JPH0453509B2 - - Google Patents
Info
- Publication number
- JPH0453509B2 JPH0453509B2 JP57138415A JP13841582A JPH0453509B2 JP H0453509 B2 JPH0453509 B2 JP H0453509B2 JP 57138415 A JP57138415 A JP 57138415A JP 13841582 A JP13841582 A JP 13841582A JP H0453509 B2 JPH0453509 B2 JP H0453509B2
- Authority
- JP
- Japan
- Prior art keywords
- exchange resin
- microorganisms
- ion exchange
- reaction solution
- tryptophan
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
Links
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims description 38
- 244000005700 microbiome Species 0.000 claims description 31
- 150000008575 L-amino acids Chemical class 0.000 claims description 19
- 238000000034 method Methods 0.000 claims description 19
- 239000003729 cation exchange resin Substances 0.000 claims description 10
- 230000002378 acidificating effect Effects 0.000 claims description 9
- 238000004519 manufacturing process Methods 0.000 claims 1
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 42
- 239000000243 solution Substances 0.000 description 34
- 239000003456 ion exchange resin Substances 0.000 description 31
- 229920003303 ion-exchange polymer Polymers 0.000 description 31
- SIKJAQJRHWYJAI-UHFFFAOYSA-N Indole Chemical compound C1=CC=C2NC=CC2=C1 SIKJAQJRHWYJAI-UHFFFAOYSA-N 0.000 description 26
- 229940024606 amino acid Drugs 0.000 description 24
- 150000001413 amino acids Chemical class 0.000 description 23
- 229960004799 tryptophan Drugs 0.000 description 21
- PZOUSPYUWWUPPK-UHFFFAOYSA-N indole Natural products CC1=CC=CC2=C1C=CN2 PZOUSPYUWWUPPK-UHFFFAOYSA-N 0.000 description 13
- RKJUIXBNRJVNHR-UHFFFAOYSA-N indolenine Natural products C1=CC=C2CC=NC2=C1 RKJUIXBNRJVNHR-UHFFFAOYSA-N 0.000 description 13
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 11
- 230000001580 bacterial effect Effects 0.000 description 9
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 6
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 6
- 239000011347 resin Substances 0.000 description 6
- 229920005989 resin Polymers 0.000 description 6
- 229960001153 serine Drugs 0.000 description 6
- MTCFGRXMJLQNBG-UHFFFAOYSA-N serine Chemical compound OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 5
- 241000588724 Escherichia coli Species 0.000 description 4
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 4
- 238000011001 backwashing Methods 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- 238000001179 sorption measurement Methods 0.000 description 4
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 241000589776 Pseudomonas putida Species 0.000 description 3
- 235000011114 ammonium hydroxide Nutrition 0.000 description 3
- 239000007795 chemical reaction product Substances 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- 239000002351 wastewater Substances 0.000 description 3
- LSNNMFCWUKXFEE-UHFFFAOYSA-M Bisulfite Chemical compound OS([O-])=O LSNNMFCWUKXFEE-UHFFFAOYSA-M 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- 239000004471 Glycine Substances 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- LCTONWCANYUPML-UHFFFAOYSA-N Pyruvic acid Chemical compound CC(=O)C(O)=O LCTONWCANYUPML-UHFFFAOYSA-N 0.000 description 2
- 229910021529 ammonia Inorganic materials 0.000 description 2
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 2
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 2
- 235000011130 ammonium sulphate Nutrition 0.000 description 2
- 239000003957 anion exchange resin Substances 0.000 description 2
- RWZYAGGXGHYGMB-UHFFFAOYSA-N anthranilic acid Chemical compound NC1=CC=CC=C1C(O)=O RWZYAGGXGHYGMB-UHFFFAOYSA-N 0.000 description 2
- 239000003125 aqueous solvent Substances 0.000 description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 239000012535 impurity Substances 0.000 description 2
- 229920000642 polymer Polymers 0.000 description 2
- 239000010802 sludge Substances 0.000 description 2
- 241000607534 Aeromonas Species 0.000 description 1
- 244000063299 Bacillus subtilis Species 0.000 description 1
- 235000014469 Bacillus subtilis Nutrition 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000588915 Klebsiella aerogenes Species 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 150000001732 carboxylic acid derivatives Chemical class 0.000 description 1
- 229940023913 cation exchange resins Drugs 0.000 description 1
- 229920001429 chelating resin Polymers 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 229910000396 dipotassium phosphate Inorganic materials 0.000 description 1
- 235000019797 dipotassium phosphate Nutrition 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 238000000921 elemental analysis Methods 0.000 description 1
- 239000003480 eluent Substances 0.000 description 1
- 238000005189 flocculation Methods 0.000 description 1
- 230000016615 flocculation Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 229910000358 iron sulfate Inorganic materials 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 229920002647 polyamide Polymers 0.000 description 1
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 229940107700 pyruvic acid Drugs 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 125000000542 sulfonic acid group Chemical group 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- 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
- C12P13/04—Alpha- or beta- amino acids
- C12P13/22—Tryptophan; Tyrosine; Phenylalanine; 3,4-Dihydroxyphenylalanine
- C12P13/227—Tryptophan
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
- C07D209/04—Indoles; Hydrogenated indoles
- C07D209/10—Indoles; Hydrogenated indoles with substituted hydrocarbon radicals attached to carbon atoms of the hetero ring
- C07D209/18—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
- C07D209/20—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals substituted additionally by nitrogen atoms, e.g. tryptophane
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/02—Separating microorganisms from their culture media
-
- 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
- C12P13/04—Alpha- or beta- amino acids
-
- 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
- C12P13/04—Alpha- or beta- amino acids
- C12P13/06—Alanine; Leucine; Isoleucine; Serine; Homoserine
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Biomedical Technology (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Description
【発明の詳細な説明】
本発明は微生物を利用して製造したＬ−アミノ
酸反応液よりＬ−アミノ酸を効率良く単離するこ
とを目的とするＬ−アミノ酸の単離方法に関する
ものである。DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for isolating L-amino acids, the purpose of which is to efficiently isolate L-amino acids from L-amino acid reaction solutions produced using microorganisms.
微生物を利用したＬ−アミノ酸を含有する反応
溶液により、イオン交換樹脂を用い反応生成物を
単離回収する方法は、従来多くの報告がなされて
いる。すなわち、カルボン酸型陽イオン交換樹脂
による塩基性アミノ酸の分離（USP2549378）、
弱塩基性陰イオン交換樹脂による酸性アミノ酸の
分離（D.T.Eaglis，H.A.Fiess.，Ind.Eng.
Chem.，36巻，6041944；R.K.Cannan，J.Biol.
chem.，152巻、4011944）、またＨ型強酸性陽イ
オン交換樹脂、OH型強塩基性陰イオン交換樹脂
を用いたアミノ酸の分離（S.M.Partridge，R.C.
Blimley，Biochem.J.，51巻、6281952）などが
知られている。 Many reports have been made on methods for isolating and recovering reaction products using ion exchange resins using microorganisms and using reaction solutions containing L-amino acids. Namely, separation of basic amino acids using carboxylic acid type cation exchange resin (USP2549378),
Separation of acidic amino acids using weakly basic anion exchange resins (DTEaglis, HAFiess., Ind.Eng.
Chem., vol. 36, 6041944; RK Cannan, J. Biol.
chem., Vol. 152, 4011944), and separation of amino acids using H-type strongly acidic cation exchange resins and OH-type strongly basic anion exchange resins (SM Partridge, RC
Blimley, Biochem.J., vol. 51, 6281952).
また、工業的にイオン交換樹脂を用い反応生成
物を単離する方法としては、糖類およびアミノ酸
含有液にポリアミド系高分子凝集剤を加え不純物
を凝集沈澱させたのち、イオン交換樹脂で精製す
る方法（特公昭39−5050）、また、Ｌ−トリプト
フアンを含むアミノ酸混合液をスルホン酸型イオ
ン交換樹脂で回収する際、DVB6％以下の架橋度
のものを用いる方法（特公昭48−21105）などが
ある。 In addition, as a method for industrially isolating reaction products using ion exchange resins, a polyamide-based polymer flocculant is added to a solution containing sugars and amino acids to coagulate and precipitate impurities, and then the impurities are purified using an ion exchange resin. (Japanese Patent Publication No. 39-5050), and when recovering an amino acid mixture containing L-tryptophan using a sulfonic acid type ion exchange resin, there is a method using a cross-linking resin with a degree of DVB of 6% or less (Japanese Patent Publication No. 48-21105). be.
しかし、これ等の方法はアミノ酸の工業的な精
製法として満足できるものではない。すなわち、
これ等の方法ではイオン交換樹脂で精製する前に
あらかじめ、反応に用いた微生物を遠心分離や、
高分子凝集剤添加による凝沈、限外過膜による
別などの方法で除去しておく必要があり、工業
的にはこの微生物除去に多くの労力を必要とされ
てきた。 However, these methods are not satisfactory as industrial purification methods for amino acids. That is,
In these methods, the microorganisms used in the reaction are centrifuged, separated, or
It is necessary to remove microorganisms by other methods such as coagulation by adding a polymer flocculant or ultrafiltration membrane, and industrially, a lot of effort has been required to remove these microorganisms.
本発明者らは、微生物を用いて製造したＬ−ア
ミノ酸溶液より効率良くＬ−アミノ酸を回収する
方法を鋭意検討した結果、アミノ酸反応液をＨ型
強酸性陽イオン交換樹脂で直接処理することによ
り、反応に用いた微生物の除去と生成物のＬ−ア
ミノ酸の単離を同時に実施できることを見出し、
本発明を完成するに到つた。 As a result of intensive research into a method for efficiently recovering L-amino acids from L-amino acid solutions produced using microorganisms, the present inventors found that by directly treating the amino acid reaction solution with an H-type strongly acidic cation exchange resin, discovered that it is possible to simultaneously remove the microorganisms used in the reaction and isolate the L-amino acid product,
The present invention has now been completed.
すなわち、本発明の最も重要な点は微生物を含
むアミノ酸反応液からアミノ酸を単離精製するた
め、Ｈ型強酸性陽イオン交換樹脂層に微生物を含
むアミノ酸反応液を通し、アミノ酸をイオン交換
樹脂に吸着させる際、アミノ酸反応液中に溶解ま
たは懸濁した状態で存在している微生物が、イオ
ン交換樹脂と接触するだけで水洗浄で容易に除去
できる状態に凝集してしまうことを見出したとこ
ろにある。 That is, the most important point of the present invention is that in order to isolate and purify amino acids from an amino acid reaction solution containing microorganisms, the amino acid reaction solution containing microorganisms is passed through an H-type strongly acidic cation exchange resin layer, and the amino acids are transferred to the ion exchange resin. It was discovered that during adsorption, microorganisms that are dissolved or suspended in the amino acid reaction solution aggregate into a state that can be easily removed by water washing just by contacting the ion exchange resin. be.
この微生物が凝集する原因は、水に溶解または
懸濁した微生物がＨ型強酸性陽イオン交換樹脂に
接触するとイオン交換樹脂上のスルホン酸基によ
り変性したためと思われる。 The reason why the microorganisms aggregate is thought to be that when the microorganisms dissolved or suspended in water come into contact with the H-type strongly acidic cation exchange resin, they are denatured by the sulfonic acid groups on the ion exchange resin.
本発明の方法は、微生物を利用してＬ−アミノ
酸を製造した、微生物を含有するＬ−アミノ酸反
応液（以下、単にアミノ酸反応液という）に適用
される。 The method of the present invention is applied to an L-amino acid reaction solution containing microorganisms (hereinafter simply referred to as amino acid reaction solution) in which L-amino acids are produced using microorganisms.
このようなアミノ酸反応液としては、例えばエ
シエリヒヤ・コリ及びシユードモナス・プチ−ダ
の存在下DL−セリンとインドールより製造した
Ｌ−トリプトフアン反応液、エシエリヒヤ・コリ
の存在下Ｌ−セリンとインドールから製造したＬ
−トリプトフアン反応液、バチルス・ズブチルス
の存在下アンスラニル酸を前駆体として製造した
Ｌ−トリプトフアンの反応液、アエロバクタ−ア
エロゲネスの存在下インドールとピルビン酸、ア
ンモニアより製造したＬ−トリプトフアン反応
液、アエロバクテリウム属細菌を用いたインドー
ル、セリン、グルコースなどから製造したＬ−ト
リプトフアン反応液、あるいは、アエロモナス、
アエロバクター、シユードモナスのメタノール資
化性菌を用いメタノールを炭素源とするグリシン
含有培地で製造されたＬ−セリン反応液、ノルカ
デイア属の微生物の存在下グリシン含有培地で製
造されたＬ−セリン反応液など、広く微生物を利
用したＬ−アミノ酸反応液の精製に適用すること
ができる。 Examples of such an amino acid reaction solution include, for example, an L-tryptophan reaction solution prepared from DL-serine and indole in the presence of Escherichia coli and Pseudomonas putida, and an L-tryptophan reaction solution prepared from L-serine and indole in the presence of Escherichia coli. L
- Tryptophan reaction solution, L-tryptophan reaction solution produced using anthranilic acid as a precursor in the presence of Bacillus subtilis, L-tryptophan reaction solution produced from indole, pyruvic acid, and ammonia in the presence of Aerobacter aerogenes, Aerobacterium L-tryptophan reaction solution produced from indole, serine, glucose, etc. using bacteria of the genus Aeromonas,
L-serine reaction solution produced in a glycine-containing medium with methanol as a carbon source using methanol-assimilating bacteria such as Aerobacter and Pseudomonas; L-serine reaction solution produced in a glycine-containing medium in the presence of Norcadea microorganisms It can be widely applied to the purification of L-amino acid reaction solutions using microorganisms.
本発明の方法で使用されるＨ型強酸性陽イオン
交換樹脂としては、例えば、Lewatit sp−120
（Bayer社製）、Lewatit sc−102（Bayer社製）、
Diaion pk−208（三菱化成製）、Diaion sk−102
（三菱化成製）、Amberlite XE−100（Rohm＆
Haas社製）などがある。 Examples of the H-type strongly acidic cation exchange resin used in the method of the present invention include Lewatit sp-120
(manufactured by Bayer), Lewatit sc-102 (manufactured by Bayer),
Diaion pk−208 (manufactured by Mitsubishi Kasei), Diaion sk−102
(manufactured by Mitsubishi Kasei), Amberlite XE−100 (Rohm &
Haas), etc.
本発明の方法においてアミノ酸反応液からＬ−
アミノ酸の単離精製は以下のように実施する。す
なわち、アミノ酸反応液をそのまま、あるいはア
ミノ酸が結晶として水溶媒中に晶出している場合
には、常温で溶解するまで水で希釈して、Ｈ型に
再生したスルホン酸型陽イオン交換樹脂層の一端
より通しアミノ酸をイオン交換樹脂に吸着させ
る。 In the method of the present invention, L-
Isolation and purification of amino acids is carried out as follows. That is, if the amino acid reaction solution is used as it is, or if the amino acid is crystallized in an aqueous solvent, it is diluted with water at room temperature until it dissolves, and the sulfonic acid type cation exchange resin layer regenerated into the H form is prepared. One end is passed through and the amino acid is adsorbed onto the ion exchange resin.
一方、反応液中の微生物はイオン交換樹脂層で
ほとんど変性し凝集状態でイオン交換樹脂上に付
着する。 On the other hand, most of the microorganisms in the reaction solution are denatured in the ion exchange resin layer and adhere to the ion exchange resin in an aggregated state.
その後、イオン交換樹脂層の他の一端から一定
流量で水を通じて逆洗してイオン交換樹脂に付着
している凝集した微生物をイオン交換樹脂層より
排水と共に流去させる。 Thereafter, the ion exchange resin layer is backwashed by flowing water at a constant flow rate from the other end of the ion exchange resin layer, so that the aggregated microorganisms adhering to the ion exchange resin are washed away from the ion exchange resin layer together with the waste water.
このアミノ酸反応液のイオン交換樹脂層でのア
ミノ酸の吸着、微生物の凝集、凝集微生物の除去
は通常イオン交換樹脂を充填した塔により実施さ
れるが、イオン交換樹脂にアミノ酸を吸着させた
のち、反応槽に排出し、水でスラツジ洗浄する方
法を用いても何ら問題ない。 The adsorption of amino acids, flocculation of microorganisms, and removal of flocculated microorganisms in the ion-exchange resin layer of the amino acid reaction solution are usually carried out in a column filled with ion-exchange resin. There is no problem in using the method of discharging the sludge into a tank and washing the sludge with water.
イオン交換樹脂塔を用いる場合は、まずイオン
交換樹脂塔の上部から一定流量でアミノ酸反応液
を流し、アミノ酸をイオン交換樹脂に吸着させ
る。この際、アミノ酸反応液に含まれている微生
物のほとんどは樹脂塔中で変性を凝集物を生成
し、樹脂上に物理的に保持されている。また微生
物の一部は樹脂塔の下部から排水と共に流出す
る。 When using an ion exchange resin tower, first, the amino acid reaction solution is flowed from the top of the ion exchange resin tower at a constant flow rate, and the amino acids are adsorbed onto the ion exchange resin. At this time, most of the microorganisms contained in the amino acid reaction solution undergo denaturation in the resin column to form aggregates, which are physically retained on the resin. In addition, some of the microorganisms flow out from the lower part of the resin tower along with the waste water.
上記の操作の後、イオン交換樹脂塔の下部から
一定流量で水を通し逆洗すると、樹脂に付着して
いる凝集した微生物が浮遊して塔上部から効率良
く流出除去される。 After the above operation, when the ion exchange resin tower is backwashed by passing water at a constant flow rate from the lower part of the tower, the flocculated microorganisms attached to the resin are suspended and efficiently flowed out from the upper part of the tower and removed.
この方法によりＬ−アミノ酸のイオン交換樹脂
への吸着時に排水と共に流出除去される微生物と
逆洗時に樹脂層より流出除去される微生物とで、
イオン交換樹脂層に供給したＬ−アミノ酸反応液
中の微生物は事実上、完全に除去することができ
る。 With this method, microorganisms are removed together with wastewater when L-amino acids are adsorbed onto the ion exchange resin, and microorganisms are removed from the resin layer during backwashing.
Microorganisms in the L-amino acid reaction solution supplied to the ion exchange resin layer can be virtually completely removed.
つぎに、Ｌ−アミノ酸を吸着したイオン交換樹
脂は通常アンモニア水などで溶離したのち、その
溶液を濃縮・晶析することにより容易に目的のＬ
−アミノ酸を単離することができる。 Next, the ion exchange resin adsorbed with L-amino acids is usually eluted with aqueous ammonia, and then the solution is concentrated and crystallized to easily obtain the desired L-amino acid.
-Amino acids can be isolated.
得られたＬ−アミノ酸はイオン交換樹脂による
精製の効果により高純度のものである。 The obtained L-amino acid has high purity due to the effect of purification using an ion exchange resin.
本発明の方法は、微生物を含んだＬ−アミノ酸
反応液をＨ型強酸性陽イオン交換樹脂で処理する
ことによりＬ−アミノ酸の単離と、通常の方法で
は工業的にその分離が極めて困難な微生物の除去
とを同時に実施することができる。 The method of the present invention isolates L-amino acids by treating an L-amino acid reaction solution containing microorganisms with an H-type strongly acidic cation exchange resin. Removal of microorganisms can be carried out at the same time.
すなわち、微生物を利用した反応生成物の精製
法としてその工業的意義は極めて大きい。 In other words, it has great industrial significance as a method for purifying reaction products using microorganisms.
以下、実施例により本発明の方法を詳細に説明
する。 Hereinafter, the method of the present invention will be explained in detail with reference to Examples.
実施例 １
リン酸−カリ、リン酸二カリ、硫安、および塩
化カルシウム、硫酸鉄などのミネラル、酵母エキ
ス、ポリペプトンなどの存在下グルコース、イン
ドールを添加しながらエジエリヒヤ・コリを含ん
だ菌体をPH７、30℃の培養条件下、空気を吹き込
みながら撹拌し、40時間培養する。最終菌体濃度
30〜35ｇ／。Example 1 In the presence of phosphoric acid-potassium, dipotassium phosphate, ammonium sulfate, minerals such as calcium chloride and iron sulfate, yeast extract, polypeptone, etc., bacterial cells containing E. coli were brought to pH 7 while adding glucose and indole. , and incubate for 40 hours at 30°C with stirring while blowing air. Final cell concentration
30-35g/.
同様にして、インドールを含まない培地でシユ
ードモナス・プチーダを含んだ菌体を培養する。 Similarly, cells containing Pseudomonas putida are cultured in a medium that does not contain indole.
培養液は通常の超遠心分離機により集菌し、含
水率75〜85％の塊として得る。 The culture solution is collected using a conventional ultracentrifuge and obtained as a mass with a water content of 75 to 85%.
つぎに、DL−セリン77.3ｇ、硫安10.5ｇ、水
486ｇを入れた反応機に29％アンモニア水でPH8.5
に調製する。さらに前述のエシエリヒヤ・コリ菌
体塊51.2ｇ、シユードモナス・プチーダ菌体
塊23.2ｇを加えよくかきまぜる。さらにインドー
ル78.4ｇを溶解したトルエン溶液392ｇを加え35
℃に保温し40時間反応させる。 Next, 77.3 g of DL-serine, 10.5 g of ammonium sulfate, water
PH8.5 with 29% ammonia water in a reactor containing 486g
Prepare to. Furthermore, 51.2 g of the aforementioned Escherichia coli bacterial mass and 23.2 g of Pseudomonas putida bacterial mass were added and stirred well. Furthermore, 392 g of toluene solution containing 78.4 g of indole was added.
Incubate at ℃ and react for 40 hours.
反応マス中のＬ−トリプトフアン生成量を液体
クロマトグラフイーで分析したところ、129.8ｇ
生成していた。収率95.0％対インドール。 When the amount of L-tryptophan produced in the reaction mass was analyzed by liquid chromatography, it was found to be 129.8g.
It was generating. Yield 95.0% vs. indole.
つぎに蒸留によりトルエンを留去した後、反応
液を水で希釈しＬ−トリプトフアンの濃度を
1.0wt％に調製しＬ−トリプトフアン結晶を完溶
する。 Next, after removing toluene by distillation, the reaction solution was diluted with water to adjust the concentration of L-tryptophan.
Adjust to 1.0wt% to completely dissolve L-tryptophan crystals.
一方、強酸性陽イオン交換樹脂Lewatit sp−
1204.84を塩酸で再生しＨ型としたものをカラ
ムに充填しその上端から先きに調製したＬ−トリ
プトフアン溶液125ｇを一定流量で通じイオン交
換樹脂にＬ−トリプトフアンを吸着させる。 On the other hand, strongly acidic cation exchange resin Lewatit sp−
1204.84 was regenerated into H-form with hydrochloric acid and filled into a column, and 125 g of the L-tryptophan solution prepared earlier was passed through the column at a constant flow rate through the top of the column to adsorb L-tryptophan onto the ion exchange resin.
水24.9ｇで逆洗することにより浮遊してくる菌
体の凝集物を流去したのちイオン交換樹脂交換容
量の２倍のアンモニア水でＬ−トリプトフアンを
溶離する。溶離液は100℃に昇温したアンモニア
を除去回収したのち室温まで冷却し析出したＬ−
トリプトフアン結晶を別、乾燥する。単離収量
10.0ｇ、純度99.8％。 After washing away floating bacterial aggregates by backwashing with 24.9 g of water, L-tryptophan is eluted with aqueous ammonia twice the exchange capacity of the ion exchange resin. The eluent was heated to 100°C, removed and recovered ammonia, and then cooled to room temperature to remove the precipitated L-
Separate the tryptophan crystals and dry. isolated yield
10.0g, purity 99.8%.
なおイオン交換樹脂処理による菌体バランスは
Ｌ−トリプトフアン吸着時の漏出液中に３％、逆
洗時の流出液中に残り97％がきていた。なおそれ
ぞれの菌体バランスは水溶液を濃縮乾固し、その
時の重量と元素分析による炭素バランスより求め
た。 The balance of bacterial cells resulting from the ion-exchange resin treatment was 3% in the effluent during L-tryptophan adsorption, and the remaining 97% in the effluent during backwashing. The bacterial cell balance for each was determined by concentrating the aqueous solution to dryness, and calculating the weight at that time and the carbon balance by elemental analysis.
実施例 ２
実施例１と同様にして培養したエシエリヒヤコ
リ菌体塊を用い水溶媒中Ｌ−セリンとインドー
ルよりＬ−トリプトフアンを製造した。反応中は
インドールによる酵素活性の低下を避けるため水
中でのインドール濃度が200ppm以下になるよう
に連続分析しながら徐々にインドールを添加する
方法で実施した。反応収率100％対インドール、
85％対Ｌ−セリン。最終Ｌ−トリプトフアン蓄積
濃度120ｇ／であつた。このＬ−トリプトフア
ン反応液を遠心脱水によりＬ−トリプトフアンと
菌体を含んだ反応塊を得る。Example 2 L-tryptophan was produced from L-serine and indole in an aqueous solvent using a mass of Eschieri Hyacoli bacteria cultured in the same manner as in Example 1. During the reaction, indole was gradually added during continuous analysis to avoid a decrease in enzyme activity due to indole, so that the indole concentration in water was 200 ppm or less. Reaction yield 100% vs. indole,
85% vs. L-serine. The final L-tryptophan accumulation concentration was 120 g/l. This L-tryptophan reaction solution is centrifugally dehydrated to obtain a reaction mass containing L-tryptophan and bacterial cells.
この反応塊を実施例１と同様な操作でイオン
交換樹脂を用い菌体除去、ならびにＬ−トリプト
フアンの単離を行つた。イオン交換樹脂としては
Lewatit sc−102のＨ型を用いた。 This reaction mass was subjected to the same procedure as in Example 1 to remove bacterial cells and isolate L-tryptophan using an ion exchange resin. As an ion exchange resin
Lewatit sc-102 H type was used.
Ｌ−トリプトフアンの単離収量11.0ｇ、純度
99.9％であり、Ｌ−トリプトフアン溶液中の菌体
はイオン交換樹脂吸着時に2.5％、イオン交換樹
脂塔逆洗時に97.5％が除去されていた。 Isolated yield of L-tryptophan 11.0 g, purity
The result was 99.9%, and 2.5% of the bacterial cells in the L-tryptophan solution were removed during adsorption with the ion exchange resin, and 97.5% were removed during backwashing of the ion exchange resin column.
Claims (1)
おいて、微生物を含有する反応液を生成物が溶解
した状態でＨ型強酸性陽イオン交換樹脂により処
理することを特徴とする微生物を利用して製造し
たＬ−アミノ酸の単離方法。1. A method for producing L-amino acids using microorganisms, characterized in that a reaction solution containing microorganisms is treated with an H-type strongly acidic cation exchange resin in a state in which the product is dissolved. Method for isolating L-amino acids.
Priority Applications (8)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP13841582A JPS5928484A (en) | 1982-08-11 | 1982-08-11 | Method for isolating l-amino acid |
| GB08334070A GB2152030B (en) | 1982-08-11 | 1983-12-21 | Isolating l-amino acids by ion exchange |
| CA000444165A CA1215069A (en) | 1982-08-11 | 1983-12-22 | Method of isolating l-trytophan |
| AU22840/83A AU567903B2 (en) | 1982-08-11 | 1983-12-23 | Method of isolating l-amino acids form culture medium using acipic cation exchange resin |
| NL8304496A NL8304496A (en) | 1982-08-11 | 1983-12-30 | METHOD FOR INSULATING L-AMINO ACIDS |
| CH8884A CH659827A5 (en) | 1982-08-11 | 1984-01-06 | METHOD FOR ISOLATING L-AMINO ACIDS. |
| FR8400302A FR2557872B1 (en) | 1982-08-11 | 1984-01-10 | PROCESS FOR ISOLATING L-AMINO ACIDS FROM A REACTION MIXTURE |
| DE19843400574 DE3400574A1 (en) | 1982-08-11 | 1984-01-10 | METHOD FOR ISOLATING L-AMINO ACIDS |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP13841582A JPS5928484A (en) | 1982-08-11 | 1982-08-11 | Method for isolating l-amino acid |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS5928484A JPS5928484A (en) | 1984-02-15 |
| JPH0453509B2 true JPH0453509B2 (en) | 1992-08-26 |
Family
ID=15221422
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP13841582A Granted JPS5928484A (en) | 1982-08-11 | 1982-08-11 | Method for isolating l-amino acid |
Country Status (8)
| Country | Link |
|---|---|
| JP (1) | JPS5928484A (en) |
| AU (1) | AU567903B2 (en) |
| CA (1) | CA1215069A (en) |
| CH (1) | CH659827A5 (en) |
| DE (1) | DE3400574A1 (en) |
| FR (1) | FR2557872B1 (en) |
| GB (1) | GB2152030B (en) |
| NL (1) | NL8304496A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007105790A1 (en) | 2006-03-15 | 2007-09-20 | Kyowa Hakko Kogyo Co., Ltd. | Method for purification of amino acid |
| WO2007119369A1 (en) * | 2006-03-15 | 2007-10-25 | Kyowa Hakko Kogyo Co., Ltd. | Process for purification of amino acid |
Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4769474A (en) * | 1985-04-10 | 1988-09-06 | Mitsui Toatsu Chemicals, Inc. | Process for purifying tryptophane |
| DE3702689A1 (en) * | 1987-01-30 | 1988-08-11 | Degussa | METHOD FOR ISOLATING L-AMINO ACIDS |
| ATE109833T1 (en) * | 1987-10-12 | 1994-08-15 | Mitsui Toatsu Chemicals | PROCESS FOR PRODUCTION OF L-TRYPTOPHANE. |
| JPH0325170U (en) * | 1989-07-20 | 1991-03-14 | ||
| US8158411B2 (en) | 2006-08-21 | 2012-04-17 | Samsung Electronics Co., Ltd. | Method of separating microorganism using nonplanar solid substrate and device for separating microorganism using the same |
| JP2008048735A (en) | 2006-08-21 | 2008-03-06 | Samsung Electronics Co Ltd | Method for separating microorganism with non-planar solid support and apparatus for separating the same |
| BR112016015718B1 (en) * | 2014-01-07 | 2021-12-07 | Novasep Process Solutions | AROMATIC AMINO ACIDS PURIFICATION PROCESS |
| CN103772086B (en) * | 2014-01-10 | 2015-04-29 | 国家海洋局第三海洋研究所 | Pretreatment process for preparing fractions of marine microorganism small molecule metabolites |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS50127879A (en) * | 1974-03-28 | 1975-10-08 |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| IT1045451B (en) * | 1966-03-23 | 1980-05-10 | Ajinomoto Kk | METHOD FOR RECOVERING LYSINE FROM FERMENTATION BROTH |
| GB1186952A (en) * | 1967-06-17 | 1970-04-08 | Kyowa Hakko Kogyo Kk | Process for producing L-Tryptophan |
| JPS5636710A (en) * | 1979-09-04 | 1981-04-10 | Fanuc Ltd | Feed speed command system |
| JPS57174096A (en) * | 1981-04-20 | 1982-10-26 | Ajinomoto Co Inc | Preparation of l-tryptophan by fermentation method |
-
1982
- 1982-08-11 JP JP13841582A patent/JPS5928484A/en active Granted
-
1983
- 1983-12-21 GB GB08334070A patent/GB2152030B/en not_active Expired
- 1983-12-22 CA CA000444165A patent/CA1215069A/en not_active Expired
- 1983-12-23 AU AU22840/83A patent/AU567903B2/en not_active Ceased
- 1983-12-30 NL NL8304496A patent/NL8304496A/en not_active Application Discontinuation
-
1984
- 1984-01-06 CH CH8884A patent/CH659827A5/en not_active IP Right Cessation
- 1984-01-10 FR FR8400302A patent/FR2557872B1/en not_active Expired
- 1984-01-10 DE DE19843400574 patent/DE3400574A1/en active Granted
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS50127879A (en) * | 1974-03-28 | 1975-10-08 |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007105790A1 (en) | 2006-03-15 | 2007-09-20 | Kyowa Hakko Kogyo Co., Ltd. | Method for purification of amino acid |
| WO2007119369A1 (en) * | 2006-03-15 | 2007-10-25 | Kyowa Hakko Kogyo Co., Ltd. | Process for purification of amino acid |
Also Published As
| Publication number | Publication date |
|---|---|
| DE3400574C2 (en) | 1987-08-06 |
| CA1215069A (en) | 1986-12-09 |
| CH659827A5 (en) | 1987-02-27 |
| NL8304496A (en) | 1985-07-16 |
| GB2152030B (en) | 1987-08-19 |
| FR2557872A1 (en) | 1985-07-12 |
| FR2557872B1 (en) | 1987-07-10 |
| DE3400574A1 (en) | 1985-07-18 |
| GB2152030A (en) | 1985-07-31 |
| GB8334070D0 (en) | 1984-02-01 |
| AU2284083A (en) | 1985-06-27 |
| AU567903B2 (en) | 1987-12-10 |
| JPS5928484A (en) | 1984-02-15 |
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