Vinyl halides are oxidized to 2-halooxiranes, which rapidly rearrange to 2-haloacetaldehydes. Both of these species can react with DNA to generate a variety of adducts, including the potentially mutagenic etheno (epsilon) products. Evidence was provided through kinetic studies that the epsilon-Gua adducts are formed primarily from 2-haloxiranes; consistent with this view, epoxide hydrolase inhibited the formation of N2,3-epsilon-Gua from vinyl chloride but alcohol dehydrogenase did not. Assignments of the NMR shifts of the etheno protons of 1,N2- and N2,3-epsilon-Gua were made with the use of 15N labeling and nuclear Overhauser effects, in revision of the literature. The H-5 proton of N2,3-epsilon-Gua showed facile exchange in acid or base; the H-7 proton of 1,N2-epsilon-Gua was exchanged at neutral or basic pH but not in acid. Reaction of Br2CHCH2OH (labeled at C1 with 2H or 13C) with Guo yielded 1,N2-epsilon-Gua and N2,3-epsilon-Gua, presumably through the intermediacy of 2-bromooxirane. 1H NMR analysis indicated that the labeled carbon was attached to the original Guo N2 atom in both cases. When N2-(2-oxoethyl)Gua was generated from a diethyl acetal or from a glycol, the major product was the cyclic derivative 5,6,7,9-tetrahydro-7-hydroxy-9-oxoimidazo[1,2-alpha]purine. This compound was also formed in considerable yield from the reaction of 2-chlorooxirane with Guo, dGuo 5'-phosphate, or DNA and is relatively stable in the presence of acid or mild base. It does not appear to be readily dehydrated to yield the etheno adducts but may be of significance as a DNA adducts in its own right.

The major DNA adduct derived from 1,2-dibromoethane is known to be S-[2-(N7-guanyl)-ethyl]glutathione; minor nucleic acid DNA adducts were characterized in view of the possibility that some might be unusually persistent or biologically active. RNA was modified in vitro by treatment with 1,2-dibromoethane and glutathione in the presence of rat liver cytosol, and bases were released by mild acid hydrolysis, which liberated greater than 99% of the bound radioactivity. One of the minor adducts was identified as S-[2-(N1-adenyl)ethyl]glutathione on the basis of its UV, mass, and NMR spectra. This adduct could be synthesized by reaction of S-(2-chloroethyl)-glutathione with adenosine. The material was desulfurized by treatment with Raney Ni to give N1-ethyladenine in low yield. The Raney Ni reaction was accompanied by considerable formation of the corresponding N6-adenine derivative via Dimroth rearrangement. Another adduct was identified as S-[2-(N7-guanyl)ethyl]cysteinylglycine by its UV, mass, and NMR spectra, but the material was demonstrated to be formed from the major DNA adduct, S-[2-(N7-guanyl)-ethyl]glutathione under conditions of mild acid hydrolysis. The imidazole ring opened derivative of S-[2-(N7-guanyl)ethyl]glutathione was synthesized and found not to be formed in DNA in vitro or in vivo. The two remaining minor adducts account for 1-2% of the total binding, but insufficient quantities were recovered to allow for structure determination; however, neither of these (uncharacterized) minor products are seen after the reaction of S-(2-chloroethyl)glutathione with guanosine or adenosine. S-[2-(N1-Adenyl)ethyl]glutathione was formed in rat liver RNA and DNA.(ABSTRACT TRUNCATED AT 250 WORDS)

The continuous withdrawal method proved to be a viable alternative to the classic intermittent sampling technique. The method should prove useful in drug discovery screening, where the evaluation of large numbers of compounds for systemic clearance or oral bioavailability is often necessary.

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