Development and Validation of the Stability indicating Assay Methodology Employing LC-MS/MS for Concurrent Quantification of Dapagliflozin Propanediol Monohydrate and Metformin Hydrochloride: Probable Degradants based on Mass Spectra

Abstract

For the concurrent measurement of dapagliflozin propanediol monohydrate (DAPA) and metformin hydrochloride (MET) in combined dosage form, a quick, accurate, specific and easy liquid chromatography tandem mass spectrometry (LC-MS/MS) approach was created. The method was performed on a column C8 RRHD Eclipse (150 × 4.60 mm, 5 μm). In 5 mM ammonium acetate buffer pH-4.0, methanol and acetonitrile were the components of the mobile phase in the ratios of 30:65:05, respectively. The effluent was detected at 227 nm at a 0.4 mL/min flow rate. The observed retention times for DAPA and MET were 7.297 and 3.230 minutes, respectively. The drug was stressed by being exposed to acid and alkali hydrolysis. From the mass spectra, it was found that two degradant peaks were observed in the standard mixture and the sample during alkali stress condition and probable degradants formed. The developed approach was validated in accordance with ICH guidelines. With correlation coefficients of 0.9969 for DAPA and 0.9975 for MET, it was discovered that the standard curve was linear over the range of 60 to 140 and 300 to 700 μg/mL for DAPA and MET, respectively. The limit of detection (LoD) was 2.959 and 8.893 μg/mL for DAPA and MET, respectively. The limit of quantification (LoQ) was 8.967 and 26.949 μg/mL for DAPA and MET, respectively. The %recovery was determined in between 98 to 102%. The precision was within the limit (Relative Standard Deviation (RSD) <2%). The proposed stability indicates that LC-MS/MS method can be successfully utilized for simultaneous estimation of DAPA and MET in combined dosage form without any prior separation of individual drugs and no interference was found due to degradant formed during stress condition.