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Method for preparing acarbose through microbial fermentation

A technology for microbial fermentation and acarbose, applied in the field of fermentation, can solve the problems of reducing maltose content and affecting the yield of acarbose, and achieves the effects of improving the fermentation level and the fermentation titer.

Active Publication Date: 2011-08-03
ZHEJIANG UNIV OF TECH +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because maltose will be directly incorporated into the structure of acarbose, the consumption of glucose in the early stage will lead to the reduction of maltose content in the later stage, which in turn affects the production of acarbose

Method used

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  • Method for preparing acarbose through microbial fermentation
  • Method for preparing acarbose through microbial fermentation
  • Method for preparing acarbose through microbial fermentation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Embodiment 1: the production of acarbose (erlenmeyer flask shaking fermentation)

[0043] Prepare seed culture medium, its culture medium composition is as follows: cornstarch 15.0g / L, soybean cake powder 40.0g / L, glycerol 20.0g / L, K 2 HPO 4 0.1g / L, CaCO 3 2.0g / L, prepared with tap water, initial pH 7.0. Sterilized at 121°C for 30 minutes.

[0044] Preparation of fermentation medium, the composition of the medium is as follows: maltose syrup 80.0g / L, glucose 20.0g / L, soybean cake powder 10.0g / L, corn steep liquor 5.0g / L, FeCl 3 0.1g / L, CaCl 2 2.0g / L, CaCO 3 6.0g / L, prepared with tap water, pH 7.0. Sterilized at 121°C for 30min.

[0045] Prepare validamycin A aqueous solution: use tap water to prepare validamycin A solution with validamycin A concentration of 6.3 g / L, and sterilize at 121° C. for 30 minutes.

[0046] Seed preparation: Aseptically transfer the CCTCC NO: M 209022 strain in a low-temperature glycerol tube to a fresh, sterile solid plate, activate...

Embodiment 2

[0052] Embodiment 2: the production of acarbose (stirred reactor fermentation)

[0053] The preparation of seed culture medium and fermentation medium is the same as in Example 1.

[0054] Preparation of validamycin A aqueous solution: use tap water to prepare validamycin A solution with validamycin A concentration of 12 g / L, and sterilize at 121° C. for 30 minutes.

[0055] Seed preparation: Aseptically transfer the CCTCC NO: M 209022 strain in a low-temperature glycerol tube to a fresh, sterile solid plate, activate it at 28°C for 2 days, pick the colony and inoculate it in a 500ml triangular flask for seed cultivation, the volume of the seed medium in the bottle 50ml, shaker rotation speed 200rpm, cultivated at 28°C for 72h to obtain seed solution.

[0056] Fermentation of Acarbose:

[0057] Take 24L of the above-prepared fermentation medium and put them into three 15L stirring fermenters respectively, each fermenter is filled with 8L fermentation medium, sterilized at 12...

Embodiment 3

[0061] Embodiment 3: the extraction of acarbose

[0062]Get 100 milliliters of fermented liquid in bottle No. 12 of Example 12, adjust the pH value to 3, centrifuge at 3000 × g for 10 minutes, discard the precipitate, and collect the supernatant; the clarified supernatant is initially separated by macroporous cation exchange. The chromatographic column is a 2.5×30 cm glass column (BIO-Rad, USA). For primary separation chromatography, UNOsphere S filler with a particle size of 80 microns was selected, and the column was packed by wet method. The bed height was about 22.3 cm, and the medium used was 5.0×10 -2 M hydrochloric acid dilute acid transformation into H + type. The flow rate of the mobile phase was set to 440 ml / hour (linear velocity 1.5 cm / min). After fully equilibrating, take 2.0 ml of concentrated solution with a concentration of 5 g acarbose / liter concentrated solution for loading, and wash 7 samples with ultrapure water. Column volume, eluted with 5.0×10 -3 M h...

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Abstract

The invention discloses a method for preparing acarbose through microbial fermentation. The method comprises the following steps: inoculating acarbose producer (CCTCC NO:M 209022) in fermentation medium which is suitable for the strain and contains carbon source, nitrogen source and inorganic salt, fermenting and culturing at 20-32 DEG C for 96-192 hours, obtaining fermentation liquor after fermentation, extracting to obtain acarbose. The method is characterized in that fermenting and culturing are performed for 0-48h and validamycin A solution is added to ensure that the one litre of fermentation medium contains 0.04-0.4g of validamycin A. By supplementing validamycin A in the fermentation process of acarbose, the fermentation level of acarbose can be increased.

Description

(1) Technical field: [0001] The invention belongs to the technical field of fermentation, and designs a method for preparing acarbose by microbial fermentation, in particular to a method for increasing the yield of acarbose by adding validamycin A during the fermentation process of acarbose. (two) background technology: [0002] Acarbose is white or off-white powder, soluble in water, pKa value is 5.1, molecular formula is C 25 h 43 NO 18 , molecular weight 645.6, chemical name O-{4,6-dideoxy-4[(1S,4R,5S,6S)-4,5,6-trimethylol-2 cyclohexenyl]-α-D -Glucopyranosyl}-(1→4)-O-α-D-glucopyranosyl-(1→4)-D-glucopyranose. Molecular structural formula is shown in following formula I: [0003] [0004] It can be seen from the chemical structural formula of acarbose that it is a complex pseudotetrasaccharide, which contains two parts, acarviose and maltose, in which acarviose is composed of valienamine and 4-amino-4,6 with an unsaturated C7-cyclic polyol structure -Dideoxyglucose ...

Claims

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Application Information

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IPC IPC(8): C12P19/00
Inventor 郑裕国王亚军薛亚平王远山沈寅初
Owner ZHEJIANG UNIV OF TECH
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