Earth Day 2026: Enhancing Analytical Methods for Detecting Organofluorines and PFAS in Municipal Wastewater

Traditional methods for assessing extractable organofluorine (EOF) in wastewater frequently underestimate the total fluorine mass, often missing trace compounds like over-the-counter fluorinated pharmaceuticals. While advanced filtering techniques have been used in the past to successfully capture a wider variety of chemicals,^1-3 they have yet to be applied to EOF testing methods. To overcome this challenge, a research team from the Man-Technology-Environment (MTM) Research Centre with the School of Science and Technology of Örebro University (Sweden) has developed an innovative analytical workflow combining multisorbent solid-phase extraction (SPE) with high-resolution mass spectrometry (HRMS). Through this advanced, highly inclusive approach, their goal was to accurately capture, trace, and quantify the diverse range of fluorinated compounds—from low-fluorinated pharmaceuticals to highly fluorinated conventional PFAS—entering and exiting municipal wastewater treatment plants.

LCGC International spoke to Leo W. Y. Yeung, Pontus Larsson, and Anna Kärrman, authors of the corresponding paper from this research,⁴ about the group’s findings.

Why is liquid chromatography–tandem mass spectrometry (LC-MS/MS) commonly used for the targeted analysis of PFAS in environmental samples such as wastewater?

LC-MS/MS is widely used because it offers high sensitivity, selectivity, and reliable quantification for trace contaminants in complex environmental samples. PFAS are often present at very low concentrations (ng/L or lower) in wastewater, and LC-MS/MS allows these compounds to be separated and detected accurately despite the complex sample matrix.

Much of the PFAS research has focused on perfluoroalkyl acids (PFAAs), including compounds such as PFOS and PFOA. Importantly, most PFAAs are strong acids that exist predominantly in their ionized form in aqueous solutions, which makes them particularly well suited for analysis by LC-MS/MS using electrospray ionization (ESI). Their pre-existing charge enables efficient ionization and therefore very sensitive detection, making LC-MS/MS a powerful technique for monitoring PFAS in environmental samples.

How does solid phase extraction (SPE) improve chromatographic analysis of PFAS in wastewater samples before LC-MS/MS detection?

SPE serves as an important sample preparation and pre-concentration step prior to chromatographic analysis. Municipal wastewater contains a complex mixture of dissolved organic matter, inorganic ions, and other contaminants that can interfere with analytical measurements. PFAS are typically present at very low concentrations (often in the ng/L range) which are much lower than many co-occurring substances in wastewater. As a result, these matrix components can affect both chromatographic separation and ionization in the mass spectrometer. SPE helps address these challenges by concentrating trace PFAS from large sample volumes while simultaneously removing matrix interferences that may suppress ionization. By improving analyte enrichment and reducing background interference, SPE enhances detection limits and increases the overall reliability and accuracy of PFAS measurements.

What is the principle behind using hydrophilic–lipophilic balance (HLB) and weak anion exchange (WAX) sorbents in SPE for PFAS extraction?

HLB and WAX sorbents are commonly used in solid phase extraction because they provide complementary retention mechanisms. HLB sorbents retain compounds through a balance of hydrophobic and hydrophilic interactions, which makes them suitable for extracting a broad range of neutral and moderately polar compounds. In contrast, WAX sorbents combine hydrophobic interactions with positively charged functional groups that selectively bind anionic species through ion-exchange interactions. This is particularly useful for capturing many conventional PFAS, such as perfluoroalkyl acids, which are negatively charged under typical extraction conditions. By combining these different interaction mechanisms, HLB and WAX sorbents help capture PFAS compounds that vary widely in polarity and functional groups.

Why might HLB or WAX sorbents fail to capture the full spectrum of organofluorine compounds present in wastewater samples?

Single-sorbent extraction approaches can be limited because they are selective toward certain chemical classes. For example, WAX sorbents primarily retain anionic PFAS but may not efficiently capture cationic or neutral compounds. Conversely, HLB sorbents are designed to extract neutral and moderately polar molecules, but they may perform poorly for very polar or strongly ionic species. Because municipal wastewater contains a wide diversity of fluorinated substances, including pharmaceuticals, metabolites, and inorganic fluorinated compounds, relying on a single sorbent may not capture the full spectrum of organofluorine compounds present in the sample.

How does multisorbent solid phase extraction (SPE) improve the chromatographic recovery and retention of a wider range of fluorinated compounds compared with single-sorbent SPE methods?

SPE combines several sorbent types in a single cartridge, typically including strong cation exchange (MCX), WAX, and HLB materials. This layered configuration allows the extraction method to retain anionic, cationic, and neutral compounds simultaneously, thereby broadening the range of fluorinated chemicals that can be captured from complex wastewater samples. HLB sorbents provide retention through balanced hydrophobic and hydrophilic interactions for neutral compounds, while WAX sorbents selectively bind anionic species such as many conventional PFAS through ion-exchange interactions. MCX sorbents, in contrast, contain negatively charged functional groups that retain positively charged compounds via strong cation-exchange interactions. This property complements WAX by enabling the extraction of cationic fluorinated compounds, including certain pharmaceuticals and their metabolites, which may not be efficiently recovered using anion-exchange sorbents alone. In our study, the multisorbent approach improved the recovery of a broader range of fluorinated substances, particularly cationic pharmaceuticals that were poorly retained when using conventional single-sorbent WAX extraction methods.

What role does high-resolution mass spectrometry (HRMS) play in suspect screening and nontargeted chromatographic workflows for PFAS detection?

HRMS plays a critical role in suspect screening and non-target workflows by enabling the detection of PFAS and other fluorinated compounds that are not included in targeted analyses. In suspect screening, HRMS data are compared against curated suspect lists compiled from sources such as literature reports, chemical databases, known-use compounds or via in-silico biotransformation prediction tools. Candidate compounds are first screened based on their accurate mass and molecular formula, and then further evaluated by examining their molecular structure and fragmentation patterns in the MS/MS spectra. Because HRMS provides high mass accuracy and detailed fragmentation information, it increases confidence in the tentative identification of compounds, even when authentic reference standards are not available. This approach allows researchers to move beyond predefined target lists and investigate previously unmonitored fluorinated substances, including pharmaceuticals, metabolites, and transformation products that may contribute to the overall organofluorine burden in wastewater.

How is fluorine mass balance analysis performed by combining combustion ion chromatography (CIC) with targeted LC-MS/MS methods?

Fluorine mass balance analysis combines targeted compound quantification with measurements of total fluorine in a sample. In this approach, LC-MS/MS is used to quantify known fluorinated compounds, such as PFAS and fluorinated pharmaceuticals, while CIC is used to measure the total amount of fluorine present in the extracted sample. Because the amount and types of organofluorine compounds that are measured depend on the extraction method used, the parameter is commonly referred to as extractable organofluorine (EOF) rather than total organofluorine. After extraction, the concentrations of identified fluorinated compounds are converted into fluorine equivalents and compared with the EOF value measured by CIC. This comparison allows researchers to determine what fraction of the extractable fluorine can be explained by known compounds and to estimate the portion that remains unidentified in the sample.

Why is chromatographic separation important before mass spectrometric detection when analyzing complex environmental matrices like municipal wastewater?

Municipal wastewater is an extremely complex chemical mixture containing thousands of organic compounds. Chromatographic separation plays an important role by resolving compounds before they enter the mass spectrometer, helping to distinguish substances that may have similar masses and preventing co-elution of analytes with other compounds in the sample. This separation also helps reduce matrix effects during ionization, which can otherwise suppress or enhance signals and affect measurement accuracy. By separating analytes from interfering substances, chromatography improves both the identification and quantification of target compounds. Without adequate chromatographic separation, co-elution and overlapping signals in the mass spectrometer could occur, making reliable detection and accurate analysis much more difficult.

How can chromatographic techniques help differentiate between conventional PFAS (such as perfluoroalkyl acids) and fluorinated pharmaceuticals in wastewater samples?

Conventional PFAS and fluorinated pharmaceuticals differ in their chemical structures, polarity, and functional groups. Chromatographic systems, when combined with MS/MS analysis, allow these compound classes to be separated and identified based on differences in retention behavior, unique fragmentation patterns, and distinct mass-to-charge ratios. For example, perfluoroalkyl carboxylic acids (PFCAs) often produce characteristic fragments such as m/z 169 (C₃F₇⁻) in MS/MS analysis, while perfluoroalkyl sulfonic acids (PFSAs) like PFOS generate diagnostic ions including m/z 99 (FSO₃⁻) and m/z 80 (SO₃⁻). These characteristic fragments help confirm the identity of conventional PFAS and distinguish them from other fluorinated compounds. In contrast, fluorinated pharmaceuticals typically produce different fragmentation patterns due to their more complex molecular structures. Together with chromatographic separation, these features enable researchers to differentiate highly fluorinated PFAS from low-fluorinated pharmaceuticals and their metabolites, even when both groups of compounds are present simultaneously in complex matrices such as municipal wastewater.

What are the analytical advantages of integrating multisorbent SPE, LC-MS/MS, HRMS screening, and CIC in a single workflow for assessing organofluorine contamination in wastewater?

Integrating these analytical tools provides a comprehensive strategy for assessing organofluorine contamination in complex environmental samples such as wastewater. Multisorbent solid phase extraction (SPE) enables the capture of a broader range of fluorinated compounds by retaining anionic, cationic, and neutral species. Targeted LC-MS/MS analysis then allows accurate quantification of known compounds, while high-resolution mass spectrometry (HRMS) screening helps identify previously unmonitored or unknown fluorinated substances. In addition, combustion ion chromatography (CIC) measures the total amount of extractable organofluorine (EOF) in the sample. Together, this integrated workflow enables both detailed chemical identification and an overall fluorine mass balance, helping researchers better understand the sources and composition of fluorinated contaminants in wastewater.

References

1. Kern, S.; Fenner, K.; Singer, H. P. et al. Identification of Transformation Products of Organic Contaminants in Natural Waters by Computer-Aided Prediction and High-Resolution Mass Spectrometry. Environ. Sci. Technol. 2009, 43 (18), 7039– 7046. DOI: 10.1021/es901979h
2. Gago-Ferrero, P.; Schymanski, E. L.; Bletsou, A. A. et al. Extended Suspect and Non-Target Strategies to Characterize Emerging Polar Organic Contaminants in Raw Wastewater with LC-HRMS/MS. Environ. Sci. Technol. 2015, 49 (20), 12333– 12341. DOI: 10.1021/acs.est.5b03454
3. Köke, N.; Zahn, D.; Knepper, T. P. et al. Multi-Layer Solid-Phase Extraction and Evaporation─Enrichment Methods for Polar Organic Chemicals from Aqueous Matrices. Anal. Bioanal. Chem. 2018, 410 (9), 2403– 2411. DOI: 10.1007/s00216-018-0921-1 
4. Larsson, P.; Kärrman, A.; Yeung, L. W. Y. Elucidating Unknown Organofluorine in Municipal Wastewater: A Mass Balance Approach including Fluorinated Pharmaceuticals. Environ Sci Technol. 2026, 60 (8), 6623-6634. DOI: 10.1021/acs.est.5c13161