EN 17203:2021
(Main)Foodstuffs - Determination of citrinin in food by HPLC-MS/MS
Foodstuffs - Determination of citrinin in food by HPLC-MS/MS
This document describes a procedure for the determination of the citrinin content in food (cereals, red yeast rice (RYR)), herbs and food supplements by liquid chromatography tandem mass spectrometry (LC-MS/MS).
This method has been validated for citrinin in red yeast rice and in the formulated food supplements in the range of 2,5 μg/kg to 3 000 μg/kg and in wheat flour in the range of 2,5 μg/kg to 100 μg/kg.
Laboratory experiences have shown that this method is also applicable to white rice, herbs such as a powder of ginkgo biloba leaves and the formulated food supplements in the range of 2,5 μg/kg to 50 μg/kg.
Lebensmittel - Bestimmung von Citrinin in Lebensmitteln mit HPLC-MS/MS
Dieses Dokument beschreibt ein Verfahren zur Bestimmung des Citriningehalts in Lebensmitteln (Getreide, rotem Reis [RYR, en: red yeast rice]), Kräutern und Nahrungsergänzungsmitteln mittels Flüssigchromato¬graphie und Tandem Massenspektrometrie (LC MS/MS).
Dieses Verfahren wurde für Citrinin in rotem Reis und in entsprechend zusammengesetzten Nahrungsergänzungsmitteln im Bereich von 2,5 μg/kg bis 3 000 μg/kg sowie für Citrinin in Weizenmehl im Bereich von 2,5 μg/kg bis 100 μg/kg validiert.
Laborerfahrungen haben gezeigt, dass dieses Verfahren auch auf weißen Reis, auf Kräuter wie Ginkgo-Biloba-Blätter in Pulverform und entsprechend zusammengesetzte Nahrungsergänzungsmittel im Bereich von 2,5 μg/kg bis 50 μg/kg anwendbar ist.
Produits alimentaires - Dosage de la citrinine dans les produits alimentaires par CLHP-SM/SM
Le présent document décrit un mode opératoire de détermination de la teneur en citrinine dans les aliments [céréales, levure de riz rouge (LRR)], les herbes aromatiques et les compléments alimentaires par chromatographie liquide couplée à une spectrométrie de masse en tandem (CL-SM/SM).
Cette méthode a été validée pour la citrinine dans la levure de riz rouge et dans les compléments alimentaires à base de levure de riz rouge dans la gamme de concentration de 2,5 μg/kg à 3 000 μg/kg et dans la farine de blé dans la gamme de concentration de 2,5 μg/kg à 100 μg/kg.
Les expériences en laboratoire ont montré que cette méthode est également applicable au riz blanc, aux herbes aromatiques telles que les feuilles de Ginkgo biloba en poudre et aux compléments alimentaires formulés dans la gamme de concentration de 2,5 μg/kg à 50 μg/kg.
Živila - Določevanje citrinina v živilih z metodo HPLC-MS/MS
General Information
Relations
Standards Content (Sample)
SLOVENSKI STANDARD
01-september-2021
Nadomešča:
SIST EN 17203:2019
Živila - Določevanje citrinina v živilih z metodo HPLC-MS/MS
Foodstuffs - Determination of citrinin in food by HPLC-MS/MS
Lebensmittel - Bestimmung von Citrinin in Lebensmitteln mit HPLC-MS/MS
Produits alimentaires - Dosage de la citrinine dans les produits alimentaires par CLHP-
SM/SM
Ta slovenski standard je istoveten z: EN 17203:2021
ICS:
67.050 Splošne preskusne in General methods of tests and
analizne metode za živilske analysis for food products
proizvode
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
EN 17203
EUROPEAN STANDARD
NORME EUROPÉENNE
May 2021
EUROPÄISCHE NORM
ICS 67.050 Supersedes EN 17203:2018
English Version
Foodstuffs - Determination of citrinin in food by HPLC-
MS/MS
Produits alimentaires - Dosage de la citrinine dans les Lebensmittel - Bestimmung von Citrinin in
produits alimentaires par CLHP-SM/SM Lebensmitteln mit HPLC-MS/MS
This European Standard was approved by CEN on 19 March 2021.
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. Up-to-date lists and bibliographical references
concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN
member.
This European Standard exists 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, Turkey and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2021 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN 17203:2021 E
worldwide for CEN national Members.
Contents Page
European foreword . 3
Introduction . 4
1 Scope . 5
2 Normative references . 5
3 Terms and definitions . 5
4 Principle . 5
5 Reagents . 5
6 Apparatus and equipment . 8
7 Procedure. 9
7.1 Preparation of the test sample . 9
7.2 Extraction of citrinin . 9
7.3 Spiking procedure . 10
7.4 Preparation of sample test solutions . 10
7.5 LC-MS/MS analysis . 10
7.5.1 General . 10
7.5.2 LC-MS/MS operating conditions . 11
7.6 Identification . 12
8 Calculation . 12
8.1 General . 12
8.2 Calculation with the internal standard . 12
9 Precision . 14
9.1 General . 14
9.2 Repeatability . 14
9.3 Reproducibility . 14
10 Test report . 14
Annex A (informative) Example conditions for suitable LC-MS/MS systems . 15
Annex B (informative) Typical chromatograms . 17
Annex C (informative) Precision data . 20
Bibliography . 23
European foreword
This document (EN 17203:2021) has been prepared by Technical Committee CEN/TC 275 “Food
analysis - Horizontal methods”, the secretariat of which is held by DIN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by November 2021, and conflicting national standards shall
be withdrawn at the latest by November 2021.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document supersedes EN 17203:2018.
This document has been prepared under a standardization request given to CEN by the European
Commission and the European Free Trade Association.
The alterations to the version of 2018 are as follows:
— 5.24 The necessity to prepare calibration solutions freshly every day was deleted.
— 6.14.2 The requirement for cross contamination below 1 % was deleted.
— 7.5.1 First sentence was re-worded in better language.
— 7.5.2 The last part “when negative ion mode is used”of second para before Table 2, starting with
“When an adduct ion is used as precursor ion” was deleted as not applicable for this method.
— 7.5.2 Table 2, the last column for the 2nd qualifier was re-introduced again.
— 7.6 Para 2 line 1 was re-worded in a clearer way.
— 8.1 Para 3 was aligned with other standards of CEN/TC 275/WG 5.
According to the CEN-CENELEC Internal Regulations, the national standards organisations of the
following countries are bound to implement this European Standard: 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, Turkey and the United
Kingdom.
Introduction
The mycotoxin citrinin is a polyketide secondary metabolite produced mainly post-harvest in food and
feed by several fungi of the genera Penicillium (e.g. P. citrinum), Aspergillus (e.g. A. candidus), and
Monascus (e.g. M. purpureus). Citrinin occurs mainly in stored grains like rice, maize, wheat, barley, oats,
and rye. Citrinin can be found as a contaminant in red fermented rice with Monascus purpureus and its
formulated dietary supplements.
WARNING 1 — Suitable precaution and protection measures need to be taken when carrying out working
steps with harmful chemicals. The latest version of the hazardous substances ordinance, Regulation (EC)
No 1907/2006 [5] should be taken into account as well as appropriate National statements.
WARNING 2 — The use of this document can involve hazardous materials, operations and equipment.
This document does not purport to address all the safety problems associated with its use. It is the
responsibility of the user of this document to establish appropriate safety and health practices and
determine the applicability of regulatory limitations prior to use.
WARNING 3 — Citrinin is known to have nephrotoxic properties, damaging the proximal tubules of the
kidney [6].
1 Scope
This document describes a procedure for the determination of the citrinin content in food (cereals,
red yeast rice (RYR)), herbs and food supplements by liquid chromatography tandem mass spectrometry
(LC-MS/MS).
This method has been validated for citrinin in red yeast rice and in the formulated food supplements in
the range of 2,5 µg/kg to 3 000 µg/kg and in wheat flour in the range of 2,5 µg/kg to 100 µg/kg.
Laboratory experiences have shown that this method is also applicable to white rice, herbs such as a
powder of ginkgo biloba leaves and the formulated food supplements in the range of 2,5 µg/kg to
50 µg/kg.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
EN ISO 3696, Water for analytical laboratory use - Specification and test methods (ISO 3696)
3 Terms and definitions
No terms and definitions are listed in this document.
ISO and IEC maintain terminological 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
4 Principle
A test portion is humidified with a hydrochloric acid aqueous solution and extracted with ethyl
acetate/acetonitrile/glacial acetic acid mixture for 60 min. Magnesium sulfate and sodium chloride are
added to the extract, agitated and centrifuged in order to expel water and allow phase separation from
the mixture. An aliquot of supernatant is collected, filtered, internal standard (ISTD) solution is added
and analysed by reversed phase LC-MS/MS. Quantification is based on matching citrinin/citrinin- C
ratios and citrinin concentrations.
5 Reagents
Use only reagents of recognized analytical grade and water complying with grade 1 of EN ISO 3696,
unless otherwise specified. Commercially available solutions with equivalent properties to those listed
may also be used.
5.1 Ethyl acetate, analytical grade or higher.
5.2 Acetonitrile, LC-MS grade.
5.3 Glacial acetic acid (CH COOH), analytical grade or higher.
5.4 Glacial acetic acid (CH COOH), LC-MS grade.
5.5 Magnesium sulfate; anhydrous (MgSO ), analytical grade or higher.
5.6 Sodium chloride (NaCl), analytical grade or higher.
5.7 Hydrochloric acid solution (HCl), analytical grade or higher, volume fraction φ(HCl) = 37 %
(acidimetric).
5.8 Water (H O), deionised (Ultrapure).
5.9 Water (H O), LC-MS grade.
5.10 Methanol (MeOH), LC-MS grade.
5.11 Ammonium acetate (CH COONH ), LC-MS grade.
3 4
5.12 Extraction solution 1.
Add 10 ml of glacial acetic acid (5.3) to 990 ml of water (5.8) and mix (water + glacial acetic acid, 99+1,
v+v). Dissolve 100 g of sodium chloride (5.6) in 1 l of this mixture and add 16 ml of hydrochloric acid
solution (5.7). This solution can be used for 1 month if stored at room temperature.
5.13 Extraction solution 2.
Mix 240 ml of acetonitrile (5.2) with 750 ml of ethyl acetate (5.1) and 10 ml of glacial acetic acid (5.3).
This solution (ethyl acetate + acetonitrile + glacial acetic acid, 75+24+1, v+v+v) can be used for 1 month
if stored at room temperature.
5.14 Dilution solution.
Mix 80 ml of methanol (5.10), 18 ml of water (5.9) and 2 ml of glacial acetic acid (5.4). This solution
(methanol + water + glacial acetic acid, 80+18+2, v+v+v) can be used for 1 month if stored at room
temperature.
5.15 Ammonium acetate/glacial acetic acid in water.
Dissolve 9,5 g of ammonium acetate (5.11) in 12,5 ml of water (5.9), then add 12,5 ml of glacial acetic acid
(5.4) and mix thoroughl
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