Catalase plays an important role in protecting organisms against oxidative damage caused by reactive oxygen species (ROS) by degrading surplus hydrogen peroxide. Addition of exogenous catalase can alleviate injuries caused by ROS. Thus, production of human catalase through genetic engineering will meet the increasing therapeutic demand for this enzyme. In this study, we successfully expressed the recombinant gene in mouse mammary gland, and biologically active human catalase was secreted into the milk of the transgenic mice. The peroxisomal targeting sequence (PTS) within the catalase gene had no significant negative effect on the secretion of the recombinant protein. Intake of the transgenic milk by the pups was found to decrease lipid peroxidation, increase the total superoxide dismutase (T-SOD) activity in the brain, and enhance the total antioxidative capacity (T-AOC) of brain, liver, and serum. To our knowledge, this is the first example of efficient production of biologically active human catalase in the milk of transgenic animals. Our study suggests that scaled-up production in transgenic farm animals would yield sufficient human catalase for biomedical research and clinical therapies.