Objective:To establish an HPLC method for content determination of vaccinoside in leaves of Vaccinium bracteatum Thunb.. MethodsBy tracking the dyeing activity,the active ingredients were isolated and identified. The content determination was performed by HPLC with a Cosmosil-5C18-PAQ column (4.6 mm×250 mm,5 μm). The mobile phase was 0.3% acetic acid- acetonitrile with gradient elution at a flow rate of 1.0 ml/min. The wavelength of detection was 320 nm.  Results:Vaccinoside was separated and identified as the main active ingredient. The chromatographic analysis showed a good separation. There was a good linearity in the range of 0.052 08-0.520 8 g/L(r=0.999 8). The recovery rate was 98.61%.  Conclusion:The determination method is quick and accurate,which could provide reference for the quality control of vaccinoside in leaves of Vaccinium bracteatum Thunb..