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Method for preparing ginsenoside Rd by using microbial transformation

A technology of microbial transformation and ginsenoside, applied in the field of fermentation, can solve the problems of large amount of enzyme, high cost of substrate, non-specific and obvious inhibition of transformation products, etc., and achieves the effect of high transformation rate and expansion of drug resource sources.

Inactive Publication Date: 2010-03-24
SHANGHAI HAITAI PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are only research reports on the conversion of ginsenoside Rd by enzymes derived from microorganisms. In these studies, the conversion products are not specific and the enzymes are significantly inhibited by ginsenosides in the conversion process, and the conversion substrates are all high-purity Rb1. As a result, these conversion methods require a large amount of enzymes, and the cost of substrates is high, which greatly limits industrial production.
[0006] Due to the difficulty in the application of the enzyme conversion method, the preparation of Rd monomer by the microbial conversion method has more research value and application prospects, but so far there has been no report on the specific preparation of ginsenoside Rd by microbial fermentation

Method used

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  • Method for preparing ginsenoside Rd by using microbial transformation
  • Method for preparing ginsenoside Rd by using microbial transformation
  • Method for preparing ginsenoside Rd by using microbial transformation

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0031] To the tolerance test of notoginseng stem and leaf saponins of the original bacteria of example 1

[0032] Strain: Paecilomyces sp.4685, the screened strain grows well and is used as the starting strain.

[0033] Culture medium: potato culture medium (20% of potato, 2% of glucose is mixed with soup liquid), sub-package in 10 * 150cm test tube, every tube 4ml, add Panax notoginseng stem and leaf saponin according to Table 1, plug the test tube mouth with silica gel stopper, Sterilize at 121°C for 20 minutes.

[0034] Inoculation: the rejuvenated fresh slant was washed with 0.9% saline to prepare 10 7 spores / ml of the spore suspension, add 0.1ml of the spore suspension to each test tube equipped with medium, cultivate for 6 days at 28°C, and observe the growth of the bacteria (Table 1).

[0035] Table 1 Bacterial growth in different concentrations of notoginseng stem and leaf saponins liquid medium

[0036]

[0037] +++When no saponin is added, the bacteria grow ver...

example 2

[0039] Example 2 Substrate-induced screening of high substrate tolerance, high specificity of the production strain Paecilomyces --Paecilomyces sp.4685

[0040] Strain: Paecilomyces Bainier sp.4685 strain with good growth condition was selected as the starting strain.

[0041] Culture medium: potato culture medium (20% of potato, 2% of glucose is mixed with soup liquid), sub-package in 10 * 150cm test tube, every tube 4ml, add different amounts of Panax notoginseng stem and leaf saponin respectively, make final concentration be 30, 35, 40, 45, 50, 55, 60mg / ml, etc., plug the mouth of the test tube with a silica gel stopper, and sterilize at 121°C for 20 minutes.

[0042] Induction: It can be seen from the tolerance experiment of strains that high concentration of saponins can inhibit the growth of bacteria, therefore, the concentration of saponins in stems and leaves of notoginseng should be continuously increased to select resistant strains.

[0043] Take 0.1ml of the bacter...

example 3

[0044] The optimization of example 3 transformation medium

[0045] Seed medium: 3% sucrose, 3% soybean flour, 0.1% ammonium sulfate, 0.1% magnesium sulfate, 0.1% calcium chloride, 0.5% bran.

[0046] Fermentation medium: 3% starch, 3% cottonseed flour, 0.1% sodium nitrate, 0.1% magnesium sulfate, 0.1% calcium chloride, 0.5% bran.

[0047] All media were packed in 250ml Erlenmeyer flasks with a volume of 30ml, and sterilized at 121°C for 20min.

[0048] After the seed bottles were inoculated, they were cultured on a shaker at 28° C. and 220 r / min for 36 hours to obtain a seed bacterial liquid. Insert 6% of the inoculum into the fermentation bottle at 28°C and cultivate on a shaker at 220r / min for 36h, add 2% notoginseng saponin and 0.6% Tween 80 solution, and continue to cultivate at 28°C on a shaker at 220r / min 72h.

[0049] Carbon source optimization: The effects of glucose, sucrose, maltose, lactose, potato, and starch on the conversion rate of Rd were investigated respe...

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Abstract

The invention relates to the technical field of medicine, in particular to a method for preparing a ginsenoside Rd by using the microbial transformation. The invention provides a method for preparinga ginsenoside Rd by using the microbial transformation, including the following steps: adding the ginsenoside into a nutrient solution, and fermenting after Paecilomyces sp being inoculated; obtainingthallines, adding alcohol liquid into the thallines to extract, and obtaining alcohol extract; extracting the alcohol extract with saturated n-butanol; and separating and purifying by chromatographiccolumn. The method disclosed by the invention has high conversion rate, which reaches 70 to 90%, the fermented product ginsenoside Rd has the purity of 98%, moreover, the microbial industrializationpreparation of ginsenoside Rd can be realized, the medicament resource source of ginsenoside Rd can be expanded, and the wide health care market requirement can be met.

Description

technical field [0001] The invention relates to the field of fermentation technology, in particular to a method for preparing ginsenoside Rd through microbial transformation. Background technique [0002] Ginsenoside Rd has good pharmacological effects on cardiovascular, immune system, nervous system, etc. Some pharmacological effects are unique to ginsenoside Rd and have no biological activity in other monomeric saponins. [0003] The content of ginsenoside Rd in ginseng is low, only about 0.2%. Because of its complex structure, chemical synthesis has not been successful so far. At present, the Rd monomer is extracted from the roots, stems and leaves of plants such as ginseng and Panax notoginseng. The extraction method is due to Its yield is low, and cost is higher, has influenced the further expansion of medical care market demand, so it is necessary to develop a kind of yield high, good purity, and the method for preparing ginsenoside Rd that is simple and feasible. [...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P33/00C12R1/79
Inventor 周珮史训龙冯美卿周超群黄维方平孙宇张亚丽
Owner SHANGHAI HAITAI PHARMA
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