The antioxidant activities of representative calcium antagonists, including amlodipine, verapamil, and diltiazem, were measured in hepatic microsomal membranes by the Fe-catalyzed, hydroxyl radical-producing system (dihydroxyfumarate + Fe3+) and assessed by malondialdehyde (MDA) formation. Despite the absence of L-type calcium channels in this membrane preparation, the calcium antagonists showed dose-dependent antioxidant activity. The biophysical mechanism for calcium-antagonist antioxidant activity was evaluated using radioligand binding assays, high-resolution differential scanning calorimetry, and small-angle x-ray diffraction approaches. These analyses demonstrated that calcium-antagonist antioxidant potency correlated directly with the compounds' relative affinity for the membrane lipid bilayer and ability to modulate membrane thermodynamic properties (amlodipine >> verapamil > diltiazem). The charged 1,4-dihydropyridine calcium antagonist, amlodipine, had the highest affinity for the membrane lipid bilayer (Kp>10(4)) and produced the largest changes in membrane thermodynamic properties, including a reduction in thermal phase transition temperature (-11%), enthalpy (-14%), and cooperative unit size (-59%), relative to control phosphatidylcholine liposomes. Electron density profiles generated from x-ray diffraction data demonstrated that amlodipine effected a broad and dose-dependent increase in molecular volume associated with the membrane hydrocarbon core. These data indicate that lipophilic calcium antagonists inhibit lipid peroxidation in cellular membranes as a result of modulating physicochemical properties of the membrane lipid bilayer, independently of calcium channel inhibition. Amlodipine had the most potent antioxidant activity as a result of distinct biophysical interactions with the membrane lipid bilayer. The nonreceptor-mediated antioxidant activity of calcium antagonists may contribute to cytoprotective mechanisms of action in cardiovascular diseases.