title: Trapping of a cross-link formed by a major purine adduct of a metabolite of the carcinogen N-nitrosomorpholine by inorganic and biological reductants.
abstract: 3-Hydroperoxy-N-nitrosomorpholine in buffered aqueous media in the presence of calf thymus DNA was treated with a phosphine reductant to generate the transient α-hydroxynitrosamine and subsequent diazonium ion that alkylated the DNA, as previously reported. Subsequent addition of hydride donors, for 30 min, followed by acid hydrolysis of the mixture allowed detection and quantification of 6-(2-{2-[(9H-purin-6-yl)amino]ethoxy}ethoxy)-9H-purin-2-amine, the reduced cross-link formed from deposition, via the diazonium ion, of a 3-oxapentanal fragment on O(6)-Gua, and condensation with N(6)-Ade, presumably in the vicinity. Decreasing the temperature of the reaction mixtures and decreasing the pH modestly increased the yields of the trapped cross-link. Among three borohydride reductants, NaNCBH3 is superior, being ∼4 times more effective on a molar basis, as opposed to a hydride equivalent basis, than NaBH4 or Na(AcO)3BH. For trapping with NaNCBH3, it is deduced that the reaction likely occurs with the iminium ion that is in protonic equilibrium with its conjugate base imine. In an experiment in which the hydroperoxide was decomposed and NaNCBH3 was introduced after various periods of time, the amount of cross-link was observed to increase, nearly linearly, by ∼4-fold over 1 week. These data indicate that there are a minimum of two populations of cross-links, one that forms rapidly, in minutes, and another that grows in with time, over days. Reduced nicotinamide cofactors and ascorbate are observed to effect reduction (over 3 days) of the cross-links, confirming the possibility that otherwise reversible cross-links might be immortalized under biological conditions.
authors.affiliation: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County , Baltimore, Maryland 21250, United States.
authors.forename: Niangoran
authors.initials: N
authors.lastname: Koissi
authors.forename: James C
authors.initials: JC
authors.lastname: Fishbein
mesh_terms: D000818:Animals; D002273:Carcinogens / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D003432:Cross-Linking Reagents / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D018736:DNA Adducts / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D015394:Molecular Structure; D009602:Nitrosamines / Q000737:chemistry / Q000378:metabolism*; D011687:Purines / Q000737:chemistry / Q000378:metabolism*; D019163:Reducing Agents / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature
chemical_list: D002273:Carcinogens; D003432:Cross-Linking Reagents; D018736:DNA Adducts; D009602:Nitrosamines; D011687:Purines; D019163:Reducing Agents; C002741:N-nitrosomorpholine; D004247:DNA; C427203:calf thymus DNA; C030985:purine