prEN ISO 20596-2
(Main)Water quality - Determination of cyclic volatile methylsiloxanes in water - Part 2: Method using liquid-liquid extraction with gas chromatography-mass spectrometry (GC-MS) (ISO 20596-2:2021)
Water quality - Determination of cyclic volatile methylsiloxanes in water - Part 2: Method using liquid-liquid extraction with gas chromatography-mass spectrometry (GC-MS) (ISO 20596-2:2021)
This document specifies a method for the determination of certain cyclic volatile methylsiloxanes (cVMS) in environmental water samples with low density polyethylene (LDPE) as a preservative and subsequent liquid-liquid extraction with hexane containing 13C-labeled cVMS as internal standards. The extract is then analysed by gas chromatography-mass spectrometry (GC-MS).
NOTE Using the 13C-labeled, chemically identical substances as internal standards with the same properties as the corresponding analytes, minimizes possible substance-specific discrimination in calibrations. Since these substances are least soluble in water, they are introduced via the extraction solvent hexane into the system.
Wasserbeschaffenheit - Bestimmung von cyclischen flüchtigen Methylsiloxanen in Wasser - Teil 2: Verfahren mittels Flüssig-Flüssig-Extraktion und Gaschromatographie-Massenspektrometrie (GC-MS) (ISO 20596‑2:2021)
Qualité de l'eau - Détermination de méthylsiloxanes cycliques volatiles dans l'eau - Partie 2: Méthode par extraction liquide-liquide avec chromatographie en phase gazeuse-spectrométrie de masse (CG-SM) (ISO 20596-2:2021)
Kakovost vode - Določevanje hlapnih cikličnih metilsiloksanov v vodi - 2. del: Metoda s tekočinsko-tekočinsko ekstrakcijo in plinsko kromatografijo z masno selektivnim detektorjem (GC-MS) (ISO 20596-2:2021)
General Information
Standards Content (sample)
SLOVENSKI STANDARD
oSIST prEN ISO 20596-2:2022
01-julij-2022
Kakovost vode - Določevanje hlapnih cikličnih metilsiloksanov v vodi - 2. del:
Metoda s tekočinsko-tekočinsko ekstrakcijo in plinsko kromatografijo z masno
selektivnim detektorjem (GC-MS) (ISO 20596-2:2021)
Water quality - Determination of cyclic volatile methylsiloxanes in water - Part 2: Method
using liquid-liquid extraction with gas chromatography-mass spectrometry (GC-MS) (ISO
20596-2:2021)Wasserbeschaffenheit - Bestimmung von cyclischen flüchtigen Methylsiloxanen in
Wasser - Teil 2: Verfahren mittels Flüssig-Flüssig-Extraktion und Gaschromatographie
mit Massenspektrometrie (GC-MS) (ISO 20596-2:2021)Qualité de l'eau - Détermination de méthylsiloxanes cycliques volatiles dans l'eau -
Partie 2: Méthode par extraction liquide-liquide avec chromatographie en phasegazeuse-spectrométrie de masse (CG-SM) (ISO 20596-2:2021)
Ta slovenski standard je istoveten z: prEN ISO 20596-2
ICS:
13.060.50 Preiskava vode na kemične Examination of water for
snovi chemical substances
oSIST prEN ISO 20596-2:2022 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
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oSIST prEN ISO 20596-2:2022
INTERNATIONAL ISO
STANDARD 20596-2
First edition
2021-01
Water quality — Determination of
cyclic volatile methylsiloxanes in
water —
Part 2:
Method using liquid-liquid extraction
with gas chromatography-mass
spectrometry (GC-MS)
Qualité de l'eau — Détermination de méthylsiloxanes cycliques
volatiles dans l'eau —
Partie 2: Méthode par extraction liquide-liquide avec
chromatographie en phase gazeuse-spectrométrie de masse (CG-SM)
Reference number
ISO 20596-2:2021(E)
ISO 2021
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2021
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.ISO copyright office
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Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii © ISO 2021 – All rights reserved
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
Contents Page
Foreword ........................................................................................................................................................................................................................................iv
Introduction ..................................................................................................................................................................................................................................v
1 Scope ................................................................................................................................................................................................................................. 1
2 Normative references ...................................................................................................................................................................................... 1
3 Terms and definitions ..................................................................................................................................................................................... 2
4 Principle ........................................................................................................................................................................................................................ 2
4.1 Principle of preservation and extraction .......................................................................................................................... 2
5 Interferences ............................................................................................................................................................................................................ 2
5.1 Interferences with sampling and processing ................................................................................................................ 2
5.2 Interferences with GC-MS .............................................................................................................................................................. 2
5.3 Interferences determination ....................................................................................................................................................... 3
6 Reagents ........................................................................................................................................................................................................................ 3
7 Apparatus ..................................................................................................................................................................................................................... 5
8 Method detection limits ................................................................................................................................................................................ 6
9 Quality control ........................................................................................................................................................................................................ 6
10 Sampling and storage ...................................................................................................................................................................................... 6
10.1 Sampling preparation ....................................................................................................................................................................... 6
10.2 Sample collection .................................................................................................................................................................................. 6
11 Extraction and analysis .................................................................................................................................................................................. 7
11.1 Extraction .................................................................................................................................................................................................... 7
11.2 GC conditions and operation ....................................................................................................................................................... 7
12 Calibration .................................................................................................................................................................................................................. 7
12.1 General requirements ....................................................................................................................................................................... 7
12.2 Calibration calculations ................................................................................................................................................................... 8
12.3 Concentration calculations ........................................................................................................................................................... 8
12.4 Calculation of results ......................................................................................................................................................................... 9
12.5 Treatment of results lying outside the calibration range ................................................................................... 9
13 Expression of results .....................................................................................................................................................................................10
14 Test report ................................................................................................................................................................................................................10
Annex A (informative) GC-MS conditions ......................................................................................................................................................11
Annex B (informative) Method detection limit and limit of quantification ..............................................................13
Annex C (informative) Example quality control samples .............................................................................................................14
Annex D (informative) Performance data ....................................................................................................................................................15
Bibliography .............................................................................................................................................................................................................................16
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/ directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/ patents).Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see www .iso .org/
iso/ foreword .html.This document was prepared by Technical Committee ISO/TC 147, Water quality, Subcommittee SC 2,
Physical, chemical and biochemical methods.A list of all parts in the ISO 20596 series can be found on the ISO website.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/ members .html.iv © ISO 2021 – All rights reserved
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ISO 20596-2:2021(E)
Introduction
The method described in this document uses low density polyethylene to prevent volatilization of
samples during transit and storage. The samples are processed using a liquid-liquid extraction into
a non-polar solvent with subsequent injection onto a gas chromatograph-mass spectrometer for
separation and quantitation.© ISO 2021 – All rights reserved v
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oSIST prEN ISO 20596-2:2022
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oSIST prEN ISO 20596-2:2022
INTERNATIONAL STANDARD ISO 20596-2:2021(E)
Water quality — Determination of cyclic volatile
methylsiloxanes in water —
Part 2:
Method using liquid-liquid extraction with gas
chromatography-mass spectrometry (GC-MS)
WARNING — Persons using this document should be familiar with normal laboratory practice.
This document does not purport to address all of the safety problems, if any, associated with its
use. It is the responsibility of the user to establish appropriate safety and health practices and to
ensure neutralization and proper disposal of waste solutions.IMPORTANT — It is absolutely essential that tests conducted in accordance with this document
be carried out by suitably qualified staff.1 Scope
This document specifies a method for the determination of certain cyclic volatile methylsiloxanes (cVMS)
in environmental water samples with low density polyethylene (LDPE) as a preservative and subsequent
liquid-liquid extraction with hexane containing C-labeled cVMS as internal standards. The extract is
then analysed by gas chromatography-mass spectrometry (GC-MS).NOTE Using the C-labeled, chemically identical substances as internal standards with the same properties
as the corresponding analytes, minimizes possible substance-specific discrimination in calibrations. Since these
substances are least soluble in water, they are introduced via the extraction solvent hexane into the system.
This document is applicable to the measurement of the following cVMS in rivers, streams, and waste
water (influent and effluent):Table 1 — Analytes determined by this method
Analyte Formula Abbreviation CAS -RN
Octamethylcyclotetrasiloxane C H O Si D4 556-67-2
8 24 4 4
Decamethylcyclopentasiloxane C H O Si D5 541-02-6
10 30 5 5
Dodecamethylcyclohexasiloxane C H O Si D6 540-97-6
12 36 6 6
CAS-RN Chemical Abstracts Services Registration Number
This method can be used to determine cVMS from 0,1 µg/l to 250 μg/l. In well controlled laboratory
environments, where contamination is minimized, the lower end of the application range can be
diminished by a factor of up to 10.2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 3696, Water for analytical laboratory use — Specification and test methodsISO 5667-4, Water quality — Sampling — Part 4: Guidance on sampling from lakes, natural and man-made
ISO 5667-6, Water quality — Sampling — Part 6: Guidance on sampling of rivers and streams
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
ISO 5667-10, Water quality — Sampling — Part 10: Guidance on sampling of waste waters
ISO 5667-14, Water quality — Sampling — Part 14: Guidance on quality assurance and quality control of
environmental water sampling and handlingISO 8466-1, Water quality — Calibration and evaluation of analytical methods and estimation of
performance characteristics — Part 1: Statistical evaluation of the linear calibration function
ISO/TS 13530, Water quality — Guidance on analytical quality control for chemical and physicochemical
water analysis3 Terms and definitions
No terms and definitions are listed in this document.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at http:// www .iso .org ./ obp— IEC Electropedia: available at http:// www .electropedia .org/
4 Principle
4.1 Principle of preservation and extraction
The siloxane compounds (D4), (D5), and (D6) are relatively volatile and have low solubility in water
thus making accurate quantification in aqueous matrices challenging. Low density polyethylene (LDPE)
is added to samples to prevent volatilization of the cVMS through a partial physical barrier between the
water and headspace and a matrix to which the cVMS may adsorb. Hexane is then used to extract the
dissolved and sorbed fractions of cVMS. The hexane extracts are then analysed by GC-MS (Annex A).
5 Interferences5.1 Interferences with sampling and processing
Silicones, including D4, D5, and D6 are widely used in industrial applications as well as personal care
products such as conditioner, hand lotion, sunscreens, and cosmetics (not all inclusive). Persons involved
with the collection and analysis of samples should refrain from using siloxane containing products to
limit potential contamination of the sample.Additionally, the users should refrain from using collection devices, sampling containers, laboratory
equipment or consumables which may contain silicones/siloxanes. Sample contact surfaces should
be suitably rinsed with acetone or hexane and subsequently dried in a clean area of the laboratory to
remove any contamination.5.2 Interferences with GC-MS
Silicones are also commonly found in parts and consumables associated with gas chromatography
including septa for the vials and inlet. Commonly used types of GC columns are polydimethylsiloxane
based which when exposed to moisture or when heated may generate cVMS and in such a way can
contribute to background. Thus, the use of non-polydimethylsiloxane-based GC columns is highly
recommended, in particular when analysing sub-ppb concentrations. Autosampler vial septa should be
silicone free or at a minimum coated with polytetrafluoroethylene on the side exposed to the sample.
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
The inlet septum should be replaced with a Merlin MicroSeal™ to reduce background contamination
from this source. In addition, any solvents should be dried prior to injection into the GC or care should
be taken to use a solvent in which water is only soluble in the mg/l levels.5.3 Interferences determination
In order to determine the integrity of the sampling, preparation and instrumental analysis of the
samples, it is recommended to prepare quality control (QC) samples. An example of QC samples consists
of a series of blanks and spikes to identify potential sources of contamination or loss during the life
cycle of the samples.6 Reagents
It is recommended to verify the absence (or presence of only negligible amounts) or absence of cVMS
from solvents being utilized.6.1 Water, grade 1, as defined in ISO 3696.
6.2 Hexane, C H n-hexane or mixture of isomers, determined to be suitably free of cVMS.
6 146.3 Tetrahydrofuran, C H O.
4 8
6.4 Calibration stock solutions.
6.4.1 Reference substances
See Table 1.
— Octamethylcyclotetrasiloxane;
— Decamethylcyclopentasiloxane;
— Dodecamethylcyclohexasiloxane.
6.4.2 Calibration stock solution 1
Weigh 30 mg of each of the listed standards into a 25 ml volumetric flask and fill to volume with hexane
(6.2). The concentration of this solution is approximately 1 200 µg/ml.6.4.3 Calibration stock solution 2
Dilute calibration stock solution 1 (6.4.2) with hexane (6.2) in a ratio of 1:250. The concentration of this
solution is approximately 4 800 ng/ml.6.4.4 Calibration stock solution 3
Dilute calibration stock solution 2 (6.4.3) with hexane (6.2) in a ratio of 1:100. The concentration of this
solution is approximately 48 ng/ml.1) Merlin MicroSeal is the trademark of a product supplied by Sigma-Aldrich. This information is given for
the convenience of users of this document and does not constitute and endorsement by ISO of the product named.
Equivalent products may be used if they can be shown to lead to the same results.
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
6.5 Spiking stock solutions
6.5.1 Spiking stock solution 1
Dilute calibration stock solution 1 (6.4.2) with tetrahydrofuran (6.3) in a ratio of 1:100. The
concentration of this solution is approximately 12 µg/ml.6.5.2 Spiking stock solution 2
Dilute spiking stock solution 1 (6.5.1) with tetrahydrofuran (6.3) in a ratio of 1:50. The concentration of
this solution is approximately 240 ng/ml.6.6 Internal standard working solution
6.6.1 Individual internal standards
C-labelled cVMS. Typical products available from suppliers are:
13 13
— C-D4, such as 2,4,6,8- C -octamethylcyclotetrasiloxane; or
— 2,2,4,4,6,6,8,8- C -octamethylcyclotetrasiloxane;
13 13
— C-D5, such as 2,4,6,8,10- C -decamethylcyclopentasiloxane; or
— or 2,2,4,4,6,6,8,8,10,10- C -decamethylcyclopentasiloxane;
13 13
— C-D6, such as 2,4,6,8,10,12- C -dodecamethylcyclohexasiloxane.
6.6.2 Internal standard stock solution 1
Weigh 10 mg of the appropriate internal standard (6.6.1) into a 100 ml volumetric flask and fill to
volume with hexane (6.2). The concentration of this solution is approximately 100 µg/ml.
6.6.3 Internal standard stock solution 2Dilute internal standard stock solution 1 (6.6.2) with hexane (6.2) in a ratio of 1:100. The concentration
of this solution is approximately 1 000 ng/ml.6.6.4 Internal standard working solution
Dilute internal standard stock solution 2 (6.6.3) with hexane (6.2) in a ratio of 1:250. The concentration
of this solution is approximately 4 ng/ml.6.7 Calibration standards
Using Table 2 weigh the appropriate amount of calibration stock 2 (6.4.3) or calibration stock 3 (6.4.4)
into a 5 ml volumetric flask and dilute to volume with internal standard working solution (6.6.4). Weigh
the amount of internal standard working solution added and convert to volume using the density of the
solvent used. Table 2 is given as an example, the calibration range can be modified to meet the needs of
the samples. It is recommended that at least five calibration standards be used for a calibration curve.
Table 2 — Calibration standardsCalibration Volume Target con- Target concentration
stock centration (relative to 50 ml sample)
µl ng/ml µg/l
STD A 3 20 0,19 0,038
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Table 2 (continued)
Calibration Volume Target con- Target concentration
stock centration (relative to 50 ml sample)
µl ng/ml µg/l
STD B 3 50 0,48 0,096
STD C 3 125 1,2 0,24
STD D 3 275 2,6 0,53
STD E 3 550 5,3 1,1
STD F 2 15 14 2,9
STD G 2 25 24 4,8
STD H 2 50 48 9,6
STD I 2 100 96 19,2
STD J 2 300 288 57,6
STD K 2 750 720 144
STD L 2 1 500 1 440 288
7 Apparatus
WARNING — Any surfaces that come into contact with a solution to be analysed should be
suitably rinsed with acetone or hexane and allowed to dry in a clean area of the laboratory to
remove any contamination.7.1 Gas chromatograph/mass spectrometer
The gas chromatograph shall be temperature-programmable, with all required accessories including
gasses, capillary columns, autosampler, and mass spectrometric detector. The inlet should be equipped
with a Merlin MicroSeal™ to minimize contamination (5.2).The mass spectrometer should be capable of operating over the mass range of interest (200 m/z to
500 m/z) and it should be equipped with a data system capable of quantifying ions using selected
m/z values.7.2 GC columns
Recommended column is DB-WAXetr (30 m × 0,25 mm i.d., 0,25 μm film thickness). If blank levels are
proved to be sufficiently low, other columns may be used such as DB-5ms , if appropriately tested prior
to sample processing.7.3 Volumetric flasks, with inert/silicone free stopper.
7.4 Vials, glass autosampler vials with a fluorocarbon lined, non-silicone septa.
7.5 Multi-Tube Vortexer , capable of handling multiple 125 ml jars.2) Merlin MicroSeal is the trademark of a product supplied by Sigma-Aldrich. This information is given for the
convenience of users of this document and does not constitute and endorsement by ISO of the product named.
Equivalent products may be used if they can be shown to lead to the same results.
3) DB-WAXetr and DB-5ms are the tradenames of products supplied by Agilent Technologies. This information is
given for the convenience of users of this document and does not constitute and endorsement by ISO of the product
named. Equivalent products may be used if they can be shown to lead to the same results.
4) Multi-Tube Vortexer is the tradename of a product available commercially. This information is given for the
convenience of users of this document and does not constitute and endorsement by ISO of this product.
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oSIST prEN ISO 20596-2:2022
ISO 20596-2:2021(E)
7.6 Jars, 125 ml wide mouth glass jars with a PTFE lined lid.
7.7 Centrifuge, Alternative phase separation can be obtained by centrifuging the samples for 5 min at
2 500 rpm.7.8 Low density polyethylene (LDPE), 51 µm to 63 μm thick clear base film. Film is cut into 2,5 cm ×
2,5 cm squares and subsequently washed twice with hexane (6.2) and allowed to dry.
8 Method detection limitsAny laboratory performing this test will determine the limit of detection and the limit of quantification
as defined in ISO/TS 13530. The method detection limit and limit of quantification should be assessed
as defined in Annex B.9 Quality control
Quality control samples shall be used to verify the integrity of the samples during sampling, processing
and analysing by indicating any potential contamination or loss of analyte (ISO 5667-14). This is most
important when analysing concentrations in the sub-μg/l range, where relevant contamination with
cVMS is more likely. Typically, a series of blanks and spiked samples are used for quality control.
Reference examples of these quality control samples, and their use are described in Annex C.
NOTE The initial interlaboratory trial conducted when developing this document, for sub-ppb concentrations
did not meet the ISO performance criteria, mainly because of an un-discovered contamination issue.
Blank levels and sample concentrations shall differ sufficiently to qualify sample concentrations as
relevant. Where concentrations reported are close to the detection limits, replicates of blank samples
and of actual samples may be measured and 2-sample t-test can be used to check for a statistically
significant difference.10 Sampling and storage
10.1 Sampling preparation
Ensure that all sample contact surfaces have been washed with acetone or hexane and allowed to dry.
And ensure that all LDPE squares have been solvent washed with hexane in duplicate and allowed to dry.
Add 8 LDPE squares to each 125 ml uniquely labelled sample jar with lid. Take a weight of the sample
jar. This weight will be used to later determine the weight and volume of the sample.
10.2 Sample collectionTake samples in accordance with ISO 5667-4, ISO 5667-6, ISO 5667-10, ensure sampling collection
devices are clean and free of siloxanes. In order to collect a homogenous sample, the sample collection
device should be sufficient to contain a bulk sample so that sufficient replicate samples can be poured
or drawn from the same source. It is advisable to take two samples, one to be retained in the event of a
repeat analysis is required.If field blanks are utilized, it is appropriate to open the field blanks as described in Annex C.
Rinse the sampling device three times with the intended sample and collect the bulk sample. Then
aliquot the bulk sample into the 125 ml sample jars so that each jar is approximately ½ full. This allows
for the solvent extraction step to be performed in the jar. The LDPE helps prevent volatilization of cVMS
into the headspace.Close the samples and place them in a storage/shipping container to be maintained at a temperature
(5 ± 3) °C. These samples should be extracted within 14 d of collection.6 © ISO 2021 – All rights reserved
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11 Extraction and analysis
11.1 Extraction
Weigh the sample jar to determine the amount of sample collected. Add 10 ml of internal standard working
solution (6.6.4) to the samples. The samples are placed on the Multi-Tube Vortexer (7.5) and vortexed at
maximum speed for 30 min. The extracted sample in the jar is allowed to settle to separate the organic
phase, showing clear phase separation, or alternatively phase separation can be obtained by centrifuging
(7.7) the samples. The organic phase is removed and placed in a previously cleaned storage vial. An aliquot
is removed from the storage vial and placed into an autosampler vial for analysis by GC-MS.
NOTE To reduce negative effects caused by water traces being present in the organic phase, it can be
advantageous to add approximately 1 g of dry MgSO into the vial.11.2 GC conditions and operation
Example operating conditions can be found in Annex A.
If multiple injections of a sample, blank, or calibration solution are to be made, it is recommended to
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