EN 17958:2024
(Main)Food authenticity - Determination of the δ13C value of mono- (fructose and glucose), di-, and trisaccharides in honey by liquid chromatography-isotope ratio mass spectrometry (LC-IRMS)
Food authenticity - Determination of the δ13C value of mono- (fructose and glucose), di-, and trisaccharides in honey by liquid chromatography-isotope ratio mass spectrometry (LC-IRMS)
This document specifies a method for the determination of the ratio of stable isotopes of carbon (13C/12C) of sugars contained in honey by using liquid chromatography coupled to an isotope ratio mass spectrometer (LC-IRMS) for compound separation and subsequent determination of the 13C/12C ratio of mono-, di-, and trisaccharides. These ratios can be used to assess honey authenticity by comparing them to guidance values of genuine honey, which have been previously agreed by subject matter experts, as the 13C/12C ratios of sugars of genuine honey and sugars contained in adulterants (syrups made from starch-rich plants or from sugar cane or sugar beet) differ to a certain extent.
The compliance assessment process is not part of this document.
Lebensmittelauthentizität - Bestimmung des δ13C-Wertes von Mono- (Fructose und Glucose), Di- und Trisacchariden in Honig durch Flüssigchromatographie-Isotopenverhältnis-Massenspektrometrie (LC‑IRMS)
Dieses Dokument legt ein Verfahren zur Bestimmung des Verhältnisses stabiler Kohlenstoffisotope (13C/12C) von in Honig enthaltenen Zuckern mittels Flüssigchromatographie gekoppelt mit einem Isotopenverhältnis-Massenspektrometer (LC-IRMS) zur Trennung der Verbindungen und anschließenden Bestimmung des 13C/12C-Verhältnisses von Mono-, Di- und Trisacchariden fest. Diese Verhältnisse können zur Beurteilung der Authentizität von Honig verwendet werden, indem sie mit den Richtwerten für echten Honig verglichen werden, die zuvor von Fachexperten vereinbart wurden, da sich die 13C/12C-Verhältnisse von Zuckern in echtem Honig und Zuckern in Verfälschungsmitteln (Sirupe aus stärkereichen Pflanzen oder aus Zuckerrohr oder Zuckerrüben) in gewissem Maße unterscheiden.
Der Prozess der Konformitätsbewertung ist nicht Teil dieses Dokuments.
Authenticité des aliments - Détermination de la valeur du δ13C des mono- (fructose et glucose), di-, et trisaccharides présents dans le miel par chromatographie en phase liquide spectrométrie de masse de rapports isotopiques (CL-SMRI)
Le présent document spécifie une méthode de détermination du rapport des isotopes stables du carbone (13C/12C) des sucres contenus dans le miel par chromatographie en phase liquide couplée à la spectrométrie de masse de rapports isotopiques (CL SMRI). La méthode présente la séparation des composés et la détermination des teneurs en 13C/12C des mono-, di- et trisaccharides. Ces rapports isotopiques peuvent être utilisés pour évaluer l’authenticité du miel par comparaison avec des valeurs recommandées pour le miel authentique et convenues par des spécialistes du sujet, car les rapports 13C/12C des sucres du miel authentique, et ceux des sucres contenus dans les produits adultérants du miel (sirops fabriqués à partir de plantes riches en amidon ou à partir de sucre de canne ou de betterave) sont, dans une certaine mesure, différents les uns des autres.
Le processus d’évaluation de la conformité n’est pas traité dans le présent document.
Pristnost živil - Določanje vrednosti δ13C mono- (fruktoza in glukoza), di- in trisaharidov v medu s tekočinsko kromatografijo - z masno spektrometrijo z izotopskim razmerjem (LC-IRMS)
General Information
Standards Content (Sample)
SLOVENSKI STANDARD
oSIST prEN 17958:2023
01-april-2023
Avtentičnost hrane - Določanje vrednosti d13C mono- (fruktoza in glukoza), di- in
trisaharidov v medu z masno spektrometrijo, tekočinsko kromatografijo in
izotopskim razmerjem (LC-IRMS)
Food authenticity - Determination of the d13C value of mono- (fructose and glucose), di-,
and trisaccharides in honey by liquid chromatography-isotope ratio mass spectrometry
(LC-IRMS)
Lebensmittelauthentizität - Bestimmung des δ13C-Wertes von Mono- (Fructose und
Glucose), Di- und Trisacchariden in Honig durch Flüssigchromatographie-
Isotopenverhältnis-Massenspektrometrie (LC-IRMS)
Authenticité des aliments - Détermination de la valeur du δ13C des mono- (fructose et
glucose), di-, et trisaccharides présents dans le miel par chromatographie en phase
liquide spectrométrie de masse de rapports isotopiques (CL-SMRI)
Ta slovenski standard je istoveten z: prEN 17958
ICS:
67.180.10 Sladkor in sladkorni izdelki Sugar and sugar products
oSIST prEN 17958:2023 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
oSIST prEN 17958:2023
oSIST prEN 17958:2023
DRAFT
EUROPEAN STANDARD
prEN 17958
NORME EUROPÉENNE
EUROPÄISCHE NORM
February 2023
ICS 67.180.10; 67.180.20
English Version
Food authenticity - Determination of the δ C value of
mono- (fructose and glucose), di-, and trisaccharides in
honey by liquid chromatography-isotope ratio mass
spectrometry (LC-IRMS)
Lebensmittelauthentizität - Bestimmung des δ C-
Wertes von Mono- (Fructose und Glucose), Di- und
Trisacchariden in Honig durch
Flüssigchromatographie-Isotopenverhältnis-
Massenspektrometrie (LC-IRMS)
This draft European Standard is submitted to CEN members for enquiry. It has been drawn up by the Technical Committee
CEN/TC 460.
If this draft becomes a European Standard, CEN members are bound to comply with the CEN/CENELEC Internal Regulations
which stipulate the conditions for giving this European Standard the status of a national standard without any alteration.
This draft European Standard was established by CEN in three official versions (English, French, German). A version in any other
language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC
Management Centre has the same status as the official versions.
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, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and
United Kingdom.
Recipients of this draft are invited to submit, with their comments, notification of any relevant patent rights of which they are
aware and to provide supporting documentation.
Warning : This document is not a European Standard. It is distributed for review and comments. It is subject to change without
notice and shall not be referred to as a European Standard.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2023 CEN All rights of exploitation in any form and by any means reserved Ref. No. prEN 17958:2023 E
worldwide for CEN national Members.
oSIST prEN 17958:2023
prEN 17958:2023 (E)
Contents Page
European foreword . 3
Introduction . 4
1 Scope . 5
2 Normative references . 5
3 Terms and definitions . 5
4 Principle . 6
5 Reagents . 6
6 Apparatus . 7
7 Procedure. 8
7.1 Preparation of the test sample . 8
7.2 Preparation of the test sample solution . 8
7.3 LC-IRMS setup and performance qualification . 8
7.3.1 LC-IRMS interface setup . 8
7.3.2 IRMS calibration . 8
7.4 Determination and analytical sequence . 8
7.5 Calculations . 8
7.5.1 General . 8
7.5.2 Calculation of compound specific δ C values using two-point linear normalization . 8
8 Precision . 9
8.1 General . 9
8.2 Repeatability . 9
8.3 Reproducibility . 9
Annex A (informative) Experimental conditions used by the participants in the
interlaboratory comparison . 10
Annex B (informative) Precision data . 11
Bibliography . 15
oSIST prEN 17958:2023
prEN 17958:2023 (E)
European foreword
This document (prEN 17958:2023) has been prepared by Technical Committee CEN/TC 460 “Food
Authenticity”, the secretariat of which is held by DIN.
This document is currently submitted to the CEN Enquiry.
oSIST prEN 17958:2023
prEN 17958:2023 (E)
Introduction
Honey is a natural sweetener having attractive sensory properties. Demand has increased over the
years, partially due to population increase, but also due to the preference of consumers for natural and
unprocessed food. It is a globally traded commodity following complex trade routes, which makes
quality and authenticity control difficult. Honey is among the commodities most vulnerable to fraud [1].
The EU Honey Directive lays down the composition and authenticity criteria for honey [2]. A set of
analytical methods have been standardized by the International Honey Commission [3], which allows
enforcement of the provisions of the Directive. Those methods are widely used but are not always
appropriate for assessing the authenticity of the product: specifically, adulteration of honey by non-
declared dilution with foreign sugars/sweeteners, which is among the most frequently encountered
cases.
Syrups that mimic the composition of honey that are produced by chemical and/or enzymatic
modification of starch or sucrose are difficult to detect [4]. If the starting product is obtained from a
plant using the Hatch–Slack pathway for carbon fixation (C4 plant), such as maize or sugar cane, stable
carbon isotope ratio analysis (SCIRA) using a combination of an elemental analyzer and an isotope ratio
mass spectrometer (EA-IRMS) offers a possibility to detect additions down to a level of 7 % [5]. Sugars
originating from C3 plants, which use the Calvin–Benson cycle, such as beet root or sugars generated
from rice or wheat starch escape detection by SCIRA. Combining liquid chromatography (LC) with IRMS
(LC-IRMS) offers new possibilities for detecting honey adulteration with sugars derived from C3 plants
as well as increasing the sensitivity for detecting C4 sugars [6][7]. The method has gained popularity
but has never been subjected to multi-laboratory validation, which is a prerequisite for further
developing it into a standard by a Standards Developing Organization.
13 12
An LC-IRMS method for the determination of the C/ C isotope ratios of glucose, fructose, glycerol
and ethanol in products of viti-vinicultural origin was collaboratively studied by the International
Organization of Vine and Wine (OIV) and endorsed for inclusion in the Compendium of International
Methods of Analysis of Wines and Musts (OIV-OENO resolution 479-2017).
This document provides the basis for the analytical method. The setup of the required apparatus
depends to a large extent on its design principles, and the specific recommendations of the
manufacturers should be followed. It is intended to serve as a frame in which the analyst can define
their own analytical work in accordance with the standard procedure.
oSIST prEN 17958:2023
prEN 17958:2023 (E)
1 Scope
This document specifies a method for the determination of the ratio of stable isotopes of carbon
13 12
( C/ C) of sugars contained in honey by using liquid chromatography coupled to an isotope ratio
13 12
mass spectrometer (LC-IRMS) for compound separation and subsequent determination of the C/ C
ratio of mono-, di-, and trisaccharides. These ratios can be used to assess honey authenticity by
13 12
comparing them to published guidance values of genuine honey as the C/ C ratios of sugars of
genuine honey and sugars contained in adulterants (syrups made from starch-rich plants or from sugar
cane or sugar beet) differ to a certain extent. The compliance assessment process is not part of this
document.
2 Normative references
There are no normative references in this document.
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
ISO and IEC maintain terminology databases for use in standardization at the following addresses:
— IEC Electropedia: available at https://www.electropedia.org/
— ISO Online browsing platform: available at https://www.iso.org/obp
3.1
isotope delta
δ
stable isotope ratio of a sample expressed relative to a reference
Note 1 to entry: For carbon, this expression is given in Formula (1):
13 12
R C/ C
( )
sample
13 12
δ C/ C − 1 (1)
( )
ref
13 12
R C/ C
( )
reference
13 12 13
Note 2 to entry: The term 𝛿𝛿 ( C/ C) is often changed from the IUPAC format to 𝛿𝛿 C ; this document uses the
ref ref
IUPAC format for familiarity.
Note 3 to entry: To ensure international comparability of isotope delta values, a common reference is used; this
reference is an international measurement standard assigned by convention with isotope delta value exactly equal
to zero.
Note 4 to entry: Carbon isotope delta values for natural isotopic abundance in food materials are small and
expressed in permille (‰) rather than in their native form.
3.2
Vienna Peedee Belemnite
VPDB
international measurement standard for 𝛿𝛿 C
Note 1 to entry: 𝛿𝛿 C is a virtual carbonate.
=
oSIST prEN 17958:2023
prEN 17958:2023 (E)
3.2.1
VPDB carbon isotope delta scale
exact isotope delta value assigned to the calcium carbonate reference material NBS 19 of
𝛿𝛿 C = +1,95 ‰
VPDB
Note 1 to entry: To ensure traceability to the VPDB scale, carbon isotope delta values are to be normalized using
two or more reference materials to account for scale effects during measurement.
3.3
mass-to-charge ratio
m/z
dimensionless quantity formed by dividing the mass number of an ion by its charge number
3.4
working gas
gas consisting of the same molecule as the analyte gas (i.e. CO ) but introduced directly into the mass
spectrometer from a high-pressure cylinder rather than being created from the sample
3.5
sequence
continuous set of analyses including reference materials for normalization, QA/QC materials,
procedural blanks and samples prepared and analyzed together
4 Principle
Honey is diluted with water, filtered and injected into a liquid chromatography system for separation
into mono-, di-, tri- and oligosaccharides. The column effluent is fed into an interface where organic
compounds are oxidized to CO . CO isotopologues with m/z 44, 45 and 46 are separated in the
2 2
spectrometer and detected using Faraday cups. Compound specific δ C (‰) values are then calculated
according to Formula (1).
5 Reagents
Unless otherwise stated, use only reagents of recognized analytical grade (purity ≥ 99 %) and ultrapure
−1
water with a resistivity > 18 MΩ cm .
5.1 Working gas, carbon dioxide, 99,995 %.
5.2 Helium, 99,999 %.
5.3 Certified reference materials, at least two reference materials with certified 𝛿𝛿 C values
traceable to VPDB and sp
...
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