The incidence rate of colitis and conversion of colitis into colorectal cancer is increasing. However, the results of drug treatments are inconsistent with variable side effects; therefore, it is necessary to find alternative ways of treating colitis, e.g. through dietary supplements. One such dietary supplement could be sulfur-containing amino acids, which are known to have anti-inflammatory, antioxidant, and gut microbiota homeostasis effects. Therefore, the aim of the present study was to explore the effect of methionine supplementation in the diet of mice on experimental dextran sulfate sodium (DSS)-induced colitis. Here, 24 male C57BL/6J mice were split into three experimental treatment groups in such a way that each treatment group had four replicates and each replicate had two mice. The control group was colitis-free, while colitis was induced by the administration of DSS in the DSS groups. In the DSS and DSS plus methionine (DSS + Met) groups, DSS was provided in drinking water containing 3% DSS on days 1-5 and later provided with purified water on days 6-7. It was found that supplementing with methionine could activate pathways like Nrf2, and inhibit pathways like TLR4 and Nlrp3 to realize anti-inflammatory and antioxidant effects. Moreover, methionine could alter the microbiota of the gut in the experimental mice, whereby exploration of the gut microbiota demonstrated that methionine supplementation in the diet increased the abundance of parabacteroides and the production of propionate and butyrate. The current study shows that the dietary prophylactic supplementation of methionine has a beneficial effect on resisting colitis, providing new insights for the prevention of colitis.