Flavonoids are a large heterogeneous group of benzo-gamma-pyrone derivatives, which are abundantly present in our diet. In this study we investigated the effects of six flavonoids (apigenin, genistein, quercetin, rutin, naringenin and catechin) on the gastric tone in mouse isolated stomach. The mechanical activity was recorded as changes of intraluminal pressure. All flavonoids tested produced a concentration-dependent relaxation, which was reversible after washout. The relative order of potency of the flavonoids was apigenin ≥ genistein > quercetin > naringenin ≥ rutin > catechin. Analysis of the chemical structure showed that the relaxant activity was progressively diminished by the presence of hydroxyl group at C-3, saturation of the C-2, C-3 double bound, saturation of the C-2, C-3 double bound coupled with lack of the C-4 carbonyl and glycosylation. The flavonoid-induced relaxations were not modified in the presence of tetrodotoxin, a voltage-dependent Na+-channel blocker, Nω-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase, indomethacin, an inhibitor of cycloxygenase or tetraethylammonium, a non-selective blocker of potassium channels. In conclusion, this study provides the first experimental evidence for gastric relaxant activity of flavonoids. This action is influenced to a great extent by the structure of the molecules and it seems not to be dependent on neural action potentials, NO/prostaglandin production or activation of K+ channels.