A simple RP-HPLC method for quantification of columbianadin in rat plasma and its application to pharmacokinetic study.

A rapid and sensitive reversed-phase high-performance liquid chromatographic (RP-HPLC) method was developed to investigate pharmacokinetics of columbianadin, one of the main bioactive constituents in the roots of Angelica pubescens f. biserrata, in rat plasma after intravenous administration to rats at two doses of 10 and 20 mg/kg. The method involves a plasma clean-up step using liquid-liquid extraction by diethyl ether, followed by RP-HPLC separation and detection. Separation of columbianadin was performed on an analytical Diamonsil ODS C(18) column, with a mobile phase of MeOH-H(2)O (85 : 15, v/v) at a flow-rate of 1.0 mL/min, and UV detection was set at 325 nm. The retention time of columbianadin and scoparone (internal standard) was 6.7 and 3.5 min, respectively. The calibration curve was linear over the range of 0.2-20.0 microg/mL (r(2) = 0.9986) in rat plasma. The lower limits of detection and quantification were 0.05 and 0.1 microg/mL, respectively. The extraction recovery from plasma was in the range of 81.61-89.93%. The intra- and inter-day precisions (relative standard deviation) were between 1.01 and 9.33%, with accuracies ranging from 89.76 to 109.22%. The results indicated that the method established was suitable for the determination and pharmacokinetic study of columbianadin in rat plasma.