Circadian disruption (CD) aggravates metabolic dysfunction-associated steatohepatitis (MASH), but supplementation with prebiotics inulin and oat β-glucan may mitigate its effects. However, their impact on colonic architecture and hepatic proteome remains unclear. We aimed to investigate the effects of prebiotics inulin and oat β-glucan on colonic architecture and hepatic proteome in mice with CD-aggravated MASH. CD was induced by weekly reversal of the light-dark cycle to simulate shift work. Male C57BL/6J mice were subjected to non-shifted chow, non-shifted fructose, palmitate, cholesterol, and trans-fat (FPC) diet, shifted chow, or shifted FPC diet (SFPC) for 26 weeks. Prebiotics inulin and oat β-glucan supplementation were provided to the SFPC group in the final 10 weeks. Distal colon and serum samples were collected for histological examination and endotoxemia evaluation, respectively. Liver samples were collected for proteomic mass spectrometry analysis. Mice with CD-aggravated MASH were found with colonic crypt loss and a distinct hepatic proteome structure compared to mice with non-CD MASH. Notably, inulin showed better effects than oat β-glucan in preserving colonic crypts in mice with CD-aggravated MASH. Furthermore, inulin supplementation restored the hepatic proteome structure similar to that of non-CD MASH mice, a benefit not observed with oat β-glucan. Given our prior findings showing oat β-glucan's superior ability to enrich gut bacterial species associated with MASH improvement under CD, this study highlights inulin's unique benefits for colonic architecture and hepatic proteome regulation in CD-aggravated MASH.