Lactobacillus Bulgaricus' Decomposing Of Urea,Creatinine By Directive Reduction And Mutation

Obeject:Applying the directive induction and mutation to upgrade thecapacity of Lactobacillus Bulgaricus for decomposing uremic toxins,inorder to provide an alternative strain of probiotics for gastrointestinalbacteriotherapy in CRF.Method:1. Suspend the cells of original L.B in urea and creatinine solution tosee it's capacity for urea or creatinine removal.2. Inoculum and incubate the L.B in 3 different uremia—toxins—rich MRS broth medium for several circles.After 20th generation,testevery 5 generation till the positive result turns out. Havest the bacteria tobe tested and then suspend the cells in urea and creatinine solution tosee their capacity of urea or creatinine removal.5 groups areset:①bacteria after circles of incubation in urea—rich medium②bacteria after circles of incubation in creatinine—rich medium③bacteria after circles of incubation in urea—and—creatinine—richmedium④bacteria after circles of incubation in urea and creatinine—rich medium and then be deactivated⑤no bacteria3. mutation1) Determine the logarithmic growth phase.2) Determine the mixed time of DES with L.B.3) Prepare the cell suspension.4) Original cells are mixed with DES and then dilluted andspreader on the screen plate.5) Harvest the cells and suspend them in the uremic toxinsolution,determine the Residual concentration as the preliminaryscreening6) Select the strains which could degrade the concentration ofuremic toxin for second screening7) The genetic properthy of the new mutant is tested bypass-generation tests.Result: 1 There is no significant change in urea and creatinine solution altersuspension with L.B without any diposal.2 There are significantly lower concentrations in the three groups ofurea and creatinine solution afer suspension with all alive L.B whichwere incubated in 3 different uremia—toxins—rich medium compared tono bacteria group and dead bacteria group.3 But no significant difference between these three groups inthemselves.As to urea residual concentration,the lowest one is theurea+creatinine group;the highest one is the creatinine group. As tocreatinine Residual concentration,the lowest one is the urea+creatininegroup;the highest one is the urea group.4 Mutation by DES, a strain are screened out which has much morecapacity of urea removal than original one.5 But after passage in urea-rich medium,in the 2ndgeneration,there is no significant difference between the mutant and theoriginal one.Conclusion:1 L.B without any treatment has no capacity of decomposing ureaand creatinine.2 Repeated being incubated in high level of uremia toxins—richmedium can induce the capacity of decomposing urea and creatinine;ureaand cteatinine has interaction on each other's induction;adding both ofthem in the same time would enhance the expression of enzyme.3 DES mutation could enhance the capacity of decomposingurea,but the mutant's genetic property is under promoting.