SIST ISO 17734-2:2015
Determination of organonitrogen compounds in air using liquid chromatography and mass spectrometry - Part 2: Amines and aminoisocyanates using dibutylamine and ethyl chloroformate derivatives
Determination of organonitrogen compounds in air using liquid chromatography and mass spectrometry - Part 2: Amines and aminoisocyanates using dibutylamine and ethyl chloroformate derivatives
This part of ISO 17734 gives general guidance for the sampling and analysis of airborne amines and
aminoisocyanates in workplace air. It is strongly recommended that the determination of amines and
aminoisocyanates is made together with the determination of isocyanates in air, using DBA as a reagent
(see ISO 17734-1).
The method can be used for simultaneous determinations of amines, such as 4,4’-methylenediphenyldiamine
(4,4’-MDA), 2,4- and 2,6-toluenediamine (2,4- and 2,6-TDA), and 1,6-hexamethylenediamine (1,6-
HDA), and compounds containing both isocyanate and amine groups, such as 4,4’-methylenediphenyl
aminoisocyanate (4,4’-MAI), 2,4-, 4,2-, and 2,6-toluene aminoisocyanate (2,4-, 4,2-, and 2,6-TAI), and
1,6-hexamethylene aminoisocyanate (1,6-HAI). The method is suitable for collecting amines and
aminoisocyanates in both the gas and particle phases. The instrumental detection limit for the amines
is about 5 nmol/sample and for the aminoisocyanate, it is about 0,3 nmol/sample. For a 15 l air sample,
this corresponds to 0,4 ng⋅m–3 for TDA and 0,03 ng⋅m–3 for TAI.
Détermination des composés organiques azotés dans l'air par chromatographie liquide et spectrométrie de masse - Partie 2: Amines et aminoisocyanates par les dérivés de la dibutylamine et du chloroformate d'éthyle
Določevanje organskih dušikovih spojin v zraku s tekočinsko kromatografijo in masno spektrometrijo - 2. del: Določevanje aminov in aminoizocianatov preko derivatov dibutilamina in etilkloroformiata
Ta del standarda ISO 17734 podaja splošne napotke za vzorčenje in analizo aminov ter aminoizocianatov v zraku na delovnem mestu. Zelo se priporoča, da se določevanje aminov in aminoizocianatov izvaja skupaj z določevanjem izocianatov v zraku, pri čemer se za reagent uporablja DBA
(glej ISO 17734-1).
Metodo je mogoče uporabiti za hkratno določevanje aminov, kot so 4,4'-metilendifenildiamin
(4,4'-MDA), 2,4- in 2,6-toluendiamin (2,4-, 2,6-TDA) in 1,6-heksametilendiamin (1,6-HDA), ter spojin, ki vsebujejo skupine izocianatov in aminov, kot so 4,4'-metilendifenil aminoizocianat (4,4'-MAI), 2,4-, 4,2- in 2,6-toluen aminoizocianat (2,4, 4,2, 2,6-TAI), 1,6-heksametilen aminoizocianat (1,6-HAI). Metoda je ustrezna za zbiranje aminov in aminoizocianatov v plinski fazi in fazi delcev. Meja detekcije instrumentov za amine je približno 5 nmol/vzorec, za aminoizocianate pa 0,3 nmol/vzorec. For a 15 l air sample, this corresponds to 0,4 ng⋅m–3 for TDA and 0,03 ng⋅m–3 for TAI.
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INTERNATIONAL ISO
STANDARD 17734-2
Second edition
2013-12-01
Determination of organonitrogen
compounds in air using liquid
chromatography and mass
spectrometry —
Part 2:
Amines and aminoisocyanates
using dibutylamine and ethyl
chloroformate derivatives
Détermination des composés organiques azotés dans l’air par
chromatographie liquide et spectrométrie de masse —
Partie 2: Amines et aminoisocyanates par les dérivés de la
dibutylamine et du chloroformate d’éthyle
Reference number
ISO 17734-2:2013(E)
©
ISO 2013
---------------------- Page: 1 ----------------------
ISO 17734-2:2013(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2013
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 2013 – All rights reserved
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ISO 17734-2:2013(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Principle . 1
4 Reagents and materials . 2
5 Standard solutions. 3
5.1 Reference compounds . 3
5.2 Amine and deuterium-labelled amine derivatives . 3
5.3 Aminoisocyanate derivatives . 4
5.4 Thermal decomposition products of polyurethane (PUR). 4
5.5 Stability of the amine and aminoisocyanate derivatives . 4
6 Apparatus . 5
7 Air sampling . 7
7.1 Pre-sampling laboratory preparation . 7
7.2 Pre-sampling field preparations . 7
7.3 Collection of air samples . 7
7.4 Blanks . . 8
7.5 Raw material . 8
7.6 Shipment of samples . 8
8 Laboratory sample preparation . 9
8.1 Sample sequence . 9
8.2 Work-up procedure .10
9 Instrumental settings .10
9.1 HPLC program (LC-MS) .10
9.2 HPLC program [LC-chemiluminescent nitrogen detector (LC-CLND)] .10
9.3 Mass spectrometer .10
10 Data handling .11
10.1 Identification .11
10.2 Calibration curves .11
10.3 Quantification .11
11 Determination of performance characteristics.11
11.1 General .11
11.2 Relevant uncertainty contributions and criteria .12
11.3 Assessment of performance characteristics (following the detailed approach in Reference
[12]) .12
Annex A (informative) Performance characteristics
.20
Annex B (informative) Examples .22
Bibliography .27
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ISO 17734-2:2013(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www.iso.org/directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www.iso.org/patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation on the meaning of ISO specific terms and expressions related to conformity
assessment, as well as information about ISO’s adherence to the WTO principles in the Technical Barriers
to Trade (TBT) see the following URL: Foreword - Supplementary information
The committee responsible for this document is ISO/TC 146, Air quality, Subcommittee SC 2,
Workplace atmospheres.
This second edition of ISO 17734-2 cancels and replaces ISO 17734-2:2006, which has been
technically revised.
ISO 17734 consists of the following parts, under the general title Determination of organonitrogen
compounds in air using liquid chromatography and mass spectrometry:
— Part 1: Isocyanates using dibutylamine derivatives
— Part 2: Amines and aminoisocyanates using dibutylamine and ethyl chloroformate derivatives
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ISO 17734-2:2013(E)
Introduction
In many applications, when considering isocyanates as a workplace contaminant, there is also a need to
investigate the presence of aminoisocyanates and amines. During thermal decomposition of polyurethane
[1][2][3][4][5][6]
(PUR), not only isocyanates, but also amines and aminoisocyanates, are formed.
The determination of isocyanates in the work environment using DBA as a reagent has been
demonstrated to be a robust method (see ISO 17734-1). Using the DBA method and derivatization with
ethyl chloroformate in the following work-up procedure makes simultaneous determination of amines,
[6][7]
aminoisocyanates, and isocyanates possible.
For quantification of amine and aminoisocyanate derivatives, reference compounds are necessary,
but are only available for a few diamines. Aminoisocyanates cannot be analysed directly because they
react with themselves. In this method, a nitrogen-specific detector has been used for quantification of
amine and aminoisocyanate derivatives in reference solutions. This technique has been demonstrated
to be a useful tool, together with MS characterization, in greatly facilitating the production of reference
[6]
solutions.
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INTERNATIONAL STANDARD ISO 17734-2:2013(E)
Determination of organonitrogen compounds in air using
liquid chromatography and mass spectrometry —
Part 2:
Amines and aminoisocyanates using dibutylamine and
ethyl chloroformate derivatives
1 Scope
This part of ISO 17734 gives general guidance for the sampling and analysis of airborne amines and
aminoisocyanates in workplace air. It is strongly recommended that the determination of amines and
aminoisocyanates is made together with the determination of isocyanates in air, using DBA as a reagent
(see ISO 17734-1).
The method can be used for simultaneous determinations of amines, such as 4,4’-methylenediphenyldiamine
(4,4’-MDA), 2,4- and 2,6-toluenediamine (2,4- and 2,6-TDA), and 1,6-hexamethylenediamine (1,6-
HDA), and compounds containing both isocyanate and amine groups, such as 4,4’-methylenediphenyl
aminoisocyanate (4,4’-MAI), 2,4-, 4,2-, and 2,6-toluene aminoisocyanate (2,4-, 4,2-, and 2,6-TAI), and
1,6-hexamethylene aminoisocyanate (1,6-HAI). The method is suitable for collecting amines and
aminoisocyanates in both the gas and particle phases. The instrumental detection limit for the amines
is about 5 nmol/sample and for the aminoisocyanate, it is about 0,3 nmol/sample. For a 15 l air sample,
–3 –3
this corresponds to 0,4 ng⋅m for TDA and 0,03 ng⋅m for TAI.
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 16200-1, Workplace air quality — Sampling and analysis of volatile organic compounds by solvent
desorption/gas chromatography — Part 1: Pumped sampling method
ISO 5725-2, Accuracy (trueness and precision) of measurement methods and results — Part 2: Basic method
for the determination of repeatability and reproducibility of a standard measurement method
3 Principle
The method permits the simultaneous sampling and analysis of amines, aminoisocyanates, and
isocyanates. Only amines and aminoisocyanates are discussed in this part of ISO 17734, because
isocyanates are considered in ISO 17734-1.
Samples are collected by drawing a known volume of air through a midget impinger flask followed by a
–1
filter. The impinger contains 10 ml of 0,01 mol⋅l of di-n-butylamine (DBA) in toluene, and the filter is a
glass fibre filter with no binder. After sampling, deuterium-labelled amine-ethyl chloroformate (ET) and
isocyanate-DBA derivatives (used as internal standard) are added to the sample solutions. The excess
reagent and solvent are evaporated, and the samples are dissolved in acetonitrile. The samples are
analysed using reversed-phase liquid chromatography (LC) and electrospray (ESP) mass spectrometric
(MS) detection, monitoring positive ions. Quantification is made by quantifying selected ions.
Quantification and qualitative determinations can be performed using different LC-MS techniques. LC-
CLND (chemiluminescent nitrogen detection) or, for aromatic isocyanates, aminoisocyanates, and amines,
LC-UV (ultraviolet detection) can be used for the determination of higher concentrations. Reference
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ISO 17734-2:2013(E)
materials can be characterized using LC-MS/CLND. For characterization of volatile compounds, a GC-
thermoionic specific detector (TSD) can also be used.
Gas phase and large particles (>1,5
Sampling
µ m ) are collected in the impinger
with impinger and filter flask. Small particles (0,01 µm
to 1,5 µm) are collected . on the filter
Standard solution:
Calibration curve Transfer of sample solution Isocyanate-DBA, amine-ET and
aminoisocyanate-DBA-ET
from standard solution and filter to test tube
derivatives
Internal standard, IS:
- Similar in retention time
- Similar in molecular structure
Addition of IS
i.e. aromatic or aliphatic
- Preferably deuterium-labelled
derivative
-Extraction of analytes on the filter
-Derivatisation of the amine groups
with ET in a two phase
Work-up procedure
derivatisation procedure
- Change of solution media
- Enrichment of the analytes
+
- SIM of molecular ion [ MH ]
Analysis with LC-MS
of analytes and IS
- Calculation of the area ratio of the
analyte and IS
Evaluation of data
- Comparison between area ratio of
the air sample and calibration plot
Figure 1 — Principle of the described method
4 Reagents and materials
4.1 DBA reagent.
Analytical grade di-n-butylamine is commercially available.
4.2 Ethyl chloroformate reagent.
Analytical grade ethyl chloroformate is commercially available.
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ISO 17734-2:2013(E)
4.3 Reagent solution.
In a 1 l volumetric flask, dilute 1,69 ml of DBA in toluene and make up to the mark. The solution is stable
and no special care during storage is necessary.
–1
4.4 Sodium hydroxide, 5 mol⋅l .
Dissolve 200 g of NaOH in water in a beaker, then transfer the solution to a 1 l volumetric flask and make
up to the mark.
4.5 Pyridine, analytical grade.
4.6 Solvents.
The reagent solvent, typically toluene, and other solvents, such as acetonitrile and methanol, should be
of liquid chromatographic quality.
4.7 Formic acid, concentrated formic acid, analytical grade.
4.8 Ethanol, absolute, extra pure 99,5 %.
4.9 HPLC mobile phases.
4.9.1 LC-MS.
The weak mobile phase (mobile phase A) consists of water/acetonitrile (95/5 volume fraction) and
0,05 % formic acid. The strong mobile phase (mobile phase B) consists of water/acetonitrile/methanol
(5/70/25 volume fraction) and 0,05 % formic acid. The mobile phases are degassed prior to use.
4.9.2 LC-CLND.
The weak mobile phase (mobile phase C) consists of water/methanol (95/5 volume fraction) and 0,05 %
formic acid. The strong mobile phase (mobile phase D) consists of water/methanol (5/95 volume fraction)
and 0,05 % formic acid. The mobile phases are degassed prior to use.
5 Standard solutions
5.1 Reference compounds
Reference compounds are necessary for LC-MS determination. For the commercially available amines,
the ethyl chloroformate (ET) derivatives are easily prepared by direct derivatization with ET for
use as calibration standards. The aminoisocyanate derivatives are prepared by reacting one of the
isocyanate groups with DBA and the other group with ethanol. The mixed derivatives formed shall be
characterized before use as calibration standards. Isocyanate, aminoisocyanate, and amine derivatives
for compounds that are not commercially available can be made from the bulk material or from the
thermal decomposition of PUR. Alternatively, standard solutions can be purchased.
5.2 Amine and deuterium-labelled amine derivatives
Calibration standards are made by spiking accurately weighed amounts (ca 0,1 mmol) of amines in
–1
100 ml of toluene. The solution is further diluted to ca 0,01 µmol⋅ml . 5 ml toluene solutions are spiked
with volumes of the amine solutions appropriate for the construction of a calibration curve. The work-
up procedure is then performed; this is described in 8.2.
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ISO 17734-2:2013(E)
The procedure for the synthesis of derivatives is as follows.
1) Dissolve a 10 mmol aliquot of the amines and the deuterium-labelled amines in 20 ml of toluene.
–1
Thereafter, add 150 µl pyridine and 40 ml of 5 mol⋅l NaOH. Then add 1,5 ml of ET dropwise under
continuous stirring.
2) After 10 min, separate the toluene phase.
3) Evaporate the reaction mixture to dryness in a rotating evaporator, and dry the residue under vacuum.
5.3 Aminoisocyanate derivatives
5.3.1 Preparation
Two procedures, A and B, are used to enrich different mixed aminoisocyanate derivatives. The isocyanate
groups in, for example, 2,4-TDI have different reactivity and two different derivatives can be formed.
In procedure A: Dissolve 0,5 mmol of the isocyanates (HDI, 2,4- and 2,6-TDI, and 4,4’-MDI) in 50 ml
isooctane. Add 0,5 mmol of DBA dissolved in isooctane under continuous stirring to the isocyanate
solutions. After 30 min, add excess ethanol to the solutions. Allow the mixtures to react for 16 h.
Evaporate the solutions to dryness and dissolve in methanol.
In procedure B: Dissolve 0,5 mmol of the isocyanate (2,4-TDI) in 50 ml of isooctane; 0,5 mmol of ethanol
dissolved in isooctane is added under continuous stirring to the isocyanate solution. After 16 h, excess
DBA dissolved in isooctane is added to the solution. The solution is allowed to react for 1 h. The solution
is evaporated under a gentle stream of nitrogen. The residue is dissolved in methanol.
The solutions are characterized as described in 5.3.2.
5.3.2 Characterization
Dilute the solutions in methanol to appropriate concentrations and characterize them on the LC-MS
and quantify them on the LC-CLND. This technique is nitrogen specific and any nitrogen-containing
compound (e.g. caffeine) can be used as external standard. The technique is used in several applications.
[8][9][10]
5.4 Thermal decomposition products of polyurethane (PUR)
5.4.1 Preparation of mixed isocyanate, amine, and aminoisocyanate derivatives
During the thermal decomposition of PUR, isocyanates, aminoisocyanates, and amines are formed
that are not commercially available. PUR-based material can be thermally decomposed at appropriate
temperatures. Collect emitted degradation products in impinger flasks (filters in series) containing
–1
0,5 mol⋅l DBA and follow this by the work-up procedure described in 7.2. The solution is characterized
as described in 5.4.2.
5.4.2 Characterization
Qualitative data are obtained with LC-MS. The obtained structural data together with the LC-CLND
data make it possible to calculate the concentrations of different components in the solution. The
characterized and diluted sample solution is used as a calibration standard for LC-MS.
5.5 Stability of the amine and aminoisocyanate derivatives
Solutions of amine-ET and ET-DBA-aminoisocyanate derivatives (MDA, 2,4- and 2,6-TDA, HDA, MAI,
2,4-, 4,2-, and 2,6-TAI, and HAI) have been found stable in toluene, acetonitrile, and methanol for 6 mo
(stored in a dark fridge).
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ISO 17734-2:2013(E)
6 Apparatus
6.1 Sampler.
Sample the air with an impinger flask followed by a filter.
6.1.1 Filter.
Use a 13 mm glass fibre filter (binder free) with a pore size of 0,3 µm.
6.1.2 Filter holder.
Use a 13 mm polypropylene filter holder with luer-lock connections.
6.1.3 Midget impingers.
A midget impinger consists of a tapered inlet tube. Match the two parts so that the distance between the
inlet and the receiver bottom is 1 mm to 2 mm. The filter holder is attached to the outlet of the impinger,
by using an impinger with a luer-lock fitting on the outlet. Alternatively, the filter holder is attached to
the outlet of the impinger by flexible tubing.
6.1.4 Sampling pump, complying to the requirements of ISO 13137, capable of maintaining the flow rate
–1 –1
at 1 l⋅min for impinger-filter sampling and 0,2 l⋅min for solvent-free sampling during the sampling time.
6.1.5 Tubing.
Use rubber tubing of suitable length and of appropriate diameter to ensure a leak-proof fit to both the
pump and the sampler outlet.
6.1.6 Vapour trap.
Use a vapour trap with an internal diameter of 17 mm and a length of 140 mm filled with charcoal (with
a median particle size <3 mm) between the sampler and the sampling pump.
6.2 Flow meter.
Use a portable flow meter capable of measuring the appropriate flow rate with acceptable accuracy.
6.3 Liquid chromatographic system.
In this method, a micro-LC system is used in order to improve the sensitivity, to minimize the maintenance
on the MS, and to minimize the consumption of the mobile phase. The micro-LC system is described in
the following paragraphs. If desired, this system can be replaced by a conventional LC system.
6.3.1 Autosampler.
6.3.1.1 LC-MS.
On-column focusing is performed by partially filled loops (typically 10 µl total volume) of 2 µl loop
injections between 4+4 µl of 50/30/20 water/acetonitrile/methanol. Any commercially available
autosampler capable of making partially filled loop injections and making sample injections of acceptable
accuracy and precision can be used.
6.3.1.2 LC-CLND.
On-column focusing is performed by partially filled loops (typically 10 µl total volume) of 2 µl loop
injections between 4+4 µl of 50/50 methanol/water. Any commercially available autosampler capable
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ISO 17734-2:2013(E)
of making partially filled loop injections and making sample injections of acceptable accuracy and
precision can be used.
6.3.2 Pumping system (LC-MS and LC-CLND).
–1
An HPLC pump capable of gradient elution with a flow rate of 100 µl⋅min is required.
6.3.3 Analytical column (LC-MS and LC-CLND).
An HPLC column capable of separating the different analytes is required.
1)
®
EXAMPLE An example of a suitable column is a PepMap C (50 mm × 1,0 mm with 3 µm particles).
18
6.3.4 Tubing.
Use short (<40 cm) tubing with a small internal diameter (typically ID <0,1 mm).
6.3.5 Detectors.
6.3.5.1 LC-MS.
Any modern MS equipped with a robust and stable electrospray interface will have the necessary
performance. The MS detection is performed with atmospheric pressure ionization, monitoring positive
ions. For quantification, selected ions are monitored. Full spectra are obtained using continuous scans
(typically 50 amu to 1 500 amu) for identification of unknown analytes. If wanted, a UV detector can be
used in series, prior to the MS. The UV detector needs to be equipped with a micro flow cell (typically
300 nl) to minimize peak band broadening.
6.3.5.2 LC-CLND.
Use a detector which is specific for bound nitrogen.
6.4 Ultrasonic bath.
Sonication of samples is necessary to make sure that isocyanate-DBA derivatives are dissolved in
the extraction solution and that the sample remaining after evaporation is properly dissolved in the
added solvent.
6.5 Evaporator.
Equipment for the evaporation of the sample solvent is necessary, preferably a vacuum centrifuge. A
gentle evaporation procedure is desirable since there is a risk that a tough evaporation can result in
losses of the most volatile isocyanate-DBA derivatives.
6.6 Glassware, glass beakers and volumetric flasks (volumetric flasks should conform to ISO 1042).
®
1) PepMap is an example of a suitable product available commercially. This information is given for the convenience
of users of this part of ISO 17734 and does not constitute an endorsement by ISO of this product.
6 © ISO 2013 – All rights reserved
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ISO 17734-2:2013(E)
7 Air sampling
7.1 Pre-sampling laboratory preparation
7.1.1 Cleaning of sampling equipment
Impingers should be taken apart and soaked in alkaline cleaning solution for a minimum of 2 h. The
upper part shall be rinsed with an alkaline cleaning solution, pure water, and finally deionized water. If
the nozzle is clogged, place it in an ultrasonic bath, and then continue with the cleaning procedure. The
lower part should be cleaned in a laboratory dishwasher. Both parts should be dried in an oven.
The filter cassettes and the gaskets should be immersed in ethanol in a glass beaker, sonicated for at
least 15 min, rinsed with deionized water, and dried in an oven.
7.1.2 Preparation of the reagent solution and extraction solution tubes
–1
Prepare test tubes containing 10 ml of 0,01 mol⋅l DBA as the reagent solution for the impingers. If the
gas phase and the particulate phase are to be collected separately, prepare test tubes containing 10 ml
–1
of 0,01 mol⋅l DBA as the extraction solution tubes for the filters.
7.2 Pre-sampling field preparations
Assemble the sampling system with the filter cassette containing the glass fibre filter coupled to the
outlet of the impinger. Transfer the reagent solution to the impinger.
Calibrate the pumps with the impinger-filter sampling system in line, using a portable flow meter. Fill
the impinger with the appropriate amount of reagent solution during calibration. The sampling rate
–1
should be 1 l⋅min .
7.3 Collection of air samples
7.3.1 Measurement task
In order to relate measurement results to occupational exposure limit values, take samples in the
worker’s breathing zone. In order to illustrate risks of being exposed, take stationary samples at
every place at the work-site where isocyanates can be emitted into the air and workers are potentially
exposed. It is also important to include operations that are not frequently performed, for example
repair and maintenance. Differences in materials and batch-to-batch variations are factors that also
should be taken into account when sampling. Collect a
...
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SIST ISO 17734-2:2015
01-marec-2015
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Determination of organonitrogen compounds in air using liquid chromatography and
mass spectrometry - Part 2: Amines and aminoisocyanates using dibutylamine and ethyl
chloroformate derivatives
Détermination des composés organiques azotés dans l'air par chromatographie liquide
et spectrométrie de masse - Partie 2: Amines et aminoisocyanates par les dérivés de la
dibutylamine et du chloroformate d'éthyle
Ta slovenski standard je istoveten z: ISO 17734-2:2013
ICS:
13.040.30 Kakovost zraka na delovnem Workplace atmospheres
mestu
71.040.50 Fizikalnokemijske analitske Physicochemical methods of
metode analysis
SIST ISO 17734-2:2015 en
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
---------------------- Page: 1 ----------------------
SIST ISO 17734-2:2015
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SIST ISO 17734-2:2015
INTERNATIONAL ISO
STANDARD 17734-2
Second edition
2013-12-01
Determination of organonitrogen
compounds in air using liquid
chromatography and mass
spectrometry —
Part 2:
Amines and aminoisocyanates
using dibutylamine and ethyl
chloroformate derivatives
Détermination des composés organiques azotés dans l’air par
chromatographie liquide et spectrométrie de masse —
Partie 2: Amines et aminoisocyanates par les dérivés de la
dibutylamine et du chloroformate d’éthyle
Reference number
ISO 17734-2:2013(E)
©
ISO 2013
---------------------- Page: 3 ----------------------
SIST ISO 17734-2:2015
ISO 17734-2:2013(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2013
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 2013 – All rights reserved
---------------------- Page: 4 ----------------------
SIST ISO 17734-2:2015
ISO 17734-2:2013(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Principle . 1
4 Reagents and materials . 2
5 Standard solutions. 3
5.1 Reference compounds . 3
5.2 Amine and deuterium-labelled amine derivatives . 3
5.3 Aminoisocyanate derivatives . 4
5.4 Thermal decomposition products of polyurethane (PUR). 4
5.5 Stability of the amine and aminoisocyanate derivatives . 4
6 Apparatus . 5
7 Air sampling . 7
7.1 Pre-sampling laboratory preparation . 7
7.2 Pre-sampling field preparations . 7
7.3 Collection of air samples . 7
7.4 Blanks . . 8
7.5 Raw material . 8
7.6 Shipment of samples . 8
8 Laboratory sample preparation . 9
8.1 Sample sequence . 9
8.2 Work-up procedure .10
9 Instrumental settings .10
9.1 HPLC program (LC-MS) .10
9.2 HPLC program [LC-chemiluminescent nitrogen detector (LC-CLND)] .10
9.3 Mass spectrometer .10
10 Data handling .11
10.1 Identification .11
10.2 Calibration curves .11
10.3 Quantification .11
11 Determination of performance characteristics.11
11.1 General .11
11.2 Relevant uncertainty contributions and criteria .12
11.3 Assessment of performance characteristics (following the detailed approach in Reference
[12]) .12
Annex A (informative) Performance characteristics
.20
Annex B (informative) Examples .22
Bibliography .27
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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 146, Air quality, Subcommittee SC 2,
Workplace atmospheres.
This second edition of ISO 17734-2 cancels and replaces ISO 17734-2:2006, which has been
technically revised.
ISO 17734 consists of the following parts, under the general title Determination of organonitrogen
compounds in air using liquid chromatography and mass spectrometry:
— Part 1: Isocyanates using dibutylamine derivatives
— Part 2: Amines and aminoisocyanates using dibutylamine and ethyl chloroformate derivatives
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Introduction
In many applications, when considering isocyanates as a workplace contaminant, there is also a need to
investigate the presence of aminoisocyanates and amines. During thermal decomposition of polyurethane
[1][2][3][4][5][6]
(PUR), not only isocyanates, but also amines and aminoisocyanates, are formed.
The determination of isocyanates in the work environment using DBA as a reagent has been
demonstrated to be a robust method (see ISO 17734-1). Using the DBA method and derivatization with
ethyl chloroformate in the following work-up procedure makes simultaneous determination of amines,
[6][7]
aminoisocyanates, and isocyanates possible.
For quantification of amine and aminoisocyanate derivatives, reference compounds are necessary,
but are only available for a few diamines. Aminoisocyanates cannot be analysed directly because they
react with themselves. In this method, a nitrogen-specific detector has been used for quantification of
amine and aminoisocyanate derivatives in reference solutions. This technique has been demonstrated
to be a useful tool, together with MS characterization, in greatly facilitating the production of reference
[6]
solutions.
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INTERNATIONAL STANDARD ISO 17734-2:2013(E)
Determination of organonitrogen compounds in air using
liquid chromatography and mass spectrometry —
Part 2:
Amines and aminoisocyanates using dibutylamine and
ethyl chloroformate derivatives
1 Scope
This part of ISO 17734 gives general guidance for the sampling and analysis of airborne amines and
aminoisocyanates in workplace air. It is strongly recommended that the determination of amines and
aminoisocyanates is made together with the determination of isocyanates in air, using DBA as a reagent
(see ISO 17734-1).
The method can be used for simultaneous determinations of amines, such as 4,4’-methylenediphenyldiamine
(4,4’-MDA), 2,4- and 2,6-toluenediamine (2,4- and 2,6-TDA), and 1,6-hexamethylenediamine (1,6-
HDA), and compounds containing both isocyanate and amine groups, such as 4,4’-methylenediphenyl
aminoisocyanate (4,4’-MAI), 2,4-, 4,2-, and 2,6-toluene aminoisocyanate (2,4-, 4,2-, and 2,6-TAI), and
1,6-hexamethylene aminoisocyanate (1,6-HAI). The method is suitable for collecting amines and
aminoisocyanates in both the gas and particle phases. The instrumental detection limit for the amines
is about 5 nmol/sample and for the aminoisocyanate, it is about 0,3 nmol/sample. For a 15 l air sample,
–3 –3
this corresponds to 0,4 ng⋅m for TDA and 0,03 ng⋅m for TAI.
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 16200-1, Workplace air quality — Sampling and analysis of volatile organic compounds by solvent
desorption/gas chromatography — Part 1: Pumped sampling method
ISO 5725-2, Accuracy (trueness and precision) of measurement methods and results — Part 2: Basic method
for the determination of repeatability and reproducibility of a standard measurement method
3 Principle
The method permits the simultaneous sampling and analysis of amines, aminoisocyanates, and
isocyanates. Only amines and aminoisocyanates are discussed in this part of ISO 17734, because
isocyanates are considered in ISO 17734-1.
Samples are collected by drawing a known volume of air through a midget impinger flask followed by a
–1
filter. The impinger contains 10 ml of 0,01 mol⋅l of di-n-butylamine (DBA) in toluene, and the filter is a
glass fibre filter with no binder. After sampling, deuterium-labelled amine-ethyl chloroformate (ET) and
isocyanate-DBA derivatives (used as internal standard) are added to the sample solutions. The excess
reagent and solvent are evaporated, and the samples are dissolved in acetonitrile. The samples are
analysed using reversed-phase liquid chromatography (LC) and electrospray (ESP) mass spectrometric
(MS) detection, monitoring positive ions. Quantification is made by quantifying selected ions.
Quantification and qualitative determinations can be performed using different LC-MS techniques. LC-
CLND (chemiluminescent nitrogen detection) or, for aromatic isocyanates, aminoisocyanates, and amines,
LC-UV (ultraviolet detection) can be used for the determination of higher concentrations. Reference
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materials can be characterized using LC-MS/CLND. For characterization of volatile compounds, a GC-
thermoionic specific detector (TSD) can also be used.
Gas phase and large particles (>1,5
Sampling
µ m ) are collected in the impinger
with impinger and filter flask. Small particles (0,01 µm
to 1,5 µm) are collected . on the filter
Standard solution:
Calibration curve Transfer of sample solution Isocyanate-DBA, amine-ET and
aminoisocyanate-DBA-ET
from standard solution and filter to test tube
derivatives
Internal standard, IS:
- Similar in retention time
- Similar in molecular structure
Addition of IS
i.e. aromatic or aliphatic
- Preferably deuterium-labelled
derivative
-Extraction of analytes on the filter
-Derivatisation of the amine groups
with ET in a two phase
Work-up procedure
derivatisation procedure
- Change of solution media
- Enrichment of the analytes
+
- SIM of molecular ion [ MH ]
Analysis with LC-MS
of analytes and IS
- Calculation of the area ratio of the
analyte and IS
Evaluation of data
- Comparison between area ratio of
the air sample and calibration plot
Figure 1 — Principle of the described method
4 Reagents and materials
4.1 DBA reagent.
Analytical grade di-n-butylamine is commercially available.
4.2 Ethyl chloroformate reagent.
Analytical grade ethyl chloroformate is commercially available.
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4.3 Reagent solution.
In a 1 l volumetric flask, dilute 1,69 ml of DBA in toluene and make up to the mark. The solution is stable
and no special care during storage is necessary.
–1
4.4 Sodium hydroxide, 5 mol⋅l .
Dissolve 200 g of NaOH in water in a beaker, then transfer the solution to a 1 l volumetric flask and make
up to the mark.
4.5 Pyridine, analytical grade.
4.6 Solvents.
The reagent solvent, typically toluene, and other solvents, such as acetonitrile and methanol, should be
of liquid chromatographic quality.
4.7 Formic acid, concentrated formic acid, analytical grade.
4.8 Ethanol, absolute, extra pure 99,5 %.
4.9 HPLC mobile phases.
4.9.1 LC-MS.
The weak mobile phase (mobile phase A) consists of water/acetonitrile (95/5 volume fraction) and
0,05 % formic acid. The strong mobile phase (mobile phase B) consists of water/acetonitrile/methanol
(5/70/25 volume fraction) and 0,05 % formic acid. The mobile phases are degassed prior to use.
4.9.2 LC-CLND.
The weak mobile phase (mobile phase C) consists of water/methanol (95/5 volume fraction) and 0,05 %
formic acid. The strong mobile phase (mobile phase D) consists of water/methanol (5/95 volume fraction)
and 0,05 % formic acid. The mobile phases are degassed prior to use.
5 Standard solutions
5.1 Reference compounds
Reference compounds are necessary for LC-MS determination. For the commercially available amines,
the ethyl chloroformate (ET) derivatives are easily prepared by direct derivatization with ET for
use as calibration standards. The aminoisocyanate derivatives are prepared by reacting one of the
isocyanate groups with DBA and the other group with ethanol. The mixed derivatives formed shall be
characterized before use as calibration standards. Isocyanate, aminoisocyanate, and amine derivatives
for compounds that are not commercially available can be made from the bulk material or from the
thermal decomposition of PUR. Alternatively, standard solutions can be purchased.
5.2 Amine and deuterium-labelled amine derivatives
Calibration standards are made by spiking accurately weighed amounts (ca 0,1 mmol) of amines in
–1
100 ml of toluene. The solution is further diluted to ca 0,01 µmol⋅ml . 5 ml toluene solutions are spiked
with volumes of the amine solutions appropriate for the construction of a calibration curve. The work-
up procedure is then performed; this is described in 8.2.
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The procedure for the synthesis of derivatives is as follows.
1) Dissolve a 10 mmol aliquot of the amines and the deuterium-labelled amines in 20 ml of toluene.
–1
Thereafter, add 150 µl pyridine and 40 ml of 5 mol⋅l NaOH. Then add 1,5 ml of ET dropwise under
continuous stirring.
2) After 10 min, separate the toluene phase.
3) Evaporate the reaction mixture to dryness in a rotating evaporator, and dry the residue under vacuum.
5.3 Aminoisocyanate derivatives
5.3.1 Preparation
Two procedures, A and B, are used to enrich different mixed aminoisocyanate derivatives. The isocyanate
groups in, for example, 2,4-TDI have different reactivity and two different derivatives can be formed.
In procedure A: Dissolve 0,5 mmol of the isocyanates (HDI, 2,4- and 2,6-TDI, and 4,4’-MDI) in 50 ml
isooctane. Add 0,5 mmol of DBA dissolved in isooctane under continuous stirring to the isocyanate
solutions. After 30 min, add excess ethanol to the solutions. Allow the mixtures to react for 16 h.
Evaporate the solutions to dryness and dissolve in methanol.
In procedure B: Dissolve 0,5 mmol of the isocyanate (2,4-TDI) in 50 ml of isooctane; 0,5 mmol of ethanol
dissolved in isooctane is added under continuous stirring to the isocyanate solution. After 16 h, excess
DBA dissolved in isooctane is added to the solution. The solution is allowed to react for 1 h. The solution
is evaporated under a gentle stream of nitrogen. The residue is dissolved in methanol.
The solutions are characterized as described in 5.3.2.
5.3.2 Characterization
Dilute the solutions in methanol to appropriate concentrations and characterize them on the LC-MS
and quantify them on the LC-CLND. This technique is nitrogen specific and any nitrogen-containing
compound (e.g. caffeine) can be used as external standard. The technique is used in several applications.
[8][9][10]
5.4 Thermal decomposition products of polyurethane (PUR)
5.4.1 Preparation of mixed isocyanate, amine, and aminoisocyanate derivatives
During the thermal decomposition of PUR, isocyanates, aminoisocyanates, and amines are formed
that are not commercially available. PUR-based material can be thermally decomposed at appropriate
temperatures. Collect emitted degradation products in impinger flasks (filters in series) containing
–1
0,5 mol⋅l DBA and follow this by the work-up procedure described in 7.2. The solution is characterized
as described in 5.4.2.
5.4.2 Characterization
Qualitative data are obtained with LC-MS. The obtained structural data together with the LC-CLND
data make it possible to calculate the concentrations of different components in the solution. The
characterized and diluted sample solution is used as a calibration standard for LC-MS.
5.5 Stability of the amine and aminoisocyanate derivatives
Solutions of amine-ET and ET-DBA-aminoisocyanate derivatives (MDA, 2,4- and 2,6-TDA, HDA, MAI,
2,4-, 4,2-, and 2,6-TAI, and HAI) have been found stable in toluene, acetonitrile, and methanol for 6 mo
(stored in a dark fridge).
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6 Apparatus
6.1 Sampler.
Sample the air with an impinger flask followed by a filter.
6.1.1 Filter.
Use a 13 mm glass fibre filter (binder free) with a pore size of 0,3 µm.
6.1.2 Filter holder.
Use a 13 mm polypropylene filter holder with luer-lock connections.
6.1.3 Midget impingers.
A midget impinger consists of a tapered inlet tube. Match the two parts so that the distance between the
inlet and the receiver bottom is 1 mm to 2 mm. The filter holder is attached to the outlet of the impinger,
by using an impinger with a luer-lock fitting on the outlet. Alternatively, the filter holder is attached to
the outlet of the impinger by flexible tubing.
6.1.4 Sampling pump, complying to the requirements of ISO 13137, capable of maintaining the flow rate
–1 –1
at 1 l⋅min for impinger-filter sampling and 0,2 l⋅min for solvent-free sampling during the sampling time.
6.1.5 Tubing.
Use rubber tubing of suitable length and of appropriate diameter to ensure a leak-proof fit to both the
pump and the sampler outlet.
6.1.6 Vapour trap.
Use a vapour trap with an internal diameter of 17 mm and a length of 140 mm filled with charcoal (with
a median particle size <3 mm) between the sampler and the sampling pump.
6.2 Flow meter.
Use a portable flow meter capable of measuring the appropriate flow rate with acceptable accuracy.
6.3 Liquid chromatographic system.
In this method, a micro-LC system is used in order to improve the sensitivity, to minimize the maintenance
on the MS, and to minimize the consumption of the mobile phase. The micro-LC system is described in
the following paragraphs. If desired, this system can be replaced by a conventional LC system.
6.3.1 Autosampler.
6.3.1.1 LC-MS.
On-column focusing is performed by partially filled loops (typically 10 µl total volume) of 2 µl loop
injections between 4+4 µl of 50/30/20 water/acetonitrile/methanol. Any commercially available
autosampler capable of making partially filled loop injections and making sample injections of acceptable
accuracy and precision can be used.
6.3.1.2 LC-CLND.
On-column focusing is performed by partially filled loops (typically 10 µl total volume) of 2 µl loop
injections between 4+4 µl of 50/50 methanol/water. Any commercially available autosampler capable
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of making partially filled loop injections and making sample injections of acceptable accuracy and
precision can be used.
6.3.2 Pumping system (LC-MS and LC-CLND).
–1
An HPLC pump capable of gradient elution with a flow rate of 100 µl⋅min is required.
6.3.3 Analytical column (LC-MS and LC-CLND).
An HPLC column capable of separating the different analytes is required.
1)
®
EXAMPLE An example of a suitable column is a PepMap C (50 mm × 1,0 mm with 3 µm particles).
18
6.3.4 Tubing.
Use short (<40 cm) tubing with a small internal diameter (typically ID <0,1 mm).
6.3.5 Detectors.
6.3.5.1 LC-MS.
Any modern MS equipped with a robust and stable electrospray interface will have the necessary
performance. The MS detection is performed with atmospheric pressure ionization, monitoring positive
ions. For quantification, selected ions are monitored. Full spectra are obtained using continuous scans
(typically 50 amu to 1 500 amu) for identification of unknown analytes. If wanted, a UV detector can be
used in series, prior to the MS. The UV detector needs to be equipped with a micro flow cell (typically
300 nl) to minimize peak band broadening.
6.3.5.2 LC-CLND.
Use a detector which is specific for bound nitrogen.
6.4 Ultrasonic bath.
Sonication of samples is necessary to make sure that isocyanate-DBA derivatives are dissolved in
the extraction solution and that the sample remaining after evaporation is properly dissolved in the
added solvent.
6.5 Evaporator.
Equipment for the evaporation of the sample solvent is necessary, preferably a vacuum centrifuge. A
gentle evaporation procedure is desirable since there is a risk that a tough evaporation can result in
losses of the most volatile isocyanate-DBA derivatives.
6.6 Glassware, glass beakers and volumetric flasks (volumetric flasks should conform to ISO 1042).
®
1) PepMap is an example of a suitable product available commercially. This information is given for the convenience
of users of this part of ISO 17734 and does not constitute an endorsement by ISO of this product.
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7 Air sampling
7.1 Pre-sampling laboratory preparation
7.1.1 Cleaning of sampling equipment
Impingers should be taken apart and soaked in alkaline cleaning solution for a minimum of 2 h. The
upper part shall be rinsed with an alkaline cleaning solution, pure water, and finally deionized water. If
the nozzle is clogged, place it in an ultrasonic bath, and then continue with the cleaning procedure. The
lower part should be cleaned in a laboratory dishwasher. Both parts should be dried in an oven.
The filter cassettes and the
...
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