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In Vitro Toxicity Assessment Technique for Volatile Substances using Cytochrome P450 Isozyme-specific metabolic pathways
Citation:
Winters, B., M. Angrish, A. Wallace, AND M. Madden. In Vitro Toxicity Assessment Technique for Volatile Substances using Cytochrome P450 Isozyme-specific metabolic pathways. PittCon, Chicago, IL, March 05 - 09, 2017.
Impact/Purpose:
A high through put toxicity screening system is in development. The system will be able to quickly and inexpensively examine the relative toxicity of volatile pollutants. The final model of the system will allow the Agency to help meet the goals of assessing the toxicity of thousands of chemicals the Agency is tasked with monitoring and testing.
Description:
The U.S. Environmental Protection Agency is tasked with evaluating the human health, environmental, and wildlife effects of over 80,000 chemicals registered for use in the environment and commerce. The challenge is that sparse chemical data exists; traditional toxicity testing methods are slow, costly, involve animal studies, and cannot keep up with a chemical registry that typically grows by at least 1000 chemicals every year. In recent years, High Throughput Screening (HTS) has been used in order to prioritize chemicals for traditional toxicity screening or to complement traditional toxicity studies. HTS is an in vitro approach of rapidly assaying a large number of chemicals for biochemical activity using robotics and automation. However, no method currently exists for screening volatile chemicals such as air pollutants in a HTS fashion. Additionally, significant uncertainty regarding in vitro to in in vivo extrapolation (IVIVE) remains. An approach to bridge the IVIVE gap and the current lack of ability to screen volatile chemicals in a HTS fashion is by using a probe molecule (PrM) technique. The proposed technique uses chemicals with empirical human pharmacokinetic data as PrMs to study toxicity of molecules with no known data for gas-phase analysis. We are currently studying the xenobiotic-metabolizing enzyme CYP2A6 using transfected BEAS-2B bronchial epithelial cell line. The CYP2A6 pathway activity is studied by the formation of cotinine from nicotine and tert-butyl alcohol (TBA) from methyl tert-butyl ether (MTBE). Modulation of the formation of metabolites due to the introduction of a toxicant indicates an alteration of cellular homeostasis. To measure perturbations in metabolite formation, cotinine is analyzed by standard liquid phase immunoassay (ELISA), while TBA is measured using gas-chromatography-mass spectrometry (GC-MS). [This is an abstract of a proposed presentation and may not reflect official US EPA policy.]