Non-conventional fermentation at laboratory scale of cocoa beans: Using probiotic microorganisms and substitution of mucilage by fruit pulps

In a non-conventional lab-scale fermentation of cocoa beans using probiotic microorganisms and substituting the cocoa pulp for fruit pulp, physicochemical, microbiological and quality parameters were investigated. Two hundred grams of beans were fermented in a controlled environmental chamber (temperature ramp rate of 25°C for 48 h; 35°C for 48 h and 45°C for 48 h; and 65% HR). pH, titratable acid, citric, lactic and acetic acids, as well as sugars and ethanol were measured. A cut test was also performed on the cocoa beans fermented 5 and 6 days. As the fermentation time progressed, citric acid concentration decreased until 0.53 g kg⁻¹, whereas both lactic and acetic acids increased until 0.44 and 16.58 g kg⁻¹, respectively. Sucrose content decreased from 12.26 g kg⁻¹ (in fresh) to 6.54 g kg⁻¹ on the 6th day. Fructose and glucose contents increased in the cotyledons from day five, reaching a maximum concentration of 1.14 and 1.01 g kg⁻¹, respectively, on day six. Yeasts were the main microorganisms during the first 24–48 h (8.4 log CFU g⁻¹), while bacterial counts reached its highest number (7.8 log CFU g⁻¹) on day four. Beans fermented 5 and 6 days resulted in more fermented beans (>81%) and less violet ones (<18.4%) than the control.     

Effect of post-harvest treatments on the occurrence of ochratoxin A in raw cocoa beans

Cocoa beans are the principal raw material for chocolate manufacture. Moulds have an important place in the change in the quality of cocoa beans due to their role in the production of free fatty acids and mycotoxins, namely ochratoxin A (OTA). This study investigated the impact of the key post-harvest treatments, namely the fermentation and drying methods on OTA contamination of raw cocoa beans. Analytical methods for OTA detection were based on solid–liquid extraction, clean-up using an immunoaffinity column, and identification by reversed-phase HPLC with fluorescence detection. Of a total of 104 randomly selected cocoa samples analysed, 32% had OTA contents above 2 µg kg^–1. Cocoa sourced from pods in a bad state of health had a maximum OTA content of 39.2 µg kg^–1, while that obtained from healthy pods recorded 11.2 µg kg^–1. The production of OTA in cocoa beans increased according to the pod-opening delay and reached 39.2 µg kg^–1 after an opening delay of 7 days after harvest, while 6.1 and 11.2 µg kg^–1 were observed when pods were opened after 0 and 4 days. OTA production also seemed to depend considerably to the cocoa fermentation materials. When using plastic boxes for bean fermentation, the OTA production was enhanced and reached an average OTA content of about 4.9 µg kg^–1, while the raw cocoa treated in banana leaves and wooden boxes recorded 1.6 and 2.2 µg kg^–1 on average respectively. In parallel, the OTA production was not really influenced by either the mixing or the duration of the fermentation or the drying materials.     

Oxidation of polyphenols in unfermented and partly fermented cocoa beans by cocoa polyphenol oxidase and tyrosinase

Unfermented and partly fermented dried cocoa beans have an excessively astringent and bitter flavour owing to their high polyphenol content. Studies on the remaining polyphenol oxidase activity and the oxidation of polyphenols in these beans have been conducted in relation to two factors, ie incubation time (0, 1, 2, 4, 8 and 16 h) and pH of incubation (3.5, 4.5, 5.5 and 6.5). Owing to unsuccessful polyphenol oxidation during incubation of partly fermented beans, incubation–enrichment treatments were carried out by combining incubation time (0, 8, 16, 24 and 32 h) and addition of crude cocoa polyphenol oxidase and purified tyrosinase from mushroom at 88 and 8800 units g^?1 and pH 5.5. Results showed that the remaining polyphenol oxidase activities of unfermented and partly fermented dried beans were 1 and 0.08% with specific activities of 9 and 1% respectively. The polyphenols of unfermented beans were effectively oxidised by incubation at 45 °C and pH 3.5–6.5 without enzyme enrichment; while those of partly fermented beans required enzyme enrichment. Both crude cocoa polyphenol oxidase and tyrosinase could be used for enzyme enrichment, but tyrosinase seemed to be more effective. © 2002 Society of Chemical Industry     

Improved Fermentation of Cocoa Beans with Enhanced Aroma Profiles

The cocoa (Theobroma cacao) fermentation was carried out using starter consortia of Saccharomyces cerevisiae, Lactobacillus plantarum, and Acetobacter aceti with 10% inoculum. The fermented beans were soaked with sterile water (1:3 wt/v) for the duration of 1, 3, and 5 h. The treated beans were dried to the moisture content of 7 ± 1%. The samples were then roasted, ground and subjected to steam distillation. The volatiles were extracted and identified using GC-MS. The samples soaked for 1 h scored 100% cut score along with 0.85% fermentation index. The volatile and non-volatiles were associated with beans treated in water after fermentation. Volatile compounds were classified as esters, alcohols, acids, aldehydes, ketones, pyrazines, hydroperoxides, and hydrocarbons. Most of the oxygenated aliphatic compounds and aromatics were potentially derived via an acetic acid and shikimic acid pathways, respectively. Untreated cocoa beans contained 12 sesquiterpene hydrocarbons potentially derived through the mevalonic acid pathway and contributed to harsh notes. Also, these metabolites were completely absent in the treated samples indicating depletion while soaking. Thus, the treated samples (1 h) had enhanced quality with high flavor notes with 8.5 rating on sensory evaluation. Thus, these soaking treatments with fermentation lead to improvement of flavor profiles which confers improved cocoa quality.     

Quantification of Catechin and Epicatechin in Foods by Enzymatic-Spectrophotometric Method with Tyrosinase

Catechin and epicatechin are important phenolic compounds found mainly in tea leaves, cocoa beans, and in the peels and seeds of fruits. The reliable detection of these compounds by spectrophotometry is limited by the interference with substances that absorb light at similar wavelengths; thus, an effective alternative is to detect the oxidation products of these compounds at more specific wavelengths. This study aimed to develop an analytical method to quantify total catechin and epicatechin in some food samples (cocoa beans, guarana powder, apples, and tea leaves) by enzymatic oxidation with tyrosinase. The samples were extracted in an ultrasonic bath and the purified extracts were oxidized with tyrosinase solution. The quinones formed in the reaction were detected by spectrophotometry. The method was selective and presented linearity between 1.21 and 7.26 mg g^?1, limit of detection of 0.48 mg g^?1, and limit of quantification of 1.61 mg g^?1. The accuracy of the proposed method was reasonable, with recoveries between 84.3 and 90.7% in the fortified matrices. High precision was observed, with low coefficients of variation for repeatability (3.17 to 3.68%) and intermediate precision (3.27%). In cocoa beans, the total catechin and epicatechin level was 1.65 mg g^?1 (3.58 mg g^?1). The successful application of the method was also demonstrated in guarana powder. Therefore, the method can be applicable for spectrophotometric analysis of catechin and epicatechin in samples containing above 1.61 mg g^?1 of these compounds.     

Physicochemical properties,antioxidant content,volatile organic compounds and sensory profile of cocoa beans fermented with yeast starter cultures

This study determined the influence of selected yeast starters on the antioxidant content, volatile organic compounds and sensory qualities of fermented cocoa beans. Cocoa beans fermented with Hanseniaspora thailandica (MH979675), Pichia kudriazevii (MH979681), or a mixture of both species in fermentation (20 kg) were highly reproducible based on the physicochemical parameters (temperature, pH of nibs, fermentation index and cut test). The usage of H. thailandica improved the total polyphenols content of cocoa beans with fermentation index around 1.24 and 1.33. The correlation analyses suggested that phenolics and flavonoids compounds may not be the main constituents that led to the antioxidant activities of the fermented samples. Principal Component Analysis showed distinct differences in VOC profiles between spontaneous and fermentation using yeast. Sweet and spicy notes were only found in sample fermented with yeast. The inoculation of yeast in fermentation influenced the sensorial and antioxidant properties of cocoa beans.     

Effects of incubation and polyphenol oxidase enrichment on colour, fermentation index, procyanidins and astringency of unfermented and partly fermented cocoa beans

Incubation of unfermented and partly fermented cocoa beans in acetate buffer, pH 5.5, at 45 °C increased yellowness, total colour differences and fermentation index value of the cocoa bean powders and decreased cocoa procyanidins (monomers to pentamers), and their astringency. Fermentation index and (–)‐epicatechin content, equivalent to those of fully fermented beans, were reached by unfermented beans after 4–8‐h incubation, but not by partly fermented beans even after 16 h. During incubation of partly fermented cocoa beans enriched with polyphenol oxidase, yellowness and fermentation index value were increased, whilst (–)‐epicatechin was decreased. Tyrosinase had a less significant effect in yellow colour formation, but showed a significant reduction of (–)‐epicatechin and increase in fermentation index compared with crude cocoa polyphenol oxidase. However, both enzymes have similar effects on procyanidin degradation and astringent taste reduction. Incubation of cocoa beans for 16 h increased the cut test score of unfermented and partly fermented beans by 50 and 30%, respectively.     

Fungi Level Analysis of Cocoa Beans Based on Fermentation Box Type and Duration

In certain location, it is undeniable that there are qualities variations of fermented cocoa beans depend on processing techniques. In addition, its cocoa bean's production central at Yogyakarta have their specific characteristic own regarding the use of fermentation box (box type). The purpose of this research was to analyze the effects of three types of fermentation box and duration to fungi level of fermented cocoa beans. CRD (completely randomized design) with two factors and three replications is applied as assessment method. The first factor is the type of fermentation box (basket, storey box and single box with a hand crank) and the second factor is the length of fermentation time (3 and 5 days). The result of the research explained that the lowest levels of the fungi achieve within 5 days of fermentation processes. Meanwhile, a single box with a hand crank provide the lowest of fungi's level. Furthermore, the combination of those two results may give the most favorably expected condition of fungi level.     

Effect of milk fat replacement by polyunsaturated fatty acids on the microbiological,rheological and sensorial properties of fermented milks

A modified milk (W3DD) where fat had been replaced by oils enriched in ω‐3 polyunsaturated fatty acids was used for the manufacture of a set‐type fermented product. In order to improve the organoleptic properties of the product, 30 g l^?1 whey protein concentrate (WPC) was added during the manufacturing process. Samples were fermented employing a commercial probiotic starter culture (ABT‐2), which contained Streptococcus thermophilus ST‐20Y, Lactobacillus acidophilus LA‐5 and Bifidobacterium lactis BB‐12. The acidification process was dependent on the WPC addition, which favoured the increase of viable counts, but fermentation was not influenced by the milk fat composition. The highest counts of the probiotic strains, L acidophilus LA‐5 (3.3 × 10⁵ cfu g^?1) and B lactis BB‐12 (5.5 × 10⁷ cfu g^?1), after 21 days of storage at 4 °C, were found in fermented products derived from W3DD supplemented with WPC. Addition of WPC also increased the firmness of the products and reduced syneresis. No apparent colour changes due to fat composition or WPC supplementation were observed in the products. Milk fat replacement by oils rich in ω‐3 polyunsaturated fatty acids had a negative influence on the product texture but did not affect the typical yoghurt flavour. These defects were overcome by the addition of 30 g l^?1 WPC, which improved the appearance, texture and general acceptability scores in the product. Copyright © 2004 Society of Chemical Industry     

Ghanaian Cocoa Bean Fermentation Characterized by Spectroscopic and Chromatographic Methods and Chemometrics

Abstract: Export of cocoa beans is of great economic importance in Ghana and several other tropical countries. Raw cocoa has an astringent, unpleasant taste, and flavor, and has to be fermented, dried, and roasted to obtain the characteristic cocoa flavor and taste. In an attempt to obtain a deeper understanding of the changes in the cocoa beans during fermentation and investigate the possibility of future development of objective methods for assessing the degree of fermentation, a novel combination of methods including cut test, colorimetry, fluorescence spectroscopy, NIR spectroscopy, and GC-MS evaluated by chemometric methods was used to examine cocoa beans sampled at different durations of fermentation and samples representing fully fermented and dried beans from all cocoa growing regions of Ghana. Using colorimetry it was found that samples moved towards higher a* and b* values as fermentation progressed. Furthermore, the degree of fermentation could, in general, be well described by the spectroscopic methods used. In addition, it was possible to link analysis of volatile compounds with predictions of fermentation time. Fermented and dried cocoa beans from the Volta and the Western regions clustered separately in the score plots based on colorimetric, fluorescence, NIR, and GC-MS indicating regional differences in the composition of Ghanaian cocoa beans. The study demonstrates the potential of colorimetry and spectroscopic methods as valuable tools for determining the fermentation degree of cocoa beans. Using GC-MS it was possible to demonstrate the formation of several important aroma compounds such 2-phenylethyl acetate, propionic acid, and acetoin and the breakdown of others like diacetyl during fermentation. Practical Application: The present study demonstrates the potential of using colorimetry and spectroscopic methods as objective methods for determining cocoa bean quality along the processing chain. Development of objective methods for determining cocoa bean quality will be of great importance for quality insurance within the fields of cocoa processing and raw material control in chocolate producing companies.     

On-farm implementation of a starter culture for improved cocoa bean fermentation and its influence on the flavour of chocolates produced thereof

Cocoa bean fermentations controlled by means of starter cultures were introduced on several farms in two different cocoa-producing regions (West Africa and Southeast Asia). Two starter culture mixtures were tested, namely one composed of Saccharomyces cerevisiae H5S5K23, Lactobacillus fermentum 222, and Acetobacter pasteurianus 386B (three heaps and one box), and another composed of L. fermentum 222 and A. pasteurianus 386B (seven heaps and one box). In all starter culture-added cocoa bean fermentation processes, the inoculated starter culture species were able to outgrow the natural contamination of the cocoa pulp-bean mass and they prevailed during cocoa bean fermentation. The application of both added starter cultures resulted in fermented dry cocoa beans that gave concomitant milk and dark chocolates with a reliable flavour, independent of cocoa-producing region or fermentation method. The addition of the lactic acid bacterium (LAB)/acetic acid bacterium (AAB) starter culture to the fermenting cocoa pulp-bean mass accelerated the cocoa bean fermentation process regarding citric acid conversion and lactic acid production through carbohydrate fermentation. For the production of a standard bulk chocolate, the addition of a yeast/LAB/AAB starter culture was necessary. This enabled an enhanced and consistent ethanol production by yeasts for a successful starter culture-added cocoa bean fermentation process. This study showed possibilities for the use of starter cultures in cocoa bean fermentation processing to achieve a reliably improved fermentation of cocoa pulp-bean mass that can consistently produce high-quality fermented dry cocoa beans and flavourful chocolates produced thereof.     

ITS-RFLP characterization of black <Emphasis Type="Italic">Aspergillus</Emphasis> isolates responsible for ochratoxin A contamination in cocoa beans

In the present study, ochratoxigenic mycobiota in cocoa beans was identified at species level by digestion of the ITS products using the endonucleases HhaI, NlaIII and RsaI. Of the 132 isolates of Aspergillus section Nigri collected from cocoa beans, 89 were identified as A. tubingensis, 27 as A. niger, 10 as A. tubingensis-like and 6 as A. carbonarius. No variation was observed between RFLP patterns (C, N, T1 and T2) described previously for grape isolates and those of the cocoa isolates analysed. With respect to OTA-producing fungi, a high percentage of black aspergilli (50.7%) was able to produce OTA. Additionally, most of the OTA-producing isolates were of moderate toxigenicity, producing amounts of OTA from 10 μg g⁻¹ to 100 μg g⁻¹. Percentages of OTA-producing isolates in the A. niger aggregate were higher than in other substrates, ranging from 30% to 51.7%. Furthermore, the detected levels of OTA production in the A. niger aggregate, particularly in A. tubingensis species was higher than in A. carbonarius, ranging from 0.7 μg g⁻¹ to 120 μg g⁻¹ (mean 24.55 μg g⁻¹). Due to the high occurrence, percentage of ocratoxigenic isolates and their ability to produce OTA, isolates belonging to the A. niger aggregate could be considered as the main cause of OTA contamination in cocoa beans used for manufacturing cocoa products.     

Aflatoxins and ochratoxin A: occurrence and contamination levels in cocoa beans from Brazil

In the present study, the occurrence of aflatoxins (AFs) and ochratoxin A (OTA) was evaluated in 123 samples of cocoa beans produced in five Brazilian states. The presence of these mycotoxins was determined by high-performance liquid chromatography with fluorescence detection (HPLC-FLD) after immunoaffinity column clean-up. The mean level of total AFs in cocoa beans samples was 5.7 μg.kg^?1. Four (3.3%) samples exceeded the maximum limit of 10 μg.kg^?1 established by the Brazilian legislation for total AFs. The mean level of OTA contamination was 1.2 μg.kg^?1, and none of the samples exceeded the maximum limit established by the Brazilian legislation. The co-occurrence of AFs and OTA was observed in 4.9% of the samples. The results of the present study demonstrated that, in relation to the levels of AFs and OTA established by the Brazilian legislation, most samples of cocoa beans analyzed are safe for consumption. This is the first report on the occurrence and levels of AFs and OTA in cocoa beans from the five main Brazilian states producing cocoa. The data in this study provide important information for farmers, traders, industry, consumers and law enforcement agencies.     

Effect of fermentation by Bacillus subtilis on antioxidant and cytotoxic activities of black rice bran

Black rice bran was fermented with Bacillus subtilis KU3 isolated from Korean traditional food, Kimchi. Antioxidant and cytotoxic activities of the fermented black rice bran were investigated. Total phenolic and anthocyanin contents decreased from 171.54 mg GAE g^?1 and 2.31 mg g^?1 to 139.13 mg GAE g^?1 and 2.12 mg g^?1, respectively, after fermentation. Antioxidant activities determined by 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) radical scavenging, β‐carotene bleaching and ferric thiocyanate assay were correlated with total phenolic and anthocyanin contents. Non‐fermented black rice bran extract (NFBE) showed greater antioxidant activities than fermented black rice bran extract (FBE). Cytotoxic activities measured by MTT assay showed that both NFBE and FBE had over 50% activities. The cytotoxic activities of FBE against MCF‐7 and HeLa cells were 71.65% and 68.07%, respectively, at 8.0 mg mL^?1, but those of NFBE were lower than 50%. These results suggested that the cytotoxic activity of black rice bran improved through fermentation, while antioxidant activity reduced.     

Activation of remaining key enzymes in dried under-fermented cocoa beans and its effect on aroma precursor formation

Incubation-activation of remaining key enzymes in dried under-fermented cocoa beans and its effect on aroma precursor formation has been studied using defatted unfermented and partly fermented cocoa bean powders. Results of the study showed that aspartic endoprotease, carboxypeptidase and invertase were significantly inactivated during fermentation and drying, and the effect of fermentation was significantly lower than that of drying. The enzyme activities remaining in these beans were still sufficient to carry out enzymatic reaction during incubation. Peptide patterns, resulting from incubation of unfermented and partly fermented beans powders, were quite similar to the well-fermented patterns. Meanwhile, free amino acid concentrations of the unfermented beans were significantly increased during the first 4 h of incubation and then remained constant; however, with partly fermented beans, the formation continued and the hydrophobic and total free amino acid concentrations reached the value of well-fermented beans after 24 h of incubation. Reducing sugar concentrations of both unfermented and partly fermented cocoa beans could reach the level of well-fermented beans by incubation.     

Aflatoxins and ochratoxin A in different cocoa clones (Theobroma cacao L.) developed in the southern region of Bahia,Brazil

Brazil is the sixth largest producer of cocoa beans in the world, after Côte d’Ivoire, Ghana, Indonesia, Nigeria and Cameroon. The southern region of Bahia stands out as the country’s largest producer, accounting for approximately 60% of production. Due to damage caused by infestation of the cocoa crop with the fungus Moniliophthora perniciosa, which causes ‘witch’s broom disease’, research in cocoa beans has led to the cloning of species that are resistant to the disease; however, there is little information about the development of other fungal genera in these clones, such as Aspergillus, which do not represent a phytopathogenicity problem but can grow during the pre-processing of cocoa beans and produce mycotoxins. Thus, the aim of this work was to determine the presence of aflatoxin (AF) and ochratoxin A (OTA) in cocoa clones developed in Brazil. Aflatoxin and ochratoxin A contamination were determined in 130 samples from 13 cocoa clones grown in the south of Bahia by ultra-performance liquid chromatography with a fluorescence detector. The method was evaluated for limit of detection (LOD) (0.05–0.90 μg kg^?1), limit of quantification (0.10–2.50 μg kg^?1) and recovery (RSD) (89.40–95.80%) for AFB₁, AFB₂, AFG₁, AFG₂ and OTA. Aflatoxin contamination was detected in 38% of the samples in the range of ?1, with AFB₁ in 25% of the total samples, whereas ochratoxin A was positive in 18% of the samples in the range of ?1.     

Fermentation of cocoa beans: influence of microbial activities and polyphenol concentrations on the flavour of chocolate

BACKGROUND: Spontaneous cocoa bean fermentation is characterised by a succession of microbial activities. Cocoa flavour precursors are developed during fermentation and drying of cocoa beans. Polyphenols and alkaloids contribute to astringency and bitterness of cocoa and chocolate. RESULTS: Population dynamics, metabolite target analyses, and chocolate production were performed for seven independent spontaneous cocoa bean heap fermentations in Ghana. Although the same micro‐organisms were involved in these heaps, carried out at different farms or in different seasons, heap temperatures and microbial metabolite concentrations were different. This could be due to heterogeneity and size of the heaps, but was mainly ascribed to microbial variability. Indeed, differences in microbial activity could be linked with the flavour of chocolates made from the corresponding dried, fermented cocoa beans. Whereas the polyphenol and alkaloid contents of cocoa beans were crop‐ and heap‐dependent, epicatechin and theobromine levels decreased during fermentation due to diffusion out of the bean cotyledons and polyphenol oxidation and condensation. Residual levels were responsible for the degree of bitterness of the final chocolates. CONCLUSION: Differences in microbial activities between different heap fermentations can result in dried fermented cocoa beans and chocolates with different flavour characteristics. Hence, fermentation control may direct the flavour of chocolate. Copyright © 2008 Society of Chemical Industry     

发酵和焙烤对可可豆多酚、黄酮和风味品质的影响

本实验采用高效液相色谱仪（high performance liquid chromatography,HPLC）、色差仪、电子鼻分别检测未发酵、发酵和焙烤海南可可豆的总酚含量、总黄酮含量、色度和可可豆风味差异。结果表明：未经焙烤的未发酵豆总酚（464.03 mg/10 g）和总黄酮（126.86 mg/10 g）含量明显高于发酵豆总酚（211.86 mg/10 g）和总黄酮（61.98 mg/10 g）含量。105～145 ℃焙烤30 min,未发酵豆总酚和总黄酮含量分别为419.5～129.8 mg/10 g和77.8～16.8 mg/10 g;发酵豆总酚和总黄酮含量分别为182.53～86.25 mg/10 g和34.7～7.0 mg/10 g。其中,125 ℃焙烤20～40 min,未发酵豆总酚和黄酮含量分别为353.74～289.45 mg/10 g和42.86～32.20 mg/10 g;发酵豆总酚和黄酮含量分别为152.08～123.55 mg/10 g和25.12～21.14 mg/10 g。在105～145 ℃的温度焙烤下没食子酸含量变化显著。可可豆色度值的范围：L*值集中在40.0～47.0之间,a*值集中在5.0～6.8之间,b*值集中在4.0～8.5之间。未发酵和发酵可可豆之间,以及不同温度焙烤可可豆之间的电子感官风味分析结果差异较大。     

Development of a spray dried probiotic yoghurt containing Lactobacillus paracasei NFBC 338

The aim of this study was to monitor viability of probiotic Lactobacillus paracasei NFBC 338 during: (a) two-stage yoghurt fermentation with starter cultures Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, (b) following spray drying, and (c) during storage for 42 days. During the initial fermentation phase (10 h), probiotic Lactobacillus numbers increased 7-fold to 3.9 × 10⁹ cfu g^?1 and these numbers were maintained during fermentation for a further 3 h in the presence of the yoghurt starters. Following spray-drying, the probiotic culture survived best, followed by S. thermophilus and L. delbrueckii subsp. bulgaricus (yielding 3.4 × 10⁸, 1.2 × 10⁸ and 4.0 × 10⁵ cfu g^?1 powder, respectively). L. paracasei NFBC 338 and S. thermophilus were stable during storage at 4 °C and 15 °C (for 42 days) with viable counts exceeding 10⁷ cfu g^?1, while viability of L. delbrueckii subsp. bulgaricus decreased considerably throughout storage.     

Characteristics of fermentation and bioavailability of isoflavones in Korean soybean paste (doenjang) with application of Bacillus sp. KH-15

We attempted to increase the fibrinolytic activity and aglycone contents of isoflavone in doenjang prepared with Bacillus sp. KH‐15. An initial rapid increase of viable cells was observed in the first 30 days followed by a gradual decrease to 1.78 × 10⁸ CFU g^?1 wet weight. The amino type nitrogen sharply was increased after 15 days, and showed the highest level (788.7 mg%) after 90 days. The total nitrogen was increased from 2.21% to 2.52% of final content after 90 days. Soluble carbohydrate was increased to 32.9 mg g^?1 for 60 days, and after 60 days, the content was slowly decreased to 29.5 mg g^?1. A rapid increase in fibrinolytic and caseinolytic activities were observed in the first 30 and 45 days of fermentation followed by a gradual decrease to 2.54 and 144.5 U g^?1, respectively. β‐Glucosidase activity also showed a similar change pattern of the caseinolytic and fibrinoytic activities. Daidzein and genistein were increased to the maximum level of 764.5 and 561.4 mg kg^?1 at day 30, respectively. At day 90, the contents of daidzein and genistein were 661.5 and 524.9 mg kg^?1, respectively. The rats were fed a diet supplemented with either doenjang powder prepared with Bacillus sp. KH‐15 (D‐1 group) or commercial doenjang powder (D‐2 group) for 40 days. The level of serum total isoflavones in the D‐1 group was significantly higher than that in the D‐2 group.