NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
is necessary to ascertain carefully how far the phenomenon is dependent on
mere solution of the bodies of erythrocytes or on this along with various degrees
of reduction in the respiratory property of hæmoglobin. Consequently, almost
all the experiments dealt with in the present instance included both the
microscopic examination of specimens of blood, which had been fixed at various
periods subsequent to contact with venom, and a corresponding study of the
respiratory phenomena presented by thin films of blood under the influence of
exposure to air.
The treatment to which the blood was subjected in the prepar-
ation of microscopic preparations, included the brief exposure of cover-glass films
to the fumes of 2 per cent. solution of osmic acid, and subsequent double-staining,
first with Delafield's solution of hæmatoxylin, and, second, with saturated aqueous
solution of eosin. Treatment of blood with osmic acid has frequently been
objected to as likely to interfere with efficient staining, but such interference
apparently only arises in connection with an unnecessarily prolonged exposure
to the acid, and very brief exposures, whilst completely fixative, in no way
interfere with perfectly successful staining. For some time double-staining
with eosin and anilin-dyes was employed, Plehn's fluid, or successive treat-
ment with methylene blue and eosin being used ; but the results were too
uncertain to be satisfactory, and this method was consequently soon abandoned
in favour of the other, which not only yields excellent contrast-effects, but is
also practically free from any risk of the occurrence of discharge of the nucleo-
plastic stain by the eosin. Eosin is of special value in such experiments., because
it not only serves as a good contrast-stain, but also as a delicate index to the
occurrence of the earlier stages of solution of erythrocytes, owing to the great
affinity which it has for hæmoglobin and the consequent staining which liquor
sanguinis or serum containing any appreciable quantity of the latter in solution
acquires under its influence.
In order to test the respiratory value of any specimen of blood, thin layers
were spread out on the surface of white porcelain freely exposed to the influence
of the air ; and, by employing a slab, such as those which are commonly used
in water-colour drawing, including a number of adjoining spaces, separated by
narrow ridges, numerous specimens, treated with various quantities of venom
and at various periods subsequent to treatment, could readily be compared with
one another and with samples of normal blood.
The following table illustrates the principal phenomena which presented
themselves in eight specimens of the blood of fowls, either pure, or mixed with
various quantities of distilled water corresponding with those which were em-
ployed as vehicles when the blood was treated with venom, and may be taken
as affording a standard of comparison with the results attending contact with the
latter under otherwise like conditions :—
TABLE I.
Phenomena present in specimens of blood of fowls, alone or with the addition of water.
No. of
Experi-
ment.
Nature of specimen.
Coagulation.
Colour.
Solution of bodies of
erythrocytes.
Reduction in respira-
tory property of
hæmoglobin.
Putrefaction.
Blood alone
.
. Immediate : No contraction of
clot.
21 days later ; surface scarlet;
substance plum-coloured.
Erythrocytes well pre-
served 21 days.
Entirely absent 21 days
later.
Absent 21 days later.
I
I I Blood alone
.
. Immediate : very considerable
contraction : serum, after 24
hours, limpid, straw-coloured.
24 hours later; surface scar-
let; substance, plum-colou r -
ed.
No solution, 48 hours
later.
Entirely absent 24 hours.
III Blood, 10.5 c. c.
Water, 1 c. c.
Firm within 2 minutes : 24
hours later, contraction con-
siderable; serum,
24—72
hours, limpid, straw-coloured.
24 hours later ; surface vivid
scarlet; substance deep
plum-coloured.
No solution, 24 hours
later.
Entirely absent up to 72
hours.
IV Blood, 10.5 c. c.
Water, 1 c. c.
Firm within 2 minutes : abund-
ant limpid, straw-coloured
serum present 24 hours later.
24 hours later ; surface scarlet;
substance bright, plum-
coloured.
An abundance of intact
erythrocytes present 48
hours later.
Entirely absent up to 48
hours.
V Blood, 10.5 c. c.
Water, 0.22 c. c.
Immediate : a considerable
amount of limpid, straw-
coloured serum present 48
hours later.
72 hours later ; surface scar-
let ; substance deep plum-
coloured.
Entire absence of solution
48 hours later.
Entirely absent up to 12
0
hours.
VI Blood, 10.5 c. c.
Water, c. c.
Firm within 2 minutes: abund-
ance of limpid, straw-coloured
serum 24 hours later.
48 hours later ; surface scar-
let ; substance plum-colour-
ed.
Considerable solution
present 24 hours later.
Entirety absent up to 4
8
hours.
VII Blood, 9 c. c. Water,
1 c . C.
Firm in 2½ minutes : a consider-
able amount of reddish serum
24 hours later.
24 hours later ; surface scar-
let ; substance deep plum-
coloured.
Very considerable solu-
tion 24 hours later.
Entirely absent.
VIII Blood, 9 c. c. Water,
1 c. c .
Firm in 1 minute .
. . 72 hours later; surface scar-
let ; substance deep plum-
coloured.
48 hours later ; majority
of erythrocytes still in-
tact.
Entirely absent up to 48
hours.
4
Scientific Memoirs by
The phenomena characteristic of this series of specimens were the following:
rapid and firm coagulation ; conspicuous contraction of the resultant clot ; pro-
longed retention of a vivid scarlet tint by the surface, and of a plum-coloured one
by the substance of the mass ; prolonged preservation of the bodies of the
erythrocytes ; and, finally, absence of any appreciable reduction in the respiratory
capacity of the hæmoglobin until very conspicuous evidences of advanced
putrefactive decomposition had made their appearance.
Deviations from the standard were, however, present in certain instances.
In specimen I there was an entire and permanent absence of any appreciable
contraction of the coagulum, but this may be readily accounted for, because,
as Lord Lister long ago demonstrated, contraction is absent in cases in which
no septic organisms have gained access to the fluid, a condition which the
behaviour of the material in other respects showed to have been present in this
instance. Minor variations were present in the case of the remaining specimens,
and specially in those to which water had been added, in respect to the precise
period at which appreciable evidences of the occurrence of processes of solution
of erythrocytes made their appearance, and may fairly be ascribed to the
presence of corresponding variations in degree of septic contamination. Even
in those specimens in which processes of solution set in relatively early, there
was no indication of any coincident appreciable reduction of the respiratory
property of the hæmoglobin, but this cannot be regarded as evidence that the
relative acceleration of solution was not determined by the incidence of putrefac-
tion, for, when the phenomena attending contact with venom come to be
considered, it will be found that the earlier stages of putrefactive change, in
place of favouring the occurrence of reduction, are in fact accompanied by a
recovery of respiratory property from the loss primarily induced by the action
of the venom.
Table II illustrates the phenomena characteristic of twenty-five experiments
in which various proportions of cobra-venom were brought into contact with
specimens of the blood of common domestic fowls.
TABLE II
Phenomena present in specimens of blood of fowls treated with various quantities of Cobra-venom.
No. of
Experi-
ment.
Nature of specimen.
Coagulation.
Colour.
Solution of bodies of
erythrocytes.
Reduction in respiratory
property of hæmoglobin.
Putrefaction.
I Blood, 76 c. c. Venom,
0.001 gramme. Water,
1 C. C.
Slow; clot tremulous ; a
large amount of deep
red opaque serum in 24
hours.
Black in substance,
dull red on surface in
24 hours.
Almost quite complete within
24 hours.
Perceptible in 48 hours.
II Blood, 76 c. c. Venom,
0.002 gramme. Water,
1 C. C.
Loose, 5 minutes; tremu-
lous, 24 minutes ; firm,
32 minutes.
Substance of clot rapid -
ly darkening ; black
in 24 hours.
About half of the erythrocytes
dissolved within 24 hours,
and remaining ones deform-
ed or partly dissolved.
Absent, 24 hours ; slight, 72
hours, absent, 96 hours.
III Blood, 9 c. c. Venom,
0.002 gramme. Water,
1 c. C.
Firm, 4 minutes; no con-
traction.
Black, 1 hour . . Complete, 1 hour . .
Perceptible, 24 hours.
IV Blood, 9 C. c. Venom,
0.00075 gramme.
Water, 0.75 c. c.
10 minutes ; no contrac-
tion of clot.
Darkening very rapidly;
almost black in 52
minutes ; perfectly
black in 2½ hours.
Almost complete, 1 hour ;
complete, 3 hours.
Distinct in 24 hours; very con-
siderable, 48 hours; almost
absent, 72 hours ; entirely
absent, 96 hours.
V Blood, 11 c. c. Venom,
0.0005
gramme. Water,
0.5 C. C.
4 minutes; a considerable
amount of deep red
serum in 24 hours.
Black within 24 hours. Considerable, 2 hours; almost
complete, 24 hours.
Barely perceptible, 24 hours ;
absent, 72 hours.
VI Blood, 9 c. c. Venom,
0.002 gramme. Water,
1 C. C.
15 minutes ; no contrac-
tion.
Black in 3 hours
. About half of the erythrocytes
completely dissolved and
the rest deformed, 1 hour ;
complete, 24 hours.
Very conspicuous, 24 hours ;
almost complete, 48 hours ;
complete, 72 hours; greatly
diminished, 120 hours.
Slight odour, 96
hours.
VII Blood, 10.5 c. c. Venom,
0.001 gramme. Water,
1 C. C.
Absent up to 2o minutes;
clot soft, incoherent;
no contraction.
Black in 24 hours
. Very considerable, 2 hours;
very greatly increased, 24
hours.
Barely perceptible, 24 hours;
absent, 72 hours.
VIII Blood, 10.5 c. c. Venom,
0.001 gramme. Water,
1 C. C.
Within 6 minutes; no
contraction.
Darkening rapidly ;
black 24 hours.
Very extensive in 2½ hours ;
complete, 24 hours.
Absent, 24 hours.
IX Blood, 10 c. c. Venom,
0.01 gramme. Water,
1 c. c.
Within a few minutes;
no contraction.
Black in 3o minutes . Complete in 50 minutes
. Complete in 24 hours.
M e d i c a l O f f i c e r s o f t h e A r m y o f I n d i a . 5
6 Scientific Memoirs by
TABLE II.—concld.
Phenomena present in specimens of blood of fowls treated with various quantities of Cobra-venom.
No. of
Experi-
ment.
Nature of specimen.
Coagulation.
Colour.
Solution of bodies of
erythrocytes.
Reduction in respiratory
property of Hæmoglobin.
Putrefaction.
X Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Loose, within 2 minutes;
no contraction.
Black in 22 minutes . Almost complete, in 22
minutes.
Complete in 24 hours; very
considerably diminished, 48
hours.
XI Blood, 11 c. c. Venom,
0.01 gramme. Water,
1 c. c .
Greatly delayed; clot,
soft, semi-fluid ; no con-
traction.
Black, 1h o u r
. . Complete in 2 hours .
. Complete, 24 hours; diminish-
ed, 48 hours; absent,96 hours.
XII Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 c. c.
Absent, 1¼ hours; loose,
soft clot, 24 hours ; no
contraction.
Black, 3¼ hours . Almost complete, 3¼ hours . Complete, 24 hours ; consider-
ably diminished, 48 hours.
XIII Blood, 10.5 c. c. Venom,
0 .01 gramme. Water,
1 c. c.
Absent, 44 minutes ;
loose, soft clot, 1 hour;
no contraction.
Black, 24 minutes
. Complete, 1¼ hours
. .
Extreme, 24 hours; diminished,
48 hours; absent, 72 hours.
XIV Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 c. c.
Absent, 1½ hours; soft,
incoherent clot, 24 hours;
no contraction.
Almost black, 2½h o u r s Almost complete, 24 hours • Very considerable, 24 hours ;
diminished, 48 hours ; almost
absent, 72 hours.
XV Blood, 10.5 c.c. Venom,
0.01 gramme. Water,
1 C. C .
Absent, 25 minutes;
semi-fluid, 24 hours ;
no contraction.
Black, 25 minutes
. Complete, 2½ hours .
Considerable, 24 hours ; almost
complete, 48 hours; diminish-
ed, 72 hours; absent, 120
hours.
XVI Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 c. c.
Rapid; contraction ab-
sent.
Black, 2½ hours
. Numerous fusiform or
partly solved erythrocytes
still present, 2½ hours.
Very considerable, 24 hours;
absent, 48 hours.
XVII Blood, 10.5 c. c. Venom,
0.01
gramme. Water,
1 c. c .
Absent, 15 minutes ; clot
soft, semi-fluid,24 hours;
no contraction.
Black, 2½ hours . . Almost complete, 2½ hours. Absent, 48—72 hours.
XVIII Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Loose, 1 hour ; firm
1
hour, 6 minutes ; no
contraction.
Almost black, 21/3 hours Almost complete, 2½ hours;
complete, 24 hours.
Very considerable, 24 hours ;
increased, 48 hours ; complete,
72 hours; progressively dimi-
nished, 96—144 hours; absent,
168 hours.
XIX Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 c. c.
2 minutes; no contrac-
tion.
Black, 2 hours,
1 0
minutes.
Complete, 2 hours,
10
minutes.
Very considerable, 24 hours ;
almost absent, 96 hours;
absent, 120 hours.
XX Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Immediate, clot soft ; no
contraction.
Black, 3 hours .
. Complete, 3 hours
•
. • Great, 24 hours ; absent, 48
hours.
XXI
Blood, 10.5 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Almost immediate ; no
contraction.
Darkening rapidly ;
black, 16 hours.
. . . . . .
Very conspicuous, 16 hours ;
diminished, 20 hours ; absent,
40 hours.
XXII Blood, 9 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Immediate; clot soft ; a
certain amount of con-
traction.
Almost black,
2 0
minutes.
. . . . . .
Almost complete, 24 hours ;
diminished, 48 hours ; absent,
96 hours.
XXIII Blood, 9 c. c. Venom,
0.01 gramme. Water,
1 C. C.
Loose, 10 minutes; firm,
33 minutes; no contrac-
tion.
Black, 33 minutes
. Complete, 33 minutes . . Very great, 24 hours; diminished
48 hours ; almost absent, 72
hours; absent, 96 hours.
XXIV Blood, 9 c. C. Venom,
0 .01 gramme. Water,
1 C. C.
Slow and imperfect; no
contraction.
Black, 11 minutes
•
Complete, 35 minutes .
. Almost complete, 24 hours;
absent, 48 hours.
XXV Blood, 9 c. c. Venom,
0.1 gramme. Water,
1 C. C.
Very soft and loose, 3
hours; no contraction.
Almost black, 5
minutes.
Complete, 5 minutes .
. Almost complete, 3 hours; very
greatly diminished, 24 hours.
Putrefactive odour
and gaseous bubbles;
present, 24 hours.
Medical Officers of the Army of India. 7
8
Scientific Memoirs by
These experiments were conducted as follows :—A test-tube, or, in excep-
tional cases in which an unusually large quantity of blood was employed, a
small flask, was carefully cleaned, sterilised and plugged with cotton-wool ; the
requisite quantity of venom, in solution in distilled water, was next introduced ;
and, finally, the blood was drawn directly into the tube or flask and shaken well
up so as to mix it thoroughly with the solution of venom. In carrying out
such experiments it was, of course, impossible to provide effectually against the
occurrence of more or less septic contamination, because any means, such as
prolonged exposure to high temperatures, adapted to effect sterilisation of the
solutions of venom were prohibited owing to the coincident reduction of both
lethal property and of capacity to give rise to hæmatic change which they
occasion in them.
A comparison of the data of Table II with those of Table I is sufficient to
demonstrate that the behaviour of samples of the blood of such susceptible
animals as fowls, containing even fractional ratios of cobra-venom, is very.
different from that of specimens of pure blood or of blood mixed with bulks of
water equivalent to those employed as vehicles for the venom. The distinctive
features presented by the envenomed blood, are the following:—delay, or more
or less complete suppression of coagulation ; absence of contraction of the
coagulum ; very rapid darkening of colour, so that the substance of the specimen
soon appears black and the surface of a very deep lake tint ; very rapid solution
of the bodies of the erythrocytes, and consequent liberation of nuclei where such
bodies are present, and the occurrence of various degrees of reduction in the
respiratory property of the hæmoglobin quite unconnected with that attending
the later stages of putrefactive decomposition.
There was unequivocal evidence of an abnormal delay in the occurrence of
coagulation in by far the greater number of the experiments, even where the ratio
of venom to blood was very low, and, where the ratio was a relatively high one,
coagulation was not only greatly retarded, but also very imperfectly carried out.
In one or two exceptional cases (XV, XVIII, XXII) coagulation took place
rapidly, but, in them, the phenomenon appeared to he related to the fact that the
blood, in place of entering the tube in a full stream, only did so drop by
drop, a condition which naturally tended to interfere with the rapid and
thorough mixture of the entire mass of it with the venom. The action of the
venom in interfering with the normal contraction of the coagulum was even more
conspicuous ; there was an entire absence of any appreciable contraction in the
great majority of instances, a phenomenon which is all the more remarkable as
evidence of the action of specific property in the venom, because the introduc-
tion of the latter must almost inevitably provide for the occurrence of septic
contamination, which, as has already been pointed out, is normally related to the
incidence of contraction.
Medical Officers of the Army of India. 1 3
fowl's blood at intervals of two, twenty-four, forty-eight, seventy-two and
ninety-six hours after mixture with 0.01 gramme of dried cobra-venom during a
period when the temperature of the laboratory in which the experiment was
conducted was comparatively high, that of the outer air having had a range of
71.2° to 89.9° F., and demonstrates that, under these conditions, an absolute
loss of respiratory property had occurred within the course of twenty-four, and
complete recovery within ninety-six, hours from the time at which the mixture
was effected. Figs. 7 and 9 show the results of similar experiments in which the
maximal laboratory temperatures to which the mixtures of blood and venom were
exposed were respectively 71° F. and 68.9° F., and very clearly illustrate the
delay both in reduction and recovery of respiratory property in the hæmoglobin
incident on depression of temperature, In the experiment illustrated by Fig.
7 maximal reduction was present forty-eight hours, and complete recovery
one hundred and twenty hours, after the mixture had been made, whilst in that
corresponding to Fig. 9 these phenomena presented themselves respectively at
72 and 168 hours. Fig. 8 illustrates the phenomena which characterised an
experiment, which was conducted coincidently with that corresponding to Fig. 9,
but under exposure to a temperature of 98° F. in an incubator, and demonstrates
the acceleration of both reduction and recovery of respiratory property under
such circumstances ; very appreciable reduction having been present within two
and half hours, and partial recovery within twenty-four hours from the beginning
of the experiment.
The results of the experiments which have hitherto been considered appear
to afford conclusive evidence that cobra-venom possesses a very great power
of giving rise to very important changes in specimens of blood with which it
comes into contact outside the body, and it now remains to be seen how far
there is any evidence of the occurrence of similar changes when the contact
takes place within the vascular system.
The phenomena which presented themselves in the course of a series of
experiments dealing with this question are illustrated in the following table : —
TABLE III.
Phenomena present in the blood of fowls treated with injections of Cobra-venom.
No. of
experi-
ment.
Nature of experiment.
Coagulation.
Colour.
Solution of bodies of
erythrocytes.
Reduction in respira-
tory property of
hæmoglobin.
1 Subcutaneous injection .
.
Venom, 0.01 gramme.
Convulsions, 33 minutes.
Blood taken, 38 minutes after
injection.
2 minutes ; contrac-
tion well developed.
Very dark at first, but
becoming scarlet.
Numerous free nuclei and
partly dissolved erythro-
cytes, solitary or in large
masses, in the blood when
taken ; serum pale red, 24
hours.
None appreciable, 24
hours after the blood
was taken.
16 Scientific Memoirs by
to blood was relatively high, it underwent rapid brightening on exposure to air
This, of course, clearly showed that the original tint was not entirely dependent
on any actual loss of respiratory property in the hæmoglobin, but was to a
great extent merely owing to defective aëration for some time before the period
at which death occurred or the specimen was removed from the body. On the
other hand, as has been already pointed out in my previous paper,* the
phenomenon by no means affords any satisfactory proof of an entire absence of
respiratory reduction, seeing that a loss of 33 per cent. of the respiratory property
in the hæmoglobin of the blood is sufficient to cause death, so that 67 per cent,
may, even in fatal cases, remain intact to account for the occurrence of very
considerable change of colour on exposure to sources of supply of oxygen. The
original colour of the blood in such cases was clearly owing to a great extent to
antecedent defect in supply of oxygen, but, as in those instances in which the
ratio of venom was exceptionally high there was conspicuous evidence of varying
degrees of permanent reduction, it is probable that this defect was not the sole
factor at work, but that, even where the ratio of venom was lowest, a certain
amount of permanent toxic change was also present to contribute towards the
induction of the sum of the reduction actually present.
But another factor, which, as has already been pointed out, exerts an im-
portant influence in modifying the colour of massive specimens of blood, quite
independently of the presence of any temporary or persistent reduction in the
oxygen-content of the hæmoglobin, and merely owing to the alterations in
reflective and refractive properties of the fluid, was also invariably present, for
in all cases, there were unequivocal evidences of the occurrence of extensive
processes of solution of the bodies of erythrocytes. (Plate I, figs. 1-5). The
out-come of such processes of solution did not, however, save in those cases in
which the ratio of venom was a very high one and death had immediately
followed its introduction into the vascular system, correspond precisely with that
attending instances of mixtures of blood and venom effected outside the body,
for, whilst in the latter the free, or partially freed, nuclei were isolated and more
or less evenly diffused, in the present case the majority of them were aggregated
into masses of irregular outline, and often of such considerable bulk as to
suggest the possibility that they may occasionally give rise to embolic
obstructions in the course of the vessels.
The reason for this difference, and for its absence in cases in which death
had immediately succeeded the introduction of the venom, is not hard to find.
Where a mixture of blood and venom is effected outside the body, the particulate
elements of the fluid subsequently remain at rest, and there can be no special
reason why those which are undergoing solution should tend to adhere to one
another. But, in cases in which solution is initiated within the vascular system
* "Scientific Memoirs by Medical Officers of the Army of India," Part IX, page 3.