Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease widespread. Increasing evidence suggest that the alterations in diet composition is associated with the epidemic of NAFLD. However, how dietary fat exposure influences the progression of NAFLD is smeared. In the present study, we aimed to explore the effects of dietary fat on NAFLD via metabolic profiles and gut microbiota. Male Wistar rats were placed on either normal chow diet (NCD) or high fat diet (HFD) for eight weeks. Liver histopathology and biochemical examinations of serum and liver were determined. GC/MS were used to elucidate the metabolic profiles of the serum, urine, liver and feces, and 16S rDNA sequencing were applied to determine the structure of gut microbiota. Typical NAFLD phenotypes in rats were identified with significant hepatic steatosis and dyslipidemia upon high dietary fat exposure. Metabolomics and microbiota analysis inferred that HFD disturbed energy metabolism and amino acid homeostasis, suppressed fatty acids oxidation, enhanced triglyceride synthesis, and promoted insulin resistance and inflammation. Our data demonstrated that dietary fat played a pivotal role in NAFLD development, and was related to glucose metabolism, lipid metabolism, amino acid metabolism and inflammation.