Angiotensin-converting enzyme (ACE, EC 3.14.5.1) is found in particulate fractions of the epididymis but not in soluble epididymal fractions or in the testis of 4-week-old rats. [3H]Captopril autoradiography of testis and epididymis from 4-week-old rats confirms the association of ACE with epididymal ducts but not the testis. ACE appears in the testis between 4 and 6 weeks of age. Soluble ACE is not detectable in the epididymis until 6-7 weeks of age. Within the epididymis, regions closest to the testis develop soluble ACE activity about 1 week before those nearest to the vas deferens. Hypophysectomy of 10 week-old-rats depletes greater than 95% of ACE activity from the testis and soluble fractions of the epididymis, with little change in ACE levels from particulate fractions of the epididymis. [3H]Captopril autoradiography after hypophysectomy reveals luminal and epithelial ACE in the epididymis. The presence of particulate ACE in the epididymis under conditions where there is no testicular ACE indicates that the two forms are synthesized separately. However, soluble ACE from the epididymis might be derived from the membrane-associated ACE of the testis. Such a relationship is supported by the lag of 1 week between the development of ACE in the initial segment of the epididymis and the tail of the epididymis, and by the occurrence of soluble epididymis ACE only in those animals with testicular ACE activity.