Abstract: OBJECTIVE To establish a high-performance liquid chromatographic method for the detection of 10 related substances in 20(S)-protopanaxadiol bulk drug. METHODS Used an ACQUITY UPLC BEH C₁₈ chromatographic column with ultrapure water(A)-acetonitrile(B) as mobile phase. The flow rate was 0.20-0.25 mL·min^-1, the detection wavelength was set at 203 nm, the column temperature was maintained at 35℃, and the injection volume was 5 μL. RESULTS 20(S)-protopanaxadiol and various impurities had reached an effective separation with a good linear relationship; the sample recovery(n=9) was ranging from 94.8% to 102.2%. The RSD values were <2.0%; the intra-day and inter-day precision RSD values were<2.0%, with a good precision; the related substances C, F, H, and I were stable in solution with RSD values<2.0% in 48 h at the room temperature; while impurities A, B, D, E, G and J were unstable under the same condition with RSD>2.0%. CONCLUSION The method is simple, sensitive and accurate, which can simultaneously determine 10 related substances in API 20(S)-protopanaxadiol, which can effectively identify the relevant substances in API and provide reference for the improvement of its production process and quality control.