This dataset provides comprehensive insights into the impact of barley-based diets on inflammation and gut microbiota in Balb/cB&J mice. 50 mice were allocated into five different diets over six weeks: standard diet (SD), white rice (RD), whole-grain barley (WGB), anthocyanin-rich barley bran (BB), and beta-glucan-rich endosperm (PG). Each group comprised 10 animals, 5 males and 5 females. Serum samples were analyzed to measure six pro-inflammatory markers. Histological analysis of brain tissue was performed to evaluate inflammation in three distinct regions using F4/80 as a microglial activation marker. Colon samples were processed with immunohistochemistry to assess MUC2 expression. Faecal and ileum content samples were collected at multiple time points for microbiome analysis via 16S rRNA gene sequencing obtaining data on microbial alpha-diversity, beta-diversity and differential abundance analysis.