Semi-synthetic triterpenoids, bearing cyano enone functionality in ring A, are considered to be novel promising therapeutic agents with complex inhibitory effects on tissue damage, inflammation and tumor growth. Previously, we showed that the cyano enone-containing 18βH-glycyrrhetinic acid derivative soloxolone methyl (SM) effectively suppressed the inflammatory response of macrophages in vitro and the development of influenza A-induced pneumonia and phlogogen-stimulated paw edema in vivo. In this work, we reported the synthesis of a novel 18βH-glycyrrhetinic acid derivative trioxolone methyl (TM), bearing a 2-cyano-3-oxo-1(2)-en moiety in ring A and a 12,19-dioxo-9(11),13(18)-dien moiety in rings C, D, and E. TM exhibited a high inhibitory effect on nitric oxide (II) production by lipopolysaccharide-stimulated J774 macrophages in vitro and dextran sulfate sodium (DSS)-induced colitis in mice, displaying higher anti-inflammatory activity in comparison with SM. TM effectively suppressed the DSS-induced epithelial damage and inflammatory infiltration of colon tissue, the hyperproduction of colonic neutral mucin and TNFα and increased glutathione synthesis. Our in silico analysis showed that Akt1, STAT3 and dopamine receptor D2 can be considered as mediators of the anti-colitic activity of TM. Our findings provided valuable information for a better understanding of the anti-inflammatory activity of cyano enone-bearing triterpenoids and revealed TM as a promising anti-inflammatory candidate.