thus the
incoming air is conducted underneath the heating
coil rather than through it.
3. When the air is pulled through the heating coil,
there is more resistance in the system than when the
air is by-passing the coil, so that adjustments of the
flow are necessary. This regulation is controlled by
dampers on the discharge side of the exhaust fan.
When the flow rate drops below 5,600 Ipm, these
dampers open and permit more air to lie drawn out
of the ehamlier; as the flow rate rises, the dampers
close and cut down the flow. The flow rate usually
oscillates between 5.300 and 5,900 1pm.
4. Ordinarily the fluctuations in the amount of
air discharged would produce variations in the pres-
sure inside the chamber. Such variations are elimi-
nated by automatic control of the dampers on the
discharge side of the supply fan. The pressure con-
troller is set for a differential of 0.1 inch of water;
when the pressure in the chamber increases, the con-
trol damper effects an opening of the dampers to the
room, so that less air is passed into the chamber.
Similarly, when the inside pressure falls to a value
lower than 0.1 inch of water Inflow the outside pres-
sure, the dampers close* to jiermit a larger volume of
air to enter the ehamlier.
SECRET GASSING CHAMBERS
283
An air lock is equipped with motor-driven ports by
means of which fresh air may be diverted through the
air lock when men wearing contaminated clothing
are leaving the chamber.
Measurements of the wind speeds in the chamber
showed that the velocities vary from less than
0.4 mph in the corners to over 8 mph in front of the
fans, with an average of 2.5 mph for 32 positions.
Constant-Flow Uhamokbs Smallkk than 100 I,
The Microline.1 The 100- and 800-1 standard cham-
bers were found to be unsuited for “screening” new
agents of which only small amounts were available,
and for working with unstable substances such as the
arsenical*. The mierolmo together with its ancillary
chambers was designed to provide a small chamber
through which a relatively high How of air at con-
trollable humidity could be sent.
The influent air is delivered via two parallel series
of bubblers and absorbers. One of these delivers dry
air, the other saturated. These arc mixed in the de-
sired proportions and passed through a dispersing
bubbler or an impinging atomizer containing the
agent and thence into the chamber.
The first chamber used with this mieroliue was a
10-1 screw-capped wide-mouthed bottle. A cylindrical
cage fastened to the bakelito screw cap contained six
mice. A U-shaped manifold, both ends of which
passed through the screw cap, was user! for body ex-
posures. The heads of mice were stuck through holes
in the manifold while fresh air was circulated through
it, and a concentration of toxic agent was set up in
the chamber. A branched manifold for testing toxic-
ity by inhalation could be substituted for the cham-
ber. This enabled 8 mice to inhale the agent while
their bodies were exposed to room air.
These chambers and manifolds had several draw-
backs. Only mice could lx? used in the chamber, and
not more than six of these. The agent flowed linearly
through the chamber, so that if the first animal af-
fected the composition of the agent the last might,
get a lowered dose. The inhalation and body expo-
sure manifolds could hold only 8 and 6 mice, re-
speetively, and were difficult to manipulate.
Later a commercial Lectrodryer unit8 was in-
stalled to supply adequate amounts of dry com-
pressed air. The size of the water-saturators was in-
creased proportionately. An 11.5-1 chamber was con-
structed of plate glass cemented together and sup-
ported inside of an angle iron framework. This was
designed to assure equal distribution of the toxic-air
mixture directly to each animal. To do this the
material is conducted into an H-sha|ied channel,
each arm of which has an opening connected with a
slit in the glass side of (he chamber, 0.1 rum wide and
extending from front to back. The channel is de-
signed to give uniform flow through the whole length
of !>olh slits, which form two horizontal lines on each
side of the chamber and are so centered that, when
the mouse cage is placed in the chandler, the animals
am directly opposite the slits in line with the flow of
the material. This permits a high degree of uniform-
ity in exposure of the mice. The effluent is carried off
by an identical arrangement on the other side of the
ehaml)er. This chamber has a capacity of 20 mice,
3 rats, or 3 guinea pigs.
A body exposure manifold which holds 20 mice and
fits into the chamber and a separate inhalation mani-
fold holding 16 mice have also boen-construct<*d.
Using an aerosol of HN3, recoveries of about. 65 per
• cent were obtained from the chamber in the absence
of animals and of about 75 per cent from the inhala-
tion manifold. Recoveries of more volatile materials
are well above 90 j>er cent.
Small Smoke Chamber. This was essentially a vastly
simplified mieroliue. Dry air was passed through a
dispersing bubbler or impinging atomizer. The re-
ducing valve on the compressed air tank, with the
atomizer connected, was calibrated in liters per min-
ute versus pressure. Auxiliary air could he bled in
through a Y tube to bring the flow to the desired
value. The air flow was then led through a water or
steam jacketed condenser into the wide-mouthed
bottle used as a gassing chamber. The toxic material
was blown out of the chamber into the air of the
hood in which the whole setup was placed. This
chamber has been used in a thermostated water hath
for exposures alxjve or Ixdow room temperatures.
This setup, which required a minimum of appara-
tus, proved to bo quite useful for screening materials
of low vapor pressure. Its use; was limited to materi-
als which would melt without decomposing, so that
they could lx> dispersed with an impinging atomizer.
Mice, guinea pigs, and rats were (he only animals
that could Ixs fitted into the chamber.
A mollification of (his chamber was used to set up
very high concentrations of gasoline vapor.*’* All air
entering the chamber was blown through a concen-
tric atomizer, and then passed through a steam-
heat ed Friederieh’s condenser. A trap in the line re-
moved non volatilized material Indore it entered the
chamber.
SECRET APPARATUS AND TECHNIQUES IN TOXICOLOGICAL STUDIES
The Wind Tunnel*1' The UCTL became inter-
ested in the relation of particle size to vesication on
bare skin and through clothing and in the relative
efficiencies of the vapor and aerosols of the same ma-
terial as vesicants. It was necessary to construct a
wind tunnel in which the arms of human subjects
could be exposed to airborne agents moving at vari-
ous and variable velocities.
The tunnel, circular in cross section, is 11 feet long
and 2J2 feet in diameter in (he largest places. It is
fashioned after certain Port on models30 designed to
give an even distribution of droplets across the work-
ing sect hip. It differs from st reamline tunnels, in
which markedly higher velocities exist at the center
of the stream than at the edges.
The wind tunnel proper (Figure 1) is of cylindrical
cross section. Two truncated cones (li and C) are
placed base to base with a short base diameter cylin-
der in between. This assembly precedes a longer
cylindrical section (£)), IS inches in diameter and
3 feet long. The source of vapor or particulate spray
is an atomizer or bubbler orifice located at (he mouth
of the tunnel. In order to mix the narrow plume of
agent with the main air stream, the diameter of the
tunnel is increased (li) to produce turbulence. The
expanding cone is followed by a reducing cone (C) to
give approximately constant velocity across the
stream in the cylindrical working section (D). The
flow through the working section is somewhat turbu-
lent at 7 mph. This turbulence can lie decreased by
placing a hardware cloth screen in the reducing cone,
l»ut such a screen causes an increase in concentrat ion
of the larger particulate droplets in the center of the
stream. Without the screen the droplet distribution
is quite homogeneous across the tunnel. Turbulence
creates vortexes in the working section, producing
differences of about 10 per cent (at 7 mph) in the
velocities at opposite sides. This difference could
probably lie decreased by increasing the length of (he
cylindrical section between the two cones B and C.
With particulate clouds of nonvolatile droplets in
which 30 |xu' cent of the mass is in the size range
10-30 fx> there is a just perceptible loss on the walls;
the loss is negligible wit h smaller droplets. With drop-
lets of 150 + 50 fi in diameter there is a slightly
greater loss on the bottom of the tunnel than on the
top. Most of the loss occurs in the reducing cone (C).
The source of suction is the room ventilation which
leads via filters to the incinerator stack. The flow is
regulated by adjustable louvers. To obtain velocities
above 25 mph a tul>e with its own reducing cone and
Specialized Chambers
The Explosion Chambers. Chambers were required
to assay the action of high explosives on the toxicity
of certain chemical warfare agents. It was necessary
to construct a rugged chamber in which small
amounts of explosives could be set off and the toxicity
of the resulting airborne material assayed.
The first of these was a 1-ton shipping container
for war gases such as mustard, A port (12-inch di-
ameter) was welded on (his. It was fitted w ith a steel
cover 2 inches thick, bolted down with 1-inch bolts.
T1 iis chamber was used while a specially designed
-chamber w as being built.21*
The latter was constructed from 18-8 stainless
steel. The interior is polished to a No. 4 finish. The
vessel is 48 inches in outside diameter and approxi-
mately 8 h'et high. The volume is 2 cu m. It is
mounted over a concrete pit in a specially const meted
laboratory and is shielded by heavy concrete walls.
The dome-shaped top of the chamber is held down
by SO bolts under spring tension to act as a safety
valve yielding at SO psi. Easy access to the interior is
provided by an 18-inch manhole with single-screw
closure. There are eight 4-inch ports which can lie
closed with :t4-inch Pyrex or stainless-steel plates;
additional ports are provided for valves and electri-
cal leads. The chamber is equipped with a shower for
Hushing. The walls and interior fittings are designed
to permit complete drainage to the valve at the bot-
tom. This permits maximum recovery of the prod-
ucts.
Steam lines lead to the chamber for decontamina-
tion. The residual gases in the chamber may be
drawn through a 200 cfm collective protective
canister.
When metal bombs are exploded they are sur-
rounded by stainless-steel baffles to protect the walls
of the chamber. This is not necessary for glass or
plastic bombs. The resultant gas-smoke mixture is
drawn through Pyrex glass piping to a small glass
constant-How exposure chamber. The effluent from
t he exposure chamber is filtered and absorbed in the
usual fashion.
This chamlier is somewhat small for testing the
effects of high explosives on chemical warfare agents.
Twenty-five grams of explosive is the maximum that
can lie detonated. It would lie desirable to have
means of heating and cooling the chamber walls.
Other than this, the chamlier has proved quite satis-
factory. It is the only known explosion chamber per-
mitting recovery and analysis of the entire residue.
SECRET METHODS OF DISPERSING AGENTS INTO CHAMBERS
285
A Entry port equipped with ancinosiat.
H Expansion cone.
C Reducing cone.
I) Exposure chamber (working section).
K Removable, Inch vi‘locityf reducing cone.
F Velocity control damper.
Cl Kjfil to stack.
Figure 1. Wind tunnel, elevation.
smaller working section is available. Fairing of the
incoming air stream is accomplished with a commer-
cial “auemostat” with the three central vanes re-
moved. The working section is provided with a door,
windows, and sampling ports for introduction of
animals, arms, and instruments. Air speeds are meas-
ured with a commercial Velometer.
The wind tunnel has been employed in studies on
vesication by particulates and in the development of
methods of assessment of particulates (see Chap-
ter In).
16.3 METHODS OF DISPERSING AGENTS
INTO CHAM HERS
16.3.1 General
Liquids may be dispersed as vapors or as aerosols.
Solids may have been previously comminuted or it
may he required to subdivide them in the process of
dispersal. The choice of method to be employed
should Ixi based on the following criteria.
1. The dispersing technique must not produce am
chemical change in the material.
2. The delivery rate must be constant during the
experiment.
3. The rate of delivery should be readily measur-
able in onler to provide a measure of the nominal
concentration.
4. The rate of delivery should be readily adjust-
able to provide an adequately wide range of con-
cent ration.
5. The material must he dispersed in particles of
desired size.
16.;l.2 Techniques of Dispersal
The UCTL has had to test (fie toxicity of materi-
als ranging in volatility from gases to metals. The
physical state of a substance determines the method
of dispersal to ho used.
Compounds Boiling Below 0 C.# Those substances
are usually available compressed in small steel or
copper cylinders. A pressure-reducing valve is at-
tached. A capillary or orifice flowmeter is then cali-
brated for (lie rate of flow of the gas by the liquid
displacement method. A liquid in which the gas is
insoluble is used in the flowmeter as well as in the
pneumatic trough. The gas is delivered at the desired
rate through the flowmeter directly into the gassing
chamber. A nominal concentration, as a cheek on
that derived from the rate of flow, is obtained by
weighing the cylinder before and after each run. Un-
stable gases (e.g., ketone21*) have been generated
directly into the chamber.
Liquids Boiling Between 0 C and Room Temper-
ature. These compounds may be dispersed as gases
or, w ith proper cooling, as liquids. If they are to be
treated as gases they are distilled into a glass am-
poule. A calibrated flowmeter and a reducing valve,
are attached as described. The nominal concentra-
tion is obtained by determining the volume dis-
placed during a run or by weighing.
It is usually more convenient to treat such com-
pounds as liquids. With proper cooling, a solution
may be made up. Any dispersing device for liquids
which ran Ixi adequately chilled can then be list'd.
Such devices are concentric and impinging atom-
SECRET APPARATUS AM) TECHNIQUES IN TOXICOLOGICAL ST I DIES
izers, bubblers, and the small Benesh and constant-
delivery atomizers.
Liquids Boiling Above Boom Temperature. These
may lx* dispersed by vaporizing or spraying. They
are vaporized by passing nitrogen through a bubbler.
The volatility of the material determines the size of
bubbler and the degree of heating or cooling required.
In general it is desirable to use as little heating as
possible. Heat is supplied by a water bath at a tem-
perature somewhat higher than that desired for the
liquid in the bubbler.
Liquids may lx* atomized either undiluted or in
solution. Solutions should not lx* dispersed from im-_
pinging atomizers since the solute and solvent are
usually refluxed to different degrees with correspond-
ing changes in concentration of the solution in the
atomizer.
Solids Which Can Be Dissolved or Which Melt with-
out Decomposition. A solution of a solid can be
sprayed in the usual fashion. The volatility of the
solvent is important. If too volatile it may evaporate
sufficiently rapidly at the atomizer tip to produce
clogging.
Agents which melt without decomposition can l>e
dispersed from a direct or impinging atomizer im-
mersed in a water or oil bath.
Solids Which Cannot Be Dissolved and Which De-
compose When Melted. In most eases these materials
must Ik* ground to the desired particle size before
dispersal. They can lx* dis|x*rsed from the dry duster
(see Section 1(1.3.3 under “The Dispersal of Par-
ticulates”).
Very fine aerosols of metals have been produced by
means of an high-tension arc, using the metal as one
of the electrodes.
16..1.3 Apparatus for Dispersal
Dispersing Bubblers 6 i3
A method of dispersing liquids with appreciable
vapor pressures is to bubble a non reactive gas
through them. The output is controlled by varying
the flow of the gas and the temperature of the water
bath in which the bubbler is immersed. The gas pass-
ing through (he liquid is broken up into small bubbles
by passage through a sintered glass disk (coarse
porosity) or a Folia bulb.
The type of bubbler used depends on the volatility
of the toxic agent. Agents boiling lx*low 50 C are kept
in bubblers with stopcocks at both inlet and outlet to
minimize the possibility of leakage when the bubbler
and contents are being weighed at room tempera-
turc. Compounds with high boiling points arc kept
in bubblers with outlets large enough that the rapid
How of the gas mixture does not blow out material
condensing in the outlet nozzle. When small amounts
of agent are to lx* dispersed the bubbler should be
kept small and light enough to I>e weighed on an
analytical balance. (This also applies to atomizers.)
hen substances are vaporized from bubblers it is
desirable to keep the bath temperature as low as
practical. This minimizes the decomposition of ther-
molabile agents. To get high concentrations in such
cases increased gas flows ate employed in large
bubblers through which as much as 12 1pm of gas
can lx.- passed. *
The use of bubblers in toxicity determinations is
limited by the purity of the substance available and
the amount of air (or nitrogen) which can l>c passed
through them. If the toxic material is quite pure and
stable the amount of substance volatilized per vol-
ume of gas passing through is quite constant . A very
slight degree of impurity will, if the impurity is
volatile, result in a changing-output from the bubbler.
As a result it. is always necessary to make a series of
preliminary runs to bring the output down to “con-
stant volatility.” Bubblers designed to hold 10 to
50 ml of toxic agent usually permit the passage of
gas at a maximum flow rate of 2 1pm. The most con-
slant operating conditions are obtained when the
rale of gas flow through the bubbler is sufficiently
slow to permit at least 95 per cent saturation of the
gas with the vapor.
Atomizers
An atomizer functions on the Bernoulli principle.
A tube is positioned in the center of a jet of air. This
tnl)e is immersed in the liquid to Ik* dispersed. The
liquid is aspirated up the tube and sheared off the
end. The size of droplets produced depends on the
diameter of the tubing at its orifice, the viscosity of
the liquids, and the rate of flow of air. The tulx; sup-
plying liquid may be concentric with the jet of air or
at right angles to it.
Concentric Atomizer*." These are commonly made
of glass, A capillary tube {A, Figure 2) is drawn
down to a tip and bent at right angles and sealed into
a bulb B. The tip of the capillary is adjusted so that
it is precisely centered in the orifice 0 of the bulb.
The annular space, between the orifice O and the tip
of the capillary is drawn to such dimensions that the
desired delivery is obtained at air pressures of 5 to
20 psi.
SECRET METHODS OF DISPERSING AG ENTS INTO CHAMBERS
287
the whole chamber is raised by a rack and pinion.
When the chamber is lowered, a gaslight seal is main-
tained by rubber gaskets. A constant air flow of
ISO 1pm is maintained by a combination pump and
meter driven by a synchronous motor. This is also
geared to the mechanism for delivery of the toxic
liquid, so that even if the motor should vary the same
proportion of toxic agent to air would be maintained.
The agent is displaced from a buret by a rising
column of mercury. It flows through stainless-steel
tubing to a small stainless-steel concentric atomizer.
The mercury column is connected through a U tnl>e,
omitted in Figure 3, to a brass cylinder filled with
oil. A stainless-steel piston, 0.250 inch in diameter,
is driven into the cylinder at a known constant
rate. This drives oil into one leg of the U tube, and
mercury out of the other. The piston is driven by
a lead screw, connected through a change gear box
to the synchronous motor. Three hundred and
eighty-five gear changes are provided.-A high-s|>eed
motor is belted to the lead screw for rapid return of
the piston.
The main air stream is divided so that 158 !pm
goes directly into the chamber while 22 1pm enters a
compressor which feeds the atomizer. The spray
from the atomizer enters a spiral ev aporator which is
provided with a flow of hot water of controlled tem-
perature; the air stream of the atomizer may also be
heated. Less volatile materials arc condensed on and
evaporated from the spiral coils.
An inverted mercury-water U tube provides an
estimate of the nominal concentration and a check on
the accuracy of displacement. In use, the mercury
level is set at the zero mark in the fust leg of the
U- tube. During the run, mercury is driven into this
bulb, displacing water, which displaces mercury from
the next bulb into the third bulb containing the solu-
tion to be displaced. At the end of the run the mer-
cury in the first bulb is drained off back to the zero
mark, and weighed. When the machine is free from
leaks, letter than 99 per cent recovery is obtained.
The Benesh machine is used as follows. Either the
density of the agent is determined, or a solution of
known density and concentration is made up. From
this is calculated the revolutions per minute needed
to give the desired .concent ration. The change gears
are then set to give the correct rpm. It. is usually
possible to select a gear setting such that the rate of
delivery is within 2 per cent of the amount desired.
Revolution counters on the carriage are set to give
the number of revolutions needed for a run of the
A Capillary. C Air inlrt.
II Hulb. H Rinjt mil.
O Orifice.
FmviiE 2. Concentric atomizer.
Maximum efficiency is obtained when the tip of
the capillary either extends slightly beyond or is
w ithdrawn slightly into the outer orifice. The adjust-
ment is most readily made by heating the atomizer
in the region of II (Figure 2).
The delivery rate and particle size are determined
by the dimensions of the t ip and the orifice.
A constant head device can be applied to the flask
so that the delivery does not vary with the level of
the fluid.
Concentric Atomizers — Constant Delivery Type.
Concentric atomizers supplied with liquid solely by
the Bernoulli effect are subject to variations in their
delivery rate. The delivery rate decreases as the
liquid level falls. Furthermore, the delivery rate is
influenced by fluctuations in pressure of the gas
driving the atomizer. At the UCTI. certain atomizing
units have been constructed which are provided with
liquid by a constant-flow motor-driven pump. Two
of these- are called Benesh machines, after M. E.
Benesh, Chief Engineer in charge of Research and
Testing of the People’s Gas, Light and Coke Com-
pany, who designed them.
1. The small Benesh machine (Figure 3).l-* This
machine consists of a chamber and an atomizer built
into one compact unit. The all glass chamber has a
volume of IS I. It can hold 10 mice, 7 rats, or 7 guinea
pigs. It is built with double walls between which a
suction of IJ4 inches of water is maintained. This
prevents leakage of toxic material. To insert animals,
SECRET 288
\PPAKATUS AND TECHNIQUES IN TOXICOLOGICAL STUDIES
Figure 3. Small Benesh machine, diagrammatic.
desired time. The animals are placed on the chamber
floor and the chamber lowered. The toxic agent is
placed in the buret. The machine is placed in gear
and started.
The exposure does not start until the first revolu-
tion counter is tripped. Prior to this, the agent is dis-
persed into the system, but exhausted before reach-
ing the ehamljer. When the first revolut ion counter is
tripped solenoids are actuated which turn off the ex-
haust valve and open a stopcock to admit mercury
into the measuring buret. The exposure continues
until the second revolution counter is reached and
thrown. This automat ically disengages t he motor and
turns on the exhaust valve. The animals are then re-
SFX’RRT METHODS OF DISPERSING AGENTS INTO CHAMBERS
289
moved. If a different concentration of the same, agent
is to be tested, it is only necessary to change the gear
and revolution counter settings. It is possible to
make as many as five 10-minute inns on the same
solution within an hour.
The maximum concentration that can theoreti-
cally be attained with any substance is one-eighth of
its equilibrium volatility at the temperature of the
heating coils. Since the air flow through the coils is
too rapid for saturation to take place, the actual con-
centration obtainable is somewhat loss.
The machine should not lx- used with substances
which attack mercury and stainless steel. In practice,
materials which react slowly with mercury can be
used.
2. The large Benesh atomizer. The atomizer on the
small Benesh machine was permanently connected to
one small chamber. It could not be used with aero-
sols, which came to form an increasingly important
part of the work. The large Benesh machine was
built to retain the advantages of the smaller machine
in a somewhat more flexible form. The essential de-
sign was retained, but the following changes were
made.
a. The volume of the buret containing the
toxic liquid was increased from 25 to 130 ml.
This permitted longer runs or higher'con-
centrations to lx- used.
1). The oil-mercury system was somewhat
cumbersome and prone to leakage. It was
made necessary by the use of a brass cylin-
der. Changing the size of piston used was a
major operation. In the large machine direct
displacement of mercury was made possible
by the use of an all-steel system.
Three concentric pistons, 1 inch, inch,
and ’4 inch in diameter are used. The two
larger pistons can be quickly locked down
and used as cylinders for the next smaller
size. The pistons and lead screw are mounted
vertically on a section of channel which can
lx- inverted for removing air bubbles from
(he cylinder.
The largest piston displaces mercury at
the rate of 0.0435 ml per revolution of the
lead screw; the other pistons displace in
proportion to their areas. The stroke is
about 10 inches, the maximum displacement
about 130 ml.
c. In place of the change-gear box on the small
machine, three pairs of standard loose
change gears connected l»v idlers are used.
These are changed by hand, with a wrench.
Twenty sizes of change gears an- available.
By using the several gears and pistons
available several thousand rates of displace-
ment are theoretically possible, ranging
from 0.008 to 7.000 nil min. In practice
both extremes are avoided, because of the
inaccniacy of the first and the high pres-
sures produced by the second.
The ease of changing geais in the small
Benesh machine made it convenient to make
up one solution and change concentrations
by changing the rate of displacement. With
the large Benesh atomizer it was frequently
more convenient to leave the gears set at a
certain ratio and make up different solu-
tions for the desired concentrations,
d. The atomizer, driver by a refrigerator com-
pressor operating at 15 to 70 psi, sprays
directly into the chamber rather than into
a condenser. This makes it possibleTo use
aerosols. Solut ions of agents of low vola) ility,
such as glycerine, may be used to produce
aerosols of a size determined by the concen-
tration of the solution. Thus 0.1 per cent
solution gives clouds of mass median di-
ameter jTM MD] 0.3 n and 5 per cent gives
an MMD of 4.0 n from droplets initially 7 p.
o. This atomizer has none of (he automatic
controls used on the small Benesh machine.
Return of the pistons is made by a hand
crank geared to the lead screw.
This atomizer cannot be used with ma-
terials with low boiling points, inasmuch as
no provisions were made for cooling the
storage buret. It should not lx- used with
materials which react with mercury or
stainless steel.
3. The small, constant-floic atomizer.2,: Many of the
features of the Benesh atomizers can be retained in a
very simple apparatus.
Standard, all-glass syringes are used as pistons and
cylinders. These an- connected to a small glass eon-
centric- atomizer. Connection to the atomizer may be
made by an all-glass system, but a short piece of rub-
ber tubing is preferred to prevent breakage. Syringes
ranging from 1 to 100 ml may Ik? used. They are held
in two metal brackets by rubber collars.
The lead screw is geared by bevel gears to a 1/150
hp Bodine synchronous motor geared down to
SECRET APPAKATUS AND TECHNIQUES IN TOXICOLOGICAL STUDIES
3 rpm. No provisions for changing the gear ratio are
made, and changes in concentration must lx* made
by changing the syringe size and the concentration
of solution. The machine is made to give IS minutes
of running, which allows for equilibration time and a
10-minutc exposure. The machine will deliver from
0.011 to 1.71 ml min with the various syringes. The
piston is returned by reversing the mot or and running
the machine I tack wants. The syringes can lie .cooled
by laying rubber bags filled with ice water across
them. This cooling is adequate for a 75 (ter cent solu-
tion of hydrogen cyanide in ethanol. This machine is
quite satisfactory. Nominal concentrations can i>e
estimated from the change in reading on thcAvringe.
It can Ihj set up on almost any chamber and used
with almost any agent. Slightly mom complex design
would provide changeable gear ratios and a quick
return device.
4. Modified Sinks atomizer.2" For work with the
wind tunnel an atomizer was needed that would give
an estimate of the amount of agent delivered at any
time during a run. A commercial spray nozzle was
modified for this purpose. The nozzle was a Binks
f 174 humidifier nozzle obtained from the Binks Mfg.
Company of Chicago. The body of this is a bronze
casting which contains a needle valve concentric to a
conical air passage. An indicator arm fitted with a
hairline was attached to the handle of the needle
valve. A 360-degree protractor dial was fitted to the
body of the nozzle. With these, precise and repro-
ducible settings of the needle valve could be made.
The fluid feed inlet was fitted with a 25-ml burnt ce-
mented into a brass sleeve threaded to fit the laxly of
the valve. Compressed air at. 10-25 psi was supplied
through a corrugated host* fitting threaded into the
lower inlet. At 15 psi and with a fluid feed of 2.0 ml/
min. a cloud of MMD 8.0 ju was obtained. Finer
sprays were obtained with higher air pressures and
slower feeds.
The principal advantage of this atomizer is the
direct reading of the amount of solution delivered
during a run. The output tends to vary somewhat
with the hydrostatic head in. the system. The atomizer
cannot lie used with substances which attack brass.
Impinging Atomizers.* In these atomizers the jet
of spray from the atomizer strikes a baffle plate.
Larger particles stick to the wall and run back to the
liquid reservoir, whereas smaller ones remain air-
borne and are swept out of the chamber, either by the
air blast from the atomizer itself or by an auxiliary
air supply.
Theoretically those devices could use either a con-
cent ric or a right-angle atomizing unit. To save space
the right-angle unit is commonly used. This unit
must bo adjusted before sealing into its container.
The size of particles obtained (and inversely the out-
put of the atomizer) is largely determined by the
distance of the, jet from the baffle plate. The wall of
the container may l>e used as a baffle, or a small plate
may be fitted in front of the orifice.
1. impinging atomizers.-1' These im-
pinging atomizers work at a very low efficiency. Per-
haps 5 per cent of the output of the atomizing unit
passes out of the nozzle as aerosol. This results in a
quite low delivery. It was necessary to develop an
atomizer which would set up larger amounts of ma-
terial as an aerosol for use in the wind tunnel. Im-
pinging atomizers of partially metal construction
were developed. The atomizing unit consists essen-
tially of a hollow brass tube, about an inch in diam-
eter, with the lower end plugged and the up|>er end
connected to the air line. Near the lower end #55
holes are spaced equally around the circumference.
A small brass tube, with the upper end machined, is
soldered to the body of the cylinder at right angles
to the axis of each hole. The lower end of t his tube
dips into (he liquid to l>e dispersed; the upper end is
centered in the jet of air from the hole. The most
successful of the atomizers has eight of these jets. A
shield in the form of a truncated cone is soldered
base down around the units to form a baffle. This
does not greatly affect particle size but it facilitates
return of fluid to the bottom of the bowl.
The vessel in which these are placed consists of a
1-1 Florence flask to the neck of which is sealed a side
arm of the same diameter. The shape and diameter of
(he side arm determines the particle size. 'Phis side
aim is fitted with a trap which returns liquid to the
reservoir, A bulge on the bottom of the flask provides
for efficient scavenging of small amounts of liquid.
The diameter of all tubing through which the aerosol
passes is kept as large as possible.
The eight-jet atomizer, operated with 5 cfm of air
at 20 lb pressure delivers from 1.5 to 2.3 g 'min with
agents of low volatility. These atomizers can produce
clouds of MM I) from 2.0 to 3.5 m-
These atomizers overcome the main drawback of
impinging atomizers, i.e., low delivery. Impinging
atomizers cannot be used with binary systems, as
they fractionate them, the more volatile component
distilling over. Impinging atomizers tend to give a
flat and fairly linear curve for output versus
SF.CRKT METHODS OF DISPERSING AGENTS INTO CHAMBERS
291
pressure, which makes fine adjustments in output
practical.
The Dispersal of Par tic elates
Certain of the atomizers used above may be used
for the dispersal of particulates, as solutions or
molten solids, as well as for vapors. There are in addi-
tion several methods peculiarly adapted to the dis-
jiersal of particulates.
The Dry Dusting Atomizer.'* '11' *1'' It became neces-
sary. to test the toxicity of dry dusts in comparison
with that of atomized droplets of solutions. The
“dry-duster” was developed for this purpose. It is
essentially an atomizer for dispersing dry powders.
The body of this atomizer is a straight tube, 25 mm
in diameter. It is separable in the middle by a ground-
glass joint, for ease in filling, A glass nipple is sealed
to the lower end. A sintered glass disk (40 60 mesh)
is sealed across the bottom of (he lower member, just
al>ovo the constriction. The powder is placed on this
sintered disk. A side arm, constricted distally, is
sealed to the upper member. A tube side of smaller
diameter is ring-sealed through the, opposite wall to
extend concentrically into the side arm.
In operation this device is charged with powder,
assembled, and placed in a flexibly mounted clamp.
A clamp attache's the assembly to an eccentric
mounted on the shaft of a small electric motor. The
vigorous agitation so provided tends to prevent
channeling, and to ensure a uniform rate of disper-
sion. The two concentric tubes sealed to the upper
portion constitute an atomizer. The Venturi vacuum
produced by passage of compressed air through the
inner member draws a current of room air through
the sintered disk. This current draws the particles up
to (he atomizer and into the chamber. Much closer
regulation of the output is possible if instead of re-
lying on the v acuum, a slow current (1 1pm) of dry
nitrogen is passed through the lower inlet.
A good estimate of the nominal concentration is
provided by the weight loss of this duster. The ap-
paratus disperses particles at. approximately their
original size. The shearing action of the air blast
shatters aggregates to a certain extent.
Fractionating Devices. As it is difficult to obtain
clouds of a desired particle size, a fractionating de-
vice is sometimes introduced between the atomizer
and the exposure chamber. Two forms of fraction-
ators have been used.
1. Fractionating lower. This device makes use of
the fact that the mass and volume of a particle de-
termine its rale of settling. This relation is formu-
lated in Stokes’ law. Hy passing a current of air up a
vertically mounted tube those particles which fall at
a velocity greater than that of the air current will
settle out; smaller particles with slower rates of fall
will lie swept up the tube. Such a tower may be used
to reject either small or large particles, depending on
whether the outlet for desired particles is placed at
the top or the bottom of the tube.
A tower of this sort was used for work with one
particulateS1h to exclude all particles al>ove 5 y in
diameter. One has been used for work with another
agent dispersed from an impinging atomizer in the
molten state.2"1 In this tower particles below 75 y
were sucked upward and rejected, while the larger
particles wore allowed to fall downward into a small
wind tunnel.
2. Rotating macro-impingcr.2" One of the princi-
ples widely used in analytical instruments for aero-
sols has been that of impingement. A jet of aerosol-
laden air is driven at high velocity against a surface.
The larger the particle, the better its chance of
sticking.
An attempt was made to use this method on a
larger scale to reduce the mass median particle di-
ameter of a dry aerosol (see Chapter 12). This was
done by impacting the dispersed agent at a high
velocity against a moving kymograph drum which
had been coated with vaseline. A moving surface was
used for impaction to prevent overloading. The
smaller particles which did not stick to the drum
were passed into an exposure chamber.
This equipment was able to reduce the MMD only
from 0.5 to 3.8 y. It was somewhat bulky and its use
was abandoned. However, this method of reducing
the MMD has certain inherent possibilities.
3. Serial mac ro-impi tigers.-'1 Large impingers (see
Figure 4) operated in series have been successfully
used to reduce the MMD of a NallCO* cloud below
1.0 y. Filtration flasks lined with vaseline are used,
with a central tube extending down from the top.
The size fraction taken out is regulated by adjusting
the position of this tube and varying the air flow.
This method is less cumbersome and more effective
than the rotating impinger.
Electrical Atomization.-1* This method of dispersal
is peculiarly well adapted to the dispersal as aerosols
of metals and other conductive, heat-stable materials
which are not obtainable in a finely powdered form.
The material to be dispersed is used as an electrode
for a high-voltage arc. If the material is to Im» dis-
SECRET 292
APPARATUS VXD TECHNIQUES IX TOXICOLOGICAL STUDIES
A Critical pressure orifice from rompressed air line at l.'i fisj (de-
livers 38 1pm).
B Manometer (maintained at 3 psi).
1 Flask boldine powder to be dispersed.
2 Air jet (directed to side to facilitate mixing).
3 Settling column (130 rm high, 5 cm in diameter).
4 Vaseline-coated impinger.
5 Mncro-imping«>r (FliliT filter fla>k runt ui nine Vasrlino and oil
mixture).
tl Maera-impinger (500-ral flo.sk as above). _
7 Mixing flask (2-15torK
8 Manifold.
*.♦ Stationary fan blades of o|)|>osing rotation.
10 Cotton plug (permits cmcapc of exeess air).
II;-12 Exits to mask and to sampler.
Figure 4. Dry cloud apparatus.
persed as an oxide, the arcing is carried on in an at-
mosphere of oxygen; otherwise helium or hydrogen
may be used. The aerosol is mixed with dry air and
led into a small chamber; regulation of the concen-
tration is achieved by varying the amount of diluent
air.
Aerosols obtained by this method are extremely
fine, somewhat less than 0.3 m in diameter. The out-
put of the are is quite constant, and regulation of the
concentration is made by regulating the air flow.
16.t SAMPLING EQUIPMENT
It is usually necessary to know not only the con-
centration of agent put up in a chamber, referred to
as the “nominal” concentration, but also to know
the concentration actually existing in (he chamber.
This concentration is determined by chemical or
physical methods anil referred to as the “analytical”
concentration. With vapors, it is only necessary to
know how much of the material is dispersed in a cer-
tain volume of air. With particulates, in addition to
this information, it is necessary to know something
about the size of the individual particles.
16.1.1 Equipment for the Sampling of
Vapors
Most of the apparatus for determining the concen-
tration of vapors in gassing chambers is of common
use iii the study of air pollution.34 UCTL practices
are as follows.
1. Withdrawing of the gas sample is effected by
either a water aspirator or an electrically driven
pump.
2. Measurement of the-vohttne- withdrawn is usu-
ally made by a wet test meter. This meter is cali-
brated by the positive, displacement of a known vol-
ume of air through it. Familiarity with methods used
in field trials led to the use of critical pressure orifices
for regulating sampling rates,
3. The type of absorbing bubbler most commonly
used in this laboratory is made of glass. A coarse
sintered disk is used to break up the gas into bubbles.
An investigation was made of.the efficiency of
three types of bubblers,2,i the sintered disk type,6
the Bushnell type, which has a plain inlet tube ex-
tending about 5 cm into the absorbing liquid, and
the Kdgewood type, which is filled with glass beads.
The absorption of H was studied, and a Northrup
titrimeter used to measure the slippage. The sintered
disk type was found to be (he most efficient. The fol-
lowing conclusions governing the ust* of bubblers
were drawn from this study and coincide with others
independently obtained.'5
1. The absorbing solvent should have a low vapor
pressure.
2. If possible the solvent should react with the
absorbate to give a nonvolatile compound.
SECRET suipum; equipment
293
3. The absorbent should dissolve water vapor if
the air has an appreciable humidity.
1. A solvent which foams considerably is to lx*
preferred to a nonfoaming solvent, other factors lx*-
ing equal.
5. Low tem|x*ratures are conducive to lx*tter ab-
sorption.
6. The flow rate of the gas should lx* kept as low
as possible.
7. The kind of bubbler is of less importance than
has usually been assumed.
Two devices for the analysis of vapors have been
developed at the UCTL. They are described below.
f.ow-1{(.Asia nee Absorber.27* Investigations of the
hydrogen cyanide content of expired air (“precision”
gassing) required a bubbler that would combine low
resistance, small volume of absorbent, and small
deail space with high efficiency at intermittently high
flow rates. It was necessary to design a new type of
bubbler to meet these specifications.
This absorber consists essentially of a pair of con-
centric glass tubes. The outer is 37 cm long and
2.2 cm inside diameter. Inside, it is a tulx* with both
ends closed, 33 cm long and of such a size as t o leave
a 1-mm annular space between it and the outer tulx*
as an air passage. The outer tulx- is fitted at its ends
with male ball joints so (hat it can l>e freely and con-
tinuously rotated about its long axis (Figure 5). It is
cent, when tin.' (low into is 30 Ipiii. Only 30 nil of wash
liquid arc needed, as compared to 500 ml needed for
a bead bubbler of somewhat lower efficiency. The
absorber and its motive power form a fairly compact
unit.
An Electronic Interval Timer for the Northrup Ti~
trimeter.'2 The Northrup titrimeter is an electro-
chemical analytical instrument* for the quantitative
determination of the airborne concentrations of
chemical warfare agents (sec Chapter 30). A sample
of contaminated air is drawn in at a constant rate.
At intervals it is titrated with a dilute bromine solu-
tion. The titration is carried on in one half of an
Ag AgNOj Br2 Br cell, with a platinum indiffer-
ent elect rislc. When oxidation is complete, an excess
of bromine creates an electrical potential, which is
recorded on a galvanometer. The amount of bromine
solution needl'd is determined by the time required
for it to flow from a constant-head burnt.
This instrument is made in two forms. In the sim-
pler field model the bromine solution is run in by the
operator, who shuts off the flow when the galva-
nometer shows a positive reading. In the automatic
model the galvanometer mirror reflects a beam of
light on a photocell when the titration is complete.
The photocell actuates relays which shut off the
buret and start another sampling period. The length
of sampling period is controlled by fixed cams, which
give a choice of four periodicities; from I minute
sampling and 2 titrating to 50 minutes sampling and
10 titrating. Bet ween the end of one titration and the
start of the next sampling period (he cell is kept in
equilibrium — the agent sampled during this period
is balanced by intermittent addition of bromine. The
owning and closing of the bromine buret is recorded
by a relay-actuated marking pen writing on a paper
record wound around a kymograph drum.
In this form the automatic model was incapable of
accomplishing some of the determinations that were
desired at L’C'TL. In particular, the shortest time
interval available (cycle repeated every 3 minutes)
was too long for showing variations in concentration
occurring at a frequency greater than that, whereas
the provision for 1-hour sampling periods was un-
necessary. The cyclic rate could have been increased
by cutting another cam. However, it was desirable to
eliminate the time lost between the end of.one titra-
tion period and the start of the next sampling period
which results from the use of the cam timing mech-
anism.
An electronic method was adopted. The sampling
Fin cue 5. Low-rcsistancc absorber.
rotated by two micarta pulleys, 2 inches in diameter,
boitil out to fit the outer tube, and cemented to it.
These pulleys rest on t he rollers of a small ball mill
(Fisher Minimill). The whole assembly is mounted
on two rods attached to the sides of the ball mill.
Two brackets hold the corresponding female ball
joints flexibly. The absorber is held down against the
rollers by helical springs attached to slip rings.
In use S ml of absorbent are poured into the ab-
sorber. This is more than enough to wet all exposed
surfaces when the absorber is rotated. Thereby the
absorbing surface is continually being renewed. Tests
have shown resistance to lx* very low, about 1 cm of
water at an air flow of 30 1pm. When 3 per cent
NaOH in ethanol is used as an absorbent, the absorp-
tion of hydrogen cyanide is 100 per cent from air con-
taining 2. t mg 1 and flowing at 7 1pm. It is 05 per
SECRET \PPAR ATI'S AND TECIIMQL'ES IN TOXICOLOGICAL STID1ES
time interval is governed by the time required to dis-
charge a condenser of high capacity through a high
resistance. The resistance was controlled by a po-
tentiometer; changing this set ting varied the time of
discharge, ami hence the sampling rate. The sizes of
the elements used were such as to give continuous
variation between 0.25 and 5.8 minutes; a longer
sampling jx'riod proved unnecessary but use of larger
condensers would provide it. At the end of the sam-
pling period titration starts and continues until all
the agent collected during the pretit ration period
plus, that collected during the titration period is
titrated. Thereupon the titration stops and essen-
tially instantaneously a new cycle begins.
This addition to the laboratory model Northrop
titrimeter has the following advantages.
1. Continuous variation in sampling times is avail-
able merely by turning a knob.
2. As soon as one titration-period has been com-
pleted, a new sampling period begins.
8. By using short time intervals the concentration
of agent in the absorption cell is kept very low at all
times, thus reducing the loss of material by slippage.
4. The original instrument is now adapted for use
in determining concentration changes in gassing
chambers over short periods of time.
Ih. 1.2 Equipment for the Sampling of
[’articulatesfi
Filters. One of the simplest ways to determine the
concentration of a smoke is to draw a measured sam-
ple of air t hrough an efficient filter and determine its
gain in weight. At UCTL much early work with
smokes was done with cotton-asbestos mats (40 per
cent cotton — 00 per cent asbestos) I to 2 mm thick,
pressed into perforated or sintered glass filtering fun-
nels (25 mm in diameter). Suction of from 0.0 to
2.0 inches of mercury was needed to pull 3 to I 1pm
through these.
Work on certain types of aerosols (see Chapter 12)
introduced several new requirements for a filter.11'1
Since the determinations involved a micro procedure,
it was necessary that the filter material have a low
blank (less than 20 /ig). The filter chosen should not
lie clogged by as much as 10 mg of a standard prepa-
ration and should offer low resistance to air flow.
Several filter papers were tried Ix-fore one made from
cellulose acetate was found to l>e satisfactory. This
paper contained no material simulating the material
determined in the analytical procedures. Insoluble
material could lie completely floated off, ami (he bait
could lx1 completely dissolved in a suitable organic
solvent, leaving the particles unaffected and ready
for counting.
In use. both in the laboratory and in the field disks
of the paper are stamped out. They are held in brass
holders in which they are backed with a wire screen.
Precipitators. Particles have been removed from
the air by direct precipitation. A Watson 40 thermal
precipitator has been constructed and used. In this
instrument the air passes over a Niehrome win- kept
at 100 C. Smokes are precipitated on cover slips
backed by brass blocks.
Electrostatic precipitation has also been used in a
small Cottrel-type precipitator.33 This is essentially a
long glass cylinder. Copper screening is wrap|xnl
around the outside, and attached to one terminal of
a I 5,000-volt transformer. A wire in the tube at its
long axis forms the other pole.
Impinging Devices, An impinging device is one in
which the smoke-laden air is drawn or driven at high
velocity against a prepared surface. The particles
may be trapped on the baffle plate or absorbed by
some liquid medium. Impinging devices may housed
either to collect all of the particles in one stage or to
fractionate the particles by using jots of various
speeds.
1. The atomizing imping/r.21' This unit (see Fig-
ure 6) consists of a concentric atomizer mounted in-
A Inlet f uix-
B Battle (supported by ted from .W not shown).
C Capillary.
r> Distanrr- between cajiillary and battle.
K K\it to vacuum.
F Flui.l to be atomized.
Fiiiimt; R. Atomizing irnpinger.
side of a glass vessel in such fashion that its jot
strikes a baffle plate and drains down to a sump from
which it is re-atomized. Dust particles in the incom-
ing air ring strike the baffle plate and are trapped.
A straight tube is ring-sealed concentrically
through tho top of a side arm test tube. The inner
tube is constricted at its inner end to give a jet. A
secret SAMPLING EQUIPMENT
295
smaller tube is ring-sealed through the wall of this
inner tube in such a fashion that its outer end ex-
tends at right angles, and its inner end is concentric
to and extends through the constricted portion,
forming a concentric atomizer. Tins outer end dips
down into a bulge* on the outer tube which acts as a
sump. The apparatus is mounted horizontally. A
glass arm supports a baffle plate which is carefully
positioned in front of (he jet. If this plate is too close
to the orifice the atomizer will not function; if too far
away the unit will act as an atomizer and not as an
impinger. A source of vacuum is attached to the side
aruuand the inlet tube connected to the charal>er.
This pattern has proved 1)0 per cent efficient in the
collection of clouds which had 1 »eon allowed to settle
for 30 40 minutes and contained particles with an
MMD id 6 ju. A practical advantage of the apparatus
is that the collecting volume is small and, conse-
quently, small amounts of toxic agent are not diluted
too much for injection into animals.
The Cascade Impactor. The construction, method
of use*, experimental results, and theoretical principles
of the cascade impactor are fully described by K. it.
May.’*' Experiments at the UCTL have emphasized
the desirability of the instrument with dry particles
and have employed slightly different, calculation pro-
cedures, such as (hi* substitution of the MAID on
each plate for the effective drop size [EDSj used by
Port on. With dry clouds a total sample of about
0.350 mg represents the maximum which can be ob-
tained without overloading if the cloud is distributed
on all four slides. W ith dry powders the presence of
aggregates in the airborne cloud complicates the
calculation of the MAID. These aggregates fre-
quently break up upon impaction so that they can-
not be measured microscopically. Since the density
of an aggregate is lower than that for unitary parti-
cles, the aggregate is impacted along with unitary
particles of smaller size. 'Phis property leads to the
recommendation (Chapter 15) that particulates
should be assessed in terms of impactibility rather
than in terms of diameter.
1. .4 device for increasing the load on cascade im-
pactor slides.-'11 The amount of a particulate impacted
upon slides No. 3 and No. 4 of the cascade impactor
must l>e kept very small to prevent overloading. The
quantity obtained on one streak is barely within the
limits of the available analytical methods. To allow
collection of a larger sample, a method of moving tin;
slides at intervals during sampling was devised so
that it is now possible to obtain eight streaks on
slide No. 4 and four streaks on slide No. 3 during one
sampling period. A heavier cap is screwed on to the
appropriate tubes of the cascade. In the center of (his
cap a hole is drilled and tapped for a '.pinch bolt.
The Iwdt is passed through this cap. The inner end
bears on the slide. This bolt is turned by band at in-
tervals during the run to move the slide. The distance
it is moved .is determined by the numlxuof rotations
of the screw.
2. .1 modified cascade impactor for use with small
particulates.-1'' The cascade impact or was originally
designed to handle the range of drop sizes set up by
munitions in (he field. In work with nasal filtration
of small droplets it was necessary to work in a range
of much smaller sizes. The larger drops in this range-
were of about the smallest size that the standard
cascade impact or would handle at 17.5 1pm. The
standard cascade with critical orifice impinger and
filter )tacking would trap the particles in this range
but would not fractionate them.
A modified cascade was constructed which frac-
tionated the drops which slipped past slide No. I and
were caught by the impinger. The standard cascade
lavs a rate of flow through its jets of 5, 30, 50, and
SO mph, with the impinger giving 700 mph. The mod-
ified cascade has jet velocities of 56, 80, 177, and
700 mph, the last slit being a critical orifice. (A criti-
cal orifice gives a speed of How equal to the speed of
sound — approximately 700 mph.) ft will lie noted
that the first two jets of this cascade correspond,
roughly, to the last two jets of the standard cascade,
while the last two correspond to the impinger, as used
with the standard instrument, and an intermediate
value. This modified impactor has proved capable of
efficiently fractionating a cloud which slips past the
standard instrument. This modified instrument re-
quires a backing filter to collect all material.
The present experimental model is blown of glass.
The four separate sections fit together with rubber
stoppers. The slides arc held in place between in-
dentations in the walls. Wall losses can be easily
detected by inspection.
Particle Fractionators. Drop traps, chemical noses.
In an attempt to simulate the characteristics of nasal
passages with respect to particulates, various devices
have been made to fractionate the cloud into a lung
fraction and a nose fraction.
Glass tubing in the form of Z’s and S’s such as
were used by British workers for liquid droplets were
coated with a sticky film of alkyd resin in an effort
to fractionate dry particulates.-^30 Better results
SECRET VI’IVVR \Tl S VM) TECIIMQI KS tX TO \ I COM )<; ICA L STUDIES
were obtained with selectors which were essentially
the first stage of a cascade impactor backed by a
filter.2"
Wires fur Sampling Particulates. The use of slides,
tubes, and wires of different dimensions for the de-
termination of particle size and cloud concentration
has been described in detail.27“ b e
16.5 MKT]lODS FOR "PRECISION
GASSING’’
In the usual gassing procedure no account is taken
of the effect of the toxic agent on the respiratory
volume or rate. Consequently, there is no means of
determining the inhaled LIh» from the LC:,„. Some
species, especially rabbits, hold their breath to agents
which are apparently undetected by other species.
Methods which take account of the actual respira-
tion during exposure have been termed “precision
gassing” methods. A tracheal cannula and Douglas
bag were employed in studying the effects of phos-
gene on the respiratory pattern of dogs.211' In some
instances the respiration was modified by the use
of CO;.-,f
For the early investigations on the effects of hy-
drogen cyanide, the animal was mounted in a body
plethysmograph attached to a Rrodie bellows.
More precise methods were later developed 27b r-r
in which a mask was fitted to the animal. A valve
with minimal dead space was used and a special low-
resistance absorber const ructed.
16.5.1 Equipment
The Mask. The first mask tried was made of
vinylite sheeting, shaped in a truncated cone. The
ai>ex of this was cemented to (he male half of a 16 22
standard taper joint. This was used with dogs. The
animal’s snout was taped shut, and the cone bound
over it so that the apex of the cone was in contact
with the nostrils.
When this facepiece was applied in the usual man-
ner, an average leakage of 17.5 j>er cent was found.
When it was applied very tightly, the leak was re-
duced to an average of 1.4 per cent. In order to
achieve this low leakage the facepiece had to be ap-
plied with sufficient pressure to embarrass respira-
tion seriously.
This facepiece subsequently was replaced by a pair
of nasal tubes leveled at one end to facilitate in-
sertion. These tubes (1 '4 inch long, and 4 mm inside
diameter) were attached by paragum rubber tubing
to the diverging arms of a glass V tube scaled onto
tiio end of a 15 20 male standard taper glass joint.
The tube ends were closely approximated to the
Y arms so as to expose as little rubber surface as
possible to the agent. After gassing no adsorbed agent
could be detected on the inside of the nosepiece.
It was possible to handle animals so intubated with
local anesthesia alone (cocaine or butyn) but this
was not entirely satisfactory. Therefore intubation
was carried out on lightly anesthetized animals (for
dogs, 20 mg kg of Nembutal intravenously) after a
swab of 1 per cent cocaine or butyn had been applied
to the nostrils to prevent sneezing. The month was
closed with elastic bandage and the tubes were then
inserted and fixed with additional taj>e. No leakage
could be detected in 11 of 12 animals tested, and the
other had only 1.9 per cent leakage.
The Valve System. The first valve used was a
copy of the all glass valve designed by Weston and
Tobias.'5 The laxly of this valve was made from two
female and one male 16 22 joints sealed together in
a 1'. The long arm was made from a male and female
joint, and the side arm from a female joint. In use
the long arm is mounted vertically with the male
joint at top. The valves proper are composed of glass
disks ground flat, of such diameter as to fit inside the
female joint. The end of the male joint is ground flat
and serves as a valve seat. The disks are held down
in place by their own weight. The assembly of male
joint and disk is inserted in a female joint which
keeps the disk in line. Such a valve will pass air in
the direction away from the male joint.
In use, the dog’s snout is connected to the female
joint on the side arm. The lower valve is connected
to a gassing chamber, and the upper, through a suit-
able absorber, to a spirometer. On inspiration the
lower valve opens and permits contaminated air to
enter. On expiration, the lower valve closes, and the
upper valve opens and passes air to the absorber.
It is desirable to minimize the dead space as much
as possible. The use of glass imposes limits on the re-
duction which can be made. In addition the convul-
sions of exposed animals place great strain on the
valve.
It was possible to machine a valve from brass
which would have less dead space. The valve is of the
same general design, except that both the laxly of
the valve and the flaps arc made from brass. The
valve lias only 13 ml of dead space as compared to
25 ml in the glass valve, and is practically unbreak-
able. It was found that while untreated brass ab-
SECRET METHODS FOll "PRECISION GASSIXo”
297
sorlied appreciable quantities of cyanide, brass
“blued” by immersion in a hot solution of As.()3 in
HC1 did not react with cyanide.
The Absorber. The use of an ahsorlier to collect
gases from air, as expired, imposes certain peculiar
requirements. It must be efficient at intermittently
higli flows (as much as 30 1pm), it mus‘ have low re-
sistance (conventional absorliers have a resistance of
several inches of mercury under these conditions),
and it must lie possible to rinse it out with a small
volume of liquid, since very small quantities of agent
are present in the expired air.
The first absorber used was the Edge wood low-
resistance, glass-bead type.25 This absorber is filled
with fluid and drained just before use. Thus, the gas
passes over the surface film and not through liquid.
However about 500 ml of wash liquid was needed to
transfer all the ahsorlied agent to the titration vessel.
With the very small amounts of material present,
this large volume of solution led to appreciable titra-
tion error. The UCTL low-resistance absorber-7'1
described (Section 16.1.1) proved more satisfactory.
16.5.2 Determination of Inhalation
Toxicity of Particulates
As descrilied in Chapter 15, the effect of particle
size on physiological action is appreciable. In order
to determine, whether an inhaled particle was trapped
in the nose or in the lung or whether it was exhaled
again, various methods were devised similar in prin-
ciple to those employed in “precision gassing” experi-
ments with vapors but moilifieri for the assessment of
particles.
1. One procedure for use with animals involved
exposure to toxic clouds of controlled particle size.
It was used with agents which are highly toxic in the
lung but of negligible toxicity in the nose. From these
experiments the relation of particle size to toxicity in
mice, rats, and rabbits was determined (Chapter 15).
2. Correlated with these experiments were meth-
ods for the determination of toxicity by intrapul-
monary instillation of solutions of the toxic agent.276
In general the trachea of an anesthetized animal is
canmilatcd, and the solution instilled into the trachea
through a tube which fits inside the cannula. The
method has previously been used for rats.35 A small
catheter is used. The neck is transilluminated with a
Spencer microscope lamp to aid obscuration of the
trachea.
For mice this method was modified as follows. A
cannula whic h fits snugly into (lie trachea of a 20-g
mouse is made by rounding off the beveled (ip of a
1 j-2-ineli 18-gauge needle. A lJ4-ineh handle is at-
tached to the hub. The solution to be instilled is con-
tained in a '4-1111 tuberculin syringe tipped with a
25-gauge needle. The mouse is etherized and tied on
its back. The mouth is held open and the tongue held
against the mandible with a pair of blunt forceps.
The throat is transilluminated and the larynx is
visible as a bright spot opening and closing with the
respiratory movements. The cannula is inserted into
the trachea through the larynx. When the cannula is
situated correctly, it is possible to cause a pulsation
of the chest, by blow ing gently into the cannula. 'The
25-gauge needle is then inserted into the cannula,
and the solution instilled. When in position, the tip
of the 25-gauge needle should ice Hush with the (ip of
(lie 18-gauge cannula. If the cannula is in the right
position, the breathing becomes labored upon in-
sertion of (lie 25-gauge needle, and returns to normal
when the needle is removed.
16.5..1 Nasal Filtration and Lung
Retention 21fc 1
The mensuration of these factors involves setting
up a particulate cloud and determining its particle
size with cascade impactors before and after passage
through portions of the human respiratory system.
Obviously these tests could be done only with non-
toxic materials. Either non hygroscopic or hydrophi-
lic aerosols may be used; dyed corn oil and calcium
phosphate were used for the former, and NaHCO*
for the latter (see Chapter 15). The basic techniques
involved in these determinations are (lie same; differ-
ences will be discussed under the subheadings.
Selling up the Particulate Cloud. In the first work
com oil (Mazda) dyed with Sudan Red was sprayed
from the large Benesh atomizer. When NaHCO;) was
used, it was dispersed from a 250-rnl Erlenmeyer
flask with a two-hole stopper. A glass tube drawn
out to a fine tip extended through the stopper to
near the bottom of the Ha.sk. When compressed air
was forced through this jet, a cloud of NaHCOa dust
emerged from a tube in the other hole of the stopper.
This cloud was directed into a 12-1 bottle, and the
large particles allowed to settle out for 5-10 seconds.
The particles remaining airborne were then drawn
into the common entrance of the Y tube. It was
found early in the experiments that the cloud dis-
persed as described was quite heterogeneous. Though
SECRET VPl'AK ATU S AND TECHNIQUES IN TOXICOLOGICAL STUDIES
passing through a common tube, samples drawn si-
multaneously from the two arms of the Y tube
showed markedly different distributions on the im-
paclor slides. This was remedied by placing two
small, oppositely oriented, metal propellers in the
common tube. This stirred the passing air sufficiently
to make the cloud uniform.
The same setup was used for calcium phosphate
smokes. Full her fractionation was provided by the
use of serial Tnaero-impingers, lined with Vaseline
(see Figure 4).
1. Dry rlnini apparatus. The dry cloud of NallCOj
was set up by the apparatus shown in Figure 4. Fif-
teen pounds pressure in excess of atmospheric applied
to a critical pressure orifice for IS 1pm (at atmos-
pheric pressure) gives a flow of about 38 Ipm, suffi-
cient to supply the lung and the control without di-
lution by unfiltered room air. The material enters the
settling column. The agitation here is adequate to
maintain the cloud at-nearly the same concentration.
The pressure in the manometer w as about 3 psi,
practically all of it living due to impinger No. 6. The
jet in impinger No. 4 was simply a large glass tube.
In No. 5, the end of the tube was somewhat flattened-,
whereas in No. 6 the orifice in the end of the tube
was about 1x5 min. For smaller clouds a still smaller
jet may be used, the pressure in the column-being"
larger. To maintain the same flow rate only a slight
shift in the initial pressure is required. If one starts
with considerably higher initial pressures with a
proportionately smaller orifice in No, 1, changes in
pressure in the column may l>e ignored.
Between runs the impingers were warmed to re-
surface the bottoms of the flasks and the mixing
flask 7 and tube 8 were blown clean. At the end of a
run the last flask should not be too heavily coated.
Passage of the Cloud through a Portion of the Human
Respiratory System .The expo rimen t s conduc ted were;
1. Nasal filtration with corn oil. The oil was
sprayed from the Benesh atomizer into the 700-1
chamlier. This was operated at a flow of 300 Ipm and
acted as a settling chamber. A glass Y tube. 22 mm
in diameter. led from the chamber. One branch of
(his Y led to a mask which fit ted tight ly over the nose
and mouth of the subject. This mask was from a com-
mercial dust respirator; an inflatable rubber tube
formed a tight gasket with the subject’s face. Pro-
truding into the mask was a second exit tulx*
about which the subject closed his mouth. The
exit tube led through a cascade impaetor, backed
by an impinger, to the pump. The second branch
of the Y tube led to another impaetor, also hacked
by an impinger and the pump. This impaetor sam-
pled the incoming cloud. The tubing between it
and the fork of the V tube was comparable in
length and shape to the tubing which led to and
from the mask. As far as could be controlled, the only
difference between the two air streams was that one
passed through the subject’s nose and the other did
not. The impingers backing the impactors were of
such size as to be critical orifices, with a flow of
17 Ipm. Ivtch experiment was done in duplicate with
the positions of (he control cascade with its corre-
sponding impinger, and the mask exit cascade with
its impinger, reversed during the second experiment.
Th is canceled out instrumental errors.
Sealed to the mask exit tube-and preceding the
cascade impaetor was a small glass tulie through
which air could be drawn from or added to the sys-
tem. This could lx1 used to increase or decrease How
rate through the mask while maintaining the same
flow through the cascade impaetor. A similar tul>e
preceded the control cascade. A second small opening
in the tube which connected the mask to the cascade
impaetor led to a manometer which indicated the
reduction in pressure caused by the resistance of the
nose when air was flowing through it.
During each experiment, which lasted 30 seconds,
the subject held his breath so that there was no ap-
preciable passage of the aerosol into and out of his
lungs. For the flow rate of 10 1pm, the 30-second rim
was repeated immediately in order to obtain a larger
sample. The nasal resistance was found to be very
low excel)! in individuals whose nasal passages were
congested or who were not sufficiently relaxed during
the experiment, [f a person is tense, his posterior
nares may become constricted, with a marked in-
crease in resistance. Subjects were used in the experi-
ments only after they had had sufficient practice on
t he apparatus to allow a 10 1pm flow with a resistance
of 0.3 inch waf er, 17 Ipm with 0.5 inch, and 2!) 1pm
with 1.0 inch or less. In the case of some subjects this
low resistance could often be achieved only after in-
halation of benzedrine, or the nasal instillation of
neosynephrine solid ion.
2. Nasal filtration with NnllCO* particles. Com-
mercial powdered NaHCOj was chosen as a nonirri-
tating, non aggregating powder containing particles
of a size range from 1 to 15 a (microns). The same
assembly of mask, Y tube, impactors, critical pres-
sure impingers, and pump was used as with the corn
oil work. The impaetor slides were covered with
SKCKET METHODS FOK TESTING VESICANTS
299
alkyd resin, and the NaTICOj on them analyzed
elect romet rically.
The hygroscopic!tv of XallC’Oa introduced several
difficulties In-cause of the moisture picked up by
passage through the nose. It was necessary to oven-
warm the impactor Indore use, and to warm the cloud
from the nose by passing it through an 8-inch length
of 15-mm tubing, electrically heated to give an
emergent air stream of approximately 90 C. A dupli-
cate heating device was used in the cent ml air stream.
The wetting and subsequent drying of the cloud
passing through the nose compacted and rounded
the particles in it. It was necessary to humidify the
control cloud also. The humidifier was a metal tube
(1 foot long, 1 inch inside diameter) lined with a
water-soaked blotter, placed in a thermostated water
bath. This humidifier was also an inefficient im-
pactor, taking out about 20 per cent of the airlmrnc
material in the cloud going through it, A similar tula-,
lined with Vaseline, had to be placed in the path of
the cloud to be transmitted through the nose. After
these modifications quantitative agreement lx-tween
the NallCOj contents of the dry and humidified
cloud could l>e obtained. With the use of cellulose
acetate filters instead of the cascade impactors it was
unnecessary to have the various driers.
3. Retention of particles in human Jungs. In these
experiments the subject’s nose was plugged. Stop-
cocks were placed in each of the two sampling tubes
between the critical orifice and the filter.
During an experiment the subject inhaled for a
fixed period, the beginning and end of w hich were in-
dicated to him by an operator. During the inhalation
period a sample of the incident cloud was drawn
through filter A, by opening the corresponding stop-
cock (the vacuum pump operated continuously).
During exhalation, which was also for a fixed number
of seconds, the exhaled cloud was drawn through
filter H. The cycle was repeated 10-15 times,depend-
ing on the length of inhabit inn and exhalation periods.
Since the incident and exhaled clouds were sampled
at the same rate and for equal periods, the material
found in the exhaled cloud represented the unre-
tained fraction of an inhaled quantity equal to that
on the other filter.
The total volume breathed during an experiment
was governed by the rate of withdrawal of the ex-
haled cloud from the mouth. The volume withdrawn
during each period was considered to be the tidal air.
The exhaled tidal air was, of course, constant from
period to period since it was controlled by the sam-
pling instrument. The inhalotl tidal air, however,
varied from period to period depending on whether
or not the subject inhaled a volume which exactly
compensated for the amount withdrawn during the
previous exhalation. Over a number of cycles, of
course, the average volume inhaled had to be equal
to the volume exhaled.
This method has been studied with smokes of
NatiCOj and calcium phosphate. Since calcium
phosphate is nonhydroscopic it is possible to dis-
pense with the humidifying and drying sections of
t he apparatus,
16,6 METHODS FOR TEST I ISO VESICANTS
The usual method for testing the vesicancy of an
agent is to put a known amount of it on the skin of
the forearm and to observe the results at a later time.
The agent may be put on as either a liquid or a vapor;
it may be either still or flowing.
16.6.1 Testing Vesicants as Liquids
The Edgcwood Rods* n One of the simplest meth-
ods for testing compounds for vesicant action in-
volves the use of a series of stainless-steel rods of
standard weight with tips varying from 0.0 to 2.OS
mm in diameter.” With the exception of the smallest
rod, all of them deliver 0.022 to 0.029 mg of H per
square millimeter. These rods, usually known as
“Edgewood rods,” are touched to the surface of a
pad saturated with the vesicant anti then applied to
the skin. Liquids were used either undiluted or dis-
solved in diphenyl ether. Solids were also dissolved
in diphenyl ether.
The method is simple and although all the material
on the surface of the rod is not delhered to the skin,
easily reproducible burns result from the iise of a
given rod with a given compound. In general the
rods have not proved satisfactory for comparison of
vesicants since a separate calibration is required for
each compound tested. It is not possible to test oint-
ments with these rods, since the droplet cannot satis-
factorily 1m‘ delivered to the surface of ointment-
covered skin without breaking the covering. Further,
the method is not desirable for compounds that react
with steel, although it has been "used with lewisite.
Nonstandard sets of glass rods have also been made.
”Drod" It was necessary to devise some form of
micropqx't which would deliver small, known vol-
umes of vesicant. Trevan v> used a standard microm-
eter caliper to drive a 1-cc syringe. A modification 2
SECT! ET 300
APPARATUS VXD TECHNIQUES IN TOXICOEOC.fCAL STUDIES
called the Drod used a specially constructed microm-
eter head to drive the plunger of a 1 j-ee tuberculin
syringe. A spring click bears on 12 longitudinal
grooves on the barrel of the head, each click corre-
sponding to 31) degrees of rotation and the delivery of
about 0.2 cu mm of liquid. (The amount would Ik*
constant for each syringe, but commercial syringes
are not interchangeable, being individually ground
to fit. The diameters of the pistons, and hence the
volume delivered, vary between syringes.) A 27-
gauge needle, w ith the tip ground flat and square, is
used to deliver the liquid.
The instrument is sturdy and portable. It delivers
an accurately measured small dose, which is not so
dependent on the physical properties of the agent as
is the case with the Edge wood rods. This apparatus
requires considerable time to fill, and the change
from one vesicant to the other n*qwires decontamina-
tion of the syringe and tip, making it unsuitable for
use when many different liquids are to be handled in
one day. The amount of liquid delivered per click is
large, with (he result that dilutions must frequently
be used.
The Modified Drod/ An attempt was made to
modify the original Drod to make it. more suitable.
Several modifications were made on the driving head.
It was redivided, so that a click occurred for each
7.5 degrees of revolution (48 clicks per revolution).
The instrument then delivered 0,065 mg of H per
click, instead of the original 0.2 mg. A 6-inch indi-
cator disk, with 192 divisions, and a pointer arm at-
tached to the head made it possible to split- the clicks
in half, and possibly into four. These are equivalent
to 0.032 mg and 0.016 mg of mustard.
The 1 pee tuberculin syringe was retained. It is
filled w ith mercury, which is used to expel the agent
from a removable delivery tip. The syringe and screw
are attached by a ground joint to a three-way stop-
cock. With the sto(>eock in one position, the agent in
the tip may lie expelled by turning the micrometer.
With the stopcock in the other position, the lip may
be filler! or washed by liquid which enters from a side
arm. A platinum or graphite surfaced stopcock is
iisihI to avoid fouling the agent.
It is possible with this modification to remove one
vesicant, decontaminate the apparatus, and load an-
other vesicant in less than I minute. It was found
that dividing the clicks into four did not give repro-
ducible lesions, but 0.032 mg of mustard, correspond-
ing to half clicks, can he delivered quite accurately.
This amount, although small, was not small enough
for some purposes. The increments were too coarse
to discriminate I>etween vesicants of nearly similar
potencies.
Other Pipits. Capillary tubes have been used for
the application of measured amounts of vesicant.24
The capillaries were however rather fragile and the
method is not adapted to testing large numbers of
men. A device for blowing drops of measured size off
a microburet tip was developed at Port on.3' 32
The Be tush Micropipet.1* In the I) rod type of
micrometer syringe the piston was of a diameter
equal to the'bore of the cylinder. To achieve a smaller
displacement with the same pitch lead screw, it was
necessary to reduce both bore and piston diameters.
The 1 j-cc syringe already in use was the smallest
available size. Micropipets capable of delivering
smaller quantities of liquid have previously been de-
scril »ed.3r*8 Various features in their design were not,
however, suited to vesicant testing.
The lienesh micropipet was based on a some-
what different displacement principle. The piston
was a steel wire 0.0122 inch in diameter. This entered
a mercury chamberthnaigh a Neoprene gasket, The
volume of mercury displaced was equal to the volume
of wire which entered the chamber, but. since the
piston worked by displacement it was unnecessary
for it to be tightly fitted to a cylinder. This scheme
avoided the difficulties of accurately machining such
a small size hole. The wire piston is driven by a
micrometer head, somew hat larger t han usual, but of
standard design. Twenty-five grooves are cut on the
thimble actuating a spring click. The lead screw has
the standard micrometer pitch of 40 threads to the
inch. The. dimensions of the wire and (he pitch of the
lead screw are such that each click (1 25th revolu-
tion) advances 0.002 cu mm (2.5 gamma) of II.
The mercury chamber communicates with a re-
movable tip, made out of capillary tubing. The end
of the tip is optically polished. The instrument is
mounted to move up and down on a rod screwed to
a wooden base. The base forms the bottom of the
carrying ease, with the rod serving as a tie rod to
hold the top and bottom of the ease together. The
instillment can bo transported as easily as a com-
pound microscope.
The apparatus is durable and simple. It lues been
found to give reproducible lesions. The principal de-
fect is incomplete delivery of all the material ad-
vanced to the capillary tip. It is necessary that each
subject’s arm come in contact with the tip with the
same pressure. The extent of loss duo to the evaj>ora-
SECRET METHODS FOR TESTING VESICANTS
301
tion of the compound between the time it gets to the
tip and the time that it is applied to the subject is
unknown. It is minimized by maintaining a regular,
rapid rate of application. The instrument can best be
list'd by a trained pair of operators, one operating
the micropipet, the other holding the men’s arms
against the tip. A regular rhythm soon leads to both
speed and accuracy.
It was found that the necessity for counting a
number of clicks repeatedly let! to personal error.
An attachment was made for the pipet that made
it possible to advance the desired amount in a single
motion, rather than by counting a number of clicks.
A brass plate with 25 holes equally spaced around
the periphery was attached to the instrument. An
index arm was attached by a ratchet to the lead
screw. By placing taper_pins in the appropriate holes
any number up to 25 clicks can lie delivered without
counting.
Liquid Vesicant Cup.'11 Occasion arose to compare
the action of 11N3 as a liquid with saturated 1IN3
vapor. The vapor concentrations were set up in vapor
cups (set' Section 16.6.2). The apparatus employed
for the application of liquid consisted of a small cup,
12 mm outside diameter and 8 mm inside diameter,
with two capillaries leading from it. One capillary
leading directly upwards from the cup, was attached
to a safety flask and a charcoal column aspirator; the
other tube, coming from near the base of the cup at
a 45-degree angle, is connected with a three-way
stopcock. A pear-shaped bulb with a small hole in
the side is sealed to the vertical arm of this stopcock.
The liquid vesicant is placed within the bulb, and
the stopcock is turned so that the vertical arm is con-
nected with the cup. The cup is placed on the sub-
ject’s arm, and the vesicant is drawn by suction out
of the bulb, through the capillary, and into the cup
until the area on the arm is covered with a continu-
ous layer of vesicant. At the end of the exposure
period 5 per cent hydrochloric acid is sucked through
the instrument, followed by water; by applying the
suction intermittently, the surface, of the arm is
flushed and decontaminated. In control tests with
fat-soluble dyes all visible dye Was removed within
5 seconds.
16.6.2 Testing Vesicants as V apors
For proper evaluation of the vesicancy of a com-
pound the vapor hazard must also be determined.
Edgewood Vapor Cups.'0 One of the simplest ways
of producing vapor burns is by the use of small glass
cups with a Hat rim. A pad of filter paper or some
other absorlient material is placed in the bot tom and
moistened with the liquid vesicant. The cups are
then taped on to the arm of the subject for the de-
sired length of time.
The amount of vapor (and its effectiveness) in the
cups will vary as a result of the interplay of outside
temperature, skin temperature, amount of moisture
under the cup, and (he presence or absence of sun-
light on it. The actual concentration in the cup is un-
determinable and may be changed by cooling or
warming the cups. In addition to vapor burns, “rim
burns” sometimes occur. These are the result of con-
densation of liquid agent on the lip of the cup. The
use of these cups permits the application of approxi-
mately saturated concent rations of vapor.
Modified cups have been devised which permitted
the application of subsaturation concentrations, pro-
vided for circulation of the vapor, and eliminated
rim burns.17 IS —
The Vapor Train.11 Some of the objections raised
to the use of the Edgewood cups are similar to those
raised against the use of “static” chambers. A dy-
namic method of exposure was devised to overcome
some of these difficulties.
This apparatus consists of the following essential
parts. (1) A bubbler from which the agent is vaporized.
(2) A serum I bubbler in which water is vaporized,
(it) A )’ tube that unites the streams of vapor-laden
air from the two bubblers. (4) Glass tubing which is
branched and rebranched to divide the vapor-air
stream into four identical streams. This tubing,
20 mm inside diameter, is in several sections that are
joined by 29 12 standard taper joints. (5) Four appli-
cator orifices. Each may be described as an open cup
with a delivery tube for conducting the vapor-laden
air to it and a side arm that serves as an outlet. The
cup is formed from a 24/40 male standard taper
joint. The delivery tube, 8 mm inside diameter, enters
the cup at the bottom through a ring seal and pro-
trudes to within 3 mm of the upper, open end. The
vapor-air stream, therefore, flows upward through
the delivery tube, impinges upon the skin of the arm
which a subject holds over the opening of the cup,
and out through a side arm. The velocity of this jet
is about 5 mph. (6) Four slain less-steel adapters for
the applicators, each with an 8-mm hole in the cen-
ter. Use of these; adapters reduces the area of skin
exposed anti thus minimizes the severity of the re-
sultant lesion. Each cap has a small ridge at its outer
edge (1 32-inch deep) to prevent an arm from mov-
8ECRET APPARATUS AM) TECHNIQUES IN TOXICOLOGICAL STUDIES
ing around during exposure. The caps are held in
place by rubber bands. (7) A branched and rebranched
glass tube, identical with (4) but used for uniting the
effluent streams from the applicators. (8) A tube to
conduct the combined effluent into a suitably venti-
lated duct. (9) A sampling apparatus to draw a meas-
ured volume of the effluent through a suitable ab-
sorber for determination of the analytical concentra-
tion of the vapor (see above). (10) Platforms upon
which subjects rest their arms while holding them
over the applicators. These are small tables of ap-
propriate height with holes through which the appli-
cators protrude alwait 14 inch. The skin is thus held
firmly against the cap of the applicator without any
possibility of excessive pressure, and the arm rests
comfortably during the exposure.
This apparatus, with a volume of 2 1 and an air
flow of 20 1pm, can be classified as a small, high-flow
chamlter. Concentrations of agent can l>e used up to
saturation; the humidity of the air can be varied
from dryness to saturation, (loud analytical-nominal
ratios are obtained. The apparatus is rapid and con-
venient to use.
i’se of Dynamic Chambers for Body Exposures.
Almost all of the standard type chambers in this lab-
oratory have, at one time or another, been equipped
for body exposures. The bodies of the animals are
exposed to contaminated air, while their heads are in
fresh air. A gasket around their necks prevents leak-
age of the noxious air and its inhalation. In one of
the earliest methods1 for use with a small smoke
chamber the animals were placed in the chamber and
provided with a manifold through which pure un-
circulated. It has been more common practice to
place the bodies of the animals in the chamber and
let their heads protrude. The first chamlxT to have
built-in provisions for body exposures was the 200-1
chamber.6
I sc of Wind Tunnel for Testing Vesicants on Man. ''1
The wind tunnel (p. 285) is equipped with ports
through which arms can be inserted perpendicular
to the air stream. Since turbulent flow occurs, it was
necessary to expose an annular space around the
arm.
The arm was prepared for exposure by wrapping
(he hand and wrist to a point 5 cm above the distal
end of the ulna with oilcloth sealed with adhesive
tape. A piece of adhesive tape 2 inches wide was
placed around the forearm leaving an exposed an-
nulus of skin 1 cm wide between the wrist covering
and the adhesive tape. Another piece of oilcloth cov-
ered the. remainder of the forearm and elbow, leaving
a second (proximal) annulus Indween the 2-inch (ape
and the elbow covering. To deliver two doses to the
same arm the distal (wrist) annulus was left exposed
for the whole exposure period; the proximal (elbow)
annulus was kept covered with oilcloth except for the
appropriate terminal fraction of the exposure period.
At the end of the exposure the coverings were re-
moved and discarded.
The use of the wind tunnel permits testing the
relative efficiencies of aerosols and s apors at various
wind speeds. Temperature and humidity of the air
stream can bo controlled only by controlling the
temperature and humidity of the laboratory.
The (ircal Lakes Man-ChamberThis apparatus
for test ing effects of vesicant vapor on masked men
has been described in Section 16.2.2.
SECRET Chapter 17
PHYSIOLOGICAL MECHANISMS CONCERNED IN THE PRODUCTION
OF CASUALTIES BY EXPOSURE TO HEAT
Alan it. Moritz
17.1 INTKODL CTION
At a MKKTixti called at the instigation of the
Technical Division of (he Chemical Warfare
Service on March 22, 1944. certain deficiencies in the
existing state of knowledge concerning the casualty-
producing effectiveness of the flame thrower were
discussed. Attention was called to the fact that, al-
lhough both heat and the inhalation of irrespirable
or poisonous gases probably contribute in varying
degrees to these effects, little was known regarding
their relative importance.
It was recommended that the physiological section
of Division 9 of the National Defense Research Com-
mittee [NDRC] investigate the various mechan-
isms by which flame thrower action may cause dis-
ability and death. In this chapter are reviewed the
studies that were made of the mechanisms by which
excessive environmental heat may lead to early dis-
ability and death.
17.2 PILOT EXPERIMENTS TO EXPLORE
CASL ALTY-PRODUCING ATTRIBUTES
OF GASOLINE CONFL VGR VITONS
A certain amount of general information concern-
ing the thermal and chemical attributes of gasoline
conflagrations was prerequisite to the planning of an
experimental program. For the purpose of orienta-
tion, certain exploratory investigations were made
of the rate, magnitude, and duration of the changes
that occur in the temperature as well as of those that
occur in the atmospheric concentrations of oxygen,
carbon dioxide, and carbon monoxide incident to the
tanning of measured quantities of flame thrower fuel
in both closed and ventilated spaces.
17.2.1 Experimental Procedure
A series of experiments 1 were accordingly under-
taken in which gasoline was burned in a fireproof
room having a capacity of 14.4 cu m. The construc-
tion of the room was such that it could be cither
closed or ventilated at will. The fuel was poured into
shallow metal pans which completely covered the
floor, which measured 1.6x3 m. Approximately 1 li-
ters were burned during each conflagration.
To measure the changes in temperature, 10 gauge
iron-const an tan spot-welded thermocouples were
suspended in the center of the chamber. The thermo-
electric potentials provided by the thermocouples
were amplified by means of an electronic optical
bridge circuit.1- It was found that the use of a split
circuit is capable of amplifying a 1-rnv input poten-
tiometrieally to a 5-ma output in less than 0.2 sec-
ond. Since this amplifier was a null-point, instru-
ment, it was independent of all the electronic tube
characteristics, of the intensity of the light beam
focused on the photocell, and of the input resistance
of the thermocouple leads. Two such amplifiers were
const ructod.
Two recorders were used. One was an Esterlinc-
Angus recording milliampere meter (5 mil, full scale)
with a response time of 0.5 second. The other was a
General Electric photoelectronic recording milli-
ampere meter with a response time of 0.2 second.
Both recorders had 12 inch per minute chart drives.
By means of a selector switch the sensitivities of
the amplifiers were usually set so that a 40-mv input
produced full-scale deflections of the recording pen.
Method of obtaining samples of atmosphere for
gas analysis: Three long tubes, each having an in-
ternal diameter of 2 mm, extended from the outside
to the center of the conflagration chamber. These
tubes passed through the wall at, the bottom, middle,
and top of the room. Samples of 300 ml were with-
drawn as desired by attaching evacuated flasks with
ground joints to the ends of these tubes. The gas
samples obtained in this manner were analyzed for
()2. CC2, and CO by means of a standard Orsat ap-
paratus.
SECRET 304
ST I DIES OF THERMAL INJURY— CUTANEOUS AM) SYSTEMIC
IT.2.2 Temperatures Developed during
(.asoline Conflagrations
Unventilatod conflagrations: In these experiments
the door was kept closed during the fire. Oxygen de-
pletion resulted in extinction of the conflagration in
about 30 seconds after ignition. Approximately’ half
of the gasoline contained in each pan remained un-
burned. When the door was opened following the
premature extinction of the file, the room was found
to lie filled with dense black smoke and there was a
strong odor of gasoline.
the lower. The sharp peaks in the temperature curve
of (he upper thermocouple are also due to convert ion
currents. The average temperatures recorded by the
two thermocouples over a 30-second period were ap-
proximately the same, namely, about 500 C. At the
termination of the combustion, the ambient temper-
atures fell rapidly and uniformly. The curves shown
in Figure IA are typical of all experiments in which
the conflagration was unventilated.
Ventilated conflagrations: Figure IB shows a con-
tinuous temperature recording of a thermocouple
which was situated about 1.5 m above the floor dur-
ing a conflagration in which ventilation sufficient to
maintain complete combustion was provided.
The temperatures obtained were about the same
as those recorded during unventilated conflagrations.
The duration of the high-temperature plateau de-
pended on the length of time that the door was left
open. In (lie experiment in which the record shown
in Figure. IB was made, the door was left open for
50 seconds,
IT.2.3 Extrapolation of Experimental
Temperature Changes to Conditions Likely
to Prevail in Bunkers and Pillboxes
Incident to Flame Thrower Attack
It was judged that the circumstances which pre-
vailed in the experiments just described probably
predisposed to the development of maximal temper-
ature elevations. It is regarded as unlikely that
higher temperatures would be developed in hunkers
or pillboxes incident to flame thrower attacks in
which gasoline was used as fuel. Due allowance
should lx> made for the toleraneo of commercial re-
cording instruments in the interpretation of data
pertaining to temperature changes in bunkers and
pillboxes incident to field tests of the effectiveness of
flame thrower equipment. Thermocouples of the
usual size and potentiometers or millivolt, meters of
the usual period are not capable of following the
rapid temperature fluctuations that occur in nn-
ventilated or incompletely ventilated gasoline con-
flagrations. Furthermore, temperature observations
made by such apparatus may be lower than the
actual temperatures obtained by as much as 500 C.
I7.2.t Exposures of \nimals to Burning
Gasoline
Adult dogs (6-8 kg) and young pigs (7 12 kg)
wore exposed in various ways to burning gasoline.
Figckb 1. Continuous temporal hit recording dining
burning of gasoline in rectangular (fixlOx 10ft)combus-
tion chamber.
(A) No ventilation. Two thermocouples, one 5 ft
and the other 3 ft above floor level. Distance from floor
to ceiling was 10 ft.
(B) Room ventilated for 50 seconds. One thermo-
couple 5 ft alxivc floor.
Figure 1A shows continuous temperature records
provided by the two thermocouples, one of which
was hung midway between the floor and ceiling in
the center of t he 3 m high conflagration chamber, and
the other approximately 0.9 in above die floor.
Because of rapid convection currents, the upper
thermocouple reached higher temperatures than did
SECRET \TTRIBLTKS OK GASOLINE CONFLAGR VTIONS
305
Tabi.r 1.
Effects of temperature and com Oust ion products resulting from gasoline conflagrations on animals.
Conflagration
thermal exposure
combustion
Comp. (* f ) of air
r ate of animal
Blood
Kef.
Avg
Temp
C
Body
Body
only
With
After
after fire
Dead
Survival
expt
CO sat
sec
face
only
fire
fire
():
COj
CO
min
or days
SC
1
30
320
+
4"
5 ruin
14.0
4.0
o.s
+
30
2
40
600
' +
+
Xo
16.2
4.0
0.8
+
7
3
30
400
4-
No
No
+
. .
4
30
370
+
No
Xo
+
5
73
700
+
Nil
Xo
+
k
30
350
Xo
5 min
14.7
4.6
1.0
+
6
7
30
350
+
+
4 min
15.7
3.5
0.3
4~
T race
8
30
450
+
—4"
4 min
16.1
3.6
0,7
4-
37
9
30
500
+
+
2 min
+
32
The animals were anesthetized by the intraix'iitonoal
injoetion of sodium pentobarbital ami fastened by
asbestos tape to an iron frame situated in the center
of the conflagration room 54 inches above the floor.
The principal data pertaining to these experiments
arc included in Table 1.
COMBINED CTtaNEOES AND RESPIRATORY EXPOSURE
Animals 1 and 2 were exposed to the full effects
(cutaneous and respiratory) of the burning gasoline.
Throughout the entire exposure of animal 1, the door
of the conflagration chamber .remained closed. The
fire burned out in about 30 seconds because of in-
sufficient oxygen. The average temperature of the
air surrounding the animal during tins period was
320 C. The animal was allowed to breath the at-
mosphere of the unveatilated room for 5 minutes
after the fire was extinguished.
Samples of the atmosphere were taken for gas
analyses as soon as the fire had burned out. The
mean concentration of CO in the atmosphere was
0.8 per cent, and the oxygen concentration was
14.6 per rent. The CO saturation of a .sample of the
animal’s blood taken 5 minutes later was 30 per cent .
Although there was no indication that the fire had
resulted in a dangerously low oxygen or a danger-
ously high CO. concentration, it did appear likely
that, the animal would have died of CO poisoning if
it had remained much longer in the unventilated
room.
Although animal 1 had been severely burned, it
did not develop early shock, required several post-
exposure injections of Nembutal to keep it quiet, and
was beginning to become restless with returning con-
sciousness when sacrificed 6 hours later. Its air pas-
sages contained an excessive amount of mucus but
them was neither clinical nor pathological evidence
of significant thermal or chemical injury of the
larynx, air passages, or lungs.
In the case of animal 2, the door remained open
during the first 10 seconds of the conflagration, with
(he result that a larger amount of gasoline burned
and a higher tem|H‘iature was achieved and was main-
tained for a longer period of time than was (he case
in the first experiment. At the end of 10 seconds, (he
door was closed with the result that the file was ex-
tinguished very soon thereafter. Samples of the at-
mosphere were then taken for gas analyses and the
animal removed. This dog was moribund when re-
moved to (he o[X‘ii air. In view of the fact (hat the
atmospheric concentration of CO was similar to that
observed in the preceding experiment, it was sur-
prising to find that (lie CO saturation of the blood
was only 7.0 per cent. The explanation of this dis-
parity probably lies in the fact that animal I breathed
the atmosphere of the conflagration chamber for a
total of 6-7 minutes, whereas animal 2 was moribund
at the end of 2 minutes.
Two factors may have contributed to the ex-
tremely rapid death of dog 2. One is systemic hyper-
thermia caused by overheating of the blood as it cir-
culated through the extensive sii|x*rficial network of
subcutaneous vessels. The. other is respiratory ob-
struction due to pharyngeal edema. That a signifi-
cant degree of hyperthermia had occurred was indi-
cated by the finding of a rectal temperature of 41.2 C
when the animal was autopsied 5 hours after the ex-
posure. That obstruction to respiration may have
contributed was indicated by the presence of severe
burning of the mouth and pharynx with what ap-
peared to he obstructive edema of the latter. The
trachea and bronchi contained abundant mucus
SECRET STL DIES OF THERMAL INJURY—CUTANEOUS AND SYSTEMIC
mixed with carbon particles. The lungs were hy-
peremie.
The results ol the first two exposures dealing with
the effects on animals of burning gasoline indicated
that, even in circumstances considered to be particu-
larly favorable to the production of CO and to the
exhaustion of oxygen, the concentration of these
gases was not sufficiently altered to cause uncon-
sciousness or death within 5-6 minutes. Although the.
results of the two experiments were not construed as
proof that neither fatal anoxia noi fatal CO poisoning
could result from a gasoline fire, they did indicate
that such an exposure can cause rapid death from
thermal injury alone.
( Vtax Kors Exposehe
The next three experiments shown In Table I were
undertaken to ascertain the effect of protecting the
respiratory tract against heat and combustion prod-
ucts during the time thaf~l.be body was being ex-
pose* I. To investigate this question, animals 3, 4,
and 5 wore light-fitting asliestos-eovereil rubber
masks through which a continuous stream of un-
heated air was circulated during their exposure to
heat. The first two animals of this series (3 and 4)
were exposed to an un vent dated conflagration of
about 30 seconds duration and average atmospheric
temperatures of 400 and 370 C, respectively. Al-
though both animals showed extensive burning of the
skin, they survived the immediate effects of heat and
were in reasonably good condition when killed 6
hours Inter. In the ease of animal 5. the door of the
room was left open for the firsl minute of the fire and
for 65 seconds the temperature of the room was in ex-
cess of KK) C. Within 15 seconds after the door was
closed the fire went out and the animal was removed.
This animal died immediately on reaching the ojicn
air and showed severe burning of all the body surface
except where the skin had been protected by the
mask.
'These experiments provided evidence that a rela-
tively brief (75 seconds) exposure of the skin to a
sufficiently high temperature could cause almost im-
mediate death independently of other factors.
Kespiratokv Kxposvhe
'The last four experiments shown in 'Table 1 were
undertaken in an attempt to investigate further the
effects on animals produced by the breathing of I lie
combustion products of a gasoline conflagration. In
each experiment the door was kept closed throughout
the entire conflagration. By this procedure post con-
flagration mixing of outside air with the combustion
products was reduced to a minimum. The skin of the
body was protected against excessive overheating by
enclosing the animals to the neck in a heavy asbestos
sack. With the exception of No, 6 the animals were
free to breath the burning gases and hot air during
the fire as well as the smoke which remained in the
chamber after the fire. Dog No. (i breathed outside
air circulated through the mask during the fire; as
soon as the temperature in the room had dropped to
200 (' the mask was detached by remote control and
for the next 5 minutes only the hot smoke and air of
the combustion chamber were available for respira-
tion.
None of these four animals showed either clinical
or pathological ev idence of thermal injury of the air
passages or lungs. Two of them (6 and 7) may have
held their breath throughout most oral! the exposure
period. That animals No. 7 and 8 breathed during
some of the lime that they were in the combustion
chandler is indicated by their carboxyhemoglobin
concentrations of 37 and 32 j)er cent, respectively.
It is possible, of course, that even these two animals
held their breath during the conflagration and ac-
quired their carlnin monoxide by breathing during
the interval between the time that the fire went out
and the time that they were removed from the
chamber.
17.2.5 Summary
The ignition within a simulated pillbox or bunker
of a well-spread layer of gasoline leads within 10 sec-
onds to a temperature rise of be)ween 800 ami
1000 The duration of such a conflagration and t he
temperature increase caused by it varied according
to the oxygen supply and the fuel. In a closed room
having a capacity of 14.4 cu m and measuring l.Gx
3x3 m, the fire was extinguished within 20 seconds
and considerably less than 250 ml of gasoline was
consumed for each cubic meter of air space. For ap-
proximately 10 seconds of the burning time the tem-
perature fluctuated between 500 and 900 C. If such
a room is ventilated, the initial temperature rise is
similar to that which occurs in a closed room, but the
fire continues to bum until the fuel is exhausted, re-
sulting in a temperature fluctuation of between 500
and 1000 C. In both instances, convection currents
established by the conflagration resulted in marked
fluctuations in the temperature at any given place
within the room. During the period of rapid com-
SEC11ET BASIC CHARACTERISTICS OF HEAT AND HEAT TRANSFER
307
bust ion the tem|x*ratures were highest near the ceil-
ing and lowest near the floor.
In none of the experiments conducted in this par-
ticular tyj)e of conflagration chandler did the oxygen
content drop below 11 per cent. The carbon dioxide
level did not rise higher than 5 per cent nor the carbon
monoxide level above 1 per rent.
'Pbe most important information gained from these
exploratory experiments was the observation that
animals as large* as dogs and pigs when exposed to
tire kind of a conflagration for more than 30 seconds
may receive injuries that are almost immediately
fatal. Such fatalities were not necessarily contributed
to by asphyxia, carbon monoxide poisoning, or in-
halation of flame. It apjx’ared that the rapid death
may result from systemic disturbances caused by the
impact of heat energy on the surface of the laxly. It
was obviously in order to conduct additional and
better controlled ex|X'iiment.s to investigate the
physiological mechanisms concerned in the produc-
tion of casualties through the thermal effects of
flame thrower attack.
IT.3 It \SIC CHARACTERISTICS OF
IIKVP \M) HEAT TRANSFER*
It could lie inferred from the results of the pilot
experiments reported in the preceding section that
heat independently of other factors was an impor-
tant, if not the most important, casualty-producing
attribute of flame thrower action. This being., the
case, consideration should be given to the nature of
beat and to the factors which determine the transfer
of heat from one medium to another and from one
place to another within the same medium.
IT.3.1 Theoretical Considerations
The Nature of Heat
The concept of temperature rises from the sensa-
tions of hotness and coldness. Experience has shown
that when two or more substances of different tem-
perature are kept free of all outside disturbances, the
hotter bodies will get colder and the colder bodies
hotter; and that ultimately these substances will
reach a state of complete thermal equilibrium (ident i-
cal temperature). The hotter bodies are said to have
lost heat, and the colder bodies are said to have
gained heat. This concept of heat becomes quantita-
tive by defining a unit of heat, the calorie, as the
amount of heat gained by a 1 g of liquid water under
atmospheric pressure when the temperature increases
from I t o (' to 15.5 C.
This gain in heat, which is discernible through a
rise in temperature, is associated with an increase in
the infra- and intermodular motion. Thus heat can
be considered as the energy stored in a substance by
virtue of the state of its molecular motion. Certain
manifestations of this increase in energy are readily
observable, for example, melting, vaporization, de-
composition, alteration in rate of diffusion and in
chemical reaction.
Reside the definition of a calorie, there are other
physical concepts pertaining to heat which are
requisite to an understanding of the.general problem
of thermal injury.
Heat Capacity
Heat capacity or specific heat of a substance is the
amount of heat which is .required to raise the tem-
perature of the substance 1C.
The importance of heat capacity (Cp) in thermal
injury is readily seen by considering the respective
injury propensity of 1 gof water (CP — 1.00) and 1 g
of silver (C,, — ().()(») both at 100 C placed in contact
with 1 g of thermally insulated skin (Cp ~ 0.7) at
35 C. After equilibrium is reached In the former ease,
the tem|)c.rature of the skin is increased to 73 C,
whereas in the latter case it is increased only to
42 C.
It is apparent, that, if the skin were to equilibrate
rapidly enough when placed in contact with a hot
body, there is insufficient heat in 1 g of silver at
100 C to produce injury to 1 g of skin. Actually, of
course, the skin, because of its thermal insulating
properties, does not equilibrate rapidly enough and
the portion of skin nearest (he silver docs reach a
sufficiently high temperature to produce injury be-
fore thermal equilibrium is reached. Hence another
physical property of importance is heat transfer.
11eatTransfer
In the experiments to lx* described beat was trans-
ported to the skin by three mechanisms: namely,
convection, radiation, and conduction. In the ease of
convection and radiation, heat reaches the skin under
such circumstances that the heat uptake is primarily
determined by the heat source. In the east; of con-
duction, the amount of heat absorbed by the skin is
primarily determined by the properties of the heat
absorber, namely, the skin itself.
* By F. C. Hcnriques, Jr.
SECRET 308
STUDIES OF THERMAL INJURY— CUTANEOUS AND SYSTEMIC
('ONVKCTION
Convection is the mechanism by which hot air
transports heat to a cooler surface because of the
eddying currents that arise. The air velocities of the
eddy currents are about 1.6 km per hour. An equa-
tion has been developed for the transfer of ambient
heat by natural convect ion from a large envelope of
hot air surrounding cylindrical objects about ;{f) cm
in diameter.29 34 This equation shows that 7, (he
caloric uptake per minute |>er square centimeter of
surface, can l>e expressed as follows:
7 = 0.0026(7’,, - 7\)J “ (I)
where T„ is the air temperature in C and 7’, is the
surface temperature in (’.
Thus, with a skin temperature of 40 C, air at
100( ' and 400 C will transport to the skin about
0.4 and 4 cal cm2 min. It is also apparent that as
this heat is absorbed by the skin the surface temper-
ature of the skin will rise and the caloric uptake of
the animal will decrease with time.
It is of interest to compare with this the caloric
uptake rate of skin at 40 (’ when an atmosphere of
steam maintained at 100 C is substituted for the air.
Under these conditions, about 300 cal enr min
would be absorbed by the skin 36 if the surface tem-
perature could be maintained at 40 C. This 800-fold
increase in caloric bombardment as compared with
that produced by air is due to the latent heat of con-
densation of steam. This, of course, is why steam is
an enormously greater hazard than hot air in the
production of heat injury.
Radiation
All substances give off heat in the form of radiant
energy in amounts that are predetermined by the
surface temperature of the substance. When this
radiation impinges upon another body, a certain
fraction is absorbed and changed into heat. Thus, if
two substances at different temperatures are placed
in an enclosure, there is a continual exchange of
energy, the hotter body radiating more energy than
it absorbs and the colder body absorbing more heat
than it radiates.
In the special case of an animal completely en-
closed in a large box of source temperature 7%, the
caloric uptake rate 7 of the animal, due to this inter-
change of radiant energy between the skin and the
wall of the box, is expressed by the following equa-
tion.30»
7 = scJUT, + 273)4 - (T, + 273)4] (2)
where s is the radiation constant and is equal to
8.2 X 10~u calorie, per square centimeter per Tx
per minute, e is the effective emissivity of the hot
walls of the box, and / is the absorptivity of the skin,
to radiation emitted at Tr. Under experimental con-
ditions to l»c described, the product rf can be taken
as about 0.8. Thus, when the skin temperature is
40 C, the hot walls at 100 (' or 100 C will radiate to
the skin about 0.7 or 13 cal cm- min, respectively.
Conduction
Conduction is defined as the transfer of heat from
the hotter portion of a substance to a colder portion
of the same substance, or from a hot laxly in physical
contact with a cold body, where in each case there is
no appreciable displacement of any of the molecules
comprising these substances. It is the latter restric-
tion that differentiates conduction from convection.
In certain experiments to be described heat was
conducted from either a hot solid or a hot liquid to
the skin. In these experiments, the purpose of both
the solid and liquid heat source was to maintain the
tenqierature of the skin surface at a predetermined
constant value and hence the conduction of heat
through the heat source need not lx* considered. In
the hot air experiments, thermal conduction through
air is small as compared with convection, and this
small contribution is included in equation (1). Thus
it is only necessary to consider conduction of heat
through the skin.
In all cases of heat flow by conduction, a temper-
ature gradient must exist within the substance. If
this temperature gradient varies with time, the rate
of heat flow will also vary with time. The type of
heat flow where temperature is a function of both
position within the body and time is called heat con-
duction in the unsteady state. Heat conduction in
the steady state refers to all cases where the temper-
ature at any point within a substance does not de-
pend upon time. Under these conditions the amount
of heat flow through the medium is determined by
this tenqierature gradient and the ability of the
body to conduct heat (thermal conductivity). The
latter case will be considered first . The equation for
steady-state heat conduction inside a rectangular
homogeneous body is based upon Fourier’s law111
and is as follows:
7 = f (T, - T„) (3)
where K, the thermal conductivity, is expressed in
SECRET BASIC CHARACTERISTICS OF MEAT AMI HEAT TRANSFER
309
calories jxm- minute, per square centimeter perpen-
dicular to (lie direction of heat flow per unit temper-
attire gradient in (' per centimeter length of path.
L is the path length through which the heat flows
and T, and T„ are the temperatures in C at the be-
ginning and end of the path, respectively; q has lieen
previously descrilied.
This equation permits the ex|>erimental determina-
tion of the in vitro thermal conductivity of the four
respective sections of tissue, namely epidermis, der-
mis, fat. and muscle, and also of any combination
thereof,
Glnehal Tiikouv ok Heat Flow thkovgii Skix
By making use* of the preceding brief definitions of
the various physical factors involved in the transport
of heat to and through the skin it is possible to con-
sider how the application of heat affects the time-
temperature relationship within a given skin site. It
is-apparent that in order to make heat flow inward
from the skin surface it is necessary to raise the tem-
perature of the skin surface to an extent that over-
comes the normal existing gradients. This can be ac-
complished by means of an external source, of heat
through conduction, convection, or radiation. Once
the skin surface temperature is sufficiently high, the
heat will start to flow inward, resulting in a general
rise in tenqierature within the skin site.
din's initial heat flow inward (and thus the rate of
temperature rise within) will depend primarily upon
two physical factors; namely (1) the heat capacity of
the skin or the ability of the skin to absorb the beat,
and (2) the thermal conductivity of the skin or the
ability of the skin to transport the heat. After a cer-
tain interval of time the amount of heal entering the
skin site will be balanced by the amount of heat
leaving the skin site, and the skin will l>e ‘‘heat-
saturated.’’ In this state the new temperature dis-
tribution within the skin site will become invariant
with time and the amount of heat flowing through
the skin will depend only upon (2) and the skin sur-
face temperature.
It is to be recognized that this picture involves not
only the solution of the steady state of heat conduc-
tion but also the solution of the initial unsteady state
of heat flow. In order to solve even the “idealized”
picture, it would lx? necessary to know the initial
temperature gradients within the tissue, the thick-
nesses. densities, thermal conductivities, and heat
capacities of the various layers, and the skin surface
temperature as a functiyp of time.
The solution of such a problem involves the follow-
ing Fourier heat equation:"
(4)
pC„iA /- / di K 1
where TIt is the temperature time, t, at a dis-
tance. x. within the skin measured from the skin
surface, p is density, and the remaining symbols
have been previously defined.
The solution of equation (4) subject to these con-
ditions is exceedingly complicated. Vet superim-
posed upon this arc the numerous indeterminate in
vivo factors which arise when we go from the idealized
picture to the living animal. It is useful to enumerate
the most important, of these various indeterminate
factors.
1. Site variations in the respective thickness of
epidermis, corium, fat, and muscle.
2. Variation of existing temperature gradients
within the skin with respect to time and or position
of site.
3. Unknown average rate of blood flow through
the various skin layers, and the unknown variations
of the unknown rate of flow with respect to position
of site and temperatures within the site.
4. The appearance of edema fluid in variable
quantities which brings forth indeterminate altera-
tion in the density, heat capacity, thickness, and
thermal conductivity of the various layers of skin so
affected.
It is obvious from this discussion that any general
solution of the time-temperature relationship within
a skin site, when heat is applied, is not possible. How-
ever, with certain of the experiments to Ire described
later in detail, it is possible to derive to a first ap-
proximation the time-temperature relationship in
the layer of basal epidermal cells. These experiments
were either (a) so conducted to bring immediately to,
and maintain the skin surface at, a predetermined
temperature level until the threshold of irreversible
epidermal injury was reached, or (b) (he entile ani-
mal was completely surrounded by an envelope of
ambient and radiant heat. These experimental con-
ditions at the boundary of the skin surface and source
of heat are expressed by the following equation;
q = II{T - T.) (5)
where q and T, have been previously defined (see
Section 17.3.1 under “Convection”). T is the temper-
ature of the heat source in C and //, in cal/cnr/min,
is known as the heat transfer coefficient. Conditions
SECRET 310
STlDIES OF THEKMAL I NJURV CL’TAM EOL'S AM) SYSTEMIC
under experiments (a) were tantamount to an infinite
heat transfer coefficient (// = «>); with experi-
ments (b), the heat transfer coefficient is finite and
the numerical value readily obtained by combining
the radiant and ambient contributions to heat trans-
fer coefficient as computed by equations (1) and (2),
respectively. In order to solve equation (1) under the
boundary condition expressed by equation (5), it is
necessary to assume that the ratio of the total tissue
thickness to the epidermal thickness (approximately
80u) is infinite rather than finite. This assumption
will lead to slightly longer time intervals for “heat
saturation” of (he epidermis than are to be experi-
mentally expected. The integration * of equation (1)
under the above conditions results in equation (<»).
j> —Tt f >1 hl:ki\+(hua^kt\~
T. ~ r„ = Ly/tJ
!1-#[^( (,i)
where
r
0(Y) = f e~'dx (0a)
V JT •'
0
and y is computed by means of equation (0b).
y~2 ” (0b)
1 pCp -
Ti is the temperature of the basal epidermal cells
at the time I in seconds. T, is the temperature of the
heat source. T„ is the temperature of the skin surface
previous to the exposure to heat. 1. is the distance of
the basal cells from the skin surface, p is the density
of the basal epidermal layer. The other symbols lane
been previously defined and are experimentally de-
terminable. The integral that defines 0(1) (equa-
tion Oa) is respectively equal to y/V 2 and zero when
T is infinite (t ~ 0) and )' is zero (/ = <»). For other
values of V, the numerical value of the integral is
tabulated.59
The time-temperature relationships at the basal
epidermal layer during an exposure of the animal to
a source of constant ambient and radiant heat are
evaluated by means of these equations in Sec-
tion 17.3.2 (see also Section 17.9.2 under “Measure-
ment of Heat Transfer”).
In the experiments in which the skin surface was
brought immediately to and maintained at a pre-
determined constant temperature, //, the heat trans-
fer coefficient, is nearly infinite, and equation (0)
reduces to
m-a.)
where, as before, 0 is given by equation (Oa). It is to
be noted that in this case T, can be taken as the skin
surface temperature during the entire heat exposure,
since tlie temperature of the heat source is identical
with the surface temperature once heat exposure
begins.
It is to be noted that equation (tic) results in a
basal layer temperature which becomes, after a cer-
tain time interval, essentially identical with the skin
surface temperature. Actually, a small but finite
temperature gradient will always exist between the
surface and the basal cell layer. This steady-state
gradient can be experimentally determined by means
of equation (3), and the true temperature of the basal
layer can be quite accurately computed for any
time t by using equation (tic) until the steady-state
temperature obtained through equation (3) is
reached. Computations using equation (tic) to ascer-
tain basal epidermal temperature are given in Sec-
tion 17.3.2, and the experimental justification for
this theory will be considered in Section 17.0.5 (see
also I7.fi.6).
17.3.2 An Kxperiinenlal Investigation
of Quantities Involved in Botli Steady
and Unsteady State of Heat Con-
duction through Skin 3"
It is apparent that certain types of special appa-
ratus were necessary for the evaluation and assess-
ment of the various physical factors involved in the
time-temperature relationship to thermal injury.
The description of these apparatuses will now follow
in detail.
Heat Capacity Apparatus
The apparatus used for the determination of the
heat capacity of the various skin layers need not lie
described in detail since these specific heals were de-
termined by the well-known method of mixtures.4*
Briefly, this procedure consists of heating a known
weight (about 10 g) of tissue in a brass container to
100 C and rapidly dropping it into a water calorim-
eter. The heat capacity of the tissue was readily
computed from the temperature rise of the water as
measured with a Beckmann thermometer.
SECRET BASIC CHARACTERISTICS OF HEAT AND HEAT TRANSFER
311
Automatic Exkrgv Recohdintj Applicator
In order to measure the rate at which heat energy
was taken up by the skin during the entire exposure
period at any predetermined skin surface tempera-
ture, the following apparatus was constructed to
simulate an infinite source of heat at any given
temperature.
The effect of bringing the skin in contact with a
source of heat having infinite capacity and constant
temperature is shown schematically in Figure 2. The
temperature of the surface of the skin immediately
reaches and is maintained at the temperature of (he
heat source. The rate of caloric uptake by the skin at
the time of the initial contact is essentially infinite
ami as the skin approaches its new state of temjier-
ature equilibrium the rate of energy transfer dimin-
ishes and finally- reaches a nearly constant value.
Thus the curve representing rate of energy transfer
is similar to that shown in Figure 2.
gain as much heat from the surrounding area as it
would lose to it. Thus the calorie uptake from the
central area of the source (Figure 2) would he a meas-
ure of the per|>endicular flow of energy through the
directly subjacent skin if a sufficiently large sur-
rounding area were in contact with the same or a
similar source of energy.
A scale drawing of the caloric applicator is shown
in Figure 3A. It consisted primarily of three separate
parts a crown, a brim, and an applicator disk. The
crown and brim were brass, whereas the applicator
disk was copper. The three units wen' maintained at
(he same constant temperature by independent elec-
trical heating units. The temperature of the crown
and brim were controlled manually by means of Gen-
eral Radio Variac transformers. The purpose of the
crown was to prevent any leakage of heat from the
applicator disk except via the exposed face. The
brim compensated for the lateral spread of the heat
from the surface of skin directly underneath the ap-
plicator. The applicator was heated by means of an
auxiliary' electronic apparatus which automatically
recorded the wattage required for continuous main-
tenance of the face of the applicator at a specified
temperat ure T. The temperatures of the crown, brim,
and disk were measured by means of three calibrated
10-mil iron-constant an Fiberglas duplex (Leeds «fe
Northrup) thermocouple wires. The wire heating-
units were of single, silk-insulated No. 40 manganin
wire (negligible temperature coefficient, of resistance).
This wire was held in the indicated spiral grooves
with a thin coat of glyptal. (’upper le:ul wires were
soldered to the ends of the manganin and the joints
were electrically insulated from the metal parts with
fine glass bushings. The electrical resistances of (he
brim, crown, and applicator were .390, 277, and
71.75 ohms respectively.
Three fine phonograph needles rigidly located the
applicator disk inside the crown. The disk was held
firmly against the needles by a rubber bushing under
compression. A steel spring, (he tension of which
could be regulated by a hard rubber screw, controlled
the pressure of the applicator against the skin. This
pressure could be set between 5 and 50 g/cm2.
Guides served to keep the numerous lead and ther-
mocouple wires apart, so that the pressure regulation
was reproducible. The two lead wires to the heating
unit of the applicator were held fast against the sides
of the Fiberglas thermocouple wire by wrapping
with thread for the first 10 cm and then with scotch
tape.
Fuu-re 2. Diagrammatic represent.'ition of rate of
ealoric uptake of skin from heat source at constant tem-
|>e rat ore, of infinite thermal conductivity and of in-
finite heat capacity.
The steady state of caloric uptake is a measure of
the thermal conductance of the skin. The unsteady
state is a measure of the ratio of the thermal capacity
to the thermal conductivity of the skin.
Unless the animal were completely enclosed by an
infinite source of heat, there would be considerable
lateral spread of energy from the application area
(see Figure 2). It was apparent, however, (hat be-
cause of lateral spread the skin in the center of the
application area would, under certain conditions.
SECRET 312
STUDIES OF THERMAL INJURY — CUTANEOUS VM) SYSTEMIC
Floras 3A. Cross section of automatic energy re-
corder applicator—
A AplinUor disk (copper).
B Brim (brass). —
C Crown (brass).
D Fiber washer.
F Heater lead wire*.
F Heater wires (3-mil silicic silk mancaiiini.
(J Fine phonograph needles (three),
I! Brass spider for boldine needlea.
I Stainless steel screw.
4 Hard rubber dow'el.
K Fiber handle.
L Iron-constantan Fiherglaa duplex thermocouple wire.
M Threaded hard rubber eup (Jot adjust inc spring pressure).
N Rubber collar (for holding applicator disk tight against needles.)
O Hraa« cup for rubber collar.
P Thin stainless steel tube.
Q Steel spring
Figure 3TF Diagram of electronic apparatus that con-
trols and measures wattage input into disk of automatic
energy recording applicator;
plate potential; thus, when the plate was positive the
grid was always sufficiently negative to pre/ent the
thyratron tube from firing. When light struck the
photocell, the resistance of this part of the grid cir-
cuit decreased sufficiently to alter the phase rela-
tionship of the grid and plate circuit and the grid
was not sufficiently negative to prevent the tube
from firing during a portion of the cycle when the
plate was positive. Once the tube fired, the grid lost
control (gas-filled tube) and the tube conducted dur-
ing the remainder of the positive plate cycle.
When the plate became negative, the plate cur-
rent became zero and the grid again gained control of
the thyratron. Thus the amount of current which
flowed during the positive plate cycle depended upon
the phase angle between the grid and plate voltage.
This phase relationship was a fimetion of the re-
sistance of the photocell, which in turn depended
upon the amount of light striking the photocell.
Hence, the amount of light striking the photocell
The electronic apparatus which controlled and
measured the wattage necessary to maintain the face
of the applicator at a constant temperature T is
shown schematically in Figure 3B.
The basic principle of the circuit was phase con-
trol of the four-element (GE FG95) thyratron tube.40
In order to obtain sufficient filtered power at the
moment that the applicator first touched the skin, it
was necessary to operate the plate circuit with the
220-v alternating current that was available from a
commercial power line. The grid circuit operated on
220-v alternating current from a radio transformer.
This transformer was connected to produce a J 10-v
potential between point .1 and point B. When no
light was striking the photocell there was nearly a
180-degree phase difference between the grid and
SECRET BASIC CIIAll ACTER1STICS OF HEAT AM) MEAT TRANSFER
313
gave a continuously variable control of the power
output of the plate circuit. The 50-megohm re-
sistance shunting the photocell added stabilization
to the circuit. The 250-aaf variable condenser
“tuned" the phase angle of the grid circuit to the
l>est operating conditions. These conditions were
that a 1-nun deflection of the light beam reflected
from the galvanometer would give full control of the
plate wattage.
The purpose' of the capacities and chokes in the
plate circuit was to filter the pulsating thyratron
output into steady direct current. The values of the
condensers and chokes were necessarily large lie-
cause of the high current requirements of the appli-
cator heater. Oscillograph Tests showed no appre-
ciable ripple current in the filtered output. The wat-
tage or caloric input rate into the applicator heater
was measured with an appropriately shunted Ester-
line-Angus recording milliammeter (5 mil, full scale
and :i4 to 12 inch per minute chart drive). Six scales
were provided by a selector switch with full-scale
deflections of I, 2, 5, 10, 20, and 50 cal min cm2 of
applicator surface area respectively. The highest
value corresponded to a filtered output of 30-v across
the applicator heater terminals.
Operation. In use the galvanometer zero was set
to provide sufficient illumination on the photocell to
generate about 1 eaL min inside the applicator.
The potentiometer was now set to the predeter-
mined millivoltage (temperature). By turning off
and on the low-sensitivity shunt button of the type K
potentiometer the photocell was kept fully illumi-
nated until the galvanometer started to deflect in the
opposite direction. The high-sensitivity button was
now locked down and the instrument was on auto-
matic control. Thus, if the temperature of the appli-
cator face as measured by the applicator thermo-
couple tended to gel either hotter or colder, the gal-
vanometer mirror moved in a direction that either
decreased or increased the illumination on the photo-
cell, which in turn either decreased or increased the
wattage through the applicator. Thus by means of
the thermocouple in the face of the applicator the
output of the thyratron tube was thermally “locked”
to a predetermined temperature of the applicator
face.
The sensitivity of the galvanometer was set to give
a deflection of 4 mm for an 0.1 C change in tempera-
ture. This deflection was sufficient to produce the
maximum available power of 50 cal cm2/min. This
was the maximum sensitivity that could be obtained
without producing periodic heating and cooling of
the applicator face (slow oscillations of the recorder
tracings of caloric uptake rate). These oscillations in
the power output were due to the short but finite
time for the heat generated in the heater wire to
affect the thermocouple.
The heat losses of the applicator disk under the
conditions of usage were determined by placing the
disk and brim on a “perfect” insulator. The perfect
insulat or consisted of a flat-bottomed, thin glass cone
which was silvered on the inside, pumped out to
I0~7 mm of mercury while being heated to 450 C for
8 hours, and then sealed off. All heat losses from the
inside surface of the glass were prevented by the
bright silver surface (no radiant loss) and t he vacuum
(no molecular heat conduction). Lateral heat loss
through the glass was prevented by maintaining the
brim at the same temperature as the applicator disk.
Heat losses from the applicator disk were deter-
mined at two temperatures, namely 45 and 00 C.
The results are given in Table 2.
Table 2
. —
Applicator
'I’eroperaluir C
disk heat loss in
Exp
Crown
Brim
Disk
cal/em!/'nin
a
15
•15
45
0.020
b
45
Not heated
45
0.45
<*
tiO
60
00
0.035
d
61
00
00
0.000
e
59
00
(iO
0.0H0
r
00 -
Not heated
60
II
.
These data showed that when all three units wore
heated to the same temperature the heat loss of the
disk was trivial as compared with the caloric uptake
of the skin at similar temperatures (see Table 5).
The slow rate of heat transfer from the crown to the
applicator disk was indicated by comparison of ex-
periments c, , and e. These data showed that the
exact setting of the crown temperature was not criti-
cal. A comparison of tin- data a, b, c, and / showed
the importance of the brim in preventing lateral
heat leakage from the applicator disk.
Needle Thermocouple for Determining Tissue
Temperature beneath Sites of Cutaneous
Exposure
It was desirable to be able to measure the temper-
ature of the tissue at various distances beneath the
surface of the skin before, during, and after exposure
to heat. For this purpose a needle thermocouple was
SECRET 314
STUDIES OF THERMAL INJURY CUTANEOUS \ NT> SYSTEMIC
constructed by threading a single silk-insulated
3-mil constantan wire through 4 feet of a No. 27
gauge trochar. The bimetallic junction was then
made by honing down the end of the trochar and
wire to a 45-degree angle; this removed the silk in-
sulation from the constantan wire and permitted it
to be surface soldered to the steel hypodermic needle.
Through experimentation it was found possible to
insert laterally a No. 22 gauge trochar along the
natural cleavage plane of the dermis-fat interface,
until a point directly underneath the surface area
to be exposed was reached. Then the No. 27 gauge
thermocouple needle was inserted into the No 22
gauge trochar until skin resistance could be per-
ceived, and the No. 27 gauge couple was withdrawn
about 1 cm. After the heat exposure was terminated,
the skin was cut to the needle depth and the distance
from the muscle-corium interface to the skin surface
was ascertained with a depth gauge. This depth be-
fore the application of heat was ascertained by a con-
trol experiment on a neighboring site.
The thermal emf of the steel-const ant an couple
was read on a Leeds & Northrop Type K2 potentiom-
eter and high-sensitivity galvanometer. The steel-
const ant an emf seemed to be very reproducible in
this temperature range 0 to 80 (.'. It was about 30 per
cent lower than the iron-const antan emf. Tempera-
ture differences of 0.1 C were readily determined.
Thermocouple foh Measuring Surface
Temperature of Skin
The surface temperature of skin exposed to air de-
pends upon two factors, namely, the rate at which
heat reaches the skin surface from the underlying
tissue and the rate at which (he skin surface loses
heat to the atmosphere. When the surface tempera-
ture of the skin reaches a steady state, these two
rates must be identical.
The use of the usual insulated thermocouples 19
for the measurement of skin temperature necessarily
alters these conditions. Upon first applying an in-
sulated thermocouple, no matter how perfect the
insulation, the temperature measured will be con-
siderably lower than the true surface temperature
because of the relatively high heat capacity of the
insulator. When a steady state of temperature is
finally reached (in some cases a matter of hours), the
temperature recorded by the insulated couple must
be greater than the true skin temperature, since the
skin site is no longer losing heat directly to the air.
Thus an accurate measurement of surface tempera-
tore by any apparatus similar to that just described
would be fortuitous.
A thermocouple for measuring the surface temper-
ature of the skin in this investigation consisted of a
bare 2 mil iron-constantan junction. The 2-mil wires
were prepared by dissolving (by nitric acid) the ends
of 15-mil iron and constantan thermocouple wires
(Leeds A Northrop) for a distance of 5 mm. The re-
duced ends of the two wires were then soldered end
to end and stretched tightly by means of a bow made
of brass tubing (see Figure 4). The heat capacity of
the junction was trivial as compared with that of
the skin.
Fioube 4. Thermocouple for measuring skin surface
temperature.
In use the junction was placed on the skin for
lateral contact and after It) seconds a reading was
made. The couple was then completely surrounded
by skin by pinching the neighboring epidermis and a
second reading was taken within 5 seconds. Numer-
ous such pairs of readings were recorded and in no case
has there been any significant difference between the
temperature of lateral and that of circumferential
contact. Thus a bare fine wire rapidly reaches skin
temperature (10 seconds) when it is in contact with
the skin.1*
The sensitivity of an iron-const an tan thermo-
couple is such that with a Leeds A Northrop Type K2
potentiometer and high-sensitivity galvanometer
'• A theoretical objection to the unprotected or bare wire
junction has been that it is partially exposed to the air and
thus will reach a temjieralure somewhere intermediate bet ween
the air and the skin temperature. It should l>e kept in mind
that the skin is also exposed to air. At normal air tempera-
tures, the heat transfer coefficient for both wire and skin to
air is quite small. Since the heat capacity of a fine wire is
small and its thermal conductivity high, one would expect the
wire rapidly to attain true skin temperature.
SECRET BASIC CHARACTERISTICS OF HEAT AND MEAT TRANSFER
temperature differences of 0.05 C were readily meas-
ured.
Determination of Heat Capacity of Four
Pertinent Tissues (in vitro)
The heat capacity of pig epidermis, dermis, fat,
and muscle were determined on approximately 10-g
samples of each tissue by the procedure given in
Section 17.3.2 under “Heat Capacity Apparatus.”
Determinations were made on each tissue of two
10-kg pigs. In order to obtain pure epidermis for
these determinations the following method was used.
After the hair was shaved as closely as possible, the
pig was immersed in water at 55 C for about I min-
ute, then removed, and the skin was carefully dried.
It was then possible to remove strips of pure epi-
dermis by scraping with a knife. The remaining tis-
sues were readily obtained in a relatively pure state
hy dissection.
The values of the heat capacities of these tissues
are given in Table 3.
the thermal conductivity. The temperature of the
tissue-cylinder interface was measured by means of
an iron-const an tan thermocouple soldered into the
face of the copper cylinder. The average t issue thick-
ness was determined by measuring (he distance of the
face of the applicator from the face of the cylinder.
The thermal conductivities of all the tissues except
epidermis were obtained by this procedure, since in
view of the epidermal thinness the above method
was not adaptable.
The method of difference was used with epidermis.
A section of well-shaved skin tissue consisting of
dermis and epidermis was rigidly clamped to the
copper cylinder, water at 55C was poured over the
skin, and the excess water was removed by blotting.
The clamps prevented lateral contraction of the
heated tissue and the hot water facilitated subse-
quent removal of the epidermis. The conductivity
determination was now made, the epidermis was then
scraped off, and the determination repeated. As a
further check, in certain experiments, a strip of in-
tact epidermis was placed over the denuded dermis
and the measurement repeated. The thickness of
numerous pig epidermal strips was determined with
a micrometer. Tire thickness was about 80 + 10 g.
At least triplicate determinations were made on
each of the four tissues of three different pigs (ap-
proximately 10 kg). The average values of the thermal
conductivities obtained on each of these tissues are
given in Table 4.
Table 3. Heat capacity of pig tissue
gram |ier C.
in calories per
Epidermis
Dermis
Fat
Muscle
Heat capacity
0.887
0.785
0.538
0.890
0.845
0.753
0.573
0.926
Average value
0.86
0.77
0.55
0.91
In view of the similar heat capacities of dry tissue,
the above variations of the different tissues are prob-
ably due to water content of tissue. In this respect
the high value for pig epidermis (0.80) is understand-
able since it was found experimentally that the water
content, in spite of the presence of the cornified layer,
averaged about 70 per cent.
Determination ok Thermal Conductivities of
Tissues (in vitro)
The experimental determinations of thermal con-
ductivities of pig epidermis, corium, fat, and muscle,
were based on equation (3) of Section 17.3.1. The
respective tissues were placed on a copper cylinder
2 inches in diameter and 4 inches high. The auto-
matic energy recording applicator was now placed
over and in contact with the tissue. Thus when the
tissue became “heat-saturated,” the knowledge of the
calorie input into the tissue, the temperatures of the
tissue-applicator (approximately 48 C) and tissue-
cylinder (approximately 30 C) interfaces, and the
thickness of the tissue permitted the computation of
Table 4.
K given
In vitro thermal conductivities K of pig tissue,
in (cal — cm)/(cm* — min — degrees C) units.
Kpidermis Dermis
Fat
Muscle
K
0036
0.051
0.021
0.064
0.023
0.053
0.024
0.062
0.032
0.051
0.023
0.073
K
0.03
0.053
0.023
O.OCG
In view of the thinness and uncertainty in the
thickness of the pig epidermis, the wide variation in
the epidermal thermal conductivity was to be ex-
pected. The data pertaining to the other tissues were
considerably more reproducible.
It is of interest to compare some of these data with
those of Brener,8 who determined the respective
thermal conductivities of both muscle and fat of cow,
horse, pig. and dog. This investigator found that the
conductivities of pig muscle and fat, expressed in the
above units, were 0.000 and 0.021 respectively; fur-
thermore essentially the same values were found for
SECRET STUDIES OF THERMAL INJURY — CUTANEOUS \MJ SYSTEMIC
the muscle and fat of the other three animals. In view
of the excellent agreement between Brener’s value
and the present one for pig muscle and fat, it is diffi-
cult to understand the value, 0.03, that Hardy and
Soderstrom 15,22 report for both cow muscle and fat.
Unfortunately no description of their experimental
method was given. In order to investigate this dis-
crepancy, the thermal conductivity of beef muscle
was redetermined and an average value of 0.057,
which checks Brener, was obtained.
In view of the numerous indeterminate factors
(Section 17.3.1) which enter into the in vivo conduc-
tion of heat through pig skin, the in vitro thermal
conductivities of these four tissues are not of them-
selves too useful. They do however serve as a base-
line-in the interpretation of certain experiments to be
described.
Observations {in vivo) of Caloric Uptakk of Pm
Skin and Rise in Temperature at Dermal-Fat
Interface as a Function of Both Time and Skin
Sir fac e Temperate re
It was of interest to ascertain (he caloric uptake of
the skin when the epidermal surface was maintained
at various temperature levels Iretween 15 and 100 C.
Numerous such experiments have been done and as
was to lie expected (see Section 17.3.1) the data were
subject to wide variations and are extremely difficult
to interpret in detail. Thus only a small fraction of
these data Mill be reported and the variations to be
expected will be indicated. During these experiments
the temperature at the dermal-fat interface was also
ascertained.
A pig under Nembutal anesthesia Mas clipped and
shaved. The hypodermic needle thermocouple Mas
introduced laterally into the dermal-fat interface.
The skin temperature at the chosen site Mas deter-
mined and the automatic energy recording appli-
cator Mas applied. Thus a continuous record of the
caloric uptake of the. skin at a predetermined epi-
dermal surface temperature Mas obtained. The tem-
perature at the dermal-fat interface Mas determined
either intermittently vith a & Northrup Type
K2 potentiometer or continually xx ith a Ceneral
Electric photoelectric recording potentiometer.
Caloric uptake rate of pig skin: Typical caloric up-
take data as a function of time and epidermal surface
temperature are presented in Table 5.
The data given in Table 5 are a composite of at
least three determinations on the lateral thoracic
area of different pigs; five pigs in all Mere used. As
Table A guide ( +30 |xt cent1 to the caloric uptake* of
the skin as a function
of time and surface
temperature as
determined l»v the automatic energy recori
ling applicator.
Time inter-
Skin surface temperature
val in min
45 ('
501’
55 ('
-
dcrmal fat in lateral thoracic area of a 10-kg pig lies
about 2 mm below the skin surface. Ten-minute ex-
posures to surface temperatures of 50 to 70C in-
creased significantly the thickness of the dermis. This
increase in thickness was due to the accumulation of
edema fluid in the dermis and the effect was maximal
when the skin surface was maintained at about GO C.
Skin surface temperatures of 45 C or below do not
activate the mechanism which gave rise to edema.
Skin surface temperatures equal to, or greater than,
SO C denature the curium so rapidly that (he mechan-
ism by which edema fluid accumulated in the curium
was destroyed.
2. Although the continual caloric uptake by the
skin (ended to increase the dermal temperature, the
appearance of relatively cool edema fluid tended to
decrease it. At skin surface temperatures of 50 C and
TOC, these two effects nearly counterbalanced each
other, and after the first minute of heat exposure the
dermal-fat interface temperature remained essen-
tially constant. With skin surface temperatures be-
tween 55 and Go C t he rapid appearance of a large
amount of edema fluid more than compensated for
caloric uptake, and the temperature at the interface
between dermis and fat was temporarily lowered.
This effect was maximal when the skin surface was
maintained at about GO C.
3. When the skin surface temperature was main-
tained at 45 C, and probably at all other tempera-
tures that fail to cause edema, the dermis becomes
“heat-saturated” after about 5 minutes of exposure.
When edema fluid w as produced, the time for dermal
heat saturation was essentially indeterminate, but it
apparently was greater than 10 minutes.
4. Histological examinations showed that com-
plete primary injury to the dermis was obtained in all
experiments where the skin surface temperature was
maintained at 65° or higher. These limited (5)
time-temperature-injury data at the dermal-fat in-
terface tended to indicate a quantitative relation-
ship very similar to that found for epidermal injury'
(see Section 17.7).
5. By making t he reasonable assumption that the
dermis is essentially “heat-saturated” at the end of a
SECRET 318
STUDIES OF THERMAL INJURY CUTANEOU S \ M) SYSTEMIC
10-minute heat exposure, the in vitro thermal con-
ductivities of dermis can be computed by substitut-
ing the approximate caloric uptake (Table 5), dermal-
fat interface and skin surface temperatures, and the
final dermal thickness into equation (3) (Section
17.3.1); the neglect of the epidermal temperature
drop introduced no appreciable error. Table 5 also
shows the results of these calculations.
A comparison of these values with the experi-
mentally determined in vitro values (Table 4) for pig
dermis indicated that the presence of edema fluid
increased the thermal conductivity of dermis two-
to threefold. This increase in conductivity, however,
was slightly more than compensated by the swelling
of the dermis; an edematous dermis is thus a some-
what better heat barrier to the underlying tissues
than normal dermis. A comparison of the in vivo
thermal conductivity obtained at 45 C with the
in vitro value of 0.053 (see Table 4) tends to indicate
that intact circulation probably increased the effec-
tive thermal conductivity of dermis by about 15 per
cent.
Estimation of Temperature Changes at
Epidermal-Dermal Interface during Exposure
of the Skin Surface to Heat
In view of the thinness (~80g) of the pig’s epi-
dermis, the experimental measurement of (he time-
temperature relationships at the epidermal-corium
junction was not feasible.
There are certain facts, however, that allowed the
estimation of this time-temperature relationship with
a considerable degree of certainty. In view of the ex-
treme thinness of epidermis, the temperature of the
basal layer was largely determined by skin surface
temperature, which was an accurately known quan-
tity. This is most readily seen by solving heat con-
duction equation (3) for steady-state temperature of
the basal epidermal layer. Of the four necessary ex-
perimental quantities, namely, skin surface tempera-
ture, epidermal thickness (alxnit HO/d, epidermal
thermal conductivity (Table 4), am 1 caloric uptake
of the skin at the requisite skin surface temperature
(Table 5), only the last two were subject to a con-
siderable variation (±30 pier cent). Fortunately, even
variations of this magnitude resulted in uncertain-
ties of less than 0.2 C in the steady-state tempera-
tun' of the basal epidermal layer.
Basal Epidermal Temperatures When the. Skin Sur-
face is Immediately Brought to amt Maintained at a
Temperature between C and 100 C. Before the
steady-stale temperature is attained, the time-tem-
perature relationship at (his epidermal-dermal junc-
tion is given under these conditions to a good approx-
imation by equation (tie) of Section 17.3.1, where y
has the following numerical value:
y — 0.15
if the time I is expressed in second*.
The numerical constant, 0.15, is not subject to the
experimental uncertainties of the quantities requisite
to computation by equation (0a), since it can la1
quite accurately determined empirically fiom the
temperature-time-epidermal injury data (see Sec-
tion 17.0.5 for details). An identical value for y can
also he.directly computed by substituting into equa-
tion (0c) the experimentally determined values for
heat capacity, thermal conductivity, and thickness
of epidermis, and by assuming an epidermal density
of 0.8 g cc (a most reasonable value). In view of (he
two completely independent, methods, one of which
was in vivo and the other in vitro, considerable confi-
dence could be placed in the -adaptation of the in-
finite body picture (see Section 17.3.1) to the solution
of the time-tomperature relationship at the epi-
dermal-dermal junction during (he unsteady state
jjeriod of heat How.
The computation of the temperature of the basal
cell layer of the epidermis as a function of both time
and skin surface temperature is given in Table 7A.
Table 7A. The computed time-temperature relation-
ships for (lie epidermal-dermal interface when the skin
surface is immediately brought to and maintained at a
specific temperature.
Time
in
seconds
Surface temperature, C
45 55 05 SO
TemiMTature at basal epidermal
100
layer* 0
0
35.0
35.0
35.0
35.0
35.0
0.01
30.3
37.0
0.02
38.9
10.9
13.4
0.05
41.8
45.2
50.3
57.1
0.1
40.1
45.2
50.3
57.9
68.2
0.2
41.3
4 7.0
53.9
63.3
75.9
0.5
12.7
50.4
58.1
09.6
85.1
1.0
13.3
51.0
60.0
72.4
89.1
20
43. S
52.6
61.4
74.0
92.3
5
44.2
53.5
62.7
70.6
95.1
10
44.5
53.9
03.4
77.6
96.0
30
44.7
54.4
01.1
78.6
98.0
tiOCl min)
44.S
51.6
64.4
79.0
98.6
120(2 min)
44.9
54.9
64.5
79.4
99.2
300 (5 min)
44.9
54.9
64.7
79.5
99.3
000 (10 min)
44.9
54.9
64.8
79.7
99.6
Steady
44.8
54.5
61.2
—
stale f
* Computed by
equation
(t»c) and nrpprimpnt&I <
lata of Section 3.2.
t Computed by equation
(3) and experimental <
lata of Section 3.2.
SECRET BASIC CHARACTERISTICS OF HEAT AM) HEAT TRANSFER
319
Table 7H. The computed time-temperature relation-
ships for (lie epidermal-dermal interface when an
entire
animal {^-
30 cm
in diameter) is surrounded by
an on-
velojie of i
unbient
and radiant heat that
results from a
constant (em|ierature source.
(
'ircumambient temperature, C
80
100 125
1.50
175
Time
Heat transfer coefficient //* in
in
cal/cm*/tn in |ier f
seconds
0.015
0.019 0.021
0.024
0.020
Temjierature at basal epidermal layer, ft,}
0
35
35 35
35
35
10
37
39 40.5
44
40
20
40.5
lit
30
38.5
41.5 44
49
52
40
51
54.6
50
30.5
43.5 40.5
53
57
70
to
44 48
56
00
100
41
45.5 50
59
04
130
42
47 52
01
100
42.5
48.5 54.5
03
200
43
50 50
05
'
300
15
52.5 59
400
40
55 03
500
47
GOO
48
800
50
1,000
50.5
—
1,200
51
• In order U
> make these data directly comparable to the. ex peri mental
i(ivestiga lions t
»f StTtion
17.0, the radiant contribution to // wa
* computed
by using a sour
ce temperature 20 j»er cent in excess of the air temperature.
t Computed by mean
s of equations (o). (6), (fia), !
and (6b) and exjieri-
mental data of Section 1
t Beeau.se of hoth the
thinness of the epidermis and the slow rate of heat
transport to the skin.
there is no appreciable diffe
renee between these
tenijjeratures a
nd 1 howe i
r>f the skin surface after the first 20 sore
»mls of heat
ex|)0»urc.
temperature and the skin surface temperature was
trivial.
It must he re-emphasized that these data apply
only to situations in which the heat transfer coeffi-
cient II from the temperature source to the skin sur-
face is infinite.® In all cases where II is finite an analy-
sis similar to that given below is required.
liasnl Epidermal Temperatures When the Entire
Animal Is Surrounded by an Envelope of Ambient and
Radiant Ileal between SO and 175 In the previous
section, the time-temperature relationships at the
epidermal-dermal junction depended'only upon the
rate of heat transfer through the skin and the con-
stant temperature of the heat source. To this must
now be added the slow rale at which heat is trans-
ported from the heat source to the skin surface via
air conduction, air convection, and infrared radi-
ation. The mathematical solution of this problem is
given by equation (0), where the only quantity that
requires further consideration is //, the heat transfer
coefficient from the heat source to the skin surface.
This quantity is readily computed through the sub-
stitution of equation (I), heat transfer by convection,
and equation (2), heat transfer by radiation, into
equation (5). The numerical values of the heat trans-
fer coefficient which were obtained at certain source
or air temperatures are shown in Table 7B. A com-
parison of the numerical values of //, 0.015 to 0.020
calorie per square centimeter per minute per C, with
epidermal thermal conductance K/L (Table 4) nu-
merically equal to 4 in the same units, indicates the
slow rate at which ambient and radiant heat is trans-
ferred to the skin surface as compared with the rate
this heat flows through the epidermis.
Table 7B also gives tlie estimated temperature of
the basal epidermal cell layer as function of source or
air temperature as calculated by means of equation
(6). These data show the ext reme slowness of tem-
perature rise at this epidermal-dermal junction. In
fact, under these conditions, the epidermal tempera-
ture even after a heat exposure of 15 minutes is far
lower than the temperature of the heat source, and
one would expect an animal to succumb to hyper-
thermia long before the temperature of the skin
approached that of the air.
Although the data for the time-temperature rela-
tionship at the skin surface are not given, they can be
The data given in Table 7A show that (here was a
rapid rise in the temperature of the basal epidermal
layer when the skin surface was immediately brought
to and maintained at a specified constant tempera-
ture. A comparison of the unsteady-state data com-
puted from equation (0c) with the steady-state data
obtained by means of equation (it) showed that the
epidermis under the above conditions became essen-
tially “heat-saturated” after a heat exposure of
0.5- to 1.0-minute duration.
Actually, only the unsteady-state time-tempera-
ture relationship as given by equation (6c) need be
considered to elucidate the irreversible epidermal in-
jury threshold data of Section 17.6.5; since these ex-
perimental tinne-temperature-epidermal injury rela-
tionships were such that for all skin surface tempera-
tures above 50 C the epidermis never reached heat
saturation, and for all temperatures below 50 (' the
difference between the steady-state basal epidermal
* Under (lie experimental conditions to be dcscrilicd in
Section 17.6 (hot water experiments), II is not infinite 34 hut
rather about 105 cal/eml/min j>er C. In these computations
t tie sulwtitulion of for l(p is of no significance.
SECRET’ 320
STUDIES OF THERMAL INJURY CUTANEOUS AND SYSTEMIC
readily computed by putting L (the thickness of the
epidermis) equal to zero in equation ((»). If this be
done, it will be found that, except for the first, 20
seconds of heat exposure, the skin surface tempera-
ture Is not significantly different from the values re-
corded in Table 7B for the basal epidermal tempera-
ture. This is due to the fact that heat transfer to the
skin is the controlling factor. Thus, these data can
also be taken as the temperature of the skin surface
as a function of time.
A comparison of Tables 7A and 7B indicates the
importance to the epidermal time-temperature rela-
tionships of the mode of imparting heat to skin sur-
face. Thus,fora given source temperature, a mecha-
nism that enables the surface temperature to be im-
mediately brought to and maintained at the source
temperature has, on a time basis, at least a thousand
times greater injury propensity to epidermis than a
heat source which raises the skin temperature by
means of radiation, conduction, and convection of
relatively immobile air. (See Section 17.9.2 under
“Measurement of Heat Transfer ”)
17.3.3 Summary
The various physical factors which determine the
transfer of heat energy to and through (he skin and
the temperatures attained thereby have been de-
fined and discussed.
A general theory of heat flow through the epi-
dermis is developed.
Experimental observations pertaining to the rate
at which heat energy is taken up by the skin during
surface exposures of varying intensity and the sub-
surface thermal gradients established therein have
been presented.
The time-temperature relationship at the dermal-
epidermal junction is computed under two greatly
different experimental conditions: (1) when (he skin
surface temperature is immediately brought to and
maintained at the Temperature of the heat source,
and (2) when the entire skin surface is exposed to a
specified circumambient and eircumradiant temper-
ature. These data indicate the extreme importance of
the mode of applying heat to the skin surface to the
time-temperature relationships within the epidermis.
ITT EFFECTS OF INHALED HEAT
It was inferred from the results of the pilot experi-
ments (Section 17.2.5) that, so far as rapid neutral-
ization of enemy personnel by flame thrower attack
is concerned, the effects of heat on the surface of the
laxly are probably of greater importance than are its
effects on the air passages and lungs. The implication
of this assumpt ion is too great to accept at face value
the small amount of evidence provided by the pilot
experiments.
A search of the literature failed to disclose any re-
liable information concerning the effects on the lungs
and air passages of inhaled heat or the circumstances
in which thermal injuries of the respiratory tract
may be sustained. The following investigation was
accordingly undertaken.*7
IT.1.1 Experimental Procedure
In order to study the effects of heat on the respira-
tory tract independently of the secondary changes
that might result from concomitant burning of the
skin, dogs were caused to breath hot air which was
conducted directly to the trachea through an in-
sulated transoral cannula.
In some experiments heated air was pumped di-
rectly into the air passages and in others it was in-
haled by the respiratory efforts of the animal. The
inner end of the cannula extended below the vocal
folds of the larynx. Three types of inhalation experi-
ments were performed. In the first the animals
breathed room atmosphere heated to temperatures
as high as 500 C in an oven. In the second, flame
from a blast burner at temperatures estimated to be
it! the vicinity of 1000 C was directed into the ex-
ternal end of the cannula. In the third, a mixture of
live steam and air was breathed from a generator
(see Figure 5). All experiments were conducted un-
der anesthesia induced by the intravenous or intra-
peril oneal injection of sodium pentobarbital.
The external temperature of the air available for
respiration in each type of experiment was measured
either by a thermometer or a platinum-rhodium
thermocouple. Thermocouples (10 gauge eopper-
constantan) were installed in the airway, one at the
laryngeal end of the transoral cannula and the other
at or near the bifurcation of the trachea, to measure
the rate at which the inhaled air was cooled. I-eads
from these thermocouples were connected with a
Mold galvanometer having a period of 0.2 second.
The excursions of the galvanometer wore observed
directly and recorded manually.
IT.T2 Kate of Cooling of Inhaled Air
When the superheated air was inhaled, the tem-
perature recorded by both the laryngeal and the
SECRET EFFECTS OF INHALED HEAT
321
Fkii uf. 5. Experimental procedure used to investigate effects of inhaled heat on air passages and lungs. In all instances,
insulated cannula conveyed hot air, flame, or steam from outside to animal’s larynx. Position of intra-laryngeal and
deep tracheal thermocouples is shown. Top left view: Animal breathed room temperature heated in oven to 350 Top
right view: Room temperature was pumped into animal’s lungs from combustion oven which was heated to 500 C,
Bottom tefl view: Flame and combustion products of blast burner were projected into cannula during each inspiration.
Bottom right view: A 400 ml blast of mixture of live steam and air was released into Iransoral cannula at the beginning
of each inspiratory effort. Results of these exjjcrimonts arc shown in Table 8.
Table 8. Results of experiments in
breathing of hot air
_
Kind of
atmosphere
breathed
Xo,
Animal
Xo.
Original pre-
inspiratory
temp of air
(C)
(approximate)
No. of
breaths
Max
temperature -
recorded (C)
Laryn-
geal Lower
cannula trachea
Recovery
period
(hours)
Site and severity of injury
Upper Lower
trachea trachea Tilings
Air from dry-
1
423
350
46
182
19
Mild
None
None
ing oven.
2
420
350
52
ISO
19
Mild
None
None
See Fig. 5A
3
391
350
103
159
30
Mild
None
N one
4
390
350
lOti
175
Not
(Complete clinical recovery —■
killed
no autopsy)
Air from com-
5
392
500
GO
267
4
Mild
None
None
bust ion oven.
G
432
.500
44
327
50
7
Moderate
None
None
See Fig. 511
7
42G
500
22
291
24
Mild
None
None
8
431
17
135
7
Moderate
Mild
None
Flame from
9
433
10
327
51
S
Severe.
Moderate
Mild
blast burner.
10
454
16
540
100
11
Severe
Mild
None
Sec Fig. 5C
11
455
24
550
65
24
Moderate
Mild
None
12
405
14
510
64
Not
(Complete clinical recovery -
killed
no autopsy)
Steam from
13
456
Over 100
27
106
59
- 6
Moderate
Mild
None
generator.
14
519
Over 100
18
98
79
7
Severe
Moderate
None
See Fig. 5D
15
4S1
Over 100
20
94
53
10
Severe
Severe
Severe
)G
475
Over 100
16
99
94
10
Severe
Severe
Severe
17
524
Over 100
10
90
24
Severe
Severe
Moderate
IS
522
Over 100
12
75
18
Severe
Severe
Mild
tracheal thermocouples rose throughout inspiration
and fell during expiration. In each situation the high-
est point in the temperature curve was reached at or
near the end of inspiration. The inhaled gas lost most
of its heat before reaching the lungs. When the in-
haled gases were relatively dry, the intratracheal
temperature rose to a sharp peak and fell away
rapidly during expiration. When steam was inhaled,
the curve described a plateau rather than a peak,
probably because of the condensation of hot water on
the thermocouple. The results of these experiments
are shown in Table 8.
When air heated to bet ween 350 and 500 C was in-
haled, the temperature fell to about half of its ex-
SECRET 322
STUDIES OF THERMAL INJURY — CUTANEOUS AND SYSTEMIC
Finnic 7. Thermal tracheitis and pneumonitis.
Photograph of respiratory tract of dog 10 hours after
inhalation of steam, showing severe tracheobronchitis
with dilatation of bronchi. There is central hemorrha-
gic pneumonitis with generalized pulmonary edema and
hyperemia.
Finnic ft. Thermal laryngitis and tracheitis without
pulmonary injury. Photograph of respiratory tract of
dog 24 hours after inhalation of flame. Sufficient heal
had licen conducted through wall of cannula to cause
mild degree of laryngeal edema which may be recog-
nized by bilateral olive-shaped mucosal protrusions
from ventricular recesses. There was extensive destruc-
tion of mucosa of upjKT trachea, diminishing rapidly to
mild catarrhal inflammation in lower third Xo abnor-
mality of bronchi or lungs of this animal was recog-
nized. —
of the trachea in such experiments was 135 C. When
a mixture of live steam and air was inhaled, the in-
spiratory peaks recorded at the laryngeal opening of
the cannula ranged between 04 and 106 C and those
by the deep tracheal thermocouple, between 53 and
94 C.
17.4.3 Effects on Animals
The mildest thermal exposure used in the inhala-
tion experiments was more than sufficient to cause
severe injury to the skin. Every animal included in
Table 8 would have sustained severe cutaneous in-
jury if the skin had been exposed for more than a few
ternal level by the time it reached the larynx, despite
the fact (hat it was conducted through the mouth by
means of an insulated cannula. By the time it, had
reached the bifurcation of the trachea, the tempera-
ture hail dropped to approximately 50 C. Flame and
combustion products of a blast burner directed into
the external end of the transoral cannula were de-
livered to the larynx at temperatures between 300
and 550 C. The highest recording at the bifurcation
SECRET 323
EFFECTS OF INHALED HEAT
seconds to such temperatures. Circumambient air
temperatures as low as 300 C produce severe injury
of unprotected skin within a few seconds. Mixtures
of steam and air at 100 C destroy epidermis even
more quickly.
Early in the investigation it was found that if ani-
mals were to survive the inhalation experiments long
enough to develop reactive changes in the lower air
passages it was necessary to protect the larynx.
Otherwise they died prematurely of asphyxia due to
laryngeal edema. For this reason the transoral can-
nula was inserted well lx*low the glottic folds.
Primary thermal injury of the lungs occurred in
none of the 7 animals that breathed hot air, in only 1
of the 5 animals that inhaled flame from a blast
burner, and in 1 of the 0 animals that inhaled live
steam. In the remaining animals thermal injury to
the respiratory tract was confined to the upper air
passages. In no instance did an animal die as a result
of thermal injury of the lungs within the first 24
hours. All animals that sustained thermal injuries of
the respiratory tract would, under nonexperimental
conditions, have received severe cutaneous burns.
Mucosal necrosis with desquamation of surface
epithelium o< sirred in alt instances where the blast of
hot atmosphere first struck the lower portion of the
larynx and the upper portion of the trachea. In the
case of hot air the injury was usually localized ami
represented by shallow ulceration associated with
catarrhal inflammation of the upper third of the
trachea (Figure 6). Inhalation of flame or steam led
to extensive destruction of the trachea with edema
of the peritracheal areolar tissue of the neck and
mediastinum and detachment of large casts ot ne-
crotic mucous membrane, which were either expelled
by coughing or subsequently inhaled into the lower
portions of the respiratory tract (Figure 7).
The portions of the lungs most vulnerable to in-
jury' were the centrally located alveolar ducts and
their communicating alveoli (Figure 8). Atmosphere
not hot enough to damage the mucosa of the large
bronchi or the alveoli of the more peripheral portions
of the lungs was in some instances capable of causing
central pulmonary edema and both intra-alveolar
and interstitial hemorrhage. After more severe ex-
posures the lungs became diffusely edematous and
hemorrhagic. Focal patches of atelectasis and em-
physema were observed and in some instances were
obviously due to aspiration of mucus or mucosal
debris. Bronchopneumonia was commonly' observed
in animals that had received tracheal burns. It ap-
pea red that, regardless of the mildness of the pri-
mary thermal injury of the lungs, if the inhaled air
was hot enough to damage the trachea it usually
predisposed the animal to pneumonia.
IT.t.l Discussion
Tf was apparent, from the foregoing observations
that air hot enough to burn the skin can be inhaled
without causing damage to the trachea or lungs and
that if the temperature of the air is high enough to
damage the respiratory passages it will inevitably
have caused burning of the surface of the body.
This observation seemed paradoxical in view of the
fact that the mucosa of the air passages is much
thinner than the skin and should therefore be more
vulnerable to thermal injury. The explanation of the
experimental findings lies in the fact that the quan-
tity of heat that can lie stored in the volume of gas
that constitutes a breath is remarkably small. At any
given air temperature the number of calories that can
be transferred to the respiratory tract incident to the
inhalation of a breath of hot air is limited by the vol-
ume of that breath, whereas convection currents are
capable of bringing a practically unlimited volume of
hot air in contact with the skin. An infinitely greater
caloric transfer can occur for each unit of surface
exposed.
Not only is the amount of heat energy available
for transfer to the skin greater than that which is
available for transfer to the respiratory membranes
but also there are Important time differences be-
tween cutaneous and respiratory exposures. In the
case of the skin the exposure is virtually continuous,
whereas the lining of the air passages is exposed in-
termittently as each new breath is inhaled.
An instructive illustration is provided by calcu-
lating the potential heat trmisfer to the respiratory
tract that might occur if air were inhaled at 142 C.
Ix*t it be assumed that the amount inhaled with each
breath would be sufficient to increase the pulmonary
volume by 500 ml, that the air was dry when inhaled
and saturated with moisture when exhaled, and that
it was cooled to body temperature by the time it left
the body. Approximately 13 cal of heat energy
could be released within the hotly by cooling of one
such breath from 142 to 38 C. Theoretically this
amount of heat would be sufficient to raise the tem-
perature of 1 g of tissue by approximately 13 degrees,
providing none of it was carried away by the blood
circulating in the subsurface capillaries. Actually no
change in the net temperature of the respiratory
SECRET 324
STUDIES OF THERMAL INJURY — CUTANEOUS UVD SYSTEMIC
Fic.cue S, Primary thermal pneumonitis. Photomicrograph of lower lol>e of dog’s lung 24 hours after inhalation of
steam. Although there was severe tracheitis, primary and secondary bronchi showed remarkably lit tie change. Evidence
of pulmonary injury was confined largely to the central {tortious of lobes and consisted of hyperemia, edema, and partial
atelectasis.
tract would occur in such circumstances because the
gain of 13 cal would be offset by a loss of 13 cal inci-
dent to the evaporation of the 23 mg of water that
would be required to saturate that amount of dry air.
This is not to imply that the inhalation of air
heated to 142 C would be necessarily harmless. Desic-
cation would probably occur near t he portal of entry
even though there were no net change in the temper-
ature of the respiratory' tract as a whole. The calcu-
lation serves to emphasize how important the heat
capacity of the inhaled gas is in relation to the prob-
lem of thermal injury of the lungs. A rise in tissue
temperature is prerequisite to the occurrence of
thermal injury and the amount (hat the tissue tem-
poraturc is raised incident to any given exposure will
depend in part on the magnitude of temperature dif-
ferential and in part on the amount of heat energy
that the inhaled gas is capable of storing.
A more important attribute of an inhaled hot gas
than its temperature in relation to its capacity to
cause thermal injury is its water content. When
steam or a mixture of steam and air comes in contact
with a cool surface such as the skin or the lining of
the respiratory tract, water is condensed on the sur-
face with liberation of a relatively large amount of
heat.
Thus the cording of a 500-rnI mixture of equal
parts of steam and air from 125 to 38 C would lead
SECRET COMPAlii.SON OF PORCINE VM) III MAN SK.IN
325
to the condensation of about 800 mg of water. The
heat energy liberated by this amount would lx* in the
neighborhood of 175 cal. There is little doubt but
that the sudden liberation of 175 cal to the lining of
the air passages or on the surface of the skin would
!x> capable of causing some injury.
17.1.5 Summary
It was apparent from these experiments that ther-
mal injury of the lungs is probably a negligible factor
in the causation of disability or death incident to
exposure trreonflagrations such as might result from
Hame thrower action. A thermal exposure of suffi-
cient, intensity to cause direct injury of the lungs was
more than sufficient not only to cause extensive burn-
ing of unprotected skin but also to result in rapidly
fatal obstructive edema of the glottis. In the case of
externally unburned or mildly burned casualties of a
flame attack it can be assumed that no significant
thermal injuries of the respiratory tract have been
sustained.
17.5 COMPARISON OK PORCINE AND
HUMAN SKIN
The original choice of the pig as a suitable subject
for this investigation was based on the fact that no
other readily available animal has skin that bears so
close an anatomical resemblance to that of man.
A comparison of the structural characteristics of
porcine and human skin at this point seems desirable
in view of the extent to which the pig was used in ex-
periments designed to provide information regarding
( I) the reciprocal relat ionship of time to temperature
in the production of cutaneous injuries in man, and
(2) the local and systemic disturbances in man which
cutaneous hyperthermia may be capable of causing.
Like that ol man the surface of the pig's body is
covered by three layers of tissue. Progressing from
outside in, these are the epidermis comprising strati-
fied epithelial cells, the dermis comprising fibrous
connective tissue, the hypodermis comprising fibrous
connective tissue, and the hypodermis comprising
fit>roadipose tissue (see Figures 9 and 10).
17.5. i Epidermis
The epidermis of the pig varies in thickness, the
average over the lateral body surface of immature
animals (8 12 kg) Ix-ing approximately 0.1 mm,
which is slightly less than that from corresponding
areas of adult human subjects. As with man there
are irregularities in contour of both the upper and
lower surfaces of the epidermis, those on the upper
surface being due to an intricate system of intercom-
municating linear depressions and those on the lower
surface corresponding to the dermal papillae over
which it is moulded. The hairs penetrating the epi-
dermis of the pig are thicker and more numerous
than those of man.
Microscopic appearance of epidermis: Like that of
man the outermost zone of epidermis or stratum
coraeu m of the pig consists of several loosely con-
nected layers of the desiccated and intensely baso-
philic remains of keratinized epithelial cells.
The second or granular layer is thin and consists of
several layers of dead or dying squamous cells, the
acidophilic cytoplasm of which contains many fine,
deeply basophilic kerato-hvaline granules. Many of
these cells have lost their nucleuses. Others contain
shrunkenTiyperchromatic nucleuses or Feulgen nega-
tive nuclear ghosts...
The third zone is comprised of several layers of
aging squamous cells which nolonger have any direct
cytoplasmic attachment to the dermis. The cyto-
plasm is dense, deeply acidophilic, and ap|>ears des-
iccated. The cells are so closely packed that neither
intercellular bridges nor spaces can be recognized.
Many of the nuclei are relatively small and more
densely packed with chromatin granules than those
of the deeper cells.
The fourth zone consists of cells in transition lie-
tween the squamous and the basal cell layer. The
transitional cells are large and polyhedral and many
of them still have an attenuated footlike cytoplasmic
attachment to the dermis. It is in this zone that in-
tercellular bridges of tonofibrils are most readily
visualized. The cytoplasm is moderately basophilic.
The cell outlines are distinct and the intercellular
spaces are clearly defined. The nuclei arc larger and
rounder than those of the more superficial cells and
contain several coarse and many fine granules of
chromatin.
The fifth zone is comprised of the basal cells,
which, except for their cuboidal or columnar shape
and their palisadelike arrangement on the dermis,
are essentially similar to the overlying transitional
cells. Projecting from the inferior surface of the basal
epidermal cells of the pig are many robust tono-
fibrils which appear to be embedded in the dense felt
work of fine collagen fibrils that comprise the super-
ficial zone of dermis. No such fibrillar anchorage of
epidermis to dermis can be seen in human skin (see
Figures 19 and 20).
SECRET 326
STUDIES OK THERMAL INJURY — CUTANEOUS AND SYSTEMIC
Appearance of porcine (Figure 9) and human (Figure 10) skin under low magnification, stained with phloxinc-methylenc
blue. Sections are representative of lateral thoracic region of pig and lateral abdominal region of man. Epidermis is
slightly thicker in man, and dermal papillae are broader in pig. Collagenous bundles in dermis of pig are heavier than
those in man. Glands shown in hypoderrnis of pig do not secrete sweat.
Figure 9
Figure 10
Tlte microscopic appearance of ( he epidermis of
hot it man and pig suggests that there is a progressive
loss of intracellular water as the epithelial cells grow
older anrcine skin which
measured 2\2x2 mm. Series of thick (50m) benzidine-treated horizontal and vertical sections were mounted in such a
way as to show distribution of veins, arteries, and capillaries at various levels beneath surface. No. I shows capillary
plexus lying in most superficial (50m) portion of dermis. No. 6 shows vessels in most superficial layer of adi|«isc tissue of
hypodermis.
only slightly less compact than the reticular zone.
The deeper portion of the reticular connective tissue
sends trabecular extensions into the underlying adi-
pose hypodermis.
Blood Vessels of Porcine Skin
It was observed in ordinary histological prepara-
tions that the appearance of the capillaries in the
dermal papillae of the body skin of the pig is similar
to that in corresponding regions of man. In recog-
nition of the fact that it is difficult or impossible to
get an accurate impression of so complicated a
structure as a capillary network by two-dimensional
visualization, a modification of the Pick worth tech-
nique 10 was employed in order that the dermal blood
vessels could lie studied in three dimensions.
Maximum cutaneous hyperemia was induced be-
neath a circumscribed area of the lateral body sur-
face of the pig by exposure to water at 50 C for
20 minutes. After such an exposure the erythrocytes
were so densely parked in (he distended capillaries
that there was practically no loss of blood from them
when the skin was excised. Skin and subcutaneous
tissue treated in this way was excised to a depth of
8 mrn, fixed in 10 per cent formalin, cut in thick
sections, and treated with benzidine.
The benzidine combined with the hemoglobin to
impart a dark blue color to the contents of the en-
gorged vessels. After skin treated in this manner was
cleared, a three-dimensional study of its blood vessels
could be made by use of a binocular microscope.
The appearance of the dermal vessels of porcine
skin at various levels below the surface is shown in
Figure 11. To prepare this illustration a block of
benzidine-treated skin was cut serially and parallel
to the surface in sections measuring 50 p in thick-
ness. Another block of the same skin was cut serially
and at right angles to the surface. Photographs were
made of both series and the prints were mounted in
such a manner as to orient the horizontal sections in
SECRET 328
STUDIES OF THERMAL INJURY CUTANEOUS AND SYSTEMIC
relation to the depth below (he surface that each
represented.
The epidermis was removed from the surface of the
block of skin shown in Figure 11. The excised skin
was not clamped prior to fixation and postexcisional
contraction resulted in an accentuation both in the
height of the dermal papillae and also in the thick-
ness of the dermis. It may be seen that (he fibrous
dermis including the papillae measures approxi-
mately 2 mm in thickness and that broad septa of
fibrous connective tissue extend down from the der-
mis at more or less regular intervals into the under-
lying fat.
In approaching the surface the blood vessels to the
skin followed an oblique course through the hypo-
dermis and after reaching the lower layer of the
fibrous dermis branched horizontally with multiple
intervenal and interarterial anastomoses. From these
first approximately horizontal plexuses originated a
series of broad vascular loops that penetrated to the
mid-portion of the dermisT Interarterial and inter-
venal anastomoses between these loops served to
establish a mid-dermal plexus. From this mid-dermal
plexus originated numerous hairpin-shaped capillary
loops which extended upward into the dermal papil-
lae. These capillary loops anastomosed freely with
one another and constituted the most superficial or
papillary plexus. It was apparent that capillary com-
munications between the arterioles and venules oc-
curred at different levels. Some followed a course
that brought them to within a few microns of the
basal epithelial cells over the tips of the papillae.
Still others followed an almost horizontal course to
establish communications between the arterioles and
venules of the intermediate plexus. At all levels
through the dermis there were numerous vascular
communications with the mantlelike mesh work of
capillaries that surrounded the hair follicles and
dermal glands.
As may he seen in Figure 11 the number, size, dis-
tribution, and communications of the dermal blood
vessels of the pig are remarkably similar to those de-
scribed by both Lewis” and Spalteholz M in human
skin. The similarity of blood vessels in human and
porcine skin was found to be so great that it was with
difficulty that one could be distinguished from the
other in Pickworth preparations.
It is not intended to imply that the anatomical re-
semblance between t he vessels of human and porcine
skin implies an equal degree of functional similarity.
Certainly the vascularization of both indicates that
ample and similar mechanical facilities exist either
for the transfer of body heat to the surface to facili-
tate its dissipation, or tor the conduct of surface heat
to the interior to raise the internal temperature of
the body.
SweaTGlands and Sweating
Several types of glands arc encountered in the
dermis and hypodermis of the pig and, although one
of them bears some resemblance to the sudoriferous
glands of human skin, it does not secrete a significant
amount of sweat.
The fact that the pig does not sweat was verified
by a series of experiments in which the water loss
from the skin of living pigs was measured at various
environmental temperatures, with and without the
administration of pilocarpine (see Table 9).
Tabi.K 9. Rate of water loss from surface of human and porcine "skin. Amount of water loss determined by
weight of Mg (CICMj contained in base of weighing bottle during the time that the neck of the brittle was held in
the skin.
accretion in
contact with
Water uptake (mg/em’/min) during a period of 10 minutes
Temp 21 C — Humidity 30-40% Temp 36 C -—Humidity 30—10%
No, of No. of
tests Min — Max Mean tests Min Max Mean
Dead pig (lateral thoracic region) 4
Live pig (lateral thoracic region)
0.016 0.026
0.019 1
0.023
0.031
0.027
Without pilocarpine 5
Live pig (lateral thigh)
0.016 0.020
0.021 0 -
0.020
0.032
0.028
Without pilocarpine
1
0.018
0.028
0.024
With pilocarpine*
(1 mg/kg bwt)
Live man (forearm)
4
0.021
0.030
0.027
Subject #1 (A.R.) 1
• • • • . . . .
0.027 1
0.180
Without pilocarpine
—
Subject #2 (A M.) 2
Without pilocarpine
0.028 0.038
0.033 - 2
0.280
0.360
0.320
* Iodine color test negative.
SECRET RECIPROCAL RELATIONSHIPS OF TIME ANI TEMPERATURE
329
It was found that the water loss from the skin of a
live pig does not differ significantly from that of one
that is dead. In a cool environment the water loss
per square centimeter per minute is approximately
the same in man and pig. At higher environmental
temperatures, the rate of water loss from human
skin is tremendously augmented, whereas the corre-
sponding increase in water loss from the skin of a pig
is relatively small and is due to more rapid evapora-
tion of tissue water rather than to sweating.
17.5.3 Summary
So far as can be judged by anatomic criteria the
pig should be a suitable experimental subject from
which to derive certain types of information regard-
ing the effects of heat on human skin. Its various
layers are of comparable thickness and structure. Its
blood vessels are similar in size, number, and distri-
bution. As will he show n later in Sections I7.(i and
17.7, its susceptibility and reactions to control epi-
sodes of hyperthermia are remarkably similar.
Since a pig does not sweat, allowance should be
made for the inability of porcine skin to lose heat
through the vaporization of moisture derived from
sweating. The significance of heat loss through vapor-
ization of moisture in respect to cutaneous burning
will be discussed in greater detail in Section 17.9.
17.6 RECIPROCAL RELATIONSHIPS OF
TIME AND TEMPERATURE*
The most direct mechanism by which exposure of
the body surface to excessive heat results in injury” is
the transfer of heat energy to the skin at so rapid a
rate that its temperature is raised to a level incom-
patible with cellular survival. Such localized thermal
injuries are commonly referred to as burns. Although
it is common knowledge that there is an inverse re-
lationship between temperature and the amount of
time required to produce a burn, there is remarkably
little precise information regarding the rate at which
burning occurs at any given temperature.
Because of the experimental difficulties inherent in
the making of accurate measurements of either the
time or the temperature characteristics of thermal
exposures so intense that they are capable of burning
the skin in a fraction of a second, it was decided to
establish by experimentation the reciprocal relation-
ships of time and temperature necessary to destroy
cells at lower temperatures and to extrapolate from
these data the time curve that should represent the
minimum cell-destroying exposures for higher de-
grees of temperature.
17.6.1 Method of Controlling Surface
Temperature
Direct exposure of the surface of the skin to a
rapidly flowing stream of hot liquid was chosen as
the method best adapted for the acquisition of these
data. With this type of exposure, the surface of the
skin could he maintained at the temperature desired
without the establishment of an appreciable gradient
between it and the source of heat. There was no in-
sulation of the surface by a static layer of gas, liquid,
or solid, no heat loss through vaporization of surface
moisture, and no diminution of subsurface heat con-
duction due to vascular occlusion by the application
of pressure on the surface. The method was simple to
.operate and led to remarkably reproducible, cut anc-
ons effects.
The applicator by which a running st ream of hot
water was brought into direct contact with the skin
consisted of a metal cup, the brim of which was cov-
ered with a pad of closed-cell sponge rubber to insure
a watertight contact. By means of an electric pump,
water was circulated from a large constant-temper-
ature reservoir through the cup, the open end of
which was applied to the skin. The rate of flow was
regulated by a screw clamp on the inlet, tube and by
the height of t he outlet tube (see Figure 12).
Fine he 12. Drawing of hot water applicator.
Tangential flow of a liquid produced no vertical
component of force and thus no vertical pressure.
Vertical water pressure w ithin the cup could be var-
ied between 70 and 8(i cm of mercury by suitable
adjustments of the aperture of the inlet and the
height of the outlet tubes. A copper-eonstantan ther-
mocouple measured the temperature of the water
flowing next to the skin. During any period of ex-
posure the temperature of the water flowing over the
skin could lie controlled to within 0.1 ('.
SECRET 330
STUDIES OF TURK MAE INJURY — CUTANEOUS AND SYSTEMIC
Two methods were used to equilibrate the appara-
tus Ixffore applying it to the skin. In one, the appa-
ratus was applied to a block of linoleum, adjusted to
the desired pressure, and transferred to the skin site
to be exposed as soon as the temperature equilibrium
was reached. In the other, the applicator was allowed
to remain immersed in the hot water reservoir with
the pump turned on until thermal equilibrium was
established. The cup was then transferred immedi-
ately to the skin and adjusted to the desired water
pressure.
Provision was made in the construction of this ap-
paratus for studying the relation of the size of the
area of exposure to the intensity of the resultant in-
jury, This was accomplished by making the brim of
the cup removable so that the area of skin to be ex-
postal could be varied according to the aperture size
trf the brim selected for use. Thus, in the same region
on the same animal and under identical conditions of
time, temperature, and pressure, circular targets
having a diameter of either 7 or 25 mm could be
exposed.
Individual bums in the animal experiments were
25 mm in diameter. This was larger than desirable
for human subjects and the diameter of the aperture
of the cup was accordingly reduced to 7 mm for the
human experiments. Before this was done, however,
it was established by animal experimentation that
the reduction in the size of the exposure area did not
make any appreciable difference in the effect of such
exposures on the epidermis.
Water was employed as the source of heat in all of
the experiments summarized in Table 10. Because the
question was raised as to whether or not a hypotonic
fluid such as water might modify the effects of heat,
a series of comparable exposures were made in which
oil was substituted for water. There was no appre-
ciable difference between the injury-producing po-
tentiality of rapidly flowing streams of water and oil
on either animal or human skin so long as the temper-
ature and duration of exposure were the same.
IT.6.2 Experiments on Pigs
The primary purpose of this investigation was to
obtain information relating to the tolerance of hu-
man skin to episodes of hyperthermia of varying
duration and of varying degrees of intensity, and the
direct approach would have l>een to make all experi-
ments on human subjects. For various reasons, this
was not feasible, and it was decided to acquire the
basic data from experiments on pigs. From an ox-
tensive series of observations on pigs, it was thought
that a relatively small number of critical exposures
of human skin would establish the extent to which
the more comprehensive animal data were applicable
to man.
Closely clipped young (8 to 12 kg) white pigs were
used. It was found that different portions of the body
surface of the pig vary slightly in respect to their
susceptibility to thermal injury. The largest uni-
formly reacting area was the lateral body surface be-
ginning in front of the thighs and extending forward
over the shoulders. The skin of the neck and for
about 10 cm to either side of the spine had a slightly
higher thermal tolerance than ( hat of the lateral body
surface. The skin covering the thighs, the buttocks,
the inguinal folds, and the mid-portion of the chest
and abdomen had a slightly lower thermal tolerance.
Results of experiments on pigs: The surface tem-
pera! ure, duration, and results of 179 hot water appli-
cations to the lateral body surface of young white
pigs are summarized in Table 10.
The surface temperatures at which these exposures
were made ranged between 11 ami 100 C. The dura-
tion of exposures varied between I second and 7
hours. The majority of the exposed sites were kept
under observation until the reaction had subsided or
the lesion had healed. In the ease of borderline re-
actions duplicate exposures were made and excised
at the end of 24 or 48 hours for microscopic study.
As indicated in Table 10, a wide variety of reac-
tions were observed. These ranged in severity from
evanescent erythema to deep ulcers.
In the beginning certain difficulties were encoun-
tered in recognizing differences in the severity of cer-
tain lesions. Although there was no difficulty in recog-
nizing the difference between a reaction whose total
effect was a mild and transient erythema and one
that led to deep coagulative necrosis, it was not al-
ways easy to recognize by clinical observations
whether a given lesion represented a severe first-
degree reaction with incomplete or focal epidermal
destruction or a relatively mild second-degree re-
action in which the epidermal destruction was com-
plete.
Apart from the microscopic appearance, the most
reliable criteria by which to recognize (ransepidermal
necrosis were (I) the ease with which dead but still
intact epidermis could be displaced by friction on the
second and third days after exposure, and (2) the de-
velopment of complete encrustation of such a lesion
within a week.
SECRET RECIPROCAL RELATIONSHIPS OF TIME VM) TEMPER ATI’RE
Taiii.k
10. Time-surface temjierature thresholds for thermal
injury of porcine skin.
Threshold
Threshold
Suhthreshold
and supra-
Subthresbold
and supra-
oxjH)sures
t hreshold
exposures
threshold
__
exposures
exposures
1 reactions
2 and 3°
1" reactions
2"and 3'
react ions
reactions
Focal
Complete
Focal
Complete
Hyperemia
epidermal
epidennal
Hyperemia epidermal
epidermal
Xo.
only
necrosis
necrosis
Xo.
only necrosis
necrosis
Trill)
Time
of
Seal- Small
Red Pale
Temp
Time
of
Seal- Small
Red
Pale
('
Min See csp(
Mild Severe
ins ulcers
burn burn
C
Min
See expt
Mild Severe ing ulcers
bum
burn
-14
420
1
+
j
52
30
1
+
45
130
1
+
/
1
2
45
1
+
ISO
I
30
1
1
1
+
46
45
1
+
3
00
+
'.<0
1
+
53
20
1
+
30
45
+ ,
+
40.5
45
00
1
1
+
+
1
2
2
+
+
+
47
35
45
1
1
+
+
1
2
30
3
1
, .
50
fiO
I
+
-f
54
15
1
+
—
25
35
48
10
3
+
1
1
+
42
14
1
2
+
55
5
1
1
1
+
14
15
1
2
+
+
10
15
+
F
10
IS
1
1
+
20
25
1
I
+
+
20
_ 1
' '
+
30
3
F
4!)
3
+
56
10
1
+
+
4
5
+
15
1
5
2
•
20
1
+
0
5
+
58
5
1
+
0
2
+
10
1
+
0
2
+
60
2
1
+
7
7
7
8
8
8
0
2
1
1
4
1
2
11
+
+
+
+
+
©
+
10
2
3
5
7
7
10
1
1
1
1
2
1
F
+
+
+
+
+
+
10
5
+
65
1
+
50
1
2
i
i
+
+
2
3
1
1 _
+
+
4
i
+
+
10
1
+
5
i
70
1
2
+
5
3
+
-—_
2
1
+
5
5
2
2
+
3
2
+
75
0 2
0 30 2
+
+
1
5
1
1
+
4-
51
4
1
» 2
2
+
+
'
80
i
5
I
1
+
+
1 30 2
+
So
1
1
+
2
I
+
+
5
1
+
3
2
90
i
1
■f
3
3
2
2
+
+
5
1
+
4
5
2
I
+
+
95
i
3
1
1
+
+
5
1
+
100
1
1
+
10
2
+
3
1
+
SKCRET 332
STUDIES OF THERMAL INJURY—CUTANEOUS AND SYSTEMIC
Figure 13. Photograph of right and left sides of pig with temperature and duration of each exposure indicated. Lesions
on right side were 24 hours old and those on left side 7 days old.
All exposures sufficient to cause vascular reaction
but insufficient to destroy the full thickness of the
epidermis throughout Ihe entire target area were
designated as subthreshold. The entire range of cu-
taneous responses to subthreshold exposures were
characterized as first-degree reactions. The shortest
time at any given temperature that was capable of
causing transepidermal necrosis constituted a thresh-
old exposure. The effect of a threshold exposure on
the skin was characterized as a second-degree re-
action. All exposures which were of longer duration
or higher temperature than was necessary to cause
complete epidermal destruction were designated as
suprathreshold and their effects as third-degree re-
act ions.
The macroscopic appearance of different degrees of
cutaneous reaction to hyperthermia may he seen in
(lie photographs of the right and left sides of pig 924
shown in Figaro 13. At tbo time t ho photographs
wore made, the lesions on (ho right sido wore 21 hours
old and those on the loft were 7 days old. It is ap-
parent from those photographs that the duration of
exposure at any given temperature was remarkably
critical in relation to the kind of reaction evoked. It
is equally apparent (hat the time required to produce
a given degree of reaction varied inversely with the
temperature.
it.6.3 Experiments on Human Subjects
In order lo determine the extent lo which the re-
ciprocal relationships of time and temperature in the
production of cutaneous hums in pigs were applicable
to human skin, a series of exposures similar to those
described on pigs were made on human subjects.
Some were made to the skin of the anterior thoracic
region and others on the ventral aspect of the fore-
SECRKT RECIPROCAL RELATIONSHIPS OF TIME \ ND TEMPER ATL’HE
333
arm. The applications were made with the apparatus
shown in Figure 12.
As in the case of the pig experiments, three de-
grees of skin reaction were observed. Reactions char-
acterized as first-degree were those that fell short of
complete destruction of the epidermis. At one ex-
treme a first-degree, reaction consisted of a faint and
transient erythema. At the other, extreme erythema
was severe and prolonged and miliary vesicles formed
but failed to coalesce. Lesions in which there was
complete destruction of the epidermis over the entire
target area w ere designated second- or third-degree
reactions, depending on the extent to which the
dermis was involved. As in the ease of tfie pig, a
threshold exposure represented the shortest time at
any given temperature that caused complete de-
struction of the epidermis. ~-
That a given exposure of human skin had resulted
in transepidermal necrosis was usually but not al-
ways recognized by complete vesication of the target
area. Although vesication resulting from heat indi-
cates t hat the full thickness of the epidermis has been
destroyed, absence of vesication does not necessarily
indicate that the epidermis has escaped complete
destruction. Transepidermal necrosis without vesica-
tion was observed after certain suprathreshold ex-
posures. The explanation of this phenomenon w ill Ire
discussed in Section 17.8.8.
The results of the human experiments have been
summarized in Table 11.
17.6.1 Relative Vulnerability of Porcine
and Human Skin to Thermal Injury
To facilitate comparison of the data included in
Tables 10 and 11, certain ol the more critical observa-
tions have been depicted graphically in Figure 14.
The solid line w as established by points representing
the lime and temperature of exposures that caused
minimal second-degree reactions of porcine skin. The
points by which this line was established are repre-
sented by crosses. Each cross represents the shortest
time at (he temperature indicated that resulted in
transepidermal necrosis of the entire target area. The
more that (lie time of any given exposure placed it to
the right or that the temperature of any given ex-
posure placed it above the solid line, the greater the
depth to w hich the skin was dest royed. All exposures
t hat were situated a significant distance above and to
the right of the solid line were suprathreshold and all
those situated a significant distance below and to the
Table
11. Time-surface temperature thresholds for
thermal injury of human skin.
Threshold
Sub-
and supra-
—
threshold
1 hrcshold
exposures
exposures
1 reac-
2° and 3°
Term
lions
reactions
at
Duration
Hyperemia
sur-
of
without
Complete
face
exposure
loss of
epidermal Sub-
Xo.
C
Hr Min Sec
epidermis
necrosis
ject
Date
1
14
5
+
BF
270
2*
5
+
BK
2 23
3
0 ..
+
BF
2/0
4*
0 ..
4-
BF
2/23
5*
45
2
+
+
KL
2/16
«•
3 ..
KL
2/3
7
T-»
3
+
HA
2 4
8*
47
.. 18 ..
.... —
f
UKt
2/13
9*
.. 20 ..
+
+
KL
2 25
10*
.. 20 ..
4"
AM
2 26
n*
.. 20
+
Ft;
2 20
12
.. 25
+
UKt
1 78
13*
40
_p
AM
2/26
14
.. 40 ..
+
PC
2/26
15
rr-
45
+
UKt
1/8
10
48
15
+
PG
7/19
17
15
+
AH
7/19
18
.. IS ..
4"
AM
6 20
19*
49
8 ..
+
AM
2/10
20
8
+
AM
0/26
21
9 30
4
AM
6/26
22*
.. 10 ..
. ±..
AM
0/20
23
.. 11 ..
+
AM
0/26
24
15
4-
AM
6/26
25
51
2 ..
+
AM
6/26
20
4 ..
+
AM
6 20
27
fi
+
AM
6/26
28
53
.. .. 30
-F
AM
6/26
29
1 30
+
AM
6/26
30
55
+
PG
7/19
31
.. .. 30
+
AH
7/19
32*
00
.. .. 3
+
FH
2/1
33*
+
FH
2/1
* Oil list'd instead of water as source of heat.
t Subject RK was atypical in
that his threshold for thermal injury was
significantly lower than that of other experimental subjects.
left of the solid line were subthreshold. The range of
variation is shown in Table 10.
The extent to which the results of human exposure
corresponded with those of the more comprehensive
animal experiments is indicated by the open and solid
circles in Figure 14. The open circles represent the
maximum exposure at the temperature indicated
that failed to destroy human epidermis and the
closed circles represent the minimum time at the
temperature indicated that resulted in complete
destruction of human epidermis.
The broken line in Figure 14 represents the ap-
proximate threshold at which the first morphological
SECRET STUDIES OF THRUM A L lAJl'RY— CUTANEOUS AM) SYSTEMIC
of epidermal injury 12 as determined I»y histological
examination is produced. T, is the temperature in ('
at the time, t. at the basal epidermal layer; A* is the
gas constant and is equal to 2 calories per C per mole;
and both .1 and AE are constants evaluated from the
experimental data.
Kquation (7) can also be expressed as an integral
equation, namely
i
12 . .4 fe-W*T‘+™\U (8)
0
where if 7’,, the dependence of the basal epidermal
temperature on time, is known the integral can be
evaluated.
In all eases where the temperature of the basal
layer of epidermis can be considered as independent
of time of heat exposure,-equalion (8) can l>e inte-
grated to equation (9).
12 = Ae ~ *K/KtT *■ 2T3)t. _ (9)
An examination of the transepidermal threshold
data depicted in Figure 14 and the epidermal time-
temperature data given in Table 7 (Section 17.3.2)
and illust rated in Figure 15 shows that equation (9)
is applicable in all heat exposures where the skin
surface temperature is less than 50 C; furthermore
the skiti surface temperature can be substituted for
the steady-state basal epidermal temperature since
the differences (<0.3 C) between these two values
are negligible in this temperature range.
Thus, by using equation (9) in this temperature
range, it is possible to evaluate numerically A and
\E by standard graphical procedures from the data
for the threshold of complete transepidermal necro-
sis; and the following equations are obtained,
IE » 150,000 cal/mole (10)
and
A = 3.1 X 10” sec"1. (11)
This value of A depends upon the arbitrary choice
of the value of unity for 42. Thus, when the threshold
of complete epidermal necrosis is reached,
it s 1. (12)
By again making use of equation (9) a similar
analysis can be made of the time-temperature rela-
tionship depicted by the broken line of Figure I 1.
Since these data arc not so complete as those used
above, it Is best to use the same numerical yahies
given by equations (10) and (11) for \E and A, and
solve for the numerical value of 42.
These data are found to be best represented by
U = 0.53 (13)
1’ira RE 14. Graph showing thresholds for porcine skin
at wliich microscopic evidence of spidermal injury is
first apparent tbroken line) and at which transcpidermal
necrosis is complete (solid line). Crosses indicate criti-
cal individual experiments and show shortest time at
temperature indicated at which transcpidermal necrosis
of entire target area occurred. ()|>en and solid cir-
cles show effects of heal on human skin.- Open circles
represent longest exposure at temperature indicated
that failed to destroy epidermis. Solid circles repre-
sent shortest exposure at temperature indicated that re-
sulted in transcpidermal necrosis.
evidence of thermal damage to porcine epidermis was
recognized. Exposures sit uated below t he broken line
caused no appreciable change in the microscopic ap-
pearance of the epidermis. Exposures lying between
the broken and solid lines resulted in varying de-
grees of epidermal damage short of transepidermal
neci osis. Since the reactions of human skin to con-
trol episodes of hyperthermia were not examined
microscopically, no inferences can be drawn as to the
reciprocal relations of time and temperature at
which microscopic evidence of injury to human epi-
dermis was first recognizable.
lT.6.3 Mathematical Predictability of
Epidermal Destruction by Exposure to Ileal ’
From a kinetic standpoint, the reciprocal relation-
ships of time and temperature in the production of
transepidermal necrosis follow the general pattern of
rate processes. If the reaction leading to thermal
death of epithelium conforms to t hat of most physical
and chemical rate processes,*" it should be quanti-
tatively predictable by the following equation:
4 -AE/lf(T, 1-273) (7)
(It ~ K
where dil dt is the rate at which an arbitrary function
J By F. C. Henriques, Jr.
SECRET RECIPROCAL RELATIONSHIPS OF TIME AND TEAIDER ATL'HE
335
when the upper limit of exposure which can be toler-
ated without the occurrence of transepidermal ne-
crosis is reached.
Although the values given by equations (.10) to
(13) for ,1, AE, and ft were obtained through the use
of equation (9), which requires that the epidermal
temperature can be considered constant during the
entire heat exposure, those numerical values should
permit the computation of the two thresholds of
transepidermal injury under all conditions by means
of equation (8), so long as T, is known.
Under the experimental conditions that, the data
depicted in Figure 14 were obtained, namely, con-
stant skin surface temperature during the entire heat
exposure, it is possible to ascertain the time de-
pendence of basal epidermal temperature by means
of equation (tie). Referring to equation (6e), it is
found that the evaluation of T, depends upon two
parameters, T , and y. An examination of the equa-
tion show s that T, is very insensitive to variations in
T„, the original epidermal temperature; 35 (’ is taken
as the original skin surface temperature (see Table 6
of Section 17.3.2);
In view of the uncertainties which enter into this
direct experimental evaluation of y by means of
equation (6b), it is best to evaluate it empirically by
obtaining the best fit to the complete transepidermal
necrosis data. —
It is then found that
7 = 0.15 (14)
if I, the lime during the heat exposure, is expressed
in seconds. This numerical value checks well with
that obtained by direct substitution of the experi-
mental values for the thermal conductivity, heat
capacity, density, and thickness of epidermis (see
Section 17.3.2) into equation (6b).
A consideration of equations (6a), (6c), and (8)
together with the requisite numerical values given by
equations (10), (11), and (11) shows that the experi-
mental data given in Table 10 and depicted in Fig-
ure 14 are completely described by the following
equation;
<
ft = 3.1 X 10-f.e ~ 75000 T‘+ m 0.5 and T, < 50 C the time dependence
of T, can be ignored and T, put equal to T,; equa-
tion (15) can then be integrated and takes the form
of equation (9), which greatly facilitates the compu-
tation of ft. For all T, > 50 C and ft < 1, the time
dependence of T, cannot In* neglected, and the evalu-
ation of ft by means of equation (15) requires one of
the standard methods of numerical integration.17
This numerical determination of ft from the two
experimental parameters, t and T„ permits the pre-
diction of the degree of epidermal injury, since mi
ft < 0.53 results in a time-temperature relationship
that can IxT tolerated without the occurrence of
< ransepidermal necrosis, and ft > 1.0 results in a
time-temperature relationship which produces com-
plete epidermal necrosis.
The success of equations (15) and (15a)In predict-
ing these time-temperature relationships is shown in
Table 12.
It can be seen that the agreement of the experi-
mental data of Section 17.6.3 with this equation is,
in general, excellent, and, thus, that t he tacit assump-
tion throughout this section of the applicability of
equation (7) is justified. In the four cases where there
is appreciable variance between experiment, and
prediction, either the experimental data are in-
sufficient or the duration of heat exposure was too
short to preclude considerable experimental error.
Thus, equations (15) and (15a) probably give a more
accurate prediction of epidermal injury thresholds
than the dotted and solid lines of Figure 14.
The numerical computations resulting from equa-
tion (9) are also included for comparative purposes.
For the reasons stated above there is no appreciable
difference, under these specific experimental condi-
tions, between this equation and equation (15) for all
surface temperatures below 50 (Kquation (9) cor-
responds to an experimental condition in which the
basal epidermal layer is immediately brought to and
maintained at a constant temperature. If this were
feasible at 70 (', complete epidermal necrosis would
result in 3 ten-thousandths of a second. The 2,000-
fold difference between this value and 0.5 second
predicted by equations (15) and (15a) indicate the
extreme importance of the heat capacity of t he skin
during the early period of heat exposure.
y = 0.15 (14)
SECRET 336
STL DIES OF THERMAL INJURY — CUTANEOUS VND SYSTEMIC
Tabi k 12. A comparison of the experimental time-temperature
Figure 14 with those obtained from equations (9) and (15).
relationship for (ransepidermal
injury as depicted by
Minimum time in seconds for complete
Maximum
time in seconds
for subthreshold
t ransepidermal necrosis
I ransepidermal necrosis
U se I
SI = 0.53
Experimental
Surface
Experimental
Equation
Equation
solid line
temp
dotted line
Equation
Equation
(»)*
(15)
Figure 14
C
Figure 14
(15)
(9)*
23,000
23,000
25,000
44
I8,000t
12,000
12,000
11,000
11,000
11,000
45
7,200f
5,900
5,800
5,100
5,200
5,000
46
3,000
2,800
2,700
2,400
2,500
2,400
47
1,300
1,380
1,300
1,100
1,200
1,100
48
560
6.50
t»0
580
630
570
49
260
340
310
270
325
300
50
130
165
110
130
165
160
51
75
90
68
65
91
90
52
44
52
35
is
31
35
54
IS
19
8
4.4
13
16
56
8.3
8.1
23
0.25
3.0
5
60
2.6
2.3
0.13
0.009
1.0
2t
65
1.0
0.7
0.005
0.0003
0.5
It
70
0.4
0.0002
♦ Above 50 C equation (9) has no experimental siunifieanca?.
t Experimental value
uncertain.
These tabulated values, resulting from the solution
of equation (15), are of course only valid under spe-
cific experimental conditions, namely, only when the
skin surface temperature is immediately brought to
and maintained at a constant value during the entire
heat exposure. However, equation (15) should ac-
curately predict the epidermal injury to l>e expected
from all conceivable kinds of heat exposures, so long
as the temperature dependence of the skin surface
temperature as a function of time is known, since the
time-temperature relationship at the basal epidermal
layer can be predicted quite accurately by making
use of the “infinite body” heat theory u implicit in
equation (15a). (See Section 17.3.1.)
1T.6.6 Vulnerability of Ischemic Skin to
Thermal Injury
One of the reasons that a running stream of hot
water was employed as the source of heat in these ex-
periments was that by this technique there would be
no mechanical interference with the circulation of
blood through the dermal capillaries. The exposures
were made at atmospheric pressure. It was felt, that
circulation of relatively cool blood through the der-
mal capillaries would probably tend to protect the
skin against burning and that any method employed
for the production of uniform burns should be one
which did not cause mechanical interference with
capillary circulation. —
The following experiments were undertaken for the
purpose of determining the extent to which local im-
pairment in blood flow may increase the vulnerability
of the epidermis to thermal injury.
A control series of burns were made on each of
three pigs by exposing various skin sites to hot water
at atmospheric pressure. The predetermined time and
temperature of each exposure were such that severe
first-degree or mild second-degree reactions could he
anticipated. See Table 13.
Tablk 13. Effects of thermal exposures with and
pressure ischemia.
without
Ani-
mal
num-
ber
Kxpt
temp
C
Expo-
sure
dura-
tion
(min)
Excess
pres-
sure
on skin
(mm I Ik )
Xo. of
exp
made
Number of lesions
Without With
transepi- traasepi-
dcrmal dermal
necrosis necrosis
887
40
7
0
5
5
0
40
9
0
5
0
5
49
7
80
5
5
0
899
49
7
0
4
4
0
49
8
0
4
2
2
40
9
0
4
0
4
40
7
80
4
4
0
49
8
80
4
3
1
!K)1
51
2
0
3
3
0
51
3
0
3
2
1
51
4
0
3
0
3
51
2
80
3
3
0
51
3
so
3
I
2
It was found that all 7-minute exposures at 49 C
and all 2-minute exposures at 51 C made at atmos-
pheric pressure were subthreshold in the sense that
SECRET REC1PROC A1, RELATIONSHIPS OF TIME WO TEMPERATURE
337
they failed to cause complete transepidermal ne-
crosis. That they were close to threshold was indi-
cated by the fact that all 9-minute exposures at 4!) ('
and all 1-minute exposures at 51 C did cause (rans-
epidcrmal necrosis.
After it was established that the position of the
threshold for transepidermal necrosis in these ani-
mals was between 7 and 9 minutes at 19 (' and lx-
t ween 2 and 4 minutes at 51 C for exposures made at
atmospheric pressure, a second series of exposures
were now made during which the water pressure was
increased by an amount corresponding to 80 mm of
mercury. With this amount of pressure on the sur-
face of the skin during the time that it was exposed
to heat, there was no instance in which the reaction
to a 7-minute exposure at 19 C or to a 2-minute ex-
posure at 51 (' was increased in severity.
It is apparent from the data summarized in
Table 13 that the application of pressure sufficient
to collapse superficial dermal capillaries during a
period of exposure does not cause appreciable aug-
mentation in the vulnerability of epidermis to ther-
mal injury.
In view of the extreme thinness of the epidermis,
these results-were to be expected, since, for reasons
given in Section 17.3.2, the epidermal temperature is
primarily determined by the skin surface tempera-
ture. Thus the dermal temperature gradients, which
may be appreciably altered in ischemic as compared
with normal skin during thermal exposure, would
have little effect on the time-temperature relation-
ship that exists at the epidermal-dermal interface.
17.6.7 Latent Thermal Injury and Cumulative
Effects of Repeated Subthreshold Exposures
If the data summarized graphically in Figure 1 I
are recalled it is apparent that recognizable damage
to the epidermis occurred only during (he terminal
phase of the subthreshold exposures represented in
these experiments. Xot until the duration of any
given episode of hyperthermia was such as to bring
it. to the level indicated by the interrupted line in
Figure 14 was there recognizable evidence of epi-
dermal injury. This phenomenon is even more readily
apparent in the photographs shown in Figure 13.
In these, it may lie seen that the 7-minute exposure
at 49 C on the left side of the animal shows only a
trace of residual erythema, whereas both of the sites
of 9-rninute exposures at that temperature show
t ransepidermal necrosis.
Does this indicate that no epidermal injury had
boon sustained during (lie first 7 minutes, or does it.
mean that injury was present hut morphologically
latent ?
In order to gain more information concerning this
point, the experiments summarized in Table 11 were
undertaken. Thermal exposures were made with a
running stream of hot water at 19 C and at atmos-
pheric pressure. Three young pigs were used.
The first series of exposures (1 to IS) were for con-
trol purposes and served to establish the reproduci-
bility of reactions to single exposures at this temper-
ature. It may be seen that there was not a single in-
stance in which an exposure for less than 7 minutes
caused recognizable necrosis of the epidermis and
that in every instance in which exposures as long as
9 minutes were given, there was complete necrosis of
the epidermis. Skin sites receiving 7-minute expo-
sures recovered without loss of epidermis, whereas
skin sites receiving 9-minute exposures underwent
complete ulceration.
The control exposures were followed by a series
(19 to 39) in which repeated exposures, individually
incapable of causing recognizable epidermal injury,
were applied to the same area. It was found, for in-
stance, that, although a single 3-minute exposure at
19 C caused no recognizable change in the epithelial
cells, three such exposures separated by recovery
periods as long as 21 minutes had the same total de-
structive capacity as a single continuous 9-ininutc
exposure.
It was clear that a certain amount of epidermal in-
jury was sustained during the first 3 minutes and
that at least 21 minutes were required before there
was an appreciable recovery from this injury. That
complete recovery occurred after between 2 and
f hours was indicated by experiments 30 and 31.
Experiments 31 to 39 showed what might have
been expected, namely, that recovery from the latent
injury of a 2-minute exposure was more rapid and
that from a 5-minute exposure less rapid than was
the case after a 3 minute exposure.
Further discussion of the implications of these ex-
perimental results will be found in Section 17.8 of
this chapter.
17.6.8 Summary
The reciprocal relationships of time and temper-
ature in the causation of transepidermal necrosis are
similar for similar skin areas of man and pig.
Thermal injury or burning of the skin was ob-
served to occur at surface temperatures as low as
SECRET 338
STUDIES OF THERMAL INJURY — CUTANEOUS VND SYSTEMIC
Table 14. The cumulative effects of repeated suhthreshold thermal exposure
on the skin
of the pig.
All exposures were made to water at 4!) C.
Effect of exposure on skin
No evidence of
Duration
epidermal injury
Epidermal
necrosis
of each
No. of
Interval
Mild
Severe
Complete
exposure
exposures at
1 ict ween
vase.
vase.
and
Expt
(min)
same site
exposures
react ion
reaction
Focal
irreversible
No.
3
1
+
1
.. —
1
+
2
1
+
3
4
1
+
4
5
1
+
5—
6
1
+
6
1
+•
7
1
+
* .
8
. 7
1
+
.-r ■
9
1
+
10
8
1
+
11 -
I
12
I
. . -
+
13
9
1
+
14
1
. . —
+
15
1
+
16
1
+■
17
1
+
IS
3
3
3 min
+
19
3
3 min
f
20
3
3 min
.
+
21
3
3
6 min
+
22
3
3
12 min
r.
+
23
3
3
24 min
+
24
3
3
48 min
+
25
3
48 min
+
26
3
3
72 min
. .
+
27
3
72 min
+
28
3
3
96 min
- +
29
3
3
120 min
+
30
3
3
240 min
+
31
3
3
24 hr
+
32
3
3
48 hr
+
33
2
5
2 min
. .
+
34
5
30 min
+
35
5
60 min
+
36
3
2
12 min
+
37
5
2
60 min
38
-
2
240 min
+
39
14 C and it, can be inferred from the shape of the
lime-temperature curve that the threshold at which
hyperihernial cellular injury is first sustained is not
far above the level that is normal for the blood.
The rate at which injury occurs increases rapidly
if the temperature is raised. The progress of injury
at any given temperature is determined by the dura-
tion of the hyperthermic episode. Thus, the amount
of time required to convert a reversible into an irre-
versible cellular injury is different for each degree of
temperature and in the case of the epidermis can be
computed if the surface temperature as a function of
time is known.
The existence of latent or morphologically unrecog-
nizable cellular injury after certain apparently harm-
less thermal exposures and the fact that the time re-
quired for recovery from such latent injuries becomes
longer the nearer they approach the threshold of
microscopic visibility were demonstrated experi-
mentally.
t7.7 PATHOLOGY OF CUTANEOUS
BURNS AND THEIR PATHOGENESIS
In the foregoing section, measurements of the re-
ciprocal relationships of time and surface tempera-
ture with respect to the capacity of thermal exposures
SECRET PATHOLOGY OF CUTANEOUS BIHXS AM) THEIR PATHOGENESIS
339
to destroy the epidermis were reported. The following
studies concern the pathological characteristics of
cutaneous injuries caused by thermal exposures at
different temperatures and for different durations,
and a comparison of the pathogenesis of cutaneous
burns in man and pig.
17.7.1 Experimental Procedure
Much of the material used in this investigation
was derived from the experiments described in Sec-
tion 17.6 of this chapter. It was added to from sev-
eral sources (see Table 15)7 Since most of the lesions
produced in the experiments reported in Sect ion 17.6
were not excised until they had been under clinical
observation for days or weeks, many duplicate ex-
posures were made and excised in order to observe in
(lie various types of lesions the sequence of micro-
scopic changes that took place between injury and
repair. To~aequire this material, approximately 60
additional hot water exposures of pigs were made and
examined microscopically after recovery periods
ranging between a few seconds and several weeks.
Additional material comprised a series of bums of
porcine skin made by exposure to hot air at temper-
atures varying between SO and 1)00 ('. There were
( wo series of human burns, one comprising 33 experi-
mentally produced lesions which were studied clini-
cally but were not excised for microscopic examina-
tion, and the other comprising a collection of skin
specimens obtained after death from victims of con-
flagrations.
Sections of tissue for microscopic examination were
cut from specimens that had been fixed in Zenker-
formol or JO per cent formaldehyde. Phloxine-
methylcne blue stains were made routinely and were
augmented by sections stained with hemotoxylin and
eosin or by Poliak’s modification of Masson's tri-
chrome method. Many sections were stained by (he
Feulgen technique.
IT.7.2 (General Consideration of Quantitative
and Qualitative Effects of Ileal on Skin
A cutaneous injury caused by hyperthermia may
be characb rized quantitatively according to the
depth to which (he tissue has been destroyed, or qual-
itatively according to the nature of the changes that
have occurred. The characterization in Section 17.0
of hyperthermic episodes as subthreshold, threshold,
and suprathreshold referred to their quantitative
capacities for injury production, the determining
factor being the capacity of the exposure to cause
complete destruction of the epidermis.
Thus, any exposure that failed to cause complete
destruction of the epidermis was designated as sub-
threshold and any reaction short of t ransepidermal
necrosis was one of the first-degree. A second-degree
reaction was one caused by the shortest exposure at
any given temperature, or by the lowest temperature
at any given time, that resulted in full-thickness'de-
struction of the epidermis. Although it was not pos-
sible to destroy the entire thickness of the epidermis
without some damage to the underlying connective
tissue, dermal necrosis was a relatively insignificant
feature of a truly threshold exposure. A third-degree
reaction was one caused by an exposure that was
suprathreshold in respect to time or temperature
and was accordingly one in which a significant degree
of dermal necrosis usually accompanied the destruc-
tion of t he epidermis.
Slope of Transcutaneous Temperature Gradi-
ent in Relation to Depth and Quality of Burn
If account is taken of the potential variations in
the intensity and duration of the different thermal
exposures that are capable of producing burns of
similar depth, if becomes apparent why thermal
lesions of approximately the same depth may be
qualitat ively dissimilar.
This fact is more readily appreciated by reference
to Figures I a and lt>. The critical temperature, so far
as the ultimate fate of the epidermis is concerned, is
that attained at the interface between epidermis and
dermis rather than that of the surface. In Figure 15
are shown the estimated changes" in temperature
that would occur at the basal cell level during the
course of thermal exposures at three different surface
temperatures if each were terminated at a time calcu-
lated to be just adequate to destroy the epidermis. In
Figure 16 are shown the temperatures that would
Table 1 5. Sources and kinds of material used for study
of the pathogenesis of cutaneous burns.
Sub-
ject
Source
of
heat
flange
of
temp
C
Hangeof
exjxtsnre
llange of
recovery
period
Pin
Water
44-100
0.5 sec -7.5 hr
1 min-1 weeks
PiR
Air
SO-000
0.5 min-45 min lmin-3davs
Man*
Water
44- 00
3 sec 0 hr
1 min 4 weeks
or oil
Man
Air
7
?
Less than 1 hr
• f-rsions not excised for microscopic study.
"Calculations based on data presented in Section 17.3. 340
STUDIES OK THERMAL INJURY—CUTANEOUS AND SYSTEMIC
would extend to, hut not far beyond, the basal coll
layer. That qualitative differences in the resulting
reactions might exist despite their quantitative simi-
larity can be inferred from the fact that in the ex-
posure shown in Figure ISA the epidermis was de-
stroyed by a 3-hour episode of hyperthermia the in-
tensity of which at no time rose above 14.8 t’ at the
basal cell level. Approximately the same amount of
irreversible change would l>e sustained as the result
of the exposure depicted in lot'. In I lie latter in-
stance, the epidermis was destroyed in approxi-
mately 0.1 second by an episode of hyperthermia in
which the temperature at the basal cell level rose
sharply and briefly to 70 C. The exposure depicted
in I5B falls about midway between these extremes.
Although the total amount of irreversible injury is
about the same in each, it is not surprising that tlie
three lesions produced in (his manner differed quali-
tatively.
Since certain qualitative characteristics of thermal
reactions arc dependent on the degree to which the
temperature of the tissue has been raised, it follows
that the longer any given episode of tissue hyper-
thermia is maintained, the greater the likelihood that
the qualitative attributes of the reaction will reflect
the intensity of the exposure.
Such was found to be true: The more severe t he ex-
posure, the greater were the qualitative differences
between the reactions produced at high and low sur-
face temperatures.
An additional reason for the occurrence of quali-
tative differences in quantitatively similar reactions
to thermal exposures of different intensity is shown
in Figure Ki. There are depicted the calculated trans-
cutaneous thermal gradients to a depth of 2 mm that
would exist at the moment of completion of the same
three episodes of hyperthermia illustrated in Fig-
ure lo. In each instance, irreversible thermal injury
would extend to, but not appreciably beyond, the
basal cell layer. In the exposure depicted in A (Fig-
ure 10), the temperature of the dermis to a depth of
about 2 rnm was elevated above normal for at least
2 hours. In the exposure depicted in (', the transcu-
taneous thermal gradient, was so steep that the re-
sulting temperature changes in the dermis were ex-
ceedingly brief and superficial. It is apparent why the
epithelial cells would be destroyed in C with rela-
tively little disturbance of the dermis, whereas, in .4,
the same or even a lesser degree of epidermal injury
might, be accompanied by a severe and persistent
vascu lar distu rbance.
Figure 15. Curves depicting changes in temperature
at interface between dermis and epidermis during sur-
face exposures of 15 (A), 55 (TV>, and 100 (C) C. Kacli
of these was threshold exposure in that 3 hours, 0.4
minute, and 0.1 second, respectively, arc estimated to
lx* shortest time at indicated temperature that would
cause transepidermal necrosis, (Estimates derived
from measurements reported in Section 17.3.2.)
Fim re 16. Solid line traversing each chart from left to
right depicts tcm|>eraturcgradient through first 2nun of
skin at conclusion of exposures estimated to Ire just
sufficient to cause transcpidermal necrosis. Inter-
rupted line traversing each chart from left to right de-
pic's original pre-exposure temperature gradient
through skin to depth of 2 nun. The 0 vertical line in
each represents surface of skin. Interrupted vertical
line at depth of approximately 0.1 min indicates depth
of dermal-epidermal interface. In A, surface tempera-
ture of 45 C had liecn maintained for 3 hours. In It,
surface temperature of 55 (' had licen maintained for
0.4 minute. In C, surface temperature had been
maintained at 100 C for 0,1 second. (Estimates de-
rived from measurements reported in Section 17.3.2.)
prevail at different depths below the surface of each
at the moment that the duration of the exposure was
just sufficient to cause irreversible injury of the en-
tire thickness of the epidermis.
In each instance, the effects would be quantita-
tively similar, in (hat irreversible cellular injury PATHOLOGY OF CU TANEOUS BURNS AND THEIR PATHOGENESIS
341
lT.7.4 First-Degree Reactions
HvCKRKMiA, Edema, and Cyanosis
Sufficient dilatation of the superficial capillaries to
cause erythema characteristically accompanied and
frequently preceded damage to the epithelium. One
exception to this generalization represented in the
material upon which (his investigation was made was
provided by the effects of heat on the skin of animals
suffering from circulatory failure. In the presence of
circulatory failure, there was frequently such a pro-
found depression of vasomotor irritability that injuri-
ous episodes of either high- or low-intensity hyper-
thermia failed to elicit vascular reactions even though
extensive epidermal injury was sustained. Other cir-
cumstances in w hich thermal damage of the epider-
mis was sustained with little or no accompanying
vascular reaction included exposures of sufficient in-
tensity to burn the stratum eorneum, but so brief as
to cause little or no rise in dermal temperature.
Attention has already l>een called to the fact that
the duration of an episode of low-intensity hyper-
thermia must be greatly prolonged if it is to produce
an injury quantitatively comparable to one resulting
from a high-intensity exposure. Since the dermal
blood vessels are far more responsive to temperature
changes than are the epithelial cells, it can be under-
stood why severe and persistent vascular reactions
were often elicited by protracted episodes of low-
intensity hyperthermia that failed to harm the epi-
dermis (see Figure 13).
There was considerably greater individual vari-
ation among human subjects than there was among
pigs in respect to the vascular reactions to cutaneous
hyperthermia. The variability of dermal vascular re-
actions in human subjects was so great and tbc num-
ber of reactions studied in this investigation was so
few that little could be inferred as to the extent to
which the animal data apply to human skin. The im-
pression was gained that the thermal stimulus neces-
sary to cause visible erythema in most human sub-
jects was substantially lower than that required to
elicit erythema in pigs. In man the change in skin
color was usually more intense and of longer duration
than that in the pig after an identical exposure.
That an active circulation of blood was maintained
through the dilated capillaries of an evanescently
erythematous skin was indicated in part by the pink
nr red color of the surface and in part by the fact that
the surface temperature during such a reaction was
characteristically between 0.5 and 1.5 degrees higher
than that of the adjacent skin.
An evanescent erythematous reaction to heat could
not as a rule lie recognized in sections prepared for
microscopic examination. Vessels, the seat of physi-
ological dilatation, usually contracted during or im-
mediately after excision, and it was difficult or im-
possible to distinguish a sample of physiologically
hypcremic skin from one that was normal or
ischemic.
If cutaneous hyperthermia was prolonged to be-
tween 40 ami 00 per cent of (he minimum time re-
quired for the production of transcpidermal necrosis
in cither man or pig, ir characteristically resulted in a
more severe and pathological vascular disturbance
which led to edema and cyanosis and which persisted
for flays rather than minutes or hours. That the How
of blood through the dilated capillaries was slowed
was Indicated by the blue or purple color of the sur-
face in cont rast to the pink or red color caused by the
moit' evanescent active hyperemia. The surface tem-
perature of such a lesion during the first few hours
was frequently found to be from 0.5 to 2 degrees be-
low that ol-thc adjacent normal skin. That the re-
action was pathological rather than physiological was
also indicated by the fact that in both man and pig it
was almost invariably accompanied by cutaneous
edema. Within the first hour after (he onset of a
vascular injury of this grade, the water content of
the dermis was observed to increase by as much as
100 per cent.
Microscopic examination of reactions of this type
at varying periods after exposure in ■ pig confirmed
the clinical observation that heat may cause a severe
disturbance of the dermal blood vessels in both pig
and man without causing recognizable damage to the
epidermis. The capillary loops of the dermal papillae
became dilated and elongated and filled with closely
packed masses oferythrocytes. Separation of collagen
fibers by edema fluid was obvious and perivascular
mantles of extravasated erythrocytes were often
seen. The escape and extravascular deterioration of
erythrocytes in such a lesion was often sufficient to
result in brown discoloration of the target area for as
long as a week. Ext ravascular fibrin was not encoun-
tered nor di«l collagen fibrils appear to be swollen.
Between 12 and 24 hours after such an injury was
sustained, occasional polymorphonuclear leucocytes
were found in the edema fluid. Neither thrombosis
nor visible alteration in the vascular endothelium
was seen, despite the fact that superficial vessels
were filled by static, sausagelike agglomerates of red
blood cells. 342
STUDIES OF THERMAL INJURY — CUTANEOUS AND SYSTEMIC
Reversible Impairment of Epidermal Antiiorau.e
During most, and possibly all, injurious episodes of
cutaneous hyperthermia in which the temperature of
the dermis was maintained for a sufficient time at
49 C or higher, there was a brief interval subjacent to
the threshold for transepidermal necrosis in which
the adhesion of epidermis to dermis was impaired.
The attainment of this degree of injury was recog-
nized by the ease with which the epidermis could l>e
dislodged by friction. If the exposure was discontin-
ued before further injury was sustained and if the
loosened epidermis was not dislodged either by
trauma or vesication, the change was often reveisible
in the case of the pig and after 12 or 18 hours the
original firm anchorage of the epidermis was usually
regained. Unless the exposure had been excessive,
such injuries subsided without further evidence of
cell death.
If skin altered in this manner was protected against
mechanical artifact, there was no microscopic evi-
dence either in the basal epithelial cells or in the un-
derlying dermis by which impairment of the epi-
dermal anchorage could be recognized. If, however,
sufficient friction was applied to the temporarily in-
secure porcine epidermis to cause its detachment,
microscopic examination revealed a fringe of up-
rooted or fractured lonofibrils protruding from the
lower ends of the detached basal epithelial cells. The
protruding fibrils appeared to have been pulled out
of their anchorage in the superficial dermal felt work
of collagen fibers. It was not determined whether the
essential change responsible for such epidermal in-
stability was a deterioration of the extracellular ex-
tensions of the tonofibrils which predisposed them to
rupture or a softening of the dermal collagen in which
they were embedded. The latter was considered the
more plausible explanation of the phenomenon. In
man it is doubtful that the tonofibrils of the epi-
dermal cells have much if anything to do with the
attachment of epidermis to dermis. In human skin,
the epidermis appeared to be cemented to, rather
than rooted in, the dermal collagen.
It has already lieen indicated that when porcine
skin sustained this type of cutaneous bum, recovery
sometimes took place within 24 hours without death
of cells, providing (he damaged area was protected
against mechanical disturbance during that period
when its anchorage to the dermis was insecure.
Too few appropriate specimens of human bums
were available for microscopic examination to permit
conclusions regarding the threshold at or the fre-
quency with which this particular type of first-degree
thermal injury occurs in man. The opinion was gained
from clinical observations of human burns that ther-
mal exposures insufficient to cause primary epider-
mal necrosis may result in a temporary impairment
in the adhesion between epidermis and dermis. If
such a temporarily insecure layer of epidermis is de-
tacher! by friction or vesication, the detached cells
would undoubtedly die. Thus, it is entirely possible
that the phenomenon of vesication results, in some
instances, in secondary destruction of human epi-
thelial cells that would otherwise survive. If this be
true, and if the thermal exposure has been insuffi-
cient to cause primary transepidermal necrosis, the
immediate institution of pressure to prevent epi-
dermal displacement by vesication should predispose
to early and uncomplicated healing of what might
otherwise become an open lesion.
Irreversible Thermal Injury of Epidermal
Cells
Material for microscopic study was available from
almost every conceivable kind, grade, and stage of
thermal injury of the skin of the pig. All hough a wide
range of experimental thermal injuries of human
skin was studied clinically, most of the burns that
were available for microscopic examination were ob-
tained from autopsies. Thus, (here was no direct in-
formation regarding the intensity or duration of the
thermal exposures that were responsible for most of
the burns of human skin that were studied micro-
scopically. The impression was gained, however,
that, apart from the phenomenon of vesication, the
cytological changes induced by heat in the epidermis
of man were similar, if not identical, to those ob-
served in (he pig (see Figures 17 to 22). Attention
lias already been directed to the fact that the time-
temperature threshold for the destruction of epi-
dermisJs almost identical in human and porcine skin
(see Figure 14).
The first manifestation of irreversible thermal in-
jury of the epidermis was a change in the distribu-
tion of water and solids within the nuclei of the cells
of the intermediate zone. As the nuclei swelled, their
chromatin granules coalesced to form compact eres-
centric masses immediately beneath and attached to
one side of the nuclear membranes (Figure 20). When
the swollen nucleus ruptured, the peripherally dis-
tributed chromatin contracted to form a dense and
irregularly shaped central mass which remained sep-
arated from the surrounding cytoplasm by clear fluid.
SECRET PATHOLOGY OF CUTANEOUS BURNS AND THEIR PATHOGENESIS
313
Figure 17
Figure 18
Photographs of severe first-degree burns of porcine (Figure 17) and human (Figure 18) skin showing degenerative
changes in In Figure 17, there is generalized pyknosis of nuclei and it is not likely that any epidermal cells
included in picture would have recovered. In Figure 18, changes are focal rather than general and most of altered nuclei
are swollen and show periphraal displacement of chromatin. This tyjx* of nuclear change precedes that shown in Figure 17.
Both specimens were excised within an hour after injury was sustained. In both instances, epidermal attachment to
dermis was insecure and lesion shown in Figure 18 would probably have gone on to vesication in normal course of events.
Magnification 100 X. _
Photographs of early second-degree burns of porcine (Figure 19) and human (Figure 20) skin showing early spontaneous
detachment, of epidermis from dermis. Vacuolar cytoplasmic disintegrat ion of basal cell layer has been added to nuclear
changes similar to those shown in Figures 17 and 18. In Figure 19, lonofibrils that were uprooted from their anchorage
in dermis can lx* seen projecting from detached basal cells. Magnification 400X.
Figure 19
Figure 20
This fluid, whether extruded into the nuclear lacuna
or contained within the distended nuclear membrane,
was faintly basophilic and sometimes contained a few
fine Feulgen-positive fragments of chromatin.
Pyknosis of nuclei was by no means pathogno-
monic of thermal injury. Spontaneous nuclear pykno-
sis was sometimes seen in the upper zone of normal
unheated epidermis and was caused by injuries other
than heat. —
In the ease of subthrcshold exposures sufficient, to
injure tin* upper layers of epidermal cells but insuffi-
cient to cause t ransepidermal necrosis, the types of
nuclear change which have 1 teen described were fre-
quently focal and difficult to distinguish from quali-
tatively similar changes in control material. Even
though it could be plausibly assumed that all of flic
cells at a given level were exposed to the same degree
of hyperthermia, il was not uncommon to find groups
of cells with normal appearing nucleuses interspersed
among those that showed advanced degenerative
change (Figure 18). The reason for this apparent dif-
ference in the susceptibility of cells in the same layer
to heat was not apparent.
If the thermal exposure was of sufficient intensity 344
STL DIES OF TIIEUMAL 1 NJIRY — CUTANEOUS AND SYSTEMIC
Photograph *if pseudo-vesicle of |H>reiue skin ( Figure 21) and early true vesicle of human skin (Figure 22). In each, trails*
epidermal necrosis apjx'ars to lie complete. In porcine skin, detached epidermis would have remained in situ as flaccid
membrane. In human skin, detached epidermis would have Iieen elevated by collection of edema fluid between it and
dermis. Magnification IOOX.
Fkjcbk 21
Fionite 22
or duration to cause irreversible cellular injury,
nuclear changes of the kinds described in the fore-
going paragraphs were usually apparent immediately
after the conclusion of the episode of hyperthermia.
This was not invariably the case, however, and after
certain exposures at relatively low temperature (un-
der 47 C) a post exposure interval of between (» and
12 hours was sometimes required for the develop-
ment of recognizable nuclear alterations. Moreover,
if the exposure was of sufficient severity to cause
pseudo vesication in the pig or I rue vesication in man,
many of the nuclei which were apparently undam-
aged at the conclusion of the exposure disintegrated
during the next 21 hours.
If the episode of hyperthermia was such as to
cause visible alterations in nuclear structure, there
was inhibition of mitotic division throughout the en-
tire area of exposure for many hours. No evidence
was derived from the microscopic study of subthresh-
old exposures to indicate that hyperthermia predis-
posed to acceleration of mitotic activity. The impres-
sion was gained that nuclear swelling with coales-
cence of chromatin granules constituted evidence of
an irreversible cellular change and invariably led to
premature death and desquamation of the altered
cells.
In the pig, the irreversibly damaged epidermal cells
were gradually desquamated over a period of a week
or 10 days in the form of thin brown scales.
Alterations in the appearance of nuclei in the upper
and intermediate layers of epithelium wen* thought,
to provide the earliest morphological evidence of
primary thermal injury of (he epidermis and were
frequently encountered will tout perceptible damage
to the cells of the basal layer, ('baracteristieally, the
earliest change in the basal cell layer caused by hy-
perthermia. was cytoplasmic rather than nuclear.
The injured basal cells swelled and their cytoplasm
became vacuolated and increasingly acidophilic
(Figures ID and 20). The vacuolization appeared to
be due in part to imbibition of fluid and in part to
redistribution of water and solids within the cells.
The fluid contained within the cytoplasmic vacu-
oles was clear, nonsudanophilic, and sometimes con-
tainer] a delicate mesh of granular amphophilic debris.
With severe* injury there was widespread rupture
and disintegration of the lower ends of t lie basal cells.
17.7.1 Second-Degree Reactions
TUANSEPIULKMAL NECROS1S
The time-temperature characteristics of exposures
just sufficient to cause transcpidermal necrosis in
both man and pig arc indicated in Figure 14. In man,
whether or not a thermal exposure has destroyed the
epidermis can usually be determined by the occur-
rence or nonoccurrence of vesication within the first
24 hours. To recognize with certainty during the first
day or two after a near-threshold exposure whether
or not porcine epidermis has been destroyed, the skin
must be excised and examined microscopically. When
the area of injury was 7 mm in diameter and when
the duration and intensity of the exposure was at or
not far above the threshold required for transepi-
dermal necrosis, the time usually required for com-
plete healing was between 5 and 10 days in the pig
and between 1 and 2 weeks in man. PATHOLOGY OF Cl TA N FOPS BURNS AND THEIR PATHOGENESIS
345
In the pig, microscopic evidence that an exposure
had been sufficient to cause transepidermal necrosis
Wits provided by the changes that had occurred at
the basal cell level. With the disintegration of the
cytoplasm of the proximal or lower ends of the in-
jured basal cells, there was at first focal and later
extensive spontaneous detachment of the epidermis
from the dermis. In the pig, the amount of fluid that
collected beneath the loosened epidermis was never
sufficient to produce true vesication.
With still more severe hyperthermia, the cyto-
plasmic disintegration of (he basal cells was followed
by nuclear changes similar to those seen in the more
superficially located cells. If the epidermal detach-
ment was incomplete, stretching and attenuation of
the remaining bridging cells and their nuclei was
often seen. Such at tenuated masses of chromatin were
often stretched to two to three times the original
length of the entire cell. —
In the event that the surface temperature of the
epidermis was brought rapidly to a level of 55 C or
higher and maintained at that level for a period
longer than that necessary to cause cell death, trans-
epidermal coagulation was likely to occur so quickly
as to inhibit recognizable redistribution of intra- and
extracellular fluids. In such an event, neither the
cytoplasm nor the nucleuses of the epithelial cells
appeared swollen (Figure 25), On microscopic ex-
amination, both appeared shrunken, the former being
intensely and uniformly acidophilic and the latter
small and homogeneously basophilic.
Vksicatiox
Attention has already been called to the fact that
a common effect of heat on the skin of both man and
pig is impairment of the attachment of the epidermis
(Figures 21 and 22) to the dermis, and the opinion
expressed that this may be due either to a change in
the physical state of the superficial dermal collagen
or to disruption of the basal layer of epithelial cells.
A common collateral effect of cutaneous hyperther-
mia, and one that is essential to tme vesication, is an
outpouring of fluid from the dermal capillaries.
When a thermal exposure of human skin was suffi-
cient to impair the attachment of the epidermis, the
amount of edema fluid that collected between it and
the dermis was usually sufficient to elevate and
stretch the entire layer of dead, dying, and living
cells and to form a vesicle. Although vesication of
human skin was usually an almost immediate re-
sponse to a thermal exposure of sufficient severity to
cause primary epidermal injury, there were several
circumstances in which it was either delayed or in-
hibited.
Delayed vesication was most frequently seen after
long-time exposures at low temperatures or after
flash exposures at high temperatures. In both cir-
cumstances it seemed likely that the delay was due
to the fact that the vascular damage was relatively
mild, and that hours rather than minutes were re-
quired for enough fluid to collect beneath the dam-
aged epidermis to form a vesicle.
Another circumstance in which vesication was de-
layed or prevented was when the injury was so over-
whelming that the dermis and its superficial capil-
laries were almost immediately coagulated. With
such-thermal injury, the level at which edema de-
veloped was too deep to result in vesication.
Thus, in man, the nonoccurrence of vesication
after a thermal exposure sufficient to cause severe
injury of the epidermis may mean that the dermal
hyperthermia was either inadequate to result in
edema or that it was so overwhelming that the super-
ficial capillaries were almost immediately occluded.
In no instance was true vesication of porcine skin
observed. This was true despite the fact that many of
the injuries met at least two prerequisites to vesicle
formation: namely, sufficient vascular injury to re-
sult in dermal edema and sufficient, impairment of
the adhesion between epidermis and dermis to per-
mit easy mechanical detachment of the former (Fig-
ure 21). Failure of the pig to vesicate appeared to be
due to the fact that the amount of edema fluid that
penetrated the surface of the dermis was never suffi-
cient to elevate the epidermis. In the absence of evi-
dence to the contrary, a tenable explanation for
non vesication in the pig is that an episode of hyper-
thermia that is sufficient to impair the attachment
of the epidermis to the dermis characteristically al-
ters either the superficial felt work of dermal collagen
fibers or the walls of the capillaries contained by it
in such a way that they become virtually imperme-
able to edema fluid.
The nature or, for that matter, the existence of
(his theoretical alteration in the permeability of the
collagen or the capillary walls was not disclosed by
microscopic examination. When the severity of an
exposure was considerably in excess ot that required
to destroy the epidermis, swelling of the superficial
dermal collagen and occlusion of its capillaries could
be recognized. There was, however, a wide range of
exposures between the threshold tor epidermal ne- 346
STUDIES OF THERMAL INJURY' CUTANEOUS AND SYSTEMIC
Figure 23
Figure 24
Photographs«f mild (Figure 23) and moderately severe (Figure 24) third-degree thermal reactions in porcine epidermis
24 hours after injury. Both injuries were produced by episodes of hyperthermia that were of low intensity (under 55 C)
and long duration. In both instances, irreversibly injured dermal tissue will undergo autolysis and organization.
Magnification 4tX)X. —
crosis and that for recognizable swelling of collagen
or occlusion of capillaries in which the microscopic
examination of the pig’s skin disclosed no explana-
tion for the failure of porcine skin to vesicate.
17.7.5 Third-Degree Reactions
The more a thermal exposure exceeded the thresh-
old required for destruction of the epidermis, either
in respect to temperature or time, the deeper the in-
jury and the longer the recovery period necessary for
repair and regenerat ion. In both pig and man, several
weeks represented the minimum healing time if a
significant degree of dermal injury had been sus-
tained.
Further Changes l\ Epidermis
In the case of the pig, prolonged exposure at a rela-
tively low surface temperature (under 55 C) caused
relatively litlle additional change in the microscopic
appearance of the epidermis. In the higher range of
surface temperatures, significant prolongation of the
rate of duration of exposure beyond the time neces-
sary lo destroy the epidermis modified the quality of
(lie superficial changes both in human and porcine
skin. In man, vesication was permanently inhibited
and in both man and pig I he loosened epidermis be-
came reattached to the damaged dermis. Early solidi-
fication and contraction, both cytoplasmic and nu-
clear, occurred before there was opportunity for the
development of the retrogressive changes observed
in first- and second-degree reactions. The higher the
temperature, the shorter the time required to cause
transepidermal coagulation. With exposures to super-
heated air, desiccation was superimposed on the
effects of heat, and, soon after the temperature rose
above 300 C, carbonization of the dry tissue began
to take place.
Red axd Pale Burns
The surface color of third-degree burns ranged
from pale gray through red, purple, and brown to
SECRET PATHOLOGY OF CUTANEOUS BURNS AND THEIR PATHOGENESIS
347
Figure 25. Third-degree thermal reaction in porcine
skin showing coagulation of epidermis and dermis 24
hours after injury. Bundles of denatured dermal colla-
gen apjH'ar swollen and homogeneous and become in-
creasingly basophilic. Thermal reactions of this type
were encountered only where surface temperature had
heen brought to and maintained at level of 55 C or
higher. Magnification 400X.
Figure 26. Photograph of third-degree thermal reac-
tion in porcine skin 72 hours after injury. Exudative
cells have migrated into interstices between bundles of
coagulated collagen. Precise level at which this injury
will be stabilised is not yet apparent. Healing will be
slow because of resistance of denatured collagen to
autolysis and organization. Magnification 400 X.
black, depending on certain attributes of the ex-
posures responsible for their production.
A black’or carbonized surface resulted from ex-
posures at temperatures in excess of 200 C (Fig-
ure 28). The precise temperature at which carboniza-
tion began was not determined.
A red, purple, or brown surface, due to the presence
of blood in (he superficial layer of the skin, resulted
from exposures in which the dermal temperature was
raised slowly enough to permit advanced engorge-
ment of the superficial capillary plexus before the
occurrence of coagulation.
A gray or ischemic surface indicated that the upper
portion of the dermis had undergone thermal coagu-
lation before the superficial capillaries had become
fully engorged.
The reciprocal relationships of time and tempera-
ture as they relate to the visibility of hemoglobin
beneath (he surface of a third-degree thermal re-
action is shown in Table 10. It was found that at at-
mospheric pressure and at surface temperatures of
65 C l or lower bums appeared superficially hyperemie
regardless of the duration of exposure. When a 70 C
surface exposure was interrupted at the end of 2 sec-
onds, the lesion remained red, but, if it were pro
longed to 3 minutes, (he zone of reactive hyperemia
became overlaid by so thick a layer of coagulated
tissue that it was no longer visible. Above 70 C all
exposures of a second or longer coagulated the super-
ficial plexus of dermal capillaries so rapidly that most
or all of the blood contained in them was displaced to
a level too deep to bo seen from the surface.
POOUNG OF BloOO IX 11 VJ’KKEMK' BURNS
A qualitative impression of the pooling of blood in
the dilated cutaneous vessels after an injurious epi-
sode of hyperthermia was derived from (lie photo-
micrographs shown in Figure 11. In order to learn
something of the actual amount of blood that was
present in such lesions, samples of both normal and
hyperemie skin were excised for chemical examina-
tion. Samples of skin and subcutaneous tissue having
an area of 25 cm2 and extending to the deep fascia
were taken from the lateral thoracic area of each of
nine pigs and their iron content was determined.
Two of the samples represented normal skin and
the other throe were from areas of hyperemie burn-
ing.
It is apparent from the results of the experiments
shown in Table Hi that a relatively large proportion
SECRET 34 H
STl DIES OF THERMAL INJIRX - CFTVXF.OFS \MD SYSTEMIC
Florins 27. Transcutaneous coagulation resulting in
deep ischemic burn. Five-minute exposure to air at
200 C. Surface temperature of skin not known but
probably in excess of 55 C. Epidermis h:is become re-
al Inched to dermis. Magnification H5X-
of the total circulating blood of an animal may be
pooled in the skin and subcutaneous tissue as a re-
sult of thermal injury. Calculations based on the
amount of recoverable iron per unit of surface area
Fioitkk 28- Carbonization of surface and intense baso—
pliilia of coagulated dermis due to 2.5-ininufe exposure
of skin at I (Vi ('. Effects of ambient heat augmented by
radiant energy. Magnification 85X.'
posiife to heat did not increase the vulnerability of
the epidermis to thermal injury. It was found, how-
ever, that compression of the skin was capable of
modifying the superficial color of the burn even
though there was no quantitative increase in its
severity. To determine the circumstances in which
compression of the skin during an episode of hyper-
thermia may modify subsequent surface color of the
lesion, the experiments summarized in Table 17 were
undertaken. In some, hot water was applied at .at-
mospheric pressure; in others, it was applied with a
compressive force of 120 mm of mercury.
The results of these exposures indicated that the
color of burns resulting from surface temperatures
lower than 55 C was not affected by pressure, hut
that an increase in pressure during exposures at sur-
face temperatures of <50 C or higher determined
whether the surface of the resulting burn would he
ischemic or hyperemic. Thus, an exposure at at-
mospheric pressure at 00 C produced a red burn even
though it was extended for as long as 5 minutes.
With increase in pressure, a 2-minute exposure at the
same temperature resulted in a pale burn and yet the
Table 16. Poe
due (o thermal
iling of blood in the
injury.
sal m it a neons vessels
Condition of
Mr iron in
Est. cc of blood
skin
25 em2 sample
in 25 em* sample
Normal
0.06
0.11
0.10
0.18
Moderate
0.14
0.25
hyjieremia
0.20
0.51
Severe
0.56
1.00
liypcremia
0.60
1.10
0 40
0.70
0 JO
0.70
o:j7
0.67
of humeri skin in relation to the body weight indi-
cated that as much as 30 per cent of the erythrocytes
of an animal suffering from generalized cutaneous
hyperemia could he accounted for in the skin and
subcutaneous fat.
Effect of Compressive Hypkktiikrmia ox Cot-or
of a Bfkx
In a preceding section attention was called to the
fact that compression of the skin surface during ex- PATHOLOGY OF CITANEOUS ni l!AS AMI THEN! PATHOGENESIS
319
Table 17. Experiments to determine the circumstances
in which compression of 1 lie skin during an episode of
hyperthermia may modify subsequent surface color of
the lesion.
Temp
of
surface
(C)
Duration of
hyperthermia
(seconds)
Pressure
on
skin
(nun Tig)
Kxlernal appearance
of burn 24 hours
after exjMisure
Ischemic ITyperemic
70
5
0
5
120
+
65
30
0
4"
30
120
+
GO
120
-F
1.200
0
+
GO
GO
0
+
GO
120
4-
120
120
+
300
0
+
55
1,800
0
+
1,800
120
+
bundles tended to collapse the dermal blood vessels
and the loose areolar tissue that surrounded them.
Visible edema- receded in advance of this type of al-
teration as though the fluid were imbibed or dis-
placed by the denatured collagen, Xot until 24 or
IS hours had elapsed was it possible by microscopic
examination to recognize the line of demarcation be-
tween reversible and irreversible dermal injury (fig-
ures 23 and 21).
From the intact blood vessels of the deeper and
relatively uninjured tissues, leucocytes migrated up-
ward through the perivascular interstices and into
the zone of denatured collagen. A frontier was even-
tually established between the tissue capable of re-
generation and repair and that destined to be seques-
tered in the form of a desiccated crust. The deeper
the lesion, the longer the time required for the stabi-
lization of such a frontier. The transition between the
obviously necrotic tissue of the upper dermis and the
least disturber! tissue of the deepest portion of the
zone of hyperthermia was a gradual one. Exudation
of leucocytes occurred within a few hours and within
24 hours usually served to delineate the zone within
which the plane of irreversible injury would eventu-
ally become st abilized. Within 2 or 3 days fibroblasts
and new capillaries began to push up toward the sur-
face in the interfascicular interstices of the denatured
collagen. The least affected connective tissue at the
base of the reaction zone recovered quickly and with-
out apparent loss of fixed t issue cells. The fate of the
more severely injured collagen varied according to
the extent to which it had been denatured. Thermal
denaturation of collagen at temperature levels under
55 C did not usually result in the kind of coagulalive
change that made the collagen resistant to subse-
quent autolysis and organization (Figure 2(>). Col-
lagenous denaturation at temperatures over 55 C
often resulted in an irreversible type of coagulation
which resisted lysis and eventually led to sequestra-
tion en masse. Thus, deep and severe burns resulting
from surface exposures lower than 55 C were likely
to remain soft and red and the necrotic tissue was
susceptible to organization. Deep burns resulting
from higher temperatures were characteristically
firm and pale and the necrotic tissue was seques-
tered rather than organized. After exposures to
temperatures between these two extremes the dead
and damaged connective tissue was infiltrated by
leucocytes and pcnel rated by granulation tissue and
its necrotic elements were gradually resorbed and
replaced by new connective tissue.
depth to which the tissue had been destroyed in the
latter was less than (hat to which it had been de-
stroyed in the former.
At 70 C a 5-second exposure at atmospheric pres-
sure resulted in a red burn, but, with an additional
pressure of 120 mm of mercury, the resulting burn
appeared ischemic.
Microscopic examination of these lesions provided
evidence that the color of a burn was not a reliable
criterion by which to judge its depth. After hyper-
thermic episodes of comparable duration and at the
same surface pressure, a red surface color usually
indicated that the lesion was less severe than one-
having a gray surface. Without knowledge of time,
temperature, or surface pressure during the period of
exposure, it is not possible to estimate the relative
severity of burns on a basis of surface color.
Other Effects of Heat on Dermis
After edema and pericapillary extravasation of
erythrocytes the earliest recognizable extravascular
alteration of the dermis was swelling of the collagen
fibers. Tins occurred first in its most superficial layer
where, in the case of porcine skin, the projecting
tonofibrils of the basal epithelial cells were imbedded
in the collagen of the subjacent connective tissue.
As the intensity and duration of the hyperthermia
increased, the corium tended to lose its fibrillar char-
acter and was converted into a thin lamella of homo-
geneous acidophilic material as though its individual
fibers had been converted to a gel. With increasing
exposure the swelling of collagen became apparent at
greater and greater depths in the underlying con-
nective tissue (Figure 25). Expansion of collagen
SECRET 350
STUDIES OF THERMAL INJURY CUTANEOUS V M) SYSTEMIC
During the time required to establish the level of
irreversible injury, tentative tonguelike masses of
new epithelial cells grew out from the margins of the
lesion and from the viable roots of partially destroyed
hair follicles as thoi igh they were seeking a suffi-
ciently well-stabilized layer of connective tissue to
provide support and nutrition. Repeated crops of
such new epithelial rolls extended over or into the
granulation tissue and failed to survive, for reasons
not disclosed by microscopic examination.
The number of experimentally produced deep
burns of human skin was not great enough to draw
any definitive conclusion regarding the relative rates
of healing of such lesions in man and pig. The impres-
sion was gained, however, that lesions of similar area
and depth heal more rapidly in the pig.
17.T.6 Summary
Comparison of effects of heat on human and por-
cine skin; In a previous section of this chapter it was
shown that the quantitative, relationships between
temperature, duration of hyperthermia, ami depth
of injury were similar in human and porcine skin.
In this section it has been shown that there is a strik-
ing qualitative similarity between the microscopic
alterations that are caused in human and porcine
skin by hyperthermia. The most important quali-
tat ive difference is that true vesication was not ob-
server! in the pig, whereas in man it is a character-
istic cutaneous reaction to certain types of thermal
injury. The reason for this difference has been dis-
cussed. Attention was called to (he fact tliat vesica-
tion is an undesirable phenomenon in that it may re-
sult in the separation and death of viable epidermal
cells and that there is reason to believe that healing
of certain burns in man would be hastened if vesica-
tion could be prevented.
Sequence of changes caused by harmful episodes of
hyperthermia: The earliest changes are latent, in the
sense that they are not associated with visible altera-
tion in the appearance of the damaged cells. Such
changes are reversible.
Beyond the stage of latent injury, (he pathological
changes produced by exposure to heat are of two
kinds: those that represent the reaction of living
tissue to injury and those that represent the effects
of excessive heat on cells and intercellular substances
that have already sustained irreversible injury. The
former may or may not Lie reversible and differ in
nature according to the type of cell or tissue in which
the reaction has occurred. The latter are of impor-
tanco principally with respect to the extent to which
such postvitill thermal cleiiaturation interferes with
t lie organization and disposal of the necrotic tissue.
Both types of reactions have been described in detail.
17.« CONSIDERATION OF THE N\TUHE
OF PHYSIC\L \ND CHEMICAL CH \NGES
INDUCED IN TISSUE BY
IIVPERTHERMl \ f
IT.».i Introduction
Ideally, an attempt to elucidate the precise nature
of the changes produced by heat on the skin should
be based on a knowledge of the various physical and
chemical phenomena that are normally essential to
the survival and functional integrity of the living
cells that comprise cutaneous tissue. If it were then
possible to observe the alterations of each of these
physical and chemical functions with temperature, a
direct solution of the problem of how heat injures the
skin might be readied. Unfortunately, detailed in-
formation regarding the basic physical and chemical
properties of the skin or the effects of temperature
thereon does not exist. In fact, very little qualitative
and almost no quantitative data arc available on
even the general physical and chemical attributes of
skin constituents as of to date. It is apparent, then,
that any consideration of temperature-induced physi-
cal and chemical changes which may lead to thermal
death must be based on the known in vitro effects of
temperature on substances that are akin in function
and/or properties to those which probably occur in
cutaneous t issues.
Since nearly all the quantitative experimental data
derived from this investigation deal with epidermis,
the ensuing discussion will be limited primarily to
this1 tissue. In Sections 17.0.5 and 17.9.3, these data
are shown to be quantitatively predictable by the
standard form of a rate equation,19 specifically, equa-
tion (7). In this equation there appear two empirical
and experimentally determinable constants, namely,
-4 and YE; any theoretical consideration of the cause
of thermally induced transepidermal necrosis should
take into account, at least qualitatively, the numeri-
cal values of these quantities. Aside from certain
general conclusions regarding the entropy of the
overall process,20 little specific information can be
obtained from the numerical value of A, since this
constant is intimately connected with the as yet un-
f By F. C. Ilcnriques, Jr.
SECRET PHYSICAL VXD CHEMICAL CHANCES INDUCED RV HYPERTHERMIA
351
known detailed physical and chemical properties and
functions of the epidermal constituents. This i.-> not
the case, however, with SE, and thus, before proceed-
ing further, a brief general consideration of the na-
ture of IE, the activation energy in calories per mole,
is in order.
17.8.2 Thermal Injur) and Energy of
Activation 20
In general, the kinetics of any given physical
amt Dr chemical process depends upon the total
energy content of the constituents involved. If this
energy content is less than a certain critical value,
known as the activation energy, the process cannot
take place; if the energy content is equal to or greater
than this critical value, the process may take place.
Thus the rate of the process will be proportional to
the fraction of these constituents which, collectively
considered, possess an energy content at least equal
to the activation energy. This fraction is deduced
from the Maxwell-Boltzmann energy distribution
law, which states that
f==e-*S/K(T + 2n) (1(})
where / is this fraction, and the remaining symbols
have heen previously defined. Equation (16) deter-
mines only the temperature coefficient of a rate
process, since, as shown by equation (7), the rate of
a process is also proportional to one other factor that
is essentially nondependent on temperature, namely
A.
Thus, the rate of any conceivable process that may
result in cell death, whatever it may be, depends
upon a critical energy content of the participants.
The fraction of the participants, collectively consid-
ered, having this energy is determined by the activa-
tion energy and the temperature [equation (Hi)].
The availability of this fraction is requisite but not
in itself sufficient to allow the process to proceed.
An inspection of equation (16) shows that the
temperature coefficient of any kinetic process is a
strong function of the activation energy; for example,
in the neighborhood of 50 C, the rate of a process
with an activation of I, 10, or 100 kcal mole will be
altered by alxmt 0.4 per cent, 7 per cent, or 70 per
cent, respectively, per unit change in temperature
in C.
The kinetics of a considerable number of physical
and chemical phenomena have been studied in detail
and it is possible to classify all rate processes and,
hence, in particular, those mechanisms which may lie
of considerable importance in the general considera-
tion of thermal injury, according to the order of
magnitude of their activation energy.
During the past 50 years, numerous theorieshave
been proposed to explain thermally induced injuries
in living organisms. Before applying the above cri-
teria to the mechanisms involved in these theories,
it is necessary briefly to characterize the attributes
of a living cell.12 _
The living cell appears to consist of a semirigid
relatively nonsoluble framework (e.g., nucleus, nu-
clear wall, and cell wall) that is pr imarily protein in
nature. This aggregate is bathed in an aqueous intra-
cellular fluid which contains both particulate (e.g.,
micellar) and soluble constituents ranging from
simple ions to proteins of extraordinary complexity.
Aside from certain purely physical attributes (e.g.,
permeability, contraetibilify, elasticity,
rigidity, and tensile strength), this protoplasmic
entity respires, excretes, synthesizes all imaginable
types of molecules, utilizes ami liberates energy, and
reproduces in a manner that perpetuates its own
kind. This exceedingly complex metabolic activity is
apparently both catalyzed and precisely controlled
by a multiplicity of enzymatic proteins and function-
ally allied molecules which contribute to both the
structural framework and the cytoplasmic, fluid.
In view of this complex picture, no theory of ther-
mal injury can be considered tenable unless it takes
into account these completely integrated and pre-
cisely balanced phenomena, which, taken as a whole,
comprise cell life. Unfortunately our knowledge of
these phenomena is as yet meager and is limited to
isolated observations on living protoplasm (e.g., cell
respiration, mitosis, diffusion of a few substances
through cell walls) and to certain chemical and physi-
cal properties and functions of a few of the molecules
that can be extracted in a presumably unaltered
state from dead cell brei.
Nevertheless, even on the basis of this limited in-
formation, it is interesting to speculate with regard
to the general kinds of mechanisms that may be of
importance in explaining the quantitative time-
temperature relationship that results in irreversible
epidermal injury as judged morphologically. These
injury data (Sections 17.6.5 and 17.9.3) showed that
episodes of transepidermal injury are quantitatively
predictable by a rate equation with an activation
energy of 150 kcal/mole over the entire experi-
mental skin temperature range (44 to 70 C).
The theories* that have been advanced to explain
SECRET 352
STIOI CS OF Til Fit MAL INJI'KY CITAYEOUS VXD SYSTEMIC
thermal injury may be classified into three general
groups.
1. Thermal alterations in proteins. In view of the
many varied functions of proteins in the maintenance
of normal cell life, it is obvious that even minor ther-
mally induced alterations of these molecules may
result in profound irreversible injuries. Thus, for
example, these thermal protein changes could pro-
duce an increased permeability of the nuclear and or
cell wall, structural alterations in the nucleus itself,
disintegration of the protein mitochondria present
in the cytoplasm, inactivation of enzymes.
.Many quantitative studies 44 have been made
of the effects of temperature on proteins, and altera-
tions that proceed at a measurable rate between 0 to
100 C with activation energies in excess of 50 kcal
mole are not unusual. The heat inactivation of in-
verbase (A A' — lit) kcal at pH 1 and A A’ — 52 kcal
at pll 5.7) and of peroxidase (AK ~ 189 kcal), and
the heat denatoration of egg albumin — 132 kcal
at ydl 5) and of hemoglobin (AE = 70 kcal at />H 0.8)
are a few of the many examples.
Thus, the morphological observations of protein
dissolution and or coagulation on which the quanti-
tative judgment of transepidermal necrosis is based
may well be directly due to the thermal alterations
of as yet unknown proteins present in epidermal cells.
2. Other passable alterations in metabolic processes.
Since temperature affects, to a greater or lesser de-
gree, the kinetics and thermodynamics of all chemi-
cal and physical phenomena, he at may cause altera-
tions in metabolism irrespective of its effect on pro-
teins. For example, the entire metabolic equilibrium
may be upset because of concent ration changes in
some of (he individual constituents as a result of
temperature variations both in rate of diffusion and
formation and degradation of the chemical reactants
comprising the process; in fact, because of this ab-
normal functioning, certain metabolites normally
present may completely disappear and or others
abnormal and toxic in character may arise. There
can lie no doubt that these phenomena do take place
and that they may cause cell death.
Many of these metabolic reactions,91' both en-
zyme- and nonenzyme-catalyzed, have lieen studied
as in vitro processes, and activation energies usually
between 10 and 20 kcal mole are found. In certain
instances the activation energies are less than 10
kcal mole but none have been found to exceed 50
kcal.
Thus, to date, there is no experimental evidence
(hat these typos of reactions can lead to a tempera-
ture coefficient for thermal injury which corresponds
to that found experimentally for transepidermal
necrosis.
3. X on prate in-inti need alterations in the physical
characteristics »f cells. In this group are placed all
physical phenomena that are characteristic of proto-
plasm but are not primarily effected by the thermal
alterations of proteins contained therein. For exam-
ple, diffusion of me(abolites.through a cell wall that
has not undergone chemical alteration is a member
of this class, whereas changes in diffusion rates that
are the result of an increased cell wall permeability
due to the degradation of structural protein are spe-
cifically excluded, since this phenomenon is classified
under group (1).
~ All of the biophysical rate processes that have been
studied, such as diffusion through liquids and mem-
branes, viscosity, rigidity, tensile strength, lique-
faction, possess activation energies that are usually
less than 5 kcal mole, and never in excess of 15 kcal
mole.
Although these types of mechanisms arc undoubt-
edly potentially capable of causing cell death, they
are not the instigators of the morphological changes
that are observed in irreversible epidermal injury.
Since many fat like substances are known to melt
around 45 C, the liquefaction of lipoids has received
considerable consideration as a potential instigator
of thermal injury.* From a kinetic viewpoint, the
rate of melting is a physical process with essentially
a zero activation energy. This theory would predict
a sharp temperature threshold for injury, with the
injury rate becoming nearly a linear function of the
increment in temperature above threshold value.
Hence, although liquefaction might account for the
quantitative epidermal thermal relationships at skin
surface temperatures between 45 C and 48 C, there
would be extreme variance with the experimental
data at the higher skin temperatures. The extent to
which thermal liquefaction of lipoid substances may
contribute to cell death in tissues other than the
epidermis was not investigated.
In view of the preceding discussion, it can be con-
cluded that the only biokinetic phenomena known
to date that can account for epidermal cell death are
the thermally induced changes in protein structure
which have an activation energy in the neighborhood
of 150 kcal mole. This in no way excludes the in-
jury propensity of the innumerable mechanisms im-
plied above, but merely states that all quantitative
SECRET PHYSICAL AND CHEMICAL CHANCES IMJECED BY IIYFEUTIIBHM1A
353
studies made in this investigation indicate that the
morphological changes (see Section 17.7.3 and Sec-
tion 17.7.4) observed in the epidermal tissue can lx*
ascribed to these protein alterations.
As to the number anti kinds of proteins involved,
the specific nature of thermally induced reactions,*
and the individual rate of each protein alteration,
nothing can be stated. Further, it is probable that
at any given hypothermic level any one of these
numerous protein alterations is potentially capable
of producing cell death.
1T.K.3 Thermal Injury and Entropy
and Free Energy of Activation
With no intention u'hal.smver of implying that the
thermal effects on living protoplasm can be ascribed
to the alteration of any single protein, it is of value
to make for the moment this extreme oversimplifi-
cation in order to interpret the significance of the
numerical value of A in the empirical rate equa-
tion (7) which predicts completely the thresholds of
transepidermal necrosis.
In vitro studies on both enzymatic and uonenzy-
matic proteins have shown that the rate of thermally
induced changes is first-order and the quantity of
degraded protein is given by 2,1
hi(—) = -T/’ *' 273) (*«"“) ( SE < 20 kcal can lead
to any overall phenomena with an activation energy less than
10 kcal or greater than 20 kcal). Thus, the interpretation in
the text is valid.
SECRET 354
STl'DIES OF THERM VI, 1NJI RV - CI TWEOI S AND SYSTEMIC
cytoplasmic proteins which have activation energies
for thermal degradation in the neighborhood of
150,000 cal, mole.
iT.a.t talent Thermal Injury
In Section 17.6.7, the existence of latent or mor-
phologically unrecognizable epidermal cellular in-
jury after certain apparently harmless thermal ex-
posures was proved by (he repeated applications of
subthreshold exposures. Furthermore, the time re-
quired for recovery from these latent exposures be-
came longer the nearer they approached the thresh-
old of microscopic visibility.
The concept of an unknown but definite fraction of
certain of the cellular proteins that must lx1 thermally
altered in order to result in morphologically recog-
nizable injury is in accord with these experimental
dat a.
During a heat exposure that results in latent in-
jury, a noncritical fraction of these proteins is al-
tered. At the termination of the heat exposure, the
epidermis rapidly approaches normal temperature
(Section 17.3.2) and at least partial cell function is
resumed. Thus, during the recovery period, the ther-
mally altered proteins arc replenished to a degree
which depends, in part, upon the length of the re-
covery period, and, in part, upon the duration of the
heat, exposure which produced unrecognizable injury.
17.8.5 Summary
The numerical constants of an experimental equa-
tion, w hich quantitatively predict s ) he morphological
episodes incident to 1 ransepidermal necrosis, have
been subjected to theoretical analyses. It is deraon-
st rated that, of all of the known biokinetic phenomena,
only thermal alterations in cellular proteins that have
an energy and entropy of activation of 150 keal mole
and 395 entropy units, respectively, will account for
the experimental observations. This theory is also in
agreement with the latent thermal injury data given
in Section 17.6.7.
17.9 E\post UK TO HOT AIR \N1)
RADIANT IIKAT
17.9.1 Introduction
In preceding sections of this chapter it. has I>een
shown that a very brief exposure of an animal to ex-
cessive circumambient heat may cause rapid circula-
tory collapse and death. It was found that transfer of
heat to and through the skin was a more important
causo of such casualties than was the effect of heat on
♦ lie respiratory tract. In a quantitative and patho-
logical study of the effects of hot water on the skin it
was shown that certain predictable and reproducible
reciprocal relationships exist between the intensity
and duration of an episode of hyperthermia and its
capacity to destroy the epidermis.
These findings suggest that similarly reproducible
and predictable relationships may exist between the
intensity and duration of an episode of hyperthermia
and casualty production by exposures to hot air and
radiant heat such as may occur incident to a con-
flagration or to a (lame thrower attack.
To determine whether or not such is the case a
series of experiments was undertaken in which pigs
received generalized cutaneous exposures for varying
periods of time to circumambient (and circumradi-
ant) temperatures that varied between 70 and 550 C.
The cutaneous and systemic effects of these expo-
sures on animals were correlated with exposure time
and source temperature.
17.9.2 Experimental Procedure
Previously clipped and anesthetized pigs were
fastened on a platform in the manner shown in Fig-
ure 29 and a preheated oven was lowered over them.
In most of the experiments the snout of the animal
protruded through an aperture in the bottom of the
platform. There were two advantages in this arrange-
ment, one being protection of the respiratory tract
and the other being that it was possible thereby to
determine the time of death of animals that suc-
cumbed during the |x*riod of exposure.
The source of heat was a bottomless oven con-
structed of iron and firebrick and having a capacity
of approximately 1,100 1. The box weighed 2,700
kg and its internal measurements were 89x91x130
cm. Chromel alumel (10 gauge) thermocouples
welded onto the inside plate of the box provided in-
formation as to the source temperature during the
period of pre-exposure heating as well .os during the
period that the animal was living exposed. To heat
the box, it was lowered into a vertical gun annealing
furnaceh (Watertown Arsenal). When it had become
thoroughly heat-soaked and was at a slightly higher
temperature than that at which it was desired to ex-
pose the animal, the oven was quickly withdrawn
from the furnace by an overhead crane and lowered
over the platform on which the animal was sus-
b These facilities at the Watertown Arsenal were made
available through the courtesy of the War Department. EXPOSURE TO HOT AIK AND RADIX XT MEAT
355
temperature of the right auricular blood was taken
for comparison with that, of the rectum.
In a numlter of experiments a 2S gauge iron-con-
stantan thermocouple contained in a venipuncture
needle was inserted into the dermis to record the
temperature of the subepithelial connective tissue
during and after exposure.
Temperature of air in different parts of exposure
chamber: Values given in the text for ambient tem-
perature refer to the mean temperature of the air in
which the-animal was enveloped. The thermocouples
by which the ambient temperature was measured
were routinely placed in approximately the same po-
sitions in relation to the animal in all experiments.
One was fastened to the skin just below the base of
the tail and one on each side of the mid-portion of the
body. It was regularly observed that the mean ambi-
ent temperature was approximately 20 per cent lower
than that measured by the thermocouples incor-
porated in the wall of the exposure chamber. Al-
though the rate of cooling of tlie exposure chamber
(and the air contained by it) varied according to the
magnitude «>1 the initial difference between its tem-
perature and that of the room, the drop was never in
excess of 5 per cent in experiments last ing 15 minutes
or less.
Because of the Convection currents that resulted
from the difference between the temperature ot the
surface of the animal and that of the air surrounding
it, the temperatures recorded in various parts of the
exposure chamber showed remarkably little vari-
at ion. Thus in the mid-horizontal axis of the chamber
difference in temperature was less than 5 per cent
from a point 15 cm internal to the wall to a point.
15 cm external to the animal. In the mid-vertical
axis there was less than 15 per cent difference in the
temperature of the air between a point 15 cm below
the roof and a point 15 cm above the floor of the ex-
posure chamber. —
Measurement of Heat Transfer
Under these conditions, there were three mecha-
nisms by which heat could be transferred from the hot
walls of the box to the surface of the animal, namely,
air conduction, air convection, and infrared radi-
ation. The energy transferred by conduction and con-
vection is hereafter designated as ambient, and that,
transferred by radiation as radiant. Although the
relative importance of these two types of heat trans-
fer can be directly computed by means of equations
(1) and (2) of Section 17.3, it was decided to verify
FiciTRE 29. Method of exposing animals to hot air and
radiant heat at Watertown Arsenal. Heavy iron and
firebrick box was preheated in gun annealing furnace
and lowered over platform.
pended. The interval required for the descent of the
box from the top of the tripod to the floor ot the plat-
form was bet ween 3 and 4 seconds.
The platform supporting the tripod upon which
the animal was suspended was elevated 75 cm above
the floor and covered by a layer of dry sand. In addi-
tion to the aperture to accommodate the snout of the
animal, there were other openings in the platform
through which wires could be passed to the temper-
ature recording equipment.
Three 28 gauge iron-constantan thermocouples
connected in parallel wen- fastened to the surface of
the animal in such a way that the junctions were
separated from the skin by a distance of between 2
and 5 cm. These prov ided for a continuous recording
of ambient temperature.
Rectal temperatures were taken routinely. In
some experiments a rectal thermocouple provided
for a continuous record. In others the temperature
was taken by thermometer before and at intervals
after exposure. On several occasions the postexposure
SECRET 356
STL DIES OF THERMAL INJURY CUTANEOUS AND SYSTEMIC
these calculations under the conditions that pre-
vailed in these experiments. Unfortunately, direct
determinations of the ambient and radiant caloric
uptakes of animals were not feasible and it was neces-
sary to measure these values by means of calorim-
eters suspended in the center of the exposure cham-
ber preliminary to animal experimentation.
The calorimeters consisted of copper cylinders
which measured 2.5 cm in diameter and 5 cm in
length. ( )ne of each pair of cylinders was gold-plated
and the other blackened with colloidal graphite
faquadag). Thus, the former measured only ambient
energy, whereas the latter determined both ambient
and radiant energy.
The caloric uptake rate of (he calorimeters was
readily calculated from their known heat, capacity
and surface area and the experimentally determined
rate of temperature rise as measured by an iron-
constant an thermocouple soldered within the calorim-
eter. Because of the discrepancy between the size of
these calorimeters and that, of the pigs (approxi-
mately 30x75 cm), it was necessary to multiply the
ambient calorimetric measurements by a numerical
factor equal to 0.5. Since the skin is known to lx* a
nearly perfect black body for the radiation emitted
under these experimental conditions and since the
dimensions of the exposure chamber were large with
respect to those of the animal, the radiant caloric
measurements are directly applicable.
Actually, these data, so corrected, apply to a
metallic cylinder of dimensions similar to those of a
pig. Since it has been shown that under the condi-
tions of experimentation these data would be equally
applicable to both smooth and rough and to metallic
and nonmetallic surfaces, it is believed that they
represent a true estimation of (he caloric uptake
rate of pig skin.
The data given in 'Fable IS are an estimation of
the radiant and ambient caloric uptake rate per
square centimeter per minute of pig skin when the
surface temperature is 35 C. It is obvious that during
the heat exposure the surface temperature increases
with time, resulting thereby in a corresponding de-
crease in the rate of caloric uptake. For skin surface
temperatures not greater than (IOC, caloric uptake
rates are directly proportional to the difference be-
tween the temperature of the surrounding air and
that of the surface of the animal. Thus, for surface
temperatures below 00 C the requisite caloric uptake
rates can be computed from these data. Further ex-
amination of Table IS shows that the infrared radi-
at ion from the inside walls of the box was the princi-
pal source of heat energy absorbed by the animals.
Under conditions that produced an air temperature
of 70 C, this contribution was 50 per cent, whereas at
500 C it was 85 per cent. These percentages remained
nearly invariant throughout the entire time of a
given heat exposure. As previously indicated, these
values for the nonradiant and radiant contribution
to calorie uptake rate can be directly computed from
equations (1) and (2) and, if this is done, it will be
found that they agree with the experimental values
to within about 15 per cent.
Tabi.e 18.
. Estimated e
aloric uptake
for pitr
when skin
surface temjierature is 3.1
C.
Air
Caloric uptake in cal cm- min
Per cent
tem(H*rature
Nonradiant
of total
C
(ambient)
Radiant *
Total
radiant
70
02
0.2
0.4
50
100
0.5
0.0
1.1
Hi)
150
1.0
1.4
2.4
_ 58
200
1.7
2.6
4.3
61
2,50
2.2
4.2
6.4
65
300
3.0
6.2
9.2
68
350
3.8
9.8
13.6
72
400
4.5
17.0
21.5
79
450
5.5
24.0
29.5
81
300
6.5
35.0
41.5
85
* BiTausf of tin* difference bet
ween the air and
source temperature when
animals an* placed in the exposure chamber, these radiant
data refer to a
source tern lx* nature 20 j>er cent in
i excess of the tabulated ambient tempera-
ture.
17.9.3 Effects on Animals
The results of 71 individual exposures of pigs are
shown in Figure 30. Il was at first intended to present
in this chart only the data derived from 49 experi-
ments in which pigs of uniform weight (7 to 18 kg)
received generalized (approximately 90 per cent)
cutaneous exposures to heat. The additional 22 ex-
periments included those in which large animals (in
excess of 15 kg) were used, those in which hot air was
breathed during the time that the skin was being
exposed, and those in which the animals were anes-
thetized after rather than before exposure. When it
was found that there were no significant differences
in the experimental results that could be related to
the body weight of the animals (7 and 32 kg) or to
anesthesia it was decided to present all experimental
data in one chart.
The temperature and duration of each exposure is
indicated by the position of the individual experi-
ments on the grid. The vertical points of reference on
the left are in logarithmic progression and represent
the internal temperature of the exposure chamber,
SECRET EXPOSURE TO HOT AIR AM) RADIANT HEAT
357
ing. The second line (II) represents the approximate
threshold at which generalized second-degree burning
occurred. The third line (III) represents the approxi-
mate threshold at which (he burned skin and sub-
cutaneous tissue underwent ischemic coagulation.
The skin of most pigs that received exposures lying
above this threshold was pale and the loss of elas-
ticity of the coagulated superficial tissues resulted in
the formation of deep fissures when the extremities
were Hexed. The uppermost line (IV) represents the
approximate threshold at which rapidly fatal sys-
temic hyperthermia occurred. Most pigs receiving
exposures in excess of this threshold died within a
few minutes (usually under 15 and occasionally as
long as 30) after the oven had been lifted from the
plat form.
Comparison of effects of hot air and hot water ex-
posures: Injury by heat is determined by the degree
and duration of the rise in tissue temperature. It will
be shown that for the same kind of skin the produc-
tion of a given degree of thermal injury depends only
on the time-temperature relationships within the
tissue irrespective of the source of (he heat. Since
threshold II in the hot air experiments (see Figure 30)
depicts the occurrence of transepidermal necrosis, it
can be inferred that for the same source temperature
actual tissue temperatures attained were consider-
ably lower than those in hot water experiments (sec
Figure 11).
In Figure 31 are depicted the source temporature-
time relationships that were required to produce
1 ransepidcrmal necrosis in both I he air and water ex-
posures, where in the latter case the surface of the
skin was maintained at essentially the same temper-
ature as that of the source. A comparison of (he two
curves shows that a 15-minute exposure to water at
48 C was sufficient to produce approximately the
same degree of injury as that w hich resulted from a
15-minute circumambient and radiant exposure at
75 C. A hot wafer exposure for 1 minute at 53 C pro-
duced about the same degree of injury as resulted
from a 1-minute exposure at 100 C to ambient and
radiant heat. It is apparent, therefore, that the actual
surface temperatures responsible for the kind of irre-
versible injury observed at threshold II in Figure 30
were considerably lower than the recorded ambient
temperatures at which they were produced. In the
hot. water exposures the change in tissue temperature
with time was determined by the rate of heat flow-
through the skin, whereas in the oven exposures it
was limited by the rate of heat I ransfer to the surface.
INDIVIDUAL ANIMALS
# GEN BU»N;NC AND FATAL rtTR upper limits of exposures which pigs survived
without either cutaneous injury or severe physiologi-
cal disturbance are indicated by the line (I) that
traverses the grid from left to right. Exposures lying
below this line failed to cause cutaneous burning.
Exposures lying between the first and second lines
characteristically resulted in mild or localized burn-
SECRET 358
STUDIES OF THERMAL INJURY - CUTANEOUS AND SYSTEMIC
of numerically integrating 17 equation (15) following
the .substitution of the epidermal time-temperature
relationships which result from an exposure to ambi-
ent and radiant heat as computed by equation ((»)
and recorded in Table 7B. These calculations were
made for air temperatures of SO, 100, 125, 150, and
175 C, respectively. It is to be observed that the con-
cordance of these computations with the experi-
mental data is excellent . Considerable confidence can
thus be placed both in the statement of the previous
paragraph and in the “infinite body picture” (Sec-
tion 17.3.1) which permitted the estimation of the
temperatures attained in the epidermis as a function
of time.
Probaiu.e Effects ok Comcvkaiu.k Exposmts
ox M ax
So far as the skin effects of ambient and radiant,
heat are concerned, the reactions in man and pig
should be similar if the time-temperature relation-
ships within the epidermis were t he same in each in-
stance (see Figure 14, Section 17,6.4). However, a
predictable difference in these relationships during
identical heat exposures of this type arises from the
fact that sweating of human skin can undoubtedly
increase the time threshold at which cutaneous burn-
ing occurs.
That sweating can afford considerable protection in
the case of relatively low-intensity hot air exposures
can be assumed, from the fact that man may lose
moisture by this mechanism at the rate of approxi-
mately a liter j>er hour. This could result in heat loss
at the rate of bet ween 0.5 to 1.0 cal/min/cm2 of skin
surface. Heat loss by porcine skin through vaporiza-
tion of moisture is relatively slight (approximately
0.1 cal cm2 min). See Section 17.5. Thus, in view of
the caloric uptake data presented in Table 18, it is
possible that the time threshold for cutaneous burn-
ing in man is appreciably longer than that for the pig
for all circumambient and radiant temperatures
lower than about 120 C. That such a degree of pro-
tection would l>e afforded at higher air temperatures
is unlikely since it would be necessary to assume that
sweating was already established at a significant level
at the moment of exposure and that all of the sweat
excreted was vaporized. No experiments were con-
ducted to establish the quantitative extent to which
sweating may be capable of protecting human skin
against thermal injury to either low or high ambient
and/or radiant temperatures.
It should be emphasized tliat these data refer only
Fiocre 31. Solid curve depicts time-source temperature
relationships requisite to complete trausepidermal
necrosis when skin site is exposed to flowing water of
constant temperature. Dotted curve shows time-air
temporal ure relationships (curve 11 of figure 30) that
produce similar degree of injury when skin surface is
surrounded by an envelope of radiant and ambient heat
(oven experiments). Often circles show values which
were computed by means of equations (I), (2), (Section
17.3) and (15) (Section 17.7).
The actual time-temperature relationships within
the epidermis under these experimental conditions
have Ikh'H computed by equation (6), which results
from the application of the general theory of heat to
this problem (Section 17.3.1), and are reported in
Table 7H (Section 17.3.2). These data show the rate
of increase in the epidermal temperature incident to
an exposure to an envelope of radiant and ambient
heat. It is apparent in the case of a generalized ex-
posure that long before the temperature of the sur-
face of the skin would approach (hat of the air, the
animal would have succumbed to a generalized by-
pert hermia.
In Section 17.6.5, the degree of epidermal destruc-
tion was shown to be mathematically predictable by
means of equation (15), so long as T,, the time de-
pendence of the basal epidermal temperature, is
known, 'this equation was developed empirically
from data pertaining to the degree of epidermal in-
jury when the skin surface was immediately brought
to and maintained at a constant temperature (hot
water experiments). It was stated that equation (15)
should predict the time required to produce all ther-
mally induced transepidermal injuries which result,
from any conceivable type of heat application, so
long as the time dependence of the temperature at
the dermal-epidermal junction during the heat ex-
posure is known.
The five circles depicted in Figure 31 are the result
SECRET EXPOSURE TO HOT AIR VXD RADIANT II GAT
359
to unclothed animals. It is possible to estimate the
degree of protection afforded by clothing by a knowl-
edge of their impedance to the heat reaching the skin
surface, but since this thermal impedance is so de-
pendent upon the physical characteristics of the
fabrics involved, upon tightness of fit, and upon the
type of heat exposure, further consideration of this
problem is not warranted in this chapter. The method
of obtaining these thermal protect ivities of clothing
under specific experimental conditions is given in
detail elsewhere.u
Death ok Bigs '
It may be seen from Figure 30 that rapidly fatal
physiological disturbances resulted from a wide range
of thermal exposures and that at any given temper-
ature within the range investigated survival or death
was determined by (he duration of the exposure
period. Observations were made on the various patho-
logical and physiological changes resulting from sub-
let hal and lethal cutaneous exposures to heat.
Pathological Changes
There was no apparent relationship between the
occurrence of early deat h and the severity of the cu-
taneous injury. Some animals that died during or
soon after exposure at relatively low temperatures
showed remarkably little evidence of cutaneous in-
jury. Others that received extensive third-degree
burns at higher temperatures survived many hours
alter exposure and showed no systemic evidence of
impending death at the time they were sacrificed. T(
was obvious that t he cutaneous lesion per se was not
responsible for early collapse and death.
Apart from cutaneous burning there were no sig-
nificant differences in the pathological changes ob-
served in animals that died following short exposures
at high temperatures and in those that died following
longer exposures at lower temperatures. The most
constant post-mortem finding in all animals that
died within 30 minutes after exposure to heat was the
presence of widely disseminated small and large
focuses of hemorrhage throughout the internal vis-
cera. These were seen most frequently and promi-
nently beneath the endocardium of the right and
left ventricles. Another site of predilection for such
hemorrhages was (he gastric and duodenal mucosa.
The right auricle was characteristically dilated and
filled with dark red unclutted blood. The impression
was gained that the ventricles were more frequently
found in the state of contraction after high- than
after low-intensity exposures.
The lungs of pigs (hat died during or soon after
cutaneous exposures to excessive heat rarely showed
more than a mild degree of pulmonary edema, in con-
trast to those of dogs and goats, in which systemic
hyperthermia characteristically led to moderate or
severe pulmonary edema.
Animals sacrificed 12 to 21 hours after seveie cu-
taneous burns had been sustained frequently showed
severe parenchymatous degeneration of adrenal
cortex, liver, and renal tubular epithelium. Hemo-
globin easts were sometimes observed in the collect-
ing tubules of the kidneys and the urine of burned
animals regularly contained large amounts of blood
pigment.
Changes in Blood
Examination of the blood of burned animals regu-
larly showed intravascular hemolysis. That intra-
vascular hemolysis was not a determining factor in
survival was indicated by its absence in animals that
died after low-intensity exposures. A more complete
discussion of the relationship between intensity of
thermal exposure and hemolysis will bo found in
Section 17,10.
Examination of wet and dn smears of blood of
severely burned animals disclosed microspherocytosis
and disintegration of erythrocytes (sec Figure 32).
These changes were similar to those observed in the
blood of burned human subjects, by Shen, Ham, and
Fleming.42 They were not observed in the blood of
animals that died after low-intensity thermal ex-
posures. In severely burned animals there was an
increase both in the clotting time and in the fragility
of erythrocytes.
Plasma Turbidity. The observation of turbidity of
the plasma toget her with the finding in some fatally
burned animals of small agglomerates of protein and
enmeshed cells in wet smears of blood led to a re-
invest igation of a phenomenon described by Rabat
and Levine.27 These observers reported that t he in-
travenous injection into a cat of 4 ml of heated cil-
rated plasma caused immediate death. After een-
(rifugalization, they found (hat the supernatant
fluid of such plasma produced no ill effects, whereas
death resulted from the intravenous injection of the
resuspended sediment.
A repetition of the experiments of Ivabat and
■ Scvcrnl goats and dogs received exposures estimated to he
lethal or sublet hat for pigs and the impression was gained that
their susceptibility to fatal systemic hyperthermia did not
differ significantly from that of the pig.
SECRET 360
STUDIES OF THERMAL INJURY—CUTANEOUS AMI SYSTEMIC
aortic pressure (Ilg manometer), systemic minute
output (total output less coronary flow), ventricular
volume (Henderson cardiomelcr with a Kie.se volume
recorder), and oxygen consumption (spirometer).
The preparation had been list'd earlier fora study
of the metabolic effects of alloxan; the heart was
failing spontaneously at the time the blood from the
burned animal was introduced into the perfusing
system (about 3 hours after the preparation had been
isolated). Fifty milliliters of the blood of the heated
animal were injected into the venous return during
a period of 1 minute. The minute output was about
130 ml min at the time of i\f injection and the in-
jected blood reached the heart diluted two or three
times with the original blood of the preparation.
Six minutes later 100 ml of the blood of the burned
animal were injected again in a period of 1 minute.
This was diluted no more than once wit h the blood of
the preparation.
Six minutes later the blood in (he venous reservoir
was removed and replaced with 200 ml of blood of
the burned animal. Following this last addition the
heart-lung preparation was living perfused almost
entirely by the blood of the heated animal.
In none of the three t rials was there any significant
change in the pressure, the minute output, the heart
rate, or the oxygen consumption. Although coronary
How was not recorded, any great increase in it such
as might have been expected if the blood had con-
tained as much as 0.5 mg of histamine would have
been recognized by an increase in the discrepancy
between the stroke volume as recorded by the cardi-
ometer and the stroke volume as calculated from
minute output and heart rate. Such a change was not
observed.
No deleterious effect resulted from perfusing the
heart-lung preparation with t he blood of the burned
dog. Actually there was slight evidence of a beneficial
effect, such as would be expected from the addition
of any fresh blood after 3 hours of perfusion.
Relation ok Systemic Hyperthermia to Survival
There appeared to be a definite correlation be-
tween survival and the height to which the internal
body temperature was raised. Most of the animals
that died soon after exposure were found to have a
marked elevation of rectal temperature. In the case
of exposures of long duration and low intensity the
rectal temperature was only slightly lower than that
of the blood within the right auricle. In animals that
died within a few minutes after exposures of short
Flue re 32, Blood smears of pig No. 856 ( 0.1 kg) 1*>-
fore ( A) and 3 minutes afti*r(B)5-minute exposure to hot
air and radiant heat at ambient tenifierature of 180 C.
Animal received third-degree hums of about 85 per cent
of hody surface and died 3 minutes later-witli rectal
temperature of 43.5 C. Examination disclosed intra-
vascular hemolysis, plasma potassium concentration of
10.1 milliequiv 1 and disintegration of erythrocytes ns
shown in (BP During exposure, temperature at inter-
face In'tween dermis and suMermal fat, as recorded by
needle thermocouple, rose to maximum of 03 C.
Levine resulted in the observation that blood presr
sure fell rapidly and (hat sometimes animals died
following the intravenous inject ion of a small amount
of heated citraleA plasma. However, when heparin
was used as an anticoagulant instead of cit rate, ani-
mals tolerated relatively large intravenous injections
of heated plasma without ill effects and without sig-
nificant change in blood pressure. Slight lowering of
blood pressure was observed in a few animals alter
injection of heated heparinized plasma or the sedi-
ment of heated plasma. No deaths occurred, how-
ever, even when amounts as great as 15 ml were used.
It was concluded (hat (he particulate masses in
preheated blood described by Rabat and Levine may
lie deleterious to a slight degree and in combination
with sodium citrate (250 mg 10 ml of blood) may
cause death if injected rapidly. It is not believed,
however, that these masses contributed significantly
to the hyperthermic deaths observed in these ex-
periments.
Perfusion Experiments. A heart-lung preparation
(Starling method) was perfused with the blood of a
dog that had died of circulatory failure 7 minutes
after lining immersed in hot water at 70 C. Continu-
ous records of the heart-lung preparation included
SECRET EXPOSURE TO HOT AIK AND RADIANT HEAT
3«I
duration and high intensity there was characteristi-
cally a difference of several degrees between rectal
and blood temperature (see Section 17.11).
The correlation between severity of systemic hy-
perthermia and the occurrence of early death is
shown in Figure 33. With one exception all pigs that
Pathological Physiology 1
Prior to the exposure of several pigs to hot air, in-
sulated electrocardiographic loads wore connected
with the extremities and a carotid cannula was in-
(roduced. The effect of the exposure on the rale of
respiration, the pulse rate, the arterial blood pres-
sure, and the conduction system of the heart of
those animals was observed.
Within a few seconds after exposure, there was a
sharp increase both in blood pressure and in rate of
respiration. The respiratory rate continued to in-
crease and remained rapid for some time after the
exposure was terminated. Soon after the initial rise
there was a tall in blood pressure to or slight ly below
the pro-exposure level. In some animals, the pressure
was well maintained at that level until within a few
minutes before death, whereas, in others, there was a
gradual and progressive decline beginning immedi-
ately at the conclusion of the initial rise, inability to
control the movements of the animal during the
period of exposure made it impossible to secure satis-
factory records of venous pressure.
Elect rocardiographic abnormalities were observed
in some animals soon after the beginning of exposure,
whereas, in others, such changes did not develop un-
til well after the onset of circulatory failure. Abnor-
malities observed in a few instances soon after the
beginning of the exposure (within 2 or 3 minutes) in-
cluded increase in rate, reduction in t be voltage of
the (>RS complex, and inversion of the T waves.
Ventricular extra-systoles were observed and as the
exposure was prolonged there were greater disturb-
ances in rhythm. Such animals developed vent ricular
tachycardia followed by fibrillation and death.
Although abnormalities in the electrocardiogram
were sometimes observed before there was evidence
of respiratory failure, the. terminal and agonal fall in
blood pressure usually occurred at about the same
time that tachypnea gave way to intermittent peri-
ods of apnea. —
Although the results of these experiments indicated
that there were two types of hyperthermic circula-
tory failure, one central and the other peripheral, it
was obvious that further and more rigidly cont rolled
physiological experimentation was required. Such
studies were not feasible in the circumstances in
which the hot air experiments were conducted (see
Section 17.11).
Changes in filood Potassium. Samples of blood were
withdrawn by cardiac puncture before, during, and
after lethal exposures of four pigs to hot air. It was
Ficcke 33. Distribution of animals according to maxi-
mum 30-inimitc rise in rectal temperature following
exposure to hot air and radiant heat. Initial tempera-
tures were low liccause of pentobarbital sodium an-
esthesia. Open portions of columns represent animals
that survived; shaded portions, animals that died dur-
ing or within 30 minutes after exposure. It is apparent
that there is close correlation between systemic hyper-
thermia and death.
died during the early postexposure period were those
that developed rectal or heart's blood temperatures
of 42.5 C or higher. Xo pig whose rectal temperature
rose to 44 C or higher survived for more than a few
minutes. Eleven of the 15 that developed rectal
temperatures between 13 and 14 C and 4 of the 13
with rectal temperatures between 12 and 43 C died
during the episode of hyperthermia.
SECRET 362
STUDIES OF THERMAL INJURY —CUTANEOUS AND SYSTEMIC
found that the potassium concentration of pig’s
blood is approximately 50 milliequiv 1. The partition
of potassium between erythrocytes and plasma is
approximately 50 to 1.5. The post exposure plasma
levels in these four animals were 7.3, 10.0, 17.4, and
19.0 milliequiv 1, respectively. The observation that
cutaneous hyperthermia was capable of causing the
plasma potassium to, rise to 17 milliequiv ! and
higher suggested acute potassium poisoning as a po-
tential cause of death. Further investigation of the
importance of potassium release to the occurrence of
circulatory failure and death following exposure to
heat will be discussed in Sections 17.10 and 17.11.
I7.«).t Summary
The time-temperature relationships responsible
for varying degrees of cutaneous injury and for acute
circulatory collapse and death incident to exposures
to circumambient and circumradiant heat similar to
those that may result from a conflagration or from a
flame t hrower attack have been determined for t he pig.
At relatively low air temperatures (under 120 C)
man, because ofhis ability to sweat, is undoubtedly
less susceptible to injury than the pig. It is doubtful,
however, that sweating provides a significant degree
of protection at higher temperatures in which the
rate of heat transfer to the skin is considerably more
rapid than the rate at which it can be dissipated by
vaporizat ion of sweat .
It should be borne in mind that the relationships
of source temperature to injury production derived
from these experiments apply to unprotected skin
and are not valid for exposures in which the skin is
protected by hair or clothing.
It has been shown that the time-tissue tempera-
ture relationships responsible for t ranscpidermal
necrosis (second-degree burning) by exposure to hot
water as given in equation (15) (Section 17.6) are
equally applicable to exposures to circumambient
and eireumradiant heat.
The severity of the immediate physiological dis-
turbances resulting from exposure to excessive heal
is frequently disproportionate to the severity of cu-
taneous burning. Rapid circulatory collapse and
death may result from exposures of such low intensity
that little or no burning of the skin is sustained. Ex-
posures ot short duration at. higher temperatures may
cause severe and generalized cutaneous burning with
remarkably little systemic physiological reaction
during the early postexposure period.
The severity of the immediate physiological dis-
(urbanees resulting from exposure to excessive heat
bears a quantitative relationship to the extent to
which the body temperature is increased. Pigs in
which the recta! temperature failed to rise above
12 0 rarely and those in which it rose as high as 44 C
invariably died of acute circulatory failure. In ani-
mals that died within a few minutes after exposure
to excessively high environmental temperatures, the
temperature of heart's blood was consistently higher
than that recorded by a rectal thermometer. The
shorter the interval between onset of exposure and
death, the greater was the difference between the
temperature in the rectum and that in the heart.
Although the precise physiological mechanisms re-
sponsible for hyperthermic circulatory failure were
not fully elucidated by these experiments, it was ap-
parent that the early death of some burned animals
was caused or contributed to by hyperpotassemia.
Perfusion experiments failed to disclose the pres-
ence of injurious humoral agents (other than po-
tassium) in the blood of recently burned animals.
Pathological examination of the bodies of animals
that died during or soon after an episode of acute
systemic hyperthermia disclosed evidence of capil-
lary endothelial damage in tin’ form of disseminated
visceral petechiae. Int ravascular hemolysis and al-
terations in the form and fragility of erythrocytes
were observed in animals that had sustained severe
cutaneous burning.
17.10 HYPEKPOTASSEMU CAUSED RY
EXPOSURE TO HEAT
it.to. l Introduction
In Section 17.1), it was observed in some experi-
ments that cutaneous exposure of pigs to excessive
heat resulted in rapidly fatal circulatory failure that
was associated with marked electrocardiographic ab-
normalities and a sharp rise in plasma potassium to
levels ordinarily considered incompatible with life.
The implication of t hese observations was such as
to warrant further study of the effect of cutaneous
hyperthermia on the concentration of potassium iu
the blood.
17.10.2 Experimental Procedure
Samples of blood for chemical analysis were ob-
tained from the heart by means of an inlying jugular
cannula.
Potassium determinations were carried out on t he
trichloroacetic acid filtrate of plasma and lysed
blood according to the method of Lowry and 1 fast-
SECRET HVPERI’OTASSEMIA CAUSED BY EXPOSURE TO HEAT
303
ings ® as modified by Cohn and Tibbetts. Hema-
tocrit was determined in \\ hit robe tul>es after cen-
trifuging for 30 minutes at 2500 rpm. The method of
Bing,33 et al, as modified by Ham,34 was used for de-
termining plasma hemoglobin. Whole blood hemo-
globin was determined on 0.1 ml of 1 5 dilution of
blood in 6 ml of dilute ammonia by the Ivlett-Sum-
merson colorimeter.
appears that the observed decrease in the concen-
tration of intracellular potassium from 132 to 128 mil-
licquiv I was probably due to swelling of red cells
rather than to loss by leakage. Since the actual po-
tassium content of the erythrocytes did not appear
to have dropped and since there was no hemolysis, it
was inferred that (he porassium in the plasma had
been increased by diffusion from extravasoular
sources.
The need for taking blood promptly after death, if
reliance is to be placed on analytical results, is illus-
trated by the rise in plasma potassium that occurred
during the first hour post mortem. At death, the
plasma concentration of potassium was 9.3 milli-
equiv 1, wheivas 1 hour later it was 1(5.8. Although
there is no evidence in the data presented in Table 19
as to the source of this increment, other observations
indicated that both leakage from red blood cells and
diffusion from extra vascular tissues may cause a
post-mortem rise in plasma potassium. So far as the
significance of this experiment in providing control
data is concerned, it is apparent that a twofold rise
in plasma potassium may occur as a result of severe
systemic anoxia.
In order to” correlate chemical data with known
degrees of cutaneous hyperthermia, it was decided
to submerge animals in hot water rather t han expose
them to hot air. By the former method, the tempera-
ture of the surface of the skin could be controlled
wit h greater precision than was possible by the lat ter.
The experimental procedure that was followed in
submerging animals in hot water is described in de-
tail in Section 17.11. The animals were anesthetized
with pentobarbital sodium and between 00 and 75
per cent of the total body surface was raised to the
desired level. The effects on the blood of exposing
four pigs to water at 47 C and eight pigs to water
at 75 C are shown in Table 20.
Exposure at 17 C: Although all these animals de-
veloped an acute and rapidly fatal systemic hyper-
thermia, none showed a rise in plasma potassium sig-
nificantly greater than that which may result from
anoxia independently of hyperthermia. In none of
these was the magnitude of the increase comparable
to that which was observed in some of the severely
burned animals reported in Section 17.9.
In the first two animals, it appeared that the po-
tassium increase in the plasma was derived from ex-
travascular sources. In the third animal the increase
was due to leakage in only one sample. In the fourth
animal it may have been due in part to leakage from
17.10.3 Animal Experiments 1
Before undertaking further investigation of the
relationship of hyperthermia to the development of
hyperpotassemia, an experiment was undertaken to
determine the effect, of systemic anoxia on the po-
tassium concentration of the plasma independently
of hyperthermia (Table 19).
Tabi.e 1ft. Changes in the blood of a pig during and after
death
by strangulation
Hemoglobin
in
Vol-
Hemo-
plasma
Blood
ume
globin
% Potassium
Potassium
with-
packed
in cells
heinoly- in red cells
in plasma
drawn
cells
g 100 ml sis millicquiv/1
milliequiv/l
Control
45
83
0 132
5.2
0 min
Trurhm damped
4 min
4ft
31
0 128
ft.l
8 min
48
34
0 130
9.3
8 min
Animal died
68 min
?
?
0 ?
16.8
A cont rol sample of blood was taken from an S.2-kg
pig. The trachea w as then exposed and clamped and
after 4 and 8 minutes additional samples of blood
were obtained. The animal died at the end of 8 min-
utes and was allowed to remain on the operating
table at room temperature for an hour thereafter, at
w hich time, the fourth and last sample of blood was
withdrawn. The analytical results are shown in
Table 19.
It may be seen that the plasma potassium level
was almost doubled during the 8 minutes that elapsed
between the onset of asphyxia and death. Most of the
increase occurred during the first 4 minutes of this
period. There are two obvious sources from which the
increment may have been derived, one being the
erythrocytes and the other the extravascular tissue.
A comparison of hematocrit and hemoglobin content
of cells at the end of the 1-minute period indicates
that swelling of erythrocytes had occurred. The
hematocrit rose from 45 to 49, whereas the hemo-
globin dropped from 33 to 31 g per 100 ml of cells. It
SECRET 364
STUDIES OF THERMAE INJURY — CUTANEOUS AND SYSTEMIC
intact erythrocytes, and in part to diffusion from kind produced in those animals did not result in a
extravascular tissue. Cutaneous hyperthermia of the significant amount of intravascular hemolysis.
T
MILE 20.
Effects on the blood of exposing pigs
to hot water.
Potassium in plasma — milliequiv/l
Hemoglobin
Increment
lltood
He mo-
in Potassium
Potential
from
Thermal
Body
Time
samples
Volume
globin
plasma
in red
increment
source*
I*i(5
Time
ex pusure
temp
of
time
parked
in cells
% hemol-
nils
from
other than
No.
. min
c
c
death
taken
erlla
g/100 ml
ysis milliequiv/1
Total
Change
hemolysis
hemolysis
877
Control
34.3
Control
32
29
0
145
3.8
0
Started 1
10
10 min
33
30
0
1.58
6.2
+2.1
0
2 4
14
‘47
14 mm
33
30
0.
154
6.9
+3.1
0
3 1
24
44 3
21 min
31
32
0.
1.58
8.2
+4.4
0
44
20
Stopped
+
1(157
Control
37.0
Control
35
35
0.1
115
4.1
0
Starter! 1
— -
20
47
20 min
36
35
0.0
125
7.0
+ 2.6
0
2.6
30
Stopped
15.5
+
36 min
36
35
0.2
120
10,2
+5.8
0.2
5.6
1056
Coni rol
37.8
Control
33
37
0.1
118
4 7
0
Started '
10
10 min
33
36
0.1
114
59
+ 1.2
0
1.2
15
47
15 min
35
33
0.1
113
7,2
, +2.5
0
2.5
34
34 min
36
35
0.2
US
7.1
+ 2.4
0.1
2.3
43
Stopped
45.5
+
923
Control
?
Control
48
33
0.0
f
3.8
0
Started [
—
13
13 min
47
14
0.1
124
5.5
+ 1.7
0.1
1.6
23
23 min
46
40
0.2
120
5,5
+ 1.7
0.2
1-5
34
■1/
34 min
55
32
. 0,1
113
6.2
+2.4
0.2
2.2
42
42 min
55
32
0.1
112
6.5
+ 2.7
0.2
2.5
47
47 min
36
33
0.1
7.5
+3.7
0.2
3.4
SO
Slopped
?
+
809
Control
37.4
Control
38
34
0.4
139
3,6
0
Started
u.
1
Stopped
75
—
5
5 min
48
31
3,0
109
10.2
+ 6.6
3.7
2.9
10
16 min
37
33
80
117
6.9
+3.3
0.5
40
46 min
39
32
7.5
IIS
4.2
+0.6
6 2
76
39.2
76 min
37
35
6.7
122
7.4
+3.8
5.2
918
Control
30.0
Control
34
41
0.0
131
3.7
0
Started
>75
7*
_
3
Stopped
4
4 min
51
30
2.5
98
11.0
+ 7.3
2.6
4.7
a
11 min
45
42
4.4
no
9.5
+5.8
4.2
1.6
17
17 ruin
44
35
5 9
102
9.5
+ 5.8
5.1
0.7
37
406
37 min
10
48
5.6
103
9.4
+5.7
4 0
1.7
55
+
919
Control
Control
43
33
0,8
118
4 2
0
Started
37.1
— . . .
4
>75
1 min
36
29
7.8
81
25.5
+21.3
8.6
12.7
3
Stopped
8
8 min
47
26
255
67
21.4
+ 17.2
20.2
10
10 min
40
32
22.2
*»
18.3
+ 14,1
T
M
14 min
35
37
23.1
77
17.0
+ 12 8
12.6
17
44.3
17 min
33
31
30.1
72
17.5
+ 133
15.2
18
+
913
Control
380
Control
26
37
0.0
no
3 5
....
0
Started
- —
2
1.5
2 min
35
32
12.3
103
14.2
+10.7
7.7
3.0
6
6 min
32
33
24.5
06
17,7
+ 14.2
14.7
7
Stopped]
8
10.8
+
8 min
30
32
25.3
in
17.4
+ 13.9
16.0
...
907
Control
37.3*
Control
12
34
0.6
125
3,5
0
.Started
8
„o
8 min
53
31
2.7
100
17.4
+ 13.9
3.1
10.8
10
Stopped]
42.5*
4-
*
Right heart temperature.
SECRET HYPERPOTASSEMI V CAUSED UV EXPOSURE TO HEAT
305
Table 20 (Continued).
Pi*
Ko,
Time
min
Thermal
exposure
c
Body
temp
C
Time
of
death
Blood
samples
time
taken
Volume
parked
fells
Hrmo-
idobin
in eells
*100 n a
Hemoglobin
in Potassium
plasma in red
% hemol* eells
j sw mitliequiv. I
Potassium in plasma milliequiv/l
Increment
Potential from
increment sources
from other than
Total Change hemolysin hemolysis
910
Cont rol
36 8
Control
r
7
3.0
0
Stalled
2
2 min
7
7
19.1
+ 16.1
5
5 min
?
7
181
+ 15.1
7
40
. . .
7 min
?
J
... *
24.0
1 21.0
13
+
.
14
Stopped
13.7
14 min
... -
17.3
+ 14.3
908
Cont rol
*
Cont rol
32
f
7
106
3.8
0
Started
—
—
4
4 min
?
7
7
*
16.7
+ 12.9
»
i75
9 min
33
?
7
98
18.5
+ 14.7
. .
11
11 min
32
f
7
90
17.1
+ 13.3
H
Stopped
7
+
912
Control
36.0
Control
33
37
0.0
125
4.1
0
Started
1
I rain
4 a
31
1-9
102
16.7
+ 12.6
1.6
11.0
4
...
4 min
33
37
19.2
7
7
?
5
5 min
34
31
21 2
100
164
+ 12.3
16.5
10
10 min
40
31
19 9
85
16,4
+ 12.3
14.2
14
Stopped
43.1
+
Exposure at 75 C: The chemical changes in this
group were of a different order of magnitude from
those observed in animals exposed at 47 C. All ani-
mals exposed for 5 minutes, or longer, at 75 C de-
veloped plasma potassium levels in excess of 10 milli-
equiv/1. In most instances, such levels were reached
during the first few minutes of exposure and were
either maintained or increased as the period of ex-
posure was prolonged. If the pig survived for more
than a few minutes after the termination of the ex-
posure, there was a slow decline in plasma potassium
concentration. Thus, in animal 919 the plasma po-
tassium rose from 4.2 to 25.5 rnilliequiv during the
first 4 minutes of exposure, and during the next
4 minutes declined to 17.4.
The rapidity with which an excessively high
plasma potassium level may be lowered by extra-
vascular diffusion is indicated by the discrepancies
that were observed between estimated increments
by hemolysis and total amounts present. Thus, it
may be seen in the case of pig 913 that with an incre-
ment by hemolysis of 7 milliequiv/1 between the 2-
and 0-minute samples, the actual plasma level rose
by only 3.5 rnilliequiv. Similarly, in pig 912 the in-
crement by hemolysis between the 1- and 5-minute
samples was 14.9 rnilliequiv 1, whereas the total
plasma potassium actually changed from 10.7 to 10.4
during this period.
In most of the animals exposed at 75 t', there was
some increase in the volume of packed cells. The
comparison of cell volume and hemoglobin content
indicated that most, if not all, of the early increase in
cell volume was due to swelling of erythrocytes
rather than to loss of plasma or mobilization of cells
from storage depots.
It is of interest to note that plasma hemoglobin
values as high as 24 per cent hemolysis were observed
as early as 5 minutes after the onset of cutaneous
hyperthermia. It was estimated that during this
period the temperature in the vicinity of the most
superficial blood vessels probably rose to approxi-
mately 70 C.
Chemical changes in the blood of dogs caused by
cutaneous hyperthermia; It was inferred from the
foregoing experiments on pigs that most of the po-
tassium responsible for these potentially fatal plasma
levels either leaked out of intact red blood cells or
escaped from hemolyzed cells. If this inference is cor-
rect, fatal hyperpotassemia due to cutaneous hyper-
thermia would occur only in animals having a high
concentration of potassium in the erythrocytes, such
as man or pig. Its occurrence could not be expected
in an animal having a low cellular concentration of
potassium, as is the case in dog’s blood.
To test this assumption, samples of blood were
taken from each of five dogs before and during im-
mersion in hot water. The results of these experi-
ments are shown in Table 21.
The animals were exposed at temperatures ranging
between 55 and 75 C until death occurred. The high-
SECRET 366
STUDIES OF THEUMAL INJl RY — CUTANEOUS AM) SYSTEMIC
Table 21.
Changes in blood of dogs caused by immersion
in hot water.
Blood
Time
Thermal
Body
Time
samples
Volume
Hemoglobin
I lemoglobin
Potassium
Potassium
Hog
in
exposure
temp
of
time
packed
in cells
in plasma
in red cells
in plasma
Xo,
min
c
C
death
taken
cells
g 100 ml
% hemolysis
milliequiv 1
milliequiv !
931
Control
35.4
Control
35
37
0
9.4
2.8
0
Started
5
5 min
41
36
0
8.1
5.2
13
55
13 min
57
32
0
10.7
4.7
21
41.4
21 min
57
33
0
11.2
6.9
23
Slopj)ed)
+
930
Control
36.9
Control
49
34
0.1
4.3
4.0
0
Started
5
. - .
5 min
66
27
17.9
6.4
33
8
60
8 min
65
28 J
20.2
5.5
4 7
11
39.1
11 min
62
28 /
23.8
6.1
5.3
17
Stopped
+
929
Coni rol
37.2
Control
49
34
0.3
6.3
3.9
0
Started
3
3 min
57
29
26.1
7.0
4.8
9
7 5
9 min
42
37
31.8
5.7
6.1
13
44.1
13 min
39
34
35.8
7.9
8:2
14
Stopj)cd
+
.. —
* * * *
922
Control
—
37.9
Control
42
35
0,2
8.8
3.1
0
Started
3
3 min
_ 47
30
22.9
8.9
5.8
7
75
,
7 jnin
47
30
29.5
12.6
6.4
10
10 min
43
29
33.5
7.9
5.8
15
Sloped
39.3
+
15 min
45
30
31.4
8.0
6.8
931
Coni ml
34.6*
Control
41
35
0.1
5.6
3.1
a
Started 1
25
Stopped/
43.5*
+
25 min
40
34
31,9
6.5
6.9
♦ HiKht heart temperature.
17.10.1 In I iiro Effects of Heat on
Pig’s Blood
It was thought (hat more precise information re-
garding the reciprocal relationships of temperature,
time, and the release of potassium from erythrocytes
could be obtained by heating samples of pig’s blood
in vitro.
Heart’s blood was collected from normal pigs by
cardiac puncture in a heparinized syringe, where it
was mixed and then discharged into heparinized
glass-stoppered vials. One vial was kept at room
temperature as a control; the others were strapped
to a mechanical mixer and immersed in a constant
temperature bath. Exposure temperature's ranged be-
tween 44 and 63 C; during exposure the blood was
mechanically decanted from one end of the vial to
the other at a rate of six times per minute. It re-
quired approximately 2 minutes for the temperature
of the blood to reach that of the water bath. As soon
as a sample was removed from the water bath, it was
immediately cooled in ice water and analyzed.
It is apparent that there was a progressive increase
in the rate at which potassium passed out of the
est potassium concentration observed in the erythro-
cytes in control samples of blood from these animals
was 9.4 millieqniv ly hi contrast to the pig, whose
erythrocyte concentrations ranged I»etween 106 and
145 millieqniv 1. The greatest potassium increase
that occurred in the plasma of the dogs that died as
a result of cutaneous exposure to heat was from 3.9 to
S.2 milliequiv/1.
T1 ie increments to the plasma potassium that were
observed in these animals could not be accounted for
by loss of potassium from the erythrocytes. The po-
tassium content of the red blood cells of the dogs
characterist ically rose during exposure in contrast to
the loss of potassium that occurred from the erythro-
cytes of the pig. As in the case of the pig, there was
severe intravascular hemolysis in animals exposed at
75 C until death occurred.
It can be inferred, therefore, that the development
ot a potentially fatal level of hyperpotassemia fol-
lowing cutaneous exposure to heat results from the
rapid release of potassium from thermally injured
red blood cells and that a high erythrocyte content
of potassium is essential to its occurrence.
SECRET HYPERPOTASSEM1A CAUSED BY EXPOSURE TO HEAT
Tabi.k 22. In vitro effects of heat
on pig's blood.
Potassium in plasma — milliequiv/1
Time
Volume
Hemoglobin
Hemoglobin
Potassium
Increment
Increment
S|)oci-
Temp
in
packed
in cells
in plasma
in red cells
from
from
mm
C
minutes
cells
g 100 ml
"% hemolysis
miliicqiiiv/1
Total
Change
hemolysis
leakage
1 047
Control
Control
30
34
0.1
105
3.2
40
15
30
34
0.1
113*
3.5
(0.3
0.3
30
30
32
0.3
111*
3.5
+ 0.3
0.1
0.2
60
30
34
0.1
102
3.8
+0.6
0.6
2 049
Cont rol
Cont rol
32
33
0
99
3.2
44
15
31
33
0.1
107*
3.9
+0.7
0.7
30
31
34
0.1
102*
4.0
+ 0.8
0.8
60
31
33
0.3
97
4.8
+ 1.6
0.1
1.5
3 940
Control
Control'
31
34
0
lot
4.6
48
15
32
31
0.1
101
7.5-
+2.9
2.9
30
32
32
0.1
90
9.2
+ 4.6
4.6
00
32
29
0.4
90
11.0
+6.4
0.1
6.3
1-930
Control
Control
33
32
0.0
109
4.3
51
15
35
81
0.8
96
10.2
+5.9
0.4
5.6
30-
34
34
0.5
98
11.8
+7.5
0.2
7.3
(40
36
31
0.7
92
10.7
+6.4
0.4
6.0
5 050
Control
Control
34
35
0.1
120
4.2
52
15
35
34
0,8
103
10.0
+5.8
0.5
5.3
—
. . .»
30
35
32
2.7
101
10.4
+6.2
1.5
4.7
60
- 36
32
2.7
100
10.9
+6.7
1.6
~5.1
6-1*47
Control
Cont ml
31
33
0.1
109
4.2
55
15
40
28
13
85
7.5
+3.3
0.7
2.6
30
37
30
5.7
87
12.1
+ 7.9
3.0
4.9
60
37
30
9.6
71
18.5
-t 14.3
4.4
9.9
7-1052
Control
Control
38
33
0.0
119
3.6
60
5
48
28
1.4
S3
12.6
+9.0
1.2
7.8
8-1052
Control
Control
36
35
0.1
121
4,1
61
5
45
28
3.5
76
20.8
+ 16.7
2.4
14.3
9 10.52
Cont rol
Control
34
36
0.0
122
3.9
62
5
33
34
11.7
69
30.8
+26.9
4.5
22.4
10-1052
Control
Control
30
36
0.1
121
4.1
63
5
26
34
31.6
58
40.2
+36.1
9.6
26.5
♦ These values must be due to analytical errors.
erythrocytes and into the plasma of the blood as its
temperature was raised ('fable 22). The amounts of
the plasma increment at the end of 1 hour’s exposure
at ft), Id, 18, 51, 52, and 55 C wen* respectively 0.G,
l.G, 6.1, 6.4, 6.7, and 14.3 milliequiv 1. At the lower
temperatures (51 C and under), the increments were
due almost entirely to leakage from intact cells. At
the end of 30 minutes of exposure at 52 and 55 0,
the proportion of the plasma increment contributed
by hemolysis was 24 and 38 per cent, respectively.
Unequivocal ev idence of swelling of erythrocytes
was first observed at 55 C, although there may have
been some swelling in all specimens exposed for more
than 30 minutes at IS O and higher.
The rate of change in the blood was much more
rapid during exposures at 60 C and higher. In these
experiments the blood remained in the bath for only
5 minutes and the actual time during which it was at
the temperature of (he water was approximately
3 minutes. The rise in plasma potassium after such
brief periods at GO, 61, 02, and 63 C was, respectively,
9.0, 10.7, 20.9, and 30.1 imiiiequiv/1. The biood was
totally hemolyzed at 05 0'.
Not until blood was heated at GO or higher in a test
tube, were the observed increases in plasma potas-
sium comparable with those that occurred in living
pigs after cutaneous exposures at 75 C. This is not to
imply that the effects of hyperthermia on blood in a
test tube are necessarily similar to those effects in a
living animal. Attention has already been called to
the fact that asphyxia without rise in temperature
may cause hyperpotassemia in a living animal. Al-
though the mean temperature of the blood of a living
pig is never raised to GO C, most or all of its blood
may in the course of its circulation through the over-
heated dermis be brought to a much higher temper-
ature than would be recorded by a rectal thermom-
eter or intracardiac thermocouple. It will be recalled
from the calculations made in Section 17.3 that the
superficial portion of the dermis of a living pig
SECRET STUDIES OF THERMAE INJURY — CUTANEOUS AND SYSTEMIC
reaches a temperature of GO C within a second after
the surface of the skin has been brought to 75 ('. It
would appear quite possible then that the temper-
ature of most or all of the blood of an animal that
had received an extensive cutaneous exposure to
water at 75 (’ for as long as 5 minutes would be
raised briefly during its passage through the sub-
cutaneous tissue to the neighborhood of 60 C.
Not until the temperature of the bath was raised
to 62 C did a 5-minute exposure of blood in a test,
tube result in hemolysis comparable with that ob-
served in living pigs exposed at 75 C.
Attention has already been directed to the fact
that unequivocal swelling of erythrocytes was first
observed in a test tube after a 15-minute exposure
at 55 C. So far as could he judged by the hemoglobin-,
hematocrit ratios, swelling of erythrocytes continued
through 61 C, beyond which if was not observed.
17.10.5 Summary
These experiments have established that severe
and extensive cutaneous burning may result in a
rapid rise in plasma potassium to levels ordinarily
considered incompatible with life. Such levels are
obtained when a large proportion of body surface is
maintained at 75 C for more than a few minutes.
That lower surface temperatures may also be re-
sponsible for fatal hyperthermia is suggested by the
fact that potassium is released rapidly from blood
cells in vitro at temperatures of 60 C or over. In pari
because of the slowness with which potassium is re-
leased at lower temperatures and in part because of
the rapidity with which excess potassium leaves the
blood stream, it is not likely that thermal exposures
of insufficient intensity to cause severe cutaneous
burning could cause sufficient damage to t he erythro-
cytes to produce dangerously high plasma levels.
In vitro experiments on pig’s blood indicate that
rapid leakage of potassium from erythrocytes oc-
curred when its temperature was raised over GO C
and that rapid hemolysis occurred when its temper-
ature was raised above 62 C. Leakage was accom-
panied by swelling at temperatures ranging between
55 and til C. Above that temperature, so far as could
be judged by the hemoglobin content of cells, rapid
release of potiissium occurs without cell swelling.
It was demonstrated that leakage from and lysis
of red blood cells were the principal sources of the
potassium increments of plasma. At the lower tem-
peratures (47 C in vivo and 4S C in vitro) hemolysis
was negligible. The increase in plasma potassium
in vivo at these tcnqKTaturcs was due either to dif-
fusion from extra vascular sources or to leakage from
erythrocytes. It was obvious in the lower-temper-
ature in vitro experiments that leakage from erythro-
cytes was the only source, of the plasma increment.
Although leakage alone could be sufficient to account
for potentially fatal plasma levels (in excess of
16 milliequiv 1), no such increases were observed
without accompanying hemolysis. When blood was
heated in vitro leakage contributed more than hemol-
ysis to the attainment of such levels. In thermal
exposures in vivo of sufficient duration and intensity
to produce comparable levels, hemolysis was the
more important factor. .
I7.il PHYSIOLOGICAL DISTURRANCKS
FROM KXCKSSIVE I1K\TJ
17.11.1 Introduction
9
In Sect ion 17.9 of 1 his chapter, at ten tion was callci 1
to the fact that acute hyperthermic circulatory fail-
ure in some animals was accompanied by, and un-
doubtedly contributed to by, large increases in the
potassium concentration of the plasma. An investi-
gation of (he circumstances in, and the sources from
which, thermally induced rises in plasma potassium
occur has been described in Section 17.10.
Although it appeared that central circulatory fail-
ure caused by hyperpot assemia was one of t he mech-
anisms responsible for death incident to cutaneous
exposure to heat, it was apparent that this was not
the sole cause of death during hyperthermia. The
following investigations2 were undertaken for the
purpose of determining the precise nature of the
various kinds of circulatory disturbances which may
result from cutaneous exposure to excessive heat.
The acute physiological disturbances caused by
systemic hyperthermia have at tracted the attention
of a number of investigators. 1 leymans 26 injected
methylene blue into dogs anesthetized with chloral-
ose. This produced a gradually mounting rectal tem-
perature which reached the lethal level of 43.7 to
44.8 C in 1 to 1} _> hours. The heart rate rose gradu-
ally from 90 120 to 300-330 per minute. At first the
respirations were deep and rapid (less than 200 per
minute); after the temperature had risen to 41.5-
43.5 C they became very shallow and even more
rapid (over 300 per minute). Systolic pressure rose
and diastolic pressure fell. Respiration almost always
failed first, and artificial respiration enabled the
J By Alticrl Rons.
SECRET PHYSIOLOGIC AL DISTURBANCES FROM EXCESSIVE MEAT
heart to continue for a longer time. Reflexes per-
sisted up to the time of respiratory standstill. IJveno45
produced hyperthermia in eats, anesthetized with
urethane, by exposing them to water of 11-42 C or
to a high environmental air temperature. During the
30 minutes of exposure the rectal temperature rose
from 35 to 39 C. There was little increase in heart-
rate, but a pronounced rise in minute-volume out-
put, Shortly after exposure the respiratory rate in-
creased to an average of 2tH) per minute. This breath-
ing was very shallow (tidal air 2 3 ee per minute)
and sometimes resulted in a 29 per cent, drop in
arterial oxygen saturation, (’beer11 placed dogs
anesthetized with morphine and barbital in a cabinet
heated hy electric light bulbs. In 2 to 3 hours a lethal
(rectal?) temperature of 43 15 C was reached. The
heart rate increased progressively unt il a temperature
of 42 1IC wjls reached, when the heart slowed
rather suddenly. Before this stage electrocardio-
graphic abnormalities were limited to slight abbrevi-
ation of the 1*1! interval, slight changes in the QRS
complex, and Inversion of the T wave. The terminal
bradycardia was due to the development either of
nodal rhythm or of various other types of ventricular
rhythm. Systolic and diastolic pressures remained
fairly constant up to 41 C, then both dropped;"the
former more than the latter. The respiratory rate
also increased. Respiratory standstill usually oc-
curred before cardiac arrest, vagotomy delaying
respiratory failure. A progressive decrease of the
blood carbon dioxide was found associated with
slight alkalosis and rise of oxygen content, which
were ascribed to the increased pulmonary ventila-
tion. From the same laboratory, Wiggers and Orias 47
reported observations on the effects of short radio
waves on dogs. The cardiac acceleration, increase in
rate and depth of 1 lie respiration, and primary fail-
ure of the respiration were identical with the findings
of Cheer.13 However, instead of a decrease in blood
pressure, a rise of systolic and diastolic pressure was
observed which progressed until death.
Clinical observations on the effect of hyperthermia
were made by Ferris et al,15 Patients with heat stroke
whose rectal temperatures varied from 39.9 to 11.0 C
exhibited a hot dry skin, a normal or elevated sys-
tolic pressure, which dropped to low levels only in
the terminal stage, and venous pressures of from 2 to
12 cm of saline. Their respiratory rate was 28 to 50
per minute. Of 29 patients (all comatose) whose
temperatures exceeded 41.5 C, 17 died; all others
recovered.
Attempts to analyze the disturbances observed in
the intact organism by elevating the temperature in
one organ have been made since 1872. Fiek 16 heated
the bio nl as it passed through the carotid arteries of
the dog and noticed marked hyperpnea without
change in heart rate or blood pressure. Cyon 14 iso-
lated the circulation of a dog’s head. Perfusion of the
head with heated blood produced bradycardia and a
drop in blood pressure. Kahn 2* warmed the carotid
arteries of unanesthetized dogs without producing a
rise in rectal temperature. He observed the develop-
ment of tachycardia and a moderate rise in blood
pressure. Moorhouse M heated the carotids and simul-
taneously cooled the jugular veins in dogs. Phis re-
sulted in tachycardia, rarely preceded by brady-
cardia, ascribed respectively to increased sympa-
thetic and vagal activity. Coincidentally, tachypnea
and peripheral vasodilatation were observed. liey-
mans and Ladon severed all connections except the
vagal nerves between head and trunk of dogs anes-
thetized with chloralose. Artificial respiration was
applied and the circulation in the head maintained
by connecting it to a donor dog. The sublingual tem-
perature of the preparation rose to to C in 1J o hours.
There was no change in the heart rate which had
risen to I GO after severance of the cervical cord. The
head exhibited a progressive and pronounced in-
crease in respiratory rate which persisted until a
sublingual temperature of 15 C was reached, when
the rate rapidly decreased and the reflexes of the
head, which had been active up to that time, dis-
appeared. .
The effect of hyperthermia on the heart was in-
vestigated by Kuowlton and Starling,*1 using the
innervated heart-lung preparation perfused with
heated blood. From 2G C t o approximately 45 C the
heart rate was a linear function of the blood temper-
ature, the rate at 45 C being 180 per minute. Above
this temperature marked slowing occurred and the
heart soon stopped. Arrhythmias occurred above
40 C.
To summarize these data, it can be said that in the
dog the highest rectal temperature compatible with
life lies between 43 and 15 C, when this temperature
is reached in I to 3 hours. Respiratory failure often
seems to precede circulatory failure. Tachypnea,
tachycardia, and peripheral vasodilatation seem to
be, in part at least, of cerebral origin.
The physiological changes of rapidly developing
hyperthermia leading to death within half an hour
have not been heretofore studied. As high environ-
SECRET 370
STUDIES OF THERMAL INJURY — CUTANEOUS AND SYSTEMIC
Tabi.k 23. Rectal temperature, arterial pressure, and electrocardiogram of 12 pigs immersed in hot water.
A - Normal sinus rhythm (normal rate, tachycardia, or bradycardia).
Normal duration of QRS complex.
A' — First or second degree A V block. Normal duration of QHS complex.
A" — Complete A V block. Normal duration of QliS complex.
H — Slight
BB Moderate Widening of QRS complex without P wave.
HUB — Pronounced j
BBB -- Can often be interpreted as ventricular fibrillation.
Time
rnin sec
Rectal
temp
C
Arterial
pressure
(mm Hg) F.CG
Rectal Arterial
Time temp pressure
min sec C (mm Hg)
KCCi
Pig 876 (7.7 kg) 48 C. Died after 26.5 min.
Control 34.3 118 A
1G .. 44.0 GG A
24 :t0 45.2 42 A
26 30 45.7 7G BB
Pig 875 (6.4 kg) 48 50 C. Died after 35 min.
Control 35.0 130 A
27 30 42.2 64 A
29 .. 42.8 64 A
34 15 43.7 26
Pig 878 (12,0 kg) 47 C. Died after 50 min.
Control ... 410 A
29 .. ~ ... 70 A
37 20 ... ■ 50 A
49 .30 44.9 30 A
Pig 879 (11.8 kg) 44-47 C. Died after 106.5 min.
Control 36.8 106 A
33 .. 43.1 5-1 A
Out of hot bath* from 33.5 to 48.5 min.
49 53 420 116
79 30 44.1 86 A
105 .. 44.5 14 A
Pig 895 (18.0 kg) 49 C. Curare. Died after 32 min.
Control 37.8 148 A
15 .. 41.9 ~*172 A
25 30 43.7 76 A
31 30 44.0 10 A'f
Pig 943 (8.3 kg) 47 C. Curare. Died after 36 min.
Control 37.7 126 A
17 .. 42.6 126 A
29 .. 44.5 136 A
Pig S!17 (16.4 kg) 47 C. Curare. Died after 56 min.
Control 37.9 — 146
24 .. 43.5 146 A
47 .. 44.0 90 A
55 ., ... 36 A'
Pig 946 (9.5 kg) 47 C for 23 min. Curare. Died after 42 min.
Control 40.1 82 A
17 .. 43.0 — 120 A
26 .. 44.4 40 A
34 30 44.6 26 A
Pig 944 (10.4 kg) 47 C for 25 min. I>icd after 99 min,—
Control 38.1 108 A
14 .. 43.5 120 A
26 ,r~ 45.4 100 A
37 ... 44.1 90 A
Pig 867 (7.3 kg) 64 65 C. Died after 15 min.
Control ... 146 A
5 30 ... 72 A
10 30 ... 72 — A
15 .. 46.0 12 _ BB
Pig 872 (7.3 kg) 64 65 C. Died after 11 min.
Control 150 A
7 ... 50 A
10 30 ,.. 50 BB
10 45 ... 40 BBB
Pig 871 (9.1 kg) 70 73 C. Died after 12 min.
Control ... 100 ...
5 30 ... - - 74 A
6 10 ... 74 BB
9 30 ... 54 BBB
12 .. 44.5 24 BBB
* Skin temperature lowc
t Occasional ventricular
red hy exposure to cool water between two cpisod
extra-systole.
s of cutaneous hyperthermia.
mental temperatures am needed for such experi-
ments, the results are necessarily complicated by the
damaging effect of heat, on the skin directly. More-
over, these high temperatures will produce damage
to the red blood cells that are circulating in the small
vessels of skin and underlying (issues.42
iT.li.2 Experimental Procedure
Young pigs weighing from (>.4 to 18 kg and adult
dogs weighing from 7.1 to 8.5 kg were used as ex-
perimental animals. They were anesthetized with
pentobarbital sodium (32 mg kg intraperitoneally),
shaved, and tied to a wooden animal board. This
was lowered into a galvanized iron tank (92x10x41
cm). 4'he head of the hoard rested on a metal bar"
in the tank, so that it was slightly higher than the
loot. A similar tank, placed on a high table, partly
projected over Ihe former. This tank was filler! with
water steam-heated to the desired temperature. In
the bottom of the projecting part was a circular
opening 13 cm in diameter that could be closed
with a heavy rubber and metal stopper, resulting
in full immersion in 8 to 10 seconds. During im-
mersion, the temperature ot the water, which was
continuously stirred, was kept within narrow limits
by intermittent introduction of steam. Drainage of
SECRET PHYSIOLOGICAL DISTURBANCES FROM EXCESSIVE HEAT
371
Table 24.
Rectal temperature, arterial pressure, electrocardiogram, hematocrit, and hemoglobin and potassium content
of plasma
md of red bio
id cells of 15 pigs immersed in hot water.
A
— Normal sinus rhythm (normal rate, tachycardia or
bradycardia).
Normal
duration of QitS complex.
A
First or second degree A- V block
. Normal duration
of QRS complex.
A
"— Complete A-V
block. Normal duration of QRS complex.
B
— Slight
)
BB — Moderate
Widening of QRS complex without P wave.
HUB — Pronounced 1
BBB —- Can
>ften lx
interpreted as v
entrieular fibrillation.
Rectal
Rectal
Time
temp
Arterial
K plasma
Time
temp
Arterial
K plasma
min sec
C
pressure
i:cc.
milliequiv/1
min see
C
pressure
KCG
millicquiv/1
Pi a
S77 (7.0 kg) 47 C. Died after 26
min.
Pig 905 (12
7 kg) 75
C. Curare.
Died after
23 min.
Control
34.3
96
A
3.8
Control
. . .
94
A
1.8
10 20
41.6
136
A
6.2
16 30
41.6
78
BB_
11 5
42.5-
112
A"
6.9
22 40
42!
32
BBB
17.3
24 10
44.3
56
A"
8.2
Pig 921 (16.8 kg) 75
C. Curare.
Died after
27 min.
Pig 923 (13.6 kg) 47 C. Died after 50 min.
C’ont rol
122
A
3 2
Control
116
A
3.8
3 30
66
A
5.1
13 15
146
A
5.5
8
58
BB
11.6
22 30
146
A
5.5
IS
36
B
11.9
34 15
102
A
6.2
26 45
28
B
10.2
42
56
A
A
6.5
Pig 906 (13.0 kg) 70-7.
C. ('urare. Died aftr
r 70 min.
40 33
66
— 7.5
Control
38.6
102
A
4.0
Pit?
057 (8.0 kg) 17 C. Died after 36
5 min.
10 50
41.4
112
BBB
Control
37.0
.. t
A
4.4
16 35
42.3
62
BB
17.4
19 50
A
7.0
25 20
43.0
92.
BBB
15.2
30 15
: T., ■
A
10.2
44 35
44.6
72
BB
13.3
30 30
45.5
O
46 40
44.8
16
A
Pin 1056 ( 7.0 kg) 47 C. Died
ifter 44.5 min.
48 29
45.0
46
BB
Control
37.8
A
4.7
65
46.8
46
BBB
9 30
A
5.9
Pig 913 (8.2 kg) 75 C
or 6.5 rnin
Died after 7.5 min.
15 7
.
A
7-2
Control
38.6
1 CM)
A
3,5
34
A
7.1
2 25
37.9
100
B
14.2
14 30
45.5
O
6 15
40.5
50
BBB
17.7
Pic 910 (9.5 kgi 72 75 C. Died after 12,5 min.
7 45
40.8
15
0
17.4
Control
36.8
148
A
3.0
Pig 919 (9.1 kg) 75 C
for 5 min.
Died after IS min.
2 15
40.7
100
A
19.1
Control
37.1
138
A
4.2
4 40
10,7
86
BB
18.1
I 15
41.1
78
BB
25.5
7 20
41.5
71
BBB
24.0
7 15
42.3
28
A
21.4
13 52
43.7
10
O
17.3
10 10
43.2
26
A
18.3
Pie 912 (10.0 kg) 7
2-75 C. Died after
14 min.
14
44.2
30
B
17.0
Control
36.0
88
A
4.1
16 45
14.3
14
B
17.5
1 20
35.4
154
A
16.7
Pig 918 (8.7
kg) 75 C for 3 rain.
Died after
55 min.
3 35
37.0
'.18
BB
Control
36.6
70
A
3.7
5 7
37.1
74
BB
16.4
4 25
38.7
56
A
11.0
9 45
40.8
74
BBB
16.4
II
39.7
62
A
9.5
13 10
43.1
30
BBB
17 5
40.3
70
A
9.5
Pig 908 (9.1 kg) 75 C. Died :
ifter 13.5 min.
37
40.6
70
A
9.4
Control
96
A
3.8
Pig 899 (13.6 kg) 75 Cf
>r 1 min. Sacrificed after 77 min.
3 40
96
BB
16.7
Control
37.4
142
A
3.6
S 55
. . .
60
BBB
18,5
5 15
40.5
30
A
10.2
11 10
52
BBB
17.1
16 5
40.5
76
A
6.9
Pig
907 (10.4 kg) 75 C. Died after 10 min.
45 45
40.3
76
A
A
4,2
Control
37.1
37.3*
116
A
3.5
76
39.2
76
7.4
6
39.0
42.7*
48
BBB
_
7 30
39.2
42.5*
32
BBB
17.4
♦ High! heart temperature.
SECRET 372
STUDIES OF THERMAL INJ LKY — CUTANEOUS AND SYSTEMIC
(he water and termination of exposure could also be
accomplished in S to 10 seconds. Temperatures rang-
ing from I 1 to 75 C were used.
Previous to exposure, all animals were heparinized
(3 mg kg intravenously). Because ol spasmodic
closure of the glottis on immersion, a tracheal can-
nula was inserted. The carotid pressure was recorded
with a mercury manometer. The rigid auricular pres-
sure was measured by means of a rubber catheter
introduced into the superior vena cava or right
auricle by way of the external jugular vein and con-
nected with a water manometer. The level of the
right auricle as determined by opening (he chest at
the end of the experiment was taken as the point of
reference. In pigs the hydrostatic pressure did not
influence the auricular pressure. In dogs immersion
resulted in- a considerable rise in recorded auricular
pressure, so t hat only changes occurring during ex-
posure could be compared Pneumograms were ob-
tained by means of a copper cannula thrust between
the ribs Into the pleural space and connected by
means of a rubber tube to a writing tambour. In
other experiments, a tracheal cannula provided with
a sealed-in side tube connected to the tambour was
used. Electrocardiograms were taken with an ampli-
fier type of electrocardiograph. It was only possible
to take the first standard lead, as the hind legs of the
animal were underwater. In some experiments, cu-
rarized animals were used and artificial respiration
was applied throughout the exposure. Intocostrin
(Squibb) 1 mg kg diluted with saline was slowly in-
jected intravenously. The side reactions were limited
to a short (20 to 30 seconds) period of mild excitation.
The drug had no effect on the arterial pressure. A
second smaller dose usually had to be given 20 to
10 minutes later, A Palmer respiration pump for
small animals, which allows the air to escape spon-
taneously on expiration, was used. When venous
pressures were recorded the animals were immersed
in such a manner that most of the anterior thorax
remained above the water level. This was sufficient
to abolish artifacts produced by the increased re-
sistance to the inflow of air. Temperatures were re-
con led with a thermocouple introduced to a depth
of 7 to 9 cm into the rectum, \Vhich had been cleaned
by repeated enemas. The anus was closed around the
couple. In three experiments, heart temperatures
were also recorded by means of a thermocouple in-
troduced through the external jugular vein into the
right auricle. In some experiments only initial and
final rectal and final heart temperatures were meas-
ured with a sensitive thermometer. In a considerable
number of animals blood was withdrawn from the
jugular vein both before and during exposure for the
determination of the hematocrit and of hemoglobin
and potassium content of red cells and plasma (Sec-
tion 17.10). In most instances, immersion was con-
tinued until death. In some experiments exposure
was temporarily interrupted, and, in a few cases, im-
mersion was terminated at a time when the animal
was still living.
In addition to these observations, three pigs were
infused with an isotonic (1.12 per cent) solution of
KC1. Frequent electrocardiograms (lead I or 111
wen* taken. In one ol these pigs, the arterial and
right auricular pressure and respirations were also
recorded. The latter animal received the solution in
the subclavian vein,.the other two in the jugular
vein. Blood samples for the determination of potas-
sium were taken from the carotid artery.
17.11.3 Results of Experiments
In Table 2d are shown the results of 12 experiments
in which pigs were exposed for varying periods of
time at temperatures ranging between II and 73 C.
Changes in rectal temperature, arterial pressure, and
electrocardiogram arc indicated.
In Table 21 are shown the resnltsof id experiments
in which pigs were exposed at temperatures ranging
between 47 and 75 C. The changes that occurred in
(he potassium concentration of the plasma are indi-
cated in relation to changes in rectal temperature,
arterial pressure, and electrocardiogram.
In Table 25 are shown the results of 5 experiments
in which dogs were exposed for varying periods of
lime at temperatures ranging between 55 and 75 C.
The changes that occurred in the potassium concen-
tration of the plasma are indicated in relation to
changes in rectal temperature, arterial pressure, and
electrocardiogram.
In Table 2b are shown the results of 3 experiments
in which pigs received intravenous infusions of
isotonic potassium chloride. The changes in the
plasma concentration of the plasma and the erythro-
cytes are indicated in relation to changes in hemato-
crit, arterial pressure, and electrocardiogram.
Arterial blood pressure. The immediate effect of
immersion in water of 00-75 G n[M»n the mean ar-
terial pressure of pigs was a rise which sometimes
amounted to as much as I 10 mm Hg. This rise also
occurred in curarized animals or when hot water was
SECRET PHYSIOLOGICAL DISTURBANCES FROM EXCESSIVE HEAT
Fin cub 34. Plot of thermocouple recordings showing rate of change in rectal and right auricular blood temperatures
during immersion in low (47 (') and high (73 C) temperature water Paths.
47 C Pig 882 (13.2 kg) 75 C Pig 1)07 (10.5 kg)
It may l>e seen that, although right auricular blood temperature rises rapidly after immersion, there is considerable lag in
temperature rise in rectum. The higher the temperature of the bath, the greater is the difference between the two.
splashed on the skin. It was absent at immersion
leni|)eratines of 45-47 C.
At temperatures of 44-59 C the blood pressure was
maintained at or above preimmersion level for 16 to
26 minutes. It began to fall at variable times during
exposure, and reached half of the original value in
17.5 41 minutes. The rectal temperature at this time
had risen from 31.3 40.1 C to 12-14 C. These ani-
mals died after 25.5 to 50 minutes with rectal temper-
atures of 43.9—15.8 C, the heavier pigs surviving
somewhat longer than the lighter ones. Heart tem-
peratures were within a few tenths of a degree of
these values (Figure 34).
In pigs exposed to water of 60-75 C, the arterial
pressure was maintained for 1 6 minutes, and reached
half of its original value in 5.5-11 minutes. The ani-
mals died after S-15 minutes with rectal tempera-
tures varying from 39.4-46.0 C. However, the dis-
crepancy between heart and rectal temperature often
was considerable (Figure 34).
The possible reversibility of the fall in arterial
pressure was investigated. Immersion of a pig at
17 (' for 33 minutes produced a fall in blood pressure
from 104 to 40 mm Ilg (Figure 35). Exposure to cool
water brought the pressure back to its original level
and lowered the rectal temperature from 43.3 to
42.0 C. Re-exposure to 47 C again resulted in a fall
in blood pressure, and death occurred at a rectal
Tap us 25.
Rectal
1 temperature, arterial pressure, electro-
ran
liogram
, hematocrit, and hemoglobin
and
potassium
content of
plasma and of
red blood cell.'
s of i
i dogs im-
merged in hot water.
A-
- Xormal sinus rhythm
(normal rate,
tachycardia or
hradv
cardia)
. Nonna
1 duration of i
QRS
complex.
B-
- Slight
widen
ing of QNS complex wit
hont
F wave.
Rectal
Arterial
rime
temp
pressure
K plasma
niir
i sec
C
mm 1 Ig 1
;t (;
millioquiv/1
Dog 031 (7.4
kg) 55
(’. Died after 23 min.
C
ontrol
55.4
112
A
2.8
5
10
37.0
02
A
5.2
13
15
40.0
58
A
4.7
20
45
41.4
IS
A
0.0
Dog !1
i30 (7.5
kg) 00
C. Died
after 10.5 min
C
ontrol
30.0
1(X)
A
4.0
4
45
37.4
80
A
3.3
7
55
38.0
04
A
4.7
10
40
30.1
00
A
5.3
Dog 022 (8.5
kg) 75
C. Died
after 15 min.
C
ontrol
37.0
118
A
3.1
2
55
37.0
00
A
5.8
6
30
38.4
08
A
6.4
It)
20
30.0
70
A
5.8
15
30.3
30
A
0.8
Dog 1)
20 (8,2
kg ) 75
C. Died
after 13.5 min
c
ontrol
37.2
130
A
3.0
3
10
38.5
130
A
4.8
8
30
42.1
120
A
6.1
12
45
44.1
74
B
8.2
Dog 9
34 (7.0
kg) 75
C. Died
after 25 min.
Control
34.0*
148
A
3.1
15
16
41.7*
100
A
24
45
43.5*
72
A
0.0
♦ Right heart
temperature.
SECRET 374
STUDIES OF THERMAL INJURY—CUTANEOUS VM> SYSTEMIC
Figure 35. Effect of two episodes of cutaneous hyperthermia on pig 870 (11.8 kg) caused by immersion in water at
47 C. First period of immersion lasted for 33.3 minutes and is indicated by words on first and second
segments of kymograph record. Fifteen minutes after end of first period of hot water immersion and bet ween second and
third segments of record, animal was immersed again at 47 C and allowed to remain in hath until dead (56.5 minutes).
Between two episodes of hot water immersion, skin temperature was lowered by exposure to cool water. Total duration
of experiment was 105 minutes, t'pjier, middle, and lower tracings on the kymograph record represent resj>cctively
pneumogram, carotid pressure, and right auricular pressure. The numlicrs under the electrocardiograms correspond to
those under the kymograph tracings; C = control period.
temperature of 44,5 C. In another instance exposure
to water of 75 C for I minute reduced the pressure
from 140 to 20 mm Hg in 5 minutes. During subse-
quent exposure to room air the pressure recovered,
and reached 130 mm Hg after 73 minutes. The ani-
mal was still alive after more than 2 hours. Exposure
of one animal to water of 75 C for 5 minutes resulted
in a fall in blood pressure from 138 to 78 immedi-
ately after immersion. The pressure continued to
fall, and the animal died after 18 minutes.
The arterial pressure in dogs behaved in a way
comparable with that in pigs at the same tempera-
ture. Animals immersed at 60-75 C survived for
13.6 25 minutes.
Right auricular pressure; Intra-auricular pres-
sures of pigs before immersion varied from +32 to
— 66 mm H;0 (average —23 mm IbO). In only three
out of fifteen animals was the pressure in the right
auricle higher than atmospheric (+13, 20, and
32 mm 1FO). In most instances, a slight rise occurred
following immersion, the control level being regained
in 0.5 to 3 minutes. In five of the six animals im-
mersed at 4-1—19C, this was followed by a gradual
drop of 4-20 mm H20. There was no rise in venous
pressure until 1 or 2 minutes before death. In the
sixth pig, immersion did not influence the auricular
pressure (Figure 35).
In seven of the nine pigs exposed to water of GO-
75 C, a gradual rise of the right auricular pressure
was observed, beginning in the middle of or even
early in exposure and continuing until death. Tins
rise amounted to 15-45 mm IT.O and occurred at a
time when both arterial pressure and respiration
were still adequate (Figure 36). In some instances, it
SECRET PHYSIOLOGICAL DISTL’RRAXCES FROM EXCESSIVE HEAT
375
Kiouhk 36. Effect on pig H71 (9.1 kg) of immersion in water bath at 70-73 (' for 12 minutes, Upper, middle, and
lower tracings on kymograph record represent respectively pneumogram, carotid pressure, and right auricular pressure.
Sequence in which electrocardiograms were taken is indicated.
was preceded by a fall of 20-30 mm H20 which rap-
idly developed 1-3 minutes after the exposure had
started. In two animals, this fall was the only change
in auricular pressure that was observed until 1 min-
ute before death, when it rapidly rose.
One pig, exposed for only 1 minute to water of
75 C, showed an abrupt fall of 40 mm H20. During
the following 70 minutes the auricular pressure grad-
ually returned to the preimmersion level, coinci-
dentally with recovery of the arterial pressure.
The auricular pressure of four dogs was lower than
that of the pigs. It ranged from —77 to —108 mm
II2(). Because of hydrostatic effects the auricular
pressures Indore and during immersion could not be
compared. However, neither in the two dogs exposed
to 73 C nor in those exposed to 55 and 00 C w as there
observed any change in the recorded auricular pres-
sure during the period of immersion.
Because of the possible contributions of the type
or rate of breathing to the observed pressure changes,
some experiments were, performed on eurarized pigs.
Artificial respiration was applied throughout the ex-
periments. The course of the auricular pressure was
found to be identical with that of the spontaneously
breathing animals. At 17-49 C a slow and moderate
fall was observed; exposure at 75 C resulted in a rise,
beginning early during exposure.
Respiration; In agreement with earlier writers it
was found that a rise in body temperature was asso-
ciated with a pronounced increase in respiratory
rate. In the pig the immediate effect of immersion
was usually a short period of very deep and fairly
rapid respirations, followed by a variable episode of
only moderately increased breathing Irate 20-40).
In the animals exposed to the lower temperature
range the onset of respiratory rates of 170-200 was
often sudden, and occurred in t he first 10 minutes of
exposure, at rectal temperatures of 39-41 C. Deep
gasps interrupted this shallow tachypnea. The ar-
terial blood maintained its bright red color. The
tachypnea gradually increased, and rates of 300 were
not infrequently reached. When the rectal temper-
ature had mounted to 13-44 C, breathing abruptly
slowed to 10 40 per minute and became much
deeper. Additional slowing usually continued until
death. In the dog, immersion was immediately fol-
lowed by a tachypnea of 100 150 per minute, which
gradually increased. Rates over 200 were not en-
countered.
It is difficult to estimate whether the respiratory
SECRET STL DIES OF THERMAL INJURY — CUTANEOUS VNL) SYSTEMIC
Table 26.
Physiological and
chemical changes in three pigs intravenously infused
with an
isotonic (1.
2%) solution of KC1.
A — Normal sinus rhythm (normal rate,
tachycardia or
bradycardia).
Normal duration of QKS complex.
B — Slight
]
HU — Moderate
) Widening of Qh'S complex without P wave
—
BBB — Pronounced 1
BBB — Can often be interpreted as ventricular fibrillation.
Arterial
Time
pressure
K plasma
K cells
min
sec
mm Hg ECG
Hematocrit
milliequiv/l milliequiv/l
Pig 901 (14.8 kg)
Bate of infusion 0.6 cc/kg min. Died after 50 min.
font r«l
A* (lead 1)
36
4.3
123
11
CHI
A*
36
9.0
125
16
(Ml
A*
37
9.5
124
IS
00
BB
38
11.2
121
26
00
BBB
37
15.5
132
Infusion stopjsed
-I
26
10
O
...
35
00
O
36
00
A
_ 11
00
A
38
8.7
139
41
30
Infusion started again. Bate 0.7 ee kg min.
50
00
BBB
35
17.7
136
Pig 911 (8.7 kg).
Bate of infusion 0.9 cc/kg/min. Died after 22.5 min.
Control
A (lead II)
35
3.2
127
11
00
Af
34
8.7
122
14
00
B
35
lO.ti
122
16
00
BBB
35
12.7
125
20
00
BBB
31
27.0
22
00
0
28
38.0
127
Pig 925 (15.9 kg).
Bate of infusion 0.6 cc/kg/min. Died
ifter 39 min.
Control
76 A (lead I!)
33
3.5
112
6
08
76 A
33
5.7
117
12
40
76 A
32
10.6
111
19
37
76 At
34
12.7
110
24
50
76 B
37
15.7
10!)
35
18
24 BBB
37
26.1
Ill
♦ P wave not clearly shown.
t P wave getting blunt.
X P wave very flat.
or the circulatory system failed first in these animals.
If hradypnea is considered as the first manifestation
of failing respiration it might he said that the cardio-
vascular system survived somewhat longer, as judged
by the presence of an appreciable arterial blood pres-
sure. However, at least in the beginning of hradyp-
nea, the pulmonary ventilation certainly was as ade-
quate as during the control period. If the onset of
prolonged apnea is considered as the end point of
adequate respiratory function, l>ot h systems failed
simultaneously. In three animals, artificial respira-
tion was applied at a time when the arterial pressure
was st ill appreciable (80-1)0 mm Hg), without having
the slightest effect upon its downward course. More-
over, the final rectal and heart temperatures of the
curarized pigs fell well within the range of those of
spontaneously breathing animals.
Exposure of pigs to (>0 75 C produced an increase
in respiratory rate which did not exceed 80 00 per
minute. The breathing remained deep until the
terminal episode of hradypnea, ending in occasional
deep gasps. In dogs the respiratory changes were
essentially the same as those encountered at the
lower temperatures.
Electrocardiographic changes: In both pigs and
dogs, the first, change, beginning immediately after
immersion, consisted of a progressive increase in
heart rate to levels of 300 350 per minute. Associ-
ated with this increase, changes occurred in the (JRS
complex, consisting of decrease in amplitude of the
l{ wave and deepening of the S wave or vice versa
with maintenance of the normal QffS interval; and
inversion of the T wave. The changes in the initial
vent ricular deflection might in part at least be due to
SECRET PHYSIOLOGICAL DISTURBANCES FROM EXCESSIVE HEAT
377
Figcue 37. Helutionsliip hctwwn plasma potassium level and changes in electrocardiogram (lead 1) during immersion of
pig 010 (0.5 kg) in water bath at 72-75 C. Plasma potassium values are given in milliequiv 1. DeaIh occurred 12.5
minutes after beginning of experiment.
variations in typo of breat hing with resulting changes
in the position of the heart. (I (arris,2*) They occurred
only to a minor degree in curarized animals.
In the pig, the abnormalities following this sinus
tachycardia varied markedly with the temperature
of exposure. Of all animals exposed to water at 44-
50 C (Tables 23 an I 24) only one showed appreciable
widening of the QRS complex and loss of P wave.
This occurred I minute before death. Another animal
showed disappearance of the P waves.
The changes in the remaining pigs were limited to
sinus bradycardia and sinus arrhythmia, which be-
came most pronounced 2 or 3 minutes before death
(Figure 35). Occasionally, auricnloventricular block
of varying degree was seen during this period.
In contrast, eleven pigs continuously exposed to
temperatures of 64-75 C (Tables 23 and 24) all
showed the gradual development of exceedingly wide
ventricular complexes with very large T waves, and
the gradual disappearance of the P wave.k The gen-
eral shape of these complexes resembled that of the
original supraventricular ones. Their development
was usually associated with definite slowing, al-
though the heart rate remained regular, in some
eases, the transitional phase consisted of salvos of
fairly rapid and wide vent ricular complexes, which
interrupted a still-existent sinus bradycardia. In the
terminal stage, the initial ventricular deflection
could not Ik- separated from the final one. The elec-
trocardiogram consisted either of very slow, ex-
tremely wide ventricular waves, separated from each
other by isoelectric intervals of 0.2-1.0 second, or of
more rapid variations at 100-240 per minute, in
which one wave merged with the next. The latter
state might be called ventricular fibrillation (see
Figures 36 and 37).
In nine of the eleven pigs, these changes made their
first appearance early during immersion, at rectal
temperatures of 37.0 to 41.6 C and at a time when
the arterial pressure and respiration were still ade-
quate. In four of these, the blood pressure at the
k During tachycardia, actual ot enervation of this disap-
pearance was impossible because of overlapping of P and
preceding T waves. In these instances, it was assumed that
the same changes had taken place as in the instances where
the P wave could !«■ followed through it stage of decreasing
amplitude to disappearance, as subsequent slowing of the
teat similarly revealed the absence of auricular complexes.
SECRET 378
STI DIES OF THERMAL IMIRY — CUTANEOUS \M> SYSTEMIC
time of onset of the wide complexes was actually
equal to or higher than that before immersion. In
only two animals were the abnormalities first noticed
when the pressure had fallen to low levels, and it is
possible that they would have been demonstrated
earlier if more electrocardiograms had been taken.
Exposure for <1.5 and 5 minutes similarly resulted in
marked widening of the QKS complex, whereas ex-
posure for 3 minutes and 1 minute did not produce
deviations other than those at lower temperatures.
In the dog, the electrocardiographic changes at
high temperatiires were in no way different from
those encountered at 11-50 C (Table 25). They were
limited to an increase in rate and to minor changes in
the ventricular complex. No widening occurred and
the auricular manifestations remained present until
the end.
Chemical Changes. For a complete discussion of the
effect of temperature on the potassium concentra-
tion of the plasma, see Section 17.10 of this chapter.
The potassium concentration of the plasma of fifteen
pigs in which physiological studies were made are
shown in Table 24. 3’lie initial plasma levels ranged
between 3.0 and 4.8 milliequiv 1. The potassium con-
centration of the red blood cells ranged from 113 to
145 milliequiv/1. The course of these concentrations
during immersion varied markedly with the tem-
perature.
Immersion of four pigs at 47 C produced a gradual
and sustained rise in plasma potassium. Ten minutes
exposure resulted in levels of about 6.0 milliequiv/l.
During the rest of the exposure, the level increased
by an additional I to 4 milliequiv. The highest level
was 10.2 milliequiv 1 obtained 30 seconds before
death.
On the other hand, continuous exposure at 70
75 C characteristically resulted in an enormous rise
in the plasma potassium level. This increase was
found to take place with surprising rapidity. In five
pigs, tlu- plasma after I to 4 minutes of exposure con-
tained 1 1.2 to 25 5 millicquiv/1 of potassium. A sam-
ple drawn in this period from one curarized pig was
still essentially normal and the peak observed in this
animal was only 11.0 milliequiv. Peaks from 1G.7 to
25.5 milliequiv were observed in six pigs during ex-
posure. Curare did not prevent rises in this range in
two pigs; however, no early observations were made
on these animals. In some instances, the potassium
level fell toward the end. However, it remained
markedly elevated.
In some experiments, the exposure was terminated
before the animal had expired. Immersion for (>.5 and
5 minutes similarly resulted in a tremendous rise of
plasma potassium. At the time of death, the level
was still very high. Immersion for 3- and I-minute
periods produced a less pronounced increase; at the
time of death, the level was only 2 2.5 times the
normal one.
17.11.4 Discussion
These observations show that the physiological dis-
turbances leading to death in pigs exposed to water
at 4G 50 C are of a different nature from those en-
countered in animals exposed to temperatures of
GO 75 C,
In pigs immersed at the lower temperatures, tlu1
occurrence of a gradual fall in right auricular pres-
sure followed by a fall in mean arterial pressure indi-
cates a progressive decrease in venous return to the
heart. That this decrease, at least during a major
part of the exposure, was due to an increase in ca-
pacity of the peripheral vascular bed, rather than to
loss of intravascular fluid, is evident, from the fact
t hat the changes in circulat ory dynamics were found
to be reversible to a considerable degree. As the ex-
posure continued, the detrimental effects of the
heated blood upon the heart muscle were added to
the peripheral effects, and both factors undoubtedly
contributed to the lethal ending.
It is difficult to say whether cardiovascular failure
or respiratory insufficiency was the immediate cause
of death. Profound arterial hypotension and pro-
nounced bradypnea were usually encountered at the
same time. It can be said, however, that the mean
arterial pressure fell considerably before any impair-
ment in respiratory function was evident. Artificial
respiration applied at a time when the arterial pres-
sure was still appreciable had no effect upon its
downward course. Moreover, curarized pigs did not
survive longer than spontaneously breathing ani-
mals; all but one animal died after 25 to 51 minutes
of continuous immersion. The plasma potassium
level increased by GG-250 per cent; the highest level
found was 10.2 millieqiiiv/1. No profound changes in
cardiac function, as judged by the electrocardio-
gram, occurred. As will Ire shown, plasma potassium
levels up to 10 milliequiv I do not produce significant
changes in intraventricular conduction.
At immersion temperatures of GO-75 C, the pigs
survived for only 8 to 15 minutes. In the middle of
the exposure, or even earlier, at a time when the
respiration was still adequate and the mean arterial
SECRET PHYSIOLOGICAL DISTURBANCES FROM EXCESSIVE HE YT
379
Figure 38. Effect of continuous intravenous infusion of 1.12 percent KClat the rate of 0.6 kg/rain. Upper, middle, and
lower tracings on kymograph record represent respectively pneumograin, carotid pressure, and right auricular pressure.
Time in minutes is shown at base of record. Time at which blood samples were taken is indicated by symbols K-, Kj, A»,
and K ,. The times at which the sequence of electrocardiograms (k to z) were taken are indicated by arrows. See pig 925,
Table 26, for corresponding potassium levels.
pressure was still considerable, pronounced changes
in cardiovascular function made their appearance,
They consisted of a rise in right auricular pressure,
and electrocardiographic changes in (he form of dis-
appearance of the P wave and progressive widening
ot the QRS complex, often terminating in ventricular
SECRET STL DIES OF THERMAL INJURY—El TWEOTS AND SYSTEMIC
fibrillation. At the same time, the potassium concen-
tration of the plasma reached values of Hi 19 milli-
equiv 1. This was associated with a striking destruc-
tion of red blood cells.
These observations strongly suggest that (he hy-
perpotassemia was responsible for the disturbances
in cardiac mechanism and for the subsequent myo-
cardial failure evidenced by the rise in auricular
pressure. That t he damaging effects of a rising plasma
potassium level first of all manifest, themselves in the
heart is well known. In rabbits and dogs, the infusion
of a solution of a potassium salt produces a sequence
of electrocardiographic changes similar to those ob-
served in pigs during ex posuretohigh temperatures.a-4*
IT was found that an identical sequence of changes
takes place in infused pigs (Table 20). In two animals,
infusion rates were maintained that were likely to
produce death in approximately the same time as in
the burned pigs. It is evident (hat potassium levels
of less than 10 milliequiv, 1 failed to produce either
changes in the P wave or widening of the QPS com-
plex, just as was the case in burned pigs. Higher
levels resulted in a succession of changes which were
similar in all respects to those observed at high tem-
peratures (Table 24). In the one animal (Figure 38)
in which arterial and right auricular pressure and
respirations were recorded, the auricular pressure
began to rise 19 minutes after the infusion had
started. The potassium level was 12.7 millicqulvTTf
the P waves had begun to flatten 3 minutes before
and had disappeared. Three minutes later widening
of the QRS complex began. The arterial pressure
and respiration remained normal for another 10
minutes.1
That the cardiac changes due to the potassium ion
are reversible to a remarkable degree is clear from
experiment 901 (Table 26). The usual succession of
electrocardiographic changes was observed until,
some seconds after a potassium level of 15.5 milli-
equiv/l had been reached, the string shadow re-
mained resting. The infusion was stopped. No ('lee-
trie or auscultatory evidence of cardiac activity could
lx* demonstrated for the following 10 minutes, al-
though the animal continued to breathe at a very
slow rate. Then heart action returned and respira-
tion became more rapid. The electrocardiogram had
returned to normal. A plasma sample taken 5 min-
utes thereafter contained S.7 milllequb' I of potas-
sium. Infusion was started again, the well-known
changes were again observed, ami the pig died with
a potassium level of 17.7 milliecpiiv, 1.
The rapidity with which potassium is removed
from the plasma makts it imperative that the release
of the ion into the circulation he intensive enough and
he continued for a sufficiently long time to lead to
death. This actually occurs in the burned pigs. The
filtration of potassium often occurred at so rapid a
rate that there was a lag between the rise in potas-
sium and the electric changes. Thus, in pig 5)10 a
level of 19.0 milliequiv 1 was reached in 2 minutes,
whereas more than 4 minutes were required to pro-
duce the typical widening. Animals exposed to high
temperatures for only I or 3 minutes did not release
sufficient potassium to produce a characteristic effect
on the heart, whereas exposure for 0.5 minutes was
adequate in this respect. Kxposure to 75 C for 5 min-
utes resulted in a tremendous rise in potassium and
in electrocardiographic changes, but even here both
manifestations diminished in intensity during tin*
following 14 minutes.
Although it is clear that in pigs exposed to high
(00 to 75 C) temperatures the most striking physi-
ological disturbances are those which result from the
release of excessive amounts of potassium, cont inued
exposure results in a progressive and generalized rise
Tn body temperature which undoubtedly causes dis-
turbances other than those due to hyperpotassemia.
Thus, the peripheral and central factors that were
the cause of death at lower temperatures also come
into play at t hese high temperatures.
In order to evaluate the relative contributions of
red blood cells and fixed body cells to the increase in
plasma, potassium experiments were performed on
dogs (Table 25). Whereas the potassium concentra-
tion of (heir fixed cells is similar to that of the pig,
their red cells contain only small amounts. Immer-
sion at 75 C resulted in an intense hemolysis, but the
potassium level did not rise above tha't encountered
in pigs at 47 C and electrocardiographic changes
characteristic of hyperpotassemia were not seen.
The distribution of the potassium in human blood
is similar to that in pig’s blood, the potassium con-
centration of the red cells being approximately
110 milliequiv/1, that of the plasma approximately
4 5 millieqiiiv/l.*2-41 High plasma potassium levels
should therefore be expected in human beings in
whom a major part of the body surface has been ex-
posed to high environmental temperatures. Several
1 The rate of infusion was slow enough so lhat the rise in
venous pressure could not lie ascribed to the administration
of the isotonic salt solution per se.h
SECRET IMIVSIOEOEICA! DfSTl RIUNCES FROM EXCESSIVE HEAT
381
minutes of exposure would probably he required to
result in the very high levels encountered in these
experiments. It is also probable that, if the immedi-
ate effects of the exposure were survived, a markedly
elevated plasma potassium occurring immediately
folk »\\ ing the injury would fall within the next hour.
It should be remembered, of course, that a rise in
plasma potassium is a normal post-mortem phe-
nomenon.
I7.li.r> Summary
The re arc two principal mechanisms by which ex-
posure of the surface of the lxidy to excessive heat
may cause rapid circulatory failure and death.
In one, the systemic hyperthermia due to con-
duction of heat to the interior of the body by way of
the blood stream leads to a rapid and progressive de-
cline in blood pressure and failure of circulation due
to peripheral vascular collapse.
In the other, the circulatory failure is principally
central and is due to the effect on the heart of an ex-
cessively high concentration of potassium in the
plasma. Centred-circulatory failure is likely to occur
when the overheating of the skin and subcutaneous
tissue is so intense, prolonged, and generalized that
potassium js released from the erythrocytes with
such rapidity and in such large amounts sis to main-
tain plasma levels in excess of 11 milliequiv 1.
In the case of thermal exposures of low intensity,
peripheral circulatory failure may occur without suf-
ficient rise in tissue (and blood) temperature to cause
a functionally significant rise in plasma potassium.
When a thermal exposure has been of sufficient sever-
ity to cause fatal hyperpot assemia, the central circu-
latory effects are likely to I>e complicated by periph-
eral vascular collapse.
It is essential to the development of acute hyper-
thermic potassium poisoning that the erythrocytes
have a high original concentration of this element.
Thus, fatal hyperpot assemia, Hue to hyperthermia,
occurs in (he pig but not in the dog. Since man and
pig have similar potassium concentrations in erythro-
cytes, it is inferred that they are probably similarly
susceptible to the development of fatal hyperpo-
tassemia following cutaneous exposures to excessive
heat.
Although thermally induced respiratory disturb-
ances undoubtedly contribute to either type of
circulatory failure, maintenance of pulmonary ven-
tilation by artificial respiration does not prevent
death or cause significant prolongation of the sur-
vival period.
SECRET Chapter 18
MISCELLA\E01TS TOXICOLOGICAL STUDIES
Birdsey Remhaw
I8.i 1NTRODLCTION
Division' 9 has carried out, in its laboratories
operated for toxicological and immunological
studies on chemical warfare agents, a limited number
of investigations with materials which were not con-
sidered for use as war gases but whose toxicological
properties were for other reasons of interest to the
Army, Navy, or other National Defense Research
Committee [NDRC] divisions. In this chapter are
summarized the results of four such investigations:
(I) the pathological changes caused by prolonged ex-
posures to oil screening smokes, (2) the toxic effects
of gasoline fumes, (3) (he toxicity of Salcomine dusts,
and (1) the hypersensitivity and dermatitis caused
by hexanitrodiphenylamine and enemy explosives
containing it.
18.2 TOXICITY OF OIL SCREENING
SMOKES
With the development by NDRC Division 10 of
generators for the production of oil screening smokes,
the question arose whether personnel exposed for
prolonged periods in such smoke clouds (consisting
of fine droplets of unburned hydrocarbon oils) would
1)0 subjected to health hazards. Although no informa-
tion was available concerning the toxicity of oil
clouds for animals or man, there were on record
marly 200 cases of “lipid pneumonia” attributed to
aspiration of mineral oilA Inasmuch as lipid pneu-
monia may occur whenever an exogenous oil reaches
the pulmonary tissues and remains for a sufficient
lime to cause irritation, the possibility existed that
this potentially debilitating condition might result
from the inhalation of the screening smokes. At the
request of the Chemical Warfare Service, an experi-
ment was performed in which mice were exposed for
prolonged periods to clouds of atomized lubricating
oil;1 the continuing interest of the Service led to the
extension of the tests4 to include the exposure of
monkeys to clouds both of lubricating oil and of fog
oil standardized for use in the Langmuir-type gener-
ator. The results of these tests with animals afforded
no basis for supposing that prolonged exposures of
military personnel to oil screening smokes in the field
would In' dangerous. By now the actual use of oil
screening smokos in military operations has lieen ex-
tensive and no evidence has been forthcoming that,
health hazards are involved.
The experiments were pertormed with animals
kept for 100 days in a large closed chamber into
which for 30 minutes of every hour air containing oil
fog was passed at a rate of 0.8 chamber volume per
minute. In the experiments with lubricating oil (Penn
Oil, SAE Xo. 10), the nominal concentration was
132 gg I and the droplets varied in diameter from
about 0.3-1,5n; the mass median diameter was I I ju.
In (he ease of fog oil (Texas Company, S(JF No. 1
Oil) I lie analytical concentration was do i.
The death rate among the mice exposed to the
clouds of atomized lubricating oil was not signifi-
cantly different from that in the normal colony and
the animals showed no serious pathological changes
during or at the end of the exposure.* No free oil was
ever seen in the alveoli or bronchi, and chemical
analyses at the end of the 100 days revealed that
relatively little had accumulated, there being in the
lungs 1.05 mg per mouse (0. t per cent of the total
lung weight). Occasional oil-containing macrophages
could be seen after the experiment had been in prog-
ress for a week. These increased in number during
the first 35 days, after which time almost every
alveolus contained at least one such cell, but they
did not become significantly more numerous during
the subsequent-two-thirds of the exposure period.
The tracheo-bronchial lymph nodes of mice sacri-
ficed after 3 weeks in the chamber showed accumu-
lations of oil-containing macrophages, but there was
no reaction to them.
These essentially negative results with mice led to
repetition of the experiments with the Rhesus mon-
key— a species which, in terms of posture and size
of respiratory passages, more closely resembles man.4
Chemical analyses of the lungs of exposed animals
revealed a progressive accumulation of oil to a maxi-
mum of about 10 per cent of the dry weight, or 2 per
cent of (he wet weight, at the end of the 100 days;
approximately one-half of this accumulation had dis-
appeared a year after the start of the exposure.
Microscopic examination revealed some free oil, and
SECRET TO\H'lTV OF SU.COMIXK DUSTS
383
oil-laden macrophages were scattered throughout the
lung, in the alveoli, snbpleurally, and in the bronchial
and pleural lymphatics. However, little inflammatory
reaction attributable to the oil occurred, and subse-
quent to the exposure the fibroplastic reaction to the
remaining oil was slight. The one conspicuous extra-
pulmonary effect was loss ol hair during the pro-
longed exposure, and its subsequent regrow th.
In so far as the animal findings may be applied to
man, the failure of large amounts of oil to accumulate
and the absence ot severe acute and chronic reactions
make it improbable that significant pulmonary ef-
fects would bo produced by any exposures likely to
be encountered.
The results of exposure to fog oil (SCF No. 1)
were similar, with one important exception. Six of
seven monkeys died, apparently of starvation, dur-
ing or shortly after the termination of the exposure.
Examination of the stomachs revealed acute or hy-
pertrophic gastritis and, in those dying after the
greatest delays, the pict ure of an adenoma malignum
superimposed on hyperplastic gastritis. These serious
pathological changes are believed to have been in-
duced by carcinogenic agents present in ingested oil;
carcinogens have been found in petroleum oils 11 and
presumptive evidence compatible with their presence
in S(iF No. 1 oil was obtained. Thus, the possibility
exists that cancer might result from prolonged ex-
posure to oil smokes. However, it should be noted
that the monkeys, their surroundings, and their food
were continually covered with oil, and the animals
therefore undoubtedly ingested much oil in addition
to that w hich they breathed and swallowed.
l« :i TOXICITY OF GASOLINE FUMES
Early in 1911 reports were received that among the
individuals killed in flame thrower attacks upon en-
closed fortifications were some who had not sus-
tained severe burns. The Chemical Warfare Service
was interested in determining the cause of these
deaths and, as a small part of a larger program, re-
quested that the effects of short exposures to the
vapors from unburned flame thrower fuel be de-
termined.
As it did not prove feasible to set up toxic concen-
trations of vapor from thickened flame thrower fuel,
the tests5* were limited to experiments with un-
(hiekened gasoline compounded to meet Federal
Specification VV.M-504. The gasoline was atomized
into an airstream which was heated to vaporize the
droplets Indore passing to an animal chamber in
which the air temperature was about 35 C. The
/J(('t);.,i’s for 5-minute exposures of mice, rats, and
guinea pigs were very high, in (lie order of 3U0 mg I
{('I = 1,500,000 mg min m:t, analytical). This con-
centration was somewhat above the sat uration value
at the temperature of the chamber, and a dense cloud
formed. The mice and rats surviving the exposure
period remained narcotized for 10 minutes but ap-
peared normal within one-half hour; no gross patho-
logical changes were produced. The guinea pigs ex-
hibited rapid, shallow breathing with forced inspira-
tions; autopsies revealed bronehospasm and emphy-
sema. It is probable that the action of the gasoline
fumes on mice and rats was purely narcotic, and that
in guinea pigs this action was augmented by broneho-
spasm.
The above-mentioned concentration is above the
upper explosive limit for gasoline and could not be
built up in the presence of flame. There is no doubt,
on the other hand, that concentrations of gasoline
vapor rapidly lethal for man as well as animals can
be attained in closed spaces in the absence of flame,10
and there is no information as to whether or not
sensitivity to the vapors is markedly enhanced at
greatly elevated ambient temperatures.
18.1 TOXICITY OF SALCOM INF DUSTS
Karly in 1942 the success of NDRC Division 11
in developing Saleomine * oxygen generators for use
on shipboard and elsewhere led to the need for an
investigat ion of the possible industrial hazards which
might be involved in manufacturing and working
w ith this compound.
* Salicylaldchyde ethylenediimine col wit, known as Sal-
«nmne, has the following structure:
-° '> <3
I V - I
\ /• \ /
IU \ :• i!
I
ICjC » a,
This material has the property of absorbing oxygen (about
4 per cent by weight) when exposed to air, and of releasing the
absorb'd oxygen when heated. Since this Cycle may l>e re-
lated many times, it is possible to construct systems employ-
ing Salcomine for the separation of atmospheric oxygen. For
details the reader is referred to the Summary Technical
Report of NDRC Division 11, Section 11.1. No doubt the
Salcomine samples used in the toxicological studies were
partially oxygenated.
SECRET MISCELL V N EDITS TOMCOIOCICAL STL DIES
UNCLASSIFIED
Preliminary tests 2 revealed that Salcomine dust is
toxic upon inhalation and clearly indicated the neces-
sity of taking precautions to protect workers. Mice
exposed for several hours to the dust at nominal con-
centrations of 0.4 2.1 mg 1 (undoubtedly the actual,
or analytical, concentrations were much lower) fre-
quently died within Hi days. Autopsies revealed
many pathological changes attributable to the Sal-
comine: there was generalized degeneration and
localized necrosis of the epithelium of the trachea
and principal bronchi; the lungs were hyperemie and,
particularly in the peripheral portions of the lobules,
edematous; the thymus gland and lymph nodes con-
tained fragmenting lymphocytes in moderate num-
ber; and fat stains revealed fatty changes in the
liver.
In 1044 (he occurrence of a number of clinical eases
of poisoning, presumably-due to Salcomine dusts,
occurred *' and prompted further animal studies.''1’
The results left no doubt that Salcomine is both a
respiratory and a systemic poison and that precau-
tions must he taken against the inhalation of its dust.
A single exposure to a high concentration killed
guinea pigs immediately and mice after varying
latencies. The lungs of (he guinea pigs were markedly
distended with air and microscopic examination re-
vealed that the bronchi and bronchioles were strongly
constricted. Mice dying soon after such an exposure
exhibited no visible changes which would account for
death; those dying after 1 day or more exhibited a
diffuse pneumonitis, suppurative tracheobronchitis,
and occasionally jaundice and eoagulative necrosis
of the liver.
More important from (he standpoint of the health
hazard and for revealing the generalized toxic effects
of Salcomine are prolonged exposures to low concen-
trations. Accordingly, animals were exposed for
1 hour daily in a chamber to air which contained the
finer particles of Salcomine dust at concentrations in
the order of 100 ag h Three to six such exposures,
corresponding to a total dosage of about 20,000
mg min m:!, sufficed to kill approximately one-half of
the exposed, mice and rats;c rabbits probably wen*
not much more resistant, but guinea pigs proved to
bo considerably less sensitive. The mice, developed
a diffuse pneumonitis and t racheobronchitis, paren-
chymatous degeneration of the renal tubular epi-
thelium, jaundice, and liquefying eoagulative ne-
crosis of the liver. Autopsies of rats sacrificed daily
during the exposures revealed a gradually developing
diffuse pneumonitis and tracheobronchitis; focal
hepatitis with occasional necrosis also developed and
was followed by the appearance of intracellular fat;
parenchymatous degeneration of the. renal tubular
epithelium occurred, followed by the appearance of
severe fatty changes: and in the duodenum and
jejunum the epithelial cells of the mucosal glands
began to show vesiculation, swelling, and many
mitoses. These changes subsided after the exposures
were discontinued. Clinical pathological studies on
rats revealed an increase in the urinary output after
the first and subsequent exposures, the development
of a mucoid diarrhea, a rise in hemoglobin and red
cell count, and a 5 per cent loss in body weight. A
leucocytosis also developed during the exposures
and subsided rapidly upon their cessation.
b Medical examinations of eleven men* exposed to small
amounts of Salcomine dust revealed that the compound pro-
duced irritation of the eyes, nose, larynx, and bronchi. The
symptoms, which apjieared shortly after exposure and re-
sembled those of an upper respiratory infection, cleared up
after removal from exposure. Signs (xtssibly indicative of
mild systemic effects — muscular aches, nausea, and vomit-
ing — apjHXired after latency of 5-24 hours in some of the
subjects. In general the respiratory symptoms disappeared
within a day but the digestion was sometimes upset for 3
days. There may have been some slight cumulative effect , be-
cause it was reported that chronic exposure led to anemia,
lack of energy, and need for increased sleep. Xo permanent
effects were noted and it was concluded that, with reasonable
precautions including use of dust respirators, no marked
industrial hazard was involved. One ease with much more
severe systemic effects has l>een reported.5 An emergency
obliged the subject to work without a mask for a short jteriod
in an atmosphere laden with Salcomine dust. On the evening
of the exposure there were no pronounced symptoms other
than discomfort in breathing, but the following day abdominal
pains of sufficient severity to require hospitalization and treat-
ment with morphine developed. The subject had nausea,
vomiting, and a fever. A tentative diagnosis of acute duode-
nitis was made and his liver became progressively more en-
larged and lender. Tests jierformed 48 hours after admission
revealed definite liver damage. The liver condition with ac-
companying jaundice gradually improved but the abdominal
signs persisted. Penicillin was utilized. An exploratory lapa-
rotomy 2 months after exposure revealed a retroperitoneal
abscess in the left lower quadrant; this was removed, as was
a second similar abscess which formed on the right side
3 months later. It was suspected that other abscesses were
present deep in the liver tissue, but none required drainage.
Definite hardening of the liver due *o scar tissue persisted.
Attending physicians assumed that the inhaled and ingested
dust was responsible for the acute digestive disturbances
that followed exposure and led to disease of the liver, duo-
denum, and retrojieritoneal tissues. Compare results of ex-
[H-rimental animal exposures. _
c To illustrate the toxic potency of the dust it may be noted
that for O-hour ex|Misures the Ij(('1)m of mustard gas vapor for
mice is i,J(li\jmfnipi\ and for rats, 1.500 mg inin/m*.*
ON i'h AiNolj' >r UNCLASSIFIED
HYPERSENSITIVITY CYISED »Y HEX VNITRODIPHENYLAMI \E
18.3 11Y PKRSKNSITIVITY VND DERM V-
TITIS CAUSED HY HKWNITRO-
DlPIfENYLAMINE
In 1943 the Navy Department reported the occur-
rence of acute dermatitis in jrersonnel of both British
and United States armed forces who had come in
physical contact with enemy explosives containing
hexanitrodiphenylamine, and requested NDHC to
investigate the cause of the dermatitis and methods
for its prevention and treatment.
A survey revealed numerous statements in the
literature that hexanitrodiphenylamine is a powerful
dermal‘.tic agent, but the factual basis for this im-
pression proved to lie weak. Furthermore, dinitro-
chlorobenzene, a potent dermatitic and sensitizing
agent for both man and the guinea pig,11-1* is em-
ployed in the manufacture of hexanitrodiphenyla-
mine, and there is the possibility that this or other
intermediates or by-products may have been respon-
sible for those cases of dermatitis which have been
observed. At du Pont Company plants, which manu-
factured limited quantities of hexanitrodiphenyla-
mine in 1918 and 1910, care was taken to avoid ex-
posure to the substance mid no noteworthy or severe
cases of dermatitis occurred.
Inasmuch .as no samples of enemy explosives
known to have produced dermatitis were available,
the investigation 3 was confined to studies with a
highly purified laboratory-prepared sample of hexa-
nitrodiphenylamine and with a preparation from a
Japanese torpedo booster. Crystallographic analysis
revealed the latter to contain about 75 per cent hexa-
nitrodiphenylamine, about 25 percent trinitrotoluene
(TNT), and no dinicrochlorobenzene; if any minor
constituents were present, they totalled less than
I per cent.
After rigorous applications of both preparations
had failed to produce irritation or sensitization in
guinea pigs and swine, tests were carried out on inert.
Neither the pure material nor the Japanese explosive
proved to he a primary irritant when applied in hot
weather to skin of the forearm as a saturated solution
in acetone or as a powder covered by an occlusive
dressing. In 2 of 29 men treated with the purifier!
hexanitrodiphenylamine and in I of 31 treated with
the Japanese explosive, however, ‘‘Hareup” derma-
titis developed about a week after the second of two
applications. The dermatitis cleared up under simple
symptomatic treatment within 7 10 days. Patch
tests later showed that these 3 men had become
markedly hypersensitive, whereas the 57 other sub-
jects had not .
The residue from an incompletely detonated sam-
ple of hexanitrodiphenylamine was innocuous to
hypersensitive skin. Prolonged treatment with excess
potassium sulfide likewise rendered the explosive
harmless, but treatment of acetone solutions of it
with sodium hydrosulfite did not alter its ability to
cause inflammation of hypersensitive skin.
Although the findings indicated that with gross
contaminations of large numbers of men, instances of
skin reaction of varying degree are to l>e expected, it
was clear that hexanitrodiphenylamine is not a pri-
mary skin irritant and that it occupies a low position
among the skin-sensitizing substances. Practical pre-
ventive measures were considered to be avoidance of
unnecessary contact with the substance and, inas-
much as the reactions are delayed, use of an organic
solvent and soap and water as soon as possible after
contamination to remove the substance from the
skin and from objects with which the skin can come
in contact.