This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Hydrolytic and oxidative metabolism of pesticides contribute to their detoxification and excretion from exposed organisms. To enable development of physiologically based pharmacokinetic (PBPK) models that predict the disposition of pesticides in humans, the relative contribution of each biotransformation pathway to pesticide clearance requires quantitative characterization. The central hypothesis to be addressed in this project is that biotransformation of pyrethroid insecticides by carboxylesterases (CES) and cytochrome P450 (CYP) monooxygenases largely influence the pharmacokinetics of these pesticides. This study will investigate the role of rodent and human CES and CYP enzymes in the in vitro metabolism of permethrin and resmethrin (type I pyrethroids) and deltamethrin and cypermethrin (type II pyrethroids), which have not been investigated with respect to their human metabolism. Two major carboxylesterase isoforms have been identified in human liver and are termed hCE-1 and hCE-2.
Specific aim #1 will involve investigating the substrate specificity of baculovirus-expressed hCE-1 and hCE-2 toward the pyrethroid compounds. Hydrolysis products of the pyrethroids will be detected and quantified by LC-MS methods. The kinetic parameters (Km, kcat) that describe the rates of cleavage of the ester-containing pyrethroids will be estimated for each isozyme to determine which compounds are most effectively hydrolyzed.
Specific aim #2 will use recombinant human CYP isozymes and determine their substrate specificity and kinetic parameters toward the pyrethroid compounds. The catalytic efficiencies of the human CYP and CES enzymes will be directly compared so that the relative contribution of each enzyme to pyrethroid biotransformation can be determined. The in vitro toxicokinetic studies described in this proposal will complement and support in vivo studies in the development of PBPK models of pyrethroids. The knowledge of pesticide structure-activity relationships with respect to esterase and oxidative metabolism will be critical for predicting the metabolism and pharmacokinetics of pesticides in humans.
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