Analysis of adducts in hepatic DNA of rats treated with N-nitrosopyrrolidine.

N-Nitrosopyrrolidine (NPYR) is a hepatocarcinogen in rats. It is metabolically activated by cytochrome P450 enzymes in the liver leading to the formation of 4-oxobutanediazohydroxide (4) and related intermediates that react with DNA to form adducts. Because DNA adducts are thought to be critical in carcinogenesis by NPYR, we analyzed hepatic DNA of NPYR-treated rats for several adducts: N2-(tetrahydrofuran-1-yl)dGuo (N2-THF-dGuo, 13), N6-THF-dAdo (14), N4-THF-dCyd (17), and dThd adducts 15 and 16. The rats were treated with NPYR in the drinking water, 600 ppm for 1 week, or 200 ppm for 4 or 13 weeks. Hepatic DNA was isolated, enzymatically hydrolyzed, and analyzed by capillary LC-ESI-MS-SIM, which indicated the presence of adducts 13, 14, and 17. Because these adducts can be unstable at the deoxyribonucleoside level, further analyses were carried out using DNA treated with NaBH3CN, which converts adducts 13-17 to N2-(4-hydroxybut-1-yl)dGuo [N2-(4-HOB)dGuo, 18], N6-(4-HOB)dAdo (19), O2-(4-HOB)dThd (20), O4-(4-HOB)dThd (21), and N4-(4-HOB)dCyd (22). [15N]-Labeled analogues of adducts 18-20 and 22 were synthesized and used in this analysis, which was performed by capillary LC-ESI-MS/MS-SRM. Convincing evidence for the presence of adducts 18-22 was obtained. Levels of 18, 19, 20, and 21 were (mumol/mol dGuo): 3.41-5.39, 0.02-0.04, 2.56-3.87, and 2.28-5.05, respectively. Compound 22 was not quantified due to interfering peaks. These results provide the first evidence for tetrahydrofuranyl-substituted DNA adducts in the livers of rats treated with NPYR. The finding of dAdo and dThd adducts is of particular interest since previous studies have shown that NPYR causes mutations at AT base pairs in DNA of rat liver.

[1]  S. Hecht,et al.  Identification of adducts formed in the reaction of alpha-acetoxy-N-nitrosopyrrolidine with deoxyribonucleosides and DNA. , 2007, Chemical research in toxicology.

[2]  S. Hecht,et al.  Identification of an acetaldehyde adduct in human liver DNA and quantitation as N2-ethyldeoxyguanosine. , 2006, Chemical research in toxicology.

[3]  S. Hecht,et al.  Mass spectrometric analysis of relative levels of pyridyloxobutylation adducts formed in the reaction of DNA with a chemically activated form of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone. , 2005, Chemical research in toxicology.

[4]  K. Kanki,et al.  In vivo mutational analysis of liver DNA in gpt delta transgenic rats treated with the hepatocarcinogens N‐nitrosopyrrolidine, 2‐amino‐3‐methylimidazo[4,5‐f]quinoline, and di(2‐ethylhexyl)phthalate , 2005, Molecular carcinogenesis.

[5]  S. Hrudey,et al.  Detecting N-nitrosamines in drinking water at nanogram per liter levels using ammonia positive chemical ionization. , 2004, Environmental science & technology.

[6]  Shana J Sturla,et al.  Identification of adducts formed by pyridyloxobutylation of deoxyguanosine and DNA by 4-(acetoxymethylnitrosamino)-1-(3-pyridyl)-1-butanone, a chemically activated form of tobacco specific carcinogens. , 2003, Chemical research in toxicology.

[7]  S. Hecht,et al.  Reactions of α-acetoxy-N-nitrosopyrrolidine with deoxyguanosine and DNA , 2001 .

[8]  S. Hecht,et al.  Reactions of 2,6-dimethyl-1,3-dioxane-4-ol (aldoxane) with deoxyguanosine and DNA. , 2001, Chemical research in toxicology.

[9]  S. Hecht,et al.  Identification of DNA adducts of acetaldehyde. , 2000, Chemical research in toxicology.

[10]  S. Hecht,et al.  Identification of paraldol-deoxyguanosine adducts in DNA reacted with crotonaldehyde. , 2000, Chemical research in toxicology.

[11]  M. Oehme,et al.  Structure Elucidation of 2,4-Dinitrophenylhydrazone Derivatives of Carbonyl Compounds in Ambient Air by HPLC/MS and Multiple MS/MS Using Atmospheric Chemical Ionization in the Negative Ion Mode. , 1998, Analytical chemistry.

[12]  S. Hecht,et al.  A cyclic N7,C-8 guanine adduct of N-nitrosopyrrolidine (NPYR): formation in nucleic acids and excretion in the urine of NPYR-treated rats. , 1997, Chemical research in toxicology.

[13]  T. Severin,et al.  Reaction of 2′-deoxyguanosine with glucose , 1995 .

[14]  T. Severin,et al.  Reaction of guanosine, 2′-deoxyguanosine and guanosine-5′-monophosphate with glucose , 1994 .

[15]  S. Hecht,et al.  Formation of 7-(4-oxobutyl)guanine in hepatic DNA of rats treated with N-nitrosopyrrolidine. , 1992, Carcinogenesis.

[16]  William Lijinsky,et al.  Chemistry and Biology of N-Nitroso Compounds , 1992 .

[17]  R. C. Shank,et al.  Formation and persistence of a DNA adduct in rodents treated with N-nitrosopyrrolidine. , 1991, Carcinogenesis.

[18]  B. Spiegelhalder,et al.  Mean daily intake of volatile N-nitrosamines from foods and beverages in West Germany in 1989-1990. , 1991, Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association.

[19]  S. Hecht,et al.  Detection of cyclic 1,N2-propanodeoxyguanosine adducts in DNA of rats treated with N-nitrosopyrrolidine and mice treated with crotonaldehyde. , 1989, Carcinogenesis.

[20]  S. Hecht,et al.  Detection of exocyclic guanine adducts in hydrolysates of hepatic DNA of rats treated with N-nitrosopyrrolidine and in calf thymus DNA reacted with alpha-acetoxy-N-nitrosopyrrolidine. , 1989, Cancer research.

[21]  J. Hotchkiss.  Preformed N-nitroso compounds in foods and beverages. , 1989, Cancer surveys.

[22]  S. Hecht,et al.  Identification of crotonaldehyde as a hepatic microsomal metabolite formed by alpha-hydroxylation of the carcinogen N-nitrosopyrrolidine. , 1988, Chemical research in toxicology.

[23]  R. C. Shank,et al.  Evidence for DNA adducts in rat liver after administration of N-nitrosopyrrolidine. , 1982, Biochemical and biophysical research communications.

[24]  S. Hecht,et al.  The Metabolism of Cyclic Nitrosamines , 1981 .

[25]  G. Eisenbrand,et al.  Volatile nitrosamines in food. , 1980, Oncology.

[26]  A. Sinskey,et al.  Activation of nitrosomorpholine and nitrosopyrrolidine to bacterial mutagens. , 1974, Mutation research.