WO2006123778A1 - Procede de fabrication de carbonate de cadaverine, et procede de fabrication de polyamide l'utilisant - Google Patents

Procede de fabrication de carbonate de cadaverine, et procede de fabrication de polyamide l'utilisant Download PDF

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WO2006123778A1
WO2006123778A1 PCT/JP2006/310037 JP2006310037W WO2006123778A1 WO 2006123778 A1 WO2006123778 A1 WO 2006123778A1 JP 2006310037 W JP2006310037 W JP 2006310037W WO 2006123778 A1 WO2006123778 A1 WO 2006123778A1
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cadaverine
carbonate
lysine
producing
solution
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PCT/JP2006/310037
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Japanese (ja)
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Masakazu Sato
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Ajinomoto Co., Inc.
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Priority claimed from JP2005147171A external-priority patent/JP2008193898A/ja
Priority claimed from JP2005147172A external-priority patent/JP2008193899A/ja
Application filed by Ajinomoto Co., Inc. filed Critical Ajinomoto Co., Inc.
Publication of WO2006123778A1 publication Critical patent/WO2006123778A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P13/00—Preparation of nitrogen-containing organic compounds
    • C12P13/001—Amines; Imines

Definitions

  • the present invention relates to a method for producing cadaverine carbonate.
  • the present invention also relates to a process for producing cadaverine dicarbonate or cadaverine.
  • the invention further relates to a process for producing polyamide.
  • Petroleum fuels such as naphtha are often used as raw materials for so-called plastic production.
  • Non-petroleum raw material power Various types of plastics, including polylactic acid, are being studied. These plastics have different physical properties such as heat resistance depending on their raw materials. Of these, the development of plastics derived from non-petroleum materials with particularly high heat resistance is suitable for the use of polylactic acid under high temperature conditions! /, Point power.
  • polyamide rosin is polyamide rosin.
  • V ⁇ is used in a large amount of hexamethylene diamine, which is a 6-carbon diamine, and adipic acid, which is a 6-carbon dicarboxylic acid.
  • Nylon 66 which is a polymer having a molar ratio of 1: 1.
  • hexamethyldiamine is produced from benzene, propylene, or butanegen, which also provides naphtha power, and production methods from non-petroleum materials are known!
  • pentamethylenediamine having 5 carbon atoms is also called cadaverine and is known to be produced from lysine, one of the amino acids, by lysine decarbonase (LDC) (Non-Patent Document 1). ). Therefore, it can be used under high temperature conditions using non-petroleum raw materials by producing polyamide resin using cadaverine having 5 carbon atoms as raw material instead of hexamethylenediamine having 6 carbon atoms. It is possible to supply plastic materials. Although cadaverine is expected to be used for pharmaceutical intermediates in addition to polyamide sallow, its price is high. Therefore, development of an inexpensive manufacturing method is indispensable for further spread.
  • adipic acid which is a dicarboxylic acid, which is often an inorganic or organic acid such as hydrochloric acid, sulfuric acid or phosphoric acid as a neutralizing agent for pH adjustment
  • Patent Documents 3 and 4 the ability of cadaverine salt to be generated
  • the cadaverine salt power that is produced also produces a by-product of the acid-derived salt added as a neutralizing agent when purifying cadaverine, giving it a significant environmental impact.
  • dicarboxylic acids such as adipic acid are added when cultivating lysine-fermenting microorganisms, and the resulting lysine / dicarboxylic acid solution is neutralized by acting lysine decarboxylase.
  • cadaverine dicarboxylic acid Patent Document 3
  • Non-Patent Document 2 a method in which polycondensation is carried out by heating cadaverine dicarboxylate under melting conditions.
  • cadaverine dicarboxylate it may be possible to produce cadaverine dicarbonate by cadaverine salt exchange obtained by using inorganic acid or organic acid as neutralizing agent.
  • a by-product salt derived from the neutralizing agent is generated, which becomes an environmental burden.
  • this by-product salt is recovered and reused, a large amount of recovery equipment is required.
  • dicarboxylate is used as a neutralizing agent, cadaverine dicarboxylate can be obtained directly.
  • organic solvent crystallization is required for the purification process. It is necessary to reinforce the equipment when collecting the waste.
  • Patent Document 1 JP 2002-223770 A
  • Patent Document 2 JP 2002-223771
  • Patent Document 3 Japanese Patent Application Laid-Open No. 2004-208646
  • Patent Document 4 JP-A-2005-006650
  • Non-Patent Document 1 Enzyme Handbook First Edition 636 pages Asakura Shoten
  • Non-Patent Document 2 Introduction to New Polymer Chemistry, page 22, Chemistry Doujin
  • the present invention minimizes by-product salt generated in the production process of cadaverine carbonate, reduces capital investment for recovery of by-product salt and organic solvent, and provides inexpensive and efficient cadaverine carbonate. It is an object to provide a manufacturing method. Another object of the present invention is to provide a method for producing cadaverine carbonate power cadaverine or cadaverine dicarboxylate, and to provide a method for producing polyamides inexpensively and efficiently using these.
  • cadaverine carbonate can be produced efficiently by carrying out enzymatic decarboxylation of lysine while supplying carbon dioxide to produce cadaverine carbonate such as lysine carbonate.
  • cadaverine dicarboxylate can be efficiently produced by adding dicarboxylic acid to the obtained cadaverine carbonate.
  • cadaverine can be efficiently produced by concentrating the obtained cadaverine carbonate and releasing carbonate ions or hydrogen carbonate ions, which are counter ions of cadaverine, as diacid carbon.
  • the present invention is as follows.
  • Enzymatic decarboxylation of lysine was performed while adding diacid-carbon to an aqueous solution of lysine carbonate so that the pH of the aqueous solution was maintained at a pH suitable for enzymatic decarboxylation of lysine.
  • a method for producing cadaverine carbonate comprising producing cadaverine carbonate.
  • Cadaverine carbonate is produced by any one of the methods (1) to (9), and cadaverine dicarboxylate is formed by adding dicarboxylic acid to the resulting aqueous solution of daverine carbonate.
  • a process for producing cadaverine dicarboxylate comprising:
  • a method for producing polyamide comprising producing cadaverine dicarboxylate by any one of the methods (10) to (12) and polycondensing the obtained cadaverine dicarboxylate.
  • a method for producing cadaverine which comprises producing cadaverine carbonate by the method according to any one of (1) to (9), and concentrating an aqueous solution of the obtained force daverine carbonate to produce cadaverine.
  • a method for producing polyamide comprising producing cadaverine by the method of (14) and polycondensing the obtained cadaverine with a dicarboxylic acid.
  • the method for producing cadaverine carbonate of the present invention comprises adding lysine carbon to an aqueous solution of lysine carbonate while adding diacid-carbon so that the pH of the solution is maintained at a pH suitable for enzymatic decarboxylation of lysine. Carrying out an enzymatic decarboxylation reaction of to produce cadaverine carbonate.
  • lysine may be L-lysine or D-lysine as long as it generates cadaverine by enzymatic decarboxylation, but L-lysine is usually good.
  • carbonate includes both carbonate and bicarbonate.
  • an aqueous solution of lysine carbonate is used.
  • the lysine salt contained in this aqueous solution need not be 100% free lysine carbonate, and may partially contain other lysine salts such as lysine hydrochloride and lysine sulfate.
  • An aqueous solution of lysine carbonate can be obtained, for example, by dissolving lysine carbonate in water.
  • a lysine carbonate fermentation solution (see, for example, JP-A-2002-65287) can also be used as an aqueous lysine carbonate solution.
  • a free lysine base (lysine base) may be dissolved in water, and diacid carbon (CO 2) may be added to this aqueous solution to form an aqueous lysine carbonate solution.
  • diacid carbon CO 2
  • the free lysine base may be a purified lysine base because of the raw material, or may be liquid lysine for feed (Japanese Patent Laid-Open No. 2000-256290).
  • a highly purified raw material with a small amount of compounds other than carbonate ions and a small amount of compounds other than lysine is preferred.
  • An isolated and purified free lysine base is more preferable.
  • a lysine fermentation broth obtained by culturing microorganisms (see, for example, WO95 / 016042, WO95 / 023864, WO96 / 040934, WO00 / 056858, WO00 / 077172, WO01 / 002547, or WO01 / 053459)
  • a lysine carbonate aqueous solution may also be used.
  • the decarboxylation reaction is performed using the lysine carbonate solution prepared as described above.
  • Lysine decarboxylation is performed by adding lysine decarboxylase (LDC) to the lysine carbonate solution.
  • LDC lysine decarboxylase
  • the LDC is not particularly limited as long as it acts on lysine to produce cadaverine.
  • LDC a purified enzyme may be used, or various cells such as microorganisms, plant cells or animal cells that produce LDC may be used.
  • the number of LDCs or cells producing them may be one type or two or more types.
  • Examples of the LDC protein include a protein having the amino acid sequence of SEQ ID NO: 12, or an amino acid sequence in which one or several amino acids are substituted, deleted, inserted or added in the amino acid sequence of SEQ ID NO: 12. And a protein having an activity of decarboxylating lysine.
  • a protein having an activity of decarboxylating lysine preferably 1 to 50, preferably 1 to 20, more preferably 1 to LO.
  • the activity of LDC can be measured according to a known method.
  • the above substitution of amino acid residues in LDC is a conservative substitution so that the activity of LDC protein is maintained.
  • a substitution is a change in which at least one residue in the amino acid sequence is removed and another residue is inserted therein.
  • the amino acids that replace the original amino acids of the LDC protein and are considered conservative substitutions are: Ala to Ser or Thr, Arg to Gln, His or Lys, Asn to Glu, Gln, Lys , His or Asp substitution, Asp to Asn, Glu or Gin substitution, Cys to Ser or Ala substitution, Gin to Asn, Glu, Lys, His, Asp or Arg substitution, Glu to Gly, Asn, Gln, Lys or Asp substitution, Gly to Pro substitution, His to Asn, Lys, Gln, Arg or Tyr substitution, lie to Leu, Met, Val or P he substitution, Leu to Ile , Met, Val or Phe, Lys to Asn, Glu, Gln, His or Arg, Met to
  • the LDC protein has an activity to decarboxylate lysine, it is 80% or more, preferably 90% or more, more preferably 95%, particularly preferably 98, with the amino acid sequence of SEQ ID NO: 12. It may be a protein having a homologous amino acid sequence of more than%. Amino acid sequence homology was determined using, for example, the algorithm BLAST (Pro. Natl. Acad. Sci. US A, 90, 5873 (1993) by Karlin and Altschul and FASTA (Methods EnzymoL, 183, 63 (1990) by Pearson. can do.
  • BLAST Pro. Natl. Acad. Sci. US A, 90, 5873 (1993) by Karlin and Altschul and FASTA (Methods EnzymoL, 183, 63 (1990) by Pearson. can do.
  • cells that express LDC may be used as they are.
  • a cell-treated product containing LDC may be used.
  • the cell treatment product include a cell culture solution, a cell disruption solution, and a fraction thereof.
  • an enzyme reaction is performed using cells such as microorganisms, plant cells, or animal cells
  • use of cells treated with organic solvents or surfactants may improve the permeability of the substrate and improve the reactivity. It is generally known.
  • the reactivity can be enhanced by treating the cells producing LDC with an organic solvent or a surfactant.
  • Triton X-100 Tween 20, sodium cholate and CHAPS can be used as the surfactant to be treated, and acetone, xylene and toluene can be used as the organic solvent. More specifically, when Triton X-100 is used, a concentration of 0.01% to 1.0% (wZv) is added, and treatment at 0 ° C to 37 ° C for 2 minutes to 1 hour is appropriate. .
  • microorganism examples include Escherichia bacteria such as E. coli, coryneform bacteria such as Brevibacterium lactofermentum, Bacillus bacteria such as Bacillus subtilis, Serratia marcescens (Serratia marcescens) of Serratia bacteria such as bacteria include eukaryotic cells such as Saccharomyces' cerevisiae (Saccharomyces cervis i ae). Of these, bacteria, particularly E.coK, are preferred.
  • the microorganism may be a wild strain or a mutant strain as long as it produces LDC.
  • Recombinant cells modified to increase LDC activity include, for example, increasing the copy number of the gene encoding LDC or regulating the expression of the gene so that the expression of the gene is enhanced. Examples include recombinant cells that have been modified so that LDC activity is increased by modifying the sequence.
  • LDC As a gene encoding LDC, for example, it has the base sequence of SEQ ID NO: 11 of E. coli DNA can be used. As long as it encodes a protein having LDC activity, a DNA that hybridizes with a polynucleotide having a complementary sequence of the nucleotide sequence of SEQ ID NO: 11 under stringent conditions can also be used.
  • stringent conditions refers to conditions under which so-called specific hybrids are formed and non-specific hybrids are not formed. Specifically, it is a condition for washing normal Southerno and Hybridisation60. C, 1 X SSC, 0.1% SDS, preferably 60. C, 0.1 X SSC, 0.1% SDS, more preferably 68. The conditions include washing at a salt concentration and temperature corresponding to C, 0.1 X SSC, 0.1% SDS, more preferably 2 to 3 times.
  • a gene encoding LDC can be obtained by PCR using a nucleotide sequence-specific primer or probe of the LDC gene or a hybridization method.
  • Increasing the number of copies of the gene encoding LDC can be achieved by, for example, transforming a cell with a plasmid containing the gene encoding LDC, or integrating the gene encoding LDC into the host cell chromosome by homologous recombination. This can be done by dragging.
  • the plasmid for introducing the gene encoding LDC is not particularly limited as long as it has replication ability in the host cell. For example, in the case of Escherichia coli, for example, pSTV 29 (manufactured by Takara Bio Inc.). ), RSF1010 (Gene vol.
  • vectors that function in coryneform bacteria include pAM330 (Japanese Patent Laid-Open No. 58-067699), pHM1519 (Japanese Patent Laid-Open No. 58-77895), and pSFK6 (Japanese Patent Laid-Open No. 2000-262288).
  • Transformation using a plasmid and homologous recombination can be performed according to known methods.
  • the expression regulatory region of the LDC gene to be introduced may be modified.
  • expression control regions include promoters, and strong promoters include, for example, lac promoter, trp promoter, tr c promoter, tac promoter, lambda phage PR promoter, PL promoter, tet promoter, amyE promoter, Examples include a spac promoter and an acid phosphatase promoter.
  • mutant strains with enhanced LDC activity may be used for the decarboxylation reaction.
  • a mutant strain is obtained by subjecting a parent strain or a wild strain to normal mutation treatment, that is, irradiation with X-rays or ultraviolet rays, Or N-methyl ⁇ 'nitro ⁇ nitrosoguanidine, etc., treated with a mutagen, etc., and obtained from the obtained mutant strain by selecting a strain with enhanced LDC activity be able to.
  • Culture for obtaining LDC protein or microorganisms or cells with enhanced LDC activity may be performed by a method suitable for LDC production depending on the microorganisms or cells used.
  • the medium used for the culture may be a normal medium containing a carbon source, a nitrogen source, inorganic ions, and other organic components as required.
  • Carbon sources include sucrose, dulcose, latatoose, galactose, fructose, arabinose, maltose, xylose, trehalose, sugars such as ribose and starch-calyzed hydrolyzate, alcohols such as glycerol, mannitol and sorbitol, darcon Organic acids such as acid, fumaric acid, succinic acid and succinic acid can be used.
  • Nitrogen sources include inorganic ammonium salts such as ammonium sulfate, salt ammonia, and ammonium phosphate, organic nitrogen such as soybean hydrolysate, ammonia gas, and aqueous ammonia. Etc. can be used.
  • organic micronutrients it is desirable to contain an appropriate amount of required substances such as vitamins such as vitamin B1, nucleic acids such as adenine and RNA, or yeast sex.
  • vitamins such as vitamin B1
  • nucleic acids such as adenine and RNA
  • yeast sex a small amount of calcium phosphate, magnesium sulfate, iron ions, manganese ions, etc. is added as necessary.
  • the culture temperature is controlled to 20-45 ° C and the culture pH is controlled to 5.0-8.0.
  • inorganic or organic acidic or alkaline substances, ammonia gas, etc. can be used for pH adjustment.
  • an inducing agent is added to the medium.
  • the cells can be collected from the culture medium using a centrifuge or a separation membrane.
  • the cells may be used as they are, but when those treatments containing LDC are used, the cells can be disrupted by ultrasonication, French press or enzymatic treatment to extract the enzyme and used as an enzyme extract. .
  • LDC when purifying the LDC, LDC can be purified by using ammonium sulfate salting-out and various chromatographies according to conventional methods. Purified LDC can be obtained using a carrier. It can also be used in a state where it can be fixed or contacted with the reaction solution through a membrane or the like.
  • a decarboxylation reaction is carried out using the cells expressing LDC protein or LDC gene obtained as described above or a treated product thereof.
  • lysine carbonate as a substrate may be further added according to the progress of the reaction.
  • LDC protein and LDC gene-expressing cells and cell treatment solution may be added directly to the reaction solution at the start of the reaction, or may be added in portions according to the progress of the reaction.
  • the pH is adjusted to a pH suitable for the enzymatic decarboxylation reaction of lysine.
  • This pH is adjusted by adjusting the purity, flow rate and pressure of the enzyme reaction system.
  • the pH is usually 9.0 or less, preferably pH 5.0-9.0, more preferably pH 7.0-9.0.
  • Cadaverine is produced by decarboxylation of lysine. At this time, lysine, which is a monovalent cation, decarboxylation, cadaverine, which is a divalent cation, is converted into cadaverine, and the carbonic acid present in the aqueous solution becomes a counter ion, and cadaverine carbonate is obtained in the reaction solution.
  • reaction pH can be maintained within the above pH range, strict pH adjustment is not necessary during the decarboxylation reaction.
  • the carbon dioxide released from the lysine force is released as the reaction solution and the pH rises. Therefore, carbon dioxide is added to the reaction solution and adjusted so that the pH of the reaction solution falls within the above range.
  • the carbon dioxide to be added may be a gas, a liquid, or a solid (dry ice), but is preferably stored as a gas.
  • the diacid carbon may be a mixed gas containing other gases, but is preferably 100% pure carbon dioxide.
  • the carbon dioxide addition may be continuous or intermittent.
  • a neutralizing agent other than carbon dioxide may be added at the same time. However, in order not to generate a double salt, it is preferable that the neutralizing agent only contains carbon dioxide.
  • the reaction temperature of the decarboxylase reaction is preferably within the range where the enzyme reaction is maximized, the release of carbon dioxide from the solution is minimized, and the reaction pH is kept within the above pH range. 20-50 ° C, more preferably 25-45 ° C.
  • the enzymatic decarboxylation reaction can be accelerated by adding vitamin B6, which is a coenzyme.
  • vitamin B6 which is a coenzyme.
  • pyridoxine, pyridoxamine and pyridoxal phosphate may be preferable. More preferably pyridoxal phosphate (PLP)
  • PBP pyridoxal phosphate
  • the addition concentration is not particularly limited, but is preferably a concentration of O.lmM or more.
  • cadaverine is recovered.
  • Dicarboxylates can be produced. That is, cadaverine dicarboxylate is formed in the solution by a salt exchange reaction between carbonic acid and dicarboxylic acid. On the other hand, carbonate ions are released out of the system as carbon dioxide.
  • the dicarboxylic acid added at this time should be equimolar with cadaverine.
  • the dicarboxylic acid to be added is preferably a dicarboxylic acid having 4 to 10 carbon atoms!
  • dicarboxylic acid having 4 to 10 carbon atoms examples include succinic acid, maleic acid, fumaric acid, glutaric acid, adipic acid, sebacic acid, terephthalic acid, and isophthalic acid. More preferred are 6-carbon adipic acid or 8 carbon-terephthalic acid.
  • the obtained cadaverine dicarboxylate can be easily separated and recovered by, for example, the following method. First, after sterilization by centrifugation, decolorization is preferably performed by activated carbon treatment to remove PLP and impurities. Next, cadaverine dicarboxylate crystals can be obtained by concentration under reduced pressure.
  • Cadaverine can be produced by concentrating an aqueous solution of cadaverine carbonate obtained by the method for producing cadaverine carbonate of the present invention. Cadaverine can be easily separated and recovered by, for example, the following method.
  • the aqueous solution of cadaverine carbonate is decolorized by treatment with activated carbon to remove PLP and impurities after removing cells by centrifugation or the like.
  • concentration carbonate ions and hydrogen carbonate ions are released into the atmosphere as carbon dioxide and carbon dioxide, and cadaverine can be obtained after water evaporation.
  • Concentration is preferably performed under reduced pressure, and can be efficiently concentrated by heating.
  • the heating temperature is preferably 40 to 100 ° C.
  • the method of the present invention does not use a large amount of water in the refining process, does not generate by-product salt in the process of resin regeneration, Melting This is an excellent and simple method that does not use a medium.
  • Polyamides can be produced using cadaverine dicarboxylate or cadaverine obtained by the above method.
  • Examples of the polyamide production method include a method of performing a polycondensation reaction using cadaverine dicarboxylic acid salt and a method of performing a polycondensation reaction using cadaverine and dicarboxylic acid.
  • the polycondensation reaction can be carried out according to a known method (see, for example, “Plastic Materials Course [16] Polyamide resin” (Nikkan Kogyo Shimbun)).
  • cadaverine dicarboxylate is mixed with water and the mixture is heated and dehydrated for condensation.
  • cadaverine and dicarboxylic acid may be mixed with water, and the mixture may be polycondensed while being dehydrated by heating.
  • the molecular weight can be increased by solid phase polymerization.
  • Solid-phase polymerization proceeds by heating in a vacuum or in an inert gas in the temperature range from 100 ° C to the melting point, and polyamides with insufficient molecular weight can be converted to high molecular weight by heating polycondensation. .
  • polyamides can be produced depending on the type of dicarboxylic acid used for polycondensation.
  • the dicarboxylic acid that can be used for polycondensation is not particularly limited as long as it can be used for the production of polyamide.
  • succinic acid, maleic acid, fumaric acid, glutaric acid, adipic acid, sebacic acid, terephthalic acid And isophthalic acid For example, when adipic acid is used for polycondensation, 5,6-nylon can be obtained, and when terephthalic acid is used for polycondensation, 5, T-nylon can be obtained.
  • the pH of the L-lysine solution depends on the CO flow rate at the beginning of the reaction. The larger the addition flow rate, the lower the pH.
  • the solution should have enough CO dissolved in the solution to neutralize L-lysine.
  • E.coli-derived LDC gene (cadA) (N. Watson et al., Journal of bacteriolog y, (1992) vol. 174, 530-540; SY Neng and GN Bennet, Journal of bacteriology (19 92) vol 174, 2659-2669), and designed a PCR primer having the base sequence shown in 5, -gtcgacactgcacacggctggcgg-3, (SEQ ID NO: 1) and 5, -gttagcggcacgtacacctgcctgg-3, (SEQ ID NO: 2).
  • a DNA fragment containing cadA was amplified by PCR using the E. coli W3110 (ATCC39936) chromosome as a saddle.
  • the amplified DNA fragment was cleaved with Kpnl and Sphl, and the obtained fragment (2,468 bp) was inserted into the Kpnl and Sphl cleavage sites of pUC18 (Tacarano) to prepare plasmid pcadA.
  • reaction solution containing 1 unit at 94 ° C for 30 seconds, then 94 ° C for 15 seconds, 55 ° C for 30 seconds, 68 ° C for 2 minutes 30 seconds 25 Repeated PCR was performed to amplify the cadA gene portion.
  • PCR was performed under the same conditions using the above plasmid pEAM330 as the cage type, 5′-gctctagaattttttcaatg tgattt-3 ( ⁇ ⁇ ⁇ ⁇ 3 ⁇ 4 ⁇ No. 5) and a-gtgattcaatattgcaataacgttcatctacatttccttacggtgtta-. (SEQ ID NO: 6) oligonucleotide as primers.
  • the promoter sequence portion of acid phosphatase was amplified.
  • the reaction solution was subjected to agarose gel electrophoresis, and each amplified DNA fragment was recovered using a Microspin column (manufactured by Amersham's Pharmacia Biotech).
  • the amplified fragment mixture is made into a bowl shape, and oligonucleotides of SEQ ID NO: 3 and SEQ ID NO: 5 are used as primers, and 94 ° C for 15 seconds, 55 ° C for 30 seconds, 68 ° C in a reaction solution having the same composition.
  • PCR was repeated 25 times in a cycle of 2 minutes and 30 seconds to construct a chimeric enzyme gene.
  • Each amplified DNA fragment was recovered using a Microspin column (Amersham 'Farmasia' Biotech) and digested with Xbal and Pstl. This was ligated to the Xbal-Pstl site of plasmid pUC119.
  • the CadA expression plasmid was constructed by force and named pcadA202.
  • plasmid pcadA202 is used as a cocoon, and as a primer, 5, -ggacatataacaccgtaagg PCR was performed using aggaatgtagatgaacgttattgc-3, (self-sequence number 7) and 5, -gcaataacgttcatctacattcctccttacggt gttatatgtcc-3, (SEQ ID NO: 8) oligonucleotide according to the method described in the manual to construct plasmid pcadA210.
  • PCR was performed using the plasmid pcadA210 as a saddle type, and using 5, -gaattttttcaatgtgattttg acatttacttccagatgac- ⁇ (item ti column 3 ⁇ 4 ⁇ No. 9) and 5-gtcatctggaagtaaatgtcaaaatcacattgaaaa attc-3 ′ (SEQ ID NO: 10) oligonucleotide as primers.
  • a plasmid was constructed. This CadA high expression plasmid was named pcadA220.
  • pcadA220 is designed to enhance LDC expression by modifying the constitutive expression promoter and ribosome binding site of the acid phosphatase gene of the genus Enterobacter.
  • E. coli JM109 strain (Takara Bio) was transformed with plasmid pcadA220, and the resulting transformant was named Escherichia coli cadA220.
  • the constitutive expression promoter of the Enterobacter genus acid phosphatase gene and the ribosome binding site were modified and succeeded in higher expression of LDC.
  • Escherichia coli cadA220 strain is inoculated on LB medium plate for 1 ase and cultured at 26 ° C for ⁇ , and then cultured cells are sown for 1 ase and inoculated into 50 mL of liquid LB medium at 28 ° C. C. Shaking culture was performed for 8 hours under conditions of 150 rpm, and a culture solution was obtained in advance.
  • the pre-culture solution obtained was inoculated into 10 mL of the pre-culture medium shown below, and pre-cultured at a total volume of 300 mL under the conditions of 28 ° C, 700 rpm, pH 7.0, aeration rate of 300 mL / min. .
  • Ammonia was used to adjust the culture pH.
  • the culture was terminated to obtain a preculture solution.
  • the medium was mixed and adjusted to pH 5.0 with KOH aqueous solution.
  • the main culture was performed at 300 ° C., 30 ° C., 700 rpm, pH 7.0, and aeration rate of 300 mL / min.
  • the culture pH was adjusted with ammonia, and when the sugar consumption in the main culture medium was completed, the culture was terminated to obtain Escherichia coli cadA220 cells.
  • the obtained Escherichia coli cadA220 strain was subjected to the pretreatment shown below before the enzyme reaction.
  • the bacterial cell culture solution was centrifuged at 6,000 rpm for 10 minutes at 4 ° C to obtain precipitated bacterial cells.
  • a 1/5 volume of 0.1% Triton X_100, 0.2 M Tris-HCl (pH 7.4) solution of 1/5 of the culture solution used for the centrifugation was added to the precipitated cells to suspend the precipitated cells uniformly. After suspending, the solution was ice-cooled for 10 minutes to obtain an enzyme solution.
  • the obtained cadaverine adipic acid solution was centrifuged (12,000 rpm, 10 minutes) to remove the cells and their residues, and a supernatant fraction was obtained.
  • cadaverine adipate solution 200 mL of the obtained cadaverine adipate solution was concentrated under reduced pressure using an evaporator. By this operation, water in the solution was removed and finally 64.0 g of cadaverine adipate was obtained.
  • the cadaverine carbonate solution obtained in Example 2 was first centrifuged at 12,000 rpm for 10 minutes at 4 ° C. The microbial cells and microbial cell residues in the reaction solution were removed, and the supernatant fraction was obtained.
  • Cadaverine was purified by concentrating 300 mL of the obtained cadaverine carbonate solution under reduced pressure at 40 ° C. using an evaporator. This operation removed carbon dioxide and water from the solution, and 28.7 g of cadaverine was finally obtained.

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  • Polyamides (AREA)

Abstract

L'invention concerne un carbonate de cadavérine, fabriqué en utilisant du carbonate de lysine en tant que substrat, en ajustant le pH par ajout de dioxyde de carbone, puis en effectuant une décarboxylation enzymatique de la lysine. Un sel d'acide dicarboxylique est ajouté au carbonate de cadavérine résultant, et un sel d'acide dicarboxylique de cadavérine est obtenu par une étape d'isolation, après une réaction d'échange de sel avec de l'acide carbonique. La cadavérine est obtenue en concentrant une solution de carbonate de cadavérine, afin d'éliminer le dioxyde de carbone du système. Un polyamide est obtenu en utilisant la cadavérine ou le sel d'acide dicarboxylique de cadavérine résultant.
PCT/JP2006/310037 2005-05-19 2006-05-19 Procede de fabrication de carbonate de cadaverine, et procede de fabrication de polyamide l'utilisant WO2006123778A1 (fr)

Applications Claiming Priority (4)

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JP2005147171A JP2008193898A (ja) 2005-05-19 2005-05-19 カダベリンの製造法
JP2005-147171 2005-05-19
JP2005-147172 2005-05-19
JP2005147172A JP2008193899A (ja) 2005-05-19 2005-05-19 リジン炭酸塩を用いたカダベリンジカルボン酸塩の製造法

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Cited By (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008084849A1 (fr) * 2007-01-11 2008-07-17 Mitsubishi Chemical Corporation Sel de cadavérine, solution aqueuse de sel de cadavérine, résine de polyamide, article moulé et procédé de fabrication du sel de cadavérine et d'une solution aqueuse de sel de cadavérine
JP2008189661A (ja) * 2007-01-11 2008-08-21 Mitsubishi Chemicals Corp カダベリン塩水溶液の製造方法、カダベリン塩水溶液、カダベリン塩、ポリアミド樹脂及び成形品
JP2008187963A (ja) * 2007-02-06 2008-08-21 Mitsubishi Chemicals Corp カダベリン及び／又はカダベリン塩の溶液及びその製造方法
JP2008220195A (ja) * 2007-03-08 2008-09-25 Mitsubishi Chemicals Corp カダベリン及び／又はカダベリン塩の製造方法
WO2010002000A1 (fr) * 2008-07-03 2010-01-07 三菱化学株式会社 Procédé de fabrication de pentaméthylènediamine, et procédé de production de résine de polyamide
US20100292429A1 (en) * 2008-01-23 2010-11-18 Basf Se Method for Fermentatively Producing 1,5-Diaminopentane
JP2010275516A (ja) * 2008-07-03 2010-12-09 Mitsubishi Chemicals Corp 精製ペンタメチレンジアミンの製造方法及びポリアミド樹脂の製造方法
EP2415801A1 (fr) * 2009-03-30 2012-02-08 Toray Industries, Inc. Résine de polyamide, composition de résine de polyamide et article moulé le comprenant
CN105164101A (zh) * 2014-04-04 2015-12-16 Cj第一制糖株式会社 有机胺的精制方法
WO2016106367A1 (fr) 2014-12-23 2016-06-30 Genomatica, Inc. Procédé de production et de traitement de diamines
US20190322995A1 (en) * 2016-05-16 2019-10-24 Ningxia Eppen Biotech Co., Ltd Method for fermentation-production of pentanediamine comprising carbon dioxide stripping technique

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2002223771A (ja) * 2001-02-01 2002-08-13 Toray Ind Inc カダベリンの製造方法
JP2003000294A (ja) * 2000-08-21 2003-01-07 Showa Denko Kk 微生物によるｌ−フェニルアラニン誘導体の製造方法
JP2005006650A (ja) * 2003-05-26 2005-01-13 Ajinomoto Co Inc カダベリン・ジカルボン酸塩の製造法

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003000294A (ja) * 2000-08-21 2003-01-07 Showa Denko Kk 微生物によるｌ−フェニルアラニン誘導体の製造方法
JP2002223771A (ja) * 2001-02-01 2002-08-13 Toray Ind Inc カダベリンの製造方法
JP2005006650A (ja) * 2003-05-26 2005-01-13 Ajinomoto Co Inc カダベリン・ジカルボン酸塩の製造法

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
MENG S-Y. ET AL.: "Nucleotide sequence of Escherichia coli cad Operon: a System for Neutralization of Low Extracellular pH", J. BACTERIOL., vol. 174, no. 8, 1992, pages 2659 - 2669, XP000978358 *

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JP2008189661A (ja) * 2007-01-11 2008-08-21 Mitsubishi Chemicals Corp カダベリン塩水溶液の製造方法、カダベリン塩水溶液、カダベリン塩、ポリアミド樹脂及び成形品
WO2008084849A1 (fr) * 2007-01-11 2008-07-17 Mitsubishi Chemical Corporation Sel de cadavérine, solution aqueuse de sel de cadavérine, résine de polyamide, article moulé et procédé de fabrication du sel de cadavérine et d'une solution aqueuse de sel de cadavérine
JP2008187963A (ja) * 2007-02-06 2008-08-21 Mitsubishi Chemicals Corp カダベリン及び／又はカダベリン塩の溶液及びその製造方法
JP2008220195A (ja) * 2007-03-08 2008-09-25 Mitsubishi Chemicals Corp カダベリン及び／又はカダベリン塩の製造方法
US8906653B2 (en) 2008-01-23 2014-12-09 Basf Se Method for fermentatively producing 1,5-diaminopentane
US20100292429A1 (en) * 2008-01-23 2010-11-18 Basf Se Method for Fermentatively Producing 1,5-Diaminopentane
CN102056889B (zh) * 2008-07-03 2016-01-20 三菱化学株式会社 五亚甲基二胺的制造方法和聚酰胺树脂的制造方法
JP2014185156A (ja) * 2008-07-03 2014-10-02 Mitsubishi Chemicals Corp 精製ペンタメチレンジアミンの製造方法及びポリアミド樹脂の製造方法
JP2010275516A (ja) * 2008-07-03 2010-12-09 Mitsubishi Chemicals Corp 精製ペンタメチレンジアミンの製造方法及びポリアミド樹脂の製造方法
WO2010002000A1 (fr) * 2008-07-03 2010-01-07 三菱化学株式会社 Procédé de fabrication de pentaméthylènediamine, et procédé de production de résine de polyamide
EP2415801A1 (fr) * 2009-03-30 2012-02-08 Toray Industries, Inc. Résine de polyamide, composition de résine de polyamide et article moulé le comprenant
EP2415801A4 (fr) * 2009-03-30 2013-07-17 Toray Industries Résine de polyamide, composition de résine de polyamide et article moulé le comprenant
CN105164101A (zh) * 2014-04-04 2015-12-16 Cj第一制糖株式会社 有机胺的精制方法
US9963421B2 (en) 2014-04-04 2018-05-08 Cj Cheiljedang Corp. Refining method of organic amine
WO2016106367A1 (fr) 2014-12-23 2016-06-30 Genomatica, Inc. Procédé de production et de traitement de diamines
US20190322995A1 (en) * 2016-05-16 2019-10-24 Ningxia Eppen Biotech Co., Ltd Method for fermentation-production of pentanediamine comprising carbon dioxide stripping technique
US11060080B2 (en) * 2016-05-16 2021-07-13 Heilongjiang Eppen New Materials Co., Ltd. Method for fermentation-production of pentanediamine comprising carbon dioxide stripping technique

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