The disruption of the intestinal mucosal barrier is strongly associated with the onset of various diseases, including inflammatory bowel disease. Exopolysaccharides (EPS) support the functionality of the intestinal barrier. Weissella Cibaria (W. cibaria), belonging to the lactic acid bacteria, exhibits a significant capacity for EPS production. However, the specific mechanisms by which the EPS produced by W. cibaria confers intestinal barrier protection remain unexplored. Here, we characterized the polysaccharide, EPS-2, produced by W. cibaria isolated from the feces of healthy infants. EPS-2 was a novel dextran composed of α-(1 → 6) and α-(1 → 3,6) glycosidic linkages with a molecular weight of 845 kDa. EPS-2 alleviates intestinal mucosal barrier dysfunction in a mouse model of colitis, via a mechanism specifically reliant on the gut microbiota and their metabolic products, which is different from the well-known direct protective effects of other EPS on the intestinal barrier. EPS-2 reversed colitis-induced reductions in Muribaculaceae and propionate levels, thereby enhancing colonic goblet cell function and mucin content. Additionally, EPS-2 decreased the number of LPS-producing bacteria, such as Escherichia_Shigella. EPS-2 alleviated dextran sulfate sodium-induced intestinal inflammation and barrier damage. Therefore, EPS-2 shows promise as a postbiotic treatment for diseases associated with intestinal barrier dysfunction.