Study On Biotransformation Of Ginsenoside Rb1 By Immobilized ?-glucosidase

Significant physiological activity in antibacterial,anti-inflammatory,blood pressure reduction,blood lipid reduction,immune enhancement,and central nervous system improvement,in particular,the activity of rare ginsenosides in anti-tumor is better than that of major ginsenosides.High efficiency and good selectivity in the preparation of rare ginsenosides by enzymatic transformation of major ginsenosides,and the immobilized enzyme technology can solve the problem that free enzyme is difficult to be recycled,after immobilization,the enzyme has more advantages,such as enhanced stability,higher selectivity,high-temperature resistance,acid,and alkali resistance.In this paper,?-glucosidase was immobilized on amino chromatography stationary phase and functional magnetic Fe₃O₄ powder as the carrier to transform ginsenoside Rb1.A hydrophilic interaction chromatography was established for the rapid and simultaneous determination of 14 ginsenosides.Using UG80-CAPCELL PAK SCX?4.6 mm I.D.×250 mm,5 ?m?column as stationary phase,ginsenoside Rb1,Rb2,Rb3,Rc,Rd,Rf,Re,Rg1,Rg2,Rg3,Rh1,Rh2,F2,C-K can be separated quickly and well.The method has high accuracy and excellent reproducibility.The results of the HILIC model showed that the molecular weight,polarity,and capacity factors of ginsenoside were positively correlated.The transformation pathway of ginsenoside Rb1 was studied by three ?-glucosidase extracted from Aspergillus niger,almond and our laboratory,the ?-glucosidase was immobilized on amino silica gel as carrier to transform ginsenoside Rb1.The optimum reaction temperature,p H value,thermal stability,acid and alkaline resistance and stability of enzyme were studied.The results showed that the optimal p H of free enzyme and immobilized enzyme was the same,both were 5.5,the optimal reaction temperature of free enzyme was 40 ?,and the optimal reaction temperature of immobilized enzyme was 45 ?;Before and after enzyme immobilization,the Michaelis constant km was 0.503 mmol·L^-1 and 0.360 mmol·L^-1,respectively.The activity of the immobilized enzyme was 56.4% after repeated use 40 times.The magnetic Fe₃O₄ powder was used as raw material,modified by3-aminopropyltriethoxysilane,and the material was characterized.?-glucosidase was immobilized by the biotin-streptavidin method.The optimum reaction temperature,p H value,thermal stability,acid,and alkali resistance,and stability of enzyme werestudied.The results showed that the optimal temperature of free enzyme was the same as that of immobilized enzyme,which was 40 ?,the optimal p H of free enzyme was5.5,and the optimal p H of immobilized enzyme was 6.0;the Michaelis constant km before and after enzyme immobilization was 0.503 mmol·L^-1,0.064 mmol·L^-1,respectively.After 25 times of repeated use,the enzyme activity was 50.9%.The stability of enzyme immobilized by a biotin-streptavidin method is greatly improved,and it has a broad prospect in practical application.