Reactive Naphthalene Metabolite Binding to Hemoglobin and Albumin

Abstract

Earlier work has shown that the murine Clara cell cytotoxi cant, naphthalene, is metabolized to reactive metabolites which deplete glutathione or, in the absence of sufficient glutathione, become bound covalently to tissue macromolecules. Correlations between bound metabolite levels in the lung with injury suggests an association between reactive metabolite binding and toxicity. In this study we examine the formation of covalent naphthalene adducts with hemoglobin and albumin in mice to determine whether these serve as useful indices of exposure and metabolism for a chemical which shows a glutathione threshold. Covalent binding of radioactivity from [3]naphthalene to both albumin and hemoglobin was dose dependent and a glutathione threshold was observed. At early times after naphthalene administration, the formation of albumin adducts was 10- to 30-fold higher than that of hemoglobin adducts. Hemoglobin and albumin adduct levels decreased by apparent first-order processes with half-lives of 11.5 and 1.8 days, respectively. These half-lives are consistent with the turnover of these blood proteins in the mouse. Pretreatment with buthionine sulfoximine resulted in higher levels of albumin adduct but in no alteration of hemoglobin adduct levels in comparison with control. In contrast, diethylmaleate pretreatment increased the level of hemoglobin adduct but not albumin adduct. The antibody to naphthalene mercapturates recognized the hemoglobin adduct(s) but not the albumin adduct(s). Comparison of the data from ELISA (standardized using hydroxymercaptodihydronaphthalene) and radiochemical analysis yielded curves with identical slopes; the absolute levels of adduct found by ELISA were approximately half those measured with radiochemical techniques. These studies indicate that high levels of reactive naphthalene metabolites are bound to hemoglobin and albumin at doses above the glutathione threshold. Although hemoglobin adduct levels can be monitored sensitively with ELISA, the presence of the glutathione threshold suggests that monitoring exposed populations may not provide a useful measure of exposure at environmentally relevant exposure levels.