Analysis of Per- and Polyfluoroalkyl Substances (PFAS) in Aqueous Samples Per EPA Draft Method 1633

Importance of the Topic

Per- and polyfluoroalkyl substances (PFAS) are widely used industrial chemicals that resist environmental degradation and accumulate in water resources. Reliable, sensitive, and reproducible analytical methods are essential for monitoring PFAS in drinking water, wastewater, and other aqueous matrices to meet tightening regulatory requirements and protect public health.

Objectives and Study Overview

This study evaluates the United States EPA draft Method 1633 for quantifying 40 PFAS in aqueous samples. The goals were to apply an optimized solid-phase extraction (SPE) workflow, coupled with LC/MS/MS analysis on an Agilent 1290 Infinity II HPLC and 6470 triple quadrupole mass spectrometer, and to verify method performance in terms of extraction recovery, linearity, sensitivity (method detection limits, MDLs), precision, and applicability to real wastewater effluent.

Methodology

• Sample Preparation and SPE

• Volume: 500 mL unpreserved reagent water or effluent in PFAS-certified HDPE bottles.
• Spiking: Addition of extracted internal standards (EIS) and adjustment of pH to 6.0–7.0.
• SPE Cartridges: 150 mg weak anion exchange (WAX) cartridges with silanized glass wool.
• Conditioning: 15 mL 1% methanolic ammonium hydroxide, 5 mL 0.3 M formic acid.
• Loading: 5 mL/min flow, rinse with water and 1:1 formic acid/methanol, dry under vacuum.
• Elution and Cleanup: Elution with 1% methanolic ammonium hydroxide, acidification with acetic acid, carbon cleanup, centrifugation, and nylon syringe filtration into tubes containing nonextracted internal standards (NIS).

• LC/MS/MS Analysis

• Chromatography: Agilent ZORBAX Eclipse Plus C18 guard and analytical columns, PFC delay column, 40 °C, 0.4 mL/min gradient from 2% to 95% acetonitrile with 2 mM ammonium acetate.
• Mass Spectrometry: Agilent 6470B triple quadrupole with Jet Stream ESI in negative mode using dynamic multiple reaction monitoring (dMRM).
• Software: Agilent MassHunter Acquisition and Quantitative Analysis.

Used Instrumentation

• Agilent 1290 Infinity II LC System (High Speed Pump, Multisampler, Multicolumn Thermostat)
• Agilent ZORBAX Eclipse Plus C18 guard (2.1×5 mm) and analytical (2.1×100 mm) columns
• Agilent InfinityLab PFC Delay Column (4.6×30 mm)
• Agilent 6470B Triple Quadrupole MS with Jet Stream ESI source
• Agilent MassHunter Acquisition and Quantitative Analysis Software

Main Results and Discussion

• Calibration and Linearity: Eight-point quadratic calibration (1/x weighting) achieved R2 > 0.998 and relative standard errors < 10% for all PFAS. Retention time RSDs were < 3%.

• Sensitivity: Initial MDLs for 32 compounds were < 1 ng/L, with the highest MDL (7:3 FTCA) at 3.12 ng/L—meeting or exceeding draft method criteria.

• Recoveries and Precision: Reagent water spikes (n = 23) yielded surrogate recoveries of 66–122% (RSDs < 15%), with 22 surrogates between 70–130%. Mid-level precision and accuracy in reagent water showed RSDs < 9% and recoveries of 75–125%.

• Wastewater Effluent: Four replicate effluent samples spiked with all analytes produced recoveries of 64–159% (accuracy) and RSDs of 3–12%. Background PFAS levels were subtracted prior to recovery calculations.

• Chromatographic Separation: The required ≥ 1 min separation between bile salts (TDCA) and PFOS was consistently achieved. The MRM chromatogram demonstrated clear resolution of all target analytes at low-level calibrator concentrations.

Benefits and Practical Applications

This work demonstrates that EPA draft Method 1633 can be implemented with high sensitivity, robust precision, and excellent linearity using standard Agilent SPE and LC/MS/MS workflows. The method supports high-throughput PFAS monitoring in environmental and wastewater laboratories, facilitating compliance with emerging regulatory guidelines and enhancing water quality assurance.

Future Trends and Potential Applications

• Expanded Analyte Lists: Incorporation of newly identified PFAS and transformation products.
• High-Resolution Techniques: Coupling with time-of-flight or Orbitrap instruments for non-target screening.
• Automation: Further streamlining of SPE and sample processing to increase throughput.
• Multi-Laboratory Validation: Collaborative studies to finalize Method 1633 and establish interlaboratory reproducibility.

Conclusion

The optimized SPE and LC/MS/MS approach on the Agilent 1290 Infinity II and 6470 TQ system meets or exceeds EPA draft Method 1633 performance requirements. It delivers low detection limits, reliable recoveries, and reproducible quantitation of 40 PFAS in aqueous matrices, offering a robust tool for routine environmental monitoring.

References

• United States Environmental Protection Agency. Draft Method 1633: Analysis of Per- and Polyfluoroalkyl Substances (PFAS) in Aqueous, Solid, Biosolids, and Tissue Samples by LC-MS/MS; Office of Water; August 2021.
• PFAS Water Testing. Get Ahead. Stay Ahead. Start-to-Finish Workflows for PFAS Analysis; Agilent Technologies, Inc.; Retrieved from Agilent website, 2022.