Constipation and the use of laxatives are relatively common in patients with diabetes mellitus. However, the mechanisms responsible for the constipation are unclear. Even though autonomic neuropathy is regarded as one of the important mechanisms of constipation, it requires further clarification. In addition, the colonic function in diabetic patients requires further investigation. The aim of this study was to compare the colonic transit time between patients with diabetes mellitus and healthy subjects, and correlate it to the presence of cardiovascular autonomic neuropathy. The colonic transit time was measured by a noninvasive, radio-opaque marker method, and the presence of cardiovascular autonomic neuropathy was evaluated by the beat-to-beat variation and the orthostatic hypotension. Constipation was defined by the Rome II criteria. The mean total colonic transit time of the 28 diabetic patients (34.9 ± 29.6 h, mean ± S.D.) was significantly longer than that of the 28 healthy subjects (20.4 ± 15.6 h, p < 0.05). Among the diabetic patients, 9/28 (32%) had constipation and 14/28 (50%) had cardiovascular autonomic neuropathy. The diabetic patients with constipation showed longer total, left and recto-sigmoid colonic transit times than those without constipation. However, the mean colonic transit time of diabetic patients with and those without cardiovascular autonomic neuropathy was similar. In conclusion, other mechanisms than the mere presence of cardiovascular autonomic neuropathy might be more relevant to the development of constipation in patients with diabetes mellitus.