Objective To investigate the antioxidant effect of berberine in a rat model of nonalcoholic steatohepatitis (NASH) and an in vitro model of oxidative stress induced by H2 O2. Methods High-fat diet was used to establish a rat model of NASH. After three weeks of treatment with berberine 18 mg/kg/d by gavage, HE staining was used to observe liver pathological changes, and the activities of glutathione (GSH) and superoxide dismutase (SOD) and the level of malondialdehyde (MDA) in liver tissue were measured. An in vitro model of oxidative stress induced by reactive oxygen species (ROS) due to H2 O2 was established; flow cytometry was used to measure the level of ROS, and the activities of SOD, GSH, lactate dehydrogenase (LDH) , and chloramphenicol acetyltransferase (CAT) in hepatocytes were measured. A one-way analysis of variance was used for comparison of continuous data between groups, and the Bonferroni test was used for further comparison between two groups; the t-test was used for comparison of continuous data between two groups. Results NASH rats had significant improvements in the extent of infiltration of inflammatory cells, focal necrosis, and ballooning degeneration of hepatocytes after berberine treatment. Compared with the NASH model group, the berberine group had significant increases in the activities of GSH (29. 8 ±2. 4 U/mg vs 19. 9 ± 1. 3 U/mg, P < 0. 001) and SOD (26. 6 ± 1. 9 μg/mg vs 19. 7 ± 1. 4 μg/mg, P < 0. 001) and a significant reduction in the level of MDA (19. 8 ± 1. 5 nmol/mg vs 24. 0 ± 2. 0 nmol/mg, P < 0. 001) . In the in vitro model of oxidative stress induced by H2 O2, the H2 O2 group had a significant increase in the level of ROS compared with the berberine + H2 O2 group (69. 8% ± 1. 9% vs 50. 4% ±6. 5%, t = 24. 42, P = 0. 008) . Compared with the H2 O2 group, the berberine + H2 O2 group had significant increases in the activities of SOD (25. 5 ± 1. 3 μg/mg vs 18. 8 ± 1. 0 μg/mg, P < 0. 001) and GSH (27. 1 ± 0. 6 U/mg vs 16. 79 ± 3. 8 U/mg, P < 0. 001) in hepatocytes; there was no significant difference in the level of LDH between the berberine + H2 O2 group and the H2 O2 group (P = 0. 103) , and there was no significant difference in the activity of CAT across all groups (F = 3. 76, P = 0. 060) . Conclusion Both in vivo and in vitro models show that berberine has a good antioxidant effect, possibly by increasing the activities of antioxidant enzymes.