SIST-TS CEN/TS 14537:2003

SLOVENSKI STANDARD
SIST-TS CEN/TS 14537:2003
01-november-2003
äLYLOD'RORþHYDQMHQHRKHVSHULGLQGLKLGURKDONRQD
Foodstuffs - Determination of neohesperidin-dihydrochalcon
Lebensmittel - Bestimmung von Neohesperidin-Dihydrochalcon mit
Hochleistungsflüssigkeitschromatographie (HPLC)
Denrées alimentaires - Dosage de la néohespéridine-dihydrochalcone par
chromatographie liquide de haute performance (CLHP)
Ta slovenski standard je istoveten z: CEN/TS 14537:2003
ICS:
67.050 Splošne preskusne in General methods of tests and
analizne metode za živilske analysis for food products
proizvode
SIST-TS CEN/TS 14537:2003 en
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST-TS CEN/TS 14537:2003

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SIST-TS CEN/TS 14537:2003
TECHNICAL SPECIFICATION
CEN/TS 14537
SPÉCIFICATION TECHNIQUE
TECHNISCHE SPEZIFIKATION
April 2003
ICS 67.050
English version
Foodstuffs - Determination of neohesperidin-dihydrochalcon
Lebensmittel - Bestimmung von Neohesperidin-
Dihydrochalcon mit
Hochleistungsflüssigkeitschromatographie (HPLC)
This Technical Specification (CEN/TS) was approved by CEN on 5 January 2003 for provisional application.
The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN will be requested to submit their
comments, particularly on the question whether the CEN/TS can be converted into a European Standard.
CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS available. It
is permissible to keep conflicting national standards in force (in parallel to the CEN/TS) until the final decision about the possible
conversion of the CEN/TS into an EN is reached.
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 United
Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: rue de Stassart, 36  B-1050 Brussels
© 2003 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 14537:2003 E
worldwide for CEN national Members.

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SIST-TS CEN/TS 14537:2003
CEN/TS 14537:2003 (E)
Contents
1 Scope .4
2 Normative references .4
3 Principle.4
4 Reagents.4
5 Apparatus .5
6 Procedure .6
7 Calculation.7
8 Precision.7
9 Test report .8
Annex A (informative) Precision data.9
Annex B (informative) Alternative HPLC-Systems.10
Annex C (informative) Example for a chromatogram .11
Bibliography .13
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SIST-TS CEN/TS 14537:2003
CEN/TS 14537:2003 (E)
Foreword
This document (CEN/TS 14537:2003) has been prepared by Technical Committee CEN/TC 275 “Food analysis -
Horizontal methods”, the secretariat of which is held by DIN.
According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following
countries are bound to announce this Technical Specification: 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.
Annexes A, B and C are informative.
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SIST-TS CEN/TS 14537:2003
CEN/TS 14537:2003 (E)
1 Scope
This Technical Specification specifies an HPLC-method for the determination of neohesperidin-dihydrochalcon
(NHDC) in foodstuffs.
It has been validated in a collaborative test with cherry yoghurt containing 42,7 mg/kg and on a multi vitamin drink
containing 35,6 mg/l NHDC [1].
The method has been successfully applied to a range of other foods including marzipan, bakery products, cream,
custard powder, chocolate, ice cream.
2 Normative references
This Technical Specification 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 listed hereafter. For
dated references, subsequent amendments to or revisions of any of these publications apply to this Technical
Specification only when incorporated in it by amendment or revision. For undated references the latest 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 Principle
The sample is diluted with methanol or extracted with a methanol/water – mixture and possibly filtered. NHDC is
separated by HPLC on a reversed phase, detected spectrometrically and determined by the external standard
method [1].
4 Reagents
4.1 General
During the analysis, unless otherwise stated, use only reagents of recognised analytical grade and water of at least
grade 1 according to EN ISO 3696 or use distilled water.
4.2 tetra-n-butyl ammonium hydrogen sulfate-solution (mobile Phase A) for HPLC recommended
Substance concentration c = 0,01 mol/l
4.3 Methanol (mobile Phase B)
4.4 HPLC mobile phase
For the mobile phase, tetra-n-butyl ammonium hydrogen sulfate-solution (4.2) and methanol (4.3) are applied in
appropriate ratios (gradient, e.g. as given in 6.3.1). Filter through a membrane filter (5.2) before use.
4.5 Standard substance
The NHDC standard substance shall fulfil the requirements according to EU directive 95/31/EU [2].
Commercially available NHDC can contain 4 mol of water. If using this commercially available substance, take its
water content into account. The molecular weight of dried NHDC is 612,6 g/mol.
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SIST-TS CEN/TS 14537:2003
CEN/TS 14537:2003 (E)
4.6 Standard test solutions
Dissolve an appropriate amount of NHDC with a mixture of methanol/water (50 + 50, volume fraction) and dissolve
this solution again with a mixture of methanol/water (50 + 50, volume fraction) to obtain standard solutions which
will give peaks comparable to those obtained with the sample solution.
This solution may be stored for 3 months in a refrigerator set at + 4 °C.
5 Apparatus
Use usual laboratory apparatus and, in particular, the following:
5.1 Filter device, e.g. filtration unit for vacuum, glass, consisting of glass sinter plate D2 (diameter 50 mm),
250 ml funnel and 1 l conical flask, each with ground joint.
5.2 Membrane filter for filtering the mobile phase, with pore size of 0,45 mm (or less).
NOTE Filtering of the mobile phase as well as of the sample solution through a membrane filter prior to use or injection is
supposed to increase longevity of the columns.
5.3 Ultrasonic bath
5.4 Stomacher, optional, e. g. for extraction and preparation of solid samples.
5.5 HPLC system, consisting of
 a pump;
 a sample injecting device;
 a UV detector with variable wavelength setting;
 an evaluating system.
5.6 HPLC column
Analytical reversed phase column, acid resistant, diameter 4 mm, length 250 mm, filled with particle size 5 mm, e.g.
®
1)
LiChrospher 60 RP-select B , or another acid resistant RP phase. To protect the analytical column, it is
recommended to use a pre-column of similar characteristics.
Column dimensions other than those specified in this Technical Specification may be used (e.g. of diameter
3,0 mm to 4,6 mm, length 100 mm to 250 mm). Separation parameters (flow, injection volume) have to be adapted
to guarantee equivalent results.
6 Procedure
6.1 Prepara
...