| 1,3-Butadiene (BD) is colorless gas and an important industrial chemical, whichis mainly used in the manufacture of rubber and plastics. BD is an air pollutant thatis mainly produced through incomplete combustion of fossil fuels.The carcinogenicity of BD stems from the in vivo metabolites. BD has twometabolic pathways. One leads to the formation of epoxides, inchluding3,4-epoxy-1-butene,3,4-epoxy-1,2-butanediol, and1,2,3,4-diepoxybutane. The otherpathway gives1-chloro-2-hydroxy-3-butene (CHB) as the initial product in thepresence of myeloperoxidased (MPO), H2O2, and high concentrations of Cl-. CHBcan be further metabolized to1-chloro-3-butene-2-one (CBO) by alcoholdehydrogenas.CHB and CBO have cytotoxicity and genotoxicity. CBO is100-fold more toxicthan CHB. CBO can directly react with DNA and form DNA adducts. Studies havefound that CBO can react with2’-deoxycytidine or2’-deoxyadenosine under in vitrophysiological conditions (pH7.4,37℃) and generate a variety of products. In thepresent study, we investigated the reaction of CBO with2’-deoxyguanosine (dG)under in vitro physiological conditions. The results showed that six products wereformed in this reaction, which were designated as dG-1, dG-2, dG-3, dG-4, dG-5and dG-6according to their retention times. dG-5and dG-6were identified as a pairof diastereomers. They were transformed to dG-5/6b in0.2M HCl at75℃.dG-5/6b was converted to dG-5/6a under in vitro physiological conditions. dG-1was hydrolyzed to dG-1a in0.2M HCl at37℃). dG-5/6a and dG-1a weredetermined to actually be the same compound. dG-3and dG-5/6b were structurallycharacterized through NMR.Isoprene is an analogue of BD, which is a possible carcinogen. It is apetrochemical and natural product with the major source being emission from plants.It is biotransformed to two monoepoxides by cytochrome P450, 2-ethenyl-2-methyloxirane (IP-1,2-O) and2-(1-methylethenyl) oxirane (IP-3,4-O),which can further be metabolized to the diepoxide (2-methyl-2,2-bioxirane, MBO).IP-1,2-O and IP-3,4-O are non-mutagenic, whereas MBO exhibits mutagenicpotential. IP-1,2-O is genotoxic; however, the genotoxicity of IP-3,4-O and MBO orthe crosslinking potential of MBO have not been investigated. In the present study,we used the comet assay to investigate the genotoxicity profiles of the threemetabolites and the cross-linking capability of MBO in human hepatocyte L02cells.All three metabolites showed positively concentration-dependent profiles withthe rank order of potency being IP-3,4-O> MBO> IP-1,2-O when the exposure timewas1h. Interestingly, their time-dependent genotoxicity profiles (from0.5to4h)were different from each other: the DNA migrations caused by IP-1,2-O (1000M),IP-3,4-O (200and500M), and MBO (200M) exhibited negative, neutral, andpositive profiles, respectively. DNA migration caused by1000M MBO increasedover time during0-2h but decreased with time during2-4h. Further experimentindicated that the negative profile of IP-1,2-O was caused by its rapid hydrolysis, thenegative profile of1000M MBO during2-4h was attributed to DNA cross-linking,which was confirmed by another experiment. MBO could induce DNA cross-linkingonly under the conditions of high concentrations and long incubation time (500μM,6h or1000μM,4h). It was important to note that the genotoxicity of IP-3,4-O wasthe strongest in the three metabolites, suggesting that IP-3,4-O may play animportant role in the toxicity of isoprene.dG-5/6b may be used as a potential biomarker for exposure of BD. Studies ongenotoxicity of three isoprene metabolites may give us a better understanding of themechanism of isoprene toxicity. |
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