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Method for preparing nucleotide from enzymolysis liquid of ribonuclease

A ribonucleic acid enzymatic hydrolysis solution and strong acid technology, which is applied in the preparation of sugar derivatives, chemical instruments and methods, sugar derivatives, etc., can solve the problems of production methods that cannot meet the needs, high requirements for equipment and environment, and long elution and separation time, etc. problem, to achieve good clarity, shorten the separation and purification time, and facilitate separation and purification

Inactive Publication Date: 2009-06-10
北京燕京中科生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method uses two negative columns and four carbon columns to separate four kinds of nucleotides. This method has a slow separation speed and takes a long time; the use of carbon columns and formic acid elution has high requirements on equipment and environment, high cost, environmental protection and safety. Poor; the alcohol-alkali elution of the carbon column will lead to a long elution and separation time, the regeneration of the carbon column is complicated, and the subsequent purification is difficult
[0007] With the increase of clinical demand for nucleic acid drugs and people's increasing requirements for quality of life, the application of nucleotides and their derivatives is becoming more and more extensive, and the original production methods can no longer meet the needs.

Method used

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  • Method for preparing nucleotide from enzymolysis liquid of ribonuclease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] The ribonucleic acid hydrolysate was passed through two ultrafiltration membrane equipment to remove solid particles and proteins, and the filtration recovery rate was greater than 90%. The MWCO of the two ultrafiltration membranes were 20,0000 and 6,000 Daltons respectively; after ultrafiltration, the filtrate protein was 0.005mg / ml; adjust the pH of the ultrafiltrate to 1.5 with 6N HCl, and separate it on an ion exchange column, which contains 2065g of nucleotides, 5′-AMP (2065×28%) grams; 5′-GMP (2065×26%) grams; 5'-CMP (2065 x 20%) g; 5'-UMP (2065 x 26%) g.

[0040] The ultrafiltrate containing the four mononucleotides was separated into UMP, GMP and CMP mixed solution, AMP three part.

[0041] Cation exchange resin loading column: pH 1.5, sample loading flow rate 1.5m / h, nucleotide loading amount is 6.0% of the total exchange capacity of cation resin; product loss rate in column separation: within 5%.

[0042] Contrast value and concentration for identification:...

Embodiment 2

[0051] The ribonucleic acid hydrolysate containing 2080 grams of nucleotides was subjected to plate and frame pressure filtration-ultrafiltration membrane equipment to remove solid particles and proteins, and the filtration recovery rate was greater than 95%. The ultrafiltration membrane MWCO was 10,000 Daltons; the filtrate after ultrafiltration Protein 0.02mg / ml; adjust the pH of the ultrafiltrate to 2.0 with 12NHCl, and separate it on an ion exchange column, which contains 2080g of nucleotides, 5′-AMP (2080×26.8%) grams; 5′-GMP (2080×28.2%) ) grams; 5′-CMP (2080×20.1%) grams; 5′-UMP (2080×24.6%) grams.

[0052] The nucleotide loading of the 732 cation exchange resin column was 6.0% of the total exchange capacity of the resin, and the separation linear velocity was 1.75 m / h. The loading volume of other concentration columns is: Dowex-1 negative column AMP loading volume is 45%; GMP+CMP separation column loading volume is 12%; CMP25%; UMP18%, using acid salt eluent (0.5% sal...

Embodiment 3

[0056] The ribonucleic acid enzymatic hydrolysate was passed through two ultrafiltration membrane equipment to remove solid particles and proteins, and the filtration recovery rate was 95%. The MWCO of the two ultrafiltration membranes were 100,000 and 10,000 Daltons respectively; after ultrafiltration, the filtrate protein was 0.01mg / ml; adjust the pH of the ultrafiltrate to 1.0 with 10NHCl, and separate it on a JK008 cation exchange column, which contains 2770g of nucleotides, 5′-AMP (2770×25%) grams; 5′-GMP (2770×28%) grams; 5'-CMP (2770 x 18%) g; 5'-UMP (2770 x 25%) g.

[0057] The nucleotide loading of the 001×8 ion exchange resin column was 8.0% of the total exchange capacity of the resin, and the separation linear velocity was 1.0 m / h. The loading volume of other concentration columns is: Amberlite IRA-400 negative column AMP loading volume is 40%; GMP+CMP separation column loading volume is 15%; CMP30%; UMP20%, using acid salt eluent (1 % salt, pH 2) for separation a...

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Abstract

The invention discloses a making method of 5'-nucleic acid from enzymolytic liquid of ribonucleic acid, which comprises the following steps: predisposing enzymolytic liquid of ribonucleic acid; adopting ionic exchange to separate and purify four nucleic acids; condensing; spraying; drying to obtaining the product.

Description

technical field [0001] The present invention relates to a method for preparing mononucleotide, in particular to a method for preparing mononucleotide from ribonucleic acid enzymatic hydrolysate using ion exchange resin column in an orderly manner. Background technique [0002] 5'-mononucleotide is an important chemical raw material, which can be used as pharmaceutical intermediates, food additives and health food raw materials. It can be used to prepare a variety of biochemical raw materials, such as ATP, CTP, CDPC, 5-FU, etc. [0003] Generally, ribonucleic acid (RNA) can be extracted from yeast in industry. 5'-AMP (5'-adenosine), 5'-GMP (5'-guanylate) can be obtained by enzymatic or alkaline hydrolysis of RNA, and then separation, purification and purification of the enzymatic or alkaline solution. , 5'-CMP (5'-cytidylic acid) and 5'-UMP (5'-uridylic acid) four nucleotides. [0004] At present, some literatures have reported its separation and preparation methods. For e...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07H1/06C07H19/167C07H19/067
Inventor 姜波赵鹏赵晜黎高沃
Owner 北京燕京中科生物技术有限公司
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