Early Time Sequence in Pregnenolone Metabolism to Testosterone in Homogenates of Human and Rat Testis

Abstract

The time sequence of the metabolism of [4-14C] pregnenolone to testosterone in homogenates of human and rat testis was studied with special emphasis on the chain of events in the early 15 min of incubation. The incubations were performed at 32 C in the presence of NAD and a NADPH-generating system. The various intermediate steroids were separated by means of HPLC using a silica aliphatic diol column. Correction for procedural losses was performed by dual labeling. The present study confirms earlier reported results which showed that in the rat metabolism of pregnenolone to testosterone proceeds via the delta 4 pathway. However, this discloses for the first time that the conversion of pregnenolone proceeds very fast: progesterone, 17 alpha-hydroxyprogesterone, and 17 alpha-hydroxypregnenolone as the only important delta 5 intermediate, peak and decline again to almost undetectable levels within the first 15 min of incubation. Androstenedione and testosterone start to accumulate from 1 min on under the conditions used. In contrast, in the human testis, homogenates metabolism of pregnenolone to testosterone proceeds comparatively slowly and almost exclusively via the delta 5 intermediates dehydroepiandrosterone and androstenediol. Testosterone makes its appearance only after about 8 min of incubation. The data illustrate the importance of short-term incubations in evaluating the metabolism of steroids.