The natural alkaloid berberine is being studied as a drug candidate for the treatment of ulcerative colitis (UC). Fingolimod is an immunomodulator approved for the treatment of multiple sclerosis. Whether fingolimod use can be extended to UC and how it interacts with berberine remain unclear. In the present study, the anti-inflammatory efficacies of berberine, fingolimod, and a combination of half-doses of them was examined in mice with dextran sulfate sodium-induced colitis. In mice with subchronic colitis, 14-day oral administration of fingolimod had greater efficacy than berberine in ameliorating the disease clinical severity and colon shortening. However, in mice with chronic colitis, 30-day oral administration of berberine was more effective than fingolimod except on splenic swelling. Notably, the combination of half-doses of each drug was equally effective as the superior single drugs for two models and resulted in reduced splenic swelling in the chronic colitis model. The inhibition of cytokine expression and STAT3 activation, as well as binding to the sphingosine 1-phosphate receptor by both drugs, contributed to the combination efficacy. Our findings suggest that fingolimod in combination with berberine at reduced doses represents a novel therapy for UC that attains satisfactory efficacy with reduced potentials for adverse effects.