JPH0371880B2 - - Google Patents
Info
- Publication number
- JPH0371880B2 JPH0371880B2 JP58243118A JP24311883A JPH0371880B2 JP H0371880 B2 JPH0371880 B2 JP H0371880B2 JP 58243118 A JP58243118 A JP 58243118A JP 24311883 A JP24311883 A JP 24311883A JP H0371880 B2 JPH0371880 B2 JP H0371880B2
- Authority
- JP
- Japan
- Prior art keywords
- fermentation
- immobilized
- gel
- solution
- microbial cells
- 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
- 238000000855 fermentation Methods 0.000 claims description 49
- 230000004151 fermentation Effects 0.000 claims description 49
- 239000007788 liquid Substances 0.000 claims description 30
- 230000000813 microbial effect Effects 0.000 claims description 28
- 239000000758 substrate Substances 0.000 claims description 23
- 239000000243 solution Substances 0.000 claims description 21
- 239000000463 material Substances 0.000 claims description 14
- 238000000034 method Methods 0.000 claims description 12
- 238000004519 manufacturing process Methods 0.000 claims description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 6
- 239000000661 sodium alginate Substances 0.000 claims description 6
- 235000010413 sodium alginate Nutrition 0.000 claims description 6
- 229940005550 sodium alginate Drugs 0.000 claims description 6
- 235000013555 soy sauce Nutrition 0.000 claims description 6
- -1 Farcellulan Substances 0.000 claims description 3
- 239000007853 buffer solution Substances 0.000 claims description 3
- 239000002994 raw material Substances 0.000 claims description 3
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 2
- 239000000679 carrageenan Substances 0.000 claims description 2
- 235000010418 carrageenan Nutrition 0.000 claims description 2
- 229920001525 carrageenan Polymers 0.000 claims description 2
- 229940113118 carrageenan Drugs 0.000 claims description 2
- 230000003301 hydrolyzing effect Effects 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 2
- UHVMMEOXYDMDKI-JKYCWFKZSA-L zinc;1-(5-cyanopyridin-2-yl)-3-[(1s,2s)-2-(6-fluoro-2-hydroxy-3-propanoylphenyl)cyclopropyl]urea;diacetate Chemical compound [Zn+2].CC([O-])=O.CC([O-])=O.CCC(=O)C1=CC=C(F)C([C@H]2[C@H](C2)NC(=O)NC=2N=CC(=CC=2)C#N)=C1O UHVMMEOXYDMDKI-JKYCWFKZSA-L 0.000 claims description 2
- 210000004027 cell Anatomy 0.000 description 37
- 239000000499 gel Substances 0.000 description 28
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 24
- 210000005253 yeast cell Anatomy 0.000 description 15
- 239000004310 lactic acid Substances 0.000 description 12
- 235000014655 lactic acid Nutrition 0.000 description 12
- 241000894006 Bacteria Species 0.000 description 11
- 244000005700 microbiome Species 0.000 description 10
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 8
- 239000003349 gelling agent Substances 0.000 description 8
- 102000004190 Enzymes Human genes 0.000 description 7
- 108090000790 Enzymes Proteins 0.000 description 7
- 230000001580 bacterial effect Effects 0.000 description 7
- 229940088598 enzyme Drugs 0.000 description 7
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 6
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 5
- 239000007864 aqueous solution Substances 0.000 description 5
- 239000011248 coating agent Substances 0.000 description 5
- 238000000576 coating method Methods 0.000 description 5
- 239000011259 mixed solution Substances 0.000 description 5
- 150000003839 salts Chemical class 0.000 description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 4
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 4
- 235000019270 ammonium chloride Nutrition 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 238000003756 stirring Methods 0.000 description 4
- 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 3
- 239000007983 Tris buffer Substances 0.000 description 3
- 239000001110 calcium chloride Substances 0.000 description 3
- 229910001628 calcium chloride Inorganic materials 0.000 description 3
- 235000011148 calcium chloride Nutrition 0.000 description 3
- 229940041514 candida albicans extract Drugs 0.000 description 3
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 239000000741 silica gel Substances 0.000 description 3
- 229910002027 silica gel Inorganic materials 0.000 description 3
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 3
- 239000012138 yeast extract Substances 0.000 description 3
- FTOAOBMCPZCFFF-UHFFFAOYSA-N 5,5-diethylbarbituric acid Chemical compound CCC1(CC)C(=O)NC(=O)NC1=O FTOAOBMCPZCFFF-UHFFFAOYSA-N 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- 235000008331 Pinus X rigitaeda Nutrition 0.000 description 2
- 235000011613 Pinus brutia Nutrition 0.000 description 2
- 241000018646 Pinus brutia Species 0.000 description 2
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 2
- OSGAYBCDTDRGGQ-UHFFFAOYSA-L calcium sulfate Chemical compound [Ca+2].[O-]S([O-])(=O)=O OSGAYBCDTDRGGQ-UHFFFAOYSA-L 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 235000021107 fermented food Nutrition 0.000 description 2
- 238000011049 filling Methods 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000007654 immersion Methods 0.000 description 2
- 239000010410 layer Substances 0.000 description 2
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 2
- 235000019341 magnesium sulphate Nutrition 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 2
- 235000019796 monopotassium phosphate Nutrition 0.000 description 2
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 2
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 2
- 238000005096 rolling process Methods 0.000 description 2
- 238000011218 seed culture Methods 0.000 description 2
- 239000007921 spray Substances 0.000 description 2
- 238000009495 sugar coating Methods 0.000 description 2
- 241000186361 Actinobacteria <class> Species 0.000 description 1
- 108090000145 Bacillolysin Proteins 0.000 description 1
- 108091005658 Basic proteases Proteins 0.000 description 1
- 241000221198 Basidiomycota Species 0.000 description 1
- 241000195493 Cryptophyta Species 0.000 description 1
- 239000004278 EU approved seasoning Substances 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 239000005909 Kieselgur Substances 0.000 description 1
- 108091005507 Neutral proteases Proteins 0.000 description 1
- 102000035092 Neutral proteases Human genes 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 241000209140 Triticum Species 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 239000008351 acetate buffer Substances 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 235000013334 alcoholic beverage Nutrition 0.000 description 1
- 108090000637 alpha-Amylases Proteins 0.000 description 1
- 102000004139 alpha-Amylases Human genes 0.000 description 1
- 229940024171 alpha-amylase Drugs 0.000 description 1
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 description 1
- DIZPMCHEQGEION-UHFFFAOYSA-H aluminium sulfate (anhydrous) Chemical compound [Al+3].[Al+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O DIZPMCHEQGEION-UHFFFAOYSA-H 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 229960002319 barbital Drugs 0.000 description 1
- 108010019077 beta-Amylase Proteins 0.000 description 1
- 238000005842 biochemical reaction Methods 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 229940088623 biologically active substance Drugs 0.000 description 1
- 210000001124 body fluid Anatomy 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 229910021538 borax Inorganic materials 0.000 description 1
- VSGNNIFQASZAOI-UHFFFAOYSA-L calcium acetate Chemical compound [Ca+2].CC([O-])=O.CC([O-])=O VSGNNIFQASZAOI-UHFFFAOYSA-L 0.000 description 1
- 239000001639 calcium acetate Substances 0.000 description 1
- 235000011092 calcium acetate Nutrition 0.000 description 1
- 229960005147 calcium acetate Drugs 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 238000007444 cell Immobilization Methods 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000000919 ceramic Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000004927 clay Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000007667 floating Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000011194 food seasoning agent Nutrition 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- NLYAJNPCOHFWQQ-UHFFFAOYSA-N kaolin Chemical compound O.O.O=[Al]O[Si](=O)O[Si](=O)O[Al]=O NLYAJNPCOHFWQQ-UHFFFAOYSA-N 0.000 description 1
- 229910052622 kaolinite Inorganic materials 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 238000003760 magnetic stirring Methods 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 229910044991 metal oxide Inorganic materials 0.000 description 1
- 150000004706 metal oxides Chemical class 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000005373 porous glass Substances 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000002250 progressing effect Effects 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 235000010339 sodium tetraborate Nutrition 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 239000002344 surface layer Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 239000000057 synthetic resin Substances 0.000 description 1
- 229920003002 synthetic resin Polymers 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- WUUHFRRPHJEEKV-UHFFFAOYSA-N tripotassium borate Chemical compound [K+].[K+].[K+].[O-]B([O-])[O-] WUUHFRRPHJEEKV-UHFFFAOYSA-N 0.000 description 1
- BSVBQGMMJUBVOD-UHFFFAOYSA-N trisodium borate Chemical compound [Na+].[Na+].[Na+].[O-]B([O-])[O-] BSVBQGMMJUBVOD-UHFFFAOYSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Description
本発明は固定化微生物菌体を用いる発酵方法に
関し、その目的とするところは液体基質の発酵効
率を長期間にわたつて著しく高めた発酵方法を提
供することにある。
近年、固定化微生物菌体を種々の発酵に用いる
バイオリアクター（生化学反応装置）は、食品工
業、医薬品工業等においてその利用が種々試みら
れ、又該リアクターに用いられる微生物の固定化
法に関する開発も急速に進んでいる。
しかしながら、従来のバイオリアクターを用い
る発酵法においては、該リアクターに用いられる
固定化微生物菌体の耐久持続性並びに液体基質の
発酵効率の点等、種々難点が残されている。
そこで本発明者等は、このような従来技術の欠
点を解消するため、鋭意検討を重ねた結果、多孔
質の粒状支持体の表面に、微生物菌体とゲル基材
との混合液を塗布し、これをゲル化剤と接触させ
ると著しくゲル強度が大きく、しかも著しく変
形、膨潤し難い固定化微生物菌体が得られるこ
と、更に該固定化微生物菌体を液体基質に分散、
浮遊させつつ接触させれば著しく発酵効率が促進
されること等を知り、これら知見に基いて本発明
を完成した。
即ち、本発明は、微生物菌体又はこれに水もし
くは緩衝液を加えたものをゲル基材含有液に混合
した液を、多孔質の粒状支持体の表面に塗付し、
これをゲル化剤と接触させて得られる球状の固定
化微生物菌体を、液体基質に連続的もしくは断続
的に分散、浮遊させつつ接触させて発酵を行なう
ことを特徴とする発酵方法である。
以下、本発明について詳細に説明する。
先ず、本発明に用いられる微生物菌体として
は、酵母、細菌、かび、放線菌、担子菌、単細胞
藻類、ウイルス等、如何なる種別の菌体でも良
く、又これらの微生物の培養物、該培養物より抽
出して得られる酵素、あるいは生物組織から抽出
して得られる酵素であつても良い。なお、上記微
生物菌体のうち、発酵食品の製造に用いられる微
生物、例えば、醤油製造用酵母、醤油製造用乳酸
菌等が特に好適な例として挙げられる。
本発明では、上記微生物菌体又はこれに水もし
くは緩衝液を加えたものを、ゲル基材含有液に加
え、混合して微生物菌体及びゲル基材を含有する
混合液を得る。
これに用いる緩衝液としては、例えば酢酸緩衝
液、マツキルブエイン緩衝液、リン酸緩衝液、ト
リス緩衝液、ベロナール緩衝液等が用いられる。
又、ゲル基材としては、通常、ゲル化剤の存在
なしでは常温でゲル化しないものが用いられ、例
えばアルギン酸ナトリウム、フアーセルラン、ポ
リビニルアルコール、イオータカラギーナン等が
好適な例として挙げられ、これらのゲル基材を
水、緩衝液等を用いて通常0.1〜10％（Ｗ／Ｖ）
程度のゲル基材含有液とする。
次に、上記のようにして得られた微生物菌体及
びゲル基材を含有する混合液を、多孔質の粒状支
持体の表面に、好ましくは0.1〜２mm程度の厚さ
に塗付し、これをゲル化剤と接触させて球状の固
定化微生物菌体を得る。
これに用いる多孔質の粒状支持体としては、例
えば多孔質ガラス、シリカゲル、アルミナ、活性
炭、カオリナイト、酸性白土、珪藻土、リン酸カ
ルシウム、金属酸化物、セラミツクス、イオン交
換樹脂、多孔質合成樹脂、鋸くず等の無機質もし
くは有機質の粒状支持体が代表例として挙げられ
る。そしてこの粒状支持体の大きさは任意で良
く、望ましくは直径が0.3〜８mm程度、特に２〜
５mm程度が最も好ましい。
微生物菌体及びゲル基材を含有する混合液を、
多孔質の粒状支持体の表面に塗付する手段として
は、例えば該混合液中に直接粒状支持体を適当時
間浸漬する方法、又大量にしかも均一にコーテイ
ングする手段として糖衣機、マツシユルーム・ミ
キサー等のパンコーテイング装置を用いる方法等
が挙げられるが、これらの装置を用いる場合、該
装置内の傾斜軸を中心に回転し、転動するパン内
に粒状支持体を入れ、これに上記混合液を加えて
塗付する方法、あるいは又流動層コーテイング装
置を用いる場合には、該装置内の静置多孔板もし
くは回転多孔板より圧縮空気を吹上げることによ
り流動化させた粒状支持体に、上方のスプレーノ
ズルより上記混合液を噴霧し、塗付する方法等が
挙げられる。
また、ゲル化剤としては、塩化カルシウム、塩
化カリウム、塩化アンモニウム、硫酸アルミニウ
ム、酢酸カルシウム、ホウ酸カリウム、ホウ酸ナ
トリウム、塩化マグネシウム等が用いられ、これ
らは通常１〜10％（Ｗ／Ｖ）程度の濃度の溶液と
して用いられる。
次に、上記微生物菌体及びゲル基材を含有する
混合液を塗付した粒状支持体をゲル化剤と接触さ
せる手段としては、任意の手段をとることができ
るが、通常、該支持体をゲル化剤含有溶液中に投
下、浸漬することにより、球状に固定化された微
生物菌体を得る。
上記操作により得られた固定化微生物菌体は、
粒状支持体の表面に、膜状の微生物菌体含有ゲル
を強固に付着させた球状の固定化微生物菌体であ
り、著しく機械的ゲル強度が大きく、しかも著し
く変形、膨潤し難いものである。
なお、後述の発酵操作に先だつて、該固定化微
生物菌体の菌体数が不足する場合には、予じめ目
的とする微生物の培養に適した培養培地で該固定
化微生物菌体を適当時間、前培養を行なつて菌体
を増殖させても良い。
次に、上記のようにして得た球状の固定化微生
物菌体を、液体基質に分散、浮遊状態に保持し得
る発酵容器、例えば撹拌槽、流動層等の発酵槽に
投入し、これに液体基質を導入して連続的もしく
は断続的に分散、浮遊させつつ該固定化微生物菌
体を液体基質に接触させて発酵を行なう。
本発明に用いられる液体基質としては、例えば
発酵による調味料の製造、発酵による酒類の製
造、アルコール発酵、有機酸発酵、アミノ酸発
酵、核酸発酵、生理活性物質発酵、抗生物質発酵
等、種々の発酵法において通常用いられるもので
あれば、何れの液体基質も用いることができる。
これらの液体基質のうち、特に好ましいものと
して、発酵食品の製造に用いられる液体基質、例
えば醤油製造用原料を酵素的もしくは化学的に加
水分解したものを、PH３〜７の液体状態としたも
の、あるいはこれに酵母を加えて予じめ発酵させ
た発酵中ないしは発酵後のPH３〜７の液体基質等
が挙げられる。
また、本発明においては、上記発酵容器の固定
化微生物菌体を、連続的もしくは断続的に分散、
浮遊させて液体基質と接触させることが必須で、
これにより発酵効率が著しく高められ、ひいては
発酵期間を著しく短縮させることが出来る。
なお、上記固定化微生物菌体と液体基質との接
触、発酵条件は、目的とする微生物、発酵生成物
等の種別により、発酵基質中のPH、発酵温度、発
酵時間等を適宜選択すれば良い。
上述した如く、本発明によれば著しくゲル強度
の大きい固定化微生物菌体をバイオリアクター内
で使用しているため、長期間の発酵に耐えるこ
と、更に該固定化微生物菌体を液体基質に分散、
浮遊させつつ接触させて発酵を行なうことによ
り、著しく基質の発酵効率を高め、発酵に要する
処理期間を短縮させることが出来る等、本発明は
産業上極めて有意義である。
以下、実施例を挙げて本発明をさらに具体的に
説明する。
実施例 １
生醤油10％（Ｖ／Ｖ）、グルコース７％（Ｗ／
Ｖ）、食塩８％（Ｗ／Ｖ）、リン酸１カリウム0.1
％（Ｗ／Ｖ）、硫酸マグネシウム0.05％（Ｗ／
Ｖ）、硫酸カルシウム0.01％（Ｗ／Ｖ）、及び酵母
エキス0.1％（Ｗ／Ｖ）を含む増殖培地（PH＝
5.3）１を３容振盪フラスコに入れ、常法に
よりオートクレーブで殺菌した後、該培地に酵母
サツカロミセス・ルキシーATCC13356の種培養
液20mlを接種し、30℃で72時間振盪培養した。得
られた培養液１を12000rpmで15分間遠心分離
して集菌した（総菌体数9.5×109／ml）。得られ
た濃縮菌体液30mlを、加熱殺菌した1.5％（Ｗ／
Ｖ）アルギン酸ナトリウム水溶液1500mlに懸濁し
た。
次に、予め２％塩化カルシウム溶液に浸漬後、
表面の液を切つた直径３mmのポリエチレン多孔質
小球〔筒中プラスチツク工業(株)製・サンロイドセ
ル〕160ｇをコーテイング用パン（菊水製作所
製・糖衣機）に入れ、容器を転動させつつ開口部
よりスプレーノズルを用いて上記の菌体懸濁アル
ギン酸ナトリウム溶液550mlを噴霧して多孔質小
球表面に均一に膜状に塗布した。かくして得られ
た表面に約１mm厚みの菌体懸濁アルギン酸ナトリ
ウム層を形成した小球を１個ずつにほぐしなが
ら、冷却した２％塩化カルシウム水溶液に落下さ
せてゲル化させ、さらに一昼夜放置して十分ゲル
層を硬化させ、直径４mmの球状の酵母菌体の固定
化ゲルを得た（実施例１の固定化酵母菌体）。
一方、対照として、上記酵母菌体懸濁アルギン
酸ナトリウム溶液950mlを定量ポンプを用いて内
径２mmのノズルより冷却した２％塩化カルシウム
溶液中に滴下させてゲル化し、一昼夜放置して直
径４mmの球状の酵母菌体ゲルを調製した（対照の
固定化酵母菌体）。
次に、内径54mm、高さ437mmのジヤケツト付ガ
ラス製カラム（内容積1000ml）２本に、各々固定
化酵母菌体（実施例１の固定化酵母菌体、及び対
照の固定化酵母菌体）を充填したら、その充填高
さはほぼ等しく270mmとなつた。これらの固定化
酵母菌体を、殺菌した酵母発酵基質溶液〔グルコ
ース10％（Ｗ／Ｖ）、食塩７％（Ｗ／Ｖ）、リン酸
１カリウム0.11％（Ｗ／Ｖ）、硫酸マグネシウム
0.03％（Ｗ／Ｖ）、塩化カルシウム0.02％（Ｗ／
Ｖ）、酵母エキス0.32％（Ｗ／Ｖ）、乳酸0.2％
（Ｖ／Ｖ）、PH5.3〕で３回リンスした後、カラム
空間を満たし、引続き各カラム底部より１時間当
り10分間の割合で200ml／分の除菌空気を断続的
に導入して固（固定化酵母菌体）・液・気の三相
流動層を形成させた。ジヤケツトに30℃の恒温水
を通し、各カラムに250ml／時間の上記酵母発酵
基質溶液を通液（S.V.＝0.25／Hr）しつつ１ケ
月間の連続発酵を実施した。
発酵終了時の固定化酵母菌体ゲルを観察したと
ころ、実施例１の固定化酵母菌体には何ら変化が
認められなかつたが、対照の固定化酵母菌体はや
や膨潤の傾向にあり、はじけて割れたゲルが約15
％あつた。経時的な発酵の経過を第１表に示す。
The present invention relates to a fermentation method using immobilized microbial cells, and an object thereof is to provide a fermentation method that significantly increases the fermentation efficiency of a liquid substrate over a long period of time. In recent years, various attempts have been made to use bioreactors (biochemical reaction devices) that use immobilized microbial cells for various fermentations in the food industry, pharmaceutical industry, etc., and there has also been development of methods for immobilizing microorganisms used in such reactors. is also progressing rapidly. However, in the conventional fermentation method using a bioreactor, various drawbacks remain, such as the durability of the immobilized microbial cells used in the reactor and the fermentation efficiency of the liquid substrate. In order to overcome these drawbacks of the conventional technology, the inventors of the present invention, as a result of extensive studies, applied a mixed solution of microbial cells and a gel base material to the surface of a porous granular support. , when this is brought into contact with a gelling agent, immobilized microbial cells that have a significantly high gel strength and are not easily deformed or swell can be obtained; furthermore, the immobilized microbial cells are dispersed in a liquid matrix;
The present invention was completed based on the knowledge that the fermentation efficiency was significantly promoted when the materials were brought into contact with each other while being suspended. That is, in the present invention, a solution obtained by mixing microbial cells or a mixture thereof with water or a buffer solution into a solution containing a gel base material is applied to the surface of a porous granular support,
This is a fermentation method characterized by carrying out fermentation by contacting spherical immobilized microbial cells obtained by contacting the microbial cells with a gelling agent while continuously or intermittently dispersing and suspending them in a liquid substrate. The present invention will be explained in detail below. First, the microorganism used in the present invention may be any type of microorganism such as yeast, bacteria, mold, actinomycetes, basidiomycetes, unicellular algae, viruses, etc., and cultures of these microorganisms and cultures. The enzyme may be an enzyme obtained by extraction from a biological tissue, or an enzyme obtained by extraction from a biological tissue. Among the microorganisms mentioned above, microorganisms used in the production of fermented foods, such as yeast for soy sauce production, lactic acid bacteria for soy sauce production, etc., are particularly preferred. In the present invention, the above-mentioned microbial cells or the mixture thereof with water or a buffer solution are added to the gel base material-containing solution and mixed to obtain a mixed solution containing the microbial cells and the gel base material. Examples of buffers used for this purpose include acetate buffer, pine kilbuein buffer, phosphate buffer, Tris buffer, and veronal buffer. In addition, the gel base material is usually one that does not gel at room temperature without the presence of a gelling agent, such as sodium alginate, facellulan, polyvinyl alcohol, iota carrageenan, etc. Gel base material is usually 0.1 to 10% (W/V) using water, buffer, etc.
The liquid contains a gel base material of about 100%. Next, the mixed solution containing the microbial cells and gel base material obtained as described above is applied onto the surface of the porous granular support to a thickness of preferably about 0.1 to 2 mm. is brought into contact with a gelling agent to obtain spherical immobilized microbial cells. Porous granular supports used for this purpose include, for example, porous glass, silica gel, alumina, activated carbon, kaolinite, acid clay, diatomaceous earth, calcium phosphate, metal oxides, ceramics, ion exchange resins, porous synthetic resins, and sawdust. Typical examples include inorganic or organic granular supports such as . The size of this granular support may be arbitrary, preferably about 0.3 to 8 mm in diameter, particularly 2 to 8 mm in diameter.
A thickness of about 5 mm is most preferable. A mixed solution containing microbial cells and gel base material,
Examples of means for coating the surface of the porous granular support include a method of directly immersing the granular support in the mixed solution for an appropriate period of time, and means for uniformly coating a large amount such as a sugar coating machine, pine room mixer, etc. When using these devices, a granular support is placed in a rolling pan that rotates around an inclined axis within the device, and the above mixed liquid is poured into the pan. In addition, if a fluidized bed coating device is used, compressed air is blown up from a stationary perforated plate or a rotating perforated plate in the device to fluidize the granular support. Examples include a method of spraying and applying the above-mentioned liquid mixture from a spray nozzle. In addition, as gelling agents, calcium chloride, potassium chloride, ammonium chloride, aluminum sulfate, calcium acetate, potassium borate, sodium borate, magnesium chloride, etc. are used, and these are usually 1 to 10% (W/V). It is used as a solution with a certain concentration. Next, any means can be used to bring the granular support coated with the mixture containing the microbial cells and gel base material into contact with the gelling agent, but usually, the support is brought into contact with the gelling agent. By dropping and immersing in a gelling agent-containing solution, spherically immobilized microbial cells are obtained. The immobilized microbial cells obtained by the above procedure are
This is a spherical immobilized microbial cell formed by firmly adhering a membranous microbial cell-containing gel to the surface of a granular support, which has extremely high mechanical gel strength and is extremely resistant to deformation and swelling. If the number of immobilized microorganisms is insufficient prior to the fermentation operation described below, the immobilized microorganisms may be grown in advance in a culture medium suitable for culturing the desired microorganism. The bacterial cells may be grown by pre-culturing for a period of time. Next, the spherical immobilized microbial cells obtained as described above are placed in a fermentation vessel that can be dispersed and maintained in a suspended state in a liquid substrate, such as a fermentation tank such as a stirring tank or a fluidized bed. Fermentation is carried out by introducing a substrate and bringing the immobilized microbial cells into contact with the liquid substrate while continuously or intermittently dispersing and suspending it. The liquid substrate used in the present invention can be used in various fermentations, such as the production of seasonings by fermentation, the production of alcoholic beverages by fermentation, alcohol fermentation, organic acid fermentation, amino acid fermentation, nucleic acid fermentation, biologically active substance fermentation, antibiotic fermentation, etc. Any liquid substrate commonly used in the process can be used. Among these liquid substrates, particularly preferred are liquid substrates used in the production of fermented foods, such as those obtained by enzymatically or chemically hydrolyzing raw materials for the production of soy sauce and made into a liquid state with a pH of 3 to 7. Alternatively, a liquid substrate having a pH of 3 to 7 during or after fermentation may be used, which is pre-fermented by adding yeast to the substrate. Further, in the present invention, the immobilized microbial cells in the fermentation container are continuously or intermittently dispersed,
It is essential that it be suspended and brought into contact with a liquid substrate;
This significantly increases fermentation efficiency and, in turn, allows the fermentation period to be significantly shortened. The contact between the immobilized microbial cells and the liquid substrate and the fermentation conditions may be appropriately selected depending on the type of target microorganism, fermentation product, etc., such as the pH of the fermentation substrate, fermentation temperature, fermentation time, etc. . As mentioned above, according to the present invention, since immobilized microbial cells with extremely high gel strength are used in the bioreactor, the immobilized microbial cells can withstand long-term fermentation, and furthermore, the immobilized microbial cells can be dispersed in a liquid substrate. ,
By carrying out fermentation by contacting the substrate while floating it, the fermentation efficiency of the substrate can be significantly increased and the processing period required for fermentation can be shortened, and the present invention is extremely significant industrially. Hereinafter, the present invention will be explained in more detail with reference to Examples. Example 1 Raw soy sauce 10% (V/V), glucose 7% (W/V)
V), salt 8% (W/V), monopotassium phosphate 0.1
% (W/V), magnesium sulfate 0.05% (W/V)
V), calcium sulfate 0.01% (W/V), and yeast extract 0.1% (W/V) (PH=
5.3) 1 was placed in a 3-volume shaking flask and sterilized in an autoclave according to a conventional method. The medium was inoculated with 20 ml of a seed culture of the yeast Saccharomyces luxii ATCC13356, and cultured with shaking at 30°C for 72 hours. The obtained culture solution 1 was centrifuged at 12,000 rpm for 15 minutes to collect bacteria (total number of bacterial cells: 9.5×10 9 /ml). 30ml of the obtained concentrated bacterial body fluid was heated and sterilized to 1.5% (W/
V) Suspended in 1500 ml of sodium alginate aqueous solution. Next, after pre-immersion in a 2% calcium chloride solution,
Put 160 g of polyethylene porous spheres with a diameter of 3 mm (Sunroid Cell, manufactured by Tsutsunaka Plastic Industries Co., Ltd.) from which the surface liquid has been drained into a coating pan (manufactured by Kikusui Seisakusho, sugar coating machine), and open the container by rolling it. Using a spray nozzle, 550 ml of the above sodium alginate solution containing bacterial cell suspension was sprayed to uniformly coat the surface of the porous globules in the form of a film. The thus-obtained globules with a layer of sodium alginate suspended in bacterial cells formed on the surface having a thickness of about 1 mm were loosened one by one and dropped into a cooled 2% calcium chloride aqueous solution to form a gel, and then left to stand overnight. The gel layer was sufficiently hardened to obtain a spherical yeast cell immobilization gel with a diameter of 4 mm (immobilized yeast cells of Example 1). On the other hand, as a control, 950 ml of the above yeast cell suspension sodium alginate solution was dropped into a cooled 2% calcium chloride solution using a metering pump through a nozzle with an inner diameter of 2 mm, gelatinized, and left overnight to form a spherical solution with a diameter of 4 mm. A yeast cell gel was prepared (immobilized yeast cells as a control). Next, the immobilized yeast cells (the immobilized yeast cells of Example 1 and the immobilized yeast cells of the control) were placed in two jacketed glass columns (inner volume 1000 ml) with an inner diameter of 54 mm and a height of 437 mm. After filling, the filling height was almost the same, 270 mm. These immobilized yeast cells were added to a sterilized yeast fermentation substrate solution [glucose 10% (W/V), salt 7% (W/V), monopotassium phosphate 0.11% (W/V), magnesium sulfate.
0.03% (W/V), calcium chloride 0.02% (W/V)
V), yeast extract 0.32% (W/V), lactic acid 0.2%
(V/V), pH 5.3] three times, fill the column space, and then intermittently introduce sterilized air from the bottom of each column for 10 minutes per hour to solidify. A three-phase fluidized bed of (immobilized yeast cells), liquid, and air was formed. Constant temperature water at 30° C. was passed through the jacket, and 250 ml/hour of the above yeast fermentation substrate solution was passed through each column (SV=0.25/Hr), while continuous fermentation was carried out for one month. When observing the immobilized yeast cell gel at the end of fermentation, no change was observed in the immobilized yeast cells of Example 1, but the control immobilized yeast cells tended to swell slightly. Approximately 15 gels were popped and cracked.
% hot. The progress of fermentation over time is shown in Table 1.
【表】
第１表より明らかな如く、実施例１の固定化酵
母菌体を断続的に分散、浮遊させつつ発酵させる
と、対照の固定化酵母菌体を用いた場合に比しア
ルコール生産量が著しく向上、しかも発酵期間が
長期にわたつてもアルコール生成能は高水準に保
存されることが認められた。
実施例 ２
生醤油10％（Ｖ／Ｖ）、グルコース１％（Ｗ／
Ｖ）、食塩８％（Ｗ／Ｖ）、酢酸ナトリウム3.5％
（Ｗ／Ｖ）、及び酵母エキス0.3％（Ｗ／Ｖ）を含
む増殖培地（PH＝7.0）300mlを１容三角フラス
コに入れ、常法によりオートクレーブで殺菌した
後、該培地に乳酸菌ペデイオコツカス・ハロフイ
ルスIAM1693の種培養液10mlを接触し、30℃で
170時間静置培養した。得られた培養液300mlを
1200rpmで15分間遠心分離して湿潤乳酸菌菌体を
得た。
この湿潤乳酸菌菌体をトリス緩衝液（PH7.0）
に懸濁して遠心分離を行う洗滌操作を３度くり返
して洗滌乳酸菌菌体を得た。この洗滌菌体を上記
トリス緩衝液（PH7.0）30mlに懸濁し、これを加
熱殺菌した２％フアーセルラン水溶液270mlに混
合して乳酸菌懸濁液（総菌数1.2×109／ml）とし
た。
次に、予め1.5％塩化アンモニウム水溶液に浸
漬後、表面の液を切つた直径３mmの球状シリカゲ
ル〔水沢化学(株)製〕30ｇを上記乳酸菌懸濁フアー
セルラン溶液110mlに浸漬混合し、表面に約１mm
厚みに塗膜を形成した。得られたコーテイング済
みシリカゲルを１個ずつほぐしながら、冷却した
1.5％塩化アンモニウム水溶液に落下させて、表
層をゲル化させて一昼夜放置し、直径4.2mmの球
状の乳酸菌菌体の固定化ゲルを得た（実施例２の
固定化物）。
一方、対照として、上記乳酸菌懸濁フアーセル
ラン溶液190mlを注射筒を用いて、冷却した1.5％
塩化アンモニウム溶液中に滴下させてゲル化し、
一昼夜放置して直径4.2mmの乳酸菌菌体ゲルを調
製した（対照の固定化物）。
次に、脱脂大豆4.5Kgと小麦4.2Kgの混合物に水
4.5を加え、これを60容密閉容器に入れ、１
Kg／cm2・Ｇの水蒸気で30分間加熱後、よくほぐ
し、さらに１Kg／cm2・Ｇの水蒸気で45分間加熱処
理した後、冷却した。
一方、販売酵素製剤α−アミラーゼ〔三共製薬
(株)製〕50ｇ、β−アミラーゼ〔長瀬産業(株)製〕50
ｇ、アルカリプロテアーゼ〔長瀬産業(株)製〕50
ｇ、中性プロテアーゼ〔生化学工業(株)製〕50ｇ、
及び酸性プロテアーゼ〔盛進製薬(株)製〕50ｇを20
の水に溶解して得た酵素液をSA−451型無菌
過機〔日本水機工業(株)製〕で過して無菌酵素
液を得た。
この無菌酵素液13.8を上記冷却原料の全量に
加え、撹拌しつつ40℃で48時間酵素分解した。得
られた加水分解物に食塩2.37Kgを加えて食塩濃度
10％（Ｗ／Ｖ）とした後、圧搾して21.5の酵素
分解液汁を採取した〔液汁成分値：TN1.86％
（Ｗ／Ｖ）、RS10.31％（Ｗ／Ｖ）、TA1.82、PH
5.44〕。
次に、マグネツト式撹拌翼を備えたジヤケツト
付の内容積300mlガラス製密ぺい式反応器に、上
記酵素分解液汁で３回リンスした上記実施例２の
固定化物の全量を充填し、次いで該反応器に上記
酵素分解汁を投入した。そしてジヤケツトに30℃
の恒温水を通し、撹拌数を15rpm（一定）で撹拌
しながら該撹拌槽反応器に25ml／時間の上記酵素
分解液汁を通液しつつ液面を175ml相当の高さに
保ち（S.V.＝0.14／Hr）実施例２の固定化物を
浮遊、分散させて17日間の連続発酵を実施した。
一方、対照として、内容積175mlのジヤケツト
付カラムに、上記酵素分解液汁で３回リンスした
上記対照の固定化物の全量を充填し、上記酵素分
解液汁で空間を満たした。引続きジヤケツトに30
℃の恒温水を通した該充填カラム反応器に25ml／
時間の上記酵素分解液汁を通液しつつ（S.V.＝
0.14／Hr）17日間の連続発酵を同様に実施した。
発酵終了時の固定化菌体ゲルを観察したとこ
ろ、実施例２の固定化物は何ら変化が認められな
かつたが、対照の固定化物は、軟化が目立ち、約
10％のゲルは原形をとどめず溶解状態にあつた。
この経時的な発酵の経過を第２表に示す。[Table] As is clear from Table 1, when the immobilized yeast cells of Example 1 were fermented while being intermittently dispersed and suspended, the amount of alcohol produced was higher than when the immobilized yeast cells of Example 1 were used. It was observed that the alcohol production ability was significantly improved, and that the alcohol production ability was maintained at a high level even over a long fermentation period. Example 2 Raw soy sauce 10% (V/V), glucose 1% (W/
V), salt 8% (W/V), sodium acetate 3.5%
(W/V) and 300 ml of a growth medium (PH=7.0) containing yeast extract 0.3% (W/V) was placed in a 1-volume Erlenmeyer flask, and after sterilizing it in an autoclave in a conventional manner, the medium was added to Contact with 10ml of IAM1693 seed culture solution and heat at 30℃.
Static culture was performed for 170 hours. 300ml of the obtained culture solution
Wet lactic acid bacteria cells were obtained by centrifugation at 1200 rpm for 15 minutes. The wet lactic acid bacteria cells are mixed with Tris buffer (PH7.0).
The washing operation of suspending and centrifuging was repeated three times to obtain washed lactic acid bacteria cells. The washed bacterial cells were suspended in 30 ml of the above Tris buffer (PH7.0) and mixed with 270 ml of a heat-sterilized 2% Farcellulan aqueous solution to obtain a lactic acid bacteria suspension (total number of bacteria 1.2 x 10 9 /ml). . Next, 30 g of spherical silica gel (manufactured by Mizusawa Chemical Co., Ltd.) with a diameter of 3 mm, which had been immersed in a 1.5% ammonium chloride aqueous solution and drained of the liquid on the surface, was mixed by immersion in 110 ml of the above lactic acid bacteria-suspended Farcellulan solution, and the surface was coated approximately 1 mm.
A thick coating was formed. The coated silica gel obtained was loosened one by one and cooled.
The gel was dropped into a 1.5% ammonium chloride aqueous solution to gel the surface layer and left overnight to obtain a spherical gel with immobilized lactic acid bacteria cells having a diameter of 4.2 mm (immobilized product of Example 2). On the other hand, as a control, 190 ml of the above lactic acid bacteria suspended Farcellulan solution was added to the cooled 1.5%
Dropped into ammonium chloride solution to form a gel.
A lactic acid bacteria cell gel with a diameter of 4.2 mm was prepared by leaving it for a day and night (control immobilized product). Next, add water to a mixture of 4.5 kg of defatted soybeans and 4.2 kg of wheat.
Add 4.5, put this in a 60 volume airtight container, and add 1
After heating with water vapor of Kg/cm 2 ·G for 30 minutes, it was thoroughly loosened, further heat treated with steam of 1 kg/cm 2 ·G for 45 minutes, and then cooled. On the other hand, the sales enzyme preparation α-amylase [Sankyo Pharmaceutical Co., Ltd.
[manufactured by Nagase Sangyo Co., Ltd.] 50 g, β-amylase [manufactured by Nagase Sangyo Co., Ltd.] 50
g, alkaline protease [manufactured by Nagase Sangyo Co., Ltd.] 50
g, neutral protease [manufactured by Seikagaku Kogyo Co., Ltd.] 50 g,
and acidic protease [manufactured by Seishin Pharmaceutical Co., Ltd.] 50g for 20
The enzyme solution obtained by dissolving in water was passed through a sterile filter model SA-451 (manufactured by Nippon Suiki Kogyo Co., Ltd.) to obtain a sterile enzyme solution. 13.8 of this sterile enzyme solution was added to the total amount of the above-mentioned cooled raw material, and enzymatically decomposed at 40° C. for 48 hours while stirring. Add 2.37 kg of salt to the obtained hydrolyzate to determine the salt concentration.
After reducing the ratio to 10% (W/V), 21.5 enzymatically decomposed sap was collected by squeezing [Juice component value: TN 1.86%
(W/V), RS10.31% (W/V), TA1.82, PH
5.44〕. Next, the entire amount of the immobilized product of Example 2, which had been rinsed three times with the enzymatic decomposition liquid, was charged into a sealed glass reactor with an internal volume of 300 ml and equipped with a jacket and equipped with a magnetic stirring blade, and then the reaction The enzymatically decomposed juice was poured into a container. And 30℃ in the jacket
25 ml/hour of the enzymatically decomposed liquid was passed through the stirred tank reactor while stirring at a constant stirring speed of 15 rpm, while maintaining the liquid level at a height equivalent to 175 ml (SV = 0.14 /Hr) The immobilized product of Example 2 was suspended and dispersed, and continuous fermentation was carried out for 17 days. On the other hand, as a control, a jacketed column with an internal volume of 175 ml was filled with the entire amount of the control immobilized material that had been rinsed three times with the enzymatically decomposed liquid, and the space was filled with the enzymatically decomposed liquid. Still 30 on the jacket
25 ml/ml was added to the packed column reactor through constant temperature water at ℃.
While passing the enzymatically decomposed juice for an hour (SV=
0.14/Hr) Continuous fermentation for 17 days was carried out in the same manner. When the immobilized bacterial cell gel was observed at the end of fermentation, no change was observed in the immobilized product of Example 2, but the immobilized product of the control was noticeably softened and approximately
The 10% gel did not retain its original shape and was in a dissolved state. The progress of this fermentation over time is shown in Table 2.
【表】
第２表より明らかな如く、実施例２の固定化物
を連続的に分散、浮遊させつつ発酵させると、対
照の固定化物を用いた場合に比し乳酸生成量が著
しく向上、しかも発酵期間が長期にわたつても乳
酸生成能は高水準に保存されることが認められ
た。[Table] As is clear from Table 2, when the immobilized product of Example 2 was fermented while being continuously dispersed and suspended, the amount of lactic acid produced was significantly improved compared to when the control immobilized product was used. It was observed that lactic acid production ability was maintained at a high level even over a long period of time.
Claims (1)
えたものをゲル基材含有液に混合した液を、多孔
質の粒状支持体の表面に塗付し、これをゲル化剤
と接触させて得られる球状の固定化微生物菌体
を、液体基質に連続的もしくは断続的に分散、浮
遊させつつ接触させて発酵を行なうことを特徴と
する発酵方法。 ２ 液体基質が、醤油製造用原料を酵素的もしく
は化学的に加水分解したものを、PH3.0〜7.0の液
体状態としたものである特許請求の範囲第１項記
載の発酵方法。 ３ ゲル基材がアルギン酸ナトリウム、フアーセ
ルラン、ポリビニルアルコール及びイオータカラ
ギーナンより選ばれた少なくとも１種である特許
請求の範囲第１項記載の発酵方法。[Claims] 1. A solution prepared by mixing microbial cells or a mixture thereof with water or a buffer solution into a gel base material-containing solution is applied to the surface of a porous granular support to form a gel. A fermentation method characterized by carrying out fermentation by contacting spherical immobilized microbial cells obtained by contacting with a liquid substrate while continuously or intermittently dispersing and suspending them in a liquid substrate. 2. The fermentation method according to claim 1, wherein the liquid substrate is obtained by enzymatically or chemically hydrolyzing a raw material for soy sauce production into a liquid state with a pH of 3.0 to 7.0. 3. The fermentation method according to claim 1, wherein the gel base material is at least one selected from sodium alginate, Farcellulan, polyvinyl alcohol, and iota carrageenan.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP58243118A JPS60137293A (en) | 1983-12-24 | 1983-12-24 | Fermentation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP58243118A JPS60137293A (en) | 1983-12-24 | 1983-12-24 | Fermentation |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS60137293A JPS60137293A (en) | 1985-07-20 |
| JPH0371880B2 true JPH0371880B2 (en) | 1991-11-14 |
Family
ID=17099060
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP58243118A Granted JPS60137293A (en) | 1983-12-24 | 1983-12-24 | Fermentation |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS60137293A (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101861831B1 (en) | 2011-11-02 | 2018-05-29 | 엘지전자 주식회사 | A refrigerator comprising a vacuum space |
-
1983
- 1983-12-24 JP JP58243118A patent/JPS60137293A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS60137293A (en) | 1985-07-20 |
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