Chlorpromazine quantification in human plasma by UPLC–electrospray ionization tandem mass spectrometry. Application to a comparative pharmacokinetic study

In the present study a method to quantify chlorpromazine in human plasma using cyclobenzaprine as the internal standard (IS) is described. The analyte and the IS were extracted from human plasma by a liquid–liquid extraction with diethyl ether/dichloromethane (70/30, v/v) and analyzed by an ultra performance liquid chromatography (UPLC) coupled to an electrospray tandem triple quadrupole mass spectrometer in positive mode (UPLC–ES+-MS/MS). Chromatography was performed isocratically on an Aquity UPLC BEH C18 1.7 μm (50 mm × 2.1 mm i.d.) operating at 40 °C. The mobile phase was a mixture of 65% water + 1% formic acid and 35% of acetonitrile at a flow-rate of 0.5 mL/min. The lowest concentration quantified was 0.5 ng/mL and a linear calibration curve over the range 0.5–200 ng/mL was obtained, showing intra-assay precisions from 2.4 to 5.8%, and inter-assay precisions from 3.6 to 9.9%. The intra-assay accuracies ranged from 96.9 to 102.5%, while the inter-assay accuracies ranged from 94.1 to 100.3%. This analytical method was applied in a relative bioavailability study in order to compare a test chlorpromazine 100 mg simple dose formulation versus a reference in 57 volunteers of both sexes. The study was conducted in an open randomized two-period crossover design and with a fourteen days washout period. Plasma samples were obtained over a 144-h interval. Since the 90% CI for both Cmax, AUClast and AUC0–inf were within the 80–125% interval proposed by the Food and Drug Administration and ANVISA, it was concluded that chlorpromazine 100 mg/dose was bioequivalent to the reference formulation, according to both the rate and extent of absorption.

► UPLC–APCI-MS method to quantify chlorpromazine in human plasma.
► Method was applied in a relative bioavailability study in order to compare a test chlorpromazine 100 mg simple dose formulation.
► The chlorpromazine was extracted from human plasma by a simple liquid–liquid extraction.
► This method agrees with the requirements proposed by the US Food and Drug Administration of high sensitivity, specificity and high sample throughput in comparative pharmacokinetic assays such as bioequivalence.