Isolation Of Acetoin-producing Microbial Community In Vinegar Pei And Its Application In Vinegar Brewing Process

Ligustrazine is an active compound with effect of cardiovascular protection effect in traditional fermented vinegar.This study focused on the formation of acetoin-an important precursor of tetramethylpyrazinet in Zhenjiang aromatic vinegar and used molecular biology technology to isolate microorganisms with acetoin-producing function.Then,the acetoin-producing ability of microbial community was enhanced through batches domestication in wine mash medium.It provided research foundation for acetoin and liquid vinegar producing based on microbial community.Setting different inoculum volume of starter Pei(last batch of vinegar Pei on day 7)to disturb the fermentation system.Dynamic changes of metabolites and microbial community were analyzed.The results showed that the main microorganisms in starter Pei included Lactobacillus and Acetobacter,it can be viewed as a functional microbial community to start fermentation.The greater the amount of inoculum volume,the faster the process of fermentation and the stronger the synthetic ability of acetoin.Using different medium(wine mash,water-bran extract,wine mash-bran extract)and different culture methods(the agitate culturing;the static culturing;the intermittent culturing)to separate different functional microbial communities associated with the production of acetoin.The analysis showed that wine mash could provide abundant carbon sources,mainly including ethanol and lactic acid,and the nitrogen sources were less while bran could provide more sugar and nitrogen sources.More acetoin could be accumulated with the method of intermittent culturing.With this culture method,the concentration of acetoin in each substrate system increased significantly in the first 2-3 days.Moreover,the content of acetoin was comparatively higher in wine mash medium and wine mash-bran extract medium than water-bran medium,indicated that wine mash may be the main substrate for microorganisms to produce acetoin.High-throughput sequencing method was used to analyze the bacterial community structure during the fermentation with intermittent culture method in each substrate system.The results showed that the structure of microbial community was different in different medium.The analysis revealed that microorganisms associated with acetoin-producing mainly included Acetobacter sp.WS8 ? Acetobacter pasteurianus ? Lactobacillus sp.D76 ?Lactobacillus helveticus?Lactobacillus sp.strain L7?Lactobacillus sp.strain L8 in wine mash mediun;Acetobacter sp.WS8?Lactobacillus acetotolerans?Lactobacillus sp.strain L7?Lactobacillus buchneri?Acetobacter pasteurianus?Acetobacter malorum?Lactobacillus casei in water-bran extract medium;Acetobacter sp.WS8 ? Lactobacillus acetotolerans and Acetobacter pasteurianus in wine mash-bran extract medium.The experiment of exogenous addition of lactic acid verified that A.pasteurianus in the community could utilize lactic acid to produce acetoin.It also demonstrated that wine mash with rich lactic acid may be the main substrate for microbial metabolism to acetoin,and in functional microbial communities,the presence of Lactobacillus may contribute to providing lactic acid,which can promote the accumulation of acetoin by Acetobacter.In addition,the acetoin and acid producing ability of microbial community was enhanced through batches domestication in wine mash system.The dynamic changes of bacterial community structure in the first three batches were analyzed.The results showed that a functional microbial community mainly including Acebacter sp.WS8,Lactobacillus sp.stain L7.Lactobacillus sp.strain L8 and Uncultured Lactobacillus sp.clone 22 was finally formed during the process of batches domestication.