Development of Amphibian Per- and Polyfluoroalkyl Substances Toxicity Reference Values for use in Ecological Risk Assessment at Aqueous Film-Forming Foam Sites

Dr. Maria Sepulveda | Purdue University




The use of aqueous film-forming foams containing poly- and perfluoroalkyl substances (PFASs) at Defense Environmental Restoration Program (DERP) fire, crash, and training sites has potentially contributed to PFAS contamination of soil, water, and sediment. The overall objective of this SERDP effort is to develop amphibian toxicity reference values (TRVs) to support ecological risk assessment of PFASs in contaminated DERP sites. Such data are needed for supporting cleanup and/or exposure mitigation decisions. Specifically, the project team aims to: 1) develop toxicity reference values for perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) in amphibians, and 2) determine the potency of two additional PFASs in amphibians: 6:2 fluorotelomer sulfonate and perfluorohexane sulfonate.

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Technical Approach

TRVs will be developed following standard published toxicity test procedures for Northern Leopard Frogs (Lithobates pipiens), American Toads (Anaxyrus americanus), and Eastern Tiger Salamanders (Ambystoma tigrinum). Toxicity testing will be conducted under steady state conditions, which will be determined prior to testing through range finding studies. Route of exposure, length of exposure, and life stage for these toxicity tests will include: 1) Aquatic Chronic Larvae 21 d; 2) Sediment Subchronic Larvae 10 d; 3) Terrestrial Dermal Chronic Adults 21 d; and 4) Terrestrial Oral Chronic Adults 42 d. Each assay will be validated through a partnering laboratory. Experiments will be conducted using larvae and juveniles reared from egg masses collected from natural amphibian populations located in West Lafayette, Indiana. To generate juveniles for the experiments, a subset of the species will be moved into outdoor mesocosms. Exposure levels will be determined from the range-finding studies (from 10X above background concentrations to no greater than 1,000 μg/L). Based on these results, 5 concentrations will be selected and each treatment will be replicated 4 – 8 times for a total of 20 – 40 experimental units per assay. Experiments will begin at Gosner stage 30 for the anurans and Harrison stage 46 for the salamanders. Although PFASs are expected to be very stable, multiple water and/or sediment samples will be collected over time to assess the amount of breakdown that has occurred. Dose-response curves will be calculated for growth, development (time to, and effects on metamorphosis), and survival. Then TRVs will be calculated and compared across species and compounds. At the end of the chronic exposures, chemical concentrations will be quantified from whole-animal tissue on a set of the experimental samples for expression of TRVs based on body burdens. PFAS extraction will be conducted following protocols established and modified for optimal recoveries and extracts analyzed by ultra-high pressure liquid chromatography tandem mass spectrometry in negative electrospray ionization.

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The core outcome of these studies will be the development of amphibian TRVs for PFASs that can be used in the derivation of surface water, sediment and soil risk-based environmental criteria for assessments at PFAS-contaminated sites. The ultimate goal is not only to develop TRVs that are protective of the most sensitive amphibian species, but also that can be used for a more refined identification of surrogate species for the evaluation of specific amphibian taxa. The project team will work to answer the following questions:

  1. What amphibian species and what life stages are most sensitive to PFASs?
  2. What is the range of amphibian TRVs for PFASs varying in environmental persistence and bioavailability?
  3. What is the range of bioaccumulation factors for PFASs varying in environmental persistence and bioavailability; and
  4. Are there differences in TRVs between different exposure routes (oral, dermal, gills)?

(Anticipated Project Completion - 09/2021)

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Hoover, G.M., M.F. Chislock, B.J. Tornabene, S.C. Guffey, Y.J. Choi, C.D. Perre, J.T. Hoverman, L.S. Lee, and M.S. Sepulveda. 2017. Uptake and Depuration of Four Per/Polyfluoroalkyl Substances (PFASS) in Northern Leopard Frog Rana pipiens Tadpoles. Environmental Science Technology Letters, 4(10):399-403. 

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Points of Contact

Principal Investigator

Dr. Maria Sepulveda

Purdue University

Phone: 765-496-3428

Program Manager

Environmental Restoration