🍎 Quantification of cocoa butter equivalents (CBEs) in cocoa butter and plain chocolate — 4–5 g of grated chocolate is hydrolysed for 15 min in boiling hydrochloric acid, the residue dried and extracted with light petroleum in a Soxhlet apparatus for 4 h (fat content in g/100 g), the fat in a 0.5 mg/ml solution is separated by high-resolution capillary gas chromatography with FID into the triacylglycerols POP, POS, POO, SOS and SOO, calibrated against the certified cocoa butter IRMM-801, and the CBE content in cocoa butter is calculated with the PLS regression equation: 37.439 + 1.175 POP − 1.939 POS − 0.121 POO + 0.982 SOS − 0.097 SOO (g/100 g) and converted to chocolate, PN-EN ISO 23275-2

Food chemistry PN-EN ISO 23275-2

Overview

WHAT THE STANDARD COVERS. We read the text of the standard in the iTeh sample of ISO 23275-2:2006 (English text): title page, contents, foreword, introduction, Clauses 1–8 and Clause 9 up to 9.3 (CBE content in cocoa butter) — the sample ends on page 5. Parts of 5.3 and 7.2 (the first words) and the heading 8.1/8.2 in the contents are missing in the text. We do not read 9.4 (conversion to chocolate), the requirements, precision, test report or Annex A; we know the titles from the contents.

ACCORDING TO THE TEXT OF ISO 23275-2:2006. The standard was prepared by ISO/TC 34 “Food products”, Subcommittee SC 11 “Animal and vegetable fats and oils”; the series has two parts — 1: detection of the presence, 2: quantification of cocoa butter equivalents. Introduction (shared with Part 1): cocoa butter equivalents (CBEs) are fats replacing cocoa butter in chocolate, very similar to it and therefore extremely difficult to quantify and sometimes even to detect; low in lauric acid, rich in the symmetrical monounsaturated triacylglycerols POP, POS and SOS, obtained only by refining and fractionation; in the EU under Directive 2000/36/EC fats from illipe, oil palm, sal, shea, kokum gurgi and mango kernel may be used; Part 2 allows these fats to be reliably quantified at the 5 % level, consistent with the limit of that directive; the “CoCal-1” toolkit includes the validated methods of both parts, the certified cocoa butter IRMM-801 and a spreadsheet for the calculations. Scope (1): quantification of CBEs in cocoa butter and plain chocolate by high-resolution capillary gas chromatography (HR-GC) of triacylglycerols and partial least squares (PLS) regression; note: the presence of CBEs down to 0.6 % (at 30 % fat in chocolate) can be established by the procedure of Part 1 — the methods differ in the number of triacylglycerols and the way of calculation: detection is linear regression on three main fractions, quantification — PLS regression on five. Definitions (2): CBEs — fats detected in cocoa butter and chocolate; CBE content of cocoa butter — in g/100 g of cocoa butter; CBE content of chocolate — in g/100 g of chocolate (both determined by the procedure of this part). Principle (3): cocoa butter or chocolate fat is separated by HR-GC into triacylglycerol fractions according to molecular mass and unsaturation, and the amount of added CBEs is estimated by PLS regression on the proportions of the individual fractions. Reagents (4): IRMM-801; non-chlorinated solvent (e.g. diethyl ether, n-heptane, isooctane); hydrochloric acid 8 mol/l; fluted filter papers 15 cm. Apparatus (5): balance reading to 0.1 mg; oven at 55 °C (or heating block); grater — food processor with the motor above the bowl; 20 ml volumetric flasks; 1 ml pipettes; microsyringe up to 10 µl or autosampler; chromatograph with cold on-column injection and FID (other injectors if they give the results of 10.2); proven conditions: column 25–30 m × 0.25 mm, 50 % polyphenylmethylsiloxane, film 0.1–0.15 µm, 100 → 340 °C at 30 °C/min, helium or hydrogen ≥ 99.999 % (other columns and conditions from the interlaboratory test in Annex A); data system; Soxhlet apparatus (siphon about 100 ml, thimble 33 mm × 88 mm, 250 ml conical flask, heating mantle). Sampling (6): not part of the method, recommended procedure in ISO 5555. Preparation (7): melt IRMM-801 in the oven, mix for at least 20 s by inversion, transfer to a tightly closed vial, keep in the cold; cool the chocolate until hard, grate into fine granules, mix, keep in the cold in a tightly closed bottle. Extraction and fat (8.1): 4–5 g of chocolate in a beaker, slowly 45 ml of boiling water to a homogeneous suspension, 55 ml of HCl 8 mol/l and boiling chips, under a watch glass slowly to the boil and gentle boiling for 15 min; rinse the watch glass with 100 ml of water, filter through a fluted filter, wash with water until the filtrate is free of chlorides; dry the filter with the residue in the thimble for 2 h at 100 °C, cover with glass wool; dry the flask with boiling chips for 1 h at 100 °C, cool in a desiccator and weigh; extraction in the Soxhlet apparatus — the beakers and watch glass are rinsed with three portions of 50 ml of light petroleum into the thimble, heating for 4 h with more than 30 siphonings; evaporate the solvent, dry the flask at 102 °C to constant mass (1.5 h; constant mass — loss in a further hour of drying < 0.05 % fat), cool and weigh; two determinations shall agree within 0.1 % fat; fat content cfat = mfat × 100 / m (g/100 g) to two decimal places; other extractions (accelerated solvent, supercritical CO2, microwaves) acceptable if they give the same results. Chromatography (8.2, 8.3): samples melted completely in the oven, filtered in the oven if a sediment is present, pipettes warmed to about 55 °C (avoiding partial fractionation); about 0.2 g in 20 ml of solvent, 1 ml to 20 ml, injection of 0.5–1.0 µl (0.5 mg/ml) on-column; identification of POP, POS, POO, SOS and SOO by retention times against IRMM-801. Calculations (9): area proportion Pref,i = Aref,i / ΣAref,i × 100 %, response factor Fi = wref,i / Pref,i (wref,i — mass proportion from the IRMM-801 certificate), to two decimal places; proportions of the five triacylglycerols in the sample wtest,i = Fi × Atest,i / Σ(Fi × Atest,i) × 100 %; CBE content of cocoa butter (with POP + POS + POO + SOS + SOO = 100 %): cCBE,CB = 37.439 + 1.175 × POP − 1.939 × POS − 0.121 × POO + 0.982 × SOS − 0.097 × SOO, in g/100 g to two decimal places; the equation was established on a calibration set of 798 mixtures of cocoa butter with CBEs containing 10 %, 15 % and 20 % CBE; mandatory use of IRMM-801 for calibration and system check ensures comparability of results between laboratories; in 99 % of cases, when commercial fats were used for the mixtures, the prediction error does not exceed ± 2.60 % relative to cocoa butter.

STATUS (as of 03.10.2026). In the PKN search (query “23275-2”, 03.10.2026) PN-EN ISO 23275-2:2010 (Polish version, “Wprowadza: EN ISO 23275-2:2008 [IDT], ISO 23275-2:2006 [IDT]”) carries no withdrawal note; PN-ISO 23275-2:2007 (Polish version) — “Wycofana i zastąpiona przez PN-EN ISO 23275-2:2009 – wersja angielska”, and that one — withdrawn and replaced by PN-EN ISO 23275-2:2010 in the Polish version. Card: publication date 25.01.2010, 19 pages, KT 92 Nasion Roślin Oleistych, Tłuszczów Roślinnych i Zwierzęcych oraz ich Produktów Ubocznych, no withdrawal note. In the iTeh catalogue (read 03.10.2026) ISO 23275-2:2006 has the status “Published”, stage 90.92 (International Standard to be revised), dated 24.09.2026.

HOW MANY LABORATORIES AND IN WHAT FORM (copy of the accreditation scope database, data up to 17.09.2026, read 03.10.2026). The number 23275-2 appears in 4 records at 3 laboratories: AB 285, AB 370 (2), AB 705. In the current PCA documents (read 03.10.2026; AB 285 issue no. 28 of 13.01.2026, AB 705 issue no. 21 of 06.05.2026, AB 370 issue no. 25 of 04.08.2026) the number appears in 3 lines at the same 3 laboratories; in each document the number of page markers equals the number of PDF pages, without repetitions; a search of all PCA documents downloaded from the BIP on 03.10.2026 found no laboratories outside the database copy. The form is “PN-EN ISO 23275-2:2010” — alone (AB 705) or together with PN-EN ISO 23275-1:2009 and the report “Report EUR 22666 EN:2007” (AB 285, AB 370). None of these items is suspended. The “Laboratories” tab (query “PN-EN ISO 23275-2”) shows all 3 laboratories — checked by a tab query on the labcoda.pl production server on 03.10.2026.

WHAT THE LABORATORIES TEST (items in PCA). Object: plain chocolate (AB 705), chocolate without additives (AB 285), chocolates, couvertures, cocoa (AB 370). Characteristic and range: content of cocoa butter equivalents (CBE) (1.0–7.0) % (AB 705); CBE content (1.30–5.00) g CBE/100 g of chocolate and milk fat (3.80–28.27) g/100 g of fat and in g/100 g of product by calculation (AB 285); presence and content of CBE and milk fat (4–50) g/100 g of fat, also in g/100 g of product by calculation (AB 370). Technique: GC-FID.

WHERE A SEEMINGLY CORRECT RESULT IS EASY TO GET. Calibration: the PLS equation was established on mixtures with 10 %, 15 % and 20 % CBE (9.3) — AB 370 gives a range up to 50 g/100 g of fat, AB 705 from 1.0 %, AB 285 from 1.30 g/100 g of chocolate; outside this interval the part of the standard we read says nothing about the prediction error. Error: ± 2.60 % relative to cocoa butter in 99 % of cases (9.3) — the result has to be read together with this error, especially close to the 5 % limit. Unit: the content in cocoa butter (g/100 g of cocoa butter) and in chocolate (g/100 g of chocolate) are different quantities (2.2) — AB 370 gives g/100 g of fat, AB 285 g/100 g of chocolate, AB 705 “%”. Fat: conversion to chocolate requires the fat content from Soxhlet extraction after acid hydrolysis (8.1); two fat determinations shall agree within 0.1 %. Matrix: the standard covers cocoa butter and plain chocolate (1); couvertures and cocoa (AB 370) and milk fat (AB 285, AB 370) go beyond its scope — for milk fat both items also cite the report EUR 22666 EN:2007, which we have not read. Presence: detection (YES/NO) is Part 1, based on three triacylglycerols and linear regression — the number from Part 2 does not replace the decision from Part 1.

WHAT WE DO NOT GIVE. The conversion of CBE content to chocolate (9.4), the system suitability requirements, precision data and the results of the interlaboratory test are not quoted — they lie outside the sample. We do not give the content of Directive 2000/36/EC or of the report EUR 22666 EN:2007.

Method principle

The fat is isolated from chocolate after hydrolysis with hydrochloric acid by extraction with light petroleum in a Soxhlet apparatus (with determination of the fat content). A 0.5 mg/ml fat solution is separated by high-resolution capillary gas chromatography with FID into the triacylglycerols POP, POS, POO, SOS and SOO, with response factors from the certified cocoa butter IRMM-801. The proportions of the five triacylglycerols (sum 100 %) are inserted into the PLS regression equation, which gives the CBE content in g/100 g of cocoa butter; conversion to chocolate takes the fat content into account.

Applications

• Quantification of cocoa butter equivalents in cocoa butter and plain chocolate, at the level of the 5 % limit of Directive 2000/36/EC (introduction, 1)
• Fat content of chocolate after acid hydrolysis and Soxhlet extraction (8.1)
• Plain chocolate, chocolate without additives, chocolates, couvertures and cocoa — items in the accreditation scopes of 3 laboratories in PCA

Key parameters

Standard

Standard number

PN-EN ISO 23275-2:2010

Title (PL)

Oleje i tłuszcze roślinne oraz zwierzęce — Odpowiedniki tłuszczu kakaowego w tłuszczu kakaowym i w czekoladzie naturalnej — Część 2: Oznaczanie ilościowe odpowiedników tłuszczu kakaowego

Title (EN)

Animal and vegetable fats and oils — Cocoa butter equivalents in cocoa butter and plain chocolate — Part 2: Quantification of cocoa butter equivalents

Step-by-step procedure

1. Preparation

   IRMM-801 melted and mixed; chocolate cooled and grated (7). PN-EN ISO 23275-2:2010 with no withdrawal note in PKN (03.10.2026).

2. Hydrolysis and fat

   4–5 g + 45 ml boiling water + 55 ml HCl 8 mol/l, 15 min boiling, filtration, drying 2 h at 100 °C, Soxhlet 4 h, 102 °C to constant mass; cfat in g/100 g (8.1).

3. Chromatography

   0.5 mg/ml, injection 0.5–1.0 µl; identification of POP, POS, POO, SOS, SOO against IRMM-801 (8.2, 8.3).

4. Proportions

   Fi = wref,i / Pref,i from IRMM-801; wtest,i for the five triacylglycerols, sum 100 % (9.1, 9.2).

5. Result

   PLS equation → g CBE/100 g of cocoa butter; conversion to chocolate according to 9.4 (outside the sample) (9.3).

Required equipment and apparatus

Reagents, media and consumables

Health and safety (OHS)

• Boiling hydrochloric acid 8 mol/l — work in a fume hood, eye and hand protection
• Light petroleum and diethyl ether — flammable; extraction away from ignition sources (warning in 4)

Frequently asked questions