Highland barley (HB) displays a series of properties including regulation of lipid metabolism and attenuation of liver injury. Our study aimed to investigate the effect of modified highland barley (MHB) including fluidized highland barley (HB-1), extruded and puffed highland barley (HB-2), and ultrafine pulverized highland barley (HB-3) on lipid metabolism, liver inflammation, gut microbiota and metabolite profiles in mice fed with a high-fat/cholesterol diet (HFCD). 6 treatment groups were fed a normal control diet or an HFCD with or without MHB supplementation for 10 weeks. Results showed that MHB significantly improved lipid parameters, liver function and injury and blood glucose indexes related to hyperlipidemia compared with the HFCD group. In addition, MHB recovered the disorder of gut microbiota by increasing the Bacteroidetes/Firmicutes ratio and Lactobacillus and Allobaculum abundances and decreasing Proteobacteria abundance related to lipid metabolism bacteria. MHB reversed the decrease of short-chain fatty acid levels caused by the HFCD. Fecal metabolomics analysis showed that the important differential metabolites between HB-1 and HFCD were deoxycholic acid, myclobutanil and dibutyl phthalate, and the important differential metabolic pathways were arachidonic acid metabolism, ABC transporters and biosynthesis of unsaturated fatty acids. Results suggested that MHB especially HB-1 were better effective dietary intervention candidates to ameliorate hyperlipidemia compared with HB.