Quantification of brazilein in rat plasma following intravenous administration was achieved by reversed-phase high-performance liquid chromatography using a mobile phase of acetonitrile-0.05 m potassium dihydrogen phosphate water (containing 0.5% triethylamine, pH 3.0; 20:80 v/v) and UV detection at 445 nm. The method was linear (determination coefficient, r(2) = 0.9992) within the tested range (0.313-5.0 microg/mL). Intra- and inter-day precision coefficients of variation and accuracy bias were acceptable (maximal CV value was 2.06% for intra-day and 1.71% for inter-day) over the entire range. The recoveries were 81.48, 84.61 and 82.83% for concentrations of 0.313, 1.25 and 5.0 microg/mL, respectively. The concentration-time curve of brazilein after intravenous administration was fitted to the two-compartment model. This is the first time that brazilein in rat plasma was detected by HPLC-UV method and its pharmacokinetic characteristic was comprehensively studied.