OBJECTIVE： To establish a method for simultaneous determination of paeoniflorin， berberine hydrochloride and ammonium glycyrrhizinate in Xiaoer huadu powder. METHODS： HPLC method was adopted. The separation was performed on Waters SunFireTM-C18 column with mobile phase consisted of acetonitrile-0.1％ phosphoric acid （gradient elution） at the flow rate of 1.0 mL/min. The detection wavelength was set at 238 nm， and column temperature was 25 ℃. The sample size was 10 μL. RESULTS： The linear ranges of paeoniflorin， berberine hydrochloride and ammonium glycyrrhizinate were 8.808-88.08 μg/mL（r＝0.999 8）， 1.778-17.78 μg/mL（r＝0.999 6）， 2.533-25.33 μg/mL（r＝0.999 9）， respectively. LOQ were 4.404， 0.889， 2.533 μg/mL； LOD were 1.101， 0.445， 1.267 μg/mL. RSDs of precision， stability and reproducibility tests were all lower than 2％. The recoveries were 95.08％-99.61％（RSD＝1.77％，n＝9）， 96.93％-99.94％（RSD＝0.92％，n＝9），98.33％-102.05％（RSD＝1.27％，n＝9）. CONCLUSIONS： The method is simple， accurate and reproducible， and can be used for simultaneous determination of paeoniflorin， berberine hydrochloride and ammonium glycyrrhizinate in Xiaoer huadu powder.