A simple HPLC method with ultraviolet detection was developed for the simultaneous determination of baicalein and its major metabolite, baicalein 7-glucuronide (baicalin) in rat plasma. Following solid phase extraction with HLB cartridge, the analytes were separated using a gradient mobile phase consisting of methanol–acetonitrile–phosphate buffer. The flow-rate was set at 1ml/min and the eluent was detected at 320nm. The method is linear over the studied range of 1–10 and 0.05–1μg/ml for baicalein and baicalin, respectively. The intra-day and inter-day variations of the analysis were less than 6.38%, with relative error ranging from −3.6 to 6%. The limit of quantification in plasma was 0.05μg/ml for both baicalein and baicalin. The developed method proves to be an improved and more reliable method because of its capability for determining baicalein and baicalin simultaneously in a single chromatographic run.