The research work aims to develop a bioanalytical method using liquid chromatography and validated for the determination of canagliflozin by using an internal standard. Department of Pharmaceutical Chemistry, Smt. Kishoritai Bhoyar College of Pharmacy, New Kamptee, Nagpur (MS). Isocratic chromatography separation was achieved on an LC system with PDA detector on an ACE C18 (150mm× 4.6mm × 5µm) column using a mobile phase composition of acetonitrile: ammonium acetate buffer in the ration of 50:50 v/v (pH 4.5), orthophosphoric acid is used to adjust pH of mobile phase and the flow rate at 1.0ml/ min. and estimation was carried out at 291 nm. The retention time of a drug was 4.633 minutes. The method was validated for several parameters (specificity, linearity, precision, and accuracy) and also successfully applied for the pharmacokinetic in female rats. Calibration plot was linear (r2 > 0.9973) over the concentration range of 5-30 µg/ml for canagliflozin. The high recovery and low relative standard deviation (%RSD) confirm the suitability of the method. The result of Limit of Detection (LOD) and Limit of Quantitation (LOQ) were found to be 0.1099 μg/ml and 0.3331 μg/ml, respectively. The new RP-HPLC method can be conveniently adapted for examining canagliflozin concentration in rat plasma after oral administration.