ISO 13913:2014

INTERNATIONAL ISO
STANDARD 13913
First edition
2014-02-01
Soil quality — Determination of
selected phthalates using capillary
gas chromatography with mass
spectrometric detection (GC/MS)
Qualité du sol — Détermination de phthalates sélectionnés en
utilisant la chromatographie gazeuse capillaire avec détection par
spectrométrie de masse
Reference number
ISO 13913:2014(E)
©
ISO 2014

---------------------- Page: 1 ----------------------
ISO 13913:2014(E)

COPYRIGHT PROTECTED DOCUMENT
© ISO 2014
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized otherwise in any form
or by any means, electronic or mechanical, including photocopying, or posting on the internet or an intranet, without prior
written permission. Permission can be requested from either ISO at the address below or ISO’s member body in the country of
the requester.
ISO copyright office
Case postale 56 • CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2014 – All rights reserved

---------------------- Page: 2 ----------------------
ISO 13913:2014(E)

Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Principle . 2
4 Interferences . 2
4.1 General . 2
4.2 Interferences during sampling . 2
4.3 Cross contamination . 2
4.4 Interferences in gas chromatography . 2
5 Reagents . 2
5.1 General . 2
6 Apparatus . 4
6.1 General . 4
7 Sampling and sample storage . 6
8 Procedure. 6
8.1 Preparation of glass apparatus . 6
8.2 Drying of the sample . 6
8.3 Extraction . 7
8.4 Clean-up . 7
8.5 Gas chromatography . 8
8.6 Blank monitoring . 8
8.7 Identification of individual compounds . 8
9 Calibration . 9
9.1 General . 9
9.2 Calibration with internal standard . 9
9.3 Verification of calibration .10
10 Calculation .11
11 Expression of results .12
12 Precision .12
13 Test report .12
Annex A (informative) Repeatability and reproducibility data .13
Annex B (informative) Example of a reference solution for multipoint calibration .16
Annex C (informative) Examples for gas chromatographic conditions .18
Annex D (informative) Flow scheme of the determination of phthalates .20
Bibliography .21
© ISO 2014 – All rights reserved iii

---------------------- Page: 3 ----------------------
ISO 13913:2014(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.
iv © ISO 2014 – All rights reserved

---------------------- Page: 4 ----------------------
ISO 13913:2014(E)

Introduction
This International Standard is applicable and validated for several types of matrices as indicated in
Table 1 (see also Annex A for the results of the validation).
Table 1 — Matrices for which this International Standard is applicable and validated
Matrix Materials used for validation
Sludge Municipal sludge
Biowaste Fresh compost
Soil Sludge amended soil
© ISO 2014 – All rights reserved v

---------------------- Page: 5 ----------------------
INTERNATIONAL STANDARD ISO 13913:2014(E)
Soil quality — Determination of selected phthalates using
capillary gas chromatography with mass spectrometric
detection (GC/MS)
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 the determination of selected phthalates in sludge,
treated biowaste, and soil, after extraction and gas chromatographic analysis with mass spectrometric
detection.
The method is applicable for the determination of phthalates (see Table 2) at the lowest mass content of
0,1 mg/kg to 0,5 mg/kg (expressed as dry matter), depending on the individual substance.
The applicability of the method to other phthalates not specified in Table 2 is not excluded except the
isomeric mixtures, e.g. DiNP (Di-isononylphthalate), but shall be verified in each case.
Table 2 — Phthalates that can be determined according to this International Standard
Molar mass
a
No Name Formula Abbreviation CAS-RN
g/mol
1 Dimethylphthalate C H O DMP 194,2 00131-11-3
10 10 4
2 Diethylphthalate C H O DEP 222,2 00084-66-2
12 14 4
3 Dipropylphthalate C H O DPP 250,3 00131-16-8
14 18 4
4 Di-(2-methyl-propyl)phthalate C H O DiBP 278,4 00084-69-5
16 22 4
5 Dibutylphthalate C H O DBP 278,4 00084-74-2
16 22 4
6 Butylbenzylphthalate C H O BBzP 312,4 00085-68-7
19 20 4
7 Dicyclohexylphthalate C H O DCHP 330,4 00084-61-7
20 26 4
8 Di-(2-ethylhexyl)phthalate C H O DEHP 390,6 00117-81-7
24 38 4
9 Dioctylphthalate C H O DOP 390,6 00117-84-0
24 38 4
10 Didecylphthalate C H O DDcP 446,7 00084-77-5
28 46 4
11 Diundecylphthalate C H O DUP 474,4 03648-20-2
30 50 4
a
Chemical Abstracts Service Registry Number.
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 5667-13, Water quality — Sampling — Part 13: Guidance on sampling of sludges
© ISO 2014 – All rights reserved 1

---------------------- Page: 6 ----------------------
ISO 13913:2014(E)

ISO 5667-15, Water quality — Sampling — Part 15: Guidance on the preservation and handling of sludge
and sediment samples
ISO 10381-2, Soil quality — Sampling — Part 2: Guidance on sampling techniques
ISO 11465, Soil quality — Determination of dry matter and water content on a mass basis — Gravimetric
method
ISO 14507, Soil quality — Pretreatment of samples for determination of organic contaminants
ISO 22892, Soil quality — Guidelines for the identification of target compounds by gas chromatography and
mass spectrometry
3 Principle
The dried sample, dried by freeze drying or with sodium sulfate, is extracted with ethyl acetate on the
shaking device. An aliquot of the extract is cleaned with aluminium oxide (if necessary) followed by
gas chromatographic separation using capillary columns and identification and quantification of the
phthalates by mass spectrometry.
4 Interferences
4.1 General
Due to their use as plasticizer agents, phthalates are ubiquitous. The sources of phthalates are multiple
and shall be checked and reduced by every laboratory, itself. Therefore, special attention shall be paid
to avoid contaminations.
4.2 Interferences during sampling
In order to avoid interferences and cross contaminations, do not use plastic materials (e.g. pipes, etc.).
4.3 Cross contamination
Chemicals and analytical equipment can be of various qualities. Cross contamination is likely to occur
with laboratory air. Therefore, remove, as far as possible, plastic materials from the laboratory. Cleaning
agents often contain phthalates and can severely contaminate the laboratory air if in use regularly.
Therefore, refrain from using these agents during application of this procedure.
Using plastic gloves during pretreatment can increase the contamination.
4.4 Interferences in gas chromatography
Phthalates can bleed from the septa of the injector into the gas chromatograph; therefore, use septa that
are not likely to contaminate the system.
Fittings, e.g. of syringes, or equipment and septa of the sampling bottles (6.5) can also contain phthalates.
5 Reagents
5.1 General
All reagents shall be of recognized analytical grade.
Use only reagents with negligibly low concentration of phthalates and verify by blank determinations
and, if necessary, apply additional cleaning steps.
2 © ISO 2014 – All rights reserved

---------------------- Page: 7 ----------------------
ISO 13913:2014(E)

5.2 Nitrogen, N , of high purity, at least a volume fraction of 99,9 % for drying and, if necessary, for
2
concentration by evaporation.
5.3 Helium, He, of high purity, at least a volume fraction of 99,999 %.
5.4 Ethyl acetate, C H O , phthalate-free, high purity.
4 8 2
5.5 Methanol, CH OH.
3
5.6 Isooctane, C H (2,2,4-trimethylpentane).
8 18
5.7 Quartz wool, heated to 400 °C for at least 4 h.
5.8 Aluminium oxide, Al O , neutral, 50 µm to 200 µm particle size, heated to 400 °C for at least 4 h.
2 3
Store heat-treated aluminium oxide in covered flask or desiccator. Use within five days after heat-
treatment.
1)
®
NOTE Alternative materials, like Florisil or silica can be used, provided their properties and capacity to
separate are similar to aluminium oxide and their properties are checked according to 8.6.
5.9 Internal standards.
For example:
— deuterated di-n-butylphthalate, “D4-ring-DBP”;
— deuterated D4-C H O ;
16 22 4
— deuterated di-(2-ethylhexyl) phthalate “D4-ring-DEHP”;
— deuterated D4-C H O ; di-n-octylphthalate, “D4-ring-DOP”;
24 38 4
— D4- C H O ;
24 38 4
13
— C-labelled standards can also be used, if available.
5.10 Reference substances.
Table 2 gives a list of phthalates with defined mass concentrations for the preparation of reference
solutions for the gas chromatographic procedure.
5.11 Solutions of the single substances.
In a 10-ml volumetric flask (6.13), transfer e.g. 10 mg of each of the reference substances (5.10) in ethyl
acetate (5.4) and bring to volume with ethyl acetate (5.4) (concentration: 1 g/l).
Store the solutions in glass bottles at –18 °C, protected from light, and check the concentration at least
every three months.
5.12 Stock solution.
In a 10-ml volumetric flask (6.13), dissolve between 100 µl and 500 µl of the single substance solutions
(5.11) and bring to volume with ethyl acetate (5.4) (concentration: 10 mg/l to 50 mg/l).
1) This information is given for the convenience of users of this document and does not constitute an endorsement
by ISO TC 190/SC 3 of the product named. Equivalent products may be used if they can be shown to lead to the same
results.
© ISO 2014 – All rights reserved 3

---------------------- Page: 8 ----------------------
ISO 13913:2014(E)

Store the solution in a glass bottle at –18 °C, protected from light, and check the concentration at least
every three months.
5.13 Reference solutions for multipoint calibration (see Annex B).
Prepare solutions by adequate dilution of the stock solution (5.12) and internal standards (5.9) with
ethyl acetate (5.4).
Store the solutions in glass bottles at –18 °C, protected from light, and check the concentration at least
every three weeks.
5.14 Solution of the internal standards (see Annex B).
5.14.1 Internal standard solution of D4-phthalates.
Weigh e.g. 0,1 g of an internal standard phthalate (D4) (5.9) in a 10-ml volumetric flask (6.13) filled with
about 5 ml of ethyl acetate (5.4) and bring to volume with ethyl acetate (5.4). Store the solution in a glass
bottle at −18 °C.
5.14.2 Solution I internal standard mix.
Combine the solutions of the single internal standard phthalates (5.9) e.g. by dilution 1:100 as follows:
Transfer with a syringe 0,1 ml (6.15) of each solution into a 10-ml volumetric flask (6.13) filled with
about 5 ml of ethyl acetate (5.4). Bring to volume with ethyl acetate. The final concentration of di-n-
octylphthalate (D4), di-n-butylphthalate (D4), and di-(2-ethylhexylphthalate) is 100 mg/l in ethyl
acetate (5.4).
5.14.3 Solution II internal standard mix.
Take from this 1:100 dilution (5.14.2) e.g. 250 µl, transfer into a 250-ml volumetric flask (6.13), filled
with 250 ml of ethyl acetate (5.4).
The final concentration of di-n-octylphthalate (D4), di-n-butylphthalate (D4), and di-(2-
ethylhexylphthalate) is 0,1 mg/l in ethyl acetate (5.4).
5.14.4 Solution III internal standard mix.
Dilute the solution I internal standard (5.14.2.) 1:10. Pipette 1 ml of the solution (5.14.2.) in a 10-ml
volumetric flask (6.13) filled with about 5 ml of ethyl acetate (5.4). Bring to volume with ethyl acetate. The
final concentration of di-n-octylphthalate (D4), di-n-butylphthalate (D4), and di-(2-ethylhexylphthalate)
is 10 mg/l in ethyl acetate.
5.15 Sodium sulfate, Na SO , heated to 400 °C for at least 4 h.
2 4
6 Apparatus
6.1 General
Equipment or parts of it which are likely to come into contact with the sample or its extract shall be free
from phthalates. This can be achieved by thorough cleaning of all glass apparatus and checked by the
blank determination.
6.2 Wide-neck flat bottomed flasks with glass stoppers, preferably brown glass, volumes 500 ml
and 1 000 ml.
6.3 Drying oven, capable of maintaining at a temperature of (105 ± 5) °C.
4 © ISO 2014 – All rights reserved

---------------------- Page: 9 ----------------------
ISO 13913:2014(E)

6.4 Muffle furnace, adjustable, up to temperatures of (400 ± 10) °C, with capacity of e.g. at least 60 l.
6.5 Sampling vial, glass, with inert stopper, e.g. septum, lined with polytetrafluoroethylene (PTFE) for
storage of the extracts, and sampling bottles, glass, with inert septum, 2 ml, for storage of the extracts for
auto sampler operation.
6.6 Vacuum device for clean-up, vacubox, extraction box.
6.7 Stainless steel cock, with stainless steel cone or polytetrafluoroethylene (PTFE) cock with Luer
connection for separate vacuum connection.
6.8 Glass cartridges, with Luer cone.
6.9 Polytetrafluoroethylene (PTFE) frits, for cartridges, 6 ml.
6.10 Aluminium foil, heated to 400 °C.
6.11 Stainless steel reservoir, for storage of smaller glass apparatus.
6.12 Measuring cylinders, volumes 50 ml and 100 ml.
6.13 Volumetric flasks, volumes 10 ml, 25 ml, and 250 ml.
6.14 Pasteur pipettes, e.g. 2 ml.
6.15 Syringes, 2 µl, 5 µl, 10 µl, 50 µl, 100 µl, and 500 µl, maximum permitted error ±2 %.
6.16 Gas chromatograph, with capillary column, temperature controlled, with mass spectrometric
detection.
6.17 Operating gases, for gas chromatography/mass spectrometer of high purity and in accordance
with manufacturer’s specifications.
6.18 Fused silica columns, with non-polar stationary phase (see Annex C for examples).
Check the quality of the column e.g. by injecting the reference solution (5.13) and ensure that the
separation is satisfactory.
6.19 Glass tubes, graduated 5 ml or 10 ml.
6.20 Nitrogen device, for drying the glass cartridges (6.8).
6.21 Beaker, volumes 50 ml and 100 ml.
6.22 Erlenmeyer flask, volume 250 ml.
6.23 Shaking device, horizontal shaking movement.
6.24 Freeze drying apparatus.
6.25 Metal spoon.
© ISO 2014 – All rights reserved 5

---------------------- Page: 10 ----------------------
ISO 13913:2014(E)

6.26 Agate mortar.
6.27 Metallic clamp, for stopper.
6.28 Balance, e.g. range 0,001 g to 100 g.
6.29 Pipette, volumes 20 ml, 25 ml, and 50 ml.
7 Sampling and sample storage
Collect, preserve, and handle samples in accordance with ISO 5667-13, ISO 5667-15, and ISO 10381-2.
Use for sampling pretreated sampling bottles (6.2) and make sure that the stoppers are pretreated as
well.
In general, sampling should be carried out using stainless steel containers or glass vessels. In order to
avoid contaminations, do not use any plastics material (tubes and other).
Dry the sample as soon as possible after sample collection. If storage is unavoidable, store the samples
in the dark at 4 °C. Dried samples are found to be stable for a longer period.
Determine the dry matter content in accordance with ISO 11465.
8 Procedure
8.1 Preparation of glass apparatus
Clean all glass apparatus, except the syringes, used during analysis in the dishwasher with water and
subsequently dry in the oven (6.3) at 105 °C.
Heat the pre-rinsed glass apparatus in the muffle furnace (6.4) for at least 4 h at 400 °C.
Subsequently let the apparatus cool to room temperature within 12 h.
NOTE Glassware for volumetric purposes can change its properties due to the heating process.
Close the cooled glass apparatus (bigger vessels) with the respective stoppers or with aluminium foil
(6.10). Store smaller glass apparatus in decontaminated (heated) and appropriately closed stainless
steel containers (6.11).
In order to avoid losses by adsorption on the walls, rinse the walls with isooctane (5.6) by using Pasteur
pipettes (6.14). Discard the solvent.
Let residual solvent evaporate under a fume hood.
Carry out this deactivation of the surface after heating and cooling or immediately prior to use.
8.2 Drying of the sample
8.2.1 General
Pretreat the samples according to ISO 14507, if not otherwise specified.
Depending on the water content and the matrix, dry the sample either with sodium sulfate (5.15) or by
freeze drying (see also Annex D).
Samples (i. e. soil, waste) with a dry matter > 80 % can be dried with sodium sulfate (see 8.2.3).
6 © ISO 2014 – All rights reserved

---------------------- Page: 11 ----------------------
ISO 13913:2014(E)

Sludge with high water content shall be dried by freeze drying (see 8.2.2).
8.2.2 Freeze drying
Freeze a part of the homogenized sample or a representative part of the sample at –18 °C. Afterwards,
lyophilize at about 5 kPa until the constant mass is achieved.
Homogenize the freeze-dried sample with the aid of an agate mortar (6.26).
8.2.3 Drying with sodium sulfate (Na SO )
2 4
Depending on the expected phthalate content of the sample, weigh between 1 g to 10 g of the wet sample
into a beaker (6.21) and give as much sodium sulfate (5.15) into the wet sample until a trickle mixture
is achieved. Mortar the mixture in an agate mortar (6.26). When a free-flowing mixture is obtained, the
humidity is bound. In parallel, determine the dry matter content of the sample (see Clause 7).
8.3 Extraction
Transfer between 1 g to 10 g, referred to the dry matter content and the expected phthalate concentration,
of the mixture of the sample and sodium sulfate (prepared according to 8.2.3) or the freeze-dried
sample (prepared according to 8.2.2) into a 250-ml Erlenmeyer flask (6.22) and give e.g. 20 ml ethyl
acetate (5.4) with internal standard (5.9) to the sample. If a high amount of phthalates is expected, the
extraction solvent can be doubled or multiplied (see Table 3). Since the concentration of phthalate in a
blank is unknown, dilutions of the sample extract shall be avoided. Close the Erlenmeyer flask (6.22)
with a stopper and fix the stopper with a metallic clamp (6.27).
Extract the sample for at least 30 min on the shaking device (6.23). Make sure that a good thorough
mixing of the sample and the solvent is obtained. After the extraction, take approximately 1 ml with a
pipette (6.14) and transfer the extract into a GC vial (6.5), or approximately 3 ml are required if a clean-
up (see 8.4) is necessary. Place the heated aluminium foil (6.10) between vial and caps in order to avoid
a contamination by phthalates from the septa. The extract can be analysed by GC-MS directly.
If a clean-up is necessary, e.g. due to interferences of the target analyte in the GC-MS chromatography,
see 8.4.
Table 3 — Examples of sample intake and ratio dry matter/solvent volume
Sample intake Ratio dry
Matrix Remark
g matter:solvent
Sludge (sewage) 1 to 10 < 1:80 A high amount of DEHP is expected
Sediment / suspended solid 2 to 10 < 2:20 DEHP is expected
Compost 2 to 10 < 2:20 Low to high concentration of DEHP
Soil 2 to 10 < 2:20 Low to high concentration of DEHP
Take care that the amount of solvent is sufficient for collecting the extract (at least 3 ml).
The described method of extraction (shaking) is recommended due to the small contamination potential.
Using Soxhlet extraction or accelerated solvent extraction (ASE), comparable amount of phthalates can
be achieved, but the contamination risk is higher. Moreover, the extraction relation (solvent and sample
intake) should be adjusted to the respective extraction method. For blank criteria, see 8.6.
8.4 Clean-up
A clean-up is only necessary if interferences in the GC-MS chromatogram, originating from matrices,
are expected; otherwise, it should be avoided due to the additional risk of contamination. After the
extraction, take approximately 3 ml of the extract with a pipette (6.14) and clean the extract with
aluminium oxide (5.8).
© ISO 2014 – All rights reserved 7

---------------------- Page: 12 ----------------------
ISO 13913:2014(E)

Clean the extracts as follows.
— Place 1 g of activated aluminium oxide (5.8) in the cartridges (6.8) between two PTFE frits (6.9).
— Clean the aluminium oxide (5.8) with one cartridge volume of ethyl acetate (5.4).
— Dry with nitrogen (5.2) for 1 min.
— Fix the cleaned cartridge with stainless steel cock (6.7) and place it on the vacuum device (6.6).
— Let the extract run through the cartridge and collect it in a glass tube (6.19).
— Transfer the extract to GC vials (6.5). Attach heated aluminium foil (6.10) between vial and cap in
order to avoid a contamination by phthalates from the septa.
8.5 Gas chromatography
Optimize the GC apparatus (6.16) according to the instrument manufacturer’s manual.
Use capillary columns (see Annex C and 6.18) for separation.
In order to clean the inlet system free from phthalates, inject ethyl acetate (5.4) at least five times from
various GC vials (see Clause 4) before measuring the sample extracts or calibration solutions.
8.6 Blank monitoring
Check the proper conditions of instruments and reagents by blank measurements at regular intervals.
For the blank measurements, treat sodium sulfate (5.15) in the same way as the sample (see 8.2 and 8.3).
Weigh nearly as much sodium sulfate as is needed to dry the samples. DEP, DiBP, DBP, and DEHP are the
most
...