Monday, August 12, 2013
Pavilion (Sheraton San Diego)
Puerarin, an isoflavone from Puerarin thomsonii, is used to treat cardio-cerebrovascular disease. However, the bioefficacy of puerarin is limited by its low hydrophilicity. We previously reported that transformation of puerarin to puerarin-7-O-glucoside and purarin-7-O-fructoside by Microbacterium oxydans (M. oxydans) CGMCC 1788 increases its water solubility. Here, we further report that transformation efficiency of puerarin is improved greatly by immobilized glycosidase from M. oxydans CGMCC 1788. Immobilization of enzyme was conducted by incubation of enzyme with DEAE-52 cellulose at 6 mg: 0.5 g ratio for 6 hours at 4oC and pH 6.5, which enzyme activity is 2.4 units /g DEAE-52 cellulose and has 90.4% recovery yield. For biotransformation, the optimal substrate concentration was 2 mg/mL; the optimal pH was 6.0, and the optimal temperature was 30oC - 40oC. Under such optimal conditions, 71.8% of puerarin was transformed to puerarin-7-O-glucoside by immobilized glycosidase after 12 hours of incubation. Transformation efficiency of puerarin by immobilized enzyme is 7.6 fold greater than that by cells, while producing only minimal amounts of by-products. Furthermore, both thermal and pH stability of the immobilized enzyme are significantly improved compared to free enzyme. And the immobilized enzyme also showed better handling and storage stability as it retained 80% and 48% of the enzymatic activity after being reused 5 and 10 times, respectively, and retained 50% of its activity after storage at 4oC for 45 days. Thus, this research offers a more efficient means by which to improve the biotransformation of puerarin.