Objective To investigate the effect of Rhizoma Atractylodes macrocephala polysaccharide on ulcerative colitis (UC) mice by highlighting the Wnt/β-catenin signaling pathway.Methods Forty mice were randomly divided into control group, model group, Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group, with 10 mice in each group. UC was induced by dextran sodium sulfate (DSS), and mesalazine and Rhizoma Atractylodes macrocephala polysaccharide were administered by gavage to mice in the Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group, respectively. The body mass, intestinal propulsion rate and colon histopathological score were recorded. The expressions of Wnt1, β-catenin, C-MYC and cyclin D1 were detected via Western blotting. Serum levels of tumor necrosis factor- α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β), motilin (MTL), and gastrin (GAS) were determined via ELISA. The percentages of regulatory T cells (Treg), T helper 1 cells (Th1), T helper 2 cells (Th2), and T helper 17 cells (Th17) were detected.Results The body mass of mice in the four groups before and 1 week, 2 weeks and 3 weeks after the experiment were compared, and the results revealed that the body mass was different among the time points (F = 7.442, P = 0.001) and the groups (F = 35.614, P = 0.000), and that the body mass in the control group, Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group was higher than that in the model group. Besides, the change trends of the body mass were different among the four groups (F = 22.375, P = 0.000). The histopathological score of the model group was higher than that of the control group, the Western medicine group and the Rhizoma Atractylodes macrocephala polysaccharide group (P < 0.05), while the intestinal propulsion rate of the model group was lower than that of the control group, the Western medicine group and the Rhizoma Atractylodes macrocephala polysaccharide group (P < 0.05). The protein expressions of Wnt1, β-catenin, C-MYC and cyclin D1, serum levels of TNF-α, IL-6 and IL-1β, and percentages of Th1, Th2 and Th17 cells in the control group were lower than those in the model group, Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group (P < 0.05), whereas the expression of MTL, the serum level of GAS, and the percentage of Treg cells in the control group were higher than those in the rest groups (P < 0.05). The relative protein expression of Wnt1, β-catenin, C-MYC and cyclin D1, serum levels of TNF-α, IL-6 and IL-1β, and percentages of Th1, Th2 and Th17 cells in the model group were higher than those in the Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group (P < 0.05), while the expression of MTL, the serum level of GAS, and the percentage of Treg cells in the model group were lower compared with the Western medicine group and Rhizoma Atractylodes macrocephala polysaccharide group (P < 0.05).Conclusions Rhizoma Atractylodes macrocephala polysaccharide can inhibit DSS-induced UC, which may be achieved through down-regulating the Wnt/β-catenin signaling pathway to modulate immune function, mitigate inflammation, and promote the repair of damaged intestinal mucosa.