Method for producing lipstatin through microbial fermentation and culture medium
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A technology for microbial fermentation and fermentation medium, which is applied in the field of microbial fermentation to produce lipistatin and medium, can solve problems such as low fermentation level, and achieve the effects of increasing titer, increasing yield and reducing cost
Inactive Publication Date: 2014-05-28
ZHEJIANG UNIV OF TECH +1
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European patent EP0803567 describes the process of improving liprestatin by adding linoleic acid, octanoic acid and N-f
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[0023] Slant medium preparation: soluble starch 10g, wheat flour 5g, yeast extract 1g, casein 1g, potassium dihydrogen phosphate 0.5g, magnesium sulfate 0.5g, agar 20g, add water to 1L, adjust the pH value to 7.1, 121~ Sterilize at 125°C for 20 minutes. This medium is used for strain activation.
[0024] Preparation of seed medium: soybean cake powder 20g, glycerin 20g, yeast extract 5g, add water to 1L, adjust pH to 7.1, sterilize at 121-125°C for 20min. This medium is used for seed liquid preparation.
[0025] Initial fermentation medium: soybean cake powder 32.5g, corn gluten powder 8g, sunflower oil 100g, soybean lecithin 25g, glycerin 22.5g, Tween-80 0.05g, calcium carbonate 0.8g, vitamin E 0.025g, vitamin C 0.125g , add water to make up to 1L, adjust the pH to 7.1, sterilize at 121-1...
Embodiment 2
[0053] Slant medium preparation: soluble starch 10g, wheat flour 5g, yeast extract 1g, casein 1g, potassium dihydrogen phosphate 0.5g, magnesium sulfate 0.5g, agar 20g, add water to 1L, adjust the pH value to 7.1, 121~ Sterilize at 125°C for 20 minutes. This medium is used for strain activation.
[0054] Preparation of seed medium: soybean cake powder 20g, glycerin 20g, yeast extract 5g, add water to 1L, adjust pH to 7.1, sterilize at 121-125°C for 20min. This medium is used for seed liquid preparation.
[0055] Preparation of fermentation medium: soybean cake powder 32.5g, corn gluten powder 8g, sunflower oil 100g, soybean lecithin 25g, glycerin 22.5g, Tween-80 0.05g, calcium carbonate 0.8g, vitamin E 0.025g, vitamin C 0.125g , MgSO 4 12 mmol, CoSO 4 ·7H 2 O1mmol, ZnSO 4 ·7H 2 O0.25mmol, add water to make up to 1L, adjust pH to 7.1, sterilize at 121-125°C for 20min. This medium is used for fermentative production of riprestatin.
[0056] Slope culture: the slant medi...
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Abstract
The invention provides a method for producing lipstatin through microbial fermentation, and a used fermentation culture medium. The method comprises the following steps: inoculating streptomyces toxytricini into the fermentation culture medium, adding Mg<2+>, Co<2+> and Zn<2+> with the final concentrations of 10-15 mmol/L, 0.5-2.0 mmol/L and 0.1-0.5 mmol/L respectively in the culture medium, conducting fermental cultivation at 220 r/min and at the temperature of 28 DEG C, adding aminocaproic acid on the fourth day of fermental cultivation to enable the final concentration of the aminocaproic acid to be 40-50 mmol/L, adding n-caprylic acid on the sixth day of fermental cultivation to enable the final concentration of the n-caprylic acid to be 30-40 mmol/L, and keeping the total fermental cultivation time to be 150-180 hours, so as to obtain the lipstatin from the fermentation liquid after the fermentation is completed. According to the invention, the addition combination of a precursor and metal ions affecting the fermentation and synthesis of the lipstatin is optimized, and the best combination of the precursor and the metal ions is found, so that the yield of the lipstatin is remarkably increased, the potency is remarkably improved after the method and the culture medium are applied to the industrial production of the lipstatin, and the cost is lowered.
Description
(1) Technical field [0001] The invention relates to a method for producing riprestatin by microbial fermentation and a used fermentation medium. (2) Background technology [0002] Lipstatin is a metabolite produced by fermentation of Streptomyces toxytricini, and its tetrahydro derivative is orlistat, a weight-loss drug successfully developed by Roche ( Xenical), Orlistat is the first gastrointestinal lipase inhibitor, which inactivates the enzyme by forming a covalent bond with the active serine site of gastric lipase and pancreatic lipase in the lumen of the stomach and small intestine, reducing Decomposition and absorption of fat, so as to achieve the role of weight control. Orlistat is currently the only weight-loss drug that does not enter the blood and does not act on the central nervous system. [0003] Orlistat can be synthesized by total chemical synthesis or by first producing liprestatin by fermentation synthesis followed by hydrogenation. Due to the relatively...
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