The inhibitory effect of various gestagens on the pregnenolone 3.beta.-ol-dehydrogenase-.DELTA.5-4-isomerase system in 10,000 .times. g supernatants of human corpora lutea of the menstrual cycle was investigated in in vitro experiments. The Km value for pregnenolone was 1.3 .times. 10-6 M. All gestations reduced the rate of conversion of pregnenolone to progesterone. As compared with controls, average relative rates of reaction with additives were as follows: progesterone, 47%; 17-hydroxyprogesterone, 64%; norethisterone, 33%; chlormadinone acetate, 47%; medroxyprogesterone acetate, 53%; dydrogesterone, 62%; and allylestrenol, 74%. The data substantiate that natural or synthetic gestagens, at least in vitro, inhibit the conversion of pregnenolone to progesterone by human corpora lutea and that natural progestrone itself is a potent inhibitor which appears to act noncompetitively (Ki = 2.2 .times. 10-5 M).