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Liquid Chromatography Tandem mass spectrometry method for Quantification of Buproprion and Hydroxy Buproprion in Human Plasma and its application to Bioequivalence Study

A rapid, specific and robust assay based on solid phase extraction and liquid chromatography-electronspray ionization tandem mass spectrometry (LC-ESI MS-MS) has been developed and validated for the quantitative analysis of Buproprion (a drug used for smoking cessation) in human plasma using Buproprion D9 as internal standard (ISTD). The precursors to product ion transitions of m/z 240.20/183.90 and m/z 256.10/237.90 were used to measure the analyte (Buproprion and hydroxy Buproprion) and the precursor to product ion transition of m/z 249.20/131.00 was used to measure the ISTD. The method was validated over a concentration range of 1.00ng mL-1 to 304.65ng mL-1 for Bupropion and 3.00ng mL-1 to 801.78ng mL-1 for hydroxy Bupropion. The method was validated over the various parameters like selectivity, matrix effect, sensitivity, linearity, precision, accuracy, stabilities (bench-top stability, standard stock solution stability in refrigerator and at room temperature, stock dilution stability, auto-sampler stability, freeze thaw stability, long-term stability at -65°C ± 10°C & -22°C ± 5°C, reagent stability, dry-extract stability, wet-extract stability and blood stability), effect of potentially interfering drugs, dilution integrity, recovery, reinjection reproducibility, ruggedness, extended batch verification, ion-suppression through infusion and inter-conversion check etc. The mean percent recovery of Bupropion was found 58.443% with a precision of 1.01% whereas the mean percent recoveries of hydroxy Bupropion and Bupropion D9 were found 62.327% and 66.513% with a precision of 2.99% and 3.91% respectively. The RSD percent of intra-day and inter-day assay was ;15%. The application of this assay was demonstrated in a bioequivalence study and it was found suitable for a study of sample size as big as sixty enrolled volunteers.

Author(s): Peeyush Jain, Nishant Paliwal, Naveen Dubey, Sandeep Sharma and Bhawani Singh

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