Work by Dakin, West, and Associates

In 1935, Dakin and West reported a different method of fractionating Cohn s fraction G (22). A preliminary purification was obtained by treating the extract with calcium acetate or chloride in 75-80 per cent alcoholic solution, the active material appearing in the filtrate. The active material was precipitable by Reinecke salt in acid solution. In the regeneration of such a precipitate, the bulk of the Reinecke acid was separated from a weak alcoholic solution of the active material as the sparingly soluble salt of a tertiary base—e.g., dimethyl aniline. The small amoimt of additional unprecipitable Reinecke acid was removed with solvents such as amyl alcohol w hich did not remove the active material. These procedures were carried out at virtually neutral reaction. The steps involved in the preparation described are diagrammed in Table III. 

Very little chemical separation was effected by treatment of Product I with picric acid. However, flavianic acid precipitated 65-75 per cent of Product I as an oily, somewhat unstable substance, which contained the active principle and thus constituted definite progress in the course of 

Commercial Liver Extract treated with calcium acetate in 75% alcohol 

Removed calcium by ammonium oxalate, and alcohol by evafmration under reduced pressure, precipitated with Rei-necke acid and regenerated by dissolving in CHiOH and precipitating the Reinecke complex with dimethyl aniline (clinically active) 

Salted out in neutral or feebly acid solution (aqueous) with (NHdsSO (i or more saturated), repeating process several times 

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