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Production method of inosine nucleotide

A technology of inosine nucleotides and production methods, applied in the field of biosynthesis, can solve problems such as the difficulty of separating and extracting inosine nucleotides, and achieve the effects of easy separation and purification, sufficient supply, and reduced equipment requirements

Active Publication Date: 2016-03-16
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the accumulation of inosine acid in flask fermentation and 5L fermenter can reach 19.1g / L and 70.3g / L respectively. Even so, it is still quite difficult to separate and extract inosine nucleotide from the 7.03% solution

Method used

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  • Production method of inosine nucleotide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Pretreatment of resin: AmberliteIRA-900 resin is pretreated according to the following method steps:

[0061] Step 1a, weigh 200g of AmberliteIRA-900 resin into a 1L Erlenmeyer flask, add 600ml of absolute ethanol, soak overnight, wash with deionized water until no ethanol remains in the eluent, and then suction filter;

[0062] Step 1b, soak the resin obtained in step 1a with 600ml of 1M NaOH solution overnight, wash with deionized water until the eluent is pH neutral, and then suction filter;

[0063] Step 1c, soak the resin obtained in step 1b with 600ml of 1M HCl solution overnight, wash with deionized water until the eluent is pH neutral, and then suction filter;

[0064] Step 1d, soak the resin obtained in step 1c with 600ml of 1M NaOH solution overnight, wash with deionized water until the eluent is pH neutral, and then suction filter;

[0065] In step 1e, soak the resin obtained in step 1d with 600ml of 0.5M NaCl solution overnight, wash with deionized water un...

Embodiment 2

[0069] Resin pretreatment: AmberliteIRA-93 resin was pretreated according to the same method as in Example 1.

[0070] Catalytic reaction: Weigh 10g of the above-mentioned pretreated AmberliteIRA-93 resin into a 100ml Erlenmeyer flask, then add 50ml of adenylate deaminase solution (10U / ml, add acetate buffer solution, adjust the pH to 5~ 6) Place the flask on a shaker at 20°C and 150rpm for 2 hours, recover the immobilized enzyme, and wash the immobilized enzyme with deionized water until there is no protein and enzyme activity in the eluate. Add the obtained immobilized enzyme (13.17U / g) to 100ml of adenylic acid solution (100g / L, pH5~6) preheated at 40°C, and make the adenylic acid The deamination reaction is carried out, and the pH is controlled to be 5-6 by dropwise adding 1M hydrochloric acid solution. After about 2.5 hours, OD265 no longer drops, liquid phase detection (liquid phase conditions: mobile phase is 0.02M ammonium dihydrogen phosphate solution, containing 3.5...

Embodiment 3

[0072] Resin pretreatment: Amberlite IRA-400 resin was pretreated according to the same method as in Example 1.

[0073] Catalytic reaction: Weigh 10g of the above-mentioned pretreated AmberliteIRA-400 resin into a 100ml Erlenmeyer flask, then add 50ml of adenylate deaminase solution (10U / ml, add acetate buffer solution, adjust the pH to 5~ 6) Place the flask on a shaker at 20°C and 150rpm for 2 hours, recover the immobilized enzyme, and wash the immobilized enzyme with deionized water until there is no protein and enzyme activity in the eluate. Add the obtained immobilized enzyme (14.37U / g) into 100ml of adenylic acid solution (100g / L, pH5~6) preheated at 40°C, and make the adenylic acid at 40°C, under the condition of mechanical stirring at 60rpm The deamination reaction is carried out, and the pH is controlled to be 5-6 by dropwise adding 1M hydrochloric acid solution. After about 3.25h, OD265 no longer drops, liquid phase detection (liquid phase conditions: mobile phase i...

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Abstract

The invention provides a production method of inosinic acid. The method comprises the step of adopting adenosine deaminase fixed in anion resin to catalyze the generation of the inosinic acid from adenylic acid. According to the method provided by the invention, the adenylic acid is taken as a substrate, the supply of raw materials is sufficient, the requirements for equipment are low, the separation and extraction are convenient, the semi-continuous production of the inosinic acid can be realized, the substrate is convenient and easy to obtain, the transformation is thorough, and the cost is relatively low.

Description

technical field [0001] The invention belongs to the field of biosynthesis, and in particular relates to a production method of inosine nucleotide. Background technique [0002] Inosine nucleotide, also known as inosinic acid, inosinic acid or hydroxypurine nucleotide, referred to as 5'-IMP, is an important intermediate metabolite of purine nucleotide metabolism in the body, and is a combination of adenylic acid and guanosine The link of acid metabolism can realize the interconversion of adenosine and guanosine, so as to maintain the balance of purine nucleotide metabolism in the body. Inosine nucleotide participates in energy metabolism and nuclear protein synthesis in the body, activates the pyruvate oxidase system, increases the activity of coenzyme A, enables tissue cells in low-energy and hypoxic conditions to continue to metabolize smoothly, and activates the liver Function, speed up the repair of damaged liver cells, and stimulate the body to produce antibodies, suita...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P19/32
Inventor 应汉杰曹治陈勇陈晓春吴菁岚谢婧婧
Owner NANJING TECH UNIV
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