SYNTHESIS, CHARACTERIZATION, AND IN-VITRO QUANTITATION OF N-7-GUANINEADDUCTS OF DIEPOXYBUTANE

Diepoxybutane (DEB) is an important metabolite of 1,3-butadiene (ED), a high-volume industrial chemical classified as a probable human carci nogen. Rodent inhalation studies show strikingly high sensitivity of m ice to carcinogenic effects of butadiene compared to rats, which has b een linked to differences in metabolism Both species convert ED to 3,4 -epoxy-1-butene (EB), but mice further oxidize a significantly greater part of EB to DEB. DEB is a potent bifunctional genotoxic agent which is 100-fold more mutagenic than EB and is likely to be involved in ED -induced carcinogenesis. Identification of specific BD-induced DNA add ucts is critical to understanding the mechanism of its biological acti vity. We have previously described reactions of EB with guanine and ad enine as nucleobases, nucleosides, and constituents of DNA. In this wo rk, DEB-induced guanine adducts were isolated and structurally charact erized by UV spectroscopy, mass spectrometry, and nuclear magnetic res onance. When guanosine was reacted with DEB in glacial acetic acid fol lowed by hydrolysis:in hydrochloric acid, three products were isolated : N-7-(2',3',4'-trihydroxybut-1'-yl)guanine (DEB-Gua I, major adduct), N-7-(2',4'-dihydroxy-3'-chlorobut-1'-yl) (DEB-Gua II), and N-7-(2',3' -dihydroxy-4'acetoxybut-1'-yl)guanine (DEB-Gua III). The suggest initi al formation of the N-7-(2'-hydroxy-3',4'-epoxybut-1'-yl)guanine inter mediate followed by nucleophilic substitution at the 3',4'-epoxy ring with hydroxide, chloride, or acetate anions to give DEB-Gua I, II, or III, respectively. DEB-Gua I and the epoxy intermediate were also isol ated from hydrolysates of DEB-exposed calf thymus DNA (CT DNA). N-7-Gu anine adducts are known to undergo spontaneous and enzymatic depurinat ion producing apurinic sites. If not repaired before DNA replication, apurinic sites can give rise to mutations and ultimately cancer. The e xtent of alkylation at the N-7 of guanine in DEE-exposed DNA (58.7 +/- 1.1 adducts/10(3) normal guanines) was similar to that previously rep orted for CT DNA exposed to EB at the same molar ratio. Since EB and D EB appear to induce comparable levels of overall DNA alkylation at the conditions applied in this work, other factors; such as formation of DNA cross-links by DEB but not EB or differences in repair of EB and D EB adducts, may be responsible for the differences in mutagenicity.