There is a paucity of normative data on hormonal levels among HIV-infected women. Hormonal levels may influence fertility and HIV-related immunological and virological factors. The objective of this study was to determine progesterone and estradiol levels during the menstrual cycle in HIV-seropositive women compared with high-risk seronegative women. The study enrolled 55 HIV-infected and 10 high-risk uninfected women with self-reported regular menstrual cycles (25-30-day cycles). Progesterone and estradiol levels were determined on a weekly basis for 8 weeks. The analysis included evaluations from the first complete menstrual cycle for the 54 HIV-infected and 9 uninfected women who had at least one complete cycle. The median age was 35 years for HIV-infected women and 36 years for uninfected women. The median CD4+ count for HIV-seropositive women was 210 cells/mm3. The median menstrual cycle length was 28 days (range 22-49 days) for HIV-infected women and 25 days (range 24-44 days) for uninfected women. The maximum progesterone level during the luteal phase was normal (>3.0 ng/ml) for 52 (96%) of 54 HIV-seropositive women and 7 (78%) of 9 HIV-seronegative women (p = 0.09, Fisher's exact test). The median maximum progesterone level was 12.2 ng/ml in HIV-seropositive women and 7.2 ng/ml in HIV-seronegative women (p = 0.07, Wilcoxon test). The median maximum estradiol value during the follicular phase was 148 pg/ml for HIV-seropositive women and 111 pg/ml for HIV-seronegative women (p = 0.04, Wilcoxon test). Among HIV-infected women, there were no significant differences in progesterone and estradiol levels by antiretroviral therapy, baseline plasma viral load, or median CD4+ cell count. We conclude that HIV-infected women with self-reported normal menstrual cycles have normal levels of progesterone and estradiol during the menstrual cycle.