UPLC-MS/MS METHOD FOR SIMULTANEOUS DETERMINATION OF METFORMIN AND GLIMEPIRIDE IN HUMAN PLASMA: A GREEN APPROACH TO ENVIRONMENT

Abstract

Reducing hazardous chemicals in chromatography and improving the fast, safe sample preparation process is the goal of an environmentally friendly and green approach. A fast and specific UPLC-MS/MS technique has been developed and validated to simultaneously determine metformin (MET) and glimepiride (GLM) in human plasma. Glipizide was used as an internal standard (IS), and analytes were extracted using the protein precipitation method with acetonitrile containing 0.1% formic acid. Chromatographic separations were performed on the BEH C18 column (1.7 µm, 2.1 mm x 50 mm) and pre-column. Elution was carried out through a gradient program using water and acetonitrile; both added 0.1% formic acid. Tandem mass spectrometry and positive ion-electrospray ionization, and the multiple reaction monitoring modes were used to detect fragment ions of the active ingredients. Precursor ion to fragment ion transitions included 130.0 → 70.9 m/z, 491.3 → 352.1 m/z, and 446.2 → 99.9 m/z for MET, GLM, and IS, respectively. The method was improved and verified in accordance with US Food and Drug Administration guidelines. Linear curves range from 1-100 ng/mL for MET and 0.25-25 ng/mL for GLM. The approach has quantitative limits of 1 ng/mL for MET and 0.25 ng/mL for GLM. The obtained recoveries were higher than 85%, while the coefficient of variation (%CV) was below 15% for all analytes. This study has provided an environmentally friendly solution, and improved sensitivity, and analysis time so it can be used to monitor drug concentration in plasma.