Doxorubicin (DOX) has a wide spectrum of antitumor activity with dose-related cardiotoxicity as a major side effect. This cardiotoxicity has been suggested to result from the generation of oxygen-free radicals. The objective of the present study was to investigate the influence of the antioxidant, thymoquinone (TQ) on cardiotoxicity and antitumor activity of DOX in mice. TQ (8 mg/kg/day, p.o.) administered with drinking water starting 5 days before a single i.p. injection of DOX (20 mg/kg) and continuing during the experimental period ameliorated the DOX-induced cardiotoxicity in mice. This finding was evidenced by significant reductions in serum lactate dehydrogenase and creatine kinase elevated levels and further supplemented by histopathological examination of cardiac tissue. TQ did not alter the plasma and heart DOX levels as monitored by fluorometric analysis. In in vivo study on mouse Ehrlich ascites carcinoma tumor, it could then be shown that TQ does not interfere with the antitumor activity of DOX. The current data support TQ as a potentially selective cytoprotective agent, which may ameliorate cardiotoxicity without decreasing DOX antitumor activity.