Non-alcoholic fatty liver disease (NAFLD) is intricately linked to dysregulated hepatic lipid metabolism and gut microbiota imbalances. This study explores the potential mechanisms by which AEN, a lignan-rich extract from Myristica fragrans Houtt., alleviates NAFLD by affecting gut bacteria, bile acid metabolism, and fecal chemical composition. We demonstrate that AEN prevents weight gain, lipid accumulation, and enhances liver function in mice subjected to a high-fat diet. Utilizing a multi-omics strategy that includes microbiome analysis and metabolomics, we observed significant shifts in gut microbial composition and fecal metabolites, notably an increase in lithocholic acid, taurocholic acid, and the ratio of non-12-OH to 12-OH conjugated bile acids. These metabolic changes were accompanied by a decrease in bile acid metabolism-related enzymes, such as bile salt hydrolase, in the AEN group compared to the high-fat diet group. Moreover, AEN downregulated the farnesoid X receptor-fibroblast growth factor 15 pathway in the intestine, promoting the alternative bile acid synthesis pathway. The elucidation of the correlation between changes in the microbiome and fecal metabolites suggests that AEN modulates specific gut microbes, thereby influencing bile acid metabolism. This modulation was evident in the increased abundance of beneficial bacteria related to bile acid production, such as Parabacteroides and Bilophila, and the decreased abundance of harmful bacteria like Helicobacter and Streptococcus. Network pharmacology and GC ¡Á GC-TOF-MS suggested that licarin A and B could be key bioactive lignans in AEN. These findings underscore the potential of AEN in countering NAFLD by targeting the gut-liver axis, offering a novel therapeutic avenue for NAFLD management.