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Preparation method of stable isotop labelled 15N-L-asparagic acid

A stable isotope, aspartic acid technology, applied in the direction of fermentation, to achieve the effect of improving extraction process, improving market competitiveness and improving product quality

Inactive Publication Date: 2008-06-25
SHANGHAI RES INST OF CHEM IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a stable isotope-labeled 15 Process for the preparation of N-L-aspartic acid by enzymatic production using free whole cells 15 The technical route of N-L-aspartic acid aims to ease the 15 The problem of decreased N abundance, and try to improve 15 N Utilization

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The Escherichia coli cultured on the slant for 18 hours was inserted into the fermentation medium, filled with 50ml medium in a 500ml Erlenmeyer flask, and cultivated in a constant temperature shaker at 30°C for 20 hours with a rotation speed of 180rpm. Cultivate the obtained fermented liquid and centrifuge at 4800rpm for 30 minutes, pour off the supernatant, and add 0.5 g of the obtained wet thalline to the substrate concentration of 10% and the abundance of 10.39at%. 15 In 100 ml of a substrate solution of 7.7% by weight of N-ammonium sulfate and 0.01% by weight of magnesium sulfate, the pH value was adjusted to 7.9 with NaOH, and the reaction was carried out at a constant temperature of 35° C. for 23 hours. The obtained reaction solution was centrifuged, and after the supernatant was concentrated to pH 7.5 by ammonia, 0.2 gram of activated carbon was added to decolorize for 30 minutes to obtain a colorless solution, which was concentrated to 25 ml, and sulfuric acid w...

Embodiment 2

[0033] Get 2.0 grams of wet thalline cultured as in Example 1, add to the substrate concentration and be 11.6% by weight, containing abundance is 99.53at% 15 In 200 ml of a substrate solution of 7.4% by weight of N-ammonium chloride and 0.01% by weight of magnesium sulfate, the pH value was adjusted to 9.0 with 10M NaOH, and the reaction was carried out at a constant temperature of 39° C. for 27 hours. The obtained reaction solution was centrifuged, and the supernatant was concentrated to pH 6.5 with ammonia, and 0.6 g of active carbon was added to decolorize for 35 minutes to obtain a colorless solution, which was concentrated to 55 ml, and hydrochloric acid was added to adjust the pH to 3.00, and a large amount of white crystals were precipitated. Cooling, suction filtration and washing the next day, to obtain snow-white crystals, drying to obtain 23.65 grams of product, with an abundance of 99.18 at%, which is only 0.35 at% absolute and 0.35 at% relative to the raw material,...

Embodiment 3

[0035] The Escherichia coli cultured on the slant for 20 hours was inserted into the fermentation medium, 50ml of the medium was filled in a 500ml Erlenmeyer flask, and cultured on a constant temperature shaker at 32°C for 24 hours at a rotation speed of 200rpm. Cultivate the obtained fermented liquid and centrifuge at 4800rpm for 30 minutes, pour off the supernatant, and add 0.5 g of the obtained wet thalline to the substrate concentration of 10% and the abundance of 98.50at%. 15 In 100 ml of a substrate solution of 8% by weight of N-ammonium sulfate and 0.01% by weight of magnesium sulfate, the pH value was adjusted to 8.3 with NaOH, and the reaction was carried out at a constant temperature of 37° C. for 25 hours. The obtained reaction solution was centrifuged, and the supernatant was concentrated to pH 6.8 by catching up with ammonia, and after adding 0.25 g of active carbon for decolorization, a colorless solution was obtained for 40 minutes, concentrated to 20 ml, added s...

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Abstract

The invention relates to a 15N-L-aspartic acid manufacture method marked by stable isotopes. It includes the following technology: fermenting the microorganism Escherichia coli ATCC 11303 or CGMCC 1.881; slant cultivating; fermenting cultivation; enzyme reacting, distilling and recycling 15N. The method lowers the producing cost, improves the product quality and has high distill ratio.

Description

technical field [0001] The present invention relates to a 15 The preparation method of N-L-aspartic acid, specifically relates to a kind of free whole cell of Escherichia coli as enzyme source to prepare stable isotope labeling 15 The method of N-L-aspartic acid belongs to the field of stable isotope labeling compound production, and involves microbial enzymes and biological downstream separation technology. Background technique [0002] because 15 N than normal 14 N has one mass unit more, and its nuclear spin I=1 / 2, has nuclear magnetic resonance signal, so its mass effect and isotope effect can be used to obtain valuable structural information by means of mass spectrometry and other testing techniques. Therefore, non-radioactive stable isotope-labeled amino acids have been widely used because of their unique tracer effect. 15 N-labeled amino acids are indispensable tracers for studying human protein metabolism and individual amino acid metabolism, as well as transamin...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P13/20
Inventor 侯静华周震李良君杜晓宁宋明鸣
Owner SHANGHAI RES INST OF CHEM IND