Studies On Isolation And Identification Of Acetobacter And Culture Of Kombucha

Kombucha is a kind of folk acid health drink,and has many benificial effects to human health.There are many disavantages to culture kombucha in traditional way.There are many problems to produce kombucha beverage with traditional method,as long period, easy contaminations and low yield rate would happen.Firstly,experiments were designed to isolated acetic acid bacteria from vinegar paste. According to the characteristics of acetic acid bacteria in kombucha,kombucha culture media with different pH were designed to enrichment acetic acid bacteria strains,and supplement of a yeast strain and ethanol were applied to accelerate the growth of the acetic acid bacteria.The fifteen candidate strains were isolated from vinegar paste by the results of gram stain,acetic acid production quantitatively,membrane production.All these fifteen strains were common in gram-negative stain,producing acetic acid and producing membrane.Eleven carbon utilization tests and fourteen physiological biochemical tests were used to identify these strains,the results showed that the fifteen acetic acid strains belong to Acetobacter Xylinum,Acetobacter aceti subsp,Acetobacter pasteurianus subsp Lovaniensis,Acetobacter pasteurianus subsp Pasteurianus.Strains A.xylinum C203 and S.cerevisiae SC03 were inoculated in medium respectively,and then cultured in static and shook conditions in kombucha culture medium. In the pure culture test,the growths of C203 and SC03 were faster in shook flask than that in static flask.The time that the strains reached the peak were shortened obviously,and the maximum cell numbers of the two strains could reach 5.1~*10~7/ml and 3.9~*10~7/ml respectively,and were 3.7 times and 2.7 times than those of in static cultivation, respectively.Then strains of C203 and SC03 were inoculated into the kombucha medium together,and grew in static and shook condition respectively,The cell numbers of strains, content of the total acid and total sugar concentrations were determined.The results showed that the growth of the strains were accelerated obviously when in shook cultivation, and the time that the cell numbers reached the peak shortened obviously;the maximum number of strains could reach to 8.7~*10~7/ml and 6.6~* 10~7/ml respectively,and were 4 times and 3 times of those in static condition,respectively.The total acid reached 6.5g/L after 10d in static cultivation,but the speed of the total acid production was accelerated obviously in shook flask reaching to the peak with 3.5 days period.Due to having no membrane production,the sugar consumption in shaked cultivation was less than that in static cultivation.When the fermentation ended,the yield(total acid/sugar consumption) were calculated,and the yield of shook cultivation was 0.69,two times of the yield in static cultivation.There would be such advantgae as time saving and cost reduction when the method of prepation kombucha in shaking condition was used in industry production.In order to ensure whether the isolated acetic acid bacteria are suitable to produce kombucha beverage,the fifteen acetic acid bacteria were co-cultured respectively with SC03 in the kombucha culture medium,determining the major metabolites in each culture medium.The results showed that treatment 5(A5 strain) had the lowest total acid of 14.0g/L,treatment 14 had the highest total acid of 21.5g/L.All of the treatments could produce about the equal amout acid in mature kombucha;treatment 12 had the lowest total sugar of 14.5g/L,and treatment 13 had the hightest total sugar of 26.1g/L.Comparison being made for the yields,treatment A7 had the lowest yield of 0.46,and treatment A10 had the highest yield of 0.66.The taste of each kombucha by different strain were different, and the taste is closely related to the content of major metabolites in culture medium.So suitable strains could be choose to produce kombucha beverages to satisfy the different demands of people,.The content of DSL in each treatment were determined,and the results showed that there wasn't DSL in treatment 9,and there were satisfied concentration of DSL in the rest of all treatments with other strains.Kombucha with strain A2 had the lowest content of 0.38mg/ml,and highest content of 1.18mg/ml for strain A14.The fourteen acetic acid bacteria would be the ideal strains to produce kombucha drinks with detoxification and cancer prevention functions.