A high-performance liquid chromatographic method for the determination of rifapentine in human serum was developed. The method utilized a Spherisorb C18 column, ultraviolet detection (336 nm), rifampin as internal standard and a calibration curve (C = 7.010 As/Ain +/- 0.156, r = 0.999) with reproducibility studies which yield a coefficient of variation (C.V.) of intra-day and inter-day assays lower than 10%. The average recovery of rifapentine from serum in the concentration range of 0.5 to 30 micrograms/ml was 92.93 +/- 9.704%.