PURIFICATION OF CORTICOTROPHIN-RELEASING FACTOR (CRF) FROM POSTERIOR PITUITARY POWDER OF HOG ORIGIN

Abstract

For the investigation of corticotrophin-releasing factor (CRF), it is necessary to prepare pure CRF, especially in a vasopressin-free form, since vasopressin itself can release ACTH in vivo and in vitro. Attempts were made to purify vasopressin-free CRF from the posterior pituitary powder of hog origin in the present study. The acetone-ether precipitate of posterior pituitary powder was subjected to CMC column chromatographandy CRF-vasopressin mixture was obtained. This crude material was purified by partition chromatography on Sephadex G-25 and separation of CRF from vasopressin was satisfactorily achieved. This purified CRF (PB-III) which was still contaminated with 15IU/mg of vasopressin was active in ACTH release at a dose of 0.04 [mu]g in dexamethasone-Nembutal treated rats and yielded a single spot on a paper chromatogram. For further purification, gel filtration on Sephadex G-2 was carried out on the purified CRF. Very highly purified CRF could be obtained with 7.5IU/mg of pressor contamination; this was active in ACTH release between 0.01 and 0.08 [mu]g per rat with a linear dose response. This is the highest purification of CRF ever reported. Specific activity of CRF and contamination of vasopressin were listed on several CRF preparations obtained at various stages of purification and the methods of purification of CRF from vasopressin were discussed.