Abstract: Objective:To establish an HPLC method for simultaneous determination of 2α-hydroxy-ursolic acid, oleanolic acid and ursolic acid in Linearstripe Rabdosia Herb from different species. Methods:The HPLC system consisting of Synergi 4u Fusion RP 80A C18 column (250mm×4. 6mm, 5μm, Phenomenex company) and a solution system of methanol-phosphoric acid (V∶ V=9∶ 1) as the mobile phase with isocratic elution was adopted. The flow rate was set at 0. 6mL·min-1, the detection wavelength was 210nm and the column temperature was 30℃. Results: The linear ranges of 2α-hydroxy-ursolic acid, oleanolic acid and ursolic acid were 0. 111～1. 111mg/mL (r=0. 9999) , 0. 026～0. 256mg/mL (r=0. 9999) , 0. 307～3. 067mg/mL (r=0. 9999) respectively. Conclusion:The average recoveries (n=6) were 97. 72% (RSD=1. 24%) , 97. 08% (RSD=1. 62%) , 97. 46% (RSD=1. 25%) respectively. The method was quick, simple and accurate with good reproducibility and could be used for determination and quality control of the three main components in Linearstripe Rabdosia Herb.