It is reported that various diseases such as non-alcoholic fatty liver disease (NAFLD) are associated with imbalance of microbiome. And FXR has been well investigated in liver diseases. The objective of this study was to identify the role of farnesoid X receptor agonist obeticholic acid via targeting gut microbiota in NAFLD. Male C57BL/6 mice were fed either a normal-chow diet or a high-fat diet (HFD). Obeticholic acid(30mg/(kg·d)) and/or a combination of antibiotics were administered orally by gavage to mice for 12 weeks. Gut microbiota profiles were established through 16S rRNA amplicon sequencing. The effects of obeticholic acid on liver inflammation, the gut barrier, endotoxemia, gut microbiome and composition of the bile acid were also investigated. Obeticholic acid treatment can significantly improve obesity, circulation metabolism disorders, liver inflammation and fibrosis, and intestinal barrier damage caused by HFD. Removal of normal commensal bacteria can weaken the effect of obeticholic acid. The gut microbial structure was changed, and abundance of Blautia was increased significantly after treated with obeticholic acid. After obeticholic acid treatment, the concentration of taurine-bound bile acid caused by HFD was reduced in the liver. Taken together, these data suggest that obeticholic acid has aprotective effect on NAFLD via changing the components of gut microbiota, specifically increasing the abundance of Blautia.