Labs Gear Up: Advanced PFAS Detection in Water Meets New Regulations

The reach of per- and polyfluoroalkyl substances (PFAS) into water sources is emerging into stark visibility. For instance, the EPA's 2023 UCMR5 (Unregulated Contaminant Monitoring Rule) findings¹ revealed PFAS in a third of major urban water systems, impacting an estimated 70 million residents². These studies, testing just 29 out of an estimated 15,000 PFAS compounds, likely only hint at the full magnitude of contamination.

The carbon-fluorine (C-F) bond, among the strongest in organic chemistry, endows PFAS with the ability to repel oil and water and resist degradation. The stability of these chemicals leads them to persist in water sources and bioaccumulate in the human body. They particularly target protein-rich organs, such as the liver³ and kidneys⁴, due to their dual affinity for polar and nonpolar substances. The structural features of PFAS molecules, including the head's charge and the tail's length, dictate their bioaccumulation. Exposure to PFAS is linked to serious health outcomes, ranging from reproductive issues⁵, immunotoxicity⁶, endocrine disruption⁷, and an elevated risk of cancer⁸.

Navigating PFAS testing challenges in water

As the health impacts of PFAS came to light, the necessity for water testing swiftly escalated. Environmental laboratories, spanning both governmental entities and private sectors, stepped up to meet this demand. However, initial efforts to manage PFAS faced challenges due to the lack of enforceable federal guidelines on the handling, manufacturing, disposal, and removal of these chemicals. The EPA provided only guidance, not mandatory directives. Consequently, regulatory responsibilities and certifications largely devolved to individual states, leading to a patchwork of regulations.

By 2019, the EPA released Method 533⁹ to accompany Method 537.1¹⁰, providing laboratories with validated tests for 29 PFAS compounds in drinking water. Method 533 targets 25 short-chain PFAS (4 to 12 carbons) using polystyrene divinylbenzene with a positively charged diamino ligand for extraction, differing from 537.1's styrene-divinylbenzene media. It leverages a weak anion exchange SPE cartridge and isotopic dilution for enhanced accuracy.

Method 533 offers logistical benefits by reducing sample volume requirements and minimizing measurement interference, streamlining the detection process. Meanwhile, Method 537.1 identifies four unique PFAS compounds beyond those covered by 533. The overlapping analytical frameworks offered to labs underscore a fundamental challenge: the diverse properties of PFAS compounds necessitate specialized methods for accurate detection, especially at low concentrations. The EPA’s proposed drinking water limits of four parts per trillion for perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) require high-resolution techniques such as liquid chromatography/tandem mass spectrometry (LC-MS-MS).

Sample collection for PFAS analysis requires meticulous attention to avoid contamination, as numerous items can introduce unwanted PFAS into samples.

Expanding PFAS testing beyond drinking water

The limitations of testing methods designed solely for drinking water left environmental labs in a quandary for analyzing PFAS in soil, wastewater, and other samples. Enter lab-modified methods, often bearing the "537M" designation, which adapted Method 537 for broader use. Though offering some consistency, these non-standardized approaches left data vulnerable to challenges.

In January 2024, the EPA released Method 1633¹¹, a standardized approach for analyzing 40 PFAS compounds in diverse samples, including wastewater, groundwater, and fish tissue. This method, developed with extensive stakeholder feedback, resolves the challenges of non-standardized testing and empowers labs to expand this list further.

To handle PFAS in multiple matrices, Method 1633 describes a comprehensive process involving sample preparation, solid-phase extraction cleanup, and LC-MS-MS analysis. It outlines tailored procedures for aqueous, solid, and tissue samples, emphasizing homogenization, dilution, and specific reagent use for accurate PFAS extraction. Instrumental analysis leverages ultrahigh-performance liquid chromatography and mass spectrometry, detailed with quality control steps for reliable PFAS quantification and identification.

While Method 1633 significantly improves PFAS detection accuracy and extends testing capabilities across various sample types, challenges persist, particularly with complex samples containing high solids. This complexity can strain labs striving to meet the method's rigorous acceptance criteria and necessitate additional resources for quality control and broader analyte characterization. Consequently, these factors might escalate both the duration and expense of analysis.

Future advancements are being developed to mitigate these challenges, focusing on refining sample preparation techniques and enhancing analytical instruments for more efficient handling of complex samples. Innovations such as automated sample processing and improved SPE methods are poised to streamline the workflow, reduce potential errors, and lower analysis costs.

Exploring the dark matter of PFAS

The “dark matter” of PFAS encompasses the undetected compounds in the environment beyond the few dozen identified by standard tests. The Total Oxidizable Precursor (TOP) assay¹², employing hydroxyl radicals to convert unknown precursors into detectable perfluoroalkylic acids via HPLC-MS, aims to uncover this hidden spectrum. However, its efficacy is limited by the oxidizability of compounds and the analytical range of C4 to C14 chain lengths, potentially missing some PFAS precursors.

Alternatively, the Total Organofluorine (TOF) assay¹³ measures the total fluorine from organic compounds in samples via combustion ion chromatography. This process transforms organofluorine to sodium fluoride (NaF), allowing for an estimation of organofluorine content. However, it cannot specify individual PFAS compounds or their chain lengths. Converting TOF results into estimated PFAS content requires consideration of the entire molecular structure, not just the fluorine content.

Key steps for reliable PFAS sample collection

Sample collection for PFAS analysis requires meticulous attention to avoid contamination, as numerous items can introduce unwanted PFAS into samples. Avoid new, unwashed clothing, clothes treated for water resistance, and garments washed with fabric softeners. Also, avoid of consuming food in wrappers, applying personal care products such as sunscreen or insect repellent, using items with Teflon or PTFE, and wearing latex gloves.

Identifying PFAS-coated items can be challenging, necessitating testing for some materials. Specific protocols and materials, such as wearing nitrile gloves and using provided polypropylene bottles with preservatives, are recommended to minimize risks. Flushing the tap or water source before sample collection and ensuring preservatives are dissolved are crucial for obtaining accurate results. Additionally, direct collection into designated bottles ensures consistent results.

Regarding procedural rigor, including a field reagent blank in each sample set is essential for validating the absence of contamination. Different methods, such as 537.1 and 533, slightly vary in handling field reagent blanks and preservatives. All bottles, whether for samples or blanks, undergo testing at the lab before distribution to confirm they are PFAS-free. Proper handling, such as not touching the inside of the bottle or lid, helps maintain sample integrity. Finally, transporting samples back to the lab in coolers with actual ice (to avoid PFAS contamination from substitutes) and ensuring they arrive at or below 10°C within 48 hours are critical steps.

Future trends in PFAS water testing

With increased demand for PFAS analysis services, labs are adopting automated systems and advanced technology to streamline detection. High-resolution mass spectrometry, featuring Orbitrap and quadrupole time-of-flight, enhances the precision and accuracy necessary for detecting PFAS at low concentrations. Moreover, LC-MS-MS methodologies, utilizing electrospray ionization combined with scheduled multiple reaction monitoring and polarity switching, significantly improve selectivity and sensitivity in PFAS identification.

Regarding procedural rigor, including a field reagent blank in each sample set is essential for validating the absence of contamination.

Automation and cheminformatics are revolutionizing PFAS testing, from sample preparation to data analysis. SPE robots¹⁴ automate extraction processes in water samples, efficiently handling tasks like sample loading and analyte elution. Concurrently, the analysis of PFAS, with its vast structural diversity, benefits from advanced computational tools. The recent development of 129 PFAS-specific fingerprints¹⁵ facilitates streamlined profiling and categorization, bypassing the need for expert input and enhancing computational models. Furthermore, an approach termed FluoroMatch Flow 2.0¹⁶ optimizes the discovery process in LC-HRMS by automating data processing, employing a comprehensive confidence criterion to accurately identify PFAS compounds and reduce inaccuracies.

As analytical methodologies for testing PFAS in water become more advanced and widespread, the focus will likely shift towards developing worldwide standards, ensuring consistency and comparability of data across borders. Collaborations between regulatory bodies, research institutions, and industry will be crucial in pooling resources and knowledge. The future promises innovations beyond enhanced detection to solutions for its mitigation and removal, safeguarding water quality and public health.

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