A sensitive assay for quantitative determination of the vinyl chloride (VC)-induced cyclic DNA adduct N2,3-ethenoguanine (EG) was developed. The method is based on the detection of EG as its di-pentafluorobenzyl derivative (3,5-PFB2-EG). This compound exhibited good gas chromatographic properties and was detected with high sensitivity by gas chromatography with electron capture detection (limit of detection 300 amol/microliters injected solution) or with negative ion chemical ionization mass spectrometry monitoring the [M-181]-fragment ion at m/z 354 (GC-NICI-MS, limit of detection 190 amol/microliters injected solution). EG, its 13C-labeled analog [13C4]-EG and 3,5-PFB2-EG were synthesized and characterized by UV and fluorescence spectrophotometry, 1H- and 13C-NMR spectroscopy and mass spectrometry. The standards were used to optimize the isolation of EG and its derivatization with pentafluorobenzyl bromide (electrophore labeling) at fmol quantities. DNA solutions were spiked with EG, the DNA was depurinated by mild acid hydrolysis, and EG was isolated from the hydrolysates by low-pressure strong cation exchange chromatography with subsequent C18 solid-phase extraction. The extracted EG was electrophore labeled and 3,5-PFB2-EG was detected using GC-NICI-MS. [13C4]EG served as internal standard. 3,5-PFB2-EG was quantitated relative to its 13C-labeled analog by measuring the ion ratio m/z 354/358. The limit of detection for the complete method was 60 fmol EG/mumols guanine. The method was applied to liver DNA from young Sprague-Dawley rats exposed to 600 p.p.m. VC from day 10 through day 14 after birth. The EG concentration in these samples was 1.8 +/- 0.3 pmol/mumols guanine.

1,3-Butadiene (BD), an important commodity chemical used in the production of synthetic rubber, is carcinogenic in B6C3F1 mice and Sprague-Dawley rats, raising concern for potential carcinogenicity in humans. Mice are more sensitive than rats to the carcinogenic effects of BD. Metabolic activation of BD to form the putative DNA-reactive metabolites, butadiene monoxide (BMO) and butadiene diepoxide (BDE), is mediated by cytochrome P450. Detoxication of the epoxides occurs by glutathione S-transferase-catalyzed conjugation with glutathione and hydrolysis by epoxide hydrolase. Species differences in metabolic activation and detoxication most likely contribute to the difference in carcinogenic potency of BD by modulating the circulating blood levels of the epoxides. This study measured the in vivo concentrations of BD, BMO and BDE in the blood of male Sprague-Dawley rats and B6C3F1 mice during and following 6 h nose-only exposure to inhaled BD at 62.5, 625 or 1250 p.p.m. BD. Blood samples for BD and BMO (> or = 3 samples/time point) were collected at 2, 3, 4 and 6 h of exposure. Blood samples for BDE were collected at 3 and 6 h of exposure. After exposure, blood samples for BD, BMO and BDE were collected at 2-10 min intervals up to 30 min post-exposure. BD was quantified by gas chromatography using a vial headspace equilibration technique. BD epoxides were extracted into methylene chloride and quantified by gas chromatography-mass spectrometry. The concentration of BD in blood was not directly proportional to the inhaled concentration of BD, suggesting that the uptake of BD was saturable at the highest inhaled concentration. In both rats and mice, BD and BMO blood levels were at steady-state at 2, 3, 4 and 6 h of exposure, and declined rapidly after removal from exposure to BD. Steady-state blood concentrations of BD were 2.4, 37 and 58 microM in mice and 1.3, 18 and 37 microM in rats exposed to 62.5, 625 and 1250 p.p.m. BD respectively. Both species formed BMO from BD. In mice the respective steady-state BMO concentrations in blood were 0.6, 3.7 and 8.6 microM, compared to BMO blood concentrations in rats of 0.07, 0.94 and 1.3 microM. Mice, but not rats, had quantifiable levels of BDE in the blood. The peak concentrations of BDE in the blood of mice at 6 h were 0.65, 1.9 and 2.5 microM.(ABSTRACT TRUNCATED AT 400 WORDS)

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Human exposure to acrylonitrile (ACN), a carcinogen in rats, may occur in industrial settings, through waste water and tobacco smoke. ACN is an electrophilic compound and binds covalently to nucleophilic sites in macromolecules. Measurements of adducts with hemoglobin could be utilized for improved exposure assessments. In this study, a method for quantification of N-(2-cyanoethyl)valine (CEVal), the product of reaction of ACN with N-terminal valine in hemoglobin has been developed. The method is based on the N-alkyl Edman procedure, which involves derivatization of the globin with pentafluorophenyl isothiocyanate and gas chromatographic-mass spectrometric analysis of the resulting thiohydantoin. An internal standard was prepared by reacting valylglycylglycine with [2H3]ACN, spiked with [14C]ACN to a known sp. act. Levels of CEVal were measured in globin from rats exposed to 3-300 p.p.m. ACN in drinking water for 105 days and from humans (four smokers and four non-smokers). CEVal was detected at all exposure levels in the drinking water study. The relationship between adduct level and water concentration was linear at concentrations of 10 p.p.m. (corresponding to an average daily uptake of c. 0.74 mg ACN/kg body wt during the 65 days prior to sacrifice) and below, with a slope of 37.7 pmol CEVal/g globin/p.p.m. At higher concentrations, adduct levels increased sublinearly, indicating saturation of a metabolic process for elimination of ACN. Comparison of adduct formation with the estimated dose (mg/kg/day) of ACN indicated that at low dose (0-10 p.p.m.) CEVal = 0.508 x ACN dose + 0.048 and at high dose (35-300 p.p.m.) CEVal = 1.142 x ACN dose - 1.098. Globin from the smokers (10-20 cigarettes/day) contained about 90 pmol CEVal/g, whereas the adduct levels in globin from non-smokers were below the detection limit. The analytical sensitivity should be sufficient to allow monitoring of occupationally exposed workers at levels well below the current Occupational Safety and Health Administration standard of 2 p.p.m.

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