CEN/TS 16190:2012

SLOVENSKI STANDARD
01-junij-2012
%ODWRREGHODQLELRORãNLRGSDGNLLQWOD'RORþHYDQMHGLRNVLQRYLQIXUDQRYLQ
GLRNVLQRPSRGREQLKSROLNORULUDQLKELIHQLORYVSOLQVNRNURPDWRJUDILMR]PDVQR
VHOHNWLYQLPGHWHNWRUMHPYLVRNHORþOMLYRVWL+5*&06
Sludge, treated biowaste and soil - Determination of dioxins and furans and dioxin-like
polychlorinated biphenyls by gas chromatography with high resolution mass selective
detection (HR GC-MS)
Schlamm, behandelter Bioabfall und Boden - Bestimmung von Dioxinen und Furanen
sowie Dioxin vergleichbaren polychlorierten Biphenylen mittels Gaschromatographie und
hochauflösender massenspektrometrischer Detektion (HR GC-MS)
Boue, biodéchet traité et sol - Détermination des dioxine et furane et biphényls
polychlorés de type dioxine par chromatographie en phase gazeuse-spectrométrie de
masse (HR GC-MS)
Ta slovenski standard je istoveten z: CEN/TS 16190:2012
ICS:
13.030.20 7HNRþLRGSDGNL%ODWR Liquid wastes. Sludge
13.080.10 .HPLMVNH]QDþLOQRVWLWDO Chemical characteristics of
soils
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

TECHNICAL SPECIFICATION
CEN/TS 16190
SPÉCIFICATION TECHNIQUE
TECHNISCHE SPEZIFIKATION
February 2012
ICS 13.030.01
English Version
Sludge, treated biowaste and soil - Determination of dioxins and
furans and dioxin-like polychlorinated biphenyls by gas
chromatography with high resolution mass selective detection
(HR GC-MS)
Boues, biodéchets traités et sols - Détermination des Schlamm, behandelter Bioabfall und Boden - Bestimmung
dioxines et furanes et polychlorobiphényles de type dioxine von Dioxinen und Furanen sowie Dioxin vergleichbaren
par chromatographie en phase gazeuse avec spectrométrie polychlorierten Biphenylen mittels Gaschromatographie und
de masse à haute résolution (CG-SMHR) hochauflösender massenspektrometrischer Detektion (HR
GC-MS)
This Technical Specification (CEN/TS) was approved by CEN on 24 April 2011 for provisional application.

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

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

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

EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

Management Centre: Avenue Marnix 17, B-1000 Brussels
© 2012 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 16190:2012: E
worldwide for CEN national Members.

Contents Page
Foreword .4
Introduction .5
1 Scope .6
2 Normative references .7
3 Abbreviations .7
4 Principle .7
5 Reagents .8
5.1 Chemicals .8
5.2 Standards .8
6 Apparatus and materials .8
6.1 General .8
6.2 Equipment for sample preparation .8
6.3 Soxhlet extractor.8
6.4 Clean-up apparatus .8
6.5 Concentration apparatus .9
6.6 Other equipment .9
7 Sample storage and sample pretreatment . 10
7.1 Sample storage . 10
7.2 Sample pretreatment . 10
8 Extraction and clean-up . 10
8.1 General . 10
8.2 Extraction . 10
8.3 Clean-up . 11
8.3.1 General . 11
8.3.2 Gel permeation chromatography . 12
8.3.3 Multilayer column . 12
8.3.4 Sulfuric acid treatment . 12
8.3.5 Activated carbon column . 12
8.3.6 Aluminium oxide column . 12
8.3.7 Removal of sulfur. 12
8.4 Final concentration of cleaned sample extract . 12
8.5 Addition of recovery standard . 13
9 HRGC/HRMS analysis . 13
9.1 General . 13
9.2 Gas chromatographic analysis . 13
9.3 Mass spectrometric detection . 13
9.4 Minimum requirements for identification of PCDF/PCDD and PCB . 15
9.5 Minimum requirements for quantification of PCDF/PCDD and PCB . 16
9.6 Calibration of the HRGC/HRMS system . 17
9.6.1 General . 17
9.6.2 Calibration for 2,3,7,8-congeners . 17
9.6.3 Calibration for sum of homologue groups . 17
9.7 Quantification of HRGC/HRMS results . 18
9.7.1 Quantification of concentrations of 2,3,7,8-congeners . 18
9.7.2 Quantification of recovery rates of C-labelled standards . 19
9.7.3 Quantification of sum of homologue groups . 19
9.7.4 Calculation of the toxic equivalent . 19
9.7.5 Calculation of the limit of detection and the limit of quantification . 20
10 Precision. 20
11 Test report . 20
Annex A (informative) Toxic equivalent factors. 21
Annex B (informative) Repeatability and reproducibility data . 23
B.1 Materials used in the interlaboratory comparison study . 23
B.2 Interlaboratory comparison results . 24
Annex C (informative) Examples of extraction and clean-up methods . 27
C.1 Example A . 27
C.1.1 General . 27
C.1.2 Chemicals . 27
C.1.3 Procedure . 28
C.2 Example B: Approved clean-up methods . 33
Annex D (informative) Examples of operation of GC/HRMS determination . 35
D.1 Example . 35
D.1.1 General . 35
D.1.2 Gas chromatographic analysis . 35
D.1.3 Mass spectrometric detection . 36
Bibliography . 39

Foreword
This document (CEN/TS 16190:2012) has been prepared by Technical Committee CEN/TC 400 “Project
Committee - Horizontal standards in the fields of sludge, biowaste and soil”, the secretariat of which is held by
DIN.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights.
The preparation of this document by CEN is based on a mandate by the European Commission
(Mandate M/330), which assigned the development of standards on sampling and analytical methods for
hygienic and biological parameters as well as inorganic and organic determinants, aiming to make these
standards applicable to sludge, treated biowaste and soil as far as this is technically feasible.
According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following
countries are bound to announce this Technical Specification: Austria, Belgium, Bulgaria, Croatia, Cyprus,
Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy,
Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia,
Spain, Sweden, Switzerland, Turkey and the United Kingdom.

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 years until the end of the 1990s for the purpose
of different use in open and closed systems, e. g. as electrical insulators or dielectric fluids in capacitors and
transformers, as specialised hydraulic fluids, as a plasticizer in sealing material. Worldwide more than one
million tons of PCBs were produced.
PCDD/F as well as PCBs are emitted during thermal processes as e. g. waste incineration. In 1997 a group of
experts of the World Health Organisation (WHO) fixed toxicity equivalent factors (TEF) for PCDD and twelve
PCBs, known as dioxin-like PCBs (see Annex A). These twelve 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 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 Technical Specification.
This Technical Specification is applicable for several types of matrices and validated for municipal sludge (see
also Annex B for the results of the validation).
WARNING — Persons using this Technical Specification should be familiar with usual laboratory
practice. This Technical Specification does not purport to address all of the safety problems, if any,
associated with its use. It is the responsibility of the user to establish appropriate safety and health
practices and to ensure compliance with any national regulatory conditions.
IMPORTANT — It is absolutely essential that tests conducted according to this Technical
Specification be carried out by suitably trained staff.
1 Scope
This Technical Specification 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 Technical Specification 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 Technical Specification,
limits of detection better than 1 ng/kg (expressed as dry matter) can be achieved.
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 Technical Specification 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.
EN 16179, Sludge, treated biowaste and soil — Guidance for sample pretreatment
3 Abbreviations
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
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 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 Technical Specification 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 PCB.
C -labelled PCDD/F and PCB congeners are added to the sample prior to extraction and HRGC/HRMS
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 HRGC/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 may overload the separation method, disturb the quantification or otherwise 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/F and PCBs is based on quantification by the isotope-
dilution technique using HRGC/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
 C-spiking solution for PCDD/F (internal Standard);
 C-spiking solution for PCB (internal Standard);
 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.
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:
 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.
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 the use with capillary columns, and an oven temperature programme which enables isothermal hold.
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 of twelve 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.

1) Kuderna Danish is an example of a suitable product available commercially. This information is given for the
convenience of users of this Technical Specification and does not constitute an endorsement by CEN of this product.
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 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 cold (< 8 °C) and in the dark. The sample pretreatment should take place within
three days of sampling. Alternatively, samples may 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 EN 16179, 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 Technical Specification 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 C or
any suitable alternative procedures.
The determination of PCDDs/PCDFs is based on quantification by the isotope-dilution technique using
HRGC/HRMS. C -labelled 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 HRGC/HRMS analysis. However, due to possible differences in the binding and
adsorption characteristics between the native PCDDs/PCDFs 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 C.
The main purpose of the clean-up procedure of the raw sample extract is the removal of sample matrix
components, which may 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 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 may 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 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 or
approximately 12 h.
Other solvents or other methods like 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/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
1,2,3,7,8,9- C -HxCDD
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
C -PCB 118
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
2,3,4,6,7,8- C -HxCDF
1,2,3,7,8,9- C -HxCDF
1,2,3,4,6,7,8- C -HpCDF
1,2,3,4,7,8,9- C -HpCDF
C -OCDF
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 PCDDs/Fs and dioxin-like PCBs 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 C.
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 PCDDs/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 silica with different activity grades and surface modifications.
Compounds with different chemical properties than PCDDs/Fs and dioxin-like PCBs can be removed.
8.3.4 Sulfuric acid treatment
A direct treatment of the sample extract with sulfuric acid is possible but is not recommended due to risk of
accident. If applied, this shall be carried out very carefully to avoid losses of PCDDs/Fs and dioxin-like PCBs
on the formed carboniferous surfaces.
8.3.5 Activated carbon column
Column adsorption chromatography using activated carbon may be used to separate planar PCDD/F and
coplanar PCB molecules from mono-ortho PCB and other interfering non-planar molecules.
8.3.6 Aluminium oxide column
Column liquid chromatography on aluminium oxide of different activity grade and acidity/basicity. Interfering
compounds with small differences in polarity or structure compared to PCDDs/Fs and dioxin-like PCBs can be
removed.
Additionally aluminium oxide columns can be used to separate PCDDs/Fs from dioxin-like PCBs.
8.3.7 Removal of sulfur
The removal of sulfur can be achieved by refluxing the extract with powdered copper or by gel permeation
chromatography.
8.4 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 solvent with
a high boiling point.
Though PCDDs/Fs have rather high boiling points (> 320 °C) vapour phase transfer mechanisms and aerosol
formation during solvent evaporation might 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) Rotary evaporators
Losses might 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) Counter gas flow evaporators
Volumes should not be reduced to less than 1 ml.
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.
d) Kuderna Danish
To avoid initial losses pre-wet the column with about 1 ml of solvent. Boiling chips should be added and adjust
the vertical position of the apparatus. At the proper rate of distillation, the balls of the column will actively
chatter but the chambers will not flood. Adjust the water bath temperature accordingly. When reaching an
extract volume of 1 ml remove the evaporation flask, replace the snyder column by a smaller one and
continue the evaporation.
8.5 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 just prior to the quantification procedure. Samples with the recovery
standard added which could not be analysed due to operational reasons (instrument failure), should be stored
as briefly as possible and any further uncontrolled solvent evaporation shall be avoided.
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 about 25 µl is
acceptable.
9 HRGC/HRMS analysis
9.1 General
GC/MS analyses of PCDD/Fs and dioxin-like PCBs shall be carried out on a high resolution GC/MS
instrument equipped with a high resolution gas chromatograph, an autosampler, a high resolution mass
spectrometer and a data system for instrument control, data acquisition and processing.
9.2 Gas chromatographic analysis
Gas chromatographic separation shall be carried out in such a way that sufficient separation of all PCDD/F
and dioxin-like PCB congeners is achieved and the quality criteria specified in 9.4 and 9.5 are met.
For PCDD/F 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 separate all twelve dioxin-like PCB
congeners from all other non-dioxin-like PCB congeners.
9.3 Mass spectrometric detection
A high resolution mass spectrometer at a minimum resolution of 10 000 is used for the detection of PCDD/F
and dioxin-like PCB. This allows the use of C -labelled congeners as internal standards for all 17 PCDD/F
congeners and twelve dioxin-like PCB congeners of interest.
The mass spectrometer is used in the MID-Mode (multiple ion detection), the GC column is directly coupled to
the man 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 3 and Table 4 shall be recorded in MID mode.
For each PCDD/F 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/F congeners can
be used.
Table 3 — Masses for the detection and quantification of PCDD/F

Substance Dibenzofurans Dibenzo-p-dioxins
12 13 12 13
C C C C
Tetra-CDD/F 303,9016 315,9419 319,8965 331,9368
305,8987 317,9389 321,8937 333,9339
Penta-CDD/F 339,8598 351,9000 355,8547 367,8949
341,8569 353,8970 357,8518 369,8919
Hexa-CDD/F 373,8208 385,8610 389,8157 401,8559
375,8179 387,8580 391,8128 403,8529
Hepta-CDD/F 407,7818 419,8220 423,7767 435,8169
409,7789 421,8190 425,7738 437,8140
Octa-CDD/F 441,7428 453,7830 457,7377 469,7779
443,7399 455,7801 459,7348 471,7750

Table 4 — Masses for the detection and quantification of PCB
12 13
Homologue groups
C C
Trichloro-PCB 255,9613 268,0016
257,9584 269,9986
Tetrachloro-PCB 289,9223 301,9626
291,9194 303,9597
Pentachloro-PCB 325,8804 337,9207
327,8775 339,9177
Hexachloro-PCB 359,8415 371,8817
361,8385 373,8788
Heptachloro-PCB 393,8025 405,8427
395,7995 407,8398
Octachloro-PCB 427,7635 439,8038
429,7606 441,8008
Nonachloro-PCB 461,7245 473,7648
463,7216 475,7618
Decachloro-PCB 497,6826 509,7229
499,6797 511,7199
9.4 Minimum requirements for identification of PCDF/PCDD and PCB
9.4.1 The isotope ratio between the two ions of the molecular isotope cluster which are recorded shall
match the theoretical value within ± 15 % (see Table 5).
Table 5 — Limits of isotope ratios
Substance
Isotope ratio Isotope ratio Isotope ratio
lower limit theoretical value upper limit
TCDD/F 0,65 0,77 (M/M+2) 0,88
PeCDD/F 0,55 0,64 (M+4/M+2) 0,75
HxCDD/F 0,69 0,81 (M+4/M+2) 0,94
HpCDD/F 0,83 0,96 (M+4/M+2) 1,13
OCDD/F 0,74 0,89 (M+2/M+4) 1,009

9.4.2 The retention time of a native 2,3,7,8-chlorine substituted isomer (Cl - to Cl -congeners) shall be
4 6
within a time window of +3 s to –3 s based on the retention time of the corresponding C -labelled isomer in
the sample. For the identification of low concentrations (S/N < 10) a time window of ± 10 s is acceptable.
Alternatively, relative retention times based on the recovery standard (e. g. C -1,2,3,4-TCDF) can be
calculated. The difference shall not be more than 0,3 % compared with the calibration standard.
9.4.3 The signal-to-noise ratio of the raw data shall be at least 3:1 for three consecutive scans for the signal
used for identification. The base line noise shall be measured in front of the signal of the native congener
within a signal-free window corresponding to ten times the signal width at half height. Peak-to-peak values are
taken.
9.5 Minimum requirements for quantification of PCDF/PCDD and PCB
9.5.1 For PCDD/F analysis there is no chromatographic column available at present that is able to separate
all 2,3,7,8-chlorine substituted congeners from all other, non-2,3,7,8-chlorine substituted congeners. Complete
separation can only be achieved by multi-analysis of the sample on different columns of different nature
(polarity).
Single column data may therefore be reported by this method, however in cases where a regulatory limit is
exceeded or congener specific data are needed, a confirmatory analysis should be performed on a second
column.
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 separate all twelve dioxin-like PCB
congeners from all other non-dioxin-like PCB congeners. The use of one relatively non-polar column (e. g.
DB-5) is the common technique. The separation of congener PCB-123 is the crucial point of the gas
chromatographic separation. But due to the minor contribution to the overall TEQ this leads to an inessential
increase of the uncertainty of the method.
9.5.2 The peak shape of the gas chromatographic signal of a congener shall contain ten or more sampling
points (scanning units).
9.5.3 2,3,7,8-TCDD shall be separated from all other interfering isomers within a 25 % valley below the top
of the minor peak with respect to the height of that peak.
9.5.4 The recovery rate of each individual 2,3,7,8-chlorine substituted PCDD/PCDF of the internal
standards in each sample shall be within:
 50 % to 130 % for the tetra- to hexachlorinated congeners;
 40 % to 130 % for the hepta- and oct
...