Preliminary Study On Biotransformation Of Levo-fosfomycin By Dextral-fosfomycin

Fosfomycin is a kind of micromolecule, broad-spectrum antibiotic. It has a role in killing against Gram-positive bacteria and Gram-negative bacteria. With the gradual and further studies of pharmacological research, the application domain of fosfomycin expands. The industrialization preparation of fosfomycin is by chemical synthesis methods currently, due to its simple chemical structure. But the total yield of this method is less than 35%, and the products are raceme including equal amount of levo-fosfomycin and non-bioactive dextral-fosfomycin. This will not only lead to high production costs and resources waste, but also cause environmental pollution at the same time. Therefore, the re-utilization dextral-fosfomycin is an effective way to reduce production costs, to increase output and to reduce waste discharge.The studies are focused on microbial transformation of dextral-fosfomycin. The research aimes at screening for positive strains which have the activity to biotransfore dextral-fosfomycin to levo-fosfomycin by primary and secondary screening from 2856 fungi strains. Fifteen fungi strains were identified as positive strains by the method of thin-layer chromatography (TLC) combining with bioautography. These positive strains were also identified based on morphological characteristics of colonies, conidium and conidiophore, as well as ITS-5.8S sequence, they were ten species of Penicillium, three of Aspergillus, one of Botrytis cinerea, one of Fusarium avenaceum. Strain 2221 (identified as Penicillium camemberti) was selected to use for the future experiments because of its higher conversion efficiency.The transformational products were separated from culture broth. Firstly, conversion solution was adjusted to pH 7.5, then it was filtered through activated 717 anion-exchange resin column, eluted with2%NH4Cl. Next, after chiral separation and purification, the product compound was analysed by capillary electrophoresis, mass spectrum, optical rotation(-1.5°) and thin-layer chromatography (TLC) combining with bioautography, finally, it was identified as levo-fosfomycin.The transformation processes were optimized in this research. The optimum conditions are as fellows:seed medium is defined as PDA fluid medium including 1%glucose, transformation medium is defined as PDA fluid medium including 0.3%glucose and 0.3%substrate, pH natrue, the 250 mL flasks is filled with 60 mL transformation media, seeds culture for 24 hours, inoculation amount at 10%, culture tempreture 25℃-28℃, conversion for 4 days. The biotransformation rate attained to 22%.