Mutation Of The Strain And Its Optimization Of Culture Conditions For The Conversion Of Dextral-fosfomycin

Fosfomycin is a new kind of broad-spectrum antibiotics with simple structure and low molecular weight,which possesses bactericidal effect on Gram-positive and Gram-negative.It is referred as a new kind of antibiotics of the twenty-first century by international pharmaceutical industry.At present,the total yield of fosfomycin produced by chemical synthesis is less than 35%.Among the chemical products,only 50%levo-fosfomycin has antibiotic activity,and the other half products of dextral-fosfomycin has non-antibiotic activity.This not only wastes resources,increases product cost,but also pollutes the environment.Therefore,it is of great significance to the industrial production of fosfomycin by conversing the dextral-fosfomycin in order to obtain the effective use of dextral-fosfomycin.Microbial conversion method has a highly specificity,and the reaction condition is mild,the process is simple,the cost is low.Microbial conversion method can complete the reactions which are difficult to finished by chemical method.By using the microbial transformation method,a part of the dextral-fosfomycin can be converted into an available levo-fosfomycin using microbial strain having a transformation ability.In this study,one strain of Bacillus amyloliquefaciens was screened out and subjected to physical and chemical compound mutagenesis.The optimal time of mutagenesis of Bacillus amyloliquefaciens was 90s.After UV mutagenesis,a mutant strain of UV2 was selected and fermented.It was found that the transformation ability of mutant strain UV2 was improved.The conversion rate was 11.76%.UV2 compared with the original strain,the conversion rate increased by 3.84%.The UV2 was further subjected to diethyl sulfate(DES)mutagenesis,the optimal mutagenesis time was determined as 40min.The conversion ability of the mutant strain U-D2 was improved,the conversion rate was 13.2%which was 1.44%higher than that of UV2.U-D2 increased the conversion rate by 5.25%compared with the original strain of Bacillus amyloliquefaciens.The mutant strain of U-D2 had good genetic stability by culturing 6 generations,which determined by the broth inhibition test with Staphylococcus aureus as a indicator strain.The nutrient composition of U-D2 was optimized by single factor test combined with the design method of CCD center in the response surface design method.The optimal medium concentration was as follows:beef extract 3.62 g/L,glucose 12.39 g/L,peptone 7.19 g/L,MgSO40.42 g/L,K2HPO40.85 g/L.The experimental results showed that the diameter of the inhibition zone was 37.4mm determined with inhibition test using the U-D2 culture broth under the optimal conditions,and the conversion of U-D2 was 15.51%,compared with the medium before optimization,the conversion rate increased by 2.31%.