The anthracycline Adriamycin is known to form adducts with DNA, but requires prior activation by formaldehyde. In contrast, the anthracycline barminomycin is also able to form adducts with DNA, but does not require activation by formaldehyde. Barminomycin, therefore, appears to function as a pre-activated form of Adriamycin. The DNA adducts formed by both anthracyclines are bound covalently to only one strand of DNA, but both also stabilise duplex DNA sufficiently that they can be detected as virtual interstrand crosslinks in heat denaturation electrophoretic crosslinking assays. The barminomycin-DNA adducts form extremely rapidly with DNA, and at exceedingly low concentrations (approximately 50-fold lower than with Adriamycin in the presence of excess formaldehyde), both characteristics consistent with barminomycin being in a pre-activated state, hence, undergoing a bimolecular reaction with DNA compared with the trimolecular reaction (drug, formaldehyde and DNA) required with Adriamycin. Surprisingly, barminomycin-DNA adducts are substantially more stable (essentially irreversible) than Adriamycin-DNA adducts (half life of approximately 25 h at 37 degrees C). Due to this understanding of the reactivity of barminomycin and its exceptional cytotoxicity (1000-fold more cytotoxic than Adriamycin), detailed structural studies of barminomycin-DNA adducts are now warranted, both in vitro and in tumour cells.