Abstract: High-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS) was used for separation and identification of flavonoid metabolites in rat urine，feces and bile after oral administration of the Citrus sinensis extraction.The HPLC experiments used a reversed-phase Zorbax SB-C ₁₈ column with the column temperature of 35 °C and a binary mobile phase system consisting of aqueous formic acid (0.1%) and methanol-acetonitrile (3∶2) using a gradient elution with the flow rate of 1.0 mL/min.The ESI-MS/MS was operated with a triple-quadruple mass spectrometer in negative ion mode.A total of 34 flavonoid compounds in vitro and 40 flavonoid compounds including 14 parent compounds and 26 metabolites in vivo were detected，and they were tentatively identified.The identification of these metabolites provided essential data for further pharmacological and clinical studies of Citrus sinensis Osbeck.and related preparations.