SIST EN 12014-4:2005

Foodstuffs - Determination of nitrate and/or nitrite content - Part 4: Ion-exchange chromatographic (IC) method for the determination of nitrate and nitrite content of meat products

This European Prestandard specifies an ion-exchange chromatographic method for the determination of the nitrate and nitrite contents of meat products having a nitrate content of 50 mg/kg to 300 mg/kg as nitrate ions and a nitrite content of approximately 40 mg/kg as nitrite ion. Note: Validation data obtained from interlaboratory studies show that this method may also be applied to the determination of nitrate in vegetables and baby food, see (1), (2). Furthermore, the method may be applied for the determination of nitrite in meat products having a nitrite content of greater than 40 mg/kg.

Lebensmittel - Bestimmung des Nitrat- und/oder Nitritgehaltes - Teil 4: Ionenchromatographisches Verfahren (IC) für die Bestimmung des Nitrat- und Nitritgehaltes in Fleischerzeugnissen

Diese Europäische Norm legt ein Ionenaustausch-chromatographisches Verfahren zur Bestimmung des Nitrit-
und Nitratgehaltes von Fleischerzeugnissen fest und wurde an verschiedenen Fleischerzeugnissen mit einem
Nitratgehalt von 50 mg/kg bis 300 mg/kg als Nitrat-Ion und einem Nitritgehalt von etwa 40 mg/kg als Nitrit-Ion
validiert.
ANMERKUNG Zuverlässigkeitsdaten aus Ringversuchen zeigen, dass dieses Verfahren auch für die Bestimmung von
Nitrat in Gemüse und Säuglingsnahrung angewendet werden kann, siehe [1], [2]. Des Weiteren darf das Verfahren für die
Bestimmung des Nitritgehaltes in Fleischerzeugnissen mit einem Nitritgehalt von mehr als 40 mg/kg angewendet werden.

Produits alimentaires - Détermination de la teneur en nitrates et/ou en nitrites - Partie 4 : Détermination par chromatographie ionique (CI) de la teneur en nitrates et en nitrites dans les produits carnés

Cette Norme européenne spécifie une méthode de détermination par chromatographie ionique de la teneur en
nitrates et en nitrites dans les produits carnés, et a été validée pour différents produits carnés pour une teneur
en nitrates comprise entre 50 mg/kg et 300 mg/kg, exprimée en ion nitrate et une teneur en nitrites d�environ
40 mg/kg, exprimée en ion nitrite.
NOTE Les données de validation issues des essais interlaboratoires montrent que cette méthode peut s'appliquer
également pour la détermination des nitrates dans les légumes et les aliments pour bébés [1], [2]. De plus, cette méthode
peut être utilisée pour la détermination des nitrites dans les produits carnés ayant une teneur en nitrites supérieure à
40 mg/kg.

Živila – Določevanje nitratov in/ali nitritov - 4. del: Metoda ionske izmenjalne kromatografije za določevanje nitratov in nitritov v mesnih proizvodih

General Information

• Status: Published
• Publication Date: 31-Aug-2005
• ICS: 67.050 - General methods of tests and analysis for food products
• ICS: 67.120.10 - Meat and meat products
• Technical Committee: KŽP - Agricultural food products
• Current Stage: 6060 - National Implementation/Publication (Adopted Project)
• Start Date: 01-Sep-2005
• Due Date: 01-Sep-2005
• Completion Date: 01-Sep-2005
• Ref Project: EN 12014-4:2005 - Foodstuffs - Determination of nitrate and/or nitrite content - Part 4: Ion-exchange chromatographic (IC) method for the determination of nitrate and nitrite content of meat products

Relations

• Replaces: SIST ENV 12014-4:1999 - Foodstuffs - Determination of nitrate and/or nitrite content - Part 4: Ion-exchange chromatographic (IC) method for the determination of nitrate and nitrite content of meat products
• Effective Date: 01-Sep-2005

Overview

EN 12014-4:2005 (CEN) specifies an ion‑exchange chromatographic (IC) method for the determination of nitrate and nitrite content in meat products. Validated by interlaboratory studies, the method covers nitrate concentrations of about 50–300 mg/kg (as NO3–) and nitrite concentrations of approximately 40 mg/kg (as NO2–). The standard also notes that the method may be applied to nitrate in vegetables and baby food and to nitrite in meat products with >40 mg/kg.

Keywords: EN 12014-4, nitrate nitrite determination, ion-exchange chromatography, meat products, food analysis, CEN.

Key Topics and Requirements

• Principle: Hot‑water extraction of sample, acetonitrile treatment to remove interferences, separation by ion‑exchange chromatography and UV detection at 205 nm.
• Sample prep: Homogenize sample, extract with 50 ml water (50–60 °C) per 10 g test portion, add acetonitrile, dilute to volume, clarify by fluted filter and membrane filters (0.45 µm and, if necessary, 0.22 µm).
• Chromatography:
  • Anion‑exchange analytical column (polymethacrylate resin with quaternary ammonium functional group; 4.6 × 150 mm, 10 µm) with precolumn.
  • Mobile phase based on lithium borate gluconate buffer + acetonitrile, adjusted to pH 6.5 ± 0.1 and filtered (≈0.22 µm).
  • Typical conditions: flow 1.0 ml/min, injection ≥40 µl, UV detection at 205 nm.
• Calibration & quantification: Use prepared standard solutions from a stock nitrate/nitrite solution; plot calibration graph and check linearity; calculate mg/kg using provided equations and dilution factors.
• Quality & precision: Precision established in Annex B (interlaboratory data). The standard gives repeatability (r) and reproducibility (R) limits for matrices such as corned beef, sausage and salami (examples: nitrate r ~6 mg/kg at ~61 mg/kg; reproducibility R up to ~37 mg/kg for some matrices).
• Reporting: Test report must identify sample, method reference, results, sampling dates, and any deviations or observations.

Applications and Users

This standard is intended for:

• Food testing laboratories performing regulatory or QA analyses for nitrate and nitrite in meat products.
• Meat processors and manufacturers implementing quality control and compliance checks.
• Regulatory authorities and inspectors assessing food safety, labeling and legal limits.
• Research groups validating analytical procedures for food contaminants.

Practical uses include routine monitoring, compliance testing, method validation and comparative interlaboratory studies.

Related Standards

• EN 12014 series (Parts 1–7) - other methods for nitrate/nitrite in vegetables, baby food and alternative detection techniques.
• EN ISO 3696 - water quality for analytical laboratory use (referenced for reagent water quality).

For laboratories implementing EN 12014-4, follow the method steps, validation limits and reporting requirements in the standard to ensure reliable nitrate/nitrite testing by ion‑exchange chromatography.

Standards Content (Sample)

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.YRGLKLebensmittel - Bestimmung des Nitrat- und/oder Nitritgehaltes - Teil 4: Ionenchromatographisches Verfahren (IC) für die Bestimmung des Nitrat- und Nitritgehaltes in FleischerzeugnissenProduits alimentaires - Détermination de la teneur en nitrates et/ou en nitrites - Partie 4 : Détermination par chromatographie ionique (CI) de la teneur en nitrates et en nitrites dans les produits carnésFoodstuffs - Determination of nitrate and/or nitrite content - Part 4: Ion-exchange chromatographic (IC) method for the determination of nitrate and nitrite content of meat products67.120.10Meso in mesni proizvodiMeat and meat products67.050Splošne preskusne in analizne metode za živilske proizvodeGeneral methods of tests and analysis for food productsICS:Ta slovenski standard je istoveten z:EN 12014-4:2005SIST EN 12014-4:2005en01-september-2005SIST EN 12014-4:2005SLOVENSKI
STANDARDSIST ENV 12014-4:19991DGRPHãþD

EUROPEAN STANDARD NORME EUROPÉENNE EUROPÄISCHE NORM
EN 12014-4
May 2005 ICS 67.120.10 Supersedes ENV 12014-4:1998 English version
Foodstuffs - Determination of nitrate and/or nitrite content - Part 4: Ion-exchange chromatographic (IC) method for the determination of nitrate and nitrite content of meat products
Produits alimentaires - Détermination de la teneur en nitrates et/ou en nitrites - Partie 4: Détermination par chromatographie ionique (CI) de la teneur en nitrates et en nitrites dans les produits à base de viande
Lebensmittel - Bestimmung des Nitrat- und/oder Nitritgehaltes - Teil 4: Ionenchromatographisches Verfahren (IC) für die Bestimmung des Nitrat- und Nitritgehaltes in Fleischerzeugnissen This European Standard was approved by CEN on 1 April 2005.
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 Central Secretariat 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 Central Secretariat has the same status as the official versions.
CEN members are the national standards bodies of Austria, Belgium, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Slovakia, Slovenia, Spain, Sweden, Switzerland and United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION COMITÉ EUROPÉEN DE NORMALISATION EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: rue de Stassart, 36
B-1050 Brussels © 2005 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 12014-4:2005: ESIST EN 12014-4:2005

Example for an IC chromatogram of corned beef
(Sample from the interlaboratory test).10 Annex B (informative)
Precision data.11 Annex C (informative)
Further applicability.13 Bibliography.16
Dilute 15 ml of hydrochloric acid of 20 (HCl) = 1,18 g/ml in a 100 ml volumetric flask to the mark with water and mix. 4.7 Hydrochloric acid, c (HCl) ≈ 0,1 mol/l Dilute 5 ml of hydrochloric acid (4.6) in a 100 ml volumetric flask to the mark with water and mix.
1) c is the substance concentration SIST EN 12014-4:2005

NOTE Gluconic acid sometimes has a dark brown colour. In this case it is recommended to clarify it until a slight yellow colour is obtained. Generally two cartridges activated with methanol are necessary. 4.11 Lithium borate gluconate buffer solution To 500 ml of water in a 1000 ml volumetric flask add 34,00 g of boric acid (4.5) and 19,6 ml of gluconic acid solution (4.10). Dissolve completely in the solution 11,00 g of anhydrous lithium hydroxide or 19,26 g of lithium hydroxide monohydrate (4.4). Add 125 ml of glycerol (4.3), dilute to the mark with water and mix well. This solution is stable for 6 months if stored in a refrigerator at approximately 4 °C. 4.12 Mobile phase To 500 ml of water in a 1000 ml volumetric flask, add 17 ml of buffer solution (4.11) and 125 ml of acetonitrile (4.2). Dilute to the mark with water. Mix well. Adjust the pH to 6,5 ± 0,1 by adding hydrochloric acid (4.6 then 4.7). Filter through a membrane filter of pore size of approximately 0,22 µm (5.6). This solution is stable for not longer than one week and the pH value shall be checked every day before use. If the pH value falls outside the stated range, prepare a new solution.
2) This solution is commercially available. SIST EN 12014-4:2005

5.3 Magnetic stirrer (optional) 5.4 Fluted filter paper (nitrate/nitrite free), care is needed in the choice of filter paper as some can absorb nitrate. 5.5 Membrane filter, for aqueous and acetonitrile solutions, with a pore size of approximately 0,45 µm. 5.6 Membrane filter, for aqueous and acetonitrile solutions, with a pore size of approximately 0,22 µm. 5.7 Filter holder for membrane filter, with suitable syringe. 5.8 pH meter
5.9 Solid phase extraction column RP C18, with suitable syringe (optional). 5.10 IC apparatus, comprising the following 5.10.1 Isocratic high performance liquid chromatography unit, with a UV detector, capable of measuring at a wavelength of 205 nm and an evaluation unit (e.g an integrator with plotter). 5.10.2 Analytical separating column, Anion exchanger, 4,6 mm × 150 mm, packing material; polymethacrylate resin with a quarternary ammonium functional group, particle size of 10 µm, capacity (30 ± 3) µ eq/ml with a precolumn, e.g. 20 mm, having the same packing to protect the analytical column. 6 Procedure
6.1 Sample preparation Homogenize the laboratory sample with the appropriate equipment (5.2). Take care that the temperature of the sample material does not rise above 25 °C. If a mincer is used, pass the sample at least twice through the equipment. Weigh, to the nearest 10 mg, 10 g of the homogenized sample into e.g. a 150 ml wide neck conical flask (test portion).
6.2 Extraction and clarification
The analysis should be performed in a single working day. Add 50 ml of water at a temperature of 50 °C to 60 °C to the test portion in the wide neck flask. Mix thoroughly with the homogenizer (5.2). Rinse the homogenizer with water and add the washings to the flask. Quantitatively transfer this slurry into a 200 ml volumetric flask by rinsing the wide neck flask with water.
...