ISO 13914:2013
(Main)Soil quality — Determination of dioxins and furans and dioxin-like polychlorinated biphenyls by gas chromatography with high-resolution mass selective detection (GC/HRMS)
Soil quality — Determination of dioxins and furans and dioxin-like polychlorinated biphenyls by gas chromatography with high-resolution mass selective detection (GC/HRMS)
ISO 13914:2013 specifies a method for quantitative determination of 17 2,3,7,8-chlorine substituted dibenzo-p-dioxins and dibenzofurans and dioxin-like polychlorinated biphenyls in sludge, treated biowaste, and soil using liquid column chromatographic clean-up methods and GC/HRMS.
Qualité du sol — Détermination des dioxines et furanes comme biphényls polychlorés par chromatographie en phase gazeuse avec spectrométrie de masse à haute résolution (CG/SMHR)
General Information
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Standards Content (Sample)
INTERNATIONAL ISO
STANDARD 13914
First edition
2013-12-01
Soil quality — Determination of
dioxins and furans and dioxin-like
polychlorinated biphenyls by gas
chromatography with high-resolution
mass selective detection (GC/HRMS)
Qualité du sol — Détermination des dioxines et furanes comme
biphényls polychlorés par chromatographie en phase gazeuse avec
spectrométrie de masse à haute résolution (CG/SMHR)
Reference number
ISO 13914:2013(E)
©
ISO 2013
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ISO 13914:2013(E)
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ISO 13914:2013(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 2
3 Abbreviated terms . 2
4 Principle . 2
5 Reagents . 3
5.1 Chemicals . 3
5.2 Standards . 3
6 Apparatus and materials. 3
6.1 General . 3
6.2 Equipment for sample preparation . 3
6.3 Soxhlet extractor . 3
6.4 Clean-up apparatus . 4
6.5 Concentration apparatus . 4
6.6 Other equipment . 4
7 Sample storage and sample pretreatment . 5
7.1 Sample storage . 5
7.2 Sample pretreatment . 5
8 Extraction and clean-up . 5
8.1 General . 5
8.2 Extraction . 6
8.3 Clean-up . 6
8.4 Final concentration of cleaned sample extract. 7
8.5 Addition of recovery standard . 8
9 GC/HRMS analysis . 8
9.1 General . 8
9.2 Gas chromatographic analysis . 8
9.3 Mass spectrometric detection . 8
9.4 Minimum requirements for identification of PCDF/PCDD and PCB . .10
9.5 Minimum requirements for quantification of PCDF/PCDD and PCB .10
9.6 Calibration of the GC/HRMS system .11
9.7 Quantification of GC/HRMS results .13
10 Precision .15
11 Test report .15
Annex A (informative) Toxic equivalent factors.16
Annex B (informative) Repeatability and reproducibility data .18
Annex C (informative) Examples of extraction and clean-up methods .21
Annex D (informative) Examples of operation of GC/HRMS determination .29
Bibliography .33
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ISO 13914:2013(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 on the meaning of ISO specific terms and expressions related to conformity
assessment, as well as information about ISO’s adherence to the WTO principles in the Technical Barriers
to Trade (TBT) see the following URL: Foreword - Supplementary information
The committee responsible for this document is ISO/TC 190, Soil quality, Subcommittee SC 3, Chemical
methods and soil characteristics.
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ISO 13914:2013(E)
Introduction
Two groups of related chlorinated aromatic ethers are known as polychlorinated dibenzo-p-
dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). They consist of a total of 210 individual
substances (congeners): 75 PCDDs and 135 PCDFs.
A group of chlorinated aromatic compounds similar to polychlorinated dibenzo-p-dioxins (PCDDs) and
polychlorinated dibenzofurans (PCDFs) is known as polychlorinated biphenyls (PCBs) which consist of
209 individual substances.
PCDDs and PCDFs can form in the combustion of organic materials. They also occur as undesirable by-
products in the manufacture or further processing of chlorinated organic chemicals. PCDDs/PCDFs
enter the environment via these emission paths and through the use of contaminated materials.
In fact, they are universally present at very small concentrations. The 2,3,7,8-substituted congeners
are toxicologically significant. Toxicologically much less significant than the tetrachlorinated to
octachlorinated dibenzo-p-dioxins/dibenzofurans are the 74 monochlorinated to trichlorinated
dibenzo-p-dioxins/dibenzofurans.
PCBs have been produced over a period of approximately 50 y until the end of the 1990s for the purpose
of different uses in open and closed systems, e.g. as electrical insulators or dielectric fluids in capacitors
and transformers, as specialized hydraulic fluids, or as a plasticizer in sealing material. Worldwide,
more than 1 million tons of PCBs were produced.
PCDD/Fs as well as PCBs are emitted during thermal processes such as waste incineration. In 1997, a group
of experts of the World Health Organization (WHO) fixed toxicity equivalent factors (TEF) for PCDDs and
12 PCBs, known as dioxin-like PCBs (see Annex A). These 12 dioxin-like PCBs consist of four non-ortho
PCBs and eight mono-ortho PCBs (no or only one chlorine atoms in 2-, 2’-, 6- and 6’-position), having a
planar or mostly planar structure. Dioxin-like PCBs can contribute considerably to the total WHO-TEQ.
Only skilled operators who are trained in handling highly toxic compounds should apply the method
described in this International Standard.
This International Standard is applicable for several types of matrices and validated for municipal sludge
(see Annex B for the results of the validation).
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INTERNATIONAL STANDARD ISO 13914:2013(E)
Soil quality — Determination of dioxins and furans
and dioxin-like polychlorinated biphenyls by gas
chromatography with high-resolution mass selective
detection (GC/HRMS)
WARNING — Persons using this International Standard should be familiar with usual 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.
IMPORTANT — It is absolutely essential that tests conducted according to this International
Standard be carried out by suitably trained staff.
1 Scope
This International Standard specifies a method for quantitative determination of 17 2,3,7,8-chlorine
substituted dibenzo-p-dioxins and dibenzofurans and dioxin-like polychlorinated biphenyls in sludge,
treated biowaste, and soil using liquid column chromatographic clean-up methods and GC/HRMS.
The analytes to be determined with this International Standard are listed in Table 1.
Table 1 — Analytes and their abbreviations
Substance Abbreviation
Tetrachlorodibenzo-p-dioxin TCDD
Pentachlorodibenzo-p-dioxin PeCDD
Hexachlorodibenzo-p-dioxin HxCDD
Heptachlorodibenzo-p-dioxin HpCDD
Octachlorodibenzo-p-dioxin OCDD
Tetrachlorodibenzofuran TCDF
Pentachlorodibenzofuran PeCDF
Hexachlorodibenzofuran HxCDF
Heptachlorodibenzofuran HpCDF
Octachlorodibenzofuran OCDF
Polychlorinated biphenyl PCB
Trichlorobiphenyl TCB
Tetrachlorobiphenyl TeCB
Pentachlorobiphenyl PeCB
Hexachlorobiphenyl HxCB
Heptachlorobiphenyl HpCB
Decachlorobiphenyl DecaCB
The limit of detection depends on the kind of sample, the congener, the equipment used, and the quality
of chemicals used for extraction and clean-up. Under the conditions specified in this International
Standard, limits of detection better than 1 ng/kg (expressed as dry matter) can be achieved.
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ISO 13914:2013(E)
This method is “performance based”. It is permitted to modify the method if all performance criteria
given in this method are met.
NOTE In principle, this method can also be applied for sediments, mineral wastes, and for vegetation. It is
the responsibility of the user of this International Standard to validate the application for these matrices. For
measurement in complex matrices like fly ashes adsorbed on vegetation, it can be necessary to further improve
the clean-up. This can also apply to sediments and mineral wastes.
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.
ISO 14507, Soil quality — Pretreatment of samples for determination of organic contaminants
3 Abbreviated terms
PCB polychlorinated biphenyls
PCDD/PCDF or PCDD/F polychlorinated dibenzo-p-dioxins/dibenzofurans
I-TEF NATO/CCMS international toxic equivalent factor, proposed by NATO-CCMS in 1988 (for a
detailed description, see Annex A)
I-TEQ international toxic equivalent, obtained by multiplying the mass determined
with the corresponding I-TEF including PCDDs and PCDFs (for a detailed
description, see Annex A). Should only be used for comparison with older
data
WHO-TEF toxic equivalent factor, proposed by WHO in 2005 (for detailed description,
see Annex A)
WHO-TEQ toxic equivalent, obtained by multiplying the mass determined with the
corresponding WHO-TEF including PCDD, PCDF, and PCB (for detailed
description, see Annex A). WHO-TEQ , WHO-TEQ should be used to
PCB PCDD/F
distinguish different compound classes
4 Principle
This International Standard is based on the use of gas chromatography/mass spectrometry combined
with the isotope dilution technique to enable the separation, detection, and quantification of PCDD/PCDF
and dioxin-like PCB in sludge, biowaste, and soil. For the isotope dilution method, 17 labelled PCDD/F
and 12 labelled PCB internal standards are used. The extracts for the GC-MS measurements contain
one or two recovery standards. The gas chromatographic parameters offer information which enables
the identification of congeners (position of chlorine substitutes) whereas the mass spectrometric
parameters enable the differentiation between isomers with different numbers of chlorine substitutes
and between dibenzo-p-dioxins, furans, and PCBs.
13
C -labelled PCDD/F and PCB congeners are added to the sample prior to extraction and GC/HRMS
12
measurement. Losses during extraction and clean-up are detected and compensated by using these
added congeners as internal standards for quantification together with recovery standards which are
added just before the GC/HRMS analysis. For the determination of these substances, it is necessary to
separate PCBs from PCDDs/PCDFs and vice versa.
The main purpose of the clean-up procedure of the raw sample extract is the removal of sample matrix
components, which can overload the separation method, disturb the quantification, or otherwise
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ISO 13914:2013(E)
severely impact the performance of the identification and quantification method and the separation of
PCDD/F from dioxin-like PCB. Furthermore, the enrichment of the analytes in the final sample extract
is achieved. Extraction procedures are usually based on Soxhlet or equivalent extraction methods of
dried, preferably freeze-dried, samples. Sample clean-up is usually carried out by multi-column liquid
chromatographic techniques using different adsorbents. The determination of PCDD/Fs and PCBs is
based on quantification by the isotope dilution technique using GC/HRMS.
5 Reagents
5.1 Chemicals
Solvents used for extraction and clean-up shall be of pesticide grade or equivalent quality and checked
for blanks. Adsorbents like aluminium oxide, silica gel, diatomaceous earth, and others used for clean-up
shall be of analytical grade quality or better and pre-cleaned and activated if necessary.
NOTE See Annex C for a specific list of solvents and chemicals.
5.2 Standards
13
— C -spiking solution for PCDD/F (internal standard);
12
13
— C -spiking solution for PCB (internal standard);
12
— calibration solutions PCDD/F;
— calibration solutions PCB;
— recovery standard PCDD/F;
— recovery standard PCB.
NOTE See Annex C for examples of concentration of the standard solutions.
6 Apparatus and materials
6.1 General
The apparatus and materials listed below are meant as minimum requirements for “conventional”
sample treatment with Soxhlet extraction and column chromatographic clean-up. Additional apparatus
and materials may be necessary due to different methods of sample extraction and clean-up methods.
6.2 Equipment for sample preparation
6.2.1 Laboratory fume hood, of sufficient size to contain the sample preparation equipment listed below.
6.2.2 Desiccator.
6.2.3 Balances, consisting of an analytical type capable of weighing 0,1 mg and a top-loading type
capable of weighing 10 mg.
6.3 Soxhlet extractor
6.3.1 Soxhlet, 50 mm internal diameter, 150 ml or 250 ml capacity with 500 ml round bottom flask.
6.3.2 Thimble, 43 mm × 123 mm, to fit Soxhlet.
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ISO 13914:2013(E)
6.3.3 Hemispherical heating mantle, to fit 500 ml round-bottom flask.
6.4 Clean-up apparatus
6.4.1 Disposable pipettes, either disposable Pasteur pipettes, or disposable serological pipettes.
6.4.2 Glass chromatographic columns, of the following sizes:
— 150 mm length × 8 mm internal diameter, with coarse-glass frit or glass-wool plug, 250 ml reservoir,
and glass or polytetrafluoroethylene (PTFE) stopcock;
— 200 mm length × 15 mm internal diameter, with coarse-glass frit or glass-wool plug, 250 ml reservoir,
and glass or PTFE stopcock;
— 300 mm length × 25 mm internal diameter, with coarse-glass frit or glass-wool plug, 300 ml
reservoir, and glass or PTFE stopcock.
6.4.3 Oven, capable of maintaining a constant temperature (±5 °C) in the range of 105 °C to 450 °C for
baking and storage of adsorbents.
6.5 Concentration apparatus
6.5.1 Rotary evaporator, equipped with a variable temperature water bath and:
— a vacuum source for the rotary evaporator equipped with a shutoff valve at the evaporator and
vacuum gauge;
— a recirculating water pump and chiller, providing cooling water of (9 ± 4) °C (use of tap water for
cooling the evaporator wastes large volumes of water and can lead to inconsistent performance as
water temperatures and pressures vary);
— round-bottom flask, 100 ml and 500 ml or larger, with ground-glass fitting compatible with the
rotary evaporator.
6.5.2 Nitrogen blowdown apparatus, equipped with either a water bath controlled in the range of
30 °C to 60 °C or a heated stream of nitrogen, installed in a fume hood.
1)
6.5.3 Kuderna-Danish concentrator.
6.5.4 Sample vials, of the following types:
— amber glass, nominated volume 2 ml to 5 ml, with PTFE-lined screw cap;
— glass, 0,3 ml, conical, with PTFE-lined screw or crimp cap.
6.6 Other equipment
6.6.1 Gas chromatograph, equipped with a splitless or on-column or temperature-programmed
injection port for use with capillary columns, and an oven temperature programme which enables
isothermal hold.
1) Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this International Standard and does not constitute an endorsement by ISO of this product.
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ISO 13914:2013(E)
6.6.2 GC column for PCDDs/PCDFs and for isomer specificity for 2,3,7,8-TCDD (e.g. 60 m
length × 0,32 mm internal diameter; 0,25 µm; 5 % phenyl, 94 % methyl, 1 % vinyl silicone bonded-phase
fused-silica capillary column).
6.6.3 Mass spectrometer, 28 eV to 80 eV electron impact ionization, capable of repetitively selectively
monitoring 12 exact masses minimum at high resolution (>10 000) during a period of approximately 1 s.
6.6.4 Data system, capable of collecting, recording, and storing mass spectrometric data.
7 Sample storage and sample pretreatment
7.1 Sample storage
Samples should be stored in suitable containers with an appropriate closure material such as
polytetrafluoroethylene (PTFE). Samples to be frozen can be stored in aluminium containers pre-
cleaned by heating to 450 °C for a minimum of 4 h or by rinsing with a non-chlorinated solvent.
Samples should be kept cold (<8 °C) and in the dark. The sample pretreatment should take place within
3 d of sampling. Alternatively, samples can be frozen (–18 °C) directly after sampling and kept frozen
before sample pretreatment.
7.2 Sample pretreatment
Drying and homogenization should be carried out according to ISO 14507, if not otherwise specified.
Store the ground material in a desiccator or a tightly closed glass container.
8 Extraction and clean-up
8.1 General
In this International Standard, the minimum requirements for extraction and clean-up to be met are
described as well as examples of operation. The analyst can use any of the procedures given below and
in Annex C or any suitable alternative procedures.
The determination of PCDDs/PCDFs is based on quantification by the isotope dilution technique using
13
GC/HRMS. C -labelled 2,3,7,8-chlorine substituted PCDD/PCDFs congeners are added at different
12
stages of the whole method. Losses during extraction and clean-up can be detected and compensated by
using these added congeners as internal standards for quantification together with recovery standards
which are added just before the GC/HRMS analysis. However, due to possible differences in the binding
13
and adsorption characteristics between the native PCDDs/PCDFs and the C -labelled congeners,
12
which are added during analysis, complete substantiation of the extraction efficiency and compensation
of losses during clean-up is not ensured. Therefore, in addition, the applied methods shall be validated
thoroughly. Examples of well-proven extraction and clean-up methods are given in Annex C.
The main purpose of the clean-up procedure of the raw sample extract is the removal of sample matrix
components, which can overload the separation method, disturb the quantification, or otherwise
severely impact the performance of the identification and quantification method and to separate
dioxin-like PCB from PCDD/F. Furthermore, an enrichment of the analytes in the final sample extract is
achieved. Extraction procedures are usually based on Soxhlet extraction of the <2 mm fraction of the
dry and ground or sieved solid sample. Sample clean-up is usually carried out by multi-column liquid
chromatographic techniques using different adsorbents.
In principle, any clean-up method can be used which recovers the analytes in sufficient quantities.
Furthermore, the final sample extract shall not affect adversely the performance of the analytical system
or the quantification step. However, all applied methods shall be tested thoroughly and shall pass a
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ISO 13914:2013(E)
set of method validation requirements before they can be employed. In addition, the verification of the
method performance for each single sample shall be part of the applied quality assurance protocol.
8.2 Extraction
The sample amount used for extraction can vary from 5 g to 50 g depending on the expected level of
contamination.
13
The internal standard consisting of C -labelled congeners listed in Table 2 shall be added directly
12
into the sample before extraction.
The extraction procedure is carried out using Soxhlet extraction with toluene. The duration of extraction
should be adjusted according to the kind and amount of sample used. The minimum requirement is
50 extraction cycles or approximately 12 h.
Other solvents or other methods like pressurized liquid extraction can also be used but shall be of
proven equal performance.
13
Table 2 — C -labelled congeners included in the internal standard
12
13
C -spiking solution — internal standard
12
PCDD/F congeners PCB congeners
13 13
2,3,7,8- C -TCDD C -PCB-77
12 12
13 13
1,2,3,7,8- C -PeCDD C -PCB-81
12 12
13 13
1,2,3,4,7,8- C -HxCDD C -PCB-126
12 12
13 13
1,2,3,6,7,8- C -HxCDD C -PCB-169
12 12
13
1,2,3,7,8,9- C -HxCDD
12
13 13
1,2,3,4,6,7,8- C -HpCDD C -PCB-105
12 12
13 13
C -OCDD C -PCB-114
12 12
13
C -PCB-118
12
13 13
2,3,7,8- C -TCDF C -PCB-123
12 12
13 13
1,2,3,7,8- C -PeCDF C -PCB-156
12 12
13 13
2,3,4,7,8- C -PeCDF C -PCB-157
12 12
13 13
1,2,3,4,7,8- C -HxCDF C -PCB-167
12 12
13 13
1,2,3,6,7,8- C -HxCDF C -PCB-189
12 12
13
2,3,4,6,7,8- C -HxCDF
12
13
1,2,3,7,8,9- C -HxCDF
12
13
1,2,3,4,6,7,8- C -HpCDF
12
13
1,2,3,4,7,8,9- C -HpCDF
12
13
C -OCDF
12
8.3 Clean-up
8.3.1 General
Clean-up methods shall prepare the sample extract in an appropriate manner for the subsequent
quantitative determination. Clean-up procedures shall concentrate PCDD/Fs and dioxin-like PCBs in the
extracts and remove interfering matrix components present in the raw extract.
Proven clean-up procedures shall be used including usually two or more of the following techniques which
can be combined in different orders. A detailed description of some of the procedures is given in Annex C.
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ISO 13914:2013(E)
Other methods can also be used but shall be of proven equal performance as the techniques described below.
8.3.2 Gel permeation chromatography
The interesting molecular weight range for PCDD/Fs and dioxin-like PCBs of 200 g/mol to 500 g/mol
can be isolated from larger molecules and polymers which might overload other clean-up methods. This
method can also be used for the removal of sulfur.
8.3.3 Multilayer column
Multilayer column liquid chromatography using s
...
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