The Use of Protein Adducts to Investigate the Disposition of Reactive Metabolites of Benzene

Abstract

Benzene is metabolized to a number of electrophilic species that are capable of binding to both DNA and proteins. We used adducts of hemoglobin (Hb) Although both mice and rats showed dose-related increases in Hb and bone marrow protein adducts of BO and of the two benzoquinones, large intertissue and interspecies differences were noted, suggesting different preferences in metabolic pathways. The highest levels of adducts in mice were of 1,4-BQ (10-27% of all cysteine adducts), while in rats, BO adducts predominated in Hb (73% of all cysteine adducts) and 1,2-BQ adducts predominated in the bone marrow (14% of all cysteine adducts). High background levels of 1,2-BO and 1,4-BQ adducts were also detected in both species, indicating that the toxic effects of quinone metabolites may only be important at high levels of benzene exposure.

The Role of Metabolism and Specific Metabolites in Benzene-Induced Toxicity: Evidence and Issues

“…In humans, a possible interpretation is that these individuals, while not occupationally exposed to benzene, were exposed via other mechanisms such as smoking or gasoline. A possible toxicological implication of relatively high levels of background benzene oxide or benzoquinone adducts is that these metabolites may only be relevant for toxicity at high levels of benzene exposure (McDonald et al, 1994;Rappaport et al, 1996;Yeowell-O'Connell et al, 1998). …”

Highly Reactive Cysteine Residues in Rodent Hemoglobins

“…This suggests that benzoquinone forms adducts with these residues before reacting with any other nucleophile on the protein such as Cys-93␤ and the N-terminal valine, both of which play potential roles in the allosteric and regulatory properties of hemoglobin. This high reactivity could explain the high background levels of hemoglobin adducts, presumably due to the endogenous formation and dietary consumption of phenolic compounds (28). Independent of its specific role in benzene metabolism, benzoquinone serves as a model for electrophilic, carcinogenic metabolites in general.…”

“…Quinones even by themselves are thought to be toxic through adduct formation and other quinone mediated processes (25). Benzoquinone DNA adducts are potentially mutagenic (26) and hemoglobin adducts, S-(2,5-dihydroxyphenyl) conjugates which form via a Michael Addition (27), have been found in vivo (28). Assays incubating highly reactive cysteines with benzoquinone in vitro show that the electrophile quantitatively conjugates with hemoglobin leading to an instantaneous 1:1 decrease in the amount of such residues (Fig.…”

Fractionation of protein adducts in rats and mice dosed with [ 14 C]pentachlorophenol

“…Several studies have successfully applied Raney Ni to cleave cysteinyl adducts in various tissues, i.e., those of styrene 7,8-oxide styrene in blood proteins (Ting et al 1990;Rappaport et al 1993;Yeowell-O'Connell et al 1996), of benzoquinone in blood and bone-marrow proteins Rappaport et al 1996), of chlorinated quinones from PCP in blood and liver proteins (Lin et al 1999;Tsai et al 2001;Waidyanatha et al 1996), and of polychlorinated biphenyl quinones in liver and brain proteins (Lin et al 2000). By administering [ 14 C/ 12 C 6 ]PCP to animals along with the sequential counting scheme shown in Fig.…”