Study On Characteristics And Isolation,screening,Identification Of Bacillus Natto

Bacillus natto is a probiotic which is isolated from natto,its effectiveness is not only related to the active ingredients by secreting in its own metabolism,but also associates with the viable count that reaches the human intestine.The research focused on metabolites of natto at present,there were few reports on high-activity bacillus natto and stability in vitro.There was a strain highly-activity bacillus natto obtained by isolation,screening and identification from natto.Then the natto which was rich of bacillus natto was obtained after optimitized the solid fermentation process.And the flavor was improved by adding spices.Finally,the bacillus natto was used to investigate the stability in simulated gastrointestinal fluid.The main results were as follows:?1?Isolation,Screening and Identification of Bacillus natto and Comparison Its Fermentation Characteristics.Bacillus natto A1,B2,C2 with high activity of protease activity were obtained after colonies morphology characteristics and skim milk hydrolysis test.The result of initial fermentation with soybean showed that the fermentation of Bacillus natto C2 was the best,its proliferation ability?multiplication ratio 116?,mucus yield?5.93±0.70%?,and drawing length?14.80±3.56 cm?are superior to the other two strains.Its optimal growth temperature was 37?,optimal growth pH was 7.0 in liquid culture conditions.?2?Optimized for solid fermentation of bacillus natto C2.With the viable count as index,the optimal solid fermentation conditions were determined by single factor and orthogonal experiment.The results showed that the best solid fermentation process was inoculation temperature of 60?,soaking water pH of7.0,inoculation size of 7%,and fermentation time of 36 h.The viable count was reached 2.6×10¹⁰ CFU/g which was nearly 6 times higher than before under this condition.Spicy natto formula:4‰thirteen fragrant powder,1.0%salt,2.5%chili powder,2.0‰MSG.Sour and sweet natto formula:2.0‰citrus flavors,4.0‰sour plum powder,2.0%sucrose,4.0%orange powder.?3?Study on the stability of bacillus natto C2 in simulated gastric juice.The bacillus natto C2 of the solid and liquid carriers were inoculated into the simulated gastric juice which had different pH and viscosity?changed by the amount of agar?,and the stability mechanism in the simulated gastric juice was analyzed.The results showed that under the same conditions,the lower pH value,the lower survival rate of bacillus natto C2;The survival rate was related to the viscosity of simulated gastric juice,the survival rate of bacillus natto was the highest in the simulated gastric juice with 0.3%agar addition;the survival rate of bacillus natto in the solid carrier was greater than the survival rate of bacillus natto on the liquid carrier;After 3 h in gastric juice,the minimum survival rate of bacillus natto C2 of the solid and liquid carriers?pH 1.5,0%agar addition?were 52.1±0.3%and 41.3±0.1%,We can thought bacillus natto C2 has good stability in gastric juice.?4?Study on the stability of bacillus natto C2 in simulated intestinal fluid.The effects of trypsin,bile salt and viscosity on bacillus.natto C2 under solid and liquid carrier conditions were investigated.The results showed that pancreatic enzyme and bile salt had damage to bacillus natto C2,and the damage was synergistic.In the simulated intestinal fluid,the survival rate of bacillus natto C2 decreased with the increase of viscosity,the survival rate of bacillus natto was the highest in the simulated gastric juice with no agar addition,the survival rate of bacillus natto in the solid carrier was greater than the survival rate of bacillus natto on the liquid carrier;After 4 hours in intestinal fluid,the lowest survival rate of bacillus natto C2 of solid and liquid carriers?adding trypsin,bile salt,agar addition 0%?were 20.0±0.3%and 16.3±0.1%,It can be determined that bacillus natto C2 has stability in intestinal fluid.We finally obtained a Bacillus natto C2 which with high protease ability and excellent fermentation.The minimum survival rates of bacillus natto?solid and liquid carriers?after gastrointestinal fluid were 10.4%and 6.7%,If the natto were ate which inrich bacillus natto,and the viable count in the intestine can reach 2.9×10⁹ CFU/g,which can ensure enough bacillus natto enters to intestines.