Protopine is a commonly available alkaloid widely distributed in several plant families. Previous studies reported its antiallergic, intestinal inhibitory, antispasmodic, antiarrhythmic, antibacterial, and GABA receptor binding-enhancing activities. This study investigated the anticholinergic and antihistaminic properties of protopine and some derivatives on isolated guinea-pig ileum. Cumulative dose-response curves for acetylcholine were displaced rightward by protopine in a parallel manner (competitive antagonism), with a pA₂ value of 5.87±0.21. Atropine, with a pA₂ of 8.69±0.23, was about 660 times more potent. Longer preincubation times (10 and 20 min) slightly increased protopine's pA₂, and its activity was easily washed out. The styrene derivative of protopine had slight contractile activity and noncompetitive effects on acetylcholine (pD'₂=4.18±0.34), 10 times less potent than papaverine. Protopine showed no true antihistaminic properties against histamine. Other derivatives (protopine N-oxide and dibenzoxazacyclo-undecine) lacked competitive antagonism toward acetylcholine and caused mild contractions (<20% of max). In conclusion, protopine is a weak anticholinergic alkaloid, and its presence in plants may explain some medicinal plants' spasmolytic effects.