| The paper mainly studied the microorganism from panax plantation of Changbaishan China which can produce ginsenoside Rb1-hydrolase,the ferment condition of the bacteria,the purification and characterization of hydrolase.Also focused on the enzyme character and specifieness.From the investigation,it confirmed the composing of culture medium is proteose peptone 10g,yeast powder 5g,NaCl 10g and water 1L.Add to 3.2mg total saponins of Panax ginseng per milliliter culture medium as revulsant.The temperature of culture is 28?.The thalli addition is 250?L per 20mL culture medium.And the best enzyme production time is 20 to 25h.The ginsenoside Rb1-hydrolase was purified using DEAE-cellulose column,the pure enzyme showed a single protein band by SDS-PAGE,and the molecular weight was estimated to be about 47KDa.From the enzyme character's investigation,it confirmed the optimum pH of enzyme reaction is 4,and stable in pH2.2 to pH10;the optimum temperature is 25?,and stable in 20??60?.Fe3+ has active influence on the reaction,the enzyme reaction was markedly inhibited by Cu2+,and other ions have no affection.The equation of enzyme reaction is V=0.0423[S]/(1.045+[S]),Km and Vmax values is 1.045mmol/Land 0.0423mmol/L·h.From the enzyme specifieness's investigation,it confirmed the ginsenoside Rb1-hydrolase can hydrolyze Protopanaxadiol Type Ginsenosides and icariin,hydrolyze the glucoses on C20 of Protopanaxadiol Type Ginsenosides one by one,and the ginsenoside Rb1 is hydrolased from ginsenoside Rd to ginsenoside Rg3.The enzyme can't hydrolyze?-amylase?pNP-?-D-Gal?pNP-P-D-Glc?pNP-?-D-Glc and pNPR except pNP-?-D-Gal,so the ginsenoside Rb1-hydrolase was a special hydrolase. |
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