OBJECTIVE: To establish a method for the determination of antiseptics in Anshen bunao oral liquid. METHODS: HPLC method was adopted. The determination was performed on Agilent Zorbax SB-C18 column with mobile phase consisted of acetonitrile-0.05% phosphorice acid solution (gradient elution) at the flow rate of 1.0 mL/min. The detection wavelength was set at 236 nm, and column temperature was 30 ℃. The injection volume was 10 μL. RESULTS: The linear range of benzoic acid and ethylparaben were 12.85-128.54 μg/mL(r=0.999 9) and 4.05-40.48 μg/mL(r=0.999 8), respectively. The limits of quantitation were 0.514, 1.012 μg/mL. The limits of detection were 0.171, 0.353 μg/mL. RSDs of precision, stability and reproducibility tests were all lower than 1.0%. The recoveries were 99.38%-100.21% (RSD=0.31%,n=6), 97.96%-100.72%(RSD=0.91%,n=6). CONCLUSIONS: The method is simple and accurate, and can be used for content determination of antiseptics in Anshen bunao oral liquid.