Detection and quantitation of PFAS in animal tissue using Orbitrap Exploris 120 high-resolution mass spectrometer

Significance of the Topic

The widespread use of per- and polyfluoroalkyl substances (PFAS) across industrial and consumer products has led to persistent environmental contamination and bioaccumulation in food chains. Analytical methods capable of detecting PFAS at ultra-trace levels in complex biological matrices such as animal tissue are essential for food safety monitoring, regulatory compliance, and understanding human exposure pathways.

Study Objectives and Overview

This application note describes the development of a robust liquid chromatography–high resolution accurate mass (LC-HRAM) method using an Orbitrap Exploris 120 mass spectrometer coupled with a Vanquish Flex UHPLC system. The goal was to extract, identify, and quantify 34 target PFAS compounds at low parts-per-trillion (pg/g) levels in pork muscle tissue. Method performance was evaluated against FDA and SANTE guidelines to ensure accuracy, precision, and sensitivity.

Methodology

The sample preparation employed a QuEChERS-based extraction followed by dispersive solid-phase extraction (dSPE) cleanup. Five grams of homogenized pork muscle were spiked with isotopically labeled standards, extracted with acetonitrile/water/formic acid, and cleaned using HyperSep dSPE salts (MgSO₄, PSA, graphitized carbon). A solvent sandwich injection technique on the Vanquish Split Autosampler minimized peak distortion by bracketing sample plugs with aqueous mobile phase. Chromatographic separation used an Accucore C18 analytical column with a saw-tooth gradient to reduce carryover. Full MS with data-independent acquisition (DIA) collected both precursor and fragment ions across the m/z 100–1000 range.

Used Instrumentation

• Thermo Scientific Vanquish Flex Binary UHPLC system with PFAS Analysis Kit (PEEK tubing, PFAS trap column)
• Thermo Scientific Orbitrap Exploris 120 high-resolution mass spectrometer with OptaMax NG source
• HyperSep dSPE Centrifuge and Clean-up Tubes for QuEChERS extraction
• Thermo Scientific Accucore C18 analytical column (100 × 2.1 mm, 2.6 μm)
• TraceFinder software and myLibrary Enterprise for data processing and spectral library management

Results and Discussion

Calibration curves for 34 PFAS compounds showed excellent linearity (r² > 0.995) over 5–5000 pg/mL, with average RSD < 7%. Method limits of quantitation (LOQ) in pork matrix were below 50 pg/g for most analytes. Recovery studies at 25–500 pg/g demonstrated 60–130% recoveries and RSD ≤ 25% for the majority of targets. A few long-chain PFAS exhibited lower recoveries, likely due to sorption on graphitized carbon. Full MS with DIA enabled reliable identification based on precursor and MS² fragment ion matching within 5 ppm, further confirmed by spectral library searches.

Benefits and Practical Applications

• Sub-ppt sensitivity in complex animal tissues without extensive concentration steps
• Broad PFAS coverage with retrospective untargeted analysis potential
• High confidence identification via HRAM and curated spectral libraries
• Automated data processing streamlining routine laboratory workflows

Future Trends and Applications

Advances in HRAM instrumentation and cloud-based spectral library platforms will facilitate non-target screening of emerging PFAS in diverse food and environmental matrices. Integration of ion mobility separation and machine-learning algorithms could further enhance compound identification and reduce interferences. Expanding the certified standard library and applying the method to fish, dairy, and plant tissues will support comprehensive PFAS surveillance programs.

Conclusion

The LC-Orbitrap Exploris 120 method combined with QuEChERS extraction and solvent sandwich injection delivers sensitive, accurate, and high-throughput analysis of multiple PFAS in animal tissue. This approach meets regulatory requirements and offers a scalable platform for monitoring legacy and emerging PFAS in food safety and environmental studies.

Reference

1. Perfluoroalkyl and Polyfluoroalkyl Substances (PFAS); National Institute of Environmental Health Sciences.
2. FDA Method C-010.01 for PFAS in Processed Food by LC-MS/MS.
3. SANTE/12682/2019 Guidelines on Pesticide Analysis.
4. Universal LC-MS method for minimized carryover in bioanalysis.
5. Thermo Fisher AN73186: Custom injection programs for peak optimization.