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Enzymatic conversion method of L-ornithine

A technology of ornithinase and arginase, which is applied in the field of L-ornithine enzymatic conversion and preparation, can solve the problem of high cost of L-arginine, and achieve reduced production costs, good economic benefits and social benefits , The effect of reducing production costs

Active Publication Date: 2015-08-12
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0014] In summary, among the methods for enzymatic preparation of L-ornithine, the arginase hydrolysis method is the closest to industrial application, but the cost of the reaction substrate L-arginine is relatively high
In view of the development of L-ornithine fermentation technology, it is difficult for the current industrial technology to prepare L-ornithine by enzymatically producing L-ornithine with high purity L-arginine to compete with the production technology of L-ornithine fermentation for a long time, so looking for The fundamental purpose of the present invention is to obtain L-arginine industrial raw materials with abundant sources and low cost and to achieve a breakthrough in the key technology of enzymatically producing L-ornithine

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] The preparation method of L-ornithine enzymatic transformation consists of the following steps:

[0030] (1) The strains Enterococcus faecalis or Alcaligenes or Bacillus or Micrococcus inhabiting with arginase activity are cultivated in a culture medium. The carbon source of the medium is glucose, maltose, sucrose and / or fructose. The mass concentration of total carbon source is 1-30g / L; the nitrogen source is beef extract, yeast extract, corn steep liquor, peptone and / or soybean cake hydrolysate, and the mass concentration of total nitrogen source in the medium is 1-30g / L, which produces high enzyme Live arginase

[0031] (2) Fermentation broth containing arginine with a mass concentration of 10~150g / L, or arginine ion exchange eluate containing arginine with a mass concentration of 10~150g / L, or arginine The crude arginine or crude arginine salt with a weight ratio of 50%-95% is prepared by adding water to prepare a substrate solution. The prepared crude substrate aqueous...

Embodiment 2

[0033] 1. Culture Enterococcus faecalis ATCC 8043 in the following 1000ml medium (g / ml): corn syrup 2.0%, protein hydrolysate 0.5%, beef extract 1%, maltose 1.0%, lactose 0.5%, NaCl 0.5%, KH 2 PO 4 0.5%, MgSO 4 ·7H 2 O 0.03%, pH 7.2. Shaking culture at 37°C for 16 hours, centrifugation at 4000 r / min for 15 minutes to obtain 18 g of wet cells.

[0034] 2. Add the wet bacteria to 1000ml of conversion solution, the conversion solution is arginine fermentation broth containing 150g / L L-arginine, pH 9.0, then add 1.0g / L TritonX-100, 40℃ enzymatic reaction At 18h, after the reaction, a sample was taken to detect that the mass concentration of L-ornithine in the conversion solution was 112.7 g / L, and the molar conversion rate to L-arginine was 99%.

[0035] 3. Centrifuge the transformation solution to remove the bacteria. The supernatant was neutralized with 6mol / L hydrochloric acid to pH 6.0, heated to 80°C, activated carbon decolorized, vacuum filtered, the filtrate was concentrated unti...

Embodiment 3

[0038] 1. Cultivate Enterococcus faecalis CGMCC NO.1786 in the following 1000ml medium (g / ml): corn steep liquor 0.5%, NaCl 0.5%, KH 2 PO 4 0.05%, MgSO 4 ·7H 2 O 0.02%, beef extract 2%, glucose 1%, maltose 0.5%, pH 7.2. Shaking culture at 37°C for 16 h, centrifugation at 4000 r / min for 15 min to obtain 15 g of wet cells.

[0039] 2. Add the wet bacteria to 1000ml conversion solution, the conversion solution is arginine fermentation broth containing 100g / L L-arginine, pH 9.0, then add 0.1g / L Tween 80, 40℃ enzymatic reaction After 15 hours, after the reaction, a sample was taken to detect that the mass concentration of L-ornithine in the conversion solution was 75.1 g / L, and the molar conversion rate to L-arginine was 99%.

[0040] 3. Centrifuge the transformation solution to remove the bacteria. The supernatant was neutralized with 6mol / L hydrochloric acid to pH 6.0, heated to 80°C, activated carbon decolorized, vacuum filtered, the filtrate was concentrated until a small amount of c...

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Abstract

The invention belongs to the field of bio-techniques and particularly relates to an enzymatic conversion method of L-ornithine. The method includes: adding bacterial cells of arginase activity or crude enzyme extract into liquor containing arginine, or crude arginine, or crude arginine salt, serving as substrate preparation enzyme conversion liquid, allowing enzymatic reaction at 25 DEG C to 55 DEG C under pH of 7 to 11, and performing separation by the combination of isoelectric point crystallization and ion exchanging so as to obtain a conversion product, L-ornithine salt. The method has the advantages such that the L-ornithine salt of high added value is synthesized by a substrate enzymatic method with the low-cost L-ornithine liquor, the range of material sources is wide, enzymatic reaction is efficient, operating is convenient, and production cost is low.

Description

1. Technical Field [0001] The invention belongs to the field of biotechnology, and specifically relates to a preparation method of L-ornithine enzymatic conversion. 2. Background technology [0002] L-Ornithine (L-Ornithine) is a non-protein amino acid, mainly found in meat, fish, and cheese. It is also a component of bacterial cell membranes and peptide antibiotics. It mainly exists in the liver in a free state in the human body. The main physiological function of L-ornithine is to participate in the urea cycle and has a strong ammonia detoxification function. Therefore, it is widely used in the development of drugs for the treatment of hepatic encephalopathy, hepatitis and liver protection, such as L-ornithine-L- Aspartic acid, L-ornithine phenylacetic acid, etc. In addition, L-ornithine has many other biological functions, such as stimulating growth hormone secretion, improving sleep, alleviating fatigue, and can also be used as a food additive to improve food flavor. With t...

Claims

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Application Information

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IPC IPC(8): C12P13/10C12R1/05C12R1/07C12R1/265C12R1/465
Inventor 焦庆才刘均忠高亮陆阳刘茜
Owner NANJING UNIV
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