618 , 'RECENT WORK ON INSULIN
lowed by the report of many modifications and improvements.
At least twelve groups of observers have already reported on
this aspect of the subject. I will not attempt to discuss the
details of all the various methods, but you may be interested to
hear in a general way of our experience with certain significant
new developments.
The first solutions of insulin were made by watery extrac-
tion of pancreas. Alcohol was introduced as a solvent by the
original investigators (2) and has remained the most popular
extractive up to the present time. This solvent has been used
continuously in Toronto in the routine preparation of material
for clinical use, with the exception of a few inonths in the spring
of 1922 whlien acetone vwas employed. Doisy, Somogyi and
Shaffer (3! have used alcohol as the extractive in their method.
These investigators introduced'the precipitation of insulin from
watery solution by half saturation of ammonium sulphate, and,
coincident with Walden of the Eli Lilly and Company, the
isoelectric method of purification. Fisher (4) has slightly modi-
fied this method by employing a higher concentration of alcohol
than used by the previous investigators to fractionally precipi-
tate the inert protein material. He claims to eliminate by this
procedure a larger amount of those substances in the pancreas
which antagonize the action of insulin. The existence of these
substances was recognized very early in the work on insulin.
Dudley (5) has retained the alcoholic extraction of the earlier
methods as the first stage of his procedure for the preparation
of insulin and has developed the so-called picric acid method of
purification. In this method insulin is precipitated from watery
solution, as the picrate, by the addition of picric acid. The
picrate is subsequently treated with hydrochloric acid in 75 per
cent alcohol and insulin is liberated as the hydrochloride. The
hydrochloride is precipitated from alcoholic solution by the
addition of ether or acetone. This method has been extensively
used by manufacturers in England. Dudley (6) has recently
published certain modifications of his method. Hle has obtained
very satisfactory results by using alkaline alcohol as the extrac-
tive. Krogh (7), of Copenhagen, uses alcohol as the extractive.
Hlie believes that improved results are obtained when the glands
are frozen as soon as possible after collection. The frozen mate-