Determination of levo-tetrahydropalmatine in rat plasma by HPLC and its application to pharmacokinetics studies

The objective of this study was to establish a high performance liquid chromatography (HPLC) method for the determination of levo-tetrahydropalmatine (l-THP) in rat plasma, and to investigate the pharmacokinetics after oral administration of l-THP in rats. The plasma samples were extracted by ethyl acetate. The mobile phase consisted of a mixture of phosphoric acid (0.1%) and methanol (60: 40 v/v). The flow rate was 1.0 ml/min and the ultraviolet detection wavelength was at 280 nm. Plasma concentrations at different time were determined after oral administration at the dose of 20, 40 and 80 mg/kg. The data of concentration-time were fitted and the pharmacokinetics parameters were calculated with 3p97 program (Chinese Pharmacology Society). The limit of quantitation was 0.02 µg/ml, the linear range was 0.02-20.0 µg/ml (R2= 0.9989). The mean absolute recoveries of l-THP at three different concentrations (0.04, 5.00 and 20.00 μg/ml) were 97.5 ± 4.9, 98.2 ± 3.6 and 99.2 ± 3.2%, respectively. The relative standard deviations (RSD) of intra-day and inter-day were both less than 10%. The parameters of low, middle and high doses were as follows: t1/2α were (0.79 ± 0.04), (0.66 ± 0.02), (4.42 ± 0.07) h, t1/2β were (20.26 ± 1.21), (19.28 ± 1.04), (31.96 ± 0.85) h, while AUC were (6.95 ± 0.98), (9.91 ± 1.11), (19.19 ± 3.35) mg·h/L, respectively. The proposed method was found to be convenient, accurate and reliable, and it can be used for determination of l-THP in rat plasma. The pharmacokinetics studies also provided the theoretical foundation and reference for the safe and reasonable clinic exploitation of l-THP.   Key words: High performance liquid chromatography (HPLC), levo-tetrahydropalmatine, pharmacokinetics, rat.