PROJECT SUMMARY Accumulating evidence suggested that gut microbiota-derived metabolites of dietary flavonoids areimportant for their biological actions in the colon such as anti-inflammation. However, currently, there is onlya poor understanding of the formation and biofunctions of microbiota-derived flavonoid metabolites, whichgreatly limits our ability to develop dietary flavonoid-based strategies for inhibiting colonic inflammation.Consumption of citrus fruits and their components has been found to associate inversely with inflammation-related chronic diseases in humans. Polymethoxyflavones (PMFs), a unique class of citrus flavonoids,displayed potent anti-inflammatory properties in the colon in our animal studies. We found that gutmicrobiota mediated the production of an array of colonic metabolites of PMFs after their oral administrationin mice, and these metabolites possessed much stronger anti-inflammatory effects than their parentalPMFs. Importantly, our results showed that oral intake of PMFs by human volunteers resulted in theproduction of these bioactive metabolites in human stool. Furthermore, we identified multiple strains ofPMF-metabolizing bacteria from human stool and found that dietary PMFs modulated the abundance andmetabolic functions of these bacteria in mice with colitis. Overall, our results provided a strong basis for theapplication of citrus PMFs in the prevention of colonic inflammation and associated diseases. The objectiveof this project is to elucidate the mode of interaction between PMFs and gut microbiota, and its implicationin inhibiting colonic inflammation. Based on our preliminary results, we hypothesize that gut microbiotamediates the production of bioactive PMF metabolites, and these metabolites are critical for the anti-inflammatory actions of PMFs in the colon. To test our hypothesis, we will pursue the following 3 specificaims: 1) Identify novel microbiota-derived metabolites of PMFs in the colon and characterize their tissueprofiles in PMF-fed mice; 2) Determine the role of microbiota-derived PMF metabolites in inhibiting colonicinflammation; and 3) Characterize the interaction between PMFs and PMF-metabolizing fecal bacteria inboth healthy mice and mice with colitis. Our rationale is that the successful completion of this project willcontribute to the development of effective dietary strategies for amelioration of colonic inflammation andassociated diseases through the PMF/microbiota interaction.