The serum concentration of estradiol (E2), testosterone (T), dihydrotestosterone (DHT) of 12 patients with hirsutism and amenorrhea were measured by radioimmunoassay, percent unbound E2 and T by equilibrium dialysis and concentration of sex hormone binding globulin (SHBG) by competitive ligand-binding assay. These were compared with values of 10 normal men and with values obtained throughout the follicular phase of 5 normal women. Only 4 of the 12 patients with hirsutism and amenorrhea had elevated serum T concentrations, the mean serum T concentration of this group being only 35% higher than the normals (p < 0.05). The mean serum E2 concentration (41.8 ± 6.0 pg/ml) was only 35% that of normal follicular phase E2 levels. The mean serum SHBG concentration of hirsute patients was 40% that of normal women and their mean unbound unconjugated E2 and T were 130% and 160%, respectively, of normal. Despite the high unbound fraction, the mean plasma unbound unconjugated E2 concentration was one-half the mean follicular phase levels. Nine of 12 patients with hirsutism had higher than normal concentrations of unbound unconjugated T resulting in a mean two times that of normal women. Because the ratio of unbound E2/T was abnormally low in 11 of 12 patients, it is suggested that the relative and absolute hypoestrogenism which is common in these patients may contribute to the hirsutism by potentiating the effect of T on hair growth. Similarly, subnormal serum SHBG concentrations in patients with hirsutism may also be a reflection of low serum unbound E2/T ratio.