SIST EN ISO 23275-2:2009

Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain chocolate - Part 2: Quantification of cocoa butter equivalents (ISO 23275-2:2006)

This part of ISO 23275 specifies a procedure for the quantification of cocoa butter equivalents (CBEs) in cocoa butter (CB) and plain chocolate by high-resolution capillary gas chromatography (HR-GC) of triacylglycerols, and subsequent data evaluation by partial least-squares regression analysis.

Tierische und pflanzliche Fette und Öle - Kakaobutter-Äquivalente in Kakaobutter und Zartbitterschokolade - Teil 2: Quantifizierung von Kakaobutter-Äquivalenten (ISO 23275-2:2006)

Dieser Teil von ISO 23275 legt ein Verfahren zur Mengenbestimmung von Kakaobutter-Äquivalenten (CBE) in
Kakaobutter und Zartbitterschokolade durch hochauflösende Kapillar-Gaschromatographie (HR-GC) von
Triacylglycerinen und anschließende Datenauswertung durch partielle Regressionsanalyse nach der Methode
der kleinsten Quadrate fest.
ANMERKUNG Das Vorliegen von CBE in Kakaobutter und Zartbitterschokolade kann noch bis zu einem Gehalt von
0,6 % (der angenommene Fettgehalt der Schokolade beträgt 30 %) durch Anwendung des in ISO 23275-1 erläuterten
Verfahrens bestimmt werden. Unterschiede bei der Durchführung der beiden Verfahren bestehen in der Anzahl der
einzelnen Triacylglycerine, die für die Datenauswertung verwendet werden, und im Prinzip der mathematischen
Auswertung der Daten. Das Vorliegen von CBE wird durch lineare Regressionsanalyse nachgewiesen, die auf die
relativen Anteile der drei hauptsächlichen Triacylglcerin-Fraktionen des analysierten Fettes angewendet wird. Die Menge
des CBE-Zusatzes wird durch partielle Regressionsanalyse nach der Methode der kleinsten Quadrate abgeschätzt, die
auf die relativen Anteile der fünf hauptsächlichen Triacylglcerine angewendet wird.

Corps gras d'origines animale et végétale - Équivalents au beurre de cacao dans le beurre de cacao et dans le chocolat de ménage - Partie 2: Quantification des équivalents au beurre de cacao (ISO 23275-2:2006)

L'ISO 23275-2:2006 spécifie un mode opératoire permettant la quantification 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 par la méthode des moindres carrés partiels.

Živalske in rastlinske maščobe in olja - Ekvivalenti kakavovega masla v kakavovem maslu in navadni čokoladi - 2. del: Kvantitativno določevanje ekvivalentov kakavovega masla (ISO 23275-2:2006)

General Information

• Status: Published
• Publication Date: 24-Feb-2009
• ICS: 67.190 - Chocolate
• ICS: 67.200.10 - Animal and vegetable fats and oils
• Technical Committee: KŽP - Agricultural food products
• Current Stage: 6060 - National Implementation/Publication (Adopted Project)
• Start Date: 16-Feb-2009
• Due Date: 23-Apr-2009
• Completion Date: 25-Feb-2009
• Ref Project: EN ISO 23275-2:2008 - Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain chocolate - Part 2: Quantification of cocoa butter equivalents (ISO 23275-2:2006)

Standards Content (Sample)

SLOVENSKI STANDARD
SIST EN ISO 23275-2:2009
01-april-2009
äLYDOVNHLQUDVWOLQVNHPDãþREHLQROMD(NYLYDOHQWLNDNDYRYHJDPDVODY
NDNDYRYHPPDVOXLQQDYDGQLþRNRODGLGHO.YDQWLWDWLYQRGRORþHYDQMH
HNYLYDOHQWRYNDNDYRYHJDPDVOD,62
Animal and vegetable fats and oils - Cocoa butter equivalents in cocoa butter and plain
chocolate - Part 2: Quantification of cocoa butter equivalents (ISO 23275-2:2006)
Tierische und pflanzliche Fette und Öle - Kakaobutter-Äquivalente in Kakaobutter und
Zartbitterschokolade - Teil 2: Quantifizierung von Kakaobutter-Äquivalenten (ISO 23275-
2:2006)
Corps gras d'origines animale et végétale - Équivalents au beurre de cacao dans le
beurre de cacao et dans le chocolat de ménage - Partie 2: Quantification des équivalents
au beurre de cacao (ISO 23275-2:2006)
Ta slovenski standard je istoveten z: EN ISO 23275-2:2008
ICS:
67.190 ýRNRODGD Chocolate
67.200.10 5DVWOLQVNHLQåLYDOVNH Animal and vegetable fats
PDãþREHLQROMD and oils
SIST EN ISO 23275-2:2009 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST EN ISO 23275-2:2009

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SIST EN ISO 23275-2:2009
EUROPEAN STANDARD
EN ISO 23275-2
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 2: Quantification of
cocoa butter equivalents (ISO 23275-2:2006)
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 2: Quantification des 2: Mengenbestimmung von Kakaobutter-Äquivalenten (ISO
équivalents au beurre de cacao (ISO 23275-2:2006) 23275-2:2006)
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 23275-2:2008: E
worldwide for CEN national Members.

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SIST EN ISO 23275-2:2009
EN ISO 23275-2:2008 (E)
Contents Page
Foreword.3

2

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SIST EN ISO 23275-2:2009
EN ISO 23275-2:2008 (E)
Foreword
The text of ISO 23275-2:2006 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
23275-2:2008 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 23275-2:2006 has been approved by CEN as a EN ISO 23275-2:2008 without any
modification.

3

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SIST EN ISO 23275-2:2009

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SIST EN ISO 23275-2:2009

INTERNATIONAL ISO
STANDARD 23275-2
First edition
2006-11-01

Animal and vegetable fats and oils —
Cocoa butter equivalents in cocoa butter
and plain chocolate —
Part 2:
Quantification 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 2: Quantification des équivalents au beurre de cacao




Reference number
ISO 23275-2:2006(E)
©
ISO 2006

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(E)
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ii © ISO 2006 – All rights reserved

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(E)
Contents Page
Foreword. iv
Introduction . v
1 Scope .1
2 Terms and definitions .1
3 Principle.1
4 Reagents and materials .2
5 Apparatus .2
6 Sampling.3
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 .4
8.3 Identification.4
9 Calculation.4
9.1 Determination of response factors .4
9.2 Calculation of percentages of triacylglycerols.5
9.3 Calculation of CBE content in cocoa butter .5
9.4 Calculation of CBE content in chocolate .6
10 Procedural requirements .6
10.1 General.6
10.2 System suitability .6
11 Precision.6
11.1 Interlaboratory test .6
11.2 Repeatability.7
11.3 Reproducibility.7
12 Test report .7
Annex A (informative) Results of interlaboratory test.8
Bibliography .13

© ISO 2006 – All rights reserved iii

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(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 23275-2 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

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(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).
ISO 23275-1 specifies a procedure for the detection of these fats (restrictions are only made for pure illipé fat
samples) in cocoa butter and plain chocolate. This part of ISO 23275 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.
© ISO 2006 – All rights reserved v

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SIST EN ISO 23275-2:2009

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SIST EN ISO 23275-2:2009
INTERNATIONAL STANDARD ISO 23275-2:2006(E)

Animal and vegetable fats and oils — Cocoa butter equivalents
in cocoa butter and plain chocolate —
Part 2:
Quantification of cocoa butter equivalents
1 Scope
This part of ISO 23275 specifies a procedure for the quantification of cocoa butter equivalents (CBEs) in
cocoa butter (CB) and plain chocolate by high-resolution capillary gas chromatography (HR-GC) of
triacylglycerols, and subsequent data evaluation by partial least-squares regression analysis.
NOTE The presence of CBEs in CB and plain chocolate down to a level of 0,6 % (fat content of chocolate assumed
to be 30 %) can be determined by using the procedure explained in ISO 23275-1. Differences in the procedure of the two
methods exist in the number of individual triacylglycerols used for data treatment and in the mathematical evaluation
principle of the data. The presence of CBEs is detected by linear regression analysis applied to the relative proportions of
the three main triacylglycerol fractions of the fat analysed. The amount of the CBE admixture is estimated by partial least
squares regression analysis applied to the relative proportions of the five main triacylglycerols.
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
2.2
CBE content of cocoa butter
mass fraction of substances in cocoa butter determined by the procedure specified in this part of ISO 23275
NOTE It is expressed in grams per 100 g of cocoa butter.
2.2
CBE content of chocolate
mass fraction of substances in chocolate determined by the procedure specified in this part of ISO 23275
NOTE It is expressed in grams per 100 g of chocolate.
3 Principle
Cocoa butter, or the fat obtained from plain chocolate, is separated by HR-GC into triacylglycerol fractions
according to their molecular mass and degree of unsaturation. The added amount of CBEs is estimated by
partial least squares regression analysis applied to individual triacylglycerol fractions of the fat analysed.
© ISO 2006 – All rights reserved 1

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(E)
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).
4.3 Hydrochloric acid, c = 8 mol/l.
1)
4.4 Fluted filter paper ,15 cm.
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, a kitchen blender with a design featuring the motor above the receiving container to avoid
2)
melting the samples .
5.4 Volumetric flasks, of capacity 20 ml.
5.5 Pipettes, of capacity 1 ml.
5.6 Microsyringe, with maximum volume 10 µl, graduated to 0,1 µl, or automatic sampler.
5.7 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 quantification have proven 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 %).

1) S&S 589 medium filter paper is an example of a suitable product available commercially.
2) Philips HR2833 is an example of suitable equipment commercially available.
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 these products.
2 © ISO 2006 – All rights reserved

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SIST EN ISO 23275-2:2009
ISO 23275-2:2006(E)
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.
5.8 Chromatographic data system.
5.9 Soxhlet extractor
...