The peroxidase-catalyzed decomposition of 3-hydroperoxy-1-butene (1), 2,3-dimethyl-3-hydroperoxy-1-butene (2), tert-butyl hydroperoxide (3), ethyl oleate hydroperoxide 4, and linoleic acid hydroperoxide 5 was applied as a chemical model system to assess whether lipid hydroperoxides may cause DNA damage under peroxidase catalysis. For this purpose, the Coprinus peroxidase (CIP), horseradish peroxidase (HRP), and the physiologically important lactoperoxidase (LP) were tested. Indeed, hydroperoxides 1-5 induce strand breaks in pBR 322 DNA upon peroxidase catalysis. For the nucleoside dG, the enzymatic decomposition of hydroperoxides 1-4 led to significant amounts of 4, 8-dihydro-4-hydroxy-8-oxo-2'-deoxyguanosine (4-HO-8-oxo-dG) and guanidine-releasing products (GRP), whereas 7, 8-dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dG) was not obtained. In isolated calf thymus DNA, the efficient conversion of the guanine base (Gua) was observed. Peroxyl radicals, which are generated in situ from the hydroperoxides by one-electron oxidation with the peroxidases, are proposed as the active oxidants on the basis of the following experimental facts. (i) Radical scavengers strongly inhibit the guanine oxidation in dG and DNA and strand-break formation in the latter. (ii) EPR spectral studies with 5, 5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin trap confirmed the formation of peroxyl radicals. (iii) The release of molecular oxygen was demonstrated, produced through the disproportionation of peroxyl radicals. The biological relevance of these findings should be seen in the potential role of the combined action of lipid hydroperoxides and peroxidases in damaging cellular DNA through peroxyl radicals.