Study Objective. To characterize the concentration-time profiles of zidovudine and zidovudine-glucuronide in semen and serum of men infected with the human immunodeficiency-1 virus (HIV-1). Design. Open-label observational study. Setting. University-affiliated teaching hospital and research center. Patients. Four asymptomatic HIV-1-infected men. Interventions. Zidovudine administration was followed by an 8-hour intensive pharmacokinetic study on day 1. Over the next 8 days, a dose administration and timed single-sample strategy was employed to determine serum and semen concentration time profiles simultaneously. Measurements and Main Results. Zidovudine and zidovudine-glucuronide concentrations were uniformly higher in semen than in serum except at 1 hour after the dose. The median area under the curve ratio (semen AUC0-48: serum AUC(0-∞)) was 3.31 for zidovudine and 15.04 for zidovudine-glucuronide. Conclusion. Zidovudine and zidovudine-glucuronide reach high levels in seminal plasma relative to serum. The virologic, pharmacodynamic, and public health implications of distribution to this compartment require further study.