The role played by oxidative stress in amiodarone-induced mitochondrial toxicity is debated. Dronedarone shows pharmacological properties similar to those of amiodarone but several differences in terms of toxicity. In this study, we analyzed the effects of the two drugs on liver mitochondrial function by administering an equivalent human dose to a rat model. Amiodarone increased mitochondrial H2O2 synthesis, which in turn induced cardiolipin peroxidation. Moreover, amiodarone inhibited Complex I activity and uncoupled oxidative phosphorylation, leading to a reduction in the hepatic ATP content. We also observed a modification of membrane phospholipid composition after amiodarone administration. N-acetylcysteine completely prevented such effects. Although dronedarone shares with amiodarone the capacity to induce uncoupling of oxidative phosphorylation, it did not show any of the oxidative effects and did not impair mitochondrial bioenergetics. Our data provide important insights into the mechanism of mitochondrial toxicity induced by amiodarone. These results may greatly influence the clinical application and toxicity management of these two antiarrhythmic drugs. ► Amiodarone is the most administered antiarrhytmic agent but shows liver toxicity. ► Dronedarone has been developed to avoid toxic effects of amiodarone. ► We studied the effects of both drugs on liver mitochondrial function. ► Amiodarone induces mitochondrial oxidative stress prevented by N-acetylcysteine. ► Dronedarone acts as an uncoupler but does not induce mitochondrial toxicity.