Baltic Marine Environment Protection CommissionHELCOM EXPERT NETWORK ON HAZARDOUS SUBSTANCES (EN-HZ)

Document prepared by HELCOM EXPERT NETWORK ON HAZARDOUS SUBSTANCES (EN-HZ) 2019

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Technical annex on the determination of Perfluoroalkylated substances (PFAS) in biota

1. Background

Perfluorooctane sulphonate (PFOS), perfluoro octanoic acid (PFOA) and other perfluorinated compounds are considered global environmental contaminants. PFOS and PFOA are chemically and biologically inert and very stable (Poulsen et al. 2005). PFOS meets the P (Persistent) and vP (very Persistent) criteria due to slow degradation. PFOS is also bioaccumulative (B) and toxic (T) (OSPAR 2005). PFOA is considered as very persistent (vP) and toxic (T), but not bioaccumulative (Van der Putte et al. 2010). It has a capacity to undergo long-range transportation. These guidelines concentrate on the sampling and extraction of perfluoroalkylated substances from seawater and also address special aspects of the sampling matrix. These guidelines provide advice for the analysis of perfluoroalkylated substances (PFAS) in biota which basically includes the following steps:

• sampling;

• pretreatment;

• analytical determination.

The extraction of the PFAS enables enrichment of the analytes which is a crucial step in the procedure as the concentrations are low and biota matrix complicated. Extraction techniques vary, but determination is usually performed by separation with high performance liquid chromatography (HPLC) and mass spectrometric (MS) detection. All the steps of the procedure are susceptible to insufficient recovery and contamination. Therefore, regular quality control measures must be applied in order to monitor method performance. These guidelines are intended to encourage and assist analytical chemists to critically reconsider and improve established methods and associated quality control measures, where necessary.These guidelines are not intended as complete laboratory manual. If necessary, guidance should be sought from specialized laboratories. Laboratories should demonstrate validity of each methodological step. Moreover, use of an alternative method, carried out concurrently to the routine procedure, is recommended for validation. The participation in analytical proficiency tests is also highly recommended.

1.1 IntroductionPFOS related substances and PFOA are members of the larger family of perfluoroalkylated substances (PFAS). In the past, PFASs were often referred to as “PFCs” (per- and polyfluorinated chemicals), but this term “PFCs” can also be understood as perfluorocarbons, which contain only carbon and fluorine, therefore, a term PFAS would be more proper to use.OECD has updated the list of PFAS in 2017: “ “Lists of PFOS, PFAS, PFOA, PFCA, Related Compounds and Chemicals That May Degrade to PFCA”. In total, 4730 PFAS-related CAS numbers have been identified and manually categorised in that study, including several new groups of PFASs that fulfil the common definition of PFASs (i.e. they contain at least one perfluoroalkyl moiety) but have not yet been commonly regarded as PFASs. In particular, it focuses on those

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PFASs, including perfluorocarbons, that contain a perfluoroalkyl moiety with three or more carbons (i.e. –CnF2n–, n ≥ 3) or a perfluoroalkylether moiety with two or more carbons (i.e. –CnF2nOCmF2m−, n and m ≥ 1) (OECD 20181).However, the identified PFASs are diverse in terms of structure and other categorization elements. The categorization is challenging, as well as the terminology related to PFASs and also many of the compounds are not commercially available. Perfluorooctyl sulfonate compounds are all derivatives of PFOS and can degrade to PFOS, also called as PFOS-related compounds. Some 100–200 PFOS-related compounds have been identified (KEMI 2006). The Swedish Chemicals Agency (KEMI) estimated that there are 3000 available PFAS-substances on the global market(KEMI2015).

Table 1 provides an overview of PFCAS that are environmentally relevant in water and biota and provides information on chemical names, acronyms, formula, and Chemical Abstracts Service (CAS) numbers.

Table 1. List of possible perfluoroalkylated substances monitored.ANALYTE ACRONYM FORMULA CAS-

NUMBER

Perfluorobutanate PFBA C3F7COO- 375-22-4

Perfluoropentanate PFPeA C4F9COO- 2706-90-3

Perfluorohexanate PFHxA C5F11COO- 307-24-4

Perfluoroheptanate PFHpA C6F13COO- 375-85-9

Perfluorooctanate PFOA C7F15COO- 335-67-1

Perfluorononanate PFNA C8F17COO- 375-95-1

Perfluorodecanate PFDA C9F19COO- 335-76-2

1 TOWARD A NEW COMPREHENSIVE GLOBAL DATABASE OF PER- AND POLYFLUOROALKYL SUBSTANCES (PFASs): SUMMARY REPORT ON UPDATING THE OECD 2007 LIST OF PER- AND POLYFLUOROALKYL SUBSTANCES (PFASs) Series on Risk Management No. 39http://www.oecd.org/officialdocuments/publicdisplaydocumentpdf/?cote=ENV-JM-MONO(2018)7&doclanguage=en

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Perfluorobutane sulfonate PFBS C4F9SO2O- 29420-49-3

Perfluorohexane sulfonate PFHxS C6F13SO2O- 3871-99-6 (potassium salt)

Perfluorooctane sulfonate PFOS C8F17SO2O- 2795-39-3

(potassium salt)

6:2 fluorotelomer sulfonate 6:2 FtS (THPFOS)

C6F13C2H4SO3- 27619-97-2

Perfluorooctane sulfonamide PFOSA (same as FOSA)

C8F17SO2NH2 754-91-6

Perfluorododecanoic acid PFDoA C12HF23O2 307-55-1

Perfluoroundecanoic acid PFUA C11HF21O2 2058-94-8

Over the past decades, regulatory agencies and researchers have mainly focused on long chain perfluoroalkyl carboxylic acids (PFCAs) and perfluoroalkyl sulfonic acids (PFSAs), because of their higher bioaccumulative potential compounds. And the main attention has been on PFOA and PFOS. However, as a result of their potential toxicity to the environment and humans, the production and sale of long-chained PFASs have been phased out (due to banning also). Although environmental concentrations of these long-chained compounds are decreasing, concentrations of other PFASs will be rising and illustrating the importance of environmental monitoring of PFASs. Based on the literature overview, most commonly accumulated PFAS in biota with high concentrations include: PFOS, PFDA, PFUnDA, (P)FOSA, PFNA, PFHxS. In addition, accumulation of PFOA, PFDoDA, PFTeDA, PFBS and PFDS has also been observed.For monitoring purposes, the chemicals PFOS and PFOA are considered to be the most important PFASs. Although most studies have focused on PFOS and PFOA, it is suggested that PFAS with longer and shorter carbon chain lengths should also be included in the analysis. Long chain PFAS‐ (≥ C8) should be included because of their potential to bioaccumulate and to adsorb onto sediment. Perfluorobutane sulfonate (PFBS) and perfluorobutanate (PFBA), the short chain‐ substitutes of PFOS and PFOA, should preferentially be monitored in water as they may bioaccumulate to a lesser extent.Under the Third Unregulated Contaminant Monitoring Rule (UCMR3), EPA collected data for chemicals that are suspected contaminants in drinking water but do not have health-based standards set under the SDWA. In the UCMR3, there were six PFAS included for monitoring including: perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), perfluorohexanesulfonic acid (PFHxS), perfluoroheptanoic acid (PFHpA) and perfluorobutanesulfonic acid (PFBS).

1.2 Purpose and aims

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The aim of monitoring is to identify spatial variations and temporal trends of perfluoroalkylated substances in biota. The presence of PFAS in biota samples provides information on the contaminant load of the Baltic Sea. To provide information for the core indicator “Perfluorooctane sulphonate (PFOS)”.

Good status is achieved when the concentration of perfluorooctane sulphonate (PFOS) in fish muscle is below 9.1 µg/ kg fish wet weight. The threshold value is an environmental quality standard (EQS), derived at EU level as a substance included on the list of priority substances under the Water Framework Directive (2000/60/EC and 2013/39/EC). Good environmental status within the MSFD is defined a ‘concentrations of contaminants at levels not giving rise to pollution effects'. EQSs are derived from ecotoxicological studies to protect freshwater and marine ecosystems from potential adverse effects of chemicals, as well as adverse effects on human health via drinking water and food from aquatic environments. (2018 Helcom core indicator report2).

2. Monitoring methods2.1 Monitoring features The main focus of monitoring will be @@.The PFCs can bind to the plasma protein serum albumin and accumulate in blood and organs (Jones et al., 2003). Accumulation in the marine foodweb starts from the bottom of the food chain, with invertebrates such as zooplankton and molluscs, followed by crustaceans, and then fish (Van de Vijver et al., 2003). The highest PFC concentrations have been found in blood and internal organs (e.g. liver, kidney) of top predators, such as marine mammals (Martin et al., 2004b; Ahrens et al., 2009c) and fish eating birds‐ (Kannan et al., 2001). Fish species widely used for monitoring of organic contaminants are also suitable for PFAS analysis. Liver is the first choice of tissue for monitoring purposes but PFAS have been also detected in other organs (e.g. blood and muscle). (ICES, 2010)

2.2 Time and areaMonitoring covers the entire Baltic area, and is performed throughout the year.

2.3 Monitoring procedure 2.3.2 Sampling method and equipment

The sampling should be carried out by trained personnel who are aware of the risk of contamination of samples posed by incorrect handling. When analyzing PFASs from biologically active samples, it is highly recommended to prepare the samples for analysis as soon as possible after sampling without further manipulation or store them in closed containers below -20 °C until sample preparation. Moreover, samples should not be transferred into further containers before extraction. Handling time at room temperature should be minimal in order to prevent the possible degradation (Rhoads et al., 2008).Homogenization of the samples is necessary and a homogenizator with non-fluorinated plastic dispersing components (e.g. polycarbonate and polysulfone) should be used in order to obtain samples free of contamination. Samples can be homogenized and then kept below -20 °C until extraction.Materials and clothes that contain, or may adsorb, fluorinated compounds, such as polytetrafluoroethylene (PTFE), must be avoided during sampling and sample processing. In particular, the containers that come into direct contact with the sample must not contain any fluorinated polymers (e.g. PTFE). Containers and equipment made of polypropylene (according to

2 http://www.helcom.fi/Core%20Indicators/Perfluorooctane%20sulphonate%20PFOS%20HELCOM%20core%20indicator%202018.pdf

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ISO 25101:2009), polyethylene, glass, or stainless steel should be used. However, especially glass and polypropylene sampling and storage containers should be carefully checked for PFAS, as longer chained PFASs tend to adsorb to glass walls, and cleaned before use (e.g. the sample container or filtration equipment must be rinsed with a polar solvent, such as methanol and dried before use) in order to minimize contamination. Every material that may come into contact with the sample must be free of fluorinated compounds. The highest contamination risk was observed for PFOA and perfluorononanate (PFNA; Theobald et al., 2007). Dissection of samples should be carried out with stainless-steel tools

2.3.3 Sample handling and analysisThe analysis of PFAS in biota generally includes extraction with organic solvents, clean up, and‐ liquid chromatography (LC) with mass spectrometric (MS) detection.Spiking with internal standardsSamples should be spiked with internal standards (see Table 2) before extraction, at concentrations close to the environmental level, in order to correct for losses during extraction, extract clean up and concentration, and for matrix effects during analysis. It is advisable that there‐ is a corresponding internal standard for each of the PFAS as the recoveries and matrix effects differentiate between analytes. After spiking and before extraction, in case of biota samples, these should be left to equilibrate for approximately 30 minutes to 1 hour at 4 °C (Kaupmees, 2017). Isotopically labelled internal standards help to take into account recovery as well as matrix effects.However, there might not be internal standards available for all analytes of interest, Groffen et al 3 found that using the ISTD closest in terms of functional group and carbon chain length is a suitable method for the quantification of PFASs that lack a corresponding ISTD.

Table 2. Possible internal standards for PFAS analysis.INTERNAL STANDARDS

Perfluoro-n-(1,2,3,4-13C4)butanate [13C4]-PFBA (2,3,4-13C3)F713COO- n.a.

Perfluoro-n-(1,2-13C2)hexanate [13C2]-PFHxA C4F9(2-13C)F213COO- n.a.

Perfluoro-n-(1,2,3,4-13C4)octanate [13C4]-PFOA C4F9(2,3,4-13C3)F613COO- n.a.

Perfluoro-n-(1,2,3,4,5,6,7,8-13C8)octanate

[13C8]-PFOA (2,3,4,5,6,7,8-13C7)F1513COO-

n.a.

Perfluoro-n-(1,2,3,4,5-13C5)nonanate

[13C5]-PFNA C4F9(2,3,4,5-13C4)F813COO-

n.a.

Perfluoro-n-(1,2-13C2)decanate [13C2]-PFDA C8F1713CF213COO- n.a.

Perfluoro-n-(1,2-13C2)undecanate [13C2]-PFUnDA

C9F1913CF213COO- n.a.

Perfluoro-n-(1,2-13C2)dodecanate [13C2]-PFDoDA C10F2113CF213COO- n.a.

Perfluoro-1-hexane(18O2)sulfonate [18O2]-PFHxS C6F13S(18O2]O- n.a.

3

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Perfluoro-1-(1,2,3,4-13C4)octanesulfonate

[13C4]-PFOS C4F9(1,2,3,4-13C4)F8SO2O-

n.a.

Perfluoro-1-(1,2,3,4,5,6,7,8-13C8)octanesulfonate

[13C8]-PFOS (1,2,3,4,5,6,7,8-13C8)F17SO2O

n.a.

Perfluoro-1-(1,2,3,4,5,6,7,8-13C8)octanesulfonamide

[13C8]-PFOSA (1,2,3,4,5,6,7,8-13C8)F17SO2NH2

n.a.

N-deuterioethylperfluoro-1-octanesulfonamidoacetic acid

d5-N-EtFOSAA C8F17SO2N(C2D5)CH2CO2H

n.a.

Blanks and contaminationSince concentrations of PFAS in samples can be very low, in order to minimize the risk of sample contamination, sample treatment and processing should be carried out on a clean bench or in a clean room containing no fluorinated compounds (e.g. PTFE) and it is strongly recommended to pretreat all used glassware and adsorption material, rinsing with HPLC grade water and methanol has been used before (Kaupmees, 2017). This also includes washing syringes and syringe filters used for filtration of extracts prior chromatographic analysis. Moreover, due to low concentration levels, it might be necessary to purchase water for cleaning and preparation of standard solutions, as the inhouse purified water might contain PFAS. This should be checked for. Moreover, when using a commercial bottled water, this should also be tested for PFAS and additionally, a dispenser should be used so that constant opening and closing of the water bottle would not contaminate the water. Again, the use of dispenser must be checked for PFAS.However, due to the low PFAS concentrations and significant handling during sample preparation, every sample batch should include a procedural blank that has been prepared in the same way as the samples. If measurable blanks occur, the analytical instrumentation and every sample preparation step must be checked for contamination and appropriate measures taken before continuation of analysis. Solvents, including ultrapure water and the internal standard spiking solution, should be of high purity and must be tested for contamination prior to use. If the sources of contamination is known and constant for all the samples, the blank concentrations can be subtracted from the samples. This can be done only if the blank concentrations are constant and tested during the validation procedure.Extraction and clean-upThree methods are commonly used for the extraction of PFCs from biota samples.

1 ) ion pair extraction with tetrabutylammonium (TBA) and the extraction solvent methyl tert butyl ether (MTBE; Hansen ‐ et al., 2001) 2) ultrasonic extraction (UE) with subsequent clean up (Powley ‐ et al., 2005)

It is advisable to keep sample preparation as simple as possible in order to avoid any contamination. One of the oldest and widely employed PFAS sample preparation methods for biota, is based on ion pairing of PFAS with tetrabutylammonium (TBA), followed by liquid-solid extraction (LSE) with methyl-tert-butylether (MTBE), often referred as IPE. Contrary to several other methods, the IPE method avoids contamination by skipping the clean-up step. The matrix-effects of a dirtier extract are taken into account by using IS-s (Kaupmees, 2017). The UE method includes a minimum of three repeated extractions using acetonitrile or methanol, each with a tenfold solvent of the sample amount and 30 min extraction time. The three extracts

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are combined for clean up. Following extraction, a clean up of the extracts is necessary to prevent‐ ‐ matrix effects when carrying out instrumental analysis. Lipids can be removed from methanol or acetonitrile extracts by precipitation at − 20 °C. The extract is then centrifuged for 1 min and the supernatant is decanted into a clean vial. After lipid removal, additional clean up steps may be‐ required, depending on sample type and analyte concentrations (as described by Powley et al., 2005). (ICES, 2010)Eluates are usually subjected to an additional clean-up with an solid phase extraction (SPE) cartridge containing HLB, WAX or ENVI-Carb resin4 and such as by So et al (So et al., 20065), but with such methods analysis of many PFAS is compromised due to blank interferences eg procedural blanks have high concentration of the analytes and this results in poor limits of quantifications. Therefore, methods containing any type of SPE should be avoided due to high risk of contamination.Since there isn’t one universal and standardised method, but there is a need for a good analysis, new analysis methods for PFAS in complex matrices are in constant development and published. Therefore, it is recommended to look for new methods in peer-reviewed articles. For example there is a new QuEChERS application for PFAS analysis in fish with using styrene-divinylbenzene beads. This new technique discards the traditional SPE cartridge but has the advantage of removing more interferences than simple extraction when applying an additional sorbent.6

Sample extracts should be concentrated in order to meet the required detection limits. Concentration techniques at low temperature (< 40 °C) and controlled pressure conditions are preferred in order to avoid losses of volatile PFCs. Evaporation to dryness should be avoided.Before analysis, extracts have to be filtered and it has been shown (So 2006; Kaupmees 2017) syringe filters contain trace amounts of PFAS. Therefore, it is best to wash these with water and methanol prior to analysis. Instrumental analysisLiquid chromatography coupled with a tandem mass spectrometer and interfaced with an electrospray ionization source in negative ion mode (LC/(−)ESI MS/MS; Hansen ‐ ‐ et al., 2001) and LC coupled with an ESI quadrupole time of flight mass spectrometer (LC/ESI QTOF MS) have both‐ ‐ ‐ ‐ been used for PFAS analysis (Berger and Haukas, 2005). Tandem MS and QTOF MS have the‐ advantage of providing low signal to noise ratio and high selectivity on low concentrations. ‐ ‐The solvent composition of the final extract should correspond to the mobile phase of the LC method in order to obtain a satisfactory peak shape of the compounds, in particular of short chain‐ PFCs eluting early from the column. If necessary, sonication of the samples can be used for better dissolving and extracts should be filtered before analysis. Unless the samples are analysed immediately, the vials should be kept at a temperature below 4 °C. If glass vials are used, PFCs can be adsorbed onto the glass surface with solvents containing a greater amount of water. It is therefore advisable to use polypropylene vials.Chromatographic determinationThe C8 or C18 reversed phase columns may be used for the LC separation of PFAS. The use of a‐ guard column is recommended in order to maintain chromatographic performance and extend the lifetime of the chromatographic column. To overcome separation problems (e.g. co eluting matrix‐ compounds), it may be helpful to use reversed phase columns with polar groups instead of C 8 or‐ C 18 columns. Mixtures of water and either methanol or acetonitrile can be used as the mobile phase, in each case with 2 – 10 mM ammonium acetate as an ionization aid. Gradients ranging

4

5

6

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from 10 % to 100 % methanol or acetonitrile are required for the separation of the compounds listed in Table 1. To ensure stability of retention times, the use of a temperature controlled‐ column oven is recommended. For solvents, use of PFC trapping column is highly recommended as it delays analyte peaks originating from eluents (Stone et al. 2010).Mass spectrometryThe most widely used technique for detection of PFAS in tandem MS (MS/MS) operated is multiple reaction monitoring (MRM) mode. Mass spectrometry parameters, such as collision energy, clustering potential, and cone voltage, must be optimized for each individual compound and each instrument. The sensitivity of MS/MS is usually approximately one order of magnitude higher than that of QTOF-MS (Berger et al., 2004).MS analysis suffers from matrix effects (signal suppression by co-eluting compounds) and to overcome this problem, quantitation can be done using isotope dilution or matrix-matched calibration curve. Loss of sensitivity caused by ionisation suppression results in an increase in non-detection. The most promising approach for minimising matrix effects is reducing the volume of the initial sample and the amount of extract injected.Co elution of matrix constituents (e.g. taurocholate bile salts) with the same transition ions as‐ perfluorohexane sulfonate (PFHxS) and PFOS may lead to a significant bias in the quantification of these compounds in biota samples. However, bias in PFHxS and PFOS levels can be avoided by separating the interferences from the target analytes and/or by using the interference free‐ transition to a mass to charge ratio (m/z) of 119 in order to verify results obtained with the‐ ‐ product ions at m/z 80 and 99. The PFCAs and PFSAs are almost completely dissociated in environmental matrices. If salts are used for the preparation of calibration standards, quantification results should be calculated for the corresponding acids. (ICAS, 2010)Limits of quantitation, limits of detectionRecent review on the methods reveals that methods have LoQ-s in the mid pg/g range.7

3. Data reporting and storageData is reported annually to the HELCOM COMBINE database, hosted by ICES. Data must be flagged as for HELCOM Combine.

4. Quality control4.1 Quality control of methodsQuality assuranceA number of measures should be taken to ensure sufficient quality of the analysis. Six main areas can be identified:

a) extraction efficiency and clean-up;

b) calibrant and calibration;

c) system performance;

d) long-term stability;7

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e) internal standards (Table 2); and

f) frequent participation in interlaboratory proficiency testing schemes and analysis of certified reference materials (CRM). At the moment only proficiency testing schemes for PFAS in sediment biota, or drinking water are offered, but not for seawater. Reference materials are available for PFAS in water and fish.

a) Extraction efficiency and clean-upThe use of internal standards is recommended in order to take any losses or matrix effects into account. For the determination of the recovery rates of the clean-up and concentration steps, it is recommended to pass a standard solution through the entire procedure. If major losses have occurred, the results should not be reported. In addition, blanks should be run.

b) Calibrant and calibrationCalibration solutions should be stored in ampoules at a cool, dark place. Weight loss during storage should be recorded for all standards. Risk of contamination during the storage must be monitored and suitable ampoules used. Moreover, due to the adsorption on the glass, it is recommended to keep stock solutions in the plastic vials and prepare calibration standards as needed.For PFAS determination calibration solutions from certified PFAS solutions should be used. Some PFSAs and sulfonamides demonstrate more than one peak in the chromatogram, owing to the presence of branched isomers. The ratio of linear and branched isomers can differ between the calibration standard and environmental samples. Branched isomers should be quantified separately if calibration standards are available. If there are no proper calibration standards and the peak area of the branched isomer exceeds 10 % of that of the linear isomer, it is advisable to estimate its concentration based on the response factor of the linear standard. However, the response factors of the linear and branched isomers may be different. It must be indicated with the results whether the reported concentration refers to the sum of the linear and branched isomers, or to the linear or branched isomer only. (ICAS, 2010) When using a PFOS standard solution of mixed isomers, it should be determined whether the nominal concentration corresponds to the linear PFOS or to the total amount of isomers (Kaupmees & Rebane, 2017).System performanceThe performance of the LC system can be monitored through regularly analyzing the resolution of two closely eluting compounds. A decrease in resolution indicates deteriorating LC conditions. The signal-to-noise ratio of a low concentrated standard gives information on the condition of the detector. For example, a dirty MS-source can be recognized by the presence of a higher background signal, together with a reduced signal-to-noise ratio.

c) Long-term stabilityOne laboratory reference sample should be included in each series of samples. A quality control chart should be recorded for selected PFAS compounds. If warning limits are exceeded, the method should be checked for possible errors and the obtained sample results should not be reported.

d) Internal standardsInternal standards should be added to all standards and samples either in a fixed volume or by weight and should not interfere with the target analytes. A number of deuterated PFAS compounds were proven to be suitable for LC-MS analysis (Table 2).

e) Duplicates, triplicatesThere are also applications where works use triplicates and duplicates.

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4.2 Quality control of data and reportingMeasurement uncertainty should be estimated using ISO 11352:2012 or according to the Nordtest report TR537. Estimation should be based on within-laboratory reproducibility, data from proficiency tests, IRM, and, when available, CRM.Data must be flagged if normal QA routines or recommended storage conditions cannot be followed.

5. Contacts and references5.1 Contact persons@@@5.2 ReferencesAhrens, L., Plassmann, M., Xie, Z., and Ebinghaus, R. 2009. Determination of polyfluoroalkyl compounds in water and suspended particulate matter in the River Elbe and North Sea, Germany. Frontiers of Environmental Science and Engineering in China, 3: 152 – 170.Arp, H. P. H., and Goss, K U. 2009. Gas/particle partitioning behavior of perfluorocarboxylic acids ‐with terrestrial aerosols. Environmental Science and Technology, 43: 8542 – 8547.Berger, U., and Haukas, M. 2005. Validation of a screening method based on liquid chromatography coupled to high resolution mass spectrometry for analysis of perfluoroalkylated ‐substances in biota. Journal of Chromatography A, 1081: 210 – 217.Berger, U., Langlois, I., Oehme, M., and Kallenborn, R. 2004. Comparison of three types of mass spectrometer for high performance liquid chromatography/mass spectrometry analysis of ‐perfluoroalkylated substances and fluorotelomer alcohols. European Journal of Mass Spectrometry, 10: 579 – 588.Huset, C. A., Chiaia, A. C., Barfosky, D. F., Jonkers, N., Kohler, H P. E., Ort, C., Giger, W., Field, J. A. ‐2008. Occurrence and mass flows of fluorochemicals in the Glatt Valley watershed, Switzerland. Environmental Science and Technology, 42: 6369 – 6377.ICES - Ahrens, L., Vorkamp, K., Lepom, P., Bersuder, P., Theobald, N., Ebinghaus, R., Bossi, R., Barber, J. L., McGovern, E. 2010. Determination of perfluoroalkyl compounds in water, sediment, and biota. ICES Techniques in Marine Environmental Sciences No. 48. 16 pp.ISO. 2009. Water quality – Determination of perfluorooctanesulfonate (PFOS) and perfluorooctanoate (PFOA) – Method for unfiltered samples using solid phase extraction and liquid chromatography/mass spectrometry. International Organization for Standardization, ISO 25101:2009 (TC 47/ SC 2). 19 pp.Kaupmees, S., Rebane, R. International Journal of Environmental Analytical Chemistry, 2017 Vol. 97, No. 8, 695–709https://doi.org/10.1080/03067319.2017.1356922 .KEMI (2006) Perfluorinated substances and their uses in Sweden. Swedish Chemical Agency (KEMI) Report 7/06. 58 pp.Moody, C. A., and Field, J. A. 1999. Determination of perfluorocarboxylates in groundwater impacted by fire fighting activity. Environmental Science and Technology, 33: 2800 – 2806. ‐OSPAR (2005) OSPAR background document on perfluorooctane sulphonate. 46 pp. OSPAR Commission. Updated in 2006.Poulsen, P., Jensen, A., Wallström, E. (2005) More environmentally friendly alternatives to PFOS-compounds and PFOA. Environmental Project No. 1013. Danish Environmental Protection Agency. 162 pp.

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Schultz, M. M., Barofsky, D. F., and Field, J. A. 2006. Quantitative determination of fluorinated alkyl substances by large volume injection liquid chromatography tandem mass spectrometry‐ ‐ ‐characterization of municipal wastewaters. Environmental Science and Technology, 40: 289 – 295.P.K.W. Stone, L. Cote, J. Gushue, R.J. Letcher and S. Chu, Agilent App. Note, 5990 (2010). Taniyasu, S., Kannan, K., Yeung, L. W. Y., Kwok, K. Y., Lam, P. K. S., and Yamashita, N. 2008. Analysis of trifluoroacetic acid and other short chain perfluorinated acids (C2 – C4) in precipitation by ‐liquid chromatography–tandem mass spectrometry: Comparison to patterns of long chain ‐perfluorinated acids (C5 – C18). Analytica Chimica Acta, 619: 221 – 230.Theobald, N., Gerwinski, W., Caliebe, C., and Haarich, M. 2007. Development and validation of a method for the determination of polyfluorinated organic substances in sea water, sediments and biota. Occurrence of these compounds in the North and Baltic Seas. Umweltforschungensplan des Bundesministeriums fur Umwelt, Naturschutz, und Reaktorsicherheit, Forschungsbericht 202 22 213, UBA FB 00 001409. 133 pp. (in German, with English abstract)‐Van der Putte, I., Murin, M., Van Velthoven, M., Affourtit, F. (2010) Analysis of the risks arising from the industrial use of Perfuorooctanoic acid (PFOA) and Ammonium Perfluorooctanoate (APFO) and from their use in consumer articles. Evaluation of the risk reduction measures for potential restrictions on the manufacture, placing on the market and use of PFOA and APFO. RPS Advies B.V. 82 pp. + annexes.van Leeuwen, S. P. J., Kärrman, A., Van Bavel, B., De Boer, J., and Lindström, G. 2006. Struggle for quality in determination of perfluorinated contaminants in environmental and human samples. Environmental Science and Technology, 40: 7854 – 7860.

5.3 Additional literature

Ahrens, L., Vorkamp, K., Lepom, P., Bersuder, P., Theobald, N., Ebinghaus, R., Bossi, R., Barber, J. L., McGovern, E. 2010. Determination of perfluoroalkyl compounds in water, sediment, and biota. ICES Techniques in Marine Environmental Sciences No. 48. 16 pp.

KEMI 2016a. Strategi för att minska användningen av högfluorerade ämnen, PFAS.

Stockholm: Kemikalieinspektionen. Rapport 9/16. ISSN 0284-1185

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