Determination of asiatic acid in beagle dog plasma after oral administration of Centella asiatica extract by precolumn derivatization RP-HPLC.
A novel precolumn derivatization reversed-phase high-performance liquid chromatography (RP-HPLC) method with UV-vis detection for the quantitative determination of total concentration of asiatic acid (AA) in beagle dog plasma is described. AA was extracted with n-hexane-dichloromethane-2-propanol (20:10:1, v/v/v) from plasma, which had been hydrolyzed by acid and derivatized with p-Toluidine. Chromatographic separation was achieved on a C(18) column using gradient elution in a water-methanol system. Detection was set at UV wavelength of 248nm. A calibration curve ranging from 0.01 to 1.5microg/mL was shown to be linear, and the lower limit of quantification (LLOQ) was 0.01microg/mL. The intra- and inter-day precisions which were determined by three different concentrations (0.05, 0.2 and 0.8microg/mL) ranged from 4.4% to 13.1% and 4.6% to 14.2%, respectively. Mean extraction recoveries were no less than 65% for AA and ursolic acid (IS). Plasma samples containing asiatic acid were stable for 30 days at -20 degrees C. The method was successfully applied to a pharmacokinetic study in beagle dogs after oral administration of Centella asiatica extract, and the main pharmacokinetic parameters obtained were: T(1/2), 4.29h; T(max), 2.70h; C(max), 0.74microg/mL; AUC(0-t) and AUC(0-infinity), 3.74 and 3.82microgh/mL, respectively.
Zheng
Xiao-Chun
XC
Institute of Materia Medica, Zhejiang Academy of Medical Sciences, Hangzhou 310013, China.
Wang
Sheng-Hao
SH
D000284:Administration, Oral; D000818:Animals; D028041:Centella; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004285:Dogs; D004355:Drug Stability; D006868:Hydrolysis / Q000187:drug effects; D053978:Pentacyclic Triterpenes; D010936:Plant Extracts; D015203:Reproducibility of Results; D013997:Time Factors; D014315:Triterpenes / Q000008:administration & dosage* / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics*
C526146:Centella asiatica extract; D053978:Pentacyclic Triterpenes; D010936:Plant Extracts; D014315:Triterpenes; C017032:asiatic acid