| Soy whey is the main by-product of the traditional soybean production process.At present,most of soybean production enterprises in our country directly discharge a large amount of soy whey as waste-water during the production process.Not only the chemical oxygen demand(COD)and biochemical oxygen demand in five days(BOD5)of soy whey are very high,but also it is rich in variety of nutrients and functional substances,which can not only be a waste of resources,but also be a great pollution to the environment.The traditional tea fungus,namely kombucha(KBC),is a kind of acidic beverage,which is sweeten black tea fermented by kombucha consortium including yeasts and acetic acid bacteria.It is considered to be a health drink with a variety of health benefits.In recent years,the application and research of kombucha in European and American countries and Japan are so popular.Especially its abilities of anticancer,detoxication and increasing immunity have attracted much attention.In this study,we choose soy whey as new fermentation substrates to kombucha consortium,called kombucha-fermented soy whey(KFSW).This fermentation can improve the nutrition and the functional properties and influence the microorganism community.Finally,we find a new way for the utilization of soy whey resources.The main research contents and results as follow:1.Effects on the chemical characteristics and determination of KFSWTerminal point of fermentation with kombucha was determined by measuring microbiological analysis,pH values,titration acidity,reducing sugar,organic acids and sensory characteristics during fermentation.The results showed that the kombucha consortium can grow well in soy whey which providessufficient nutrients.The cell counts of yeast and acetic acid bacteria showed a trend of growth with the prolonging of fermentation time until the sixth day,and then it remain stable or slightly down.The pH value decreased from 4.89 to 3.24.The changes of reducing sugar and organic acids content during fermentation were analyzed.The results showed that the content of reducing sugar was slightly increased at beginning of fermentation,and then it decreased from 9.69 mg/mL to 1.37 mg/mL.There were four kinds of organic acids detected by high performance liquid chromatography(HPLC),including formic acid,acetic acid,citric acid,furmaric acid.The results showed that acetic acid was the main organic acid produced during fermentation.Its content reached 5.786 g/L in the sixth day.The content of formic acid increased to 2.496 g/L and then decreased to 0.778 g/L.Compared with traditional kombucha fermented sweeten black tea,it can detect citric acid in the fermented soy whey,whose content is about 1.1-1.3 g/L.There is no distinct change in fumaric acid content.The sensory evaluation showed that the fermentation to the sixth day,the appearance,taste and other sensory indicators of fermented soy whey have reached the best state.Considering with previous results above,we choose the sixth day as the terminal point of fermentation.Using HS-SPME/GC-MS to determinate volatile substances in soy whey during fermentation.The results showed that,almost of the indigenous aroma compounds in soy whey were metabolized to low or undetectable levels,meanwhile the new aroma,esters and aldehydes especially nonanal and undecanal were more formed after fermentation.Hence the beany off-flavor of soy whey was replaced by the new fruity and floral flavor,which was given a new pleasant smell.2.Analyze the microbial diversity by high throughput sequencingExtract total microbial DN A of broth and layer samples from KBC and KFSW,employ the V1F/V3R and ITS1F/ITS2R as PCR amplification primers,amplify the V1-V3 region of bacterial 16S rRNA and ITS1-ITS2 region of fungus 18S rRNA,recover、library construction、sequencing and statistical analysis.The Alpha diversity of bacterial is higher than fungus,and their microbial composition has similarity and variability.Bacterial in samples belong to Firmicutes、Proteobacteria in phylum level and to acetobacter,komagataeibacter and gluconobacter in genus level,the komagataeibacter existe in all samples;Fungi in samples can be classified in Ascomycota and Zygomycota in phylum level and dekkera、pichia and zygosaccharomyces in genus level,the dekkera existe in all samples.3.Changes of functional susbtances and bioactivity of KFSWUsing 80%methanol and ultrasound to extract of unfermented soy whey and soy whey fermented by kombucha,to measurethe total flavonoids content(TFC),the result show that the TFC of 80%ethanolic extracts from fermented soy whey was increased with prolonging the fermentation times until sixth day,TFC increased from 183.19 mg RE/L to 264.44 mg RE/L The changes of specific flavonoids compositionof 80%ethanolic extracts from unfermented soy whey and fermented soy whey were quantitatively determined by HPLC.The results showed that the contents of aglycones were significantly increased in the fermented soy whey,and the content of glycitein and genistein were increased from 24.11 mg/L to 77.61 mg/Land from 24.63 mg/L and 73.34 mg/L,respectively.Accordingly,the contents of glycosides were reduced,the content of glycitin and genistein were decreased from 55.53 mg/L to 22.63 mg/L and from 70.98 mg/L to 26.22 mg/L respectively.D-glucuronic acid is a special functional substance in kombucha,whose yield in KFSW was up to 2.766 g/L in the sixth day.By using DPPH radical scavenging ability,ABTS radical scavenging ability,ferric reducing antioxidant power and reducing power as the in-vitro antioxidant capacity of fermented soy whey,the results showed that:80%ethanol extracted more phenol and flavonoids compounds from fermented soy whey,due to the transformation from glycosides to aglycosides,which showed much more bioactivity,thus the fermented soy whey showed a stronger antioxidant activity.And we selected the Staphylococcus aureus,Escherichia coli and Bacillus subtilis as the tested strains,the results showed that KFSW contained more effective antibacterial substances,thus it has a strong antimicrobial activity.At the same time,the fermented soy whey neutralized by NaOH and heating-denatured fermented soy whey exhibited also a weaker antimicrobial ability against Staphylococcus aureus and Escherichia coli,this showed that there are another antibacterial substances in addition to acetic acid and macromolecular protein in fermented soy whey,the antimicrobialability of fermented soy whey is not only due to its organic acid content,but also to other bioactive compounds produced during fermentation time.In addition,because of higher organic acids contents,KFSW can degradate nitrite in vitro from 394.57±1.95 mg/L to 16.64±0.43 mg/L in 48h. |
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