Ulcerative colitis (UC) is a typical type of inflammatory bowel disease, which is often recurrent and directly related with colorectal cancer. Therefore, early prevention and treatment for UC is very necessary. Therefore, it is necessary to find efficient substances to treat the UC with less side effects than drugs. Exopolysaccharides (EPSs) are important bioactive constituents of lactic acid bacteria. The study evaluated the effects of EPS1 produced by Levilactobacillus brevis M-10 on UC by determining the weight, the disease activity index (DAI) and the physiological and biochemical indexes. The pathological structures of colon were observed. The gut microbiota and the short-chain fatty acids were analyzed. The results demonstrated high dose (HD) of EPS1 (400 mg/kg-BW) that had the best effects on UC mice. The HD group restored body weight, decreased DAI and alleviated shortening of the length of the colon, recovered liver tissue, declined lipopolysaccharide, and myeloperoxidase. Also the HD group showed that the expression of tight junction proteins increased, IL-10 up-regulated, IL-6, IL-1beta, and TNF-alpha down-regulated, and the gut microbiota dysbiosis balanced. The HD group markedly elevated the relative abundance of Lachnospiraceae_NK4A136_group, Unclassified-Lachnospiraceae, and Unclassified- Muribaculaceae. Acetic acid, propionic acid, and n-butyric acid were significantly increased in the HD group (P < 0.05). The study could provide a theoretical basis and material support for the exploration of safe functional food in alleviating and preventing UC.