Generation of hydroxyl radicals during the enzymatic reductions of the Fe 3+-ADP-Phosphate-adriamycin and Fe 3+-ADP-EDTA systems : Less involvement of hydroxyl radical and a great importance of proposed perferryl ion complexes in lipid peroxidation

Abstract

A system which contains NADPH, purified cytochrome P-450 reductase (enzyme) and Fe 3+-ADP-adriamycin complex in Tris-HCl buffer does not produce hydroxyl radical, but possesses a strong lipid peroxidation activity on exogenously added phospholipid micelles. Fe 3+-ADP-adriamycin complex, a tightly coordinated complex in Tris-HCl buffer, could be dissociated to Fe 3+-ADP-phosphate complex and adriamycin in phosphate buffer. Hydroxyl radical, which can be detected by a spin trapping method using N-tert-buryl-α-phenylnitrone, is produced during the enzymatic reduction of a mixture of Fe 3+-ADP-phosphate complex and adriamycin or of Fe 3+-ADP-EDTA complex while it is not involved in phospholipid peroxidation under the conditions used. With hydroxyl radical-generating systems, little or no quenching of hydroxyl radical in Tris-HCl buffer could be demonstrated. The oxidative cleavage of phospholipid is initiated by the proposed perferryl ion complex, which may be generated by the interaction of Fe 2+-ADP-adriamycin complex with O 2. A similar perferryl ion complex is also produced during the enzymatic reduction of Fe 3+-ADP-EDTA complex with a molar ratio of 2 for [Fe 3+)/[EDTA] in the presence of air. This is also able to catalyze lipid peroxidation.