Research On Producing Enantiopure Glycidyl Azide By Biotransformation

This topic of epoxide hydrolase producing strains mainly use racemic glycidyl azide as substrate to screen. It was analyzed by high performance liquid and obtained a high enantioselective strain of Aspergillus niger from special soil which collected around the country, its ITS gene sequences was detected and it was named Aspergillus niger ZJUTZQ208.Optimized the fermentation conditions of Aspergillus niger ZJUTZQ208 and investigated the conditions such as the carbon and nitrogen sources, metal ions, the fermentation temperature, fermentation time, initial pH and substrate-induced which effected the cell biomass and enzyme activity. Through this single factor experiments determined when the temperature is 30℃, initial pH 7.0 and fermentation time 72h, biomass of cell and enzyme activities achieved to the best, and know that produced epoxide hydrolase by the strain is the constitutive enzyme. At the same time, the optimal concentrations of carbon source, nitrogen source and metal ions were determined by response surface methodology. To the two methods, the optimal fermentation medium of the strains was ultimately determined as follows:soluble starch 15.23g/L, soybean powder 10.88g/ L, K2HPO4·3H2O 0.8g/L, KH2PO40.4g/L, NaCl 10g/L, MgSO4·7H2O 0.98g/L. Under the optimal conditions, the cell’s enzyme activity reached 128.1U/L.The bioresolution conditions were optimized, and ultimately determined the best resolution conditions:0.1M pH 7.2 potassium phosphate buffer, cell/substrate ratio 8:1, temperature 30℃, time 10h, could give the ees value of substrate is 98.5%, the E value 21.2.The substrate specificity of propylene oxide carbon substituents substrate, the strain on the enantioselectivity greatly reduced. Under the optimal bioconversion conditions, enantiopure glycidyl azide was separated, and ultimately synthesis the linezolid.