Dynamics and species diversity of communities of lactic acid bacteria and acetic acid bacteria during spontaneous cocoa bean fermentation in vessels

Lefeber, Timothy; Gobert, William; Vrancken, Gino; Camu, Nicholas; Vuyst, Luc De

doi:10.1016/j.fm.2010.10.010

Cited by 110 publications

(73 citation statements)

References 25 publications

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“…In addition, Acetobacter ghanensis and Acetobacter senegalensis are found during spontaneous cocoa bean fermentation (9,13,17,18). Further studies provided first insights into the basic microbiological properties of such strains and macroscopic dynamics during cocoa pulp fermentation (12,15,(18)(19)(20). At this point, it appears to be relevant to resolve the metabolic contribution of AAB in greater detail.…”

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confidence: 94%

“…Obviously, the most prevalent AAB species is Acetobacter pasteurianus (13,(15)(16)(17). In addition, Acetobacter ghanensis and Acetobacter senegalensis are found during spontaneous cocoa bean fermentation (9,13,17,18). Further studies provided first insights into the basic microbiological properties of such strains and macroscopic dynamics during cocoa pulp fermentation (12,15,(18)(19)(20).…”

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confidence: 99%

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The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Adler

Frey

Berger

et al. 2014

Appl Environ Microbiol

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c Acetic acid bacteria (AAB) play an important role during cocoa fermentation, as their main product, acetate, is a major driver for the development of the desired cocoa flavors. Here, we investigated the specialized metabolism of these bacteria under cocoa pulp fermentation-simulating conditions. A carefully designed combination of parallel 13 C isotope labeling experiments allowed the elucidation of intracellular fluxes in the complex environment of cocoa pulp, when lactate and ethanol were included as primary substrates among undefined ingredients. We demonstrate that AAB exhibit a functionally separated metabolism during coconsumption of two-carbon and three-carbon substrates. Acetate is almost exclusively derived from ethanol, while lactate serves for the formation of acetoin and biomass building blocks. Although this is suboptimal for cellular energetics, this allows maximized growth and conversion rates. The functional separation results from a lack of phosphoenolpyruvate carboxykinase and malic enzymes, typically present in bacteria to interconnect metabolism. In fact, gluconeogenesis is driven by pyruvate phosphate dikinase. Consequently, a balanced ratio of lactate and ethanol is important for the optimum performance of AAB. As lactate and ethanol are individually supplied by lactic acid bacteria and yeasts during the initial phase of cocoa fermentation, respectively, this underlines the importance of a well-balanced microbial consortium for a successful fermentation process. Indeed, AAB performed the best and produced the largest amounts of acetate in mixed culture experiments when lactic acid bacteria and yeasts were both present.A cetic acid bacteria (AAB) play an important role in cocoa fermentation (1). During fermentation of the pulp that surrounds the cocoa beans, they form acetate. Acetate then diffuses into the beans (2-4), where it initiates a cascade of chemical and biochemical reactions leading to precursor molecules for cocoa flavor (2, 5, 6). Potential substrates for AAB are lactate and ethanol, which are individually produced by lactic acid bacteria (LAB; mainly Lactobacillus fermentum) and yeasts (diverse yeasts such as Saccharomyces cerevisiae, Hanseniaspora opuntiae, and Candida krusei), respectively, during the fermentation process (6-12). Hereby, the degradation of lactate by AAB is desired, since the remaining lactate may provide an off flavor in the final cocoa product (11,13,14). In recent years, AAB have been extensively analyzed for their contribution to cocoa fermentation. Obviously, the most prevalent AAB species is Acetobacter pasteurianus (13,(15)(16)(17). In addition, Acetobacter ghanensis and Acetobacter senegalensis are found during spontaneous cocoa bean fermentation (9,13,17,18). Further studies provided first insights into the basic microbiological properties of such strains and macroscopic dynamics during cocoa pulp fermentation (12,15,(18)(19)(20). At this point, it appears to be relevant to resolve the metabolic contribution of AAB in greater detail. The basic know...

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confidence: 94%

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confidence: 99%

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Adler

Frey

Berger

et al. 2014

Appl Environ Microbiol

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“…During fermentation, bacterial growth (CFU per ml) was quantified through plating of 10-fold serial dilutions of the samples in saline (0.85% [wt/vol] NaCl solution) on CPSM agar (CPSM containing 1.5% [wt/vol] agar, pH 5.5) that was incubated at the appropriate fermentation temperature for 24 h. Metabolite concentrations were determined through high-performance anion-exchange chromatography using a standard addition protocol (glucose, fructose, mannitol, and citric acid) (15,23) and high-performance liquid chromatography using external standards (lactic acid, acetic acid, and ethanol) (16). Cell count and metabolite (with external standards) measurements were performed on three independent samples.…”

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confidence: 99%

“…The errors on the measurements are represented as standard deviations. Gas chromatography (GC) was used for the qualitative determination of 2,3-butanediol and acetoin (15). Concentrations of carbon dioxide in the fermentor gas effluents were determined online through GC (13).…”

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Interesting Starter Culture Strains for Controlled Cocoa Bean Fermentation Revealed by Simulated Cocoa Pulp Fermentations of Cocoa-Specific Lactic Acid Bacteria

Lefeber

Janssens

Moens

et al. 2011

Appl Environ Microbiol

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Among various lactic acid bacterial strains tested, cocoa-specific strains of Lactobacillus fermentum were best adapted to the cocoa pulp ecosystem. They fermented glucose to lactic acid and acetic acid, reduced fructose to mannitol, and converted citric acid into lactic acid and 2,3-butanediol.Fermented dry cocoa beans are the basic raw material for chocolate production. Cocoa beans are the seeds of the cocoa tree, Theobroma cacao L. The key microorganisms for successful cocoa bean fermentation processes are yeasts, lactic acid bacteria (LAB), and acetic acid bacteria (AAB) (1, 3-5, 14, 18-21). Although the LAB species diversity involved in the onset of any spontaneous cocoa bean fermentation is wide, not much is known about how cocoa-specific LAB species, such as Lactobacillus fermentum, adapt physiologically and what their targeted functional roles are during fermentation (1,3,4,14,(19)(20)(21). The present study aimed at the kinetic investigation of carbohydrate fermentation and citric acid conversion by various LAB strains to unravel this.The LAB strains used throughout this study are listed in Table 1. Monoculture fermentations were performed in 1.5 liters of a cocoa pulp simulation medium (CPSM) for LAB (16) in Biostat B-DCU fermentors (Sartorius AG/B. Braun Biotech International, Melsungen, Germany) anaerobically for 48 h. Inoculum build-up, fermentor setup, online control of temperature (Table 2), pH profile, agitation, and sampling were as described previously (13, 16). All fermentations were performed in duplicate. The results and figures presented hereinafter are representative for both fermentations.During fermentation, bacterial growth (CFU per ml) was quantified through plating of 10-fold serial dilutions of the samples in saline (0.85% [wt/vol] NaCl solution) on CPSM agar (CPSM containing 1.5% [wt/vol] agar, pH 5.5) that was incubated at the appropriate fermentation temperature for 24 h. Metabolite concentrations were determined through high-performance anion-exchange chromatography using a standard addition protocol (glucose, fructose, mannitol, and citric acid) (15, 23) and high-performance liquid chromatography using external standards (lactic acid, acetic acid, and ethanol) (16). Cell count and metabolite (with external standards) measurements were performed on three independent samples. The errors on the measurements are represented as standard deviations. Gas chromatography (GC) was used for the qualitative determination of 2,3-butanediol and acetoin (15). Concentrations of carbon dioxide in the fermentor gas effluents were determined online through GC (13). The carbon recovery (CR, expressed as percentage) was calculated by dividing the total amount of carbon recovered in the metabolites by the total amount of carbon present in the carbon sources.All LAB strains tested were able to grow in CPSM ( Fig. 1 and T fermented fructose but not glucose and converted citric acid. Due to the initial low pH (3.5), Enterococcus casseliflavus M484 and Lactobacillus amylovorus DCE 471 were not able to...

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“…In fermentation, the molecular identification of microbial species [6]- [8] has to elucidate the metabolic mechanisms of the degradation of the mucilage and formation of metabolites. In addition, during the drying stage potential species have been identified (G. xylinum, Clostridium sp) which can be biological markers [9].…”

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confidence: 99%

Effect of Pectin Lyase Enzyme on Fermentation and Drying of Cocoa (<i>Theobroma cacao</i> L.): An Alternative to Improve Raw Material in the Industry of Chocolate

Garzón¹,

Orrego²,

Cadena³

et al. 2016

FNS

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Cocoa (Theobroma cacao), in all its presentations, is consumed all over the world and is one of the main drivers of the economic in several countries. The world's Cocoa tendency is focused on developing special beans. This category is subject to postharvest processes of utmost importance such as the fermentation and dry, which are currently carried out with traditional and poorly effective devices, which need to be improved to obtain a high quality product. The aim of this study was to evaluate the influence of the pectin lyase enzyme (E.C.4.2.2.10) on the postharvest cocoa process. We evaluated the enzyme dosage (1.0% and 0.5%) in fermentation and its effect on the variables temperature, acidity and drying time by convection at 60˚C. The Pectin lyase activity during fermentation does not cause a significant effect on the variables of temperature and acidity; however, the drying process time required to achieve 7.0% moisture was reduced. The enzyme dosage of 1.0% was the best result, the amount of exudate obtained (115 ml) during fermentation and the best degree of fermentation (77% ± 3.8) were increased and further shows a change in porosity facilitating the scale surface and internal moisture diffusion. The drying rate (Nw) expressed in kgwater/m 2 * min was determined based on the empirical model of Newton, where the higher speed was obtained during the falling period. In conclusion, enzyme dosage 1% was the best concentration evaluated because weaken grain husk, which allowed an adequate fermentation,

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Dynamics and species diversity of communities of lactic acid bacteria and acetic acid bacteria during spontaneous cocoa bean fermentation in vessels

Cited by 110 publications

References 25 publications

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Interesting Starter Culture Strains for Controlled Cocoa Bean Fermentation Revealed by Simulated Cocoa Pulp Fermentations of Cocoa-Specific Lactic Acid Bacteria

Effect of Pectin Lyase Enzyme on Fermentation and Drying of Cocoa (<i>Theobroma cacao</i> L.): An Alternative to Improve Raw Material in the Industry of Chocolate

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Dynamics and species diversity of communities of lactic acid bacteria and acetic acid bacteria during spontaneous cocoa bean fermentation in vessels

Cited by 110 publications

References 25 publications

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Interesting Starter Culture Strains for Controlled Cocoa Bean Fermentation Revealed by Simulated Cocoa Pulp Fermentations of Cocoa-Specific Lactic Acid Bacteria

Effect of Pectin Lyase Enzyme on Fermentation and Drying of Cocoa (&lt;i&gt;Theobroma cacao&lt;/i&gt; L.): An Alternative to Improve Raw Material in the Industry of Chocolate

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Effect of Pectin Lyase Enzyme on Fermentation and Drying of Cocoa (<i>Theobroma cacao</i> L.): An Alternative to Improve Raw Material in the Industry of Chocolate