SIST EN ISO 19343:2017

Microbiology of the food chain - Detection and quantification of histamine in fish and fishery products - HPLC method (ISO 19343:2017)

ISO 19343:2017 specifies a high performance liquid chromatography (HPLC) method to analyse histamine in fish and fishery products (fish sauces, fish maturated by enzyme in brine, etc.) intended for human consumption.

Mikrobiologie der Nahrungskette - Nachweis und Bestimmung von Histamin in Fisch und Fischereierzeugnissen - HPLC-Methode (ISO 19343:2017)

Dieses Dokument legt eine Hochleistungsflüssigkeitschromatografie Methode (HPLC) fest, um Histamin in Fisch und Fischereierzeugnissen (Fischsauce, in Salzwasser eingelegter, durch Enzyme gereifter Fisch usw.) für die menschliche Ernährung zu untersuchen.

Microbiologie de la chaîne alimentaire - Détection et quantification de l'histamine par méthode HPLC (ISO 19343:2017)

L'ISO 19343:2017 spécifie une méthode de chromatographie en phase liquide à haute performance (CLHP) pour analyser l'histamine dans le poisson et les produits de la pêche (sauces au poisson, poissons ayant subi un traitement de maturation enzymatique dans la saumure, etc.) destinés à la consommation humaine.

Mikrobiologija v prehranski verigi - Odkrivanje prisotnosti in kvantifikacija histamina v ribah in ribjih proizvodih - Metoda HPLC (ISO 19343:2017)

Ta standard opisuje način odkrivanja prisotnosti in kvantifikacijo histamina.

General Information

• Status: Published
• Public Enquiry End Date: 19-Jun-2016
• Publication Date: 15-Aug-2017
• ICS: 07.100.30 - Food microbiology
• ICS: 67.120.30 - Fish and fishery products
• Technical Committee: KŽP - Agricultural food products
• Current Stage: 6060 - National Implementation/Publication (Adopted Project)
• Start Date: 01-Aug-2017
• Due Date: 06-Oct-2017
• Completion Date: 16-Aug-2017
• Directive: 882/2004 - Regulation (EC) No 882/2004 of the European Parliament and of the Council of 29 April 2004 onofficial controls performed to ensure the verification of compliance with feed and food law, animal health and animal welfare rules
• Mandate: M/381 - Mandate for standardisation addressed to CEN in the field of Methods of analysis of foodstuffs concerning Food Hygiene
• Ref Project: EN ISO 19343:2017 - Microbiology of the food chain - Detection and quantification of histamine in fish and fishery products - HPLC method (ISO 19343:2017)

Overview

EN ISO 19343:2017 (ISO 19343:2017) is an international/European standard for the detection and quantification of histamine in fish and fishery products using a validated HPLC method. It applies to fresh and processed seafood (including fish sauces and products matured in brine) intended for human consumption and supports regulatory food safety control, notably in relation to scombroid (histamine) poisoning.

Key topics and technical requirements

• Scope and principle: Specifies a high performance liquid chromatography (HPLC) procedure that separates histamine from other biogenic amines. Samples are extracted with perchloric acid, pre-column derivatized with dansyl chloride, and analyzed by HPLC with UV detection.
• Reagents and materials: Lists required solvents and reagents (e.g., acetone, acetonitrile, toluene, perchloric acid, dansyl chloride) and recommends laboratory-grade water complying with ISO 3696.
• Internal standard and calibration: Uses an internal standard (recommended: 1,7-diaminoheptane) and calibration curves for histamine quantification based on peak-area ratios.
• Apparatus and LC conditions: Defines typical equipment (grinder/homogenizer, refrigerated centrifuge, nitrogen evaporator, 0.2 µm filters) and an LC system (column oven, UV detector at λ = 254 nm). An example column is Kromasil® C18 5 µm, 25 cm × 4.6 mm.
• Procedure steps: Covers sample preparation, extraction, derivatization, purification, LC separation, and calculation methods.
• Performance and precision: Includes annexes with recommended HPLC separation and method performance characteristics, reporting repeatability, reproducibility and interlaboratory study results (method validation).

Practical applications

• Food testing laboratories: Routine monitoring of histamine levels in raw, processed and fermented fish products.
• Quality control for seafood producers and processors: Ensures products meet safety criteria and limits set by regulatory authorities.
• Regulatory and public health agencies: Enforces compliance with microbiological food safety regulations (e.g., EU Regulation 2073/2005) and investigates scombroid poisoning incidents.
• Research and method development: Reference method for comparative studies on biogenic amines in seafood.

Who should use this standard

• Analytical chemists and microbiologists in food testing labs
• Seafood quality assurance teams
• National reference laboratories and regulatory bodies
• Accreditation bodies assessing laboratory competence in histamine testing

Related standards

• ISO 3696 - Water for analytical laboratory use (referenced normative document)
• EN ISO 19343:2017 is the CEN-endorsed version of ISO 19343:2017 for use across Europe

This standard is essential for consistent, reproducible histamine detection and quantification in the seafood sector and supports food safety risk management for histamine-related poisoning.

Standards Content (Sample)

SLOVENSKI STANDARD
01-september-2017
Mikrobiologija v prehranski verigi - Odkrivanje prisotnosti in kvantifikacija
histamina v ribah in ribjih proizvodih - Metoda HPLC (ISO 19343:2017)
Microbiology of the food chain - Detection and quantification of histamine in fish and
fishery products - HPLC method (ISO 19343:2017)
Mikrobiologie der Nahrungskette - Nachweis und Bestimmung von Histamin in Fisch und
Fischereierzeugnissen - HPLC-Methode (ISO 19343:2017)
Microbiologie de la chaîne alimentaire - Détection et quantification de l'histamine par
méthode HPLC (ISO 19343:2017)
Ta slovenski standard je istoveten z: EN ISO 19343:2017
ICS:
07.100.30 Mikrobiologija živil Food microbiology
67.120.30 Ribe in ribji proizvodi Fish and fishery products
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EN ISO 19343
EUROPEAN STANDARD
NORME EUROPÉENNE
July 2017
EUROPÄISCHE NORM
ICS 07.100.30
English Version
Microbiology of the food chain - Detection and
quantification of histamine in fish and fishery products -
HPLC method (ISO 19343:2017)
Microbiologie de la chaîne alimentaire - Détection et Mikrobiologie der Lebensmittelkette - Nachweis und
quantification de l'histamine dans le poisson et les Bestimmung von Histamin in Fisch und
produits de la pêche - Méthode par CLHP (ISO Fischereierzeugnissen - HPLC-Methode (ISO
19343:2017) 19343:2017)
This European Standard was approved by CEN on 8 June 2017.

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, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania,
Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, 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: Avenue Marnix 17, B-1000 Brussels
© 2017 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 19343:2017 E
worldwide for CEN national Members.

Contents Page
European foreword . 3

European foreword
This document (EN ISO 19343:2017) has been prepared by Technical Committee ISO/TC 34 “Food
products” in collaboration with 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 January 2018 and conflicting national standards shall
be withdrawn at the latest by January 2018.
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 mandate given to CEN by the European Commission and the
European Free Trade Association.
According to the CEN-CENELEC Internal Regulations, the national standards organizations of the
following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria,
Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia,
France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta,
Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland,
Turkey and the United Kingdom.
Endorsement notice
The text of ISO 19343:2017 has been approved by CEN as EN ISO 19343:2017 without any modification.
INTERNATIONAL ISO
STANDARD 19343
First edition
2017-06
Microbiology of the food chain —
Detection and quantification of
histamine in fish and fishery products
— HPLC method
Microbiologie de la chaîne alimentaire — Détection et quantification de
l’histamine dans le poisson et les produits de la pêche — Méthode CLHP
Reference number
ISO 19343:2017(E)
©
ISO 2017
ISO 19343:2017(E)
© ISO 2017, Published in Switzerland
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized otherwise in any form
or by any means, electronic or mechanical, including photocopying, or posting on the internet or an intranet, without prior
written permission. Permission can be requested from either ISO at the address below or ISO’s member body in the country of
the requester.
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ii © ISO 2017 – All rights reserved

ISO 19343:2017(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 1
5 Reagents and materials . 1
6 Apparatus . 2
7 Procedure. 3
7.1 Sample preparation . 3
7.2 Extraction . 3
7.3 Derivatization . 3
7.4 Purification . 3
7.5 LC condition . 4
7.6 Range of standard sample . 4
8 Calculation . 4
8.1 Calibration line (curve) . 4
8.2 Histamine quantification . 5
9 Precision . 5
9.1 Interlaboratory study . 5
9.2 Repeatability limit . 5
9.3 Reproducibility limit . 5
Annex A (informative) Recommendations for HPLC separation . 7
Annex B (informative) Performance characteristics of the method . 9
Bibliography .14
ISO 19343:2017(E)
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 ISO technical committees. Each member body interested in a subject for which a technical
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described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the
different types of ISO documents 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).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
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URL: w w w . i s o .org/ iso/ foreword .html.
This document was prepared by the European Committee for Standardization (CEN) Technical
Committee CEN/TC 275, Food analysis — Horizontal methods, in collaboration with ISO Technical
Committee TC 34, Food products, Subcommittee SC 9, Microbiology, in accordance with the agreement
on technical cooperation between ISO and CEN (Vienna Agreement).
iv © ISO 2017 – All rights reserved

ISO 19343:2017(E)
Introduction
Histamine is a causative agent of scombroid poisoning or histamine fish poisoning. Histamine can
be present mainly in Scombridae (tuna, mackerel) and Clupeidae (herring, sardine), species which
contain a high level of free histidine. Histamine is formed through the decarboxylation of histidine by
microbiological histidine decarboxylase.
Histamine [2-(1H-imidazol-5-yl)ethanamine] is defined as a biologically active low molecular weight
basic nitrogenous molecule. The consumption of food containing significant concentration of histamine
can cause symptoms similar to those associated to food allergies.
This document was developed in response to the need to standardize a method for histamine detection
[1]
and quantification in fish and fishery products, in particular for European Regulation 2073/2005 on
microbiological criteria for foodstuffs.
INTERNATIONAL STANDARD ISO 19343:2017(E)
Microbiology of the food chain — Detection and
quantification of histamine in fish and fishery products —
HPLC method
WARNING — Persons using this document should be familiar with normal laboratory practice.
This document does not purport to address all of the safety problems, if any, associated with its
use. It is the responsibility of the user to establish appropriate safety and health practices.
1 Scope
This document specifies a high performance liquid chromatography (HPLC) method to analyse
histamine in fish and fishery products (fish sauces, fish maturated by enzyme in brine, etc.) intended
for human consumption.
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.
ISO 3696, Water for analytical laboratory use — Specification and test methods
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 http:// www .electropedia .org/
— ISO Online browsing platform: available at http:// www .iso .org/ obp
4 Principle
This method enables the separation of histamine among biogenic amines in fish and fishery products.
The sample is extracted by mixing with perchloric acid. Precolumn derivatization is performed using
dansyl chloride. The biogenic amines and the components in the solution are separated by HPLC with
an appropriate column, using UV detection. Histamine mass concentration is calculated from the peak
area ratio of histamine and internal standard with a calibration curve.
5 Reagents and materials
Use only reagents of recognized analytical grade and water complying with grade 1 of ISO 3696, unless
otherwise specified. Solvents shall be of quality for HPLC analysis, unless otherwise specified.
5.1 Acetone, CH COCH .
3 3
5.2 Acetonitrile, CH CN.
5.3 Toluene, C H .
7 8
ISO 19343:2017(E)
5.4 Water, HPLC grade.
5.5 Water, distilled or equivalent.
5.6 Nitrogen, gas.
5.7 Perchloric acid, c(HClO ) = 0,2 mol/l (recommended).
Dilute 19,5 ml of HClO 65 % or 17,2 ml of HClO 70 % to 1 000 ml of water (5.5). The solution is stable
4 4
for six months if stored at room temperature (15°C to 25°C).
5.8 Saturated sodium carbonate solution, Na CO .
2 3
Dissolve 110 g of sodium carbonate in about 150 ml of water (5.5). The solution is in excess and stable
for three months if stored at 5 °C ± 3°C.
5.9 Dansyl chloride solution, ρ(C H ClNO S) = 7,5 mg/ml.
12 12 2
Dissolve 0,375 g of dansyl chloride in 50 ml of acetone (5.1). The solution is stable for three weeks if
stored in the dark at a temperature lower than < −18 °C.
5.10 L-proline solution, ρ(C H NO ) = 100 mg
...