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Method for synthesizing vidarabine by enzymic method

A technology of adenosine vidarabine and adenosine vidarabine, which is applied in the field of enzymatic synthesis of adenosine vidarabine, which can solve the problems of low conversion rate of target substances, large consumption of substrates, pollution of chemical synthesis, etc.

Active Publication Date: 2017-07-07
上海鑫欣生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In the above work, two kinds of enzymes are mixed and used to synthesize the target in one step. Although the pollution problem caused by chemical synthesis is solved, there is a reverse reaction in this method—that is, when the substance at one end of the reaction reaches a certain concentration, it will reverse to the reverse reaction. direction, resulting in unequal molar substrate dosage, that is, in the above reaction, the molar concentration of adenine is 3 times that of arabouridine; the substrate consumption is large; the shortcoming of low conversion rate of the target substance cannot fundamentally replace chemical synthesis

Method used

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  • Method for synthesizing vidarabine by enzymic method
  • Method for synthesizing vidarabine by enzymic method
  • Method for synthesizing vidarabine by enzymic method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Embodiment 1, two-step method reaction 1

[0091] The reaction proceeds in two steps, as follows:

[0092] Response one:

[0093] In the 300mM sodium phosphate solution of 1000ml pH6.8, add 0.04 mole arabouridine to make its concentration reach 40mM, then add 1.56g thallus (every gram contains 32200U of uridine phosphorylase) to make the concentration of uridine phosphorylase reach 50U / ml; place the mixture in a shaker and shake at 55°C for 8 hours.

[0094] Take out the reaction solution and cool it down to 10°C, add 34g of anhydrous calcium chloride solid, and stir for 30 minutes, remove the precipitated uracil, calcium phosphate and other insoluble matter by ultrafiltration, the insoluble matter includes bacteria containing uridine phosphorylase Keep the ultrafiltration solution at 5-10°C, collect the ultrafiltration solution, wash the membrane with 500ml double distilled water after completion, wash out the remaining reaction solution remaining in the membrane bag...

Embodiment 2

[0099] Embodiment 2, two-step method reaction 2

[0100] The reaction proceeds in two steps, as follows:

[0101] Response one:

[0102] In the 600mM sodium phosphate solution of 1000ml pH6.8, add 0.2 mole arabouridine to make its concentration reach 200mM, then add 6.21g thalline (every gram contains 32200U of uridine phosphorylase) to make the concentration of uridine phosphorylase reach 200U / ml; place the mixture in a shaker and shake at 55°C for 8 hours.

[0103]Take out the reaction solution and cool it to 10°C, add 1000ml of double distilled water to dilute the reaction solution; add 68g of anhydrous calcium chloride solid, and stir for 30 minutes, ultrafiltration to remove the precipitated uracil, calcium phosphate and other insoluble matter, the insoluble The substances include bacteria containing uridine phosphorylase; keep the ultrafiltration solution at 5-10°C, wash the membrane with 500ml double distilled water after completion, wash out the remaining reaction so...

Embodiment 3

[0108] Embodiment 3, comparative example 1

[0109] Using the same method as in Example 1, the main difference is that in "Reaction 1" after "shaking at 55°C for 8 hours", adenine is directly added to carry out Reaction 2, excluding low-temperature ultrafiltration and "Reaction 1" in "Reaction 1". Two" in the double distilled water dilution step.

[0110] The reaction proceeds in two steps, as follows:

[0111] Response one:

[0112] In the 300mM sodium phosphate solution of 1000ml pH6.8, add 0.04 mole arabouridine to make its concentration reach 40mM, then add 1.56g thallus (every gram contains 32200U of uridine phosphorylase) to make the concentration of uridine phosphorylase reach 50U / ml; place the mixture in a shaker and shake at 55°C for 8 hours.

[0113] After the mixed solution obtained above was diluted 3 times, it was quantitatively analyzed by high performance liquid phase (ELSD), and the peak area of ​​arabinose-1-phosphate was 72974.04. This figure was substitut...

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Abstract

The invention relates to a method for synthesizing vidarabine by an enzymic method, and provides a method for preparing vidarabine by a two-step method. According to the method, different functions of uridine phosphorylase and purine nucleoside phosphorylase are utilized fully, so that reaction is carried in one direction; reverse reaction is eliminated; and conversion rate of the vidarabine is increased remarkably.

Description

technical field [0001] The invention belongs to the field of biochemical industry, and more specifically, the invention relates to a method for enzymatically synthesizing adenosine vidarabine. Background technique [0002] Adenosine Arabinoside (araA) is a natural antiviral compound with broad-spectrum antiviral activity and has been widely used clinically to treat diseases caused by cytomegalovirus, hepatitis B virus and herpes virus. [0003] In the prior art, the synthesis method of vidarabine includes chemical synthesis and biotransformation synthesis. The chemical method has complicated process, harsh reaction conditions, high cost of raw materials, and the use of heavy metal reagents, which pollutes the environment. The basic principle of the biotransformation method is that arabinogluridine and phosphate ions generate arabinose 1-phosphate and corresponding bases under the action of uridine phosphorylase (EC 2.4.2.3); Vidarabine and phosphate ions are generated unde...

Claims

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Application Information

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IPC IPC(8): C12P19/40
CPCC12P19/40
Inventor 沈波魏民志
Owner 上海鑫欣生物科技有限公司
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