The Study Of Bioconversion And Purification Of The Nucleoside Intermediate

2’-deoxyadenosine is a natural nucleoside, it is both important raw materials for biochemical drug and genetic engineering, but also the intermediate of anti-tumor and antiviral nucleoside drugs (cladribine, etc.), therefore, there is extensive demand in the market. Chemical synthesis of 2’-deoxyadenosine need some groups protection、 deprotection and the activation of sugar, the steps are very complex, and need to split the pair of enantiomers; compared with the traditional chemical synthesis, microbiotransformation is high selectivity、 mild conditions、 low cost and pollution, so the research of the microbial transformation of dA is very meaningful.In this paper, a high-yield strain producting 2’-deoxyadenosine were got Brevibacterium acetylium HE-4. Firstly, The optimization of fermentation conditions for HE-4 were carried out. and the optimal conditions were:glucose 0.2%. corn steep liquor 3.0%. ammonium chloride 0.5%, disodium hydrogen phosphate 0.3%, magnesium sulfate 0.075%, sodium chloride 0.4%; the original fermentation pH is7.2; fermentation temperature is 37℃ strain age is16h. inoculation quantity is 3%, volume is 100mL/500mL, the rate is 210r/min and fermentation time is 24h, the yield of bacteria was doubled.10L magnified fermentation was carried out after optimizing the fermentation conditions. The result was that:temperature 33 ℃,constant pH 6.9, moderate ventilatory capacity 7.5L/min, strain growth is better in the fermenter.After got the high-yield and stable strains, conversion conditions were optimized:the best conversion conditions were:temperature 58℃, strain concentration 6%, pH 7.2 phosphate concentration 30mmol/L, convertion time 8h; substrate ratio(A/dT) 125%, concentration of dT 72.8..ol/L,stir rate 60r/min. adding MnSO40.003%.Finally, the product purification was discussed, and purification process is as follows: firstly, after the initial disposal, the liquid was through D101 resin column to separate the remaining dA, the optimized conditions were as follow:ample flow rate was 2.0mL/min,eluting with 4.5BV 0.1mol/L Na2CO3、3BV 30% ethanol orderly, flow rate was 1.5mL/min,the purity of eluting solution is 73.5%,yield is 93.9%, then crystallize and recrystallize the eluting solution, the purity of product was 99% by HPLC, the purification yield is 82％.