Method development, validation and bioequivalence of varenicline in human plasma by liquid chromatography tandem mass spectrometry

• LC–MS/MS method for the analysis of varenicline in human plasma has been developed.
• Varenicline method was successfully validated.
• This validated method was successfully used in a bioequivalence study.

A method based on liquid chromatography coupled to tandem mass spectrometry was developed for quantitative determination of varenicline in human plasma. Varenicline and the internal standard (25.0 ng/mL of Clarithromycin) were extracted from human plasma by liquid–liquid extraction, using methyl tertiary butyl ether as the organic solvent. The chromatographic separation was achieved using C8 column with isocratic elution using a mixture of acetonitrile:0.001 M ammonium acetate (adjusted to pH 4.0) (70:30%, v/v). The method was validated over the concentration range of 0.1–10.0 ng/mL by investigating specificity, sensitivity, linearity, precision, accuracy, recovery, matrix effect and stability according to United State Food and Drug Administration guideline. The validated bioanalytical method was successfully applied to evaluate bioequivalence of two commercial products of 1 mg varenicline single dose.