ISO/PRF 13914

ISO/DIS PRF 13914:2025(en)
ISO/TC 190/SC 3/WG 6
Secretariat: DIN
Date: 2025-11-2412-15
Soil, treated biowaste and sludge — Determination of dioxins and
furans and dioxin-like polychlorinated biphenyls by gas
chromatography with mass selective detection (high resolution mass
spectrometry, HRMS, and tandem mass spectrometry, MS/MS)")
PROOF
ISO #####-#:####(X/PRF 13914:2025(en)
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ii
ISO/DIS PRF 13914:2025(en)
Contents
Foreword . iv
Introduction . v
1 Scope . 1
2 Normative references . 2
3 Terms and definitions . 2
4 Abbreviated terms . 3
5 Principle . 3
6 Reagents . 4
6.1 Chemicals . 4
6.2 Standards . 4
7 Apparatus . 4
7.1 General . 4
7.2 Equipment for sample preparation . 4
7.3 Soxhlet extractor . 4
7.4 Clean-up apparatus . 5
7.5 Concentration apparatus . 5
7.6 Other equipment . 6
8 Sample storage and sample pretreatment . 6
8.1 Sample storage . 6
8.2 Sample pretreatment . 6
9 Extraction and clean-up . 6
9.1 General . 6
9.2 Extraction . 7
9.3 Clean-up . 8
9.4 Final concentration of cleaned sample extract . 9
9.5 Addition of recovery standard . 9
10 GC/MS (HRMS or MS/MS) analysis . 10
10.1 General . 10
10.2 Gas chromatographic analysis . 10
10.3 Mass spectrometric detection . 10
10.4 Minimum requirements for identification of PCDF/PCDD and PCB . 13
10.5 Minimum requirements for quantification of PCDF/PCDD and PCB . 14
10.6 Calibration of the GC/MS system (HRMS or MS/MS) . 15
10.7 Quantification of GC/MS (HRMS or MS/MS) results . 16
11 Expression of results . 19
12 Precision . 19
13 Test report . 19
Annex A (informative) Toxic equivalency factor (TEF) . 21
Annex B (informative) Example of extraction and clean-up methods . 23
Annex C (informative) Examples of operation of GC/HRMS determination . 32
Annex D (informative) Repeatability and reproducibility data . 36
Bibliography . 46

iii
ISO #####-#:####(X/PRF 13914:2025(en)
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
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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).
ISO draws attention to the possibility that the implementation of this document may involve the use of (a)
patent(s). ISO takes no position concerning the evidence, validity or applicability of any claimed patent rights
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related to conformity assessment, as well as information about ISO'sISO’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 190, Soil quality, Subcommittee SC 3, Chemical
and physical characterization.
This third edition cancels and replaces the second edition (ISO 13914:2023), which has been technically
revised.
The main changes are as follows:
— — tandem mass spectroscopy (MS/MS) has been added as an alternative gas chromatographic analysis
to high resolution mass spectrometry (HRMS);
— — validation results for MS/MS have been added;
— — document has been editorially revised.
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.
© ISO #### 2025 – All rights reserved
iv
ISO/DIS PRF 13914:2025(en)
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 PCDD and 135 PCDF of which 17 have chlorine substitution in the 2,3,7,8-positions.
A group of chlorinated aromatic compounds similar to polychlorinated dibenzo-p-dioxins (PCDDs) and
polychlorinated dibenzofurans (PCDFs) is known as polychlorinated biphenyls (PCBs) which consists of 209
individual substances.
PCDD and PCDF 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. PCDD/PCDF enter the environment
mainly 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.
PCB have been produced over a period of approximately 50 years 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 and as a plasticizer in sealing material. Worldwide more than one
million tons of PCB were produced.
PCDD/PCDF as well as PCB are emitted during thermal processes, for example waste incineration. In 1997 a
group of experts of the World Health Organization (WHO) fixed toxicity equivalent factors (TEF) for PCDD and
twelve PCB, known as dioxin-like PCB (dl-PCB, see Annex AAnnex A).). These twelve dioxin-like PCB consist
of four non-ortho PCB and eight mono-ortho PCB (no or only one chlorine atoms in 2-, 2’-, 6- and 6’-position),
having a planar or mostly planar structure. Dioxin-like PCB 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 document.
v
DRAFT International Standard ISO/FDIS 21068-2:2025(en)

Soil, treated biowaste and sludge — Determination of dioxins and
furans and dioxin-like polychlorinated biphenyls by gas
chromatography with mass selective detection (HRMS orhigh
resolution mass spectrometry, HRMS, and tandem mass spectrometry,
MS/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.
IMPORTANT — It is absolutely essential that tests — Tests conducted according to this document shall
be carried out by suitably trained staff.
1 Scope
This document 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 gas chromatography/high resolution mass
spectrometry (GC/HRMS). Detection by tandem mass spectrometry (MS/MS) can be used in an equivalent
way.
The analytes that can be determined with the method specified in this document are listed in Table 1Table 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
a
Trichlorobiphenyl ( ) TCB
Tetrachlorobiphenyl TeCB
Pentachlorobiphenyl PeCB
Hexachlorobiphenyl HxCB
Substance Abbreviation
Heptachlorobiphenyl HpCB
a
Decachlorobiphenyl ( ) DecaCB
(a ) Groups containing no dl-PCB, given for informative purpose only.
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 document, limits of detection
better than 1 ng/kg (expressed as dry matter) can be achieved.
This method is “performance based”. The method can be modified if all performance criteria given in this
method are met.
This document is applicable for several types of matrices and validated for municipal sludge (see also
Annex AAnnex A)) for the results of the validation).
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 document to validate the application for these matrices. For measurement in complex
matrices such as 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 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.
EN 15934ISO 11465, Sludge, treated biowaste, soil and waste — Calculation of dry matter fraction after
determinationsolid environmental matrices — Determination of dry residue or water content and calculation of
the dry matter fraction on a mass basis
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
ISO and IEC maintain terminology databases for use in standardization at the following addresses:
— — ISO Online browsing platform: available at https://www.iso.org/obp
— — IEC Electropedia: available at https://www.electropedia.org/
3.1 3.1
internal standard
C -labelled analyte analogue added to samples prior to extraction against which the concentrations of
native analytes are calculated
[SOURCE: ISO 18073:2004, 3.1.5]
, modified — “2,3.2,7,8-PCDD/PCDF” and “PCDDs and PCDFs” were changed to “analytes”.]
3.2
recovery standard
C -labelled dl-polychlorinated biphenyl (PCB) and polychlorinated dibenzo-p-dioxin/polychlorinated
dibenzofuran (PCDD/PCDF,), added before injection into the GC
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
[SOURCE: ISO 18073:2004, 3.1.12], modified — “2,3,7,8-chloro-substituted PCDD/PCDF” was changed to “dl-
PCB and PCDD/PCDF”.]
4 Abbreviated terms
dl-PCB dioxin-like polychlorinated biphenyls
MRM multiple reaction monitoring
PCDD/PCDF or PCDD/F Polychlorinated dibenzo-p-dioxins/dibenzofurans
International toxic equivalent
I-TEQ
Note: the I-TEQ is obtained by multiplying the mass determined with the
corresponding I-TEF including PCDD and PCDF (for 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 D). WHO-TEQ , WHO-TEQ should be used to
PCB PCDD/PCDF
distinguish different compound classes
Note: WHONOTE 1 The I-TEQ PCBis obtained by multiplying the mass determined with the corresponding
I-TEF including PCDD and PCDF (for detailed description, see Annex A). Should only be used for comparison with older
data.
NOTE 2 For detailed description of WHO-TEF, see Annex A.
NOTE 3 For detailed description of WHO-TEQ, see Annex D PCDD. WHO-TEQPCB and WHO-TEQPCDD/PCDF should
be used to distinguish different compound classes.
5 Principle
This document 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, at least 16 out of 17 labelled PCDD/PCDF and
12 labelled PCB internal standards are used. The extracts for the GC-MS measurements contain at least one
recovery standard per group, i.e. at least 1one 13C12-PCDD/PCDF and 1one 13C12-PCB. The use of more
recovery standards is recommendable. 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 PCB.
C -labelled PCDD/PCDF and PCB congeners are added to the sample prior to extraction and GC/MS (HRMS
or MS/MS) 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 analysis. For the determination of these substances it is necessary to separate PCB from
PCDD/PCDF 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 severely
impact the performance of the identification and quantification method and the separation of PCDD/PCDF
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/PCDF and PCB is based on quantification
by the isotope-dilution technique using GC/MS (HRMS or MS/MS).
6 Reagents
6.1 Chemicals
Solvents used for extraction and clean-up shall be of pesticide grade or equivalent quality and checked for
blanks. Adsorbents such as 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 BAnnex B for a specific list of solvents and chemicals.
6.2 Standards
— — C -spiking solution for PCDD/PCDF (internal standard);
— — C -spiking solution for PCB (internal standard);
— — calibration solutions PCDD/PCDF;
— — calibration solutions PCB;
13 13
— — C-spiking solution as recovery standard for PCDD/PCDF (remark: typical C-congeners used as
13 13 13
recovery standards are C -123789-HexaCDD, C -1234-TetraCDD, C -1234-TetraCDD or -DF);
12 12 12
— — C -spiking solution as recovery standard for dl-PCB.
NOTE See Annex BAnnex B for examples of concentration of the standard solutions.
7 Reagents and Apparatus
7.1 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
can be necessary due to different methods of sample extraction and clean-up methods.
7.2 Equipment for sample preparation
7.2.1 7.2.1 Laboratory fume hood, of sufficient size to contain the sample preparation equipment listed
below.
7.2.2 7.2.2 Desiccator.
7.2.3 7.2.3 Balances, consisting of an analytical type capable of weighing 0,1 mg and a top-loading type
capable of weighing 10 mg.
7.2.4 7.2.4 Snyder column.
7.3 Soxhlet extractor
Alternatively, other automated Soxhlet-like devices can be used, if applicable. It is the responsibility of the
user of this document to validate the application for these alternative devices.
7.3.1 7.3.1 Soxhlet, 50 mm internal diameter, 150 ml or 250 ml capacity with 500 ml round bottom flask.
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
7.3.2 7.3.2 Thimble, 43 mm × 123 mm, to fit Soxhlet.
7.3.3 7.3.3 Hemispherical heating mantle, to fit 500 ml round-bottom flask.
7.4 Clean-up apparatus
7.4.1 7.4.1 Disposable pipettes, either disposable Pasteur pipettes, or disposable serological pipettes.
7.4.2 7.4.2 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.
7.4.3 7.4.3 Oven, capable of maintaining a constant temperature (±5 °C) in the range of 105 °C to 450 °C
for baking and storage of adsorbents.
7.5 Concentration apparatus
Alternatively, other evaporation devices can be used, if applicable. It is the responsibility of the user of this
document to validate the application for these alternative evaporation devices.
7.5.1 7.5.1 Rotary evaporator, equipped with a variable temperature water bath and:
— — vacuum source for rotary evaporator equipped with shutoff valve at the evaporator and vacuum
gauge;
— — 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.
7.5.2 7.5.2 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 or of another suitable inert gas, installed in a fume hood.
1 1)
7.5.3 7.5.3 Kuderna-Danish concentrator.
7.5.4 7.5.4 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.

Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this document and does not constitute an endorsement by ISO of this product.
1)
Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this document and does not constitute an endorsement by ISO of this product.
7.6 Other equipment
7.6.1 7.6.1 Gas chromatograph, equipped with a splitless or on-column or temperature programmed
injection port for the use with capillary columns, and an oven temperature programme which enables
isothermal hold.
7.6.2 7.6.2 GC column for PCDD/PCDF 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).
7.6.3 7.6.3 Mass spectrometer, 28 eV to 80 eV electron impact ionization, capable of repetitively
selectively monitoring of twelve exact masses minimum during a period of approximately 1 s either at high
mass resolution (≥10 000 at 10 % peak valley or equivalent) or 70 eV electron impact ionization following
analysis utilizing ana triple quad MS/MS system on specific multiple reaction monitoring (MRM) (see
Table 5Table 5).).
7.6.4 7.6.4 Data system, capable of collecting, recording, and storing mass spectrometric data.
8 Sample storage and sample pretreatment
8.1 Sample storage
Samples should be stored in suitable containers with an appropriate closure material such as
polytetrafluoroethylene (PTFE). Samples to be frozen may be stored in aluminium containers pre-cleaned by
heating to 450 °C for minimum 4 h or by rinsing with a non-chlorinated solvent.
Samples should be kept cool and dark where necessary in order to avoid alteration of sample constitution e.g.
in combination with water content determination.
8.2 Sample pretreatment
Drying and homogenization should be carried out according to EN 16179 [20],, if not otherwise specified.
Store the ground material in a desiccator or a tightly closed glass container.
Determination of water content shall be carried out according to EN 15934ISO 11465 or any equivalent
method which is internationally standardized.
9 Extraction and clean-up
9.1 General
In this document, the minimum requirements for extraction and clean-up to be met are described as well as
examples of operation. The analyst may use any of the procedures given below and in Annex CAnnex C or any
suitable alternative procedures.
The determination of PCDD/PCDF and dl-PCB is based on quantification by the isotope-dilution technique
using GC/MS (HRMS or MS/MS). C -labelled dl-PCB and 2,3,7,8-chlorine substituted PCDD/PCDF congeners
are added at different 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/MS analysis. However, due to possible differences in the binding
and adsorption characteristics between the native analytes and the C -labelled congeners, 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 CAnnex C.
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
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/PCDF. Furthermore, an enrichment of the analytes in the final sample extract is achieved. Extraction
procedures are normally 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 set of method
validation requirements before they can be employed (see Annex DAnnex D).). In addition, the verification of
the method performance for each single sample shall be part of the applied quality assurance protocol.
9.2 Extraction
The sample amount used for extraction can vary from 5 g to 50 g depending on the expected level of
contamination.
The internal standard consisting of C -labelled congeners listed in Table 2Table 2 shall be added directly
onto the sample before extraction.
The extraction procedure is carried out using Soxhlet extraction with toluene. Duration of extraction should
be adjusted according to kind and amount of sample used. The minimum requirement is 50 extraction cycles
(approximately 12 h).
Other solvents or other methods such as pressurized liquid extraction can also be used but shall be of proven
equal performance.
Table 2 — C labelled congeners included in the internal standard
C-spiking solution – internal standard
PCDD/PCDF congeners PCB congeners
13 13
2,3,7,8- C12-TCDD C12-PCB 77
13 13
1,2,3,7,8- C12-PeCDD C12-PCB 81
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 a
1,2,3,7,8,9- C -HxCDD
13 13
1,2,3,4,6,7,8- C12-HpCDD C12-PCB 105
13 13
C12-OCDD C12-PCB 114
C -PCB 118
13 13
2,3,7,8- C -TCDF C -PCB 123
12 12
13 13
1,2,3,7,8- C12-PeCDF C12-PCB 156
13 13
2,3,4,7,8- C12-PeCDF C12-PCB 157
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
2,3,4,6,7,8- C12-HxCDF
1,2,3,7,8,9- C12-HxCDF
C-spiking solution – internal standard
PCDD/PCDF congeners PCB congeners
1,2,3,4,6,7,8- C12-HpCDF
1,2,3,4,7,8,9- C12-HpCDF
13C12-OCDF
a 1,2,3,7,8,9- C -HxCDD is traditionally widely used as a recovery standard. Its use as internal standard is therefore
not mandatory.
9.3 Clean-up
9.3.1 General
Clean-up methods shall prepare the sample extract in an appropriate manner for the subsequent quantitative
determination. Clean-up procedures shall concentrate PCDD/PCDF and dioxin-like PCB in the extracts and to
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 DAnnex D.
Other methods can also be used but shall be of proven equal performance as the techniques described below.
9.3.2 Gel permeation chromatography
The interesting molecular weight range for PCDD/PCDF and dioxin-like PCB 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 sulphur.
9.3.3 Multilayer column
Multilayer column liquid chromatography using silica with different activity grades and surface modifications
is used. Compounds with different chemical properties than PCDD/PCDF and dl-PCB can be removed.
9.3.4 Sulphuric acid treatment
A direct treatment of the sample extract with sulphuric acid is possible but is not recommended due to risk of
accident. Furthermore, this shall be carried out very carefully to avoid losses of PCDD/PCDF and dl-PCB on
the formed carboniferous surfaces.
9.3.5 Activated carbon column
Column adsorption chromatography using activated carbon can be used to separate planar PCDD/PCDF and
coplanar PCB molecules from mono-ortho PCB and other interfering non-planar molecules.
9.3.6 Aluminium oxide column
Column liquid chromatography on aluminium oxide of different activity grade and acidity or basicity is used.
Interfering compounds with small differences in polarity or structure compared to PCDD/PCDF and dioxin-
like PCB can be removed.
Additionally, aluminium oxide columns can be used to separate PCDD/PCDF from dioxin-like PCB.
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
9.3.7 Removal of sulphur
The removal of sulphur can be achieved by refluxing the extract with powdered copper or by gel permeation
chromatography.
9.4 Final concentration of cleaned sample extract
To achieve sufficient detection limits, the cleaned sample extract shall be concentrated to a volume in the order
of 25 µl to 100 µl before quantification. The final solvent shall be nonane, toluene or another high boiling
solvent.
Though PCDD/PCDF and dl-PCB have rather high boiling points (>320 °C) vapour phase transfer mechanisms
and aerosol formation during solvent evaporation can lead to substantial losses when concentrating volumes
below 10 ml. Depending on the method to be used for solvent volume reduction the following precautions
shall be taken into consideration.
a) a) Rotary evaporators:
Losses can be substantial when reducing solvent volumes below 10 ml. Counter measures are the use of
controlled vacuum conditions according to the vapour pressure and boiling point of the solvent, addition
of a high-boiling solvent as a keeper as well as the use of specially shaped vessels (e.g. V-shaped).
b) b) Counter gas flow evaporators:
Volumes should not be reduced to less than 1 ml.
c) c) Nitrogen flow:
An excessive flow of nitrogen which disturbs the solvent surface should be avoided. The vial shape has
also some influence on possible losses. V-shaped vials or vial inserts shall be used for volume reductions
below around 200 µl.
2 2)
d) d) Kuderna Danish : :
To avoid initial losses pre-wet the column with about 1 ml of solvent. Boiling chips should be added.
Adjust the vertical position of the apparatus. At the proper rate of distillation, the balls of the column
actively chatter but the chambers do not flood. Adjust the water bath temperature accordingly. When
reaching an extract volume of 1 ml remove the evaporation flask, replace the Snyder column (7.2.4(7.2.4))
by a smaller one and continue the evaporation.
9.5 Addition of recovery standard
The very last step before quantification is the addition of the recovery standards for calculation of the recovery
rates of the internal standards.
Recovery standards shall be added after the final volume reduction. Any further direct volume reduction shall
be avoided. A slow evaporation at room temperature from the open sample vial to a volume of
aboutapproximately 25 µl is acceptable. After addition of the recovery standards, sample extracts should be
stored as briefly as possible before measurement and any further uncontrolled solvent evaporation until
dryness shall be avoided.
Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this document and does not constitute an endorsement by ISO of this product.
2)
Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this document and does not constitute an endorsement by ISO of this product.
10 GC/MS (HRMS or MS/MS) analysis
10.1 General
GC-MS analyses of PCDD/PCDF and dioxin-like PCB shall be carried out on a GC-MS instrument equipped with
a high resolution gas chromatograph, an auto sampler, a mass spectrometer (high resolution or MS/MS) and
a data system for instrument control, data acquisition and processing.
10.2 Gas chromatographic analysis
Gas chromatographic separation shall be carried out in such a way that sufficient separation of all PCDD/PCDF
and dioxin-like PCB congeners is achieved, and the quality criteria specified in 10.410.4 and 10.510.5 are met.
For PCDD/PCDF there is no capillary column available at present that allows the separation of all 2,3,7,8-
substituted congeners from all other non-2,3,7,8-substituted congeners. Complete separation can only be
achieved by analysing a sample on different capillary columns of different polarity.
For dioxin-like PCB analysis similar problems exist for the separation of all coplanar and mono-ortho
congeners. There is no column available at present which is able to completely separate all twelve dioxin-like
PCB congeners from all other non-dioxin-like PCB congeners.
For routine purposes, a state-of-the-art single-GC-column analysis is usually acceptable unless otherwise
demanded.
10.3 Mass spectrometric detection
10.3.1 General
Both a high resolution mass spectrometer or aan MS/MS system on specific MRM can be used for
determination of PCDD/PCDF and dioxin-like PCB.
10.3.2 highHigh resolution mass spectrometry (HRMS)
A high resolution mass spectrometer at a minimum resolution of ≥ 10 000 is used for the detection of
PCDD/PCDF and dioxin-like PCB. This allows the use of C-labelled congeners as internal standards for the
2,3,7,8-PCDD/PCDF congeners and dioxin-like PCB congeners of interest.
The mass spectrometer is used in the MID-Mode mode (multiple ion detection), the GC column is directly
coupled to the mass spectrometer. The ion source temperature should be between 250 °C to 270 °C depending
on type of instrument. To achieve appropriate sensitivity the detection capability should be at least 200 fg for
2,3,7,8-TCDD.
For identification and quantification, the masses given in Table 3Table 3 and Table 4Table 4 shall be recorded
in MID mode. For each PCDD/PCDF or PCB congener of interest at least two ions of the molecular isotope
cluster shall be recorded for both the native and the added C-labelled congeners.
In addition, masses for quality control of the mass calibration shall be measured depending on the type of
instrument, e.g. lock mass, calibration mass, lock mass check.
The time slots for the MID windows shall be defined by a calibration standard in a way that all congeners of
interest elute within the related MID-window. In the case the sum of the concentrations of isomer groups are
needed, the retention time window for all isomers of an isomer group shall be defined by measuring a standard
mixture containing the first and last eluting isomers of each isomer group corresponding to the used GC
column. As an alternative, a fly ash extract or any other solution containing all native PCDD/PCDF congeners
can be used.
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
Table 3 — Masses for the detection and quantification of PCDD/PCDF in [amu]
Substance Dibenzofurans Dibenzo-p-dioxins
12 13 12 13
C C C C
303,901 6 315,941 9 319,896 5 331,936 8
Tetra-CDD/F
305,898 7 317,938 9 321,893 7 333,933 9
339,859 8 351,900 0 355,854 7 367,894 9
Penta-CDD/F
341,856 9 353,897 0 357,851 8 369,891 9
373,820 8 385,861 0 389,815 7 401,855 9
Hexa-CDD/F
375,817 9 387,858 0 391,812 8 403,852 9
407,781 8 419,822 0 423,776 7 435,816 9
Hepta-CDD/F
409,778 9 421,819 0 425,773 8 437,814 0
441,742 8 453,783 0 457,737 7 469,777 9
Octa-CDD/F
443,739 9 455,780 1 459,734 8 471,775 0
Table 4 — Masses for the detection and quantification of PCB in [amu]
Homologue groups 12C 13C
255,961 3 268,001 6
Trichloro-PCB
257,958 4 269,998 6
289,922 3 301,962 6
Tetrachloro-PCB
291,919 4 303,959 7
325,880 4 337,920 7
Pentachloro-PCB
327,877 5 339,917 7
359,841 5 371,881 7
Hexachloro-PCB
361,838 5 373,878 8
393,802 5 405,842 7
Heptachloro-PCB
395,799 5 407,839 8
427,763 5 439,803 8
Octachloro-PCB
429,760 6 441,800 8
461,724 5 473,764 8
Nonachloro-PCB
463,721 6 475,761 8
497,682 6 509,722 9
Decachloro-PCB
499,679 7 511,719 9
10.3.3 Tandem mass spectroscopy (MS/MS)
In addition to the high -resolution mass spectroscopy as described in 10.3.29.3.1, also, a tandem mass
spectroscopic system can be used (MS/MS). The system should support multiple reaction monitoring (MRM)
mode for mass analysis and quantification purposes. Typical MRMs can be seen in Table 5Table 5 below. For
minimum identification criteria see ISO 22892 [21]. In general MRM quan should be used for quantification
and MRM qual in order to secure identification.
Table 5 — Typical MRMs for the detection and quantification of PCDD/PCDF
precursor product CE
(collision
energy)
precursor product
[amu] [amu] CE [V]
quan 289,9 219,9 24
DL PCB 77
qual 291,9 221,9 24
quan 289,9 219,9 24
DL PCB 81
qual 291,9 221,9 24
quan 323,9 253,9 24
DL PCB 105
qual 325,9 255,9 24
quan 323,9 253,9 24
DL PCB 114
qual 325,9 255,9 24
quan 323,9 253,9 24
DL PCB 118
qual 325,9 255,9 24
quan 323,9 253,9 24
DL PCB 123
qual 325,9 255,9 24
quan 323,9 253,9 24
DL PCB 126
qual 325,9 255,9 24
quan 359,8 289,9 24
DL PCB 156
qual 357,8 287,9 24
quan 359,8 289,9 24
DL PCB 157
qual 357,8 287,9 24
quan 359,8 289,9 24
DL PCB 167
qual 357,8 287,9 24
quan 359,8 289,9 24
DL PCB 169
qual 357,8 287,9 24
quan 393,8 323,9 26
DL PCB 189
qual 395,8 325,9 26
quan 457,7 394,8 20
OCDD
qual 459,7 396,8 20
quan 360,8 423,8 20
1,2,3,4,6,7,8-HpCDD
qual 362,8 425,8 20
quan 389,8 326,9 20
1,2,3,4,7,8-HxCDD
qual 387,8 324,9 20
quan 389,8 326,9 20
1,2,3,6,7,8-HxCDD
qual 387,8 324,9 20
1,2,3,7,8,9-HxCDD quan 389,8 326,9 20
© ISO #### 2025 – All rights reserved
ISO/DIS PRF 13914:2025(en)
precursor product CE
(collision
energy)
precursor product
[amu] [amu] CE [V]
qual 387,8 324,9 20
quan 355,9 292,9 20
1,2,3,7,8-PeCDD
qual 357,9 294,9 20
quan 319,9 256,9 20
2,3,7,8-TCDD
qual 321,9 258,9 20
quan 441,8 378,8 26
OCDF
qual 443,8 380,8 26
quan 407,8 344,8 26
1,2,3,4,6,7,8-HpCDF
qual 409,8 346,8 26
quan 407,8 344,8 26
1,2,3,4,7,8,9-HpCDF
qual 409,8 346,8 26
quan 373,8 310,9 26
1,2,3,4,7,8-HxCDF
qual 371,8 308,9 26
quan 373,8 310,9 26
1,2,3,7,8,9-HxCDF
qual 371,8 308,9 26
quan 373,8 310,9 26
1,2,3,6,7,8-HxCDF
qual 371,8 308,9 26
quan 373,8 310,9 26
2,3,4,6,7,8-HxCDF
qual 371,8 308,9 26
quan 339,9 276,9 26
1,2,3,7,8-PeCDF
qual 341,9 278,9 26
quan 339,9 276,9 26
2,3,4,7,8-PeCDF
qual 341,9 278,9 26
quan 303,9 240,9 26
2,3,7,8-TCDF
qual 305,9 242,9 26
...