substances used for this purpose are: crystalline sulphate of lime,
boric acid, mannite, sugar, starch, salicine, stearic acid, and the
sulphates of cinchonine and quinidine. These bodies are detected as
follows:
_a._ Upon slightly warming 2 grammes of sulphate of quinine with 120
grammes of alcohol of 21° B., the pure salt completely dissolves; if,
however, starch, magnesia, mineral salts, or various other foreign
substances are present, they are left as insoluble residues.
_b._ Those mineral substances that are soluble in alcohol are detected
by calcining the suspected sample: pure sulphate of quinine is
completely consumed; whereas, the mineral substances present remain
behind as a residue.
_c._ In presence of salicine, the salt acquires a deep red color, when
treated with concentrated sulphuric acid.
_d._ Stearic acid remains undissolved upon treating sulphate of quinine
with acidulated water.
_e._ To detect sugar and mannite, the sample is dissolved in acidulated
water, and an excess of hydrate of baryta added: a precipitate,
consisting of quinine and sulphate of baryta, is produced. Carbonic acid
is then passed through the fluid, in order to precipitate the excess of
baryta as insoluble carbonate, the fluid saturated with ammonia, to
throw down the quinine which may have been re-dissolved by the carbonic
acid, and the mixture filtered. If the salt be pure, no residue will be
obtained upon evaporating the filtrate; a residue of sugar or mannite is
formed, if these substances are present.
_f._ Sulphate of quinine invariably contains 2 or 3 per cent. of
cinchonine, originating, not from a fraudulent admixture, but from an
incomplete purification of the salt. One of the best methods for
detecting the respective quantities of quinine and cinchonine, present
in a sample of the sulphate, is the following: Several grammes of
ammonia and ether (which has previously been washed with water) are
added to one or two grammes of the salt under examination, the mixture
thoroughly agitated, and then allowed to remain at rest. The supernatant
etherial solution contains all of the quinine; the cinchonine, which is
almost completely insoluble, both in water and ether, remaining
suspended between the layers of the two fluids. The ether is next
removed by means of a stop-cock funnel, evaporated to dryness, and the
weight of the residue obtained determined. The operation is then
repeated, the ether being replaced by chloroform in which both quinine
and cinchonine are soluble. The residue, formed by the evaporation of
the second solution, will be heavier than the first residue: the
difference between the two weighings gives the weight of the cinchonine
present.
_g._ The detection of the presence of sulphate of quinidine is based
upon the difference in the solubilities of the oxalates of quinine and
quinidine. Oxalate of quinidine is sufficiently soluble in cold water
not to be precipitated by double decomposition when solutions of oxalate
of ammonia and sulphate of quinidine are mixed. Under the same
circumstances, quinine is almost completely thrown down. The test is
applied as follows:
The suspected salt is dissolved in water, a slight excess of oxalate of
ammonia added, and the precipitate formed separated by filtration. If
the salt be pure, the filtrate is scarcely rendered turbid by the
addition of ammonia; when, however, sulphate of quinidine is present, it
will be entirely contained in the filtrate, in which ammonia will
produce an abundant precipitate.
EXAMINATION OF BLOOD STAINS.
This branch of legal chemistry formerly gave but very unreliable
results. It is scarcely ten years since the reactions that are now
regarded as only secondary and confirmative in their character, and far
from conclusive, were the only ones in use: these are the tests based
upon the presence of iron and albumen in the blood. Since then, great
progress has been made in the methods employed. It must not be
understood, however, that the question under consideration always admits
of an easy and decisive solution: the stains are sometimes too greatly
altered to be identified; but in cases where the distinctive reactions
of blood can be produced, the real nature of the stains under
examination can, at present, be determined with certainty.
The tests more recently introduced consist in the production of small
characteristic crystals, termed _haemin_ crystals, and in the use of the
spectroscope. Crystals of haemin (first discovered by _Teichman_) are
formed when dry blood is dissolved in concentrated acetic acid, and the
solution evaporated to dryness: they are of a brownish-red color.
_Brücke_ first suggested an analytical method, based upon this property
of blood, which is equally characteristic and sensitive: It is only
necessary to dissolve a minute portion of the matter to be examined
(dried blood, or the residue left by the evaporation of the fluid
obtained by treating the stain, or the dried blood, with cold water) in
glacial acetic acid and evaporate the solution to dryness in order to
obtain crystals of haemin, which can be readily recognized by means of a
microscope having a magnifying power of 300 diameters. If the crystals
originate from fresh blood, they appear as represented in Fig. 21;
crystals from old blood are represented in Fig. 22.
[Illustration: Fig. 21.]
[Illustration: Fig. 22.]
The former possess a reddish-brown, the latter a lighter color.
The various methods now employed to produce haemin crystals were
proposed by _Hoppe-Seyler_, by _Brücke_ and by _Erdman_. Whichever
process is used, the suspected stains are at first carefully separated
from the material upon which they are deposited. If they are present on
linen, or other fabrics, the stained portions, which always remain
somewhat stiff, are cut off: they will present a reddish-brown color, in
case the cloth is not dyed: if the stains are on wood, they are removed
by means of a sharp knife; if on stone or iron, they are detached by
scraping.
In case Hoppe-Seyler's method is used, the stains, separated as
directed above, are macerated with a little _cold_ water (warm water
would coagulate the albumen present, and consequently prevent solution
taking place): the stains become soft, striae and brown or reddish
clouds are observed, especially when the dried blood is fresh, and, at
the same time, the objects upon which the stains were deposited are
decolorized. Upon allowing the fluid obtained in this way to
spontaneously evaporate on a watch-glass, a reddish brown or brownish
residue is left, from which the crystals of haemin are prepared in the
following manner: An almost imperceptible amount of common salt is added
to the residue, then, six to eight drops of concentrated acetic acid,
and the mass thoroughly mixed by stirring with a small glass rod. The
mixture is at first heated over a small gas flame, then evaporated to
dryness by the heat of a water-bath. If the stains were produced by
blood, a microscopic examination of the residue will reveal the presence
of haemin crystals. This method presents an objection: if the stained
objects have been washed with warm water previously to the examination,
the albumen will be coagulated, and the blood rendered insoluble; in
this case, cold water will fail to dissolve anything, and the residue
will not produce crystals when treated with acetic acid.
In order to remedy this difficulty Brücke operates directly upon the
stained woven or ligneous fibre, or the matter removed from the stone or
iron: The materials are boiled in a test-tube with glacial acetic acid,
the fluid decanted or filtered, a trace of common salt added, and the
liquid then evaporated on a watch-glass at a temperature between 40 and
80°. If the stains really originated from blood, haemin crystals will
now be easily perceptible upon examining the residue obtained under the
microscope.
The stained fabric, the matter removed from the stone or iron, or the
residue left by the solution with which the stains have been treated, is
placed on the glass, a trace of chloride of sodium added, and the whole
covered with a thin glass plate. A drop of acetic acid is then placed at
the edge of the plates--between which it is soon introduced by capillary
attraction--and the mixture allowed to rest in the cold for a few
moments. The mass is next brought into solution by slightly heating, and
is then evaporated by holding the plate at a considerable distance above
a gas burner. The fluid is examined from time to time under the
microscope: when it is sufficiently concentrated, crystals, presenting
the appearance represented in Figs. 21 or 22, will be observed. These
are especially well-defined, if an insoluble substance is also present
between the plates--which prevents their adhering. The fluid collects by
capillary attraction at the points of contact of the plates as a more or
less colored layer, in which the crystals are deposited.
Should the above test fail to present distinctive indications at first,
one or two fresh drops of acetic acid are introduced between the plates,
and the examination is repeated. The result is not to be regarded as
negative, until several trials have proved fruitless, as the stained
portions are but slowly soluble, and crystallization may have been
prevented by the too rapid evaporation of the acetic solution.
Haemin crystals, once seen, can hardly be confounded with other
substances; still, it is well to identify them by confirming their
insolubility in water, alcohol, and cold acetic acid, as well as their
instantaneous solubility in soda lye.
The addition of common salt is ordinarily superfluous, as it is
normally contained in the blood; but it is possible, if the stains were
washed with warm water, that, in addition to the coagulation of the
albumen, the solution of the salt may have taken place, in which case
crystals will fail to form. The addition of salt is to remedy this
possible contingency; albeit, the delicacy of the test is not affected,
even if crystals of chloride of sodium are produced, as these are easily
soluble in water, and are readily distinguished from those of haemin by
aid of the microscope.
The indications furnished by means of the spectroscope are less reliable
than those given by the production of haemin crystals; moreover, the
spectroscopic examination requires favorable weather for its execution.
Still, the test should be employed in all possible instances. The course
pursued is the following:
The aqueous fluid, with which the stains have been treated, is placed in
a watch glass, and evaporated _in vacuo_ over sulphuric acid; the last
remaining portion of the fluid being united in the bottom of the glass
by causing it to collect in a single drop. When the evaporation of fluid
is completed, the watch-glass is placed before the narrowed slit of a
spectroscope, and a ray of diffused light (or better, light reflected
from a heliostat) made to pass through the part of the glass containing
the residue. If the stains originate from blood, the absorption lines of
_haemoglobin_, consisting of two large dark bands, to the right of the
sodium line (_Frauenhofer's_ line D), will be observed in the spectrum.
In case both of the above tests fail to give positive results, it is
almost certain that the stains examined were not caused by blood. If, on
the contrary, the reactions were produced, scarcely any doubt exists as
to the presence of blood. Under these circumstances it is advisable to
confirm the results by means of the tests that have been previously
spoken of as being formerly exclusively employed; these are the
following:
_a._ 1/2 to 1 c. c. of ozonized oil of turpentine, _i. e._ turpentine
which has been exposed to the air sufficiently long to acquire the
property of decolorizing water that is slightly tinted with indigo--is
introduced in a test-tube, and an equal volume of tincture of guaiacum
added (the latter tincture is prepared by treating an inner portion of
the resin with alcohol, until its brownish color is changed to a
brownish-yellow).
If upon adding some of the substance under examination to the above
mixture a clear blue coloration ensues, and the insoluble matter thrown
down possesses a deep blue color, the presence of coloring matter of the
blood is indicated. The mixture also imparts a blue color to moistened
spots from which the blood stains have been as completely extracted as
possible. Unfortunately sulphate of iron gives the same reaction.[X]
[X] Fresh gluten, gum arabic, and caseine also cause the blue
coloration.--_Trans._
_b._ Upon heating the fluid obtained by treating the stains with cold
water in a test-tube, its brown or reddish color disappears, and
greyish-white flakes of coagulated albumen are thrown down. The
precipitate acquires a brick-red color, when treated with an acid
solution of nitrate of mercury containing nitrous acid. The albumen is
also coagulated by the addition of nitric acid: it assumes a more or
less yellow color, if heated with a slight excess of the acid.
Chlorine-water, especially upon heating, likewise precipitates albumen
in the form of white flakes.
_c._ If the fluid is acidulated with a few drops of acetic acid, and a
drop of ferrocyanide of potassium added, a white precipitate, or, at
least, turbidity is produced.
_d._ The flakes of albumen, separated by heating, dissolve in caustic
alkalies to a solution, from which they are re-precipitated by nitric
acid, or chlorine water.
_e._ Upon treating blood stains with chlorine-water, a solution which
contains chloride of iron, and acquires a red coloration by the addition
of sulphocyanide of potassium, is formed.
_f._ Should the stains have failed to be affected by cold water (which,
as has already been remarked, is the case when they have been previously
washed with hot water), they are treated with weak soda lye. Nitric
acid, hydrochloric acid, and chlorine water will produce in the solution
so obtained a white precipitate, which exhibits the general properties
of albumen previously described. In case the stains are deposited upon
linen, it is necessary to replace the soda by ammonia, in order to avoid
dissolving the fabric.
_g._ Solutions of the alkalies, which dissolve the albumen, leave the
coloring matters intact, and consequently do not decolorize the fabric.
If the latter is afterwards subjected to the action of hydrochloric
acid, the coloring matter is dissolved, forming a solution that leaves
upon evaporation to dryness a residue containing iron, which gives a
blue coloration with ferrocyanide of potassium, and a red coloration
with sulphocyanide of potassium.
_h._ The coloring matter of blood dissolves in boiling alcohol, to which
sulphuric acid has been added, to a brown dichroic fluid (appearing
green by transmitted light, and red by reflected light). A mixture of
rust and blood exhibits the same phenomenon.
_i._ If substances containing blood are heated in a dry tube, an odor
resembling that of burnt horn is emitted. In case the stained fabric is
a substance that would produce this odor, (such as wool, silk, or hair),
the test naturally loses all value.
_j._ If the fluid obtained by treating the stains either with water or
alkali is evaporated with a little carbonate of potassa, and the residue
heated, at first at 100°, then to redness, in a glass tube to which a
fresh quantity of carbonate of potassa has been added, cyanide of
potassium is formed. When cold, the tube is cut above the part
containing the fused mixture, the mass heated with iron-filings and
water, the fluid filtered, and the filtrate then acidulated with
hydrochloric acid: ferrocyanide of potassium will be present in the
fluid, and upon adding a drop of solution of perchloride of iron a
green, or blue, color will be produced, and a precipitate of Prussian
blue gradually thrown down.
If the stained cloth is non-nitrogenous (_per ex._: hemp, linen, or
cotton), instead of treating it with water, it may be heated until
pulverulent, mixed with carbonate of potassa, the mixture calcined, and
the operation then completed as just described. This test having given
affirmative results, the operations should be repeated with an unstained
portion of the cloth, to remove all doubt that the indications obtained
do not really originate from the fabric.
In the present state of science, it is impossible to discriminate
chemically between human and animal blood. _M. Barruel_, it is true, is
able, not only to accomplish this, but also to distinguish the blood of
the various species of animals by its odor! But this test has a somewhat
hypothetical value for scientific purposes. In regard to the crystals of
haemin, they do not present sufficient difference to allow the blood of
different animals to be distinguished. We have not yet treated of the
globules. It often occurs that these minute organs are so altered as to
be no longer recognized in the microscopic examination; when, however,
the stains are tolerably recent, they may be detected by examining the
moistened stained cloth, directly under the microscope: a discrimination
between animal and human blood is then possible: corpuscules of human
blood possess the greater size: those of the sheep, for instance, have
only one-half the diameter of the former. It is, however, but seldom
that this distinction can be made use of.[Y]
[Y] _Menstrual blood_ is recognized by the presence of epithelial
cells.--_Trans._
EXAMINATION OF SPERMATIC STAINS.
In cases where attempt at violence, rape or pederasty is suspected, the
expert may be required to determine the nature of stains found on
clothing, sheets, etc. The fact that the stains were produced by semen,
may often be regarded, _per se_, as criminating evidence. This class of
investigation possesses, therefore, considerable importance.
_External appearance of the stains._--Dry spermatic stains are thin, and
exhibit a greyish or, occasionally, a citron-yellow color, if present on
white cloth. In case the fabric is colored, they appear whitish and, if
on linen, present a glossy aspect. They are translucid, when observed by
transmitted light. If the fabric, upon which the stains are deposited,
is of a heavy texture, they are visible only on one side: under all
circumstances, their circumference is irregular and undulated. These
indications, however, are not conclusive, but vary according to whether
the stains were produced by the thick semen of a vigorous man, or the
aqueous seminal fluid of an aged and diseased person, or by semen more
or less mixed with the prostatic fluid. Upon moistening spermatic
stains, the distinctive stale odor of fresh semen is sometimes emitted,
but this characteristic is usually obscured by the presence of foreign
substances.
Semen stains are soluble in water, forming a gummy fluid, in which
chlorine, alcohol, bichloride of mercury, acetate and subacetate of lead
produce a white precipitate, but which fails to be coagulated by
heating. Plumbate of potassa does not impart a fawn-color to these
stains, at a temperature above 20°, as is the case with those produced
by albuminous substances.
Persulphate of iron imparts to spermatic stains a pale yellow color,
Sulphate of copper, a bluish grey color,
Cupro-potassic tartrate, a bluish grey color,
Nitrate of silver, a pale grey color,
Nitric acid, a pale yellow color.
The above reactions, separate or united, are insufficient; they are not
very delicate, and are likewise produced by stains originating from the
other varieties of mucus: the indications furnished by a microscopic
examination of the stains are alone conclusive.
_Microscopic examination._--Semen contains as its principal and
fecundating constituent, peculiar vibratory filaments, (_spermatozoa_),
held suspended in a viscous fluid. These filaments, when preserved in a
warm and moist place, retain their activity for a considerable time: it
is even possible that they may exhibit vitality in the organs, into
which they have been voluntarily or forcibly ejaculated, for ten, or
even twenty-four hours. When exposed to cold air, the spermatozoa
quickly expire; still, they preserve their form for some time, and, as
this is very characteristic, it is then easy to identify them; moreover,
since they originate exclusively in the testicles, their detection may
be considered as certain evidence of the presence of semen. In stains
produced by aged persons, and by persons enfeebled by excesses, the
spermatozoa fail to be presented; in case they are discovered, this fact
evidently does not affect the certainty of the spermatic origin of the
stains. The contrary conclusion is never absolutely certain: still, if
the use of the microscope fails to establish the presence of
spermatozoa, it is almost certain that the stains were not produced by
semen.
Of the various methods for obtaining from the stains a preparation
adapted to the microscopic examination, the one proposed by M. Charles
Robin is the most simple and reliable.
A strip, 1 c. c. in size (comprising the entire stain, if this be small,
containing its inner portion, if it be large), is cut from the fabric
under examination, care being taken that the two extremities of the
sample extend beyond the stained portion.
One end of the cloth is then immersed in a capsule, or watch-glass,
containing pure water: the stains become moistened by capillary
attraction, and, in a space of time varying from twenty minutes to two
hours, acquire the appearance of fresh semen. As soon as the stained
portion becomes swollen and softened, the surface of the cloth is gently
scraped with a spatula, and the substance removed placed on the slide of
the microscope. The particles are next slightly detached, a drop of
water added, if necessary, and the whole covered with a small plate of
very thin glass. The preparation is then examined by a microscope,
having a magnifying power of from 500 to 600 diameters. In this way, the
presence of either entire or broken spermatozoa is readily detected.
Their existence is rendered still more apparent, if the mucus present is
dissolved by adding a drop of acetic acid to the preparation.
Entire spermatozoa consist of long slender filaments, having a length
of 0.04041 to 0.04512 millimetre; the anterior extremity presents an
oval enlargement, either round or pyriform, exhibiting a double outline,
when magnified to 500 diameters. This enlarged end is termed the "head;"
the entire remaining portion being regarded as the "tail." In case the
spermatozoa are broken, they are severed either near the head or in the
middle of the tail, and a mass of detached fragments will be observed in
the microscopic examination. The spermatozoa are not the only
corpuscules revealed by the microscope; other substances, entirely
different in character, are often observed. Although the detection of
these bodies is, in itself, of no value, it will be well to enumerate
and characterize them; they are:
_a._ Oily globules.
_b._ Leucocytes, or spherical and finely granulous globules of mucus.
_c._ Corpuscules, originating from the seminal vesicles, termed
sympexions. These are rounded or ovoid, possess an irregular outline,
and are usually mixed with the spermatozoa and globules of mucus.
_d._ Crystals of phosphate of magnesia, varying greatly in size; the
largest are from 0.mm. 001 to 0.mm. 002 in length. The crystals formed
upon cooling the semen, present the form of an oblique prism, with a
rhomboidal base. Occasionally they are elongated and flattened; they
then assume the form of a rhomboid.
_e._ Epithelial cells; originating from the mucous follicles of the
urethra.
_f._ Irregular grains of dust; soluble in acetic and hydrochloric acids,
with gaseous evolution.
_g._ Brownish-red grains of rust; only slightly soluble in acetic acid,
but easily soluble in hydrochloric acid.