Ion exchange chromatography of purified posterior pituitary preparations.

The methods employed previously in this laboratory for the isolation of oxytocin and vasopressin depend upon the use of several procedures to arrive at a highly purified sample. From the acetone powder of the posterior pituitary gland, the extraction and fractionation method of Kamm and coworkers (1) leads to a fraction rich in oxytocin and one rich in vasopressin, each at a stage of intermediate potency. Further purification involves the use of countercurrent distribution (2, 3). In the isolation of lysine vasopressin additional purification is obtained by the use of zone electrophoresis, followed by a final purification by countercurrent distribution (4). Purification by the technique of partition chromatography was applied to preparations of oxytocin and vasopressin (5). In the present work a study was undertaken on the ion exchange chromatography of posterior pituitary fractions of varying degrees of purity to obtain data on the complexity of these samples. It was also possible to determine whether this technique can be used to shorten the over-all purification. Basic proteins have been successfully chromatographed on the carboxylic acid ion exchange resin, IRC-50, and peptides resulting from the enzymatic hydrolysis of ribonuclease have been separated on the sulfonic acid ion exchange resin, Dowex 50 (6). Preliminary studies on the ion exchange chromatography of vasopressin and oxytocin were previously reported by Taylor (7). In continuing this work, conditions were investigated which could be employed for both analytical and preparative experiments. The majority of the experiments reported in this paper deal with the chromatography of vasopressin. The chromatography of lysine vasopressin at constant pH and ionic strength will be reported as an analytical procedure for the assessment of purity of preparations. The high resolving power of this procedure permitted the separation of lysine vasopressin from partially inactivated material. Conditions for the simultaneous separation of the two hormones, oxytocin aud lysine vasopressin, were found