Traditional Fermented Soybean Research Articles

Lactococcus lactis subsp. lactis LJL7m20 strongly inhibits growth of the Bacillus cereus group from sufu

Sufu, a traditional fermented soybean curd, is prone to contamination by Bacillus cereus. In this study, 133 lactic acid bacteria (LAB) strains were initially screened from spring and winter Mao-tofu samples for their ability to inhibit the growth of B. cereus ATCC 14579T. Among these strains, five Lactococcus lactis strains exhibited a good safety profile and were capable of growing in pH 4–9 and salt concentrations of 6%–8%. Antimicrobial experiments conducted on the B. cereus group from sufu factories showed that L. lactis LJL7m20 strain had the strongest inhibitory effect. Whole-genome sequencing revealed the presence of three genes associated with high osmotic stress response and eight distinct two-component systems that may contribute to the strain's tolerance, enhancing its survival in high-salt environments. The strain was further identified as L. lactis subsp. lactis through a combination of phenotypic and genotypic analysis. Additionally, bacteriocins nisin Z and nisin A/Q were identified using the BAGEL4 and antiSMASH databases. ELISA further confirmed the strain's ability to produce nisin. A fermentation validation study demonstrated that inoculation with the strain contributed to reduce the number of B. cereus group in sufu pehtzes. This study provides a theoretical foundation for the use of new biocontrol bacteria to improve the safety of traditional fermented soybean foods.

Thai Fermented Soybean (Thua-Nao) Prevents Early Stages of Colorectal Carcinogenesis Induced by Diethylnitrosamine and 1,2-Dimethylhydrazine Through Modulations of Cell Proliferation and Gut Microbiota in Rats.

Thua-nao is a traditional fermented soybean product widely consumed in the northern areas of Thailand. There has been little research on the biological activity of Thua-nao, particularly its anticancer properties. The objective of this study was to examine the cancer chemopreventive effects of dried Thua-nao on liver and colorectal carcinogenesis induced by carcinogens in rats. Rats were injected with diethylnitrosamine (DEN) and 1,2-dimethylhydrazine (DMH) to induce preneoplastic lesions. Rats orally received dried Thua-nao for 13 weeks. The preneoplastic lesions, including glutathione S-transferase placental form (GST-P)-positive foci and aberrant crypt foci (ACF), were evaluated in the liver and colon, respectively. The cancer chemopreventive mechanisms of dried Thua-nao on liver and colorectal carcinogenesis were examined. Dried Thua-nao administration suppressed colorectal aberrant crypt foci. Moreover, dried Thua-nao reduced proliferation cell nuclear antigen (PCNA)-positive cells in the colon. Interestingly, dried Thua-nao modulated the gut microbiota in DEN- and DMH-induced rats. Isoflavones, including genistein and daidzein, represent promising chemopreventive agents in dried Thua-nao. In conclusion, these results highlight the cancer chemopreventive effect of dried Thua-nao in DEN and DMH-induced colorectal carcinogenesis through cell proliferation reduction and gut microbiota modulation.

A Review on Ethnic Manipuri Cuisine of Northeast India: A Description of its Traditional Ingredients, Therapeutic Potential and Food Preparation Techniques

Manipur is nestled in the northeastern part of India. The word Manipur means “land of diamonds” in Sanskrit. Jawaharlal Nehru, India’s first prime minister, referred to Manipur as the country’s “jewel.” It has a valley in the middle and is surrounded by nine mountain ranges. There are several different ethnic groups there. Because of its ethnic diversity and diverse landscape, Manipur is home to innumerable ethnic food that embodies the rich heritage and traditional flavors of the state. People from Manipur, among other things, have a propensity to consume a variety of leafy vegetables, which are common in the region. Amidst all the subsequent changes in eating patterns, Manipuri people continue to practice their culinary tradition. Traditional foods usually provide a good balance of bioactive compound and health benefits. The major ingredients of ethnic food have historically included rice, vegetables, fat (from ghee or oil) and some protein sources from traditional fermented soybean and fish. They are free from chemicals and abundant in nutrients like healthy fat, carbohydrate, vitamins and minerals. With the help of locally accessible vegetables, herbs and fermented food products, this review article aims to describe traditional knowledge of most popular food ingredients, therapeutic potential and nutritional properties of Manipur’s ethnic culinary culture.

Characterization of the Geographical Origin of the Traditional Chinese Fermented Soybean Douchi by Gas Chromatography-Mass Spectrometry (GC-MS) and Chemometrics

Douchi is a highly appreciated product due to its unique flavor, high nutritional value, and health benefits. Douchi from different regions exhibits distinct characteristics. The objective of this study was to investigate the volatile profiles in douchi produced from different geographical origins using gas chromatography-mass spectrometry (GC-MS) and chemometrics. The volatiles were extracted by headspace solid-phase micro-extraction (HS-SPME) and subsequently determined by GC-MS. A comparative analysis was further conducted to identify the variations in volatile compounds among the samples. The correlation between the samples was investigated by employing cluster analysis (CA) and principal component analysis (PCA). An orthogonal partial least squares-discriminant analysis (OPLS-DA) model was established to discriminate samples from different regions and identify key differential compounds. A total of 46 volatile compounds were detected, among which terpenes, alcohols were the main compounds. Furthermore, 14 key differentiating compounds were screened out based on the OPLS-DA modle. The application of GC-MS combined with chemometrics offers a theoretical framework for douchi quality control and classification.

Transcriptomic analysis of the metabolic characteristics of Aspergillus oryzae during the fermentation of doenjang-meju, a traditional Korean fermented soybean brick

To investigate the fermentative traits of Aspergillus oryzae in doenjang-meju, a traditional Korean fermented soybean brick, we prepared doenjang-meju bricks inoculated with both A. oryzae and Bacillus velezensis––recognized as crucial microbes in doenjang-meju fermentation, and then the transcriptomes of A. oryzae were analyzed throughout the entire fermentation period. The transcriptomic analysis of carbohydrate-active enzyme (CAZy) genes revealed that A. oryzae exhibited robust expression of CAZy genes involved in the decomposition of various carbohydrates, particularly cellulose, starch, and hemicellulose, indicating the active breakdown of soybean carbohydrates by A. oryzae during fermentation. Furthermore, KEGG analysis, metabolic pathway reconstruction, and their transcriptomic analysis provided a comprehensive overview of the decomposition patterns of various carbohydrates, lipids, and proteins and the metabolic features of various carbon compounds during fermentation. Additionally, the transcriptomic analysis of biosynthetic gene clusters indicated the potential production of various secondary metabolites by A. oryzae during fermentation. Genes related to carbohydrate and carbon compound metabolisms and secondary metabolite biosynthesis displayed unique and different transcriptional expression patterns depending on the specific gene and fermentation stage. Overall, our findings provide comprehensive insights into the metabolic characteristics of A. oryzae, thus contributing to our understanding of soybean fermentation.

A novel ACE inhibitory peptide from Douchi hydrolysate: Stability, inhibition mechanism, and antihypertensive potential in spontaneously hypertensive rats

Angiotensin I-converting enzyme (ACE) regulates blood pressure through the renin-angiotensin system. Douchi, a traditional fermented soybean condiment, may have antihypertensive effects, but research on ACE inhibitory peptides from Douchi hydrolysates is limited. We hypothesized that enzymatic treatment could enhance ACE inhibitory peptide diversity and efficacy. We tested ten single enzymes and four combinations, finding pepsin-trypsin-chymotrypsin most effective. Hydrolysates were purified using Sephadex G-15 and reversed-phase HPLC, and peptides were identified via LC-MS/MS. Five peptides (LF, VVF, VGAW, GLFG, NGK) were identified, with VGAW as the most potent ACE inhibitor (IC50 46.6 ± 5.2 μM) showing excellent thermal and pH stability. Lineweaver-Burk plots confirmed competitive inhibition, and molecular docking revealed eight hydrogen bonds between VGAW and ACE. In hypertensive rats, VGAW significantly reduced blood pressure at 12.5, 25, and 50 mg/kg. These findings highlight Douchi as a source of ACE inhibitory peptides and suggest VGAW as a promising functional food ingredient.

Effect of drying techniques on the stability of volatile compounds and antioxidative activities of dried Thai fermented black soybean (thua nao)

Thua nao, a traditional fermented soybean product in Thailand, is widely consumed as a plant-based food or a condiment due to its distinct flavor, taste, and nutritional benefits. In this research, thua nao samples were produced from black soybeans. After fermentation, various drying methods, including sun drying (SUN), hot air drying (HOT), microwave vacuum drying (MIC), and vacuum drying (VAC) were employed to produce dried thua nao. The dried samples were stored under ambient conditions for 8 months. Results revealed lipid oxidation and the Maillard reaction occurred during storage, leading to a decline in bioactive compounds and antioxidant activities. Volatile aroma compounds were analyzed using solid-phase microextraction coupled with gas chromatography-mass spectrometry, while principal component analysis (PCA) evaluated aroma profile changes during storage. PCA indicated that HOT, MIC, and VAC -treated samples remained stable for up to 4 months, while SUN-treated samples changed after 2 months. It was found that SUN treatment prominently promoted the formation of oxidized volatile compounds associated with fatty-waxy and green attributes. The HOT treatment contributed more to aroma volatile compounds associated with nutty-roasted, floral, woody-smoky, and sweet attributes. While, both MIC and VAC treatments showed a noticeable loss of aroma volatile compounds, they retained higher level of bioactive compounds compared to the other treatments. This information aids in selecting suitable drying techniques for thua nao products and enhances understanding of chemical transformations induced by drying.

Reduction of aflatoxins and ochratoxin A by addition of commercial Koji during fermentation of the Korean traditional soybean paste, Doenjang

Contamination by aflatoxins and ochratoxin A of traditional Korean fermented soybean paste (Doenjang) is a serious food safety issue. In this study, we investigated the inhibitory effects of commercial Koji, a microbial starter culture used for fermentation, on mycotoxin contamination during Doenjang production at in vitro and scaled-up fermentation levels. A commercial Koji dominated by Aspergillus oryzae JI-4 exhibited strong antifungal and antimycotoxigenic activities against aflatoxigenic Aspergillus flavus and ochratoxigenic Penicillium verrucosum both in vitro and in situ. Furthermore, co-inoculation with Koji JI and mycotoxigenic fungi in the early stage of Doenjang fermentation significantly suppressed aflatoxins and ochratoxin A production by more than 93% (p < 0.05) and improved the nutritional properties and sensory qualities of the final product. These results suggest that the addition of commercial Koji could be an effective and practical approach to reduce aflatoxins and ochratoxin A contamination during the fermentation of soybean paste.

Monitoring and risk assessment of biogenic amines in commercial Thua-Nao, a traditional Northern Thai fermented soybean paste

Thua-Nao, a traditional fermented soybean product in Thailand, is susceptible to contamination with biogenic amines (BAs), thus increasing the risk to human health. The present study evaluated the contamination, formation, and risk assessments of BAs in two types of Thua-Nao. The elution conditions were optimized and used to analyze thua nao BAs with good correlation coefficients and high recovery. The average total BA content in dried Thua-Nao (D-TN; 342 mg/kg) was greater than that in fresh Thua-Nao (F-TN; 239 mg/kg), but the difference was not significant. Tyramine (TYM) was the dominant BA in thua nao, accounting for more than half of the total BAs in the majority of the samples, and it also exhibited a significant positive correlation with soluble protein and total BAs. The calculated highest estimated daily intake (EDI) and index of food safety (IFS) of TYM were 1.51 μg/kg/day and 0.008, respectively, among the age groups who consumed thua nao, indicating a low risk to human health. However, the acute risk index of TYM in F-TN and D-TN among patients taking monoamine oxidase inhibitors (MAOIs) was greater than 80 %. Thus, these results suggested that an adverse health effect is unlikely for the general population after excessive consumption of TYM-contaminated Thua-Nao, but it may have serious health implications for patients taking MAOIs.

Whole genome sequencing of the poly-γ-glutamic acid-producing novel Bacillus subtilis Tamang strain, isolated from spontaneously fermented kinema

Kinema, a traditional fermented soybean food from the Himalayas, is well-liked for its sticky texture and flavourful umami taste. Among 175 bacterial strains from spontaneously fermented kinema samples, Bacillus subtilis Tamang strain stood out for its high stickiness and viscosity. The strain’s Poly-γ-glutamic acid (γ-PGA) contains various groups of glutamic acid and has a molecular weight of 660 kDa. It demonstrates the ability to solubilize iron, preserve ferritin in Caco-2 cells, and exhibit antibacterial properties. The genome of B. subtilis Tamang is devoid of plasmid elements but does feature nine insert elements. Noteworthy is the presence of unique secondary metabolites with potential antimicrobial effects, such as amyloliquecidin GF610, bogorol A, and thermoactinoamide A. A total of 132 carbohydrate-active enzymes (CAZy) were identified, hinting at possible prebiotic characteristics. The genome analysis revealed genes responsible for γ-PGA production via the capBCA complex. Furthermore, genes associated with fibrinolytic activity, taste enhancement, biopeptides, immunomodulators, and vitamins like B12 and K2 were found, along with probiotics and various health benefits. The genetic material for L-asparaginase production, known for its anti-cancer properties, was also detected, as well as CRISPR-Cas systems. The absence of virulence factors and antimicrobial resistance genes confirms the safety of consuming B. subtilis Tamang as a food-grade bacterium.

Characterization and correlations of dominant microorganisms and volatile compounds in fermentation process of Yangjiang douchi.

Yangjiang douchi (YD) is a traditional fermented soybean product, which is popular in Chinese cuisine for its unique flavor. However, due to its high salt content and unstable flavor, its competitiveness in the international market is gradually weakening. Microorganisms have a key role in the production process of YD because it is a fermented food but the effect of microorganisms on the volatile compounds of YD is also not currently clear. In this paper, aroma compounds and microbial diversity in different fermentation stages of YD were analyzed using gas chromatography-mass spectrometry/olfactometry (GC-MS/O) and IlluminaMiseq system sequencing. A total of 78 aroma-active compounds were detected throughout the fermentation process and they influenced the formation of flavor in YD. Fungi flora were relatively single in YD, and bacteria were rich and varied. A total of 418 species of bacteria were present during fermentation, with unclassified_Staphylococcus, Staphylococcus_kloosii, and Bacillus_velezensis_Bacillus predominating. There were 25 species of fungi at the species level, and Aspergillus minisclerotigenes (OTU 4) played a dominant role in the whole fermentation process. Staphylococcus and Bacillus in the bacterial genus were strongly correlated with most flavor compounds detected, and A. minisclerotigenes in the fungi were more relevant to flavor compounds. This research provides a theoretical basis for the enhancement of the flavor of traditional fermented douchi in China. © 2024 Society of Chemical Industry.

Evaluation of Menopausal Syndrome Relief and Anti-Obesity Efficacy of the Korean Fermented Food Doenjang: A Randomized, Double-Blind Clinical Trial.

Foods that help improve menopausal syndrome are being studied worldwide. Doenjang is a traditional Korean fermented soybean food with potential health benefits for menopausal women. In this clinical trial using Doenjang, we aimed to compare the effectiveness of traditional Doenjang and commercial Doenjang in menopausal women. Furthermore, we compared whether Doenjang has a better effect if the number of beneficial microbes is higher. The analyses included the following groups: traditional Doenjang containing either a high dose (HDC; n = 18) or low dose (LDC; n = 18) of beneficial microbes and commercial Doenjang (CD; n = 20). The Kupperman index and hematological changes were examined before and after the use of Doenjang pills. The effects of Doenjang on obesity and body composition were studied before and after ingestion. Lastly, the microorganisms and short-chain fatty acid changes in the stool were compared. The Kupperman index decreased after Doenjang consumption in all three groups, with the greatest decrease in the LDC group. Only the groups that took traditional Doenjang pills exhibited reduced LDL cholesterol. No changes in obesity and inflammation-related indicators were observed. The number of Firmicutes, associated with obesity, decreased in the CD group but the numbers of Bacteroidetes increased in the HDC and CD groups. Thus, traditional Doenjang is more effective in alleviating menopausal syndrome than commercial Doenjang. Further research on the anti-obesity effect or changes in microbiomes and short-chain fatty acids in feces is needed.

Characteristic flavor compounds and bacterial community of different gray sufu, a traditional Chinese fermented soybean curd

ABSTRACT Gray sufu is a traditional Chinese fermented soybean curd, and its flavor is significantly influenced by bacterial community. In this study, volatile flavor compounds (VFCs) were examined by headspace solid‐phase microextraction (HS-SPME) in conjunction with gas chromatography and mass spectrometry (GC‐MS). Additionally, bacterial community was analyzed using high-throughput sequencing. The results showed that, a total of 156 VFCs were tentatively identified, in which 8 esters, 6 S/N-containing compounds, 3 alcohols, 3 aldehydes, 1 ketone and 3 other compounds were recognized as the main characteristic VFCs. The abundant S/N-containing compounds are the most important flavor characteristics of gray sufu. A total of 4 phyla and 12 genera were revealed as dominant bacteria. Firmicutes and Proteobacteria were the most abundant phylum, whereas Bacillus, Tetragenococcus and Lactococcus were the prevailing genera. According to the KEGG pathway annotation, amino acid metabolism (26.98%) was the highest metabolic pathway. Among them, aromatic amino acids metabolism accounted for the highest proportion, up to 34.94%. Correlation analysis showed that Lactobacillus, Tetragenococcus, Bacillus, Pseudomonas and Halanaerobium were the top five positive flavor-producing bacteria. The results will be helpful for understanding the fermentation mechanism of gray sufu and improving flavor quality by developing microbial regulation strategies.

Unraveling umami complexity: From exploring umami peptides in fermented soybean curd to molecular elucidation of taste mechanisms

Sufu is a traditional umami fermented soybean curd (FSC). We isolated and identified 25 potential umami peptides using ethanol precipitation, dialysis desalination, resin adsorption recovery, nano-HPLC-MS/MS and virtual docking prediction. The results of sensory experiments showed that 23 of the synthetic peptides presented a lower umami threshold than that of MSG (0.50 mg/mL), and 10 peptides had obvious effects of increasing umami and saltiness. Seven peptides were quantified in FSC by UPLC-QQQ-MS, and the content of one novel umami tripeptide ADL (0.51 ± 0.03 mg/mL) was four times its umami threshold (0.125 mg/mL). Taste reconstitution experiments revealed that the umami peptides enhanced the umami and saltiness of the simulated system by 24.53% and 24.00%, respectively. Molecular dynamics proved that hydrogen bonding is the main interaction between peptide and receptors. This study is important for understanding and improving the taste quality of FSC.

Correlation between monosaccharide, oligosaccharide, and microbial community profile changes in traditional soybean brick (meju) fermentation

Meju is essential for making diverse traditional fermented soybean foods in Korea. To understand the changes in carbohydrates during fermentation, we aimed to identify autochthonous microorganisms from spontaneously fermented meju and compare the alterations in monosaccharides and oligosaccharides throughout the fermentation process. Microbial diversity was determined using a metabarcoding approach, and monosaccharide and oligosaccharide profiles were obtained by HPLC-Q-TOF MS and HPLC-MS/MS analyses, respectively. The dominant bacterial genera were Weissella, Lactobacillus, and Leuconostoc, while Mucor was highly abundant in the fungal community. The total monosaccharide content increased from Day 0 to Day 50, with the highest amount being 4.37 mg/g. Oligosaccharide profiling revealed the degradation of soybean dietary fiber during fermentation, and novel oligosaccharide structures were also discovered. Correlation analysis revealed that the fungus Mucor was positively related to pentose-containing oligosaccharides, galactose, and galacturonic acid, indicating that Mucor may degrade soybean dietary fibers such as xylogalacturonan, arabinogalactan, and rhamnogalacturonan. The negative relationships between the abundances of Weissella and oligo- and monosaccharides suggested that the bacteria may utilize saccharides for fermentation. These findings provide insights into the mechanisms underlying carbohydrate degradation and utilization; the key components involved in saccharide transformation that contribute to the characteristics of traditional meju were subsequently identified.

Nutraceutical potential of traditional fermented soybean (Hawaijar)

Fermentation not only reduces the toxins but also it is an excellent processing method for improving nutritional and functional properties of soybean due to the increased content of small bioactive compounds. The NEH region of India also has similar traditional fermented soy products. ACE inhibitory activity was found to increase during fermentation and exhibited higher ACE inhibition percent in starter culture strain (H) than starter culture strain (S) of both the soybean used (local small variety and JS-335). Fermentation of soybean with different micro-organisms improve the bio-functional properties due to the increase in free isoflavones and peptides.

Identification, Characterization, and Receptor Binding Mechanism of New Umami Peptides from Traditional Fermented Soybean Paste (Dajiang).

Dajiang, a traditional Chinese condiment, is made from fermented soybeans. It is highly popular among consumers as a result of its delicious umami flavor, which mainly originates from umami peptides. To examine the mechanism of umami taste in Dajiang, we selected Dajiang samples with strong umami taste and subjected them to purification and identification analysis using ethanol precipitation, gel chromatography, reversed-phase high-performance liquid chromatography, and ultraperformance liquid chromatography-tandem mass spectrometry. Subsequently, on the basis of toxicity and umami prediction analysis, we screened, synthesized, and characterized three novel bean umami peptides in Dajiang: TLGGPTTL, 758.4174 Da; GALEQILQ, 870.4811 Da; and HSISDLQ, 911.4713 Da. Their sensory threshold values were 0.25, 0.40, and 0.17 mmol/L, respectively. Furthermore, molecular docking results showed that hydrogen-bonding and hydrophobic interactions are important interaction forces in the binding of umami peptide to taste receptors. Ser147 and Glu148 of the T1R3 taste receptor are important amino acid residues for binding of the three umami peptides. This study uncovers the mechanism of umami-peptide-driven flavor in fermented soybean products.

Complete genome sequence of Arthrobacter sp. KFRI-F3372, a strain isolated from Korean soybean paste Doenjang.

Arthrobacter sp. KFRI-F3372 is a Gram-positive bacterium with a high G + C content of 65.7%, which was isolated from Doenjang, a traditional Korean fermented soybean paste. In this report, we introduce the complete genome sequence of Arthrobacter sp. KFRI-F3372.

Safety Analysis of Korean Cottage Industries' Doenjang, a Traditional Fermented Soybean Product: A Special Reference to Biogenic Amines.

The typical Korean diet contains a significant quantity of doenjang owing to its unique taste and health benefits. However, the presence of anti-nutritional and toxic substances, such as biogenic amines and microbial pathogens, in doenjang has resulted in a loss of revenue and poor consumer health. The present study focused on the identification and quantification of different biogenic amines, pathogenic Bacillus cereus, and yeast counts in 36 doenjang products (designated as De-1 to De-36, 500 g each) procured from the different cottage industries situated in different parts of the Republic of Korea. The results indicated, only three samples were contaminated with B. cereus, exceeding the recommended limit (4 log CFU/g) suggested by the national standards of Korea. A total of six distinct yeasts were identified in different doenjang samples, whose comprehensive enzymatic profiling suggested the absence of harmful enzymes such as N-acetyl-β-glucosaminidase, α-chymotrypsin, and β-glucuronidase. The biogenic amines were detected in the range of 67.68 mg/kg to 2556.68 mg/kg and classified into six major groups based on hierarchical cluster analysis. All doenjang samples contained tryptamine, putrescine, cadaverine, histamine, and tyramine, while 94.44% were positive for spermidine and spermine. The results documented the analysis of traditional cottage industry doenjang and suggest the need for constant monitoring to ensure the safety of food for the consumer.

Microbial community succession and its correlation with the dynamics of flavor compound profiles in naturally fermented stinky sufu.

Wuhan stinky sufu is a traditional fermented soybean product with a short ripening period and unique flavor. The aim of this study was to explore the characteristic flavor compounds and core functional microbiota of naturally fermented Wuhan stinky sufu. The results indicated that 11 volatile compounds including guaiacol, 2-pentylfuran, dimethyl trisulfide, dimethyl disulfide, acetoin, 1-octen-3-ol, (2E)-2-nonenal, indole, propyl 2-methylbutyrate, ethyl 4-methylvalerate, nonanal were characteristic aroma compounds, and 6 free amino acids (Ser, Lys, Arg, Glu, Met and Pro) were identified as taste-contributing compounds. 4 fungal genera (Kodamaea, unclassified_Dipodascaceae, Geotrichum, Trichosporon), and 9 bacterial genera (Lysinibacillus, Enterococcus, Acidipropionibacterium, Bifidobacterium, Corynebacterium, Lactococcus, Pseudomonas, Enterobacter, and Acinetobacter) were identified as the core functional microbiota with positive effects on the production of flavor compounds. These findings would enhance the understanding of core flavor-producing microorganisms in naturally fermented soybean products and potentially provide guidance for enhancing the quality of sufu.