Quantitation of six androgens by combined high performance liquid chromatography and radioimmunoassay

A technique was developed to separate six androgens (testosterone, dihydrotestosterone, androstenedione, 5 alpha-androstane-3 alpha, 17 beta-diol, 5 alpha-androstane-3 beta, 17 beta-diol, and androsterone) by high performance liquid chromatography prior to quantitation by specific radioimmunoassay systems. Methanol:water (60:40 v:v) was used as the solvent system with a C18 reversed-phase column. The method was verified and used to quantitate the androgens in serum from adult rams bled every 20 minutes for 6 hours and yearling bulls bled every 30 minutes for 8 hours. Concentrations of all 6 androgens varied in an episodic manner with testosterone being the dominant androgen.