added, the solution filtered with suction, and the precipitate disearded. In order to assure precipitation of the enzyme by the further addition of aleo- hol, 1.25 ce. of saturated NaCl is added to each 100 ce. of 15% alcohol fillzate. The alcohol concentration is then slowly increased to 45%, the temperature of the solution being gradually lowered to about — 10°C. The precipitate is allowed to settle overnight and the supernatant is siphoned off. The precipitate is ventrifuged compactly in a refrigerated centrifuge. ‘The bulk of the enzyme activity is in the precipitate Second Ethanol Fractionation. he precipi ice-cold water to give an extinction of 20 at a wavelength of 280 my in a Lem, cell. This corresponds to a protein concentration of about. 1 protein /ec. Di nt 10% alcohol (8.5 ce. per 100 cc. of enzyme solution) is added at a gradually diminishing temperature to bring the concentration to 8 volumes %. The inactive precipitate is centrifuged off in a refrigerated centrifuge. Alvohol is then added to the supernatant to bring the alcohol concentration to ‘The temperattue is meanwhile lovered to —7 to —8°C. ‘The solution is centrifuged and the supernatant discarded. Third Ethanol Fractionation. ‘The precipitate is taken up in suffivient cold water to give an extinction of 24 at 280 my ina Lem. cell (15 mg. pro tein/ce.). Disregarding the alcohol content of the precipitate, sufficient alcohol is aclded to bring the concentration to 8%; the precipitate is centii- fuged off and discarded. The pH of the supernatant, which is about 5.6, is then adjusted to 6.8 by the careful addition of 5% sodium carbonate. Alcohol is added to 23% and the precipitate, containing the activity, centrifuised off. garding the alcohol in the precipitate, suffic