| Based on the actual problem of high alcohol content in the original wine of a Luzhou-flavor winery in the western area of Inner Mongolia,this thesis selects three new cellars with a one-year age(EG group)and three old cellars with a three-year age(CG group)to carry out a tracking test for one fermentation cycle,taking the fermented grains taken at different fermentation time points as the research object,the dynamic changes of the microbial flora,physical and chemical indexes and main higher alcohol content in the fermented grains during the fermentation process were mainly studied.Further research was carried out on the correlation between the microbial flora and the physical and chemical indicators of the fermented grains and the main higher alcohol.Based on this,the reasons for the high content of higher alcohol were found and corresponding solutions were formulated.The main conclusions are as follows:(1)The dynamic changes of physical and chemical indicators of fermented grains during the whole fermentation period were detected by near-infrared spectrometer,and the following rules were found by comparative analysis: the water content of EG and CG groups increased sharply from 0-7 days,and from the 7th day to the end of the fermentation period,the water content of the EG group was significantly lower than that of the CG group;the acidity of the fermented grains in both groups gradually increased with the increase of the fermentation days.After the acidity entered a period of rapid increase;the residual sugar content of the two groups of fermented grains reached the highest value on the 7th day,and then gradually decreased.The starch content in the CG group decreased gradually with the increase of the fermentation days.After 7 days until the end of the fermentation,the starch content in the CG group was significantly lower than that in the EG group,indicating that the starch utilization rate of the EG group was not as high as that of the CG group.(2)The content of higher alcohol in the fermented grains and the original wine was detected by gas chromatography detection technology.Through comparative analysis,it was found that the changes of n-propanol in the fermented grains of the EG group and the CG group were basically the same during the fermentation process,and the content of n-propanol in the original wine was basically the same.The difference was not significant(P>0.05).While the change trends of sec-butanol,isobutanol,n-butanol and isoamyl alcohol in the two groups of fermented grains were basically the same,there were significant differences in the content.The content of n-butanol in the EG group was twice that of the CG group,and the content of sec-butanol,isobutanol and isoamyl alcohol in the CG group was significantly higher than that in the EG group,which were 106.9%,48.8%and 43.4% higher,respectively.(3)Using bioinformatics method to analyze the microbial diversity of the fermented grains,it was found that the bacterial flora of the fermented grains in the EG group and the CG group had the characteristics of diversity,and showed a certain dynamic evolution with the fermentation time.The number of bacterial OTUs in the EG group was 1033,while that in the CG group was 1034,and the number of fungal OTUs in both groups was 1243.Both EG and CG groups identified 25 bacterial phyla and 10 fungal phyla,of which 445 bacterial genera and 413 fungal genera were identified in EG group,while 443 bacterial genera and 413 fungal genera were identified in CG group.The common dominant bacterial genera of EG group and CG group are Lactobacillus,Azospirillum,Acetobacter,Escherichia-Shigella,uncultured＿bacterium＿f＿Muribaculaceae,etc.The common dominant fungal genera are Thermomyces,Thermoascus,Saccharomyces,Aspergillus,Fusarium,Cladosporium,Kazachstania,Mortierella.The dominant fungal genera unique to the EG group were Trichosporon and Botryotrichum,while the dominant fungal genera unique to the CG group were Candida and Penicillium.(4)Calculate the correlation between bacterial and fungal microbial flora and physicochemical indexes of fermented grains according to the Spearman correlation calculation method.The results showed that there was a good correlation between the microbial flora in the fermented grains and the physical and chemical indicators of the fermented grains,that is,the physical and chemical indicators of the fermented grains had an important impact on the distribution and succession of the microbial flora.The water content of fermented grains in the EG group was significantly positively correlated with three fungal genera including Fusarium,the bacterial genera Staphylococcus,the fungal genera Thermoascus and Thermomyces were significantly negatively correlated.However,in the CG group,the water content of the fermented grains was only significantly positively correlated with Kazachstania,the bacterial genera Staphylococcus and Acetobacter were significantly negatively correlated,and the fungus was only Thermomyces;the CG group was significantly correlated with the acidity of the fermented grains.The positively related fungal genera included 12 genera Fusarium,Mortierella,Cladosporium,and in addition to these 12 genera,there were 5 more genera Aspergillus and Candida in the EG group.The fungal genera that were significantly negatively correlated with the acidity of the two groups of fermented grains were Thermoascus and Thermomyces,and these two fungal genera were significantly positively correlated with the starch and residual sugar content.At the same time,17 fungal genera Aspergillus were significantly positively correlated with the acidity of fermented grains in the EG group,but were significantly negatively correlated with the starch content and residual sugar content.In the CG group,most of the 12 fungal genera including Fusarium,which had a significant positive correlation with the acidity of the fermented grains,had a significant negative correlation with the starch content.(5)Calculate the correlation between the bacterial and fungal microbial flora and the main higher alcohol content in the fermented grains according to the Spearman correlation calculation method.The results showed that there was a good correlation between the microbial flora in the fermented grains and the higher alcohol content in the fermented grains.(1)The content of n-propanol(NPA)in the fermented grains in the EG group was significantly positively correlated with two bacterial genera Rhodococcus,uncultured＿bacterium＿c＿Subgroup＿6 and two fungal genera Kazachstania and Saccharomyces,while no positive correlation was found in the CG group.Propanol had a significantly associated flora.(2)The content of sec-butanol(SBA)in fermented grains in EG group was significantly positively correlated with 13 fungal genera Fusarium,Trichoderma,and 3bacterial genera Azospirillum,Dorea and Thermomyces,Saccharomyces and Thermoascus three fungal genera were significantly negatively correlated;and the CG group was also associated with 13 fungal genera that were significantly positively correlated,most of which were the same as the EG group,and the CG group was associated with the unique fungal genera are Mortierella and Plectosphaerella,and the endemic fungal genera associated with the EG group are Candida and Coniochaeta.The EG group was not associated with significantly positively correlated bacterial genera,while the CG group was associated with three significantly positively correlated bacterial genera Sphingomonas and uncultured＿bacterium＿o＿Chloropla.Significant negatively correlated bacterial genera differed between the two groups,with the CG group only associated with Acetobacter.There was one more genus,Rasamsonia,in the CG group than in the EG group,which had a significant negative correlation.(3)The isobutanol(ISOB)content in the fermented grains of the CG group was significantly negatively correlated with one bacterial genus Pediococcus and three fungal genera Thermomyces,Rasamsonia and Thermoascus,and was significantly negatively correlated with Fusarium,Cladosporium,Aspergillus and other 13 fungal genera were significantly positively correlated.However,in the EG group,there was no association with the flora significantly associated with isobutanol.(4)The content of n-butanol(NBA)in the fermented grains of the EG group was significantly positively correlated with 17 fungal genera Fusarium,Candida,Aspergillus,and 3 bacterial genera Staphylococcus,Azospirillum,Dorea and Thermomyces,Thermoascus,Saccharomyces three fungal genera were significantly negatively correlated.However,the n-butanol content in the fermented grains of the CG group was only significantly negatively correlated with one bacterial genus Pediococcus,and was not associated with other significantly related flora.(5)The isoamyl alcohol(ISOA)content in the fermented grains in the EG group was significantly positively correlated with 7 fungal genera including Kazachstania and Fusarium,while the CG group had a significant positive correlation with 9 fungal genera Fusarium,Aspergillus,etc.There were four genera of fungi,Fusarium,Chaetomium,Russula,and Plectosphaerella.Significant negatively correlated bacterial genus was found in both groups Staphylococcus.The EG group had a significantly negatively correlated bacterial genus Weissella,while the CG group had a significantly negatively correlated bacterial genus Pediococcus.There were 2 fungal genera that were significantly negatively correlated with isoamyl alcohol content in the EG group,while 3 in the CG group.The common fungal genera of the two groups were Thermoascus and Thermomyces.It is Rasamsonia.(6)Based on the analysis of the above experimental results,the measures to reduce the content of higher alcohol in liquor are as follows:(1)Measures to reduce the content of n-butanol in the new pit pool(EG group)include:increasing the amount of Northern Luzhou-flavor Daqu to increase the relative abundance of Thermomyces and Thermoascus and other bacterial groups that are significantly negatively correlated with n-butanol;Appropriately lower the temperature of entering the cellar so that beneficial bacteria such as yeast can grow and reproduce better;add a certain amount of active dry yeast to increase the inoculum of yeast;moderately reduce the amount of auxiliary materials(rice husk)to reduce the content of fermented grains.The acidity of fermented grains can be adjusted by controlling the acidity entering the pit and increasing the ratio of grain to grain;the strains that reduce the production of n-butanol can be screened in the old pit mud of the CG group,and then the n-butanol of the EG group can be reduced by microbial disturbance.(2)Measures to reduce the content of sec-butanol,isobutanol and isoamyl alcohol in the old pit Pool(CG group): increase the dosage of Luzhou-flavor Daqu or reasonably mix with Thermomyces and Thermoascus with higher relative abundances The "Chuanpai" Daqu is used to increase the relative abundance of dominant bacteria that are significantly negatively correlated with these three alcohols;appropriately reducing the temperature of entering the pit is beneficial to the proliferation of beneficial bacteria such as yeast,and inhibits the reproduction of some acid-producing bacteria;control the acidity of the fermented grains entering the pit can directly regulate the flora abundance of thermophilic fungi and other significantly negatively correlated fungi;reducing the amount of auxiliary materials(rice husk)and increasing the intensity of stepping on the pit can reduce the oxygen content in the fermentation process.Affect the proliferation algebra of yeast;add active dry yeast,and at the same time,a certain amount of saccharification enzyme can be appropriately added to make the fermentation power and saccharification power more coordinated within a certain range,which can effectively reduce higher alcohol effect. |
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