Objective To investigate the protective effects of berberine(BBR)against drug-induced liver injury(DILI)in mice and the change profiles in gut microbiota,and to elucidate the underlying mechanism by which BBR alleviates DILI through modulation of the gut microbiota.Methods Acetaminophen(APAP)administration was performed on C57BL/6J mice with or without pre-treatment of ampicillin-neomycin-metronidazole-vancomycin(ABX,depletion of gut microbiota)to establish a DILI model.Based on body weight,12 mice were randomly divided into an APAP group and a BBR group,with 6 animals in per group.Serum and liver tissue samples were collected from both groups to determine the effects of intragastrical administration of BBR on serum transaminase levels,inflammatory indicators,and liver histopathology.Fresh fecal samples were also collected to profile the gut microbiota using 16S rRNA gene sequencing.Then the alpha and beta diversity,as well as the overall microbial community structure,were compared between the 2 groups.Pearson correlation analysis was employed to analyze the relationships between differentially abundant bacteria and the serological indicators.Results Intragastrical administration of BBR significantly reduced APAP-induced serum levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST),and ameliorated the proportion of hepatocellular necrosis caused by APAP.The results of 16S rRNA gene sequencing revealed that there were no statistical differences in the alpha diversity indices(Chao1 and Shannon)between the APAP-BBR and APAP groups,but a significant difference was observed in beta diversity(P<0.05).Compared to the APAP-BBR group,the relative abundances of Alloprevotella and Ruminococcaceae_UCG_014 were significantly increased in the APAP group(P<0.05).Conversely,the relative abundance of Parabacteroides was significantly higher in the APAP-BBR group than the APAP group(P<0.05).Pearson correlation analysis demonstrated that Alloprevotella was strongly correlated with AST level(P<0.05),while both Ruminococcaceae and Parabacteroides showed strong correlations with ALT and AST levels(P<0.05).Conclusion Intragastric administration of BBR can mitigate the progression of DILI in mice,potentially through modulating the gut microbiota.