STUDIES ON THE ASSAY OF ALDOSTERONE: CHROMATOGRAPHY AND CHEMICAL DETERMINATION1

Abstract

Chlorofbrm extracts from urines were prepared for the determination of aldosterone. These were subjected to various types of paper chromatography and the purified extracts evaluated by the ultra violet absorption at 240 mu and by the blue tetrazolium color reaction. These results were compared with those obtained by bioassay. A sequence of 4 chromatographic systems was considered necessary for most of the urines to obtain a satisfactory agreement between the chemical and the biological determinations. The chromatographic systems were applied in the following sequence: (1) chloroform-formamide; (2) toluene-ethyl acetate-methanol-water; (3) isooctane-tertiary butanol-water and (4) benzene-methanol-water. With this method, aldosterone was isolated from pregnancy urine and identified by the infra red spectra of the isolated material and its lactone. The comparison between the results of the bioassay and the chemical determination at various stages of the chromatography indicated that some urine extracts contained, even after separation of hydrocortisone, cortisone and corticosterone from aldosterone, one or possibly more substances which caused a lowering of the results obtained by the bioassay. One such compound appears to be separated from aldosterone by development in the E2B system.