This application focuses on the kinetics, metabolism and toxicology of two metabolites of trichloroethylene (TCE): dichloroacetate (DCA) and chloral hydrate (CH). The studies are predicated on several novel findings we made during the previous grant period that include 1) orally administered CH is catabolized to DCA in children, 2) CH and DCA alter each other's metabolism, 3) DCA is biotransformed to glyoxylate, 4) DCA inhibits maleyacetoacetate isomerase (MAAI, also known as GST- zeta), a key enzyme in tyrosine metabolism 5) MAAI inhibition by DCA may cause accumulation of intermediates in the tyrosine catabolic pathway that may be responsible for the hepatotoxicity and neurotoxicity of DCA and 6) this inhibition may also be responsible for prolonging the elimination half-life of subsequent DCA doses. The following specific aims capitalize on these discoveries: (1): Determine the in vivo kinetics and biotransformation of CH in health adults and the influence of CH and DCA on each other's metabolism and toxicity. We will examine several postulates regarding CH and DCA biotransformation in humans. [13C] CH and DCA will be administered at environmentally (mug/kg) and clinically (mg/kg) relevant doses to adults and the kinetics and metabolism of these chemicals will be studied. (2): Quantify the effect of DCA and CH on tyrosine catabolism in children and adults by measuring the plasma and urinary concentrations on tyrosine, its metabolites and the heme precursor aminolevulinate. We will test hypotheses relevant t the perturbation of the tyrosine catabolic pathway by xenobiotics, via inhibition of MAAI. Such inhibition should lead to accumulation to accumulation of potentially hepatotoxic and neurotoxic metabolites. (3): Compare and contrast the in vivo human data with similar experiments performed in intact cells. We will address hypotheses concerning CH and DCA kinetics and biotransformation and their effects on tyrosine metabolism in animals. (4): Elucidate the molecular mechanisms of DCA and CH biotransformation, and their effects of biotransformation enzymes, using cellular and subcellular fractions of rat and human liver. We will test postulates regarding early molecular events involved in DCA biotransformation and their effects of biotransformation enzymes, using cellular and subcellular fractions of rat and human liver. We will test postulates regarding early molecular events involved in DCA biotransformation and the mechanism by which DNA influences MAAI activity.
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