Manual Extraction of PFAS in Drinking Water Following DIN 38407-42

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 1

Manual Extraction of PFAS in Drinking Water 

Following DIN 38407-42

Authors
William Jones, Biotage, Salem, NH, USA
Matthew Harden, Biotage, Salem, NH, USA

Application Note AN968

Introduction
Per- and polyfluorinated alkyl substances (PFAS) have been used abundantly since 
their inception in the twentieth century and have become a closely monitored class of 
compounds within environmental testing. This application note outlines a procedure for 
those seeking to follow DIN 38407-42 for large volumes of water. The data presented 
was generated using a Biotage® VacMaster™ vacuum manifold with a PFAS free Biotage®
VacMaster™ Large Volume Extraction (LVE) kit in conjunction with EVOLUTE® PFAS SPE 
columns and a TurboVap® LV system.

Agilent

» InfinityLab PFC Delay Column, 4.6 x 30 mm, p/n 5062-8100

» ZORBAX RRHD Eclipse Plus C18, 95 Å, 

2.1 x 50 mm, 1.8 µm, p/n 959757-902

Sigma-Aldrich

» Ammonium Acetate, ACS Reagent 

Grade ≥ 97%, p/n 238074-25G

» Acetic Acid, Glacial, ReagentPlus®, ≥ 99% p/n A6283
Honeywell

» Water, ACS Certified, HPLC Grade, p/n AH365-4

» Methanol, Burdick & Jackson™, LC-MS Grade, p/n LC230-4
VWR

» 15 mL Polypropylene Centrifuge Tubes 

with Caps, p/n 21008-670

» 250 mL Polypropylene Wide Mouth Bottles, p/n 414004-125

Equipment and Materials Used
Biotage

» Biotage® VacMaster™ 20 Sample Processing Station (with 

15 mL rack), p/n 121-2015ML, fitted with polypropylene 
(PFAS free) stopcocks (p/n 121-0009-PP)

» Biotage® VacMaster™ LVE Kit (PFAS) 

for 1, 3, 6 mL SPE Columns (p/n 121-2190)

» EVOLUTE® PFAS 500 mg/6 mL SPE Columns, p/n 614-0050-CP

» EVOLUTE® PFAS 150 mg/6 mL SPE Columns, p/n 614-0015-CP

» TurboVap® LV Automated Solvent 

Evaporation System, p/n 415000

» TurboVap® LV Multi Rack (48 Positions, 

10–20 mm Tubes), p/n 414964

Wellington Laboratories

» ISO 21675:2019 Labelled Stock Solution, 

1.2 mL, p/n ISO 21675-LSS

» ISO 21675:2019 Native Stock Solution, 

1.2 mL, p/n ISO 21675-NSS

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 2

Analytes
Table 1. Listing of Target Analytes and Internal Standards.

Target Analyte

Acronym

CAS

Perfluorobutanoic acid

PFBA

375-22-4

Perfluoropentanoic acid

PFPeA

2706-90-3

Perfluorohexanoic acid

PFHxA

307-24-4

Perfluoroheptanoic acid

PFHpA

375-85-9

Perfluorooctanoic acid

PFOA

335-67-1

Perfluorononanoic acid

PFNA

375-95-1

Perfluorodecanoic acid

PFDA

335-76-2

Perfluorobutanesulfonic acid

PFBS

375-73-5

Perfluorohexanesulfonic acid

PFHxS

355-46-4

Perfluorooctanesulfonic acid

PFOS

1763-23-1

Perfluoroundecanoic acid1

PFUnA

2058-94-8

Perfluorododecanoic acid1

PFDoA

307-55-1

Perfluoroheptanesulfonic acid1

PFHpS

375-92-8

Perfluorodecanesulfonic acid1

PFDS

335-77-3

1H,1H,2H,2H-perfluorooctanesulfonic acid1

H4PFOS

27619-97-2

Internal Standard

Perfluoro-n-[1,2,3,4-13C4]butanoic acid

13

C4-PFBA

Perfluoro-n-[1,2,3,4,5-13C5]pentanoic acid

13

C5-PFPeA

Perfluoro-n-[1,2-13C2]hexanoic acid

13

C2-PFHxA

Sodium perfluoro-1-[1,2,3-13C3]hexanesulfonate

13

C3-PFHxS

Sodium perfluoro-1-[1,2,3-18O2]hexanesulfonate

18

O2-PFHxS

Perfluoro-n-[1,2,3,4-13C4]heptanoic acid

13

C4-PFHpA

Perfluoro-[1,2-13C4]octanoic acid

13

C4-PFOA

Sodium perfluoro-1-[1,2,3,4-13C4]octanesulfonate

13

C4-PFOS

Perfluoro-n-[13C5]nonanoic acid

13

C5-PFNA

Perfluoro-n-[1,2-13C2]decanoic acid

13

C2-PFDA

1

Target compounds are from the expanded list given in DIN 38407-42 Section 2.1.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 3

Solution Preparation
Ammonia/Methanol Solution
1.  Add 400 μL of NH4OH for every 100 mL of methanol to  

a clean beaker.

2.  Agitate to homogenize.
3.  Prepare new solution daily.
Acetate Buffer
1.  Measure out 499.5 mL of reagent water in a clean beaker.
2.  Add 0.193 g of NH4Ac.
3.  Sonicate the solution for 5 minutes until the salt is  

fully dissolved.

4.  Add 570 μL of glacial acetic acid.
5.  Agitate to homogenize the solution.
Working Spiking Solution
1.  Dilute 100 μL of the native stock solution with 900 μL of 

methanol to achieve a 10 ppt solution.

Summary of SPE Method
SPE Column Format
EVOLUTE® PFAS 500 mg/6 mL or EVOLUTE® PFAS 150 mg/6 mL

Sample Pre-Treatment
Adjust the pH of each sample to 3 using glacial acetic acid. Add 
targets and internal standards.

Conditioning
Condition each column with 0.1 % NH4OH in methanol (10 mL) 
followed by methanol (10 mL).

Equilibration
Equilibrate each column with reagent water (10 mL).

Sample Loading
Load sample at a flow rate of 5 mL/min.

Wash
Rinse the sample container with acetate buffer solution (10 mL) 
and load onto the column. Repeat using reagent water (10 mL).

Dry
Dry the column for 5 minutes at a flow rate of 5 mL/min.

Elution
Rinse the sample container with methanol (5 mL) and use to 
elute the analytes from the column at a flow rate of 2 mL/min. 
Repeat using 0.1 % NH4OH in methanol (5 mL).

Post Extraction
Concentrate the extract to a volume of 1 mL. Add IS and mix 
prior to analysis.

Note: EVOLUTE® PFAS 60 mg/3 mL columns may be used 
however sample and solvent volumes should be adjusted.  
Refer to DIN 38407-42 for the appropriate amounts.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 4

Sample Preparation Procedure
1.  Clean all parts of the Biotage® VacMaster™ system per the 

procedure given in Appendix A.

2.  Set up and fill new sample containers with water; 

250–500 mL are typical for this method.

3.  Add glacial acetic acid to each of the sample containers to 

reduce the pH to 3 (approximately 100 μL for 250 mL sample 
volumes and 200 μL for 500 mL sample volumes).

4.  Verify the pH of the sample is 3 using pH paper. To reduce 

the possibility of contamination, a duplicate volume was 
collected and adjusted to the appropriate pH and the same 
volume of acid was added to the sample container.

5.  Prepare for the determination of the initial sample volume by 

either marking the level of the sample on the container or by 
weighing the sample container.

6.  Add 20 μL of the undiluted Labeled Stock Solution to each 

of the sample containers. If desired, fortify a sample using 
target analytes: the addition of 125 μL or 37.5 μL of the native 
stock solution will yield either 50 ppt or 15 ppt concentra-
tions respectively, while the addition of 50 μL of the working 
spiking solution will yield a 2 ppt concentration. If the mixes 
used were different than the ones outlined in this note, 
adjust the concentration or spiking amounts as needed.

7.  Load the desired EVOLUTE® PFAS columns onto the Biotage® 

VacMaster™. Seal any unused positions using VacMaster Port 
Sealing Plugs (p/n 121-0005)

8.  Rinse each column with 10 mL of 0.1 % NH4OH in methanol 

and apply vacuum at 10 mL/min to pull it to waste. Do not 
allow the sorbent to go dry.

9.  Rinse each column with 10 mL of methanol and apply 

vacuum at 10 mL/min to pull it to waste. Do not allow the 
sorbent to go dry.

10. Rinse each column with 10 mL of reagent water and apply 

vacuum at 10 mL/min to send it to waste. Do not allow the 
water level to drop below the top of the packing.

11.  Using the Biotage® VacMaster™ LVE Kit, place one end of 

the cleaned tubing into the bottom of each of the sample 
containers, and secure in position using the clips provided.

12. Load the samples onto the columns using a flow rate of 

5 mL/min.

13. Once the sample has been fully loaded, rinse the sample 

containers using 10 mL of acetate buffer solution, swirl to 
ensure the full rinsing of the container, and load the aliquot 
onto the column at a rate of 5 mL/min.

14. Rinse the sample containers using 10 mL of reagent water, 

swirl to ensure the full rinsing of the container, and load the 
aliquot onto the column at a rate of 5 mL/min.

15. Dry the column for 5 minutes at a rate of 5 mL/min.
16. Load 15 mL centrifuge tubes into the rack corresponding to 

each of the column positions and load into the VacMaster.

17.  Rinse each sample container using 5 mL of methanol and 

swirl to ensure the full rinsing of the container. Load the 
aliquot through the appropriate column and collect at a 
dropwise rate.

18. Rinse each sample container using 5 mL of 0.1% NH4OH 

in methanol and swirl to ensure the full rinsing of the 
container. Load the aliquot through the appropriate column 
and collect at a dropwise rate.

19. Determine the initial sample volume by either using a gradu-

ated cylinder and filling the sample container to the original 
mark or by taking an additional weight of the container.

20. Transfer the centrifuge tubes to the TurboVap® LV system and 

concentrate the samples to just under 1 mL using nitrogen 
according to the parameters in Table 2.

21. Bring the final extract to 1 mL and transfer to an  

autosampler vial.

22. Load the extract onto a calibrated LC-MS/MS system and 

process using the conditions given in the below sections.

Table 2. TurboVap® LV Concentration Protocol.

Bath Temp:

60 ˚C

Evaporation Mode

Method (Ramp Gradient)

Manifold Setup

48 positions

Rack Row Height

120 mm*

Step 1:

1.5 L/min for 20 min

Step 2:

3.0 L/min for 15 min

Step 3:

3.5 L/min for 45 min

*The nozzle position was adjusted such that it was as far to the right as 

possible to give the user a clear view of the vortex within the tube.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 5

LC-MS/MS Conditions
Agilent 1290 Infinity II LC System

 » 1290 Infinity II Multicolumn Thermostat, G7116B

 » 1290 Infinity II Multisampler, G7167B

 » 1290 Infinity II High Speed Pump, G7120A

 » InfinityLab PFC-free HPLC Conversion Kit, 5004-0006

Columns

 » InfinityLab PFC Delay Column, 4.6 x 30 mm, p/n 5062-8100

 » ZORBAX RRHD Eclipse Plus C18, 95 Å, 

2.1 x 50 mm, 1.8 µm, p/n 959757-902

Mobile Phases

 » A:  20 mM Ammonium Acetate in Water

 » B: Methanol

Table 3. LC Gradient.

Time (min)

%A

%B

0.50

95.00

5.00

3.00

60.00

40.00

16.00

20.00

80.00

18.00

20.00

80.00

20.00

5.00

95.00

20.50

0.00

100.00

25.00

0.00

100.00

26.00

5.00

95.00

 » Flow Rate: 0.2 mL/min

 » Injection Volume: 5 μL

 » Column Temperature: 50 ˚C

Agilent 6470 MS/MS, G6470B

 » Gas Temperature: 230 ˚C

 » Gas Flow: 4 L/min

 » Nebulizer: 20 psi

 » Sheath Gas Temperature: 375 ˚C

 » Sheath Gas Flow: 12 L/min

 » Capillary Voltage (Positive): 3500 V

 » Capillary Voltage (Negative): 3500 V

 » Nozzle Voltage (Positive): 500 V

 » Nozzle Voltage (Negative): 0 V

For a complete listing of MRM Transitions, see Appendix B

Results
System Calibration
For the work being done here, a total of six points were used 
in the calibration covering a range of 0.2-20 ppt. The lowest 
three points were below the calculated MRL. The curve was 
forced through zero and achieved excellent linearity across the 
calibration range.

PFBS

PFHxS

Figure 1. Calibration curves for PFBS and PFHxS. Calibration curves for 

the remaining target analytes in Table 1 are shown in Appendix C.

Determination of the Minimum Reporting 

Level (MRL) and Detection Limits (DL)
A target MRL of 2 ng/L was selected and at least seven replicate 
laboratory fortified blanks (LFBs) were created and run at that 
concentration. Figure 2 below illustrates the results of this test 
for both the 150 mg and 500 mg columns using 250 mL sample 
volumes; all compounds were recovered within 15% of the 
spiked amount and had less than 10% CV.

Figure 2. MRL and DL Recoveries. Those compounds with an asterisk were 

used in salt form.

The data for individual compounds is shown in Appendix D.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 6

Demonstration of Low System Background
An investigation into the background of the complete process 
was done in three steps. The first step was to run blank 
injections of methanol on the analytical system (system blank). 
The second step was to load centrifuge tubes containing a 
similar volume of methanol as would result from the extraction 
process onto the evaporation system, allowing them to 
concentrate and then run on the analytical system (evaporation 
blank). The third and final step was to create a full Laboratory 
Reagent Blank (LRB), extract and concentrate it, and run it on 
the analytical system. By separating the process into three 
steps it becomes easier to determine what, if any, contribution 
to the overall background each of the steps has. The result of 
these tests are given in Appendix E and selected data are shown 
below in Figures 3–5.

Figure 3. Contribution of the TurboVap® LV to the PFAS Background.  

Those compounds with an asterisk were used in salt form.

Figure 4. PFAS Background for full LRB using EVOLUTE® PFAS 500 mg/6 mL 

columns. Those compounds with an asterisk were used in salt form.

Figure 6. Initial Demonstration of Accuracy (15 ng/L, n=4).  

Those compounds with an asterisk were used in salt form.

Figure 7. Initial Demonstration of Precision (15 ng/L, n=4). Those 

compounds with an asterisk were used in salt form.

Figure 5. PFAS Background for full LRB using EVOLUTE® PFAS 150 mg/6 mL  

columns. Those compounds with an asterisk were used in salt form.

For those results which were generated using only the analytical 
system, all target analytes were N.D. (unable to be separated 
from the noise in the baseline) and so were not listed out in the 
previous tables.

When examining the data resulting for both the TurboVap® LV 
and the full LRB tests (which includes the Biotage® VacMaster™ 
manifold, PFAS Free Large Volume Loading Kit, and the 
EVOLUTE® cartridges as well as the TurboVap® LV) there are clear 
indications of the presence of a PFAS background. However, 
even at the highest concentrations detected, all levels are much 
lower than the 1/3 MRL limit indicating that the background is 
acceptable and will not interfere with future sample runs.

Initial Demonstration of Precision 

and Accuracy (IDP, IDA)
To determine the precision and accuracy of the sample prepara-
tion process, four LFB samples were prepared at concentrations 
of 15 ppt. The data is given in Appendix F and illustrated in 
Figures 6 and 7.

The results show that the average recovery for each target 
analyte was within 15% of the nominal value and that the 
coefficient of variation (CV) for each analyte fell under 10% 
on average.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 7

Examination of System Carryover
To simulate an influent sample, four LFB samples were 
created with concentrations which were above the range of 
the calibration curve. These samples were extracted, and 
the cleanup procedure given in Appendix A was run three 
times. To ensure that the system background was adequately 
reduced, a set of four LRB samples were extracted immediately 
afterwards and analyzed. The LRB data is presented in 
Appendix G and illustrated in Figure 8.

Figure 8. Results of carryover study following four, 50 ng/L LFB samples 

using EVOLUTE® PFAS 500 mg/6 mL columns. Those compounds with an 

asterisk were used in salt form.

Conclusion
With the scrutiny being given to the presence of PFAS 
compounds in the environment, it is essential to find reliable 
products which can meet the requirements of DIN 38407-42.  
This application note has shown that the VacMaster™ vacuum 
manifold with PFAS free accessories, EVOLUTE® PFAS SPE 
columns and the TurboVap® LV can be used to easily meet and 
exceed the demands of the method.

Ordering Information

Part Number

Description

Qty

121-2015ML

Biotage® VacMaster™ 20 Sample Processing 

Station With 15 mm Rack

1

121-2190

Biotage® VacMaster™ LVE Kit (PFAS) for 1, 3, 

6 mL SPE Columns

1

121-0009-PP

Polypropylene (PFAS) Stopcocks

10

614-0050-CP EVOLUTE® PFAS 500 mg/6 mL columns

30

614-0015-CP

EVOLUTE® PFAS 150 mg/6 mL columns

30

614-0006-BP EVOLUTE® PFAS 60 mg/3 mL columns

50

415000

TurboVap® LV Automated Solvent 

Evaporation System

1

414964

TurboVap® LV Multi Rack (48 Positions, 

10–20 mm Tubes)

1

The graph shown in Figure 8 shows a clear indication that the 
cleaning procedure in Appendix A was successful in reducing 
the background of PFAS compounds to below the 1/3 MRL limit.  
For further reductions in the background, additional cleaning 
steps could be employed.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 8

Appendix A

Biotage® VacMaster™ Cleaning Procedure

For the best results, it is recommended that this procedure be 
completed before the use of the VacMaster™ each day and at the 
end of each extraction prior to proceeding with the next set of 
samples.

1.  Ensure that a column and column adapter is installed onto 

each VacMaster™ position slated to be cleaned.

2.  Fill a clean beaker with 50 mL of methanol and place no more 

than four of the LVE Kit lines into the beaker.

3.  Apply vacuum to the manifold and pull the methanol through 

the positions into the waste container.

4.  Remove the column and discard.
5.  Using methanol in a squeeze bottle, clean the exterior of the 

LVE Kit’s lines, the column adapters, the stopcock, and the 
metal cannula. Discard all rinsate.

6.  Repeat this up to three times for all positions which  

require cleaning.

Note: In situations where the previous sample was highly 
concentrated, the above cleaning procedure may need to be 
repeated multiple times. If there is concern regarding potential 
carryover contamination regardless of the cleaning procedure, 
a laboratory reagent blank should be run in that position to 
ensure its cleanliness.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 9

Appendix B

MRM Transitions

Table 4. MRM Transitions for Agilent 6470 MS/MS. 

Cpd Name

ISTD?

Prec 

Ion

MS1 Res

Prod 

Ion

MS2 Res

Frag 

(V)

CE 

(V)

Cell Acc 

(V)

Ret Time 

(min)

Ret 

Window

Polarity

H4PFOS

No

427

Unit/Enh (6490)

406.8

Unit/Enh (6490)

125

24

5

13.3

1.37

Negative

H4PFOS

No

427

Unit/Enh (6490)

80.9

Unit/Enh (6490)

125

40

5

13.3

1.37

Negative

C3-PFHxS

No

402

Unit/Enh (6490)

80

Unit/Enh (6490)

100

49

5

11.7

1.5

Negative

C4-PFBA

No

217

Unit/Enh (6490)

172

Unit/Enh (6490)

60

8

5

5.1

1.45

Negative

C4-PFHpA

No

367

Unit/Enh (6490)

322

Unit/Enh (6490)

72

0

5

11.6

1.2

Negative

C5-PFHxA

No

318

Unit/Enh (6490)

273

Unit/Enh (6490)

70

8

5

9.7

1.14

Negative

C5-PFPeA

No

268

Unit/Enh (6490)

223

Unit/Enh (6490)

60

20

5

7.4

1.55

Negative

C6-PFDA

No

519

Unit/Enh (6490)

474

Unit/Enh (6490)

81

4

5

16.2

1.65

Negative

C8-PFOA

No

421

Unit/Enh (6490)

376

Unit/Enh (6490)

80

8

5

13.4

1.38

Negative

C8-PFOS

No

507

Unit/Enh (6490)

80

Unit/Enh (6490)

100

50

5

15

1.53

Negative

C9-PFNA

No

472

Unit/Enh (6490)

427

Unit/Enh (6490)

66

4

5

14.9

1.52

Negative

PFBA

No

213

Unit/Enh (6490)

168.9

Unit/Enh (6490)

60

8

5

5.1

1.48

Negative

PFBS

No

298.9

Unit/Enh (6490)

98.9

Unit/Enh (6490)

100

29

5

7.9

1.41

Negative

PFBS

No

298.9

Unit/Enh (6490)

80

Unit/Enh (6490)

100

45

5

7.9

1.41

Negative

PFDA

No

513

Unit/Enh (6490)

469

Unit/Enh (6490)

81

4

5

16.2

1.65

Negative

PFDA

No

513

Unit/Enh (6490)

218.7

Unit/Enh (6490)

100

16

5

16.2

1.65

Negative

PFDoA

No

613

Unit/Enh (6490)

569

Unit/Enh (6490)

79

5

5

18.2

1.84

Negative

PFDoA

No

613

Unit/Enh (6490)

268.7

Unit/Enh (6490)

100

20

5

18.2

1.84

Negative

PFDS

No

599

Unit/Enh (6490)

99

Unit/Enh (6490)

100

40

5

17.15

1.75

Negative

PFDS

No

599

Unit/Enh (6490)

80

Unit/Enh (6490)

100

40

5

17.15

1.75

Negative

PFHpA

No

362.9

Unit/Enh (6490)

319

Unit/Enh (6490)

72

0

5

11.6

1.2

Negative

PFHpA

No

362.9

Unit/Enh (6490)

169

Unit/Enh (6490)

72

12

5

11.6

1.2

Negative

PFHpS

No

448.9

Unit/Enh (6490)

98.7

Unit/Enh (6490)

100

44

5

13.5

1.39

Negative

PFHpS

No

448.9

Unit/Enh (6490)

79.7

Unit/Enh (6490)

100

52

5

13.5

1.39

Negative

PFHxA

No

313

Unit/Enh (6490)

268.9

Unit/Enh (6490)

70

8

5

9.7

1.12

Negative

PFHxA

No

313

Unit/Enh (6490)

119

Unit/Enh (6490)

70

18

5

9.7

1.12

Negative

PFHxS

No

398.9

Unit/Enh (6490)

99

Unit/Enh (6490)

100

45

5

11.7

1.5

Negative

PFHxS

No

398.9

Unit/Enh (6490)

80

Unit/Enh (6490)

100

49

5

11.7

1.5

Negative

PFNA

No

463

Unit/Enh (6490)

419

Unit/Enh (6490)

66

4

5

14.9

1.52

Negative

PFNA

No

463

Unit/Enh (6490)

219

Unit/Enh (6490)

66

17

5

14.9

1.52

Negative

PFOA

No

413

Unit/Enh (6490)

369

Unit/Enh (6490)

69

4

5

13.4

1.38

Negative

PFOA

No

413

Unit/Enh (6490)

169

Unit/Enh (6490)

69

12

5

13.4

1.38

Negative

PFOS

No

498.9

Unit/Enh (6490)

99

Unit/Enh (6490)

100

50

5

15

1.53

Negative

PFOS

No

498.9

Unit/Enh (6490)

80

Unit/Enh (6490)

100

50

5

15

1.53

Negative

PFPeA

No

263

Unit/Enh (6490)

218.9

Unit/Enh (6490)

60

8

5

7.4

1.77

Negative

PFUnA

No

563

Unit/Enh (6490)

519

Unit/Enh (6490)

73

5

5

17.2

1.75

Negative

PFUnA

No

563

Unit/Enh (6490)

269

Unit/Enh (6490)

100

20

5

17.2

1.75

Negative

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 10

Appendix C

Calibration Curves

Figure 9. Calibration curves for the target analytes in Table 1, covering a 

concentration range of 0.2-20 ppt.

PFBA

PFHpS

H4PFOS

PFOA

PFOS

PFPeA

PFHxA

PFHpA

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 11

PFNA

PFDA

PFDS

PFUnA

PFDoA

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 12

Appendix D

MRL and DL Data

Table 6. MRL and DL Recoveries for EVOLUTE® PFAS 150 mg/6 mL columns.

Conc. 

(ng/L)

1 

(ng/L)

2 

(ng/L)

3 

(ng/L)

4 

(ng/L)

5 

(ng/L)

6 

(ng/L)

7 

(ng/L)

8

(ng/L)

x̅

(ng/L)

x̅  

(%)

s 

(ng/L)

CV 

(%)

DL 

(ng/L)

PFBA*

1.77

1.88

1.83

1.87

1.94

1.83

1.86

1.79

1.83

1.85

104

0.05

2.56

0.14

PFPeA

2.00

1.96

1.92

1.96

2.05

1.97

1.93

1.86

1.96

1.95

98

0.05

2.79

0.16

PFBS

2.00

1.87

1.76

1.84

1.73

1.77

1.81

1.73

1.75

1.78

89

0.05

2.83

0.15

PFHxA

2.00

2.09

2.00

2.04

2.10

2.03

2.09

1.98

2.08

2.05

103

0.05

2.20

0.14

PFHpA

2.00

2.08

1.96

1.98

1.91

2.04

1.95

2.01

2.01

1.99

100

0.06

2.76

0.17

PFHxS*

1.90

1.75

1.83

1.80

1.85

1.66

1.74

1.77

1.74

1.77

93

0.06

3.34

0.18

PFHpS*

1.91

1.77

1.78

1.85

1.79

1.64

1.66

1.69

1.64

1.73

91

0.08

4.69

0.24

H4PFOS*

1.90

1.91

1.84

1.97

1.92

2.05

2.06

2.01

2.11

1.98

104

0.09

4.58

0.27

PFOA

2.00

1.96

1.99

2.01

2.08

1.97

2.05

1.91

1.97

1.99

100

0.05

2.59

0.15

PFOS*

1.92

2.00

1.89

1.93

2.01

1.95

1.95

1.98

1.96

1.96

102

0.04

1.99

0.12

PFNA

2.00

2.08

1.94

2.02

2.01

2.04

2.08

2.11

2.02

2.03

102

0.05

2.59

0.16

PFDA

2.00

1.99

1.89

1.96

1.99

1.91

2.03

2.07

2.04

1.98

99

0.06

3.12

0.19

PFDS

2.00

1.92

2.08

1.81

2.04

2.01

1.93

2.04

1.90

1.97

98

0.09

4.61

0.27

PFUnA

2.00

2.09

1.97

1.93

1.95

1.89

1.97

1.95

1.98

1.97

98

0.06

2.94

0.17

PFDoA

2.00

2.08

1.91

2.02

2.05

2.02

2.03

2.04

2.04

2.02

101

0.05

2.52

0.15

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

Table 5. MRL and DL Recoveries for EVOLUTE® PFAS 500 mg/6 mL columns.

Conc. 

(ng/L)

1 

(ng/L)

2 

(ng/L)

3 

(ng/L)

4 

(ng/L)

5 

(ng/L)

6 

(ng/L)

7 

(ng/L)

x̅

(ng/L)

x̅  

(%)

s 

(ng/L)

CV 

(%)

DL 

(ng/L)

PFBA*

1.77

1.67

1.71

1.67

1.68

1.81

1.60

1.80

1.71

96

0.08

4.43

0.24

PFPeA

2.00

1.85

1.83

1.92

1.97

1.88

1.75

1.91

1.87

94

0.07

3.81

0.22

PFBS

2.00

1.73

1.90

1.82

1.70

1.73

1.65

1.95

1.78

89

0.11

6.12

0.34

PFHxA

2.00

1.80

1.92

1.89

1.80

1.98

1.75

1.92

1.87

93

0.08

4.50

0.26

PFHpA

2.00

1.96

2.06

2.04

1.94

2.01

1.80

2.08

1.99

99

0.09

4.75

0.30

PFHxS*

1.90

1.85

1.78

1.77

1.70

1.82

1.73

1.77

1.77

94

0.05

2.84

0.16

PFHpS*

1.91

1.90

1.90

1.86

1.65

1.92

1.51

1.73

1.78

93

0.16

8.79

0.49

H4PFOS*

1.90

1.80

1.87

1.88

1.86

1.87

1.68

1.86

1.83

96

0.07

4.02

0.23

PFOA

2.00

1.80

1.86

1.93

1.91

1.94

1.67

1.94

1.86

93

0.10

5.34

0.31

PFOS*

1.92

1.96

1.89

1.79

1.89

1.92

1.69

1.89

1.86

97

0.09

4.98

0.29

PFNA

2.00

1.88

1.92

1.95

1.90

1.94

1.79

2.07

1.92

96

0.08

4.30

0.26

PFDA

2.00

1.87

1.89

1.87

1.83

2.01

1.79

2.01

1.89

95

0.09

4.55

0.27

PFDS

2.00

2.10

2.06

1.95

2.14

1.70

1.64

1.93

1.93

97

0.19

9.98

0.61

PFUnA

2.00

1.89

1.91

1.90

1.81

1.88

1.75

1.94

1.87

93

0.07

3.51

0.21

PFDoA

2.00

1.85

1.96

1.96

1.81

2.00

1.75

1.96

1.90

95

0.10

5.04

0.30

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 13

Appendix E

PFAS Background Study

Table 7. Results of PFAS Background Study for EVOLUTE® PFAS 500 mg/6 mL columns (recoveries in ng/L).

TurboVap® LV

Laboratory Reagent Blanks

Replicate

1

2

3

4

5

6

7

8

1

2

3

4

PFBA*

0.00

0.00

0.00

0.00

0.00

0.10

0.05

0.04

0.09

0.06

0.06

0.06

PFPeA

0.02

0.03

0.03

0.03

0.03

0.06

0.02

0.03

0.04

0.04

0.02

0.04

PFBS

0.01

0.00

0.00

0.00

0.01

0.00

0.01

0.01

0.00

0.00

0.00

0.06

PFHxA

0.01

0.02

0.00

0.01

0.00

0.02

0.01

0.02

0.02

0.02

0.02

0.02

PFHpA

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

PFHxS*

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.01

0.00

0.01

0.01

0.01

PFHpS*

0.00

0.02

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

H4PFOS*

0.02

0.00

0.00

0.00

0.00

0.00

0.00

0.01

0.05

0.06

0.07

0.07

PFOA

0.02

0.00

0.00

0.00

0.03

0.00

0.01

0.00

0.00

0.00

0.00

0.00

PFOS*

0.00

0.00

0.00

0.00

0.03

0.00

0.00

0.02

0.01

0.02

0.01

0.01

PFNA

0.00

0.00

0.00

0.01

0.00

0.00

0.00

0.00

0.02

0.00

0.01

0.00

PFDA

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.02

0.00

0.00

0.00

0.00

PFDS

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

PFUnA

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

PFDoA

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

Table 8. Results of PFAS Background Study for EVOLUTE® PFAS 150 mg/6 mL columns (recoveries in ng/L).

TurboVap® LV

Laboratory Reagent Blanks

Replicate

1

2

3

4

1

2

3

4

PFBA*

0.06

0.02

0.01

0.00

0.11

0.09

0.09

0.10

PFPeA

0.03

0.03

0.03

0.02

0.05

0.06

0.04

0.04

PFBS

0.01

0.01

0.00

0.00

0.00

0.01

0.01

0.01

PFHxA

0.02

0.02

0.01

0.00

0.03

0.03

0.02

0.02

PFHpA

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

PFHxS*

0.01

0.01

0.00

0.00

0.01

0.01

0.01

0.01

PFHpS*

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

H4PFOS*

0.01

0.00

0.00

0.00

0.03

0.03

0.04

0.02

PFOA

0.00

0.02

0.03

0.00

0.00

0.00

0.05

0.00

PFOS*

0.00

0.00

0.01

0.01

0.00

0.00

0.00

0.00

PFNA

0.00

0.01

0.00

0.00

0.00

0.00

0.01

0.00

PFDA

0.00

0.02

0.00

0.00

0.00

0.00

0.00

0.02

PFDS

0.00

0.00

0.01

0.00

0.00

0.00

0.00

0.00

PFUnA

0.00

0.00

0.00

0.00

0.02

0.00

0.00

0.00

PFDoA

0.00

0.06

0.00

0.00

0.00

0.00

0.00

0.00

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 14

Appendix F

IDP and IDA Data

Table 9. Results of IDP and IDA for EVOLUTE® PFAS 500 mg/6 mL columns (15 ng/L, n=4).

Replicate

1 (%)

2 (%)

3 (%)

4 (%)

x̅  (%)

s (%)

CV (%)

PFBA*

93.97

93.52

97.66

94.56

94.93

1.87

1.97

PFPeA

91.10

99.84

99.34

92.16

95.61

4.62

4.83

PFBS

88.73

88.20

91.76

86.48

88.79

2.20

2.47

PFHxA

94.83

95.12

99.13

95.85

96.23

1.98

2.06

PFHpA

104.38

106.43

104.54

106.18

105.38

1.07

1.02

PFHxS*

95.20

99.75

101.57

95.96

98.12

3.04

3.10

PFHpS*

89.31

87.18

98.22

79.57

88.57

7.67

8.66

H4PFOS*

96.43

102.56

100.83

101.53

100.34

2.70

2.69

PFOA

94.28

94.77

96.59

92.79

94.61

1.56

1.65

PFOS*

91.58

93.17

98.48

89.66

93.22

3.79

4.06

PFNA

93.76

98.27

100.96

96.89

97.47

2.99

3.07

PFDA

96.19

91.59

101.20

100.51

97.38

4.45

4.57

PFDS

96.50

102.88

108.47

98.56

101.60

5.29

5.21

PFUnA

88.89

85.05

87.96

90.74

88.16

2.37

2.69

PFDoA

96.65

99.00

102.74

98.79

99.30

2.53

2.55

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

Table 10. Results of IDP and IDA for EVOLUTE® PFAS 150 mg/6 mL columns (15 ng/L, n=4).

Replicate

1 (%)

2 (%)

3 (%)

4 (%)

x̅  (%)

s (%)

CV (%)

PFBA*

102.37

101.69

101.80

103.26

102.28

0.72

0.70

PFPeA

101.89

102.16

101.32

100.30

101.42

0.82

0.81

PFBS

99.30

102.15

99.56

102.63

100.91

1.72

1.71

PFHxA

100.58

103.25

99.56

100.27

100.92

1.61

1.60

PFHpA

102.78

102.88

102.64

102.10

102.60

0.35

0.34

PFHxS*

97.07

98.52

98.87

97.40

97.96

0.86

0.88

PFHpS*

91.57

92.08

90.92

93.18

91.94

0.95

1.04

H4PFOS*

99.40

99.12

104.89

102.46

101.47

2.74

2.70

PFOA

104.83

105.70

104.59

106.65

105.44

0.94

0.89

PFOS*

101.18

103.31

101.24

102.55

102.07

1.04

1.02

PFNA

102.52

103.28

101.34

101.57

102.18

0.90

0.88

PFDA

102.85

104.63

101.29

103.65

103.10

1.41

1.37

PFDS

105.46

102.18

103.54

106.38

104.39

1.89

1.81

PFUnA

98.55

97.67

97.27

98.90

98.10

0.76

0.77

PFDoA

105.74

103.29

103.54

105.36

104.48

1.25

1.19

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.

© Biotage 2022

Manual Extraction of PFAS in Drinking Water Following DIN 38407-42 | Page 15

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Appendix G

Carryover Data

Table 11. Results of carryover study following four, 50 ng/L LFB samples using EVOLUTE® PFAS 500 mg/6 mL columns.

Replicate

1 

(ng/L)

2 

(ng/L)

3 

(ng/L)

4 

(ng/L)

x̅  

(ng/L)

PFBA*

0.11

0.09

0.09

0.09

0.09

PFPeA

0.04

0.04

0.04

0.04

0.04

PFBS

0.02

0.01

0.02

0.01

0.02

PFHxA

0.02

0.02

0.03

0.03

0.03

PFHpA

0.00

0.00

0.00

0.00

0.00

PFHxS*

0.02

0.00

0.02

0.01

0.01

PFHpS*

0.00

0.00

0.00

0.00

0.00

H4PFOS*

0.04

0.04

0.04

0.04

0.04

PFOA

0.09

0.04

0.07

0.00

0.05

PFOS*

0.04

0.03

0.04

0.04

0.04

PFNA

0.00

0.00

0.00

0.00

0.00

PFDA

0.02

0.02

0.02

0.02

0.02

PFDS

0.03

0.00

0.00

0.00

0.01

PFUnA

0.00

0.00

0.03

0.01

0.01

PFDoA

0.00

0.00

0.00

0.00

0.00

*Analytes were used in salt form and calculated concentrations were corrected to compensate where needed.