A precise and reproducible HPLC method has been established and validated for determination of salvianolic acid B (SaIB) in rat plasma after oral administration of Radix Salviae Miltiorrhizae extract. Liquid-liquid extraction was adopted for the sample preparation. Separation was accomplished on a C, column with a linear gradient elution consisting of acetonitrile and aqueous phosphoric acid. Ultraviolet detection was at 280 run. The method was validated over the concentration range 10.8-259.4 mug/mL using 1 mL of plasma. The assay was linear over this concentration range with a coefficient of variation less than 7%. The extraction recovery of SaIB was within the range 71-83% with RSD 11%. The mean recovery of the internal standard was 84% (n = 6) with RSD of 5.6%. This method is suitable to determine SalB in plasma and to investigate the pharmacokinelics of SalB. Copyright (C) 2004 John Wiley Sons, Ltd.