Study of the determination and pharmacokinetics of bufadienolides in dog's plasma after administration of Liu-Shen-Wan by high performance liquid chromatography time-of-flight mass spectrometry.

A sensitive and reliable high performance liquid chromatography-tandem time-of-flight mass spectrometry method (HPLC/TOF MS) has been developed to determine three active bufadienolides from Liu-Shen-Wan (LSW) in dog's plasma. Enhanced selectivity and sensitivity in comparison with traditional HPLC/DAD method could be obtained through this method. Bufodienolides could be well separated and distinguished from its nominally isobaric endogenous components by HPLC/TOF MS, with the linear calibration range covering from 0.5 ng/mL to 100 ng/mL and Limit of Detection (LOD) being about 0.15 ng/mL. This method was also proved to be quite stable, with the intra-day precision and the inter-day precision results being lower than 6.39% and 7.44%, respectively. Meanwhile HPLC/TOF MS was successfully used in the pharmacokinetic study of LSW. For resibufogenin, the major pharmacokinetic parameters AUC0-t, Cmax and t1/2alpha were 160.72+/-21.97 ng/mL min, 2.35+/-0.71 ng/mL and 20.74+/-5.89 min, respectively, and for bufalin the corresponding parameters were 55.55+/-7.55 ng/mL min, 0.91+/-0.15 ng/mL and 25.45+/-13.28 min, respectively.