Both adriamycin and daunomycin chelate Fe 3+ , forming complexes with similar spectroscopic properties but very different redox chemistry. Anaerobic Fe 3+ -adriamycin and Fe 3+ -daunomycin complexes of identical stoichiometry give rise to identical optical absorbance and EPR spectra with maxima at 600 nm and signals at g = 4.2 and g = 2.01. In anaerobic preparations, however, the 600 nm absorption band and the EPR signals of the Fe 3+ -adriamycin complexes decrease as Fe 3+ is reduced to Fe 2+ , with the appearance of an adriamycin free-radical signal at g = 2.0035, while the spectra of Fe 3+ -daunomycin complexes remain unchanged, with no free-radical signal appearing. Polarographic measurements demonstrate that the Fe 3+ -adriamycin complexes consume O 2 while the Fe 3+ -daunomycin complexes do not. Measurements in the presence of catalase and superoxide dismutase suggest that 75% of the O 2 consumed by Fe 3+ -adriamycin is reduced to H 2 O 2 or .O 2 − . Spin-trapping experiments demonstrate that the Fe 3+ -adriamycin complexes generate .OH, while the daunomycin complexes do not. Qunatitation of .OH generation by Fe 3+ -adriamycin demonstrates that the initital rate of .OH generation approaches the rate of total O 2 consumption. DNA cleavage studies show that only the Fe 3+ -adriamycin complexes cleave DNA in the absence of exogenous added H 2 O 2 . This DNA cleavage can be blocked by catalasa or .OH radical scavengers. These results indicate that the side-chain hydroxyl group is essential for Fe 3+ reduction, subsequent O 2 reduction, as well as for the generation of the drug free radical, since adriamycin and daunomycin are structurally identical except for this hydroxyl. The difference in redox chemistry of the iron complexes of adriamycin and daunomycin is the only known mechanism which can explain the difference in anti-tumor potency and cardiotoxicity of these two structurally similar drugs.