By a combination of organic precipitation and high pressure liquid chromatography, human sperm acrosome reaction inducing activity has been purified from the fluid aspirated from preovulatory human ovarian follicles and identified as 4-pregnen-3,20-dione (progesterone) and 4-pregnen-17 alpha-ol-3,20-dione (17 alpha-hydroxyprogesterone). It is argued that progesterone is present at the site of fertilization of placental mammals in concentrations sufficient for activity, and hence provides a mechanism of inducing the acrosome reaction, an exocytotic event, in vivo.