Development of Novel Analytical Methods for Quantifying Group Concentrations of PFAS, Nitro Compounds, and Nitriles in Water

Abstract

The increasing pressures of population growth, climate change, urbanization, and water reuse have intensified the introduction of emerging micropollutants into aquatic environments and drinking water sources. Among these, per- and polyfluoroalkyl substances (PFAS, CnF2n+1–R) have emerged as a class of persistent, bioaccumulative, and widely detected anthropogenic contaminants, yet current monitoring approaches capture only a limited fraction of the vast diversity of PFAS structures. In parallel, dissolved organic nitrogen (DON) containing amine-based moieties (e.g., amines, amino acids, peptides, and proteins) involved in water disinfection or oxidative treatment processes serve as important precursors of diverse nitrogenous disinfection byproducts (N-DBPs). While nitrosamines have been extensively investigated, other N-DBP groups, such as nitro (R–NO2) and nitrile compounds (R–CN), remain underexplored despite their potential toxicity and significance in water treatment and reuse contexts. Therefore, further elucidation of the formation, transformation, and quantification of these N-DBP species is essential for comprehensive water quality assessment and management. Targeted analyses, typically employing liquid or gas chromatography coupled with mass spectrometry (LC–MS or GC–MS), allow for sensitive and selective detection of individual compounds. However, their applicability is limited by the availability of authentic standards, well-established analytical methods, and, in some cases, incomplete sample characterization. Non-targeted analyses using high-resolution mass spectrometry (HRMS) can reveal unidentified compounds but remain resource-intensive and often yield only semi-quantitative data. Consequently, both approaches may underestimate the overall burden of micropollutants and DBPs in complex water matrices, particularly in wastewater systems. To address these limitations, this study developed novel analytical methods for quantifying group concentrations of emerging micropollutants, including PFAS, nitro compounds (R–NO2), and nitriles (R–CN). By targeting their shared functional features, such as total fluorine content, anionic acid groups (for PFAS) and nitrogen-containing moieties (–NO2, –CN) in nitro and nitrile compounds, these methods provide inclusive, cost-effective, and robust analytical tools that complement conventional targeted and non-targeted analyses. In particular, the developed approaches for total PFAS quantification can substantially reduce analytical costs and technical barriers, facilitating broader implementation in water quality monitoring and management. The first topic (Chapter 3) presented the development and optimization of innovative, cost-effective methods for quantifying extractable total PFAS in water. The methods integrated persulfate preoxidation (total oxidizable precursor or TOP assay) with solid phase extraction (SPE), followed by either chemical defluorination with sodium biphenyl (SBP assay) or ion-pair formation between anionic PFAS and cationic methylene blue (MB assay). Persulfate preoxidation enhanced analytical selectivity by mineralizing interfering non-PFAS organofluorines and degrading anionic surfactants, while SPE enabled selective PFAS preconcentration and efficiently removed matrix-derived impurities, such as inorganic fluoride (F–) and residual sulfate (SO42–) from persulfate preoxidation and/or water matrices. The SBP and MB assays provide saved-cost approaches with less expertise levels, compared to benchmark methods, such as combustion ion chromatography (CIC). The developed methods demonstrated broad applicability across structurally diverse PFAS, although challenges remain for (ultra)short-chain species. With SPE, the overall methods achieved sub-ppb detection limits, 0.016 μgF/L for SBP and 0.2 μgF/L for MB, while maintaining robust recoveries across multiple PFAS in simulated water samples. Application to real water samples revealed extractable total PFAS concentrations of 5.1 μgF/L in industrial wastewater and 0.30 μgF/L in river water by the SBP assay, consistent with the reference CIC. By contrast, the MB assay produced systematically higher concentrations, 1.9–3.1 times above those obtained by SBP and CIC, indicating its suitability for rapid screening but the need for confirmatory testing with more accurate assays. Importantly, the extractable total PFAS concentrations detected in environmental waters were substantially greater than the sum of individually quantified PFAS by LC–MS, underscoring the presence of uncharacterized PFAS and the need to move beyond compound-by-compound analysis. Together, the SBP and MB assays offer practical, scalable, and cost-efficient tools for monitoring extractable total PFAS in diverse water matrices. In the second topic (Chapter 4), we developed novel methods for quantifying nitro compounds (R–NO2) in water, using a batch UV254 photolysis (15,000 mJ/cm2 fluence at pH 12) coupled with Griess colorimetry for total quantification and a liquid chromatography with post-column photolysis (LC–PCUV) for individual nitro compound analysis. The batch total nitro quantification method demonstrated its optimal conditions at 15,000 mJ/cm2 fluence and pH 12 (10 mM NaOH) sample solution. The LC–PCUV method employed a C18 column for separating nitro compounds, 10,000 mJ/cm2 fluence, and eluents containing pH 8 aqueous phase (10 mM phosphate buffer) with acetonitrile. Photolytic molar nitrite yields revealed clear structural dependence, averaging 68±17% (n = 8) for aliphatic and 11±11% (n = 5) for aromatic nitro compounds, confirming lower sensitivity towards aromatic species. Importantly, the method demonstrated robustness in environmentally relevant matrices, including river water and wastewater, particularly when integrated with solid phase extraction (SPE) to mitigate matrix effects. Detection limits in the low nanomolar range enabled sensitive monitoring under realistic conditions. Application to ozonated water samples showed nitro compound yields of 83±10% and 21±8% from aliphatic amines and amino acids, respectively, consistent with prior literature. Strikingly, more than 20% of amine moieties in natural organic matter (NOM) were converted to nitro compounds during ozonation. Field applications further revealed substantial nitro concentrations (140–180 nM) in wastewater effluents, with levels increasing upon ozonation. Overall, this chapter established simple yet effective tools for investigating the fate of reactive nitrogenous precursors and the formation of nitro compounds, including halonitroalkanes, during oxidative water treatment. The third topic (Chapter 5) focused on the development and application of analytical approaches for quantifying total nitriles (R–CN), an underexplored class of N-DBPs formed during chlorination of nitrogenous precursors. Nitriles encompass a wide structural and physical chemical range, from highly volatile haloacetonitriles (HANs) to less volatile arylalkyl/benzylic and aryl/aromatic nitriles, posing challenges for comprehensive analysis. To address this, complementary preconcentration techniques were employed: SPE for less volatile nitriles and liquid–liquid extraction (LLE) to additionally include nitriles with higher volatility (e.g., HANs). This dual strategy ensured broad analytical coverage and improved sensitivity across diverse nitrile structures. For total quantification, a cytochrome p450 enzyme–based assay was developed to liberate cyanide (CN–) from nitriles, enabling estimation of yields across aliphatic (9.5±4.2%, n = 7), arylalkyl (9.4±1.2%, n = 5), and aryl nitriles (6.5 ± 1.5%, n = 3). Despite overall modest cyanide yields and structural variability, the method provided reproducible conversion factors, revealing potential applicability for analytical method development. The developed total nitrile quantification method showed 15 nM limit of detection (with preconcentration) and favorable recovery rates for synthetic water samples (pure water and tap water spiked with selected nitriles). The method was suggested for application in chlorination of real water samples and natural organic matter (NOM) to explore the formation of a wide spectrum of nitriles and to highlight their relevance as DBPs under environmentally realistic conditions. Taken together, the methods developed in this work overcome the limitations of compound-specific analysis by transforming structurally diverse contaminants into common measurable moieties, which were subsequently quantified using derivatization or colorimetry. These approaches allow for a holistic assessment of contaminant loads across treatment processes, offering new insights into occurrence, transformation, and removal. Ultimately, the outcomes of this work contribute to improving water quality monitoring, optimizing treatment technologies, and ensuring the safety and reliability of drinking water in the context of growing water reuse.