Chemical and flavor profile changes of cocoa beans (Theobroma cacao L.) during primary fermentation.
ABSTRACT: This survey reports for the first time the changed of quality of fermented cocoa (Theobroma cacao L.) beans. The quality evaluation and simultaneous detection of amino acids, flavor, procyanidin, color, fat, protein, antioxidant activity, and enthalpy were obtained for different fermentation stages of cocoa beans. The results showed that total essential amino acids contents ranged from 2.64 g/100 g to 3.68 g/100 g. A total of 88 compounds identified at the end of the fermentation belonged to alcohols, acids, esters, ketones, pyrazines, aldehydes, and terpenoids. One of the chemical groups that were present in highest abundance in the consummation treatments was acids, representing 56.04% of the total extracted area, followed by alcohols (22.95%) and ketones (9.40%). The colors of the beans in different fermentation stages were different, from deep purple to deep red-brown. Fermented cocoa beans were shown to be 53.45% and 13.51% bean butter and protein content, respectively. The value of denaturation enthalpy (?H) ranged from 30.4 (J/g) to 43.38 (J/g). The 3-day fermented sample had the highest ?H (43.38 J/g). When the fermentation process was complete, the procyanidin concentration of the beans decreased, with the final yield of procyanidin at 6.2%. During fermentation, the antioxidant capacity of beans gradually reduced. The fermenting of cocoa beans had a significant effect on the quality formation. The findings of this study constitute a basis for further investigations on the quality formation of cocoa during fermentation.
Project description:The growth of filamentous fungi during the spontaneous cocoa bean fermentation leads to inferior cocoa bean quality and poses a health risk for consumers due to the potential accumulation of mycotoxins. We recently developed anti-fungal cultures with the capacity to inhibit the growth of mycotoxigenic filamentous fungi on cocoa beans. However, it is not clear how these anti-fungal cultures affect the fermentation process and cocoa bean quality. For that, the anti-fungal co-cultures, Lactobacillus fermentum M017-Saccharomyces cerevisiae H290 (A) and Lb. fermentum 223-S. cerevisiae H290 (B), were applied to 180-kg box fermentations in Honduras in three time-independent replications each including a spontaneous control fermentation. The comparison of inoculated and spontaneous fermentation processes revealed that the co-cultures only marginally affected the fermentation process and cocoa bean quality. Microorganisms reached maximal levels of 6.2-7.6 log CFU/g of yeasts and acetic acid bacteria and 7.9-9.5 log CFU/g of lactic acid bacteria during all fermentations and led to maximal metabolite concentrations in bean cotyledons of 4-12 mg/g ethanol, 2-6 mg/g lactic acid and 6-14 mg/g acetic acid. The fermentation and drying processes resulted in 38-90 mg epicatechin equivalents/g in the cotyledons of dried beans. However, the co-cultures led to up to ten times higher mannitol levels in cotyledons of inoculated beans compared to beans during spontaneous fermentation, and caused a slower fermentation process, detectable as up to 8-12 °C lower temperatures in the centre of the fermenting pulp-bean mass and up to 22% lower proportions of well-fermented beans after drying. Co-culture B-with Lb. fermentum 223 -led to improved cocoa bean quality compared to co-culture A-with Lb. fermentum M017 -, i.e. cocoa beans with 0.5-1.9 mg/g less acetic acid, 4-17% higher shares of well-fermented beans and, on a scale from 0 to 10, to 0.2-0.6 units lower astringency, up to 1.1 units lower off-flavours, and 0.2-0.9 units higher cocoa notes. Therefore, the anti-fungal co-culture B is recommended for future applications and its capacity to limit fungal growth and mycotoxin production during industrial-scale cocoa bean fermentation should be investigated in further studies.
Project description:The sensory quality and the contents of quality-determining chemical compounds in unfermented and fermented cocoa from 100 cacao trees (individual genotypes) representing groups of nine genotype spectra (GG), grown at smallholder plantings in the municipality of Waslala, Nicaragua, were evaluated for two successive harvest periods. Cocoa samples were fermented using a technique mimicking recommended on-farm practices. The sensory cocoa quality was assessed by experienced tasters, and seven major chemical taste compounds were quantified by near infrared spectrometry (NIRS). The association of the nine, partially admixed, genotype spectra with the analytical and sensory quality parameters was tested. The individual parameters were analyzed as a function of the factors GG and harvest (including the date of fermentation), individual trees within a single GG were used as replications. In fermented cocoa, significant GG-specific differences were observed for methylxanthines, theobromine-to-caffeine (T/C) ratio, total fat, procyanidin B5 and epicatechin, as well as the sensory attributes global score, astringency, and dry fruit aroma, but differences related to harvest were also apparent. The potential cocoa yield was also highly determined by the individual GG, although there was significant tree-to-tree variation within every single GG. Non-fermented samples showed large harvest-to-harvest variation of their chemical composition, while differences between GG were insignificant. These results suggest that selection by the genetic background, represented here by groups of partially admixed genotype spectra, would be a useful strategy toward enhancing quality and yield of cocoa in Nicaragua. Selection by the GG within the local, genetically segregating populations of seed-propagated cacao, followed by clonal propagation of best-performing individuals of the selected GG could be a viable alternative to traditional propagation of cacao by seed from open pollination. Fast and gentle air-drying of the fermented beans and their permanent dry storage were an efficient and comparatively easy precondition for high cocoa quality.
Project description:Chocolate production suffered a vast impact with the emergence of the "witches' broom" disease in cocoa plants. To recover cocoa production, many disease-resistant hybrid plants have been developed. However, some different cocoa hybrids produce cocoa beans that generate chocolate with variable quality. Fermentation of cocoa beans is a microbiological process that can be applied for the production of chocolate flavor precursors, leading to overcoming the problem of variable chocolate quality. The aim of this work was to use a cocktail of microorganisms as a starter culture on the fermentation of the ripe cocoa pods from PH15 cocoa hybrid, and evaluate its influence on the microbial communities present on the fermentative process on the compounds involved during the fermentation, and to perform the chocolate sensorial characterization. According to the results obtained, different volatile compounds were identified in fermented beans and in the chocolate produced. Bitterness was the dominant taste found in non-inoculated chocolate, while chocolate made with inoculated beans showed bitter, sweet, and cocoa tastes. 2,3-Butanediol and 2,3-dimethylpyrazine were considered as volatile compounds making the difference on the flavor of both chocolates. Saccharomyces cerevisiae UFLA CCMA 0200, Lactobacillus plantarum CCMA 0238, and Acetobacter pasteurianus CCMA 0241 are proposed as starter cultures for cocoa fermentation.
Project description:Cocoa post-harvest practices were monitored on a small-farm scale (ca. 50 kg fresh beans) at five intermediaries from four provinces in Ecuador: (A) in Manabí, (B) and (E) in Los Ríos, (C) in Cotopaxi, (D) in Guayas. Temperature, pH (pulp, cotyledon), cell counts (yeasts, lactic acid bacteria, acetic acid bacteria) were recorded daily, and cut-tests and sensory descriptive analysis evaluated end quality. An overall inconsistency and variability in processing were observed with different fermentation devices (jute/plastic bags, wooden boxes), pre-drying, turning during fermentation, fermentation duration, and different drying processes (temperatures, direct/indirect). Key parameters (maximum temperature, pH cotyledon development) revealed a significant impact of the fermentation device on the post-harvest process and, therefore, on the fermentation development. 67-74 h in jute bags without turning was sufficient to reach well-fermented cocoa beans without moldy off-flavors, whereas 133 h in plastic bags without turning resulted in 3 ± 1% moldy beans and cocoa liquor with moldy off-flavor. Drying at high temperatures (80 ± 10 °C) with direct heat contact resulted in beans roasted to burnt off-flavor. Conclusively, the whole post-harvest process was crucial for well-fermented beans without off-flavor. Plastic bags seemed unsuitable, while jute bags could be an alternative to wooden boxes.
Project description:This study aimed to identify the volatile compounds in the fermented and dried cocoa beans conducted with three distinct inoculants of yeast species due to their high fermentative capacity: <i>Saccharomyces cerevisiae</i>, <i>Pichia kudriavzevii</i>, the mixture in equal proportions 1:1 of both species, and a control fermentation (with no inoculum application). Three starter cultures of yeasts, previously isolated and identified in cocoa fermentation in the municipality of Tomé-Açu, Pará state, Brazil. The seeds with pulp were removed manually and placed in wooden boxes for the fermentation process that lasted from 6 to 7 days. On the last day of fermentation, the almonds were packaged properly and placed to dry (36 °C), followed by preparation for the analysis of volatile compounds by GC-MS technique. In addition to the control fermentation, a high capacity for the formation of desirable compounds in chocolate by the inoculants with <i>P. kudriavzevii</i> was observed, which was confirmed through multivariate analyses, classifying these almonds with the highest content of aldehydes, esters, ketones and alcohols and low concentration of off-flavours. We conclude that the addition of mixed culture starter can be an excellent alternative for cocoa producers, suggesting obtaining cocoa beans with desirable characteristics for chocolate production, as well as creating a product identity for the producing region.
Project description:Traditional fermentations of the local Ecuadorian cocoa type Nacional, with its fine flavor, are carried out in boxes and on platforms for a short time. A multiphasic approach, encompassing culture-dependent and -independent microbiological analyses of fermenting cocoa pulp-bean samples, metabolite target analyses of both cocoa pulp and beans, and sensory analysis of chocolates produced from the respective fermented dry beans, was applied for the investigation of the influence of these fermentation practices on the yeast and bacterial species diversity and community dynamics during cocoa bean fermentation. A wide microbial species diversity was found during the first 3 days of all fermentations carried out. The prevailing ethanol-producing yeast species were Pichia kudriavzevii and Pichia manshurica, followed by Saccharomyces cerevisiae. Leuconostoc pseudomesenteroides (glucose and fructose fermenting), Fructobacillus tropaeoli-like (fructose fermenting), and Lactobacillus fermentum (citrate converting, mannitol producing) represented the main lactic acid bacterial species in the fermentations studied, resulting in intensive heterolactate metabolism of the pulp substrates. Tatumella saanichensis and Tatumella punctata were among the members of the family Enterobacteriaceae present during the initial phase of the cocoa bean fermentations and could be responsible for the production of gluconic acid in some cases. Also, a potential new yeast species was isolated, namely, Candida sorbosivorans-like. Acetic acid bacteria, whose main representative was Acetobacter pasteurianus, generally appeared later during fermentation and oxidized ethanol to acetic acid. However, acetic acid bacteria were not always present during the main course of the platform fermentations. All of the data taken together indicated that short box and platform fermentation methods caused incomplete fermentation, which had a serious impact on the quality of the fermented dry cocoa beans.
Project description:Microbial metabolism drives changes in the physicochemical properties and, consequently, the sensory characteristics of fermented cocoa beans. In this context, information regarding the structure, function, and metabolic potential of microbial communities' present during cocoa pulp-bean mass fermentation is limited, especially concerning the formation of aromatic compounds. To bridge the gap, the metagenome of fermented cocoa pulp-bean mass (Criollo and Forastero) has been investigated using shotgun metagenomics coupled with physicochemical, microbiological, quality, and sensory analyses to explore the impact of microbial communities on the quality of fermented cocoa pulp-bean mass on one farm in one season and in one region under the same environmental conditions. Our findings showed that the metagenomic diversity in cocoa, the fermentation length, and the diversity and function of metagenome-assembled genomes (MAGs) greatly influence the resulting distinctive flavors. From the metabolic perspective, multiple indicators suggest that the heterolactic metabolism was more dominant in Criollo fermentations. KEGG genes were linked with the biosynthesis of acetic acid, ethanol, lactic acid, acetoin, and phenylacetaldehyde during Criollo and Forastero fermentations. MAGs belonging to Lactiplantibacillus plantarum, Limosilactobacillus reuteri, and Acetobacter pasteurianus were the most prevalent. Fermentation time and roasting are the most important determinants of cocoa quality, while the difference between the two varieties are relatively minor. The assessment of microbiological and chemical analysis is urgently needed for developing fermentation protocols according to regions, countries, and cocoa varieties to guarantee safety and desirable flavor development. <b>IMPORTANCE</b> Monitoring the composition, structure, functionalities, and metabolic potential encoded at the level of DNA of fermented cocoa pulp-bean mass metagenome is of great importance for food safety and quality implications.
Project description:Yeasts play an important role in the cocoa fermentation process. Although the most relevant function is the degradation of sugars and the production of ethanol, there is little understanding of the enzyme activities and attributes that allow them to survive even after drying. The present study explored the functional biodiversity of yeasts associated with Criollo Colombian cocoa fermented beans, able to survive after drying. Twelve species belonging to 10 genera of osmo-, acid-, thermo-, and desiccation-tolerant yeasts were isolated and identified from fermented and dry cocoa beans, with Pichia kudriavzevii and Saccharomyces cerevisiae standing out as the most frequent. For the first time, we reported the presence of Zygosaccharomyces bisporus in cocoa fermented beans. It was found that resistance to desiccation is related to the different degradation capacities of fermentation substrates, which suggests that associative relationships may exist between the different yeast species and their degradation products. Besides, the increased thermotolerance of some species was related to the presence of polyphenols in the medium, which might play a fundamental role in shaping the microbial community composition.
Project description:The aim of the current work was to use hyperspectral imaging (HSI) in the spectral range 1000-2500?nm to quantitatively predict fermentation index (FI), total polyphenols (TP) and antioxidant activity (AA) of individual dry fermented cocoa beans scanned on a single seed basis, in a non-destructive manner. Seventeen cocoa bean batches were obtained and 10 cocoa beans were used from each batch. PLS regression models were built on 170 samples. The developed HSI predictive models were able to quantify three quality-related parameters with sufficient performance for screening purposes, with external validation R2 of 0.50 (RMSEP?=?0.27, RPD?=?1.40), 0.70 (RMSEP?=?34.1?mg ferulic acid?g-1, RPD?=?1.77) and 0.74 (60.0?mmol Trolog?kg-1, RPD?=?1.91) for FI, TP and AA, respectively. The calibrations were subsequently applied at a single bean and pixel level, so that the distribution was visualised within and between single seeds (chemical images). HSI is thus suggested as a promising approach to estimate cocoa bean composition rapidly and non-destructively, thus offering a valid tool for food inspection and quality control.
Project description:The data provide information in support of the research article, "The cleavage specificity of the aspartic protease of cocoa beans involved in the generation of the cocoa-specific aroma precursors" (Janek et al., 2016) . Three different protein substrates were partially digested with the aspartic protease isolated from cocoa beans and commercial pepsin, respectively. The obtained peptide fragments were analyzed by matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF-MS/MS) and identified using the MASCOT server. The N- and C-terminal ends of the peptide fragments were used to identify the corresponding in-vitro cleavage sites by comparison with the amino acid sequences of the substrate proteins. The same procedure was applied to identify the cleavage sites used by the cocoa aspartic protease during cocoa fermentation starting from the published amino acid sequences of oligopeptides isolated from fermented cocoa beans.