Biotransformation of Protopanaxadiol-Type Ginsenosides in Korean Ginseng Extract into Food-Available Compound K by an Extracellular Enzyme from Aspergillus niger

Abstract

Compound K (C-K) is one of the most pharmaceutically effective ginsenosides, but it is absent in natural ginseng. However, C-K can be obtained through the hydrolysis of protopanaxadiol-type ginsenosides (PPDGs) in natural ginseng. The aim of this study was to obtain the high concentration of food-available C-K using PPDGs in Korean ginseng extract by an extracellular enzyme from Aspergillus niger KACC 46495. A. niger was cultivated in the culture medium containing the inducer carboxymethyl cellulose (CMC) for 6 days. The extracellular enzyme extracted from A. niger was prepared from the culture broth by filtration, ammonium sulfate, and dialysis. The extracellular enzyme was used for C-K production using PPDGs. The glycoside-hydrolyzing pathways for converting PPDGs into C-K by the extracellular enzyme were Rb1 → Rd → F2 → C-K, Rb2 → Rd or compound O → F2 or compound Y → C-K, and Rc → Rd or compound Mc1 → F2 or compound Mc → C-K. The extracellular enzyme from A. niger at 8.0 mg/ml, which was obtained by the induction of CMC during the cultivation, converted 6.0 mg/ml (5.6 mM) PPDGs in Korean ginseng extract into 2.8 mg/ml (4.5 mM) food-available C-K in 9 h, with a productivity of 313 mg/l/h and a molar conversion of 80%. To the best of our knowledge, the productivity and concentration of C-K of the extracellular enzyme are the highest among those by crude enzymes from wild-type microorganisms.

Keywords: Aspergillus niger; Panax ginseng; biotransformation; compound K; protopanaxadiol ginsenosides.

Fig. 1. Effect of inducer added during cultivation on the hydrolytic activity for ginsenosides (A) Rb1, (B) Rb2, and (C) Rc.

The inducer polysaccharide at 20 g/l was added during cultivation. Data are expressed as the means of three experiments and the error bars represent standard deviations.

Fig. 2. Biotransformation of ginsenosides (A) Rb1, (B) Rb2, and (C) Rc into compound K by extracellular enzyme from A. niger.

The extracellular enzyme from A. niger converted 1.0 mg/ml of ginsenoside Rb1, Rb2, and Rc into C-K with molar conversions of 100%, 94%, and 100%, respectively. Data are expressed as the means of three experiments and the error bars represent standard deviations.

Fig. 3. Proposed biotransformation pathways of protopanaxadiol-type ginsenosides into compound K by extracellular enzyme from Aspergillus niger.

Glc, glucose, Arap, arabinopyranose, and Araf, arabinofuranose.

Fig. 4. Biotransformation of protopanaxadiol-type ginsenosides (PPDGs) in protopanaxadiol-type ginsenoside mixture from Korean ginseng (PPDKG) into compound K by extracellular enzyme from A. niger.

(A) Time-course reactions of protopanaxadiol-type ginsenosides into compound K. (B) HPLC profiles before and after the reaction. ●, Ginsenoside Rb1; ○, ginsenoside Rc; ▼ , ginsenoside Rb2; △, ginsenoside Rd; ■ , compound O; □ , ginsenoside F2; ◆, compound Y; ◇, compound K. Data are expressed as the means of three experiments and the error bars represent standard deviations.