Ulcerative colitis is a chronic and recurrent inflammatory bowel disease mediated by immune response. Geniposide is the main active ingredient extracted from Gardenia jasminoides, which has been suggested to exert excellent efficacy on inflammatory disease. Herein, in this study, we aimed to uncover the systematic understanding of the mechanism and effects of geniposide in ameliorating inflammatory responses in colitis. In brief, the TCMSP server and GEO DataSets were used to analyze the systematic understanding of the mechanism and effects of geniposide in ameliorating inflammatory responses in colitis. Dextran Sulfate Sodium (DSS)-induced acute colitis of mice were administered with 25-100[Formula: see text]mg/kg of geniposide for 7 days by gavage. Lipopolysaccharide (LPS)-induced Bone Marrow Derived Macrophage (BMDM) cell or RAW264.7 cell models were treated with 20, 50 and 100[Formula: see text][Formula: see text]M of geniposide for 4[Formula: see text]h. Myeloperoxidase (MPO) activity and Interleukin-1[Formula: see text] (IL-1[Formula: see text] levels were measured using MPO activity kits and IL-1[Formula: see text] levels enzyme-linked immunosorbent assay (ELISA) kits, respectively. Additionally, Western blot was used to determine the relevant protein expression. As a result, Geniposide could ameliorate inflammatory responses and prevent colitis in DSS-induced acute colitis of mice by activating AMP-activated protein kinase (AMPK)/Transcription 1 (Sirt1) dependent signaling via the suppression of nod-like receptor protein 3 (NLRP3) inflammasome. Geniposide attenuated macrophage differentiation in DSS-induced acute colitis of mice. Geniposide suppressed NLRP3 inflammasome and induced AMPK/Sirt1 signaling in LPS-induced BMDM cell or RAW264.7 cell models. In mechanism studies, the inhibition of AMPK/Sirt1 attenuated the anti-inflammatory effects of geniposide in colitis. The activation of NLRP3 attenuated the anti-inflammatory effects of geniposide in colitis. Taken together, our results demonstrated that geniposide ameliorated inflammatory responses in colitis vai the suppression of NLRP3 inflammasome in macrophages by AMPK/Sirt1-dependent signaling.