Abstract
One component that contribute to the flavor and aroma of chocolate are the polyphenols, which have received much attention due to their beneficial implications to human health. Besides bioactive action, polyphenols and methylxantines are responsible for astringency and bitterness in cocoa beans. Another important point is its drastic reduction during cocoa processing for chocolate production and the difference between cultivars. Thus, the present study aimed to evaluate the modifications in monomeric phenolic compounds and methylxanthines during fermentation of three cocoa cultivars grown in southern Bahia. Cocoa beans from three cultivars were fermented and sun dried and monomeric phenolic compounds and methylxantines were determinated. The results showed that each cultivar have different amounts of phenolic compounds and the behaviour of them is different during fermentation. The amount of methylxantines varied but there was not a pattern for methylxantines behavior during process. In addition a huge reduction in phenolic compounds could be observed after drying. Differently of phenolic compounds, methylxantines did not have great modification after sun drying. So, the differences observed in this study between cultivars, take to the conclusion that the compounds studied in those cocoa cultivars have different behavior during fermentation and drying, which consequently, give to these cultivars differences in sensory characteristics.
cocoa; fermentation; polyphenols; methylxantines
1 Introduction
Beyond the “witches’ broom disease”, caused by the fungus Moniliophthora
perniciosa, which caused a great reduction in cocoa production in
southern Bahia, other problem for cocoa treading is the low quality of the beans
produced due to the poor control of fermentation and drying, obtaining a significant
amount of beans with undesirable characteristics, that will not develop the
characteristic flavor for high quality chocolate production (Cruz et al., 2013Cruz, J. F. M., Leite, P. B., Soares, S. E., & Bispo, E. S.
(2013). Assessment of the fermentative process from different cocoa cultivars
produced in Southern Bahia, Brazil. African Journal of Biotechnology, 12(33),
5218-5225. http://dx.doi.org/10.5897/AJB2013.12122.
http://dx.doi.org/10.5897/AJB2013.12122...
).
The cocoa pre-processing steps (harvest, breaking, fermentation and drying) are
important to ensure the high quality of the beans. For Lagunes-Gálvez et al., (2007)Lagunes Gálvez, S., Loiseau, G., Paredes, J. L., Barel, M., &
Guiraud, J. P. (2007). Study on the microflora and biochemistry of cocoa
fermentation in the Dominican Republic. International Journal of Food
Microbiology, 114(1), 124-130.
http://dx.doi.org/10.1016/j.ijfoodmicro.2006.10.041.
PMid:17187887
http://dx.doi.org/10.1016/j.ijfoodmicro....
fermentation is one of the
post-harvest step that most affect the quality of the products obtained from
cocoa.
During fermentation, the pulp is degraded by the successive action of microorganisms,
promoting the rise of temperature to about 50 °C and the formation of ethanol and
lactic acid, driving to the embryo death, which creates an environment for formation
of flavor and aroma precursors of chocolate, developed through roasting process
(Lopez, 1986Lopez, A. S. (1986). Chemical changes occurring during the
processing of cacao. In Proceedings of the Symposium Cacao Biotechnology,
Pennsylvania (pp. 19-54). Pennsylvania: University Park, The Pennsylvania State
University. ; Leite et al., 2013Leite, P. B., Maciel, L. F., Opretzka, L. C. F., Soares, S. E.,
& Bispo, E. S. (2013). Phenolic compounds, methylxantines and antioxidant
activityin cocoa massand chocolates produced from “witch broom disease”
resistant and non resistant cocoa cultivars. Ciência e Agrotecnologia, 37(3),
244-250. http://dx.doi.org/10.1590/S1413-70542013000300007.
http://dx.doi.org/10.1590/S1413-70542013...
).
Other components that contribute to the flavor and aroma of chocolate are the
polyphenols, which have received much attention due to their beneficial implications
to human health by preventing cancer, cardiovascular diseases and other pathologies
(Keen et al., 2005Keen, C. L., Holt, R. R., Oteiza, P. I., Fraga, C. G., &
Schmitz, H. H. (2005). Cocoa antioxidants and cardiovascular health. The
American Journal of Clinical Nutrition, 81(1, Suppl.), 298S-303S.
PMid:15640494.; Hermann et al., 2006Hermann, F., Spieker, L. E., Ruschitzka, F., Sudano, I., Hermann,
M., Binggeli, C., Lüscher, T. F., Riesen, W., Noll, G., & Corti, R. (2006).
Dark chocolate improves endothelial and platelet function. Heart, 92(1),
119-120. http://dx.doi.org/10.1136/hrt.2005.063362.
PMid:16365364
http://dx.doi.org/10.1136/hrt.2005.06336...
; Cooper et al., 2008Cooper, K. A., Donovan, J. L., Waterhouse, A. L., & Williamson,
G. (2008). Cocoa and health: a decade of research. The British Journal of
Nutrition, 99(1), 1-11. http://dx.doi.org/10.1017/S0007114507795296.
PMid:17666148
http://dx.doi.org/10.1017/S0007114507795...
). In this group of compounds, catechin and
epicatechin are the most importante, since they are easily absorbed by the
intestinal walls and brought the bloodstream, presenting a great potential for
action in the body (Richelle et al., 1999Richelle, M., Tavazzi, I., Enslen, M., & Offord, E. A. (1999).
Plasma kinetics in man of epicatechin from black chocolate. European Journal of
Clinical Nutrition, 53(1), 22-26. http://dx.doi.org/10.1038/sj.ejcn.1600673.
PMid:10048796
http://dx.doi.org/10.1038/sj.ejcn.160067...
).
Besides bioactive action, polyphenols are responsible for the development of
astringency and bitterness in cocoa beans, followed by alkaloids, some amino acids,
peptides and pyrazines (Bonvehi & Coll,
2000Bonvehi, J. S., & Coll, F. V. (2000). Evaluation of purine
alkaloids anddiketopiperazines contents in processed cocoa powder. European Food
Research and Technology, 210(3), 189-195.
http://dx.doi.org/10.1007/PL00005510.
http://dx.doi.org/10.1007/PL00005510...
).
Another important point to be observed for these compounds is its drastic reduction
during cocoa processing for chocolate production. This reduction occurs during
fermentation, drying and roasting. Furthermore, the concentration varies depending
on the cultivar used and also the type of chocolate produced. Chocolates with higher
concentrations of cocoa have higher levels of polyphenols (Miller et al., 2006Miller, K. B., Stuart, D. A., Smith, M. L., Lee, C. Y., McHale, N.
L., Flanagan, J. A., Ou, B., & Hurst, W. J. (2006). Antioxidant activity and
polyphenol and procyanidin contents of selected commercially available cocoa
containing and chocolate products in the United States. Journal of Agricultural
and Food Chemistry, 54, 4062-4068.; Payne et
al., 2010Payne, M. J., Hurst, W. J., Miller, K. B., Rank, C., & Stuart,
D. A. (2010). Impact of fermentation, drying, roasting, and Dutch processing on
epicatechin and catechin content of cacao beans and cocoa ingredients. Journal
of Agricultural and Food Chemistry, 58(19), 10518-10527.
http://dx.doi.org/10.1021/jf102391q. PMid:20843086
http://dx.doi.org/10.1021/jf102391q...
).
During fermentation, the polyphenol content decreases about 70%, and the (-) -
epicatechin, the main substrate of polyphenoloxidase, suffers a reduction of 90% of
its initial concentration. At the same time, catechins form complex tannins and
anthocyanins are hydrolysed to anthocyanins which polymerise themselves (Efraim et al., 2011Efraim, P., Alves, A. B., & Jardim, D. C. P. (2011). Polifenóis
em cacau e derivados: teores, fatores de variação e efeitos na saúde. Brazilian
Journal of Food Technology, 14(3), 181-201.
http://dx.doi.org/10.4260/BJFT2011140300023.
http://dx.doi.org/10.4260/BJFT2011140300...
).
Methylxanthines, such as caffeine and theobromine are another group of bioactive
compounds also found in cocoa beans. While caffeine has a stimulating effect on the
brain, increasing the psychomotor reactions and blood pressure, the theobromine acts
as a muscle relaxant, diuretic and blood pressure reducer (Van den Bogaard et al., 2010Van den Bogaard, B., Draijer, R., Westerhof, B. E., Van den
Meiracker, A. H., Van Montfrans, G. A., & Van den Born, B. J. (2010).
Effects on peripheral and central blood pressure of cocoa with natural or
high-dose theobromine: a randomized, double-blind crossover trial. Hypertension,
56(5), 839-846. http://dx.doi.org/10.1161/HYPERTENSIONAHA.110.158139.
PMid:20823377
http://dx.doi.org/10.1161/HYPERTENSIONAH...
; Bruinsma & Taren, 1999Bruinsma, K., & Taren, D. L. (1999). Chocolate: food or drug?
Journal of the American Dietetic Association, 99(10), 1249-1256.
http://dx.doi.org/10.1016/S0002-8223(99)00307-7. PMid:10524390
http://dx.doi.org/10.1016/S0002-8223(99)...
).
The theobromine has low detection threshold (10 mg.L–1), which contributes
to generate a bitter taste to foods. After roasting the beans, theobromine can form
chemical adducts with diketopiperazine, which have some relationship with the bitter
notes attributed to roasted cocoa (Pickenhagen et
al., 1975Pickenhagen, W., Dietrich, P., Keil, B., Polonsky, J., Nouaille, F.,
& Lederer, E. (1975). Identification of the bitter principle of cocoa.
Helvetica Chimica Acta, 58(4), 1078-1086.
http://dx.doi.org/10.1002/hlca.19750580411. PMid:1158737
http://dx.doi.org/10.1002/hlca.197505804...
).
In order to achieve the knowledge of dynamics involving the compounds that may influence the sensory characteristics of chocolate during cocoa pre-processing, the present study aimed to evaluate modifications in monomeric phenolic compounds and methylxanthines amounts during fermentation of three cocoa cultivars grown in southern Bahia.
2 Materials and methods
2.1 Cocoa beans
Cocoa beans from cultivars resistant to “witches’ broom disease”, “SR162” and “PH16”, and a blend of cultivars, composed by Pará, Parazinho and Maranhão, all of them belonging to Forastero group, no resistant to the disease were used. These Forastero cultivars is grown from decades in Bahia, considered cultivars of high productivity and high fruit quality, however, no resistant to the disease (Pinto & Pires, 1998Pinto, L. R. M., & Pires, J. L. (1998). Seleção de Plantas de Cacau Resistentes à vassoura-de-bruxa (Boletim Técnico, No. 181). Ilhéus: CEPLAC/CEPEC.).
The “PH16” is a cultivar originated from a selection made in a commercial area. They have unknowing parents and the plant was originally identified in 1996 in a population of cocoa hybrids (crosses between Amazônico group and Trinitario) from “Porto Híbrido” farm, in São José da Vitória County, Bahia, Brazil.
The “SR162” is a cultivar derived from genetic mutation of the common cocoa (Alto Amazônico group), with white seeds. The name came from the farm where there were identified, “São Roque” farm, in Itagibá County, Bahia, Brazil.
2.2 Fermentation and drying process
The fermentation was carried out in 70 × 70 × 75cm wooden boxes, with around 20 holes with 1.27cm each in the bottom and sides of the boxes, in order to flow the liquefied pulp. A total mass of 400 kg in each box were processed. Turnings were performed for oxygenation and mixing of the mass, 48 hours from the beginning of each fermentation and after the decreasing of temperature, until the end of the process. To cover the mass, banana leaves were used. After fermentation, the seeds were dried in the sun-roof surfaces with mobile timber for 5 to 7 days up to 8.0% of moisture.
2.3 Samples collection and data treatment
Samples from two fermentations were taken at the beginning and after 72, 108 and 120 hours. Cocoa beans were collected from the surface, middle and bottom of the boxes, mixed and freeze dried in a LIOTOP Freeze Dryer, L108 model in order to perform the analysis. Data were subjected to analysis of variance (oneway ANOVA) using the Tukey test at 5% significance for means comparison .
2.4 Extraction of Phenolic Compounds and Methylxanthines
Methanolic extracts were obtained according to Fantozzi & Montedoro (1978)Fantozzi, P., & Montedoro, G. (1978). Dosage dês composés phénoliques dans drupes d’olives récoltés à differents stades de maturation. Industrie Alimentaire et Agricole, 17, 1335-1339.. Ten grams of cocoa mass and chocolate were weight and defatted with petroleum ether, under shacking during 30 minutes for three times. From defatted samples, five grams were weight and 80% methanol in water (v/v) solution were added and stirring during one hour. After filtration, the methanolic extracts were stored in dark flask under nitrogen atmosphere at low temperature.
2.5 Determination of monomeric phenols and methylxanthines by HPLC
The determination of monomeric phenolic compounds (galic acid, catechin, caffeic
acid and epicatechin) and methylxantines (caffeine and theobromine) was
performed according to the method described by Elwers et al. (2009)Elwers, S., Zambrano, A., Rohsius, C., & Lieberei, R. (2009).
Differences between the content of phenolic compounds in Criollo, Forastero and
Trinitario cocoa seed (. Theobroma cacao L.)European Food
Research and Technology, 229(6), 937-948.
http://dx.doi.org/10.1007/s00217-009-1132-y.
http://dx.doi.org/10.1007/s00217-009-113...
. Ten microlitres of each sample solution was
analyzed by HPLC system (Perkin Elmer Model Flexar) equipped with VI Flow
injector, C18 column (100 mm × 4.6 mm O.D.S.-2, 3 μm) and using the
solvents (A): 2% acetic acid in water and (B): a mixture of acetonitrile, water
and acetic acid (400:90:10 v/v/v). Elution was performed with a linear gradient
showed in Table 1. The compounds were
monitored by UV detection at 280 nm wavelength. The total run time was 20 min
and the temperature was 26 °C. All standards used for quantitative
determinations were from Sigma-Aldrich, St. Louis, MO.
HPLC gradient used for separation of phenolic compounds and methylxanthines in cocoa mass and chocolates.
3 Results and discussion
3.1 Phenolic compounds and methylxantinesin cocoa beans during fermentation process
In Table 2 are showed the phenolic
profile during fermentation of three cocoa cultivars. First of all, it can be
observed that the cultivar have different amounts of phenolic compounds. The
non-resistant blend have the highest amount of phenolic compounds in the
beginning of fermentation process, 18.9mg.g–1, while PH16 have
15.3mg.g–1 and SR162 5.7mg.g–1. It was already
expected, since non-resistant cultivars were composed by healthy and “witches’
broom disease” infected fruits and the fungal infection enhance the synthesis of
phenolic compounds (Soares, 2002Soares, S. E. (2002). Phenolic acids as antioxidants. Revista de
Nutrição, 15, 71-81.). When
a plant is attacked by a potential pathogen, there is activation of the defense
responses complex, with expression of various genes, resulting in synthesis and
accumulation of secondary metabolites such as phenolic compounds (Leite et al., 2013Leite, P. B., Maciel, L. F., Opretzka, L. C. F., Soares, S. E.,
& Bispo, E. S. (2013). Phenolic compounds, methylxantines and antioxidant
activityin cocoa massand chocolates produced from “witch broom disease”
resistant and non resistant cocoa cultivars. Ciência e Agrotecnologia, 37(3),
244-250. http://dx.doi.org/10.1590/S1413-70542013000300007.
http://dx.doi.org/10.1590/S1413-70542013...
). On the other hand the
SR-162 cultivar have the lowest levels of monomeric phenolic compounds, showing
that this cultivar is not only poor in anthocyanins, but also in other phenolic
compounds.
Phenolic compounds amounts (mg.g-1) during fermentation of three cocoa cultivars* * The values shown are the average of 2 fermentation replicates and expressed in dry defatted basis. .
Other point to be observed is the predominance of (-) epicatechin in all
cultivars when fermentation started, and once more, the non-resistant blend
showed highest levels of this compound, 16.3mg.g–1, while for SR162
and PH16 were 3.9mg.g–1 and 13.7mg.g–1, respectively. Many
authors reported the highest levels of (-) epicatechin in cocoa beans, mass and
chocolate (Miller et al., 2006Miller, K. B., Stuart, D. A., Smith, M. L., Lee, C. Y., McHale, N.
L., Flanagan, J. A., Ou, B., & Hurst, W. J. (2006). Antioxidant activity and
polyphenol and procyanidin contents of selected commercially available cocoa
containing and chocolate products in the United States. Journal of Agricultural
and Food Chemistry, 54, 4062-4068.; Payne et al., 2010Payne, M. J., Hurst, W. J., Miller, K. B., Rank, C., & Stuart,
D. A. (2010). Impact of fermentation, drying, roasting, and Dutch processing on
epicatechin and catechin content of cacao beans and cocoa ingredients. Journal
of Agricultural and Food Chemistry, 58(19), 10518-10527.
http://dx.doi.org/10.1021/jf102391q. PMid:20843086
http://dx.doi.org/10.1021/jf102391q...
; Leite et al., 2013Leite, P. B., Maciel, L. F., Opretzka, L. C. F., Soares, S. E.,
& Bispo, E. S. (2013). Phenolic compounds, methylxantines and antioxidant
activityin cocoa massand chocolates produced from “witch broom disease”
resistant and non resistant cocoa cultivars. Ciência e Agrotecnologia, 37(3),
244-250. http://dx.doi.org/10.1590/S1413-70542013000300007.
http://dx.doi.org/10.1590/S1413-70542013...
).
In addition, the non-resistant blend showed more varied phenolic profile with
small amounts of galic and caffeic acids, followed by SR162 that had galic acid
and PH16 that had just catechin and epicatechin. This fact can occur since
factors as genetic, soil, growing conditions and weather characteristics can
affect the profile and concentration of these compounds (Niemenak et al., 2006Niemenak, N., Rohsius, C., Elwers, S., Ndoumou, D. O., &
Lieberei, R. (2006). Comparative study of different cocoa (Theobroma cacao L.)
clones in terms of their phenolics and anthocyanins contents. Journal of Food
Composition and Analysis, 19(6-7), 612-619.
http://dx.doi.org/10.1016/j.jfca.2005.02.006.
http://dx.doi.org/10.1016/j.jfca.2005.02...
; Meng et al., 2009Meng, C. C., Jalil, A. M. M., & Ismail, A. (2009). Phenolic and
theobromine contents of commercial dark, milk and white chocolates on the
Malaysian market. Molecules (Basel, Switzerland), 14(1), 200-209.
http://dx.doi.org/10.3390/molecules14010200. PMid:19127248
http://dx.doi.org/10.3390/molecules14010...
; Elwers et al.,
2009Elwers, S., Zambrano, A., Rohsius, C., & Lieberei, R. (2009).
Differences between the content of phenolic compounds in Criollo, Forastero and
Trinitario cocoa seed (. Theobroma cacao L.)European Food
Research and Technology, 229(6), 937-948.
http://dx.doi.org/10.1007/s00217-009-1132-y.
http://dx.doi.org/10.1007/s00217-009-113...
; Leite et al.,
2013Leite, P. B., Maciel, L. F., Opretzka, L. C. F., Soares, S. E.,
& Bispo, E. S. (2013). Phenolic compounds, methylxantines and antioxidant
activityin cocoa massand chocolates produced from “witch broom disease”
resistant and non resistant cocoa cultivars. Ciência e Agrotecnologia, 37(3),
244-250. http://dx.doi.org/10.1590/S1413-70542013000300007.
http://dx.doi.org/10.1590/S1413-70542013...
).
A significant reduction in (-) epicatechin was observed for non-resistant and PH16 cultivars during fermentation. At the end of the process, the reduction observed for non-resistant cultivar was 58% and for PH16 was 48%. Gil et al. (2011)Gil, A., Rojas, L. F., Atehortua, L., & Londono, J. (2011). Effect of Fermentation and Sun Drying on Phytochemical Composition of Native Colombian Cocoa. In Proceedings of the 6th CIGR Section VI International Symposium “Towards a Sustainable Food Chain” Food Process, Bioprocessing and Food Quality Management, Nantes, France. described higher reduction of epicatechin (80%) in colombian cocoa after fermentation and drying and it was due to this compound be the principal substrate of existing polyphenoloxidase.
Other authors suggest that the reduction of (-) epicatechin may be due to an
epimerizarion phenomena caused by changes in pH during fermentation process
(Seto et al., 1997Seto, R., Nakamura, H., Nanjo, F., & Hara, Y. (1997).
Preparation of epimers of tea catechins by heat treatment. Bioscience,
Biotechnology, and Biochemistry, 61(9), 1434-1439.
http://dx.doi.org/10.1271/bbb.61.1434.
http://dx.doi.org/10.1271/bbb.61.1434...
).
On the other hand, the SR162 cultivar showed an increase in (-) epicatechin during fermentation. This behavior is related to the hydrolysis of tannins and procyanidins and the action of polyphenoloxidase, processes that generate and oxidize, respectively, smaller phenolic compounds in the medium. Under this aspect, it can be assumed that the phenolic compounds had lower oxidation in this cultivar than the other cultivars while had more hydrolytic activity, probably due to the fermentation conditions achieved by this cultivar during the process as described by Cruz et al. (2013).
Concerning (+) catechin, the amounts remained almost the same during the
fermentation process for all cultivars, as observed by Seto et al. (1997)Seto, R., Nakamura, H., Nanjo, F., & Hara, Y. (1997).
Preparation of epimers of tea catechins by heat treatment. Bioscience,
Biotechnology, and Biochemistry, 61(9), 1434-1439.
http://dx.doi.org/10.1271/bbb.61.1434.
http://dx.doi.org/10.1271/bbb.61.1434...
, but SR162 showed slightly higher
amounts than the other cultivars, which is a goal for this cultivar, since
(+)-catechin have a well-known healt benefits (Maskarinec, 2009Maskarinec, G. (2009). Cancer protective properties of cocoa: a
review of the epidemiologic evidence. Nutrition and Cancer, 61(5), 573-579.
http://dx.doi.org/10.1080/01635580902825662. PMid:19838930
http://dx.doi.org/10.1080/01635580902825...
; Khawaja et al.,
2011Khawaja, O., Gaziano, J. M., & Djoussé, L. (2011). Chocolate and
coronary heart disease: a systematic review. Current Atherosclerosis Reports,
13(6), 447-452. http://dx.doi.org/10.1007/s11883-011-0203-2.
PMid:21894553
http://dx.doi.org/10.1007/s11883-011-020...
).
The three cultivar studied showed different amounts of theobromine in the
beginning of fermentation (Table 3), the
non-resistent blend showed the highest value with 7.96mg.g–1,
followed by PH16 and SR162 with 6.50mg.g–1and 5.45mg.g–1,
respectively. The same phenomena was observed for caffeine (Table 3), the non-resistant blend showed
the highest amount, 4.35mg.g–1, while PH16 had
3.65mg.g-1and SR162 1.32mg.g–1. This fact was already
demonstrated by Aneja & Gianfagna
(2001)Aneja, M., & Gianfagna, T. (2001). Induction and accumulation of
caffeine in young actively growing leaves of cocoa (, L.) by wounding or
infection with Theobroma cacaoCrinipellis
perniciosa.Physiological and Molecular Plant Pathology, 59(1),
13-16. http://dx.doi.org/10.1006/pmpp.2001.0337.
http://dx.doi.org/10.1006/pmpp.2001.0337...
in their studies, showing that the plant tissue infected by
fungus can generate some defence biochemical mechanisms, evidenced by higher
content of methylxantines and phenolic compounds. Besides, some authors also
reported the existing difference between different cocoa genotype and
methylxantine content (Figueira et al.,
1997Figueira, A., Lambert, S., Carpenter, D., Pires, J. L., Cascardo, J.
C. M., & Romanczyk, L. (1997). The similarity of cocoa flavour of fermented
seeds from fingerprinted genotypes of L. from Brazil and Malaysia.
Theobroma cacaoTropical Agriculture, 74,
132-139.).
Methylxantines amounts (mg.g-1) during fermentation of three cocoa cultivars* * The values shown are the average of 2 fermentation replicates and expressed in dry defatted basis. .
On the other hand, higher levels of methylxantines can affect the sensory
characteristics of the cocoa beans, since sensory attributes like bitterness and
astringency were strongly correlated with methylxantines and phenolic compounds
(Efraim et al., 2011Efraim, P., Alves, A. B., & Jardim, D. C. P. (2011). Polifenóis
em cacau e derivados: teores, fatores de variação e efeitos na saúde. Brazilian
Journal of Food Technology, 14(3), 181-201.
http://dx.doi.org/10.4260/BJFT2011140300023.
http://dx.doi.org/10.4260/BJFT2011140300...
).
During fermentation, a significative variation in methylxantines levels was found only for non-resistant blend and PH16. The theobromine content in non-resistant blend was 6.18mg.g–1 at the end of fermentation, showing a reduction of 22% when compared with the beginning of the process. PH16 showed a reduction of 26% while SR162 showed an increase of theobromine content around 9%, which was not significative (Table 3). For caffeine the reduction at the ending of fermentation was around 50% for non-resistant blend and 14% for PH16 while for SR162 was an increase of 36%.
These results showed that there was not a pattern for methylxantines behavior
during fermentation process, since this process is composed by many complex
biochemical reactions involving lipids, proteins, methylxantines and phenolic
compounds, driving all these compounds to many chemical changes (Efraim et al., 2006Efraim, P., Tucci, M. L., García, N. H. P., Haddad, R., &
Eberlin, M. (2006). Teores de compostos fenólicos de sementes de cacaueiro de
diferentes genótipos. Brazilian Journal Food Technology, 9(4),
229-236.). Specifically for
methylxantines, probably their complexation with some peptides and the
decompartmentalisation of structures within the cell, related to fermentation
conditions like temperature and pH during the process, can explain that
difference observed for these three cultivars (Cruz et al., 2013Cruz, J. F. M., Leite, P. B., Soares, S. E., & Bispo, E. S.
(2013). Assessment of the fermentative process from different cocoa cultivars
produced in Southern Bahia, Brazil. African Journal of Biotechnology, 12(33),
5218-5225. http://dx.doi.org/10.5897/AJB2013.12122.
http://dx.doi.org/10.5897/AJB2013.12122...
).
3.2 Phenolic compounds and methylxantinesfrom fermented and dried cocoa beans
A huge reduction in phenolic compounds can be observed after drying as shown in
Table 4. The reduction of
epicatechin was higher in SR162 cultivar (68%), for non-resistant and PH16 the
reduction was around 55%. For catechin the reduction was 50% for non-resistant
cultivar, 45% for SR162 and 35% for PH16. This reduction was reported for other
authors and they attributed the reduction of polyphenols during sun drying
mainly to enzymatic browing, followed by non-enzymatic reactions due to quinones
polymerization (Payne et al., 2010Payne, M. J., Hurst, W. J., Miller, K. B., Rank, C., & Stuart,
D. A. (2010). Impact of fermentation, drying, roasting, and Dutch processing on
epicatechin and catechin content of cacao beans and cocoa ingredients. Journal
of Agricultural and Food Chemistry, 58(19), 10518-10527.
http://dx.doi.org/10.1021/jf102391q. PMid:20843086
http://dx.doi.org/10.1021/jf102391q...
;
Gil et al., 2011Gil, A., Rojas, L. F., Atehortua, L., & Londono, J. (2011).
Effect of Fermentation and Sun Drying on Phytochemical Composition of Native
Colombian Cocoa. In Proceedings of the 6th CIGR Section VI International
Symposium “Towards a Sustainable Food Chain” Food Process, Bioprocessing and
Food Quality Management, Nantes, France.). The polyphenol
oxidase would not have appropriate conditions to react during fermentation, but
during sun drying, with the increase in pH and high O2 uptake, the
conditions will be appropriate to phenolic oxidation. These can also explain the
reduction differences between cultivars, since each of them presented different
genotypic characteristics and consequent specific behavior during fermentation,
and the sun drying process were extremely influenced by climactic conditions
(Niemenak et al., 2006Niemenak, N., Rohsius, C., Elwers, S., Ndoumou, D. O., &
Lieberei, R. (2006). Comparative study of different cocoa (Theobroma cacao L.)
clones in terms of their phenolics and anthocyanins contents. Journal of Food
Composition and Analysis, 19(6-7), 612-619.
http://dx.doi.org/10.1016/j.jfca.2005.02.006.
http://dx.doi.org/10.1016/j.jfca.2005.02...
; Cruz et al., 2013Cruz, J. F. M., Leite, P. B., Soares, S. E., & Bispo, E. S.
(2013). Assessment of the fermentative process from different cocoa cultivars
produced in Southern Bahia, Brazil. African Journal of Biotechnology, 12(33),
5218-5225. http://dx.doi.org/10.5897/AJB2013.12122.
http://dx.doi.org/10.5897/AJB2013.12122...
). Although losses in
phenolic compounds were observed, sun drying allows to obtain products with
better sensory quality in relation to those artificially drying (Faborode et al., 1995Faborode, M. O., Favier, J. F., & Ajayi, O. (1995). A. On the
effects of forced air drying on cocoa quality. Journal of Food Engineering,
25(4), 455-472. http://dx.doi.org/10.1016/0260-8774(94)00018-5.
http://dx.doi.org/10.1016/0260-8774(94)0...
; Efraim et al., 2010)Efraim, P., Pezoa-Garcia, N. H., Jardim, D. C. P., Nishikawa, A.,
Haddad, R., & Eberlin, M. N. (2010). Influence of cocoa beans fermentation
and drying on the polyphenol content and sensory acceptance. Ciência e Tecnologia de Alimentos, 30(1), 142-150..
Phenolic compounds amounts (mg.g-1) in fermented and dried cocoa beans of three cocoa cultivars* * The values shown are the average of 2 fermentation replicates and expressed in dry defatted basis. .
In health aspects, this reduction is not a goal, since phenolic compounds like
catechin and epicatechin play an important part in preventing some degenerative
diseases through their high antioxidant activity in biological systems (Keen et al., 2005Keen, C. L., Holt, R. R., Oteiza, P. I., Fraga, C. G., &
Schmitz, H. H. (2005). Cocoa antioxidants and cardiovascular health. The
American Journal of Clinical Nutrition, 81(1, Suppl.), 298S-303S.
PMid:15640494.; Cooper et al., 2008Cooper, K. A., Donovan, J. L., Waterhouse, A. L., & Williamson,
G. (2008). Cocoa and health: a decade of research. The British Journal of
Nutrition, 99(1), 1-11. http://dx.doi.org/10.1017/S0007114507795296.
PMid:17666148
http://dx.doi.org/10.1017/S0007114507795...
; Mostofsky et al., 2010Mostofsky, E., Levitan, E. B., Wolk, A., & Mittleman, M. A.
(2010). Chocolate intake and incidence of heart failure: a population-based
prospective study of middle-aged and elderly women. Circulation: Heart Failure,
3(5), 612-616. http://dx.doi.org/10.1161/CIRCHEARTFAILURE.110.944025.
PMid:20713904
http://dx.doi.org/10.1161/CIRCHEARTFAILU...
).
The methylxantines content in fermented and dried cocoa beans are shown in Table 5. Differently of phenolic compounds, methylxantines did not have great modification after sun drying, the amounts of either caffeine or theobromine were almost the same before and after the sun drying process. However, for all cultivars theobromine always appears as the main methylxantine in beans, around three-fold the caffeine content for non-resistant and SR162 cultivars and two-fold for PH16, but in all cultivar de theobromine content is almost the same, varying from 5.53 mg.g–1 in PH16 cultivar to 6.13mg.g–1 in non-resistant blend, with no statistical difference between them. For caffeine higher variation was found, the content varied from 1.89mg.g–1 for SR162 cultivar to 3.02mg.g–1 for PH16 cultivar, with significative difference between them.
Methylxantines amounts (mg.g–1) in fermented and dried cocoa beans of three cocoa cultivars* * The values shown are the average of 2 fermentation replicates and expressed in dry defatted basis. .
These values found for methylxantines can be discussed in two points of views:
first in technological aspect, since these compounds together with polyphenols
are responsible for bitterness and astringency of the chocolate produced (Efraim et al., 2011Efraim, P., Alves, A. B., & Jardim, D. C. P. (2011). Polifenóis
em cacau e derivados: teores, fatores de variação e efeitos na saúde. Brazilian
Journal of Food Technology, 14(3), 181-201.
http://dx.doi.org/10.4260/BJFT2011140300023.
http://dx.doi.org/10.4260/BJFT2011140300...
). The other aspect is
the effect of methylxantines in human body, since caffeine is well known for its
action in central nervous system promoting a stimulating effect, increasing the
sense of allarm, vitality, psychomotor reactions and rising the blood pressure.
On the other hand, theobromine has no stimulating effect, because the action of
theobromine in the central nervous system is very weak or almost non-existent,
but it can act as a vasodilator, a muscle relaxant, diuretic and blood pressure
reducer (Van den Bogaard et al., 2010Van den Bogaard, B., Draijer, R., Westerhof, B. E., Van den
Meiracker, A. H., Van Montfrans, G. A., & Van den Born, B. J. (2010).
Effects on peripheral and central blood pressure of cocoa with natural or
high-dose theobromine: a randomized, double-blind crossover trial. Hypertension,
56(5), 839-846. http://dx.doi.org/10.1161/HYPERTENSIONAHA.110.158139.
PMid:20823377
http://dx.doi.org/10.1161/HYPERTENSIONAH...
;
Mitchell et al., 2011Mitchell, E. S., Slettenaar, M., vd Meer, N., Transler, C., Jans,
L., Quadt, F., & Berry, M. (2011). Differential contributions of theobromine
and caffeine on mood, psychomotor performance and blood pressure. Physiology
& Behavior, 104(5), 816-822.
http://dx.doi.org/10.1016/j.physbeh.2011.07.027. PMid:21839757
http://dx.doi.org/10.1016/j.physbeh.2011...
).
4 Conclusions
A reduction in (-) epicatechin was observed for two cultivars (non-resistant and PH16) during fermentation. At the end of the process, the reduction observed was around 50% in these two cultivars and was attributed to (-)epicatechin be the principal substrate of existing polyphenoloxidase. Other hypothesis suggested was due to (-) epicatechinepimerizarion phenomena caused by changes in pH during fermentation process. On the other hand in SR162 showed an increase in (-) epicatechin during fermentation, assuming that the phenolic compounds had lower oxidation in this cultivar than the other cultivars, while had more hydrolytic activity, probably due to the fermentation conditions achieved by this cultivar during fermentation process.
During fermentation, an expressive variation in methylxantines contents was found. These results showed that there was not a pattern for methylxantines behaviour during fermentation process, and it is extremely related to fermentation conditions like temperature and pH during the process.
A huge reduction in phenolic compounds can be observed after drying, attributed mainly to enzymatic browing, followed by non-enzymatic reactions due to quinones polymerization with the increase in pH and high O2 uptake during sun drying.
Differently of phenolic compounds, methylxantines did not have great modification after sun drying, the amounts of either caffeine or theobromine were almost the same before and after the sun drying process.
Under technological aspect, methylxantines and phenolic compounds are responsible for bitterness and astringency of the chocolate produced. So, the differences observed in this study between cultivars, take to the conclusion that the cocoa cultivars have different behavior during fermentation and drying considering phenolic and methylxantines contents, which consequently, give to these cultivars differences in sensory characteristics.
Acknowledgements
To Fundação de Amparo à Pesquisa do Estado da Bahia (FAPESB) for the scholarship and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the financial support.
-
Practical Application: Observe the behavior of bioactive substances in cocoa beans during preprocessing.
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Publication Dates
-
Publication in this collection
June 2015
History
-
Received
28 Oct 2014 -
Accepted
19 Mar 2015