LC-MS Determination and Pharmacokinetic Studies of Ursolic Acid in Rat Plasma after Administration of the Traditional Chinese Medicinal Preparation Lu-Ying Extract

Sambucus chinensis L. is a native perennial herb distributed throughout China. In traditional Chinese medicine (TCM), this herb is known as Lu-Ying. Ursolic acid is the major eSective constituent of Lu-Ying. A rapid, sensitive, and accurate liquid chromatography-mass spectrometry (LC-MS) method for the determination of ursolic acid in rat plasma was developed and validated. Plasma samples taken from rats that had received Lu-Ying extract orally were aci- died with acetic acid and then extracted with a mixture of hexane-dichloromethane-2-propanol (20:10:1, v/v/v). Separation of ursolic acid was accomplished on a C18 column interfaced with a single quadrupole mass spectrometer. The mobile phase consisting of methanol and water (95:5, v/v) was delivered at a ‰ow rate of 1.0 ml/min. Atmospher- ic pressure chemical ionization was operated in negative-ion mode. Using selected ion-monitoring mode, the deprotonat- ed molecules (M-H) - at m/z 455 and 469 were used to quantify ursolic acid and glycyrrhetic acid (internal standard), respectively. The assay was shown to be linear over the range of 10―1000 ng/ml (r0.9960) with a lower limit of quan- tication of 10 ng/ml. The method was shown to be reproducible and reliable with intraday precision below 7.8% ,i nter- day precision below 8.1% ,a ccuracy within±4.3%, and mean extraction recovery excess of 83.6% ,w hich were all cal- culated from the blank plasma sample spiked with ursolic acid at three concentrations of 20, 200, and 800 ng/ml. The LC-MS method has been successfully applied to pharmacokinetic studies of ursolic acid after oral administration of Lu- Ying ethanolic extract (at a dose containing 80.32 mg/kg ursolic acid) to rats. The main pharmacokinetic parameters were: t1/2 ,4 .3 h;Ke ,0 .16 1/h; tmax ,1 .0 h;Cmax ,2 94.8 ng/ml; AUC0-t and AUC0-∞, 1007.1 ng·h/ml and 1175.3 ng·h/ ml, respectively. Key words―Lu-Ying extract; ursolic acid; pharmacokinetics; liquid chromatography-mass spectrometry