JPH03180187A - Production of l-malic acid - Google Patents
Production of l-malic acidInfo
- Publication number
- JPH03180187A JPH03180187A JP31924489A JP31924489A JPH03180187A JP H03180187 A JPH03180187 A JP H03180187A JP 31924489 A JP31924489 A JP 31924489A JP 31924489 A JP31924489 A JP 31924489A JP H03180187 A JPH03180187 A JP H03180187A
- Authority
- JP
- Japan
- Prior art keywords
- malic acid
- ustilago
- culture
- acid
- ability
- 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.)
- Pending
Links
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 title claims abstract description 59
- 229940116298 l- malic acid Drugs 0.000 title claims abstract description 26
- 238000004519 manufacturing process Methods 0.000 title claims description 7
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 claims abstract description 33
- 235000011090 malic acid Nutrition 0.000 claims abstract description 33
- 241000221566 Ustilago Species 0.000 claims abstract description 18
- 244000005700 microbiome Species 0.000 claims abstract description 7
- 238000012258 culturing Methods 0.000 claims abstract description 4
- 229940099690 malic acid Drugs 0.000 claims description 8
- 239000001630 malic acid Substances 0.000 claims description 8
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 abstract description 8
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 abstract description 8
- 150000007524 organic acids Chemical class 0.000 abstract description 5
- 239000000126 substance Substances 0.000 abstract description 5
- 229910052799 carbon Inorganic materials 0.000 abstract description 4
- 229910052757 nitrogen Inorganic materials 0.000 abstract description 4
- 241000893097 Tolyposporium Species 0.000 abstract description 3
- 239000002253 acid Substances 0.000 abstract description 3
- 150000001413 amino acids Chemical class 0.000 abstract description 3
- 239000006227 byproduct Substances 0.000 abstract description 3
- 239000001963 growth medium Substances 0.000 abstract description 2
- 229930003231 vitamin Natural products 0.000 abstract description 2
- 229940088594 vitamin Drugs 0.000 abstract description 2
- 235000013343 vitamin Nutrition 0.000 abstract description 2
- 239000011782 vitamin Substances 0.000 abstract description 2
- 230000000813 microbial effect Effects 0.000 abstract 1
- 239000002609 medium Substances 0.000 description 11
- 238000000034 method Methods 0.000 description 9
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 8
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 4
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
- 229910000019 calcium carbonate Inorganic materials 0.000 description 4
- 150000001720 carbohydrates Chemical class 0.000 description 4
- 235000014633 carbohydrates Nutrition 0.000 description 4
- 238000000855 fermentation Methods 0.000 description 4
- 230000004151 fermentation Effects 0.000 description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 4
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 4
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 238000012136 culture method Methods 0.000 description 3
- 239000002994 raw material Substances 0.000 description 3
- 235000002639 sodium chloride Nutrition 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 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 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 2
- 241000233866 Fungi Species 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
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 235000019270 ammonium chloride Nutrition 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 235000019253 formic acid Nutrition 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 235000005985 organic acids Nutrition 0.000 description 2
- 229910052697 platinum Inorganic materials 0.000 description 2
- 239000001965 potato dextrose agar Substances 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- PAWQVTBBRAZDMG-UHFFFAOYSA-N 2-(3-bromo-2-fluorophenyl)acetic acid Chemical compound OC(=O)CC1=CC=CC(Br)=C1F PAWQVTBBRAZDMG-UHFFFAOYSA-N 0.000 description 1
- ATRRKUHOCOJYRX-UHFFFAOYSA-N Ammonium bicarbonate Chemical compound [NH4+].OC([O-])=O ATRRKUHOCOJYRX-UHFFFAOYSA-N 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 241000228212 Aspergillus Species 0.000 description 1
- 241000223651 Aureobasidium Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- 235000003385 Diospyros ebenum Nutrition 0.000 description 1
- 241000792913 Ebenaceae Species 0.000 description 1
- 229930091371 Fructose Natural products 0.000 description 1
- 239000005715 Fructose Substances 0.000 description 1
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 1
- 241000222481 Schizophyllum commune Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 229910052770 Uranium Inorganic materials 0.000 description 1
- 241000041347 Ustilago coicis Species 0.000 description 1
- 241000509513 Ustilago hordei Species 0.000 description 1
- 241000952806 Ustilago syntherismae Species 0.000 description 1
- 241000007071 Ustilago trichophora Species 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 238000013019 agitation Methods 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 239000001099 ammonium carbonate Substances 0.000 description 1
- 235000012501 ammonium carbonate Nutrition 0.000 description 1
- 235000011114 ammonium hydroxide Nutrition 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 239000003957 anion exchange resin Substances 0.000 description 1
- 244000000005 bacterial plant pathogen Species 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 239000003729 cation exchange resin Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 229910000365 copper sulfate Inorganic materials 0.000 description 1
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000012259 ether extract Substances 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000011790 ferrous sulphate Substances 0.000 description 1
- 235000003891 ferrous sulphate Nutrition 0.000 description 1
- 235000013312 flour Nutrition 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- 229910000358 iron sulfate Inorganic materials 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 229940099596 manganese sulfate Drugs 0.000 description 1
- 239000011702 manganese sulphate Substances 0.000 description 1
- 235000007079 manganese sulphate Nutrition 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 235000013379 molasses Nutrition 0.000 description 1
- 239000012452 mother liquor Substances 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 125000001477 organic nitrogen group Chemical group 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 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
- 229910000160 potassium phosphate Inorganic materials 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000004317 sodium nitrate Substances 0.000 description 1
- 235000010344 sodium nitrate Nutrition 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
- 229960001763 zinc sulfate Drugs 0.000 description 1
Abstract
Description
【発明の詳細な説明】
産業上の利用分野
本発明は、Ｌ−リンゴ酸の製造法に関する。ＬＩＪンゴ
酸は天然型の有機酸で、医薬用、食用として広く利用さ
れている。DETAILED DESCRIPTION OF THE INVENTION Field of the Invention The present invention relates to a method for producing L-malic acid. LIJ malic acid is a natural organic acid that is widely used for medicine and food.
従来の技術
従来、発酵法により糖質から直接Ｌ−リンゴ酸を製造す
る方法としては、アスペルギルス属菌種を用いる方法（
特公昭３７−１５２９７、特公昭４Ｏ−６３９５）、オ
ーレオバシジウム属菌種を用いる方法（特開昭５ｌ−７
９７８３） 、シゾフィリューム・コミューンを用いる
方法（特公昭３９−７３９０）などが知られている。Conventional technology Conventionally, as a method for directly producing L-malic acid from carbohydrates by a fermentation method, a method using Aspergillus species (
Japanese Patent Publication No. 37-15297, Japanese Patent Publication No. 40-6395), method using Aureobasidium species (Japanese Patent Publication No. 51-7
9783), a method using Schizophyllum commune (Japanese Patent Publication No. 39-7390), etc. are known.
発明が解決しようとする課題
安価な糖質を原料としてＬ−リンゴ酸を直接発酵生産す
る場合、従来の方法では、生産性が低く、また副生ずる
有機酸が多いためＬ−リンゴ酸の精製が困難であるなど
の欠点を有していた。Problems to be Solved by the Invention When producing L-malic acid directly by fermentation using inexpensive carbohydrates as raw materials, conventional methods have low productivity and produce a large amount of organic acids as by-products, making it difficult to purify L-malic acid. It had drawbacks such as being difficult.
課題を解決するための手段
本発明者は、安価な糖質を原料としてＬ −リンゴ酸を
直接発酵生産するために、Ｌ −リンゴ酸生産能を有す
る菌株を種々検討した。その結果、Ｌ−リンゴ酸生産能
が高く、副生ずる有機酸が少ない菌株を得、該菌株を用
いたＬ −Ｑンゴ酸の直接発酵生産方法を確立した。Means for Solving the Problems The present inventor investigated various strains capable of producing L-malic acid in order to directly ferment and produce L-malic acid using inexpensive carbohydrates as raw materials. As a result, a strain with high L-malic acid production ability and low by-product organic acid was obtained, and a method for direct fermentation production of L-Q malic acid using this strain was established.
本発明は、ウスティラゴ属またはトリボスポリウム属に
属し、Ｌ−リンゴ酸生産能を有する微生物を培地に培養
し、培養物中にＬ−’Ｊンゴ酸を生成蓄積させ、該培養
物よりＬ −リンゴ酸を採取することを特徴とするＬ−
リンゴ酸の製造法を提供する。The present invention involves culturing a microorganism that belongs to the genus Ustilago or the genus Tribosporium and has the ability to produce L-malic acid in a medium, producing and accumulating L-'J malic acid in the culture, and producing L-malic acid from the culture. L-, which is characterized by collecting
Provided is a method for producing malic acid.
以下に本発明の詳細な説明する。The present invention will be explained in detail below.
本発明に用いる微生物としては、ウスティラゴ属または
トリボスボリウム属に属し、Ｌ−リンゴ酸生産能を有す
る菌株であればいずれでもよい。The microorganism used in the present invention may be any strain as long as it belongs to the genus Ustilago or the genus Tribosborium and has the ability to produce L-malic acid.
具体的には、ウスティラゴ・クリスーガリイ（Ｌｌｓｔ
ｉｌａｇｏ ｃｒｕｓ−ｇａｌｌｉ）に３９０． Ｋ３
９１．に３９２．に３９３゜Ｋ２１６． ウスティラ
ゴ・ホルダイ（Ｕ、ｈｏｒｄｅ ｉ） Ｎ（Ｌ １１５
、ウスティラゴ・スファエロゲナ（Ｕ、 ｓｐｈａｅｒ
ｏｇｅｎａ）Ｋ２Ｏ２，に４０２、ウスティラゴ・コイ
シス （Ｕ、ｃｏｉｃｉｓ）Ｋ３３１． Ｋ３３２、ウ
スティラゴ・スキタミネア（Ｕ。Specifically, Ustilago Crisugali (Llst
390. K3
91. 392. 393°K216. Ustilago Hordei (U, horde i) N (L 115
, Ustilago sphaerogena (U, sphaer
ogena) K2O2, ni402, Ustilago coicis (U, coicis) K331. K332, Ustilago scythaminea (U.
ｓｃｉｔａｍｉｎｅａ） Ｋ３３５． Ｋ３３６． Ｋ
３３８．に３３９、ウスティラゴ・ニスキュＬ／ンタ（
Ｕ、ｅｓｃｕｌｅｎｔａ） Ｋ２Ｏ２，Ｋ２Ｏ２、ウス
ティラゴ・シンセリスマエ（口、ｓｙｎｔｈｅｒｉｓｍ
ａｅ）Ｋ３８２．に３８３、トリポスポリウム・ブラタ
ム（Ｔｏｌｙｐｏｓｐｏｌｉｕｍ ｂｕｌｌａｔｕｍ）
にＴ３０４．にＴ３２６． ＫＴ３２７゜ＫＴ３２８な
どがあげられる。これらの菌類は、黒檀菌類に属する植
物病原菌である。scitaminea) K335. K336. K
338. 339, Ustilago Niscu L/Nta (
U, esculenta) K2O2, K2O2, Ustilago syntherismae (mouth, syntherism)
ae) K382. 383, Tolyposporium bullatum
T304. T326. Examples include KT327°KT328. These fungi are plant pathogenic bacteria that belong to the Ebony fungi.
上記菌種の菌学的性質は、以下の文献にそれぞれ記載さ
れており、該文献に基づいて、各菌株の同定を行った。The mycological properties of the above bacterial species are described in the following documents, and each strain was identified based on the documents.
Ｕ、ｃｒｕｓ−ｇａｌｌｉ： Ｂｕｌｌ、Ｔｏｒｒｅ
ｙ Ｂａｔ、Ｃ１ｕｂ ２２ 。U, crus-galli: Bull, Torre
y Bat, C1ub 22.
１７５ （１８９５）
ＩＪ、ｈｏｒｄｅｉ ： Ｍｉｔｔ、Ｂａｄｉｓｃｈ
ｅｎ Ｒｏｔ、Ｖｅｒ、、Ｐ、７０（１８８９）［１，
ｓｐｈａｅｒｏｇｅｎａ ： ５ｙｌｌ、Ｆｕｎｇ、
７． ４６８（１８８ｇ）Ｅｌ、 ｃｏｉｃｉｓ ：
［Ｉｎｔｅｒｓ、 ＧｅｓａｍｌｌＩｔ、Ｍｙｋｏｌ
、 １２．１１０（１８９５）［１，ｓｃｉｔａｍｉｎ
ｅａ ： ＾ｎｎ、 Ｍｙｃｏｌ、 ２２ 、 ２
８１（１９２４）［１，ｅｓｃｕｌｅｎｔａ ：
）Ｉｅｄｗｉｇｉａ ３４ 、１０（１８９５）Ｕ、ｓ
ｙｎｔｈｅｒｉｓｍａｅ ： Ａｎｎ、Ｒｅｐｔ、Ｎ、
Ｙ、５ｔａｔｅ ！Ｊｕｓ、２７ 。175 (1895) IJ, hordei: Mitt, Badisch
en Rot, Ver., P. 70 (1889) [1,
sphaerogena: 5yll, Fung,
7. 468 (188g) El, coicis:
[Inters, GesamllIt, Mykol
, 12.110 (1895) [1, scitamin
ea: ^nn, Mycol, 22, 2
81 (1924) [1, esculenta:
) Iedwigia 34, 10 (1895) U, s
yntherismae: Ann, Rept, N,
Y, 5tate! Jus, 27.
１０３ （１８７５）
Ｔ、ｂｕｌｌａｔｕｍ ： Ｋｒｙｐｔ、 Ｆｌ、 ５
ｃｈｌｅｓ、３．２７６（１８８７）本発明の微生物を
通常の培養方法によって培養することによって、Ｌ−リ
ンゴ酸を培養物中に生成蓄積させることができる。すな
わち、これらの微生物を適当な炭素源、窒素源、無機物
、アミノ酸、ビタミンなどを含有する通常の培地中にお
いて、好気的条件下で温度、ｐＨなどを調節しつつ培養
を行えばよい。103 (1875) T, bullatum: Krypt, Fl, 5
chles, 3.276 (1887) By culturing the microorganism of the present invention by a conventional culture method, L-malic acid can be produced and accumulated in the culture. That is, these microorganisms may be cultured in a conventional medium containing appropriate carbon sources, nitrogen sources, inorganic substances, amino acids, vitamins, etc. under aerobic conditions while controlling temperature, pH, etc.
培養培地としては炭素源、窒素源、無機物などを程よく
含有する培地ならば天然培地、合成培地のいずれでも用
いることができる。As the culture medium, either a natural medium or a synthetic medium can be used as long as it contains moderate amounts of carbon sources, nitrogen sources, inorganic substances, etc.
炭素源としてはブドウ糖、澱粉、グリセロール、マンノ
ース、フラクトース、シュークロース、糖蜜などを単独
または組み合わせて用いることができる。さらに菌の資
化能によっては炭化水素、アルコール類、有機酸などを
用いることができる。As the carbon source, glucose, starch, glycerol, mannose, fructose, sucrose, molasses, etc. can be used alone or in combination. Furthermore, depending on the assimilation ability of the bacteria, hydrocarbons, alcohols, organic acids, etc. can be used.
窒素源としては塩化アンモニウム、硫酸アンモニウム、
硝酸アンモニウム、硝酸ナトリウム、尿素などの無機も
しくは有機の窒素含有化合物、またペプトン、肉エキス
、酵母エキス、乾燥酵母、コーン・スチープ・リカー、
大豆粉、力ずミノ酸などの天然物含有窒素源を単独また
は組み合わせて用いることができる。Ammonium chloride, ammonium sulfate,
Inorganic or organic nitrogen-containing compounds such as ammonium nitrate, sodium nitrate, urea, as well as peptone, meat extract, yeast extract, dried yeast, corn steep liquor,
Natural product-containing nitrogen sources such as soybean flour and glutinous amino acid can be used alone or in combination.
無機物としては塩化ナトリウム、塩化カリウム、硫酸第
一鉄、硫酸亜鉛、硫酸マンガン、硫酸銅、炭酸カルシウ
ム、リン酸塩などの無機塩類を用いることができる。そ
のほか必要に応じてＬ−リンゴ酸の生産を促進する有機
物や無機物を適当に添加することができる。また、培養
中のｐＨの低下を抑える目的で、炭酸カルシウムを添加
することもできる。As the inorganic substance, inorganic salts such as sodium chloride, potassium chloride, ferrous sulfate, zinc sulfate, manganese sulfate, copper sulfate, calcium carbonate, and phosphates can be used. In addition, organic or inorganic substances that promote the production of L-malic acid may be appropriately added as necessary. Moreover, calcium carbonate can also be added for the purpose of suppressing a decrease in pH during culture.
培養法としては、好気的条件下、通常は液体培養法、と
くに深部攪拌培養法が用いられる。培養温度は１５〜３
５℃、特に２０〜３０℃が好適である。培地のｐＨは、
アンモニア水や炭酸アンモニウム溶液などを添加して、
ｐＨ４〜１０．特に５〜６に保ちながら培養を行うこと
が望ましい。As a culture method, under aerobic conditions, usually a liquid culture method, especially a deep agitation culture method is used. Culture temperature is 15-3
5°C, especially 20-30°C is preferred. The pH of the medium is
Add ammonia water or ammonium carbonate solution, etc.
pH 4-10. In particular, it is desirable to carry out the culture while maintaining the temperature at 5 to 6.
液体培養で通常５〜８日培養を行うと、Ｌ−リンゴ酸が
培養物中に生成蓄積する。培養物中のＬリンゴ酸生成量
が最大に達したときに培養を停止し、培養物中からＬ
−リンゴ酸を単離精製する。When liquid culture is carried out for usually 5 to 8 days, L-malic acid is produced and accumulated in the culture. When the amount of L-malic acid produced in the culture reached the maximum, the culture was stopped and L was removed from the culture.
- Isolate and purify malic acid.
培養終了後、遠心分離により菌体を除去し、得られた上
澄液を、イオン交換樹脂法やエーテルなどの有機溶媒に
よる抽出法などの公知の方法を組み合わせることによっ
て、Ｌ −リンゴ酸を分離し採取することができる。After the culture is completed, the bacterial cells are removed by centrifugation, and L-malic acid is separated from the resulting supernatant using a combination of known methods such as ion exchange resin method and extraction method using organic solvents such as ether. It can be collected.
以下に本発明の実施例を示す。Examples of the present invention are shown below.
実施例１
グルコース１２０ｇ、塩化アンモニウム１１．６ｇ１リ
ン酸−カリウム０．５ｇ、硫酸マグネシウム７永和物０
．２ｇ、硫酸鉄７永和物０．０１ｇおよび酵母エキス１
ｇを水道水ＩＩｌに溶解した溶液３０−を、３００−容
三角フラスコに入れ、１２０℃、１、２ ｋｇ ／ ｃ
ｄで１０分間殺菌して得られた培地に、予めポテトデキ
ストロース寒天の斜面培地上で生育させた、ウスティラ
ゴ・クリスーガリイに３９１を一白金耳植菌した。同時
に別に殺菌した炭酸カルシウム１ｇを加えた。この培養
液をロータリーシェーカー上で２５℃、７時間振盪培養
した。培養終了後、培養液を分析した結果、培養液１１
当り５２ｇのＬ −Ｕンゴ酸が生成していた。得られた
培養液１１を塩酸で酸性とし、遠心分離によって菌体を
除いた後、上澄液を陽イオン交換樹脂、ダイヤイオン５
ＫＩＢのＨ型（三菱化或社ＩＩ）に通し、脱カチオンし
た後、陰イオン交換樹脂ダイヤイオンＰＡ４１２のギ酸
型（三菱化成社製）に吸着させ、６Ｎギ酸により溶出し
た。この溶出液を活性炭で脱色後、減圧濃縮し、Ｌ −
Ｑンゴ酸を析出させた。乾燥後、２９ｇのＬ−リンゴ酸
を回収した。結晶母液には未晶出のＬ−リンゴ酸２３ｇ
が含まれていた。Example 1 Glucose 120g, ammonium chloride 11.6g 1-potassium phosphate 0.5g, magnesium sulfate 7-eternal salt 0
．． 2g, iron sulfate 7 permanent 0.01g and yeast extract 1
A solution of 30 g dissolved in tap water IIl was placed in a 300-capacity Erlenmeyer flask, and the mixture was heated at 120°C at 1.2 kg/c.
One platinum loop of Ustilago chrysugalii 391, which had been grown on a potato dextrose agar slant medium, was inoculated into the medium obtained by sterilizing the medium for 10 minutes. At the same time, 1 g of separately sterilized calcium carbonate was added. This culture solution was cultured with shaking on a rotary shaker at 25°C for 7 hours. After the culture was completed, the culture solution was analyzed and found that culture solution 11
52 g of L-U malic acid was produced per sample. The obtained culture solution 11 was made acidic with hydrochloric acid, and after removing the bacterial cells by centrifugation, the supernatant was treated with a cation exchange resin, Diaion 5
After decationization by passing through KIB H type (Mitsubishi Kasei Corporation II), it was adsorbed onto an anion exchange resin Diaion PA412 formic acid type (manufactured by Mitsubishi Kasei Corporation) and eluted with 6N formic acid. This eluate was decolorized with activated carbon, concentrated under reduced pressure, and L-
Q ngoic acid was precipitated. After drying, 29 g of L-malic acid was recovered. Crystal mother liquor contains 23g of uncrystallized L-malic acid.
was included.
実施例２
実施例１と同じ組成の培地３０ｒＩｉ！を３００−容三
角フラスコに入れ１２０℃、１．２ｋｇ／ｃＩ＋！で１
０分間殺菌して得られた培地に、予めポテトデキストロ
ース寒天の斜面培地上で生育させたトリポスポリウム・
ブラタムＴＫ３０４を一白金耳植菌した。Example 2 Medium 30rIi with the same composition as Example 1! into a 300-capacity Erlenmeyer flask at 120℃, 1.2kg/cI+! de1
To the medium obtained by sterilization for 0 minutes, Tryposporium spp., which had been grown on a potato dextrose agar slant medium, was added.
One platinum loop of Bratum TK304 was inoculated.
同時に別に殺菌した炭酸カルシウム１ｇを加えた。At the same time, 1 g of separately sterilized calcium carbonate was added.
この培養液をロータリーシェーカー上で２５℃、７日間
振盪培養した。培養終了後、培養液を分析した結果、培
養液１１当り５１ｇのＬ −リンゴ酸が生成していた。This culture solution was cultured with shaking on a rotary shaker at 25°C for 7 days. After the culture was completed, the culture solution was analyzed and it was found that 51 g of L-malic acid was produced per 11 culture solutions.
得られた培養液１ｊ！を遠心分離して菌体を除去し、上
澄液を硫酸で酸性とした後、減圧濃縮し、活性炭で脱色
後、エーテルで抽出した。エーテル抽出液を減圧乾固し
、Ｌ −Ｕンゴ酸３５ｇを得た。Obtained culture solution 1j! The cells were removed by centrifugation, and the supernatant was made acidic with sulfuric acid, concentrated under reduced pressure, decolorized with activated carbon, and extracted with ether. The ether extract was dried under reduced pressure to obtain 35 g of L-U malic acid.
実施例３
ウスティラゴ・クリスーガリイに３９１に替えて下記第
１表に示した菌株を用いる以外は実施例１と同様に培養
して、培養液中に生成蓄積されるＬ−リンゴ酸の量を測
定した。Example 3 Ustilago chrisugalii was cultured in the same manner as in Example 1 except that the strains shown in Table 1 below were used instead of 391, and the amount of L-malic acid produced and accumulated in the culture solution was measured. .
結果を第１表に示す。The results are shown in Table 1.
菌株
ウスティラゴ・クリス〜カリイ に３９０に３９２
に３９３
に２１６
ウスティラゴ・ネルダイ Ｎ（１１１５ウステイラ
ゴ・スフ７エロゲナ に４０１に４０２
ウスティラゴ・コイシス に３３１に３３２
ウスティラゴ・スキタミネ７ Ｋ３３５に３３６
に３３８
に３３９
ウスティラゴ・エスキｌレンタ Ｋ２Ｏ２に５０１
ウスティラゴ・シンセリスマエ に３８２に３８３
トリボスボリウム・ブラタム
にＴ３２６
０
Ｔ３２７
９
にＴ３２８
６
発
明
の効果
本発明によれば、安価な糖質を原料として、ＩＪンゴ酸
を収率よく直接発酵生産することかできる。Strains Ustilago chris-carii 390 to 392 393 to 216 Ustilago nerdai N (1115 Ustilago suf7 erogena 401 to 402 Ustilago koicis 331 to 332 Ustilago scythamine 7 K335 to 336 to 338 to 339 Ustilago eschi L Rental 501 to K2O2 382 to 383 to Ustilago synthelismae T326 to Tribosborium bratum 0 T327 9 to T328 6 Effects of the Invention According to the present invention, IJ ngoic acid can be directly produced in high yield using inexpensive carbohydrates as raw materials. It can be produced by fermentation.
Claims (1)
リンゴ酸生産能を有する微生物を培地に培養し、培養物
中にＬ−リンゴ酸を生成蓄積させ、該培養物よりＬ−リ
ンゴ酸を採取することを特徴とするＬ−リンゴ酸の製造
法。Belongs to the genus Ustilago or Tryposporius, L-
A method for producing L-malic acid, which comprises culturing a microorganism capable of producing malic acid in a medium, producing and accumulating L-malic acid in the culture, and collecting L-malic acid from the culture.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP31924489A JPH03180187A (en) | 1989-12-08 | 1989-12-08 | Production of l-malic acid |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP31924489A JPH03180187A (en) | 1989-12-08 | 1989-12-08 | Production of l-malic acid |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH03180187A true JPH03180187A (en) | 1991-08-06 |
Family
ID=18108024
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP31924489A Pending JPH03180187A (en) | 1989-12-08 | 1989-12-08 | Production of l-malic acid |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH03180187A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2011043443A1 (en) * | 2009-10-07 | 2011-04-14 | 三菱化学株式会社 | Method for producing aliphatic dicarboxylic acid |
-
1989
- 1989-12-08 JP JP31924489A patent/JPH03180187A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2011043443A1 (en) * | 2009-10-07 | 2011-04-14 | 三菱化学株式会社 | Method for producing aliphatic dicarboxylic acid |
| JP5724876B2 (en) * | 2009-10-07 | 2015-05-27 | 三菱化学株式会社 | Method for producing succinic acid |
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