Abstract Ethanolamine(EA) often occurs at a relatively high concentration within the inflamed gut of IBD patients. To investigate the role of EA in colonic inflammation and host-microbiome dysbiosis, thirty-six ICR mice were treated with 3% DSS for a week to generate acute intestinal inflammation and then supplied with 0μM, 500μM (LowEA), and 3000 μM (HighEA) in drinking water for two weeks, after that,16s RNA sequencing was applied in characterizing the changes in colonic microbiota driven by different EA levels. An inflamed colonic organoid model via 3% DSS treatment was also established for further verification of these in vivo findings.EA significantly reduced proximal colonic crypt depth but increased distal colonic villus height in HighEA group. The protein and mRNA expression of occludin and Reg3β, BD1, BD2, and MUC2were significantly up-regulated in EA treated groups. EA decreased mucosal inflammation-related cytokines levels (IL1, IL6, IL17, TNFα, and INFγ) and increased the significantly increased concentration of sIgA. Serum aspartate aminotransferase and alanine aminotransferase were significantly down-regulated in the highEA group. EA increased the relative abundance of Blautia, Roseburia, Lactobacillus, Faecalibaculum, Candidatus_Saccharimonas, Alloprevotella , and Lachnoclostridum .and thus microbial metabolic pathways including Oxidative phosphorylation, Lipopolysaccharide biosynthesis, Arginine and proline metabolism, Folate biosynthesis , and Biotin metabolism were more abundant in LowEA group than those in control. EA up-regulated the protein or mRNA expression of TLR4/MyD88 in colonic tissues and the DSS-treated colonic organoid model. This study firstly demonstrated that ethanolamine in altering host-microbiome dysbiosis, which may provide new insights into the role of dietary lipids in IBD. Importance Inflammatory bowel disease (IBD) affects ~3.1 million people in the USA and is increasing in incidence worldwide. IBD pathogenesis has been associated with gut microbiome dysbiosis characterized as a decrease in gut microbial diversity. Extensive works have demonstrated the roles of dietary fiber, short-chain fatty acids, and aromatic amino acids in altering the composition of gut microbiota to restore immune homeostasis and alleviate inflammation via diverse mechanisms in IBD. However, little is known about essential sphingolipids like ethanolamine (EA), an essential compound in the CDP-ethanolamine pathway for phosphatidylethanolamine (PE) in both intestinal cells and bacteria. PE synthesis deficiency can ultimately result in a loss of membrane integrity and metabolic disorders in IBD. Our results demonstrate that ethanolamine could improve colonic barrier functions and inflammatory immunoreactions via shifting microbiome dysbiosis, which provides new insights into the role of dietary lipids in IBD.