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International
Standard
ISO 18363-3
Second edition
Animal and vegetable fats and
2024-06
oils — Determination of fatty-acid-
bound chloropropanediols (MCPDs)
and glycidol by GC/MS —
Part 3:
Method using acid
transesterification and
measurement for 2-MCPD, 3-MCPD
and glycidol
Corps gras d'origines animale et végétale — Détermination des
esters de chloropropanediols (MCPD) et d'acides gras et des
esters de glycidol et d'acides gras par CPG/SM —
Partie 3: Méthode par transestérification acide et mesure du
2-MCPD, du 3-MCPD et du glycidol
Reference number
© ISO 2024
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Published in Switzerland
ii
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 2
5 Reagents . 2
5.1 Standard and reference compounds .2
5.2 Standard solutions .3
5.2.1 General .3
5.2.2 Stock solutions (1 mg/ml) .3
5.2.3 Working solutions .3
5.3 Other reagents .4
5.4 Reagent solutions .4
6 Apparatus . 5
7 Sample . 5
7.1 Sampling .5
7.2 Preparation of the test sample .5
8 Procedure . 6
8.1 Test sample preparation .6
8.2 Preparation of the calibration curve .7
8.3 Gas chromatography/mass spectrometry references .7
9 Expression of results . 8
9.1 Quantification of 3-MCPD esters .8
9.2 Quantification of 2-MCPD esters .8
9.3 Quantification of glycidyl esters .9
10 Precision . 10
10.1 General .10
10.2 Repeatability .10
10.3 Between-day reproducibility . .10
11 Test report . 10
Annex A (informative) Construction of the calibration curves .11
Annex B (informative) Results of the interlaboratory test .15
Bibliography . 17
iii
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out through
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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 document should be noted. This document was drafted in accordance with the editorial rules of the
ISO/IEC Directives, Part 2 (see www.iso.org/directives).
ISO draws attention to the possibility that the implementation of this document may involve the use of (a)
patent(s). ISO takes no position concerning the evidence, validity or applicability of any claimed patent
rights in respect thereof. As of the date of publication of this document, ISO had not received notice of (a)
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This document was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 11,
Animal and vegetable fats and oils, in collaboration with the European Committee for Standardization (CEN)
Technical Committee CEN/TC 307, Oilseeds, vegetable and animal fats and oils and their by-products – Methods
of sampling and analysis, in accordance with the Agreement on technical cooperation between ISO and CEN
(Vienna Agreement).
This second edition cancels and replaces the first edition (ISO 18363-3:2017), of which it constitutes a minor
revision.
The main changes are as follows:
— the text of the Introduction has been revised to be consistent with ISO 18363-4:2021.
A list of all parts in the ISO 18363 series can be found on the ISO website.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www.iso.org/members.html.
iv
Introduction
[1]
The ISO 18363 series can be used for the determination of ester-bound MCPD and glycidol. This
introduction describes the methods specified in the different parts so that the analyst can decide which
methods are suitable for application. The detailed application of each method is contained within the scope
of each individual method.
[2]
ISO 18363-1 is a differential method equivalent to DGF standard C-VI 18 (10) and identical to AOCS Official
[3]
Method Cd 29c-13 . In brief, it is based on a fast alkaline catalysed release of 3-MCPD and glycidol from the
ester derivatives. Glycidol is subsequently converted into induced 3-MCPD. It consists of two parts. The first
part (A) allows the determination of the sum of ester-bound 3-MCPD and ester-bound glycidol, whereas the
second part (B) determines ester-bound 3-MCPD only. Both assays are based on the release of the target
analytes 3-MCPD and glycidol from the ester-bound form by an alkaline catalysed alcoholysis carried
out at room temperature. In part A, an acidified sodium chloride solution is used to stop the reaction and
subsequently convert the glycidol into induced 3-MCPD. Thus, 3-MCPD and glycidol become indistinguishable
in part A. In part B, the reaction stop is achieved by the addition of an acidified chloride-free salt solution
which also prevents the conversion of glycidol into induced MCPD. Consequently, part B allows the
determination of the genuine 3-MCPD content. Finally, the glycidol content of the sample is proportional to
the difference of both assays (A – B) and can be calculated when the transformation ratio from glycidol to
3-MCPD has been determined. ISO 18363-1 is applicable to the fast determination of ester-bound 3-MCPD
and glycidol in refined and non-refined vegetable oils and fats. ISO 18363-1 can also apply to animal fats and
used frying oils and fats, but a validation study must be undertaken before the analysis of these matrices.
Any free analytes within the sample would be included in the results, but the document does not allow the
distinction between free and bound analytes. However, as of publication of this document, research has not
shown any evidence of a free analyte content as high as the esterified analyte content in refined vegetable
oils and fats. In principle, ISO 18363-1 can also be modified in such a way that the determination of 2-MCPD
is feasible, but again, a validation study must be undertaken before the analysis of this analyte.
[4]
ISO 18363-2 represents AOCS Official Method Cd 29b-13 . In brief, it is based on a slow alkaline release of
MCPD and glycidol from the ester derivatives. Glycidol is subsequently converted into 3-MBPD. ISO 18363-2
consists of two sample preparations that differ in the use of internal standards. Both preparations are used
for the determination of ester-bound 2-MCPD and 3-MCPD. In part A, a preliminary result for ester-bound
glyc
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