Determination of chlorpheniramine in human plasma by HPLC‐ESI‐MS/MS: application to a dexchlorpheniramine comparative bioavailability study

Abstract

In the present study a fast, sensitive and robust validated method to quantify chlorpheniramine in human plasma using brompheniramine as internal standard (IS) is described. The analyte and the IS were extracted from plasma by LLE (diethyl ether–dichloromethane, 80:20, v/v) and analyzed by HPLC‐ESI‐MS/MS. Chromatographic separation was performed using a gradient of methanol from 35 to 90% with 2.5 mm NH4OH on a Gemini Phenomenex C8 5 μm column (50 × 4.6 mm i.d.) in 5.0 min/run. The method fitted to a linear calibration curve (0.05–10 ng/mL, R > 0.9991). The precision (%CV) and accuracy ranged, respectively: intra‐batch from 1.5 to 6.8% and 99.1 to 106.6%, and inter‐batch from 2.4 to 9.0%, and 99.9 to 103.1%. The validated bioanalytical procedure was used to assess the comparative bioavailability in healthy volunteers of two dexchlorpheniramine 2.0 mg tablet formulations (test dexchlorpheniramine, Eurofarma, and reference Celestamine®, Schering‐Plough). The study was conducted using an open, randomized, two‐period crossover design with a 2 week washout interval. Since the 90% confidence interval for Cmax and AUC ratios were all within the 80–125% interval proposed by ANVISA and FDA, it was concluded that test and reference formulations are bioequivalent concerning the rate and the extent of absorption