Chiral method for preparing D-ornithine and putrescine or derivatives thereof

A derivative, ornithine technology, applied in the field of biotransformation and separation of chiral compounds, can solve the problems of inconvenient to obtain polyamine compounds, inability to prepare D-ornithine, environmental pollution, etc., and achieves short conversion time and low price. , the effect of reducing production costs

Inactive Publication Date: 2008-08-06
XUZHOU MEDICAL COLLEGE
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] There are a large number of synthetic routes for the synthesis of putrescine, and all these chemical routes have the following disadvantages: the starting materials are obtained from chemical raw materials that are considered to be non-renewable, and will cause environmental pollution
[0013] However, both methods have the disadvantage of not being able to produce D-ornithine, and polyamines are generally considered to be toxic to any cells or microorganisms used in biochemical production.
Therefore, until now, it is still inconvenient to obtain polyamine compounds through the method of biological fermentation
[0014] So far, using L-arginine or part of racemic arginine or its derivatives as racemization reaction raw material, DL-arginine or its derivatives are obtained through racemization reaction, followed by lysine decarboxylase or Microbes or enzymes of ornithine decarboxylase carry out biotransformation, and the method for obtaining D-ornithine, putrescine and derivatives thereof has not been reported in the literature

Method used

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  • Chiral method for preparing D-ornithine and putrescine or derivatives thereof
  • Chiral method for preparing D-ornithine and putrescine or derivatives thereof
  • Chiral method for preparing D-ornithine and putrescine or derivatives thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] 1. Put the Hafnia alvei AS 1.1009 bacterial strain in the following 1000mL culture medium: 2% sucrose, 1% peptone, 2% corn steep liquor, 0.1% yeast extract, (NH 4 ) 2 SO 4 0.1%, L-Lysine 1%. The fermentation conditions are as follows: temperature 35°C, inoculum size 6%, medium loading volume 100mL, fermentation time 14h. Wet cells were obtained by centrifugation at 4000rpm for 15min.

[0044] 2. Add cells to 600mL transformation solution containing 10% DL-ornithine obtained by racemization reaction, add 6mL 0.5% Tween-80 and 600mL pH8.0 phosphate buffer, and react at 37°C 24h. .

[0045] 3. Concentrate the conversion solution to 200 mL, add 2 g of activated carbon for decolorization. Continue to concentrate the decolorized solution to 100mL, add 200mL of 95% ethanol and stir evenly, cool and crystallize, filter and dry in vacuum to obtain a dry weight of 37.5g of crystals, [α] D 20 =-11.5 (C=5.5, H 2 (0), the gained mother liquor is adsorbed by JK008 cationic ...

Embodiment 2

[0047] 1. Select the Hafnia alvei AS 1.1009 bacterial strain, the seed culture and fermentation conditions are the same as in Example 1, and wet cells are obtained.

[0048] 2. Add the cells to 600mL transformation solution containing 10% DL-ornithine obtained by racemization reaction, add 6mL 0.1% cetyltrimethylammonium bromide and 600mL pH7.0 diphosphate Sodium hydrogen-citric acid buffer solution, react at 30°C for 36h.

[0049] 3. Concentrate the conversion solution to 200 mL, add 2 g of activated carbon for decolorization. Continue to concentrate the decolorized solution to 100mL, add 200mL of 95% ethanol and stir evenly, cool and crystallize, filter and dry in vacuo to obtain 30.1g of crystal dry weight, [α] D 20 =-10.8 (C=5.5, H 2 (0), the obtained mother liquor is adsorbed by JK008 cationic resin, eluted with 3% ammonia water, and the eluent is decolorized by activated carbon and then concentrated, and the concentrated solution is adjusted to pH 10 to obtain putresc...

Embodiment 3

[0051] 1. Put the Hafnia alvei AS 1.1009 strain in the following 1000mL medium: 2% glucose, 1% soybean cake hydrolyzate, 2% corn steep liquor, 0.1% yeast extract, (NH 4 ) 2 SO 4 0.1%, L-ornithine 0.5%. The fermentation conditions are as follows: a temperature of 35° C., an inoculum size of 5%, a culture medium loading capacity of 100 mL, and a fermentation time of 20 h. Wet cells were obtained by centrifugation at 4000rpm for 15min.

[0052] 2. Add the cells to 600mL of transformation solution containing 10% DL-ornithine obtained by racemization reaction, add 6mL of 0.5% Tween-80 and 600mL of pH9.0 borax-hydrochloric acid buffer solution, 40°C Reaction 48h.

[0053] 3. Concentrate the conversion solution to 200 mL, add 2 g of activated carbon for decolorization. Continue to concentrate the decolorized solution to 100mL, add 200mL of 95% ethanol and stir evenly, cool and crystallize, filter and vacuum dry to obtain a crystal dry weight of 35.5g, [α] D 20 =-11.2 (C=5.5, ...

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Abstract

The invention belongs to the technical field of chiral compounds biological transformation separation, in particular relates to a method for preparing chiral organic coumounds and a method for preparing D-ornithine and putrescine, and is also suitable for preparing other D-ornithine compounds and aliphatic amine. The method for preparation comprises: mixing L-arginine or salt of the L-arginine with salicylal or derivant of the salicylal according to proportion, dissolving in alkaline or acidic dissolvant, obtaining DL-ornithine after racemization reaction, taking the DL-ornithine, biotransforming with microorganism or enzyme which contains lysine decarboxylase or ornithine decarboxylase under the temperature from 25DEG C to 45DEG C, obtaining the D-ornithine and the putrescine, then, using an isoelectric point crystallization method or a cation exchange resin separation method to separate the D-ornithine and the putrescine, and obtaining optically pure D-ornithine and putrescine. A chemical racemization method of the invention main takes water as the dissolvant, the catalyst consumption is little, the dissolvant which is used does not have poison and has low cost, the microorganism is easily cultured, and the invention is suitable for large scale industrial production.

Description

technical field [0001] The invention belongs to the technical field of biotransformation and separation of chiral compounds, and in particular relates to a preparation method of chiral organic compounds, especially a method for preparing D-ornithine and putrescine, which is also applicable to other chiral D-type Preparation of amino acid compounds and aliphatic or aromatic amines. Background technique [0002] D-type amino acids have important physiological functions. With the scientific development of the pharmaceutical industry, food industry and peptide synthesis, the demand for optically pure amino acids has increased dramatically. But D-Ornithine (D-Ornithine, D-Orn) has few production methods and high production cost. It is difficult to obtain D-ornithine by fermentation, and the amino acids synthesized by chemical synthesis are mostly racemates (DL type), which must be resolved. Enzymatic biotransformation of DL-amino acids using decarboxy...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P13/10C12P13/00C12P41/00
Inventor 刘毅印晓星刘莉张玲刘玲林奇泗
Owner XUZHOU MEDICAL COLLEGE
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