Twelve patients with primary hypercholesterolemia were treated for 12 weeks with probucol (500 mg b.i.d.). For each patient low density lipoproteins (LDL), isolated by ultracentrifugation were subfractionated by ion exchange high resolution chromatography in order to evaluate the content of a more electronegatively charged LDL (LDL-), a small subfraction that probably represent a circulating oxidatively modified lipoprotein. The treatment induced a 17% reduction of total LDL and 43% reduction of LDL-. By thin layer chromatography the probucol content in LDL- was a quarter of that in normally charged LDL. Under basal conditions, native LDL incubated for 24 h with 3 microM copper sulphate shows a net increase in electrophoretic mobility, an increase in relative fluorescence intensity and a reduction in vitamin E content, thus indicating peroxidative damage. After treatment with probucol, no significant changes of electrophoretic mobility, fluorescence and vitamin E content are detectable. LDL isolated from patients treated with probucol thus become resistant to oxidation by copper ions. The observed reduction of LDL- after treatment with probucol, confirms in vivo the antioxidant role of the drug and support the hypothesis that circulating LDL- may be linked to an oxidative process occurring in vivo.