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Method for preparing <14>NL-phenylalanine with enzyme

A technology for phenylalanine and enzymatic preparation, which is applied in the fields of food, chemical industry and medicine, can solve the problems of difficulty in obtaining, complicated separation, and complicated steps, and achieves the effects of easy separation and purification, simple recovery process and simple synthesis process.

Inactive Publication Date: 2008-09-24
BEIJING UNIV OF CHEM TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Among the other common phenylalanine preparation methods that can be used for reference, the extraction method cannot introduce isotopes; the DL-phenylalanine obtained by the chemical synthesis method needs to be split, the steps are cumbersome, there are many by-products, the separation is complicated, and the yield is low
Fermentation method has many and complex nitrogen sources, is not easy to obtain, has a long preparation cycle and low yield
And because the nitrogen source is many and complex, it affects the product abundance

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] A fermentation culture of bacteria

[0025] The Rhodotorula glutinis 2.102 strain containing phenylalanine ammonia-lyase was cultured in a solid state at 30° C. for 20 hours. The components and contents of the solid medium were: agar with a mass percentage of 1.5%, 8° Be malt extract powder, The rest is water. Take one ring in the seed culture medium, the composition and mass percentage of the seed culture medium are: glucose 1.0%, peptone 1.5%, yeast extract 1.0%, K 2 HPO 4 0.15%, NaCl 0.5%, and the rest is water. Cultivate on a shaking table at 30°C for 20 hours at 150 rpm, and transfer to the enzyme-producing medium with 10% inoculum. The components and mass percentages of the enzyme-producing medium are: 1.0% glucose, 3.0% corn steep liquor, bean cake powder 2.0%, NaCl 0.4%, K 2 HPO 40.05%, L-Phe (L-phenylalanine) 0.02%, adjust the pH to 5.5 with dilute hydrochloric acid with a concentration of 3-4%, and the rest is water. Cultivate on a shaker at 30° C. and 1...

Embodiment 2

[0033] A fermentation culture of bacteria

[0034] The Rhodotorula glutinis 2.102 strain containing phenylalanine ammonia-lyase was cultured in solid at 25°C for 30 hours. The composition and content of the solid medium were: agar with a mass percentage of 2.0%, 10° Be malt extract powder , and the rest is water. Take one ring in the seed medium, the composition and mass percentage of the seed medium are: 0.5% glucose, 1.0% peptone, 1.0% yeast extract, K 2 HPO 4 0.10%, NaCl 0.5%, and the rest is water. Cultivate on a shaking table at 28° C. at 150 rpm for 24 hours, and transfer to the enzyme-producing medium with a 10% inoculum size. The components and mass percentages of the enzyme-producing medium are: 0.5% glucose, 2.0% corn steep liquor, bean cake powder 1.0%, NaCl 0.5%, K 2 HPO 4 0.10%, L-Phe 0.05%, pH5.5, the rest is water. Cultivate on a shaker at 28° C. at 150 rpm for 22 hours, centrifuge and wash twice with saline, and the obtained PAL-containing bacteria are us...

Embodiment 3

[0042] A fermentation culture of bacteria

[0043] The Rhodotorula glutinis 2.102 bacterial strain containing phenylalanine ammonia-lyase was cultured in a solid state at 28° C. for 24 hours. The composition and content of the solid medium were: agar with a mass percentage of 2.5%, 12° Be malt extract powder , and the rest is water. Take one ring in the seed medium, the composition and mass percentage of the seed medium are: glucose 2.0%, peptone 2.0%, yeast extract 0.5%, K 2 HPO 4 0.2%, NaCl0.6%, and the rest is water. 25 DEG C, under 150rpm shaker culture 24 hours, move in the enzyme-producing medium with 10% inoculum size, the composition and mass percentage content of producing enzyme medium are: glucose 0.5%, corn steep liquor 5.0%, bean cake powder 3.0% %, NaCl 0.6%, K 2 HPO 4 0.2%, L-Phe 0.1%, pH5.5, the rest is water. 25° C., cultured on a shaker at 150 rpm for 24 hours, centrifuged and washed twice with saline, and the obtained PAL-containing bacterium was used ...

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Abstract

The invention aims to provide an enzymatic preparation method of <15>NL-Phenylalanine. <15>N-ammonium sulfate and trans-cinnamic acid react to produce <15>NL-Phenylalanine under the catalyst of phenylalanine ammonia-lyase. The method of the invention has the advantages of the simple synthesis process, the higher output, compared with organic synthesis method and enzymatic method, achieving the easy substitution of N isotope, resulting in the product purity at 99 percent, the yield efficiency at 71 percent, the isotopic abundance at higher than 98 percent, the recovery rate of <15>N at 88 percent and being favorable for the large-scale production.

Description

Technical field: [0001] The invention relates to an enzymatic preparation 15 NL-Phenylalanine method. It belongs to the fields of food, medicine and chemical industry. Background technique: [0002] 15 NL-Phenylalanine is L-Phenylalanine 14 N isotope 15 The stable labeled compound substituted by N is non-radioactive and can be used without time limit. Compared with radioactive amino acids, it is safe and convenient, especially suitable for infants and young children, and is an excellent tracer. Since L-phenylalanine is one of the essential amino acids for the human body, it can be used as the basic unit of protein synthesis to synthesize enzymes and hormones with important biological activities. Therefore, using 15 As a tracer, NL-phenylalanine plays a key role in uncovering the physical and chemical processes in organisms and cells and understanding the metabolism of organisms by using the metabolism of amino acids. [0003] Mosoni et al. [20] reported [U- 14 C] Ph...

Claims

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Application Information

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IPC IPC(8): C12P13/22C12R1/85
Inventor 袁其朋岳海燕汪文川
Owner BEIJING UNIV OF CHEM TECH
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