Specific and efficient hydrolysis of all outer glucosyls in protopanaxadiol type and protopanaxatriol type ginsenosides by a β-glucosidase from Thermoclostridium stercorarium

To develop a new method for enzymatic preparation of minor ginsenosides , T. stercorarium β-glucosidase (Tsbgl) was characterized and its activities of deglycosylation towards natural ginsenosides were examined. The substrates of 1 mmol l −1 were incubated with the enzyme of 38.3 U ml −1 at 65 ℃ and pH 5.0. The K m values of Tsbgl for ginsenosides Rb 1 , Rg 1 and p NPG were 0.37 ± 0.03, 3.26 ± 0.19, and 1.24 ± 0.03 mmol l −1 , and the V max values were 183.63 ± 7.15, 85.03 ± 4.90, and 117.66 ± 1.96 μmol mg −1 min −1 , respectively. The molar conversion of ginsenosides Rb 1 , Rb 2 , Rb 3 , Rc, Re, Rg 1 , and Rf by Tsbgl within 6 h was 100%, 50.1%, 42.7%, 92.0%, 57.3%, 67.9%, and 76.8%, respectively. The yield of aglycone protopanaxadiol was 35.5 μmol l −1 h −1 for Rb 1 , while the yields of aglycone protopanaxatriol were 64.2 and 70.4 μmol l −1 h −1 for Rg 1 and Rf. Tsbgl with good organic solvent tolerance, mild reaction conditions and broad substrate specificity , could completely remove all outer glucosyls at the C-3 and C-20 hydroxyls of protopanaxadiol-type ginsenosides, and the C-6 and C-20 hydroxyls of protopanaxatriol-type ginsenosides through various pathways, providing a specific and efficient way to produce minor ginsenosides.