SIST EN 14176:2004

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Lebensmittel - Bestimmung von Domoinsäure in Muscheln mit HPLCProduits alimentaires - Dosage de l'acide domoique dans les moules par CLHPFoodstuffs - Determination of domoic acid in mussels by HPLC67.120.30Ribe in ribji proizvodiFish and fishery productsICS:Ta slovenski standard je istoveten z:EN 14176:2003SIST EN 14176:2004en01-maj-2004SIST EN 14176:2004SLOVENSKI
STANDARD



SIST EN 14176:2004



EUROPEAN STANDARDNORME EUROPÉENNEEUROPÄISCHE NORMEN 14176December 2003ICS 67.120.30English versionFoodstuffs - Determination of domoic acid in mussels by HPLCProduits alimentaires - Dosage de l'acide domoïque dansles moules par CLPHLebensmittel - Bestimmung von Domoinsäure in Muschelnmit HPLCThis European Standard was approved by CEN on 1 October 2003.CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this EuropeanStandard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such nationalstandards may be obtained on application to the 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 translationunder the responsibility of a CEN member into its own language and notified to the Management Centre has the same status as the officialversions.CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece,Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and UnitedKingdom.EUROPEAN COMMITTEE FOR STANDARDIZATIONCOMITÉ EUROPÉEN DE NORMALISATIONEUROPÄISCHES KOMITEE FÜR NORMUNGManagement Centre: rue de Stassart, 36
B-1050 Brussels© 2003 CENAll rights of exploitation in any form and by any means reservedworldwide for CEN national Members.Ref. No. EN 14176:2003 ESIST EN 14176:2004



EN 14176:2003 (E)2ContentsPageForeword.31Scope.42Normative references.43Principle.44Reagents.45Apparatus.56Sampling.57Procedure.58Calculation.69Precision.710Test report.7Annex A (informative)
Precision data.9Bibliography.10SIST EN 14176:2004



EN 14176:2003 (E)3ForewordThis document (EN 14176:2003) 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 identicaltext or by endorsement, at the latest by June 2004, and conflicting national standards shall be withdrawn atthe latest by June 2004.Annex A in this document is informative.WARNING —The use of this standard can involve hazardous materials, operations and equipment.This standard does not purport to address all the safety problems associated with its use. It is theresponsibility of the user of this standard to establish appropriate safety and health practices anddetermine the applicability of regulatory limitations prior to use.According to the CEN/CENELEC Internal Regulations, the national standards organizations of the followingcountries are bound to implement this European Standard: Austria, Belgium, Czech Republic, Denmark,Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands,Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and the United Kingdom.SIST EN 14176:2004



EN 14176:2003 (E)41 ScopeThis European Standard specifies a method for the determination of domoic acid in mussels using highperformance liquid chromatography (HPLC).The method has been successfully validated in an interlaboratory study according to AOAC guidelines onmussels containing 14,1 mg/g (spiked sample) to 186 mg/g (naturally contaminated sample) domoic acid.Laboratory experiences show that the method is also applicable on the common cockle (Cerastoderma edule),the peppery furrow shell (Scrobicularia plana), clams (Venerupis pullastra, Ruditapes decussata), oyster(Crassostrea japonica) and razor clams (Ensis spp., Solen spp) [1].2 Normative referencesThis European Standard incorporates by dated or undated reference, provisions from other publications.These normative references are cited at the appropriate places in the text and the publications are listedhereafter. For dated references, subsequent amendments to or revisions of any of these publications apply tothis European Standard only when incorporated in it by amendment or revision. For undated references thelatest edition of the publication referred to applies (including amendments).EN ISO 3696,Water for analytical laboratory use - Specification and test methods (ISO 3696:1987).3 PrincipleDomoic acid is extracted by boiling homogenized mussel tissue with a solution of hydrochloric acid. Themixture is cooled, diluted to a known volume and then centrifuged. The supernate is diluted, filtered, andanalysed by reverse-phase HPLC with ultraviolet detection.4 Reagents4.1 GeneralDuring the analysis, unless otherwise stated, use only reagents of recognized analytical grade and onlydistilled water or water of grade 1 as defined in EN ISO 3696. Solvent shall be of quality for HPLC analysis.4.2 Hydrochloric acid solution, substance concentration c(HCl) = 0,1 mol/l4.3 Phosphoric acid solution, mass fraction w(H3PO4) = 8,5 %4.4 Acetonitrile4.5 Mobile phaseMix 2 volume parts of phosphoric acid (4.3) with 873 volume parts of water. Check to ensure that pH » 2,5.Add 125 ml acetonitrile (4.4), mix and degas this solution before use. Perform a preliminary analysis of thedomoic acid standard solution (4.6) and adjust the acetonitrile concentration as necessary to give a domoicacid retention time of approximately 8 min under method conditions.4.6 Domoic acid standard solutionPrepare a standard solution at a precisely known concentration in the region of 1 mg/ml. If necessary dilutewith water. Refrigerate the solution when not in use. Warm the solution to room temperature before use.SIST EN 14176:2004



EN 14176:2003 (E)55 Apparatus5.1 GeneralUsual laboratory equipment and, in particular, the following:5.2 Blender or household-type grinder5.3 Membrane filter, of pore size of approximately 0,45 mm, attached to a 5 ml glass or disposable plasticsyringe.5.4 Centrifuge, high speed, with timer, capable of 700 g using 50 ml to 100 ml centrifuge tubes.5.5 Centrifuge tubes, of a capacity from 50 ml to 100 ml.5.6 HPLC apparatus, comprising the following:5.6.1 High performance liquid chromatograph, pump able to deliver up to 1,5 ml/min constant flow rate,equipped with an injection system with a volume of e.g.
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