JPH01247088A - Method for supporting biocatalyst with polymer membrane - Google Patents
Method for supporting biocatalyst with polymer membraneInfo
- Publication number
- JPH01247088A JPH01247088A JP63075160A JP7516088A JPH01247088A JP H01247088 A JPH01247088 A JP H01247088A JP 63075160 A JP63075160 A JP 63075160A JP 7516088 A JP7516088 A JP 7516088A JP H01247088 A JPH01247088 A JP H01247088A
- Authority
- JP
- Japan
- Prior art keywords
- bag
- biocatalyst
- membrane
- immobilized
- sealed
- 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.)
- Granted
Links
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 14
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 13
- 238000000034 method Methods 0.000 title claims abstract description 13
- 239000011942 biocatalyst Substances 0.000 title claims abstract description 8
- 229920005597 polymer membrane Polymers 0.000 title claims abstract description 7
- 239000012528 membrane Substances 0.000 claims abstract description 6
- 239000011148 porous material Substances 0.000 claims abstract description 4
- 238000004519 manufacturing process Methods 0.000 abstract description 9
- 244000005700 microbiome Species 0.000 abstract description 7
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 abstract description 6
- 235000015097 nutrients Nutrition 0.000 abstract description 5
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 abstract description 4
- 239000007788 liquid Substances 0.000 abstract description 4
- 239000001301 oxygen Substances 0.000 abstract description 4
- 229910052760 oxygen Inorganic materials 0.000 abstract description 4
- 239000007864 aqueous solution Substances 0.000 abstract description 3
- 230000000813 microbial effect Effects 0.000 abstract description 3
- 238000002360 preparation method Methods 0.000 abstract description 3
- 230000001954 sterilising effect Effects 0.000 abstract description 3
- 239000000126 substance Substances 0.000 abstract description 3
- 238000005273 aeration Methods 0.000 abstract description 2
- 230000008569 process Effects 0.000 abstract description 2
- 239000004744 fabric Substances 0.000 abstract 2
- 244000052616 bacterial pathogen Species 0.000 abstract 1
- 239000003054 catalyst Substances 0.000 abstract 1
- 239000003622 immobilized catalyst Substances 0.000 abstract 1
- 230000035755 proliferation Effects 0.000 abstract 1
- 230000001580 bacterial effect Effects 0.000 description 18
- 239000000243 solution Substances 0.000 description 14
- 239000004745 nonwoven fabric Substances 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 5
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 3
- 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
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 239000013028 medium composition Substances 0.000 description 3
- KPGXRSRHYNQIFN-UHFFFAOYSA-N 2-oxoglutaric acid Chemical compound OC(=O)CCC(=O)C(O)=O KPGXRSRHYNQIFN-UHFFFAOYSA-N 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-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
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 2
- 229910052782 aluminium Inorganic materials 0.000 description 2
- 230000004663 cell proliferation Effects 0.000 description 2
- POULHZVOKOAJMA-UHFFFAOYSA-N dodecanoic acid Chemical compound CCCCCCCCCCCC(O)=O POULHZVOKOAJMA-UHFFFAOYSA-N 0.000 description 2
- 230000005284 excitation Effects 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000011888 foil Substances 0.000 description 2
- 230000002062 proliferating effect Effects 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- ZTOKUMPYMPKCFX-CZNUEWPDSA-N (E)-17-[(2R,3R,4S,5S,6R)-6-(acetyloxymethyl)-3-[(2S,3R,4S,5S,6R)-6-(acetyloxymethyl)-3,4,5-trihydroxyoxan-2-yl]oxy-4,5-dihydroxyoxan-2-yl]oxyoctadec-9-enoic acid Chemical compound OC(=O)CCCCCCC/C=C/CCCCCCC(C)O[C@@H]1O[C@H](COC(C)=O)[C@@H](O)[C@H](O)[C@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](COC(C)=O)O1 ZTOKUMPYMPKCFX-CZNUEWPDSA-N 0.000 description 1
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 description 1
- NGHVIOIJCVXTGV-ALEPSDHESA-N 6-aminopenicillanic acid Chemical compound [O-]C(=O)[C@H]1C(C)(C)S[C@@H]2[C@H]([NH3+])C(=O)N21 NGHVIOIJCVXTGV-ALEPSDHESA-N 0.000 description 1
- NGHVIOIJCVXTGV-UHFFFAOYSA-N 6beta-amino-penicillanic acid Natural products OC(=O)C1C(C)(C)SC2C(N)C(=O)N21 NGHVIOIJCVXTGV-UHFFFAOYSA-N 0.000 description 1
- 239000005715 Fructose Substances 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 108010093096 Immobilized Enzymes Proteins 0.000 description 1
- 150000008575 L-amino acids Chemical class 0.000 description 1
- 239000005639 Lauric acid Substances 0.000 description 1
- 241000228143 Penicillium Species 0.000 description 1
- 239000004743 Polypropylene Substances 0.000 description 1
- 229920005830 Polyurethane Foam Polymers 0.000 description 1
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 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
- 238000010564 aerobic fermentation Methods 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 239000000783 alginic acid Substances 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 229960001126 alginic acid Drugs 0.000 description 1
- 150000004781 alginic acids Chemical class 0.000 description 1
- HWXBTNAVRSUOJR-UHFFFAOYSA-N alpha-hydroxyglutaric acid Natural products OC(=O)C(O)CCC(O)=O HWXBTNAVRSUOJR-UHFFFAOYSA-N 0.000 description 1
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 description 1
- 229940009533 alpha-ketoglutaric acid Drugs 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- 230000001851 biosynthetic effect Effects 0.000 description 1
- 239000000648 calcium alginate Substances 0.000 description 1
- 235000010410 calcium alginate Nutrition 0.000 description 1
- 229960002681 calcium alginate Drugs 0.000 description 1
- OKHHGHGGPDJQHR-YMOPUZKJSA-L calcium;(2s,3s,4s,5s,6r)-6-[(2r,3s,4r,5s,6r)-2-carboxy-6-[(2r,3s,4r,5s,6r)-2-carboxylato-4,5,6-trihydroxyoxan-3-yl]oxy-4,5-dihydroxyoxan-3-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylate Chemical compound [Ca+2].O[C@@H]1[C@H](O)[C@H](O)O[C@@H](C([O-])=O)[C@H]1O[C@H]1[C@@H](O)[C@@H](O)[C@H](O[C@H]2[C@H]([C@@H](O)[C@H](O)[C@H](O2)C([O-])=O)O)[C@H](C(O)=O)O1 OKHHGHGGPDJQHR-YMOPUZKJSA-L 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 239000005515 coenzyme Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 239000001630 malic acid Substances 0.000 description 1
- 235000011090 malic acid Nutrition 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- -1 polypropylene Polymers 0.000 description 1
- 229920001155 polypropylene Polymers 0.000 description 1
- 239000011496 polyurethane foam Substances 0.000 description 1
- 238000011403 purification operation Methods 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 239000011949 solid catalyst Substances 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- VOCWXFXWGYGHQH-UHFFFAOYSA-N tetradecane-1,3,4-tricarboxylic acid Chemical compound C(CC(C(CCCCCCCCCC)C(=O)O)C(=O)O)C(=O)O VOCWXFXWGYGHQH-UHFFFAOYSA-N 0.000 description 1
- 230000008719 thickening Effects 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Landscapes
- Catalysts (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
酵素活性を安定化し、また水溶性であるにもかかわらず
くり返して使用を可能にするために、酵素を物理的また
は化学的手法で水に不溶化して固体触媒の形で使用する
技術が開発され、Ｌ−アミノ酸、異性化糖、６−アミノ
ペニシラン酸及びリンゴ酸等の工業生産に利用されてい
る６さらに固定化酵素の手法を進めて酵素を包含したま
まの微生物菌体（固定化微生物）を利用すれば、抽出精
製操作を省ける、固定化または操作時の安定性がよい、
多階段酵素系の利用に有利、補酵素やＡＴＰなどの供給
が可能、などの利点がある。また、二次代謝産物の生産
のために生菌体の固定化法を用いることができ、さらに
固定化増殖微生物を用いて収量を向上させることも可能
である。[Detailed Description of the Invention] [Industrial Application Field] In order to stabilize enzyme activity and enable repeated use despite being water-soluble, enzymes are immersed in water by physical or chemical methods. A technology to insolubilize and use in the form of a solid catalyst has been developed, and is used in the industrial production of L-amino acids, high-fructose sugar, 6-aminopenicillanic acid, malic acid, etc.6 Furthermore, the method of immobilized enzymes has been advanced. By using microbial cells that still contain enzymes (immobilized microorganisms), extraction and purification operations can be omitted, and stability during immobilization or manipulation is good.
It has advantages such as being advantageous in using a multi-step enzyme system and being able to supply coenzymes, ATP, etc. Furthermore, a method of immobilizing living microorganisms can be used to produce secondary metabolites, and it is also possible to improve the yield by using immobilized growing microorganisms.
上記のために固定化技術の確立は基本的に重要であるが
、担体結合法、橋かけ法及び包括法など道、ＰＰＭ、Ｎ
ｏ、６．２８ （１９８７）、福井三部編著、「生体触
媒としての微生物」共立出版（１９７９）。For the above reasons, the establishment of immobilization technology is fundamentally important, but methods such as carrier binding method, crosslinking method, and comprehensive method, PPM, N
o, 6.28 (1987), Fukui Sanbe (ed.), “Microorganisms as biocatalysts”, Kyoritsu Shuppan (1979).
好気性菌の発酵生産技術において栄養源及び酸素の拡散
速度が目的とする生産物の生産速度に影響を及ぼす。一
般に栄養源の利用効率は低く、また菌体の増殖や生産物
の生成により培養液の粘性が増大して生合成反応が阻害
され目的とする生産物の生産速度が低下する。糸状菌の
場合特に三次元網目構造を有する画境の生成や培養液の
増粘による栄養分や酸素の不足が起こりやすい。また、
培養液から生産物を分離するのに手間がかかるなど改良
の余地が大きい。In aerobic fermentation production technology, the rate of diffusion of nutrients and oxygen influences the rate of production of the desired product. In general, the utilization efficiency of nutrient sources is low, and the viscosity of the culture solution increases due to bacterial cell proliferation and product production, inhibiting biosynthetic reactions and reducing the production rate of the desired product. In the case of filamentous fungi, shortages of nutrients and oxygen are particularly likely to occur due to the formation of borders with a three-dimensional network structure and thickening of the culture solution. Also,
There is a lot of room for improvement, as it takes time and effort to separate the products from the culture solution.
つぎに、ゲル内に固定化した菌体・増殖菌体のバイオリ
アクターへの応用技術が開発され、アルさらにそのため
の培養液との混濁による生産物の回収の困難さが生じる
。Next, a technology was developed to apply bacterial cells and proliferating bacterial cells immobilized in gel to bioreactors, but it became difficult to recover the product due to turbidity with the alkaline medium and the culture solution used for this purpose.
本発明者らは発酵生産及び固定化生体触媒（酵素及び微
生物）における物質生産において安価、耐久性、プロセ
スや操作の簡略化、生産性の向上及び安定性の高い固定
化法について鋭意研究を重ねた結果、高分子膜を用いて
生体触媒を担持することにより、表面積が大きく菌体の
過密な成育やそれに伴う画境の生成がないので栄養物や
酸素の供給が充足し、耐久性があってくり返し長期の使
用が可能であり、また増殖菌体だけでなく、休止菌体の
使用も可能であるため生産性が高く、さらに膜内に菌体
があるため培養液の滅菌が不要で且つ煩雑なゲル調製操
作を必要としないことを見出し、この知見に基づき本発
明を完成するに至った。The present inventors have conducted extensive research into immobilization methods that are inexpensive, durable, simplify processes and operations, improve productivity, and are highly stable in fermentation production and substance production using immobilized biocatalysts (enzymes and microorganisms). As a result, by using a polymer membrane to support a biocatalyst, the surface area is large, and there is no dense growth of bacterial cells or the formation of boundaries, so nutrients and oxygen are supplied sufficiently, and it is durable. It can be used repeatedly for a long period of time, and it is highly productive as it is possible to use not only proliferating cells but also resting cells.Furthermore, since the cells are inside the membrane, sterilization of the culture solution is unnecessary and complicated. It was discovered that no extensive gel preparation operation was required, and based on this finding, the present invention was completed.
ジュラガード３２０１ （孔径０．４Ｘ０．０４μｍ）
製の縦５０．横４０ｍｍの三方が閉鎖された袋の中に縦
４５、横３５ｍｍの不織布（１％Ｔ ｗｅａｎ８０水溶
液に浸漬乾燥したもの）１枚を入れた後、Ｎ／１０００
塩酸水溶液に１時間以上浸漬した。Duraguard 3201 (pore diameter 0.4x0.04μm)
Made of length 50. After putting a piece of non-woven fabric (dipped in 1% T wean 80 aqueous solution and dried) with a length of 45 mm and a width of 35 mm into a bag with a width of 40 mm and closed on three sides, N/1000
It was immersed in an aqueous hydrochloric acid solution for over 1 hour.
この後、Ｐｅｎｉｃｌｌｉｕｍ ｓｐｉｃｕｌｉｓｐｏ
ｒｕｍ ＬｅｈＬＩｌａｎＮｏｌＯ−１の菌体約１００
ｍｇを袋の中に入れ、シーラーを用いて加熱し袋の入口
を封じた。こうして作った袋１０枚を、下記の組成の滅
菌しない培地３００ｍ１の入ったＩＱ容広ロポリビンに
入れ、アルミホイルで口を覆い、通気しながら往復励振
とう培養器（振幅７０ｍｍ、１２０回／分往復）にセッ
トして３０℃にて６日間培養した。発酵の進行とともに
培養液は著しく発泡した。培養後、袋を切り開いてみる
と不織布の両面にわたって厚さ約０．５＋＋ｍの菌体が
フィルム状に付着し重量は１゜）また、膜外への培養菌
体の漏出は認められなかった。培養液中のブドウ糖量は
グルコースメーターによると２．４％であった。粘度は
調製時の培養液の粘度１０ｃｐから変化がなかった。さ
らに発酵液にエタノールを加えＩＮ水酸化ナトリウム１
００ｍＱを加えて中和後ＩＮ塩酸１００ｍＱを加えて中
和しクロロホルムにてスピクリスポール酸（４，５−ジ
カルボキシ−４−ペンタデカノリド）の開環体（１，３
，４−テトラデカントリカルボン酸）が培養液ＩＱ当り
１．３ｇ生成した。After this, Penicllium spiculispo
Approximately 100 bacterial cells of rum LehLIlanNolO-1
mg was placed in a bag, and the entrance of the bag was sealed by heating using a sealer. The 10 bags thus made were placed in an IQ wide-capacity Ropoly bottle containing 300 ml of unsterilized medium with the following composition, the mouth was covered with aluminum foil, and the reciprocating excitation incubator (amplitude 70 mm, 120 reciprocating cycles/min) was used while ventilating. ) and cultured at 30°C for 6 days. As fermentation progressed, the culture solution foamed significantly. After culturing, when the bag was cut open, bacterial cells with a thickness of about 0.5++ m were adhered to both sides of the nonwoven fabric in a film shape (weighing 1°), and no leakage of cultured bacterial cells to the outside of the membrane was observed. The amount of glucose in the culture solution was 2.4% according to a glucose meter. The viscosity did not change from the viscosity of the culture solution at the time of preparation, which was 10 cp. Furthermore, add ethanol to the fermentation liquid and IN sodium hydroxide 1
After neutralizing by adding 00 mQ of IN hydrochloric acid, the ring-opened form (1,3
, 4-tetradecanetricarboxylic acid) was produced in an amount of 1.3 g per IQ culture solution.
培地組成（水ＩＱ当り）
ブドウ糖 （無水） １００ｇＮＨ，
Ｃ１１，Ｏｇ
ＫＨ，ＰＯ４１，０ｇ
ＭｇＳＯ４・７Ｈ，００，５ｇ
次に表１により高分子膜担持による実験経過を示した。Medium composition (per water IQ) Glucose (anhydrous) 100gNH,
C11, Og KH, PO4 1,0 g MgSO4.7H, 00,5 g Next, Table 1 shows the progress of the experiment with polymer membrane support.
実施例２
実施例１と同じジュラガード３５０１ （０，４Ｘ０゜
０４ μｍ）に不織布を挟んだ袋の中にＰｅｎｉｃｌｌ
ｉｕｍ ｓｐｉｃｕｒｉｓｐｏｒｕｍ Ｌｅｈｍａｎ
Ｎｏ１Ｏ−１を１００ｍｇを入れて封じたものを３袋用
意した。培養液３００ｗＭを入れたＩＱ容広口瓶にこれ
らの３袋を入れ、アルミホイルで口を覆い、３０℃にて
実施例１と同様にして培養した。実施結果は表１の実験
Ｎｏ、２スポール酸の生産効率が低下した。Example 2 Penicll was placed in a bag with a nonwoven fabric sandwiched between the same Duragard 3501 (0.4×0°04 μm) as in Example 1.
ium spiculisporum Lehman
Three sealed bags containing 100 mg of No1O-1 were prepared. These three bags were placed in an IQ wide mouth bottle containing 300 wM of culture solution, the mouth was covered with aluminum foil, and cultured at 30° C. in the same manner as in Example 1. The implementation results were Experiment No. 2 in Table 1, in which the production efficiency of 2-sporic acid decreased.
実施例３
実施例１と同じジュラガード３５０１ （０，４Ｘ０゜
０４μｆｆ１）の５０Ｘ４０ｍｍの袋を作り、不織布を
入れずに菌体１００ｍｇを袋の中に封じた。６日後開封
すると培養開始時とほぼ同量の菌体量であった。Example 3 A 50 x 40 mm bag made of the same Duragard 3501 (0.4 x 0° 04 μff1) as in Example 1 was made, and 100 mg of bacterial cells was sealed in the bag without inserting a nonwoven fabric. When opened after 6 days, the amount of bacterial cells was almost the same as at the start of culture.
それ故、菌体を担持し増殖させるために不織布を袋の中
に入れることが必要であることがわかった。Therefore, it has been found that it is necessary to insert a nonwoven fabric into the bag in order to support and propagate the bacterial cells.
実施例４
実施例１と３の比較から分かるようにジュラガード製の
袋の中に不織布のような菌体の支持体を置くことが好ま
しい。不織布の代わりにアルギン酸カルシウムゲルを用
いてＰｅｎｉｃｉｌｌｉｕｍ ｓｐｉｃｕｌｉｓｐｏｒ
ｕｍの菌体を担持することも可能であった。すなわち、
０．８％アルギン酸水溶液またはＰｅｎ１ｃｉ１１ｉｕ
、ｍ ｓｐｉｃｕｌｉｓｐｏｒｕｍの菌体を懸濁した０
、８％ルはそのままジュラガードの袋に入れ、また空の
ビーズは菌体とともにジュラガードの袋に入れて封をし
た後、実施例１と同様にして培養を行った。Example 4 As can be seen from the comparison of Examples 1 and 3, it is preferable to place a bacterial support such as a nonwoven fabric in a Duraguard bag. Penicillium spiculispor using calcium alginate gel instead of non-woven fabric
It was also possible to carry um bacterial cells. That is,
0.8% alginic acid aqueous solution or Pen1ci11iu
, m spiculisporum cells suspended in 0
, 8% Le were placed directly in a Duraguard bag, and the empty beads were placed in a Duraguard bag along with the bacterial cells and sealed, and then cultured in the same manner as in Example 1.
６日後菌体の増殖が見られ、ゲルが菌体の支持に有用で
あることが示された。さらに、支持体としてポリウレタ
ンフォームを用いてもよく、菌体を接種して６日後菌体
量は３倍に増殖した。After 6 days, bacterial cell proliferation was observed, indicating that the gel was useful for supporting bacterial cells. Furthermore, polyurethane foam may be used as a support, and 6 days after inoculating the cells, the amount of cells increased three times.
実施例５
実施例１により得られた菌体を成育させ不織布上に固定
化した高分子膜バッグ１０袋を取り出し、ガラスロート
上に置いて培養液を自然に滴下させて除いた後水洗いを
くり返した。これらのバッグ内に固定化された菌糸体の
酵素系を利用して、基質を含む下表の培地３００ｍＱを
入れたＩＱ容広ロポリ瓶を用いてスピクリスポール酸の
生合成（バイオコンバージョン）を行った。３０℃にて
６日間往復励振とう培養器（振幅７０ｍｍ、１２０（１
： １）液に溶解した。ベンゼンを展開液として３相ク
ロマトグラフィーを行い、スピクリスポール酸の生成を
確認した。Example 5 Ten polymer membrane bags in which the bacterial cells obtained in Example 1 were grown and immobilized on a non-woven fabric were taken out and placed on a glass funnel to allow the culture solution to drip naturally and removed, followed by repeated washing with water. Ta. Utilizing the enzyme system of the mycelia immobilized in these bags, biosynthesis (bioconversion) of spicrysporic acid was carried out using an IQ wide-capacity bottle filled with 300 mQ of the medium shown in the table below containing the substrate. Ta. Reciprocating excitation incubator (amplitude 70 mm, 120 (1
: 1) Dissolved in liquid. Three-phase chromatography was performed using benzene as a developing solution, and the production of spicrisporic acid was confirmed.
培地組成（水ＩＱ当り） ＮＨ，Ｃ１１，０ｇ Ｋ Ｈ２Ｐ Ｏ−１−Ｏｇ ＭｇＳＯ４・７Ｈ２００，５ｇ コーンスチープリ力− １．０ｇ ラウリン酸 １２．８ｇ α−ケトグルタル酸 ７．３ｇ ｐＨ４，２、滅菌せずそのまま使用。Medium composition (per water IQ) NH,C11,0g K H2P O-1-Og MgSO4・7H200.5g Corn steeple force - 1.0g Lauric acid 12.8g α-ketoglutaric acid 7.3g pH 4.2, used as is without sterilization.
ることにより、ソホロリピッド：７−Ｌ−（（２’−０
−β−Ｄ−グルコピラノシルーβ−Ｄ−グルコピラノシ
ル）オキソ〕オクタデセン酸−１，４”−ラクトン類を
主成分とし、その関連化合物からなる同族体を生産する
ことが知られている（Ｎ、Ｋｏｓａｒｉｃ ｅｔ、 ａ
ｌ、、Ｊ、ａｍ、ＯｉｌＣｈｅｍ、Ｓｏｃ、、６１．１
７３５（１９８４））。実施例１と同じ手法により菌体
を高分子膜内に担持させた袋を５袋。By doing so, the sophorolipid: 7-L-((2'-0
-β-D-glucopyranosyl-β-D-glucopyranosyl)oxo]octadecenoic acid-1,4”-lactones are the main components, and it is known to produce homologs consisting of related compounds (N , Kosaric et, a.
l,,J,am,OilChem,Soc,,61.1
735 (1984)). Five bags were prepared in which bacterial cells were supported in a polymer membrane using the same method as in Example 1.
無滅菌の培養液３００ｍＱの入ったＩＱ広ロポリ瓶中に
入れて３０℃で６日間通気しながら振とうして培養した
。菌体量は接種時（１００ｍｇ）の８倍に増殖していた
。The cells were placed in an IQ wide polypropylene bottle containing 300 mQ of non-sterile culture solution and cultured at 30° C. for 6 days with aeration and shaking. The amount of bacterial cells had grown eight times that at the time of inoculation (100 mg).
培地組成（水ＩＱ当り）
ブドウＭ １００ ｇイーストエキス
Ｌｏｇ
尿素 １ｇMedium composition (per water IQ) Grape M 100 g Yeast extract Log Urea 1 g
Claims (1)
生体触媒の担持方法。(1) A method for supporting a biocatalyst having a polymer membrane with a pore size of 2 μm or less as a wall membrane.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63075160A JPH082302B2 (en) | 1988-03-29 | 1988-03-29 | Cell culture member and cell culture method using the same |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63075160A JPH082302B2 (en) | 1988-03-29 | 1988-03-29 | Cell culture member and cell culture method using the same |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH01247088A true JPH01247088A (en) | 1989-10-02 |
| JPH082302B2 JPH082302B2 (en) | 1996-01-17 |
Family
ID=13568174
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63075160A Expired - Lifetime JPH082302B2 (en) | 1988-03-29 | 1988-03-29 | Cell culture member and cell culture method using the same |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH082302B2 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114377696A (en) * | 2021-11-12 | 2022-04-22 | 天俱时工程科技集团有限公司 | Biomembrane-based BiOClxBr(1-x)/Au/MnO2Composite material and preparation method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59213389A (en) * | 1983-05-17 | 1984-12-03 | Nissin Electric Co Ltd | Preparation of microbial membrane |
| JPS6027699U (en) * | 1983-07-25 | 1985-02-25 | 日東電工株式会社 | Immobilized enzyme container |
| JPS62171672A (en) * | 1985-10-24 | 1987-07-28 | Oosakashi | Bioreactor |
-
1988
- 1988-03-29 JP JP63075160A patent/JPH082302B2/en not_active Expired - Lifetime
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59213389A (en) * | 1983-05-17 | 1984-12-03 | Nissin Electric Co Ltd | Preparation of microbial membrane |
| JPS6027699U (en) * | 1983-07-25 | 1985-02-25 | 日東電工株式会社 | Immobilized enzyme container |
| JPS62171672A (en) * | 1985-10-24 | 1987-07-28 | Oosakashi | Bioreactor |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114377696A (en) * | 2021-11-12 | 2022-04-22 | 天俱时工程科技集团有限公司 | Biomembrane-based BiOClxBr(1-x)/Au/MnO2Composite material and preparation method and application thereof |
| CN114377696B (en) * | 2021-11-12 | 2023-08-11 | 天俱时工程科技集团有限公司 | Biofilm-based BiOCl x Br (1-x) /Au/MnO 2 Composite material, preparation method and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH082302B2 (en) | 1996-01-17 |
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