Various vanilloid-type beta-adrenoceptor blockers were studied on guinea pig right atrium and trachea and rat colon. In addition, we also investigated their beta(1)-, beta(2)-, and beta(3)-adrenoceptor binding affinities. All these beta-adrenergic antagonists inhibited (-)isoproterenol-induced positive chronotropic effects of the right atrium and tracheal relaxation responses in a concentration-dependent manner. Some of these agents prevented the inhibition of rat colon spontaneous motility by (-)isoproterenol. Of the agents tested, we found that ferulidilol, eugenodilol, eugenolol, isoeugenolol, and ferulinolol, as well as propranolol and metoprolol, possessed beta(3)-adrenoceptor blocking activities, others were nearly without effectiveness. Furthermore, the binding characteristics of vanilloid-type beta-adrenergic antagonists were evaluated in [3H]CGP-12177, a beta(1)/beta(2)-adrenoceptor blocker and a beta(3)-adrenoceptor agonist, binding to beta(1)-, beta(2)-, and beta(3)-adrenoceptor sites in rat ventricle, lung, and interscapular brown adipose tissue (IBAT) membranes, respectively. Eugenodilol, eugenolol, metoprolol, isoeugenolol, and ferulinolol were less potent than both propranolol and ferulidilol in competing for the beta(3)-adrenoceptor binding sites. From the results of in vitro functional and binding studies, we suggested that propranolol, ferulidilol, eugenodilol, eugenolol, metoprolol, isoeugenolol, and ferulinolol all possessed beta(3)-adrenoceptor blocking activities. On the other hand, we also found that eugenodilol, eugenolol, metoprolol, isoeugenolol, and ferulinolol had a low lipid solubility in comparison with propranolol and ferulidilol. In conclusion, we proposed that beta(3)-adrenoceptor antagonistic actions of these vanilloid-type beta-blockers were positively correlated with their lipid solubility.