ABSTRACT This study was designed (a) to examine the involvement of the Δ5-pregnenes, pregnenolone and 17-OH-pregnenolone in the abnormal steroidogenesis present in hirsute patients; and (b) to examine the possibility that suppressed adrenal 21-hydroxylase activity may frequently underlie the abnormal steroidogenesis in hirsute women. Basal plasma pregnenolone and 17-OH-pregnenolone levels were significantly elevated in hirsute women. The levels of these steroids following treatment with dexamethasone and the decrement induced by this treatment were also significantly greater in hirsute patients than in normal subjects. These precursor steroids were also excessively responsive to stimulation with HCG in the hirsute patients. Urinary 17-hydroxycorticosteroid excretion was significantly higher in hirsute patients than in normal subjects. In contrast, the urinary excretion of pregnanetriol, either under basal conditions or following stimulation with ACTH was not significantly higher than in normal women similarly treated. In a further attempt to identify any limitation in 21-hydroxylase activity, the steroidogenic response to ACTH plus metyrapone was measured in hirsute and in normal women. Neither the urinary pregnanetriol response nor the ratio of pregnanetriol to 17-hydroxycorticosteroids was elevated in hirsute women. These observations document that plasma pregnenolone, and 17-hydroxypregnenolone are elevated in hirsute patients. The excess appears to be derived both from the adrenals and the ovaries. However, we were unable to demonstrate any limitation in adrenal 21-hydroxylase activity in hirsute patients.