Extracting and Analyzing PFAS from Human Serum

Significance of the Topic

Human exposure to per- and polyfluoroalkyl substances (PFAS) is a major public health concern due to their persistence, bioaccumulation and potential toxicity. Monitoring PFAS in human serum provides critical data on exposure levels and pathways, informing risk assessment and regulatory measures.

Study Objectives and Overview

This work aimed to develop and validate a robust workflow for extracting and quantifying 30 PFAS compounds from human serum. Key goals included optimizing solid phase extraction (SPE) using Oasis WAX µElution plates, achieving interference-free separation via UPLC, and ensuring sensitive detection with tandem quadrupole mass spectrometry.

Methodology and Instrumentation

Sample preparation combined protein precipitation (1:3 serum:acetonitrile) with SPE on Oasis WAX 96-well plates to disrupt PFAS–protein binding and remove matrix components. Chromatographic separation was performed on an ACQUITY UPLC I-Class PLUS system modified with PFAS kit components, using an ACQUITY UPLC HSS T3 column (2.1×100 mm, 1.8 µm) at 35 °C. Mobile phases consisted of water/methanol (95:5) and methanol, both containing 2 mM ammonium acetate. Detection employed a Xevo TQ-S micro mass spectrometer in negative electrospray ionization mode. Data acquisition and quantitation used MassLynx and TargetLynx software.

Key Results and Discussion

Pretreatment with 1:3 acetonitrile minimized PFAS breakthrough during loading. The HSS T3 column resolved steroid sulfate interferences that coeluted on C18 phases, notably improving PFHxS quantitation. Calibration in PFAS-free fetal bovine serum was linear (0.05–20 ng/mL) with R²≥0.996 (6:2 FTS R²=0.992) and residuals ≤15%. Recoveries for 29 compounds ranged 85–120% across six serum lots; FBSA showed lower recovery (~20%) due to its neutral chemistry. Verification with NIST SRM 1957 yielded measured values within ±10% of certified levels and RSD<4%. Analysis of pooled serum lots revealed distinct PFAS concentration profiles and branched-to-linear isomer ratios, suggesting potential for exposure source fingerprinting.

Benefits and Practical Applications

• Reliable biomonitoring workflow for diverse PFAS chemistries
• PFAS kit modifications reduce background contamination
• High-throughput SPE clean-up lowers instrument maintenance
• Accurate and reproducible quantitation across ppt–ppb ranges

Future Trends and Applications

• Extension to emerging and ultra-short-chain PFAS and precursors
• Isomer ratio analysis for source attribution and pathway elucidation
• Integration with high-resolution MS for non-targeted screening
• Automation and miniaturized SPE for large-scale epidemiological studies

Conclusion

The validated SPE-LC-MS/MS method delivers sensitive, accurate measurement of a broad PFAS panel in human serum. Its robustness and adaptability make it suitable for occupational health monitoring, environmental exposure assessment and epidemiological research.

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