Modification of the base and the sugar moieties of DNA with 4-(hydroxymethyl)benzenediazonium salt (HMBD), a carcinogen in the mushroom Agaricus bisporus, was investigated. When deoxyribonucleosides dGuo, dAdo, dThd, and dCyd were incubated with HMBD at pH 7.4 and 37 degrees C, the levels of all the nucleosides were decreased. The decrease was inhibited by ethanol or Cys. When deoxyribose was incubated with HMBD, malonaldehyde was released as assessed by the thiobarbituric acid reactivity. The release was inhibited by ethanol. Major products of the reaction of dGuo and dAdo with HMBD were isolated, and their structures were established to be 8-[4-(hydroxymethyl)phenyl]dGuo (8-HMP-dGuo) and 8-[4-(hydroxymethyl)phenyl]dAdo), respectively. Calf thymus DNA treated with HMBD was enzymatically digested into nucleosides, in which 8-HMP-dGuo and 8-HMP-dAdo were detected. Formation of the modified nucleosides in DNA was inhibited by ethanol or 2-mercaptoethanol. Malonaldehyde was released from DNA treated with HMBD, which indicated that the deoxyribose moiety of DNA had been damaged. The results indicate that the 4-(hydroxymethyl)phenyl radical generated from HMBD can directly modify the base and the sugar moieties of DNA under the mild conditions. Inhibitory effect of ethanol was ascribable to its scavenging activity for the carbon-centered radical. The inhibitory effect of Cys and 2-mercaptoethanol was found to be due to the formation of the reversible adducts between HMBD and the SH compounds.