Development and Performance Evaluation of a Method for the Analysis of Per- and Polyfluoroalkyl Substances (PFAS) in Foods of Animal Origin

Importance of the Topic

Per- and polyfluoroalkyl substances (PFAS) have emerged as persistent environmental pollutants with the ability to accumulate in animal tissues and enter the human food chain. Elevated awareness of PFAS presence in fish, meat, offal, and eggs has driven the need for reliable analytical methods capable of quantifying these compounds at sub-ng/g levels to protect public health and meet evolving regulatory limits.

Objectives and Study Overview

The study aimed to develop and validate a streamlined workflow for the extraction, cleanup, and quantification of 30 PFAS compounds in complex food matrices of animal origin. Specific goals included:

• Designing an alkaline methanol digestion coupled with solid-phase extraction (SPE) for efficient isolation of PFAS.
• Implementing LC-MS/MS analysis for sensitive and selective detection.
• Assessing method performance across six commodity types (salmon, tilapia, ground beef, beef liver, beef kidney, chicken eggs).
• Conducting an interlaboratory study on four key PFAS (PFOA, PFNA, PFHxS, PFOS) in fish tissues to evaluate reproducibility.

Applied Methodology and Instrumentation

An overview of the sample preparation and analytical workflow is as follows:

• Sample Homogenization: Frozen tissues and eggs were blended and subsampled.
• Extraction: 2 g homogenate spiked with isotope-labeled surrogates was treated with 0.02 M NaOH in methanol, shaken, and centrifuged.
• SPE Cleanup: Diluted extract was loaded onto Oasis WAX cartridges preconditioned with ammonia, methanol, and water. Wash steps removed matrix interferences, then PFAS were eluted with 2 % ammonium hydroxide in methanol.
• Concentration: Eluate was evaporated and reconstituted in 1:1 methanol–water containing internal standard.
• LC-MS/MS Analysis: Separation was achieved on a BEH C18 column (2.1×100 mm, 1.7 µm) with a 22-min gradient at 0.3 mL/min. Mobile phases were water and methanol, each with 2 mM ammonium acetate. Detection utilized electrospray ionization in multiple-reaction monitoring mode on a Xevo TQ-XS mass spectrometer.

Main Results and Discussion

• Recoveries for most PFAS ranged between 40 and 120 % with relative standard deviations ≤22 % at 1 ng/g, meeting FDA guidelines.
• Neutral sulfonamides exhibited low recovery due to loss during methanol washes, suggesting need for alternative sorbents for these analytes.
• Long-chain carboxylates showed reduced recoveries in certain matrices (eggs, fish), pointing to matrix-dependent extraction challenges.
• Limits of quantitation were generally below 0.1 ng/g, enabling detection well under EFSA dietary exposure thresholds.
• Interlaboratory study across seven labs yielded trueness between 102 and 121 %, repeatability <20 %, and reproducibility ≤30 % for PFOA, PFNA, PFHxS, PFOS in fish QC material.

Benefits and Practical Applications

The proposed method offers:

• A unified protocol for a wide class of PFAS in diverse animal-derived foods.
• Sensitivity to meet stringent regulatory requirements for dietary PFAS levels.
• Demonstrated robustness and reproducibility suitable for routine compliance testing in quality control and research laboratories.

Future Trends and Applications

• Integration of alternative SPE media (e.g., HLB sorbents) to improve recovery of neutral PFAS subclasses.
• Expansion of target analyte lists to include emerging ultra-short and ultra-long PFAS variants.
• Adoption of high-throughput automation for large-scale monitoring programs.
• Coupling with high-resolution mass spectrometry for non‐targeted screening of novel PFAS.
• Alignment with evolving global regulations and dietary exposure models.

Conclusion

The developed alkaline methanol extraction followed by WAX SPE and UPLC-MS/MS analysis provides a sensitive, accurate, and reproducible approach for quantifying a broad range of PFAS in foods of animal origin. The method meets international recovery and precision criteria, making it suitable for regulatory compliance and dietary exposure assessment.

Reference

Fujimoto G., Organtini K., Adams S., Hird S. Development and Performance Evaluation of a Method for the Analysis of Per- and Polyfluoroalkyl Substances (PFAS) in Foods of Animal Origin. Waters Corporation, 2023.