CEN/TS 16692:2015

SLOVENSKI STANDARD
01-junij-2015
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Water quality - Determination of tributyl tin (TBT) in whole water samples using solid
phase extraction (SPE) and gas chromatography with triple quadrupole mass
spectrometry
Wasserbeschaffenheit - Bestimmung von Tributylzinn (TBT) in Gesamtwasserproben
mittels Festphasenextraktion (SPE) und Gaschromatographie mit Triple-Quadrupol-
Massenspektrometrie
Qualité de l'eau - Dosage du tributylétain (TBT) dans des échantillons d'eau totale par
extraction sur phase solide (SPE) et chromatographie en phase gazeuse avec
spectrométrie de masse triple quadrupôle
Ta slovenski standard je istoveten z: CEN/TS 16692:2015
ICS:
13.060.50 3UHLVNDYDYRGHQDNHPLþQH Examination of water for
VQRYL chemical substances
71.040.50 Fizikalnokemijske analitske Physicochemical methods of
metode analysis
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

TECHNICAL SPECIFICATION
CEN/TS 16692
SPÉCIFICATION TECHNIQUE
TECHNISCHE SPEZIFIKATION
April 2015
ICS 13.060.50
English Version
Water quality - Determination of tributyltin (TBT) in whole water
samples - Method using solid phase extraction (SPE) with SPE
disks and gas chromatography with triple quadrupole mass
spectrometry
Qualité de l'eau - Dosage du tributylétain (TBT) dans la Wasserbeschaffenheit - Bestimmung von Tributylzinn (TBT)
totalité des échantillons d'eau - Méthode par extraction sur in Gesamtwasserproben - Verfahren mittels
phase solide (SPE) avec disques SPE et chromatographie Festphasenextraktion (SPE) mit SPE-Disks und
en phase gazeuse avec spectrométrie de masse triple Gaschromatographie mit Triple-Quadrupole
quadrupôle Massenspektrometrie
This Technical Specification (CEN/TS) was approved by CEN on 1 April 2014 for provisional application.

The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN will be requested to submit their
comments, particularly on the question whether the CEN/TS can be converted into a European Standard.

CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS available
promptly at national level in an appropriate form. It is permissible to keep conflicting national standards in force (in parallel to the CEN/TS)
until the final decision about the possible conversion of the CEN/TS into an EN is reached.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania,
Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United
Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels
© 2015 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 16692:2015 E
worldwide for CEN national Members.

Contents Page
Foreword .3
Introduction .4
1 Scope .5
2 Normative references .5
3 Principle .5
4 Interferences .5
5 Reagents .6
6 Apparatus .8
7 Sampling .9
8 Procedure .9
8.1 Sample preparation .9
8.2 Derivatization .9
8.2.1 General .9
8.2.2 Derivatization procedure . 10
8.3 Extraction with SPE-disks . 10
8.4 Clean-up of the extract . 11
8.5 Solvent concentration step . 11
8.6 Gas chromatograph . 11
8.7 Identification of individual compounds by GC-MS . 11
8.8 Blank value measurements . 12
9 Calibration . 12
9.1 General requirements . 12
9.2 Calibration of the GC-step . 13
9.3 Calibration of the total procedure using the internal standard . 14
9.4 Determination of procedural recovery values . 15
9.5 Determination of derivatization standard recovery . 15
10 Calculation . 16
11 Expression of results . 17
12 Test report . 17
Annex A (informative) Suitable gas chromatographic conditions and example chromatograms . 18
Annex B (informative) Repeatability and reproducibility data . 20
Annex C (informative) Example of conditions for automated solid-phase extraction . 21
Annex D (informative) Silica clean-up . 22
D.1 Silica for the clean-up column . 22
D.2 Clean-up column . 22
D.3 Eluent for cleaning extract . 22
D.4 Clean-up of extract . 22
Bibliography . 23

Foreword
This document (CEN/TS 16692:2015) has been prepared by Technical Committee CEN/TC 230 “Water
analysis”, the secretariat of which is held by DIN.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights.
This document has been prepared under a mandate given to CEN by the European Commission and the
European Free Trade Association, and supports essential requirements of the Water Framework Directive
(WFD, 2000/60/EC), and the Directive on Environmental Quality Standards (Directive 2008/105/EC).
According to the CEN-CENELEC Internal Regulations, the national standards organizations of the following
countries are bound to announce this Technical Specification: Austria, Belgium, Bulgaria, Croatia, Cyprus,
Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany,
Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland,
Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom.
WARNING — Persons using this Technical Specification should be familiar with usual laboratory
practice. This Technical Specification 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 compliance with any national regulatory conditions.
IMPORTANT — It is absolutely essential that tests conducted according to this Technical
Specification be carried out by suitably trained staff.
Introduction
Tributyltin (TBT) is a priority substance listed in Annex X of the EU Water Framework Directive (WFD,
Directive 2000/60/EC) for which Environmental Quality Standards (EQS) have been set at EU level for inland
as well as other surface waters to protect the aquatic environment against chemical pollution (Directive
2008/105/EC). With the exception of metals, the EQSs are expressed as total concentrations in the whole
water sample. Furthermore, analytical methods used in WFD monitoring need to meet certain requirements as
regards the minimum limit of quantification and the maximum tolerable measurement uncertainty (Directive
2009/90/EC). So far, there is no standardized method available for the determination of TBT in whole water
samples fulfilling those requirements. Hence, the European Commission mandated CEN to develop or
improve standards in support of the implementation of the monitoring requirements of WFD.
Directive 2008/105/EC has been amended by Directive 2013/39/EU, however this standard has been
developed for the analysis of TBT as listed in Annex A of Directive 2008/105/EC.
The annual average environmental quality standard (AA-EQS) value for TBT is 0,0002 μg/l and is defined for
the concentration in the whole water sample, including suspended particulate matter (SPM) present in the
sample. As compounds like TBT, sorb strongly to environmental solids, the fraction bound to particles may be
substantial. Therefore it is important to be able to handle whole water samples within the analytical process.
Identification and quantification of TBT at trace level concentrations often require both high sensitive
chromatographic equipment and effective enrichment steps.
1 Scope
This Technical Specification specifies a method for the determination of tributyltin cation (TBT) in whole water
samples. It is applicable to the analysis of TBT in surface water, which may contain suspended particulate
matter (SPM) up to 500 mg/l (whole water samples), groundwater, drinking water and seawater. The working
range is 0,04 ng/l to 20 ng/l. The LOQ will be mainly determined by the blank value obtained during validation
of this method.
NOTE 1 The method has been successfully applied to seawater samples during method development, but sea water
samples were not included in the interlaboratory comparison.
NOTE 2 In this document TBT is synonymous for tributyltin cation.
NOTE 3 Near the lower limit of the working range the measurement uncertainties may be higher.
2 Normative references
The following documents, in whole or in part, are normatively referenced in this document and are
indispensable for its application. For dated references, only the edition cited applies. For undated references,
the latest edition of the referenced document (including any amendments) applies.
EN ISO 3696, Water for analytical laboratory use - Specification and test methods (ISO 3696)
EN ISO 5667-3, Water quality - Sampling - Part 3: Preservation and handling of water samples (ISO 5667-3)
3 Principle
TBT in the whole water sample is derivatized while stirring thoroughly. The derivatized target analyte is
extracted using solid phase extraction with SPE disks and subsequently concentrated by evaporation. An
aliquot of the extract is brought on the gas chromatograph with programmed temperature vaporizing - large
volume injection (PTV-LVI) and identified and quantified with a triple quadrupole mass spectrometric detection
in single GC-MS reaction mode.
Water samples of 1 l are spiked with internal standards and brought to pH 4 to pH 5 with a sodium acetate
buffer. Subsequently, TBT is ethylated by adding sodium tetra ethylborate (NaEt B) to the water samples,
while rigorously stirring. Solid phase disk extraction is used for extracting ethylated TBT with mainly hexane as
a solvent. The extract is then brought to pH 12 with sodium hydroxide [1]. The final extract is then
concentrated to 300 µl. A volume of 20 µl is injected into the gas chromatograph using a PTV-LV injection
technique followed by GC-MS/MS in single GC-MS reaction mode for separation and detection.
A deuterium-labelled TBT substance is used as internal standard for quantification of TBT. A TBT-spiking
solution is used for the determination of procedural recovery values. A tri-alkylated spiking solution is used for
checking the efficiency of the alkylation process and is added to each sample. Tetra-alkylated reference
solutions are used for the calibration.
4 Interferences
The reagents can contain impurities of organotin compounds, including the derivatization reagent. It is
absolutely essential to verify contamination before use by analysis of blanks. Glassware can become
contaminated. Heat all used glassware to 450 °C, or clean the glassware with a 10 % (volume fraction) nitric
acid solution before use, by decontamination overnight. Make sure the GC-system is not contaminated.
Regularly refresh the wash solvents to ensure that no contamination occurs via the injection needle. Verify the
GC-system before analysis by analysis of blank solvent.
Commercially available solid-phase extraction disks (SPE-disks) differ frequently in quality. Variations in the
selectivity of the materials also frequently occur from batch to batch, thus possibly causing significant
deviations in extraction yield. This does not basically impair their suitability, apart from a resulting higher
detection limit of individual substances. To ensure that the measuring results show high trueness and
precision, use materials of one batch for both measurement and calibration. Avoid major fluctuations in the
extraction times and elution procedures within one sample sequence when analysing the samples. SPE -
disks may be contaminated with mono-, di- or tributyltin cation compounds. Verify the contamination of the
disks before use. Pre-cleaning can be achieved by pre-extracting the disks with a derivatization agent. To
thoroughly clean magnetic stirrers, they can be shaken in a methanol-hexane solution (9:1, v/v) with 1 ml of 2
% derivatization agent. Decontaminate overnight and rinse three times with deionized water.
5 Reagents
The reagents shall be free from impurities possibly interfering with the GC-MS analysis.
Use solvents and reagents of sufficient purity, i.e. with negligibly low impurities compared with the
concentration of analytes to be determined. As reagents use, as far as available “residual grade” or better in
order to obtain low blanks. Verify by blank determinations and, if necessary, apply additional cleaning steps.
5.1 Water, Grade 1 according to EN ISO 3696.
5.2 Operating gases for the gas chromatography mass spectrometry, of high purity and in accordance with
manufacturer’s specifications.
5.3 Nitrogen of high purity, i.e. minimum 99,996 % by volume, for concentration by evaporation.
5.4 Solvents for extraction, chromatography and preparation of reference solutions.
— hexane, C H , (boiling point: 69 °C);
6 14
— iso-octane, C H (boiling point: 99 °C);
8 18
— iso-propanol, C H OH (boiling point: 82 °C);
3 7
— methanol, CH OH (boiling point: 65 °C).
5.5 Acetate buffer solution, solution, c(CH COONa) = 4 mol/l.
5.6 Sodium hydroxide, solution, c(NaOH) = 10 mol/l.
5.7 Hydrochloric acid, w(HCl) = 36 % to 38 % mass fraction.
5.8 Nitric acid, v(HNO ) = 10 % volume fraction.
5.9 Aluminium oxide, activated at 800 °C for 4 h, (10 ± 2) % deactivated.
5.10 Sodiumtetraethylborate, w(NaEt B) = 2 % mass fraction, (CAS Registry Number 15523-24-7).
5.11 Reference ethyl-TBT stock solution
This substance is used for the calibration. Use a commercially available solution, e. g. in iso-octane or the
commercially available pure substance (Table 1). The stock solution is prepared by dissolving, e. g. 5 mg of
the reference substance (precision of 0,001 mg) in 20 g of an appropriate solvent (precision of 0,01 g) e. g.
hexane or iso-octane. Store stock solutions at temperatures between 1 °C and 5 °C, protected from light.
Stock solutions with concentrations > 1 mg/g are stable for at least 12 months and at least 3 months for
concentrations > 1 µg/g.
5.12 Reference TBT stock solution
This substance is used for spiking water samples for determination of procedural recovery values. Use a
commercially available solution of TBT, e. g. TBT-chloride in iso-octane or the commercially available pure
substance (Table 1). Stock solution is prepared as described in 5.11, with e. g. iso-propanol or methanol as
solvent. Stock solutions with concentrations > 1 mg/g are stable for at least 12 months and at least 3 months
for concentrations > 1 µg/g.
5.13 Derivatization standard stock solution
In order to check the efficiency of the derivatization process, choose a tri-alkylated cation substance not
present in the sample with similar physicochemical properties as TBT, e. g. TPrT-Cl (Table 1). The
ethylderivative of the derivatization standard, e. g. ethyl-TPrT (Table 1) is used for calibration. Prepare stock
solutions of the derivatization standard substances in the same way as specified for the reference stock
solutions (5.11 and 5.12). Stock solutions with concentrations > 1 mg/g are stable for at least 12 months and
at least 3 months for concentrations > 1 µg/g.
5.14 Internal standard solution
5.14.1 Internal standard for calculation of TBT concentration
As internal standard for quantification of TBT, a deuterium-labelled substance is used. The ethylderivative is
used in the calibration. In Table 1 internal standards are listed. Prepare stock solutions of the internal standard
substances in the same way as specified for the reference stock solutions (5.11 and 5.12).
C-enriched TBT might be used as an alternative internal standard, but suitability should be checked, as it
has not been used in the method development nor in the interlaboratory comparison.
5.14.2 Internal standard for calculation of the derivatization standard
For calculation of the derivatization standard (5.13) concentration use, e. g. tetrapropyltin (Table 1) as internal
standard or choose a substance with similar physicochemical properties as the derivatization standard
(extraction behaviour, retention time). The internal standard should be tetra-alkylated. Prepare stock solutions
of the internal standard substance in the same way as specified for reference stock solution (5.11).
5.15 Calibration working solutions
Use the ethyl derivative of TBT (5.11) and the derivatization standard (5.13) of a defined concentration, used
for gas chromatography and suitable for the preparation of calibration solutions. Prepare a minimum of seven
calibration solutions with concentrations according to the detection capability of the mass spectrometer.
Combine and dilute internal standards (5.14) with hexane and iso-octane to produce solutions for the
calibration range in a hexane - iso-octane (9:1, volume fraction) solvent. Store calibration solutions for
maximum 3 days at temperatures between 1 °C and 5 °C protected from light.
5.16 TBT reference substance solution
Use TBT (5.12) of defined concentration, suitable for the preparation of a reference solution used for spiking
water samples. Spike blank water samples for the determination of procedural recovery values (9.4).
5.17 Derivatization standard solution
Use the tri-alkylated derivatization standard (5.13) of defined concentration, suitable for the preparation of
reference solutions used for spiking water samples. Spike each sample for calculation of the recovery (9.5).
Table 1 — Reference substances, internal and derivatization standards and their ethyl-derivatives
Substance Abbreviation Molecular Molar mass a b
EC number CAS RN
formula
g/mol
Reference substance and ethyl derivative
Tributyltin chloride TBT-Cl C H SnCl 325,51 215–958–7 1461–22–9
12 27
(5.12)
Ethyltributyltin (5.11) Ethyl-TBT C H Sn 319,11 not applicable 19411–60–0
14 32
Deuterated internal standard and ethyl derivative (5.14.1)
Tributyltin- d -TBT-Cl C D SnCl 352,51 not applicable –
27 12 27
d chloride
Ethyltributyltin-d Ethyl-d -TBT C H D Sn 346,11 not applicable –
27 27 14 5 27
Tetra-alkyltin standard (5.14.2)
Tetrapropyltin TePrT C H Sn 291,05 218–536–0 2176–98–9
12 28
Derivatization standard and ethyl derivative (5.13)
Tripropyltin chloride TPrT-Cl C H SnCl 283,41 218–910–3 2279–76–7
9 21
Ethyltripropyltin Ethyl-TPrT C H Sn 277,03 not applicable 3440–79–7
11 26
a
EC Number: European inventory of existing commercial substances (EINECS) or European list of notified chemical
substances (ELINCS).
b
CAS RN: Chemical Abstracts Service Registry Number.
6 Apparatus
6.1 General
Equipment or parts of it which are likely to come into contact with the water sample or its extract shall be free
from residues causing interferences. The use of vessels made of glass is recommended. Heat all glassware to
450 °C, or clean the glassware with 10 % (volume fraction) nitric acid solution. Decontaminate overnight and
rinse three times with deionized water.
6.2 Sample flasks, e. g. brown glass, flat bottomed, with glass or PTFE coated stoppers, e. g. 250 ml or
500 ml and 1 000 ml.
6.3 Magnetic stirrer, including PTFE-coated magnetic stir bar of suitable size.
6.4 Drying ovens, capable of maintaining temperatures in the ranges of 100 °C to 800 °C for baking and
storage of clean-up materials and maintaining temperatures up to 450 °C for baking glassware.
6.5 Solid-phase extraction disks (SPE disks), wide inner diameter: e. g. 45 mm, packed with an
appropriate reversed phase adsorbent material, e. g. C -based or DVB-based adsorbent.
6.6 Vacuum device for solid-phase extraction, e. g. vacubox, extraction box or automated workstation
for solid-phase extraction procedure capable for processing SPE-disks or automatic solid disk extractor,
capable of extracting water samples up to 1 l with organic solvents and with automatic cleaning cycle.
6.7 Glass columns for chromatographic clean-up.
6.8 Evaporation device, based on nitrogen flow evaporation or rotary evaporator under reduced pressure.
6.9 Syringes, 10 µl, 25 µl, 50 µl, 100 µl, 1 000 µl, volume precision ± 2 %.
6.10 Single volume pipettes, capacities between 100 µl and 50 ml.
6.11 Glass sample vials, dark coloured glass capacity e. g. 2 ml with 300 µl inserts, with inert cap and
PTFE-coated septum.
6.12 Capillary gas chromatograph with a triple quadrupole mass spectrometer, (GC-MS/MS) using
single GC-MS reaction mode, gas supply in accordance with the respective manufacturer’s instructions.
6.13 Non-discriminating GC injector, e. g. splitless mode of a split or splitless injection system and
programmable temperature vaporiser (PTV), programmable for large volume injection (LVI).
6.14 Automatic sampler with option for large volume injection (LVI), including syringes for normal
injection (e. g. 1 µl and 2 µl) and LVI (e. g. 50 µl and 100 µl).
6.15 Capillary column, for gas chromatography, fused silica column with non-polar low bleed separating
phase (e. g. 5 % diphenyl 95 % dimethyl polysiloxane, length 20 m, inner diameter 0,18 mm and film thickness
0,18 µm).
6.16 Pasteur pipettes
6.17 pH indicator strips, for the appropriate pH-ranges.
7 Sampling
For sampling, use thoroughly cleaned, flat bottomed glass flasks (6.2). Fill the bottles completely with the
water to be examined. Samples should be brought to pH (2 ± 0,2) with hydrochloric acid (5.7) within 24 h.
If extraction requires the sample flask to be included in the working process, it is recommended to fill the bottle
to the shoulder (e.g. about 1 00
...