Development and validation of empagliflozin in human plasma using nevirapine as internal standard by liquid chromatography-tandem mass spectrometry

Abstract

The present LC-MS/MS method for the estimation of Empagliflozin in human plasma by using Nevirapine as an internal standard was established and validated as per ICH guidelines. The best response was obtained with Phenomenex Synergi,4µm,4.6×75mm, and a mobile phase containing a mixture of 5mM Ammonium Acetate buffer with 0.1% Formic Acid: Acetonitrile (50:50 v/v) was delivered at a flow rate of 0.900 mL/min by positive ion mode (API 4000Q Trap) with an injection volume of 20 µL and a run time of 3 min. Detection is performed by atmospheric pressure electrospray ionization (ESI) tandem mass spectrometry in positive ion mode. The precursor to product ion transitions is m/z451.52>71.32 for Empagliflozin and m/z 267.3>226.8 for Nevirapine (Internal standard) were used for quantization. The retention time of Empagliflozin and Nevirapine (Internal standard) was found to be 1.50 min and 2.40 min respectively. Linearity was established for Empagliflozin in the range of 2.0pg/mL to 979.9pg/mL with a correlation coefficient (r=0.999) and the overall percentage recovery was 94.63 % for Empagliflozin and 91.94 % for Nevirapine (Internal standard) respectively.