Year: 1934
Type: article
Abstract: DURING the past few years the problem of isolating insulin in a crystalline form has been of great interest to the author for several reasons.When large quantities of crystalline material are required for chemical or clinical investigations the present methods of separation leave much to be desired.In the hands of an experienced worker a fair degree of success may be attained.Nevertheless, there would seem to be some unknown condition or conditions which must be fulfilled if a good yield of crystals is to be obtained.Attempts to duplicate successful experiments do not always produce a crystalline product.This has been noted by severai workers, who have also drawn attention to the fact that different preparations of crude insulin show a marked variation in the ease with which they may be crystallised.Of further interest is the fact that it is a common belief that the chemistry of insulin is closely related to if not identical with that of proteins, and the results of an investigation concerning the exact con- ditions under which insulin crystallises would be of value for the isolation of other proteins in a pure form.Insulin was first isolated in a crystalline form by Abel [1926].The method utilised as the crystallising medium a solution which was highly buffered and consisted of several substances.In addition to ammonium acetate, it contained large quantities of brucine and pyridine.The isolation of the crystals was attri- buted to the fact that, by the use of such a complex system of weak bases, the acidity could be adjusted to the isoelectric point of insulin so slowly and so exactly that a supersaturated solution of insulin was obtained.The conditions then seemed to be such that the insulin separated in crystalline form.The problem of crystalline insulin was further investigated by Harington and the author [1929].Using a buffer solution of ammonium acetate containing saponin, they were able to prepare crystals from amorphous insulin.In microscopic appearance, physiological activity and chemical analysis, these crystals were identical with those described by Abel.In this paper we stated that we were " quite unable to offer any rational explanation of the mechanism of the action of saponin in promoting crystallisation."Nevertheless two factors were apparent.One was the power of saponin to inhibit precipitation and thus permit the preparation of a supersaturated solution of insulin at the isoelectric point.The other was the fact that saponin seemed to sharpen the isoelectric point of foreign proteins in the amorphous insulin and thus permit the removal of many of the impurities at a lower acidity than that at which the insulin crystallised.Since the publication of the saponin method, I have carried out many experi- ments on the crystallisation of insulin.In view of the fact that they have a distinct bearing upon the experimental work described later in the present paper, they should be mentioned briefly here.During the summer of 1929, many attempts were made to develop a method superior to the saponin procedure.It was felt that much would be gained if saponin could be replaced by some
Source: Biochemical Journal
DOI: 10.1042/bj0281592
Author: David Aylmer Scott
Institution: University of Toronto
Language: English
Subfield: Materials Chemistry
Field: Materials Science
Domain: Physical Sciences
Open Access status: bronze
OpenAlex ID: W4214923298