A new preparation of crystalline anterior pituitary growth hormone.

The isolation and identification of the anterior pituitary growth hormone as a pure protein were first achieved by Li, Evans, and Simpson (1, 2) in 194445. Their method is a beautiful example of the classical procedures of protein fractionation by the controlled use of salt and of hydrogen ion concentrations. It owes its distinction to the proof that the growth-promoting activity of anterior pituitary extracts, first demonstrated by Evans and Long (3), is attributable to an individual protein. The disadvantages of the method lie in the somewhat low extraction efficiency from acetone powders of the anterior pituitary glands, in the tedium of the many repeated steps of the process, in the time consumed, in the losses inevitably associated with repeated manipulations, and in the resulting low yields of product (of the order of 60 mg. per kilo of fresh anterior pituitary glands). The method attains its objective of isolating pure growth hormone, and it secures information about the properties of the protein that is essential in the development of an improved method, but is impractical for obtaining the hormone in the quantities required for extensive experimental and clinical investigation. It has been found that a fractionation of calcium hydroxide extracts of fresh bovine anterior pituitary glands, by means of ethanol at low temperatures along the lines successfully exploited by Cohn and his colleagues (4) in the separation of the plasma proteins, yields an abundance of crude fractions with high growth-promoting activity. Re-solution of these active fractions in dilute potassium chloride, followed by removal of the bulk of impurities in a precipitate formed at pH 5.0 and by a fractional precipitation with ethanol, starting at pH 8.5 to 8.7, yields a crystalline protein, electrophoretically homogeneous, which from its biological activity and other properties is identified as the anterior pituitary growth hormone. The yields of the crude primary fractions average about 33 gm. per kilo of fresh glands. The yields of the purification procedure lie between 8 and 16 per cent. On this basis, the new method can be expected to produce quantities