PFAS Testing Methods and Guidance for Sample Filtration

• What are Poly- and Perfluorylalkyl Substances (PFAS)?
• PFAS and the Regulatory Landscape
• Analytical Methods for Determining PFAS in Water Samples
• Filtration Media and PFAS Contamination
• Testing Millex^® Filtration Devices in PFAS Workflows
• Sample Preparation in PFAS Testing
• Products and Catalog Numbers
• References

What are poly- and perfluorylalkyl substances (PFAS)?

PFAS are poly- and perfluorylalkyl substances known as “forever chemicals” and comprise a group of over 4,000 varieties of long- and short-chain perfluorinated compounds.¹ PFAS are used in a variety of industries for their excellent oil-, water-, temperature-, chemical-, and fire-resistant properties and noted for their use in polymerization reactions of fluoropolymers such as Teflon® by companies such as 3M and Dupont. Products that contain PFAS and related compounds are ubiquitous in industrial and consumer products, including product packaging, cosmetics, non-stick cookware, stain repellents, polishes, paints, coatings, and firefighting foams.

The excellent properties and broad use of PFAS have led to the persistent accumulation of these man-made chemicals in environmental and biological matrices, recently linked to liver damage, cancer, weakened immune system, and high cholesterol in humans.^1-3

PFAS and the Regulatory Landscape

In response, agencies in the US and Europe have taken regulatory action. The Stockholm Convention proposed regulations for two of the most common PFAS compounds — perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) — with certain exemptions, effective in 2020. The US Environmental Protection Agency (EPA) published an Action Plan in 2019 followed by recommendations for testing water matrices for PFAS compounds under the Safe Drinking Water Act in early 2020, with a drinking water advisory concentration of 70 parts per trillion (ppt). In October 2021, the EPA published its PFAS Strategic Roadmap, which outlines their extensive approach to addressing PFAS from 2021 through 2024. Most recently, the EPA released drinking water advisories for four PFAS compounds (PFOA, PFOS, Hexafluoropropylene Oxide (HFPO) dimer acid and its ammonium salt, and PFBS and its potassium salt). The European Union (EU) drinking water directive, which includes a limit of 0.5 µg/L for all PFAS, took effect in January 2021. Additionally, the European Chemicals Agency (ECHA) submitted a restriction proposal in January 2022 for PFAS in firefighting foams, with several other proposals expected through 2023. Additional PFAS substances are on the list for evaluation under REACH (Registration, Evaluation, Authorisation and Restriction of Chemicals). In response to rapidly evolving regulatory proposals and actions, academic and industrial testing labs have developed analytical methods for testing and monitoring PFAS in a variety of matrices. These regulations are important to understanding the extent of human exposure and environmental contamination to inform future remediation efforts.

Analytical Methods for Determining PFAS in Water Samples

Currently, most analytical methods for water matrices involve solid phase extraction (SPE) sample preparation after the addition of internal standards and fortification of samples with surrogate standards, prior to liquid chromatography and tandem mass spectrometry (LS/MS/MS) for the detection of selected per- and polyfluorinated alkyl substances (usually 20-30 compounds).

Drinking water is considered a “clean” matrix and often does not require filtration ahead of sample preparation. However, methods such as SW-846 Method 8327, ASTM D7968-17a, ASTM D7979-20, and ISO 21675 involve matrices such as wastewater that could have a higher degree of particulates. One of the more recent methods, EPA Draft Method 1633, is undergoing multi-lab validation for the analysis of PFAS in a variety of high-particulate matrices including tissue samples. Particulates in solution must be removed prior to LC/MS/MS as they can be detrimental to sample analysis, column longevity, and overall instrument function. These methods identify the need for filtration using membranes in a syringe filter format.

Filtration Media and PFAS Contamination

In recent studies, there has been concern about adding PFAS contamination to samples from a variety of sources including collection bottles, storage vials, tubing components, and any other plastic that contacts the sample. This includes membrane filters and filter housing; for example, adsorption of PFAS onto filtration media is dependent on filter type and type of PFAS molecule.^4-5 Further, some filters may show trace amounts of contamination that may interfere with LC-MS/MS detection.⁶ In some cases, both contamination, and sorption onto filtration media can be reduced by washing with methanol.^6-7

Testing Millex^® Filtration Devices in PFAS Workflows

Materials and Methods

To alleviate this concern, three lots of polyethersulfone (PES), three lots of nylon, and two lots of nylon-HPF (nylon membrane with a glass fiber prefilter for high-particulate filtration) Millex® syringe filter devices were tested for PFAS extractables according to EPA Method 537.1. Some PFAS compounds not required by the method, including next-generation PFAS compounds and fluorotelomer sulfonates, were also included. The compounds tested are listed in Table 1. EPA 537.1 does not require sample filtration, but filtration was used to provide a clean sample to test extractable contamination levels from the syringe filter. A comparison of those compounds tested to the compounds required by ASTM D7979-19, SW-846 Method 8327, ISO 21675, and EPA Draft Method 1633 are also included in Table 1.

Table 1. Table 1. A comparison of the PFAS compounds used in the current study compared with EPA 537.1, ASTM D7979-19, SW-846 Method 8327, ISO 21675, and new method EPA Draft Method 1633 (high-particulate matrices)

^aSample filtration required and/or suggested by the method

Compound	Abbreviation	This Study	EPA 537.1	ASTM D7979-19a	SW-846 Method 8327a	ISO 21675a	EPA Draft 1633a
Perfluoro-n-butanoic acid	PFBA	x		x	x	x	x
Perfluoro-n-pentanoic acid	PFPeA	x		x	x	x	x
Perfluoro-n-hexanoic acid	PFHxA	x	x	x	x	x	x
Perfluoro-n-heptanoic acid	PFHpA	x	x	x	x	x	x
Perfluoro-n-octanoic acid	PFOA	x	x	x	x	x	x
Perfluoro-n-nonanoic acid	PFNA	x	x	x	x	x	x
Perfluoro-n-decanoic acid	PFDA	x	x	x	x	x	x
Perfluoro-n-undecanoic acid	PFUnDA	x	x	x	x	x	x
Perfluoro-n-dodecanoic acid	PFDoDA	x	x	x	x	x	x
Perfluoro-n-tridecanoic acid	PFTrDA	x	x	x	x	x	x
Perfluoro-n-tetradecanoic acid	PFTeDA	x	x	x	x	x	x
Perfluoro-n-hexadecanoic acid	PFHxDA	Not tested				x
Perfluoro-n-octadecanoic acid	PFOcDA / PFODA	Not tested				x
Perfluoro-n-butanesulfonic acid	PFBS	x	x	x	x	x	x
Perfluoro-n-pentanesulfonic acid	PFPeS	x			x		x
Perfluoro-n-hexanesulfonic acid	PFHxS	x	x	x	x	x	x
Perfluoro-n-heptanesulfonic acid	PFHpS	x			x	x	x
Perfluoro-n-octanesulfonic acid	PFOS	x	x	x	x	x	x
Perfluoro-n-nonanesulfonic acid	PFNS	x			x		x
Perfluoro-n-decanesulfonic acid	PFDS	x			x	x	x
Perfluoro-n-dodecanesulfonic acid	PFDoS	Not tested					x
n-Decafluoro-4-(pentafluoroethyl) cyclohexanesulfonate	PFecHS	Not tested		x
4:2 Fluorotelomer sulfonic acid	4:2 FTS 4:2 FTSA	x			x		x
6:2 Fluorotelomer sulfonic acid	6:2 FTS 6:2 FTSA	x			x	x	x
8:2 Fluorotelomer sulfonic acid	8:2 FTS 8:2 FTSA	x			x	x	x
8:2 Fluorotelomer unsaturated carboxylic acid	8:2 FTUCa/ 8:2 FOUEA	Not tested				x
3-Perfluoropropyl propanoic acid	3:3 FTCA	Not tested					x
2H,2H,3H,3H-Perfluorooctanoic acid	5:3 FTCA	Not tested					x
8:2 Polyfluoroalkyl phosphate diester	8:2 diPAP	Not tested				x
Perfluorooctanesulfonamide	PFOSA/FOSA	x			x	x	x
N-methyl-perfluorooctanesulfonamide	N-MeFOSA	Not tested				x	x
N-ethyl-perfluorooctanesulfonamide	N-EtFOSA	Not tested				x	x
N-methyl Perfluorooctanesulfonamidoacetic acid	N-MeFOSAA	x			x	x	x
N-ethyl Perfluorooctanesulfonamidoacetic acid	N-EtFOSAA	x			x	x	x
N-methyl perfluorooctanesulfonamidoethanol	N-MeFOSE	Not tested					x
N-ethyl perfluorooctanesulfonoamidoethanol	N-EtFOSE	Not tested					x
n-2-Perfluorohexyl ethanoic acid	FHEA / 6:2 FHEA / 6:2 FTCA	Not tested		x
n-2-Perfluorodecyl ethanoic acid	FDEA / 10:2 FDEA	Not tested		x
n-2H-Perfluoro-2-decenoic acid	FOUEA / 8:2 FTUCA	Not tested		x
n-3-Perfluoroheptyl propanoic acid / 3-Perfluoroheptyl propanoic acid	FHpPA /7:3 FTCA	Not tested		x			x
n-2H-Perfluoro-2-octenoic acid	FHUEA / 6:2 FHUEA	Not tested		x
Hexafluoropropylene oxide dimer acid	GenX HFPO-DA	x				x	x
4,8-Dioxa-3H-perfluorononanoic acid	ADONA DONA	x				x	x
Perfluoro-3-methoxypropanoic acid	PFMPA	Not tested					x
Perfluoro-4methoxybutanoic acid	PFMBA	Not tested					x
Nonafluoro-3,6-dioxaheptanoic acid	NFDHA	Not tested					x
9-Chlorohexadecafluoro-3-oxanone-1-sulfonic acid	9Cl-PF3ONS F-53B Major	x				x	x
11-Chloroeicosafluoro-3-oxaundecane-1-sulfonic acid	11Cl-PF3OUdS F-53B Minor	x					x
Perfluoro(2-ethoxyethane)sulfonic acid	PFEESA	Not tested					x

EPA 537.1 was used to test for PFAS contamination of filtration media in collaboration with SGS North America (Orlando, FL location). Briefly, a 250 mL PFAS-free DI water sample was spiked with surrogates. The internal standard spike of 0.08 ppb was used for QC blanks. To determine if sample filtration media contributes to PFAS contamination, the entire sample was passed through the filter and into a styrene divinylbenzene (SDVB) SPE cartridge. The sample bottles and tubes were rinsed with basic methanol, passed through the filter and into the cartridge. The entire sample was then subjected to SPE and concentrated to 1 mL in 96:4% (v/v) methanol:water prior to LC/MS/MS analysis using a C18 column. Analysis was performed using internal standards. The filters tested included three lots of Millex®-GP syringe filters (non-sterile, 33 mm filter with PES membrane) in both 0.22 µm (Cat. No. SLGP033N) and 0.45 µm (Cat. No. SLHP033N) pore sizes; three lots of Millex® nylon syringe filters (non-sterile, 33 mm filter with nylon membrane) in a 0.20 µm pore size (Cat. No. SLGN033); and two lots of Millex® nylon-HPF syringe filters (non-sterile, 25 mm filter with nylon membrane and glass fiber prefilter) in a 0.20 µm pore size (Cat. No. SLGNM25).

PFAS Testing Results

There were no detectable PFAS contaminants in any of the Millex® devices tested, even given the very low minimum detection limits (MDL) of the method (Table 2). The results were the same for the three different lots of the 0.22 and 0.45 µm PES membranes, three different lots of 0.20 µm nylon membranes, and two different lots of 0.20 µm nylon-HPF membranes. These results suggest that PES, nylon and nylon-HPF Millex® filtration devices are reliable and appropriate to utilize in the filtration of samples for the analysis of PFAS compounds in environmental matrices. In this study, consumables were rinsed with basic methanol. In the case of nylon membranes, non-specific adsorption of internally spiked standards and sample can be reduced using a methanol rinse⁶. Similar results were found using PES membranes alone (without housing and filtered using a 25 mm Swinnex® filter holder device).

Table 2. Table 2. Detection of PFAS after filtration with Millex® PES and nylon devices using LC/MS/MS according to EPA 537.1

Abbreviations: RL = reporting limit (ppb); MDL = minimum detection limit (ppb); ND = not detected below both RL and MDL; HPF = high particulate filter

Compound​

Abbreviation​

RL​ (ppb)

MDL​ (ppb)

SLGP033 0.2 µm PES

SLHP033 0.45 µm PES

SLGN033 0.2 µm Nylon

SLGNM25 0.2 µm Nylon-HPF

Lot1

Lot2

Lot3

Lot1

Lot2

Lot3

Lot1

Lot2

Lot3

Lot1

Lot2

Perfluoroalkylcarboxylic Acids

Perfluorobutanoic acid​

PFBA​

0.0040​

0.0020​

ND – not detected in filtrate

Perfluoropentanoic acid​

PFPeA​

0.0020​

0.0010​

Perfluorohexanoic acid​

PFHxA​

0.0020​

0.0010​

Perfluoroheptanoic acid​

PFHpA​

0.0020​

0.0010​

Perfluorooctanoic acid​

PFOA​

0.0020​

0.0010​

Perfluorononanoic acid​

PFNA​

0.0020​

0.0010​

Perfluorodecanoic acid​

PFDA​

0.0020​

0.0010​

Perfluoroundecanoic acid​

PFUnDA​

0.0020​

0.0010​

Perfluorododecanoic acid​

PFDoDA​

0.0020​

0.0010​

Perfluorotridecanoic acid​

PFTrDA​

0.0020​

0.0010​

Perfluorotetradecanoic acid​

PFTeDA​

0.0020​

0.0010​

Perfluoroalkylsulfonic Acids, Perfluorooctanesulfonamides, and Perfluorooctanesulfonamidoacetic Acids

Perfluorobutanesulfonic acid​

PFBS​

0.0020​

0.0010​

ND – not detected in filtrate

Perfluoropentanesulfonic acid​

PFPeS​

0.0020​

0.0010​

Perfluorohexanesulfonic acid​

PFHxS​

0.0020​

0.0010​

Perfluoroheptanesulfonic acid​

PFHpS​

0.0020​

0.0010​

Perfluorooctanesulfonic acid​

PFOS​

0.0020​

0.0010​

Perfluorononanesulfonic acid​

PFNS​

0.0020​

0.0010​

Perfluorodecanesulfonic acid​

PFDS​

0.0020​

0.0010​

PFOSA​

PFOSA​

0.0040​

0.0020​

N-MeFOSAA​

MeFOSAA​

0.0040​

0.0020​

N-EtFOSAA​

EtFOSAA​

0.0040​

0.0020​

Fluorotelomer Sulfonates and Next Generation PFAS Analytes

4:2 Fluorotelomer sulfonate​

8:2 FTS​

0.0080​

0.0020​

ND – not detected in filtrate

6:2 Fluorotelomer sulfonate​

6:2 FTS​

0.0080​

0.0020​

8:2 Fluorotelomer sulfonate​

8:2 FTS​

0.0080​

0.0020​

HFPO-DA​

GenX​

0.0040​

0.0020​

ADONA​

ADONA​

0.0080​

0.0020​

9Cl-PF3ONS (F-53B Major)​

--​

0.0080​

0.0020​

11Cl-PF3OUdS (F-53B Minor)​

--​

0.0080​

0.0020​

Sample Preparation in PFAS Testing

PFAS are a class of over 4,000 chemicals labeled as “forever chemicals” due to their persistence in the environment. Increasing concern about their threat to human health has sparked a cascade of regulatory action in the US and Europe. The primary focus is on water matrices starting with drinking water and then focusing on higher particulate water matrices and beyond. The primary analytical method for testing PFAS in water is solid phase extraction followed by liquid chromatography tandem mass spectrometry.

In all analytical methods, sample preparation should be carefully considered, especially for higher particulate samples which require filtration. However, in PFAS workflows, additional factors could complicate PFAS analyses downstream. Thus, we tested PES, nylon, and nylon-HPF membranes within a PFAS detection workflow to determine levels of contamination and found that none of the filters had any detectable levels of PFAS contamination. Therefore, when filtration of higher particulate samples is needed in a PFAS workflow, PES, nylon, and nylon-HPF Millex® filters provide a suitable option.

Alternative diameter PES Millex^® syringe filters and Millipore^® cut disc membranes are also available.

References

1. Kwiatkowski CF, Andrews DQ, Birnbaum LS, Bruton TA, DeWitt JC, Knappe DRU, Maffini MV, Miller MF, Pelch KE, Reade A, et al. 2020. Scientific Basis for Managing PFAS as a Chemical Class. Environ. Sci. Technol. Lett.. 7(8):532-543. https://doi.org/10.1021/acs.estlett.0c00255

2. Winchell LJ, Wells MJ, Ross JJ, Fonoll X, Norton JW, Kuplicki S, Khan M, Bell KY. 2021. Analyses of per- and polyfluoroalkyl substances (PFAS) through the urban water cycle: Toward achieving an integrated analytical workflow across aqueous, solid, and gaseous matrices in water and wastewater treatment. Science of The Total Environment. 774145257. https://doi.org/10.1016/j.scitotenv.2021.145257

3. Pérez F, Nadal M, Navarro-Ortega A, Fàbrega F, Domingo JL, Barceló D, Farré M. 2013. Accumulation of perfluoroalkyl substances in human tissues. Environment International. 59354-362. https://doi.org/10.1016/j.envint.2013.06.004

4. Lath S, Knight ER, Navarro DA, Kookana RS, McLaughlin MJ. 2019. Sorption of PFOA onto different laboratory materials: Filter membranes and centrifuge tubes. Chemosphere. 222671-678. https://doi.org/10.1016/j.chemosphere.2019.01.096

5. Labadie P, Chevreuil M. 2011. Biogeochemical dynamics of perfluorinated alkyl acids and sulfonates in the River Seine (Paris, France) under contrasting hydrological conditions. Environmental Pollution. 159(12):3634-3639. https://doi.org/10.1016/j.envpol.2011.07.028

6. So MK, Taniyasu S, Lam PKS, Zheng GJ, Giesy JP, Yamashita N. 2006. Alkaline Digestion and Solid Phase Extraction Method for Perfluorinated Compounds in Mussels and Oysters from South China and Japan. Arch Environ Contam Toxicol. 50(2):240-248. https://doi.org/10.1007/s00244-005-7058-x

7. Yamashita N, Kannan K, Taniyasu S, Horii Y, Petrick G, Gamo T. 2005. A global survey of perfluorinated acids in oceans. Marine Pollution Bulletin. 51(8-12):658-668. https://doi.org/10.1016/j.marpolbul.2005.04.026