CN1673257A - Prepn process of compound conductive polymer film used as biosensor fixing carrier - Google Patents
Prepn process of compound conductive polymer film used as biosensor fixing carrier Download PDFInfo
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- CN1673257A CN1673257A CN 200410026617 CN200410026617A CN1673257A CN 1673257 A CN1673257 A CN 1673257A CN 200410026617 CN200410026617 CN 200410026617 CN 200410026617 A CN200410026617 A CN 200410026617A CN 1673257 A CN1673257 A CN 1673257A
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Abstract
The compound conductive polymer film used as biosensor fixing carrier is prepared through mixing protein solution and pyrrole in certain ratio, electric polymerization inside proper buffer solution in a three electrode system, oxidation at some potential for certain time and reduction at some other potential for certain time to obtain compound conductive polymer film. Or, it may be prepared through mixing pyrrole with buffering solution, electric polymerization in a three electrode system, oxidation at some potential for certain time and adsorption in protein solution at some other potential for certain time to obtain laminated compound conductive polymer film. The compound conductive polymer film of the present invention is used as bioactive matter fixing carrier in preparing biosensor with fast response, wide linear response range, long service life and excellent repeatability and high selectivity.
Description
Technical field
Proteinic graft modification, the preparation of composite conductive high molecular film, biosensor
Background technology
Polymer is regarded as insulating material always, after having reported " behind the polyacetylene chemical doping; electricity is led rapid increase; can reach the electroconductibility of bismuth metal " in the synthetic and academic meeting of character of the low-dimensional material of holding in New York up in June, 1977, caused the very big interest of academia and industry member, the research of conducting polymer performance has become popular domain, and has obtained bigger progress.Royal Swedish Academy of Sciences has announced that Nobel chemistry Prize in 2000 authorizes the Bai Chuanying tree of Japanese University of tsukuba, the Alan of Univ Pennsylvania USA. the Alan of mark's Diarmid and California, USA university. black square you, to commend the initiative contribution that they make at this emerging field of conductive polymers.The conductive polymers conduct fixedly carrier of active substance is applied to the concern that the biosensor aspect has also obtained the scientific worker.The conductive polymers biosensor mainly is as carrier or coating material immobilizing biologically active composition (as: enzyme, antigen, antibody, microorganism etc.) with conductive polymers, and with this as sensor, again with the device of appropriate signals conversion and proofing unit be combined into.Be used as the fixedly conducting polymer composite of carrier for active principle of biosensor, it must possess following characteristics: (1) has the ability of good conduction electron; (2) can effectively prevent the loss of activeconstituents; (3) can well keep the activity of activeconstituents; (4) can prolong work-ing life of biosensor; (5) has excellent biological compatibility with activeconstituents; (6) can reduce the cost of biosensor and processing easily.
Often as biosensor fixedly the conductive polymers of carrier for active principle polypyrrole (PPY), polyaniline (PAN), poly-neighbour (or) phenylenediamine (PDAN) is arranged.PPY can accelerate the electron transport between enzyme and the material having higher electroconductibility in the PH scope widely, also owing to the positively charged of PPY own, electronegative enzyme can be fixed in the film when preparation PPY film.But make enzyme be easy to from film, run off or inactivation because PPY has comparatively loose structure, and PPY itself because of more easily by the lower compound of peroxidation generation electroconductibility, so it has lower stability, the life-span of the electrode of making is shorter,
Protein is abundant occurring in nature reserves, and its functional property has: (1) hydrated properties comprises water absorption and reservation, wettability, swelling, tackiness, dispersiveness, solubleness and viscosity.By polar group on the protein peptide chain backbone and water molecules generation hydration; (2) character relevant with protein-protein interaction comprises various other structures of precipitating action, gelation and formation (as protein dough and fiber).Protein molecule is heated and unfolds, and inner hydrophobic grouping comes out, and (passes through Ca by hydrophobic interaction (high temperature can provide this type of effect), electrostatic interaction
2+With other divalent ion bridge joints), hydrogen bond (cooling can improve this type of effect) or two sulfur-crosslinked formation space reticulated structures.Protein has good biocompatibility with the active substance (as: enzyme, antigen, antibody, microorganism etc.) that is used as biosensor, and active substance is played a very good protection, and keeps their biological activity.But the protein of pure natural is apt to deteriorate, and in water easy swelling, the biologically active substance of its embedding is run off easily.So can carry out modification or chemically modified to protein, connecting hydrophobic group on its main chain or the side chain, overcome its swollen shortcoming apt to deteriorate and easy, increase the consistency of itself and biologically active substance, prevent the loss of active substance.Proteinic method of modifying is divided into physically modified, chemical modification and enzyme modification.
Therefore, if the polypyrrole (or pyrrole derivative) of conduction is made the compound conducting polymer film with protein (comprising the protein through chemical modification and chemically modified), make it have good electron transmittability and biocompatibility, will aspect biosensor, have bigger application prospect.
Summary of the invention
Purpose of the present invention provides a kind of preparation method of composite conductive high molecular film over against the problems referred to above, and this composite conductive high molecular film is applied in the biosensor.Adopt the method for electropolymerization to prepare the composite conductive high molecular film, promptly adopt pyrroles's (or pyrrole derivative), protein (through the protein of chemical modification or chemically modified) for raw material, by the incompatible acquisition conducting polymer film of voltolisation.
Method of the present invention has utilized the good electron transmittability of polypyrrole (or pyrrole derivative) and protein and biologically active substance to have the advantage of excellent biological compatibility.Can form the uniform conductive film to polypyrrole (or pyrrole derivative) and protein by electropolymerization, chemical crosslink reaction has taken place with protein in polypyrrole (or pyrrole derivative), the polypyrrole that makes (or pyrrole derivative) is penetrated into proteinic intramolecule, formed the favorable conductive network, thereby the composite high-molecular film that makes has not only had excellent conducting performance, and also has good biocompatibility, reticulated structure between the two intersects mutually, make whole polymeric membrane fine and close more, thereby can effectively keep biologically active substance activity, also reduced their losses when contacting with solution.And, can also carry out electropolymerization to pyrroles's (or pyrrole derivative) earlier, by electro-adsorption the surface that protein is embedded in conducting film is formed sandwich layering conducting film again, also have the premium properties of even composite conductive film.The composite conductive high molecular film that makes is done the carrier of the fixedly active substance of biosensor, all effectively raise the time of response of electrode, widened the linear response range of electrode, prolonged the life-span of electrode.
The invention provides a kind of preparation method who is used as the composite conductive high molecular film of biosensor immobilization carrier, wherein the preparation method of composite conductive high molecular film is: with protein (through the protein of chemical modification or chemically modified) solution, pyrroles's (or pyrrole derivative), in the buffered soln of pH=5-8, (platinum electrode, carbon dioxide process carbon electrode or gold electrode are working electrode to adopt three-electrode system, Ag/AgCl or saturated calomel electrode are reference electrode, platinum wire electrode is a counter electrode), carry out electric polymerization reaction.Wherein proteinic part by weight is 5 to 85%, and the part by weight that is used for carrying out protein modified organic molecule or biomacromolecule is proteinic 15 to 90%, and pyrroles's's (or pyrrole derivative) part by weight is 10 to 95%.The preparation conducting polymer film adopts the electropolymerization method, and its oxidizing potential is 0.2 to 1.4V, and oxidization time is 5-90 minute, and reduction potential is-1.4 to 0V, and the recovery time is 0-60 minute.
The biosensor that adopts composite conductive high molecular film that method of the present invention obtains to prepare as the carrier of immobilizing biologically active materials, its time of response is fast, and linear response range is wide, long service life, and have good circulation ratio and highly selective.
Embodiment
Implement will be further understood that the present invention from following illustrative.
Adopt protein (through the protein of chemical modification or chemically modified) solution, pyrroles's (or pyrrole derivative), method by electropolymerization prepares the composite conductive high molecular film, and prepares biosensor with the composite conductive high molecular film that obtains as the carrier of immobilizing biologically active materials.
Embodiment 1
Proportioning raw materials is: getting concentration is the casein solution 5mL with the methyl methacrylate modification of 0.2g/mL, joins 50mL and contains 0.1mol/L pyrroles, 1mol/LKNO
3, the phosphate buffer solution of pH=6.86 adopts three-electrode system (platinum electrode is a working electrode, and Ag/AgCl is a reference electrode, and platinum wire electrode is a counter electrode), and CONTROLLED POTENTIAL is 0.9V, oxidation 30min.The combined electrode that obtains is moved in the pH=6.86 phosphate buffer soln after with the second distillation water wash, and reductase 12 0min under the current potential of-0.3V obtains the combined electrode that the composite conductive high molecular film is a carrier.Control voltage is 0.7V, and this electrode is placed the glucose oxidase solution electro-adsorption 10min of 100u/mL, just obtains glucose oxidase electrode.Come the content of glucose in the test soln with it, its time of response only is 8s, and linear response range is 1.0 * 10
-6～1.43 * 10
-2Mol/L, its limit of detection is 1.0 * 10
-7Mol/L, surpass one month work-ing life, and have good circulation ratio and highly selective.
Embodiment 2
Proportioning raw materials is: contain 0.1mol/L pyrroles at 100mL, 1mol/LKNO
3The phosphate buffer solution of pH=6.86, (platinum electrode is a working electrode, and Ag/AgCl is a reference electrode to adopt three-electrode system, platinum wire electrode is a counter electrode), CONTROLLED POTENTIAL is 0.7V, and oxidation 40min is with moving to behind the second distillation water wash in the pH=6.86 phosphate buffer soln, under the current potential of-0.5V, reduce 10min, putting into concentration again is the casein solution with the methyl methacrylate modification of 0.3g/mL, and CONTROLLED POTENTIAL is 0.9V, electro-adsorption 20min.Just obtain with the composite conductive high molecular film is the combined electrode of carrier.Control voltage is 0.7V, and this electrode is placed the glucose oxidase solution electro-adsorption 10min of 100u/mL, just obtains glucose oxidase electrode.Come the content of glucose in the test soln with it, its time of response only is 12s, and linear response range is 1.5 * 10
-6～2.43 * 10
-2Mol/L, surpass one month work-ing life, and have good circulation ratio and highly selective.
Embodiment 3
Getting concentration is the collagen solution 20mL of 0.1g/mL, joins 50mL and contains 0.1mol/L pyrroles, 0.1mol/LNaClO
4, the phosphate buffer solution of pH=6.86, (platinum electrode is a working electrode to adopt three-electrode system, saturated calomel electrode is a reference electrode, and platinum wire electrode is a counter electrode), CONTROLLED POTENTIAL is 0.9V, oxidation 15min obtains the combined electrode that the composite conductive high molecular film is a carrier.Again this electrode is put into phosphate buffer solution electro-adsorption 20min under the voltage of 0.9V of pH=6.6 of the tyrosine oxidase of 0.03mg/mL, just obtained the tyrosine oxidase electrode.To being that the solution of substrate is measured with the cresylol, its time of response is fast, and linear response range is 3.5 * 10
-8～2.0 * 10
-5Mol/L has good storage stability.
Embodiment 4
Getting concentration is the casein 10mL with the methyl methacrylate modification of 0.1g/mL, join 20mL and contain 0.15mol/L N-methylpyrrole, 0.5mol/L KCl is in the solution of 2mg/mL L-amino-acid oxidase, (platinum electrode is a working electrode to adopt three-electrode system, saturated calomel electrode is a reference electrode, and platinum wire electrode is a counter electrode), keeping temperature is 0 ℃, electrolytic solution leads to nitrogen 10min, constant potential is 0.95V, and electropolymerization 15min just obtains L-amino-acid oxidase enzyme electrodes.To being that the solution of substrate is measured with the L-phenylalanine, its time of response is fast, and linear response range is 1.2 * 10
-5～6.3 * 10
-4Mol/L, and have good circulation ratio and highly selective.
Embodiment 5
Getting concentration is the casein solution 10mL with the methyl methacrylate modification of 0.3g/mL, join 40mL and contain 0.2mol/L pyrroles, 0.5mol/L in the solution of KCl, (carbon dioxide process carbon electrode is a working electrode to adopt three-electrode system, saturated calomel electrode is a reference electrode, and platinum wire electrode is a counter electrode), electropolymerization 20min under the current potential of 0.7V, again in the HAC-NAAC of PH=4.7 buffered soln at current potential for reducing 5min under the-0.5V, just having obtained the composite conductive high molecular film is the electrode of carrier.Then this electrode is put into citric acid-sodium dihydrogen phosphate buffer of PH=5.3, enrichment 30min under the current potential of 0.6V has just obtained the urase electrode.Solution to urea is measured, and its time of response is fast, and linear response range is 4.2 * 10
-5～1.3 * 10
-2Mol/L, and have good circulation ratio and highly selective.
Claims (9)
1, a kind of preparation is characterized in that as the method for the conductive polymer composites of biosensor immobilization carrier: prepare biosensor with the protein protein of chemically modified, modification (or through) solution and pyrroles's (or derivative of pyrroles) by the immobilization carrier that the incompatible preparation composite conductive high molecular of voltolisation film also is used as this film biologically active substance (as: enzyme, antigen, antibody, microorganism etc.).
2, according to the described preparation method of claim 1, it is characterized in that: protein in the composite conductive high molecular film that electropolymerization the obtains protein of chemically modified, modification (or through) and polypyrrole (or pyrrole derivative) form the uniform conductive film, and have formed chemical bond between the two.The color of film is yellowish to brownish black.
3, according to the described preparation method of claim 1, it is characterized in that: protein in the composite conductive high molecular film that electropolymerization the obtains protein of chemically modified, sex change (or through) and polypyrrole (or pyrrole derivative) form stratified conducting film, and have formed chemical bond between the two.The color of film is yellowish to brown.
4, according to the described preparation method of claim 1, it is characterized in that: the thickness of this composite conductive high molecular film is at 10-1000nm.
5, according to the described preparation method of claim 1, it is characterized in that: synthesis material pyrroles's the part by weight of preparation conducting polymer film is 10 to 95%, and the part by weight of the protein protein of chemically modified, modification (or through) is 5 to 85%.
6, according to the described preparation method of claim 1, it is characterized in that: the preparation conducting polymer film adopts the electropolymerization method, and its oxidizing potential is 0.2V to 1.4V, and oxidization time is 5-90 minute, and reduction potential is-1.4V to 0V that the recovery time is 0-60 minute.
7, according to the described preparation method of claim 1, it is characterized in that: the electrode by the incompatible preparation composite conductive high molecular of voltolisation film comprises metal electrode (as: platinum electrode, gold electrode, silver electrode etc.) and nonmetal electrode (as: carbon dioxide process carbon electrode etc.).
8, according to the described protein of claim 1, it is characterized in that: protein can carry out chemically modified, modification to its main chain or side chain with organic molecule (as: methyl methacrylate, propylene cyanogen etc.) or biomacromolecule.
9, described according to Claim 8 preparation method, it is characterized in that: proteinic part by weight is 10 to 85%, the part by weight that is used for carrying out protein modified organic molecule or biomacromolecule is 15 to 90%.
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| CN 200410026617 CN1673257A (en) | 2004-03-26 | 2004-03-26 | Prepn process of compound conductive polymer film used as biosensor fixing carrier |
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|---|---|---|---|
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107033349A (en) * | 2017-03-30 | 2017-08-11 | 青岛科技大学 | A kind of preparation method and applications of polypyrrole/nano platinum particle composite |
| CN107799797A (en) * | 2017-09-19 | 2018-03-13 | 华南师范大学 | A kind of anode of microbial fuel cell and preparation method thereof |
| CN109737861A (en) * | 2018-11-22 | 2019-05-10 | 厦门大学 | A kind of adjustable type protein base flexibility strain transducer and preparation method thereof, application |
| CN113533473A (en) * | 2021-06-22 | 2021-10-22 | 武汉纺织大学 | Working electrode containing metal-organic framework and preparation method and application thereof |
-
2004
- 2004-03-26 CN CN 200410026617 patent/CN1673257A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107033349A (en) * | 2017-03-30 | 2017-08-11 | 青岛科技大学 | A kind of preparation method and applications of polypyrrole/nano platinum particle composite |
| CN107033349B (en) * | 2017-03-30 | 2019-08-30 | 青岛科技大学 | A kind of preparation method and applications of polypyrrole/nano platinum particle composite material |
| CN107799797A (en) * | 2017-09-19 | 2018-03-13 | 华南师范大学 | A kind of anode of microbial fuel cell and preparation method thereof |
| CN109737861A (en) * | 2018-11-22 | 2019-05-10 | 厦门大学 | A kind of adjustable type protein base flexibility strain transducer and preparation method thereof, application |
| CN113533473A (en) * | 2021-06-22 | 2021-10-22 | 武汉纺织大学 | Working electrode containing metal-organic framework and preparation method and application thereof |
| CN113533473B (en) * | 2021-06-22 | 2024-02-06 | 武汉纺织大学 | Working electrode containing metal-organic framework, and preparation method and application thereof |
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