Redefining PFAS Workflow Applications: A New Horizon

Setting a new horizon for PFAS workflow applications PFAS, a global concern Per- and polyfluoroalkyl substances (PFAS) are among the primary emerging contaminants of concern. The detection and quantification of known PFAS and the discovery of unknown PFAS substances has never been more important. Determining the best workflow for your PFAS analysis can be challenging. Optimal methods will vary depending on the matrix you are working with and goals of your analysis. There are strategies to help you with either targeted analysis of known PFAS compounds or the discovery of unknowns, from a variety of matrices. 2 3 Perfluorinated Perfluoroalkyl Sulfonamide Amino Carboxylates Perfluoroalkyl Sulfonamido Amines Perfluoroalkyl Acids (PFAAs) Poly fluorinated Perfluoroalkyl Sulfonic Acids (PFSAs) Perfluoroalkyl Carboxylic Acids (PFCAs) Fluorotelomer Sulfonates Fluorotelomer Sulfonamido Betaines Fluorotelomer Alcohols 4 Targeted analysis • Identification and quantitation based on a known candidate list of compounds • Established or regulated methods available • Confirms presence of a compound based on limit of detection Unknown screening • Discover new or unexpected compounds • Detection is independent of a candidate list • Find new compounds of interest in previously run samples by retrospective data analysis PFAS transport through the environment and analysis strategies Household products Cleaning products Personal care products Paints, waxes, polishes Food contact materials Non-stick cookware Microwave popcorn bags Fast food wrappers Manufacturing processes Commercial products Electronics Petrochemicals Fabrics Stain resistant carpets and fabrics Waterproof or resistant clothing Aqueous firefighting foam Airports Military bases PFAS sources PFAS exposure Workflow strategies PFAS PFAS PFAS PFAS PFAS PFAS PFAS Soil Runoff Leachate Animals Produce Fish Humans Plants PFAS PFAS PFAS Sample collection Sample preparation Analysis Data processing The vast uses of PFAS compounds has resulted in a great number of sources for these substances to enter the environment. Common modes of transport are runoff, especially with firefighting foams into water, or leachate from landfills, discharge from manufacturing processes or improper disposal. Once in the ecosystem, plants, animals and humans are all exposed to PFAS. The C-F bonds in the PFAS molecule are very strong, which make metabolism of these compounds very difficult. Therefore, PFAS bioaccumulates over time and causes various health concerns. The workflow for analyzing PFAS will depend on the goals of your analysis. Many labs are doing targeted analysis where a specific list of PFAS molecules are analyzed and quantified. It is estimated there are over 5000 different types of PFAS compounds; some labs will want to discover and identify unknown PFAS through untargeted screening. Water Drinking water Surface water Groundwater Wastewater PFAS Water Drinking water Surface water, groundwater, wastewater Analysis goal Targeted analysis Screening or unknown profiling Targeted analysis Screening or unknown profiling Targeted analysis Screening or unknown profiling Sample prep Solid phase extraction (SPE) Dilution, filtration, acidification Dilution, filtration, acidification or solid phase extraction Accelerated solvent extraction SPE clean-up Analytical method LC-MS/MS: Triple quad mass spectrometry LC-HRAM: High resolution accurate mass LC-MS/MS: Triple quad mass spectrometry LC-HRAM: High resolution accurate mass LC-MS/MS: Triple quad mass spectrometry LC-HRAM: High resolution accurate mass Data acquisition and processing Chromeleon CDS TraceFinder software Compound Discoverer software Soil Careful sampling is important in PFAS analysis. A representative sample must be obtained without introducing any background PFAS into the sample. PFAS can be found in many sources while sampling and in the laboratory. Items to avoid when sampling are waterproof clothing, adhesives, sampling device and materials coming into contact with vehicle upholstery, and common blue ice packs used in shipping. In the lab, avoid items such as aluminum foil, sticky notes and some materials used in the flow path of analytical instruments. It is important to use instruments with minimal fluoropolymers. Polypropylene sampling vessels must be used for water and soil samples as well as the devices used to collect the sample. PFAS workflows will be different based on the matrix you are working with and the goals of your analysis, either targeted analysis or unknown screening. Each matrix uses a different sample preparation method. For water, solid phase extraction (SPE) will be used for drinking water while other water types will use dilution, filtration and/or acidification. For soils, accelerated solvent extraction (ASE) is the optimal method. All the PFAS workflows use liquid chromatography (LC) for separation, but the best mass spectrometer is determined by the analysis type, targeted or unknown screening. The data processing software will depend on the type of workflow. Both Thermo Scientific™ Chromeleon™ Chromatography Data System (CDS) software or Thermo Scientific™ TraceFinder™ software are designed for targeted analysis and quantification. Thermo Scientific™ Compound Discoverer™ software streamlines compound identification and structural elucidation. PFAS sampling PFAS workflow 5 Everyday PFAS analysis in drinking water SPE sample preparation to mass spectrometry analysis Thermo Scientific™ Dionex™ AutoTrace™ 280 PFAS solid-phase extraction instrument • Improves recovery and reproducibility • Significantly reduces risk of background PFAS • Improves lab efficiency The AutoTrace 280 PFAS system ensures inertness and prevents PFAS contamination into samples during extraction, while at the same time delivering consistent and reliable performance. Thermo Scientific™ TSQ™ Fortis triple quadrupole mass spectrometer with the Thermo Scientific™ Vanquish™ UHPLC system fitted with the PFC free kit • Excellent quantitative performance at low dwell times • Enhanced confidence in data and outstanding robustness that prolongs instrument uptime • Simplicity and ease-of-use for users of all expertise levels Established methods: U.S. EPA 537.1 U.S. EPA 533 Orbitrap Exploris 120 Dionex ASE 350 Accelerated Solvent Extractor AutoTrace 280 H 27 x W 23 27.7 H x 21 W Orbitrap Exploris 240 Orbitrap Exploris 120 H 26.75 x W 30 TSQ Altis Triple-Stage Quadrupole TSQ Fortis Triple-Stage Quadrupole Dionex ICS-6000 HPIC system 32.25 H x 21.65 W with Tablet Interface Vanquish Core 32.25 H x 21.65 W Vanquish Core 32.25 H x 21.65 W Vanquish Core AutoTH 27 x Dionex Integrion HPIC System 24.61 H x 11.8 W TSQ Quantis Triple-Stage Quadrupole Thermo Fisher Scientific can supply and support everything you need to detect and quantify PFAS in drinking water. Many established methods, including regulatory, require collection into polypropylene bottles, followed by sample preparation by SPE and analysis using triple quad mass spectrometry. 3.75 5.0 7.5 10.0 12.5 15.0 0 2.5e4 5.0e4 8.0e4 1 2,3 4,5 22 6 7,8 9,10 11,12,13 17,18 15,16 14 25 20,21 19 24 23 Min. Counts Excellent separation of all 18 PFAS compounds and isotropically labeled standards in under 15 minutes. General targeted PFAS analysis workflow in drinking water samples based on either US EPA Method 537.1 or US EPA Method 533. Collect 250 mL sample in a polypropylene bottle and add preservative Extract with AutoTrace 280 PFAS instrument Add internal standards Data analysis with Chromeleon CDS or TraceFinder software Add surrogates or isotope dilution analogues Evaporate to dryness and reconstitute LC-MS/MS analysis using the Vanquish UHPLC system and TSQ Fortis mass spectrometer 01 03 05 04 06 07 02 6 All 18 target compounds spiked at 2 levels, show recoveries well within the method limits of 70–130% and reproducibility is under the required 20%. Preparing PFAS water samples with the AutoTrace 280 PFAS shows negligible background. PFAS blank LC-MS/MS chromatograms -2.0e2 -1.3e2 0.0e0 1.3e2 2.5e2 3.8e2 5.0e2 6.3e2 7.5e2 8.8e2 1.0e3 1.1e3 1.3e3 1.4e3 PFBA PFOA PFMPA PFPeA PFBS PFMBA PFEESA NFDHA 4:2 FTS PFHxA PFPeS HFPO-DA PFHpA PFHxS ADONA 6:2 FTS PFHpS PFNA PFOS 9Cl-PF3ONS 8:2 FTS PFDA PFUnA 11Cl-PF3OUdS PFDoA Counts 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0 16.0 17.0 Min. Peak number Analyte Fortified conc. (ng/L) Mean recovery (%) RSD Fortified conc. (ng/L) Mean recovery (%) RSD 1 PFBS 16.0 107 3.3 80.0 98.3 3.6 2,3* PFHxA 16.0 108 2.3 80.0 106 2.6 4,5* HFPO-DA 16.0 84.1 7.5 80.0 88.6 6.3 6 PFHpA 16.0 113 2.7 80.0 117 1.3 7 PFHxS 16.0 120 3.4 80.0 123 2.1 8 ADONA 16.0 117 2.5 80.0 121 1.1 9,10* PFOA 16.0 113 2.5 80.0 119 1.6 11 PFNA 16.0 114 2.9 80.0 118 2.1 12,13* PFOS 16.0 113 4.5 80.0 117 2.9 14 9Cl-PF3ONS 16.0 96.1 4.1 80.0 103 2.6 15*,16 PFDA 16.0 105 3.2 80.0 111 2.1 17*,18 NMeFOSAA 16.0 103 5.2 80.0 110 5.2 19 PFUnA 16.0 96.8 5.0 80.0 103 3.1 20*21 NEtFOSAA 16.0 100 9.9 80.0 104 2.3 22 11Cl-PF3OUdS 16.0 88.5 5.5 80.0 97.1 4.8 23 PFDoA 16.0 89.8 4.4 80.0 97.3 3.4 24 PFTrA 16.0 89.6 3.8 80.0 95.8 3.7 25 PFTA 16.0 89.0 4.8 80.0 98.1 3.3 * Designates the isotope labeled internal standard or surrogate. PFAS 7 8 Thermo Scientific™ TSQ™ Altis triple quadrupole mass spectrometer with the Vanquish UHPLC system fitted with the PFC free kit • Ultimate sensitivity in matrices ranging from simple to complex • Outstanding instrument robustness enables increased confidence with no loss of instrument uptime • Ultrafast selected reaction monitoring (SRM) increases the number of molecular quantifications in less time 27.7 H x 21 W Orbitrap Exploris 240 Orbitrap Exploris 120 H 26.75 x W 30 TSQ Altis Triple-Stage Quadrupole TSQ Fortis Triple-Stage Quadrupole Dionex ICS-6000 HPIC system 32.25 H x 21.65 W with Tablet Interface Vanquish Core 32.25 H x 21.65 W Vanquish Core Dionex AccelerExtractAutoTrace 280 H 27 x W 23 Dionex Integrion HPIC System 24.61 H x 11.8 W e-Stage 11 Table 5. PFAS recoveries in different water matrices, low and high levels at 60 and 200 ng/L, respectively Compound Recoveries % Reagent water Ground water Surface water Waste water Low level High level Low level High level Low level High level Low level High level PFBA 77% 78% 71% 75% 74% 74% 58% 75% PFPeA 84% 80% 104% 80% 115% 81% 88% 78% PFBS 87% 81% 95% 81% 95% 79% 72% 77% PFHxA 82% 81% 83% 79% 86% 80% 77% 74% 4:2 FTS 81% 82% 90% 78% 87% 79% 76% 91% PFPeS 80% 80% 82% 79% 85% 78% 80% 83% PFHpA 84% 81% 88% 80% 89% 80% 74% 81% PFHxS 81% 81% 87% 78% 94% 81% 85% 85% 6:2 FTS 84% 82% 85% 80% 87% 94% 78% 79% PFOA 83% 80% 88% 82% 123% 83% 83% 86% PFHpS 81% 81% 84% 76% 83% 78% 79% 86% PFNA 79% 81% 84% 80% 86% 80% 79% 82% PFOS 91% 82% 91% 78% 93% 81% 79% 90% 8:2 FTS 85% 80% 81% 75% 76% 79% 78% 83% PFNS 85% 75% 89% 79% 81% 76% 72% 78% PFDA 80% 81% 86% 78% 85% 79% 74% 83% NMeFOSAA 77% 81% 80% 77% 86% 81% 82% 84% PFOSA 76% 76% 87% 75% 91% 75% 79% 81% PFDS 82% 78% 89% 77% 85% 79% 72% 81% PFUnA 76% 76% 80% 81% 75% 78% 75% 83% NEtFOSAA 82% 79% 89% 77% 89% 81% 80% 85% PFDoA 79% 82% 83% 78% 85% 82% 79% 85% PFTriA 87% 86% 89% 79% 92% 91% 87% 89% PFTreA 109% 103% 112% 91% 113% 119% 100% 110% Most recoveries in different water matrices at 60 and 200 ng/L are within the 70% to 130% range. Overlaid chromatograms of all PFAS compounds included in this method. PFOS chromatogram of an injection of 1 ng/L, which is five times lower than the reporting limit of quantitation. Figure 1. Overlaid chromatograms of all PFAS compounds included in this method Excellent quantitative results for PFAS direct analysis in the low ng/L range in non-drinking water matrices using dilution and filtration for sample preparation. Direct inject targeted PFAS methods Established methods: U.S. EPA 8327 ASTM D7979 These alternative methods are sometimes preferred for groundwater, surface water and especially for wastewater. Direct injection methods, like U.S. EPA 8327 or ASTM D7979 are based on sample dilution, filtration and acidification followed by LC-MS/MS analysis. This direct analysis method is for 24 diverse types of PFAS compounds in a wide variety of non-drinking water matrices. PFAS 9 AOF-CIC provides an easy-to-use and economical way to determine if other PFAS molecules might be present in your sample. This technique can help to optimize the utilization of analytical instrumentation by selecting and only analyzing “suspicious” samples. CIC chromatogram of industrial wastewater, diluted 1:10. Total fluorine consists of both inorganic and organic components. The organic part contains compounds that are adsorbable (AOF) and extractable (EOF), both of which have a subset of analytes that are typically targeted by LC-MS/MS analysis. There are also organic compounds that are not captured be either of these approaches. Thermo Scientific™ combustion ion chromatography system • Eliminates complex sample preparation steps using an automated method • Produces fewer environmental contaminants • Highly sensitive • Easy-to-use and saves time 0 2 4 6 8 10 Fluoride 0 5 10 15 t (min) µS/cm Adsorbable organic fluorine (AOF) by combustion ion chromatography Analyzing for adsorbable organically-bound fluorine is used to determine if the mass of fluorine present in the sample exceeds that in the targeted screen. If the total amount is higher, then other PFAS may be present in the sample which were not on the target list. The use of this technique is well documented for the determination of other adsorbable organic halogen-containing components (AOX). Unknown Screening by LC-HRAM MS Could other PFAS be present? Greater mass of fluorine present than accounted for in target list No significant increase in total fluorine, all mass accounted for in targeted screen Discover compounds contributing to total mass of fluorine Quantify the amount of each compound on the target list OF Total Fluorine (TF) Inorganic Fluorine (IF) Organic Fluorine (OF) Adsorbable Organic Fluorine (AOF) Targeted PFAS analysis Extractable Organic Fluorine (EOF) Could other PFAS molecules be present? Automated combustion ion chromatography (CIC) provides a complimentary screening method indicate the possible presence of non-targeted PFAS or other fluorine containing compounds. Targeted screening by LC-MS/MS Fluorine mass balance PFAS Combustion Ion Chromatography System AOF by combustion ion chromatography For complex samples with unknown amounts of other PFASs, utilization of Compound Discoverer software can reduce the data processing time and quickly show results. Workflow example in Compound Discoverer software. Flow-chart style elements can be easily dragged and dropped into place for easy customization. Thermo Scientific™ Orbitrap Exploris™ 120 mass spectrometer • Ultra high-resolution capability • Removes concerns over false positives/negatives • Eliminates matrix interferences • High mass accuracy to generate reliable results • Significantly accelerates the path to confident results for both expert and novice MS users Orbitrap Exploris 120 0 HPIC system face Dionex ASE 350 Accelerated Solvent Extractor AutoTrace 280 H 27 x W 23 A UCMR3 sample shown having trace hits for a non-targeted compound (PFDS). Post-run identification was performed by looking at the isotopic pattern recognition, accurate mass and retention time for confirmation. Note the real-world water sample chromatogram on the left shows the linear (RT 17.5) and branched isomers (RT 17.2) of PFDS, vs the linear PFDS isomer standard on the right. U.S. EPA Method 537 target list PFAS compound Critical Level (ng/L) DL (ng/L) LCMRL (ng/L) PFBS 0.15 0.2 <0.5 PFDA 0.15 0.26 <0.5 PFDoA 0.47 0.73 PFHpA 0.09 0.15 <0.5 PFHxA 0.13 0.19 <0.5 PFHxS 1.7 2.4 PFNA 0.11 0.17 <0.5 PFOA 0.22 0.5 PFOS 0.26 0.5 PFTA 0.15 0.2 <0.5 PFTrDA 0.31 0.55 PFuNA 0.38 1 U.S. EPA Method 537 target list PFAS compound Critical Level (ng/L) DL (ng/L) LCMRL (ng/L) PFBA 0.19 0.64 PFODA 0.55 1 PFDS 0.13 0.19 <0.5 PFHxDA 0.12 0.5 PFPA 0.18 0.19 <0.5 PFAS compounds can be detected and quantitated by using a triple quadrupole mass spectrometer (MS) in targeted analysis. However, the identification and quantitation of unknown PFAS uses LC coupled to high resolution mass spectrometry (HRAM MS). With significant advances in HRAM MS comes can this move to the first column to not only increase the range of potential targets monitored, but to also increase confidence in assignments, access the power of comprehensive tools to identify unknowns and emerging contaminants, and retrospectively analyze data even when the sample no longer exists. Screening for unknown PFAS Routine quantitative workflows and non-targeted analysis can be performed in a single analysis. Input files Mark background compounds Predict compositions Search mass lists Search ChemSpider™ database search Fill gaps Merge features Align retention times Select spectra Group unknown compounds Using a full scan approach, required Detect unknown compounds detection limits or MRLs can be achieved while interrogating for other untargeted PFAS compounds. The compounds highlighted in blue are additional analytes that are not part of the original U.S. EPA Method 537 list but were found in processed drinking water from the same UCMR3 water extracts. 10 Search Thermo Scientific™ mzCloud mass spectral library 2 g of soil extracted using the ASE 350 accelerated solvent extractor SPE clean up of the extract using a styrene divinylbenzene (SDVB) LC-MS/MS analysis using the Vanquish UHPLC system and TSQ Quantis mass spectrometer Data analysis with Chromeleon CDS and TraceFinder software 01 03 04 02 11 Compound Recovery (%) 13C3-PFBS 98 13C3-PFHxS 95 13C8-PFOS 91 13C3-HFPODA 56 2 H3-NMEFOSAA 93 2 H3-NETFOSAA 90 13C8-FOSA 92 13C2 -4:2FTS 110 13C2 -6:2FTS 93 13C2 -8:2FTS 98 Thermo Scientific™ TSQ Quantis™ triple quadrupole mass spectrometer with the Vanquish UHPLC system fitted with the PFC free kit • Superb sensitivity, even in complex matrices • Outstanding robustness; prolonging instrument uptime • Reliability and reproducibility improve data quality for every run and every sample • Simplicity and ease-of-use allow users of all expertise levels to acquire high quality data with improved confidence in results Thermo Scientific™ Dionex™ ASE™ 350 accelerated solvent extractor • Automates extraction for up to 24 samples • Saves solvent—Just 50 mL used for PFAS • Saves time—Only 7 minutes per PFAS extraction H 26.75 x W 30 TSQ Altis Triple-Stage Quadrupole TSQ FoQuadru32.25 H x 21.65 W Vanquish Core 32.25 H x 21.65 W Vanquish Core 32.25 H x 21.65 W Vanquish Core Dionex Integrion HPIC System 24.61 H x 11.8 W TSQ Quantis Triple-Stage Quadrupole 27.7 H x 21 W Orbitrap Exploris 240 Orbitrap Exploris 120 Dionex ICS-6000 HPIC system with Tablet Interface Dionex ASE 350 Accelerated Solvent Extractor AutoTrace 280 H 27 x W 23 100 80 60 40 20 0 Relative abundance 0 2 4 6 8 10 12 14 16 1.38 Time (min) 2.00 2.72 3.16 3.98 4.95 5.76 6.18 4.31 6.35 7.03 8.03 8.73 9.09 10.20 10.69 11.19 11.59 12.46 11.75 12.59 13.71 14.68 15.55 15.98 16.40 15.34 14.49 13.48 12.30 10.91 9.27 7.41 5.40 3.52 2.17 1.49 9.72 C4 acid (PFBA) C14 acid (PFTeDA) PFOA 6.18 8.03 4.31 2.72 9.72 PFOS 11.19 12.46 Acids Sulfonates Fluorotelomer Sulfonamides Chromatogram of various PFAS molecules extracted from soil. Recovery of the isotopically labeled PFAS compounds. Compound Recovery (%) 13C4-PFBA 71 13C5-PFPeA 93 13C5-PFHxA 97 13C4-PFHpA 96 13C8-PFOA 94 13C9-PFNA 104 13C6-PFDA 99 13C7 -PFUdA 95 13C2 -PFDoA 97 13C2 -PFTeDA 108 Data courtesy of Pacific Rim Labs. The need to analyze PFAS in soil samples is also extremely important. Soil becomes contaminated with PFAS through runoff and leachate from landfills and other sources. Any produce or vegetation growing in this soil is at risk for accumulating PFAS. PFAS extraction and analysis in soil Accelerated solvent extraction is an effective method for extracting a wide selection of PFAS, from 4-carbon to 14-carbon fluoroalkyl chain lengths and five different polar head-groups, from soil over a wide range of concentrations. PFAS Find out more at thermofisher.com/pfas-testing For Research Use Only. Not for use in diagnostic procedures. © 2021 Thermo Fisher Scientific Inc. All rights reserved. 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