Isolation and identification of microbial species found in cocoa fermentation as microbial starter culture candidates for cocoa bean fermentation in Colombia.

Microbial activity involved in the cocoa beans fermentation process is essential to maintain and improve the organoleptic and nutritional qualities of chocolate; therefore, the aim of this investigation was to search and select microbial isolates with the potential to improve the quality of cocoa beans. Fermentation experimentswere conducted on farms located in Maceo (Antioquia), San Vicente de Chucurí (Santander), and Rivera and Algeciras (Huila), Colombia. Yeast, lactic acid bacteria (LAB), acetic acid bacteria (AAB), and mesophilic aerobic microorganisms were obtained from different fermentation batches. The growth of these microorganismswas tested in six treatments as follows: 50% cocoa pulp agar (CPA), high concentrations of glucose (10%), ethanol (5%), and acetic acid (7%), an acidic pH of 3.0, and a high temperature of 50oC for 24 h. The isolates with the highest growth were identified by 18S and 16S rRNA gene analysis, revealing a high diversity ofspecies associated with cocoa fermentation, including eight species of yeasts (Debaryomyces hansenii, Meyerozyma guillermondii, Wickerhanomyces anomalus, Pichia guillermondii, Pichia kudriavzevii, Trichosporon asahii, Candida parapsilosis, and Pichia manshurica), six species of LAB (Pediococcus acidilactici, Lactobacillus brevis, Lactobacillus plantarum, Lactobacillus farraginis, Lactobacillus rhamnosus, and Leuconostoc mesenteroides), four species of AAB (Gluconobacter japonicus, Acetobacter tropicalis, Acetobacter pasteurianus, and Acetobacter malorum/tropicalis), and three species of Bacillus spp. (Bacillusaryabhattai /megaterium, Bacillus subtilis, and Bacillus coagulans). In general, microbial populations increased in cocoa batches after 12 h of fermentation and decreased after 84-96 h. All the yeast isolates grew in 10% glucose and CPA, 85.7% in 5% ethanol, and 95% at a pH of 3.0. All the yeast isolates were affectedby 7% acetic acid and incubation at 50oC for 24 h. Eighty-five percent of the LAB grew in 10% glucose, 100% in 5% ethanol, 42.8% in CPA, 64% at a pH of 3.0, and 35.7% grew after being exposed to 50oC for 24 h; all were affected by 7% acetic acid. As for the AAB, 100% grew in 10% glucose, 71% in 7% ethanol, 100% grew in CPA, in 7% acetic acid, and at a pH of 3.0, while 100% were affected by incubation at 50oC. Three yeast isolates, W. anomalus, D. hansenii and M. guillermondii, three LAB isolates, P. acidilactici, L. brevis, and L. plantarum, and three AAB isolates, A. tropicalis, A. pasteurianus and G. japonicus, were selected as promising strains to be used in a microbial starter culture for cocoa bean fermentation to improve the organoleptic quality of cocoa.

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Changes in Physico-Chemical and Microbiological Properties in Thai Cocoa Bean Fermentation

Walailak Journal of Science and Technology (WJST) ◽

10.48048/wjst.2021.21443 ◽

2021 ◽

Vol 18 (14) ◽

Author(s):

Tiparat TIKAPUNYA

Keyword(s):

Small Volume ◽

Ph Value ◽

Acetic Acid Bacteria ◽

Cocoa Bean ◽

Cocoa Beans ◽

Cocoa Bean Fermentation ◽

Physico Chemical ◽

Cocoa Fermentation ◽

Microbiological Properties

The purpose of this research is to investigate the quality-related physical, chemical, and microbiological changes in Thai cocoa beans during fermentation in 2 types of wooden containers. The results will compose a book of guidelines for good Thai cocoa fermentation in order to educate Thai farmers. Fresh Thai cocoa beans have a low pH value (5.0 - 5.5) compared to those from other countries in general (6.0 - 7.0). However, fermented temperature is able to reach 40 - 45 °C in 6 days, which is a main criteria for finishing cocoa fermentation. The color of fresh cocoa beans changes from white to brown within 2 days; after that, cocoa beans are mixed from the top to the bottom of the containers. Three groups of microorganism are evaluated with 3 different sampling points in wooden containers. The results reveal that yeast is grown quickly in 2 days on the top of containers, and then acetic acid bacteria and lactic acid bacteria are grown by the utilized yeast’s metabolites. These behaviors were found in both of the 2 wooden containers; however, a heap of cocoa beans (200 - 250 kgs) in a wooden box showed better quality of cocoa fermentation than a small volume (40 - 50 kgs) in a wooden tray. HIGHLIGHTS Understanding changes in Thai cocoa beans fermentation based on traditional method Two type of wooden containers applied for comparing all quality-related cocoa fermentation Proper cocoa beans fermentation process composed to a book of guideline for Thai Farmers GRAPHICAL ABSTRACT

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Curing of Cocoa Beans: Fine-Scale Monitoring of the Starter Cultures Applied and Metabolomics of the Fermentation and Drying Steps

Frontiers in Microbiology ◽

10.3389/fmicb.2020.616875 ◽

2021 ◽

Vol 11 ◽

Author(s):

Cristian Díaz-Muñoz ◽

Dario Van de Voorde ◽

Andrea Comasio ◽

Marko Verce ◽

Carlos Eduardo Hernandez ◽

...

Keyword(s):

Starter Culture ◽

Agar Plate ◽

Starter Cultures ◽

Cocoa Bean ◽

Sequence Variant ◽

Cocoa Beans ◽

Pichia Kudriavzevii ◽

Fermentation Processes ◽

Wide Range ◽

Cocoa Fermentation

Starter culture-initiated cocoa fermentation processes can be applied to improve the quality of cured cocoa beans. However, an accurate monitoring of the microbial strains inoculated in fresh cocoa pulp-bean mass to assess their contribution to the cocoa bean curing process is still lacking. In the present study, eight different cocoa fermentation processes were carried out with Trinitario cocoa in vessels in Costa Rica to assess the contribution of two candidate yeast starter culture strains, namely Saccharomyces cerevisiae IMDO 050523 and Pichia kudriavzevii IMDO 020508, inoculated in combination with Limosilactobacillus fermentum IMDO 0611222 and Acetobacter pasteurianus IMDO 0506386. A multiphasic approach, consisting of culture-dependent selective plating and incubation, rRNA-PCR-DGGE community profiling of agar plate washes, and culture-independent high-throughput amplicon sequencing, combined with a metabolite target analysis of non-volatile and volatile organic compounds (VOCs), was performed on samples from the fermentation and/or drying steps. The different starter culture mixtures applied effectively steered the cocoa fermentation processes performed. Moreover, the use of an amplicon sequence variant (ASV) approach, aligning these ASVs to the whole-genome sequences of the inoculated strains, allowed the monitoring of these inoculated strains and their differentiation from very closely related variants naturally present in the background or spontaneous fermentation processes. Further, traits such as malolactic fermentation during the fermentation step and acetoin and tetramethylpyrazine formation during the drying step could be unraveled. Finally, the yeast strains inoculated influenced the substrate consumption and metabolite production during all starter culture-initiated fermentation processes. This had an impact on the VOC profiles of the cured cocoa beans. Whereas the P. kudriavzevii strain produced a wide range of VOCs in the cocoa pulp, the S. cerevisiae strain mostly influenced the VOC composition of the cured cocoa beans.

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Enhancement of Cocoa Quality By the Indigenous Yeast Candida Tropicalis KLK4 Through Cocoa Bean Fermentation

JOURNAL OF ADVANCES IN BIOTECHNOLOGY ◽

10.24297/jbt.v4i1.1640 ◽

2014 ◽

Vol 4 (1) ◽

pp. 327-335

Author(s):

Nur Arfa Yanti

Keyword(s):

Candida Tropicalis ◽

Ph Value ◽

Large Subunit ◽

Starter Culture ◽

Cocoa Bean ◽

Rrna Gene ◽

Cocoa Bean Fermentation ◽

Assimilation Capacity ◽

Indigenous Yeast ◽

Cocoa Fermentation

An indigenous yeast strain that isolated from fermented cocoa bean has a role in cocoa bean fermentation process, was characterized and identified to be member of the Candida tropicalis Â species Â based on phenotypic characteristics and the D1/D2 region of the large subunit rRNA gene. Properties of importance for cocoa bean fermentation, namely sucrose, glucose, and citrate assimilation capacity, pH-, ethanol-, and heat-tolerance, were examined for isolate. The Quality of fermented cocoa bean was analyzed by cut test. Candida tropicalis KLK4 was tolerance to low pH value, high temperature, ethanol and could assimilate citrate, reflecting it is able to adapt to cocoa fermentation environment. The utilization of Candida tropicalis KLK4 strain as a starter culture for cocoa fermentation can enhance cocoa Â quality.

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Microbiological and Physicochemical Characterization of Small-Scale Cocoa Fermentations and Screening of Yeast and Bacterial Strains To Develop a Defined Starter Culture

Applied and Environmental Microbiology ◽

10.1128/aem.01144-12 ◽

2012 ◽

Vol 78 (15) ◽

pp. 5395-5405 ◽

Cited By ~ 86

Author(s):

Gilberto Vinícius de Melo Pereira ◽

Maria Gabriela da Cruz Pedrozo Miguel ◽

Cíntia Lacerda Ramos ◽

Rosane Freitas Schwan

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Citric Acid ◽

Dominant Species ◽

Starter Culture ◽

Physicochemical Characterization ◽

Cocoa Bean ◽

Small Scale ◽

Content Type ◽

Cocoa Bean Fermentation

ABSTRACTSpontaneous cocoa bean fermentations performed under bench- and pilot-scale conditions were studied using an integrated microbiological approach with culture-dependent and culture-independent techniques, as well as analyses of target metabolites from both cocoa pulp and cotyledons. Both fermentation ecosystems reached equilibrium through a two-phase process, starting with the simultaneous growth of the yeasts (withSaccharomyces cerevisiaeas the dominant species) and lactic acid bacteria (LAB) (Lactobacillus fermentumandLactobacillus plantarumwere the dominant species), which were gradually replaced by the acetic acid bacteria (AAB) (Acetobacter tropicaliswas the dominant species). In both processes, a sequence of substrate consumption (sucrose, glucose, fructose, and citric acid) and metabolite production kinetics (ethanol, lactic acid, and acetic acid) similar to that of previous, larger-scale fermentation experiments was observed. The technological potential of yeast, LAB, and AAB isolates was evaluated using a polyphasic study that included the measurement of stress-tolerant growth and fermentation kinetic parameters in cocoa pulp media. Overall, strainsL. fermentumUFLA CHBE8.12 (citric acid fermenting, lactic acid producing, and tolerant to heat, acid, lactic acid, and ethanol),S. cerevisiaeUFLA CHYC7.04 (ethanol producing and tolerant to acid, heat, and ethanol), andAcetobacter tropicalisUFLA CHBE16.01 (ethanol and lactic acid oxidizing, acetic acid producing, and tolerant to acid, heat, acetic acid, and ethanol) were selected to form a cocktail starter culture that should lead to better-controlled and more-reliable cocoa bean fermentation processes.

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Dynamics and Biodiversity of Populations of Lactic Acid Bacteria and Acetic Acid Bacteria Involved in Spontaneous Heap Fermentation of Cocoa Beans in Ghana

Applied and Environmental Microbiology ◽

10.1128/aem.02189-06 ◽

2007 ◽

Vol 73 (6) ◽

pp. 1809-1824 ◽

Cited By ~ 187

Author(s):

Nicholas Camu ◽

Tom De Winter ◽

Kristof Verbrugghe ◽

Ilse Cleenwerck ◽

Peter Vandamme ◽

...

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Lactic Acid Bacteria ◽

Fermentation Process ◽

Denaturing Gradient Gel Electrophoresis ◽

Acetic Acid Bacteria ◽

Cocoa Bean ◽

Rrna Gene ◽

Cocoa Bean Fermentation ◽

Leuconostoc Pseudomesenteroides

ABSTRACT The Ghanaian cocoa bean heap fermentation process was studied through a multiphasic approach, encompassing both microbiological and metabolite target analyses. A culture-dependent (plating and incubation, followed by repetitive-sequence-based PCR analyses of picked-up colonies) and culture-independent (denaturing gradient gel electrophoresis [DGGE] of 16S rRNA gene amplicons, PCR-DGGE) approach revealed a limited biodiversity and targeted population dynamics of both lactic acid bacteria (LAB) and acetic acid bacteria (AAB) during fermentation. Four main clusters were identified among the LAB isolated: Lactobacillus plantarum, Lactobacillus fermentum, Leuconostoc pseudomesenteroides, and Enterococcus casseliflavus. Other taxa encompassed, for instance, Weissella. Only four clusters were found among the AAB identified: Acetobacter pasteurianus, Acetobacter syzygii-like bacteria, and two small clusters of Acetobacter tropicalis-like bacteria. Particular strains of L. plantarum, L. fermentum, and A. pasteurianus, originating from the environment, were well adapted to the environmental conditions prevailing during Ghanaian cocoa bean heap fermentation and apparently played a significant role in the cocoa bean fermentation process. Yeasts produced ethanol from sugars, and LAB produced lactic acid, acetic acid, ethanol, and mannitol from sugars and/or citrate. Whereas L. plantarum strains were abundant in the beginning of the fermentation, L. fermentum strains converted fructose into mannitol upon prolonged fermentation. A. pasteurianus grew on ethanol, mannitol, and lactate and converted ethanol into acetic acid. A newly proposed Weissella sp., referred to as “Weissella ghanaensis,” was detected through PCR-DGGE analysis in some of the fermentations and was only occasionally picked up through culture-based isolation. Two new species of Acetobacter were found as well, namely, the species tentatively named“ Acetobacter senegalensis” (A. tropicalis-like) and “Acetobacter ghanaensis” (A. syzygii-like).

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Profil Pertumbuhan Mikroorganisme pada Fermentasi Biji Kakao Aceh

Jurnal Teknologi dan Industri Pertanian Indonesia ◽

10.17969/jtipi.v9i1.5975 ◽

2017 ◽

Vol 9 (2) ◽

pp. 50-54

Author(s):

Murna Muzaifa ◽

Yusya Abubakar ◽

Faitzal Haris

Keyword(s):

Acetic Acid ◽

Fermentation Process ◽

Acetic Acid Bacteria ◽

Cocoa Bean ◽

Chemical Changes ◽

Crucial Step ◽

Growth Profile ◽

Cocoa Bean Fermentation

Fermentation process is the most crucial step in the formation of the flavor and aroma of the cocoa bean. Cocoa bean fermentation triggers an array of chemical changes within the bean.These chemical changes are vital to the development of the complex and much-loved flavour known as “chocolate”. Fermentation involves a number of specific microorganisms that play a role during fermentation. The aim of this research was to analized microorganism growth profil of Aceh cacao during fermentation. Fermentation was conducted on 6 days with different aerations (agitation every 24 and 48 hours). The result showed that growth profile of microorganism during fermentation relatively had similar trend. Yeast dominated on the early fermentation, lactid bacteria reached the higest population on day 3 and acetic acid bacteria on day 4. Better quality of fermented cacao was resulted on every 48 hours of agitation that reached 70,19% of full fermentation.

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Designing an Arduino-based Automatic Cocoa Fermentation Tool

SinkrOn ◽

10.33395/sinkron.v5i1.10611 ◽

2020 ◽

Vol 5 (1) ◽

Author(s):

Dirja Nur Ilham ◽

Balkhaya Balkhaya ◽

Rudi Arif Candra ◽

Hardisal Hardisal ◽

Hasbaini Hasbaini

Keyword(s):

Temperature Sensor ◽

Dc Motor ◽

Cocoa Bean ◽

Cocoa Beans ◽

Cocoa Bean Fermentation ◽

Unpleasant Odor ◽

Working Principle ◽

The Right ◽

Cocoa Farmers ◽

Cocoa Fermentation

Automatic cocoa fermentation design is expected to facilitate the work of cocoa farmers during the process of reversing and stirring cocoa fermentation based on the right temperature. Fermentation process is of course done in a box or sack so that chocolate quickly produces heat and is cemented. However, in certain conditions, especially when in sacks there are often obstacles in the stirring process. Often the fermented chocolate experiences weathering or moldyness due to the uneven reversal that causes chocolate to clot, causing weathering or moldiness and produce an unpleasant odor and unattractive color on the cocoa beans. To overcome this problem a tool that automatically can turn or stir the cocoa beans evenly. This device is controlled by Arduino Uno R3 with a sensor that is an LM35 temperature sensor and has an LCD output and DC motor. This tool uses Relay to adjust the delay when driving a DC motor. The working principle of this tool, when the LM35 temperature sensor receives heat conditions on the cocoa beans, the LCD will display the condition of the temperature while the relay will instruct the DC motor to move the Cocoa Fermentation rail rotating left or right. The purpose of making this tool is to create a tool that can help alleviate the work of cocoa farmers in cocoa bean stirring activities at the time of cocoa bean fermentation controlled by Arduino.

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Oxidation of Metabolites Highlights the Microbial Interactions and Role ofAcetobacter pasteurianusduring Cocoa Bean Fermentation

Applied and Environmental Microbiology ◽

10.1128/aem.03344-13 ◽

2014 ◽

Vol 80 (6) ◽

pp. 1848-1857 ◽

Cited By ~ 64

Author(s):

Frédéric Moens ◽

Timothy Lefeber ◽

Luc De Vuyst

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Starter Culture ◽

Acetic Acid Bacterium ◽

Microbial Interactions ◽

Cocoa Bean ◽

Acid Oxidation ◽

Functional Roles ◽

Cocoa Bean Fermentation ◽

Oxidation Of Ethanol

ABSTRACTFour cocoa-specific acetic acid bacterium (AAB) strains, namely,Acetobacter pasteurianus386B,Acetobacter ghanensisLMG 23848T,Acetobacter fabarumLMG 24244T, andAcetobacter senegalensis108B, were analyzed kinetically and metabolically during monoculture laboratory fermentations. A cocoa pulp simulation medium (CPSM) for AAB, containing ethanol, lactic acid, and mannitol, was used. All AAB strains differed in their ethanol and lactic acid oxidation kinetics, whereby onlyA. pasteurianus386B performed a fast oxidation of ethanol and lactic acid into acetic acid and acetoin, respectively. OnlyA. pasteurianus386B andA. ghanensisLMG 23848Toxidized mannitol into fructose. Coculture fermentations withA. pasteurianus386B orA. ghanensisLMG 23848TandLactobacillus fermentum222 in CPSM for lactic acid bacteria (LAB) containing glucose, fructose, and citric acid revealed oxidation of lactic acid produced by the LAB strain into acetic acid and acetoin that was faster in the case ofA. pasteurianus386B. A triculture fermentation withSaccharomyces cerevisiaeH5S5K23,L. fermentum222, andA. pasteurianus386B, using CPSM for LAB, showed oxidation of ethanol and lactic acid produced by the yeast and LAB strain, respectively, into acetic acid and acetoin. Hence, acetic acid and acetoin are the major end metabolites of cocoa bean fermentation. All data highlight thatA. pasteurianus386B displayed beneficial functional roles to be used as a starter culture, namely, a fast oxidation of ethanol and lactic acid, and that these metabolites play a key role as substrates forA. pasteurianusin its indispensable cross-feeding interactions with yeast and LAB during cocoa bean fermentation.

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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

Applied and Environmental Microbiology ◽

10.1128/aem.00594-11 ◽

2011 ◽

Vol 77 (18) ◽

pp. 6694-6698 ◽

Cited By ~ 33

Author(s):

Timothy Lefeber ◽

Maarten Janssens ◽

Frédéric Moens ◽

William Gobert ◽

Luc De Vuyst

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Citric Acid ◽

Lactic Acid Bacteria ◽

Starter Culture ◽

Lactobacillus Fermentum ◽

Cocoa Bean ◽

Bacterial Strains ◽

Content Type ◽

Cocoa Bean Fermentation

ABSTRACTAmong various lactic acid bacterial strains tested, cocoa-specific strains ofLactobacillus fermentumwere 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.

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PENGARUH PENAMBAHAN STARTER MIKROBA SERTA PEMERASAN PULP TERHADAP KONDISI FERMENTASI DAN MUTU BIJI KAKAO (Theobroma cacao L.)

Jurnal Penelitian Pascapanen Pertanian ◽

10.21082/jpasca.v13n3.2016.156-166 ◽

2018 ◽

Vol 13 (3) ◽

pp. 156

Author(s):

S. Joni Munarso ◽

Kun Tanti Dewandari ◽

Zahra Haifa

Keyword(s):

Acetic Acid ◽

Lactobacillus Plantarum ◽

Acid Concentration ◽

Theobroma Cacao ◽

Starter Culture ◽

Acetic Acid Concentration ◽

Acetobacter Aceti ◽

Cocoa Beans ◽

Theobroma Cacao L

Telah dilakukan penelitian yang bertujuan untuk mengetahui pengaruh penambahan starter mikroba (Acetobacter aceti, Lactobacillus plantarum dan Saccharomyces cereviceae) serta pemerasan pulp terhadap fermentasi dan mutu biji kakao. Penelitian menggunakan metode Rancangan Acak Lengkap (RAL) pola faktorial 3x5 dengan dua kali ulangan. Faktor pertama adalah fermentasi yaitu fermentasi biji kakao secara spontan (F1), Fermentasi biji kakao dengan penambahan A. aceti, L. plantarum dan S. cereviciae (F2), Fermentasi biji kakao dengan perlakuan pemerasan pulp serta penambahan A. aceti, L. plantarum dan S. cereviciae (F3). Sedangkan faktor kedua adalah lama fermentasi (H1, H2, H3, H4 dan H5). Hasil penelitian menunjukkan bahwa penambahan starter meningkatkan konsentrasi etanol pada saat fermentasi dan meningkatkan kadar asam asetat, tetapi menurunkan konsentrasi asam oksalat pada biji kakao. Penambahan starter disertai pemerasan pulp menghasilkan biji kakao dengan kadar asam asetat sebesar 0,47%, sedangkan biji kakao tanpa pemerasan kadar asam asetat 0,49%. Penambahan starter disertai pemerasan pulp menghasilkan mutu biji kakao terbaik dengan karakteristik sebagai berikut: skor nilai uji belah tertinggi (379 dari 400), mutu fisik (Golongan mutu A) serta memenuhi persyaratan mutu SNI 2008 No. 2323 tentang biji kakao dengan rasio jumlah per berat biji sebanyak 88 biji/100g; nilai pH 4,93; kadar asam asetat 0,47%, kadar lemak 34,90%, kadar air 4,47%, kadar serat kasar 3,66% dan kadar abu 4,82% dengan waktu fermentasi selama 5 hari.English Version AbstractEffect of Starter Culture Addition and Depulping on The Fermentation and Quality of Cocoa Beans (Theobroma cacao L.)S Joni Munarso, Kun Tanti Dewandari, and Zahra Haifa. 2016. Effect of Starter Culture Addition and Depulping on The Fermentation and Quality of Cocoa Beans (Theobroma cacao L.). The aimed of this study was to investigate the effect of starter culture addition (Acetobacter aceti, Lactobacillus plantarum, and Saccharomyces cerevisiae) with depulping on the fermentation and quality of cocoa beans. The experimental design of this study was conducted using a 3×5 factorial Completely Randomized Design (CRD) with duplicate replication. The first factor was fermentation condition included spontaneously fermented cocoa beans (F1), fermentation of cocoa beans with the addition of A. aceti, L. plantarum and S. cereviciae (F2), Fermented cocoa beans with depulping and addition of A. aceti, L. plantarum and S. Cereviciae (F3). The second factor was time of fermentation. The result revealed that starter addition increased ethanol concentration on the fermentation process, increased acetate acid, and citric acid concentratio, meanwhile oxalic acid decreased on cocoa beans during 5 days of fermentation. Depulping caused a slight decrease in acetic acid concentration at the end of fermentation with value of 0,47%, meanwhile the sample of cocoa beans without depulping treatment had acetic acid concentration of 0,49%. Starter culture addition and depulping treatment resulted the best characteristic of cocoa beans which visualized by the largest amounts of cut test score (379 of 400), physical quality (Grade A) and completed SNI No. 2323-2008 requirements with total beans/100 g ratio of 88 beans/100g; pH values of 4,93; acetic acid concentrations of 0,47%, content of fat 15,12%, moisture 4,47%, crudefiber 3,66% and total ash 4,82% after 5 days fermentation.

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