To establish an HPLC mehod for the analysis of pharmacokinetics of salvianolic acid B in rats. METHODS: The biological samples were extracted with acetic ether. The chromatographic conditions were as follows: Hypersil ODS column (200 mm×4.6 mm, 5μm) was used. The mobile phase was acetonitrile-water(with Ammoniom Acetate 0.25 mol/L) was set at 328 nm. RESULTS: Salvianolic acid B was injected intravenously at doses of 1.6, 3.2, 6.4 mg/kg. The terminal elimination half-life(t1/2) of α phase and β phase was (3.1±0.1) min and (31.5±3.2) min. The extents of excrement,urine and biliary excretion of salvianolic acid B were 1.43%±0.90%, 0.77%±1.01% and 8.82%±4.11%. The tissue concentration of salvianolic acid B was as followed in order: Cheart>Cliver>Clung>Cintestine>Ckidney>Cspleen>Cstomach. The plasma protein binding rate of salvianolic acid B in human plasma and in rat was similar(89.2%±1.8%,92.5%±1.5%). CONCLUSION: The method is accurate, stable and reliable, and can be used for the investigation of salvianolic acid B in pharmacokinetics research. Salvianolic acid B eliminates fast and it shows a high plasma protein binding rate, the mainly excretion way of salvianolic acid B is from biliary.