Objective: To investigate the content of catalpol and acteoside in Rehmannia glutinosa with different varieties from genuine producing area. Method: Catalpol and acteoside were reflux extraction with methanol and determined by HPLC. The chromatographic column was Dikma Diamonsil C18(4.6 mm×250 mm

5 μm); the flow rate was 1.0 mL·min-1;the column temperature was kept at 30℃;the mobile phase consisted of acetonitrile-0.1% phosphoric acid solution(1:99) and the detection wavelength was set at 210 nm for catalpol;the mobiles phase consisted of acetonitrile-0.1% acetic acid solution(16:84) and the detection wavelength was set at 334 nm for acteoside. Result: The content of catalpol and acteoside was different in different varieties from genuine producing area and the differences also exist in the fresh and dried R. glutinosa. Conclusion: The quality of varieties of R. glutinosa was different and it was necessary to screen the optimal variety.