Background: Alosetron is a potent and selective 5-HT3 receptor antagonist, which has been shown to be beneficial in the treatment of female patients with non-constipated irritable bowel syndrome. Aims: To investigate the effect of alosetron on whole gut, small bowel and colonic transit in patients with irritable bowel syndrome (Study 1) and healthy volunteers (Study 2). Subjects: Thirteen patients with irritable bowel syndrome and 12 healthy volunteers. Methods: Both studies were randomized, double-blind, placebo-controlled with a two-way crossover design, in which each subject received alosetron (2 mg b.d. administered orally) or placebo for 8 days. Mean whole gut transit was determined from the excretion of radio-opaque markers; small bowel transit was determined from rise in breath hydrogen after a meal; and colonic transit and segmental transit were evaluated from abdominal X-ray. In addition, colonic transit was calculated by subtracting small bowel transit time from whole gut transit time. Results: Alosetron increased colonic transit time by prolonging left colonic transit in both patients with irritable bowel syndrome and controls. This resulted in a tendency for the whole gut transit to be delayed in irritable bowel syndrome patients (P = 0.128), which was confirmed in controls (r = 0.047). Conclusion: Alosetron delays colonic transit by prolonging left colonic transit. These results add to the body of evidence suggesting that alosetron should have a therapeutic role in patients with non-constipated irritable bowel syndrome.