Objectives: Reactive oxygen species (ROS) have been associated with pathogenic processes including carcinogenesis through direct effect on DNA directly and by acting as a tumor promoter. Therefore, it has been regarded that ROS may be a major target for cancer prevention. The root of Paeonia lactiflora pall (PL), a traditional Chinese herb, has been a component of effective prescriptions for treatment of liver disease. Also, there are some reports about the antioxidant activities of the extracts from PL. However, little has been known about the effects of PL against oxidative damage. This work aimed to elucidate the anti-oxidant effects of Paeonia lactiflora pall (PL) in the non-cellular system and cellular system. Methods: Antioxidant activities of PL were evaluated by hydroxyl radical scavenging assay and Fe2+ chelating assay. Anti-oxidative effect of PL was evaluated by φX-174 RF I plasmid DNA cleavage assay in non-cellular system. In addition, DNA migration assay, expression level of phospho-H2AX, MTT assay and lipid peroxidation assay were performed for evaluate the anti-oxidative effect of PL in cellular system. Results: PL had a dose-dependent hydroxyl radical scavenging and Fe2+ chelating capacity. In addition, PL inhibited oxidative DNA and cell damage induced by hydroxyl radical in non-cellular system and cellular system. Conclusion: Taken together, P. lactiflora pall may be possible for the application to a potential drug for treating the oxidative diseases such as cancer.