A NEW BIOANALYTICAL METHOD DEVELOPMENT & VALIDATION FOR SIMULTANEOUS ESTIMATION OF METFORMIN HYDROCHLORIDE AND SITAGLIPTIN PHOSPHATE MONOHYDRATE IN HUMAN PLASMA BY USING RP-HPLC

A new RP-HPLC method for the quantitative determination of metformin and sitagliptin in human plasma was developed and validated as per US-FDA guidelines. The drug was spiked in the plasma and extracted with mobile phase by precipitation method. The extracted analyte was injected into Symmetry C18 (4.6 x 150 mm, 3.5μm, Make: XTerra) or equivalent, maintained at ambient temperature and effluent was monitored at 254 nm. The mobile phase consisting of potassium dihydrogen phosphate [pH 5.8]: acetonitrile [HPLC Grade] (65:35 v/v). The flow rate was maintained at 0.9 mL/min. The calibration curve for metformin and sitagliptin was linear from 10.0 to 35.0 µg/mL (r2= 0.999) and 1.0 to 3.5 µg/mL (r2= 0.998) respectively. The inter-day and intra-day precision was found to be within limits. The lower limit of quantification (LLOQ) for metformin and sitagliptin were found to be 0.026 and 0.70 μg/mL respectively. The average % recovery for metformin and sitagliptin were found to be 98.82-100.03 & 99.76-100.89 % respectively and reproducibility was found to be satisfactory. This RP-HPLC method is suiTable for determining the concentration of metformin and sitagliptin in human plasma and it can applied for routine analysis for determination of the metformin and sitagliptin from dosage form during pharmacokinetic study.