Objective: To study the role of the prenatal environment in regulating reproductive development by measuring the prospective association between umbilical cord concentrations of sex hormone binding globulin (SHBG; principal regulator of sex steroid activity), bioavailable sex steroids, and age at menarche. Design: Prospective population-based cohort. Setting: Not applicable. Patient(s): In 286 female members of the Western Australian Pregnancy (Raine) cohort, concentrations of SHBG and steroids (estrogens: estrone, estradiol, estriol and estetrol [E4]; androgens: total testosterone, Δ4-androstenedione, androstenedione and dehydroepiandrosterone) were measured by liquid chromatography-tandem mass spectrometry from archived umbilical cord blood samples collected at birth. Bioavailable concentrations of testosterone and estradiol were calculated along with total composite measures of androgen and estrogen bioactivity. SHBG was measured by ELISA. Intervention(s): None. Main Outcome Measure(s): Age of menarche was calculated from date of menarche, collected prospectively by questionnaire sent home with participants at the year 10 follow-up. Result(s): Higher maternal education, higher body mass index, and the presence of antepartum hemorrhage were all significantly associated with earlier age at menarche. The bioavailable sex steroid measures accounted for 8.3% of the variance in age at menarche. Further, both SHBG and E4 concentrations accounted for a significant proportion of unique variance in age at menarche. Conclusion(s): Lower SHBG and higher E4 concentrations in umbilical cord blood were associated with earlier age at menarche. These results suggest that the prenatal sex steroid environment contributes toward pubertal development and age at menarche.