Objective: A rapid, selective, sensitive, precise and accurate liquid chromatography in tandem with electro-spray ionization mass spectrometry method has been developed and validated for the simultaneous quantification of bupropion (BPR), hydroxyl bupropion (HBPR), erythrohydrobupropion (EHBPR) and threohydrobupropion (THBPR) in human plasma using only 100µL of human plasma sample. Methodology: Multi reaction monitoring detection was performed by electrospray ionization in the positive ion mode, conferring an additional selectivity to the method. The solid phase extraction technique was used for sample preparation. Chromatographic separation of drug and metabolites with better peak shape and resolution was achieved by using an Acquity BEH phenyl column with an isocratic elution of 42 % methanol and 58 % ammonia (0.06%, v/v) aqueous solution at a flow rate of 0.5 ml/min. Methanol was chosen because it enabled good resolution between THBPR and EHBPR as well as good peak symmetry of all the four analytes. Detection was carried out by mass spectrometry using positive electro-spray ionization mode, and the compounds were monitored using multiple reactions monitoring method. Deuterium-labeled isotopes of the compounds were used as internal standards. Results and Conclusion: No significant matrix effect was observed in the presented method. The assay method was validated over the concentration range of 1.75-500 ng/ml for BPR; 5-1000 ng/ml for HBPR; 0.5-100 ng/ml for EHBPR; and 2-500 ng/ml for THBPR as per FDA guideline and validated method was successfully applied for estimation of drug and metabolite concentration in the healthy adult volunteers, bioequivalence and pharmacokinetic study of Bupropion hydrochloride 300 mg extended release tablets under fasting condition.

A rapid and sensitive LC-MS/MS method has been developed and fully validated for simultaneous quantification of aspirin (ASP), salicylic acid (SLA), rosuvastatin (RVT), rosuvastatin lactone (RVL) and N-desmethyl rosuvastatin (DM RVT) in human plasma using a polarity switch with 400 µL of sample human blank plasma. Deuterated Internal Standards (IS) were used for each analyte. The solid phase extraction was used as sample preparation techniques. Chromatograph was monitored on a zorbax SB-phenyl column with a gradient mode with API-5500 triple quadrupole mass spectrometer as detector. The assay method was validated over the concentration range of 0.1-25 ng/mL for RVT; 50-10000 pg/mL for RVL and DMRVT; 5-2000 ng/mL for ASP; and 0.1-8 µg/mL for SLA. Intra and inter-day precision and accuracy were within acceptance limit. The mean recovery was >85% for all analytes. The analytes were stable at RT for 6 h in solution, at 2-8°C for 15 days in solution, for RT for 6 h in plasma, RT for 2 h in blood, till 3 freeze/thaw cycles, 104 h in auto-sampler at 6°C. This proposed method can be used for measurement of reliable concentration for dossier submission.The method was linear with weighing factor (1/x 2 ) in the range of 0.1-25 ng/mL for RVT, 50-10000 pg/mL for RVL and DMRVT, 5-2000 ng/mL for ASP and 0.1-8 µg/mL for SLA with intra and inter-day accuracy and precision within the acceptance criteria as per FDA and EMA guidelines (Table 4). The mean regression coefficient was >0.99 for all analytical run for all analytes.