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NMR ANALYSIS OF MALE FATHEAD MINNOW URINARY METABOLITES: A POTENTIAL APPROACH FOR STUDYING IMPACTS OF CHEMICAL EXPOSURES
EKMAN, D. R., Q. TENG, K. M. JENSEN, D. MARTINOVIC, DAN VILLENEUVE, G. T. ANKLEY, AND T. W. COLLETTE. NMR ANALYSIS OF MALE FATHEAD MINNOW URINARY METABOLITES: A POTENTIAL APPROACH FOR STUDYING IMPACTS OF CHEMICAL EXPOSURES. AQUATIC TOXICOLOGY. Elsevier Science Ltd, New York, NY, 85(2):104-112, (2007).
This task is divided into four major research areas: (1) Development of computational tools and databases for screening-level modeling of the environmental fate of organic chemicals; (2) Metabolism of xenobiotics: Enhancing the development of a metabolic simulator; (3) Metabonomics: The use of advanced analytical tools to identify toxicity pathways; and (4) Software infrastructure to support development and application of transformation/metabolic simulators.
For many chemicals, multiple transformation/metabolic pathways can exist. Consequently, transformation/metabolic simulators must utilize transformation rate data for prioritization of competing pathways. The prioritization process thus requires the integration of reliable rate data. When this data is absent, it is necessary to generate a database with metabolic and transformation rate constants based on: (1) experimentally measured values, including those requiring the use of advanced analytical techniques for measuring metabolic rate constants in vivo and in vitro; (2) rate constants derived from SPARC and mechanistic-based QSAR models; and (3) data mined from the literature and Program Office CBI. A long-term goal of this project is to build this database. This information will be used to enhance the predictive capabilities of the transformation/metabolic simulators. As indicated previously, exposure genomics, which provide early signs of chemical exposure based on changes in gene expression, will be used to guide chemical fate and metabolism studies. The incorporation of exposure genomics into fate studies will provide information concerning (1) the minimal concentrations at which biological events occur; and (2) the identification of biologically relevant chemicals(s) in mixtures.
The capability of categorizing chemicals and their metabolites based on toxicity pathway is imperative to the success of the CompTox Research Program. Metabonomics, which is the multi-parametric measurement of metabolites in living systems due to physiological stimuli and/or genetic modification, provides such a capability. The application of metabonomics to toxicity testing involves the elucidation of changes in metabolic patterns associated with chemical toxicity based on the measurement of component profiles in biofluids, and enables the generation of spectral profiles for a wide range of endogenous metabolites. Metabolic profiles can provide a measure of the real outcome of potential changes as the result of xenobiotic exposure.
The potential for profiling endogenous metabolites in urine from male fathead minnows (Pimephales promelas) to assess chemical exposures was explored using nuclear magnetic resonance (NMR) spectroscopy. Both one dimensional (1D) and two dimensional (2D) NMR spectroscopy was used for the assignment of metabolites in urine from unexposed fish. Because fathead minnow urine is dilute, we lyophilized these samples prior to analysis. Furthermore, 1D 1H-NMR spectra of unlyophilized fathead minnow and rat urine were acquired to allow comparison of relative water content and to compare rat and fish metabolite profiles. As a small proof-of-concept study, lyophilized urine samples from male fathead minnows exposed to three different concentrations of the antiandrogen vinclozolin were analyzed by 1D 1H-NMR to assess exposure-induced changes. Two partial least squares discriminant analysis (PLS-DA) models were constructed - one comparing the low-exposure class (100 g/L, nominal), and one comparing the high-exposure class (700 g/L, nominal), to controls. Weight loadings plots from these two models were used to determine changes in urinary metabolite profiles associated with the exposure. The results of this study demonstrate the potential utility of an NMR-based approach for assessing chemical exposures in male fathead minnow, using urine collected from individual fish.
Record Details:Record Type: DOCUMENT (JOURNAL/PEER REVIEWED JOURNAL)
Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL EXPOSURE RESEARCH LABORATORY
ECOSYSTEMS RESEARCH DIVISION
PROCESSES & MODELING BRANCH