Objective: To establish a HPLC method for simultaneous determination of rutin, hyperoside, quercetin in Xiang lotus. Methods: The HPLC method was performed on an Agilent ZORBAX SB-C₁₈ column (4.6 mm×250 mm, 5 m) with gradient elution of methanol and 0.6% phosphoric acid solution. The flow rate was 1.0 mL/min. The column temperature was 30 ℃ and the detection wavelength was set at 255 nm. Results: The contents of rutin, hypericin and quercetin in 20 batches of lotus samples were 1.32～2.06, 1.36～2.43, 1.41～3.26 mg/kg, respectively. The linear range of the concentration of rutin, hypericin and quercetin was 5～200 g/mL (R²=0.999 9), the three flavonoids showed a good linearity, a high sensitivity, and good serviceability. The relative standard deviation (RSD) for precision, stability, repeatability, and recoveries test were all less than 4.0%, and the average recovery rate of added standard were 96.6%, 96.9%, 97.4%, respecitvely. Conclusion: The method is convenient and accurate, and can be used for the content determination and quality evaluation of flavonoids in Xiang lotus.