Production of beta-glucosidase and hydrolysis of isoflavone phytoestrogens by Lactobacillus acidophilus, Bifidobacterium lactis, and Lactobacillus casei in soymilk

ABSTRACT:  The study determined β-glucosidase activity of commercial probiotic organisms for hydrolysis of isoflavone to aglycones in fermenting soymilk. Soymilk made with soy protein isolate (SPI) was fermented with Lactobacillus acidophilus LAFTI^® L10, Bifidobacterium lactis LAFTI^® B94, and Lactobacillus casei LAFTI^® L26 at 37 °C for 48 h and the fermented soymilk was stored for 28 d at 4 °C. β-Glucosidase activity of organisms was determined using ρ-nitrophenyl β-D-glucopyranoside as a substrate and the hydrolysis of isoflavone glycosides to aglycones by these organisms was carried out. The highest level of growth occurred at 12 h for L. casei L26, 24 h for B. lactis B94, and 36 h for L. acidophilus L10 during fermentation in soymilk. Survival after storage at 4 °C for 28 d was 20%, 15%, and 11% greater ( P < 0.05) than initial cell counts, respectively. All the bacteria produced β-glucosidase, which hydrolyzed isoflavone β-glycosides to isoflavone aglycones. The decrease in the concentration of β-glycosides and the increase in the concentration of aglycones were significant ( P < 0.05) in the fermented soymilk. Increased isoflavone aglycone content in fermented soymilk is likely to improve the biological functionality of soymilk.     

Transformation of isoflavone phytoestrogens during the fermentation of soymilk with lactic acid bacteria and bifidobacteria

In the present study, soymilk is fermented with lactic acid bacteria (Streptococcus thermophilus BCRC 14085, Lactobacillus acidophilus BCRC 14079) and bifidobacteria (Bifidobacterium infantis BCRC 14633, B. longum B6) individually, and in combination. The change in the content of various isoflavones (aglycones, glucoside, acetyl- and malonyl-glucosides) and the beta-glucosidase activity in soymilk during fermentation is investigated. It is observed that fermented soymilk contains a lower total isoflavone content (81.94-86.61 microg/ml) than soymilk without fermentation (87.61 microg/ml). Regardless of starter organism employed, fermentation causes a major reduction in the contents of glucoside, malonylglucoside and acetylglucoside isoflavones along with a significant increase of aglycone isoflavones content. The level of change in the content of various isoflavones and beta-glucosidase activity after fermentation varies with the starter organism. Among all the fermented soymilks tested, soymilk fermented with S. thermophilus showed the highest beta-glucosidase activity and the greatest increase in the contents of aglycones. The percentage of daidzein, genistein and glycitein to total isoflavone content in S. thermophilus-fermented soymilk increases from an initial 14.24%, 6.89% and 2.45%, respectively, to 36.20%, 28.80% and 12.44% after 24h of fermentation. Finally, the increase of aglycones and decrease of glucoside isoflavones during fermentation coincides with the increase of beta-glucosidase activity observed in fermented soymilk.     

Stability of β-glucosidase Activity Produced by Bifidobacterium and Lactobacillus spp. in Fermented Soymilk During Processing and Storage

ABSTRACT: Microorganisms possess endogenous enzymes, however the stability of these enzymes during storage in soymilk has not been studied. β-glucosidase is an important enzyme that could be used in the bioconversion of the predominant soy isoflavone glucosides to their bioactive aglycone forms. Fifteen probiotic microorganisms including bifidobacterium, Lactobacillus acidophilus , and Lactobacillus casei were screened for β-glucosidase activity using p-nitrophenyl-β-d-glucopyranoside as a substrate. Six strains were selected on the basis of β-glucosidase activity produced during fermentation of soymilk. The stability of the enzyme activity was assessed during incubation for up to 48 h and storage for 8 wk at frozen (-80°C), refrigerated (4°C), room (24.8°C), and incubation (37°C) temperatures. L. casei strains showed the highest β-glucosidase activity after 24 h of incubation followed by L. acidophilus strains, whereas bifidobacterium strains showedleast activity. However, p-glucosidase from Bifidobacterium animalis BB12 showed the best stability during the 48 h fermentation. Lower storage temperatures (-80°C and 4°C) showed significantly higher ( P < 0.05) β-glucosidase activity and better stability than that at higher temperatures (24.8°C and 37°C). The stability of β-glucosidase from these microorganisms should be considered for enzymic biotransformation during storage of isoflavone β-glucosides to bioactive isoflavone aglycone forms with potential health benefits.     

Enrichment of bioactive isoflavones in soymilk fermented with β-glucosidase-producing lactic acid bacteria

This study was undertaken to investigate the possible application of β-glucosidase-producing lactic acid bacteria as a functional starter cultures to obtain the bioactive isoflavones, genistein and daidzein, in fermented soymilk. Four strains – Lactobacillus plantarum KFRI 00144, Lactobacillus delbrueckii subsp. lactis KFRI 01181, Bifidobacterium breve K-101 and Bifidobacterium thermophilum KFRI 00748 – among the 31 lactic acid bacteria tested for β-glucosidase activity using ρ-nitrophenyl-β-d-glucopyranoside as the substrate were selected. Acid development, viable populations, and quantification of isoflavones using HPLC were performed at 0, 24, and 48 h of incubation at 37 °C. The significant bioconversion (P < 0.001) of the glucoside isoflavones into their bioactive aglycones in soymilk fermented with four β-glucosidase-producing strains, with an average 7.1-fold increase of aglycones (daidzein + genistein) was observed. There appeared to be correlations between the level of growth and β-glucosidase activity of each strain, and the hydrolysis of conjugated isoflavones in soymilk fermentation. Lactobacillus sp. were able to readily proliferate in soymilk than Bifidobacterium sp. (P < 0.05) and therefore completed more rapidly the hydrolysis of glucoside isoflavones.The present study indicates that four β-glucosidase-producing lactic acid bacteria have great potential for the enrichment of bioactive isoflavones in soymilk fermentation.     

Enzymic Transformation of Isoflavone Phytoestrogens in Soymilk by β-Glucosidase-Producing Bifidobacteria

ABSTRACT: Five strains of bifidobacteria were screened for β-glucosidase activity using p-nitrophenyl-β-D-glucopyranoside as the substrate, and selected strains were used to ferment soymilk. Enumeration of viable bifidobacteria and quantification of isoflavones using HPLC were performed at 0, 12, 24, 36, and 48 h of incubation. Four strains produced β-glucosidase. B. pseudolongum and B. longum -a displayed the best growth in soymilk, with an increase of 1.3 log₁₀ CFU/mL after 12 h. B. animalis, B. longum -a, and B. pseudolongum caused hydrolysis of isoflavone malonyl-, acetyl- and β-glucosides to form aglycones, and transformed daidzein to equol in soymilk. Fermentation of soymilk with Bifidobacterium sp. resulted in a significant increase (p < 0.05) in the concentration of aglycones.     

Using of Lactobacillus and Bifidobacterium to product the isoflavone aglycones in fermented soymilk

The study determined isoflavone aglycone contents in soymilk and sugar-soymilk fermented with a single culture of two strains of Lactobacillus paracasei, two strains of Lactobacillus acidophilus, and one strain of Bifidobacterium longum respectively at 37 degrees C for 24 h. Isoflavone concentration was analyzed by HPLC. The viable count of bacteria in all of the fermented soymilk ranged from 7 to 9 log CFU/ml. The isoflavone aglycone had a significant increase of 62%-96% of isoflavone in all of the fermented soymilk compared to 17% in non-fermented soymilk (P<0.05). The five strains of microorganisms produced lesser amount of isoflavone aglycones in fermented sucrose-soymilk than in other fermented soymilk.     

Fermentation of commercial soy beverages with lactobacilli and bifidobacteria strains featuring high β-glucosidase activity

An increase of isoflavone aglycone content in soy foods can be attained through fermentation with food-grade bacteria. In this study, two commercial soy beverages with distinctive chemical composition (AS and VS) were fermented by strains of lactobacilli (eight strains) and bifidobacteria (two strains) with a high β-glucosidase activity. Along fermentation, growth of the strains and isoflavone deglycosylation in the soy beverages were monitored. Large differences in growth, aglycone content and chemical parameters in AS and VS beverages fermented by different species and strains were observed. Isoflavone glycosides were completely transformed into their corresponding aglycones by most strains during fermentation of AS beverage, whereas large amounts of undeglycosilated isoflavones were still present in fermented VS. Four strains showing strong deglycosylation activity and appropriate technological properties (Lactobacillus casei LP71, Lactobacillus plantarum E112, Lactobacillus rhamnosus E41 and Bifidobacterium pseudocatenulatum C35) were proposed as industrial starters to improve functionality in soy-based fermented foods.     

Fermentation of calcium-fortified soymilk with Lactobacillus: effects on calcium solubility, isoflavone conversion, and production of organic acids

ABSTRACT:  The objective of this study was to enhance calcium solubility and bioavailability from calcium-fortified soymilk by fermentation with 7 strains of Lactobacillus , namely, L. acidophilus ATCC 4962, ATCC33200, ATCC 4356 , ATCC 4461 , L. casei ASCC 290, L. plantarum ASCC 276, and L. fermentum VRI-003. The parameters that were used are viability, pH, calcium solubility, organic acid, and biologically active isoflavone aglycone content. Calcium-fortified soymilk made from soy protein isolate was inoculated with these probiotic strains, incubated for 24 h at 37 °C, then stored for 14 d at 4 °C. Soluble calcium was measured using atomic absorption spectrophotometry (AA). Organic acids and bioactive isoflavone aglycones, including diadzein, genistein, and glycetein, were measured using HPLC. Viability of the strains in the fermented calcium-fortified soymilk was > 8.5 log₁₀ CFU/g after 24 h fermentation and this was maintained for 14-d storage at 4 °C. After 24 h, there was a significant increase ( P < 0.05) in soluble calcium. L. acidophilus ATCC 4962 and L. casei ASCC 290 demonstrated the highest increase with 89.3% and 87.0% soluble calcium after 24 h, respectively. The increase in calcium solubility observed was related to lowered pH associated with production of lactic and acetic acids. Fermentation significantly increased ( P < 0.05) the level of conversion of isoflavones into biologically active aglycones, including diadzein, genistein, and glycetein. Our results show that fermenting calcium-fortified soymilk with the selected probiotics can potentially enhance the calcium bioavailability of calcium-fortified soymilk due to increased calcium solubility and bioactive isoflavone aglycone enrichment.     

Stability of isoflavone phytoestrogens in fermented soymilk with Bifidobacterium animalis Bb12 during storage at different temperatures

Soy isoflavones in fermented soymilk with Bifidobacterium animalis Bb12 were stored at various temperatures (?80, 4, 24.8 and 37 °C) for 8 weeks and the concentration of isoflavones determined weekly using reverse‐phase high‐performance liquid chromatography (RP‐HPLC). The first‐order kinetic model was used to assess the degradation of each isomer at each storage temperature. Soymilk predominantly possesses high concentrations of isoflavone glucosides and very low concentration of bioactive aglycone component. During storage at various temperatures, concentrations of individual isoflavone isomers appeared to be significantly stable (P < 0.01). Interestingly, aglycones showed much smaller degradation constants compared with glucosides at all the storage temperatures studied. Genistein and daidzein were much more stable than glycitein and had almost similar degradation patterns, despite differences in their concentrations in the fermented soymilk. It was, however, observed that 4 °C was the most suitable storage temperature for the product in order to guarantee minimal degradation of bioactive isoflavone aglycones.     

Evaluation of enzymic potential for biotransformation of isoflavone phytoestrogen in soymilk by Bifidobacterium animalis,Lactobacillus acidophilus and Lactobacillus casei

Three strains of Lactobacillus acidophilus, two of Lactobacillus casei and one of Bifidobacterium were screened for β-glucosidase activity using ρ-nitrophenyl-β-d-glucopyranoside as a substrate and their potential for the breakdown of isoflavone glucosides to the biologically active aglycones in soymilk. Isoflavones quantification with HPLC and β-glucosidase activity were performed after 0, 12, 24, 36, and 48 h of incubation in soymilk at 37 °C. All six micro-organisms produced β-glucosidase, which hydrolysed the predominant isoflavone β-glucosides. There was a significant increase and decrease (P < 0.05) in the concentration of isoflavone aglycones and glucosides, respectively, in fermented soymilk. Based on the concentration of isoflavones during peak β-glucosidase activity, the hydrolytic potential was calculated. L. acidophilus 4461 had the highest aglycone concentration of 76.9% after 24 h of incubation, up from 8% in unfermented soymilk (at 0 h). It also had the best isoflavone hydrolytic index of 2.01, signifying its importance in altering the biological activity of soymilk.     

植物乳杆菌58发酵豆乳产大豆异黄酮苷元条件的优化

本研究通过测定植物乳杆菌58在不同碳源中的生长和产β-葡萄糖苷酶情况,筛选菌株的最适碳源,并确定接种至豆乳的最佳时间。在接种量、糖含量、发酵时间和发酵温度等单因素实验基础上,根据Box-Behnken中心组合原理进行响应面实验设计,以大豆异黄酮苷元含量为指标,进一步优化菌株58发酵豆乳产大豆异黄酮苷元条件。结果显示,植物乳杆菌58生长和产β-葡萄糖苷酶的最适碳源为乳糖,酶活达0.66 U/m L,显著高于果糖、蔗糖、葡萄糖和麦芽糖(p<0.05)。菌株在乳糖碳源中培养18 h产酶活力最佳,达0.75 U/mL,且生长情况良好。响应面优化实验得出发酵豆乳大豆异黄酮最佳转化条件为:接种量3.80%,糖含量5.80%,发酵温度38.10℃,发酵时间9.80 h。此条件下,大豆异黄酮苷元含量预测值达68.63 mg/L,与实验值68.16 mg/L相比差异不显著,表明构建二次模型的科学性和准确性,与优化前(59.64 mg/L)相比提高15.07%,有助于乳酸菌发酵豆乳中大豆异黄酮糖苷向高生物活性和利用度大豆异黄酮苷元的转化。     

Enhancement of Biological Properties of Soymilk by Fermentation

Soymilk has attracted much interest in the food industry because of its health- promoting properties. Fermentation with lactic acid bacteria is known to provide value addition to soymilk by reducing the beany flavor and content of indigestible oligosaccharides and by enhancing the bioavailability of isoflavones, resulting in a nutritious probiotic food product. In the present study, Soymilk fermentation was studied using Lactobacillus acidophilus strain isolated from ragi, and effect of soymilk supplementation on the fermentation characteristics was also investigated. Viable count of 1.99 × 10⁹ CFU/mL sample and 630.1 U/g of β-glucosidase activity were determined in soymilk after 12 h of fermentation. Soymilk supplementation with skimmed milk powder at 5% level gave best results with viable counts of 22.9 × 10⁹ CFU/ml sample or 212.9 × 10⁹ CFU/g solids; 764 U/g of β-glucosidase activity, 1.44% la titratable acidity and pH of 3.9. Daidzein and genistein profile in fermented soymilk showed that 45.8% and 57.5% of respective isoflavones existing as aglycone form, indicating enhancement of the biological property of soymilk through improvement of health-relevant bioactive forms.     

Bioactive action of β‐glucosidase enzyme of Bifidobacterium longum upon isoflavone glucosides present in soymilk

Bioconversion of isoflavone glucosides and antioxidant activity by probiotic strain (Bifidobacterium longum) during soymilk fermentation was investigated, as well as partial characterisation of the produced enzyme β‐glucosidase. The enzyme has higher affinity for genistin than for other substrates assayed. Maximum activity occurred at 42 °C and at pH 6.0; keeping 70–80% of activity for 60 days stored at low temperatures. Bifidobacterium longum grew well in soymilk (8.26 log CFU mL^?1 and pH of 3.9 at 24 h) and were produced in good quantities of organic acids. High hydrolysis degree of isoflavone glucosides (81.2%) was observed at 24 h. Enhancements in bioactivity were assessed in fermented soymilk by monitoring the radical‐scavenging activity, antioxidant activity and DNA protective action. The use of probiotic Bifidobacterium strain as β‐glucosidase producer increased bioactive isoflavone content and demonstrated that this enzyme plays a key role in the bioavailability of soymilk isoflavones, reducing the bioconversion time compared to other studies.     

Conversion of isoflavone glucosides to aglycones in soymilk by fermentation with lactic acid bacteria

ABSTRACT:  Four lactic acid bacteria (LAB), Lactobacillus paraplantarum KM, Enterococcus durans KH, Streptococcus salivarius HM and Weissella confusa JY, were isolated from humans and tested for their capabilities of converting isoflavone glucosides to aglycones in soymilk. Changes in growth, pH, and titratable acidity (TA) were investigated during fermentation at 37 °C for 12 h. After 6 to 9 h of fermentation, each population of 4 LAB reached 10⁸ to 10⁹ CFU/mL. The initial pH of 6.3 ± 0.1 decreased while the TA of 0.13%± 0.01% increased as fermentation proceeded, resulting in the final range between 4.1 ± 0.2 and 4.6 ± 0.1 for pH and between 0.51%± 0.02% and 0.67%± 0.06% for TA after the 12 h of fermentation. The glucoside concentrations were significantly decreased in soymilks fermented with either L. paraplantarum KM, S. salivarius HM, or W. confusa JY with fermentation time ( P < 0.05). L. paraplantarum KM was the best in percent conversion of glucosides to corresponding aglycones, resulting in 100%, 90%, and 61% hydrolysis of genistin, daidzin, and glycitin, respectively, in 6 h. Consequently, the aglycone concentrations in soymilk fermented with L. paraplantarum KM were 6 and 7-fold higher than the initial levels of daidzein and genistein, respectively, after 6 h of fermentation. Changes in the daidzin and genistin levels were not significant in soymilk fermented with E. durans KH. The rates of hydrolysis of glucosides varied depending on the species of LAB. Especially, L. paraplantarum KM seems to be a promising starter for bioactive-fermented soymilk based on its growth, acid production, and isoflavone conversion within a short time.     

Enhancing the Biotransformation of Isoflavones in Soymilk Supplemented with Lactose Using Probiotic Bacteria during Extended Fermentation

ABSTRACT: Soymilk (SM) lacks lactose; hence supplementation of SM with lactose is likely to enhance the growth of probiotic bacteria and biotransformation of isoflavone glycosides to isoflavone aglycones. In this study, 11 strains of probiotic bacteria including Lactobacillus rhamnosus, L. salivarius, L. plantarum, L. acidophilus, L. paracasei, HOWARU L. rhamnosus, L. delbrueckii subsp. bulgaricus, Bifidobacterium lactis type Bi-07, B. longum, HOWARU B. bifidum, and B lactis type Bi-04 were inoculated individually or as mixed cultures into SM and soymilk supplemented with lactose (SML). A total of 2% of lactose was added to 1 L of SM with the aim of improving the growth of probiotic organisms and promoting the biotransformation of isoflavone isomers to bioactive isoflavone aglycomes. Samples of SM were incubated at 37 °C and 10 mL aliquots of SM were taken at 0, 24, 48, and 72 h to monitor the growth of probiotic bacteria and changes in isoflavone contents using high-performance liquid chromatography (HPLC). Results indicated that SML fermented with probiotics had higher viable counts by >2.4 log CFU/mL than that in SM at the end of the 72 h fermentation period. Mixed cultures grew at different rates and in general Lactobacilius spp. had >1.02 log CFU/mL more cells than Bifidobacterium spp. at the end of the fermentation period. The total aglycone content in SM at 72 h of fermentation was 0.924 mg/100 mL, whereas that in SML was 1.623 mg/100 mL. Addition of lactose not only improved the growth of probiotic bacteria in SM but also enhanced the biotransformation of isoflavone glucosides to the more bioactive isoflavone aglycones. Mixed cultures did not improve the biotransformation of bioactive isoflavones when compared to single cultures.     

Effect of Lactic Acid Fermentation on Enrichment of Antioxidant Properties and Bioactive Isoflavones in Soybean

ABSTRACT: The antioxidant properties of 80% ethanolic extracts from soybean obtained with lactic acid fermentation using ABTS•⁺ [2,2'-azinobis (3-ethylbenzothiazoline-6-sulphonicacid) diammonium salt] free radical decolorization assay and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging assay were measured, and the relationship between the observed antioxidant properties and the compositional changes in the isoflavone isomers was evaluated. Fermentation of soybean with 4 bacterial strains producing β-glucosidase for 48 h at 37°C— Lactobacillus plantarum KFRI 00144, Lactobacillus delbrueckii subsp. latisKFRI01181, Bifidobacteriathermophilum KFRI00748, and Bifidobacteria breve K-101—resulted in a significant increas (P < 0.05) in the antioxidant capacity expressed as Trolox equivalent antioxidant capacity (TEAC) (mM) and % scavenging activity. The significant bioconversion (P < 0.001) of the isoflavone glucosides (daidzin + genistin) into their responding bioactive aglycones (daidzein + genistein) during soy fermentation was observed. There was a good linear correlation between the concentration of isoflavone aglycones and the scavenging activity of ABTS ( R = 0.9045) assay, and DPPH ( R = 0.8299) assay in each extract. Among each extract, B. thermophilum KFRI 00748 showed a particularly high antioxidant activity (19.8 mM TEAC) and increased by 4.1 times compared with that of the control (4.8 mM TEAC), which fermented without strains. These results indicated that fermented soybean could be regarded as a potent antioxidant and radical scavenging dietary source due to their remarkable content of bioactive isoflavones.     

Profiling and quantification of isoflavones in soymilk from soy protein isolate using extracted ion chromatography and positive ion fragmentation techniques

Dietary supplements on soy based foods and beverages are increasingly gaining prominence all over the world. In this study, liquid chromatography coupled with positive electrospray ionisation tandem mass spectrometry (LC-ESI-MS) and diode array detection was used for the quantitation and characterisation of isoflavones in fermented and unfermented soymilk made from soy protein isolate SUPRO 590. Bifidobacterium animalis ssp. lactis Bb12 was used for the fermentation of soymilk. The isoflavones were found to produce characteristic radical ions as well as molecules of H₂O, CO₂, a sugar unit, and an alcohol through collision-induced fragmentation. Product ion fragments revealed unique fragmentation pathways for each isoflavone compound. Characteristic fragmentation of different isoflavones were unequivocally identified and differentiated. The occurrence of aldehydes such as pentanal, ethanal and methanal was shown to be specifically linked with isoflavone aglycones, daidzein, genistein and glycitein, respectively. Main glycosides such as genistin, daidzin and glycitin as well as the acetyl-, and malonyl forms also showed respective aglycone ions in their spectra fragmentation. Thus positive ion fragmentation was important in the absolute confirmation of isoflavones and to reveal the occurrence of other related compounds such as aldehydes in soymilk.     

Isoflavone phytoestrogen degradation in fermented soymilk with selected beta-glucosidase producing L. acidophilus strains during storage at different temperatures

Soymilk fermented with 3 selected Lactobacillus acidophilus strains were stored at various temperatures (-80 degrees C, 4 degrees C, 25 degrees C and 37 degrees C) for 8 weeks and the concentration of isoflavones determined weekly using RP-HPLC. The decreasing concentration of isoflavones in soymilk during storage due to degradation was found to fit the first order kinetics model. Isoflavone aglycones as well as isoflavone glucosides largely appeared to be stable during storage (P<0.01). Interestingly, the aglycone forms showed much smaller degradation as compared to glucoside forms at all the storage temperatures studied. Of the isoflavone aglycones, daidzein was found to be the most stable followed by genistein, while glycitein was least stable. Isoflavone aglycones such as glycitein, daidzein and genistein showed smaller degradation constants in fermented soymilk at lower storage temperatures (-80 degrees C and 4 degrees C) and higher degradation constants at higher storage temperatures (25 degrees C and 37 degrees C) with each strain. In contrast, glucosides glycitin and daidzin showed higher degradation at lower storage temperatures (-80 degrees C and 4 degrees C) and lower degradation at higher storage temperatures (25 degrees C and 37 degrees C). Storage temperature was therefore found to be very important in regulating the rate of degradation soy isoflavones in fermented soymilk.     

Biotransformation of Isoflavones by Bifidobacteria in Fermented Soymilk Supplemented with D-Glucose and L-Cysteine

ABSTRACT: Soymilk prepared using soy-protein isolate supplemented with D-glucose and L-cysteine was fermented with 4 strains of Bifidobacterium. Enumeration of bifidobacteria and quantification of isoflavones using HPLC were performed at 0, 12, 24, 36, and 48 h of incubation. Supplementation did not significantly enhance ( p > 0.05) the growth of bifidobacteria between 0 and 12 h, but did after 12 h. The increase in concentration of isoflavone aglycones and equol was significantly lower ( p < 0.05) in supplemented soymilk after 24 h when compared to plain soymilk. Supplementation increased the concentration of aglycones by 0.796 mg/100 mL in soymilk fermented with B. animalis between 12 and 24 h, and the population by 1.27 log₁₀ CFU/mL ( p < 0.05).     

Enrichment of isoflavone aglycones in soymilk by fermentation with single and mixed cultures of Streptococcus infantarius 12 and Weissella sp. 4

Soymilk was fermented with either Streptococcus infantarius 12 (Si 12), Weissella sp. 4 (Ws 4), or their mixed cultures with different mixing ratios (Si 12:Ws 4 = 1:1, 1:3, 1:5, and 1:10, v/v) for 12 h at 37 °C. All cultures in soymilk readily proliferated and reached about 10^8–9 CFU/mL. After 12 h, pH and titratable acidity of soymilk ranged 4.19–4.47 and 0.57%–0.64%, respectively. The pH of soymilk fermented with Si 12 was the lowest while that obtained with Ws 4 the highest. A sharp increase in β-glucosidase (β-glu) activity corresponded well with a rapid decrease in isoflavone glucosides and an increase in aglycone contents. The rate of hydrolysis of isoflavone glucosides was the least with Si 12 while the highest with Ws 4, resulting in about 23%–33% and 98%–99% hydrolysis of the glucosides with Si 12 and Ws 4, respectively, after 12 h. Mixed cultures with 1:3, 1:5, and 1:10 ratios seem to be more effective starters for bioactive fermented soymilk with more aglycones and appropriate acidity in a short time than single cultures.