Abstract
Ginsenosides are the principal compounds responsible for the pharmacological effects and health benefits of Panax ginseng root. Among protopanaxadiol (PPD)-type ginsenosides, minor ginsenosides such as ginsenoside (G)-F2, G-Rh2, compound (C)-Mc1, C-Mc, C-O, C-Y, and C-K are known to be more pharmacologically active constituents than major ginsenosides such as G-Rb1, G-Rb2, G-Rc, and G-Rd. A novel ginsenoside Rc-hydrolyzing β-glucosidase (BG-1) from Armillaria mellea mycelia was purified as a single protein band with molecular weight of 121.5 kDa on SDS-PAGE and a specific activity of 17.9 U mg−1 protein. BG-1 concurrently hydrolyzed α-(1 → 6)-arabinofuranosidic linkage at the C-20 site or outer β-(1 → 2)-glucosidic linkage at the C-3 site of G-Rc to produce G-Rd and C-Mc1, respectively. The enzyme also hydrolyzed outer and inner glucosidic linkages at the C-3 site of G-Rd to produce C-K via G-F2, and inner glucosidic linkage at the C-3 site of C-Mc1 to produce C-Mc. C-Mc was also slowly hydrolyzed α-(1 → 6)-arabinofuranosidic linkage at the C-20 site to produce C-K with reaction time prolongation. Finally, the pathways for formation of C-Mc and C-K from G-Rc by BG-1 were G-Rc → C-Mc1 → C-Mc and G-Rc → G-Rd → G-F2 → C-K, respectively. The optimum reaction conditions for C-Mc and C-K formation from G-Rc by BG-1 were pH 4.0–4.5, temperature 45–60 °C, and reaction time 72–96 h. This is the first report of efficient production of minor ginsenosides, C-Mc and C-K from G-Rc by β-glucosidase purified from A. mellea mycelia.
Highlights
The result showed that C-Mc and C-K were efficiently produced from G-Rc by crude enzyme preparations from A. mellea mycelia, whereas crude enzyme preparations from G. lucidum, P. baumii, G. applanata, and P. ostreatus produced of G-Rd as a final product (Fig. 2)
The results indicate that A. mellea β-glucosidases exist in three isomeric forms that they exhibited different retention behaviors on DEAE cellulose column and different hydrolytic activity toward pNP-β-d-glucopyranoside
In the screening for mushroom mycelia containing G-Rchydrolyzing activity, we found that enzyme preparation from A. mellea mycelia could efficiently convert G-Rc to C-Mc and C-K, whereas enzyme preparations from G. lucidum, P. baumii, G. applanata, and P. ostreatus produced of G-Rd as a final product
Summary
A. Meyer, has been used as a traditional folk medicine in East Asian countries such as Korea, Japan and China for thousands of years, and has to some extent been popularized in many western countries during recent decades. The major pharmacologically active constituents of ginseng are triterpenoid saponins called ginsenosides. They can be classified into two groups by the skeleton of their aglycones, dammarane-type and oleanane-type. The dammaranetype ginsenosides can be classified into protopanaxadiol (PPD)-type and protopanaxatriol (PPT)-type
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