EN 17853:2023
(Main)Animal feeding stuff: Methods of sampling and analysis - Determination of intact glucosinolates in feed materials and compound feed by LC-MS/MS
Animal feeding stuff: Methods of sampling and analysis - Determination of intact glucosinolates in feed materials and compound feed by LC-MS/MS
This document specifies a method for the determination of individual intact glucosinolates in feed materials including oilseeds and oilseed products and in compound feeds by high performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (MS/MS).
The method specified in this document has been successfully validated by collaborative trial in the following matrices: rape seed, camelina seed, Brassica oleracea seeds, mixed oilseeds, rape seed flakes, compound feed for bovine, porcine and poultry.
The method is applicable for the quantitative determination of epiprogroitrin, glucoalyssin, glucoarabin, glucobrassicanapin, glucobrassicin, glucocamelinin, glucoerucin, glucoiberin, gluconapin, gluconapoleiferin, gluconasturtiin, glucoraphanin, glucoraphenin, glucotropaeolin, homoglucocamelinin, 4-hydroxyglucobrassicin, 4-methoxyglucobrassicin, neoglucobrassicin, progoitrin, sinalbin and sinigrin.
The concentration ranges tested in the collaborative trial for each individual glucosinolate and for the total glucosinolate content are summarized in Table 1.
[table not represented]
Futtermittel - Probenahme- und Untersuchungsverfahren - Bestimmung von intakten Glucosinolaten in Futtermittel-Ausgangserzeugnissen und Mischfuttermitteln mittels LC-MS/MS
Dieses Dokument legt ein Verfahren zur Bestimmung der einzelnen intakten Glucosinolate in Futtermittel-Ausgangserzeugnissen einschließlich Ölsaaten und Ölsaatenerzeugnissen sowie in Mischfuttermitteln durch Hochleistungsflüssigchromatographie (HPLC) in Verbindung mit Tandem-Massenspektrometrie (MS/MS) fest.
Das in diesem Dokument festgelegte Verfahren wurde im Rahmen eines Ringversuchs mit den folgenden Matrices erfolgreich validiert: Rapssamen, Leindottersamen, Brassica-oleracea-Samen, gemischte Ölsaaten, Rapssamenflocken, Mischfuttermittel für Rinder, Schweine und Geflügel.
Das Verfahren ist anwendbar für die quantitative Bestimmung von Epiprogroitrin, Glucoalyssin, Glucoarabin, Glucobrassicanapin, Glucobrassicin, Glucocamelinin, Glucoerucin, Glucoiberin, Gluconapin, Gluconapoleiferin, Gluconasturtiin, Glucoraphanin, Glucoraphenin, Glucotropaeolin, Homoglucocamelinin, 4-Hydroxyglucobrassicin, 4-Methoxyglucobrassicin, Neoglucobrassicin, Progoitrin, Sinalbin und Sinigrin.
Die im Ringversuch geprüften Konzentrationsbereiche für jedes einzelne Glucosinolat und für den Gesamtgehalt an Glucosinolaten sind in Tabelle 1 zusammengefasst.
[table not represented]
Alimentation animale : Méthodes d’échantillonnage et d’analyse - Dosage des glucosinolates intacts dans les matières premières pour l’alimentation animale et les aliments composés par CLHP MS/MS
Le présent document spécifie une méthode pour le dosage de différents glucosinolates intacts dans les matières premières pour l’alimentation animale incluant les graines oléagineuses et les produits oléagineux et dans les aliments composés pour animaux par chromatographie liquide haute performance (CLHP) couplée à la spectrométrie de masse en tandem (MS/MS).
La méthode spécifiée dans le présent document a été validée par un essai interlaboratoires avec les matrices suivantes : graines de colza, graines de cameline, graines de Brassica oleracea, mélange de graines oléagineuses, flocons de graines de colza, aliments composés pour bovins, porcins et volaille.
La méthode est applicable pour la détermination quantitative de l’épiprogroitrine, la glucoalyssine, la glucoarabine, la glucobrassicanapine, la glucobrassicine, la glucocamélinine, la glucoérucine, la glucoibérine, la gluconapine, la gluconapoléiférine, la gluconasturtiine, la glucoraphanine, la glucoraphénine, la glucotropaéoline, l’homoglucocamélinine, la 4-hydroxyglucobrassicine, la 4-méthoxyglucobrassicine, la néoglucobrassicine, la progoitrine, la sinalbine et la sinigrine.
Les plages de concentration soumises à essai dans le cadre de l’essai interlaboratoires pour déterminer la teneur des différents glucosinolates et des glucosinolates totaux sont présentées dans le Tableau 1.
[Tableau 1]
Krma: metode vzorčenja in analize - Ugotavljanje nepoškodovanih glukozinolatov v sestavinah krme in krmni mešanici z LC-MS/MS
Ta dokument opisuje metodo za določevanje posameznih nepoškodovanih glukozinatov v ogrščici s
tekočinsko kromatografijo visoke ločljivosti (HPLC) v kombinaciji s tandemsko masno spektrometrijo (MS/MS). Progoitrin, glukonapin, glukobrasikanapin, glukobrasicin, nasturtiin, neoglukobrasicin in 4-metoksiglukobrasicin se določajo kvantitativno. Druge glukozinate, kot so 4-hidroksiglukobrasicin, gluknapoliferin in glukoalisin, bo mogoče kvantitativno določiti šele, ko bodo analitični standardi komercialno dostopni.
Ta metoda je bila interno potrjena za semena ogrščice v območju 0,05–60 mmol/kg za posamezne glukozinate.
General Information
Standards Content (Sample)
SLOVENSKI STANDARD
01-september-2023
Krma: metode vzorčenja in analize - Ugotavljanje nepoškodovanih glukozinolatov
v sestavinah krme in krmni mešanici z LC-MS/MS
Animal feeding stuff: Methods of sampling and analysis - Determination of intact
glucosinolates in feed materials and compound feed by LC-MS/MS
Futtermittel - Probenahme- und Untersuchungsverfahren - Bestimmung von intakten
Glucosinolaten in Futtermittel-Ausgangserzeugnissen und Mischfuttermitteln mittels LC-
MS/MS
Alimentation animale : Méthodes d’échantillonnage et d’analyse - Dosage des
glucosinolates intacts dans les matières premières pour l’alimentation animale et les
aliments composés par CLHP MS/MS
Ta slovenski standard je istoveten z: EN 17853:2023
ICS:
65.120 Krmila Animal feeding stuffs
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
EN 17853
EUROPEAN STANDARD
NORME EUROPÉENNE
April 2023
EUROPÄISCHE NORM
ICS 65.120
English Version
Animal feeding stuff: Methods of sampling and analysis -
Determination of intact glucosinolates in feed materials
and compound feed by LC-MS/MS
Alimentation animale : Méthodes d'échantillonnage et Futtermittel - Probenahme- und
d'analyse - Dosage des glucosinolates intacts dans les Untersuchungsverfahren - Bestimmung von intakten
matières premières pour l'alimentation animale et les Glucosinolaten in Futtermittel-Ausgangserzeugnissen
aliments composés par CLHP MS/MS und Mischfuttermitteln mittels LC-MS/MS
This European Standard was approved by CEN on 3 March 2023.
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, Türkiye 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
© 2023 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN 17853:2023 E
worldwide for CEN national Members.
Contents Page
European foreword . 4
Introduction . 5
1 Scope . 6
2 Normative references . 7
3 Terms and definitions . 7
4 Principle . 7
5 Reagents . 7
5.1 Analytical standards . 7
5.2 Chemicals . 8
5.3 Standard solutions . 8
5.4 Reagents . 11
5.5 Quality control material . 11
6 Apparatus . 12
7 Procedure . 13
7.1 Sample pre-treatment . 13
7.2 Test portion . 13
7.2.1 Oilseeds and oilseed products . 13
7.2.2 Compound feed . 13
7.3 Extraction . 13
7.3.1 Oilseeds and oilseed products . 13
7.3.2 Compound feed . 13
7.3.3 Recovery sample for compound feed . 14
7.3.4 Preparation of calibration standards in blank feed extract . 14
8 LC-MS/MS analysis . 15
8.1 General. 15
8.2 Analysis sequence . 16
8.2.1 Oilseeds and oilseed products . 16
8.2.2 Compound feed . 16
9 Evaluation of results . 17
9.1 Identification . 17
9.2 Quantification . 17
9.2.1 General. 17
9.2.2 Linearity of the calibration curve . 17
9.2.3 Calculation of the glucosinolate concentration in oilseed or oilseed product . 17
9.2.4 Calculation of the glucosinolate concentration in compound feed. 18
9.3 Expression of results . 19
10 Precision . 19
10.1 General. 19
10.2 Repeatability . 19
10.3 Reproducibility . 19
11 Test report . 20
Annex A (informative) Precision data . 21
Annex B (informative) Example of LC-MS/MS conditions . 45
B.1 General . 45
B.2 Chromatographic conditions . 45
B.3 MS conditions . 46
Annex C (informative) Examples of chromatograms . 48
Annex D (informative) Glucosinolate standards from commercial sources . 50
Bibliography . 52
European foreword
This document (EN 17853:2023) has been prepared by Technical Committee CEN/TC 327 “Animal
feeding stuffs: Methods of sampling and analysis”, the secretariat of which is held by NEN.
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 October 2023, and conflicting national standards shall
be withdrawn at the latest by October 2023.
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 has been prepared under a Standardization Request given to CEN by the European
Commission and the European Free Trade Association.
Any feedback and questions on this document should be directed to the users’ national standards body.
A complete listing of these bodies can be found on the CEN website.
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, Türkiye and the United
Kingdom.
Introduction
Glucosinolates are a group of plant produced secondary metabolites predominantly found in the family
Brassicaceae (mustards and cabbages) ([1], [2], [3]). Many common vegetables such as broccoli, Brussels
sprouts, cabbage and cauliflower, belong to this plant family. At the same time species from the genera
Brassica, Camelina, Crambe, Rhaphanus and Sinapis are agricultural crops used for the production of plant
oils, such as rapeseed oils. The press cake is used as animal feed material. Glucosinolates are considered
undesirable substances in feed [4]. Glucosinolates in rapeseed and rapeseed products can also be
measured after enzymatic desulfation by high-performance liquid chromatography (HPLC) coupled with
UV detection. This method is described in standard EN ISO 9167 [5].
WARNING — This protocol does not purport to address all the safety problems associated with its use.
It is the responsibility of the user of this protocol to establish appropriate safety and health protection
measures and to ensure that regulatory and legal requirements are complied with.
1 Scope
This document specifies a method for the determination of individual intact glucosinolates in feed
materials including oilseeds and oilseed products and in compound feeds by high performance liquid
chromatography (HPLC) coupled with tandem mass spectrometry (MS/MS).
The method specified in this document has been successfully validated by collaborative trial in the
following matrices: rape seed, camelina seed, Brassica oleracea seeds, mixed oilseeds, rape seed flakes,
compound feed for bovine, porcine and poultry.
The method is applicable for the quantitative determination of epiprogroitrin, glucoalyssin, glucoarabin,
glucobrassicanapin, glucobrassicin, glucocamelinin, glucoerucin, glucoiberin, gluconapin,
gluconapoleiferin, gluconasturtiin, glucoraphanin, glucoraphenin, glucotropaeolin, homoglucocamelinin,
4-hydroxyglucobrassicin, 4-methoxyglucobrassicin, neoglucobrassicin, progoitrin, sinalbin and sinigrin.
The concentration ranges tested in the collaborative trial for each individual glucosinolate and for the
total glucosinolate content are summarized in Table 1.
Table 1 — Summary of glucosinolate concentration ranges tested in the collaborative trial
Glucosinolate Number of samples Tested concentration range
with acceptable
mmol/kg
results
Min
...
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