Conquer the Challenges of Small Molecule Analysis

Importance of the Topic

Per- and polyfluoroalkyl substances (PFAS/PFAA) pose serious environmental and health challenges due to their persistence and bioaccumulation. Developing reliable analytical workflows that combine broad-spectrum profiling with sensitive, targeted quantitation is essential for monitoring drinking water, wastewater and soil. High-resolution mass spectrometry (HRMS) enables non-targeted screening, while triple-quadrupole systems deliver precise quantitation to meet regulatory requirements.

Objectives and Study Overview

This study presents an integrated analytical strategy for the identification and quantification of a panel of PFAS/PFAA compounds. Key aims include:

• Establishing non-targeted profiling and retrospective data mining using Orbitrap HRMS.
• Developing a targeted quantitation method down to single-digit parts-per-trillion levels on TSQ triple-quadrupole platforms.
• Validating performance across multiple water matrices and evaluating method robustness against EPA draft SW-846 Method 8327 guidelines.

Methodology and Instrumentation

Sample Preparation:

• Direct injection of 5 mL water samples spiked with surrogates in methanol.
• Filtration through 0.2 µm GHP membrane syringe filters.
• Analysis in silanized vials to avoid PFAS background.

Non-Targeted Screening:

• Thermo Scientific™ Orbitrap™ Hybrid and Orbitrap ID-X™ Tribrid systems.
• Data acquisition at ultra-high resolution with sub-1 ppm mass accuracy.
• Compound Discoverer™ 3.0 with mzCloud™ library for automated spectral matching and unknown identification.

Targeted Quantitation:

• Thermo Scientific™ TSQ Fortis™, Quantis™ and Altis™ triple-quadrupole mass spectrometers.
• Reverse-phase chromatography with Accucore™ RP-MS columns.
• Calibration over 5–200 ppt, monitoring multiple reaction monitoring (MRM) transitions.

Main Results and Discussion

Non-Targeted Profiling:

• High resolution separated co-eluting isobaric ions and enabled retrospective mining for over 500 compounds.
• Accurate mass assignments reduced false positives and facilitated discovery of unknown PFAS above regulated lists.

Targeted Quantitation:

• All 24 PFAS/PFAA compounds achieved linearity (R² > 0.99) across 5–200 ppt with deviations <20%.
• Lowest limit of quantitation reached 5 ppt, outperforming draft EPA LLOQs by up to fivefold.
• Inter-batch retention time stability exhibited <1% drift; RSDs <20% in ground, reagent, surface and wastewater matrices.
• Long-chain acids (PFTriDA, PFTeDA) showed variable recoveries due to solubility constraints at high spike levels.

Benefits and Practical Applications

This combined workflow delivers comprehensive PFAS coverage with:

• Rapid screening of known and unknown small molecules in complex matrices.
• Robust, high-throughput quantitation meeting regulatory demands for environmental safety.
• Data-rich HRMS archives for retrospective analysis as new contaminants emerge.

Future Trends and Potential Applications

Emerging directions include:

• Expansion of in-silico and spectral libraries for broader non-targeted screening.
• Automation of data analysis pipelines to accelerate unknown identification.
• Miniaturized and field-deployable HRMS platforms for on-site monitoring.
• Integration with ion mobility separation to resolve isobaric PFAS isomers.

Conclusion

The described HRMS screening coupled with TSQ targeted quantitation provides a powerful, sensitive and reliable approach for PFAS monitoring in environmental samples. The workflow meets stringent regulatory guidelines, supports retrospective discovery and ensures consistent performance across diverse water matrices, positioning laboratories to address current and future PFAS challenges effectively.

Used Instrumentation

• Thermo Scientific™ Orbitrap™ Hybrid and Orbitrap ID-X™ Tribrid Mass Spectrometers
• Thermo Scientific™ TSQ Fortis™, Quantis™ and Altis™ Triple-Quadrupole MS
• Thermo Scientific™ Vanquish™ Flex Binary UHPLC System
• Thermo Scientific™ Compound Discoverer™ 3.0 Software with mzCloud™ Library