2019.01.30_CCO.p11_PFASWaterModifiedVersion8WorkInstrucution

Certain sections are replaced by the words PROPRIETARY CONTENT to allow distribution of this document.
Perfluoro-n-butanoic acid PFBA 375-22-4
Perfluoro-n-pentanoic acid PFPeA 2706-90-3
8:2 - Fluorotelomersulfonate 8:2FTS 39108-34-4
N-methylperfluoro-1-octanesulfonamidoacetic acid NMeFOSAA 2355-31-9
N-ethylperfluoro-1-octanesulfonamidoacetic acid
Perfluoroundecanoic acid
NEtFOSAA 2991-50-6
4:2-Fluorotelomersulfonate 4:2-FTS 757124-72-4
Perfluoropentanesulfonate PFPeS 2706-94-4
6:2-Fluorotelomersulfonate 6:2-FTS 27619-97-2
Perfluoroheptanesulfonate PFHpS 375-92-8
Perfluorononanesulfonate PFNS 68259-12-1
Perfluorodecanesulfonate PFDS 335-77-3
10:2-Fluorotelomersulfonate 10:2-FTS 120226-60-0
Perfluorododecanesulfonate PFDoDS 79780-39-5
Perfluorohexadecanoic acid PFHxDA 67905-19-5
Perfluorooctadecanoic acid PFODA 16517-17-6
Perfluorooctanesulfonamide PFOSA 754-91-6
2-(N-methylperfluoro-1-octanesulfonamido)-ethanol NMePFOSAE 24448-09-7
N-methylperfluoro-1-octanesulfonamide NMePFOSA 31506-32-8
2-(N-ethylperfluoro-1-octanesulfonamido)-ethanol NEtPFOSAE 1691-99-2
N-ethylperfluoro-1-octanesulfonamide NEtPFOSA 4151-50-2
Basic Principles
A 250-mL aqueous sample is fortified with isotopically-labeled extraction standards and is passed
through a solid phase extraction (SPE) cartridge to extract the analytes. The compounds are eluted
from the solid phase with a combination of solvents. The extract is concentrated to ~400-500ul with
nitrogen in a heated water bath, and then reconstituted to 1ml with methanol. Isotopically-labeled
injection internal standards are added to the sample extract and it is analyzed by LC/MS/MS
operated in negative electrospray ionization (ESI) mode for detection and quantification of the
analytes. Quantitative analysis is performed using isotope dilution.
Reference Modifications
EPA Method 537 is written specifically for the analysis of drinking water samples. The following
modifications to the method have been made to accommodate all aqueous samples.
1. A labeled isotopic analog is spiked into samples for all compounds where an isotopic analog is
commercially available. These isotopic compounds are referred to as extraction standards. For
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 4 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
Exposure to these chemicals must be reduced to the lowest possible level by whatever means
available, such as fume hoods, lab coats, safety glasses, and gloves. Gloves, lab coats, and safety
glasses should be worn when preparing standards and handling samples. Avoid inhaling solvents
and chemicals and getting them on the skin. Wear gloves when handling neat materials. When
working with acids and bases, take care not to come in contact and to wipe any spills. Always add
acid to water when preparing reagents containing concentrated acids.
All laboratory waste is accumulated, managed, and disposed of in accordance with all Federal, State,
and local laws and regulations. All solvent waste and extracts are collected in approved solvent
waste containers in the laboratory and subsequently emptied by personnel trained in hazardous
waste disposal into the lab-wide disposal facility. HPLC vials are disposed of in the lab container for
waste vials, and subsequently lab packed. Any solid waste material (disposable pipettes and broken
glassware, etc.) may be disposed of in the normal solid waste collection containers.
Personnel Training and Qualifications
All personnel performing this procedure must have documentation of reading, understanding, and
agreeing to follow the current version of this SOP and an annual documented Demonstration of
Capability (DOC).
Each chemist performing the extraction must work with an experienced employee for a period of
time until they can independently perform the extraction. Also, several batches of sample
extractions must be performed under the direct observation of another experienced chemist to
assure the trainee is capable of independent preparation. Proficiency is measured through a
documented Initial Demonstration of Capability (IDOC).
Each LC/MS/MS analyst must work with an experienced employee for a period of time until they can
independently calibrate the LC/MS/MS, review and process data, and perform maintenance
procedures. Proficiency is measured through a documented Initial Demonstration of Capability
(IDOC).
The IDOC and DOC consist of four laboratory control samples (or alternatively, one blind sample for
the DOC) that is carried through all steps of the extraction and meets the defined acceptance
criteria. The criteria include the calculation of mean accuracy and standard deviation.
Sample Collection, Preservation, and Handling
A. Sample Collection
The samples are collected in 250 mL polyethylene bottles containing 1.25 grams of Trizma, resulting
in a Trizma concentration in the sample of 5 g/L. Trizma functions as a free chlorine scavenger;
therefore, any chlorinated water supplies require the preservative. Water samples from non-
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 6 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
chlorinated water sources would not necessarily require the Trizma preservative. Keep the sample
sealed from time of collection until extraction.
NOTE: PFAS contamination during sampling can occur from a number of common sources, such as
food packaging and certain foods and beverages. Proper hand washing and wearing nitrile gloves
will aid in minimizing this type of accidental contamination of the samples.
B. Sample Storage and Shipment
1. Samples must be chilled during shipment and must not exceed 10°C during the first
48 hours after collection. Sample temperature must be confirmed to be at or below 10°C when the
samples are received at the laboratory.
2. Samples stored in the lab must be held at a temperature of 0° to 6°C, not frozen,
until extraction.
3. Water samples must be extracted within 14 days. Extracts must be analyzed within
28 days after extraction. Extracts are stored at room temperature.
Apparatus and Equipment
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 7 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
PROPRIETARY CONTENT
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 8 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
Reagents and Standards
All solvents, acids, and bases are stored in glass bottles in flammable proof cabinets or pressure
resistant steel drums. Solvents, acids, and bases are stored at ambient temperature for up to 1
year. All non-solvents are stored according to manufacturer’s storage conditions.
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 9 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
B. Standards: See SOP T-PFAS-WI13881
A. Reagents: PROPRIETARY CONTENT
If the criteria are not met, the source of the problem must be determined and corrected.
Situations may exist where the initial calibration can be used. In those cases, the data will be
reported with a qualifying comment.
7. Initial Calibration Verification (ICV)
A check standard prepared from a second source (ICV) is injected to confirm the validity of the
calibration curve/standard. The calculated amount for each analyte should be ± 30% of the true
value.
B. Continuing calibration
1. Once the calibration curve has been established, the continuing accuracy must be verified by
analysis of a continuing calibration verification (CCV) standard every ten samples and at the end of
the analysis sequence.
a. The CCV run after the initial calibration must be at the CAL3 level.
b. Subsequent CCV standards should alternate between the low, mid and high levels of the
calibration curve.
2. Acceptance criteria
a. The calculated amount for each compound (native and extraction standard) in the CCV
standard must be within ±30% of the true value. Samples that are not bracketed by acceptable CCV
analyses must be reanalyzed. The exception to this would be if the CCV recoveries are high,
indicating increased sensitivity, and there are no positive detections in the associated samples, the
data may be reported with a qualifying comment. If two consecutive CCVs fail criteria for target
analytes, two passing CCVs must be analyzed or the source of the problem determined and the
system recalibrated before continuing sample analysis.
b. The absolute areas of the injection internal standards should be within 50-150% of the
average areas measured during the initial calibration.
Procedure
A. Sample Preparation
1. Weigh full sample container on a calibrated top loading balance and record the first reading in
the automated prep entry system.
2. If required, add 1.25 grams of Trizma to a 250 ml HDPE bottle for the method blank and the
laboratory control sample (LCS) and LCSD if needed. Fill each bottle with 250 ml of Milli-Q water.
Record 250 ml as the volume for the batch QC samples on the batchlog.
3. If sample has dissolved and/or settleable solid content; i.e., is cloudy or has a layer of
sediment/solids at the bottom of the bottle, an aliquot should be taken from the original bottle and
diluted with reagent water in order to minimize difficulty passing through the SPE sorbent bed. If
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 11 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
3. Acquisition method: See attachment 3
4. Load sample vials containing standards, quality control samples, and sample extracts into
autosampler tray. Allow the instrument adequate time to equilibrate to ensure the mass spec and
LC have reached operating conditions (approximately 5 minutes) before the first injection. Analyze
several solvent blanks clean the instrument prior to sample acquisition.
5. After the initial calibration, inject a solvent blank, followed by the ICV, L/B standard, closing
Cal 3 level CCV , CCV, extraction batch QC, and samples. Bracket each set of ten samples with a
CCV standard, alternating between the Cal3, Cal4, and Cal5 levels.
6. After injections are completed, check all CCV recoveries and absolute areas to make sure
they are within method control limits. See Calibration section B.2 for acceptance criteria. Process
each chromatogram and closely evaluate all integrations, baseline anomalies, and retention time
differences. If manual integrations are performed, they must be documented and a reason given for
the change in integrations. The manual integrations are documented during data processing and all
original integrations are reported at the end of the sample PDF file with the reason for manual
integration clearly listed.
7. Quantitate results for the extraction blank.
a. No target analytes at or above the reporting limit may be found in the extraction blank for
acceptable batch results. If a target analyte is detected in the extraction blank but not detected in
the sample, the data is reported. If a target analyte is detected in the method blank at a
concentration greater than the reporting limit and also in the sample, the sample must be
reextracted. If the target analyte in the sample is detected at a concentration greater than 10 times
the amount detected in the method blank, the data is reported.
8. Calculate the recoveries of spiked analytes for the LCS, matrix spike and matrix spike
duplicate (MS/MSD) by comparing concentrations observed to the true values. The advisory QC
acceptance limits for LCS and MS/MSD recovery are 70 to 130% for each analyte. The advisory QC
acceptance limit for the relative percent difference (%RPD) between LCS/LCSD and MS/MSD is
≤30%. The limits are advisory until sufficient data points are available to determine statistical QC
acceptance limits. If LCS and/or LCSD recoveries are acceptable, proceed to sample quantitation. If
the LCS recoveries are unacceptable, the samples associated with the LCS may need to be
reanalyzed. If LCS recoveries are above the advisory QC acceptance limits, and there are no positive
detections in the sample, the data may be reported. If MS/MSD recoveries are outside QC
acceptance criteria, the associated data will be flagged or noted in the comments section of the
report.
9. Isotopically labeled extraction standards are added to all samples, extraction blank,
LCS/LCSD, and MS/MSD prior to extraction.
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 15 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
a. The recovery of the extraction standards should be within QC acceptance criteria. If the
extraction standard recovery(ies) is(are) outside the QC limit(s), consult a supervisor to determine
the appropriate course of action based on batch and sample results.
10. Isotopically labeled injection standards are added to each QC and field sample extract
prior to analysis.
a. The absolute areas of the injection standards should be within 50-150% of the average
areas measured during the initial calibration. If the internal standards are recovered outside 50-
150%, consult a supervisor to determine the appropriate course of action based on batch and
sample results.
11. Compare the retention times of all of the analytes, surrogates and internals standards. The
relative retention times should not vary by more than 0.2 retention time units.
12. The MDL standard and the linear/branch chain standard are used when assessing the
correctness of the computer generated peak integrations.
13. If the calculated concentration exceeds the calibration range of the system, dilute the
extract with MeOH and add the appropriate amount of extraction standard to match the original
concentration. Add 10 ul of injection internal standard and analyze the dilution.
Dilution Example 1/10: Mix 0.877 mL of MEOH with 0.100 mL of sample extract and
0.0225 mL of labeled extraction standard. Vortex to mix. Using an auto-pipette, transfer 200 uL of
the mixed solution into a labeled auto-sampler vial containing a plastic insert. Using a syringe, add
10 uL of labeled injection std to the 200 uL aliquot. Cap and vortex thoroughly to mix.
Calculations
A. Peak Area Ratio
B. Analyte Concentration using linear through zero curves (MQ Data processing system)
Concentration = (area ratio ÷ slope) x Dilution Factor x Internal Standard concentration
C. Sample Concentration (used only for aqueous samples using the MultiQuant data processing
system on the AB Sciex LC/MS/MS)
Sample concentration (ng/l) = Calc conc x (Sample volume ÷ Sample weight) x DF
US Eurofins US Lancaster Laboratories Environmental ­ Polyfluorinated Alkyl Substances (PFAS) in Aqueous Samples by Method 537 Version 1.1
Modified Using LC/MS/MS
Printed by: Jeremy Beckley, d. 2019/01/14 15:34 CET
Page 16 of 17
Polyfluorinated Alkyl
Substances (PFAS) in Aqueous
Samples by Method 537 Version
1.1 Modified Using LC/MS/MS
Level:
Work Instruction
Document number:
T­PFAS­WI14355
Old Reference:
1­P­QM­WI­9039651 (1­P­QM­WI­9012802) 
Version:
8
Organisation level:
5­Sub­BU  
Approved by: UDM6
Effective Date 11­SEP­2018
Document users:
6_EUUSLA_PFAS_Analyst,
6_EUUSLA_PFAS_Data_Reviewers,
6_EUUSLA_PFAS_Sample_Prep
Responsible:
5_EUUSLA_PFAS_Manager
Always check on­line for validity.
PFOA 413 369
PFOA (2)413 169
13C4-PFOS 503 80
13C8-PFOS 507 80
PFOS 499 80
PFOS (2)413 169
13C9-PFNA 472 427
PFNA 463 419
PFNA (2)463 169
13C8-PFOSA 506 78
PFOSA 498 78
PFNS 549 80
PFNS (2)549 99
13C2-PFDA 515 470
13C6-PFDA 519 474
PFDA 513 469
PFDA (2)513 169
13C2-8:2-FTS 529 81
8:2-FTS 527 507
8:2-FTS (2)527 81
d7-NMePFOSAE 623 59
NMePFOSAE 616 59
d3-NMePFOSA 515 169
NMEPFOSA 512 169
d3-NMeFOSAA 573 419
NMeFOSAA 570 419
NMeFOSAA (2)570 483
d9-NEtPFOSAE 639 59
NEtPFOSAE 630 59
d5-NETPFOSA 531 169
NEtPFOSA 526 169
PFDS 599 80
PFDS (2)599 99
13C7-PFUnDA 570 525
PFUnDA 563 519
PFUnDA (2)563 169
d5-NEtFOSAA 589 419
NEtFOSAA 584 419
NEtFOSAA (2)584 526
13C-PFDoDA 615 570
PFDoDA 613 569
PFDoDA (2)613 169
10:2-FTS 627 607
10:2-FTS (2)627 81
PFDoS 699 80
PFTrDA 663 619
PFTrDA (2)663 169
13C2-PFTeDA 715 670
PFTeDA 713 669
PFTeDA (2)713 169
PFHxDA 813 769
PFHxDA (2)813 169
PFODA 913 869
PFODA (2)913 169
Page 1 of 2

Attachment 2

PFAS Injection Standards/Extraction Standards/Native Compounds

Injection Standards
Inj Std Internal Standard/Injection
Standard
I13C3-PFBA 13C3-PFBA
I13C2-PFOA 13C2-PFOA
I13C4-PFOS 13C4-PFOS
I13C2-PFDA 13C2-PFDA

Extraction Standards
Extraction Standard Internal Standard
E13C4-PFBA
13C3-PFBA E13C5-PFPeA
E13C3-PFBS
E13C2-4:2-FTS

13C2-PFOA
E13C5-PFHxA
E13C3-PFHxS
E13C4-PFHpA
E13C2-6:2-FTS
E13C8-PFOA
E13C8-PFOS 13C4-PFOS E13C9-PFNA
E13C8-PFOSA
13C2-PFDA
E13C6-PFDA
E13C2-8:2-FTS
Ed7-NMePFOSAE
Ed3-NMePFOSA
Ed3-NMeFOSAA
Ed9-NEtPFOSAE
Ed5-NEtPFOSA
E13C7-PFUnDA
Ed5-NEtFOSAA
E13C2-PFDoDA
E13C2-PFTeDA

Page 2 of 2

Native PFAS Compounds
Native Extraction Standard PFBA 13C4-PFBA
PFPeA 13C5-PFPeA
PFBS 13C3-PFBS PFPeS
4:2-FTS 13C2-4:2-FTS
PFHxA 13C5-PFHxA
PFHxS 13C3-PFHxS PFHpS
PFHpA 13C4-PFHpA
6:2-FTS 13C2-6:2-FTS
PFOA 13C8-PFOA
PFOS
13C8-PFOS PFNS
PFDS
PFDoS
PFNA 13C9-PFNA
PFOSA 13C8-PFOSA
PFDA 13C6-PFDA
8:2-FTS 13C2-8:2-FTS 10:2-FTS
NMePFOSAE d7-NMePFOSAE
NMePFOSA d3-NMePFOSA NMeFOSAA d3-NMeFOSAA
NEtPFOSAE d9-NEtPFOSAE
NEtPFOSA d5-NEtPFOSA
PFUnDA 13C7-PFUnDA
NEtFOSAA d5-NEtFOSAA
PFDoDA 13C2-PFDoDA PFTrDA
PFTeDA
13C2-PFTeDA PFHxDA
PFODA