Sofosbuvir is used to treat specific hepatitis C viruses and it is a direct acting antiviral agent. In this study, the HPLC method for the quantification of Sofosbuvir and its three degradation impurities (DIs) were developed and validated for Sofosbuvir drug substance. The specificity of the method was achieved in analytical column Zorbax XDB C18 (150mm X 4.6 mm, 5.0 µm) using a suitable mobile phase 10 mM Ammonium acetate buffer (pH 4.0 with diluted acetic acid) and Acetonitrile in the gradient programme. The flow rate is 1.0 mL/min. the injection volume is 1 µL, detection at 210 nm in UV and total run time is 70.0 minutes. The samples were made for forced degradation under hydrolysis, oxidation, thermal and photolytic conditions. The method was validated for specific, selective, sensitive, linear, rugged, robust and accurate as per the ICH guidelines. The linearity of the method for Sofosbuvir and its three DIs were found from QL level to 400 % concentration level with the correlation coefficient (r2) > 0.999. The accuracy for Sofosbuvir and its three DIs were performed from QL to 150% level concentration, and mean recovery was found from 98-102%. The degradation and validation study results indicate its unstable nature in acidic, basic, peroxide conditions and stable nature in thermal, neutral and photolytic and mass balance was achieved in all the stress conditions. Therefore, this method could be used in routine stability studies and quality control analysis