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Preparation method for increasing CK (Compound K) yield of ginsenoside

A ginsenoside and production technology, applied in the field of biomedicine, can solve the problems of unfavorable growth of microbial cells, long production cycle, low substrate concentration, etc., and achieve the effect of improving cell permeability, increasing substrate concentration and yield of transformants

Inactive Publication Date: 2014-01-15
FUDAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The previous research of the present invention (our laboratory) has isolated and screened the highly efficient transformation strain Paecilomyces bainiersp.229 (authorized patent number: ZL20041008000.0) from the soil of wild ginseng, and selected cheap and easy-to-obtain diol-type Panax notoginseng stems and leaves Total saponin is used as the substrate, and the conversion rate has reached 72.9% through optimization of fermentation conditions. However, there are still problems of low substrate concentration and long production cycle. Because excessive saponin is not good for microbial cell growth, the substrate feeding concentration Only 0.5%

Method used

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  • Preparation method for increasing CK (Compound K) yield of ginsenoside
  • Preparation method for increasing CK (Compound K) yield of ginsenoside
  • Preparation method for increasing CK (Compound K) yield of ginsenoside

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Effect test

Embodiment 1

[0031] The mutagenesis of embodiment 1 parent strain

[0032] Strains: The ginsenoside-transforming bacteria is Paecilomyces Bainiersp.229, and the strains with good growth and high CK conversion rate are selected as parent strains.

[0033] Medium: Potato medium (20% potato, 2% glucose made into soup), divided into 10×150mm test tubes, 4ml per tube, added 30mg / ml (3%) notoginseng stem and leaf saponins and different concentrations of LiCl , 121°C, sterilized for 20min.

[0034] Mutation: The rejuvenated fresh slant was washed with 0.9% physiological saline to make 10 7 The spore suspension of 1 spore / ml is transferred to a flat plate with a diameter of 9 cm, and a paper clip is placed in the flat plate as a stirrer. Irradiate under ultraviolet light for 0, 2min, 4min, 6min and 8min respectively, take 0.1ml of the spore suspension after ultraviolet irradiation and add it to the above-mentioned sterilized medium, cultivate at 28°C, observe and record the growth of bacteria. ...

Embodiment 2

[0038] The determination of embodiment 2 substrate concentration

[0039] After the mutagenized strains were cultured on a shaking table at 28°C and 200r / min for 48 hours, substrates with a concentration of 0.5%, 1%, 1.5%, 2.0%, and 2.5% were added respectively, and the transformation was continued for 72 hours, and the concentration of CK in the bacteria was detected. content. The result is as figure 1 shown.

Embodiment 3

[0040] The selection of embodiment 3 surfactants

[0041] Since the continuous accumulation of the transformation end product in the microorganism will cause feedback inhibition and also cause certain toxicity to the bacteria, one of the solutions can be to improve the permeability of the cell membrane by adding an active agent to promote the transfer of the substrate into the cell and the transfer of the product to the cell. Extracellular transfer, thereby increasing productivity. Select Tween40, Tween80, Tween20, SDS, Span60 and Triton X-100 and other low toxicity surfactants to investigate the improvement of cell membrane permeability. Finally, the optimal surfactant Tween80 was selected.

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Abstract

The invention belongs to the technical field of medical technology, relates to a preparation method for increasing CK (Compound K) yield of ginsenoside, and in particular relates to a preparation method for increasing the CK yield of ginsenoside by increasing the yield of a converted product through adjustment and control in a microbial conversion process. According to the preparation method, the yield of the converted product and the concentration of a primer are improved through metabolic regulation. The preparation method comprises the steps of: reconstructing ultraviolet mutagenesis of a parental strain, culturing in a high-concentration pseudo-ginseng stem and leave saponin culture medium, sieving in a directional manner to obtain a strain resisting to 1-5% of the total saponins of the pseudo-ginseng stems and leaves. Through the strain mutagenesis and the metabolic regulation, the concentration of the primer and the yield of the converted product are improved obviously, the final conversion rate is up to 43.63%, and the yield of the Compound K of the ginsenoside is improved by more than 80%. The preparation method for increasing CK yield of ginsenoside has practical significance for the industrial application of the microbial conversion.

Description

Technical field: [0001] The invention belongs to the technical field of biomedicine. The invention relates to a preparation method for increasing the yield of ginsenoside CK, and specifically relates to a preparation method for increasing the yield of ginsenoside CK through regulation and control in the transformation and metabolism process of microorganisms to increase the yield of transformation products. Background technique: [0002] my country's traditional precious Chinese herbal medicine ginseng has a long history of medicinal use. Modern medical research shows that ginsenosides isolated from ginseng are the main medicinal components of ginseng, which have a wide range of pharmacological activities and have preventive effects on various diseases. Studies have shown that natural ginsenosides contain higher content of Rb1, Rg1, Re, etc., while less Rd, Rh1, Rh2, etc., ginsenoside Compound K (CK) does not exist in nature, it is the result of ginsenoside transformation i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P33/00C12N15/01C12R1/79
Inventor 冯美卿张春燕周珮周伟叶丽史训龙周超群
Owner FUDAN UNIV
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