Ciprofibrate quantification in human plasma by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry for pharmacokinetic studies

Abstract

A rapid, sensitive and specific method for quantifying ciprofibrate in human plasma using bezafibrate as the internal standard (IS) is described. The sample was acidified prior extraction with formic acid (88%). The analyte and the IS were extracted from plasma by liquid–liquid extraction using an organic solvent (diethyl ether/dichloromethane 70/30 (v/v)). The extracts were analyzed by high performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC–MS/MS). Chromatography was performed using Genesis C18 4 μm analytical column (4.6 × 150 mm i.d.) and a mobile phase consisting of acetonitrile/water (70/30, v/v) and 1 mM acetic acid. The method had a chromatographic run time of 3.4 min and a linear calibration curve over the range 0.1–60 μg/mL (r > 0.99). The limit of quantification was 0.1 μg/mL. The intra- and interday accuracy and precision values of the assay were less than 13.5%. The stability tests indicated no significant degradation. The recovery of ciprofibrate was 81.2%, 73.3% and 76.2% for the 0.3, 5.0 and 48.0 ng/mL standard concentrations, respectively. For ciprofibrate, the optimized parameters of the declustering potential, collision energy and collision exit potential were −51 V, −16 eV and −5 V, respectively. The method was also validated without the use of the internal standard. This HPLC–MS/MS procedure was used to assess the bioequivalence of two ciprofibrate 100 mg tablet formulations in healthy volunteers of both sexes. The following pharmacokinetic parameters were obtained from the ciprofibrate plasma concentration vs. time curves: AUClast, AUC0–168 h, Cmax and Tmax. The geometric mean with corresponding 90% confidence interval (CI) for test/reference percent ratios were 93.80% (90% CI = 88.16–99.79%) for Cmax, 98.31% (90% CI = 94.91–101.83%) for AUClast and 97.67% (90% CI = 94.45–101.01%) for AUC0–168 h. Since the 90% CI for AUClast, AUC0–168 h and Cmax ratios were within the 80–125% interval proposed by the US FDA, it was concluded that ciprofibrate (Lipless® 100 mg tablet) formulation manufactured by Biolab Sanus Farmacêutica Ltda. is bioequivalent to the Oroxadin® (100 mg tablet) formulation for both the rate and the extent of absorption