Traditional Chinese medicinal plant Cinnamomum osmophloeum Kaneh (Lauraceae) has been used as spices and condiments. A simple and accurate reversed-phase high-performance liquid chromatographic (RP-HPLC) separation method was developed for the determination of two bioactive flavonol glycosides, namely kaempferol-3,7-O-α-L-dirhamnoside (kaempferitrin, 1), and kaempferol 3-O-β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl-7-O-α-L-rhamnopyranoside (2), in leaves of C. osmophloeum. Separation of the two compounds was achieved with a Hypersil BDS C18 column by gradient elution using acetonitrile-water (30:70, v/v) containing 0.1% trifluroacetic acid as a mobile phase. The flow rate and detection wavelength was set at 0.8 ml/min and 265 nm, respectively. Calibration curves were found to be linear within the range of 5–100 μg/mL. The contents of two flavonoids in C. osmophloeum leaves were successfully determined with retention times of 14.8 and 15.5 min. The recovery of the two flavonoids was 98.0 and 99.3, respectively. The results indicated that the developed HPLC assay could be readily utilized as a quality control method for C. osmophloeum and its related traditional Chinese medicinal preparations.