© 2020. Korean Association for the Study of Intestinal Diseases. Background/Aims: 5-Aminosalicylic acid (ASA) causes intolerance reactions in some patients. This study was performed to examine the prognosis of patients with ulcerative colitis (UC) and 5-ASA intolerance, and to evaluate the potential interaction between 5-ASA intolerance and the intestinal microbiota. Methods: We performed a retrospective cohort study of patients with UC who visited participating hospitals. The primary endpoint was to compare the incidence of hospitalization within 12 months between the 5-ASA intolerance group and the 5-ASA tolerance group. The secondary endpoint was to compare the risk of adverse clinical outcomes after the start of biologics between the 2 groups. We also assessed the correlation between 5-ASA intolerance and microbial change in an independently recruited cohort of patients with UC. Results: Of 793 patients, 59 (7.4%) were assigned to the 5-ASA intolerance group and 734 (92.5%) were assigned to the 5-ASA tolerance group. The admission rate and incidence of corticosteroid use were significantly higher in the intolerance than tolerance group (P<0.001). In 108 patients undergoing treatment with anti-tumor necrosis factor biologics, 5-ASA intolerance increased the incidence of additional induction therapy after starting biologics (P<0.001). The 5-ASA intolerance group had a greater abundance of bacteria in the genera Faecalibacterium, Streptococcus, and Clostridium than the 5-ASA tolerance group (P<0.05). Conclusions: In patients with UC, 5-ASA intolerance is associated with a risk of adverse clinical outcomes and dysbiosis. Bacterial therapeutic optimization of 5-ASA administration may be important for improving the prognosis of patients with UC.