EN ISO 23275-1:2008

Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain chocolate - Part 1: Determination of the presence of cocoa butter equivalents (ISO 23275-1:2006)

ISO 23275-1:2006 specifies a procedure for the detection of cocoa butter equivalents (CBEs) in cocoa butter and plain chocolate by high-resolution capillary gas liquid chromatography of triacylglycerols and subsequent data evaluation by regression analysis.
The method is applicable for the detection of 2 % CBE admixture to cocoa butter, corresponding to about 0,6 % CBE in chocolate (i.e. the assumed fat content of chocolate is 30 %).

Tierische und pflanzliche Fette und Öle - Kakaobutter-Äquivalente in Kakaobutter und Zartbitterschokolade - Teil 1: Bestimmung der Präsenz von Kakaobutter-Äquivalenten (ISO 23275-1:2006)

Dieser Teil von ISO 23275 legt ein Verfahren zum Nachweis von Kakaobutter-Äquivalenten (CBE) in Kakaobutter
und Zartbitterschokolade durch hochauflösende Kapillar-Gaschromatographie (HR-GC) von Triacylglycerinen
und anschließende Datenauswertung durch Regressionsanalyse fest.
Das Verfahren ist anwendbar für den Nachweis eines Zusatzes von 2 % CBE zur Kakaobutter, was etwa
0,6 % CBE in Schokolade entspricht (d. h. der angenommene Fettgehalt der Schokolade beträgt 30 %).

Corps gras d'origines animale et végétale - Équivalents au beurre de cacao dans le beurre de cacao et dans le chocolat de ménage - Partie 1: Détermination de la présence d'équivalents au beurre de cacao (ISO 23275-1:2006)

L'ISO 23275-1:2006 spécifie un mode opératoire permettant la détection des équivalents au beurre de cacao (EBC) dans le beurre de cacao (BC) et le chocolat de ménage par analyse chromatographique en phase gazeuse/liquide capillaire haute résolution (HR-GC) des triacylglycérols, et l'évaluation ultérieure des données par analyse de régression.
La méthode s'applique à la détection de 2 % d'EBC incorporés dans du beurre de cacao, proportion correspondant à environ 0,4 % d'EBC dans le chocolat (dans l'hypothèse d'un taux de matières grasses de ce dernier égal à 30 %).

Živalske in rastlinske maščobe in olja - Ekvivalenti kakavovega masla v kakavovem maslu in navadni čokoladi - 1. del: Ugotavljanje prisotnosti ekvivalentov kakavovega masla (ISO 23275-1:2006)

General Information

• Status: Published
• Publication Date: 11-Nov-2008
• Withdrawal Date: 30-May-2009
• ICS: 67.190 - Chocolate
• ICS: 67.200.10 - Animal and vegetable fats and oils
• Technical Committee: CEN/TC 307 - Oilseeds, vegetables and animal fats and oils and their by-products - Methods of sampling and analys
• Drafting Committee: CEN/TC 307 - Oilseeds, vegetables and animal fats and oils and their by-products - Methods of sampling and analys
• Current Stage: 6060 - Definitive text made available (DAV) - Publishing
• Start Date: 12-Nov-2008
• Due Date: 25-Jan-2010
• Completion Date: 12-Nov-2008
• Ref Project: SIST EN ISO 23275-1:2009 - Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain chocolate - Part 1: Determination of the presence of cocoa butter equivalents (ISO 23275-1:2006)

Overview

EN ISO 23275-1:2008 (identical to ISO 23275-1:2006) defines a laboratory method to detect cocoa butter equivalents (CBEs) in cocoa butter and plain chocolate. The procedure uses high-resolution capillary gas-liquid chromatography (HR‑GC) of triacylglycerols followed by regression data evaluation to give a qualitative result: CBEs present / CBEs not present (YES/NO). The method reliably detects 2% CBE in cocoa butter, corresponding to about 0.6% CBE in chocolate (assuming 30% fat).

Key topics and technical requirements

• Analytical principle: Separation of triacylglycerol (TAG) fractions by HR‑GC and linear regression analysis of TAG peaks.
• Detection limit: Applicable for detecting 2% CBE admixture in cocoa butter (~0.6% in standard chocolate).
• Calibration & system suitability: Use of certified reference material IRMM-801 for calibration and checks; CoCal-1 toolbox referenced for validated methods and electronic evaluation.
• Instrumentation:
  • Gas chromatograph with cold on‑column injection and flame ionization detector (FID) (alternative injection systems allowed if equivalent results obtained).
  • Typical column: 25–30 m, 0.25 mm i.d., fused silica with 50% phenylmethylpolysiloxane, film 0.10–0.15 µm.
  • Temperature program example: start 100 °C, ramp 30 °C/min to 340 °C.
  • Carrier gas: helium or hydrogen, purity ≥ 99.999%.
• Sample preparation & procedure highlights: Sampling, fat extraction from chocolate, TAG separation by HR‑GC, identification, calculation of response factors and TAG percentages, and decision rule for presence of CBEs.
• Reagents & safety: Use analytical‑grade reagents and non‑chlorinated fat solvents (e.g., diethyl ether, n‑heptane). Follow safety regulations for hazardous materials.
• Precision & validation: Method includes system suitability, repeatability and reproducibility data from interlaboratory tests.

Applications and users

This standard is intended for:

• Food testing laboratories and quality control (QC) units in chocolate and confectionery manufacturers
• Regulatory and enforcement laboratories verifying compliance with labeling and compositional rules
• Researchers and method developers working on fat analysis and authenticity testing
• Certification bodies needing a validated detection method for cocoa butter equivalents

Use cases include routine screening of cocoa butter supplies, auditing plain chocolate for unauthorized vegetable fats, and supporting regulatory compliance (e.g., EU Directive 2000/36/EC).

Related standards

• ISO 23275‑2 - Quantification of cocoa butter equivalents (provides quantification capability, e.g., validated to 5%).
• Reference materials and analytical toolbox: IRMM‑801 and CoCal‑1 for calibration and data evaluation.

Keywords: EN ISO 23275-1:2008, ISO 23275-1:2006, cocoa butter equivalents, CBEs detection, HR‑GC triacylglycerols, IRMM-801, chocolate authenticity.

Standards Content (Sample)

SLOVENSKI STANDARD
01-april-2009
äLYDOVNHLQUDVWOLQVNHPDãþREHLQROMD(NYLYDOHQWLNDNDYRYHJDPDVODY
NDNDYRYHPPDVOXLQQDYDGQLþRNRODGLGHO8JRWDYOMDQMHSULVRWQRVWL
HNYLYDOHQWRYNDNDYRYHJDPDVOD,62
Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain
chocolate - Part 1: Determination of the presence of cocoa butter equivalents (ISO 23275
-1:2006)
Tierische und pflanzliche Fette und Öle - Kakaobutter-Äquivalente in Kakaobutter und
Zartbitterschokolade - Teil 1: Bestimmung der Präsenz von Kakaobutter-Äquivalenten
(ISO 23275-1:2006)
Corps gras d'origines animale et végétale - Équivalents au beurre de cacao dans le
beurre de cacao et dans le chocolat de ménage - Partie 1: Détermination de la présence
d'équivalents au beurre de cacao (ISO 23275-1:2006)
Ta slovenski standard je istoveten z: EN ISO 23275-1:2008
ICS:
67.190 ýRNRODGD Chocolate
67.200.10 5DVWOLQVNHLQåLYDOVNH Animal and vegetable fats
PDãþREHLQROMD and oils
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EUROPEAN STANDARD
EN ISO 23275-1
NORME EUROPÉENNE
EUROPÄISCHE NORM
November 2008
ICS 67.200.10; 67.190
English Version
Animal and vegetable fats and oils - Cocoa butter equivalents in
cocoa butter and plain chocolate - Part 1: Determination of the
presence of cocoa butter equivalents (ISO 23275-1:2006)
Corps gras d'origines animale et végétale - Équivalents au Tierische und pflanzliche Fette und Öle - Kakaobutter-
beurre de cacao dans le beurre de cacao et dans le Äquivalente in Kakaobutter und Zartbitterschokolade - Teil
chocolat de ménage - Partie 1: Détermination de la 1: Bestimmung der Präsenz von Kakaobutter-Äquivalenten
présence d'équivalents au beurre de cacao (ISO 23275- (ISO 23275-1:2006)
1:2006)
This European Standard was approved by CEN on 23 October 2008.
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 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 Management Centre has the same status as the
official versions.
CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Finland,
France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal,
Romania, 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
© 2008 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 23275-1:2008: E
worldwide for CEN national Members.

Contents Page
Foreword.3

Foreword
The text of ISO 23275-1:2006 has been prepared by Technical Committee ISO/TC 34 “Agricultural food
products” of the International Organization for Standardization (ISO) and has been taken over as EN ISO
23275-1:2008 by Technical Committee CEN/TC 307 “Oilseeds, vegetable and animal fats and oils and their
by-products - Methods of sampling and analysis” the secretariat of which is held by AFNOR.
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 May 2009, and conflicting national standards shall be withdrawn at the
latest by May 2009.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights.
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, Cyprus, Czech
Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia,
Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain,
Sweden, Switzerland and the United Kingdom.
Endorsement notice
The text of ISO 23275-1:2006 has been approved by CEN as a EN ISO 23275-1:2008 without any
modification.
INTERNATIONAL ISO
STANDARD 23275-1
First edition
2006-11-01
Animal and vegetable fats and oils —
Cocoa butter equivalents in cocoa butter
and plain chocolate —
Part 1:
Determination of the presence of cocoa
butter equivalents
Corps gras d'origines animale et végétale — Équivalents au beurre de
cacao dans le beurre de cacao et dans le chocolat de ménage —
Partie 1: Détermination de la présence d'équivalents au beurre de
cacao
Reference number
ISO 23275-1:2006(E)
©
ISO 2006
ISO 23275-1:2006(E)
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ii © ISO 2006 – All rights reserved

ISO 23275-1:2006(E)
Contents Page
Foreword. iv
Introduction . v
1 Scope .1
2 Terms and definitions .1
3 Principle.1
4 Reagents and materials .1
5 Apparatus .2
6 Sampling.2
7 Preparation of test sample.3
7.1 Preparation of cocoa butter CRM for calibration purposes and system suitability check.3
7.2 Preparation of chocolate sample .3
8 Procedure .3
8.1 Fat extraction .3
8.2 Separation of individual triacylglycerols by HR-GC .3
8.3 Identification.3
9 Calculation.4
9.1 Determination of response factors .4
9.2 Calculation of percentages of triacylglycerols.4
9.3 Decision if sample is pure cocoa butter.4
10 Procedural requirements .5
10.1 General considerations.5
10.2 System suitability .5
11 Precision.5
11.1 Interlaboratory test .5
11.2 Repeatability.6
11.3 Reproducibility.6
12 Test report .6
Annex A (informative) Results of interlaboratory test.7
Bibliography .12

ISO 23275-1:2006(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 committee has been
established has the right to be represented on that committee. International organizations, governmental and
non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the
International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
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.
ISO 23275-1 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 11, Animal
and vegetable fats and oils.
ISO 23275 consists of the following parts, under the general title Animal and vegetable fats and oils — Cocoa
butter equivalents in cocoa butter and plain chocolate:
⎯ Part 1: Determination of the presence of cocoa butter equivalents
⎯ Part 2: Quantification of cocoa butter equivalents
iv © ISO 2006 – All rights reserved

ISO 23275-1:2006(E)
Introduction
“Cocoa butter equivalents” is the general term for fats used to replace cocoa butter in chocolate. They
resemble the chemical composition and physical properties of cocoa butter very closely, making them
therefore extremely difficult to quantify and even in some cases to detect. In principle, cocoa butter
equivalents must by definition be fats low in lauric acid, rich in symmetrical mono-unsaturated triacylglycerols
of the type 1,3-dipalmitoyl-2-oleoylglycerol, 1-palmitoyl-2-oleoyl-3-stearoylglycerol and 1,3-distearoyl-2-
oleoylglycerol, miscible with cocoa butter, and obtained only by refining and fractionation.
Within the European Union, the following vegetable fats, obtained from the plants listed below, may be used
[1]
singly or in blends, according to Directive 2000/36/EC :
⎯ illipé, Borneo tallow or tengkawang (Shorea spp.),
⎯ palm oil (Elaeis guineensis, Elaeis olifera),
⎯ sal (Shorea robusta),
⎯ shea (Butyrospermum parkii),
⎯ kokum gurgi (Garcinia indica), and
⎯ mango kernel (Mangifera indica).
This part of ISO 23275 specifies a procedure for the detection of these fats (restrictions are only made for
pure illipé fat samples) in cocoa butter and plain chocolate. ISO 23275-2 specifies a procedure allowing a
reliable quantification of these fats at the level of 5 %, complying with the statutory limit laid down in Directive
[1]
2000/36/EC of the European Parliament and the Council.
To facilitate the usage of both parts of ISO 23275, an analytical toolbox named “CoCal-1” has been
established. “CoCal-1” contains the validated methods for detection (part 1) and quantification (part 2) of
CBEs in plain chocolate, and also a certified cocoa butter reference material (IRMM-801) to calibrate the
analyst’s instruments and an electronic evaluation sheet for Microsoft Excel® to calculate the final result. An
analyst working on CBE detection and quantification has only to calibrate the gas chromatographic separation
system using IRMM-801, separate the triglyceride fractions of the sample in question, and use the electronic
evaluation sheet for subsequent data treatment to detect and quantify CBEs.
Information on “CoCal-1” is available on the website of the Institute for Reference Materials and
Measurements: http://www.irmm.jrc.be.

INTERNATIONAL STANDARD ISO 23275-1:2006(E)

Animal and vegetable fats and oils — Cocoa butter equivalents
in cocoa butter and plain chocolate —
Part 1:
Determination of the presence of cocoa butter equivalents
1 Scope
This part of ISO 23275 specifies a procedure for the detection of cocoa butter equivalents (CBEs) in cocoa
butter (CB) and plain chocolate by high-resolution capillary gas liquid chromatography (HR-GC) of
triacylglycerols and subsequent data evaluation by regression analysis.
The method is applicable for the detection of 2 % CBE admixture to cocoa butter, corresponding to about
0,6 % CBE in chocolate (i.e. the assumed fat content of chocolate is 30 %).
2 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
2.1
cocoa butter equivalents
CBEs
fats detected in cocoa butter and plain chocolate according the procedure specified in this part of ISO 23275
NOTE The result is expressed as a qualitative result, i.e. CBEs present/CBEs not present (YES/NO).
3 Principle
Cocoa butter, or the fat obtained by solvent extraction from plain chocolate, is separated by HR-GC into
triacylglycerol fractions according to their molecular mass and degree of unsaturation. The presence of CBEs
is detected by linear regression analysis applied to individual triacylglycerol fractions of the fat analysed.
4 Reagents and materials
Use only reagents of recognized analytical grade, unless otherwise stated.
WARNING — Attention is drawn to the regulations which specify the handling of dangerous matter.
Technical, organizational and personal safety measures shall be followed.
[2]
4.1 Cocoa butter Certified Reference Material (CRM) IRMM-801 , for calibration purposes and system
suitability check.
4.2 Fat solvent, non-chlorinated solvents (e.g. diethyl ether, n-heptane, iso-octane).
ISO 23275-1:2006(E)
5 Apparatus
5.1 Analytical balance, with a readability of 0,1 mg.
5.2 Drying oven, maintained at 55 °C.
A dry heater block may be used.
5.3 Food grater, i.e. a kitchen blender with a design featuring the motor above the receiving container to
1)
avoid melting the samples .
5.4 Rotary evaporator.
Alternative evaporation procedures may be used.
5.5 Pipettes, of capacity 1 ml.
5.6 Volumetric flasks, of capacity 20 ml.
5.7 Microsyringe, with maximum volume 10 µl, graduated to 0,1 µl, or automatic sampler.
5.8 Gas chromatograph (GC), fitted with a cold on-column injection system and a flame ionization detector
(FID).
Alternative injection systems [e.g. a split injector, a programmed-temperature vaporizer (PTV) or a moving-
needle injector] may be used provided the same results are obtained as indicated in 10.2.
The separation and detection have been found to be satisfactory if the following experimental conditions are
followed:
⎯ GC column: 25 m to 30 m length, with 0,25 mm i.d., fused silica coated with thermostable 50 %
phenylmethylpolysiloxane to a film thickness of 0,1 µm to 0,15 µm.
⎯ temperature programme: 100 °C (initial temperature), programme rate 30 °C/min to 340 °C (final
temperature).
⎯ carrier gas: helium or hydrogen (purity W 99,999 %).
NOTE Suitable columns and alternative experimental conditions, used in an international collaborative study, are
listed in Annex A. Operating conditions may be changed to obtain optimum separation of cocoa butter triacylglycerols.
5.9 Chromatographic data system.
6 Sampling
A representative sample should have been sent to the laboratory. It should not have been damaged or
changed during transport or storage.
Sampling is not part of the method specified in this part of ISO 23275. A recommended sampling method is
given in ISO 5555.
1) Philips HR2833 is an example of suitable equipment available commercially.
This information is given for the convenience of the users of this part of ISO 23275 and does not constitute an
endorsement by ISO of this product.
2 © ISO 2006 – All rights reserved

ISO 23275-1:2006(E)
7 Preparation of test sample
7.1 Preparation of cocoa butter CRM for calibration purposes and system suitability check
Before opening and using the cocoa butter CRM (4.1), the ampoule shall be warmed in a drying oven (5.2)
until the contents have melted. When a clear solution is obtained, mix t
...