EN ISO 23161:2011

SLOVENSKI STANDARD
01-februar-2012
.DNRYRVWWDO'RORþHYDQMHL]EUDQLKRUJDQRNRVLWURYLKVSRMLQ0HWRGDSOLQVNH
NURPDWRJUDILMH,62
Soil quality - Determination of selected organotin compounds - Gas-chromatographic
method (ISO 23161:2009)
Bodenbeschaffenheit - Bestimmung ausgewählter Organozinnverbindungen -
Gaschromatographisches Verfahren (ISO 23161:2009)
Qualité du sol - Dosage d'une sélection de composés organostanniques - Méthode par
chromatographie en phase gazeuse (ISO 23161:2009)
Ta slovenski standard je istoveten z: EN ISO 23161:2011
ICS:
13.080.10 .HPLMVNH]QDþLOQRVWLWDO Chemical characteristics of
soils
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EUROPEAN STANDARD
EN ISO 23161
NORME EUROPÉENNE
EUROPÄISCHE NORM
August 2011
ICS 13.080.10
English Version
Soil quality - Determination of selected organotin compounds -
Gas-chromatographic method (ISO 23161:2009)
Qualité du sol - Dosage d'une sélection de composés Bodenbeschaffenheit - Bestimmung ausgewählter
organostanniques - Méthode par chromatographie en Organozinnverbindungen - Gaschromatographisches
phase gazeuse (ISO 23161:2009) Verfahren (ISO 23161:2009)
This European Standard was approved by CEN on 14 July 2011.

CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European
Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such national
standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member.

This European Standard exists in three official versions (English, French, German). A version in any other language made by translation
under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management Centre has the same
status as the official versions.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland,
Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland and United Kingdom.

EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

Management Centre: Avenue Marnix 17, B-1000 Brussels
© 2011 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 23161:2011: E
worldwide for CEN national Members.

Contents Page
Foreword .3

Foreword
The text of ISO 23161:2009 has been prepared by Technical Committee ISO/TC 190 “Soil quality” of the
International Organization for Standardization (ISO) and has been taken over as EN ISO 23161:2011 by
Technical Committee CEN/TC 308 “Characterization of sludges” the secretariat of which is held by AFNOR.
This European Standard shall be given the status of a national standard, either by publication of an identical
text or by endorsement, at the latest by February 2012, and conflicting national standards shall be withdrawn
at the latest by February 2012.
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.
According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following
countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech
Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia,
Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain,
Sweden, Switzerland and the United Kingdom.
Endorsement notice
The text of ISO 23161:2009 has been approved by CEN as a EN ISO 23161:2011 without any modification.

INTERNATIONAL ISO
STANDARD 23161
First edition
2009-09-01
Soil quality — Determination of selected
organotin compounds — Gas-
chromatographic method
Qualité du sol — Dosage d'une sélection de composés
organostanniques — Méthode par chromatographie en phase gazeuse

Reference number
ISO 23161:2009(E)
©
ISO 2009
ISO 23161:2009(E)
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ii © ISO 2009 – All rights reserved

ISO 23161:2009(E)
Contents Page
Foreword .iv
Introduction.v
1 Scope.1
2 Normative references.2
3 Terms and definitions .2
4 Principle.2
5 Reagents.4
5.1 General .4
5.2 Chemicals.4
5.3 Standards .5
5.4 Preparation of reagents and solutions.6
5.5 Clean-up .7
6 Apparatus.7
6.1 Requirements for glassware .7
6.2 Sampling apparatus .8
6.3 Additional apparatus.8
7 Procedure.8
7.1 Sampling and sample pretreatment .8
7.2 Sample extraction.9
7.3 Clean-up of the extract.10
7.4 Determination of dry mass .11
7.5 Measurement .11
8 Calibration.12
9 Recovery rates of the internal standard compounds .12
10 Quantification .13
11 Expression of results.14
12 Precision.14
13 Test report.14
Annex A (informative) Information about the procedure .15
Annex B (informative) Additional clean-up procedures.17

Annex C (informative) Information about typical instrumental conditions .20
Annex D (informative) Information about GC/MS identification.31
Annex E (informative) Validation data .33
Bibliography.37

ISO 23161:2009(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.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
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.
ISO 23161 was prepared by Technical Committee ISO/TC 190, Soil quality, Subcommittee SC 3, Chemical
methods and soil characteristics.
iv © ISO 2009 – All rights reserved

ISO 23161:2009(E)
Introduction
It is absolutely essential that tests conducted in accordance with this International Standard be carried out by
suitably qualified staff.
It can be noted whether, and to what extent, particular problems will require the specification of additional
boundary conditions.
INTERNATIONAL STANDARD ISO 23161:2009(E)

Soil quality — Determination of selected organotin
compounds — Gas-chromatographic method
WARNING — Persons using this International Standard should be familiar with normal laboratory
practice. This International Standard 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.
1 Scope
This International Standard specifies a gas-chromatographic method for the identification and quantification of
organotin compounds (OTCs) in soils as specified in Table 1. The method is also applicable to samples from
sediments, sludges and wastes (soil-like materials). The working range depends on the detection technique
used and the amount of sample taken for analysis. The limit of quantification for each compound is about
10 µg/kg.
Table 1 — Organotin compound, which can be determined in accordance with this International
Standard
(4−n)+
R Sn R n Name Acronym
n
a
Organotin cations
3+
BuSn Butyl 1 Monobutyltin cation MBT
2+
Bu Sn Butyl 2 Dibutyltin cation DBT
+
Bu Sn Butyl 3 Tributyltin cation TBT
3+
OcSn Octyl 1 Monooctyltin cation MOT
2+
Oc Sn Octyl 2 Dioctyltin cation DOT
+
Ph Sn Phenyl 3 Triphenyltin cation TPhT
+
Cy Sn Cyclohexyl 3 Tricyclohexyltin cation TCyT
Peralkylated organotin
Bu Sn Butyl 4 Tetrabutyltin TTBT
a
Organotin compounds are measured after derivatization.
NOTE When applying this method to the determination of other organotin compounds not specified in the scope, its
suitability is proven by proper in-house validation experiments, e.g. methyltin compounds. See Table 2. Methyltin cations
are unlikely to evaporate from aqueous solvents, but peralkylated methyltin compounds are volatile and subject to losses
(see C.3). Therefore, additional precautions are established.
Table 2 — Methyltin compounds
(4−n)+
R Sn R n Name Acronym
n
3+
MeSn Methyl 1 Monomethyltin cation MMT
2+
Me Sn Methyl 2 Dimethyltin cation DMT
+
Me Sn Methyl 3 Trimethyltin cation TMT
ISO 23161:2009(E)
Organotin cations can only be determined in accordance with this International Standard after derivatization.
The anionic part bound to the organotin cation is mainly dependent on the chemical environment and is not
determined using this method. The peralkylated organotin compounds behave in a completely different way
from their parent compounds. Tetraalkylated organotin compounds which are already peralkylated, such as
tetrabutyltin, are determined directly without derivatization.
The properties, such as particle size distribution, water content and organic matter content of the solids to be
analysed using this International Standard vary widely. Sample pretreatment is designed adequately with
respect to both the properties of the organotin compounds and the matrix to be analysed.
2 Normative references
The following referenced documents are indispensable for the application 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 methods
ISO 11465, Soil quality — Determination of dry matter and water content on a mass basis — Gravimetric
method
ISO 16720, Soil quality — Pretreatment of samples by freeze-drying for subsequent analysis
ISO 22892, Soil quality — Guidelines for the identification of target compounds by gas chromatography and
mass spectrometry
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
organotin compound
substance containing 1 to 4 Sn-C bonds
NOTE The number of Sn-C bonds is a measure for the degree of substitution.
3.2
organotin cation
part of the organotin compound (3.1) that contains all Sn-C bonds and is formally charged
3.3
organotin cation derivatives
non-dissociated tetrasubstituted organotin compounds which are produced by derivatization
3.4
solid
soil, sediment, sludge and waste (soil-like material)
4 Principle
For the ionic and the non-ionic organotin compounds (see Table 1), a different sample pretreatment and
sample preparation are necessary. For the determination of organotin cations, laboratory samples are
pretreated by freeze drying and grinding. This procedure enables homogeneity of the sample to be achieved.
The determination of non-ionic TTBT cannot be carried out with freeze-dried materials due to evaporation
losses, thus, it shall be determined in the field-moist sample. Organotin cations can only be determined after
derivatization, whereas TTBT is already peralkylated and can be determined without derivatization (see the
flowchart in Figure 1).
2 © ISO 2009 – All rights reserved

ISO 23161:2009(E)
Figure 1 — Flowchart for the pretreatment and analysis of selected organotin compounds
For the determination of organotin compounds, two alternative extraction methods are given, both followed by
in situ derivatization with a tetraethylborate compound and simultaneous extraction with hexane:
a) treatment with acetic acid;
b) treatment with methanolic potassium hydroxide.
NOTE 1 If it is necessary to take a large amount of sample, extraction and derivatization can be done in two steps. An
aliquot of the extract can be taken for derivatization. This also applies for samples with high levels of OTC contamination.
NOTE 2 During in situ derivatization, the solid phase is still present. This supports the extraction by continuous
changing of the polar organotin cations to the non-polar organotin cation derivates. In situ methods can improve the
extraction efficiency, particularly for monoalkylated organotin compounds.
NOTE 3 Other extraction techniques can be applied if a comparable extraction efficiency is achieved.
ISO 23161:2009(E)
NOTE 4 Treatment with potassium hydroxide provides some degree of digestion and is recommended especially when
the solid contains high amounts of organic and biological materials.
The internal standard mix comprises four compounds representing four alkylation states in order to mimic the
behaviour of the target compounds. After alkylation, they cover a wide range of volatility. A recovery of at least
80 % for derivatization/extraction and again 80 % for each clean-up step of the internal standard compounds
should be achieved. (For more information, see A.3.) Tetraalkylborate is very reactive and will also alkylate
other compounds in the matrix. Those compounds (and also boroxines) may interfere with the target
compounds during gas chromatographic determination and influence detection. In order to protect the column
and to reduce the interference in chromatography, it will be necessary to apply a precleaning step to most
samples. Clean-up with silica or aluminium oxide is the minimum; further clean-up steps (e.g. aluminium
oxide/silver nitrate, silica/silver nitrate, pyrogenic copper; see Annex B) may be applied if necessary.
The determination of the tetrasubstituted organotin compounds is carried out after clean-up and concentration
steps by separation with capillary gas chromatography and detected with a suitable system [mass
spectrometer (MS), (MS/MS), flame photometric detector (FPD), atomic absorption spectrometer (AAS),
atomic emission detector (AED), inductively coupled plasma/mass spectrometer ICP/MS]. The concentrations
are determined by calibration over the total procedure using aqueous multi-component calibration standard
solutions in accordance with 5.4.3.
5 Reagents
5.1 General
Use reagents of highest purity, typically of pesticide grade or better. The reagents may contain impurities of
organotin compounds. It is absolutely essential to verify the blanks.
The water shall be free of interferences. Use water in accordance with Grade 3 of ISO 3696.
5.2 Chemicals
5.2.1 Acetic acid, CH COOH, glacial.
5.2.2 Sodium hydroxide solution, NaOH, 40 % (m/V).
5.2.3 Sodium acetate, CH COONa.
5.2.4 Sodium sulfate, Na SO , anhydrous.
2 4
5.2.5 Potassium hydroxide, KOH.
5.2.6 Silica gel, grain size 0,085 mm to 0,28 mm (63 mesh to 200 mesh).
5.2.7 Aluminium oxide, Al O , alkaline.
2 3
5.2.8 Tetrahydrofurane, C H O, free of peroxides, free of water.
4 8
5.2.9 Acetone, (CH ) CO.
3 2
5.2.10 Hexane, C H .
6 14
NOTE Both n-hexane and 2-methylpentane (i-hexane) have been found to be suitable.
4 © ISO 2009 – All rights reserved

ISO 23161:2009(E)
5.2.11 Tetraethylborate compound, e.g sodium tetraethylborate, NaB(C H ) .
2 5 4
NOTE The active species during derivatization is the tetraethylborate anion. The choice of the cation is arbitrary.
Sodium tetraethylborate was chosen since it is commercially available. In principle, any other tetraethylborate compound
can be used for analysis, including complexes formed with tetrahydrofuran (THF). A simple and rapid synthesis of a
suitable derivatization agent is described in A.1.
WARNING — Sodium tetraethylborate may contain traces of triethylboron, which may cause
instantaneous combustion.
5.2.12 Methanol, CH OH.
5.2.13 Dichloromethane, CH Cl .
2 2
5.3 Standards
WARNING — Organotin compounds vary largely regarding toxicological properties towards mammals
with respect to the alkylation stage and type of alkyl group. Cautious handling of reagents is
mandatory at any time.
Table 3 lists the standards used for calibration of the target compounds (solution A), internal standards
(solution B) and injection standard (solution C). Additional information is provided concerning weighing factors
for calculation to organotin cations (for 100 % purity of the substances).
Table 3 — Standards and internal standards for calibration of target compounds
a b c
No. Standard Abbreviation Formula CAS-RN WF Solution
5.3.1 Monobutyltin trichloride MBTCl C H SnCl 1118-46-3 0,623 A
4 9 3
5.3.2 Dibutyltin dichloride DBTCl (C H ) SnCl 683-18-1 0,767 A
4 9 2 2
5.3.3 Tributyltin chloride TBTCl (C H )SnCl 1461-22-9 0,891 A
4 9 3
5.3.4 Tetrabutyltin TTBT (C H )Sn 1461-25-2 1,000 A
4 9 4
5.3.5 Monooctyltin trichloride MOTCl C H SnCl 3091-25-6 0,686 A
8 17 3
5.3.6 Dioctyltin dichloride DOTCl (C H ) SnCl 3542-36-7 0,830 A
8 17 2 2
5.3.7 Triphenyltin chloride TPhTCl (C H )SnCl 639-58-7 0,908 A
6 5 3
5.3.8 Tricyclohexyltin chloride TCyTCl (C H )SnCl 3091-32-5 0,912 A
6 11 3
Internal standards
5.3.9 Monoheptyltin trichloride MHTCl C H SnCl 59344-47-7 0,672 B
7 15 3
5.3.10 Diheptyltin dichloride DHTCl (C H ) SnCl 74340-12-8 0,817 B
7 15 2 2
5.3.11 Tripropyltin chloride TPTCl (C H )SnCl 2279-76-7 0,875 B
3 7 3
5.3.12 Tetrapropyltin TTPT (C H )Sn 2176-98-9 1,000 B
3 7 4
5.3.13 Tetrapentyltin TTPeT (C H )Sn 3765-65-9 1,000 C
5 11 4
a
Chemical Abstracts Registration Number.
b
WF= Weighing factor = Molar mass of organotin cation/molar mass of organotin compound.
c
A for the multicomponent-standard solution in methanol.
B for the solution of the internal standards in methanol.
C for the solution of the injection standards in hexane.

ISO 23161:2009(E)
5.4 Preparation of reagents and solutions
5.4.1 General requirements
Prepare the following (see also Table 3):
⎯ multicomponent standard stock solution A in methanol (e.g. 1 mg/ml);
⎯ multicomponent standard spiking solutions for calibration, by diluting solution A with methanol;
⎯ stock solution B of internal standards in methanol (e.g. 1 mg/ml);
⎯ spiking solution of the internal standards, by diluting solution B with methanol (e.g. 100 ng/ml);
⎯ stock solution C of the injection standard in methanol (e.g. 2 mg/ml);
⎯ injection standard solution, by diluting solution C (e.g. 2 µg/ml).
5.4.2 Blank solution
Add 20 ml of water (5.1) to an Erlenmeyer flask with a ground joint or a screw-capped [polytetrafluoroethylene
(PTFE) lined] vial.
5.4.3 Aqueous calibration solutions (multicomponent solution of organotin compounds in water)
For each working range, prepare at least 6 calibration solutions with appropriate concentration levels.
Add 20 ml of water (5.1) to an Erlenmeyer flask with a ground joint or a screw-capped (PTFE-lined) vial. While
stirring vigorously, pipette an appropriate volume of the respective spiking solution underneath the surface
and ensure that the spiking solution is well distributed in the water. Stir for an additional 20 min.
5.4.4 Methanolic potassium hydroxide solution
Prepare a solution of 25 % (m/V) potassium hydroxide (5.2.5) in methanol (5.2.12). This is the methanolic
potassium hydroxide solution.
5.4.5 Acetate buffer solution
Dissolve about 1 mol of sodium acetate (equal to 82 g of anhydrous sodium acetate) (5.2.3) in 500 ml of water
(5.1) in a 1 L volumetric flask. Add sufficient glacial acetic acid (5.2.1) to adjust to a pH of 4,5. Dilute to volume
with water (5.1) and mix well.
5.4.6 Solvent mixture
Prepare a solvent mixture of acetic acid, methanol and water with a volume ratio of 1:1:1.
5.4.7 Derivatization agent
Prepare an approximately 10 % (m/V) solution of tetraethylborate compound (5.2.11) in tetrahydrofurane
(5.2.8).
NOTE This solution is stable for about three months if stored under an inert-gas blanket.
6 © ISO 2009 – All rights reserved

ISO 23161:2009(E)
5.5 Clean-up
5.5.1 General requirements
A silica or aluminium oxide clean-up is the minimum requirement. Further clean-up steps (aluminium
oxide/silver nitrate, silica/silver nitrate, pyrogenic copper) may be applied if necessary (see Annex B). A
recovery of > 80 % of the internal standards and target compounds shall be achieved for each clean-up step.
5.5.2 Silica gel for the clean-up column
Heat silica gel (5.2.6) for at least 12 h at (500 ± 20) °C on a quartz plate in a muffle furnace. Ensure that the
temperature does not exceed 520 °C.
Allow the plate to cool in an oven to about 200 °C, transfer the silica to a wide-necked glass bottle and allow
cooling to room temperature in a desiccator.
Add water to the cooled silica until a mass fraction of 3 % is reached. Close the bottle and homogenize the
contents for 2 h on a shaker.
5.5.3 Aluminium oxide for the clean-up column
Activate aluminium oxide (5.2.7) by heating to 600 °C for a minimum of 24 h.
Allow to cool in the oven to about 200 °C, transfer the aluminium oxide to a wide-necked glass bottle and
allow cooling to room temperature in a desiccator.
Add water to the cooled aluminium oxide until a mass fraction of 10 % is reached. Close the bottle and
homogenize the contents for 2 h on a shaker.
5.5.4 Clean-up column
Add about 5 g of adsorbent (5.5.2 or 5.5.3) to one column, and add about 3 g of drying agent. Ensure that the
clean-up column is filled homogeneously, for example, by using hexane as a moistening agent during the
filling process.
Commercially pre-packed columns may be used as an alternative if the requirement for recovery is met.
5.5.5 Eluent for extract cleaning with silica gel
A mixture of hexane (5.2.10) with a more polar solvent can be used as an eluent to obtain a quantitative
elution of all organotin compounds. In routine work, about 5 % of acetone (5.2.9) or 20 % of dichloromethane
(5.2.13) was used successfully. The concentration of polar solvent in hexane and the volume of total eluent
should be determined prior to application.
5.5.6 Eluent for extract cleaning with aluminium oxide
Generally, hexane (5.2.10) is used as the eluent. The volume of the eluent should be determined prior to
application.
6 Apparatus
6.1 Requirements for glassware
Customary laboratory glassware shall be used.
All glassware and material that comes into contact with the sample or extract shall be thoroughly cleaned.
ISO 23161:2009(E)
6.2 Sampling apparatus
Sampling devices shall not be a source of contamination. The use of stainless steel, glass or PTFE is
recommended.
NOTE For example, poly(vinyl chloride) (PVC) can contain large amounts of organotin compounds.
Containers shall be inert and appropriate for storing and transport.
The size of the container shall be appropriate to ensure sampling of a suitable amount of solid to provide a
representative sample and facilitate a determination in accordance with this International Standard within the
calibrated working range.
6.3 Additional apparatus
Use ordinary laboratory apparatus and the following.
6.3.1 Centrifuge.
WARNING — The use of organic solvents in centrifuges needs to be assessed for safety reasons.
6.3.2 Glass column for clean-up, e.g. length 15 cm, inner diameter 1 cm, with frit, without a cock.
6.3.3 Shaker.
6.3.4 Ultrasonic bath or horn-type transducer.
6.3.5 Analytical balance, with suitable reading accuracy and range.
6.3.6 Concentration apparatus, e.g. rotary evaporator, Kuderna Danish.
6.3.7 Gas chromatograph, equipped with a high-resolution capillary column of suitable polarity and
Injector, split or splitless, preferably with an automated sampling device (C.1).
6.3.8 Detectors (for typical detector configurations, see C.2). The following detector types may be used for
the measurement of alkylated organotin compounds:
⎯ atomic absorption spectrometer (AAS), quartz oven, tin(Sn) lamp;
⎯ flame photometric detector (FPD), equipped with a cut-off filter of 590 nm or interference filter of 610 nm;
⎯ pulsed flame photometric detector (PFPD) equipped with a large pass-band filter working at 610 nm or
390 nm with a time-selective acquisition;
⎯ mass spectrometer (MS) for electron impact mode (EI-mode);
⎯ atomic emission detector (AED);
⎯ inductively coupled plasma/mass spectrometric detector (ICP/MS).
6.3.9 Data processing system, suitable for the respective detector for acquisition and data evaluation.
7 Procedure
7.1 Sampling and sample pretreatment
[4]
Sample pretreatment should be carried out according to ISO 14507 or ISO 16720.
The use of, for example, stainless steel, PTFE and glass is recommended. Store the sample until
pretreatment in a cool place.
8 © ISO 2009 – All rights reserved

ISO 23161:2009(E)
If the storage time is less than 48 h, store the sample in a dark, cool place until pretreatment.
If the storage time exceeds 48 h, the sample shall be stored frozen (< −18 °C) in the dark.
The laboratory sample should represent the field sample. The amount of sample taken depends on
homogeneity and on the resulting dry mass after preparation. If necessary, select coarse material and sieve to
particle size < 2 mm. Stir with a metal spoon.
For the preparation of freeze-dried samples take, for example, 250 g of original field-moist sample and
proceed with freeze drying in accordance with ISO 16720.
Grind the freeze-dried material, for example, in an agate centrifugal ball mill, to a homogeneous powdery
consistency. Prevent high temperatures in the mill by grinding for a short time.
Determine the dry mass of the freeze-dried material in accordance with ISO 11465.
For the determination of organotin compounds in original field-moist material, take the sieved and stirred
sample as described above. From this homogenized laboratory sample, suitable amounts of sub-samples
(test samples) are taken for subsequent analysis for the determination of organotin compounds and dry mass
in accordance with ISO 11465.
7.2 Sample extraction
7.2.1 General
Add 1 g to 5 g of solid to a container that can be closed [e.g. an Erlenmeyer flask with a ground joint or a
screw-capped vial, polytetrafluoroethylene (PTFE) lined]. It is recommended to choose two samples, varying
in size at least by a factor of 2. Ensure that the mass of analytes in the samples is covered by the working
range.
Pretreat samples of solids, blank solutions (5.4.2) and aqueous calibration solutions (5.4.3) as follows.
7.2.2 Acidic extraction and derivatization of an aliquot
Add an appropriate amount of internal standard mixture and of a solvent mixture of acetic acid:methanol:water
(1:1:1) to the freeze-dried sample to obtain a sample slurry containing 20 % or less of solid material.
Sonicate for 30 min in an ultrasonic bath.
Transfer all the slurry to a centrifuge glass tube and then centrifuge to obtain a liquid/solid phase separation.
The liquid phase is then transferred by a pipette to another container. The extraction procedure is repeated in
the same way by adding half of the volume of extraction solvent mixture used for the first extraction step. The
two extraction solutions are combined prior to derivatization.
For derivatization, add aqueous sodium hydroxide (5.2.2) to an appropriate aliquot (at least 5 ml) of the
extraction solution obtained above and adjust to pH 4,5 using acetic acid (5.2.1). After the addition of 5 ml of
hexane (5.2.10) and a 10 % solution of tetraethylborate compound (5.2.11) in tetrahydrofurane (5.2.8)
(0,5 ml/g of sample taken), the solution is immediately shaken by hand for 1 min. Afterwards, the whole
mixture is shaken for 20 min on a mechanical shaking machine. The procedure is then repeated. The hexane
phases separated are combined and dried over sodium sulfate (5.2.4) and concentrated to 1 ml.
Blank solutions and aqueous calibration solutions (5.4.2 and 5.4.3) shall be treated in the same way as the
samples.
7.2.3 Alkaline treatment and in situ derivatization
Add an appropriate amount of the internal standard solution and water (5.1) to the freeze-dried sample to
obtain a sample slurry with 20 % or less of solid material.
ISO 23161:2009(E)
Shake for about 20 min and ensure that the spiking solution is well distributed in the water or water/solid slurry.
Add methanolic potassium hydroxide solution (1,2 ml/g of sample taken) (see 5.4.4) and 20 ml of hexane
(5.2.10). Heat to 70 °C for 1 h in a closed container (ensure the tightness). Choose a volume of methanolic
potassium hydroxide solution to ensure that the slurry is alkaline. Instead of treatment at 70 °C for 1 h,
ultrasonic treatment (e.g. for a few minutes followed by 1 h of shaking) or treatment overnight at ambient
temperature may be applied.
Add acetic acid (5.2.1) to adjust to a pH of 4,5. Add 10 ml of acetate buffer solution (5.4.5) and shake for
about 1 min. To the buffered solution, add the 10 % solution of tetraethylborate compound in tetrahydrofurane
(see 5.4.6) (0,5 ml/g of sample taken). Shake for about 2 h. Repeat the derivatization procedure and shake for
1 h minimum. Ensure that the phases are well mixed.
Separate the phases using a centrifuge. Collect the hexane layer and dry it with sodium sulfate (5.2.4), and
reduce the volume of the organic phase to 1 ml using a suitable apparatus, but avoid reduction to dryness in
every case.
Blank solutions and aqueous calibration solutions (5.4.2 and 5.4.3) shall be treated in the same way as the
samples.
7.2.4 Separate determination of TTBT in the field-moist sample
The determination of TTBT can be carried out by extraction of the field-moist sample with hexane without the
derivatization step. Therefore, it is possible to take a larger amount of homogenized field-moist sample (5 g or
more) and to use only tetrapropyltin as the internal standard.
7.3 Clean-up of the extract
7.3.1 General
If necessary, sample extracts shall be subjected to an adsorption chromatography clean-up. If the
chromatographic measurements of the target compounds are disturbed by interferences, apply further
appropriate clean-up procedures (see Annex B) provided a recovery of > 80 % of the internal compounds is
achieved for each clean-up step. The reference and blank solutions shall be treated in the same way.
NOTE 1 Triphenyltin elutes later from the clean-up column than the other organotin compounds. If TPHT is not to be
analysed, the clean-up efficiency can be improved by reducing the eluent volume, the water content of the adsorbent or
the concentration of the polar solvent in hexane.
NOTE 2 Boroxins will be formed during derivatization, which can affect the gas chromatography (GC) column. These
are eliminated by silica clean-up with hexane, but can be eluted if acetone is added to the eluent. An alternative separation
method is to shake with sodium hydroxide (NaOH) solution; peralkylated organotin compounds are stable against NaOH
solution.
7.3.2 Silica and aluminium oxide clean-up
Rinse the clean-up column, freshly prepared in accordance with 5.5.4 with 30 ml of hexane (5.2.10).
Transfer the concentrated extract in hexane to the clean-up column (5.5.4). After the extract has penetrated
the top of the adsorbent layer, cautiously add the volume of eluant (5.5.5 or 5.5.6) found to be necessary.
Collect the eluate and reduce the volume of the organic phase to 1 ml using a suitable apparatus, but avoid
reduction to dryness in every case.
If the chromatography turns out to be unacceptable, apply further clean-up procedures (see Annex B).
10 © ISO 2009 – All rights reserved

ISO 23161:2009(E)
7.4 Determination of dry mass
Determine the fraction of dry mass gravimetrically in accordance with ISO 11465. The fraction of dry mass of
original field-moist samples or of freeze-dried materials is expressed as a percentage.
NOTE The following standards can be used for other solids: EN 12880 for sediments or sludges; EN 14346 for
wastes.
7.5 Measurement
7.5.1 Gas chromatographic separation
Optimize the instrument in accordance with the manufacturer's instructions. Ensure at least baseline
separation of the target peaks of interest. Higher resolution is recommended to avoid co-elution of matrix
compounds as far as appropriate (for typical gas chromatographic conditions, see C.1).
The resolution of triphenylethyltin and tricyclohexylethyltin should be at least 0,8.
Prepare injection solutions of blanks, references and samples by adding, for example, 50 µl of injection
standard (see 5.4.1) to the final extract of 1 ml.
Inject an appropriate volume of the prepared sample extracts into the injection port of a gas chromatograph.
Record retention times and the signal intensity of each compound.
Quantify the gas chromatographic signals either as peak areas or as peak heights. In the case of non-
continuous detection (e.g. mass spectrometry), evaluation using peak areas is recommended.
NOTE In this International Standard, only the evaluation using peak areas is described as an example.
7.5.2 Detection and identification
Use an appropriate detector (see 6.3.8) for monitoring the target peaks.
Independent from the detection system, identify the analytes by comparison of the retention times for samples
and references. Minimal requirements for identification are retention times within ± 0,02 min and relative
retention times within ± 0,1 % over the total run of a chromatogram.
Following the retention time criteria, three identification points are necessary. For GC/MS, this procedure is
described in ISO 22892; each individual mass meeting the criteria gives one identification point. Identification
points for other detectors are described in Table 4. If the detector does not give three identification points,
additional points can be obtained by, for instance, using a second column or by pattern recognition (see also
ISO 22892).
Table 4 — Identification points
Detector Number of identification points Remarks
FPD, PFPD 2
MS 1 for each individual mass Refer to ISO 22892
MS/MS 2 for each mass transfer
n
MS 1 for each mass transfer
AED 3 Different spectral lines
ICP/MS 3
AAS 3
ISO 23161:2009(E)
8 Calibration
Calibration is carried out by putting standards, including internal standards, through the whole procedure. The
underivatized organotin compounds are added to water to give the aqueous calibration solutions (5.4.3). The
whole procedure of derivatization, extraction, clean-up and concentration is carried out to establish calibration
curves. At least six calibration solutions at different concentrations should be used to prepare the calibration
curve.
For quantification of monobutyltin and monooctyltin compounds, use monoheptyltin trichloride (MHTCl) as the
internal standard; for dibutyltin and dioctyltin compounds, use diheptyltin dichloride (DHTCl) as the internal
standard; and for tributyltin, triphenyltin and tricyclohexyltin compounds, use tripropyltin chloride (TPTCl) as
the internal standard. The recovery of the internal standards corresponding to each group of organotin
compounds is to be checked to verify complete derivatization and extraction. For quantification of tetrabutyltin
use tetrapropyltin (TTPT) as the internal standard.
Derive from the chromatograms, by integration, the peak areas of the organotin cation derivates, TTBT and
the internal standards. Calculate, for each organotin cation and TTBT, a calibration curve according to
Equation (1) for each working range, using the least-squares linear regression
ya=⋅x+a (1)
where
x is the ratio of the masses of organotin cation, respectively TTBT (m ) and the corresponding internal
i
standard (m ) in the reference solution;
I
m
i
x = (2)
m
I
a is the slope of the calibration curve;
y is the ratio of the peak areas of organotin cation derivates or TTBT and the corresponding internal
standard in the chromatograms of the calibration solutions;
a is the intercept of the calibration curve.
9 Recovery rates of the internal standard compounds
The recoveries facilitate the recognition of bias caused by the procedure or by the matrix of the sample,
allowing indications of the reliability of the procedure to be derived. Ex
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