Production and characterization of ginsenoside-β-glucosidase from bacteria GS0202
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Abstract:
Production of the ginsenoside-β-glucosidasebybacteria GS0202 was optimized.The medium composition is proteose peptone 10 g,yeast powder 5 g,NaCl 10 g.3.2 mg/mL total saponins of Panax ginseng and water 1 L and culture temperature is 28 ℃.The opimum temperature for enzyme catalysis is 25 ℃,pH 4 and the concentration of substrate is 3 mg/mL.Keywords:
Yeast extract
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Bioconversion
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In this paper,the condition for ginsenosidase production and reaction from Arthrobacter sp.No.3 GS0202 strain was examined by TLC and HPLC.The ginsenosidase was produced in the medium containing 1% triptone,0.5% yeast extract and 1% NaCl at 30 ℃,using total ginsenoside as inducer.The optimal conditions for hydrolysis of ginsenoside Rg1 were at 30 ℃ for 24 h and the concentration of substrate was 0.1 mg/mL.The enzyme is able to hydrolyze the 20-O-b-D-glucoside of the protopanaxatriol ginsenoside Rg1 to the ginsenoside Rh1,and the yield of ginsenoside Rh1 was approximately 20% detected by HPLC.
Arthrobacter
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Ginseng (Panax ginseng) is frequently used in Asian countries as a traditional medicine. The major components of ginseng are ginsenosides. Among these, ginsenoside compound K has been reported to prevent the formation of malignancy and metastasis of cancer by blocking the formation of tumor and suppressing the invasion of cancer cells. In this study, ginsenoside $Rb_1$ was converted into compound K, via secreted ${\beta}$ -glucosidase enzyme from the Leuconostoc lactis DC201 isolated, which was extracted from Kimchi. The strain DC201 was suspended and cultured in MRS broth at $37^{\circ}C$ . Subsequently, the residue from the cultured broth supernatant was precipitated with EtOH and then dissolved in 20 mM sodium phosphate buffer (pH 6.0) to obtain an enzyme liquid. Meanwhile, the crude enzyme solution was mixed with ginsenoside $Rb_1$ at a ratio of 1:4 (v/v).The reaction was carried out at $30^{\circ}C$ and 190 rpm for 72 hours, and then analyzed by TLC and HPLC. The result showed that ginsenoside Rb1 was transformed into compound K after 72 hours post reaction.
Bioconversion
Beta-glucosidase
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The ginsenosidase was produced by Phycicoccus sp.BXN5-13 for transformation of Rb1 and other protopanaxadiol saponin to the ginsenoside Rd and Rg3.The ginsenosidase was an intracellular enzyme that produced maximally when cultured in the medium containing 10 g/L triptone,5 g/L yeast extract and 10 g/L NaCl and ginsenoside at 35 ℃ for 72 h.The optimal condition for enzymatic hydrolysis of ginsenoside was as pH 6.0 at 45 ℃ for 24 h.The transformation rate of ginsenoside-Rg3 was 44%.
Protopanaxadiol
Enzymatic Hydrolysis
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Microbial conversion of ginsenoside Rd from Rb1 by the fungus mutant Aspergillus niger strain TH-10a
Ginsenoside Rd, one of the ginsenosides with significant pharmaceutical activities, is getting more and more attractions on its biotransformation. In this study, a novel fungus mutant, the Aspergillus niger strain TH-10a, which can efficiently convert ginsenoside Rd from Rb1, was obtained through screening survival library of LiCl and ultraviolet (UV) irradiation. The transformation product ginsenoside Rd, generated by removing the outer glucose residue from the position C20 of ginsenoside Rb1, was identified through high-performance liquid chromatography (HPLC) analysis. Factors for the microbial culture and biotransformation were investigated in terms of the carbon sources, the nitrogen sources, pH values, and temperatures. This showed that maximum mycelia growth could be obtained at 28°C and pH 6.0 with cellobiose and tryptone as the carbon source and the nitrogen source, respectively. The highest transformation rate (∼86%) has been achieved at 32°C and pH 5.0 with the feeding time of substrate 48 hr. Also, Aspergillus niger strain TH-10a could tolerate even 40 mg/mL ginseng root extract as substrate with 60% bioconversion rate after 72 hr of treatment at the optimal condition. Our results highlight a novel ginsenoside Rd transformation fungus and illuminate its potentially practical application in the pharmaceutical industries.
Biotransformation
Aspergillus niger
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The study was conducted to develop an edible and low cost growth medium for cultivation of Weissella hellenica DC06, a lactic acid bacteria (LAB) and to study whether, the medium is suitable for bioconversion of major ginsenoside Rb1 into ginsenoside Rg3 through fermentation by W. hellenica DC06. Fourteen different media compositions were investigated to cultivate W. hellenica DC06. Among these, W. hellenica DC06 exhibited the highest growth in media containing 20 g/l radish, 20 g/l glucose, and 10 g/l yeast extract (Medium 3). The optical density of W. hellenica DC06 cultivated in medium 3 reached 1.8 (1.066 x 1010 CFU/ml) after 24 h of incubation. Importantly, the optimized medium was approximately four times cheaper compared to MRS medium. In addition to being economical, the new medium was also edible. Also W. hellenica DC06 showed strong fermentation ability in newly developed medium regarding on major ginsenoside Rb1 biotransformation. Ginsenoside Rb1 was converted into pharmacologically active ginsenoside Rg3 in new medium. In contrast,W. hellenica DC06 showed weak fermentation ability in MRS medium where ginsenoside Rb1 was converted intoginsenoside Rd. The transformation products were analyzed by TLC, and HPLC. Within seven days of fermentation, almost all ginsenoside Rb1 was decomposed and converted into Rg3 in optimized medium. W. hellenica DC06 hydrolyzed two glucose moieties attached to the C-20 position of the ginsenoside Rb1aglyconeand synthesized Rg3 in newly developed medium.Bangladesh J. Sci. Ind. Res. 51(4), 271-278, 2016
Biotransformation
Bioconversion
Yeast extract
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The ginsenosidase is found from a newly screened strain, Absida sp.F7. The ginsenosidase can hydrolyze 20C βGlucoside of ginsenoside Rb1 to produce ginsenoside Rd. The optimal condition for enzyme reaction is as follows: pH, 5.0;temperature, 40?℃;time,24?h; substrate concentration, 5?mg/mL.
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Protoplast
Biotransformation
Strain (injury)
Mutation Breeding
Osmotic pressure
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Ginsenoside-Rg3 is a ginsenoside monomer with high anti-cancer activity,but its content in ginseng is extremly low.The extracellular enzyme produced by new screened bacteria Arthrobacter sp.3 can transform PPD to ginsenoside Rg3.TLC analysis showed that the best inducer is PPD of 0.1 mg/mL.The optimal conditions were 40 ℃,24 h,and the substrate concentration was 0.1 mg/mL.In this condition,the content of Rg3 can reach 21% through the detection of HPLC.
Arthrobacter
Exoenzyme
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This study was conducted to isolate and characterize Paenibacillus sp. MBT213 possessing β-glucosidase activity from raw milk, and examine the enzymatic capacity on the hydrolysis of a major ginsenoside (Rb1). Strain MBT213 was found to have a high hydrolytic ability on ginsenoside Rb1 by Esculin Iron Agar test. 16S rDNA analysis revealed that MBT213 was Paenibacillu sp. Crude enzyme of MBT213 strain exhibited high conversion capacity on ginsenoside Rb1 into ginsenoside Rd proven by TLC and HPLC analyses. The API ZYM kit confirmed that Paenibacillu sp. MBT213 exerted higher β-glucosidase and β-galactosidase activity than other strains. Optimum pH and temperature for crude enzyme were found at 7.0 and 35°C in hydrolysis of ginsenoside Rb1. After 10 d of optimal reaction conditions for the crude enzyme, ginsenoside Rb1 fully converted to ginsenoside Rd. Ginseng roots (20%) were fermented for 14 d, and analyzed by HPLC showed that amount of ginsenoside Rb1 significantly decreased, while that of ginsenoside Rd was significantly increased. The study confirmed that the β-glucosidase produced by Paenibacillus sp. MBT213 can hydrolyze the major ginsenoside Rb1 and convert to Rd during fermentation of the ginseng. The β-glucosidase activity of this novel Paenibacillus sp. MBT213 strain may be utilized in development of variety of health foods, dairy foods and pharmaceutical products.
Beta-glucosidase
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