CN101908630B - Mediator type biological fuel cell anode and preparation method thereof - Google Patents
Mediator type biological fuel cell anode and preparation method thereof Download PDFInfo
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- CN101908630B CN101908630B CN200910085234XA CN200910085234A CN101908630B CN 101908630 B CN101908630 B CN 101908630B CN 200910085234X A CN200910085234X A CN 200910085234XA CN 200910085234 A CN200910085234 A CN 200910085234A CN 101908630 B CN101908630 B CN 101908630B
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Abstract
The invention discloses a mediator type biological fuel cell anode and a preparation method thereof. The electrode comprises a substrate electrode, a carbon nano material layer, a mediator layer and an enzyme layer, wherein the carbon nano material layer is coated on the substrate electrode; the mediator layer is coated on the carbon nano material layer; the enzyme layer is coated on the mediator layer; and the carbon nano material layer comprises a cation functionalized carbon nano material; and the enzyme layer comprises (a) oxidase or (b) immobilized enzyme formed by crosslinking oxidase, inert protein and a cross-linking agent or (c) immobilized enzyme formed by crosslinking oxidase, biopolymer and a cross-linking agent. The mediator type biological fuel cell anode of the invention has the advantages of easy preparation, high stability, and the like, more importantly, the electrode oxidizes a substrate under the neutral pH condition, and the used substrate is a substance widely existing in nature. Accordingly, the mediator type biological fuel cell anode prepared by adopting method has wide application prospect in the research of biological fuel cells.
Description
Technical field
The present invention relates to a kind of biological fuel cell anode, particularly relate to a kind of mediator type biological fuel cell anode.
Background technology
Biological fuel cell is a kind of novel energy reforming unit, and it is a catalyst with the microbe or the enzyme of occurring in nature, directly chemical energy is converted into electric energy.Biological fuel cell is except having fuel availability height, advantage such as pollution-free, and also have some unique advantages: 1) fuel source is extensive, and renewable organic substances such as a large amount of glucose that exist of nature, starch can act as a fuel; 2) reaction condition is gentle, can under normal temperature, normal pressure, pH neutral condition, react easy operating, control and maintenance; 3) bio-compatibility is good, can the energy be provided for the man-made organ or the biology sensor of implant into body.Yet, be as also remoter in real life in the body application of power, its main cause is that the stability of battery can not satisfy actual requirement far away.It is a variety of to cause the unsettled reason of battery to have, wherein, and for enzymatic biological fuel battery; Buried inside, the activated centre of most of enzymes at enzyme; Be difficult to carry out direct electron transfer between realization and the electrode, the research worker extensively adopts amboceptor to realize indirect electron transfer (Chemical Reviews.2004,104 between enzyme and the electrode both at home and abroad; 4867-4886); Yet often there are problems such as the amboceptor of being fixed is difficult for coming off quantitatively, easily in the fixed form of above-mentioned amboceptor, has limited the power and the stability of battery.
Summary of the invention
The purpose of this invention is to provide a kind of mediator type biological fuel cell anode (mediator type enzyme electrode), its amboceptor is fixed on the electrode through electrostatic interaction, can improve the stability of biological fuel cell anode.
Technical scheme of the present invention is following:
Mediator type biological fuel cell anode provided by the present invention comprises basal electrode, is coated on carbon nanomaterial layer on the said basal electrode, is coated on the amboceptor layer on the said carbon nanomaterial layer and is coated on the enzyme layer on the said amboceptor layer; Wherein, said carbon nanomaterial layer is made up of the carbon nanomaterial of cationic functionalization, said enzyme layer comprise following a) or b) or material c): a) oxidizing ferment; B) immobilised enzymes that is cross-linked to form of oxidizing ferment, inert protein and crosslinking agent; C) immobilised enzymes that is cross-linked to form of oxidizing ferment, biopolymer and crosslinking agent.
In said carbon nanomaterial layer, the content of carbon nanomaterial is 20-170 μ g on every square centimeter of basal electrode.Said carbon nanomaterial is a kind of or its combination in any in CNT, carbon nano-fiber and the Nano carbon balls; Said cation is a kind of or its combination in any in polypyrrole cation, polyaniline cation, glyoxaline cation and the indoles cation.
In said amboceptor layer, said amboceptor is [Mo (CN)
8]
3-/4-, [W (CN)
6]
3-/4-, [Fe (CN)
6]
3-/4-[Os (CN)
6]
3-/4-In a kind of or its combination in any.The content of amboceptor is 10-30 μ g on every square centimeter of basal electrode.
In said enzyme layer, oxidasic content is 1.4～217.2U on every square centimeter of basal electrode.Said inert protein can be a kind of or its combination in any in bovine serum albumin, human albumin and the gelatin; Said biopolymer can be a kind of or its combination in any in cellulose, starch, shitosan and the agarose; The mass fraction of the inert protein or the biopolymer aqueous solution is 0.5%～10%.According to the difference of oxidation substrates, can select to be directed against accordingly the oxidizing ferment of this substrate: for example,, select alcohol oxidase for use for ethanol; For glucose, select glucose oxidase for use.
The present invention has no special requirements to the size of nano material, and in general, the diameter of said CNT, carbon nano-fiber is 1～500nm, is preferably 1～10nm, and average length is 0.5～50 μ m, is preferably 5～15 μ m; The diameter of said Nano carbon balls is 10～500nm.
Prepare the method for above-mentioned mediator type biological fuel cell anode, may further comprise the steps:
1) described carbon nanomaterial is carried out sonicated, then the carbon nanomaterial after the sonicated is coated on the basal electrode drying at room temperature;
2) will be through 1) electrode handled is soaked in the said amboceptor solution, takes out after 10～30 minutes, and drying at room temperature forms the amboceptor layer;
3) will contain oxidasic solution and be coated on the amboceptor layer, drying at room temperature forms the enzyme layer on the amboceptor layer, promptly obtain described based on oxidasic mediator type biological fuel cell anode.
Wherein, can better be fixed on the electrode, also can contain crosslinking agent in the oxidizing ferment solution in order to make the enzyme layer.Said crosslinking agent can like glutaraldehyde, hexamethylene diamine, maleic anhydride, bisazo benzene etc., be preferably glutaraldehyde for common function reagent.In the specifically preparation process of mediator type biological fuel cell anode, various material consumptions are following on every square centimeter of basal electrode: carbon nanomaterial 10～85 μ L that contain the cationic functionalization of 2mg/mL in the carbon nanomaterial layer; Oxidizing ferment solution 14～114.2 μ L, the mass fraction that contains 100～1900U/mL in the enzyme solutions is that 0.5～10% inert protein solution or biopolymer solution 14～86 μ L, mass fraction are 0.5～6% cross-linking agent solution 7～57 μ L.
Carbon nanomaterial has caused scientific research field and industrial circle worker's extensive interest owing to have unique mechanical property, physicochemical properties and biocompatibility.Carbon nanomaterial itself and the nano material that obtains afterwards through functionalization have crucial value in the research in Electroanalytical Chemistry field.In activated centre of realizing enzyme and the research of the electronics indirect branch between the electrode, [Mo (CN)
8]
3-/4-, [W (CN)
6]
3-/4-, [Fe (CN)
6]
3-/4-[Os (CN)
6]
3-/4-Deng being the electron mediator that is widely used, can realize the transfer of electronics well.So through utilizing carbon nanomaterial and the interaction between the electron mediator after carbon nanomaterial or the functionalization, both are compounded to form multi-functional compound, can realize the indirect branch of electronics.
The present invention at first introduces one or several effects in pyrroles's cation, polyaniline cation, glyoxaline cation or the indoles cation with carbon nanomaterial through a series of chemical reaction, has prepared the carbon nanomaterial of macrocation functionalization.Then, the carbon nanomaterial of macrocation functionalization is fixed in the basal electrode surface, electrode is placed the solution that contains amboceptor, utilize the carbon nanomaterial and [Mo (CN) of macrocation functionalization
8]
3-/4-, [W (CN)
6]
3-/4-, [Fe (CN)
6]
3-/4-[Os (CN)
6]
3-/4-Deng in one or several interact, the carbon nanomaterial of preparation macrocation functionalization and the nano-complex of amboceptor are fixed in electrode surface, can be used as oxidasic electron mediator.At last, on the nano-complex of carbon nanomaterial and amboceptor, apply the enzyme layer that contains oxidizing ferment, crosslinking agent and inert protein or biopolymer composition, make mediator type biological fuel cell anode.This enzyme electrode has advantages such as easy preparation, good stability, what is more important, and this enzyme electrode carries out under condition of neutral pH the oxidation of substrate, and used substrate is the material that occurring in nature extensively exists.Therefore, adopt the mediator type biological fuel cell anode of this method preparation in the research of biological fuel cell, to have wide practical use.
Description of drawings
Fig. 1 is the structural representation of the mediator type biological fuel cell anode of embodiment 1,2,3 preparations; Wherein
represents CNT or carbon nano-fiber;
represents amboceptor;
represents oxidizing ferment;
represents crosslinking agent, and
represents basal electrode.
Fig. 2 is the mediator type biological fuel cell anode of embodiment 1 preparation, under saturation of the air state, is to the cyclic voltammogram of glucose responding in 7.4 the phosphate buffer solution in the pH value.
Fig. 3 is the mediator type biological fuel cell anode of embodiment 2 preparation, under saturation of the air state, is to the cyclic voltammogram of glucose responding in 7.4 the phosphate buffer solution in the pH value.
Fig. 4 is the mediator type biological fuel cell anode of embodiment 3 preparation, under saturation of the air state, is to the cyclic voltammogram of glucose responding in 7.4 the phosphate buffer solution in the pH value.
Embodiment
The preparation of embodiment 1, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic for 4mg glyoxaline cation functionalized carbon nanotube; Diameter range 20～30nm; Average length is 0.5～50 μ m; Analytical Chemistry 80 (17): 6587-6593) concrete building-up process is: at first, multi-walled carbon nano-tubes is ultrasonic to introduce carboxyl in 3: 1 sulfuric acid and nitric acid mixed liquor, and the product that obtains and thionyl (two) chlorine reacts the introducing acid chloride groups; Then, under the ferric trichloride catalytic effect, acid chloride groups starts epoxychloropropane multi-walled carbon nano-tubes is carried out the ring-opening polymerisation of original position cation, obtains the polyether grafting CNT; At last, with the methylimidazole reaction, obtain the glyoxaline cation functionalized carbon nanotube.It is dispersed in the 2mL distilled water continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) two hours; Draw on 5 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL; Take out after 20 minutes; Dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought SISIGMA-ALDRICH company, and cat. no is: G7141) 1.9U (1900U/mL, 1 μ L), bovine serum albumin 20 μ g (1% aqueous solution will to contain glucose oxidase; 2 μ L) and crosslinking agent (glutaraldehyde) 10 μ g (1% aqueous solution; 1 μ L) 4 μ L mixed liquors are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological fuel cell.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 80% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
This electrode performance test is as shown in Figure 2 in the description of drawings; Wherein, Solid line represents the prepared glucose oxidase electrode of embodiment 1 in pH is 7.4 phosphate buffer solution; Under the saturation of the air state, the cyclic voltammetry curve when concentration of glucose is 4mM, the cyclic voltammetry curve when the dotted line representative does not add glucose.Testing used reference electrode is the Ag/AgCl reference electrode, is platinum filament to electrode.
Add after the glucose oxidation peak current and have when not adding glucose obvious increase (oxidation peak current is electric current shown in the figure peak, middle and upper part) through seeing the comparison of solid line and dotted line among the figure, reduction peak current has significantly when not adding glucose and reduces (reduction peak current is for scheming electric current shown in the peak, middle and lower part) after adding glucose simultaneously.Therefore can reach a conclusion: prepared electrode has good catalysed oxidn for glucose.
The preparation of embodiment 2, mediator type biological fuel cell anode
The glyoxaline cation functionalized carbon nanotube of preparation among the 4mg embodiment 1 is dispersed in the 2mL distilled water; Continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) 30 minutes is drawn on 4 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode; Dry under the room temperature; Dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL, takes out after 20 minutes, dry under the room temperature; Form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company will to contain glucose oxidase 5.7U; Cat. no is: G7141) the 5 μ L mixing drops of (1900U/mL, 3 μ L) and bovine serum albumin 20 μ g (1% aqueous solution, 2 μ L) are coated on the amboceptor layer; Dry under the room temperature; Form the enzyme layer, obtain glucose oxidase electrode, this electrode can be used as the anode of biological dye battery.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 76% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
This electrode performance test is as shown in Figure 3 in the description of drawings; Wherein, Solid line represents the prepared glucose oxidase electrode of embodiment 1 in pH is 7.4 phosphate buffer solution; Under the saturation of the air state, the cyclic voltammetry curve when concentration of glucose is 2mM, the cyclic voltammetry curve when the dotted line representative does not add glucose.Testing used reference electrode is the Ag/AgCl reference electrode, is platinum filament to electrode.Contrast can draw: prepared electrode pair glucose has good catalysed oxidn.
The preparation of embodiment 3, mediator type biological fuel cell anode
The glyoxaline cation functionalized carbon nanotube of preparation among the 4mg embodiment 1 is dispersed in the 2mL distilled water; Continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) 30 minutes is drawn on 4 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode; Dry under the room temperature; Dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL, takes out after 20 minutes, dry under the room temperature; Form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141 (1900U/mL, 8 μ L), shitosan 100 μ g (10% aqueous solution will to contain glucose oxidase 15.2U; 1 μ L) and crosslinking agent (glutaraldehyde) 40 μ g (4% aqueous solution; 1 μ L) 10 μ L mixing drops are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological dye battery.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 85% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
This electrode performance test is as shown in Figure 4 in the description of drawings; Wherein, Solid line represents the prepared glucose oxidase electrode of embodiment 1 in pH is 7.4 phosphate buffer solution; Under the saturation of the air state, the cyclic voltammetry curve when concentration of glucose is 20mM, the cyclic voltammetry curve when the dotted line representative does not add glucose.Testing used reference electrode is the Ag/AgCl reference electrode, is platinum filament to electrode.Contrast can draw: prepared electrode pair glucose has good catalysed oxidn.
The preparation of embodiment 4, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic for the Nano carbon balls of 4mg glyoxaline cation functionalization; Diameter 0.5 μ m; With reference to Macromolecular Chemistry and Physics 207 (18): 1633-1639 and AnalyticalChemistry 80 (17): 6587-6593) concrete building-up process is: at first; Nano carbon balls is ultrasonic to introduce carboxyl, the product that obtains and thionyl (two) chlorine reaction introducing acid chloride groups in 3: 1 sulfuric acid and nitric acid mixed liquor; Then, under the ferric trichloride catalytic effect, acid chloride groups starts epoxychloropropane Nano carbon balls is carried out the ring-opening polymerisation of original position cation, obtains the polyether grafting Nano carbon balls; At last, with the methylimidazole reaction, obtain the Nano carbon balls of glyoxaline cation functionalization.It is dispersed in the 2mL distilled water continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) 30 minutes; Draw on 7 μ L are coated in 0.07 square centimeter through the Nano carbon balls suspension-turbid liquid of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL; Take out after 20 minutes; Dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of Nano carbon balls quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141 (1900U/mL, 8 μ L), shitosan 100 μ g (10% aqueous solution will to contain glucose oxidase 15.2U; 1 μ L) and crosslinking agent (glutaraldehyde) 40 μ g (4% aqueous solution; 1 μ L) 10 μ L mixing drops are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological dye battery.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 80% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
The preparation of embodiment 5, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic for the CNT of 4mg polypyrrole cationic functionalization; Diameter range 20～30nm; Average length is 0.5～50 μ m; Analytical Chemistry 80 (17): 6587-6593) concrete building-up process is: at first, multi-walled carbon nano-tubes is ultrasonic to introduce carboxyl in 3: 1 sulfuric acid and nitric acid mixed liquor, and the product that obtains and thionyl (two) chlorine reacts the introducing acid chloride groups; Then, under the ferric trichloride catalytic effect, acid chloride groups starts epoxychloropropane multi-walled carbon nano-tubes is carried out the ring-opening polymerisation of original position cation, obtains the polyether grafting CNT; At last, with the polypyrrole reaction, obtain the CNT of polypyrrole cationic functionalization.It is dispersed in the 2mL distilled water continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) two hours; Draw on 5 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL; Take out after 20 minutes; Dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141) 1.9U (1900U/mL, 1 μ L), bovine serum albumin 20 μ g (1% aqueous solution will to contain glucose oxidase; 2 μ L) and crosslinking agent (glutaraldehyde) 10 μ g (1% aqueous solution; 1 μ L) 4 μ L mixed liquors are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological fuel cell.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 75% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
The preparation of embodiment 6, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic for the CNT of 4mg polyaniline cationic functionalization; Diameter range 20～30nm; Average length is 0.5～50 μ m; Analytical Chemistry 80 (17): 6587-6593) concrete building-up process is: at first, multi-walled carbon nano-tubes is ultrasonic to introduce carboxyl in 3: 1 sulfuric acid and nitric acid mixed liquor, and the product that obtains and thionyl (two) chlorine reacts the introducing acid chloride groups; Then, under the ferric trichloride catalytic effect, acid chloride groups starts epoxychloropropane multi-walled carbon nano-tubes is carried out the ring-opening polymerisation of original position cation, obtains the polyether grafting CNT; At last, with the polyaniline reaction, obtain the CNT of polyaniline cationic functionalization.It is dispersed in the 2mL distilled water continuous ultrasound under the room temperature (ultrasonic power is 50W, and frequency is 50Hz) two hours; Draw on 5 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the potassium ferricyanide solution that concentration is 10mg/mL; Take out after 20 minutes; Dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141) 1.9U (1900U/mL, 1 μ L), bovine serum albumin 20 μ g (1% aqueous solution will to contain glucose oxidase; 2 μ L) and crosslinking agent (glutaraldehyde) 10 μ g (1% aqueous solution; 1 μ L) 4 μ L mixed liquors are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological fuel cell.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 72% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
The preparation of embodiment 7, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic with the glyoxaline cation functionalized carbon nanotube for preparing among the 4mg embodiment 1; Diameter range 20～30nm; Average length is 0.5～50 μ m, and Analytical Chemistry 80 (17): 6587-6593) be dispersed in the 2mL distilled water, (ultrasonic power is 50W to continuous ultrasound under the room temperature; Frequency is 50Hz) two hours; Draw on 5.9 μ L are coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the [Mo (CN) that concentration is 10mg/mL
8]
3-/4-In the solution, take out after 20 minutes, dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141) 1.9U (1900U/mL, 1 μ L), bovine serum albumin 20 μ g (1% aqueous solution will to contain glucose oxidase; 2 μ L) and crosslinking agent (glutaraldehyde) 10 μ g (1% aqueous solution; 1 μ L) 4 μ L mixed liquors are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological fuel cell.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 78% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
The preparation of embodiment 8, mediator type biological fuel cell anode
(Institute of Chemistry, Academia Sinica is synthetic with the glyoxaline cation functionalized carbon nanotube for preparing among the 4mg embodiment 1; Diameter range 20～30nm; Average length is 0.5～50 μ m, and Analytical Chemistry 80 (17): 6587-6593) be dispersed in the 2mL distilled water, (ultrasonic power is 50W to continuous ultrasound under the room temperature; Frequency is 50Hz) two hours; Draw on 0.7 μ L is coated in 0.07 square centimeter through the carbon nanotube suspension of sonicated the glass carbon substrate electrode, dry under the room temperature, dried electrode is soaked in the [Os (CN) that concentration is 10mg/mL
6]
3-/4-In the solution, take out after 20 minutes, dry under the room temperature, form the amboceptor layer, the content of amboceptor can calculate through the variation of CNT quality before and after relatively adsorbing.(glucose oxidase is bought from SIGMA-ALDRICH company, and cat. no is: G7141) 15.2U (1900U/mL, 8 μ L), bovine serum albumin 20 μ g (1% aqueous solution will to contain glucose oxidase; 2 μ L) and crosslinking agent (glutaraldehyde) 10 μ g (1% aqueous solution; 1 μ L) 4 μ L mixed liquors are coated on the amboceptor layer, and are dry under the room temperature, form the enzyme layer; Obtain glucose oxidase electrode, this electrode can be used as the anode of biological fuel cell.Prepared enzyme electrode not the time spent be kept at (temperature of setting during preservation is 4 ℃) in the refrigerator, at the same terms, be original 82% after 20 days to the catalytic oxidation electric current of same concentrations glucose.
Claims (12)
1. mediator type enzyme electrode comprises basal electrode, is coated on carbon nanomaterial layer on the said basal electrode, is coated on the amboceptor layer on the said carbon nanomaterial layer and is coated on the enzyme layer on the said amboceptor layer; Wherein, said carbon nanomaterial layer is made up of the carbon nanomaterial of cationic functionalization; Said enzyme layer comprise following a) or b) or material c): a) oxidizing ferment; B) immobilised enzymes that is cross-linked to form of oxidizing ferment, inert protein and crosslinking agent; C) immobilised enzymes that is cross-linked to form of oxidizing ferment, biopolymer and crosslinking agent.
2. mediator type enzyme electrode according to claim 1 is characterized in that: said carbon nanomaterial is a kind of or its combination in any in CNT, carbon nano-fiber and the Nano carbon balls; Said cation is a kind of or its combination in any in polypyrrole cation, polyaniline cation, glyoxaline cation and the indoles cation; Said amboceptor is [Mo (CN)
8]
3-/4-, [W (CN)
6]
3-/4-, [Fe (CN)
6]
3-/4-[Os (CN)
6]
3-/4-In a kind of or its combination in any.
3. mediator type enzyme electrode according to claim 1 and 2 is characterized in that: in the said carbon nanomaterial layer, the content of carbon nanomaterial is 20～170 μ g on every square centimeter of basal electrode; In the said amboceptor layer, the content of amboceptor is 10～30 μ g on every square centimeter of basal electrode; In the said enzyme layer, every square centimeter of the above oxidasic content of basal electrode is 1.4～217.2U.
4. mediator type enzyme electrode according to claim 1 and 2 is characterized in that: said inert protein is a kind of or its combination in any in bovine serum albumin, human albumin and the gelatin; Said biopolymer is a kind of or its combination in any in cellulose, starch, shitosan and the agarose; Forming the used crosslinking agent of said immobilised enzymes is glutaraldehyde, hexamethylene diamine, maleic anhydride or bisazo benzene.
5. mediator type enzyme electrode according to claim 2 is characterized in that: the diameter of said CNT, carbon nano-fiber is 1～500nm, and average length is 0.5～50 μ m; The diameter of said Nano carbon balls is 10～500nm.
6. mediator type enzyme electrode according to claim 5 is characterized in that: the diameter of said CNT, carbon nano-fiber is 1～10nm, and average length is 5～15 μ m.
7. prepare the method for arbitrary said mediator type enzyme electrode among the claim 1-6, may further comprise the steps:
1) arbitrary described carbon nanomaterial among the claim 1-6 is carried out continuous ultrasound and handle, the carbon nanomaterial after then continuous ultrasound being handled is coated on the basal electrode drying at room temperature;
2) will pass through the basal electrode that step 1) handles and be soaked in the solution of arbitrary said amboceptor among the claim 1-6, and take out after 10～30 minutes, drying at room temperature forms the amboceptor layer;
3) will contain following a) or b) or the solution of material c) be coated on the amboceptor layer, drying at room temperature forms the enzyme layer on said amboceptor layer, promptly obtain described mediator type enzyme electrode; Said a) is oxidizing ferment; B) be oxidizing ferment, crosslinking agent and inert protein; C) be oxidizing ferment, crosslinking agent and biopolymer.
8. method according to claim 7 is characterized in that: the continuous ultrasound of said step 1) is handled and in distilled water, acetonitrile or ethanol, is carried out.
9. according to claim 7 or 8 described methods; It is characterized in that: in the step 1) carbon nanomaterial is coated on the method on the basal electrode: the solution with carbon nanomaterial applies; The concentration of said carbon nanomaterial solution is 2mg/ml, and the amount that makes carbon nanomaterial on every square centimeter of basal electrode is 20-170 μ g.
10. according to claim 7 or 8 described methods; It is characterized in that: will contain b in the step 3)) the solution of material when being coated on the amboceptor layer, every square centimeter of basal electrode uses the mixed solution that contains following consumption: oxidizing ferment solution 14.2～114.3 μ L, the mass fraction that contains 100～1900U/mL in the enzyme solutions is that 0.5～10% inert protein solution 14.2～85.7 μ L and mass fraction are 0.5～6% cross-linking agent solution 7.1～57.1 μ L.
11. according to claim 7 or 8 described methods; It is characterized in that: will contain c in the step 3)) the solution of material when being coated on the amboceptor layer, every square centimeter of basal electrode uses the mixed solution that contains following consumption: oxidizing ferment solution 14.2～114.3 μ L, the mass fraction that contains 100～1900U/mL in the enzyme solutions is that 0.5～10% biopolymer solution 14.2～85.7 μ L and mass fraction are 0.5～6% cross-linking agent solution 7.1～57.1 μ L.
12. arbitrary described mediator type enzyme electrode is as the application of biological fuel cell anode among the claim 1-6.
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| CN102411022A (en) * | 2011-08-03 | 2012-04-11 | 中国科学院化学研究所 | Electrochemical sensor based on primary battery and preparation method thereof |
| CN103086508B (en) * | 2011-11-04 | 2014-07-23 | 中国科学院城市环境研究所 | Microbial fuel cell wastewater treatment system for improving nitrogen removal effect |
| CN103682353A (en) * | 2012-09-12 | 2014-03-26 | 苏州宝时得电动工具有限公司 | Electrode composite material as well as preparation method thereof, positive electrode and battery with same |
| WO2014098153A1 (en) * | 2012-12-19 | 2014-06-26 | トヨタ自動車株式会社 | Bioreactor comprising immobilized enzyme, activity improvement method for immobilized enzyme, and biofuel cell |
| CN103066302B (en) * | 2012-12-20 | 2015-08-26 | 华南理工大学 | A kind of biological fuel cell anode and preparation method thereof and application |
| CN103066304B (en) * | 2012-12-20 | 2015-10-28 | 华南理工大学 | A kind of enzyme biological fuel cell positive pole and preparation method thereof and application |
| CN103103178A (en) * | 2013-02-06 | 2013-05-15 | 扬州大学 | Reaction-adsorption coupling immobilization protease preparation method |
| CN103088011A (en) * | 2013-02-06 | 2013-05-08 | 扬州大学 | Preparation method of reaction-adsorption coupled immobilized lipase |
| CN103103177A (en) * | 2013-02-06 | 2013-05-15 | 扬州大学 | Reaction-adsorption coupling immobilization oxidoreductase preparation method |
| CN105244512B (en) * | 2015-08-28 | 2018-07-10 | 华南理工大学 | A kind of glycerin catalytic biological fuel cell anode and preparation method and application |
| CN105552395B (en) * | 2015-12-15 | 2018-01-16 | 华南理工大学 | A kind of biological fuel cell anode of enzymatic glycerine and preparation method and application |
| CN106450399A (en) * | 2016-11-11 | 2017-02-22 | 青岛大学 | High-performance starch/oxygen fuel cell based on microbial surface co-display sequential enzyme |
| JP6891372B2 (en) * | 2019-06-28 | 2021-06-18 | 東洋紡株式会社 | Method for predicting the presence or absence of an enzyme-electrode electron transfer enhancing effect |
| JP2021040633A (en) * | 2020-11-04 | 2021-03-18 | 東洋紡株式会社 | Method for predicting presence of effect of enhancing electron transport between enzyme-electrode |
| CN115172834B (en) * | 2022-09-08 | 2022-11-29 | 中国科学院天津工业生物技术研究所 | Flexible enzyme fuel cell, ion electro-osmosis mask driven by flexible enzyme fuel cell and preparation methods of flexible enzyme fuel cell and ion electro-osmosis mask |
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| CN101131374A (en) * | 2006-08-23 | 2008-02-27 | 佳能株式会社 | Enzyme electrode, enzyme electrode producing method, sensor and fuel cell each using enzyme electrode |
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