Abstract: OBJECTIVE To develop an HPLC method for the determination of enantiomers and diastereoisomers in tadalafil. METHODS A CHIRALPAK-IC column(250 mm×4.6 mm, 5 μm) was adopted. The mobile phase was acetonitrile-water(40:60) at the flow rate of 1.0 mL·min^-1 and injection volume of 20 μL. The UV-detection wavelength was set at 222 nm with the column temperature of 30℃. The isomer contents were calculated by self contrast method with the correction factor, and the correction factor of isomer A, isomer B and isomer C were 1.22, 1.07 and 1.25 respectively. RESULTS Calibration curves were found to be linear in the ranges of 25.3-379.5 ng·mL^-1 for isomer A, 50.6-379.5 ng·mL^-1 for isomer B and 50.4-378.0 ng·mL^-1 for isomer C(r=0.997 3-0.998 7). The average recovery(n=9) of isomer A, isomer B, and isomer C were 107.5%, 96.9%, and 98.5%, respectively. RSD of repeatability was 1.44%, 1.64% and 4.89% respectively, and the RSD of sampling precision was ≤2%. The LOD of isomer A, isomer B and isomer C were 12.7, 25.3, 25.2 ng·mL^-1. Isomer B and C were not detected (<0.01% of tadalafil) and the content of isomer A was up to 0.018% in 3 batches of samples. CONCLUSION The method is simple, accurate and reproducible. It can be reliable for the quality control of enantiomers and diastereoisomers of tadalafil.