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Analyze EPA Method 533 PFAS Reliably with Resprep WAX SPE

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Abstract

Low-level analysis of per-and polyfluoroalkyl substances (PFAS) requires high sensitivity and low background levels throughout the entire workflow from sample collection, transportation, and storage to analytical instrumentation and sample preparation. Resprep polymeric WAX SPE cartridges allow for accurate low-level quantitation of PFAS compounds in drinking water while meeting the requirements of EPA Method 533.

Introduction

Due to the low levels required by many regulatory agencies, the analysis of PFAS in drinking water often employs solid phase extraction (SPE) coupled with LC-MS/MS to reach low detection limits. EPA Method 533 [1] uses weak anion exchange (WAX) SPE to better retain short-chain acid compounds and obtain part-per-trillion (ppt) detection limits. However, due to the ubiquitous nature of PFAS compounds, the selection and screening of all materials and consumables that contact samples, including SPE cartridges, is critical for maintaining a clean background and allowing continued low-level analysis. In this assessment, Resprep WAX cartridges were used with a Resprep QR-12 vacuum manifold to extract samples for EPA Method 533 PFAS analysis using a Shimadzu Nexera LC and 8045 MS/MS. 

Experimental

Sample Preparation

Spikes and blanks were prepared in polypropylene bottles using 250 mL deionized water spiked with isotope dilution standards as per EPA Method 533, Section 7.16.1. The 6 mL SPE cartridges, which contained 500 mg of 30 µm WAX (cat.# 28291), were placed on a Resprep QR-12 vacuum manifold (cat.# 28298-VM) that was fitted with quick-replace liners (cat.# 28310-VM). Resprep sample delivery system lines (cat.# 26250) were used to transfer the samples to the SPE cartridges. While both the quick-replace liners and sample delivery lines contain PTFE, which can be a potential source of PFAS contamination, investigation of blanks taken using all lines and all ports on the manifold showed no detectable PFAS leaching. Thorough and regular blank checking of all SPE components and solvents, especially when using new lots of materials, is recommended to ensure that background contamination is below acceptable levels.

After preparation of the samples and setup of the SPE system, the samples were extracted following the instructions in EPA Method 533, Section 11.4, which is summarized in Figure 1.

Figure 1: Sample Preparation Procedure for EPA Method 533 PFAS Analysis

Flowchart showing steps for SPE cartridge conditioning, sample loading, drying, elution, and concentration

 

Analytical System

After extraction, the samples were analyzed by LC-MS/MS under the EPA Method 533 PFAS analysis conditions shown below. The use of a PFAS delay column is important to prevent any PFAS contamination upstream of the injector from coeluting with the samples. Thorough blank checking of the analytical system was performed and showed no detectable PFAS contamination. 

Instrument Conditions for EPA Method 533 PFAS Analysis
System: Shimadzu Nexera X2/Shimadzu LCMS-8045
Columns:

  • PFAS delay column (cat.# 27854)
  • Analytical column: Force C18, 1.8 µm x 50 mm x 2.1 mm (cat.# 9634252)

Injection volume: 3 µL
Mobile phase A: Water, 5 mM ammonium acetate
Mobile phase B: Methanol
Flow rate: 0.4 mL/min
Temperature: 40 °C
Gradient: 

Time (min) %B
0 20
6 95
6.6 95
6.61 20
7.5 20

Ion source: electrospray
Ion mode: ESI-
Mode: MRM

Method Detection Limits (MDL)

The method detection limit was calculated from the analysis of seven blank replicates and seven low-level spikes, as outlined in EPA’s Definition and Procedure for the Determination of the Method Detection Limit, Revision 2 [2]. The spikes were made at 0.5 ng/L using a 2 µg/mL stock solution of native PFAS compounds (EPA 533 PFAS calibration standard, cat.# 30736). The standard deviations of the spike and blank results were multiplied by the Student’s t-value of 3.143 to determine the MDL, and the higher of the results between the spikes and blanks was selected as the MDL.

Accuracy and Precision

Accuracy and precision were determined by analyzing five replicate 10 ng/L spikes. The accuracy of the spikes was calculated and compared to the recovery limits of 70-130% from EPA Method 533. The relative standard deviations of the spike results were also determined.

The recovery of the isotope dilution standards was calculated from the spike replicates and compared to the recovery limits of 50-200% from EPA Method 533 for PFAS analysis.

Results and Discussion

Good chromatographic results were obtained for all compounds, as shown in Figure 2. The MDL, accuracy, and precision results for native PFAS analytes are shown in Table I. The calculated MDLs were below the reporting limits shown in Table 7 in EPA Method 533, and the accuracy of the 10 ng/mL spikes ranged from 84 to 119% of the spiked value, well within the 70-130% recovery required by the method. The spikes showed good precision as well, with the results being ≤20% RSD.

Similarly, the recoveries for the isotope dilution standards were also within 20% of the true value and had precision ≤20% RSD. The results are shown in Table II.

 

Figure 2: Analysis of 25 ng/mL Standard for EPA Method 533

cgarm-img
LC_EV0584
PeakstR (min)Conc.
(ng/mL)
Precursor IonProduct Ion
1.Perfluoro-n-butanoic acid (PFBA)1.93325213169
2.Perfluoro-3-methoxypropanoic acid (PFMPA)2.5442522985
3.Perfluoro-n-pentanoic acid (PFPeA)3.36925263219
4.Perfluorobutanesulfonic acid (PFBS)3.5942529980
5.Perfluoro-4-methoxybutanoic acid (PFMBA)3.6692527985
6.Perfluoro(2-ethoxyethane)sulfonic acid (PFEESA)3.91925315135
7.Perfluoro-3,6-dioxaheptanoic acid (NFDHA)4.08425295201
8.1H,1H,2H,2H-Perfluorohexane sulfonic acid (4:2 FTS)4.12925327307
9.Perfluorohexanoic acid (PFHxA)4.18925313269
10.Perfluoro-1-pentanesulfonic acid (PFPeS)4.2782534980
11.Perfluoro(2-methyl-3-oxahexanoic) acid (HFPO-DA)4.36525285169
12.Perfluoroheptanoic acid (PFHpA)4.71525363319
PeakstR (min)Conc.
(ng/mL)
Precursor IonProduct Ion
13.Perfluoro-1-hexanesulfonic acid (PFHxS)4.7502539980
14.4,8-dioxa-3H-perfluorononanoic acid (ADONA)4.7825277251
15.1H,1H,2H,2H-Perfluorooctane sulfonic acid (6:2 FTS)5.09625427407
16.Perfluoro-1-heptanesulfonic acid (PFHpS)5.1322544980
17.Perfluorooctanoic acid (PFOA)5.11525413369
18.Perfluorooctanesulfonic acid (PFOS)5.4412549980
19.Perfluorononanoic acid (PFNA)5.43925463419
20.9-Chlorohexadecafluoro-3-oxanonane-1-sulfonic acid (9Cl-PF3ONS)5.58825531351
21.1H,1H,2H,2H-Perfluorodecane sulfonic acid (8:2 FTS)5.71225527507
22.Perfluorodecanoic acid (PFDA)5.71225513469
23.Perfluoroundecanoic acid (PFUnA)5.95425563519
24.11-Chloroeicosafluoro-3-oxaundecane-1-sulfonic acid (11Cl-PF3OUdS)6.04925631451
25.Perfluorododecanoic acid (PFDoA)6.15825613569
ColumnForce C18 (cat.# 9634252)
Dimensions:50 mm x 2.1 mm ID
Particle Size:1.8 µm
Pore Size:100 Å
Temp.:40 °C
Standard/SampleEPA 533 PFAS calibration standard (cat.# 30736)
Diluent:80:20 Methanol:water
Conc.:25 ng/mL
Inj. Vol.:3 µL
Mobile Phase
A:Water, 5 mM ammonium acetate
B:Methanol
Time (min)Flow (mL/min)%A%B
0.000.48020
6.000.4595
6.600.4595
6.610.48020
7.500.48020
DetectorShimadzu LCMS-8045
Ion Source:Electrospray
Ion Mode:ESI-
Mode:MRM
InstrumentShimadzu Nexera X2
NotesBranched isomers for PFOA, PFOS, and PFHxS labeled as peak number "a" and "b."
PFAS delay column used (cat.# 27854).
 
 

Table I: Results from MDL, Precision, and Accuracy Experiments for Native PFAS

Compound Abbreviation MDL (ng/L) Accuracy (%) %RSD
Perfluorobutanoic acid PFBA 8.5 95 20
Perfluoro-3-methoxypropanoic acid PFMPA 0.2 119 20
Perfluoropentanoic acid PFPeA 0.2 114 10
Perfluorobutane sulfonate PFBS 0.3 94 14
Perfluoro-4-methoxybutanoic acid PFMBA 1.1 88 8
Perfluoro(2-ethoxyethane)sulfonic acid PFEESA 0.2 84 12
Nonafluoro-3,6-dioxaheptanoic acid NFDHA 0.2 103 15
1H, 1H, 2H,2H-perfluorohexane sulfonate 4:2 FTS 0.3 97 19
Perfluorohexanoic acid  PFHxA 0.1 98 11
Perfluoropentane sulfonate  PFPeS 0.2 96 13
Hexafluoropropylene oxide dimer acid  HFPO-DA 1 89 13
Perfluoroheptanoic acid PFHpA 0.4 101 17
Perfluorohexane sulfonate PFHxS 0.3 110 15
4,8-Dioxa-3H-perfluorononanoic acid  ADONA 1.1 100 7
1H, 1H, 2H,2H-perfluorooctane sulfonate 6:2 FTS 0.7 105 8
Perfluoroheptane sulfonate PFHpS 0.5 101 12
Perfluorooctanoic acid  PFOA 0.2 112 9
Perfluorooctane sulfonate PFOS 0.3 109 5
Perfluorononanoic acid PFNA 0.5 101 10
9-Chlorohexadecafluoro-3-oxanonane-1-sulfonic acid  9Cl-PF3ONS 0.2 93 7
Perfluorodecanoic acid PFDA 0.4 111 6
1H, 1H, 2H,2H-perfluorodecane sulfonate 8:2 FTS 0.6 109 5
Perfluoroundecanoic acid PFUnA 0.8 110 6
11-Chloroeicosafluoro-3-oxaundecane-1-sulfonic acid 11Cl-PF3OUdS 0.3 95 11
Perfluorododecanoic acid PFDoA 1.1 94 7

 

Table II: Results from Precision and Accuracy Experiments for Isotope Dilution Standards

Compound Abbreviation Accuracy (%) %RSD
Perfluoro-n-[1,2,3,4-13C4]butanoic acid 13C4-PFBA 111 13
Perfluoro-n-[1,2,3,4,5-13C5]pentanoic acid 13C5-PFPeA 118 14
Sodium perfluoro-1-[2,3,4-13C3]butanesulfonate 13C3-PFBS 108 16
Sodium 1H,1H,2H,2H-perfluoro-1-[1,2-13C2]hexane sulfonate 13C2-4:2FTS 97 12
Perfluoro-n-[1,2,3,4,6-13C5]hexanoic acid 13C5-PFHxA 115 14
2,3,3,3-Tetrafluoro-2-(1,1,2,2,3,3,3-heptafluoropropoxy13C3-propanoic acid 13C3-HFPO-DA 89 14
Perfluoro-n-[1,2,3,4-13C4]heptanoic acid  13C4-PFHpA 110 14
Sodium perfluoro-1-[1,2,3-13C3]hexanesulfonate 13C3-PFHxS 114 14
Sodium 1H,1H,2H,2H-perfluoro-1-[1,2-13C2]-octane sulfonate 13C2-6:2FTS 92 7
Perfluoro-n-[13C8]octanoic acid 13C8-PFOA 113 14
Sodium perfluoro-[13C8]octanesulfonate 13C8-PFOS 102 7
Perfluoro-n-[13C9]nonanoic acid 13C9-PFNA 101 9
Perfluoro-n-[1,2,3,4,5,6-13C6]decanoic acid 13C6-PFDA 111 12
Sodium 1H,1H,2H,2H-perfluoro-1-[1,2-13C2]-decane sulfonate 13C2-8:2FTS 118 14
Perfluoro-n-[1,2,3,4,5,6,7-13C7]undecanoic acid 13C7-PFUnA 114 13
Perfluoro-n-[1,2-13C2]dodecanoic acid 13C2-PFDoA 112 13

Conclusions

EPA Method 533 PFAS analysis in drinking water can be challenging with low-level analysis complicated by background contamination. Resprep WAX cartridges have been shown to provide performance that meets or exceeds the requirements of EPA Method 533, allowing for analysis of PFAS at ng/L levels and lower. Visit www.restek.com/PFAS for additional products, methods, and technical resources.

References

  1. U.S. Environmental Protection Agency, Method 533, Determination of per-and polyfluoroalkyl substances in drinking water by isotope dilution anion exchange solid phase extraction and liquid
    chromatography/tandem mass spectrometry, November 2019. https://www.epa.gov/sites/default/files/2019-12/documents/method-533-815b19020.pdf
  2. U.S. Environmental Protection Agency, Definition and procedure for the determination of the method detection limit, revision 2, December 2016.
    https://www.epa.gov/sites/default/files/2016-12/documents/mdl-procedure_rev2_12-13-2016.pdf  
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