2.11. HPLC analysis
All HPLC analysis were carried out on a HPLC system (Agilent 1200 series, USA) equipped with a UV detector (Agilent G1314– 60,100) and reversed phase column (BDS C-18, 5 mm, 4.6 mm 250 mm). The mobile phase consisted of acetonitrile and water (40/60, v/v), delivered isocratically at a flow rate of 1.0 mL/min. The wavelength of the UV detector was set at 225nm. The HPLC analysis method was evaluated through specificity, linearity, limit of detection, intra- and inter-day precision. The chromatograms showed stable base line and no interference at the retention time of RPV. The calibration curves of RPV were linear over the concentration range of 0.1–40 mg/mL with correlation coefficient r 0.9997. The limit of detection was 0.1 mg/mL. The precision of the method indicated that all coefficients of variation were below 15m.