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Evaluation of the potential of SPME-GC-MS and chemometrics to detect adulteration of ground roasted coffee with roasted barley
Journal of Food Composition and Analysis 22 (2009) 257–261
Contents lists available at ScienceDirect
Journal of Food Composition and Analysis journal homepage: www.elsevier.com/locate/jfca
Short Communication
Evaluation of the potential of SPME-GC-MS and chemometrics to detect adulteration of ground roasted coffee with roasted barley Rafael C.S. Oliveira a,*, Leandro S. Oliveira a, Adriana S. Franca a, Rodinei Augusti b a b
Programa de Po´s-Graduac¸a˜o em Cieˆncia de Alimentos/UFMG, Av. Antoˆnio Carlos, 6627, 31270-901, Belo Horizonte, MG, Brazil Departamento de Quı´mica/UFMG, Av. Antoˆnio Carlos, 6627, 31270-901, Belo Horizonte, MG, Brazil
A R T I C L E I N F O
A B S T R A C T
Article history: Received 24 June 2008 Received in revised form 29 September 2008 Accepted 20 October 2008
The objective of the present study was to verify the feasibility of detection of coffee adulteration with roasted barley by a comparative analysis of the volatile profiles of both coffee and barley, pure and mixed, at several roasting degrees. The methodology was based on a GC-MS analysis of the headspace volatiles of several samples of ground roasted coffee and barley. The collection and concentration of the headspaces was performed by solid phase micro-extraction (SPME). The separation of the nonadulterated and adulterated samples was accomplished by application of principal component analysis (PCA) to the chromatographic data obtained. It was observed that, the highest the degree of roast, the more easily discriminated were the adulterated samples, allowing for detection of adulterations with as low as 1% (w/w) roasted barley in dark roasted coffee samples. ß 2009 Elsevier Inc. All rights reserved.
Keywords: Coffee adulteration Fraud Gas chromatography GC Mass spectrometry MS Solid-phase micro-extraction SPME Multivariate statistics Detection and quantification method Food analysis Food quality Food composition
1. Introduction Although detection of adulteration of food and food products was reported as early as the 18th century (Singhal et al., 1997), the increasing number of studies on the development of analytical tools for the detection and quantification of adulterations of food and food products in published works in recent years has demonstrated that this type of fraud has become a common practice in all food sectors all around the world (Cordella et al., 2002; Fugel et al., 2005). Adulteration is a serious problem since it promotes exploitation of consumers by non-compliance with product specifications and with regulatory standards (with consequent deterioration of product quality). Also, it leads to unfair competition, disrupting otherwise stabilized local and global economies, with defrauders gaining over reputable competitors. Adulteration with intent to deceive is usually carried out by admixture of cheaper products and materials which are usually difficult to detect by consumers and by simple routine analytical
* Corresponding author. Tel.: +55 31 91648536; fax: +55 31 34433783. E-mail address: [email protected] (Rafael C.S. Oliveira). 0889-1575/$ – see front matter ß 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.jfca.2008.10.015
techniques, with high-priced commodities being usually the main target for adulteration. Brazil is the largest coffee producer in the world and is also among the largest consumers. Coffee is the main commodity in Brazil and, as such, has been the target of fraudulent admixtures with a diversity of cheaper materials, such as twigs, coffee berry skin and parchment, spent coffee, roasted barley, maize, cocoa, soybean and others (Singhal et al., 1997). As a product of consumption, ground roasted coffee is quite vulnerable to adulteration since it presents physical characteristics such as particle size, texture and color, which are easily reproduced by roasting and grinding a variety of biological materials (cereals, seeds, roots, parchments, etc.). A plethora of techniques has been developed in order to establish suitable parameters and markers for adulterations of ground roasted coffee and instant or soluble coffee. Most of the analytical techniques were based on determination of the carbohydrate chromatographic profiles coupled to multivariate statistical analysis of chromatographic data (Blanc et al., 1989; Davis et al., 1990; Berger et al., 1991; Prodolliet et al., 1995; Bernal et al., 1996), and the overall conclusion was that it is possible to establish quantifiable limits for the presence of specific carbohydrates in the products (e.g. xylose, glucose and
R.C.S. Oliveira et al. / Journal of Food Composition and Analysis 22 (2009) 257–261
258
fructose) above which the product can be safely considered adulterated. Techniques that do not rely on the determination of specific chemical tracers, but rather on strictly physical measurements, were also proposed for the detection of adulterants in coffee, e.g. the use of photoacoustic spectroscopy by Cesar et al. (1984), of infra-red spectroscopy by Briandet et al. (1996), of scanning electron microscopy by Amboni et al. (1999), and of thermal lens spectrometry by Fontes et al. (2001). The aforementioned techniques do not allow for the identification of individual constituents or constituents in admixtures of several adulterants. In the case of roasted coffee, adulterations can usually be perceived by a trained sensory panel, thus, indicating that an analysis of the volatiles profile, which is expected to be singular for a particular adulterant, could provide a reliable method to verify adulteration of ground roasted coffee. Among the commonly used contaminants, barley is particularly difficult to detect especially at low concentrations, and even methodologies that are based on the chemical profile are not effective for this specific contaminant. The study by Godinho et al. (2003) presented a methodology based on solvent extraction followed by HPLC analysis for detection of coffee adulteration by barley and coffee husks. Results were satisfactory for the detection of coffee husks, but the chromatographic profiles of the extracts obtained for coffee and barley were quite similar, with the exception of the absence of caffeine in the chromatograms obtained for barley. Such difference would be helpful only if roasted barley were the product being adulterated, and hence it was concluded that the proposed methodology was not effective for detection of adulteration of coffee by barley. Thus, it was the aim of this work to verify the feasibility of detecting coffee adulteration with roasted barley, based on GC-MS analysis of the headspace volatiles of several samples of ground roasted coffee and barley, including tentative identification of chemical markers and multivariate statistical analysis of the chromatographic data. 2. Materials and methods Good quality crude coffee and crude barley samples were obtained from local markets. Both crude coffee and crude barley were roasted to three distinct degrees of roast in a laboratory oven at the same temperature. Preliminary tests showed that it would take temperatures higher than 200 8C to promote significant color changes in crude barley, at reasonable periods of processing time, to be considered roasted to degrees comparable to those for coffee. Hence, a roasting temperature of 300 8C was selected for the laboratory roasting tests. The degrees of roast were characterized as light, medium, and dark according to industrial color standards (Table 1). The roasting experiments were performed in duplicates. Samples were weighted before and after roasting in order to evaluate weight loss. Just before each analysis, the coffee and barley samples were ground with an electric coffee grinder (C-mill 5679-01US, Bodum, USA) for 30 s.
An SPME triple phase (divinylbenzene/carboxen/polydimethylsiloxane) 50/30 mm fiber (model 57348-U, Supelco Inc., USA) was employed for the extraction of volatiles from the roasted coffee and barley headspace, according to the methodology described by Mancha Agresti et al. (2008). Approximately 3 mL of each sample (ground coffee, barley and coffee/barley mixtures) was placed in sealed 5 mL vials (Supelco Inc., USA) and heated for 10 min at 70 8C. Afterwards the SPME needle was inserted into the vial and the fiber was exposed to the headspace above the sample for 40 min at 70 8C. After sampling, the fiber was thermally desorbed in the GC injection port for 10 min at 250 8C. GC analyses were performed in duplicates using a gas chromatograph (Trace Ultra) coupled to a mass spectrometer (PolarisQ) (ThermoElectron, San Jose, CA), a RTX-5MS column (5% diphenyl, 95% dimethyl polysiloxane) 30 m 0.25 mm I.D (Restec, Ireland) and Helium as carrier gas (1 mL/min). The injector and detector temperatures were set to 250 8C and 300 8C, respectively. The GC oven temperature was programmed from 40 8C (5 min) to 180 8C at the rate of 3 8C/min, then to 250 8C (5 min) at 10 8C/min. The GC injector was operated in the splitless mode. Mass spectra were acquired in the electron impact mode at 70 eV and using an m/z range of 50–650 and a scan time of 2 s. Tentative identification of compounds was performed by comparison of the mass spectra obtained for the peaks in the analysis with those of the NIST/EPA/NIH Mass Spectral Library, version 2.0 (available in the instrument software), considering a similarity level (RSI) higher than 800. For principal component analysis, data matrices were assembled so that the rows corresponded to roasted coffee and/ or barley samples and each column represented the peak area of a specific component. The chromatographic peaks with S/N (signalto-noise ratio) higher than 50 were included in the final data matrix regardless of substance identification. This choice was based on the extensive number of peaks detected (over 250). Some preliminary tests with smaller S/N ratio values indicated that the amount of detected substances would be prohibitive and increase the degree of redundancy in terms of multivariate statistical analysis, since most of them were present in similar amounts in all chromatograms for both roasted coffee and roasted barley. 3. Results and discussion Results for weight loss during roasting are shown in Fig. 1, with the solid lines representing linear fits. Both coffee and barley presented the same qualitative behavior for weight loss during roasting, being characterized by a slower rate at the beginning of roasting, followed by an increase in weight loss rate towards the end of the process. This behavior was satisfactorily described by two straight lines (r2 > 0.96) representing the two different rates, and has been reported by previous studies on coffee roasting (Dutra et al., 2001; Oliveira et al., 2005). The first period,
Table 1 Roasting conditions (roasting temperature: 300 8C). Sample ID Coffee CL CM CD Barley BL BM BD
Roasting degree
Roasting time (min)
light medium dark
9 10 11
light medium dark
16 17 18
Fig. 1. Weight loss during roasting.
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corresponding to the slower weight loss rate, is called drying stage, and it is due mainly to the slow release of water and volatiles from crude coffee and barley. The onset of pyrolysis can be associated with the transition between the two slopes, with the increase in weight loss rate being attributed to an intensive release of volatile organic compounds and CO2. A comparison of the weight loss curves for coffee and barley shows that the weight loss rates during both the drying and the pyrolysis stages were slower for barley in comparison to those for coffee. However, average weight loss values for a given roasting degree (strictly based on color) were higher for barley than for coffee, indicating that more severe roasting conditions are required for roasted barley to attain a similar color to that of coffee at lighter degrees of roast. Such differences could be attributed to the difference in chemical changes occurring within the barley grains during roasting. These differences indicate that, if one is to adulterate roasted coffee with roasted barley without significant alterations in roasted color, the roasting of each product should be carried out separately, since, for the same roasting conditions (time and temperature), barley will either roast to a much lighter color or not roast at all (for shorter times and lower temperatures). Also, as discussed by Cristo et al. (2006) in their study of flow regimes of coffee beans in rotating roasters, smaller particles will tend to migrate to the low velocity core of the bed transverse section. Thus, due to poor mixing (low heat transfer rates) in that region, these particles will necessarily roast to a lesser degree, which would be the case of smaller barley grains in a mixture with larger coffee beans. Given the extensive number of compounds that were detected in all roasted coffee and roasted barley samples, multivariate statistical analysis (PCA) was performed in order to verify the possibility of discrimination between roasted coffee and roasted barley based on the volatile profiles. The biplot of the coffee and barley samples submitted to roasting at 300 8C is displayed in Fig. 2. Regarding the volatile profiles of roasted coffee in comparison to roasted barley, the first two principal components (PCs) explained 93.8% and 2.4% of the chromatographic variance, respectively. A clear separation between roasted coffee and roasted barley could be obtained based on the first component alone. The substances that presented higher influence on PC1 and PC2, based on the projection of score plot, are displayed in Table 2. Pyridine (RT 4 min) was the substance with the highest influence on PC1 for roasted coffee and the relative peak intensity increased with roasting time. Pyridine formation during coffee roasting can be attributed to protein pyrolysis, trigonelline degradation, Strecker degradation and Maillard reactions (De Maria et al., 1996). Also, the
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Fig. 2. PCA scores scatter plot of normalized chromatographic SPME areas of pure coffee and barley submitted to roasting at 300 8C (PC1 vs PC2). C = coffee, B = barley, L = light roast, M = medium roast, D = dark roast.
amount of pyridine depends on the degree of roast, and this compound has been reported to be characteristic of darker roasts (Dart and Nursten, 1985; Franca et al., 2008). The other substances listed in Table 2 as characteristic of roasted coffees have been identified by previous studies in the headspace of roasted coffee (De Maria et al., 1996; Sanz et al., 2002; Rocha et al., 2003; Zambonin et al., 2005; Franca et al., 2008). The two aldehydes that were identified as characteristic of the volatile profile of barley were also detected in the roasted coffee headspace, but in much lower concentrations. Admixtures of roasted barley with roasted coffee samples were intentionally prepared and their respective volatile profiles analyzed in order to verify the capability of discrimination between adulterated and non-adulterated roasted coffee samples. The biplots for the headspace volatiles of admixtures of 1%, 5% and 50% roasted barley in roasted coffee are presented in Fig. 3. These results show that it was possible to detect coffee adulteration with roasted barley in admixtures with as low as 1% (w/w) roasted barley, but only for the highest (darker) degrees of roast. This does not pose a problem, given that adulterated samples are usually submitted to severe roasting conditions (dark roasts), so that sensory perception of the adulteration becomes more difficult. Also, as the amount of barley increased, the separation became more evident. The PCA scores of the headspace profiles for the admixtures with roasted barley are closer to those for non-
Table 2 Tentative indentification of aroma components with the most pronounced effect on PC1 and PC2 values. Retention time (min)
3.98 6.35 8.50 8.65 10.36 13.02 14.75 29.86 6.92 12.95 17.54 22.53 27.33 3.77 14.34 *
Compound Roasted Coffee pyridine 2-methyl pyrimidine* 2- furanmethanol 3-furanmethanol* 2,6-dimethyl pyrazine 5-methyl-2-furancarboxaldehyde 2-furanmethanol, acetate 4-hidroxy-3-methylacetophenone Roasted barley 3-furaldehyde 5-methyl-2-furancarboxaldehyde* Butyl-Benzene Pentyl-Benzene Hexyl-Benzene Coffe/barley admixtures Ethanedinitrile 2-furanmethanol, acetate
Detected by Franca et al. (2008) on volatile profiles of roasted coffee at 200 8C.
m/z of the most intense ions (relative abundance %)
79 (100), 80 (85), 52 (50) 94 (100), 67 (40), 95 (40) 81 (100), 98 (70), 97 (60) 81 (100), 98 (55), 97 (50) 108 (100), 109 (30), 107 (20) 109 (100), 110 (50), 53 (30) 98 (100), 81 (80), 53 (40) 150 (100), 135 (70), 107 (45) 95 (100), 96 (30), 67 (10) 109 (100), 110 (50), 53 (30) 91 (100), 92 (60), 134 (30) 91 (100), 92 (55), 148 (30) 91 (100), 92 (55), 162 (20) 53 (100), 58 (5), 85 (5) 98 (100), 81 (90), 53 (40)
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Fig. 3. PCA scores scatter plot of normalized chromatographic SPME areas of (a) 1%, (b) 10%, and (c) 50% roasted barley in roasted coffee in comparison to pure coffee and pure barley. C = coffee, B = barley, M1 = 1:99 barley coffee mixture; M10 = 10:90 barley coffee mixture; M50 = 50:50 barley coffee mixture; L = light roast, M = medium roast, D = dark roast.
adulterated coffee samples due to the fact that roasted coffee headspace profiles present higher amounts of discriminating compounds than the headspace for roasted barley. The preliminary results obtained in the present study indicate that SPME-GC-MS is a promising detection technique to be employed for detection of coffee fraud. However, further studies regarding different roasting conditions and contaminants are necessary to assure the applicability of the proposed methodology to a wider range of roasting conditions and also for simultaneous detection of different contaminants in the same sample. 4. Conclusions SPME-GC-MS was successfully employed for the detection of adulteration of ground roasted coffee with roasted barley. The methodology allowed for the detection of adulterations with as low as 1% (w/w) roasted barley in roasted coffee samples, for the darkest degrees of roast. It was observed that the higher the amount of adulterant, the more effective was the methodology in discriminating adulterated from non-adulterated samples, as expected. Also, the higher the degree of roast, the more easily discriminated were the adulterated samples. It was possible to tentatively identify volatile compounds in the headspace profile of roasted barley to be further used as chemical markers in the detection of adulteration of roasted ground coffee with roasted barley. Acknowledgements The authors acknowledge financial support from the following Brazilian Government Agencies: CAPES, CNPq and FAPEMIG.
References Amboni, R.D.M.C., Francisco, A.D.E., Teixeira, E., 1999. Detection of frauds in ground coffee with scanning electronic microscopy. Cieˆncia e Tecnologia de Alimentos 19, 311–313 (in Portuguese). Berger, A., Brulhart, M., Prodolliet, J., 1991. Detection of adulteration in pure soluble coffee by enzymatic sugar determination. LWT – Food Science and Technology 24, 59–62. Bernal, J.L., Del Nozal, M.J., Toribio, L., Del Alamo, M., 1996. HPLC analysis of carbohydrates in wines and instant coffees using anion exchange chromatography coupled to pulsed amperometric detection. Journal of Agricultural and Food Chemistry 44, 507–511. Blanc, M.B., Davis, G.E., Parchet, J-.M., Viani, R., 1989. Chromatographic profile of carbohydrates in commercial soluble coffees. Journal of Agricultural and Food Chemistry 37, 926–930. Briandet, R., Kemsley, E.K., Wilson, R.H., 1996. Approaches to adulteration detection in instant coffees using infrared spectroscopy and chemometrics. Journal of the Science of Food and Agriculture 71, 359–366. Cesar, C.L., Lima, C.A.S., Vargas, H., Miranda, L.C.M., 1984. On the use of photoacoustic spectroscopy for investigating adulterated or altered powdered coffee samples. Journal of Agricultural and Food Chemistry 32, 1355–1358. Cordella, C., Moussa, I., Martel, A.-C., Sbirrazzuoli, N., Lizzani-Cuvelier, L., 2002. Recent developments in food characterization and adulteration detection: technique-oriented perspectives. Journal of Agricultural and Food Chemistry 50, 1751–1764. Cristo, H.P., Martins, M.A., Oliveira, L.S., Franca, A.S., 2006. Transverse flow of coffee beans in rotating roasters. Journal of Food Engineering 75, 142–148. Davis, G.E., Garwood, V.W., Barfuss, D.L., Husaini, S.A., Blanc, M.B., Viani, R., 1990. Chromatographic profile of carbohydrates in commercial coffees. 2. Identification of mannitol. Journal of Agricultural and Food Chemistry 38, 1347– 1350. Dart, S.K., Nursten, H.E., 1985. Volatile components. In: Clarke, R.J., Macrae, R. (Eds.), Coffee: vol. 1, Chemistry. Elsevier Applied Science, London, pp. 223– 265. De Maria, C.A.B., Trugo, L.C., Aquino Neto, F.R., Moreira, R.F.A., Alviano, C.S., 1996. Composition of green coffee water-soluble fractions and identification of volatiles formed during roasting. Food Chemistry 55, 203–207. Dutra, E.R., Oliveira, L.S., Franca, A.S., Ferraz, V.P., Afonso, R.J.C.F., 2001. A preliminary study on the feasibility of using the composition of coffee roasting
R.C.S. Oliveira et al. / Journal of Food Composition and Analysis 22 (2009) 257–261 exhaust gas for the determination of the degree of roast. Journal of Food Engineering 47, 241–246. Fontes, A.S., Bento, A.C., Miranda, L.C.M., Baesso, M.L., 2001. Thermal lens evaluation of the presence of adulterants in brewed coffee. Analytical Sciences 17, s526– s529. Franca, A.S., Oliveira, L.S., Oliveira, R.C.S., Mancha Agresti, P., Augusti, R., 2008. A preliminary evaluation of the effect of processing temperature on coffee roasting degree assessment, Journal of Food Engineering, in press, accepted manuscript, available online 25 December 2008, doi:10.1016/j.jfoodeng.2008.12.012. Fugel, R., Carle, R., Schieber, A., 2005. Quality and authenticity control of fruit pure´es, fruit preparations and jams – a review. Trends in Food Science & Technology 16, 433–441. Godinho, A., Oliveira, L.S., Ferraz, V.P., Franca, A.S., 2003. Detecc¸a˜o e quantificac¸a˜o de impurezas em cafe´ torrado e moı´do (Detection and quantification of contaminants in roasted and ground coffee). Revista Brasileira de Armazenamento, Especial Cafe´ 7, 50–55 (in portuguese). Mancha Agresti, P.C.M., Franca, A.S., Oliveira, L.S., Augusti, R., 2008. Discrimination between defective and non-defective Brazilian coffee beans by their volatile profile. Food Chemistry 106, 787–796.
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Oliveira, S.D., Franca, A.S., Glo´ria, M.B.A., Borges, M.L.A., 2005. The effect of roasting on the presence of bioactive amines in coffees of different qualities. Food Chemistry 90, 287–291. Prodolliet, J., Bruelhart, M., Blanc, M.B., Leloup, V., Cherix, G., Donnely, C.M., Viani, R., 1995. Adulteration of soluble coffee with coffee husks and parchments. Journal of AOAC International 78, 761–767. Rocha, S., Maeztu, L., Barros, A., Cid, C., Coimbra, M.A., 2003. Screening and distinction of coffee brews based on headspace solid phase microextraction/ gas chromatography/principal component analysis. Journal of the Science of Food and Agriculture 84, 43–51. Sanz, C., Maeztu, L., Zapelena, M.J., Bello, J., Cid, C., 2002. Profiles of volatile compounds and sensory analysis of three blens of coffee: influence of different proportions of Arabica and Robusta and influence of roasting coffee with sugar. Journal of the Science of Food and Agriculture 82, 840–847. Singhal, R.S., Kulkarni, P.R., Rege, D.V., 1997. Handbook of Indices of Food Quality and Authenticity. Woodhead Publishing Ltd., Cambridge. Zambonin, C.G., Balest, L., De Benedetto, G.E., Palmisano, F., 2005. Solid-phase microextraction-gas chromatography mass spectrometry and multivariate analysis for the characterization of roasted coffees. Talanta 66, 261–265.
Contents lists available at ScienceDirect
Journal of Food Composition and Analysis journal homepage: www.elsevier.com/locate/jfca
Short Communication
Evaluation of the potential of SPME-GC-MS and chemometrics to detect adulteration of ground roasted coffee with roasted barley Rafael C.S. Oliveira a,*, Leandro S. Oliveira a, Adriana S. Franca a, Rodinei Augusti b a b
Programa de Po´s-Graduac¸a˜o em Cieˆncia de Alimentos/UFMG, Av. Antoˆnio Carlos, 6627, 31270-901, Belo Horizonte, MG, Brazil Departamento de Quı´mica/UFMG, Av. Antoˆnio Carlos, 6627, 31270-901, Belo Horizonte, MG, Brazil
A R T I C L E I N F O
A B S T R A C T
Article history: Received 24 June 2008 Received in revised form 29 September 2008 Accepted 20 October 2008
The objective of the present study was to verify the feasibility of detection of coffee adulteration with roasted barley by a comparative analysis of the volatile profiles of both coffee and barley, pure and mixed, at several roasting degrees. The methodology was based on a GC-MS analysis of the headspace volatiles of several samples of ground roasted coffee and barley. The collection and concentration of the headspaces was performed by solid phase micro-extraction (SPME). The separation of the nonadulterated and adulterated samples was accomplished by application of principal component analysis (PCA) to the chromatographic data obtained. It was observed that, the highest the degree of roast, the more easily discriminated were the adulterated samples, allowing for detection of adulterations with as low as 1% (w/w) roasted barley in dark roasted coffee samples. ß 2009 Elsevier Inc. All rights reserved.
Keywords: Coffee adulteration Fraud Gas chromatography GC Mass spectrometry MS Solid-phase micro-extraction SPME Multivariate statistics Detection and quantification method Food analysis Food quality Food composition
1. Introduction Although detection of adulteration of food and food products was reported as early as the 18th century (Singhal et al., 1997), the increasing number of studies on the development of analytical tools for the detection and quantification of adulterations of food and food products in published works in recent years has demonstrated that this type of fraud has become a common practice in all food sectors all around the world (Cordella et al., 2002; Fugel et al., 2005). Adulteration is a serious problem since it promotes exploitation of consumers by non-compliance with product specifications and with regulatory standards (with consequent deterioration of product quality). Also, it leads to unfair competition, disrupting otherwise stabilized local and global economies, with defrauders gaining over reputable competitors. Adulteration with intent to deceive is usually carried out by admixture of cheaper products and materials which are usually difficult to detect by consumers and by simple routine analytical
* Corresponding author. Tel.: +55 31 91648536; fax: +55 31 34433783. E-mail address: [email protected] (Rafael C.S. Oliveira). 0889-1575/$ – see front matter ß 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.jfca.2008.10.015
techniques, with high-priced commodities being usually the main target for adulteration. Brazil is the largest coffee producer in the world and is also among the largest consumers. Coffee is the main commodity in Brazil and, as such, has been the target of fraudulent admixtures with a diversity of cheaper materials, such as twigs, coffee berry skin and parchment, spent coffee, roasted barley, maize, cocoa, soybean and others (Singhal et al., 1997). As a product of consumption, ground roasted coffee is quite vulnerable to adulteration since it presents physical characteristics such as particle size, texture and color, which are easily reproduced by roasting and grinding a variety of biological materials (cereals, seeds, roots, parchments, etc.). A plethora of techniques has been developed in order to establish suitable parameters and markers for adulterations of ground roasted coffee and instant or soluble coffee. Most of the analytical techniques were based on determination of the carbohydrate chromatographic profiles coupled to multivariate statistical analysis of chromatographic data (Blanc et al., 1989; Davis et al., 1990; Berger et al., 1991; Prodolliet et al., 1995; Bernal et al., 1996), and the overall conclusion was that it is possible to establish quantifiable limits for the presence of specific carbohydrates in the products (e.g. xylose, glucose and
R.C.S. Oliveira et al. / Journal of Food Composition and Analysis 22 (2009) 257–261
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fructose) above which the product can be safely considered adulterated. Techniques that do not rely on the determination of specific chemical tracers, but rather on strictly physical measurements, were also proposed for the detection of adulterants in coffee, e.g. the use of photoacoustic spectroscopy by Cesar et al. (1984), of infra-red spectroscopy by Briandet et al. (1996), of scanning electron microscopy by Amboni et al. (1999), and of thermal lens spectrometry by Fontes et al. (2001). The aforementioned techniques do not allow for the identification of individual constituents or constituents in admixtures of several adulterants. In the case of roasted coffee, adulterations can usually be perceived by a trained sensory panel, thus, indicating that an analysis of the volatiles profile, which is expected to be singular for a particular adulterant, could provide a reliable method to verify adulteration of ground roasted coffee. Among the commonly used contaminants, barley is particularly difficult to detect especially at low concentrations, and even methodologies that are based on the chemical profile are not effective for this specific contaminant. The study by Godinho et al. (2003) presented a methodology based on solvent extraction followed by HPLC analysis for detection of coffee adulteration by barley and coffee husks. Results were satisfactory for the detection of coffee husks, but the chromatographic profiles of the extracts obtained for coffee and barley were quite similar, with the exception of the absence of caffeine in the chromatograms obtained for barley. Such difference would be helpful only if roasted barley were the product being adulterated, and hence it was concluded that the proposed methodology was not effective for detection of adulteration of coffee by barley. Thus, it was the aim of this work to verify the feasibility of detecting coffee adulteration with roasted barley, based on GC-MS analysis of the headspace volatiles of several samples of ground roasted coffee and barley, including tentative identification of chemical markers and multivariate statistical analysis of the chromatographic data. 2. Materials and methods Good quality crude coffee and crude barley samples were obtained from local markets. Both crude coffee and crude barley were roasted to three distinct degrees of roast in a laboratory oven at the same temperature. Preliminary tests showed that it would take temperatures higher than 200 8C to promote significant color changes in crude barley, at reasonable periods of processing time, to be considered roasted to degrees comparable to those for coffee. Hence, a roasting temperature of 300 8C was selected for the laboratory roasting tests. The degrees of roast were characterized as light, medium, and dark according to industrial color standards (Table 1). The roasting experiments were performed in duplicates. Samples were weighted before and after roasting in order to evaluate weight loss. Just before each analysis, the coffee and barley samples were ground with an electric coffee grinder (C-mill 5679-01US, Bodum, USA) for 30 s.
An SPME triple phase (divinylbenzene/carboxen/polydimethylsiloxane) 50/30 mm fiber (model 57348-U, Supelco Inc., USA) was employed for the extraction of volatiles from the roasted coffee and barley headspace, according to the methodology described by Mancha Agresti et al. (2008). Approximately 3 mL of each sample (ground coffee, barley and coffee/barley mixtures) was placed in sealed 5 mL vials (Supelco Inc., USA) and heated for 10 min at 70 8C. Afterwards the SPME needle was inserted into the vial and the fiber was exposed to the headspace above the sample for 40 min at 70 8C. After sampling, the fiber was thermally desorbed in the GC injection port for 10 min at 250 8C. GC analyses were performed in duplicates using a gas chromatograph (Trace Ultra) coupled to a mass spectrometer (PolarisQ) (ThermoElectron, San Jose, CA), a RTX-5MS column (5% diphenyl, 95% dimethyl polysiloxane) 30 m 0.25 mm I.D (Restec, Ireland) and Helium as carrier gas (1 mL/min). The injector and detector temperatures were set to 250 8C and 300 8C, respectively. The GC oven temperature was programmed from 40 8C (5 min) to 180 8C at the rate of 3 8C/min, then to 250 8C (5 min) at 10 8C/min. The GC injector was operated in the splitless mode. Mass spectra were acquired in the electron impact mode at 70 eV and using an m/z range of 50–650 and a scan time of 2 s. Tentative identification of compounds was performed by comparison of the mass spectra obtained for the peaks in the analysis with those of the NIST/EPA/NIH Mass Spectral Library, version 2.0 (available in the instrument software), considering a similarity level (RSI) higher than 800. For principal component analysis, data matrices were assembled so that the rows corresponded to roasted coffee and/ or barley samples and each column represented the peak area of a specific component. The chromatographic peaks with S/N (signalto-noise ratio) higher than 50 were included in the final data matrix regardless of substance identification. This choice was based on the extensive number of peaks detected (over 250). Some preliminary tests with smaller S/N ratio values indicated that the amount of detected substances would be prohibitive and increase the degree of redundancy in terms of multivariate statistical analysis, since most of them were present in similar amounts in all chromatograms for both roasted coffee and roasted barley. 3. Results and discussion Results for weight loss during roasting are shown in Fig. 1, with the solid lines representing linear fits. Both coffee and barley presented the same qualitative behavior for weight loss during roasting, being characterized by a slower rate at the beginning of roasting, followed by an increase in weight loss rate towards the end of the process. This behavior was satisfactorily described by two straight lines (r2 > 0.96) representing the two different rates, and has been reported by previous studies on coffee roasting (Dutra et al., 2001; Oliveira et al., 2005). The first period,
Table 1 Roasting conditions (roasting temperature: 300 8C). Sample ID Coffee CL CM CD Barley BL BM BD
Roasting degree
Roasting time (min)
light medium dark
9 10 11
light medium dark
16 17 18
Fig. 1. Weight loss during roasting.
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corresponding to the slower weight loss rate, is called drying stage, and it is due mainly to the slow release of water and volatiles from crude coffee and barley. The onset of pyrolysis can be associated with the transition between the two slopes, with the increase in weight loss rate being attributed to an intensive release of volatile organic compounds and CO2. A comparison of the weight loss curves for coffee and barley shows that the weight loss rates during both the drying and the pyrolysis stages were slower for barley in comparison to those for coffee. However, average weight loss values for a given roasting degree (strictly based on color) were higher for barley than for coffee, indicating that more severe roasting conditions are required for roasted barley to attain a similar color to that of coffee at lighter degrees of roast. Such differences could be attributed to the difference in chemical changes occurring within the barley grains during roasting. These differences indicate that, if one is to adulterate roasted coffee with roasted barley without significant alterations in roasted color, the roasting of each product should be carried out separately, since, for the same roasting conditions (time and temperature), barley will either roast to a much lighter color or not roast at all (for shorter times and lower temperatures). Also, as discussed by Cristo et al. (2006) in their study of flow regimes of coffee beans in rotating roasters, smaller particles will tend to migrate to the low velocity core of the bed transverse section. Thus, due to poor mixing (low heat transfer rates) in that region, these particles will necessarily roast to a lesser degree, which would be the case of smaller barley grains in a mixture with larger coffee beans. Given the extensive number of compounds that were detected in all roasted coffee and roasted barley samples, multivariate statistical analysis (PCA) was performed in order to verify the possibility of discrimination between roasted coffee and roasted barley based on the volatile profiles. The biplot of the coffee and barley samples submitted to roasting at 300 8C is displayed in Fig. 2. Regarding the volatile profiles of roasted coffee in comparison to roasted barley, the first two principal components (PCs) explained 93.8% and 2.4% of the chromatographic variance, respectively. A clear separation between roasted coffee and roasted barley could be obtained based on the first component alone. The substances that presented higher influence on PC1 and PC2, based on the projection of score plot, are displayed in Table 2. Pyridine (RT 4 min) was the substance with the highest influence on PC1 for roasted coffee and the relative peak intensity increased with roasting time. Pyridine formation during coffee roasting can be attributed to protein pyrolysis, trigonelline degradation, Strecker degradation and Maillard reactions (De Maria et al., 1996). Also, the
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Fig. 2. PCA scores scatter plot of normalized chromatographic SPME areas of pure coffee and barley submitted to roasting at 300 8C (PC1 vs PC2). C = coffee, B = barley, L = light roast, M = medium roast, D = dark roast.
amount of pyridine depends on the degree of roast, and this compound has been reported to be characteristic of darker roasts (Dart and Nursten, 1985; Franca et al., 2008). The other substances listed in Table 2 as characteristic of roasted coffees have been identified by previous studies in the headspace of roasted coffee (De Maria et al., 1996; Sanz et al., 2002; Rocha et al., 2003; Zambonin et al., 2005; Franca et al., 2008). The two aldehydes that were identified as characteristic of the volatile profile of barley were also detected in the roasted coffee headspace, but in much lower concentrations. Admixtures of roasted barley with roasted coffee samples were intentionally prepared and their respective volatile profiles analyzed in order to verify the capability of discrimination between adulterated and non-adulterated roasted coffee samples. The biplots for the headspace volatiles of admixtures of 1%, 5% and 50% roasted barley in roasted coffee are presented in Fig. 3. These results show that it was possible to detect coffee adulteration with roasted barley in admixtures with as low as 1% (w/w) roasted barley, but only for the highest (darker) degrees of roast. This does not pose a problem, given that adulterated samples are usually submitted to severe roasting conditions (dark roasts), so that sensory perception of the adulteration becomes more difficult. Also, as the amount of barley increased, the separation became more evident. The PCA scores of the headspace profiles for the admixtures with roasted barley are closer to those for non-
Table 2 Tentative indentification of aroma components with the most pronounced effect on PC1 and PC2 values. Retention time (min)
3.98 6.35 8.50 8.65 10.36 13.02 14.75 29.86 6.92 12.95 17.54 22.53 27.33 3.77 14.34 *
Compound Roasted Coffee pyridine 2-methyl pyrimidine* 2- furanmethanol 3-furanmethanol* 2,6-dimethyl pyrazine 5-methyl-2-furancarboxaldehyde 2-furanmethanol, acetate 4-hidroxy-3-methylacetophenone Roasted barley 3-furaldehyde 5-methyl-2-furancarboxaldehyde* Butyl-Benzene Pentyl-Benzene Hexyl-Benzene Coffe/barley admixtures Ethanedinitrile 2-furanmethanol, acetate
Detected by Franca et al. (2008) on volatile profiles of roasted coffee at 200 8C.
m/z of the most intense ions (relative abundance %)
79 (100), 80 (85), 52 (50) 94 (100), 67 (40), 95 (40) 81 (100), 98 (70), 97 (60) 81 (100), 98 (55), 97 (50) 108 (100), 109 (30), 107 (20) 109 (100), 110 (50), 53 (30) 98 (100), 81 (80), 53 (40) 150 (100), 135 (70), 107 (45) 95 (100), 96 (30), 67 (10) 109 (100), 110 (50), 53 (30) 91 (100), 92 (60), 134 (30) 91 (100), 92 (55), 148 (30) 91 (100), 92 (55), 162 (20) 53 (100), 58 (5), 85 (5) 98 (100), 81 (90), 53 (40)
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Fig. 3. PCA scores scatter plot of normalized chromatographic SPME areas of (a) 1%, (b) 10%, and (c) 50% roasted barley in roasted coffee in comparison to pure coffee and pure barley. C = coffee, B = barley, M1 = 1:99 barley coffee mixture; M10 = 10:90 barley coffee mixture; M50 = 50:50 barley coffee mixture; L = light roast, M = medium roast, D = dark roast.
adulterated coffee samples due to the fact that roasted coffee headspace profiles present higher amounts of discriminating compounds than the headspace for roasted barley. The preliminary results obtained in the present study indicate that SPME-GC-MS is a promising detection technique to be employed for detection of coffee fraud. However, further studies regarding different roasting conditions and contaminants are necessary to assure the applicability of the proposed methodology to a wider range of roasting conditions and also for simultaneous detection of different contaminants in the same sample. 4. Conclusions SPME-GC-MS was successfully employed for the detection of adulteration of ground roasted coffee with roasted barley. The methodology allowed for the detection of adulterations with as low as 1% (w/w) roasted barley in roasted coffee samples, for the darkest degrees of roast. It was observed that the higher the amount of adulterant, the more effective was the methodology in discriminating adulterated from non-adulterated samples, as expected. Also, the higher the degree of roast, the more easily discriminated were the adulterated samples. It was possible to tentatively identify volatile compounds in the headspace profile of roasted barley to be further used as chemical markers in the detection of adulteration of roasted ground coffee with roasted barley. Acknowledgements The authors acknowledge financial support from the following Brazilian Government Agencies: CAPES, CNPq and FAPEMIG.
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