These studies were undertaken to determine the principal pathway of androgen biosynthesis by the testis of the marmoset Saguinus oedipus. Testicular fragments (25 mg) were incubated at 37 degrees C in Krebs-Ringer bicarbonate buffer, pH 7.4, containing pregnenolone-7-3H (3beta-hydroxy-5-pregnen-20-one) or progesterone-7-3H. Duplicate fragments were incubated with each substrate for 30 min, one hr, three hr, or five hr, for a total of 16 separate incubations. Metabolites were separated by paper and thin-layer chromatography, with identity established by recrystallization to constant specific activities and 3H/14C ratios. Pregnenolone was readily metabolized to progesterone, 17alpha-hydroxyprogesterone, androstenedione (4-androstene-3, 17-dione) and testosterone. Progesterone was converted to 17alpha-hydroxyprogesterone, androstenedione and testosterone. 17alpha-hydroxyprogesterone was the predominant metabolite obtained from both substrates at one, three and five hrs of incubation. Neither 17alpha-hydroxypregnenolone (3beta-17-dihydroxy-5-pregnen-20-one) nor dehydroepiandrosterone (3beta-hydroxy-5-androsten-17-one) was detected in the incubates. These data suggest a predominant delta4 pathway with accumulation of 17alpha-hydroxyprogesterone in the testis of this primate specie.