The present paper deals with HPLC method development for determination of amino acid. A high performance liquid chromatography (HPLC) method with UV detector is developed for determination of individual 17 Amino acids content in bulk drug substance without derivatisation and without any pretreatment of sample. The method is developed with market available Silica column by using isocratic mobile phase 2.5 mM Potassium dihydrogen phosphate with pH 2.85 and Acetonitrile with ration 25:75. The proposed method contains the high percentage of Acetonitrile in mobile phase, so that silica column works as HILIC column for amino acid analysis. The limit of quantitation for non aromatic Amino acids is in the range 1.2 – 2.1 µg/mL and for aromatic Amino acid is in the range of 8 – 12 ng/mL. The limit of detection for non aromatic Amino acids is in the range 0.4 – 0.65 µg/mL and for aromatic Amino acid is in the range of 2.5 – 3.6 ng/mL.. The method is accurate, linear and precise from LOQ level to 150% level with respect to analyte concentration for the analysis of Amino acid in bulk drug substance. The method is specific to resolve the degradation impurity formed in Acidic, Basic and Oxidative condition.