Despite the existence of effective drugs used to treat inflammatory bowel disease (IBD), many patients fail to respond or lose response over time. Further, many drugs can carry serious adverse effects, including increased risk of infections and malignancies. Saffron (Crocus sativus) has been reported to have anti-inflammatory properties. Its protective role in IBD and how the microbiome and metabolome play a role has not been explored extensively. We aimed to establish whether saffron treatment modulates the host microbiome and metabolic profile in experimental colitis. Colitis was induced in C57BL/6 mice with 3% DSS and treated with either saffron in a dose of 20 mg/kg body weight or vehicle through daily gavage. On day 10, stool pellets from mice were collected and analyzed to assess saffron’s effect on fecal microbiota and metabolites through 16S rRNA sequencing and untargeted primary metabolite analysis. Saffron treatment maintained gut microbiota homeostasis by counter-selecting pro-inflammatory bacteria and maintained Firmicutes/Bacteroides ratio, which was otherwise disturbed by DSS treatment. Several metabolites (uric acid, cholesterol, 2 hydroxyglutaric acid, allantoic acid, 2 hydroxyhexanoic acid) were altered significantly with saffron treatment in DSS-treated mice, and this might play a role in mediating saffron’s colitis-mitigating effects. These data demonstrate saffron’s therapeutic potential, and its protective role is modulated by gut microbiota, potentially acting through changes in metabolites.