Antioxidant activity of amiodarone on human lipoprotein oxidation

Abstract

Lipoprotein oxidation is crucial in atherogenic processes. Amiodarone is a lipophilic antiarrhythmic/antianginal drug which is able to influence the physicochemical status of biological lipid components. Since oxidation of lipids is affected by their physicochemical state and amiodarone binds to lipoproteins, we hypothesized that the drug may exert an antioxidant activity on human lipoprotein oxidation. Dose-dependent effects of therapeutically achievable amiodarone concentrations (1.5, 3, 5, 7 and 10 microM) were studied on copper-catalysed oxidation of the non-HDL fraction in vitro. Amiodarone inhibited oxidation as judged by generation of thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LOOH) and fluorescent products of lipoperoxidation (FPL) as well as from the kinetics of conjugated diene formation. This antioxidant activity was significant at 1.5 microM with total inhibition at 10 microM and an IC(50) of 4 microM. The primary in vivo metabolite of amiodarone, namely desethylamiodarone, also exhibited specific antioxidant properties although it was less effective than amiodarone with an IC(50) of 7 microM. In further in vivo experiments, susceptibility to copper-mediated oxidation of the non-HDL fraction was investigated before and 4 weeks after oral amiodarone administration to humans. Following treatment, significant inhibition of TBARS, LOOH and FPL generation was observed in comparison with baseline levels and a placebo-treated control group, highlighting an effective antioxidant capacity of amiodarone in vivo. Amiodarone did not change lipoprotein vitamin E and phospholipid content in vivo and did not show scavenging effects on oxidizing species involved in lipoprotein oxidation, such as peroxyl radicals, nor metal-binding/inactivating properties, suggesting that physicochemical modifications of lipoprotein lipids induced by the lipophilic drug may be involved in its antioxidant activity. In conclusion, amiodarone, and its primary metabolite desethylamiodarone, show previously unrecognized antioxidant activity on human lipoprotein oxidation. This effect is also evident in vivo and at therapeutically achievable drug concentrations. Thus, amiodarone may act as an antioxidant/antiatherosclerotic agent in humans, although this issue warrants further clinical study.

Figure 1

Concentration-dependent inhibition of copper-mediated lipoprotein oxidation by amiodarone. The non-HDL fraction was oxidized by 5 μM CuCl₂ for 3 h at 37°C, in PBS (pH 7.4), with and without 1.5, 3, 5, 7 and 10 μM amiodarone. LOOH, FPL and TBARS were, respectively, 297.5±48 nmol LOOH mg⁻¹ non-HDL protein, 92.5±12 units of relative fluorescence mg⁻¹ non-HDL protein and 58.5±8 nmol TBARS mg⁻¹ non-HDL protein in control experiments. The results represent the means of percentage inhibition of lipoprotein oxidation calculated from seven independent experiments (the s.d. is less than 10% and was omitted for clarity). ^*P<0.05 vs control; †P<0.05 vs the lower drug concentration (ANOVA plus Student–Newman–Keuls test).

Figure 2

Antioxidant activity of amiodarone on the kinetics of copper-mediated oxidation of the non-HDL fraction evaluated by continuous spectrophotometric monitoring of absorbance increase at 234 nm due to conjugated diene formation. Trace 1: control; traces 2, 3, 4 and 5: 1.5, 3, 5 and 7 μM amiodarone, respectively. The results shown are representative of seven similar experiments. See Methods and Results sections for further explanations.

Figure 3

Spectral characteristics of 10 μM amiodarone in the absence (trace A) and presence (trace B) of 5 μM CuCl₂ in PBS (pH 7.4). See Methods and Results sections for further explanations.