JPH0549335B2 - - Google Patents
Info
- Publication number
- JPH0549335B2 JPH0549335B2 JP63075150A JP7515088A JPH0549335B2 JP H0549335 B2 JPH0549335 B2 JP H0549335B2 JP 63075150 A JP63075150 A JP 63075150A JP 7515088 A JP7515088 A JP 7515088A JP H0549335 B2 JPH0549335 B2 JP H0549335B2
- Authority
- JP
- Japan
- Prior art keywords
- capsule
- solution
- chitosan
- permeability
- membrane
- 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 - Fee Related
Links
- 239000002775 capsule Substances 0.000 claims description 52
- 229920002101 Chitin Polymers 0.000 claims description 19
- 239000012528 membrane Substances 0.000 claims description 18
- 230000035699 permeability Effects 0.000 claims description 16
- 150000003839 salts Chemical class 0.000 claims description 11
- 150000004676 glycans Chemical class 0.000 claims description 10
- 229920001282 polysaccharide Polymers 0.000 claims description 10
- 239000005017 polysaccharide Substances 0.000 claims description 10
- 238000004519 manufacturing process Methods 0.000 claims description 9
- 239000012530 fluid Substances 0.000 claims description 8
- 239000002253 acid Substances 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 5
- 150000002500 ions Chemical class 0.000 claims description 4
- 230000007935 neutral effect Effects 0.000 claims description 2
- 239000000243 solution Substances 0.000 description 33
- 229920001661 Chitosan Polymers 0.000 description 20
- 238000000034 method Methods 0.000 description 13
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 10
- 235000018102 proteins Nutrition 0.000 description 9
- 108090000623 proteins and genes Proteins 0.000 description 9
- 102000004169 proteins and genes Human genes 0.000 description 9
- 239000011248 coating agent Substances 0.000 description 8
- 238000000576 coating method Methods 0.000 description 8
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 7
- 239000001768 carboxy methyl cellulose Substances 0.000 description 7
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 7
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 7
- 229940105329 carboxymethylcellulose Drugs 0.000 description 7
- 238000006243 chemical reaction Methods 0.000 description 6
- 238000005538 encapsulation Methods 0.000 description 6
- 239000007864 aqueous solution Substances 0.000 description 5
- 238000004132 cross linking Methods 0.000 description 5
- 239000011780 sodium chloride Substances 0.000 description 5
- 108010025899 gelatin film Proteins 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 239000003094 microcapsule Substances 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 210000004027 cell Anatomy 0.000 description 3
- 238000000502 dialysis Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 229920000447 polyanionic polymer Polymers 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 210000004102 animal cell Anatomy 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 230000000850 deacetylating effect Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 210000004408 hybridoma Anatomy 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 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 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N 2,3,4,5-tetrahydroxypentanal Chemical compound OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- SQDAZGGFXASXDW-UHFFFAOYSA-N 5-bromo-2-(trifluoromethoxy)pyridine Chemical compound FC(F)(F)OC1=CC=C(Br)C=N1 SQDAZGGFXASXDW-UHFFFAOYSA-N 0.000 description 1
- 235000001674 Agaricus brunnescens Nutrition 0.000 description 1
- 229920001287 Chondroitin sulfate Polymers 0.000 description 1
- 241000238557 Decapoda Species 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000239366 Euphausiacea Species 0.000 description 1
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 229920000869 Homopolysaccharide Polymers 0.000 description 1
- 238000012695 Interfacial polymerization Methods 0.000 description 1
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 1
- OVRNDRQMDRJTHS-UHFFFAOYSA-N N-acelyl-D-glucosamine Natural products CC(=O)NC1C(O)OC(CO)C(O)C1O OVRNDRQMDRJTHS-UHFFFAOYSA-N 0.000 description 1
- OVRNDRQMDRJTHS-FMDGEEDCSA-N N-acetyl-beta-D-glucosamine Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O OVRNDRQMDRJTHS-FMDGEEDCSA-N 0.000 description 1
- 102000007562 Serum Albumin Human genes 0.000 description 1
- 108010071390 Serum Albumin Proteins 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 235000010418 carrageenan Nutrition 0.000 description 1
- 239000000679 carrageenan Substances 0.000 description 1
- 229920001525 carrageenan Polymers 0.000 description 1
- 229940113118 carrageenan Drugs 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 229940059329 chondroitin sulfate Drugs 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 229950006780 n-acetylglucosamine Drugs 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 229920002851 polycationic polymer Polymers 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 239000012679 serum free medium Substances 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 235000010413 sodium alginate Nutrition 0.000 description 1
- 239000000661 sodium alginate Substances 0.000 description 1
- 229940005550 sodium alginate Drugs 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 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 description 1
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
- B01J13/02—Making microcapsules or microballoons
- B01J13/20—After-treatment of capsule walls, e.g. hardening
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Dispersion Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Manufacturing Of Micro-Capsules (AREA)
Description
【発明の詳細な説明】
産業上の利用分野
本発明はキトサンのような可溶性キチン誘導体
を被膜形成材として用いた被膜透過性制御可能な
カプセル体の製造方法に関する。DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to a method for producing a capsule whose film permeability can be controlled using a soluble chitin derivative such as chitosan as a film forming material.
従来の技術及び解決すべき課題
バイオテクノロジーの分野において、カプセル
を利用して微生物や酵素を固定したり、動物細胞
をカプセルを用いて培養する方法（マイクロカプ
セル法）等が特に注目されている。Background Art and Problems to be Solved In the field of biotechnology, methods of immobilizing microorganisms and enzymes using capsules and culturing animal cells using capsules (microcapsule method) are attracting particular attention.
マイクロカプセル法により培養を行うと、細胞
を培養中の機械的剪断力から保護することができ
かつ、カプセル膜の透過性を制御することにより
細胞の産生する生理活性物質をカプセル内に高濃
度で蓄積させることができるので、その後の分離
回収が有利であるとともに、細胞と培養液との分
離も容易に行い得る等の優れた利点がある。 When cultured using the microcapsule method, cells can be protected from mechanical shearing forces during culture, and by controlling the permeability of the capsule membrane, physiologically active substances produced by the cells can be kept in high concentrations within the capsule. Since it can be accumulated, subsequent separation and recovery is advantageous, and it has excellent advantages such as easy separation of cells and culture medium.
しかしながら、マウクロカプセル法において
は、カプセル膜の透過性を適切に制御することが
できないなど問題となつていた。このため、例え
ば特開昭55−44387号（半透過性マイクロカプセ
ル−の製造法）の技術では、マイクロカプセルの
透過性を、その膜形成の間、、界面重合反応にお
けるパラメータを調節することにより行うとの提
案がなされている。しかしながら、この技術にお
いては方法が煩雑でありまた、種々の溶剤を使用
しなければならないなど実用的なものではなかつ
た。 However, the macrocapsule method has had problems such as the inability to appropriately control the permeability of the capsule membrane. For this reason, for example, in the technique of JP-A No. 55-44387 (method for manufacturing semi-permeable microcapsules), the permeability of microcapsules is adjusted by adjusting parameters in the interfacial polymerization reaction during membrane formation. There are proposals to do so. However, this technique is not practical as it is complicated and requires the use of various solvents.
発明の目的
従つて、本発明は、簡易な方法でカプセル膜の
透過性を自由に、かつ正確に制御できるカプセル
体の製造方法を提供することを目的とする。OBJECTS OF THE INVENTION Therefore, an object of the present invention is to provide a method for manufacturing a capsule body that allows the permeability of the capsule membrane to be controlled freely and accurately using a simple method.
本発明者は、上記目的を達成すべく鋭意検討し
た結果、カプセル化に使用する溶液のイオン強度
を予め調節してからカプセルを調製すれば、調節
したイオン強度に対応した膜透過性をもつたカプ
セル膜をつくることができることを見出し、本発
明をなすに至つたものである。 As a result of intensive studies to achieve the above object, the inventors of the present invention have found that if the ionic strength of the solution used for encapsulation is adjusted in advance before preparing capsules, membrane permeability corresponding to the adjusted ionic strength can be achieved. It was discovered that a capsule membrane could be produced, and the present invention was completed.
発明の構成
即ち、本発明は、
ポリアニオン多糖類またはその塩の単独もしく
はそれらの混合物を基材とする流動体と、可溶性
キチン誘導体の溶液とを接触してその流動体を芯
部として内包するカプセル体を製造する方法にお
いて、その可溶性キチン誘導体の溶液を透析し
て、実質的に全ての遊離酸イオンを除去し、次い
で該可溶性キチン誘導体の溶液に0.1〜0.7モル濃
度の中性塩を添加した後、流動体と可溶性キチン
誘導体の溶液とを接触するカプセル体の製造方法
に関する。Structure of the Invention That is, the present invention provides a capsule in which a fluid based on a polyanionic polysaccharide or a salt thereof or a mixture thereof is brought into contact with a solution of a soluble chitin derivative, and the fluid is encapsulated as a core. In the method of manufacturing the soluble chitin derivative solution, the solution of the soluble chitin derivative was dialyzed to remove substantially all free acid ions, and then a 0.1 to 0.7 molar concentration of a neutral salt was added to the solution of the soluble chitin derivative. The present invention then relates to a method for producing a capsule body, which involves contacting a fluid with a solution of a soluble chitin derivative.
以下、本発明について詳述する。 The present invention will be explained in detail below.
本発明においてカプセル体のゲル被膜を形成す
るのに用いるポリアニオン多糖類またはその塩は
水溶液中でポリアニオン重合体となる多糖類また
はその塩であつて、低メトキシルペクチン、カラ
ギナン、カルボキシメチルセルロース、アルギン
酸ナトリウム、コンドロイチン硫酸等を例示し得
る。これらの多糖類は単独または混合物としてい
得る。分子量としては、カプセル形成性の点で
104〜106のものが好ましい。 In the present invention, the polyanionic polysaccharide or its salt used to form the gel coating of the capsule body is a polysaccharide or its salt that forms a polyanionic polymer in an aqueous solution, and includes low methoxyl pectin, carrageenan, carboxymethyl cellulose, sodium alginate, Examples include chondroitin sulfate. These polysaccharides may be used alone or in mixtures. In terms of molecular weight, capsule-forming properties
10 4 to 10 6 is preferred.
一方、同じゲル被膜の形成に用いる可溶性キチ
ン誘導体は、本来不活性な物質であるキチンに化
学的処理を施してその反応活性を高めたものであ
つて、キチンを脱アセチル化処理して得られるキ
トサンが代表的なものとして例示し得る。 On the other hand, the soluble chitin derivative used to form the same gel film is obtained by chemically treating chitin, which is an inert substance to increase its reaction activity, and by deacetylating chitin. Chitosan can be exemplified as a typical example.
因に、キチンはカニ、オキアミ、昆虫等の甲皮
微生物の細胞壁、きのこ類等に含まれるＮ−アセ
チル−Ｄ−グルコサミンがβ（１→４）結合した
直鎖ホモ多糖体であつて、天然に豊富に生産され
るものであるが、その不活性の故にそのままで利
用できない未利用天然資源といえる。 Incidentally, chitin is a linear homopolysaccharide containing β (1→4) bonds of N-acetyl-D-glucosamine, which is found in the cell walls of carapaceous microorganisms such as crabs, krill, and insects, and in mushrooms. Although it is abundantly produced in the United States, it can be said to be an unused natural resource that cannot be used as is because of its inertness.
しかし、キチンを脱アセチル化処理して得られ
るキトサンのようなキチン誘導体は稀酸に可溶と
なり、反応活性を有するようになる。すなわち、
キトサンは下記一般式（）で表わされる構造単
位を有し、
式中のアミノ基により正に帯電し、ポリカチオ
ン重合体として反応活性を示す。 However, chitin derivatives such as chitosan obtained by deacetylating chitin become soluble in dilute acids and have reactive activity. That is,
Chitosan has a structural unit represented by the following general formula (), It is positively charged due to the amino group in the formula and exhibits reactive activity as a polycationic polymer.
キチン誘導体の分子量としては、カプセル形成
性の点で、105〜106のものが好ましい。 The molecular weight of the chitin derivative is preferably 10 5 to 10 6 from the viewpoint of capsule-forming properties.
したがつて、上述したような可溶性キチン誘導
体としてのキトサンの溶液に、上記ポリアニオン
多糖類またはその塩もしくはそれらの混合物の水
溶液を接触させると、ポリアニオン多糖類とキト
サンとの間に荷電による架橋反応、すなわち架橋
結合を起してゲル状物質を生成する。 Therefore, when an aqueous solution of the polyanionic polysaccharide, its salt, or a mixture thereof is brought into contact with a solution of chitosan as a soluble chitin derivative as described above, a crosslinking reaction due to charge occurs between the polyanionic polysaccharide and chitosan. That is, crosslinking occurs to produce a gel-like substance.
本発明において上記両溶液の接触を行なうに
は、上記ポリアニオン多糖類またはその塩を含む
水溶液をデポジツターなどによりキトサン溶液中
に攪拌下に滴下させるとよく、その際上記架橋反
応が起る。この架橋反応により一旦ゲル被膜が形
成されると、該被膜に内包されて芯部を構成する
溶液のゲル化は全くみられなくなるので所望のカ
プセル体が得られるようになる。このような現象
は、ゲル被膜が形成されると、該被膜に内包され
た溶液（すなわち芯部）中のポリアニオン重合体
およびキトサン分子がもはや上記被膜を透過でき
なくなつて、反応が芯液中で進行しなくなること
に因るものと考えられる。 In order to bring the above-mentioned solutions into contact in the present invention, the aqueous solution containing the above-mentioned polyanionic polysaccharide or its salt is preferably dropped into the chitosan solution under stirring using a depositor, and the above-mentioned crosslinking reaction occurs at this time. Once a gel film is formed by this crosslinking reaction, the solution contained in the film and constituting the core will not gel at all, making it possible to obtain the desired capsule. This phenomenon occurs because once a gel coating is formed, the polyanionic polymer and chitosan molecules in the solution (i.e., the core) contained in the coating can no longer pass through the coating, and the reaction occurs in the core solution. This is thought to be due to the fact that the disease does not progress.
本発明では、カプセル体の芯部を構成する流動
体として用いる溶液の調製に当つてはイオン強度
を調節したポリアニオン多糖類またはその塩もし
くはそれらの混合物を0.3〜1.0重量％含む水溶液
とすることが適当である。0.3重量％より少ない
と、カプセル体を形成することが困難となり、一
方、1.0重量％より多くなると、液滴形成が困難
となる。特にポリアニオン多糖類としてカルボキ
シメチルセルロースを用いるのがゲル被膜形成上
好ましい。 In the present invention, when preparing the solution used as the fluid constituting the core of the capsule, it is preferable to use an aqueous solution containing 0.3 to 1.0% by weight of a polyanionic polysaccharide, a salt thereof, or a mixture thereof with adjusted ionic strength. Appropriate. If it is less than 0.3% by weight, it will be difficult to form a capsule body, while if it is more than 1.0% by weight, it will be difficult to form droplets. In particular, it is preferable to use carboxymethyl cellulose as the polyanionic polysaccharide in terms of forming a gel film.
また、上記水溶液を接触させるキトサン溶液
は、酢酸あるいはグルタミン酸のような弱酸に
0.5〜1.0重量％の濃度に溶解したものが適当であ
る。0.5重量％より少なくなると、被膜はできに
くくなり、一方、1.0重量％より多くなると芯液
がキトサン溶液中に入りにくくなる。 In addition, the chitosan solution that the above aqueous solution is brought into contact with is a weak acid such as acetic acid or glutamic acid.
A solution dissolved at a concentration of 0.5 to 1.0% by weight is suitable. If it is less than 0.5% by weight, it will be difficult to form a film, while if it is more than 1.0% by weight, it will be difficult for the core liquid to enter the chitosan solution.
本発明において、可溶性キチン誘導体溶液のイ
オン強度を調節する方法としては、例えばその溶
液を透析用チユーブにつめ純水中で透析を行うも
のがある。この透析処理により、大部分の遊離酸
イオンを除くことができる。このようにして、遊
離酸イオンを除去した可溶性キチン誘導体溶液に
塩化ナトリウムなどの各種塩を所定量添加するこ
とにより、種々の程度のイオン強度をもつた溶液
を容易に調製することができる。そして、イオン
強度の低い溶液で調製したカプセルは、密な膜構
造をもち、カプセル膜の透過性は低い。一方、イ
オン強度を上げていくと、カプセル膜は透過性が
上がつていく。 In the present invention, as a method for adjusting the ionic strength of the soluble chitin derivative solution, for example, the solution is placed in a dialysis tube and dialyzed in pure water. This dialysis treatment can remove most of the free acid ions. By adding a predetermined amount of various salts such as sodium chloride to the soluble chitin derivative solution from which free acid ions have been removed in this manner, solutions having various degrees of ionic strength can be easily prepared. Capsules prepared from solutions with low ionic strength have a dense membrane structure, and the permeability of the capsule membrane is low. On the other hand, as the ionic strength increases, the permeability of the capsule membrane increases.
本発明では、カプセル体のゲル被膜を有機溶剤
などを使用することなく、極めて温和な条件、す
なわち、生物学的に温和な条件下で短時間に形成
し得るので、不安定な生物学的物質や機能性物質
およびカプセル体の使用目的に応じその他の種々
の添加物を、カプセル体の芯部を構成する前記流
動体に添加して分散させることができる。従つ
て、種々の有用物質を芯液に含有させたカプセル
体を提供することが可能となる。 In the present invention, the gel coating of the capsule body can be formed in a short period of time under extremely mild conditions, that is, biologically mild conditions, without using organic solvents etc. Various other additives can be added and dispersed in the fluid constituting the core of the capsule, depending on the purpose of use of the capsule. Therefore, it is possible to provide a capsule body containing various useful substances in the core liquid.
又、本発明は、ゲル被膜の形成によるカプセル
化を１工程で行ない得るので、例えば特開昭57−
197031号にみられるポリアニオンとポリカチオン
間の塩架橋を利用した公知のカプセル化法に比し
製造上有利であるといえる。 In addition, the present invention enables encapsulation by forming a gel film in one step.
It can be said that this method is advantageous in terms of production compared to the known encapsulation method that utilizes salt crosslinking between a polyanion and a polycation as seen in No. 197031.
更に、本発明ではカプセル体のゲル被膜の形成
条件をコントロールすることにより、該被膜の膜
透過性を変化させることが可能であるので、カプ
セル体の被膜の分画機能を付与することができ
る。 Furthermore, in the present invention, by controlling the formation conditions of the gel coating of the capsule body, it is possible to change the membrane permeability of the gel coating, so that the coating of the capsule body can be provided with a fractionation function.
以上のとおり、本発明によると、入手の容易な
原材料を用いて簡易な製造手段で、しかも短時間
で、広範囲な用途に供し得るカプセル体を提供し
得る利点がある。 As described above, the present invention has the advantage of being able to provide capsule bodies that can be used in a wide range of applications using easily available raw materials, by simple manufacturing means, and in a short period of time.
以下、実施例により本発明を更に具体的に説明
する。 Hereinafter, the present invention will be explained in more detail with reference to Examples.
実施例 １
カプセル膜の透過性を評価するため、分子量既
知のたんぱく質をカプセル化し、経時的にカプセ
ル内、およびカプセル外のたんぱく質濃度を測定
し、次式より計算を行つた。Example 1 In order to evaluate the permeability of the capsule membrane, a protein with a known molecular weight was encapsulated, and the protein concentration inside and outside the capsule was measured over time and calculated using the following formula.
Ｐ（％）＝CiVi／CiVi＋CoVo×100
ここで、Ci：カプセル内たんぱく質濃度
Vi：カプセル全体積
Co：カプセル外たんぱく質濃度
Vo：カプセル外液体積
カプセルは分子量が2.6×105のCMC（カルボキ
シメチル・セルロース）と、分子量が2.8×106の
キトサンで調製した。両溶液を２日間純水中で透
析した後、CMCにたんぱく質として血清−アル
ブミン（分子量約７万）を加え、カプセル化用芯
液とした。最終CMC濃度は0.5％であつた。キト
サンにはイオン強度を調節するため塩化ナトリウ
ムを0.1〜0.7M（モル濃度）添加し、種々のイオ
ン強度のキトサン溶液を調製した。 P (%) = CiVi / CiVi + CoVo × 100 where, Ci: Protein concentration inside the capsule Vi: Total volume of the capsule Co: Protein concentration outside the capsule Vo: Liquid volume outside the capsule The capsule has a molecular weight of 2.6 × 10 5 CMC (carboxymethyl Cellulose) and chitosan with a molecular weight of 2.8 x 106 . After both solutions were dialyzed in pure water for 2 days, serum albumin (molecular weight approximately 70,000) was added as a protein to CMC to prepare a core liquid for encapsulation. The final CMC concentration was 0.5%. Sodium chloride was added to chitosan at a concentration of 0.1 to 0.7 M (molar concentration) to adjust the ionic strength, and chitosan solutions with various ionic strengths were prepared.
以上のように調製した溶液でカプセル化を行つ
た結果を第１図に示す。同図から明らかなよう
に、イオン強度を上げると、カプセル膜の透過性
は上がり、たんぱく質濃度（Ｐ）の値が下がる。
つまり、カプセル膜はたんぱく質を透過しやすく
なることを意味している。 Figure 1 shows the results of encapsulation using the solution prepared as described above. As is clear from the figure, increasing the ionic strength increases the permeability of the capsule membrane and decreases the protein concentration (P).
This means that the capsule membrane becomes more permeable to proteins.
従つて、カプセル膜の透過性はイオン強度を変
えることで制御できることを示している。 Therefore, it is shown that the permeability of the capsule membrane can be controlled by changing the ionic strength.
実施例 ２
分子量1.6×106のキトサンを用い、実施例１と
同様な方法で実験を行つた結果を第２図に示す。Example 2 The results of an experiment conducted in the same manner as in Example 1 using chitosan with a molecular weight of 1.6×10 6 are shown in FIG.
実施例１と同様な結果を得た。 Similar results as in Example 1 were obtained.
実施例 ３ 本発明を細胞培養に応用した例を示す。Example 3 An example in which the present invention is applied to cell culture will be shown.
芯液として免疫たんぱく質（IgG）を産生する
動物細胞（ハイブリドーマ）を0.5％CMC溶液中
に分散した溶液を調製した。 A solution in which animal cells (hybridoma) producing immune protein (IgG) were dispersed in a 0.5% CMC solution was prepared as a core fluid.
キトサン溶液として透析した溶液及び透析
後１％塩化ナトリウムを添加した溶液をそれぞれ
調製し、上記芯液を用いてカプセル化を行つた。 A dialyzed solution as a chitosan solution and a solution to which 1% sodium chloride was added after dialysis were prepared, and encapsulation was performed using the above core solution.
カプセルは無血清倍地で11日間培養し、培養過
程でカプセル内及びカプセル外のIgG濃度を測定
し、カプセル内の保持されているIgG量を次式よ
り求めた。 The capsules were cultured in a serum-free medium for 11 days, and during the culture process, the IgG concentrations inside and outside the capsules were measured, and the amount of IgG retained within the capsules was determined using the following formula.
IgG保持率（％）＝（カプセル内IgG）×100／
（カプセル内IgG）＋（カプセル外IgG）
結果を第３図に示す。 IgG retention rate (%) = (IgG in capsule) x 100/
(Intracapsular IgG) + (Extracapsular IgG) The results are shown in Figure 3.
図中、黒丸で示したのが、の透析キトサン溶
液を用いた結果であり、白丸で示したのがの１
％塩化ナトリウムを添加したキトサンの溶液を用
いたときの結果である。 In the figure, the black circles indicate the results using the dialyzed chitosan solution, and the white circles indicate the results using the dialyzed chitosan solution.
% sodium chloride was used.
透析したキトサンを用いて調製したカプセル
ではIgGがほとんどカプセル外に漏出せず、カプ
セル内に蓄積されているが、一方塩化ナトリウム
を添加したキトサンで調製したカプセルでは、
IgGがカプセル内からカプセル外へ漏出してい
る。 In capsules prepared using dialyzed chitosan, IgG hardly leaked out of the capsule and was accumulated inside the capsule, whereas in capsules prepared using chitosan added with sodium chloride,
IgG leaks from inside the capsule to outside the capsule.
以上のように、キトサン溶液のイオン強度を調
節することにより、カプセル内にIgGを高濃度に
蓄積したり、あるいはカプセル外へ透過させるこ
とができる。つまりカプセル膜の透過性を制御す
ることができる。 As described above, by adjusting the ionic strength of the chitosan solution, it is possible to accumulate IgG at a high concentration within the capsule or to allow it to permeate outside the capsule. In other words, the permeability of the capsule membrane can be controlled.
第１図及び第２図は、イオン強度の変化に対す
るカプセル膜の透過性の変化との関係を示す図で
あり、そして、第３図はハイビリドーマ培養にお
ける免疫たんぱく質の膜透過性の結果を示す図で
ある。
Figures 1 and 2 are diagrams showing the relationship between changes in capsule membrane permeability with changes in ionic strength, and Figure 3 is a diagram showing the results of membrane permeability of immune proteins in hybridoma culture. It is.
Claims (1)
らの混合物を含む流動体と、可溶性キチン誘導体
の溶液とを接触して前記流動体を芯部とし内包し
て成るカプセル体を製造する方法において、 前記可溶性キチン誘導体の溶液を透析して、実
質的に全ての遊離酸イオンを除去し、次いで該可
溶性キチン誘導体の溶液に0.1〜0.7モル濃度の中
性塩を添加した後、前記接触を行うことを特徴と
する膜透過性制御可能なカプセル体の製造法。[Claims] 1. A method for producing a capsule body comprising a fluid containing a polyanionic polysaccharide, a salt thereof, or a mixture thereof and a solution of a soluble chitin derivative and containing the fluid as a core. , dialyzing the solution of the soluble chitin derivative to remove substantially all free acid ions, and then adding a 0.1 to 0.7 molar neutral salt to the solution of the soluble chitin derivative, followed by the contacting. A method for producing a capsule body whose membrane permeability can be controlled, characterized in that:
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63075150A JPH01245848A (en) | 1988-03-29 | 1988-03-29 | Production of capsule body with film of controllable permeability |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63075150A JPH01245848A (en) | 1988-03-29 | 1988-03-29 | Production of capsule body with film of controllable permeability |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH01245848A JPH01245848A (en) | 1989-10-02 |
| JPH0549335B2 true JPH0549335B2 (en) | 1993-07-26 |
Family
ID=13567884
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63075150A Granted JPH01245848A (en) | 1988-03-29 | 1988-03-29 | Production of capsule body with film of controllable permeability |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH01245848A (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS60175539A (en) * | 1984-02-23 | 1985-09-09 | Snow Brand Milk Prod Co Ltd | Capsule and its production |
| JPS633786A (en) * | 1986-06-23 | 1988-01-08 | Asahi Chem Ind Co Ltd | Collagen-containing capsule |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS6418440A (en) * | 1987-07-10 | 1989-01-23 | Dainippon Pharmaceutical Co | Micro-capsule |
-
1988
- 1988-03-29 JP JP63075150A patent/JPH01245848A/en active Granted
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS60175539A (en) * | 1984-02-23 | 1985-09-09 | Snow Brand Milk Prod Co Ltd | Capsule and its production |
| JPS633786A (en) * | 1986-06-23 | 1988-01-08 | Asahi Chem Ind Co Ltd | Collagen-containing capsule |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH01245848A (en) | 1989-10-02 |
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