Isolation And Identification Of Lactic Acid Bacteria From Mulberry And Its Application

Mulberry is rich in nutrition and is a good raw material for lactic acid bacteria fermentation.Commercial lactic acid bacteria are difficult to grow in Asian mulberries,which have low sugar content and low pH.In order to solve this problem,we isolated lactic acid bacteria from mulberry,and screened lactic acid bacteria with good substrate adaptability.Furthermore,the tolerance in simulated gastrointestinal solution and bacteriostatic activity,self-agglutination and hydrophobic ability of lactic acid bacteria strains were studied.The mulberry pulp was fermented using Lactobacillus plantarum which isolated from mulberry fruit.The changes in viable counts,pH value,titratable acid,organic acid,anthocyanin,total phenol content and in vitro antioxidant capacityduring fermentation were investigated.The fermented mulberry pulp was prepared into freeze-dried powder.The length of vacuum freeze drying time and the prefreezing time were determined,and three kinds of freezed dried protectants of lactic acid bacteria were screened by single factor experiment.The optimum formulation of three kinds of lyophilization protectants was obtained by response surface experiment.The main results were as follows:(1)10 strains of lactic acid bacteria were isolated from natural fermented mulberry.After microbial physiological and biochemical identification,sugar fermentation test and 16 SrDNA gene molecular identification,7 of 10 lactic acid bacteria strains were Lactobacillus plantarumum and 3 strains were Lactobacillus fermentumum.By comparing the growth and acid-producing ability of 10 strains in mulberry,Lactobacillus plantarum Lb-1 and Lb-6 were selected as the further study strains.(2)The experimental results of tolerance to artificial simulated gastrointestinal fluid showed that Lactobacillus plantarum Lb-1 and Lb-6 had certain tolerance to bile salt solution,but had better tolerance to gastric and pancreatic solution.In the bacteriostatic experiment,two isolated strains had obvious inhibitory effect on Escherichia coli,but the inhibitory effect on Staphylococcus aureus was general.The results of self-agglutination test of lactic acid bacteria showed that the rate of self-agglutination of two strains was higher than 40%.The cell surface hydrophobicity of two isolates was measured.The results showed that the two isolated strains had higher surface hydrophobicity than 50%,and had strong cell surface hydrophobic ability.(3)The results revealed that Lactobacillus plantarum has good substrate adaptability under anaerobic fermentation at 30?.After fermentation,the number of live bacteria has increase to stable above 9.5 log cfu/mL and a large amount of lactic acid was produced.Succinic acid,cyanidin3-glucoside and cyanidin3-rutinoside were the dominant organic acids and anthocyanins in mulberry pulp,respectively.During fermentation,the content of lactic acid increased while the contents of cyanidin3-glucoside,cyanidin3-rutinoside,total anthocyanin,and total phenolic contents decreased.The antioxidant capacity increased firstly,and decreased secondarily during the fermentation.Correlation analysis showed that the decrease of anthocyanin and total phenolics was the main reason for the decrease of antioxidant capacity during fermentation,and Lactobacillus plantarum affected the content of anthocyanin and total phenol and antioxidant capacity to some extent.(4)Single factor experiment results showed that the length of vacuum freeze drying time was 24 hours and the prefreezing time was 30 min at temperature-80 ?.Mycose,D-sorbitol and glucose were selected as freeze-drying protectants for lactic acid bacteria.The regression equation was optimized by Design expert.v8.0.6 software.The optimum formula of three kinds of lyophilized protectants was that the amount of D-sorbitol was 3.72%,trehalose was 4.12%,glucose was 2.28%.Under these conditions,the survival rate of lactic acid bacteria was 93.99.The survival rate of lactic acid bacteria in the control group increased from 87.37% to 93.93%,which proved that the model can be used to predict the actual production.