Isinglass, a dried product of the swim bladder, has been widely used in traditional Chinese medicine. This study attempts to identify natural antioxidant peptides after digestion and absorption of isinglass in vivo. The antioxidant effects of dietary isinglass were demonstrated by evaluating the activities of SOD, CAT and MDA contents in the mouse liver. Four novel antioxidant-related peptides (RLLWENGNLL, GSKAENPTNPGP, SPVPDLVPGSF and VPDLVPGSF) were screened based on serum peptidomics and amino acid composition. Furthermore, pretreating with four peptides significantly increased the cell viability, and SOD and CAT activities of AML12 cells with H2O2-mediated oxidative damage, meanwhile, significantly reduced the ROS level, MDA content and apoptosis rate and attenuated DNA damage. Therefore, it was concluded that pretreatment of the identified peptides had a protective effect on oxidatively damaged cells. This result can aid in the recognition of active peptides from isinglass consumption for potential application in nutraceuticals or functional ingredients in food.