Determination of oleanolic acid in human plasma and study of its pharmacokinetics in Chinese healthy male volunteers by HPLC tandem mass spectrometry.
A highly selective and sensitive HPLC-ESI-MS-MS method was developed for the determination of oleanolic acid in human plasma. The oleanolic acid and glycyrrhetinic acid (internal standard) were recovered from plasma with ethyl acetate liquid-liquid extraction. The organic extracts were dried under a stream of warm nitrogen, reconstituted in mobile phase and injected into a Zorbax-Extend ODS analytical column (150 mm x 4.6 mm i.d., 5 microm), with the mobile phase consisting of methanol-ammonium acetate (32.5 mM) (85:15, v/v) pumped at a flow rate of 1.0 ml/min, and 30% of the eluent was split into a MS system with electrospray ionization tandem mass (ESI-MS-MS) detection in negative ion mode. The tandem mass detection was performed on a Finnigan Surveyor LC-TSQ Quantum Ultra AM tandem mass spectrometer operated in selected reaction monitoring mode. The parent to product ion combinations of m/z 455.4-->455.4 and 469.3-->425.2 at 38 V 1.5 mTorr Ar CID were used to quantify oleanolic acid and glycyrrhetinic acid, respectively. The assay was validated in the concentration range of 0.02-30.0 ng/ml for oleacolic acid when 0.5 ml of plasma was processed. The precision of the assay (expressed as relative standard deviation, R.S.D.%) was less than 15% at all concentrations levels within the tested range and adequate accuracy, and the limit of quantification was 0.02 ng/ml. The established method was applied for the pharmacokinetics study of oleanolic acid capsules in 18 healthy male Chinese volunteers with the mean values of C(max), T(max), AUC(0-48), AUC(0-infinity), t(1/2,) CL/F, and V/F of oleanolic acid after p.o. a single 40 mg dose obtained were 12.12 +/- 6.84 ng/ml, 5.2 +/- 2.9h, 114.34 +/- 74.87 ng h/ml, 124.29 +/- 106.77 ng h/ml, 8.73 +/- 6.11 h, 555.3 +/- 347.7 L/h, and 3371.1 +/- 1,990.1 L, respectively.
Song
Min
M
Department of Pharmaceutical Analysis, China Pharmaceutical University, Nanjing 210009, PR China.
Hang
Tai-jun
TJ
Wang
Ying
Y
Jiang
Li
L
Wu
Xiao-luan
XL
Zhang
Zhengxing
Z
Shen
Jianping
J
Zhang
Yindi
Y
D000085:Acetates / Q000032:analysis; D000328:Adult; D019540:Area Under Curve; D002138:Calibration; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002626:Chemistry, Pharmaceutical / Q000379:methods*; D002681:China; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D006034:Glycyrrhetinic Acid / Q000032:analysis; D006801:Humans; D008297:Male; D000432:Methanol / Q000032:analysis; D008956:Models, Chemical; D009828:Oleanolic Acid / Q000032:analysis / Q000097:blood*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D013997:Time Factors
D000085:Acetates; D009828:Oleanolic Acid; D006034:Glycyrrhetinic Acid; C018824:ammonium acetate; D000432:Methanol