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ether to remove any remaining traces of alcohol and benzoic acid after which the traces of ether were boiled off. The aqueous residue was then sterilized by filtration through a Berkfeldt filter, and contained the potent substance.
We also extracted the secretion or hormone from the minced pancreas with ethyl alcohol, methyl alcohol, methylated spirits, and acetone, as in the methods of Banting, Best and Collip, and then concentrated the potent substance in the extractive filtrate by precipitating it with benzoic acid as above.
The following is an example of the alternative method by which we obtained the potent substance from the pancreas of mammalia:-
One kilogram of ground pancreas was mixed with 70 cc of 10% copper sulphate solution and 15 cc of concentrated hydrochloric acid to retard the extraction of the proteins and other injurious substances from the gland. Two litres of water, heated to 80 c., were then added to the mixture for extracting the potent substance. The resulting aqueous extractive solution was then filtered and to the filtrate was added an equal volume of saturated solution of ammonium sulphate. When the ammonium sulphate was added, a solid separated out of the solution and floated to the top. This solid contained the potent substance and some of the other extractive substances. This solid was removed from the filtrate and freed from the adhering ammonium sulphate solution. The solid was then dissolved in 100 cc of distilled water to which was separately added sodium benzoate and hydrochloric acid which formed a precipitate of benzoic acid. This precipitate united with the potent substance as by adsorption.
The benzoic acid precipitate with the adhering