Epoxybutene-hemoglobin adducts in rats and mice: dose response for formation and persistence during and following long-term low-level exposure to butadiene.

Measurement of specific adducts to hemoglobin can be used to establish the dosimetry of electrophilic compounds and metabolites in experimental animals and in humans. The purpose of this study was to investigate the dose response for adduct formation and persistence in rats and mice during long-term low-level exposure to butadiene by inhalation. Adducts of 3,4-epoxy-1-butene, the primary metabolite of butadiene, with N-terminal valine in hemoglobin were determined in male B6C3F1 mice and male Sprague-Dawley rats following exposure to 0, 2, 10, or 100 ppm of 1,3-butadiene, 6 h/day, 5 days/week for 1, 2, 3, or 4 weeks. Blood samples were collected from groups of five mice and three rats at the end of each week during the 4 weeks of exposure and weekly for 3 weeks following the end of the 4-week exposure period. The increase and decrease, respectively, of the adduct levels during and following the end of the 4-week exposure followed closely the theoretical curve for adduct accumulation and removal for rats and mice, thereby demonstrating that the adducts are chemically stable in vivo and that the elimination follows the turnover of the red blood cells. The adduct level increased linearly with butadiene exposure concentration in the mice, whereas a deviation from linearity was observed in the rats. For example, after exposure to 100 ppm butadiene, the epoxybutene-hemoglobin adduct levels were about four times higher in mice than in rats; at lower concentrations of butadiene, the species difference was less pronounced. Blood concentrations of epoxybutene, estimated from hemoglobin adduct levels, were in general agreement with reported concentrations in mice and rats exposed by inhalation to 62.5 ppm. These studies show that adducts of epoxybutene with N-terminal valine in hemoglobin can be used to predict blood concentration of epoxybutene in experimental animals.

Monitoring exposure to acrylonitrile using adducts with N-terminal valine in hemoglobin.

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.

Use of haemoglobin adducts for biomonitoring exposure to 1,3-butadiene.

Measurement of specific adducts to haemoglobin can be used to establish the dosimetry of electrophilic compounds and metabolites in experimental animals and in man. Adducts of 1,2-epoxybutene with the N-terminal valine in haemoglobin were determined in male B6C3F1 mice and Sprague-Dawley rats following exposure by inhalation to low concentrations of 1,3-butadiene (0, 2, 10 or 100 ppm, 6 h/day, 5 days/week, 4 weeks; animals killed within 1 h after the last exposure). The adduct level increased linearly with butadiene concentration in the mice, whereas a deviation from linearity was observed in the rats. After exposure to 100 ppm butadiene, the adduct levels were four times higher in mice than in rats; at lower concentrations of butadiene, the species difference was less pronounced. Adduct levels of about 1-3 pmol/g globin were recorded in human subjects (nonsmokers) who worked in a production area where butadiene levels of about 1 ppm had been recorded in a survey conducted three to nine months prior to this study. Increased adduct levels were also observed in cigarette smokers (two subjects) who were not exposed occupationally to butadiene. Although preliminary, the data suggest that the adduct levels, and consequently the doses of epoxybutene per parts per million-hour of butadiene are lower in humans than in mice and rats. The adduct levels are much lower than those seen after occupational exposures to corresponding air levels of ethylene oxide and are lower than those seen after exposure to ethylene.

Studies of biological and chemical monitoring of low-level exposure to ethylene oxide.

Ten sanitary workers exposed to concentrations of ethylene oxide below 1 ppm were studied to determine whether effects could be observed at low exposure levels. A significant increase in the number of sister chromatid exchanges in cultured lymphocytes was found only for five subjects with relatively high exposure in the sterilization area. However, it was not possible to separate clearly the effect of smoking from that of ethylene oxide exposure. No increase in the frequencies of micronuclei in lymphocytes and buccal cells was found. The level of 2-hydroxyethyl adducts to the N-terminal valines in hemoglobin responded in a reliable fashion to chronic ethylene oxide exposure and smoking. Furthermore, measurement of levels of 2-hydroxyethyl adducts to the N-terminal valines in hemoglobin made it possible to reconstruct the dynamics of a leakage of ethylene oxide which involved three workers.

Ethylene oxide doses in ethene-exposed fruit store workers.

Blood samples from 10 ethene-exposed fruit store workers and 10 referents were analyzed for the level of hydroxyethyl adducts to N-terminal valine in hemoglobin (Hb). A statistically significant difference was obtained between the nonsmoking workers (N = 7) and the nonsmoking referents (N = 6). This finding demonstrates that ethene is metabolized to ethylene oxide in man. According to this preliminary study, ethene exposure at 0.3 (uncertainty range 0.1-1) ppm during work hours increases the adduct level by 23 pmol/g Hb. This figure is compatible with a metabolic conversion of 3% (1-10%) of the inhaled ethene to ethylene oxide.