LC-MS/MS-ESI method for simultaneous quantitation of metformin and repaglinidie in rat plasma and its application to...

A highly sensitive and specific LC-MS/MS-ESI method has been developed for simultaneous quantification of metformin (MFN) and repaglinide (RGN) in rat plasma (50 μL) using phenacetin as an internal standard (IS). Simple protein precipitation was used to extract MFN and RGN from rat plasma. The chromatographic resolution of MFN, RGN and IS was achieved with a mobile phase consisting of 0.2% formic acid in water-acetonitrile (1:1, v/v) with a time program flow gradient on a Chromolith RP-18e column. The total chromatographic run time was 3.5 min and the elution of MFN, RGN and IS occurred at 1.64, 2.21 and 2.15 min, respectively. A linear response function was established for the range of concentrations 0.855-394 and 0.021-21.7 ng/mL for MFN and RGN, respectively. The intra- and inter-day precision values for MFN and RGN met the acceptance as per FDA guidelines. MFN and RGN were stable in battery of stability studies viz., bench-top, auto-sampler and freeze-thaw cycles. The developed assay was applied to a pharmacokinetic study in rats.