Journal of Engineering Research

The enzyme-induced transformation method was used, with vitamin C (VC) and glucose donor as the substrate, and recombinant E.coli BL21(DE3)-pET30a-cgt1 as the enzyme-producing strain. The enzyme-producing conditions and the synthesis conditions of 2-O-α-D-glucopylanose-L-ascorbic acid (AA-2G) were investigated. The results showed that the reaction product was determined to contain AA-2G by HPLC analysis. The induction temperature was 25 ℃, the fermentation medium was TB liquid medium, the induction time was 8 h, the inoculation amount was 4 %, the IPTG induction concentration was 50 μM, the cell density of the recombinant strain was 1.2-1.9, the initial pH value of the medium was natural pH value, the enzyme expression of the recombinant strain was the highest and the terminal bacterial cell density was the highest. With β-cyclodextrin and VC as substrates, the yield of AA-2G was 2.21 g/L and the conversion of VC was 23.02 % under the initial reaction conditions. By optimizing the transformation reaction conditions, the optimum conditions for the transformation reaction were preliminarily determined: The pH value of the conversion reaction was 5.0, the conversion reaction time was 48 h, the conversion reaction temperature was 30 ℃, the conversion reaction solvent was pure water, v(crude enzyme solution):v(total transformation reaction volume) = 1:5, m(VC):m(β-cyclodextrin)=1:2, and the addition amount of glucose amylase was 50 U/mL. When the hydrolysis time of glucose amylase was 16 h, the yield of AA-2G reached 68.61 g/L and the conversion rate of VC was 71.45 %.

α-cyclodextrin glucosyltransferase(α-CGTase); 2-O-alpha-D-glucopyranosyl-L-ascorbic acid(AA-2G); Enzymatic synthesis