JPH0223868A - Production of carrier for immobilizing physiologically active substance - Google Patents
Production of carrier for immobilizing physiologically active substanceInfo
- Publication number
- JPH0223868A JPH0223868A JP17182788A JP17182788A JPH0223868A JP H0223868 A JPH0223868 A JP H0223868A JP 17182788 A JP17182788 A JP 17182788A JP 17182788 A JP17182788 A JP 17182788A JP H0223868 A JPH0223868 A JP H0223868A
- Authority
- JP
- Japan
- Prior art keywords
- carrier
- physiologically active
- active substance
- gas atmosphere
- plasma treatment
- 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
- 239000013543 active substance Substances 0.000 title claims description 14
- 230000003100 immobilizing effect Effects 0.000 title claims description 8
- 238000004519 manufacturing process Methods 0.000 title claims description 5
- 239000007789 gas Substances 0.000 claims abstract description 11
- 238000009832 plasma treatment Methods 0.000 claims abstract description 11
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims abstract description 7
- 239000001301 oxygen Substances 0.000 claims abstract description 7
- 229910052760 oxygen Inorganic materials 0.000 claims abstract description 7
- 150000001875 compounds Chemical class 0.000 claims abstract description 4
- 239000011261 inert gas Substances 0.000 claims abstract description 4
- 229920002313 fluoropolymer Polymers 0.000 claims description 11
- 239000004811 fluoropolymer Substances 0.000 claims description 11
- MYMOFIZGZYHOMD-UHFFFAOYSA-N Dioxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 claims description 4
- 229910001882 dioxygen Inorganic materials 0.000 claims description 4
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 abstract description 5
- 229910052731 fluorine Inorganic materials 0.000 abstract description 5
- 239000011737 fluorine Substances 0.000 abstract description 5
- 229920000642 polymer Polymers 0.000 abstract description 4
- 239000003960 organic solvent Substances 0.000 abstract description 2
- 238000000465 moulding Methods 0.000 abstract 2
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 9
- 229940088598 enzyme Drugs 0.000 description 9
- 239000011347 resin Substances 0.000 description 9
- 229920005989 resin Polymers 0.000 description 9
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 8
- 102000004190 Enzymes Human genes 0.000 description 8
- 108090000790 Enzymes Proteins 0.000 description 8
- 230000000694 effects Effects 0.000 description 7
- 238000004458 analytical method Methods 0.000 description 6
- 108010051210 beta-Fructofuranosidase Proteins 0.000 description 6
- 238000006243 chemical reaction Methods 0.000 description 6
- 239000001573 invertase Substances 0.000 description 6
- 235000011073 invertase Nutrition 0.000 description 6
- BFKJFAAPBSQJPD-UHFFFAOYSA-N tetrafluoroethene Chemical group FC(F)=C(F)F BFKJFAAPBSQJPD-UHFFFAOYSA-N 0.000 description 5
- XKRFYHLGVUSROY-UHFFFAOYSA-N Argon Chemical compound [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 description 4
- 239000007864 aqueous solution Substances 0.000 description 4
- 150000001718 carbodiimides Chemical class 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 239000012528 membrane Substances 0.000 description 4
- 229910052757 nitrogen Inorganic materials 0.000 description 4
- QOSSAOTZNIDXMA-UHFFFAOYSA-N Dicylcohexylcarbodiimide Chemical compound C1CCCCC1N=C=NC1CCCCC1 QOSSAOTZNIDXMA-UHFFFAOYSA-N 0.000 description 3
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 3
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 3
- 238000010521 absorption reaction Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 description 2
- 108090000435 Urokinase-type plasminogen activator Proteins 0.000 description 2
- 102000003990 Urokinase-type plasminogen activator Human genes 0.000 description 2
- 229910052786 argon Inorganic materials 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- DDRJAANPRJIHGJ-UHFFFAOYSA-N creatinine Chemical compound CN1CC(=O)NC1=N DDRJAANPRJIHGJ-UHFFFAOYSA-N 0.000 description 2
- 125000000524 functional group Chemical group 0.000 description 2
- 229960002897 heparin Drugs 0.000 description 2
- 229920000669 heparin Polymers 0.000 description 2
- -1 polytrifluoroethylene Polymers 0.000 description 2
- 238000006557 surface reaction Methods 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 229960005356 urokinase Drugs 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- KMHSUNDEGHRBNV-UHFFFAOYSA-N 2,4-dichloropyrimidine-5-carbonitrile Chemical class ClC1=NC=C(C#N)C(Cl)=N1 KMHSUNDEGHRBNV-UHFFFAOYSA-N 0.000 description 1
- 108091006112 ATPases Proteins 0.000 description 1
- 241000186361 Actinobacteria <class> Species 0.000 description 1
- 102000057290 Adenosine Triphosphatases Human genes 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 1
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 1
- 102000000584 Calmodulin Human genes 0.000 description 1
- 108010041952 Calmodulin Proteins 0.000 description 1
- 102000016938 Catalase Human genes 0.000 description 1
- 108010053835 Catalase Proteins 0.000 description 1
- 108010089254 Cholesterol oxidase Proteins 0.000 description 1
- 108010066906 Creatininase Proteins 0.000 description 1
- 101710088194 Dehydrogenase Proteins 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 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 1
- 239000004366 Glucose oxidase Substances 0.000 description 1
- 108010015776 Glucose oxidase Proteins 0.000 description 1
- 102000016901 Glutamate dehydrogenase Human genes 0.000 description 1
- 108700023156 Glutamate dehydrogenases Proteins 0.000 description 1
- 102000005548 Hexokinase Human genes 0.000 description 1
- 108700040460 Hexokinases Proteins 0.000 description 1
- 108010093096 Immobilized Enzymes Proteins 0.000 description 1
- 102000004882 Lipase Human genes 0.000 description 1
- 108090001060 Lipase Proteins 0.000 description 1
- 239000004367 Lipase Substances 0.000 description 1
- 239000002033 PVDF binder Substances 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 102000011420 Phospholipase D Human genes 0.000 description 1
- 108090000553 Phospholipase D Proteins 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- 102000000019 Sterol Esterase Human genes 0.000 description 1
- 108010055297 Sterol Esterase Proteins 0.000 description 1
- 108010092464 Urate Oxidase Proteins 0.000 description 1
- 108010046334 Urease Proteins 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000008065 acid anhydrides Chemical class 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 230000002785 anti-thrombosis Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 239000002473 artificial blood Substances 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 229910002090 carbon oxide Inorganic materials 0.000 description 1
- 238000006482 condensation reaction Methods 0.000 description 1
- 229940109239 creatinine Drugs 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 125000001153 fluoro group Chemical group F* 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 229940116332 glucose oxidase Drugs 0.000 description 1
- 235000019420 glucose oxidase Nutrition 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 235000019421 lipase Nutrition 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000010355 oscillation Effects 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 229920002620 polyvinyl fluoride Polymers 0.000 description 1
- 229920002981 polyvinylidene fluoride Polymers 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000011282 treatment Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Landscapes
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
- Treatments Of Macromolecular Shaped Articles (AREA)
Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
本発明は、生理活性物質固定化用担体の製造法に関する
。更に詳しくは、生理活性物質固定化に用いられる含フ
ッ素重合体成形品担体の製造法に関する。DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a method for producing a carrier for immobilizing a physiologically active substance. More specifically, the present invention relates to a method for producing a fluoropolymer molded carrier used for immobilizing a physiologically active substance.
（従来の技術〕
従来から各種の生理活性物質を担体上に固定化し、例え
ばバイオセンサ用膜、アフィニティクロマトグラフィー
、バイオリアクタ用担体などの用途に用いることが行わ
れている。(Prior Art) Various physiologically active substances have been immobilized on carriers and used for applications such as membranes for biosensors, affinity chromatography, and carriers for bioreactors.
一方、従来から人工血管、カテーテル、人工臓器などの
材料として用いられている、テトラフルオロエチレン樹
脂によって代表される樹脂状またはエラストマー状の含
フッ素重合体に、例えばヘパリン、ウロキナーゼ、ＴＰ
Ａなどを固定化することができれば、生体材料の問題点
となっている抗血栓性の特性を付与することができるな
どの効果が期待できる。On the other hand, resin-like or elastomer-like fluoropolymers typified by tetrafluoroethylene resin, which have been conventionally used as materials for artificial blood vessels, catheters, artificial organs, etc., include heparin, urokinase, TP, etc.
If A and the like can be immobilized, effects such as being able to impart antithrombotic properties, which have been a problem with biomaterials, can be expected.
即ち、耐有機溶剤性、耐薬品性、耐熱性、酸素透過性な
どの性質ですぐれた特性を示す含フッ素重合体に、生化
学的特性を更に付加することができれば、それの応用範
囲が質的にもまた量的にも飛耀的に拡大することが期待
される。In other words, if biochemical properties can be added to fluoropolymers, which have excellent properties such as organic solvent resistance, chemical resistance, heat resistance, and oxygen permeability, the range of applications will increase. It is expected that the industry will expand dramatically both in terms of market and quantity.
そのための一つの手段として、まず含フッ素重合体成形
品の表面に官能性基を導入し、それを足掛りとして各種
の生理活性物質をそこに固定化することが考えられるが
、含フッ素重合体に通常の有機合成反応によって官能性
基を導入することはほぼ不可能である。One way to achieve this is to first introduce functional groups onto the surface of a fluoropolymer molded article, and use this as a foothold to immobilize various physiologically active substances there; It is almost impossible to introduce functional groups into these materials by ordinary organic synthesis reactions.
具体的には、例えばテトラフルオロエチレン樹脂ではＣ
−Ｆ結合力が大きく、またＦ原子がＣ−Ｃ結合の周囲を
くまなく埋めていて、Ｃ−Ｆ結合に対する他の原子団か
らのアタックに対する立体障害となっているため、そこ
にカルボキシル基などの官能性基を導入した上で酵素な
どを固定化させることはほぼ不可能であった。Specifically, for example, in tetrafluoroethylene resin, C
The -F bond strength is large, and the F atoms surround the C-C bond, creating a steric hindrance to attacks from other atomic groups on the C-F bond, so carboxyl groups etc. It was almost impossible to immobilize enzymes and the like after introducing functional groups.
本発明の目的は、含フッ素重合体成形品表面に有機合成
反応によってではなく、他の方法によって官能性基を形
成せしめ、生理活性物質固定化用担体を製造する方法を
提供することにある。An object of the present invention is to provide a method for producing a carrier for immobilizing a physiologically active substance by forming a functional group on the surface of a fluoropolymer molded article not by an organic synthesis reaction but by another method.
かかる目的を達成せしめる生理活性物質固定化用担体の
製造は、含フッ素重合体成形品表面を不活性ガス雰囲気
中および酸素ガスまたは含酸素化合物ガス雰囲気中で順
次プラズマ処理することにより行われる。A carrier for immobilizing a physiologically active substance that achieves this purpose is produced by sequentially plasma-treating the surface of a fluoropolymer molded article in an inert gas atmosphere and an oxygen gas or oxygen-containing compound gas atmosphere.
含フッ素重合体成形品としては、樹脂状またはエラスト
マー状の含フッ素重合体の成形品、一般には膜状、シー
ト状、板状のものなどが用いられる。成形品を形成する
含フッ素重合体として、好ましくはテトラフルオロエチ
レン樹脂（ポリトリフルオロエチレン）が用いられるが
、この他にポリフッ化ビニル、ポリフッ化ビニリデン、
ポリトリフルオロエチレン、テトラフルオロエチレン−
へキサフルオロプロペン共重合体なども用いられる。ま
た、酸素ガスと同様に用いられる含酸素化合物ガスとし
ては、二酸化炭素、−酸化炭素などが用いられる。As the fluoropolymer molded product, a resin-like or elastomer-like fluoropolymer molded product, generally in the form of a film, sheet, or plate, is used. As the fluorine-containing polymer forming the molded product, preferably tetrafluoroethylene resin (polytrifluoroethylene) is used, but in addition to this, polyvinyl fluoride, polyvinylidene fluoride,
Polytrifluoroethylene, tetrafluoroethylene
Hexafluoropropene copolymers and the like can also be used. Further, as the oxygen-containing compound gas used similarly to oxygen gas, carbon dioxide, -carbon oxide, etc. are used.
これらのガス雰囲気中でのプラズマ処理は、例えば真空
ポンプ、リークバルブおよびメインバルブに接続され、
真空計を備えたチューブ状プラズマ反応容器内嘉こ含フ
ッ素重合体成形品を収容し、反応容器内の圧力を約０．
００１〜ＩＴｏｒｒとした後バルブを開き１反応容器内
にガスを約０．００１−１０Ｔｏｒｒの圧力になる迄導
入し５このようにして反応容器内にガスを充満させたら
、高周波発生装置１１（１３，５６ＭＨｚ）およびマツ
チングユニットからなる高周波電源を用いて、有効電力
的１０〜１００１１、グロー放電時間約１〜６０分間の
条件下で、反応容器の端部細径円筒部に捲回された発振
コイルからプラズマ照射することにより行われる０反応
容器としては、上記チューブ状のもの以外に、ペルジャ
ー型なども用いることができる。また、放電電極として
は、上記コイル状のもの以外に、外部もしくは内部平行
電極板を用いることもできる。Plasma treatment in these gas atmospheres can be performed by connecting, for example, a vacuum pump, a leak valve and a main valve,
The fluorine-containing polymer molded article is housed in a tubular plasma reaction vessel equipped with a vacuum gauge, and the pressure inside the reaction vessel is maintained at about 0.
001 to I Torr, open the valve 1 and introduce gas into the reaction vessel until the pressure reaches approximately 0.001 to 10 Torr. 5 After filling the reaction vessel with gas in this way, the high frequency generator 11 ( , 56 MHz) and a matching unit, under conditions of an effective power of 10 to 10,011 and a glow discharge time of approximately 1 to 60 minutes, the oscillation was carried out around the small diameter cylindrical part at the end of the reaction vessel. In addition to the tube-shaped container described above, a Pelger type or the like can also be used as the zero-reaction container, which is carried out by plasma irradiation from a coil. Further, as the discharge electrode, in addition to the coil-shaped one described above, an external or internal parallel electrode plate can also be used.
このような一連のプラズマ処理によって本発明の生理活
性物質固定化用担体は製造されるが、この担体表面には
カルボキシル基が導入されているので、それを利用して
の生理活性物質の固定化が一般にカルボジイミド処理を
経て行われる。The carrier for immobilizing a physiologically active substance of the present invention is produced by such a series of plasma treatments, and since a carboxyl group is introduced into the surface of this carrier, it is possible to immobilize a physiologically active substance by utilizing this carboxyl group. is generally carried out via carbodiimide treatment.
ジシクロへキシルカルボジイミド、Ｎ−シクロヘキシル
−Ｎ′−２−モルホリニル−エチルカルボジイミドなど
のカルボジイミド（ＲＮ ＝ Ｃ＝ ＮＲ）は、容易に
カルボキシル基と付加体を形成し、この付加体は更にア
ルコール、アミン、酸などと縮合反応し、それぞれ相当
するエステル、アミド、酸無水物などを形成する０本発
明に係る固定化用担体の場合には、その表面に導入され
たカルボキシル基が、次式に示されるように、カルボジ
イミドを介して＃素のアミノ基と結合される６
担体−ＣＯＯＨ＋ ＲＮ ＝ Ｃ＝ ＮＲ→［１１＋酵
素−ＮＨ，−＋担体−ＣＯＮＨ−酵素＋ＲＮＨＣＯＮＨ
Ｒ生理活性物質としては、例えばインベルターゼ、ウレ
アーゼ、クレアチニンディイミナーゼ、クレアチニンア
ミドヒドロラーゼ、グルコースオキシダーゼ、パーオキ
シダーゼ、ヘキソキナーゼ、カタラーゼ、Ｇ−６−Ｐデ
ヒドロゲナーゼ、グルタメートデヒドロゲナーゼ、ウロ
キナーゼ、ウリカーゼ、コレステロールオキシダーゼ、
コレステロールエステルヒドロラーゼ、アデノシントリ
フォスファターゼ、アルカリフォスファターゼ、ホスホ
リパーゼＤ、リパーゼ、プロテアーゼ、 ＴＰＡなどの
酵素、各種酵母、糸状菌、放線菌、バクテリアなどの微
生物、抗生物質、抗原抗体、ホルモン、レセプター、ヘ
パリン、カルモジュリン、生体組織、アルブミンなどの
たん白質などが挙げられ、また人工酵素としての鉄−フ
タロシアニン錯体などにも適用される。Carbodiimides (RN = C = NR), such as dicyclohexylcarbodiimide and N-cyclohexyl-N'-2-morpholinyl-ethylcarbodiimide, easily form adducts with carboxyl groups, and these adducts can be further combined with alcohols, amines, In the case of the immobilization carrier according to the present invention, which undergoes a condensation reaction with an acid or the like to form a corresponding ester, amide, acid anhydride, etc., the carboxyl group introduced onto its surface is represented by the following formula: 6 carrier-COOH+ RN = C= NR → [11+enzyme-NH, -+ carrier-CONH-enzyme+RNHCONH, which is bonded to the # elementary amino group via carbodiimide as follows
R physiologically active substances include, for example, invertase, urease, creatinine diiminase, creatinine amidohydrolase, glucose oxidase, peroxidase, hexokinase, catalase, G-6-P dehydrogenase, glutamate dehydrogenase, urokinase, uricase, cholesterol oxidase,
Enzymes such as cholesterol ester hydrolase, adenosine triphosphatase, alkaline phosphatase, phospholipase D, lipase, protease, and TPA, microorganisms such as various yeasts, filamentous fungi, actinomycetes, and bacteria, antibiotics, antigens and antibodies, hormones, receptors, heparin, and calmodulin. , biological tissues, and proteins such as albumin, and is also applicable to iron-phthalocyanine complexes as artificial enzymes.
これらの生理活性物質の固定化は、一般にそれぞれ濃度
が約０．１〜１０ｍｇ／＋ｍ Ｑとなる量の生理活性物
質およびカルボジイミドを添加した水溶液中に約３〜５
℃または室温下にプラズマ処理担体を約１／２〜２４時
間浸漬することによって行われる。The immobilization of these physiologically active substances is generally carried out in an aqueous solution containing approximately 3 to 5 mg of a physiologically active substance and carbodiimide, each with a concentration of approximately 0.1 to 10 mg/+mQ.
The plasma treatment is carried out by immersing the plasma-treated carrier at a temperature of about 1/2 to 24 hours at a temperature of 1/2 to 24 hours.
本発明方法により、各種生理活性物質を固定化可能な含
フッ素重合体成形品担体が得られ、各種のすぐれた性質
を有する含フッ素重合体に更に生化学的特性を付加させ
ることができる。By the method of the present invention, a fluoropolymer molded carrier capable of immobilizing various physiologically active substances can be obtained, and further biochemical properties can be added to the fluoropolymer having various excellent properties.
次に、実施例について本発明を説明する。 Next, the present invention will be explained with reference to examples.
実施例１
市販テトラフルオロエチレン樹脂膜（厚さ０．２ｍｍ）
を容量結合型プラズマ発生装置内に置き、アルゴンガス
圧力０．ＩＴｏｒｒ、電力５Ｈ１時間３０分間の条件下
で、プラズマ処理を行った６次いで、酸素ガス圧力０．
ＩＴｏｒｒ、電力４５１１、時間５分間の条件下でプラ
ズマ処理を行った。いずれも、１３．５６ＭＨｚの高周
波が用いられている。Example 1 Commercially available tetrafluoroethylene resin membrane (thickness 0.2 mm)
is placed in a capacitively coupled plasma generator, and the argon gas pressure is 0. Plasma treatment was performed under the conditions of ITorr and power of 5H for 1 hour and 30 minutes.Then, the oxygen gas pressure was 0.
Plasma treatment was performed under the conditions of ITorr, power of 4511, and time of 5 minutes. In both cases, a high frequency of 13.56 MHz is used.
このようにしてプラズマ処理された樹脂膜の表面は、２
５℃、２５％ＲＨの条件下での水の接触角が、未処理物
の１１０’から８５°へと大きく変化しており、疎水性
から親水性になっていることが分る。The surface of the resin film treated with plasma in this way is 2
It can be seen that the contact angle of water under the conditions of 5° C. and 25% RH changed significantly from 110° of the untreated material to 85°, indicating that the material became hydrophilic from hydrophobic.
また、 ＦＴＩＲによる分析結果では、１４００ｃｍ−
’および１７００ｃｍ−”付近に吸収がみられ、表面に
カルボキシル基の導入されていることが確認された。Also, according to the analysis results by FTIR, 1400cm-
Absorption was observed near ' and 1700 cm-'', confirming that carboxyl groups were introduced on the surface.
次いで、この試料について、酵素の固定化を行なった。Next, enzyme immobilization was performed on this sample.
即ち、この試料を、１ｍｇ／＋ｓ Ｑ量のインベルター
ゼおよび７ｍｇ／ｍ Ｑ量のジシクロへキシルカルボジ
イミドをそれぞれ添加したｐＨ３，０の水溶液中に４℃
で２４時間浸漬した。That is, this sample was placed at 4°C in an aqueous solution at pH 3.0 to which were added 1 mg/+s Q amount of invertase and 7 mg/m Q amount of dicyclohexylcarbodiimide.
Soaked for 24 hours.
その後、 ｐＨ７，０のリン酸緩衝液で洗浄し、固定化
インベルターゼ量をケルダール窒素分析法で測定したと
ころ、樹脂板表面１−当り２２ｍｇのインベルターゼが
結合されていることが分った。Thereafter, the plate was washed with a phosphate buffer solution of pH 7.0, and the amount of immobilized invertase was measured by Kjeldahl nitrogen analysis, and it was found that 22 mg of invertase was bound per 1 inch of the resin plate surface.
また、この固定化酵素の活性を、ネルソンーソモギイ法
により測定したところ、同量の非結合酵素の活性に対す
る相対的な活性が９８％であるという結果が得られた。Furthermore, when the activity of this immobilized enzyme was measured by the Nelson-Somogyi method, it was found that the relative activity to that of the same amount of unbound enzyme was 98%.
実施例２
市販の酸素通過性テトラフルオロエチレン樹脂膜（厚さ
０．１＋＋ａ＋）について、実施例１と同一条件でのプ
ラズマ処理を行った後、７ｍｇ／ｍ Ｑのジシクロへキ
シルカルボジイミドおよび乾燥重量で１＋＊ｇ／ｍＱ量
の酵母（サツ力ロマイセツセレヴイジェ、対数増殖期集
菌）をそれぞれ添加したＰＨ７，０の水溶液中に室温下
で２４時間浸漬した。Example 2 A commercially available oxygen-permeable tetrafluoroethylene resin membrane (thickness 0.1++a+) was subjected to plasma treatment under the same conditions as in Example 1, and then treated with dicyclohexylcarbodiimide at 7 mg/m Q and dry weight. The samples were immersed for 24 hours at room temperature in an aqueous solution having a pH of 7.0 to which 1+*g/mQ of yeast (Satu Romaisetsu Cerevige, logarithmically grown bacteria) had been added.
この酵母固定化膜を十分水洗した後、クラーク型酸素電
極に装着し、１１０ｌｌ１／ＩＩＱ濃度のグルコース水
溶液に対する応答をみたところ、溶存酸素の減少がみら
れ、固定化酵母としての活性を有していることが示され
た。After thoroughly washing this yeast-immobilized membrane with water, it was attached to a Clark-type oxygen electrode and its response to a glucose aqueous solution with a concentration of 110ll1/IIQ was observed, and a decrease in dissolved oxygen was observed, indicating that it had activity as an immobilized yeast. It was shown that there is.
比較例１
実施例１において、酵素ガスプラズマ処理を行わないと
、樹脂膜表面には、ＦＴＩＲ分析によるカルボキシル基
の吸収がみられず、またインベルターゼとの固定化処理
物にも、ケルダール窒素分析法での窒素成分はみられず
、酵素活性もなかった。Comparative Example 1 In Example 1, if the enzyme gas plasma treatment was not performed, no absorption of carboxyl groups was observed on the resin film surface by FTIR analysis, and the immobilized product with invertase was also detected by Kjeldahl nitrogen analysis method. No nitrogen components were observed, and there was no enzyme activity.
比較例２
実施例１において、アルゴンガスプラズマ処理を行わな
い場合には、樹脂膜表面にＦＴＩＲ分析によるカルボキ
シル基の吸収が認められ、またケルダール窒素分析値か
らは表面１−当り５ｍｇのインベルターゼ結合量が示さ
れ、更に酵素の相対活性は６５２であることが示された
が、いずれも実施例１よりも低い値しか得られていない
。Comparative Example 2 In Example 1, when argon gas plasma treatment was not performed, absorption of carboxyl groups was observed on the resin film surface by FTIR analysis, and Kjeldahl nitrogen analysis showed that the amount of invertase bound was 5 mg per surface. was shown, and the relative activity of the enzyme was also shown to be 652, but both values were lower than in Example 1.
Claims (1)
よび酸素ガスまたは含酸素化合物ガス雰囲気中で順次プ
ラズマ処理することを特徴とする生理活性物質固定化用
担体の製造法。1. A method for producing a carrier for immobilizing a physiologically active substance, which comprises sequentially subjecting the surface of a fluoropolymer molded article to plasma treatment in an inert gas atmosphere and an oxygen gas or oxygen-containing compound gas atmosphere.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP17182788A JP2658211B2 (en) | 1988-07-12 | 1988-07-12 | Method for producing a carrier for immobilizing a physiologically active substance |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP17182788A JP2658211B2 (en) | 1988-07-12 | 1988-07-12 | Method for producing a carrier for immobilizing a physiologically active substance |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH0223868A true JPH0223868A (en) | 1990-01-26 |
| JP2658211B2 JP2658211B2 (en) | 1997-09-30 |
Family
ID=15930478
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP17182788A Expired - Lifetime JP2658211B2 (en) | 1988-07-12 | 1988-07-12 | Method for producing a carrier for immobilizing a physiologically active substance |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP2658211B2 (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02100677A (en) * | 1988-10-07 | 1990-04-12 | Nok Corp | Production of carrier for immobilizing organism substance |
| KR100269277B1 (en) * | 1992-09-25 | 2000-10-16 | 윤종용 | Manufacturing Method of Semiconductor Memory Device |
| JP2007510029A (en) * | 2003-10-30 | 2007-04-19 | ライプニッツ−インスティチュート フュア ポリマーフォルシュング ドレスデン エーファウ | Modified perfluoroplastic and process for producing the same |
| GB2585038A (en) * | 2019-06-25 | 2020-12-30 | Inst Jozef Stefan | Method |
-
1988
- 1988-07-12 JP JP17182788A patent/JP2658211B2/en not_active Expired - Lifetime
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02100677A (en) * | 1988-10-07 | 1990-04-12 | Nok Corp | Production of carrier for immobilizing organism substance |
| KR100269277B1 (en) * | 1992-09-25 | 2000-10-16 | 윤종용 | Manufacturing Method of Semiconductor Memory Device |
| JP2007510029A (en) * | 2003-10-30 | 2007-04-19 | ライプニッツ−インスティチュート フュア ポリマーフォルシュング ドレスデン エーファウ | Modified perfluoroplastic and process for producing the same |
| GB2585038A (en) * | 2019-06-25 | 2020-12-30 | Inst Jozef Stefan | Method |
Also Published As
| Publication number | Publication date |
|---|---|
| JP2658211B2 (en) | 1997-09-30 |
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