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Method for preparing citicoline

A technology for citicoline and choline chloride, applied in the field of preparation of citicoline, can solve the problems of increased operation intensity, complicated process, increased equipment and the like, and achieves improved utilization rate, high conversion efficiency and cost low effect

Inactive Publication Date: 2009-09-23
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Phosphorylcholine is formed by dehydration and condensation of choline chloride and anhydrous phosphoric acid under high temperature conditions, which increases the number of equipment, complicates the process, greatly increases the operating intensity, and increases the cost

Method used

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  • Method for preparing citicoline
  • Method for preparing citicoline
  • Method for preparing citicoline

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Yeast medium (g / L): glucose 40, urea 2.0, potassium dihydrogen phosphate 1.5, magnesium sulfate heptahydrate 0.5, zinc sulfate heptahydrate 4.0×10 -3 , ferrous sulfate heptahydrate 3.0×10 -3 , manganese chloride tetrahydrate 0.3×10 -3 , anhydrous calcium chloride 1.0×10 -3 , Biotin 0.05×10 -3 . Saccharomyces cerevisiae inoculum was 10%, cultured on a shaker at 120 rpm at 30°C for 24 hours, and centrifuged at 4000 rpm for 20 minutes. Take the yeast paste and store it at -7°C for later use.

Embodiment 2

[0045] Embodiment 2: Using CMP and choline chloride as substrates and glucose as energy donor to prepare citicoline

[0046] In a reaction tank with a capacity of 15L, prepare 60mM choline chloride, 0.20M sodium dihydrogen phosphate, 30mM CMP, 0.3M glucose, 50mM magnesium sulfate, 1mM potassium chloride, 2mM cysteine, using Example 1 2800 grams of Saccharomyces cerevisiae mud, 10 grams of cetyltrimethylamine ammonium bromide and 10 L of reaction liquid cultured by the described method were used to adjust the pH to 6.8 with sodium hydroxide, and stirred at a low speed for 10 hours at 37 ° C. , after the reaction, centrifugal precipitation, supernatant is carried out citicoline quantitative analysis, contains citicoline 14.9 grams per liter in the conversion liquid, and the conversion rate of citicoline reaches 97.4% (mol meter) .

Embodiment 3

[0047] Example 3: Preparation of Citicoline with Cytidine and Choline Chloride as Substrates and Glucose as Energy Donor

[0048] In a reaction tank with a capacity of 15L, prepare 150mM choline chloride, 0.5M potassium dihydrogen phosphate, 50mM cytidine, 0.5M glucose, 2mM magnesium sulfate, 5mM mannitol, and utilize the method described in Example 1 to cultivate 2700g of baker's yeast puree, 10g of Triton X-100 and 12L of reaction liquid, adjust the pH to 6.0 with sodium hydroxide, and stir at 32°C for 15 hours at a low speed. Quantitative analysis of citicoline was carried out, the conversion liquid contained citicoline 22.95 g / liter, and the conversion rate of citicoline reached 90% (in mol).

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Abstract

The invention discloses a method for preparing citicoline. The method comprises the following steps of taking choline chloride, phosphate ions and cytidine monophosphate or a precursor thereof as substrates, taking glucose, fructose, sucrose or maltose as an energy donor, adding a small-molecule chemical effect substance and utilizing the yeast-cell whole cells with permeability to catalyze and prepare the citicoline. By building a metabolic network model and metabolic flow analysis, adopting the small-molecule chemical effect substance to regulate and control metabolic flow so as to improve the efficiency of energy self-coupling and choline phosphorylation and utilizing the yeast cells with permeability to efficiently prepare the citicoline, the method has the advantages of greatly increasing product concentration and raising the utilization ratio of the substrates.

Description

technical field [0001] The invention belongs to the technical field of biopharmaceuticals, and in particular relates to a preparation method of citicoline. Background technique [0002] Citicoline, also known as citicoline sodium, is an activator of brain metabolism, which can promote the synthesis of lecithin in nerve cell membranes. It has the functions of repairing brain damage, resisting hypoxia, improving memory and enhancing intelligence. It is widely used in clinical practice . Therefore, the synthesis technology of citicoline is a research subject that people are more concerned about. [0003] Compared with CMP, which can be produced and supplied cheaply, although it has been reported that citicoline has chemical synthesis, enzymatic and microbial conversion synthesis, etc., the first two methods are too expensive and are no longer used. At present, the commonly used method is the microbial transformation method, which is synthesized by microbial cell enzymes, but ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P19/30C12R1/645
Inventor 应汉杰熊健汤佳鹏张磊李振江柏建新欧阳平凯
Owner NANJING UNIV OF TECH
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