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Assessment of metabolic stability using the rainbow trout (Oncorhynchus mykiss) liver S9 fraction
Johanning, K., G. Hancock, B. Escher, A. Adekola, M. Bernhard, C. Cowan-Ellsberry, J. Domoradzki, S. Dyer, C. Eickhoff, M. Embry, S. Erhardt, P. Fitzsimmons, M. Halder, J. Hill, D. Holder, R. Johnson, S. Rutishauser, H. Segner, I. Schultz, AND J. Nichols. Assessment of metabolic stability using the rainbow trout (Oncorhynchus mykiss) liver S9 fraction. Current Protocols in Toxicology. Wiley InterScience, Silver Spring, MD, 53:4.10.1-14.10.28, (2012).
Biotransformation reduces the extent to which organic chemicals accumulate in fish, but this activity is difficult to predict from chemical attributes such as molecular size or hydrophobicity (typically represented as log Kow). Instead, researchers are developing in vitro methods to measure biotransformation directly and then extrapolating this information to the intact animal. Studies conducted to date provide a proof-of-concept for this approach. However, the methods employed by different groups have varied resulting in problems of interpretation. The goal of this effort was to develop a standardized method for conducting in vitro metabolism assays with fish using trout liver S9 fractions. The paper that describes this method provides a template for conducting S9 depletion assays with fish as well as guidance on fish handling, liver processing, and chemical analyses. This research will allow greater comparability between laboratories using these methods, resulting in much greater confidence in the method and resulting data.
Standard protocols are given for assessing metabolic stability in rainbow trout using the liver S9 fraction. These protocols describe the isolation of S9 fractions from trout livers, evaluation of metabolic stability using a substrate depletion approach, and expression of the result as in vivo intrinsic clearance. Additional guidance is provided on the care and handling of test animals, design and interpretation of preliminary studies, and development of analytical methods. Although initially developed to predict metabolism impacts on chemical accumulation by fish, these procedures can be used to support a broad range of scientific and risk assessment activities including evaluation of emerging chemical contaminants and improved interpretation of toxicity testing results. These protocols have been designed for rainbow trout and can be adapted to other species as long as species-specific considerations are modified accordingly (e.g., fish maintenance and incubation mixture temperature). Rainbow trout is a cold-water species. Protocols for other species (e.g., carp, a warm-water species) can be developed based on these procedures as long as the specific considerations are taken into account.
Record Details:Record Type: DOCUMENT (JOURNAL/PEER REVIEWED JOURNAL)
Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL HEALTH AND ENVIRONMENTAL EFFECTS RESEARCH LABORATORY
MID-CONTINENT ECOLOGY DIVISION