Fermentation Of Hawthorn Vinegar Using Excellent Strains Isolated From Shanxi Aged Vinegar

Fruit vinegar fermentation is mostly carried out by natural or artificial inoculation.Natural fermentation mainly uses the bacteria carried in the air and on the surface of the fruit to ferment without adding any starter,which is easy to perform but difficult to meet people's needs due to long fermentation time,inefficiency and difficulty to control the fermentation process.Liquid fermentation for fruit vinegar in China usually are usually inoculated with Acetobacter rancens(AS1.41)or Acetobacter lovaniense(1.01)and the vinegar produced is slightly bitter,and the organic acid type and flavor substance are relatively too single.Compared with fruit vinegar,the traditional brewed cereal vinegar tastes soft and mellow,which is the result of the coexistence and metabolism of many microorganisms in the fermentation process.Shanxi aged vinegar is a kind of traditional Chinese cereal vinegar.Its acetic acid fermentation stage is a spontaneous solid-state fermentation system that is rich in microbial resources.Many microorganisms involved in fermentation are directly produced by metabolism or a combination of various substances such as alcohols,esters and organic acids formed by synthesis,which altogether give the complex and unique flavor of Shanxi aged vinegar.After long-term domestication,the functional strains involved in fermentation become more tolerant and more resistant to diseases.Based on this,this study analyzed the diversity of microorganisms in the fermentation process of Shanxi aged vinegar by high-throughput sequencing combined with pure culture technique,and selected the excellent strains suitable for the fermentation of hawthorn vinegar.And the selected excellent strains were applied to the fermentation of hawthorn vinegar by synergistic fermentation to improve the flavor characteristics and enrich the resources of the fruit vinegar.The main research results are as follows:1.At the genus level,the main bacteria involved in the acetic acid fermentation(AAF)process of Shanxi aged vinegar were Acetobacter,Lactobacillus,Weiss,Streptococcus,Bacillus,Pseudomonas,etc.,among which Lactobacillus and Acetobacter were the dominant genus.The species and abundance of bacteria was obviously higher in the samples at the prometaphase of fermentation than that in the samples at the end of the fermentation.The biomass of LAB was the highest,followed by the Acetobacter.At thespecies level,LAB included Lactobacillus helveticus,Lactobacillus mucosae,Lactobacillus panis and Lactobacillus secaliphilus,and only Acetobacter pasteurians and Gluconacetobacter europaeus were detected in AAF.2.A total of 238 microbial strains,comprising 80 yeasts and 158 strains of bacteria were isolated from the alcohol fermentation(AF)and AAF process of Shanxi aged vinegar by pure culture technique.The obtained microorganisms belonged to 31 species through colony morphology,microscopic observation and molecular identification.A total of 6 yeast species were identified,including Saccharomyces cerevisiae,Pichia scaptomyzae,Candida ethanol,Candida humilis,Pichia membranifaciens and Rhodotorula;a total of 13 LAB species were identified,including Lactobacillus fermentum,Pediococcus acidilactici,Lactobacillus plantarum and so on;a total of 11 LAB species were identified,including Bacillus amyloliquefaciens,Bacillus cereus,Bacillus licheniformis and so on.The isolated acetic acid bacteria were identified as Acetobacter pasteurianus.3.Candida ethanol has an ester-producing capacity of 33.48±0.22 g/100 m L,and it could survive in environment with 9%(v/v)ethanol concentration,p H3.0,40 g/100 m L glucose concentration,and temperature no more than 40?.The fermentation technology of hawthorn wine was obtained by response surface analysis: 18.5 g/100 m L glucose concentration,sequential inoculation of 2% C.ethanol and 1% S.cerevisiae,and fermentation at temperature 28.5? for 6 days.The fermented hawthorn wine was full-bodied,slightly acidic,rich in hawthorn aroma,with an alcohol content of 9.5% vol and a total ester content of 0.68 g/100 m L.4.A.pasteurianus was screened from 22 strains of acetic acid bacteria by acid production test and environmental tolerance analysis.The acid-producing ability of A.pasteurianus is 35.5±0.48 g/L,which could survive in the environment of temperature 42?,10%(v/v)ethanol concentration and 5.0 g/100 m L acetic acid concentration,and the genetic performance was stable.The acid-producing ability of Pediococcus pentosaceus screened by the same method was 9.56±0.23 g/L,which was able to withstand temperature 50? and 12%(v/v)ethanol concentration.5.Three excellent strains were applied to hawthorn vinegar brewing by synergistic fermentation.In the alcohol fermentation process,at temperature 28?,inoculated 2% C.ethanol and 1% S.cerevisiae,and fermented with the first 2d open and the next 5d sealing;as for the fermentation of acetic acid,inoculated5% of both P.pentosaceus and A.pasteurianus,fermented at temperature 30? for 6 days with the rotatingspeed 130 r/min,and the total acid content of hawthorn fruit vinegar produced was 4.29 g/100 m L,total ester content 1.97 g/100 m L,non-volatile acid content 0.72 g/100 m L.Finally,SPME-GC-MS technology was utilized to identify the aroma components of hawthorn vinegar fermented by different strains.The results show that about 90% of total volatile substances were acids,esters,alcohols,aldehydes and phenolic compounds,of which ethyl acetate,ethyl benzoate,acetic acid,caproic acid,furfural,benzaldehyde and eugenol were the main components of aroma.The product hawthorn fruit vinegar is brownish red and bright,and it tastes sour,soft and slightly sweet.