Clinical evidence suggests that the herbal formulation ColiMil (which contains Matricaria recutita flowers extract, Foeniculum vulgare fruit extract and Melissa officinalis aerial parts extract) is effective in the treatment of breastfed colic in infants. Therefore the effect of this phytotherapeutic formulation and its herbal constituents on upper gastrointestinal transit was investigated in mice in vivo. Oral administration of the herbal formulation (0.4-0.8 mL/mice) dose-dependently delayed upper gastrointestinal transit. Among the herbal components, Matricaria recutita extract (0.89 and 1.78 mg/mouse) and Melissa officinalis extract (6.46 and 12.92 mg/mouse), but not Foeniculum vulgare (8.21 and 16.42 mg/mouse), reduced motility significantly. These results suggest that ColiMil reduces upper gastrointestinal motility in mice, with a major contribution by Matricaria recutita and Melissa officinalis. These experimental data may be important to better understand the observation that the herbal formulation ColiMil improves colic in breastfed infants.