Abstract: This study was designed to explore the hepatoprotective effects of enzymatic degradation products derived from sturgeon roe on alcohol-damaged hepatocytes and to elucidate the underlying mechanisms. Utilizing sturgeon roe as the research material, the enzymatic process was optimized by focusing on key parameters such as enzymatic efficiency and in vitro activity. The protective effects on hepatocytes were systematically investigated by using the HepG2 cell model. Advanced computerized virtual screening technology was employed to identify potential bioactive peptides within the sturgeon roe. The findings indicated that the optimum enzymatic product of sturgeon roe was obtained under the conditions of 1:1 compounding of alkaline protease (1343 U/g) and trypsin (27 U/g) for 8 h, yielded a protein recovery of 49.30%±0.57%, a hydrolysis degree of 46.05%±0.92%, and ABTS⁺ free radical scavenging rate of 51.45%±0.66%. Notably, at a concentration of 5 mg/mL, this product significantly enhanced the activity of alcohol dehydrogenase (ADH) by 168.64%±1.42% (P<0.05). The product prepared under this enzymatic condition at 0.3~2 mg/mL significantly elevated superoxide dismutase (SOD) activity, glutathione (GSH) contents, and reduced malondialdehyde (MDA) contents in alcohol-injured HepG2 cells compared to the model group (P<0.05). In addition, the product at 0.3~2 mg/mL also significantly reduced alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities (P<0.05). Virtual screening identified 5 peptides, including LPG and FLPR, in the sturgeon roe enzymatic degradation product, showed promising hepatoprotective properties against alcohol-induced hepatocyte injury. Consequently, this study lays a solid foundation for the further development and utilization of aquatic resources, such as sturgeon roe, in hepatoprotective applications.