Per- and polyfluoroalkyl substances (PFAS) are a diverse group of chemicals that have been used as components of aqueous film-forming foam (AFFF) for decades. Early formulations used perfluorooctane sulfonate (PFOS) in large proportion. Unfortunately, long-chain PFAS like PFOS have since been found to be bioaccumulative and toxic, prompting voluntary phase-outs, establishment of drinking water guidelines by the U.S. EPA and many states, and the need to remediate contaminated water for the protection of environmental and human health. Areas where AFFF were routinely deployed during firefighting exercises, including DoD sites, have accumulated a variety of PFAS in the groundwater, and are now in need of cost effective and efficient remediation solutions. Many PFAS eventually degrade to form perfluorinated alkyl acids, which are extremely persistent and have varying levels of water solubility. Because of this, ex situ treatment technologies may be most suitable. However, currently available technologies such as adsorption with activated carbon cannot effectively treat both short- and long-chain PFAS.

The objective of Phase I of this proof-of-concept project was to exploit the propensity of PFAS to bind with proteins, and using a combination of molecular modeling and batch testing, verify whether PFAS-protein interactions could be tuned to efficiently adsorb a variety of PFAS, opening a pathway to development of bio-based PFAS sorbents for treatment of AFFF-contaminated water. Results from Phase I can be found in the Final Report and are summarized below.

In Phase II, the objectives are to:

• Quantify kinetics of PFAS sorption to select proteins, which is needed to assess viability of protein sorbents to effectively treat AFFF-contaminated groundwater under realistic treatment train conditions.
• Assess protein stability and sorption performance over time under groundwater treatment-relevant conditions.
• Investigate thermal regeneration of the protein sorbents and post-regeneration sorption effectiveness.
• Continue molecular model screening on a parallel track.

This project combined molecular simulations with in vitro experiments to identify protein-based sorbents for PFAS and evaluated the PFAS-protein binding affinities.

In Phase I, candidate protein sorbents were identified from existing literature based on interactions with PFAS and with analogous ligands such as fatty acids and acidic drugs. Protein-specific binding affinities for a set of 8 selected perfluorinated alkyl acids— PFOS, PFNA, PFOA PFHxS, PFHxA, PFHpA, PFBS, and PFBA—were then predicted using a multi-step molecular modeling framework. Finally, the most promising proteins—liver and intestinal fatty acid binding proteins, L-FABP and I-FABP, and the peroxisome-proliferator-activated receptors, PPAR-α, -γ, and -δ—were experimentally evaluated for PFAS adsorption through a series of batch tests at the bench scale to determine the equilibrium dissociation constant, K_D, and the equilibrium binding capacity, q_max. EqD experiments were performed under ideal matrix conditions with single PFAS-protein pairs, under varied pH, ionic strength and temperature conditions, as well under “realistic” conditions using AFFF-contaminated groundwater and single proteins.

Initial results were promising and suggested proteins could be effective filter media but fundamental questions remained. First, EqD experiments provided K_D values and protein-PFAS sorption capacities but did not provide sorption kinetics, which would be key for filter design and prototyping. This will be addressed in Phase II during time-resolved sorption batch and column tests. Second, it may be possible to regenerate protein sorbents via low temperature heating/cooling cycles. Initial heat regeneration experiments demonstrated release of bound PFAS at elevated temperatures (50°C), but no measurable sorption to proteins was observed after heating, presumably because the proteins were denatured. Regeneration testing will explore progressively lower temperature heating/cooling cycles to identify an optimal release/regenerate cycle. Finally, durability of protein sorbents under water filter-relevant environmental conditions has never before been tested. During Phase II testing, the durability/sustained PFAS sorption performance of selected proteins will be evaluated following extended periods of incubation in non-sterile groundwater selected to be representative protein-filter influent.

In the Phase I effort, the binding of PFAS to different proteins was found to be variable, and both chain length- and condition-specific, suggesting the ability to tune PFAS removal by the use of different proteins alone or in combination. Interestingly, the nuclear receptor PPAR-δ was found to bind strongly with both PFBA and PFHxS, while PPAR-α bound strongly to PFHxA (as well as the long-chain PFNA). This is the first report to show such a strong association of biomolecules with short-chain PFAS, raising both prospects for treatability and potential concern about toxic effects. Moreover, the binding capacities calculated from protein-PFAS data for some short-chain PFAS were substantially higher than with either granular or particulate activated carbon, illustrating the potential for targeting difficult to treat PFAS. Results of the Phase I effort confirmed the potential of protein-based sorbents for PFAS remediation by identifying, through a complementary model-experiment approach, proteins that associate strongly with both long- and short-chain PFAS. Of particular benefit is the ability to tune an adsorbent, by incorporating multiple protein-based moieties and/or by use of strategic changes in feed composition (ionic strength, pH), to address a wide variety of PFAS structures. The combination of molecular modeling and batch testing used here thus forms the basis of a robust and powerful design framework for developing protein-based sorbents for PFAS water treatment.

This project has the potential to open an entirely new area of sorbent design for PFAS remediation. Of particular benefit is the ability to tune an adsorbent, by incorporating multiple protein-based moieties, to address a wide variety of PFAS structures. The combination of molecular modeling and batch testing in this project could form the basis of a robust and powerful design framework for remediation technology. An additional benefit of this approach is that it will rely on a benign (protein-based) substance to remove AFFF contaminants without introducing additional harmful substances (e.g. harsh oxidants) into the environment.

Khazaee, M., E. Christie, W. Cheng, M. Michalsen, J. Field, and C. Ng. 2021. Perfluoroalkyl Acid Binding with Peroxisome Proliferator-Activated Receptors ⍺, γ, and δ and Fatty Acid Binding Proteins by Equilibrium Dialysis with a Comparison of Methods. Toxics, 9(3): 45.