Incubation of calf thymus DNA with 3-morpholinosydnonimine (SIN-1), which simultaneously generates nitric oxide (NO) and superoxide (O2-), induced a significant increase of 8-hydroxydeoxyguanosine (8-OH-dG). Peroxynitrite also increased 8-OH-dG in calf thymus DNA. Addition of free hydroxyl radical (.OH) scavengers inhibited the increase of 8-OH-dG by SIN-1 or peroxynitrite. Incubation of 32P-labeled DNA fragment with SIN-1 or peroxynitrite caused DNA cleavage at every nucleotide with a little dominance at guanine residues. The results suggest that NO reacts with O2- to form peroxynitrite and the peroxynitrite induces oxidative DNA damage through an active intermediate of which reactivity is similar to .OH.