Objective UPLC-MS/MS bio-analysis method was developed for the simultaneous determination ofberberine,naringin,hesperidin,and neohesperidin in plasma of rats.Methods UPLC Acquity BEH C18 (50 rmm × 2.1 mm,1.7 μm) column was used,mobile phases were containing 0.05％ formic acid and 2 mmol/L ammonium formate in water (A)-containing 0.05％ formic acid in acetonitrile (B) as the mobile phase gradient elution;SD rats were randomly divided into oral administration berberine group,Citrus aurantium extract group,and berberine and C.aurantium extract compatibility group.Results UPLC-MS/MS method could be applied to determination of berberine,naringin,hesperidin,and neohesperidin,method validation meets the requirements of biological sample analysis.When rats were administered with berberine and C.aurantium extract compatibility,the plasma concentration of berberine was much more than single dose of berberine group and the bioavailability of berberine was increased.Meanwhile,naringin and neohesperidin can be detected in rat's plasma.Conclusion The bioavailability of flavonoids is significantly improved as well compared to the single dose of C.aurantium extract.This suggests that berberine and C.aurantium extract compatibility has significant drug-drug interaction.