Cocoa butter analysis

All of the classical analytical techniques applied to oils analysis have been applied to studying the authenticity of cocoa butter and considerable attention has been paid to the issue of detecting and quantifying non-cocoa fats in mixtures with cocoa butter and as incorporated into chocolate. A substantial review of these methods has been published (Lipp and Anklam, 1998b). 

Much of the sterol component of shea butter is present as esters of cinnamic acid, which are less readily saponified than esters with glycerol (Peers, 1977) published data for this fat might therefore underestimate the hue value. Surprisingly measurement of cinnamic acid has not been used to test for the presence of shea butter in mixtures. Triterpene alcohols similar to those found in shea also occur in sal fat. Homberg and Bielefeld (1982) showed the presence of triterpene alcohols in illipe and sal fats and in commercial CBEs, and their analysis was proposed as a qualitative measure to detect cocoa butter adulteration. 

Reaction of shea triterpene alcohols with acetic anhydride and sulphuric acid to produce coloured products (Fitelson s reaction) was the basis of a sensitive early test for the presence of shea butter in cocoa butter (Fincke, 1975). Analysis of the triterpene fraction of a commercial cocoa butter by TLC fractionation followed by GC (Fincke, 1976), or argentation TLC followed by GC (Gegiou and Staphylakis, 1985), have been shown to have potential for detecting CBEs in chocolate based on the difference in levels of P-amyrin, butyrospermol and... 

Pyrolysis of food samples provides a large number of products, which, after detection by MS and analysis by advanced statistical treatment, can be used to compare different samples. The method is very rapid and does not require chromatographic separation or MS identification of the pyrolysis products. Pyrolysis MS, coupled with multivariate data analysis procedures, has been used to discriminate between cocoa butters of three different continental areas (Radovic et al., 1998). The technique could in some cases separate deodorized from non-deodorized cocoa butters and also show those that have had alkali treatment. The presence of non-cocoa fats did not affect the assay. 

Trace element analysis of foods can be carried out to check for contamination by toxic elements, such as lead and cadmium, or to determine beneficial micronutrients, or as an aid to distinguishing geographical origin. In fats, small numbers of trace elements are measured after digestion of the sample in acid followed atomic absorption spectrophotometry (AAS) or by direct graphite furnace vaporization. An AAS procedure for measuring lead in edible oils and fats has been collaboratively trialed with cocoa butter as a test material (Firestone, 1994). 

Hernandez et al. (1994) used AAS with acid digestion to measure copper, zinc, iron, magnesium and manganese in four South American and 15 African cocoa butters. The much higher levels reported (e.g. iron, 20-50 mg/kg, copper, 11-27 mg/kg) were ascribed to contamination by metals from grinding tools. Principal component analysis could not distinguish between samples from different continents or countries. 

A fluorescence under UV illumination of an unidentified compound separated by TLC has been used to detect 5% kokum (possibly unrefined) in mixtures with cocoa butter (Deotale el al., 1990). Identification of this compound and its analysis by more specific techniques might be used to improve current methods of quantifying CBEs in chocolate. 

Nuclear magnetic resonance (NMR) can be used as a rapid alternative to differential scanning calorimetry in the determination of the solid fat content and studies on the melting behaviour. The determination is based on detection of the different populations of protons in solid and liquid phases, which indicates the hardness of the fat. Hernandez and Rutledge (1994b) used low resolution pulse NMR to compare melting curves of roasted and non-roasted cocoa butters from Africa, Indonesia and South America. Discriminant analysis techniques showed... 

The advent of relatively inexpensive computers has enabled the accumulation and rapid analysis of large sets of data. By this means patterns and trends not always apparent from visual inspection of chromatograms or tables of data can be discriminated by being sorted into recognizable patterns. This approach is essential for some techniques such as pyrolysis where the quantity of data produced would otherwise be overwhelming. Several statistical approaches to exploit the information content of fatty acid and triacylglycerol patterns for the detection and quantification of CBEs in cocoa butter have been reported (Lipp et al., 2001 Simoneau et al., 1999). 

Discriminant analysis has been used in many of the analyses described in this chapter, in particular the classification of cocoa butters by origin and processing from pyrolysis MS data (Radovic et al., 1998), from triacylglycerol profiles obtained by HPLC (Hernandez et al., 1991) and from analysis of volatiles (Pino, 1992). Data from the analysis of mixtures of CBEs with cocoa butter, which model techniques for measuring CBEs in chocolate, have been treated by similar means (Anklam et al., 1996). 

Multivariate statistical techniques can be used to combine data from several classical analytical methods. In this way Simoneau et al. (2000) tested the quantification of CBEs in mixtures with cocoa butter by analysis of fatty acids by GC and triacyglycerols with both GC and HPLC. 

Anklam, E., Lipp, M. and Wagner, B. (1996) HPLC with light scatter detector and chemometric data evaluation for the analysis of cocoa butter and vegetable fats. Fett Lipid, 98(2), 55—59. 

Fincke, A. (1976) Cocoa butter and substitute fats—Chemistry and analysis (Kakaobutter und Ersatzfette—chemie und analytik). Deut. Lebensm.-Rundsch., 72(1), 6-12. 

FSA (Food Standards Agency) (2001) The estimation of cocoa butter equivalents in cocoa butter and chocolate fats by triglyceride analysis collaborative trial. Information Bulletin, No. 13, October 2001. 

Lipp, M., Simoneau, C. and Anklam, E. (1996) Analysis of triglycerides from cocoa butter, vegetable... 

Md Ali, A.R. and Dimick, P.S. (1994) Thermal analysis of palm mid fraction, cocoa butter and milk fraction blends by differential scanning calorimetry. J. Am. Oil Chem. Soc., 71(3), 299-302. 

Simoneau, C., Hannaert, P. and Anklam, A. (1999) Detection and quantification of cocoa butter equivalents in chocolate model systems analysis of triglyceride profiles by high resolution GC. Food Chem., 65, 111-116. 

Unlike olive oil, the analysis of cocoa butter is not governed by legal definitions. However, the legal definition of chocolate is specific in relation to whether cocoa butter is present alone or as the major vegetable fat, with strict limits on the presence of other vegetable fats in the product. Cocoa butter is also one of the few fats for which artificially manufactured substitutes of similar composition have been constructed and openly marketed. Because of this, analysis of the adulteration of cocoa butter probably has a greater importance than that of any fat other than olive oil, and the approaches to this analysis are described in chapter 3. 

This method can be applied directly to acylglycerols present in a mixture, as is illustrated by the mass spectrometric analysis of natural cocoa butter. [299] This analysis (Figure 8.65) allowed the determination of the complete structure of the predominant acylglycerols in the cocoa (Table 8.10). 

Fessas, D., Signorelli, M. and Schiraldi, A. (2005). Polymorphous transitions in cocoa butter -a quantitative dsc siuAy.Joumal Of Thermal Analysis And Calorimetry, 82(3) 691-702. 

Autooxidation products of triolein, trilinolein, and trilinolenin were analysed by nonaqueous RPLC with a Cjg colunrn and a acetonitrile/dichloromethane gradient and positive-ion APCl-MS [26]. Among others, mono- and dihydroxy peroxides and different types of mono- and diepoxides were observed. In a subsequent study [27], two parallel mass spectrometers, one ruiming ESl-MS, the other APCl-MS, were applied in the TAG analysis of canola oil. ESl-MS especially showed advantages over APCl-MS in the analysis of TAG oxidation products, because it yielded a more abundant intact [M+NH4]" than APCl-MS did. The ability of ESl-MS to reduce fragmentation of TAG also played an important role in the analysis of regioisomers of short-chain TAG, as reported by Kalo et al. [28]. LC-ESl-MS was also applied in the characterization of the TAG content in Malaysian cocoa butter, reported by... 

In terms of particle size and column dimensions, stationary phases with particle sizes of 5-10 /rm in 250 x 4.6-mm analytical columns are most common. HPLC columns with smaller particle sizes (2-3 /rm) have also been prepared and used. Such columns have the advantages of higher efficiency, better resolution, and shorter analysis times. For example, the triglycerides of cocoa butter were baseline separated with columns that contained 2-3-/tm particles. Research and development of columns with smaller particles is very active. 

Metin, S., and R.W. Hartel, Thermal Analysis of Isothermal Crystallization Kinetics in Blends of Cocoa Butter with Milk Fat or Milk Fat Fractions, Ibid. 75 1617-1624 (1990). 

The base used in the formulation of suppositories can often affect the rate of decomposition of the active ingredients. Aspirin decomposes in several polyoxyethylene glycols which are often incorporated into suppository bases. Degradation was shown to be due in part to transesterification, giving the decomposition products salicylic acid and acetylated polyethylene glycol. The rate of decomposition, which followed pseudo first-order kinetics, was considerably greater than when a fatty base such as cocoa butter was used. Analysis of commercial batches of 100 mg indometacin-polyethylene glycol... 

The new European Chocolate Directive [14] allows the addition of up to 5% of vegetable fats other than cocoa butter (CB), the so-called cocoa butter equivalents (CBEs), in chocolate. CBEs resemble the chemical composition and physical properties of CB very closely, making them therefore extremely difficult to quantify and even in some cases to detect (especially at very low levels). There is a perceived need within official control laboratories for reliable analytical methods for the quantification (around the 5% level) of CBEs in chocolate, as Member States laws and administrative provisions need to comply with the new Chocolate Directive before August 2003. All proposed analytical methods have been evaluated by the JRC in collaboration with EU expert laboratories [15]. The performance of several methods has been compared and a final method based on the analysis of the main components, triglycerides, has been proposed for further validation. 

Metin and Hartel (year ). Crystallization Kinetics of Blends of Cocoa Butter and Milk Fat or Milk Fat Fractions, J. Thermal Analysis 47(5), 1527-1544. 

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