Invited Perspective: Reference Standards Are Key to Environmental and Human Health Research—The Case of PFAS

Per- and polyfluoroalkyl substances (PFAS) include a broad range of ionic, neutral, and amphoteric compounds that contain fluoroalkyl moieties. The first study of occurrence of organofluorines in human blood in 1968 produced ambiguous results due to the lack of pure, authentic reference standards and the less sensitive and less specific detection technique (nuclear magnetic resonance) employed at that time.¹ More than three decades later, studies reporting global distribution and bioaccumulation of PFAS spurred renewed interest about this class of chemicals.² Although definitive and sensitive detection methods (e.g., liquid chromatography–tandem mass spectrometry) were employed in the early 2000s, the lack of adequate reference standards—and more importantly, isotopically labeled internal standards—hampered accurate quantification of these chemicals, their transformation products, and potential PFAS impurities found in biological matrices. In fact, authentic reference standards were lacking even for perfluorooctanesulfonate (PFOS) and perfluorooctanoic acid (PFOA) in 2000, leading to an outcry from scientists about the need for PFAS reference standards.³

In this issue of Environmental Health Perspectives, Trier, van Leeuwen, et al.⁴ describe an intriguing case in which the producer of a new PFAS, perfluoro ([5-methoxy-1,3-dioxolan-4-yl]oxy) acetic acid, obstructed unlimited access to a reference standard of this chemical for research. A similar issue with another PFAS, chloroperfluoroether carboxylates, occurred in 2020, as reported by Wagner and Gold.⁵ In both cases, the chemical manufacturers prevented the distribution of reference standards to research laboratories and threatened legal action alleging patent infringement.^4,5 Unfortunately, situations such as this are not uncommon,⁵ and they can obstruct scientific advances in understanding the environmental distribution, exposure, fate, and toxicity of a chemical in question.

Trier, van Leeuwen, et al.⁴ highlight the vital role that reference standards play in designing, developing, and executing environmental, chemical, and toxicological studies, which yield data that lay the foundation for formulating environmental and public health policies and regulations. Lack of a reference standard precludes the definitive identification and quantification of molecules found in the environment. I believe a well-characterized reference standard, certified for its purity and composition, is the basic requirement for delivering accurate, precise, and reproducible analytical and empirical data in environmental chemical research.

Reference standards are often purchased from companies that either procure them from manufacturers or synthesize and characterize them to meet purity and traceability standards. Most environmental scientists, toxicologists, and researchers in human health sciences rely on these companies for acquiring reference standards. Restricted access to reference standards could hinder research and thereby delay the scientific advances needed to protect humans and the environment from potential toxic threats posed by chemicals. The threat of legal action, whether based on a valid claim or not, can discourage companies, vendors, and individual chemists from synthesizing or marketing reference standards.

Another article recently published in Environmental Health Perspectives provides evidence that the lack of authentic reference standards led to inaccurate assessment of actual chemical exposures. Kotlarz et al.⁶ reported that median serum concentration of perfluoro-3,5,7,9,11-pentaoxadodecanoic acid (PFO5DoA) was underestimated by a factor of 30 due to the unavailability of a reference standard at the time they conducted an earlier investigation in 2018. In that earlier study,⁷ the authors had used a technical-grade PFO5DoA standard provided by Chemours (a fluorochemical manufacturing industry), which was later found to contain some interferences that resulted in underestimation of the actual concentration of this compound in human serum. The authors repeated analysis of serum samples in 2022, using a pure reference standard that had since become available, and found that the median concentration of PFO5DoA was $9.2\text{\hspace{0.17em}ng}/\mathrm{mL}$ instead of the $0.3\text{\hspace{0.17em}ng}/\mathrm{mL}$ previously reported.⁶

Aside from the additional cost and time involved in reanalysis of samples, the lack of a reference standard potentially altered the regulatory and public health implications of the earlier study⁷; the original study implied that PFO5DoA was minimally bioaccumulative, whereas the reanalysis showed that the concentration of this compound was the highest of all PFAS measured in the samples. This reiterates the vital significance of reference standards for the accurate assessment of exposures.

Currently, barely 100 of the $>\mathrm{12,000}$ PFAS available in commerce⁸ can be measured quantitatively in environmental and biological matrices due to a lack of reference standards.⁹ Nontargeted high-resolution mass spectrometry techniques for the analysis of “unknown” PFAS have identified $>750$ fluoroalkyl compounds belonging to $>130$ diverse classes.⁹ In the absence of reference standards, these results are qualitative. Although access to technical-grade formulations from chemical manufacturers may yield some clues to chemical information, it is important that the purity and composition of these formulations be assessed prior to use in environmental analysis. Reference standards should meet traceability criteria and should accompany certification of purity and composition.

The experiences of Trier, van Leeuwen, et al.⁴ and Wagner and Gold⁵ reflect the ease with which chemical manufacturers can block the sale of a reference standard. This sets up a precarious precedent for other chemical manufacturers. The precautionary principle—applied to all pharmaceuticals via clinical trials before they are released to the market—is not used when new chemicals are introduced into commerce. I propose that legislative reforms are needed to allow for unrestricted access to reference standards for researchers. Although regulations should still protect legitimate confidentiality interests of chemical manufacturers, one could argue that the right to challenge a patent infringement is lost when a chemical is detected in the environment or human bodies. I further propose that regulatory agencies require manufacturers to provide a reference standard for each registered chemical and make it publicly available. Most, if not all, environmental research is aimed at protecting ecosystem and human health, especially for the safeguarding of future generations, not to impair the chemical manufacturing industry. Any effort to block environmental research, such as obstruction of chemical reference standards, can be expected to result in exponentially higher costs to be paid by exposed populations or industries for generations to come.

Conclusions and opinions are those of the individual authors and do not necessarily reflect the policies or views of EHP Publishing or the National Institute of Environmental Health Sciences.

Refers to https://doi.org/10.1289/EHP12331