Metabolomics Of Soy Sauce Brewed By Aspergillus Niger And Aspergillus Oryzae

In the brewing process of soy sauce,many kinds of microorganisms grow and interact with each other,and through their rich enzyme system,the microorganisms can produce the proteins,polysaccharides and sugars in the raw materials into the soy sauce.The hydrolysis of macromolecular substances,which ultimately together constitute the unique taste and flavor of soy sauce.Metabolomics is the hotspot of current research,but it is seldom used in food fermentation research.Therefore,this project simulates the brewing process of high-salt rare state soy sauce and investigates the effects of Aspergillus oryzae brewing and its synergistic brewing with different ratios of Aspergillus niger on the taste and flavor of soy sauce.The changes of metabolites and metabolic pathways in the process of maltogenesis and brewing,combined with metabolomics and bioinformatics,are the key factors for microorganisms in the fermentation process.The study of interaction mechanisms provides new ideas.Its main conclusions are as follows.1.The experiments were carried out with the curds collected at the A.oryzae making stage up to 8h,16 h,24h,32 h and 40 h.The study identified changes in the metabolites of A.oryzae at different times: flavonoids,triazines,sulfonamides,indoles.The detection levels vary,with triazines increasing over time and flavonoids decreasing over time.The greatest variation was observed between 8 and 24 hours after the start of the malt making process,when microbial growth was strongest and metabolism was most rapid.After 24 hours of koji making,the level of metabolites was stable and microbial growth was slow.2.Metabolite characterization and multivariate analysis were carried out on the fermented kojis with two groups of single-strain inoculation and three groups of mixed-mix inoculation with different inoculation ratios.Statistical analysis of variables revealed significant differences in metabolites between the five groups,with smaller differences between the four groups inoculated with A.oryzae.The difference with the A.oryzae.inoculation group was large,and this difference was related to the time and the compound ratio,with the longer the time spent in making the koji.The greater the amount of A.niger inoculation,the greater the difference.The greatest differences between the five samples were observed between 8 and 24 hours after the beginning of the production,indicating that the growth was most intense at this time.After 24 hours the differences between the samples became smaller,indicating stable metabolite levels and slow microbial growth at this time.It was found that the access of A.niger could increase the levels of methanesulfonic acid,3-indolebutyric acid,guanidinoic acid,heavy metals and other organic acids detected in the borer.oleic acid,3-alpha-hemephenolic acid,varanic acid and other organic acids,and to reduce the content of indoleacetic acid,gallic acid.The content of organic acids such as eugenic acid and teichoic acid.The content of organic acids such as arachidonic acid,stearic acid and ?-glycine first decreased with the addition of A.niger and then rose again at 1:1.Compared with the fermentation of only A.oryzae,the addition of A.niger was able to cause significant changes in the metabolites of soy sauce making.This was mainly reflected in the changes in the content of dipeptides,flavonoids,unsaturated fatty acids,carotenoids,phenols and indoles,and was similar to the changes in the metabolites of the fermentation of A.niger.The relationship exists between the fungus complex ratios.3.In combination with the enrichment analysis of the metabolic pathway,it was found that the glycerol phospholipid metabolic pathway was the most important pathway in the mixed production process of A.oryzae and A.niger.disturbed metabolic pathways.The ?-alanine metabolic pathway,the glycerol ester metabolic pathway,the pantothenic acid and coenzyme A biosynthetic pathways are associated with a 1:1 complex ratio.The arginine biochemical synthesis pathway is somewhat disturbed;at a complex ratio of 2:1,the ?-alanine metabolic pathway,the glycerol ester metabolism pathway,pantothenic acid and coenzyme A biosynthesis pathway,lysine degradation pathway,and purine metabolism pathway are to some extent affected by the purine metabolic pathway was disturbed to some extent at the complex ratio of 3:1.The integrated metabolic pathway analysis revealed that the interference with the glycerol phospholipid metabolic pathway might be caused by the secreted bisphosphatidyl phosphatidylcholine of A.niger.Glycerol synthase,2-acylglycerol O-acyltransferase,D-serine deaminase,glycerol dehydrogenase and other enzymes that A.oryzae cannot product.4.the addition of A.niger was able to cause significant changes in the mash fermentation stage compared to the monoculture fermentation of A.oryzae.A multivariate statistical analysis of the mash fermentation metabolites of the five groups of samples revealed significant principal component differences between the five groups.This difference changed with time.It is mainly reflected in the fact that,in the first 20 days,the mixed fungal group of flavour substances in the ratio of 2:1 and 3:1(between A.oryzae and A.niger)and the pure A.oryzae were After 30 days of mash fermentation,the flavors of the three types of hybrid bacteria were closer to each other.In combination with the metabolic pathway enrichment analysis,it was found that when the mash fermentation was 80 days old,the three types of mixed bacteria were more similar in flavor than the pure A.oryzae fermentation group.Fermentation can make 2-Ethylbutanol,2-Phenylacetic acid,2-Methylpropionate,2-Phenylethyl-2-phenylacetate,2-Methyl-(2-methyl-)3-Prop-1-en-2-ylpyrazine,4-hydroxy-2,5-dimethylfuran-3-one,1-(2,5-dimethyl),4-hydroxy-2,5-dimethy lfuran-3-one,1-(2,5-dimethyl)(Furan-3-yl)ethanone,2-phenylacetaldehyde,2-phenylpropi onaldehyde,2-phenylethanol,tetradecanal,2-methyloxyethanol,hexadecyl3-(2-methylpro pyl)pyrazine,2-methoxy-3-propyl-2-pyrazine,3-ethoxy-4-hydroxybenzaldehyde.Decrease in 3,4-dimethoxybenzaldehyde,1,4-dimethylbenzene,2-ethylbutyric acid,3-methylbutyric acid and other substances.