| L-ascorbic acid (vitamin C, VC) has important physiological activities in body, but the application is limited because of its instability. Some derivatives of VC have largely improved stability while still keep their physiological activities. 2-0-glucopyranosyl-L-ascorbic acid (AA-2G), one of its derivatives, was studied in this text. The main content is about its synthesis and separation of AA-2G.The culture medium component and culture condition for producing saccharide-transferring enzyme by B. Stearothermophilis was primarily established by Single Factor design. That is Glucose 1%, Peptone 0.5%, medium volumes 30ml in 250 ml shake flask, inoculum size 4%, pH8.0 and temperature 55℃.To optimize the transformation condition, the Orthogonal Test design was used. The best condition is using CMC-Na as saccharide offer, 0.04mol/L VC as acceptor, the enzyme volume 25ml, pH4.0, reacting under 60℃ for 16hr.A high yield strain that produced this enzyme was obtained by induced-mutation. Its transformation ability increased 149.28%. And the content of AA-2G reached 274.78mg/ml.Systematic studies were carried out through Plackett-Burman design and response surface analysis (RSA). The optimum condition for producing saccharide-transferring enzyme by B. Stearothermophilis was established by the software of Origin. The result showed that the optimum condition is Glucose 2%, Peptone 0.6%, MgSO4 7H2O 0.01%, KH2PO4 0.0625%, the initial pH 7.15, culturing 48hr under 48 ℃, rotate speed 200r/min. The enzyme produced under this condition transformed VC to 375.24mg/l AA-2G, which was 41.16% higher than that of non-optimized medium.The 3.7-liter scale batch bioreactor test was carried out to improve the enzyme activity, which came to 419.40mg/L after transformation. Also the elementary separation and purification was done. The preferable result was attained by using the resin A500P and the eluent of 0.3mol/L hydrochloric acid.
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