High-pressure liquid chromatographic separation of a mixture of corticosteroids, androgens, and progestins

Fifteen steroids including corticosteroids, androgens, progestins, and their derivatives were completely separated by reverse-phase high-pressure liquid chromatography on a ChemcoPak 7 ODS-H column in 50 min. The elution procedures were first with water:methanol:acetonitrile:isopropanol 55:32:6.5:7.5 (v/v) for 15 min and followed with a linear gradient elution for 35 min from 0 to 80% of water:methanol:n-butanol 40:40:20 (v/v). The applicability of this method was successfully demonstrated in the analyses of the biological samples of carp plasma, testis, and head kidney.