Abstract: Objective: To establish the purification process of Malus doumeri pectin（MDP), and to investigate the anti-inflammatory mechanism of the pectin on ulcerative colitis (UC) mice. Methods: The weighted composite score of galacturonic acid retention and protein removal was used as the response value, the pectin purification process was optimized through esponse surface methodology. The structural features of the pectin before and after purification were comparatively analyzed, by using ultraviolet spectroscopy (UV-vis), fourier infrared spectroscopy (FT-IR), X-diffraction (XRD), and scanning electron microscopy (SEM). Dextran sulfate sodium salt (DSS) was used to establish the UC mouse model, and to investigate the protective mechanism of pectin against UC. Results: X-5 resin was used as the purification medium, and the optimal conditions for purification were as follows: loading solution concentration of 7.5 mg/mL, pH5, and elution agent concentration of 10%. The deproteinization rate was 83.25%±1.35%, and the galacturonic acid recovery rate was 88.29%±1.39%. The galacturonic acid content of the purified pectin increased from 58.81%±0.75% to 67.04%±0.42%, meeting national food safety regulations. This process effectively removed protein and nucleic acid impurities, maintained the core polysaccharide structure and crystalline nature of the pectin, and homogenized its morphology. The pectin significantly improved the disease activity and colon histopathological injury in UC mice, effectively reduced the levels of TNF-α, IL-1β, and IL-6, increased the level of IL-10, inhibited the inflammatory response, and down-regulated the expression of TLR4/MyD88/NF-κB signaling pathway. Conclusions: The purification process obtained in this study is suitable for the purification of broad Malus doumeri pectin, and the purified pectin may protective effects against UC by regulating the TLR4/MyD88/NF-κB signaling pathway. The results of this study provide a theoretical basis for the in-depth development and application of Malus doumeri pectin.