288 grams of the original poisonous material were extracted with 50 per
cent. alcohol, and this alcoholic solution was precipitated with lead
acetate in the manner already described (p. 17). The lead precipitate so
obtained was extracted with ether in Soxhlet extractors and after the
extraction was found by test to be free from poison. Therefore the
poison, if precipitated by the lead acetate, must have been extracted by
the ether. This ether solution had a dark green color, and was acid from
acetic acid brought down in the lead precipitate. The ether was
evaporated in a vacuum desiccator without heat and there remained a
small quantity of an acid mixture of water and a soft tar; the watery
part was colored green, showing that the tar was soluble to some extent
in dilute acetic acid. The mixture had the peculiar odor of the original
material. A small drop of the green watery part was applied to the
wrist, allowed to remain a few minutes and was then removed by absorbent
paper, but the spot was not washed. Itching and reddening of the skin
commenced within twenty-four hours. At the end of forty-eight hours,
there was a well developed case of poisoning. How this was cured will be
described in another place.
A small portion of the poisonous mixture was dissolved in alcohol, and
this solution was divided into three parts. The first part was treated
with ferric chloride, but it gave no color reaction. Another portion of
the alcoholic solution was diluted with water. It became turbid. The
third portion gave a dirty-green precipitate with lead acetate, which
seemed to come down more readily when the solution was diluted with
water. The main portion of the poisonous mixture was then dissolved in
95 per cent. alcohol and lead acetate in 50 per cent. alcohol was added.
The precipitate was filtered, washed, and decomposed by hydrogen
sulphide in a mixture of water and ether. The ether solution was
filtered and evaporated. The residue was a tar which, on standing in a
desiccator for some time, became dry enough to break into sticky lumps.
An alcoholic solution of this substance gave a dark color with ferric
chloride and a light colored precipitate with lead acetate.
To get more of the poisonous tar for study, 233 grams of original
material were extracted with 95 per cent. alcohol. Strong alcohol was
used in order to dissolve as much of the tar as possible. The solution
had a dark greenish color, but was somewhat yellow in thin layers. The
undissolved tar was filtered off and extracted twice again in the same
way. The tar left after the third extraction was only slightly soluble
in alcohol, and its solution was not poisonous. The three filtrates from
these three extractions were precipitated separately by lead acetate in
50 per cent. alcohol. The first precipitate was largest, darkest in
color, and carried down more tarry matter. The second was light green,
and the third was quite small, black, and was not a lead compound at
all, but some of the tar which separated out on diluting the strong
alcohol with the weaker grade containing lead acetate. It was soluble in
ether and less soluble in alcohol. The alcoholic solution of this third
lot gave no precipitate with hydrogen sulphide. The first and second
lead precipitates were filtered by suction and washed with water. They
were kept a day or two in a desiccator over sulphuric acid, but did not
become completely dry. The weight of these two moist precipitates
together was 172 grams. They were combined and extracted with ether in
Soxhlet extractors which were kept in operation during work hours for
three days.
In the meantime the alcoholic filtrates from these lead precipitates
were combined and concentrated on the water bath by distilling off two
liters of alcohol. The alcohol obtained had the peculiar odor of the
original material, but was not poisonous.
After a long extraction of the lead precipitate in the Soxhlet
extractors, the green ether solutions were combined and washed by
shaking them with water to remove lead acetate and acetic acid in case
any should have been held in the lead precipitate. The ether was
distilled off at a low temperature and there remained a soft tar, a
portion of which was not completely soluble in 95 per cent. alcohol. The
alcoholic solution had a greenish yellow color and was poisonous. The
tar was also partly soluble in acetic acid, and this solution was found
to contain lead. Thinking that the lead acetate had not been completely
washed out, the main part of the tar was dissolved in ether and shaken
with water. The wash water continued to give a test for lead as long as
the washing was continued. This indicated probably the hydrolysis of an
unstable lead compound. Hydrogen sulphide was passed into the ether
solution mixed with water to remove the lead. Lead sulphide was filtered
off, and the ether was evaporated. A small portion of the tar residue in
alcoholic solution gave a color reaction with ferric chloride. As this
may have been due to traces of lead gallate dissolved in the extraction
with ether and afterwards decomposed by hydrogen sulphide, the main
portion of the tar was redissolved in ether and shaken with water until
it no longer reacted with ferric chloride. The ether was then evaporated
and a soft, black, poisonous tar or gum of uniform consistency was left
which was shown by tests to be free from gallic acid and lead. These
experiments showed that some of the poison was precipitated as a lead
compound soluble in ether and some was brought down mechanically in the
free state. To see if the extraction with ether in the Soxhlet apparatus
was complete, the residue in the thimbles was decomposed by hydrogen
sulphide and shaken with ether. The dark colored ether solution was
freed from gallic acid by shaking with water and dilute sodium carbonate
solution, and was evaporated. A small quantity of tar was obtained which
was added to the main portion.
A solution of the poisonous tar in 95 per cent. alcohol did not reduce
Fehling solution and did not give a precipitate with lead acetate except
the separation of a small quantity of tar, which was not a lead
compound. The lead compound of the poison was apparently soluble in 95
per cent. alcohol as well as in ether, for it would not precipitate in
this medium, although it was found in the original precipitate by lead
acetate. The alcoholic solution of the tar became turbid on diluting
with water.
In order to see if the poison is volatile with vapor of acetic acid,
since this acid is found in the plant and it is thought by some that the
poison is volatile, a portion of the tar was distilled under diminished
pressure with acetic acid. It was soluble to some extent in the acid.
The temperature did not go higher than 55 deg. during the distillation. A
tube containing cotton wet with sweet oil was placed between the
receiver and the water suction so that the uncondensed vapors would have
to pass through the cotton. This cotton was rubbed on the skin and was
not poisonous. The yellow distillate collected in the receiver was also
tested and was not poisonous.