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Clonazepam quantification in human plasma by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry in a bioequivalence study
Authors:Cavedal Luiz E  Mendes Fabiana D  Domingues Claudia C  Patni Anil K  Monif Tausif  Reyar Simrit  Pereira Alberto Dos S  Mendes Gustavo D  De Nucci Gilberto
Institution:Department of Pharmacology, Faculty of Medical Sciences, State University of Campinas (UNICAMP), Campinas/SP, Brazil.
Abstract:A rapid, sensitive and specific method for quantifying clonazepam in human plasma using diazepam as the internal standard (IS) is described. The analyte and the IS were extracted from plasma by liquid-liquid extraction using a hexane/diethylether (20 : 80, v/v) solution. The extracts were analysed by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC-MS-MS). Chromatography was performed on a Jones Genesis C8 4 microm analytical column (100 x 2.1 mm i.d.). The method had a chromatographic run time of 3.0 min and a linear calibration curve over the range 0.5-50 ng/ml (r2 > 0.9965). The limit of quantification was 0.5 ng/ml. This HPLC/MS/MS procedure was used to assess the bioequivalence of two clonazepam 2 mg tablet formulations (clonazepam test formulation from Ranbaxy Laboratories Ltd and Rivotril from Roche Laboratórios Ltda as standard reference formulation).
Keywords:clonazepam  healthy volunteer  plasma  pharmacokinetic  HPLC‐MS‐MS
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