Abstract: OBJECTIVE To develop an HPLC method for simultaneous determination of tamarixetin and kaempferide in rat tissues after oral administration of the total flavonoides from Tamarix chinensis (TFT). METHODS Using Waters C18 column (4.6 mm×250 mm, 5 μm), the mobile phase was methanol-0.15% aqueous formic acid solution (pH 2.7) at a flow rate of 1.0 mL·min^-1 and detection wavelength was 254 nm, the column temperature was kept at 35 ℃. RESULTS Tamarixetin and kaempferide were simultaneous determined in several tissues, the highest level of tamarixetin was observed in lung, then in heart; and the highest concentration of kaempferide was determined in heart, then in lung. CONCLUSION The HPLC method is good in the aspect of easy operation and veracity, and can be used to study tissues distribution of tamarixetin and kaempferide.