Abstract
We have reported 1 , 2 on the use of a liver preparation in the prevention of liver necrosis from carbon tetrachloride or chloroform administration. At this time we wish to report a method for the preparation of this active principle in a crystalline form.
A concentrated aqueous extract of hog liver representing approximately 10 gm. per cc, from which the heat coagulable materials have been removed, is warmed in a water bath to a temperature of approximately 60° C. To each 1000 cc. of this solution 2400 cc. of ethyl alcohol of about the same temperature is added with stirring. The precipitate which forms is filtered off after cooling and 2000 cc. of a saturated aqueous solution of ammonium sulfate added to the filtrate. The solution is then thoroughly shaken. On standing it separates into 2 layers: above, an alcoholic layer and below, a watery layer containing a great deal of precipitated ammonium sulfate. The upper layer is syphoned off and 1500 cc. of alcohol added to it to precipitate excess ammonium sulfate. The solution is filtered after being cooled in a refrigerator for several hours. The filtrate is then evaporated under reduced pressure to approximately 170 cc. It is then placed in a refrigerator and cooled over night. The precipitate is separated by centrifuging and washed in the centrifuge tubes, first with about 60 cc. of ice cold water and then with approximately 40 cc. of cold alcohol. The residue is suspended in 40 to 60 cc. water, depending upon the amount present, and concentrated sodium hydroxide added drop by drop with thorough mixing until needle-like crystals separate out or form when a drop of the solution is placed on a microscope slide and allowed to evaporate slightly. As a rule a satisfactory pH is approximately 9.3. After the correct amount of alkali is added the solution is set aside in a refrigerator for a number of hours and then filtered with suction.
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