Non-alcoholic fatty liver disease (NAFLD) is associated with low-grade chronic inflammation and intestinal dysbiosis. In this study, we investigated the potential benefits of sinapine, a rapeseed polyphenol known to exert anti-inflammatory and anti-oxidant effects, on high-fat diet (HFD)-induced NAFLD in C57BL/6 J mice and the underlying mechanisms. Four week-old mice were randomly divided into four groups and fed a low-fat diet (LFD), a HFD, a HFD with common rapeseed oil (HFD + CRO) and a HFD with sinapine in rapeseed oil (HFD + SRO) for 12 weeks. Supplementation with sinapine reduced the body weight of HFD mice by 10.99%, and decreased the levels of TG and LDL-C by 15.67% and 73.62%, respectively. In addition, sinapine also suppressed the intestinal NF-κB and TNF-α expressions and enhanced the adipose tissue IRS-1 expression in the HFD mice (P < 0.05). In terms of effects on the gut microbiota, sinapine induced a decrease in the ratio of Firmicutes to Bacteroidetes and increased the abundance of probiotics, such as Lactobacillaceae, Akkermansiaceae and Blautia, along with metabolite short-chain fatty acid (SCFA)-mediated upregulation of G protein-coupled receptor 43 (GPR43) to inhibit expression of inflammatory factors. Our collective results strongly supported the fact that the utility of sinapine as a prebiotic agent could prevent gut dysbiosis and obesity-related chronic diseases, such as insulin resistance (IR) and NAFLD.