Objective In the present study, we investigated the antioxidant and anti-aging effects of Silybum marianum protein hydrolysate (SMPH) in D-galactose-treated mice. Methods D-galactose (500 mg/kg body weight) was intraperitoneally injected daily for 7 weeks to accelerate aging, and SMPH (400, 800, 1, 200 mg/kg body weight, respectively) was simultaneously administered orally. The antioxidant and anti-aging effects of SMPH in the liver and brain were measured by biochemical assays. Transmission electron microscopy (TEM) was performed to study the ultrastructure of liver mitochondria.Results SMPH decreased triglyceride and cholesterol levels in the D-galactose-treated mice. It significantly elevated the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and total antioxidant capacity (T-AOC), which were suppressed by D-galactose. Monoamine oxidase (MAO) and malondialdehyde (MDA) levels as well as the concentrations of caspase-3 and 8-OHdG in the liver and brain were significantly reduced by SMPH. Moreover, it increased Bcl-2 levels in the liver and brain. Furthermore, SMPH significantly attenuated D-galactose-induced liver mitochondrial dysfunction by improving the activities of Na+-K+-ATPase and Ca2+-Mg2+-ATPase as well as mitochondrial membrane potential (ΔΨm) and fluidity. TEM showed that the degree of liver mitochondrial damage was significantly decreased by SMPH.Conclusion The results indicated that SMPH protects against D-galactose-induced accelerated aging in mice through its antioxidant and anti-aging activities.