Plasma Protein Binding Evaluations of Per- and Polyfluoroalkyl Substances for Category-Based Toxicokinetic Assessment

New approach methodologies (NAMs) that make use of invitro screeningand in silico approaches to inform chemical evaluations rely on invitro toxicokinetic (TK) data to translate in vitro bioactive concentrationsto exposure metrics reflective of administered dose. With 1364 per-and polyfluoroalkyl substances (PFAS) identified as of interest underSection 8 of the U.S. Toxic Substances Control Act (TSCA) and concernover the lack of knowledge regarding environmental persistence, humanhealth, and ecological effects, the utility of NAMs to understandpotential toxicities and toxicokinetics across these data-poor compoundsis being evaluated. To address the TK data deficiency, 71 PFAS selectedto span a wide range of functional groups and physico-chemical propertieswere evaluated for in vitro human plasma protein binding (PPB) byultracentrifugation with liquid chromatography-mass spectrometry analysis.For the 67 PFAS successfully evaluated by ultracentrifugation, fractionunbound in plasma (f (up)) ranged from lessthan 0.0001 (pentadecafluorooctanoyl chloride) to 0.7302 (tetrafluorosuccinicacid), with over half of the PFAS showing PPB exceeding 99.5% (f (up) < 0.005). Category-based evaluations revealedthat perfluoroalkanoyl chlorides and perfluorinated carboxylates (PFCAs)with 6-10 carbons were the highest bound, with similar medianvalues for alkyl, ether, and polyether PFCAs. Interestingly, bindingwas lower for the PFCAs with a carbon chain length of & GE;11.Lower binding also was noted for fluorotelomer carboxylic acids whencompared to their carbon-equivalent perfluoroalkyl acids. Comparisonsof the f (up) value derived using two PPBmethods, ultracentrifugation or rapid equilibrium dialysis (RED),revealed RED failure for a subset of PFAS of high mass and/or predictedoctanol-water partition coefficients exceeding 4 due to failureto achieve equilibrium. Bayesian modeling was used to provide uncertaintybounds around f (up) point estimates forincorporation into TK modeling. This PFAS PPB evaluation and groupingexercise across 67 structures greatly expand our current knowledgeand will aid in PFAS NAM development.

Automated summary

New approaches using in vitro screening and in silico approaches, rely on in vitro toxicokinetic (TK) data to evaluate chemical substances. The evaluation of 71 PFAS compounds was conducted to understand their plasma protein binding (PPB) by ultracentrifugation with liquid chromatography-mass spectrometry analysis. Results showed that over half of the PFAS compounds had PPB exceeding 99.5%, and binding was lower for longer carbon chain length PFCAs. Bayesian modeling was used to provide uncertainty bounds for incorporation into TK modeling. This evaluation greatly expands our current knowledge and will aid in PFAS NAM development.