Study On The Optimization Of Milk Beer Strains,the Preparation Of Freeze-dried Bacterial Powder And The Fermentation Process Of Milk Beer

Milk beer is a new type of milk drink with unique style made from animal milk as raw material and fermented by lactic acid bacteria and yeast twice.Its taste is sweet,sour and refreshing,the texture is delicate and smooth,and the aroma is harmonious and full,which is more and more popular with consumers.Yeast affects the taste and aroma of milk beer.Now,milk beer production enterprises mainly use beer yeast or Saccharomyces cerevisiae for milk beer fermentation production,but in the process of milk beer fermentation,beer yeast or Saccharomyces cerevisiae need to add a lot of hops and malt and other auxiliary materials,resulting in complicated process,which significantly increases the cost.In addition,the milk beer produced by beer yeast or Saccharomyces cerevisiae fermentation has the same flavor,single variety and heavier beer taste,which makes it unable to meet the diverse needs of consumers for milk beer products.This has severely restricted the development of milk beer beverages.Although a few companies have begun to use the selected high-quality milk-source yeast to ferment the milk beer and improve the flavor quality of milk beer,the lack of active yeast for milk beer fermentation is not conducive to large-scale milk beer.Industrial production.Therefore,it is of great significance for improving the quality of milk beer and promoting the long-term stable development of milk beer industry in China to select fine quality yeast from milk source and develop the preparation of its high-activity lyophilized bacterial powder.In this study,a high-quality yeast strain was isolated and screened from different sources of kefir grains,and was identified by 5.8S r DNA sequence analysis.Secondly,the high density fermentation conditions were optimized to provide high concentration of cells for active lyophilized bacterial powder.Then the preparation process of high-quality freeze-dried yeast powder from milk was optimized.Finally,the fermentation technology and stability of milk beer were studied.The main research results obtained in the experimental research are as follows:(1)Isolation and screening of a high-quality milk-derived yeast strain-Kluyveromyces marxianus LNJ10.Wild kefir grains collected from Xizang,Heihe,Liaoning,Inner Mongolia and other regions were used as the source of yeast isolation,and a total of 68 dominant strains were isolated.After colony morphology observation,gram staining and catalase test,preliminarily determine 14 strains of yeast,the molecular biology identification of 14 strains of yeast,were identified as:6 strains of Pichia pastoris,5strains of Kluyveromyces marxianus,1 strain of Saccharomyces cerevisiae,1 strain of Kazachstania unispora and 1 strain of Issatchenkia onentalis.14 strains of yeast were fermented and screened from four aspects of sensory evaluation,genetic stability,physical and chemical health indexes and volatile components.Finally,a strain LNJ10was determined,and the quality of fermented milk beer was the best.Finally,the phylogenetic tree of LNJ10 was constructed to further determine its species and genus.(2)The high density fermentation conditions of LNJ10 were optimized to obtain a high concentration of yeast cells.Through analysis of single factor and response surface results,it was found that the optimal medium formulation of strain LNJ10 was:Malt wort was used as the basic medium,molasses 39.005 g/L,corn steep powder 15 g/L,calcium carbonate 2.028g/L,vitamin B₁0.019 g/L.Under these conditions,the maximum concentration of viable bacteria was 9.3×10⁸CFU/m L.On this basis,orthogonal experiment was conducted to optimize the culture conditions of strain LNJ10.The optimal culture conditions were as follows:temperature 30?,inoculation concentration 1.5×10⁶CFU/m L,rotation speed 200 rpm,initial p H 5.5,and the maximum viable strain concentration could reach 9.84×10⁸CFU/m L under these conditions.In a 5L fermenter,the strain LNJ10 was cultured in batches and fed in batches,and it was found that the fed batch culture could significantly promote the growth of the strain.The concentration of live bacteria in batch culture was 4.55×10⁹CFU/m L,and the dry weight of bacteria was 35.33 g/L,which was 2.46 times higher than that in batch culture(1.85×10⁹CFU/m L),and 4.62 times higher than that in shake flask culture.(3)The freeze-dried protective agent formula and freeze-drying process of Kluyveromyces marxianus strain LNJ10 were optimized,and the storage conditions of freeze-dried bacterial powder were studied.By single factor and orthogonal test,the optimal formula of lyophilized protectant was as follows:sucrose 7%,skimmed milk powder 15%,maltodextrin 17%and glycerin 4%,and the survival rate of lyophilized cells of LNJ10 strain was82.89±1.11%.The best freeze-drying conditions are determined by single factor experiment:pre-freezing at-40?for 12 h,the best mixing ratio of bacterial cells and compound protective agent is 1:1,the best balance time between the two is 40 min,freezing The dry survival rate can reach 86.07±1.03%.The lyophilized bacterial powder of strain LNJ10 was packaged in vacuum aluminum foil bags and placed in-40?refrigerator.After 3 months of storage,the lyophilized survival rate of strain LNJ10 cells was 78.81±1.28%,and the number of viable bacteria per unit bacterial powder was 5.976×10⁹CFU/g.The experiment of producing milk beer by lyophilized bacterial powder of strain LNJ10 and fresh bacterial liquid showed that the fermentation performance of lyophilized bacterial powder of strain LNJ10 was stable and the quality of milk beer was uniform.(4)The fermentation process of milk beer was optimized and its stability was studied.Through single factor and orthogonal analysis,the optimal fermentation conditions of milk beer were as follows:inoculation concentration of yeast 2.0×10⁶CFU/m L,fermentation temperature 28?,fermentation time 24 h,sucrose content 9%.On this basis,the optimal formula of compound stabilizer for milk beer was determined by orthogonal test,which was 0.15%xanthan gum,0.5%sodium carboxymethyl cellulose and 0.15%sucrose ester.The volatile component analysis of the optimized milk beer samples showed that the aroma components of the optimized milk beer were more abundant than those of the optimized milk beer samples,and the taste and aroma of the optimized milk beer were obviously improved.