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A kind of method that uridine phosphoryl compound co-produces chiral hydroxyl ester

A technology of uridine phosphoryl and compound is applied in the field of preparation of biochemical products, can solve the problems of low yield and high cost, achieve the effects of high conversion efficiency, low cost, and overcome low substrate conversion rate

Inactive Publication Date: 2012-02-01
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem to be solved by the present invention is to provide a method for the joint production of chiral hydroxy esters of uridine phosphoryl compounds, which can overcome the problems of synthesizing uridine phosphoryl compounds or chiral hydroxy esters alone in the prior art. The problem of low yield and high cost

Method used

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  • A kind of method that uridine phosphoryl compound co-produces chiral hydroxyl ester
  • A kind of method that uridine phosphoryl compound co-produces chiral hydroxyl ester
  • A kind of method that uridine phosphoryl compound co-produces chiral hydroxyl ester

Examples

Experimental program
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Effect test

Embodiment 1

[0039] Example 1: Uridine-phosphate (UMP) co-production of (S)-(-)-4-chloro-3-hydroxybutyric acid ethyl ester.

[0040] In a reaction tank with a capacity of 15L, 10 grams of orotic acid OA, 50 grams of ethyl 4-chloroacetoacetate, 500 grams of glucose, 500 grams of commercially available baker's yeast powder, magnesium sulfate 10mM, potassium dihydrogen phosphate 0.01M, 10 L of reaction solution consisting of 12 ml of xylene and water was used to adjust the pH to 8.5 with sodium hydroxide, and the reaction was stirred at 30° C. for 20 h at high speed, and the ventilation rate was 0.5 (V / V·min). After the reaction, inactivate with perchloric acid, extract (S)-(-)-4-chloro-3 hydroxybutyric acid ethyl ester with acetoacetic acid, use HPLC and GC to analyze uridine monophosphate (UMP) and (S )-(-)-4-chloro-3-hydroxybutyric acid ethyl ester is quantitatively analyzed, and the optical rotation value of product (S)-(-)-4-chloro-3-hydroxybutyric acid ethyl ester is measured with a pol...

Embodiment 2

[0041] Example 2: Coproduction of uridine diphosphate D-glucose (UDP-Glc) (S)-(+)-4,4,4-trifluoro-3-hydroxybutyric acid ethyl ester ((S)-TFHBE) .

[0042] 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% (v / v), 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.

[0043] In a reaction tank with a capacity of 15L, 100 grams of OA, 3000 grams of glucose, 500 grams of 4,4,4-trifluoro-acetoacetate (TFAAE), 5000 grams of Saccharomyces cerevisiae mud, air-dried treatment, magnesium sulfate 200mM, Sodium dihydrogen phosphate 2M, Triton X-100 10ml and 10L of reaction solution composed of water...

Embodiment 3

[0044] Example 3: Coproduction of ethyl (S)-(-)-4-bromo-3-hydroxybutyrate from uridine triphosphate (UTP).

[0045] In the reaction tank with a capacity of 15L, 60 grams of OA, 1400 grams of glucose, 200 grams of ethyl 4-bromoacetoacetate, 20 mM of magnesium sulfate, and 2700 grams of baker's yeast were cultivated by the method described in Example 2, and freeze-thawed 3 times. Disodium hydrogen phosphate 0.6M, cetyl trimethylamine ammonium bromide 10 g and water reaction solution 10L, adjust the pH to 7 with sodium hydroxide, the ventilation rate is 1.0 (V / V min), in 30 Stir the reaction at high speed for 16 hours at ℃, then stop the ventilation, reduce the stirring rate, raise the temperature to 37℃, adjust the pH to 6.5 with HCl, add 500 grams of glucose, and react for another 2 hours. After the reaction, inactivate with perchloric acid , extracted (S)-(-)-4-bromo-3-hydroxybutyrate ethyl ester with ethyl acetate, and used HPLC and GC to analyze uridine triphosphate (UTP) an...

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PUM

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Abstract

The invention discloses a method for co-producing chiral hydroxyl esters of uridine phosphoryl compounds. The method uses orotic acid and β-keto ester compounds as substrates, uses glucose as an energy donor, and utilizes permeable Simultaneous production of uridine phosphoryl compounds and chiral hydroxyl esters by yeast cells. The present invention utilizes the principles of whole-cell catalysis and metabolic engineering, establishes a metabolic network model and analyzes metabolic flow, and utilizes permeable yeast cells to co-produce uridine phosphoryl compounds and chiral hydroxyl esters, increasing the yield and reducing production costs , The synthesis time is shortened, and the utilization rate of the substrate is greatly improved.

Description

technical field [0001] The invention relates to a preparation method of biochemical products, in particular to a new method for co-producing chiral hydroxyl esters of uridine phosphoryl compounds. Background technique [0002] Uridine phosphoryl compounds and chiral hydroxyl esters are extremely important compounds. Uridine phosphoryl compounds are direct precursors for the synthesis of nucleic acids and oligosaccharides, important coenzymes and energy carriers, and play a role in the life metabolism of cells. It plays a very important role; while chiral hydroxy esters are mainly used in the synthesis of drugs, and have more applications in pesticides, insect pheromones, spices and food additives, organic materials and optical materials. [0003] The reported methods for preparing uridine phosphoryl compounds mainly include chemical synthesis and enzymatic methods, but the cost of these methods is too high. The commonly used method is the microbial transformation method, whi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12R1/78C12R1/84C12P7/04C12R1/645C12P19/30C12R1/72
Inventor 应汉杰张月圆叶齐栗西木曹厚陈勇熊健柏建新
Owner NANJING TECH UNIV
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