Abstract
The ginsenoside Rg2(S), which is one of the pharmaceutical components of ginseng, is known to have neuroprotective, anti-inflammation, and anti-diabetic effects. However, the usage of ginsenoside Rg2(S) is restricted owing to the small amounts found in white and red ginseng. To enhance the production of ginsenoside Rg2(S) as a 100 gram unit with high specificity, yield, and purity, an enzymatic bioconversion method was developed to adopt the recombinant glycoside hydrolase (BglPC28), which is a ginsenoside-transforming recombinant β-glucosidase from Pseudonocardia sp. strain Gsoil 1536. The gene, termed bglPC28, encoding β-glucosidase (BglPC28) belonging to the glycoside hydrolase family 3 was cloned. bglPC28 consists of 2,232 bp (743 amino acid residues) with a predicted molecular mass of 78,975 Da. This enzyme was overexpressed in Escherichia coli BL21(DE3) using a GST-fused pGEX 4T-1 vector system. The optimum conditions of the recombinant BglPC28 were pH 7.0 and 37°C. BglPC28 can effectively transform the ginsenoside Re to Rg2(S); the K m values of PNPG and Re were 6.36±1.10 and 1.42±0.13 mM, respectively, and the V max values were 40.0±2.55 and 5.62±0.21 µmol min−1 mg−1 of protein, respectively. A scaled-up biotransformation reaction was performed in a 10 L jar fermenter at pH 7.0 and 30°C for 12 hours with a concentration of 20 mg/ml of ginsenoside Re from American ginseng roots. Finally, 113 g of Rg2(S) was produced from 150 g of Re with 84.0±1.1% chromatographic purity. These results suggest that this enzymatic method could be usefully exploited in the preparation of ginsenoside Rg2(S) in the cosmetics, functional food, and pharmaceutical industries.
Highlights
Ginseng, an important herbal medicine, has been widely used for thousands of years in East Asia and has been popularized in the West during the past decades [1,2]
The ginseng root consists of ginsenosides, polysaccharides, peptide, polyacetylenic alcohols, and fatty acids [8]
For the production of the recombinant BglPC28, the LB medium supplemented with ampicillin (100 mg/ml) was used to cultivate the E. coli harboring pGEX-bglPC28 in a 10 L stirredtank reactor (BiotronGX, Hanil science Co., Korea) with a 5 L working volume at 500 rpm
Summary
An important herbal medicine, has been widely used for thousands of years in East Asia and has been popularized in the West during the past decades [1,2]. Ginsenosides are the major active components of ginseng, and they appear to be responsible for the principle pharmacological activities of ginseng, including vasorelaxation and anti-neoplastic, anti-diabetic, antiinflammation, and anti-oxidant effects [9,10,11]. The PPD- and PPT-type ginsenosides are further classified into subgroups based on the position and number of sugar moieties attached to the aglycon at positions C3 or C6 and C20. One of the largest PPD-type ginsenosides, Rb1, contains 4 glucose moieties, with two attached via glycosidic linkages to the C3 and C20 positions, respectively, of the aglycon (Fig. 1). They may be classified as major or minor ginsenosides based on the amount found in cultivated ginseng
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