Epidemiological studies consistently show that excessive amounts of reactive oxygen species (ROS) is an important underlying cause of the initiation of oxidative stress in various tissue injuries as well as cell death and several chronic diseases. Several compounds, such as D-Galactose (D-Gal), bromobenzene, carbon tetrachloride (CCl₄), acetaminophen, and ethanol have been implicated in the etiology of liver diseases. D-Gal is commonly used as a toxin to induce reactive oxygen species and hepatic disorders, which has been extensively used in oxidative damage animal models to evaluate the therapeutic potential of drugs and dietary antioxidants. Due to the risks of synthetic antioxidants, there is a growing interest in the use of natural antioxidants to prevent ROS related liver pathologies. Therefore, during the past several decades, human nutrition and biochemistry research focused on antioxidants derived from foods that could prevent or diminish ROS-induced damage. More recently, there is growing interest in natural hydrolysed protein antioxidants from many plant and animal sources. So, we hypothesized that the administration of antioxidant peptides might be useful for preventing various types of oxidative stress in vivo. In our previous experimental studies, Fraction I (MW <1 ku), Fraction II (MW 1–4 ku) and Fraction III (MW >4 ku) were isolated from the hydrolysates of Scomberomorous niphonius skin through ultrafiltration membrane with the cutoff of 1 and 4 ku, respectively. Among them, the FractionⅡ (1–4 ku) exhibited the highest radical scavenging activity, inhibiting both lipid and protein oxidation of cooked patties during refrigerated storage, and protecting Caco-2 cells against the toxicity caused by H₂O₂. Therefore, the aim of this study was to evaluate the antioxidant effects of Fraction II on D-Gal-induced liver damage in rats. Wistar rats were randomly divided into 6 groups, which were normal control, negative control, D-Gal+Low-dose of Fraction II, D-Gal+Middle-dose of Fraction II, D-Gal+High-dose of Fraction II, and D-Gal +V_E as positive control for seven weeks. The activities of serum asparate aminotransferase (AST), alanine aminotransferase (ALT), monoamine oxidase (MAO) and antioxidant enzyme activities including superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), total antioxidant capacity (T-AOC), nitric oxide (NO) and malondialdehyde (MDA) contents in liver were determined. The result showed that all doses of Fraction II could significantly reduce the ALT and AST level in serum of the rat, while M-dose and H-dose of Fraction could sharply decrease the MAO level. Compared with negative control group, SOD, GSH-Px, CAT activity and T-AOC of liver of the rats were increased significantly. the MDA and NO contents in liver of the rats were reduced when treated with all doses of FractionⅡ. Treatment with high dose FractionⅡ (200 mg/kg) could increase the SOD, GSH-Px activity to 236.27 and 182.23 U/mg protein, which could reach the level of normal and positive control, decrease the contents of AST, ALT and MDA to 302.47 U/L, 220.43 U/L and 138.83 nmol/mg protein, which was close to the level of normal control. Moreover, Fraction II could significantly reduce the injury score of prominent nucleus, visible central veins and necrosis in rats. These data indicate that Fraction II can protect the liver against D-Gal induced oxidative damage.