An RP-HPLC method for simultaneous separation and quantification of pantoprazole and its five main impurities in pharmaceutical formulations was developed and validated. The separation was accomplished on a Zorbax Eclipse XDB C18 column (5 m particle size, 150 4.6 mm id) using a gradient with mobile phase A [bufferacetonitrile (70 + 30, v/v)], and mobile phase B [bufferacetonitrile (30 + 70, v/v)]. The buffer was 0.01 M ammonium acetate solution with addition of 1 mL triethylamine/L of the solution, adjusted to pH 4.5 with orthophosphoric acid. The eluent flow rate was 1 mL/min, the temperature of the column was 30C, and the eluate was monitored at 290 nm. Linearity (r = 0.999), recovery (97.6105.8), RSD (0.551.90), and LOQ (0.0991.48 g/mL) were evaluated and found to be satisfactory. The proposed method can be used for simultaneous identification and quantification of the analyzed compounds in pharmaceutical formulations.