CN101573816A - Bioanode and biocathode stack assemblies - Google Patents

Abstract

A biofuel cell device for generating electrical current. The device includes a fuel manifold having a face, and at least one cavity in the face defining a fuel reservoir, an inlet in fluid communication with the reservoir for flow of fuel fluid into the manifold to fill the reservoir and an outlet in fluid communication with the reservoir for flow of fuel fluid out of the manifold. The device has an anode assembly including at least one bioanode positioned for contact with fuel fluid in the fuel reservoir, and a cathode assembly including at least one cathode positioned for flow of fuel fluid through the bioanode to the cathode. The device includes a controller operatively connected to the anode assembly and the cathode assembly for controlling the output of electrical current from the biofuel cell device.

Description

Biological anode and biological-cathode stack assemblies

Background

Relate generally to of the present invention is based on fuel cell (being also referred to as biological fuel cell) of biology enzyme and its production and use.More specifically, the present invention relates to biological anode, biological anode stacked body (stack), biological-cathode and its production and use.

Biological fuel cell be a kind of can be therein catalytic activity by living cells and/or their the enzyme Conversion of energy that will obtain from chemical reaction be the electrochemical apparatus of electric energy.It is the required hydrogen ion of water that the molecule of biological fuel cell general using complexity generates hydrogen reduction at anode, produces the free electron that is used in the electrical application simultaneously.Biological anode is the electrode that discharges electronics in the biological fuel cell herein after the oxygenated fuel, and biological-cathode is that the electronics and the proton that are used to from anode of catalyst is the electrode of water with peroxide or hydrogen reduction herein.Biological fuel cell is different from the material that the conventional fuel cell part is to be used for the catalytic electrochemical reaction.Biological fuel cell depends on biomolecule such as enzyme reacts, rather than utilizes noble metal as catalyst.

Summary of the invention

Relate to a kind of biological fuel cell equipment that is used to produce electric current in many aspects of the present invention, it comprises fuel conflux chamber (fuel manifold), anode assemblies, cathode assembly, housing and controller.Fuel conflux the chamber have face and in face, define fuel storage pond (fuel reservoir) at least one chamber (cavity), be used for fuel fluid and flow into this chamber of confluxing with the import of filling the storage pond be used for the outlet that fuel fluid flows out this chamber of confluxing.Anode assemblies comprises and is provided at least one biological anode of contacting with fuel fluid in the described fuel storage pond.Cathode assembly comprises and is provided for fuel fluid flows to negative electrode by biological anode at least one biological-cathode.Housing holds the chamber of confluxing, anode assemblies and cathode assembly.The controller Control current is from the output of this biological fuel cell equipment.

Relate to a kind of biological fuel cell equipment that is used for to load (load) supply electric power (electrical power) on the other hand, described equipment comprises at least one fuel cell, the controller that is used for the output of operational mode control fuel cell according to the rules, and the switching circuit between fuel cell and load, described switching circuit response controller is alternately to connect according to operational mode and to disconnect fuel cell and load.

Relate to a kind of biological fuel cell equipment that is used for to load supply electric power on the other hand, described equipment comprises at least one fuel cell, the controller that is used for the output of operational mode control fuel cell according to the rules, and additional power circuit, this power circuit response controller is connected with the output of fuel cell optionally will replenish power supply, replenishes the electric power of this biological fuel cell equipment to the load supply thus.

Relate to a kind of biological fuel cell equipment that is used for to load supply electric power on the other hand, described equipment comprises a plurality of fuel cells that tandem is electrically connected, be used for controller according to the output of at least a each fuel cell of control of multiple regulation operational mode, and the switching circuit between fuel cell and load, described switching circuit response controller is optionally to make one or more fuel cells be connected with load according to operational mode.

The method that electricity is adjusted (electrically conditioning) is carried out in the output that relates to a kind of one or more fuel cells to biological fuel cell equipment on the other hand, thereby this described biological fuel cell equipment is adapted to load supply electric power, described method is included between fuel cell and the load and is electrically connected switching circuit, and operation switching circuit is optionally to make one or more fuel cells be connected with load according at least a in the multiple regulation operational mode.

Another aspect of the present invention relates to a kind of electrode, it comprises that air can see through and first district of the electrical conductivity material of fuel fluid impermeable, second district of all permeable electrical conductivity material of fuel fluid and air, and the non-precious metal catalyst that can contact fuel fluid and air.The air self-respiration type half-cell (air-breathing half-cell) that comprises this electrode is at room temperature, 0.4V electrode potential and 10mg/cm ²Catalyst loading produces during operation down at least about 16,20,25,30,35,40,45,50,55,60,65,70,75,80 or bigger mA/cm ²Current density.

Another aspect of the present invention relates to that a kind of it comprises electronic conductor through heat treated electrode, and at least a can optionally be the non-precious metal catalyst of water with hydrogen reduction.The air self-respiration type half-cell that comprises this electrode is at room temperature, 0.4V electrode potential and 10mg/cm ²Catalyst loading produces during operation down at least about 16,20,25,30,35,40,45,50,55,60,65,70,75,80 or bigger mA/cm ²Current density.

Another aspect of the present invention relates to biological-cathode, and it comprises electronic conductor, can generate at least a cathode enzyme of water with oxidant reaction and can fix and stablize this enzyme and can see through the enzyme immobilization material of this oxidant.In addition, this electronic conductor comprises functionalized multi-walled carbon nano-tubes, activated carbon (activated carbon) sill or its combination.

Another aspect of the present invention relates to and contains cobalt (II) 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, the catalyst of 31H-phthalocyanine (CoPcF) and polypyrrole.This CoPcF and polypyrrole through Overheating Treatment to increase the interaction between cobalt metallic atom and the polypyrrole nitrogen-atoms.

Another aspect of the present invention relates to the particle that contains the nuclear core that immobilised enzymes coats, this enzyme be fixed in the immobilization material and have with respect to its in immobilization and the initial activity before coating at least about 0.65,0.7,0.75,0.8,0.85,0.9,0.95 or bigger activity.

Another aspect of the present invention relates to the particle that contains the nuclear core that immobilised enzymes coats, this enzyme be fixed in the immobilization material and when this enzyme continuous catalysis chemical conversion, continue to keep at least 7 days its initial catalytic activity at least about 75%.In a plurality of embodiment preferred, this enzyme when the continuous catalysis chemical conversion, kept at least 30 days its initial catalytic activity at least about 75%.

The particle that relates to the nuclear core that contains the coating of immobilized cell device on the other hand.This organelle is fixed in the immobilization material.

Relate to a kind of method for preparing the particle of immobilised enzymes or organelle coating on the other hand, this method comprises that the suspension that mixes the solution that contains enzyme or organelle and contain at least a nuclear core particle, immobilization material and liquid medium is to form mixture.Then, with this mixture spray drying.

Another aspect of the present invention relates to the enzyme that is fixed in the enzyme immobilization material, wherein this enzyme comprises that the amylase and the enzyme immobilization material that consume starch comprise the butyl shitosan that is suspended in the tert-pentyl alcohol, and perhaps this enzyme comprises the decyl modification of chitosan that the amylase that consumes maltose and enzyme immobilization material comprise intermediate molecular weight.

Relate to the self-bearing type electronic conductor on the other hand, comprise by have high surface be used for metastatic electron the first electrical conductivity material, be used to support second electrical conductivity material of electronic conductor and the individual layer that adhesive is formed.The weight ratio of the second electrical conductivity material and the first electrical conductivity material is at least 0.5: 1 in this electronic conductor, thinks that this electronic conductor provides enough rigidity to achieve self-supporting.In a plurality of embodiment preferred, the weight ratio of the second electrical conductivity material and the first electrical conductivity material is at least 1: 1 in this electronic conductor.

Description of drawings

Fig. 1 is the schematic diagram of fuel cell system of the present invention.

Fig. 2 is the perspective view of fuel cell system of the present invention.

Fig. 3 is the perspective view of taking apart of Fig. 2 equipment.

Fig. 4 is the conflux part perspective view (fragmentary perspective) in chamber of the fuel of Fig. 2 equipment.

Fig. 5 is the conflux front view in chamber of fuel.

Fig. 6 is the fuel cutaway view (cross section) of chamber along the line 6-6 of Fig. 5 that conflux.

Fig. 7 is the front view of the anode assemblies of Fig. 2 equipment.

Fig. 8 is the perspective view of taking apart of Fig. 7 anode assemblies.

Fig. 9 is the cutaway view of anode assemblies along the line 9-9 of Fig. 7.

Figure 10 is the front view of the cathode assembly of Fig. 2 equipment.

Figure 11 is the perspective view of taking apart of Figure 10 cathode assembly.

Figure 12 is the cutaway view of cathode assembly along the line 12-12 of Figure 10.

Figure 13 is the perspective view of the housing parts of fuel cell system of the present invention.

Figure 14 is the schematic diagram of fuel cell system.

Figure 15 is the schematic diagram of the controller of fuel cell system.

Figure 16 and 17 has described exemplary user interface, and this interface is used to receive user's input is used for the equipment of 8 batteries with design and simulation pattern.

Figure 18 is the perspective view of the electronic building brick of fuel cell system.

Figure 19 is the perspective view of taking apart of the fuel cell system of second embodiment of the invention.

Figure 20 is the conflux perspective view in chamber of the fuel of Figure 19 equipment.

Figure 21 is the perspective view of taking apart of Figure 19 anode assemblies.

Figure 22 is the electrode schematic diagram, and this electrode comprises that air can see through but the second area and the catalyst that can contact fuel fluid and air of all permeable electrical conductivity material of first area, fuel fluid and air of the impervious electrical conductivity material of fuel fluid.This electrode is the work electrode (WE) of air self-respiration type half-cell.This work electrode (WE) comprises zone or the layer that is labeled as first area, second area and catalyst.This half-cell also comprise near place the work electrode reference electrode (RE) and preferably precious metal network or carbon plate to electrode (CE).

Figure 23 is the electronic conductor with asymmetric pore size distribution.

Figure 24 is polarization (polarization) and a power curve, has showed the present progress of NAD-dependent form GDH biological fuel cell stacked body.

Figure 25 is two kinds of carbon gas diffusion layer materials of comparison (one-sided Elat and the gas diffusion layers that makes as described herein (GDL)) and two kinds of current collector material (SS and the Ni of the plating Au) curve to the influence of NAD-dependent form ADH anode performance.

Figure 26 is carbon carrier (gas diffusion layers) material character of comparison different materials and they curve to the influence of NAD-dependent form ADH anode performance.

Figure 27 is the curve of 6 kinds of ADH enzymes of comparison and commercially available ADH (NAD dependent form).

Figure 28 is single half-cell anode (1cm ²) power curve, show that power density is 15mW/cm ²

Figure 29 A and 29B are polarization and power curve, the present progress and the present situation of performance PQQ dependent form ADH biological fuel cell stacked body performance.

Figure 30 A and 30B are polarization and power curve, the present progress and the present situation of performance PQQ dependent form ADH biological fuel cell stacked body performance.

Figure 31 shows the polarization of air self-respiration type negative electrode in the 1M sulfuric acid solution.With platinum black (Johnson Matthey) with 5mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 10%Nafion in this catalyst layer.With CoPcFCPPy with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer, and exist or do not have phosphotungstic acid (PTA), if when existing then (PTA: CoPcFCPPy) than being 1: 10.Sweep speed: 2mV/s.

Figure 32 is the polarization curve of cathode electrode in the 1M sulfuric acid solution with 15% or 30% ethanol.With the CoPcFCPPy that as above makes with 10mg/cm ²Carrying capacity be coated onto on the Torey carbon paper, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 33 is the polarization curve of air self-respiration type negative electrode in containing the 1M sulfuric acid solution of 15% ethanol.The CoPcFCPPy that will make by embodiment 7 is with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 34 is the time dependence hydrogen reduction electric current under the 0.4V in containing the 1M sulfuric acid solution of 5% ethanol.The CoPcFCPPy that embodiment 7 is made is with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer, and exist or do not have phosphotungstic acid (PTA), if when existing then (PTA: CoPcFCPPy) than being 1: 10.With platinum black (Johnson Matthey) with 5mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 10%Nafion in this catalyst layer.For relatively, also in the 1M sulfuric acid solution, test platinum cathode.Sweep speed: 2mV/s.

Figure 35 is the polarization curve of air self-respiration type negative electrode in containing the 1M sulfuric acid solution of 15% ethanol.With platinum black (Johnson Matthey) with 5mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 10%Nafion in this catalyst layer.With CoPcFCPPy with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 36 is the polarization curve of air self-respiration type negative electrode in the 1M sulfuric acid solution.With the CoPcFCPPy catalyst with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.Before making this air self-respiration type negative electrode, with CoPcFCPPy catalyst lixiviate 0,1,20 or 120 hours in 1M sulfuric acid.

Figure 37 is the polarization curve of air self-respiration type negative electrode in the 1M sulfuric acid solution.Will through pyrolysis or without the CoPcFCPPy of pyrolysis with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 38 is the polarization curve of air self-respiration type negative electrode in the 1M sulfuric acid solution that has or do not exist 15% ethanol.Will be through the CoPcF/C of pyrolysis catalyst with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 39 is the polarization curve of air self-respiration type negative electrode in the 1M sulfuric acid solution.With CoPcFPPy and CoPcCPPy catalyst with 10mg/cm ²Carrying capacity be coated onto on the carbon cloth (E-Tek) that Teflon handled, have 30%Nafion in this catalyst layer.Sweep speed: 2mV/s.

Figure 40 is bilirubin oxidase (myrothecium verrucaria (Myrothecium verrucaria))-cyclic voltammogram (cyclic voltammogram) of multi-walled carbon nano-tubes in acetate buffer (pH 5) that is fixed on the Torey carbon paper.Sweep speed: 100mV/s.

Figure 41 is laccase (agaricus bisporus (Agaricus bisporus))-cyclic voltammogram of multi-walled carbon nano-tubes in acetate buffer (pH 5) that is fixed on the Torey carbon paper.Sweep speed: 50mV/s.

Figure 42 is the polarization curve of air self-respiration type biological-cathode in acetic acid buffer solution (pH 5).Laccase (Trametes (Trametes) species)/carboxy-functionalized multi-walled carbon nano-tubes is fixed on the waterproof carbon cloth.Sweep speed: 2mV/s.

Figure 43 is the polarization curve of air self-respiration type biological-cathode in acetic acid buffer solution (pH 5).Bilirubin oxidase (myrothecium verrucaria)/carbon nano-tube is fixed on the waterproof carbon cloth.Sweep speed: 2mV/s.

Figure 44 is the polarization curve of air self-respiration type biological-cathode in acetic acid buffer solution (pH 5).Bilirubin oxidase (myrothecium verrucaria)/carbon black or carbon nano-tube are fixed on the waterproof carbon cloth.Sweep speed: 2mV/s.

Figure 45 is the polarization curve of air self-respiration type biological-cathode in acetic acid buffer solution (pH 5).The multi-walled carbon nano-tubes of bilirubin oxidase (myrothecium verrucaria)/hydroxy-functional is fixed on waterproof carbon cloth (bilateral coating-from the DS of E-Tek; Self-control coating-carbon cloth D/XE2) on.Sweep speed: 2mV/s.

Figure 46 is the polarization curve of biological-cathode in oxygen-saturated acetic acid buffer solution (pH 5).Bilirubin oxidase (myrothecium verrucaria) is adsorbed onto common (plain) carbon cloth (triangle) to be gone up or activates on the cloth (circle).Sweep speed: 2mV/s.

Figure 47 is the schematic diagram that coats the particle of electron mediator (electron mediator), enzyme (having two subunits) and enzyme immobilization material.

Figure 48 is that spray gun is spray dried into the schematic diagram that Merlon shields with mixture.

Figure 49 is linear scan voltammogram (linear sweep voltammogram), shows that the activity as the embodiment 17 described alcohol dehydrogenase of sealing of making carbon composite electrode keeps.

Figure 50 is the diastatic relative activity figure that is fixed on the consumption starch on the different enzyme immobilization materials.

Figure 51 is the diastatic relative activity figure that is fixed on the consumption maltose on the different enzyme immobilization materials.

Figure 52 is the schematic diagram of air self-respiration type biological-cathode electrode.Electrode comprises conductibility individual layer 527.

Figure 53 is the schematic diagram of biological anode electrode.Electrode comprises conductibility individual layer 537.

Figure 54 is that comparison GDL of the present invention and GDL (Elat) are used for polarization and the power curve based on the fuel cell of biological-cathode electrode.The anode of these two batteries is the platinum black catalyst on Elat, makes fuel with hydrogen.The cathod catalyst of these two batteries is immobilization laccases, makes oxidant with oxygen.

Figure 55 is the anode GDL that makes more as described herein and the polarization and the power curve that are purchased the direct methanol fuel cell of anode GDL (Elat).The anode of these two batteries is the platinum ruthenium black catalyst on corresponding GDL material, and 5.0% methyl alcohol is made fuel.The cathod catalyst of these two batteries is the platinum black catalyst on Elat GDL, and oxygen is made oxidant.

Figure 56 is the interactional schematic diagram of enzyme/nano wire and carbon particle.

Figure 57 is the schematic diagram because of the predetermined electron path that forms interact with each other (neural net shape structure) between nano wire.

Figure 58 is the half-cell test of (non-mediated) biological carbon paste electrode in the 1M phosphate buffer of no amboceptor mediation.

Figure 59 is amboceptor mediation (mediated) anodic half-cell at pH is test in 7.2 the 1M phosphate buffer.

Figure 60 is relative platinum black anode H ₂/ O ₂Fuel cell just has the carbon filler and does not have laccase negative electrode that the situation of carbon filler carries out relatively in room temperature.

Figure 61 has described at room temperature at H ₂/ O ₂Performance when changing the carbon content in laccase/platinum black in the PEM fuel cell.

Figure 62 has described laccase under the room temperature/platinum black H ₂O ₂The performance of various dissimilar carbon in the PEM fuel cell.

Figure 63 has described laccase under the room temperature/platinum black H ₂O ₂The performance of different enzymes/TBAB carrying capacity in the PEM fuel cell.

Figure 64 has described laccase under the room temperature/platinum black H ₂O ₂The performance of different electrode holders in the PEM fuel cell.

Figure 65 has described laccase/platinum black H chitosan-immobilized under the room temperature ₂O ₂Biological-cathode PEM fuel cell performance.

The electrode that Figure 66 is based on the Nafion of TBAB modification press-fits the schematic diagram of (press package).

Figure 67 A and 67B are the steps 1 and 2 that the chitin immobilized enzyme electrode press-fits.

Appropriate section in the corresponding mark indication accompanying drawing.

Detailed Description Of The Invention

Referring now to accompanying drawing, Fig. 1 shows an embodiment of fuel cell system of the present invention, and its integral body is represented with label 1.Equipment 1 can produce and can be used for satisfying the electric current that the power consumption be labeled as 5 load requires.As an example, fuel cell system 1 can be used for providing electric power for little hand-hold electronic equipments.The fuel that is used for operational outfit 1 is provided by suitable source 7, and in use be consumed or after using slave unit 1 be discharged into suitable waste material destination (for example receiver) 9.

Fig. 2 shows the fuel cell system 1 of Fig. 1, and Fig. 3 is the separately view of each parts of display device 1.Generally speaking, this equipment comprises the fuel chamber 15 of confluxing, and this chamber of confluxing has front side 21, rear side 23, be used for fuel fluid flows into the import 29 in the chamber of confluxing and be used for fuel fluid flows into waste material destination 9 from the chamber of confluxing outlet 33 from fuels sources 7.The chamber 15 of confluxing comprises one or more 41 fuel storage ponds that respectively are labeled as.This fuel cell system also comprise generally be labeled as respectively 45 and 47 be used for fuel reaction with fuel storage pond 41 facing to the front side anode assemblies of the front side 21 in the chamber 15 of confluxing and rear side 23 and rear side anode assemblies, and generally be labeled as front side cathode assembly and rear side cathode assembly that 51 and 53 neighbour front side anode assemblies 45 and rear side anode assemblies 47 respectively.The electronic controller that is generally referenced as 71 (Figure 14) is set for the operation of this equipment of control, and this will be described below.In also can being included in, the additional power circuit that is generally referenced as 81 (Figure 15) is used for providing as required the normal output of power with the postcombustion battery.Above-mentioned these parts are contained in and are generally referenced as in 91 the housing, make this fuel cell system to hold certainly and are relatively little compact unit.Describe above-mentioned each parts below in detail.

The fuel chamber of confluxing

More specifically, shown in Fig. 4-6, shown in the fuel of the embodiment chamber 15 of confluxing comprise the body (body) or the block (block) 101 of suitable insulative materials (as acrylic compounds), it has top 105, bottom 107, opposite end 109, front 111 and the back side 115.This block 101 can form (for example molded, mechanism etc.) for any suitable shape (as rectangle or other) and preferably be made of single integral type member.As selection, it can constitute to form monolithic construction by a plurality of members that separate are fixed to one another.Front 111 and the chamber in the back side 115 (also being labeled as 41) by this block define fuel storage pond 41.In the illustrated embodiment, the front is provided with 4 this chambeies 41, and the back side is provided with 4 this chambeies, forms 8 fuel storage ponds so add up to 8 chambeies.In other embodiments, according to the number and the design of biological fuel cell required in the assembly, the chamber 15 of confluxing can comprise 1 to reaching 40 or more a plurality of chambeies.The entrance point 121 of fuel by each fuel storage 41 tops, pond enters corresponding storage pond and the port of export 125 by same Chu Chi top leaves this storage pond.Illustrate as Fig. 5 the best, the top in each fuel storage pond 41 becomes the step shape, thus entrance point 121 on the entrance point surface 127 and the port of export 125 be positioned on the port of export surface 129 of certain distance above the surface 127.As a result, the height of the port of export 125 in each storage pond all stores up the height height of the entrance point 121 in pond than each.Vertical interval between these two surfaces 127 and 129 forms chamber or the space 131 that height (as 0.10 inch) is fit to hold any air bubble (may be trapped within the fuel chambers).This set makes the danger that the air bubble of can complete filling fuel storage pond 41 and not holding back and anode assemblies 45,47 contact with cathode assembly 51,53, and this will be described below.

The conflux fuel storage pond 41 in chamber 15 connects by runner or pipeline 135, and runner or pipeline 135 can for example comprise 1/8 in the hole that is assembled to the chamber of confluxing " * 1/8 " polypropylene fluking type elbow (barbed elbow).In the layout of an expectation, fuel fluid (series connection) successively flows to the next fuel storage pond 41 of this series from fuel storage pond 41 of this series.For example, shown in Figure 4 and 5, fuel fluid flows into first fuel storage pond 41 of the front side, chamber 21 of confluxing of adjacent chamber one end 109 that confluxes from the import 29 in the chamber 15 of confluxing, then flow into second, third and the 4th storage pond 41 (in Figure 4 and 5 from left to right) of front side successively, then order flows into 4 storage ponds 41 of the chamber rear side 23 that confluxes, and leaves the chamber 15 of confluxing by exporting 33 subsequently.Other fuel flow path between the different fuel storage pond 41 also is fine.Can between the entrance point 121 in the next one storage pond 41 of the port of export 125 and this series in a storage pond 41, valve (as check-valves 141) be set, to prevent that flowing to the chamber of confluxing by a plurality of fuel storages pond 41 at fuel successively from the import 29 in the chamber 15 of confluxing exports 33 o'clock fuel and backflow from a storage pond 41 that another stores up pond.The use of check-valves (as the Poweraire valve) can interrupt any ion communication between these batteries, thereby prevents short circuit.Arranged in series check-valves 141 exists fuel input and output to fill all storage ponds 41 like this.

In the embodiment illustrated, the conflux layout in fuel storage pond 41 in chamber 15 makes and confluxes fuel storage pond on the chamber rear side 23 directly facing to the storage pond of the front side, chamber 21 of confluxing.But other layout also can.And the chamber of confluxing can have more than one import 29 and more than one outlet 33.For example in interchangeable design, an import and outlet are set being used to the fuel storage pond 41 of front side, chamber 21 of confluxing, and different imports and the outlet fuel storage pond 41 with the chamber rear side 23 that is used to conflux is set.

Anode assemblies

With reference to Fig. 7-9, front side anode assemblies 45 comprises framework 151, and this framework 151 comprises the preceding frame member 151A and the back of the body frame member 151B of a pair of coupling in one embodiment.Each has a plurality of openings 155 therein frame member 151A and 151B, and configuration and layout that these openings are constructed and arranged to the fuel storage pond 41 in the front 111 with the chamber 15 of confluxing are complementary.These frame members can be thick about 0.018 inch vinyl sheet.Other material also can use.And, framework 151 can have with shown in different configurations.

Front side anode assemblies 45 also comprises a plurality of anodes, and each anode is generally referenced as 157, remains in the framework opening 155 anode of each framework opening by framework 151.

In certain embodiments, each anode 157 comprises the collector body 161 that is fixed in the corresponding framework opening 155.In one embodiment, collector body 161 comprises thick about 0.018 inch nickel wire net sheet.Suitable electrical lead 169 (for example No. 28 conductors) is fixed on the collector body 161.Each anode 157 also is included in the gas diffusion layers 165 that is positioned at collector body 161 back sides between collector body 161 and the back of the body frame member 151B.For example, this diffusion layer 165 can be the carbon paste structure of describing at " gas diffusion layers " chapters and sections as following.Then can be by being applied to the enzyme in the cushioning liquid and the Nafion of TBuA modification

Ionomeric curtain coating (cast) mixture adds the catalyst layer (not shown) to gas diffusion layers 165.Before being attached to catalyst layer on the anode assemblies, be dried.In preferred embodiments, used enzyme is a PQQ dependent form alcohol dehydrogenase (PQQ-ADH) in the catalyst layer.

The size of each anode 157 is slightly larger than corresponding framework opening 155, and the respective side that makes the side edge portion (side edge margin) of anode extend beyond the framework opening is used for the part of overlapping frame member 151A and 151B.Anode 157 by the binding agent anterior layer 175 between the collector body 161 that former frame member 151A and anode 157 are set and behind the binding agent between the diffusion layer 165 of after-frame member 151B and anode 157 layer 181 be fixed on the frame member.Binding agent anterior layer 175 and back layer 181 comprise for example thick about 0.005 inch polyurethane hot melt adhensive membrane.Binding agent anterior layer 175 and back layer 181 are configured to have respectively the corresponding size and dimension of size and dimension with preceding frame member 151A and after-frame member 151B.When applying to this assembly (as passing through hot pressing) when hot, this binding agent fusing to be being fixed on these two frame member 151A and 151B, collector body 161 and diffusion layer 165 on the fixing toward each other position, thereby forms the front side anode construction of integral body.

In other embodiment with the biological anode catalyst carrier of self-bearing type shown in Figure 53, each anode 157 comprises the collector body 535 that is embedded in the anode.In one embodiment, this collector body has near-end and far-end, and axle extends along the longitudinal.The conductibility individual layer 537 that contacts with collector body 535 from the near-end of collector body to the coaxial extension of far-end.Suitable electrical lead 169 is attached on the collector body 535.In other embodiments, the biological anode catalyst carrier of self-bearing type does not comprise embedding collector body 535.Conductibility individual layer 537 is mixtures of the first electrical conductivity material, the second electrical conductivity material and adhesive, and can be by following " the biological anode catalyst carrier preparation of self-bearing type " the described manufacturing of chapters and sections.Then apply the catalyst layer (not shown) to the surface of individual layer 537 by the described method of following " conducting polymer base nano-wire " chapters and sections.As selection, can in the mixture that is used to form individual layer 537, add the catalyst particle (as described in following " enzyme of sealing is sneaked in the carbon paste " and " biocatalyst ink formulations " chapters and sections) that contains enzyme, perhaps this catalyst particle can be included in and then be administered in lip-deep paste of catalyst carrier or the ink (ink).

The size of each anode 157 is slightly larger than corresponding framework opening 155, and the respective side that makes the side edge portion of anode extend beyond the framework opening is used for the part of overlapping frame member 151A and 151B.Anode 157 by the binding agent anterior layer 175 between the individual layer 537 that former frame member 151A and anode 157 are set and behind the binding agent between the individual layer 537 of after-frame member 151B and anode 157 layer 181 be fixed on the frame member.Binding agent anterior layer 175 and back layer 181 comprise for example thick about 0.005 inch polyurethane hot melt adhensive membrane.Binding agent anterior layer 175 and back layer 181 are configured to have respectively the corresponding size and dimension of size and dimension with preceding frame member 151A and after-frame member 151B.When applying to this assembly (as passing through hot pressing) when hot, this binding agent fusing to be being fixed on these two frame member 151A and 151B and individual layer 537 toward each other on the fixed position, thereby forms the front side anode construction of integral body.

Front side anode assemblies 45 is fixed on the front 111 in the chamber 15 of confluxing, make framework opening 155 and anode 157 and corresponding front side fuel store up pond 41 general alignment (align), this layout makes the fuel fluid that respectively stores up in the pond 41 be adapted to contact with the respective anode 157 of front side anode assemblies 45.What may expect is that the overall dimension of each framework opening 155 and shape are near the size and dimension in corresponding fuel storage pond 45, so that the whole basically zone of anode 157 all is exposed to the fuel fluid from this fuel storage pond.Anode assemblies 45 is fixed on the chamber 15 of confluxing by the mode of the adhesive layer between the front 111 in the back side of framework 151 and the chamber 15 of confluxing (the hot melt polyurethane binder layers of for example thick 0.005 inch fusing under 128 ℃) with sealing.Provide sealing that each front side fuel storage pond 41 and corresponding anode 157 thereof are stored up pond with adjacent each front side fuel in this way and corresponding anode 157 separates.This sealing can form otherwise, for example uses one or more sealing gaskets.

Rear side anode assemblies 47 is preferably by making up with front side anode assemblies 45 essentially identical modes, and the corresponding corresponding labelled notation of parts.Rear side anode assemblies 47 is against the back side 115 in the chamber 15 of confluxing, and fuel storage pond 41 registrations (registration) of framework opening 155 and the anode 157 and the chamber rear side 23 that confluxes.Rear side anode assemblies 47 is fixed on respect on the appropriate location in the chamber 15 of confluxing and by the above back side 115 sealed engagement at front side anode assemblies 45 same way as of describing and the chamber of confluxing.Above-mentioned anode assemblies generally comprises nickel collector body, carbon based gas diffusion layer, PQQ dependent form alcohol dehydrogenase and as the PQQ of electron mediator.But those skilled in the art will understand that these parts can replace with following various substitutions of elements.

The direct electron at biological anode or biological-cathode place shifts (DET)

In another aspect of this invention, the system that utilizes direct electron to shift has been described.In the system that shifts based on direct electron, electron transfer is relevant to the catalyzed conversion of product with substrate, or takes place in this conversion process.Enzyme serves as eelctro-catalyst (electrocatalyst), is grafted on the electron transfer between nano wire on first conductive of material such as the carbon particle (pruning (tailor) with suitable surface functionalities) promotion enzyme, electronic conductor (electrode) and the substrate molecule.In certain embodiments, this process does not relate to electron mediator.When nano wire increases conductibility but do not experience reversible electron transfer, observe direct electron and shift.The DET system provides more effective electron transfer rate usually, and this is because operation and enzyme less touch disturbance reponse near the potential range of the redox potential of enzyme.Usually, the integration between biomolecule and the electrode surface (integration)/communication degree is high more, and the electron transfer rate is big more, and the power output that is provided is many more.

The performance of the high power output biofuel cell system that shifts based on direct electron depends on the many aspects of enzyme immobilization process.For example, the character of character of electronic conductor/carrier (for example electricity of the size of the chemical property of surface functionalities, nano wire and nano wire and ionic conduction character) and biomolecule and stability (immobilization technology that for example the is used for stabilized enzyme) performance that determined biological fuel cell.

In other embodiments, being grafted on nano wire on first conductive of material (pruning with suitable surface functionalities) has and is grafted on the lip-deep electron mediator of this first conductive of material.But these electron mediators can be in the inverted electron transfer of male or female experience.In multiple embodiments, the particle of nano wire grafting can be used as the nuclear core of the particle of following immobilised enzymes coating.

Preparation is described below is used in the conventional biological anode in the biological fuel cell and the process of biological-cathode electronic conductor (electrode).

The biological anode catalyst carrier of self-bearing type

Biological anode catalyst carrier as herein described is similar to the enzyme coated side of biological-cathode, and the microcellular structure of high surface is used for fixing of enzyme and mixing of electrolyte/substrate herein.Illustrating in Figure 53 of biological anode catalyst carrier.With with the similar mode of biological-cathode electrode, the first electrical conductivity material (for example carbon black) can be modified with electron mediator or dopant (for example being grafted on its surface by the conducting polymer that will mix), thereby helps electron transfer and electricity and gas ions to lead.

The biological anode catalyst carrier preparation of self-bearing type

The catalyst support layer that is used for anode electrode is made by material with carbon element, adhesive, pore former, solvent and optional catalyst such as the mixture of enzyme.By changing the ratio of these components, the character that can change this carrier structure is to provide application-specific desired performance.For example, the character that can change comprises: (1) electrical conductivity, (2) hydrophobicity/hydrophily, (3) surface area and (4) surface texture.The mixture that forms carrier structure is at first made paste, then be pressed into framework (MDS nylon 6/6 framework of filling for example, 0.020 " thick); for the electrode that does not have embedding porous metals silk screen (expanded metal) collector body, this framework adhered to have bilateral pressure bonding agent film (pressure adhensivefilm) on the interim carrier of polytetrafluoroethylene (PTFE) coated fibres glass (for example 0.020 inch thick) of (for example 0.002 inch thick).

For electrode with embedding collector body carrier, with the framework of two band pressure bonding agent films (MDS nylon 6/6 framework of filling for example, 0.010 " thick) be fixed on each one side of a slice (for example 0.005 inch thick) porous metals silk screen, and do not need the interim carrier of PTFE coated fibres glass.Remove on the framework and the unnecessary thickener at top with instrument (for example artist with cutter or spatula).When thickener is also wet, with this electrode structure by 2 block plates (6 " * 6 " * 0.065 "), 2 blocks of PTFE coated fibres sheet glass (4 " * 4 " * 0.005 ") and press-fiting in the part of constituting of 2 paper handkerchiefs (folding) to match with 2000 pounds of pressures 15 seconds.From press-fit part, take out electrode, and remove any unnecessary thickener on the framework with cutter and paper handkerchief with the artist.Then depend on desired results and preparation, whole assembly is no more than 20 minutes at the temperature sintering that is no more than 200 ℃.

After the sintering, from framework and interim carrier, shift out electrode, and with its 2 aluminium blocks (12 " * 12 " * 1 ") between cooling until solidifying, usually a few minutes only, be used for biological fuel cell afterwards.2 that use interval 1cm with milliohm table measurement conductibility.With respect to commercial gas diffusion layer (GDL) material of measuring with the same manner, the report measured value; This measured value is not the literature value of the record under the condition condition of unknown of test and instrument.For every kind of electrode type, because the variation between limited manual manufacture round, these values also provide as approximation.

If catalyst is not included in wherein when the conductibility individual layer forms, then catalyst layer can be applied on the catalyst carrier as described in following " enzyme of sealing is sneaked in the carbon paste " chapters and sections.As selection, catalyst particle (as described in following " enzyme of sealing is sneaked in the carbon paste " and " biocatalyst ink formulations " chapters and sections) adding that contains enzyme can be used to form in the mixture of individual layer 537, maybe it can be included in paste or the ink and then be applied to the catalyst carrier surface.

The material and the component that are used for the preparation of electrode and catalyst layer

Can utilize the various components of accepting to make up electrode described herein and catalyst layer.Following table has been listed various carbon blacks, pore former, adhesive material and conductibility carbon fiber and supplier thereof in detail.These materials can be used to prepare electronic conductor as herein described (electrode).

Table 1. as among the bioelectrode GDL the carbon black of the first electrical conductivity material

The carbon black trade name	Supplier
		Vulcan XC-72	Cabot
Polypyrrole, doping, compound with carbon black	Sigma Aldrich
		PrintexXE
2	Degussa AG
		KetjenBlack EC-600JD	Akzo Nobel Chemicals
KetjenBlack EC300J	Akzo Nobel Chemicals
		Raven 5000 Ultra II	Columbian Chemical Company
Raven 7000	Columbian Chemical Company
		Monarch 880	Cabot
Black Pearls 460	Cabot
		Black Pearls 1300	Cabot
ChemSorb 1505 G5	Molecular C*Chem
		Monarch
1000	Cabot
		Timrex HSAG 300CAT	Timcal-Stratmin
Black Pearls
	2000	Cabot
XPB F 138			Degussa Engineered Carbons，LP.
	Printex XE 2-B	Degussa AG
ChemSorb 1202 G5			Molecular C*Chem
	ChemSorb 1000-60 G5	Molecular C*Chem
Conductex SC Ultra			Columbian Chemical Company
	Raven 5000 Ultra III	Columbian Chemical Company
Monarch 1400			Cabot
	Black Pearls 570	Cabot
The carbon nano powder			Sigma Aldrich
	The carbon nano-fiber powder	Nanostructured&Amorphous Materials Inc.
The vitreous carbon spherical powder			Alfa Aesar
	Multi-walled carbon nano-tubes COOH is functionalized	Cheap Tubes Inc.
Multi-walled carbon nano-tubes OH is functionalized			Cheap Tubes Inc.
	Multi-walled carbon nano-tubes	Cheap Tubes Inc.
The carbon awl			n-TEC
	Carbon nano-tube, single wall	Sigma Aldrich
Inner self-control modified carbon black			Akcrmin，Inc.

Table 2. is used in the pore former among the bioelectrode GDL

Pore former (pore forming agent)	Supplier
		Ammonium carbonate	Sigma
Carbonic hydroammonium	Sigma
		Lithium carbonate	Sigma
Ammonium oxalate	Sigma
		Ethylene glycol	Sigma
Polyethylene glycol	Sigma
		Glycerine	Sigma

Table 3. is used in the adhesive material among the bioelectrode GDL

Adhesive	Supplier
		Polyvinylidene fluoride (being also referred to as Kynar)	Sigma
Polytetrafluoroethylene (being also referred to as Teflon)	Sigma

Table 4. is as the conductibility carbon fiber of the second electrical conductivity material among the bioelectrode GDL

Graphite fibre	Supplier
		DKDX	Cytec Carbon Fiber
XN-100	Nippon Graphite Fiber

Can select surface area and electrical conductivity and the electron transfer of carbon black materials to influence electronic conductor.Surface area (by form the hole in structure, increasing its surface area thus) and substrate that pore former can be used for increasing electronic conductor (electrode) are diffused into the enzyme ability of (if existence).The hydrophobic/hydrophilic character that adhesive material can be used for advantageously changing electronic conductor (electrode) increases mass transfer and influences the structural integrity of this structure.The conductibility carbon fiber can be selected with the conductibility that influences electronic conductor (electrode) and influence its structural integrity.

Alternative biological anode material

According to the biological anode of certain embodiments of the invention comprise collector body (as collector body 161), gas diffusion layers (or electronic conductor) (as gas diffusion layers 165), randomly electron mediator, randomly be used for the eelctro-catalyst of electron mediator and be fixed on enzyme in the enzyme immobilization material.In certain embodiments, these parts are adjacent one another are, mean that they pass through suitable method and physically or chemically connect.

1. collector body

Collector body (as 161) is conduction electron and the material that provides grid to support for gas diffusion layers and catalyst layer.Therefore, provide these materials with function all can be used for collector body.For various biological anode embodiments, preferred nickel or nickel-containing material (being Inconel).When PQQ dependent form alcohol dehydrogenase during as biological anode enzyme, the nickel ion that slowly dissolves generation by collector body serves as the promoter (promoter) of enzymatic reaction.

2. gas diffusion layers (GDL or electronic conductor)

Gas diffusion layers (or electronic conductor) (as 165) is the material of conduction electron.Gas diffusion layers can be the organic or inorganic material, as long as it can pass through this conduct electronics.Gas diffusion layers can be carbon-based material, stainless steel, stainless (steel) wire, metallic conductor, semiconductor, metal oxide, modification conductor or its combination.In preferred embodiments, gas diffusion layers is a carbon-based material.

Specially suitable gas diffusion layers is a carbon-based material.Exemplary carbon-based material be carbon cloth, carbon paper, carbon filament reticulated printing electrode, carbon paper (Toray), carbon paper (ELAT), carbon black (Vulcan XC-72, Etek), carbon black, carbon dust, carbon fiber, Single Walled Carbon Nanotube, double-walled carbon nano-tube, multi-walled carbon nano-tubes, carbon nano pipe array, coated with CVD (chemical vapor deposition) diamond conductor, vitreous carbon, mesoporous carbon and combination thereof.In addition, other exemplary carbon-based material graphite worm (worm) that is graphite, uncompressed, tear the purifying flake graphite (Superior of layer open

Graphite), high-performance (high performance) graphite and carbon dust (Formula BT ^TM, Superior

Graphite), high order pyrolytic graphite, pyrolytic graphite, polycrystalline graphite and combination thereof.Preferred gas diffusion layers is the carbon cloth sheet.

In other embodiments, gas diffusion layers can be made by metallic conductor.Suitable electronic conductor can be made by gold, platinum, iron, nickel, copper, silver, stainless steel, mercury, tungsten, other metal and combination thereof that is applicable to the structure electrode.In addition, when gas diffusion layers was metallic conductor, it can be made of the nano particle that makes from cobalt, carbon and other suitable metal.Other metal electron conductor can be silver-plated nickel wire reticulated printing electrode.

In addition, gas diffusion layers can be a semiconductor.Suitable semi-conducting material comprises the silicon and the germanium of other element that can mix.This semiconductor can Doping Phosphorus, boron, gallium, arsenic, indium or antimony or its combination.

In addition, gas diffusion layers can be metal oxide, metal sulfide, main group compound (being transistion metal compound), the material and the combination thereof of modifying with electronic conductor.Exemplary this gas diffusion layers is mesoporous silica and the combination thereof that collosol and gel, ruthenium carbon aerogels, conductive of material such as the carbon of aeroge, conductive of material such as the carbon modification of nano-pore titanium oxide, tin oxide coated glass, cerium oxide particles, molybdenum sulfide, boron nitride nano-tube, conductive of material such as carbon modification is modified.In some preferred embodiment, gas diffusion layers comprises carbon black, mesoporous carbon, epoxy resin (epoxy) and polytetrafluoroethylene.

In various preferred embodiments, gas diffusion layers comprises the carbon (C*Chem) of the steam activation of dried component: Monarch 1400 carbon blacks (Cabot) and Chemsorb 1505G5 porous dipping, and wet component: be dissolved in the acetone Quick Set 2-part epoxy resin (The Original SuperGlue Corp.) and in water 60% polytetrafluoroethylene (PTFE) dispersion (Sigma).To do component and grind in the food grinder, and mix wet component with ultrasonic homogenizer, then merging is done wet component and with putty knife it is mixed, and has the denseness of toothpaste until mixture.

3. electron mediator

Biological anode electron mediator plays the effect of accepting or supplying with electronics, is varied to the reduction form from oxidised form easily.Electron mediator is the compound that can be spread to and/or mix in the enzyme immobilization material.The diffusion coefficient maximization of preferred electron amboceptor.

Exemplary electron mediator is nicotinamide adenine dinucleotide (NAD ⁺), flavin adenine dinucleotide (FAD), nicotinamide-adenine dinucleotide phosphate (NADP), PQQ (PQQ), every kind equivalent and combination thereof.Other exemplary electron mediator is phenazine methosulfate, dichloropheno-lindophenol, short chain ubiquinone, high-potassium ferricyanide, protein, metalloprotein, stellacyanin and combination thereof.

When electron mediator can not experience redox reaction separately on electronic conductor, biological anode comprised the eelctro-catalyst that is used for electron mediator, and it promotes that electronics discharges at the electronic conductor place.As selection, the reversible redox with normal reduction potential 0.0V ± 0.5V is to being used as electron mediator.On the other hand, can use the electron mediator that reversible electrochemical is provided on the electronic conductor surface.This electron mediator can with organelle in contained and depend on the native enzyme combination of this electron mediator.

4. the eelctro-catalyst that is used for electron mediator

Usually, eelctro-catalyst is the material that helps the release of electronic conductor place electronics.On the other hand, eelctro-catalyst improves the reduction or the oxidation kinetics of electron mediator, makes the reduction of electron mediator or oxidation to carry out at lower normal reduction potential.Eelctro-catalyst can be produced electronics by reversible oxidation at biological anode, and therefore produces.When the eelctro-catalyst neighbour electronic conductor, eelctro-catalyst and electronic conductor were electrically connected to each other, but physical connection each other.In one embodiment, electronic conductor is a part that is used for the eelctro-catalyst of electron mediator, is connected with it or adjacent.

Usually, eelctro-catalyst can be azine, conducting polymer or electroactive polymer.The exemplary electrical catalyst is the ligand transition metal, 1 of methylene green, methylenum careuleum, luminol, nitryl fluorenone derivative, azines, phenanthroline dione osmium, youngster's naphthol-side group (pendant) terpyridyl, toluene blue, cresyl blue, Nile blue, dimethyl diaminophenazine chloride, azophenlyene derivative, tionin, reddish black A, reddish black B, blutene, acetophenone, metal phthalocyanine, Nile blue A, modification, 10-phenanthroline-5,6-diketone, 1,10-phenanthroline-5, the 6-glycol, [Re (phen-diketone) (CO) ₃Cl], [Re (phen-diketone) ₃] (PF ₆) ₂Poly-(metal phthalocyanine), poly-(thionine), the benzoquinones class, the diimine class, the diaminobenzene class, the diamino-pyridine class, phenthazine phenoxazine, toluidine blue, brilliant cresyl blue, 3, the 4-4-dihydroxy benzaldehyde, poly-(acrylic acid), poly-(reddish black I), poly-(Nile blue A), poly-(methylene green), poly-(methylenum careuleum), polyaniline, polypyridine, polypyrrole, polythiophene, poly-(thieno [3,4-b] thiophene), poly-(3-hexyl thiophene), poly-(3,4-ethylidene dioxy pyrroles), poly-(isothianaphthene), poly-(3,4-ethylidene dioxy thiophene), poly-(difluoro acetylene), poly-(4-dicyano methylene-4H-ring penta [2,1-b; 3,4-b '] two thiophene), poly-(3-(4-fluorophenyl) thiophene), poly-(dimethyl diaminophenazine chloride), albumen, metalloprotein, stellacyanin and combination thereof.In a preferred embodiment, the eelctro-catalyst that is used for electron mediator is poly-(methylene green).

5. enzyme

The enzymatic fuel fluid is in biological anodic oxidation.Usually, can use the native enzyme of native enzyme, synthetic enzyme, artificial enzyme and modification.In addition, can use by nature evolution or orthogenesis and the engineering enzyme of through engineering approaches.On the other hand, can use the organic or inorganic molecule of imitation enzymatic property in embodiments of the invention.

Particularly, the exemplary enzyme that is used in biological anode is an oxidoreducing enzyme.In certain embodiments, oxidoreducing enzyme works to the CH-OH group or the CH-NH group of fuel (alcohol, ammoniate, carbohydrate, aldehyde, ketone, hydrocarbon, aliphatic acid etc.).

In some embodiments, enzyme is a dehydrogenase.Exemplary enzyme comprises alcohol dehydrogenase, aldehyde dehydrogenase, hydrogenlyase, formaldehyde dehydrogenase, glucose dehydrogenase, glucose oxidase, lactic dehydrogenase, lactose dehydrogenase, pyruvic dehydrogenase or lipoxygenase.Preferably, this enzyme is alcohol dehydrogenase (ADH).

The energy density maximization that obtains from fuel fluid in order to make in being used in biological fuel cell may be expected the fuel complete oxidation, this means bond fission as much as possible in the molecule.The enzyme that selection is used in the biological anode will be suitable for making the energy density of special fuel fluid to maximize.For example, hydrogen fuel requires a kind of enzyme, and methyl alcohol requires three kinds.When ethanol is used as fuel, can use the tricarboxylic acid cycle enzyme.For example, can in biological anode, use aconitase, fumarase, malic dehydrogenase, succinate dehydrogenase, Succinyl-CoA synthetase, isocitric dehydrogenase, ketoglutaric dehydrogenase, citrate synthase and combination thereof.

In preferred embodiments, enzyme is a PQQ dependent form alcohol dehydrogenase.PQQ is the coenzyme of PQQ dependent form ADH, and keeps static to combine with PQQ dependent form ADH, so this enzyme can remain in the enzyme immobilization material, thereby causes being used for the life-span and the activity of the increase of biological fuel cell.PQQ dependent form alcohol dehydrogenase can extract from gluconobacter sp.When extracting PQQ dependent form ADH, it can be two kinds of forms: (1) PQQ static is attached to that PQQ dependent form ADH goes up or (2) PQQ does not have static and is attached on the PQQ dependent form ADH.Do not have static to be attached to second kind of form on the PQQ dependent form ADH for PQQ wherein, PQQ is added on the ADH when biological anode assembling.In preferred embodiments, extract PQQ dependent form ADH in conjunction with the form of PQQ from Gluconobacter with static.

6. enzyme immobilization material

Enzyme or organelle immobilization material can be used for fixing and stabilized enzyme or organelle.Should be applicable to enzyme immobilization material and organelle immobilization material about the discussion of immobilization material.In various embodiments, the enzyme immobilization material is permeable for the compound littler than enzyme, and this makes the reaction of expectation can be immobilized enzymatic.

Usually, can use the multiple reaction of enzymatic, and enzyme can be fixed in the enzyme immobilization material of not only fixing but also stabilized enzyme.As for the stabilisation of enzyme, the enzyme immobilization material can provide chemistry and mechanical barrier to stop or the obstruction enzyme denaturation.For this reason, the enzyme immobilization material is restriction enzyme physically, prevents that this enzymolysis is folding.The process that enzyme is launched by folding three-dimensional structure is a kind of mechanism of enzyme denaturation.Usually, the resolvase in the solution lost its catalytic activity in several hours to several days, and the present invention is by the enzyme of immobilization and stabilisation can be with its catalytic activity maintenance at least about 7 days to about 730 days or longer rightly.The maintenance of catalytic activity be defined as enzyme in the continuous catalysis chemical conversion, keep its initial activity at least about 75% fate.Enzymatic activity can be measured in the following way: chemiluminescence, electrochemistry, UV-Vis, radiochemistry or fluorescent test (wherein measuring the intensity of this character at initial time).Usually, utilize fluorescent test to measure enzymatic activity.In other words, stabilized enzyme of the present invention in the continuous catalysis chemical conversion at least about kept in 7 days to about 730 days its initial activity at least about 75%.In some embodiments, the enzyme of this immobilization and stabilisation is at least about 30,45,60,75,90,105,120,150,180,210,240,270,300,330,365,400,450,500,550,600,650,700,730 days or longer its initial catalytic activity of maintenance at least about 75%, preferably at least about 5,10,15,20,25,30,45,60,75,90,105,120,150,180,210,240,270,300,330,365,400,450,500,550,600,650,700,730 days or longer its initial catalytic activity of maintenance at least about 80%, 85%, 90%, 95% or more.For the present invention, if enzyme enliven catalytic chemistry transform can continue simultaneously at least about 30 days to about 730 days or longer its initial catalytic activity of maintenance at least about 75%, then this enzyme is through " stabilisation " (can be used in the purposes of the long-term relatively catalytic activity of the biological anode of biological fuel cell or biological-cathode or other needs).

And, in various embodiments, in being fixed on the enzyme immobilization material after, enzyme keep its activity before immobilization at least about 0.65,0.7,0.75,0.8,0.85,0.9,0.95 or more.

Immobilized enzyme is the enzyme that physically is limited in certain zone of enzyme immobilization material and keeps its catalytic activity simultaneously.There are various enzyme immobilization methods, comprise carrier-combination, crosslinked and hold back.Carrier-combination is that enzyme is attached on the water-fast carrier.It is crosslinked that to be that enzyme passes through between the molecule of difunctionality or multifunctional reagent crosslinked.Hold back is that enzyme is mixed in the grid of semipermeable materials.The concrete grammar of enzyme immobilization is not crucial important, as long as the enzyme immobilization material can (1) immobilized enzyme and (2) stabilized enzyme.In various embodiments, but the also little compound of this enzyme of tranmittance of enzyme immobilization material.

Immobilized organelle is the organelle that physically is confined in certain zone of enzyme immobilization material and keeps its catalytic activity simultaneously.

For with regard to the permeability of all cpds littler than enzyme or organelle, immobilization material allows substrate, fuel fluid or oxidant compound to move through it, makes compound can touch enzyme or organelle thus with regard to immobilization material.Immobilization material can be so that the mode that it comprises internal holes, passage, hole or its combination prepares, thereby allows compound to move in whole immobilization material, but enzyme or organelle are limited in basic identical space in the immobilization material.This restriction makes enzyme or organelle can keep its catalytic activity.In various embodiment preferred, enzyme or organelle be limited in enzyme or organelle have in the space of substantially the same size and dimension, this enzyme or organelle keep its whole basically catalytic activity therein.Described hole, passage, hole have the physical size that satisfies above requirement, and this physical size depends on the size and dimension for the treatment of immobilized certain enzyme or organelle.

In some embodiments, enzyme or organelle are trapped within the hole of immobilization material and with the nano wire that is grafted to first conductive of material such as carbon black particle and contact.In some cases, nano wire comprises polymeric material, oxide, organo metallic material or metal material.These materials were discussed at above " DET " chapters and sections, and " conducting polymer base nano-wire " chapters and sections discussion below.

In one embodiment, enzyme or organelle are preferably placed in the hole of immobilization material, and compound passes in and out immobilization material by transfer passage.The relative size of hole and transfer passage can so that the hole even as big as immobilized enzyme or organelle, but transfer passage is too little and it is passed through for enzyme or organelle.In addition, the diameter of transfer passage is preferably at least about 10nm.In another embodiment, bore dia and transfer passage diameter ratio were at least about 2: 1,2.5: 1,3: 1,3.5: 1,4: 1,4.5: 1,5: 1,5.5: 1,6: 1,6.5: 1,7: 1,7.5: 1,8: 1,8.5: 1,9: 1,9.5: 1,10: 1 or bigger.In another embodiment, the diameter of preferred transfer passage is at least about 2nm, and bore dia and transfer passage diameter ratio were at least about 2: 1,2.5: 1,3: 1,3.5: 1,4: 1,4.5: 1,5: 1,5.5: 1,6: 1,6.5: 1,7: 1,7.5: 1,8: 1,8.5: 1,9: 1,9.5: 1,10: 1 or bigger.

In certain embodiments, immobilization material has micella or reverse micelle structure (invertedmicellar structure).Usually, the molecule that constitutes micella is both sexes, this means that they comprise polar hydrophilic group and nonpolar hydrophobic group.These molecules can be assembled the formation micella, and wherein polar group is on the surface of aggregation, and the hydrophobicity non-polar group completely cuts off in aggregation inside.The polar group of reverse micelle and non-polar group opposite orientation.The amphiphatic molecule that constitutes this aggregation can be arranged in every way, as long as polar group is located adjacent one another and non-polar group is located adjacent one another.And these molecules can form the bilayer that non-polar group faces toward each other and polar group is carried each other.As selection, following bilayer can form, wherein polar group can be in bilayer each other facing to and non-polar group is carried each other.

In a preferred embodiment, the micella immobilization material is perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer of modification) (Nafion of modification of modification

Or the Flemion of modification ) film.The perfluorinated ion-exchange polymer film can be used than ammonium ion (NH4 ⁺) big dewatering cationic modification.This dewatering cationic plays the hole dimension of (1) regulation film and the double action that (2) serve as the chemical buffer of the pH level that helps retaining hole, and the two all makes enzyme stabilization.

About first effect of this dewatering cationic, perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer of modification the) (Nafion that mixes curtain coating generation modification of perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer) and dewatering cationic

Or Flemion ) film, immobilization material is provided, its intermediate pore size depends on the size of dewatering cationic.Therefore, dewatering cationic is big more, and hole dimension is big more.This effect of dewatering cationic make can do hole dimension big by the size that changes dewatering cationic or do little, to be complementary with certain enzyme or organelle.

About second effect of this dewatering cationic, the character of perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer) film can by dewatering cationic and proton exchange as on perfluorinated sulfonic acid-PTFE co-polymer membrane-SO ₃ ^-The gegenion of group (or the anion on the perfluorinated ion-exchange polymer) changes.The variation of this gegenion provides cushioning effect to pH because dewatering cationic than proton right-SO ₃ ^-The affinity in site is much bigger.The sort buffer effect of film makes the pH in hole keep not changing with pH value of solution substantially; On the other hand, the pH in hole can resist the pH value of solution variation.In addition, this film provides mechanical barrier, and this has protected immobilised enzymes or organelle again.

Be perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer) film of preparation modification, first step is with perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer), particularly Nafion

, with the suspension curtain coating formation film of dewatering cationic solution.Then extract dewatering cationic excessive in the membrane and salt thereof, and with film curtain coating again.Again during curtain coating, film comprise with perfluorinated sulfonic acid-PTFE copolymer (or perfluorinated ion-exchange polymer) film-SO ₃ ^-The dewatering cationic of site combination.The salt that removes the dewatering cationic in the film obtains more stable and reproducible film; If they do not remove, then excess salt may be trapped within and cause the space in the hole or in film.

In one embodiment, the Nafion of modification

Film passes through Nafion

The suspension curtain coating of the salt of polymer and dewatering cationic such as bromination quaternary ammonium solution makes.Behind the excessive bromination quaternary ammonium or hydrogen bromide in extracting membrane, with film again curtain coating form desalination membrane.The desalination meeting of film makes quaternary ammonium cation remain resident in sulfonic acid exchange site, but eliminated that excess salt may be trapped within the hole or may the film after balance in cause the complex situations in space.The chemistry of the film after the desalination and physical property measured with voltammetry, ion exchange capacity before enzyme immobilization and fluorescence microscopy characterizes.Exemplary dewatering cationic is the ammonium cation, quaternary ammonium cation, alkyl trimethyl ammonium cation, alkyl triethyl ammonium cation, organic cation phosphonium cation, triphenyl phosphonium, pyridylium, glyoxaline cation, cetyl pyridinium, second ingot (ethidium), viologen (viologen), the methyl viologen, the benzyl viologen, two (triphenylphosphine) imonium (iminium), metal complex, the bipyridyl metal complex, phenanthrolines Base Metal complex compound, [Ru (bipyridine) ₃] ²⁺[Fe (phenanthroline) ₃] ³⁺

In a preferred embodiment, dewatering cationic is the ammonium cation.Especially, dewatering cationic is a quaternary ammonium cation.In another embodiment, quaternary ammonium cation is represented by following formula 1:

R wherein ₁, R ₂, R ₃And R ₄Be the alkyl or the heterocycle of hydrogen, alkyl, replacement, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.In another embodiment, preferred R ₁, R ₂, R ₃And R ₄Be hydrogen, methyl, ethyl, propyl group, butyl, amyl group, hexyl, heptyl, octyl group, nonyl, decyl, undecyl, dodecyl, tridecyl or myristyl, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.In another embodiment, R ₁, R ₂, R ₃And R ₄Identical and be methyl, ethyl, propyl group, butyl, amyl group or hexyl.In another embodiment, preferred R ₁, R ₂, R ₃And R ₄It is butyl.Preferably, quaternary ammonium cation is tetrapropyl ammonium (T3A), four pentyl ammonium (T5A), tetrahexyl ammonium (T6A), four heptyl ammoniums (T7A), trimethyl icosyl ammonium (TMICA), trimethyl octyl-decyl ammonium (TMODA), trimethyl hexyl decyl ammonium (TMHDA), trimethyl myristyl ammonium (TMTDA), trimethyl octyl group ammonium (TMOA), trimethyldodecane base ammonium (TMDDA), trimethyl decyl ammonium (TMDA), trimethyl hexyl ammonium (TMHA), TBuA (TBA), triethyl group hexyl ammonium (TEHA) and combination thereof.

In other various embodiments, exemplary micella or reverse micelle immobilization material are the hydrophobically modified polysaccharide, and these polysaccharide are selected from shitosan, cellulose, chitin, starch, amylose, alginates, glycogen and combination thereof.In various embodiments, micella or reverse micelle immobilization material are polycationic polymers, particularly the shitosan of hydrophobically modified.Shitosan is poly-[β-(1-4)-2-amino-2-deoxidation-D-glucopyranose].Shitosan is usually by deacetylated the making of chitin (poly-[β-(1-4)-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-D-glucopyranose]).Typical commercial shitosan about 85% is deacetylated.These amine groups deacetylated or that dissociate can further be used alkyl, particularly alkyl functionalization.Thereby in various embodiments, the shitosan of micellelike hydrophobically modified is corresponding to formula 2 structures:

Wherein n is an integer; R ₁₀Be the alkyl or the hydrophobic redox mediators of hydrogen, alkyl, replacement independently; R ₁₁Be the alkyl or the hydrophobic redox mediators of hydrogen, alkyl, replacement independently.In certain embodiments of the invention, n makes that the molecular weight of polymer is about 21, and 000-about 500,000, preferred about 90,000 one is about 500,000, more preferably from about 150, and 000-is about 350,000, and more preferably from about 225, the integer of 000-about 275,000.In many embodiments, R ₁₀Be hydrogen or alkyl independently, R ₁₁Be hydrogen or alkyl independently.In addition, R ₁₀Be hydrogen or hexyl independently, R ₁₁Be hydrogen or hexyl independently.Perhaps, R ₁₀Be hydrogen or octyl group independently, R ₁₁Be hydrogen or octyl group independently.

In other multiple embodiments, the shitosan of micellelike hydrophobically modified is corresponding to the hydrophobic redox mediators chitosan modified of the micellelike of formula 2A:

Wherein n is an integer; R _10aBe hydrogen or hydrophobic redox mediators independently; R _11aBe hydrogen or hydrophobic redox mediators independently.

In addition, in multiple embodiments, the shitosan of micellelike hydrophobically modified is corresponding to the modification of chitosan of formula 2B or redox mediators chitosan modified:

R wherein ₁₁, R ₁₂With n suc as formula the definition in 2.In certain embodiments, R ₁₁And R ₁₂Be hydrogen or straight chained alkyl or branched alkyl independently, preferred hydrogen, butyl, amyl group, hexyl, heptyl, octyl group, nonyl, decyl, undecyl or dodecyl.In various embodiments, R ₁₁And R ₁₂Be hydrogen, butyl or hexyl independently.

The shitosan of micellelike hydrophobically modified can carry out in various degree modification with hydrophobic grouping.The degree of hydrophobically modified can be by being determined with respect to the percentage of the number of free amine groups in the unmodified shitosan by the free amine of hydrophobic grouping modification.The hydrophobically modified degree can be according to acid base titration and/or nulcear magnetic resonance (NMR) (NMR) particularly ¹H NMR data are assessed.This hydrophobically modified degree can change by wide region, and at least about 1,2,4,6,8,10,12,14,16,18,20,25,30,32,24,26,28,40,42,44,46,48% or higher.Preferably, the hydrophobically modified degree is about 45% for about 10%-, and about 10%-is about 35%, and about 20%-is about 35%, or about 30%-about 35%.

In other multiple embodiments, the hydrophobic redox mediators of formula 2A is an osmium, ruthenium, iron, nickel, rhodium, rhenium or cobalt and 1,10-phenanthrolines (phen), 2,2 '-bipyridine (bpy) or 2,2 ', 2 " terpyridyl (terpy); methylene green; methylenum careuleum; poly-(methylene green); poly-(methylenum careuleum); luminol, the nitryl fluorenone derivative, azines, phenanthrolines diketone osmium, catechol-side group terpyridyl, toluene blue, cresyl blue, Nile blue, dimethyl diaminophenazine chloride, the azophenlyene derivative, tionin, reddish black A, reddish black B, blutene, acetophenone, metal phthalocyanine, Nile blue A, the ligand transition metal of modification, 1,10-phenanthrolines-5, the 6-diketone, 1,10-phenanthrolines-5, the 6-glycol, [Re (phen-diketone) (CO) ₃Cl], [Re (phen-diketone) ₃] (PF ₆) ₂Poly-(metal phthalocyanine), poly-(thionine), the benzoquinones class, the diimine class, the diaminobenzene class, the diamino-pyridine class, phenthazine phenoxazine, toluidine blue, brilliant cresyl blue, 3, the 4-4-dihydroxy benzaldehyde, poly-(acrylic acid), poly-(reddish black I), poly-(Nile blue A), polyaniline, polypyridine, polypyrrole, polythiophene, poly-(thieno [3,4-b] thiophene), poly-(3-hexyl thiophene), poly-(3,4-ethylidene dioxy pyrroles), poly-(isothianaphthene), poly-(3,4-ethylidene dioxy thiophene), poly-(difluoro acetylene), poly-(4-dicyano methylene-4H-ring penta [2,1-b; 3,4-b '] two thiophene), the transition metal complex (complex) of poly-(3-(4-fluorophenyl) thiophene), poly-(dimethyl diaminophenazine chloride), or its combination.

Preferably, hydrophobic redox mediators is Ru (phen) ₃ ^{+ 2}, Fe (phen) ₃ ^{+ 2}, Os (phen) ₃ ^{+ 2}, Co (phen) ₃ ^{+ 2}, Cr (phen) ₃ ^{+ 2}, Ru (bpy) ₃ ^{+ 2}, Os (bpy) ₃ ^{+ 2}, Fe (bpy) ₃ ^{+ 2}, Co (bpy) ₃ ^{+ 2}, Cr (bpy) ₃ ^{+ 3}, Os (terpy) ₃ ^{+ 2}, Ru (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Co (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Cr (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Fe (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Os (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}Or its combination.More preferably, hydrophobic redox mediators is Ru (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Co (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Cr (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Fe (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}, Os (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}Or its combination.In various preferred embodiments, hydrophobic redox mediators is Ru (bpy) ₂(4-methyl-4 '-(6-hexyl)-2,2 '-bipyridine) ^{+ 2}

Make the immobilization material of modifier for having hydrophobic redox mediators, hydrophobic redox mediators is covalently bound on shitosan or the polysaccharide main chain usually.Usually, under the shitosan situation, hydrophobic redox mediators is covalently bound on the amine functional group of shitosan by-N-C key.Under metal complex redox mediators situation, metal complex by the alkyl on from the amido of shitosan to the one or more parts that are connected to metal complex-the N-C key is attached on the shitosan.Structure corresponding to formula 2C is the example that is attached to the metal complex on the shitosan:

Wherein n is an integer, R _10cBe hydrogen or independently corresponding to the structure of formula 2D, R _11cBe hydrogen or corresponding to the structure of formula 1D, m is the integer of 0-10 independently, M is Ru, Os, Fe, Cr or Co, and heterocycle is bipyridyl, phenanthrolines, acetylacetone,2,4-pentanedione and the combination thereof of bipyridyl, replacement.

The hydrophobic grouping that is used for modification of chitosan plays electronic environment that (1) regulation hole dimension of immobilization material and (2) change shitosan to keep the double action in acceptable orifice ring border, and the two all makes enzyme or organelle stabilisation.About first effect of this hydrophobic grouping, the shitosan of hydrophobically modified provides its intermediate pore size to depend on the immobilization material of hydrophobic grouping size.Therefore, size, shape and the modification degree of the shitosan of band hydrophobic grouping influence the size and dimension in hole.This effect of hydrophobic grouping make can be by changing hydrophobic grouping size and branching and do hole dimension big or do little or make difformity, to be complementary with certain enzyme or organelle.

About second effect of this dewatering cationic, the character of the chitosan film of hydrophobically modified can be by changing with the hydrophobic grouping modification of chitosan.This hydrophobically modified of shitosan can influence the orifice ring border with the number in exchange site by increasing proton.Except the pH that influences material, the hydrophobically modified of shitosan also provides the film as mechanical barrier, and this has protected immobilised enzymes again.

Table 5 shows the proton of the chitosan film of hydrophobically modified can use the exchange number of sites.

Table 5: the proton of every gram chitosan polymer can be with exchanging number of sites

Film	Exchange site (the x10 of every gram -4mol SO 3/g)
		Shitosan	10.5±0.8
The butyl modification	226±21
		The hexyl modification	167±45
The octyl group modification	529±127
		The decyl modification	483±110

In addition, this polycationic polymer can immobilized enzyme or organelle, and with respect to same enzyme or organelle the activity in cushioning liquid, increase the activity of the enzyme that is fixed therein.In various embodiments, polycationic polymer is the hydrophobically modified polysaccharide, particularly the hydrophobically modified shitosan.For example, measured the enzymatic activity of glucose oxidase for mentioned hydrophobically modified.Observe and have enzymatic activity high in the hexyl modification of chitosan of glucose oxidase in being suspended in tert-pentyl alcohol.For glucose oxidase, to compare in buffer solution with enzyme, these fixed films show 2.53 times of enzymatic activitys to be increased.Table 6 has been listed the glucose oxidase activity of a series of hydrophobically modified shitosans in detail.

Table 6: the glucose oxidase activity of modification of chitosan

Enzymatic activity

Film/solvent (unit/gm)

Buffer solution 103.61 ± 3.15

Unmodified shitosan 214.86 ± 10.23

The hexyl shitosan

Chloroform 248.05 ± 12.62

Tert-pentyl alcohol 263.05 ± 7.54

50% acetate 118.98 ± 6.28

The decyl shitosan

Chloroform 237.05 ± 12.31

Tert-pentyl alcohol 238.05 ± 10.02

50% acetate 3.26 ± 2.82

The octyl group shitosan

Chloroform 232.93 ± 7.22

Tert-pentyl alcohol 245.75 ± 9.77

50% acetate 127.55 ± 11.98

The butyl shitosan

Chloroform 219.15 ± 9.58

Tert-pentyl alcohol 217.10 ± 6.55

50% acetate 127.65 ± 3.02

The present invention has the hydrophobically modified shitosan of alkyl as modifier for preparation, and chitosan gel rubber is suspended in the acetate, then adds alcoholic solvent.In this chitosan gel rubber, add aldehyde (for example butyraldehyde, hexanal, octanal or capraldehyde), then add sodium cyanoborohydride.By the isolated by vacuum filtration products therefrom, and wash with alcoholic solvent.Then with modification of chitosan in vacuum furnace in 40 ℃ of dryings, and obtain the stratiform white solid.

The present invention has the hydrophobically modified shitosan of redox mediators as modifier for preparation, by with 4,4 '-dimethyl-2,2 '-bipyridine contacts with diisopropylamine lithium, then add alkylene dihalide with generate 4-methyl-4 '-(6-haloalkyl)-2,2 '-bipyridine, derivatization redox mediators part.Then make this part and Ru (bipyridine) ₂Cl ₂Hydrate contacts in the presence of inorganic base and refluxes in water-alcohol mixture, until exhausting Ru (bipyridine) ₂Cl ₂Then use ammonium hexafluorophosphate, or optional perchloric acid sodium salt or sylvite precipitated product, recrystallization subsequently.The then redox mediators that will derive (Ru (bipyridine) ₂(4-methyl-4 '-(6-bromine hexyl)-2,2 '-bipyridine) ^{+ 2}) contact with deacetylated shitosan and heat.Then make redox mediators chitosan modified precipitation and recrystallization.

The hydrophobically modified chitosan film is advantageously insoluble in ethanol.For example, above-mentioned chitosan enzyme immobilization material generally has the fixing also function of stabilized enzyme in having up to the solution of about 99wt.% or 99 volume % ethanol.In multiple embodiments, the chitosan enzyme immobilization material has 15,20,25,30,35,40,45,50,55,60,65,70,75,80,85,90, function is being arranged in the solution of 95wt.% or volume % or more ethanol.

In other embodiments, micella or reverse micelle immobilization material are polyanionic polymers, for example hydrophobically modified polysaccharide, particularly hydrophobically modified alginates.Alginates are D-mannuronic acids of comprising β-(1-4)-connect and the linearity of the L-guluronic acid residue of α-(1-4)-be connected branched polymer not.

The not protonated form of the D-mannuronic acid of β-(1-4)-connect is corresponding to formula 3A structure:

The not protonated form of the L-guluronic acid residue of α-(1-4)-connect is corresponding to formula 3B structure:

Assorted (heterogeneous) polymer that alginates are made up of the polymer blocks of the polymer blocks of mannuronic acid residue and guluronic acid residue.

Alginate polymer is modification in various manners.One type is to use greater than ammonium ion (NH ₄ ⁺) the alginates of dewatering cationic modification.This dewatering cationic plays the hole dimension of (1) regulation polymer and the double action that (2) serve as the chemical buffer of the pH level that helps retaining hole, and the two all makes enzyme or organelle stabilisation.First effect about this dewatering cationic obtains the immobilization material that its intermediate pore size depends on the dewatering cationic size with dewatering cationic modification alginates.Therefore, size, shape and the modification degree with alginates of dewatering cationic influences the size and dimension in hole.This effect of dewatering cationic make can be by changing dewatering cationic size and branching and do hole dimension big or do little or make difformity, to be complementary with certain enzyme or organelle.

About this dewatering cationic second the effect, the character of alginate polymer can by dewatering cationic be exchanged for proton as on the alginates-CO ₂ ^-The gegenion of group changes.The variation of this gegenion provides cushioning effect to pH because dewatering cationic than proton right-CO ₂ ^-The affinity in site is much bigger.The sort buffer effect of alginates film makes the pH in hole keep not changing with pH value of solution substantially; On the other hand, the variation of the pH in hole opposing pH value of solution.In addition, this alginates film provides mechanical barrier, and this has protected immobilized enzyme or organelle again.

Be the alginates film of preparation modification, first step is the suspension curtain coating formation film with alginate polymer and dewatering cationic solution.Then extracting goes out excessive dewatering cationic and salt thereof from film, and with film curtain coating again.Again during curtain coating, film comprise with the alginates film-CO ₂ ^-The dewatering cationic of site combination.The salt that removes the dewatering cationic in the film obtains more stable and reproducible film; If they are not removed, then excess salt may be trapped within and cause the space in the hole or in film.

In one embodiment, the alginates film of modification makes by the suspension curtain coating with alginate polymer and dewatering cationic salt such as bromination quaternary ammonium solution.Behind the excessive bromination quaternary ammonium or hydrogen bromide in removing film, with film again curtain coating form desalination membrane.The desalination meeting of film makes quaternary ammonium cation remain resident in carboxylic acid exchange site, but eliminated that excess salt may be trapped in the hole or may the film after balance in cause the complex situations in space.Exemplary dewatering cationic is ammonium cation, quaternary ammonium cation, alkyl trimethyl ammonium cation, alkyl triethyl ammonium cation, organic cation, phosphonium cation, triphenyl phosphonium, pyrrole pyridine phosphonium cation, miaow azoles phosphonium cation, 16 alkane base pyrrole pyridine Phosphonium, second ingot, viologen, methyl viologen, benzyl viologen, two (triphenylphosphine) imonium, metal complex, bipyridyl metal complex, phenanthrolines Base Metal complex compound, [Ru (bipyridine) ₃] ²⁺[Fe (phenanthrolines) ₃] ³⁺

In a preferred embodiment, dewatering cationic is the ammonium cation.Especially, dewatering cationic is a quaternary ammonium cation.In another embodiment, quaternary ammonium cation is represented by following formula 4:

R wherein ₁, R ₂, R ₃And R ₄Be the alkyl or the heterocycle of hydrogen, alkyl, replacement, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.In another embodiment, preferred R ₁, R ₂, R ₃And R ₄Be hydrogen, methyl, ethyl, propyl group, butyl, amyl group, hexyl, heptyl, octyl group, nonyl, decyl, undecyl, dodecyl, tridecyl or myristyl, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.In another embodiment, R ₁, R ₂, R ₃And R ₄Identical and be methyl, ethyl, propyl group, butyl, amyl group or hexyl.In another embodiment, preferred R ₁, R ₂, R ₃And R ₄It is butyl.Preferably, quaternary ammonium cation is tetrapropyl ammonium (T3A), four pentyl ammonium (T5A), tetrahexyl ammonium (T6A), four heptyl ammoniums (T7A), trimethyl icosyl ammonium (TMICA), trimethyl octyl-decyl ammonium (TMODA), trimethyl hexyl decyl ammonium (TMHDA), trimethyl myristyl ammonium (TMTDA), trimethyl octyl group ammonium (TMOA), trimethyldodecane base ammonium (TMDDA), trimethyl decyl ammonium (TMDA), trimethyl hexyl ammonium (TMHA), TBuA (TBA), triethyl group hexyl ammonium (TEHA) and combination thereof.

Studied the characteristic in hole, and the pore structure of this film being desirable for enzyme immobilization, because these holes are hydrophobic, is micella on the structure, can cushion outside pH and change, and have interconnectivity between high hole.

In another experiment, the alginates and the lauryl amine of ultra-low molecular amount placed 25% ethanol, and reflux, generate the alginates of dodecyl modification by the hydroxy-acid group amidatioon.Various alkylamines can substitute lauryl amine and generate the C that has on the reactive hydroxy-acid group of the varying number that is connected to the alginates structure ₄-C ₁₆The alkyl-modified alginates of alkyl.In various embodiments, at least about 1,2,4,6,8,10,12,14,16,18,20,22,24,26,28,30,32,34,36,38,40,42,44,46,48% or more multi-carboxy acid group and alkylamine.

Hydrophobically modified alginates film has favourable insoluble in ethanol.For example, above-mentioned alginates enzyme immobilization material generally has the function of enzyme immobilization and stabilisation in the solution that has at least about 25wt.% or 25 volume % ethanol.In multiple embodiments, alginates enzyme immobilization material has 25,30,35,40,45,50,55,60,65,70,75,80,85, function is being arranged in the solution of 90wt.% or volume % or more ethanol.

For estimating for certain enzyme or the best immobilization material of organelle, selected enzyme or organelle can be fixed in the various immobilization materials, be deposited on the electronic conductor, and be used in contain in the cushioning liquid be useful on this certain enzyme electron mediator (as NAD ⁺) and/or the solution-treated of substrate.Obtain fluorescence micrograph, be fixed on enzyme in the particular fixed formed material or organelle after the immobilization when being still the enzyme with catalytic activity, this photo demonstrates fluorescence.This is to determine that whether the particular fixed formed material can and keep a kind of mode of the catalytic activity of enzyme or organelle simultaneously with enzyme or organelle immobilization and stabilisation.For example, the amylase for consuming starch provides the enzyme immobilization material of maximum relative activity to provide by enzyme being fixed in the butyl shitosan that is suspended in tert-pentyl alcohol.For the amylase that consumes maltose, maximum relative activity can provide by enzyme being fixed in the intermediate molecular weight decyl chitosan modified.

Cathode assembly

Shown in Figure 10-12, the front side cathode assembly 51 that is positioned at the front side 21 in the chamber 15 of confluxing is constructed according to the mode that is similar to front side anode assemblies 45.Promptly, assembly 51 comprises similar framework (being generally referenced as 201) and a plurality of negative electrode (each negative electrode generally all is labeled as 207), this framework comprises the frame member 201A and the 201B of a plurality of openings 205 of having of a pair of coupling, each negative electrode remains in the framework opening 205 by framework 201, and negative electrode of each framework opening.

In some embodiments, each negative electrode 207 comprises collector body 211.In one embodiment, collector body 211 comprises thick about 0.018 inch gold-plated stainless steel cloth sheet.Suitable electrical lead 213 (for example No. 28 conductors) is fixed on the collector body 211.Negative electrode 207 also is included in the layered catalyst structure (being generally referenced as 225) on collector body 211 back sides.This layered catalyst structure comprises gas diffusion layers, catalyst layer and polyelectrolyte film (Nafion

).In some instances, cathode gas diffusion layer comprises the carbon (C*Chem) of dried component: Monarch 1400 carbon blacks (Cabot) and the steam activation of Chemsorb 1505G5 porous dipping, and wet component: be dissolved in QuickSet 2-part epoxy resin (The Original Super Glue Corp.) and 60% polytetrafluoroethylene (PTFE) dispersion (Sigma) in water in the acetone.To do component and grind in the food grinder, and mix wet component with ultrasonic homogenizer, then merging is done wet component and with putty knife it is mixed, and has the denseness of toothpaste until mixture.In some other embodiment, the layer that gas diffusion layers is not necessarily different with catalyst layer, but may be incorporated in together.When assembling, this layered catalyst structure is fixed on the back side of collector body 211.In a specific embodiment, by comprising platinum black catalyst and Nafion

The side that ionomeric ink is applied to gas diffusion layers (for example, as the LT2500W Low Temperature Elat that has microporous layers in each side E-Tek) prepares the stratiform catalyst structure.In case comprise after the ink dried of catalyst, with regard to following assembling stratiform catalyst structure: gaseous diffusion is placed on the collector body top, makes on the platinum black catalyst side direction, then with polyelectrolyte film (Nafion for example

) place on the top of gas diffusion layers and catalyst layer.This assembly is soaked in water to be fixed together by hot pressing then, obtains the negative electrode accessory (fixture) of independence (stand-alone).Polyelectrolyte film (Nafion for example

Ionomer) be impermeable to fuel fluid, but can conduction electron and proton.

The size of each negative electrode 201 is slightly larger than corresponding framework opening 205, and the respective side that makes the side edge portion of negative electrode extend beyond the framework opening is used for the part of overlapping frame member 201A and 201B.Negative electrode 201 by the binding agent anterior layer 275 between the collector body 211 that former frame member 201A and negative electrode 201 are set and be arranged on after-frame member 201B and the layered catalyst structure 225 of negative electrode 201 between binding agent after layer 277 be fixed on the frame member.Binding agent anterior layer 275 and back layer 277 comprise for example thick about 0.0005 inch polyurethane hot melt adhensive membrane.Binding agent anterior layer 275 and back layer 277 have respectively the corresponding size and dimension of size and dimension with preceding frame member 201A and after-frame member 201B through structure.When applying to this assembly (as passing through hot pressing) when hot, the fusing of this binding agent to be being fixed on these two frame member 201A and 201B, collector body 211 and catalyst structure 225 on the fixing toward each other position, thereby forms the front side cathode construction of integral body.

In other embodiment with self-bearing type biological-cathode catalyst carrier shown in Figure 52, each negative electrode 207 comprises the collector body 525 that is embedded in the negative electrode.In one embodiment, this collector body has near-end and far-end, and axle extends along the longitudinal.The conductibility individual layer 527 that contacts with collector body 525 from the near-end of collector body to the coaxial extension of far-end.Suitable electrical lead 213 is received on the collector body 525.In other embodiments, self-bearing type biological-cathode catalyst carrier does not comprise embedding collector body 525.Conductibility individual layer 527 is mixtures of the first electrical conductivity material, the second electrical conductivity material and adhesive, and can be by following in " preparation of self-bearing type biological-cathode catalyst carrier " the described manufacturing of part.One side of conductibility individual layer 527 is breathed part as the hydrophobic air of biological-cathode.Then apply the catalyst layer (not shown) that contains enzyme in the described method of " conducting polymer base nano-wire " part to the opposite side of individual layer 527 by following.Resulting like this biological-cathode comprises hydrophobic air breathing side in the one side of collector body 525, comprises at the back side of collector body 525 containing the enzyme side.As selection, can in the mixture that is used to form individual layer 527, add the catalyst particle that contains enzyme (as following as described in " enzyme of sealing is sneaked in the carbon paste " part), thereby in the conductibility individual layer, provide enzyme.

The size of each negative electrode 201 is slightly larger than corresponding framework opening 205, and the respective side that makes the side edge portion of negative electrode extend beyond the framework opening is used for the part of overlapping frame member 201A and 201B.Binding agent anterior layer 275 between one of the individual layer 527 of negative electrode 201 by former frame member 201A and negative electrode 201 the are set side and be arranged on after-frame member 201B and the offside of the individual layer 527 of negative electrode 201 between binding agent after layer 277 be fixed on the frame member.Binding agent anterior layer 275 and back layer 277 comprise for example thick about 0.0005 inch polyurethane hot melt adhensive membrane.Binding agent anterior layer 275 and back layer 277 have respectively the corresponding size and dimension of size and dimension with preceding frame member 201A and after-frame member 201B through structure.When applying to this assembly (as passing through hot pressing) when hot, this binding agent fusing to be being fixed on these two frame member 201A and 201B, collector body 525 and individual layer 527 on the fixing toward each other position, thereby forms the front side cathode construction of integral body.

Front side cathode assembly 51 is fixed on the fixing position of front side anode assemblies 45 and chamber 15 speech that confluxes, and makes anode 157 and negative electrode 207 general alignment each other, and be positioned at the conflux corresponding fuel of front side 21 in chamber of fuel and store up pond 41 general alignment.In one embodiment, front side negative electrode framework 201 is fixed on the front side anode framework 151 by binding agent or other suitable mechanical means.Between front side negative electrode framework 201 and front side anode framework 151, provide the sealing (not shown) so that each anode/cathode groups and adjacent each anode/cathode groups are separated.Can form sealing with multitude of different ways, for example by utilizing the suitable binder layer between negative electrode framework 201 back sides and anode framework 151 fronts, or by one or more containment members (for example sealing gasket), or pass through alternate manner.

The rear side cathode assembly 53 of rear side 23 that is positioned at the chamber 15 of confluxing is according to constructing with front side cathode assembly 51 essentially identical modes, and corresponding parts are with identical labelled notation.The framework 201 of rear side cathode assembly 53 is against rear side anode assemblies 47, and rear side negative electrode 201 and rear side anode 157 be registration each other, and with corresponding fuel storage pond 41 registrations of the rear side 23 that is positioned at the chamber 15 of confluxing.By the above same way as of describing at front side cathode assembly 51, rear side cathode assembly 53 is fixed on the position of fixing with respect to conflux chamber 15 and rear side anode assemblies 47.

Preferably, conflux chamber 15, front side anode assemblies 45, rear side anode assemblies 47, front side cathode assembly 51, rear side cathode assembly 53 has essentially identical overall size and shape, and this makes that they can be by stacked or pile up and form compact overall structure (as shown in Figure 6) and place housing 91.The size of this structure depends on the number of " piling up " fuel cell together and changes.(each fuel cell comprises fuel storage pond 41, anode 157 and negative electrode 201).As an example, the fuel cell system 1 with folded 8 batteries of one shown in Fig. 1-6 can have following size: 3.5 inches * 2.0 inches * 1.0 inches (about 8.9cm * 5.1cm * 2.54cm).In design of the present invention, any amount of fuel cell easily can be stacked forms a compact unit.

Should note when above-described each parts are assembled, the anode 157 and the negative electrode 201 that are exposed in the corresponding framework opening 155,205 are placed in the chamber at corresponding fuel storage 41 tops, pond or the below in space 131, thereby make any bubble that is trapped in these spaces can not contact with negative electrode with respective anode.Illustrate best among this Fig. 5 of being configured in, wherein the height of the top of Dui Qi anode and negative electrode framework opening 155,201 is pointed out by line 291.This height 291 is lower than chamber 131, perhaps is lower than the top of chamber 131 at least, thus make in the fuel storage pond 41 any bubble can on float to and the not contacted position of respective electrode structure.

Negative electrode

Negative electrode of the present invention and biological-cathode generally include catalyst, electronic conductor (gas diffusion layers) and optional collector body.Negative electrode of the present invention and biological-cathode comprise following catalyst: the oxygen in this catalyst selectivity ground reducing atmosphere, can carry out direct electron with the cathode electronics conductor and shift, and can minimum ground catalysis (if any) be used for the pure fuel fluid oxidation of fuel cell.When using enzyme as this catalyst, negative electrode also comprises the enzyme immobilization material.Negative electrode also comprises following electronic conductor (gas diffusion layers): thus its to air and fuel fluid be permeable, can promote direct electron with catalyst to shift and the reaction of control oxygen reduction in the water that produces minimize or prevent negative electrode waterflooding (flooding).In each preferred embodiment, catalyst can discharge to oxidant (oxygen) from the direct electron gain of electronic conductor and with electronics.In other embodiments, this negative electrode or biological-cathode also comprise electron mediator, its mediation electronics transfer from the electronic conductor to the catalyst.In other various embodiments, this negative electrode or biological-cathode also comprise electron mediator and be used for the eelctro-catalyst of this electron mediator, and wherein electron mediator and eelctro-catalyst promote electronics from the electronic conductor to the catalyst, and further to the transfer of oxidant (oxygen).

Air self-respiration type negative electrode

In many aspects of the present invention, relate to a kind of electrode as shown in figure 22.This electrode comprises that air can see through but first area, fuel fluid and the air of the impervious electrical conductivity material of the fuel fluid all second area and the catalyst that can contact fuel fluid and air of permeable conductive of material.Electrode shown in Figure 22 is the work electrode (WE) of air self-respiration type half-cell.This work electrode (WE) comprises zone or the layer that is labeled as first area, second area and catalyst.This half-cell also comprise place near the reference electrode (RE) the work electrode and be preferably precious metal network or carbon plate to electrode (CE).Comprise electron potential and the 10mg/cm of the air self-respiration type half-cell of kind electrode (WE) at room temperature, 0.4V ²Catalyst loading down during operation, produce at least about 16,17,18,19,20,25,30,35,40,45,50,55,60,65,70,75,80mA/cm ²Or higher current density.When air self-respiration type negative electrode of the present invention is introduced into fuel cell, found its electron potential and 10mg/cm at room temperature, 0.4V ²Catalyst loading can provide with the same fuel cell that comprises the known non-platinum cathode of air self-respiration type during operation down and compare higher current density.

In many aspects of the present invention, relate to a kind of electrode that comprises electronic conductor, at least a non-precious metal catalyst and optional carbon load polyamine (polyamine) in addition.In many preferred embodiments, thereby this electrode has passed through the interaction of heat treatment enhancing metallic atom and polyamine, and non-precious metal catalyst is a water with oxygen reduction optionally.In addition, comprise electron potential and the 10mg/cm of the air self-respiration type half-cell of this electrode at room temperature, 0.4V ²Catalyst loading down during operation, produce at least about 16,17,18,19,20,25,30,35,40,45,50,55,60,65,70,75,80 or higher mA/cm ²

In other embodiments, this electrode comprises electronic conductor, can produce enzyme, enzyme immobilization material and the optional electron mediator of water with oxidant reaction and/or is used for the eelctro-catalyst of this electron mediator.The enzyme immobilization material can immobilized enzyme and is made enzyme stable.Electronic conductor can comprise functionalized multi-walled carbon nano-tubes or based on the material of activated carbon, be used to provide the current density of comparing increase with the electrode with non-functionalized or inactive identical electronic conductor.

Go through as following, the negative electrode and the biological-cathode embodiment that wherein exist the direct electron from the electronic conductor to the catalyst to shift are preferred, but comprise electron mediator or electron mediator and eelctro-catalyst the enzymatic biological-cathode embodiment also within the scope of the invention.

When testing in air self-respiration type half-cell as mentioned above, a plurality of embodiments of negative electrode of the present invention and biological-cathode are at electron potential and the 10mg/cm of room temperature, 0.4V ²Catalyst loading down during operation, produce at least about 16,17,18,19,20,25,30,35,40,45,50,55,60,65,70,75,80mA/cm ²Or higher current density.

Cathod catalyst

Cathod catalyst can be enzyme or non-precious metal catalyst usually.Preferably, catalyst selectivity is reduced to oxidant (oxygen).This selectivity is preferred, and reason is: such cathod catalyst can not react and oxidized fuel fluid, thereby keeps the high usage of fuel and high cathode potential.In addition, anode and negative electrode needn't be separated with polymer dielectric film, thereby improve the cost and the effect of fuel cell.

When enzyme was used as catalyst, it must be at biological-cathode place reduction oxygen.Usually, can use the native enzyme of native enzyme, synthetic enzyme, artificial enzyme and modification.In addition, can use by nature evolution or orthogenesis and the engineering enzyme of through engineering approaches.In other words, can use the organic or inorganic molecule of analogue enztme character in embodiments of the invention.In a plurality of preferred embodiments of the present invention, this enzyme is bilirubin oxidase, laccase, superoxide dismutase, peroxidase or their combination.Preferably, this enzyme comprises bilirubin oxidase.

In other embodiments, cathod catalyst is a non-precious metal catalyst.Preferably, non-precious metal catalyst optionally reduces oxygen and does not react with fuel fluid.Another preferred feature of non-precious metal catalyst is it has tolerance basically to the existence of fuel fluid.When catalyst had tolerance basically to fuel fluid, along with the concentration increase of the fuel fluid that contacts with this catalyst, catalyst activity can not reduce basically.In other words, when using the electrolyte solution contain alcohol, comprise that current density that the half-cell of this catalyst produces is not less than (that is, reaching about 75% at least) same maximum current density that half-cell produced when using the electrolyte solution that contains 5% alcohol basically.For example, when fuel fluid is alcohol, about at least 75% of the maximum current density that the current density when using the electrolyte solution contain 30wt% alcohol in the half-cell is produced when being to use the electrolyte solution of the alcohol that contains 5wt%.Especially, catalyst is to the tolerance that has of methyl alcohol and ethanol.

In order to increase the current density of the negative electrode that contains non-precious metal catalyst, catalyst can be heat-treated under about 500-900 ℃.Non-precious metal catalyst can be transition metal, transition metal macrocyclic compound or their combination.When catalyst was transition metal macrocyclic compound, it can be transition metal phthalocyanine, transition metal porphyrin, its derivative or analog or their combination.The example of transition metal macrocyclic compound is FePC, phthalocyanine cobalt, PORPHYRIN IRON, Cobalt Porphyrin, its derivative or analog or their combination.Preferred transition metal macrocyclic compound comprises 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II), or derivatives thereof or analog.

Non-precious metal catalyst preferably interacts with polyamine or combines.In the particularly preferred embodiment, polyamine is the carbon loaded amine.Polyamine can be polyaniline, polypyrrole, its derivative or analog or their combination.In different preferred embodiments, polyamine comprises polypyrrole, or derivatives thereof or analog.

In order to strengthen the interaction between non-precious metal catalyst and the polyamine or to combine, carry out a heat treatment step in case destroy the interaction of non-precious metal catalyst inside and polyamine inside and cause non-precious metal catalyst and polyamine between interaction (or key).This heat treatment step preferably carries out under inert atmosphere.During heat treatment step, non-precious metal catalyst, particularly Metallomacrocycle catalyst, with situation that polyamine or carbon load polyamine contact under be heated to about 500 ℃ to about 900 ℃, preferred about 550 ℃ to about 650 ℃, more preferably from about 590 ℃ to about 610 ℃.General about 0.5 hour to about 6 hours of this heating steps duration, preferred about 0.5 hour to about 3 hours, more preferably from about 1 hour.

Go through as following, thereby the hydrophobicity that can control the electronic conductor of these electrodes is controlled at the water that produces during the oxygen reduction.For this reason, can add hydrophilic reagent, make up the hydrophobicity that reduces electronic conductor such as phosphotungstic acid, poly-(4-styrene sulfonic acid) or its.

Below table 7 show acceptable material and the process conditions that are used to prepare the electrode that contains non-precious metal catalyst and polyamine, and when these electrodes are introduced in the above-mentioned half-cell observed performance characteristic.Each material, process conditions or performance characteristic can make up with the electron conductor material that describes below, and can make up with each other materials of describing, process conditions or performance characteristic.

Table 7

Electronic conductor (gas diffusion layers)

Usually, electronic conductor is the material of conduction electron.Electronic conductor can be an organic or inorganic, as long as it can pass through this conduct electronics.In a plurality of embodiments of the present invention, electronic conductor is carbon-based material preferably.Exemplary carbon-based material be carbon cloth (E-Tek), carbon paper, carbon filament reticulated printing electrode, carbon paper (Toray), carbon paper (ELAT), carbon black (Vulcan XC-72, Cabot), the graphite worm of carbon black, carbon dust, carbon fiber, Single Walled Carbon Nanotube, double-walled carbon nano-tube, multi-walled carbon nano-tubes, carbon nano pipe array, coated with CVD (chemical vapor deposition) diamond conductor, vitreous carbon, mesoporous carbon, graphite, uncompressed, tear the purifying flake graphite (Superior of layer open

Graphite), high-performance graphite and carbon dust (Formula BT ^TM, Superior

Graphite), high order pyrolytic graphite, pyrolytic graphite, polycrystalline graphite and combination thereof.

The carbon back electronic conductor of number of different types can provide optimum performance for specific purposes.For example, it is useful for the fuel cell of use liquid electrolyte or fuel fluid with the different electronic conductors that change in the position in the electronic conductor layer to have porous carbon structure and a porosity.In addition, for biological-cathode, the direct electron of the enzyme during functionalized multi-walled carbon nano-tubes and activated carbon electronic conductor help from the electronic conductor to the biological-cathode shifts.

In some other embodiment, use following electronic conductor (gas diffusion layers): its hole in a zone of electronic conductor is greater than the hole in another zone at electronic conductor.For example, Figure 23 shows the electronic conductor with asymmetric pore size distribution.The control hole size makes the first area of electronic conductor can see through air but can not see through fuel fluid (light-colored part among Figure 23) that the second area of electronic conductor then can see through fuel fluid and air (dark part among Figure 23).In the particular of fuel cell and biological fuel cell, when a kind of major part of material (promptly, at least about 50 volume %, 55 volume %, 60 volume %, 65 volume %, 70 volume %, 75 volume %, 80 volume %, 85 volume % or more) in the time of can permeating by electronic conductor through one section special time, then electronic conductor is that this material (for example, air or fuel fluid) can see through (or permeable) substantially.For example, when the fuel fluid of about at least 50 volume % passed through electronic conductor in about 3 hours, then electronic conductor was that fuel fluid is permeable substantially.

Such electronic conductor is for using liquid electrolyte but not the fuel cell of solid electrolyte is particularly useful.Usually, this type of electronic conductor comprises carbon black.But, also can use conductive carbon material as acceptable substitute with following character: high surface, with regard to can see through air and/or air and fuel fluid with regard to all can seeing through acceptable porosity and suitable hydrophobicity.Can control hydrophobicity and control the water that generates from oxygen reduction.Can increase hydrophobicity by the polytetrafluoroethylene that in electronic conductor, adds recruitment.Therefore, the polytetrafluoroethylene that adds in electronic conductor is many more, and then the hydrophobicity of electronic conductor is big more.In addition, can be by adding more a spot of polytetrafluoroethylene or reducing hydrophobicity by the hydrophilic reagent that adds such as phosphotungstic acid or poly-(4-styrene sulfonic acid).

Can prepare electronic conductor by adding pore former with porosity gradient.Acceptable pore former diffuses in the electronic conductor (for example carbon-coating), and can easily remove from electronic conductor subsequently.Exemplary pore former is an ammonium carbonate.

In other a plurality of embodiments, electronic conductor comprises functionalized multi-walled carbon nano-tubes (MWCNT).These functionalized MWCNT are modified by hydroxyl, carboxyl, amino or thin base or their combination on nanotube surface.In a plurality of embodiment preferred, functionalized MWCNT is modified by hydroxyl or carboxyl or its combination.Functionalized MWCNT has the average diameter less than about 15nm.Preferably, functionalized MWCNT has the average diameter less than about 8nm.These functionalized MWCNT with enzyme as catalyst and to wish to set up between electronic conductor and enzyme in the biological-cathode that direct electron shifts be particularly preferred.

Other electronic conductor comprises activated carbon.As described below, many kinds of material with carbon elements can be activated.Preferably, the carbon black that is activated is the carbon black of selling with Printex XE-2 trade name that can obtain from Degussa, and it has highly structural, the carbon black of special conduction, and average particle size is 30nm, the BET surface area is about 910m ²/ g.Material with carbon element immerses in the cold water then and is activated by being heated to 600-900 ℃.Be not subject to concrete theory, it is believed that the afterwards quick cooling of heating can destroy carbon plate and form the structure that has high surface, may be similar to nanotube.

Be not subject to concrete theory, it is believed that functionalized MWCNT and activated carbon have following function when being used for enzymatic biological-cathode: (1) interacts by the cyano group acid side chain of hydrogen bond and enzyme; (2) the enzyme orientation is made the active side of biological-cathode towards enzyme; (3) in biological-cathode, provide the electron tunnel of the enzyme of favourable orientation.

Be used for the preparation of the material of air self-respiration type negative electrode

Can use following universal process to prepare above-mentioned various electronic conductor, catalyst and negative electrode.

Electronic conductor with gradient pore size

In order to prepare the electronic conductor of the porosity gradient with hope, preparation earlier has surface area, porosity and the hydrophobic electronic conductor of hope.Usually, prepare electronic conductor by a slice carbon-based supports material being immersed in the electrophoretic deposition bath that contains the electron conductor material (for example carbon black), nonionic surfactant and the water-repelling agent (for example polytetrafluoroethylene PTFE) that are hopeful.Under the known reference condition of prior art, use electrode is carried out electrophoretic deposition.After the electrophoretic deposition, water flushing electronic conductor is also dry.The coating dough sample thing (dough) of electron conductor material (for example carbon black), water-repelling agent (for example PTFE) and solvent that preparation is wished, and it is coated to the both sides of the electronic conductor for preparing by electrophoretic deposition.Solvent in the coating dough sample thing is evaporated, and forms coated film.Coated film is by hot pressing sintering then.By pore former being brushed a side of electronic conductor, make pore former diffuse in the electronic conductor again the thermal decomposition pore former then and form hole, distribution that can control hole.

Particularly, electronic conductor shown in Figure 2 passes through a slice carbon cloth (18cm ²E-Tek, B1A) be dipped into electrophoretic deposition and prepare in bathing, described electrophoretic deposition is bathed and is contained 0.60g carbon black (XE2,20mL solution Degussa), 0.60mLTriton X-100,0.67g 60% polytetrafluoroethylene (PTFE) dispersion (Aldrich) and 100mL milliQ water.Use the carbon paper conduct to electrode, making carbon paper and carbon cloth spacing is 3mm, carries out 20 minutes electrophoretic depositions under 40V.In case finish electrophoretic deposition, water flushing carbon cloth is so that remove Triton X-100, and in 100 ℃ of vacuum furnaces dry 1 hour.Preparation 0.40g carbon black (Degussa, XE2), the dough sample thing of 0.33g 60%PTFE dispersion (Aldrich) and 8mL ethanol.Then dough sample thing is coated to as mentioned above both sides by the carbon cloth of electrophoretic deposition preparation.At room temperature evaporate the solvent 30 minutes in the coated film.Then at 500 pounds of pressure and 95 ℃ of these films of following hot pressing, afterwards 380 ℃ of sintering 30 minutes.PTFE content in the coated film is about 33wt%.In addition, control the distribution of coated film mesopore by a side that 2mL pore former (the 1.0wt% solution of ammonium carbonate in isopropyl alcohol) is brushed carbon cloth.The pore former migration enters coated film and causes the formation of gradient.At 90 ℃ of following thermal decomposition pore formers, in 120 ℃, 28 inch of mercury vacuum under pressure stoves, clean carbon cloth then.

Heat treated and not heat treated non-precious metal catalyst

Such as cobalt macrocyclic compound (phthalocyanine cobalt (CoPc) and 1,2,3 for example, 4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II) are (CoPcF)) non-precious metal catalyst can place ultrasonic bath to be prepared by this non-precious metal catalyst (for example CoPc or CoPcF) is distributed in the solvent and with dispersion.Evaporating solvent then, and place crucible under 500-900 ℃ of temperature, to heat-treat the powder of drying in inert atmosphere.Resulting catalyst mixes with Nafion solution, and utilizes ultrasonic bath to disperse in solvent.This mixture slowly is coated to the active side and the drying of electronic conductor (gas diffusion layers).In some cases, hydrophilic reagent (for example phosphotungstic acid) is added in the mixture of non-precious metal catalyst and Nafion solution, and utilize ultrasonic bath to disperse, and then be coated on the electronic conductor.When using without heat treated non-precious metal catalyst, non-precious metal catalyst (for example CoPc or CoPcF) and Nafion solution are distributed in the solvent, then this solution slowly is coated to the active side and the drying of electronic conductor.

Heat treated and not heat treated non-precious metal catalyst and polyamine

Such as cobalt macrocyclic compound (phthalocyanine cobalt (CoPc) and 1,2,3 for example, 4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II) are (CoPcF)) non-precious metal catalyst can place ultrasonic bath to be prepared by this non-precious metal catalyst (for example CoPc or CoPcF) and polyamine (for example polypyrrole) are distributed in the solvent and with dispersion.Evaporating solvent then, and place crucible under 500-900 ℃ of temperature, to heat-treat the powder of drying in inert atmosphere.Resulting catalyst mixes with Nafion solution, and utilizes ultrasonic bath to be distributed in the solvent.This mixture slowly is coated to the active side and the drying of electronic conductor (gas diffusion layers).In some cases, hydrophilic reagent (for example phosphotungstic acid) is added in the mixture of CoPc or CoPcF and Nafion solution, and utilize ultrasonic bath to disperse, and then be coated on the electronic conductor.When using without heat treated non-precious metal catalyst and polyamine, non-precious metal catalyst (for example CoPc or CoPcF), polyamine and Nafion solution are distributed in the solvent, then this solution slowly is coated to the active side and the drying of electronic conductor.

Heat treated non-precious metal catalyst on the carbon

Such as cobalt macrocyclic compound (phthalocyanine cobalt (CoPc) and 1,2,3 for example, 4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II) are (CoPcF)) non-precious metal catalyst can evaporating solvent be prepared by this non-precious metal catalyst (for example CoPc or CoPcF) and material with carbon element (for example carbon black) being distributed in the solvent then in ultrasonic bath.Place crucible under 500-900 ℃ of temperature, to heat-treat in the powder of drying in inert atmosphere.Again powder is mixed with Nafion solution, and utilize ultrasonic bath to be distributed in the solvent, then this mixture slowly is coated to the active side and the drying of electronic conductor (gas diffusion layers).

The preparation of enzyme layer

Utilize ultrasonic bath that material with carbon element is distributed to and obtain the carbon suspension in the solvent.Add the enzyme in the cushioning liquid (for example BOD or laccase) solution in carbon suspension vibration then.This mixture slowly is coated to the active side and the drying of electronic conductor (gas diffusion layers).

Self-bearing type biological-cathode catalyst carrier

The flexibility that the design of biological-cathode catalyst carrier as herein described provides structural design, material to select, and can create have suitable hydrophobicity/hydrophilic sandwich construction is used for for example enzyme of biological support cathod catalyst.Illustrating in Figure 52 of biological-cathode catalyst carrier.As mentioned above, the biological-cathode catalyst carrier comprises embedding porous metals web material 525 that is used for collected current and enhancing biological-cathode catalyst carrier rigidity and the conductibility individual layer 527 that centers on collector body 525.Collector body 525 is optional features of biological-cathode catalyst carrier; If do not comprise these parts, then by being electrically connected conductibility individual layer 527 collected currents from the biological-cathode catalyst carrier.

No matter whether have embedding porous metals web material in the biological-cathode catalyst carrier, best biological-cathode electrode allows to carry out water management (promptly in the aerial respiration and the oxidant reactant side of electrode, if excessive water arranged then repel water), thus prevent that the excessive water that generates from blocking the transportation of oxygen to electrode.This generally is to realize by the overall structure hydrophobicity of giving conductibility individual layer surface or conductibility individual layer.With the aerial respiration or oxidant reactant side electrode of opposite surface of electrode on, the outer surface of high surface microporous layers with slight hydrophilic properties and conductibility individual layer is adjacent, is used for strengthening enzyme-electrode and interacts.This can realize by the biological fuel cell with electrode structure of describing among the embodiment 27.Reach in this embodiment shown in Figure 52, the microporous layers (not shown) that is used for enzyme forms on the surface of the conductibility individual layer 527 of being responsible for water management and reactant mass transfer.In certain embodiments, the first electrical conductivity material is modified so that strengthen electronics and ionic conductivity and improve DET, for example carbon black is modified with the conducting polymer that mixes with electron mediator.

In a plurality of preferred embodiments, the first electrical conductivity layer of one or more carbon blacks mixes with the second electrical conductivity layer of one or more carbon fibers and one or more adhesives and forms the conductibility individual layer.Carbon black exists to the concentration of about 45wt% with about 30wt% in the mixture, and the concentration of carbon fiber is at about 20wt% extremely between about 40wt% in the mixture, and the concentration of adhesive is at about 25wt% extremely between about 40wt% in the mixture.In a plurality of preferred embodiments, the content of carbon black is about 33wt% in the mixture, and the content of carbon fiber is about 33wt% in the mixture, and the content of adhesive is about 33wt% in the mixture.

The preparation of self-bearing type biological-cathode catalyst carrier

Be similar to biological anode, the catalyst support layer that is used for the biological-cathode electrode is made by the mixture of material with carbon element, adhesive, pore former, solvent and optional catalyst (as enzyme).By changing the ratio of these components, the character that can change this total is to provide application-specific desired character; Perhaps can utilize the combination manufacturing of these components to have the electrode that two or more zoness of different are used for water management and enzyme interacting.The example of this type of character comprises: (1) substrate permeability, (2) electronic conductivity, hydrophobicity/hydrophily, (3) surface area and (4) surface texture.

The preparation of biological-cathode catalyst carrier material and aforementioned similar at biological anode, difference is receptacle material and sintering circulation and temperature.For example, MDS nylon 6/6 receptacle material that is used for biological anode can be replaced by the PTFE coated fibres glass sheet of similar thickness.Because replace poly-(vinylidene fluoride) adhesive as adhesive, can change so be used for the sintering circulation of biological-cathode with polytetrafluoroethylene (PTFE).For the GDL based on PTFE, whole assembly (being framework and electrode material) at first is no more than 20 minutes at 200 ℃ sintering temperature at the most, and this depends on desired results and preparation.After the sintering, from framework and interim carrier, shift out electrode, and with its 2 aluminium blocks (12 " * 12 " * 1 ") between cooling until solidifying, a few minutes only usually, under 300 ℃ temperature, carry out second time sintering afterwards and be no more than 10 minutes.No matter whether have embedding porous metals web material in the biological-cathode catalyst carrier, these preparation processes all are identical.

If catalyst is not included in wherein when the conductibility individual layer forms, be applied on the catalyst carrier as described in then catalyst layer can " be sneaked into the enzyme of sealing in the carbon paste " as following chapters and sections.As selection, can with contain enzyme catalyst particle (as following chapters and sections " enzyme of sealing is sneaked in the carbon paste " and " biocatalyst ink formulations " described in) add and to be used to form in the mixture of individual layer 527, maybe it can be included in stick with paste or ink in then be applied to the catalyst carrier surface.

The technology of preparing of anode and negative electrode

The concrete steps that are used to prepare anode and cathode assembly have been described in the following embodiments in more detail.

The conducting polymer base nano-wire

For pruning nano wire, the main character that can improve the direct electron transfer of the biosystem that is used for the biological fuel cell application is: (1) electronics and ionic conductivity, (2) high aspect ratio is to penetrate into enzyme active center, (3) thermal stability of precursor material and the electrochemical stability of (4) precursor material under the fuel battery operation condition.The major function of nano wire provides following nanometer construction structure (nano-architectural structure), and this structure has the predetermined electron transfer path (seeing Figure 56 and 57) that is connected between enzyme interior oxidation reduction site and electrode surface.Figure 56 is the schematic diagram of nano-wire array, and wherein enzyme and carbon particle interact.Figure 57 shows nano wire interaction to each other, thereby forms neural net shape electron transfer network between carbon particle, and these carbon particles are the key components that are used to make base electrode.

Conducting polymer (for example polyaniline, polypyrrole, polythiophene etc.) is the exemplary polymer that is used for making by the oxidation polymerization approach nano wire.Utilize the suitable dopant (being used to improve the conductibility of polymer) and first conductive of material for example carbon black particle as being used for the nucleation site of nanowire growth, grown and grafting have the size of hope and a conductibility nano wire of thermal stability and electrochemical stability (for example electricity with ion).Some dopants have more than a kind of function, and can be in building-up process the adjustment agent of structure by serving as internal holes or interior diameter influence the size of nano wire.

Nano wire not only can be grown from polymeric material, also can be from oxide, organic metal and metal material growth in suitable synthesis step.For the direct electron transfer method, preferably, polymer precursor or oxide/Organometallic precursor form the efficiently conductibility nano wire of metastatic electron.

Biological-cathode nano wire with dried enzymatic oxygen reduction reaction

In this part, can provide the example of the conducting polymer base nano-wire that is used for biological fuel cell enzymatic oxygen reduction reaction, but the present invention is not limited to conducting polymer.Conductive oxide precursor, conductibility whisker (whisker) precursor and metal nanometer line are in the direct electron transfer process of the biosystem that can be created in biological fuel cell highly effectively the neural net shape structure, also can grow and be grafted on the various base carrier material (being carbon particle), and can be used to immobilized enzyme.

Multiple different polymer can be used as the conducting polymer that forms nano wire, and for example the Leucoemeraldine alkali of polyaniline, polypyrrole, polyacetylene, aniline, Emaraldine alkali, Pernigraniline alkali form, polythiophene, poly-(to phenylene), poly-(to the phenylene vinyl) and their binary, ternary or quaternary combination can be used as the precursor of growth conductibility nano wire.Can utilize chemical polymerization approach or electrochemical polymerization approach to come the nano wire of these base monomer of polymerization to obtain to want.In order to obtain required size, it is preferred using one or more dopants.Multiple organic sulfonic acid can be used to produce the conducting polymer nano wire.For example, the amphion complex compound (both having comprised that positively charged group also comprised electronegative group on a part) (for example hydroxide 1-(3-sulfopropyl) pyridinium inner salt, 3-(1-pyridine radicals)-1-propane sulfonate etc.) of the conjugation of naphthalene sulfonic acids (camphorsulfonic acid, naphthoquinone sulfonic acid etc.), toluenesulfonic acid (as p-methyl benzenesulfonic acid pyridiniujm, p-methyl benzenesulfonic acid pyridone salt, p-methyl benzenesulfonic acid 4-butyl ammonium etc.), band multiple charge etc. can be used as dopant.

For the conductive oxide precursor that is used as nano wire in biological fuel cell is used, this oxide should be stablized under acidity or alkali condition.For example, the oxide of titanium, ruthenium, osmium, iridium, platinum, gold, palladium, rhenium, aluminium and tin indium oxide (ITO) and their binary, ternary or quaternary combination can obtain to be applicable to the conductibility nano wire of the object of the invention.For this purpose, these oxides can modification, and this both can also can realize by the hydro-thermal synthetic route by conventional route of synthesis (chemical vapor deposition, based on the deposition of ion-plasma etc.).

The example of conductibility whisker precursor is the big ring phthalocyanine of metal complex or various phthalocyanine/porphyrins (porphine) things (speices) of porphyrin complex and nonmetal complexing, for example CuPc, phthalocyanine cobalt, phthalocyanine nickel, FePC, Phthalocyanine Zinc, porphyrin copper, Cobalt Porphyrin, porphyrin ruthenium, porphyrin palladium, porphyrin vanadium, porphyrin zinc etc. and their combination.Can use conventional deposition technique (for example evaporation, glow discharge, gas chemistry method and liquid phase chemical formation technology) long conductibility whisker in next life based on low and high vacuum.

The example of the metal nanometer line precursor that acid or alkali are stable comprises titanium, ruthenium, osmium, iridium, platinum, gold, palladium, rhenium and their binary, ternary or quaternary combination, they can make the conductibility nano wire, are used for the direct electron transferring path between biosystem and base electrode structure.Can use conventional deposition technique (for example evaporation, glow discharge, gas chemistry method and the liquid phase chemical formation technology) metal nanometer line/nano whisker of growing based on low and high vacuum.

The enzyme of sealing is sneaked in the carbon paste

In many embodiments, finish before the deposition that is manufactured on enzyme of electrode holder, this technology causes two different layers.By spray-drying process described herein, the carbon particle of immobilized enzyme can be mixed provides enzyme/carbon electrode integrative-structure in the carbon paste, and the typical loss of enzyme activity that does not cause by solvent and thermoinducible sex change.As described to other GDL preparations, the enzyme of sealing/carbon diffusion electrode carrier can be made in the mode of customization, makes them have the feature of the hope that is used for male or female.Preferably, anode can have the more hydrophilic paste of the hydrophilic component of being discussed below utilizing.In addition, thus cathode carrier can have higher Teflon content provides higher hydrophobicity reserve part hydrophily simultaneously.

Depend on specific requirement, have the electrode of wishing performance thereby there is multiple different component can provide large-scale adjustable parameter to produce to electrode.Particularly, can change concrete material with carbon element (comprising the graphite fibre that uses for rigidity), adhesive and pore former and their selections of ratio separately, thereby the electrode with a series of performances is provided.Initial test has shown maintenance and favorable mechanical stability active in the half-cell structure.

As the catalyst ink water formulation of discussing below, high surface high conductance carbon black is put into paste formulation (anode) or the bigger direct electron transfer from electrode (negative electrode) to enzyme from the enzyme to the electrode can be provided.

The manufacturing process of the enzyme of sealing

In some embodiments, enzyme is fixed on the carbon black (those that provide in for example above showing) of high conductance high surface.In case enzyme is fixed, with the carbon drying, then with graphite fibre, carbon black, alcoholic solvent, poly-(vinylidene fluoride) (PVDF) and/or PTFE adhesive and optional hydrophilic reagent (that is, ammonium carbonate, polyethylene glycol, polyvinyl alcohol or silica dioxide gel) mix.

When mixing carbon paste, all the dried components that may expect the desired amount of first weighing are in container.With mortar and pestle these dried components are mixed up to even dispersion then.Repeatedly add alcohol then on a small quantity, simultaneously slurries are stirred into even thickener.Solvent very little can make the thickener drying and can not play a role, and too many solvent will make thickener become soup shape and inhomogeneous.

Then thickener is taped against on the porous metals silk screen (normally nickel or stainless steel, but can use any porous metals silk screen), with a shape qualification framework round it so that the electrode size that obtains wishing.Can use any instrument (for example spatula, spoon, mud shovel etc.) to form and sprawl thickener.After sprawling, paper handkerchief is placed on around the electrode, electrode carries out mechanical compaction under 5000 pounds pressure, thereby they are compressed and help to remove unnecessary solvent.

After the mechanical compaction step is finished, remove shape and limit framework.Be sidelong Kimwipe in each of electrode, then they under the temperature of 5000 pounds and 125 ℃ by hot pressing 20 seconds.

The biocatalyst ink formulations

As described herein, developed a kind of being used for immobilized enzyme is sprayed to such as the method on the various particles of carbon, polymer and metal oxide.The conventional method that this process for fixation allows to be used for common PE M fuel cell system is made conventional catalyst ink and MEA.In traditional PEM fuel cell, catalyst ink is coated onto on the electrode holder material, dry and be hot-pressed onto amberplex (as Nafion ) on.When enzyme was not fixed, this manufacture method can make enzyme owing to interacting with solvent environment or being exposed to heat and sex change during hot pressing.

The more high stability of the enzyme in immobilization material allows exploitation directly to be coated onto on the commercial electrode holder or is coated onto ink formulations on the electrode holder that is used for fuel cells applications described herein.Carbon, carbon black filler and Nafion that this ink formulations is coated by enzyme

Solution constitutes.The carbon that enzyme coats is made up of the carbon particle that is centered on by enzyme, and wherein said enzyme is trapped within the immobilized polymer.

Biological-cathode catalyst ink water formulation

Many aspects of the present invention relate to the particle that contains the nuclear core that is immobilized the enzyme coating.This nuclear core can be the material that can provide support for immobilized enzyme layer, and wherein this immobilized enzyme layer is coated on the nuclear core.Immobilized enzyme layer comprises enzyme, enzyme immobilization material and optional electron mediator.This immobilised enzymes have its in immobilization and the initial activity before coating at least about 0.65,0.7,0.75,0.8,0.85,0.9,0.95 or bigger activity, and/or when this enzyme continuous catalysis chemical conversion, keep its at least about 75% initial catalytic activity at least about 1,2,3,4,5,6,7 day.In a plurality of other embodiments, this enzyme keeps it at least about 75% initial catalytic activity at least 5,10,15,20,25,30,45,60,75,90,105,120,150,180,210,240,270,300,330,365,400,450,500,550,600,650,700,730 days or more of a specified duration.Be described in more detail below the component of these particles.

Other many aspects of the present invention relate to the technology that preparation is immobilized the particle of enzyme coating.This technology comprises mixes with the suspension that comprises at least a carrier granular, immobilization material and liquid medium containing enzyme solutions, forms mixture.This mixture is formed coating particles (coatedparticle) by spray drying then.

As shown in figure 47, the particle of producing can comprise nuclear core, optional electron mediator, enzyme and enzyme immobilization material (for example polymer substrate).The effect of polymer substrate is stabilized enzyme and is fixed on the carrier that polymer substrate can be the various enzyme immobilization materials that describe below.In addition, except enzyme, multiple compound can be joined and help enzyme in the matrix and play a role.For example, electron mediator, co-factor, coenzyme can be immobilized, and are contacting or can not be leached in the liquid during cyclic washing.

In multiple preferred embodiment, enzyme covalently is not attached on the nuclear core or is adsorbed onto on the nuclear core.In addition, preferably, in the liquid medium that enzyme can not be leached into from the enzyme immobilization material with immobilized enzyme layer contacts.Typically, the immobilised enzymes particle comprises the nuclear core of the about 25wt% of about 0.1wt%-and the coating of the about 70wt% of about 0.1wt%-, and this coating comprises enzyme and the enzyme immobilization material of the about 43wt% of 0.1wt%-and the electron mediator of maximum about 29wt% of the about 29wt% of about 0.1wt%-.Typically, the overall weight percent of enzyme and electron mediator can reach the 57wt% of coating at most.

The nuclear core component

The nuclear core is any can providing support and can be by spray-dired particle for immobilized enzyme layer.For example, the nuclear core particle can be polymer particle, carbon particle, zeolite particles, metallic, ceramic particle, metal oxide particle or their combination.In some embodiments, the nuclear core particle is the inert core may particle.In multiple embodiments, the nuclear core particle is not a polymer particle.The chemical conversion that preferred nuclear core particle can not participate in the stability or the enzyme of enzyme has adverse influence.In some embodiments, the nuclear core particle has the average diameter of the about 100 μ m of about 200nm-, and this depends on the desired use that is immobilized the particle after enzyme coats.

The method for preparing coating particles

Mix with the suspension that comprises at least a nuclear core particle, immobilization material and liquid medium by the solution that will comprise enzyme or organelle, and the gained mixture is carried out spray drying, can prepare coating particles.Specifically describe this solution, suspension and spray drying step below.

The enzyme solutions that comprises enzyme and solvent is used to the coating process.Perhaps, in the coating process, use the organelle solution that comprises organelle and solvent.Enzyme merges with group of solvents and mixes up to forming solution.Acceptable enzyme and organelle have been specifically described above.Solvent can be the aqueous solution, preferably cushioning liquid, for example acetate buffer solution or PBS.The pH of cushioning liquid is designed to provide acceptable pH value for treating immobilized specific enzyme or organelle.In addition, in multiple embodiments, enzyme solutions can comprise electron mediator as mentioned above.

Mix and to prepare described suspension by examining core particle, required immobilization material and liquid medium.Exemplary nuclear core particle and immobilization material have been described in the front.Liquid medium can be solvent or cushioning liquid, for example acetate buffer solution or PBS.When using cushioning liquid as liquid medium, the pH of cushioning liquid is selected to provide acceptable pH value for treating immobilized specific enzyme or organelle.

After enzyme or organelle solution and suspension preparation were good, they may be incorporated in together and fully mix.The gained mixture is dried then.Preferred drying means is a spray drying, because this drying means also causes the coating of immobilized enzyme layer to the nuclear core particle.Can use conventional spray drying technology in the method for the invention.Comprise other common process that is used to form coating particles as spray-dired replacement scheme, for example fluidized bed prilling, spray drying granulation, rotating granulation (Rotogranulation), fluid bed/spray drying granulation, extrude with round as a ball etc.

In some other embodiment, solution comprises the enzyme of the about 15wt% of about 0.1wt%-and the solvent of the about 99.1wt% of about 85wt%-, and suspension comprises the enzyme immobilization material of the nuclear core particle of the about 50wt% of about 0.1wt%-, the about 10wt% of about 4wt%-and the liquid medium of the about 75wt% of about 50wt%-.Other method of preparation curtain coating solution comprises particle and enzyme or organelle are mixed formation suspension in cushioning liquid, adds the immobilization material that dissolves then and finishes mixing; Perhaps disposable mixing all material forms suspension.

In multiple preferred embodiment, the mixture of enzyme, enzyme immobilization material and optional electron mediator can be coated on the support particle by spraying/dry technology.For example, can use spray gun (for example, Paasche VL series) to produce the aerosol of component of mixture and they are advanced towards target.Referring to Figure 48.Aerosol is by using the compressed nitrogen of regulating with about 25psi to produce.To the surface such as the Merlon screen, the distance from the spray gun head end to screen is about 40cm to mixture by gun spraying.Spray gun can move with grating mode, moves down along the Merlon target according to zigzag in vertical direction simultaneously, and applies curtain coating solution.Utilize this process that the coating layer thickness on the screen is minimized, particle in the dry run and the interaction between the particle are minimized.Curtain coating solution on screen dry about 20 minutes is afterwards by big spatula/scraper collection.

Biological anode ink water formulation

When preparation anode ink, the identical points for attention of mentioning above need considering.Whether but electronics transports to enzyme, but electronics need transport and leaves enzyme.Because this reason, the selection meeting of electron mediator is different, but the selection of carbon black may be similar.Shift in order to observe direct electron, preferably use carbon black filler.

Electron mediator is incorporated into the ink formulations that is used for biological anode will helps to reduce needs, shown in embodiment 34 and 35 carbon black filler.For example, used hexamine ruthenium (Hexamineruthenium) (III), but also can use any electron mediator of electron transfer potentiality with expectation.For example, used electron mediator such as dicyclopentadienyl nickel, ferrocene, bipyridine cobalt and the iron cyanide.

GDE based on the TBAB polymer

By being hot-pressed onto on the solid proton conductive electrolyte film, platinum anode catalyst and laccase immobilization negative electrode make MEA.Used film is 0.005 inch thick Nafion

Membrane N115 (deriving from E.I.Du Pont de Nemours).Then on MEA and below Kapton polyimide film in the covering, this be because Kapton polyimide film durability at high temperature with and can prevent that MEA from adhering on the steel plate.By under 125 ℃, 3000 pounds pressure, using two block plates hot pressing anode and negative electrode, wherein Nafion simultaneously

Film forms the 5cm that is covered by Kapton between two electrodes ²MEA.The part that press-fits of this MEA illustrates in Figure 66.

GDE based on chitosan polymer

Make MEA by two steps, suppress the platinum anode catalyst earlier on the one side of solid proton conductive electrolyte, and then compacting laccase immobilization negative electrode is to the another side of solid proton conductive electrolyte.Used film is the Nafion of 5 mil thick

Membrane N115 (deriving from E.I.DuPont de Nemours).Then on MEA and below Kapton polyimide film in the covering because the Kapton polyimide film at high temperature durability and can prevent that MEA from adhering on the steel plate.For first step, under 125 ℃, 3000 pounds pressure, only the platinum anode electrode is hot-pressed onto Nafion

On the film.The part that press-fits of first step illustrates the A in Figure 67.Second step is the chitosan-immobilized laccase cathode electrode of hot pressing under 85 ℃, 3000 pounds pressure.The part that press-fits of second step illustrates the B in Figure 67.Because shitosan thermal decomposition under the temperature more than 85 ℃, this two process are necessary.

Electronic controller and the power circuit that replenishes

In the embodiment that illustrates, fuel cell system 1 comprises one " folding " 8 briquette fuel batteries, and each piece battery comprises fuel storage pond 41 and relevant anode assemblies and cathode assembly.Yet, should be appreciated that the number of this battery can be 1 arbitrary number that arrives greater than 1.

Figure 14 shows a kind of embodiment of equipment 1 with the form of block diagram.As shown in the figure, one or more fuel cells 321 are electrically connected electronic controller 71 by line 323.Battery 321 can be stacked as mentioned above.For example, line 323 can comprise flat ribbon cable connector or analog, thereby is 16 electrical connections that folded 8 batteries are provided to controller 71.Electronic controller 71 is controlled the output of each battery 321 according to the operational mode of hope.In the first predetermined running pattern, each battery 321 is connected in series, and controller 71 is controlled their Push And Releases together according to predetermined duty ratio (duty cycle) (for example at 1000Hz, 25%).Advantageously, can improve fuel cell performance according to predetermined duty ratio operation battery 321.Particularly, the stability that battery 321 Push And Releases can improve long-term use is switched in circulation, as measured by the current attenuation under the voltage of setting.In addition, the cycling switch battery 321 by this way, switch to open-circuit condition by giving 321 times of battery from load condition, can improve the power output of equipment 1.At open-circuit condition, the catalyst layer of battery 321 can have bigger reactivity and not oxidized, and this causes subsequently power output bigger under load.

In the second regulation operational mode, the output of controller 71 control batteries 321 makes a battery disconnect and being connected of load 5, and being connected of other battery maintenance and load 5 (for example for 8 battery pack, close 1 battery and open 7 batteries).In this embodiment, allow each battery 321 periodically to be in open-circuit condition, and all the other batteries are in load condition, this has improved the stability and the power output of equipment equally, thereby has improved fuel battery performance.

As mentioned above, the power circuit 81 of Bu Chonging can provide the normal output of power with ancillary equipment 1 as required.This situation (for example electronic equipment start or when powering up (powerup)) of drawing more substantial electric current in load 5 is particularly useful down.In exemplary embodiment, replenish power circuit 81 and move as the part of controller 71.The voltage of controller 71 monitoring batteries 321 is exported, and by comparator output voltage and minimum output reference voltage (for example 1.5V) is compared.If monitored output voltage is fallen under this threshold value, the battery auxiliary circuit just opened by controller 71 or hybrid circuit comes supplemental capacity output.In this embodiment, replenish power supply (battery (battery) 325 for example, but for example the lithium ion battery of recharge) thus provide power to replenish the output of battery 321 by line 327 to controller 71.Although illustrated additional power circuit 81 is parts of controller 71, be appreciated that it also can be the independent circuits that is connected with controller 71 with battery 325.

In the operational mode of the 3rd regulation, controller 71 all fuel cells 321 of cut-out are connected with load 5.This pattern can be used to utilize the output voltage of battery 321 that battery 325 is recharged.

The boost pressure circuit (boost circuit) 331 of the output that is used for regulating cell 321 (or the mixing of battery 321 and battery 325 output) that provides by line 329 also is provided Figure 14.Regulation and control by this way, fuel cell system 1 is supplied with constant relatively voltage at its output, and this output usually is labeled as 333 (in addition referring to Fig. 2 and Fig. 3), can be used for various load specification.Should be appreciated that output 333 can be connected with load 5 hard wires, and be connected to fuel cell system 1 by devices such as plugs; Perhaps conversely.In the embodiment that illustrates, fuel cell system 1 comprises capacitor 335, capacitor 335 chargings when system " dormancy ".Advantageously, capacitor 335 provides the extra supplemental capacity except that battery 325, or replaces battery 325 that supplemental capacity is provided.As an example, boost pressure circuit 331 comprises a DC-DC voltage up converting circuit, and the 5V voltage of modulation is provided to load 5 at its output.

Referring now to the block diagram of Figure 15, it illustrates in greater detail the each side of controller 71.As shown in the figure, processor (for example first microprocessor 337) control first switching circuit 339, the first switching circuits 339 receive the output of battery 321.Microprocessor 337 object computer executable instructions are used for coming actuating battery 321 according to the operational mode of hope.According to operational mode, switching circuit 339 response the microprocessors 337 and output of each monocell 321 is opened or closed.In one embodiment, switching circuit 339 comprises a plurality of single-pole double-throw switch (SPDT)s, its each all be connected with a battery 321, be used for optionally its output being connected with load 5 (perhaps disconnecting).Replenish power circuit 81 and comprise second microprocessor 341 and second switch circuit (usually being labeled as 343).The voltage of second microprocessor, 341 monitoring batteries 321 is exported (from switching circuit 339, online 329 places), and output voltage and minimum output reference voltage are compared.If monitored output voltage is fallen under the lowest threshold, microprocessor 341 control switch circuit 343 are electrically connected to load 5 with battery 325, thereby replenish the output of battery 321.Multiple microprocessor, for example comprise a kind of from the obtainable Programmable System of Cypress SemiconductorCorporation on Chip (programmable system on chip) mixed signal array control unit CY8C29XXX series, be suitable as microprocessor 337, a kind of from the obtainable Programmable Systemon of Cypress Semiconductor Corporation Chip mixed signal array control unit CY8C21XXX series is suitable as microprocessor 341.In addition, should be appreciated that monitoring and compare controlled function such as output voltage, control switch circuit 339, control switch circuit 343 can be by single processor execution, for example microprocessor 337 or microprocessor 341.

In one embodiment, carry out by the computer (not shown) and to be used to instruct the software of microprocessor 337, thereby come control switch circuit 339 according to a kind of operational mode of biological fuel cell equipment.Figure 16 and Figure 17 have described to show exemplary user interface on computers, and this interface receives user's input is used for 8 biological fuel cell equipment with design and simulation pattern.In Figure 16, output 1-8 represents the state of each battery (for example battery 321 of equipment 1) of fuel cell system.The outer button switch (not shown) that input 1 representative is used for switching between plurality of operating modes.The VAR2 response imports 1, represents the operational mode of equipment.For example, push input 1 value and increase to 1, then start first pattern up to VAR2.Under this pattern, output 1 is set to predetermined duty ratio (for example at 1000Hz, 25%).Because battery 321 tandem electrical ties, all batteries 321 are according to predetermined duty ratio opening and closing together.When the VAR2 value was 0, microprocessor 337 will be with the second mode control switch circuit 339.When the value of VAR2 increased to 0 (or " pass "), output 1 was set to another kind of predetermined duty ratio (for example at 1000Hz, 10%).In addition, VAR1 opens and starts phase shift (phase shift).When moving with 10% duty ratio, this phase shift provides enough delay to make in the embodiment that illustrates in 8 batteries 321 7 open and in 8 batteries 321 1 closes.Pent concrete battery 321 by turns, makes each battery can be closed the approximately uniform time in whole battery group.User interface shown in Figure 17 is corresponding to the simulation of this second operational mode, and wherein VAR2 is " pass " and VAR1 is " opening ".In the 3rd operational mode, the VAR2 value increases to 2, and this will close all output 1-8.This third pattern can be used to utilize the output voltage of battery 321 that battery 325 is recharged.

Above-described operational mode (comprising duty ratio) all is exemplary, those skilled in the art understand: depend on that the requirement of load 5 and the physique of fuel cell system 1 (for example number of battery cells) can define other operational mode, and this can not depart from scope of the present invention.

With reference to Figure 18, show electronic building brick (usually be labeled as 345, also can referring to Fig. 3), it comprises first printed circuit board (PCB) 347, and electronic controller 71 is installed on the circuit board 347, comprises replenishing power circuit 81.In addition, electronic building brick 345 comprises second printed circuit board (PCB) 349, is provided for the electrical connection of battery 325.On second printed circuit board (PCB) 349 metal receiver 353 is installed, has formed pocket or the container of shelving battery 325 when battery 325 is installed.In the embodiment that illustrates, receiver 353 has the spring member 355 of one or more deflection batteries 325, is used for remaining on battery on the appropriate location in the receiver and forms and being electrically connected an of end of battery.Conductive spacer (not shown) on the printed circuit board (PCB) 349 provides and being electrically connected of the other end of battery.Lead (not shown) or other electric installation are electrically connected first printed circuit board (PCB) 347 with second printed circuit board (PCB) 349, thereby battery 325 is connected with controller 71.In addition, a plurality of leads (usually being labeled as 357) or other electric installation can be electrically connected controller 71 with fuel cell 321.

Housing

May expect that housing 91 is multi-part structures, this helps the assembly and disassembly of fuel cell system 1.(Fig. 2 and Fig. 3) in one embodiment, housing comprise the first parts 91A and the second parts 91B, when they lump together, have formed a box, and its solvent is enough to hold closely the stack assemblies of fuel cell system 1.The first parts 91A and the second parts 91B are fixed together by one or more securing members (for example hexagon-headed bolt 275) or other mechanical means in removable mode.When parts 91A and parts 91B were fixed together, the joint between them sealed with sealing gasket or other sealing device (not shown).Housing 91 has at least one opening 277 and is used to allow fuel fluid to be delivered to the inlet 29 in the chamber of confluxing from fuels sources 7, and allows fuel to be delivered to waste material destination 9 from the chamber outlet 33 of confluxing.Thereby having a lot of apertures 281 on the wall of housing 91 allows air to come in and go out enclosure interior so that ventilate and the interior parts of cooling housing.Housing can be molded, mach or otherwise make from suitable feedstock (for example acrylic compounds).

As Fig. 3 and shown in Figure 13, projection 285 on housing parts 91A and the 91B inner surface provides the support effect of electrode structure to fuel cell system (be fuel conflux chamber 15, cathode assembly 51,53 and anode assemblies 45,47), thereby provides suitable location and support in enclosure interior for these parts.In these projections 285 some also can define the position that is used to receive electronic controller 71 and power circuit 81, and for example compartment 289,291.Alternatively or in combination, shell inner surface also can form groove and be used to receive these parts.

The operation of equipment

In when operation, utilize suitable means (for example syringe or pump) fuel to be delivered to from fuels sources 9 fuel cell system 1 is full of up to fuel storage pond 41 and fuel fluid contacts with respective anode 157.As the electrochemical cell of standard, anode is the site that fuel fluid generation oxidation reaction discharges electronics and proton simultaneously.Electronics is directed to some current consuming apparatus from anode through electric connector.Proton moves through fuel fluid and polyelectrolyte film arrives negative electrode.Electronics moves through equipment and arrives another electric connector, and this electric connector is used to the negative electrode of electron transport to biological fuel cell reduction-oxidation agent (being the oxygen from air in this example) at this electronics with proton and generates water.By this way, biological fuel cell of the present invention can be as the energy () source of its external electric load.In order to promote the redox reaction of fuel fluid, electrode comprises collector body, gas diffusion layers (electronic conductor), optional electron mediator, the optional eelctro-catalyst that is used for electron mediator, enzyme and enzyme immobilization material.

Electron mediator is the compound that can accept or provide electronics.In preferred this biological fuel cell, the oxidised form of electron mediator is in biological anode and fuel fluid and the oxidised form of enzyme reaction generation fuel fluid and the reduction form of electron mediator.Subsequently or simultaneously, the reduction form of electron mediator and the oxidised form of the oxidised form of eelctro-catalyst reaction generation electron mediator and the reduction form of eelctro-catalyst.The reduction form of eelctro-catalyst subsequently at the oxidized generation electronics of biological anode to form.Redox reaction (except the oxidation of fuel fluid) at biological anode all can be reversible, so enzyme, electron mediator and eelctro-catalyst are not consumed.Randomly, be used to the reactant that provides extra if add electron mediator and/or eelctro-catalyst, these redox reactions can be irreversible so.

Perhaps, can use electronic conductor and enzyme, wherein and biological anode contacting electronic amboceptor can be in metastatic electron between its oxidised form and the reduction form at unmodified electrode (unmodified electrode).If electron mediator can be in metastatic electron between its oxidised form and the reduction form at unmodified biological anode, then electron mediator subsequently and the reaction between eelctro-catalyst no longer are necessary, and electron mediator self can be oxidized to produce electronics and to produce thus at biological anode place.

At negative electrode, the electronics that is derived from biological anode flows into cathode current collector and gas diffusion layers.There, electronics contacts with catalyst, and this catalyst can obtain electronics and generate the oxidised form and the water of catalyst with oxidant reaction from gas diffusion layers.

Other fuel cell embodiment

Figure 19 illustrates the fuel cell system of second embodiment of the invention, and its integral body is marked as label 501.Fuel cell system 501 is similar to embodiment described above substantially, but some feature differences are arranged, and these are discussed in more detail below.The property class of the part of the equipment 501 of second embodiment and the equipment 1 of first embodiment seemingly, but these parts will with similar 500 the labelled notation that adds of first embodiment.

The fuel cell system 501 of Figure 19 is split so that each parts of equipment 501 are shown.Generally speaking, equipment 501 comprises the fuel chamber 515 of confluxing, and this chamber of confluxing has front side 521, rear side 523, be used for accepting the import 529 of fuel fluid and the fuel fluid that is used to give up flows to the outlet 533 (Figure 20) of waste material destination (not shown) from the chamber of confluxing from the fuels sources (not shown).The chamber 515 of confluxing comprises that 4 respectively are labeled as 541 fuel storage pond.This fuel cell system 501 also comprise be used for the fuel reaction in fuel storage pond 541 be labeled as 545 anode assemblies and adjacent lid 561 usually be labeled as 551 cathode assembly usually.The operation that provides the electronic controller (not shown) to come control appliance 501, and comprise that replenishing the power circuit (not shown) provides the normal output of power with the postcombustion battery as required.Provide at the rear side 523 of equipment 501 and can see through film 593 and protective panel 595.Above-described parts make fuel cell system 501 show as from the compact unit of holding.

As shown in figure 20, the fuel chamber 515 of confluxing has import 529 and outlet 533, and import 529 allows fuel fluids to flow into the chamber of confluxing from the fuels sources (not shown), and outlet 533 allows fuel fluids to leave to conflux the chamber to flow into waste material destination (not shown).Import 529 and outlet 533 can comprise and the chamber 15 similar pipe joints (tubing nipple) that conflux that are shown in first embodiment of Fig. 4.As shown in figure 20, the chamber 515 of confluxing comprises 4 fuel storage ponds 541.The fuel chamber 515 of confluxing comprises the body or the block 601 of suitable insulative materials, and it has top 605, bottom 607, opposite end 609, front 611 and the back side 615.This block 601 can be by being that chemically inert polymer-based material or non-polymeric substrate material form to used fuel.In one embodiment, this block 601 is formed by acrylic material.This block 601 can be shaped (for example molded, machining etc.) be rectangle, and preferably constitute by single integral type member.Fuel storage pond 541 is defined by the chamber in the front 611 of this block (also being labeled as 541).In the alternative embodiment (not shown) of another kind, the chamber that defines fuel storage pond can form at the front 611 and the back side 615 of block 601.In one embodiment, each chamber has about 1.74 cubic centimetres volume, but fuel storage pond 541 also can have other volume and can not depart from scope of the present invention.Fuel enters each fuel storage pond 541 and leaves same storage pond by the port of export 625 by entrance point 621.Each entrance point 621 and the corresponding port of export 625 are positioned at the opposite end of runner (being generally referenced as 627), and runner 627 extends through the inwall 637 of compartment 541.As shown in figure 20, the height of each port of export 625 is the same with the top of each entrance point 621 at least high.This layout makes and can store up pond 541 by complete filling fuel, do not contact (stating as follows) and do not have air bubble with cathode assembly with anode assemblies.Thereby in each runner 627, between the entrance point 621 in the port of export 625 in fuel storage pond 541 and next fuel storage pond 541, the check-valves (not shown) is set and prevents that fuel from flowing back to another storage pond from a storage pond, eliminate any ion communication between these batteries thus, thereby prevented short circuit.In one embodiment, each check-valves is to have 0psi cracking pressure (cracking pressure) stainless steel check-valves, and it is fitted in the runner of 2.5mm, but also can use other check-valves and can not depart from scope of the present invention.

This chamber of confluxing comprises that compartment 645 is used to receive electronic controller and power circuit.As understood by one of ordinary skill in the art, can form around the front 611 of block 601 that antelabium 647 is used for and lid 561 engagements.The dorsal part in contiguous each chamber 541 provides opening 649, discharges with the carbon dioxide that will form in the course of reaction.Confluxing also provides pore in the chamber 515 and enters fuel cell for air.The further feature and first embodiment in the chamber 515 of confluxing of second embodiment are similar, repeat no more.

Figure 21 shows anode assemblies 545, the fuel reaction in anode assemblies 545 and the fuel storage pond 541, cathode assembly 551 contiguous anode assemblies.Anode assemblies 545 comprises printed circuit plate frame body 651.This framework 651 has a plurality of openings 655, they be configured and be arranged to and the front 611 in the chamber 615 of confluxing in the configuration and the layout in fuel storage pond 541 be complementary.Anode assemblies 545 also comprises a plurality of anodes, and each anode generally all is labeled as 657, remain in the framework opening 655 by framework 651, and anode of each framework opening.Each anode 657 comprises the collector body 661 that is fixed in the corresponding framework opening 655.In one embodiment, collector body 661 comprises two gold-plated stainless (steel) wire sheets spot-welded together.The size of a net sheet can accurately match it with framework opening 655, the size of another net sheet makes itself and opening overlapping.Collector body 661 is spoted weld on the pad (pad) 663 of the opposite edges portion that is printed on opening 655.When collector body 661 is spoted weld on the pad 663, welding parameter importantly to be set make welding machine not puncture (arc).Use CD250DPSunspot-b dipulse spot welding machine, use following parameter that acceptable result can be provided: energy grade-14, pulse 1-12, pulse 2-45, welding machine energy-14.These parameters provide the welding of mechanically stable, and can not damage collector body 661.

Each anode 657 also is included in the gas diffusion layers 665 on collector body 661 back sides.This diffusion layer 665 can be the carbon paste structure, this carbon paste structure comprises the carbon (C*Chem) of the steam activation of dried component: Monarch 1400 carbon blacks (Cabot) and Chemsorb 1505G5 porous dipping, and wet component: be dissolved in the acetone Quick Set 2-part epoxy resin (The Original Super GlueCorp.) and in water 60% polytetrafluoroethylene (PTFE) dispersion (Sigma).To do component and grind in the food grinder, and mix wet component with ultrasonic homogenizer, then merging is done wet component and with putty knife it is mixed, and has the denseness of toothpaste until mixture.Then by being applied to the enzyme in the cushioning liquid and the Nafion of TBuA modification

Ionomeric curtain coating mixture adds the catalyst layer (not shown) to gas diffusion layers 665.Before being attached on the anode assemblies, catalyst layer is dried.In preferred embodiments, used enzyme is a PQQ dependent form alcohol dehydrogenase (PQQ-ADH) in the catalyst layer.As shown in the figure, anode adds hot melt layer 675 and vinyl framework 677.

The method of describing at anode assemblies 545 above each cathode assembly 551 usefulness is similar to is constructed.Cathode assembly 551 is also included within the layered catalyst structure on its collector body back side.This layered catalyst structure comprises gas diffusion layers, catalyst layer and polyelectrolyte film (Nafion

).When assembling, this layered catalyst structure is fixed on the back side of collector body.In a specific embodiment, by comprising platinum black catalyst and Nafion

The side that ionomeric ink is applied to gas diffusion layers (for example, as the LT2500W Low Temperature Elat that has microporous layers at either side E-Tek) prepares this catalyst stratiform structure.In case comprise after the ink dried of catalyst, with regard to following assembling stratiform catalyst structure: gaseous diffusion is placed on the collector body top, makes on the platinum black catalyst side direction, then with polyelectrolyte film (Nafion for example

) place on the top of gas diffusion layers and catalyst layer.This assembly is soaked in water to be fixed together by hot pressing then, obtains independently negative electrode accessory (for example, under 125 ℃, 3000 pounds pressure 35 seconds).Polyelectrolyte film (Nafion for example

Ionomer) be impermeable to fuel fluid, but can conduction electron and proton.

After preparing anode assemblies 545 and cathode assembly 551 as mentioned above, in the following way anode assemblies 545 and cathode assembly 551 are fitted together: smear welded disc (connector pad) 731 on each printed circuit plate frame body 651 with soldering paste or common spooled solder.Then eight conductor flat cables 735 are welded in one temporarily and determine on the circuit board, reserve the edge that about 0.020 inch bare exposed conductor is stretched out circuit board.Second circuit board alignd with first circuit board and cable, and firmly it is remained in the appropriate location.Thereby each conductor contacts with flatiron the soldering paste on the upper and lower circuit board/scolder refuse then, forms good electrical contact thus between circuit board and cable.Check the continuity (continuity) between each the installation pad that is used for expanded metal on flat flexible cable far-end and the framework then.Also check the short circuit between each conductor.On demand, this assembly/conductor can be by refuse up to realizing true(-)running.Welded disc 731 off-center make the pad of a circuit board and another circuit board pad and do not line up, but they all with a conductor alignment of cable 735.

Before assembling, remove solder mask (solder mask), to guarantee to obtain leak free stacked body.The removal of solder mask is finished by the following: printed circuit board (PCB) is immersed in the dichloromethane solution up to whole solder mask fluffs and can remove easily.Remove after the solder mask, circuit board is taken out and use the carrene cleaning down from dichloromethane solution.Then at other dry circuit board must parts time of preparation.

Anode assemblies 545 and cathode assembly 551 each self assembly and welded together after, they are pressed (for example, under 125 ℃, 1000 pounds pressure about 10-about 20 seconds).If desired, the stacked body of being finished can be pruned, and by the binding agent combination be sealed on the housing (for example, hot melt and under 2000 pounds of pressure compacting up to cooling).After the cooling, just anode reactant can be joined fuel storage pond.

Carbon dioxide permeable membrane 593 covers the opening 649 that is positioned at fuel storage pond 541 dorsal parts.Remove the gases that generate in the fuel storage pond 541 and can not allow unreacted fuel to leave fuel storage pond 541 thereby film 593 optionally allows carbon dioxide to see through.The example that can optionally see through the polymer film of carbon dioxide comprises polysiloxanes, styrene analog thermoplastic elastomer, the thermoplastic elastomer (TPE) based on polyamide, the thermoplastic elastomer (TPE) based on polybutadiene, EPDM rubber-based thermoplastic elastomer (TPE).Since to carbon dioxide very highly select permeability, the carbon dioxide that generates during fuel cell operation can be discharged by loose structure, and loose structure can be used as the barrier of pure fuel.

The application of coating particles and purposes

Can be with the particle that immobilised enzymes coats as the catalyst of number of chemical conversion.Enzyme can be fixed in the enzyme immobilization material and be coated on the particle and deposit in the multiple substrate with technology described herein.The chemical conversion that these immobilised enzymes particles can contact with reactant mixture then and catalysis is expected.Replace enzyme in the embodiment that particle of the present invention can be described below.

Especially, following immobilised enzymes particle can be used to catalytic reaction, and wherein the enzyme immobilization material is fixed and stabilized enzyme, and the enzyme immobilization material is permeable for the compound littler than enzyme and is micellelike hydrophobically modified polysaccharide.In multiple preferred embodiment, micellelike hydrophobically modified polysaccharide is hydrophobically modified shitosan or hydrophobically modified alginates.

In addition, the immobilised enzymes particle can be used to detect analysans.Fixing and the stabilized enzyme of enzyme immobilization material, the enzyme immobilization material is permeable for the compound littler than enzyme and is micellelike hydrophobically modified polysaccharide.In multiple preferred embodiment, micellelike hydrophobically modified polysaccharide is hydrophobically modified shitosan or hydrophobically modified alginates.

For example, the enzyme spot in the clothing dirt that can be used to degrade is for example decomposed in washing agent and is removed deproteinize from clothing.Usually, the enzyme that is used for washing agent is protease, amylase, carbohydrase, cellulase and lipase.These enzymes can be fixed in enzyme immobilization material described herein and be stable, and are scattered in the washing agent or provide the clothing dirt to handle in the aqueous carrier.These enzyme immobilization materials can make enzyme stablize and improve storage stability in other component of washing agent.Multiple enzyme-containing detergent product is described in United States Patent (USP) 7,179, and in 780,6,894,013 and 6,827,795, these patent documentations are included in here by reference.

In addition, enzyme can be used to waste water treatment and decomposes various refuses in the current.For example, except bacterium, also use lipase, cellulase, amylase and protease to remove various refuses.The degrade suitable enzyme of this waste water stream of the component of the known various concrete waste water streams of those skilled in the art and being used to.For example United States Patent (USP) 7,053, and 130,6,802,956,5,531,898 and 4,882,059 discloses the multiple enzyme that is used for waste water treatment, and these patent documentations are included in here by reference.These enzymes can be replaced improving enzyme stability by the particle that contains these enzymes of the present invention.

Another example of the industrial use of enzyme is to be used for corn or other cereal are converted into high-fructose corn syrup.Enzyme is used in three procedure of processings of high-fructose corn syrup, and these steps are: starch is converted into sugar in the saccharification of the liquefaction of corn or cereal, corn or other cereal and glucose isomerase turns to fructose.Especially, to be used to conversion of glucose be fructose to glucose isomerase.These enzymes that are used can advantageously be fixed in the enzyme immobilization material described herein.The immobilization of this kind of enzyme is favourable, and sex change does not take place because enzyme can be stabilized, thereby and enzyme can easily be removed and cause technology more controlled.Equally, the immobilised enzymes particle can easily be separated from the high-fructose corn syrup product.These procedure of processings are described in greater detail in United States Patent (USP) 5,593, and in 868 and 4,567,142, these patent documentations are included in here by reference.

In addition, enzyme can also be used for food processing, because the multiple reaction of these arts demand enzymatic protein.For example, in baking process, fungal amylase, hemicellulase, pentosanase, zytase, protease, amylopectase and acid protease are used to various purposes.In a kind of technology, fungal amylase is used to modified wheat flour and is suitable for curing and producing the more dough and the product of homogeneous.In addition, produce the shelf life that maltogenic amylase is used to prolong polytype bread, amylopectase is used as the antistaling agent in the baked product.In brewage process, amylase, protease, 1,4 beta-glucanase and acetolactate decarboxylase are used to a plurality of steps.For beer, some in these enzymes be introduced into and promote the conversion of starch to fermentable sugars in the Fructus Hordei Germinatus, thereby remove the cold muddiness of beer (chill haze) and improve filtration.For cheese and and the production of whey, the maturation stage of coagulating change and cheese that protease (for example, pepsin) and lipase are used to suckle, lactase is used to produce whey syrup.In addition, the isoflavones phytoestrogen in the beta-glucosidase enzymatic conversion soymilk.In these enzymes each can be fixed in the edible immobilization material and is advantageously used for these catalyst for reaction by method described herein.These technologies are described in greater detail in United States Patent (USP) 7,014, and in 878,6,936,289,6,830,770,4,358,462 and 6,372,268, these patent documentations are included in here by reference.

The number of chemical synthesis technique utilizes enzyme to carry out the ester exchange of the synthetic and oil of esterification, chirality and comes unstuck.For example, multiple carboxylic acid compound (comprising the polymer with carboxylic acid side group) can utilize plurality of enzymes by pure esterification.The enzyme that is used for catalytic esterification is hydrolase, particularly lipase, protease and esterase normally.Schematically enzyme comprises antarctic candida (Candida antarctica) lipase B (being produced by Novozyme), rice black wool mould (Mucor meihei) lipase IM, Pseudomonas cepacia (Pseudomonas Cepacia) lipase PS-30, pseudomonas aeruginosa (Pseudomonasaeruginosa) lipase PA, Pseudomonas fluorescens (Pseudomonas fluoresenses) lipase PF, aspergillus niger (Aspergillus niger) lipase, with column Candida (Candida cylinderacea) lipase from pig pancreas lipase.The details more specifically of these reactions is described in United States Patent (USP) 7,183, and in 086 and 6,924,129, these patent documentations are included in here by reference.These enzymes can be replaced improving enzyme stability by the particle that contains these enzymes of the present invention.

For chemical synthesis, plurality of enzymes can the catalysis number of chemical transform.For example, glutaminase is used to glutamine is converted into glutamic acid; Penioillin acylase is used to chemical synthesis; It is synthetic that chloroperoxidase is used to steroids; Aspartic acid β-decarboxylase is used to make the L-alanine from the L-aspartic acid; Cyclodextrin glycosyl transferases is used to make cyclodextrin from starch.In addition, the synthetic enzyme that also uses of multiple chipal compounds.Subtilopeptidase A is used to the chiral resolution of chemical compound or medicine; Amino-acylase can optical resolution amino acid; Alcohol dehydrogenase is used to the chirality of chemicals and synthesizes; Amino acid oxidase is used to the fractionation of racemic amino acid blend.The details more specifically of some of these reactions is described in United States Patent (USP) 6,036, and in 983,5,358,860,6,979,561,5,981,267,6,905,861 and 5,916,786, these patent documentations are included in here by reference.These enzymes can be replaced improving enzyme stability in the chemical synthesis process by the particle that contains these enzymes of the present invention.

Utilize plurality of enzymes, oil can and come unstuck by ester exchange.For example, enzymatic interesterification procedure is the effective ways of the melting characteristic of the edible oil ﹠ fat of control.This is to realize by the degree of controlling conversion/reaction.Do not use chemicals in this method, and can as in other production technology, not produce trans fats.Immobilized lipase can be used to the aliphatic acid in the oil ﹠ fat of producing margarine and shortening is carried out ester exchange.In addition, enzymatic method can be used to remove the colloid in the vegetable oil; This method is commonly referred to as comes unstuck.Enzymatic degumming uses phosphatidase hydrated lecithin (colloid) not to be converted into water miscible lysolecithin, and lysolecithin can be by centrifugation.These technologies are described in United States Patent (USP) 6,162, and in 623,6,608,223,7,189,544 and 6,001,640, these patent documentations are included in here by reference.These enzymes can be replaced improving ester exchange by the particle that contains these enzymes of the present invention and the process of coming unstuck in enzyme stability.

In addition, the immobilised enzymes particle can be used for biology sensor.Usually, these biology sensors are used for diagnostic method and detect various analysans at complex mixture (for example body fluid and cuts).For example, multiple sensors can be used for detecting urea, uric acid and the cholesterol of various body fluid.United States Patent (USP) 5,714 has been described multiple sensors in 340, and this patent documentation is included in here by reference.

The immobilised enzymes particle can be used for the multiple method of inspection, and whether and/or its concentration the existence that is used to measure multiple compound in the body fluid (for example, saliva, blood or urine) or organism.These methods of inspection use various enzymes to come to interact with analysans and enzyme inhibitor; These interact and allow the detection from one or more analyte molecules or organism to form or change into a large amount of molecules.Thus, the concentration of enzymatic activity and analysans is proportional, and allows to detect the analysans of low concentration.Analysans can be that micromolecule agricultural chemicals and other toxin arrive microbe.Can use specific analysans caught and detect the multiple check probe with high specific.Use the method for inspection that enzyme amplifies at United States Patent (USP) 6,171, describe to some extent in 802,6,383,763 and 4,067,774, these patent documentations are included in here by reference.

In addition, the immobilised enzymes particle can be used for the purifying of material and separating of mixture component.For example, enzyme can be used for from racemic mixture separating chiral molecule and isomers; Be used for removing desulfuration from oil, gas or other raw material of industry; Be used for decomposing and separating the lignocellulosic component from plant; Be used for handling and purifying liquid waste; Be used for processed food and remove undesired compound.

From oil, gas or other raw material of industry, remove the process of desulfuration and use plurality of enzymes.These enzymes are commonly called desulfurase.A kind of important desulfurase is desulfinase (desulfinase).The thieno that these desulfurases decompose in the various oil products was therefrom removed sulphur atom before the feedstock oil burning.These technologies are described in United States Patent (USP) 6,461, and in 859 and 7,045,314, these patent documentations are included in here by reference.These enzymes can be replaced improving the stability of enzyme in sweetening process by the particle that contains these enzymes of the present invention.

Lignin from various plants can be degraded by lignin peroxidase.The multiple oxidation reaction of lignin peroxidase catalysis obtains multiple pyrolysis product and other oxidation product.These oxidation reactions comprise that the carbon-carbon bond fracture of the propyl side chains of lignin, hydroxylating, the benzyl alcohol of benzyl (benzylic) methylene are oxidized to corresponding aldehydes or ketones and phenol oxidation.The details of these processes is described in United States Patent (USP) 7,049, and in 485, this patent documentation is included in here by reference.Thereby lignin peroxidase can the method according to this invention be immobilized the raising enzyme stability.

Thereby can use pectase, amylase and cellulase to handle fruit juice decomposes the various structures of fresh fruit cell and strengthens juice extraction technology.The polysaccharide that discharges from cell in the fruit course of processing is insoluble fruit juice, and makes fruit juice seem muddy.In order to make juice clarification, can use pectase and amylase to decompose these soluble polysaccharide and obtain soluble sugar.The fruit juice that this technology obtains clarifying, and owing to introduced soluble sugar and make fruit juice sweeter.These technologies are at United States Patent (USP) 5,585, more detailed description are arranged in 128,5,419,251 and 4,971,811, and these patent documentations are included in here by reference.These enzymes can be replaced improving enzyme stability in this type of technical process by the particle that contains these enzymes of the present invention.

Active surface film (Active surface film) is to comprise the coating that can carry out automatically cleaning or remove the reactive materials and/or the enzyme of various toxin from the surface with undesired pollutant reaction.For example, the active surface film can be included in and come minute toxolysin or sterilization in packaging for foodstuff, fiber, superficial layer, bandage or the filter.For example, can the degrade enzyme of various toxin can be included in the toxin that destroys in the fiber as biological weapons.In addition, the enzyme that this fiber could be able to be killed or destroy bacterium applies, thereby makes the wearer avoid bacterial infection.These enzymes can be replaced improving the enzyme stability that is used in this type of active surface film by the particle that contains these enzymes of the present invention.

In addition, can immobilization provide the multifunctional material of energy catalysis more than a kind of reaction more than a kind of enzyme.For example, the combination of above-described enzyme can be immobilized in one or more enzyme immobilization materials, thereby the product of energy catalysis more than a kind of reaction is provided.

Use immobilised enzymes described herein, can prepare and the similar electrode ink of conventional fuel cell ink.At first, enzyme is immobilized material by spray drying technology described herein and seals.Then, carbon that the enzyme after the stabilisation is coated and carbon filler, 5%Nafion

The solution mixing is coated onto on the electrode holder then.Dried electrode can be pressed onto on the exchange membrane with respective anode then, can be as H ₂/ O ₂Fuel cell equally operates.

When considering carbon black as enzyme during the filler in the catalyst based ink, whether can influence the overall performance of electrode with doped polymer grafting, conductibility and surface area.Detected several carbon blacks and can improve battery performance as filler, Monarch 1400 has shown to be the most maneuverable ink and to have best performance.But any carbon black can substitute the interaction that Monarch 1400 improves enzyme and collector body.All carbon blacks that mix with the enzyme of sealing all do very well than independent enzyme, but should consider that every kind of carbon black has different parameters optimization.

Many aspects of the present invention relate to the purposes that immobilized enzyme is used for catalytic reaction, and wherein enzyme is immobilized in the enzyme immobilization material.Fixing and the stabilized enzyme of enzyme immobilization material, the little compound of contrast enzyme is permeable, and can be the polysaccharide of micellelike hydrophobically modified.

Others of the present invention relate to the purposes that immobilized enzyme is used to detect analysans, and wherein enzyme is immobilized in the enzyme immobilization material.Fixing and the stabilized enzyme of enzyme immobilization material, the little compound of contrast enzyme is permeable, and is the polysaccharide of micellelike hydrophobically modified.

Others of the present invention relate to the purposes that immobilized enzyme is used for catalytic reaction, and wherein enzyme is immobilized in the enzyme immobilization material.Fixing and the stabilized enzyme of enzyme immobilization material, the little compound of contrast enzyme is permeable.Reaction by catalysis is selected from: (a) carboxylic acid is by pure esterification; (b) liquefaction of corn or other cereal; (c) saccharification of corn or other cereal is converted into sugar with starch; (d) glucose isomerase turns to fructose; (e) chipal compounds is synthetic; (f) You ester exchange; (g) oil comes unstuck; (h) waste water treatment (reaction); (i) juice clarificationization; (j) utilize starch explained hereafter glucose; (k) produce glucose and galactolipin from lactose; (l) synthetic compound with peptide bond; (m) produce the amino penicillic acid of 6-from benzyl penicillin; (n) sugar is converted into alcohol; (o) sulphur removal from petroleum fractions; (p) acrylonitrile is converted into acrylamide; (q) the 3-cyanopyridine is converted into niacinamide; (r) spot in the degraded clothing dirt.

Others of the present invention relate to the improvement of enzymic catalytic reaction, and described enzymic catalytic reaction is selected from: carboxylic acid is by pure esterification; The liquefaction of corn or other cereal; The saccharification of corn or other cereal is to be converted into sugar with starch; Glucose isomerase turns to fructose; Synthesizing of chipal compounds; The ester exchange of oil; Coming unstuck of oil; Waste water treatment; Juice clarificationization; Utilize starch explained hereafter glucose; Produce glucose and galactolipin from lactose; Synthetic compound with peptide bond; Produce the amino penicillic acid of 6-from benzyl penicillin; Sugar is converted into alcohol; Sulphur removal from petroleum fractions; Acrylonitrile is converted into acrylamide; The 3-cyanopyridine is converted into niacinamide; Or the spot in the degraded clothing dirt.Described improvement comprises enzyme immobilization in the enzyme immobilization material, the fixing and stabilized enzyme of described enzyme immobilization material, and the little compound of contrast enzyme is permeable.

Others of the present invention relate to the purposes that immobilized enzyme is used to detect analysans, and wherein enzyme is immobilized in the enzyme immobilization material.Fixing and the stabilized enzyme of described enzyme immobilization material, and the little compound of contrast enzyme is permeable.Detected analysans comprises urea, uric acid, cholesterol, agricultural chemicals, toxin or microbe.

Others of the present invention relate to immobilized enzyme and are used for the purposes of separating or removing material from mixture, and wherein enzyme is immobilized in the enzyme immobilization material.Fixing and the stabilized enzyme of described enzyme immobilization material, and the little compound of contrast enzyme is permeable.

Others of the present invention relate to the purposes that immobilized enzyme is used for chemism film surface, and wherein said film surface is immobilized in the enzyme immobilization material with at least a substance reaction that contacts this film surface and described enzyme.Fixing and the stabilized enzyme of described enzyme immobilization material, and the little compound of contrast enzyme is permeable.

Embodiment

Embodiment 1: biological anode performance and preparation

1. nickel and carbon electrode structure

Being used for the metal of biological anode preparation and the performance that is chosen in this anode of material plays an important role.This is shown in Figure 24, wherein also estimates as 3 electrodes of amboceptor preparation with commercial NAD-dependent form ADH enzyme and methylene green.This electrode is by gold-plated stainless steel or nickel collector body, constitutes as catalyst support layer with the commercial GDL bilateral E-lat of Etek or inner homemade epoxy resin and carbon GDL.The nickel collector body reveals optimum performance with the combination table of inner self-control GDL.

The kind electrode structure also is used for glucose dehydrogenase, and being used for glucose is the biological fuel cell of fuel.The performance of this battery is shown in Figure 25.About Figure 25, the curve that rhombus is represented is 4 batteries (the every battery 4.5cm with this biology anode and air self-respiration type platinum cathode ²) the performance curve of biological fuel cell stacked body.The curve that triangle is represented is to use the performance curve of the existing best battery of commercial GDL.

2. inner homemade gas diffusion layers (GDL)

A principal element that influences biological anode performance is the enzyme catalyst carrier layer shown in previous section.Anode with dried carbon paste carrier layer has optimum performance.The picture of this battery is shown in Fig. 7.This structure is served as catalyst carrier, reactant diffusion layer and electric contacting layer between collector body and catalyst layer.This layer is the mixture of carbon black, mesoporous carbon, PTFE, bi-component (2-part) epoxy resin, and is applied on collector body and the framework with the thickener form that water and acetone make.Before applying catalyst and modification Nafion layer with this thickener drying.With regard to accurate accommodation property, this material has many advantages as catalyst carrier.This GDL can make more hydrophobic or hydrophilic with the ratio of mesoporous carbon by regulating PTFE.Its example is shown in Figure 26.About Figure 26, the GDL material by comprise carbon sieve (carbon sieves) (rhombus is represented) make more hydrophobic, thereby performance is than the improvement that has of using nitric sulfid (triangle is represented) separately.The remainder of battery is stainless steel expanded metal collector body (for negative electrode, gold-plated), and N112 is as film, and 1% ethanol acts as a fuel, and negative electrode is 4mg/cm ²Platinum black.

3. the enzyme that biological anode is used

The another effect is purity, age and the concentration of ADH enzyme in the performance of biological anode.Figure 27 has provided 7 kinds of ADH enzymes, comprises 6 kinds of inner self-control purifying PQQ dependent form ADH enzymes and a kind of commercial NAD dependent form ADH enzyme, representative curve.The enzyme of poor performance is TB1, and it is the oldest in this BT(batch testing) enzyme (4 ℃ of following 6 weeks).That best is ML10, is the freshest in all enzymes when the enzyme of a kind of high-purity and concentration and test.Commercial NAD dependent form ADH enzyme is in the figure as illustrating with reference to point.The performance of this commercialization enzyme is suitable with 4 kinds of inner self-control enzymes.The difference of each electromotive force and 0V (vs NHE) is represented the anode overpotential of each electrode, and lower electromotive force is preferred under the given current density.Optimum performance is sample ML10, and it shows 75mA/cm under 400mV (vs.NHE) electromotive force ²Running.This performance level depends on the quality of used enzyme and the type of type and near the metal ion activation agent (as nickel) that exists of immobilised enzymes.The enzyme of observing purifying under inner (in-house) acid operating condition has optimum performance.

4. biological anodic half-cell performance

Biological anode performance in the half-cell structure, power density, an example be shown in Figure 28.Anode among this figure has 1cm ²Area, and have the 1M H of 5% ethanol ₂SO ₄Test in the solution.The maximum power density that this battery reaches is 15mW/cm ²This performance level depends on the quality of used enzyme and near the type of the metal ion activation agent (as nickel) that exists of type (enzyme through applicant's purifying has optimum performance), operating condition (acidity) and immobilised enzymes.

Embodiment 2: the performance of 4 and 8 cell stack

When described biological anode of Figure 28 and the pairing of platinum base negative electrode, and size is amplified to 4.5cm ²And when being connected in series (4 batteries), overall performance drops to 33% of performance shown in Figure 28.The electric current of 4 batteries series connection stacked bodies is shown among Figure 29 A and the 29B voltage and the electric current representative curve to power.4 batteries (every battery 4.5cm that top figure expression has biological anode and air self-respiration type platinum cathode ²) performance of biological fuel cell stacked body.The average battery voltage and the power density of each monocell of this stacked body of below figure expression.Used anode enzyme is inner oneself preparation and purifying in these batteries.

When two this 4 cell stack were connected in series, performance further reduced by half.The electric current of 8 battery series connection stacked bodies and the representative curve of voltage and electric current and power are shown among Figure 30 A and the 30B.This stacked body is the combination that two 4 cell stack (shown in the picture of front the sort of) are connected in series.8 batteries (every battery 4.5cm that top figure expression has biological anode and air self-respiration type platinum cathode ²) performance of biological fuel cell stacked body.The average battery voltage and the power density of each monocell of this stacked body of below figure expression.Along with the number of battery cells in this stacked body increases, observe low more monocell performance.

Embodiment 3: anode construction

Catalyst carrier-gas diffusion layers (GDL)

The catalyst support layer that is used for anode electrode is made by the mixture of material with carbon element, Teflon, bi-component (2-part) epoxy resin and solvent.By changing the ratio of these components, can accurately regulate the character of this structure.The example of these character comprises: (1) electrical conductivity, (2) hydrophobic/hydrophilic character, (3) surface area and (4) drying time.This mixture is made thickener, then is pressed in framework/collector body structure that Kapton is covered with, and scrape off with medical knife unnecessary.The framework both sides all are covered with, and otch are arranged to expose the collector body of exposure in a side.At this thickener bone dry with before solidifying, remove Kapton adhesive tape mask, make total dry under the halogen heating lamp.

The GDL paste formulation is done component by 3.57g Monarch 1400 carbon blacks (Cabot) and 1.43g Chemsorb1505G5 porous dipping steam activation carbon (C*Chem) and is made.These dried components are packed into impacted 60 seconds in the food grinder.Wet component prepares separately, and they are 5.00g Quick Set bi-component epoxide-resin (The Original Super Glue Corp.), 4.00g acetone and 2.00mL 60%PTFE dispersion (Sigma) in water.Before adding the PTFE dispersion, epoxy resin is dissolved in the acetone.The component that will wet was mixed 15 seconds with ultrasonic homogenizer.Merge dried wet mixture, and have the denseness of toothpaste until mixture with the putty knife mixing.

Catalyst layer

The curtain coating mixture of enzyme catalyst layer as enzyme in the cushioning liquid and TBAB modification Nafion immobilization material is applied on the GDL body structure surface of drying.With before corresponding cathode construction combines, make solution dry and solidify (cure).

The preparation that is used for the enzyme catalyst layer

The curtain coating solution of preparation certain volume is to be provided for 8 battery (4.5cm ²) q.s, and this curtain coating solution casting that will wait branch 0.45mL is to each electrode.Curtain coating solution be the PQQ dependent form ADH of 2.1mL in the PBS buffer solution (～10mg/mL) with the Nafion immobilization material of 2.1mL 5 volume % bromination tert-butyl group ammonium modifications.

Framework and collector body preparation

The parts that are used for anode current collector and frame assembly are shown in Fig. 8.This structure starts from two polyurethane-hot melt spares are melted to a side of each vinyl framework otch.Fusing by with framework with place hot melt part on the top to be exposed to 128 ℃ of temperature to finish in 30 seconds.Then remove hot melt part, and allow its cooling, just touch until binding agent and be clamminess.Wherein wire is placed into by configuration shown in Figure 8 on the frame structure through the porous metals silk screen collector body of braiding and compacting (8 tonnes), and lead-in wire is fixed in the hot melt glue along the framework side.Another framework is placed on the top of collector body/framework/adhesive structure, the colloid side is facing to porous metals silk screen collector body.Then under minimum pressure and 128 ℃ of temperature, should suppress 30 seconds by whole assembly, to seal this structure.

Embodiment 4: cathode construction

Catalyst carrier-GDL and collector body

The catalyst support layer that is used for cathode electrode is commercially available carbon cloth GDL material.E-Tek has microporous layers with this material called after LT2500W low temperature Elat in each side.The collector body that is used for this negative electrode is 0.007, and " thick stainless steel porous web material, this material is cut into certain size, Yi Bian along No. 28 insulated wires with braiding and compacting, to make flag shape structure.Before this cathode electrode of assembling, the conductive portion of the exposure of this negative electrode flag shape thing is carried out acid etching (5M HCl), and carry out electrogilding with Orthotherm HT RTU rack gold bath (Technic Inc.).

Catalyst layer

Cathode catalyst layer is 4mg/cm by carrying capacity separately ²Form with platinum black catalyst and the Nafion ionomer of 8.5wt%.This catalyst is made ink, and be coated onto one even as big as being used for 8 cube electrode (36cm ²) Elat on, make its drying.After the catalyst layer drying, it is 4.5cm that this piece is cut into area ²(each electrode of 3 * 1.5cm), and last assembling such as put aside.

The preparation that is used for cathode catalyst layer

Preparation is 8 battery (4.5cm of preparation enough ²) the curtain coating solution of certain volume.This curtain coating solution is by 158.4mg HiSPEC ^TM1000 platinum black (Alfa Aesar), 380.2 μ L deionized waters, 100.0 μ L 2-propyl alcohol and the 336.7 μ L 5wt.%Nafion ion exchange resin (Sigma Aldrich) in rudimentary aliphatic alcohol/aqueous mixtures is made.Before smearing, curtain coating solution was mixed 15 seconds with ultrasonic homogenizer.

The negative electrode preparation

Negative electrode is made a piece that comprises collector body, GDL/ catalyst layer and Nafion immobilization material.Nafion immobilization material Nafion 212 (Sigma Aldrich) is cut into the piece that is a bit larger tham the GDL layer, then it is wetting with deionized water.Catalyst/GDL is placed on the porous metals silk screen collector body top, coats 5wt%Nafion solution thin layer, and wetting with deionized water.Place wetting Nafion film at the top of this assembly, and this total placed 2 (3 * 3 ") 0.010 " between the thick Kapton (McMaster-Carr).The Kapton film serves as and press-fits and release film.Press-fit part in 3 tonnes pressure and 128 ℃ compacting 90 seconds down with whole, then " cool off between the thick aluminium sheet at 21.Remove the Kapton film, before placing framework, repair size to the 3.75 * 1.7cm of this negative electrode.

Framework preparation and cathode assembly

The parts of cathode current collector and frame assembly are shown in Fig. 8.At first, two polyurethane-hot melt spares are melted to a side of each vinyl framework otch.Fusing by with framework with place hot melt part on the top to be exposed to 128 ℃ of temperature to finish in 30 seconds.Then remove hot melt part, and allow its cooling, just touch until binding agent and be clamminess.4 prefabricated negative electrodes are placed on the frame structure by configuration shown in Figure 8, and wherein going between is fixed in the PUR along the framework side.Another framework is placed on the top of this collector body/framework/adhesive structure, the colloid side is facing to porous metals silk screen collector body.Then under minimum pressure and 128 ℃ of temperature, should suppress 30 seconds by whole assembly, to seal this structure.

Embodiment 5: chamber and fuel bath conflux

Carry the chamber of confluxing for the fuel of this device design and brought benefit for this fuel cell with electrolyte fuel solution, this electrolyte fuel solution can make battery short circuit in conventional liq fuel cell (being direct methanol fuel cell).Fuel is transported to each battery storage pond by a series of one way stop peturn valves between each battery.The use of check-valves (Poweraire) has interrupted any ion communication between the battery, thereby has prevented short circuit.Be provided with in series connection under the situation of check-valves, exist a fuel inlet and outlet with all batteries of can.With 1/8 " * 1/8 " the polypropylene fluking type elbow (SmallParts) of size these valves are connected on the battery through pruning.The picture of this set is shown in the Figure 4 and 5." fuel (a kind of method that needs additional supply) is carried and removed to internal diameter Tygon pipe to utilize syringe and 1/16.

The fuel storage cell system of this equipment is designed to store up fuel/electrolyte of eliminating between the battery in the pond separately by each battery use and contacts.The specific characteristic of this groove is the step shape top of Chu Chi, as shown in Figures 4 and 5.The bubble that this feasible can stacked body fully and elimination are held back contacts with anode.Fuel enters at the lower-order of groove, and leaves from higher-order, and any air of holding back all is pulled to the high-order place and does not contact with electrode." the thick hot melt polyurethane adhesive that melts under 128 ℃ and minimum pressure installs to Chu Chishang with one group of 4 briquette fuel battery, and anode-side is facing to fuel to use 0.005.

Embodiment 6: housing

The housing that is used for the biological fuel cell stacked body of 8 batteries is shown in Fig. 2 and 13." thick acrylic acid material processes final case design; characteristics are the supporting base that is used for electrode structure in the middle of housing half part; conflux chamber system perforate and in the perforate that is used for electronics and IO interface of each end of housing of the fuel on case side from 0.600 by machine.This stacked body is inserted between two housing half parts, and " liing 8/32 hexagon-headed bolt with the housing bolt together with 1 at each place, angle of four jiaos of stacked bodies." thick PTFE sealing gasket is sealed this two and half part around periphery with 0.020.

Embodiment 7: composite catalyst

The CoPcF that has found to be deposited in the carbon load polypyrrole (shows excellent activity for hydrogen reduction after common 5～60wt%) the in a single day at high temperature suitable pyrolysis.CoPcF-carbon load polypyrrole compound makes this composite catalyst CoPcFPPy very unique with the combination of suitably pyrolysis.In air self-respiration type half-cell, evaluate and test these catalyst in about 25 ℃ room temperature with 1.0M sulfuric acid solution (having or do not have phosphotungstic acid (PTA) promoter).This catalyst prepares according to following program.During the general preparation condition of catalyst is listed in the table below.

The Preparation of Catalyst condition

Composite catalyst	Weight ratio (CoPcF: CPPy)	Pyrolysis temperature (℃)	Pyrolysis time (hour)
				CoPcFCPPy	1∶8～6∶1	100～1000	0.5～6
CoPcCPPy	1∶8～6∶1	100～1000	0.5～6

Annotate: CoPcF-ten hexafluoro phthalocyanine cobalts; CPPy-carbon load polypyrrole; CoPc-phthalocyanine cobalt.

1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II) structure (CoPcF)

(if being doped)

Expression polypyrrole structure

Being prepared as follows of this compound stated, and preparation condition is CoPcF: the ratio of CPPy is 1: 2, and pyrolysis temperature is 600 ℃, and pyrolysis time is 1 hour.With platinum black (Johnson Matthey) with 5mg/cm ²Carrying capacity is coated onto on the carbon cloth (E-Tek) of Teflon processing, and catalyst layer has 10%Nafion.With CoPcFCPPy with 10mg/cm ²Carrying capacity is coated onto on the carbon cloth (E-Tek) that Teflon handles, and catalyst layer has 30%Nafion, and has or do not have phosphotungstic acid (PTA) (if having then ratio is 1: 10 (PTA: CoPcFCPPy)).Sweep speed is 2mV/s.The gained catalyst shows excellent activity to hydrogen reduction, and (maximum power density is about 25mW/cm ²), represent as the dotted line polarization curve of Figure 31.When phosphotungstic acid (PTA) was sneaked into CoPcFCPPy composite catalyst (weight ratio of PTA: CoPcFCPPy=1: 50～1: 2, representative weight ratio are 1: 10), (maximum power density was 36mW/cm to observe tangible performance rising ²).This performance and platinum (Pt) catalyst performance is suitable.

Have PTA and do not have the preparation procedure of the CoPcFCPPy of PTA

With 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-hexafluoro-29H, 31H-phthalocyanine cobalt (II) (CoPcF) (100mg, Sigma) and 200mg carbon load polypyrrole (PPy/C) (Sigma) in ultra sonic bath, in 4mL oxolane (Sigma), disperseed 10 minutes, follow in vacuum furnace under the room temperature solvent evaporation 1 hour.Then dry powder is placed crucible, and in the blanket of nitrogen stove in 600 ℃ of following heat treatments 1 hour.The gained catalyst is called CoPcFCPPy.With CoPcFCPPy (10mg) and 60mgNafion solution (5%, Sigma) in ultra sonic bath, in 200 μ L isopropyl alcohols, disperseed 10 minutes, mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.When adding phosphotungstic acid (PTA), dissolving 1mg PTA, and with 10mg CoPcFCPPy and 60mg Nafion solution (5%, Sigma) in ultra sonic bath, in 200 μ L isopropyl alcohols, disperseed 10 minutes, mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.

The preparation procedure of Pt negative electrode

Platinum black (5mg, Johnson Matthey HiSpec 1000) and 2mg carbon black (Monarch1000) were dispersed in ultra sonic bath in the 200 μ L isopropyl alcohols 5 minutes, then add 33mg Nafion solution (5%), subsequently with mixture sonicated 5 minutes again.Mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then place 30 minutes dryings of vacuum furnace under the room temperature to gas diffusion layers.

Embodiment 8: the alcohol resistance of CoPcCPPy compound cathod catalyst

Contain 15 or 30% as the CoPcFPPy that makes as described in the embodiment 1 at electrolyte solution and do not show loss of activity (Figure 32) during by oxygen-saturated ethanol.This shows the pure tolerance that catalyst shows.In 30% ethanolic solution, observe better result, because oxygen has higher solubility in this solution.

Embodiment 9: hot curing (heat-curing) influence of CoPcFCPPy air self-respiration type negative electrode

Because the fuel cell assembling is included in about 100 ℃ and suppresses fuel cell component down and solidify the Nafion of curtain coating again under about 80 ℃

Electrolytic polymer is so evaluated and tested catalyst activity after 100 ℃ solidify negative electrode down.Result among Figure 33 shows that performance and electrode curing temperature have nothing to do.

Embodiment 10: the stability of CoPcFCPPy air self-respiration type negative electrode

When evaluation and test platinum cathode in the 1M of 5% ethanol sulfuric acid solution, the current density of hydrogen reduction is less than 30mA/cm under the 0.4V electron potential ², and after 1 hour, decay to 19mA/cm ², as shown in figure 34.Platinum is the suitable catalyst of hydrogen reduction, but its also catalytic alcohol oxidation (ethanol).As shown in figure 35, the mixed potential of Pt negative electrode is lower, may cause the faster decay of its activity because the intermediate of oxidation of ethanol makes Pt catalyst poisoning on the platinum.Under the same potential of 0.4V, the CoPcFCPPy negative electrode has higher current density (referring to Figure 34) far away to hydrogen reduction, original treaty 65mA/cm ², decay to 58mA/cm after 1 hour ²CoPcFCPPy catalyst with PTA causes having than the CoPcFCPPy that does not have PTA even the negative electrode of high current density more.Current density after 1 hour from 82mA/cm ²Decay to 73mA/cm ²Have or do not have the not catalysis ethanol oxidation of CoPcFCPPy of PTA, as figure B.3 and B.7, and oxygen reduction reaction had high selectivity.For relatively, in not having the 1M sulfuric acid solution of ethanol, test platinum cathode.The hydrogen reduction current density of platinum cathode is about 77mA/cm ², decay to 71mA/cm after 1 hour ²This shows that the decay of these negative electrodes may not be by catalyst poisoning or due to decomposing.The CoPcFCPPy catalyst is highly stable in the sulfuric acid solution that contains 5% ethanol.

Embodiment 11: the stability of CoPcFCPPy catalyst

A possible explanation to observed decay (Figure 34) in the time lifetime test is negative electrode waterflooding (flooding).Figure 36 confirms that negative electrode waterflooding to a certain degree is possible, does not influence activity because acidleach in 1 or 20 hour is carried.After acidleach in 120 hours was carried, catalyst still showed extraordinary activity to hydrogen reduction.

Extracting condition.CoPcFCPPy (20mg) is dispersed in 1mL 1M H ₂SO ₄In the solution and allow its sedimentation.Filter out catalyst fines, and use the distilled water rinsing.Place vacuum furnace to transfer 2 hours catalyst fines in room temperature.(5%, Sigma) dispersion 10 minutes in 200 μ l isopropyl alcohols in ultra sonic bath slowly is coated onto 1cm with mixture with the catalyst (10mg) done and 60mg Nafion solution ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.

Embodiment 12: the CoPcFCPPy catalyst of pyrolysis not

Observe the high-temperature heat treatment CoPcFCPPy catalyst growth pattern of catalyst activity afterwards, as shown in figure 37.Carbon load P Py provides the carrier with high conductivity and high surface to be used for fixing CoPcF, to strengthen electro catalytic activity.Especially, PPy can provide with CoPcF in the nitrogen-atoms of cobalt atom coordination (coordinate).Because porphyrin is made up of 4 pyrroles unit, among the CoPcF among Co atom and the PPy coordination of the nitrogen-atoms of pyrroles unit can imitate the catalysis of Co porphyrin.Therefore, the CoPcFCPPy catalyst composites can be integrated CoPcF function and Co porphyrin function.Heat treatment may partly make CoPcF structure and PPy chain decompose under the high temperature, thereby provides more multimachine meeting with the nitrogen-atoms coordination of pyrroles unit for the Co atom.

The preparation of pyrolysis CoPcFCPPy is described among the above Embodiment B .1.

Come the preparation of pyrolysis CoPcFCPPy

With CoPcF (3.3mg, Sigma), 6.7mg PPy/C (Sigma) and 60mg Nafion solution (5%, Sigma) in ultra sonic bath, in 200 μ L isopropyl alcohols, disperseed 10 minutes, mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.

Embodiment13: at the carbon black loadings CoPcF of 600 ℃ of following pyrolysis

The CoPcF catalyst that contains PPy shows than the CoPcF catalyst that only loads on the carbon black (Monarch 1000) has the hydrogen reduction activity of increase.The activity of oxygen reduction reaction be lower than CoPcFCPPy's (Figure 31), even catalyst under 600 ℃ by pyrolysis (Figure 38).And, there is not the catalyst of PPy not have good alcohol resistance.

The preparation of CoPcF/C

With CoPcF (100mg, Sigma) and 200mg carbon black (Monarch 1000) in ultra sonic bath, in 4mL oxolane (sigma), disperseed 10 minutes, follow in vacuum furnace under the room temperature solvent evaporation 1 hour.Then dry powder is placed crucible, and in the blanket of nitrogen stove in 600 ℃ of following heat treatments 1 hour.The gained catalyst is called CoPcF/C.With CoPcF/C (10mg) and 60mg Nafion solution (5%, Sigma) in ultra sonic bath, in 200 μ L isopropyl alcohols, disperseed 10 minutes, mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.

Embodiment 14: the CoPc-PPy-C that under 600 ℃, handled

Except CoPcF, we have also prepared the phthalocyanine cobalt (CoPc) among the PPy of load on carbon black, and handle it down at 600 ℃.Its activity for hydrogen reduction is starkly lower than (the seeing Figure 39) of CoPcFPPy.

The preparation of CoPcCPPy

With CoPc (100mg, Sigma) and 200mg PPy/C (Sigma) in ultra sonic bath, among 4mLTHF (Sigma), disperseed 10 minutes, follow in vacuum furnace under the room temperature solvent evaporation 1 hour.Then dry powder is placed crucible, and in the blanket of nitrogen stove in 600 ℃ of following heat treatments 1 hour.The gained catalyst is called CoPcCPPy.With CoPcCPPy (10mg) and 60mg Nafion solution (5%, Sigma) in ultra sonic bath, in 200 μ L isopropyl alcohols, disperseed 10 minutes, mixture slowly is coated onto 1cm ²(ELAT, active side A7NCV2.1) then placed under the room temperature vacuum furnace 30 minutes to gas diffusion layers.

Embodiment 15: the biological-cathode that contains enzyme

The preparation of biological-cathode

Carbon black (or carbon nano-tube) (0.30mg) was disperseed 2 minutes preparation carbon dust suspension in 100 μ L isopropyl alcohols (Sigma) in ultra sonic bath.Enzyme (BOD or the laccase) solution of 10 μ L in the acetate buffer of 1mg/ml pH 5 is added in the suspension, shook subsequently 1 minute.Mixture slowly is coated onto 1cm ²The active side of gas diffusion layers (GDL) (ELAT, A7NCV2.1, or inner self-control GDL) then placed under the room temperature vacuum furnace 30 minutes.The acetate buffer of pH 5 makes by 12.0g acetate and 28.7g sodium acetate are dissolved in 1 liter of deionized water.

The direct electron of observing in the acetate buffer of nitrification between bilirubin oxidase (BOD) and the MWCNT/Torey carbon paper electrode shifts, as shown in figure 40.The about 0.4V of the redox potential of BOD.When cushioning liquid during by oxygen saturation, the catalytic current of oxygen reduction reaction begins to appear at about 0.8V, and increases fast under the electromotive force near the redox potential of BOD.

Also observe the know direct electron of laccase and shift, as shown in figure 41 at multi-walled carbon nano-tubes (MWCNT)/Torey carbon paper electrode place.Merit attention, 3 pairs of redox peaks clearly separate in the cyclic voltammogram, and they shift corresponding to the direct electron from the active copper site of T1, T2 in the laccase and T3 type.

Redox potential and primary structure by the T1 site are classified, and have 3 groups of laccases, promptly low, in and high oxidation reduction potential enzyme.The value of the redox potential in T1 site changes between 430-790mV (vs.NHE).Owing to successfully observed the direct electron displacement behavior of enzyme, estimated that suitable laccase can catalytic oxidation-reduction reaction under the more high potential that is beneficial to for fuel battery voltage.When using laccase (Trametes species), the ORR of laccase catalysis starts from 0.9V at hydroxy-functional MWCNT electrode (air self-respiration type), and can obtain 0.4mA/cm at 0.5V ²Current density, as shown in figure 42.

For the better direct electron that obtains enzyme shifts, must prune carbon nano-tube (CNT) at certain enzyme.For bilirubin oxidase (BOD), MWCNT is better than Single Walled Carbon Nanotube (SWCNT) performance.For oxygen reduction reaction (ORR) in the BOD catalysis of air self-respiration type negative electrode, among the MWCNT performance of hydroxy-functional MWCNT better than former MWCNT and carboxy-functionalized MWCNT, as shown in figure 43.

The direct electron that carbon black (for example DegussaXE2, XE2B, XPB F138 etc.) also is beneficial to enzyme shifts.For example, the ORR of the BOD catalysis of locating in XE2 base gas-diffusion electrode (GDE) with MWCNT JiGDEChu to compare in low current density areas be suitable, and good than at XE2B and XPB F138 JiGDEChu, as shown in figure 44.

At the biological-cathode of air self-respiration type, (the bilateral waterproof DS) is one of best commercially available GDE material of performance at the E-Tek of both sides coated with waterproof material carbon cloth.(E-Tek DS) only has water-proofing treatment to this GDE, and this can be used in polymer dielectric film (PEM) fuel cell to prevent the negative electrode waterflooding.But at biological-cathode, the activity of cathode enzyme must be held and be beneficial to direct electron and shift, so and GDE directly contact among the electrolyte solution GDE and have ionic conduction, should allow enough surrounding airs to spread (breathing) naturally simultaneously and pass through GDE.

Therefore, be necessary for the specific GDE of biological-cathode design.Use is fixed on the hydroxy-functional MWCNT/BOD mixture on the different GDE, and the performance of biological-cathode is influenced by these GDE obviously for ORR, as shown in figure 45.Confirmed that Degussa XE2 carbon black and MWCNT can effectively promote the direct electron of BOD to shift.These material with carbon elements and waterproof material (for example polytetrafluoroethylene (PTFE)) are used in the preparation of inner self-control GDE.The dough sample thing (dough) of material with carbon element and PTFE is by mixed carbon black or MWCNT and the formation of PTFE suspension.Then, this dough sample thing is coated on carbon cloth, carbon paper or the porous carbon sheet.Solvent in the coated film evaporates by pressure and 50-120 ℃ of this film of following hot pressing at the 50-10000 pound.PTFE content in the coated film is 0wt.%-80wt.%.Prior step is by adding pore former (for example 0.1-20wt.% sal volatile) in film, then pore former is controlled pore size distribution in the coated film 40-90 ℃ of following thermal decomposition.Best, as to obtain expecting GDE.Evaluation result shows that 30%PTFE among the carbon black XE2 base GDE shows optimum performance for the ORR of BOD catalysis, observes obviously high current density, for example 1mA/cm under the 0.8V especially under high potential ², this is far above E-Tek GDE (Figure 45).

Similarly, the ORR of laccase catalysis can obtain by the laccase (being coated on the GDE) that is fixed on the functionalized MWCNT.Functional group in the carbon nano-tube comprises hydroxyl, carboxyl, dredges base and amino.

In addition, do not exist under the MWCNT, be adsorbed onto BOD on the plain carbon cloth and do not show effective direct electron and shift, thus the ORR electric current of the BOD catalysis of this carbon cloth electrode under 0.2V less than 0.1mA/cm ²But, this carbon cloth can by with this cloth 500-900 ℃ down heating activate in 1-20 minute, then the cloth with heat is immersed in cooling cloth in the cold water (0 ℃), and then in vacuum furnace under 40-100 ℃ with its bone dry.Then, BOD is fixed on the activated cloth.BOD-activation cloth electrode shows obviously high ORR electric current (1.7mA/cm under 0.2V ²) (Figure 46).This activating technology provides the useful approach of the direct electron transfer that can promote enzyme.

Embodiment 16: poly-(styrene-be total to-divinylbenzene) of CuPCTSA-TBAB coating

In this embodiment, will gather (styrene-be total to-divinylbenzene) particle and coat the Nafion that goes up Tetrabutylammonium bromide (TBAB) modification

The mixture of immobilization material and water-soluble dye, this dyestuff serve as the electron mediator of enzyme with hydrogen reduction Cheng Shui.For preparing these particles, with 0.080g tetrasulfonic acid CuPc (II) (CuPCTSA) at 4.00mL TBAB modification Nafion Dye solution in the ethanolic solution (5wt%) and 2.00g poly-(styrene-be total to-divinylbenzene) particle suspension of particle in 4.00mL 0.5M phosphate buffer pH 7.2 merges.Dye solution is added in the particle suspension, and vortex mixed several seconds, until obtaining basic mixture uniformly.Then whole mixture is sprayed onto on the Merlon screen with spray gun.With mixture on screen dry 20 minutes, collect subsequently and dry storage in scintillation vial.

The gained coating particle is blue, and has examined the reservation of dyestuff by coating particles is loaded in the pillar and with the hundreds of ml water by it.But from the pillar wash-out flow mutually not have the dyestuff of discovery detection level.Coating layer ranges in thickness is designed to 0.07 micron thickness, and the average diameter of particle before coating is 8 microns.The surface area of particle (before the coating) is calculated as 0.75m/g ²The carrying capacity of gained electron mediator is 4wt%.

Embodiment 17: the polypyrrole/carbon black compound of the doping that alcohol dehydrogenase-TBATFB coats

On polypyrrole/carbon black compound (Sigma, catalog number (Cat.No.) 530573) particle, coat the Nafion of tetrabutyl ammonium tetrafluoroborate (TBATFB) modification through mixing

The mixture of immobilization material and alcohol dehydrogenase.Preparation 0.080g is through the solution of cryodesiccated alcohol dehydrogenase (ML57) and 2.00mL 0.5M phosphate buffer (pH 7.2).The ethanolic solution of preparation 4.00mL TBATFB modification Nafion (5wt%), 1.00g are through the polypyrrole/carbon black compound of doping and the suspension of 4.00mL 0.5M phosphate buffer (pH7.2).Then with enzyme solutions and suspension vortex mixed several seconds, until obtaining basic mixture uniformly.Then whole mixture is sprayed onto on the Merlon screen with spray gun in the technology of similar embodiment 16.With the products therefrom dry storage in 4 ℃ scintillation vial.

The gained coating particles is a black, and utilizes test of standard spectrophotometry and electrochemistry to evaluate and test the reservation of having examined enzymatic activity.Evaluation and test is carried out with respect to normal hydrogen reference electrode (NHE).See Figure 49.Gained enzyme carrying capacity is 6.25wt%.

Embodiment 18: alcohol dehydrogenase-Ru (II) (NH ₃) ₆Polypyrrole/carbon black the compound of the doping that-TBATFB coats

To wrap the Nafion of TBATFB modification through the polypyrrole/carbon black complex particle (Sigma, catalog number (Cat.No.) 530573) that mixes

, alcohol dehydrogenase and Ru (II) (NH ₃) ₆Mixture.Ru (II) (NH ₃) ₆It is the electron mediator that is used for the ethanol enzymatic oxidation.Preparation contains 0.080g through cryodesiccated alcohol dehydrogenase (ML59), 0.100g Ru (II) (NH ₃) ₆Solution with 4.00mL 0.5M phosphate buffer (pH 7.2).Also preparation contains the ethanolic solution of 6.00mL 0.5M phosphate buffer (pH 7.2), 2.00mLTBATFB modification Nafion (5wt%) and the suspension of the polypyrrole/carbon black compound of 2.00g through mixing.Solution and suspension merged and vortex mixed several seconds to form basic mixture uniformly.Then whole mixture is sprayed onto on the Merlon screen with spray gun in the technology of similar embodiment 16.With the products therefrom dry storage in 4 ℃ scintillation vial.

The gained coating particles is a black, and the standard spectrophotometry of utilization is tested the reservation of having examined enzymatic activity.Because particle surface is long-pending unknown, does not calculate coating thickness.Gained enzyme carrying capacity and electron mediator carrying capacity are respectively 10.3wt% and 12.8wt%.

Embodiment 19: the diastatic enzymatic activity that consumes starch

The enzyme test that is used for amyloglucosidase (EC 3.2.1.3) is announced by Sigma, and can be got from http://www.sigmaaldrich.com/img/assets/18200/Amyloglucosidasel. pdf; It is based on Bergmeyer, H.U., Gawehn K. and Grassl, M. (1974) Methods ofEnzymatic Analysis (Bergmeyer, H.U. edits) the 2nd edition, I volume, the document program described in the 434-435.Some slight variation of this program: use the material immobilized amyloglucosidase of expectation enzyme immobilization in the cuvette bottom, rather than amyloglucosidase is added solution.The enzyme immobilization material of being tested is the Nafion with bromination tetrapropyl ammonium (TPAB), Tetrabutylammonium bromide (TBAB), bromination triethyl group hexyl ammonium (TEHA), bromination trimethyl hexyl ammonium (TMHA), bromination trimethyl octyl group ammonium (TMOA), bromination trimethyl decyl ammonium (TMDA), bromination trimethyldodecane base ammonium (TMDDA), Tetradonium Bromide (TMTDA), bromination trimethyl cetyltrimethyl ammonium (TMHDA), bromination trimethyl octadecyl ammonium (TMODA) modification

, be suspended in the butyl modification of chitosan (Chitosan AB) in the acetate buffer and be suspended in butyl modification of chitosan (ChitosanTB) in the tert-pentyl alcohol.For the amylase that consumes starch provides the enzyme immobilization material of maximum relative activity is the butyl modification of chitosan (Chitosan TB) that is suspended in the tert-pentyl alcohol.See Figure 50.

Embodiment 20: the diastatic enzymatic activity that consumes maltose

Program described in the embodiment 19 is used for measuring the amylase that consumes maltose and is fixed on activity in the various enzyme immobilization materials.The program of being announced process is as described in example 19 above revised, and starch is replaced with maltose.The enzyme immobilization material that is tried is the Nafion with bromination tetrapropyl ammonium (T3A), Tetrabutylammonium bromide (TBAB), bromination four pentyl ammonium (T5A), bromination triethyl group hexyl ammonium (TEHA), bromination trimethyl hexyl ammonium (TMHA), bromination trimethyl octyl group ammonium (TMOA), Tetradonium Bromide (TMTDA) modification , intermediate molecular weight decyl chitosan modified (Decyl M), low-molecular-weight butyl chitosan modified (Butyl L), low-molecular-weight octyl group chitosan modified (Octyl L) and intermediate molecular weight butyl chitosan modified (Butyl M).For the amylase that consumes maltose provides the enzyme immobilization material of maximum relative activity is intermediate molecular weight decyl chitosan modified (Decyl M).See Figure 51.

Embodiment 21: biological anode catalyst carrier 1

Preparation has the biological anode catalyst carrier electrode of porous metals silk screen carrier, collector body and polymer pore former.Porous metals silk screen carrier is used in the preparation said preparation; The ratio (0.8: 2.5) of the ratio (1: 4) of graphite fibre and conductibility carbon black and polymer adhesive and carbon solid causes high conductance but structure that can not self-supporting.When not having the porous metals silk screen, carbon electrode can rupture in operating process, thereby thinks that can not be used for MEA prepares.This material that is used for this electrode is a preferred material, but carbon black and adhesive material can be replaced (table 1 and 3) with other substitute.

The dried component of GDL thickener be 150 microns long of 4.00g Monarch 1400 carbon blacks (Cabot), 1.00g poly-(vinylidene fluoride) powder of Milled XN-100 graphite fibre (Nippon Graphite Fiber), 1.60g as polymer adhesive (Sigma) and 2.00g molecular weight be 8,000 gather (ethylene glycol) as polymer pore former (Sigma).These dried components were mixed in food processing equipment 1 minute.The solvent of GDL thickener is a 4.00mL ethanol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is coated onto in porous metals silk screen and the frame structure by above-mentioned, and 180 ℃ of sintering 20 minutes.Remove framework,, change water therebetween termly electrode bubble in distilled water 30 minutes.This step is necessary for the pore former inclusion poly-(ethylene glycol) of removing in the electrode structure.Then electrode is reburned under 200 ℃ and tied 10 minutes.With the electrode cool to room temperature, and prepare as biological anode catalyst carrier and electrode.The gained electrode is rigidity, can cut off with scissors, absorbs power and water easily and separates matter.Conductivity in the plane (in-plane conductivity) is purchased GDL material～125%.

Embodiment 22: biological anode catalyst carrier 2

Preparation has the biological anode catalyst carrier electrode of porous metals silk screen carrier, collector body and polymer pore former.In said preparation porous metals silk screen carrier not necessarily because graphite fibre and conductibility carbon black equal proportion, and polymer adhesive and the ratio of carbon solid higher (1: 2).Said preparation obtains the conductibility structure more lower slightly than biological anode catalyst carrier embodiment 21, but unlike the carrier of embodiment 21, it is a self-bearing type.This electrode material can wet, but since preparation in do not have pore former and unlike the embodiment of front adsorbing electrolyte.The electrode of the type can be purchased GDL similar fashion utilization, and can consider to be used for the MEA preparation.This material that is used for this electrode is a preferred material, but can replace carbon black and adhesive material (seeing table 1 and 3 for details) with other.

The dried component of this GDL thickener is poly-(vinylidene fluoride) powder (Sigma) of long DKDX graphite fibre of 3.00g Monarch 1400 carbon blacks (Cabot), 3.00g150 micron (Cytec Carbon Fibers) and 3.00g.These dried components were mixed in food processing equipment 1 minute.The solvent of GDL thickener is the 3.00mL normal propyl alcohol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is coated onto in frame structure and the PTFE coated fibres sheet glass structure by above-mentioned, and 200 ℃ of sintering 20 minutes.With the electrode cool to room temperature, and prepare as biological anode catalyst carrier and electrode.The gained electrode is rigidity, can use scissor cut, and water can abundant wetting GDL surface.Conductivity is purchased GDL material～75% in the plane.

Embodiment 23: biological anode catalyst carrier 3

Preparation has the biological anode catalyst carrier electrode of mesoporous carbon content and pore former, is used for the enzyme catalyst load with interaction and the increase avaivable electrode surface area that promotes bigger electrolyte/electrode.Graphite fibre in the said preparation: conductive black: the ratio of mesoporous carbon is 2: 1: 1, and polymer adhesive is 1: 3 with the ratio of carbon solid.The electrical conductance of resulting structures and biological anode catalyst carrier 2 quite.Because pore former and mesoporous carbon in the preparation, this electrode material can be than other embodiment and listed GDL material wetting better and absorbed electrolyte and the enzyme curtain coating ink of being purchased.The electrode of the type can be purchased GDL similar fashion utilization, and can be used in the MEA preparation.These materials that are used for this electrode are preferred materials, but carbon black and adhesive can be replaced (table 1 and 3) with other.

The dried component of GDL thickener be DKDX graphite fibre (Cytec Carbon Fibers), 1.50g Chemsorb 1505G5 porous dipping steam activation carbon (C*Chem), poly-(vinylidene fluoride) powder (Sigma) of 2.00g and the 1.50g molecular weight of 150 microns long of 1.50g Monarch 1400 carbon blacks (Cabot), 3.00g be 8,000 gather (ethylene glycol) (Sigma).Activated carbon was used ceramic mortar and the artificial of short duration grinding of pestle before sneaking into; These dried components were mixed in food processing equipment 1 minute.Solvent is～the 3.00mL normal propyl alcohol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is taped against in frame structure and the PTFE coated fibres sheet glass structure, and 200 ℃ of sintering 20 minutes.Remove framework,, change water poly-to remove (ethylene glycol) therebetween termly electrode bubble in distilled water 30 minutes.Then electrode is reburned under 200 ℃ and tied 10 minutes.With the electrode cool to room temperature, and prepare as biological anode catalyst carrier and electrode.The gained electrode is rigidity, can use scissor cut, and absorbs power and water acceptably and separate matter.Conductivity is purchased GDL material～50% in the plane.

Embodiment 24: biological anode catalyst carrier 4

This biology anode catalyst carrier electrode is similar to biological anode catalyst carrier 2 on physical property, but different being to use carbon black conductibility component and using pore former.This biology anode catalyst carrier comprise chemical graft on the carbon black materials the conducting polymer through mixing and the pore former of thermal decomposition in sintering process.The use of new pore former has been shortened the process time of preparation and has been saved the water absorption step and second sintering step.This electrode material can be wetting as front embodiment, absorbed electrolyte, and can participate in electron transfer reaction between enzyme and the electrode.The electrode of the type can be purchased GDL similar fashion utilization, and can be used in the MEA preparation.This material that is used for this electrode is a preferred material, but carbon black and adhesive can be replaced (table 1 and 3) with other.

The dried component of this GDL thickener is poly-(vinylidene fluoride) powder (Sigma) of DKDX graphite fibre (CytecCarbon Fibers), 3.00g and the 2.00g ammonium carbonate (Sigma) that 3.00g is grafted with 150 microns long of Monarch 1400 carbon blacks, the 3.00g of poly-(pyrroles) that 8-hydroxyl pyrene trisulfonic acid mixes.These dried components were mixed in food processing equipment 1 minute.The solvent of GDL thickener is～4.00mL ethanol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is coated onto in frame structure and the PTFE coated fibres sheet glass structure by above-mentioned, and 180 ℃ of sintering 20 minutes.With the electrode cool to room temperature, and prepare as biological anode catalyst carrier and electrode.The gained electrode is rigidity, can use scissor cut, and the surface can be by water-wet.Conductivity is purchased GDL material～75% in the plane.

Embodiment 25: biological-cathode catalyst carrier 1

Preparation biological-cathode catalyst carrier electrode wherein uses the polymer pore former to be used for the surface area that enzyme reacts to each other and to improve the product of cathode reaction and the mass transfer of reactant to increase electrode.Said preparation does not need to use porous metals silk screen carrier, because higher ratio between the ratio of graphite fibre and conductibility carbon black and polymer adhesive and the carbon solid (～1: 2) cause the structure of self-bearing type.This structure also shows and is similar to the conductibility that is purchased carbon cloth GDL material.This electrode material is very hydrophobic, but can be wetting by enzyme and ionomeric alcohol mixture in the enzyme deposition process.Use pore former to improve surface area in this electrode, make enzyme electrode react to each other better.Found that usefulness is only very hydrophobic by the cathode electrode that PTFE prepares as adhesive, but in the utilization process, broken easily.This problem replaces with PVDF by the PTFE binder substance with the medium and small percentage of preparation and avoids.The electrode of the type can be purchased GDL similar fashion utilization, and be used in the MEA preparation.This material that is used for this electrode is a preferred material, but carbon black and adhesive can be replaced (table 1 and 3) with other.

The dried component of this GDL thickener is DKDX graphite fibre (Cytec Carbon Fibers), 1.50g poly-(vinylidene fluoride) powder (Sigma), 3.00g polytetrafluorethylepowder powder (Sigma) and the 2.00g carbonic hydroammonium (Sigma) of 150 microns long of 4.00g Monarch 1400 carbon blacks (Cabot), 4.00g.These dried components were mixed in food processing equipment 1 minute.The solvent of GDL thickener is～the 3.00mL normal propyl alcohol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is coated onto in frame structure and the PTFE coated fibres sheet glass structure by above-mentioned, and 200 ℃ of sintering 20 minutes.With the electrode cool to room temperature, and then under 300 ℃, reburn the knot 10 minutes.With the electrode cool to room temperature, and prepare as biological-cathode catalyst carrier and electrode.The gained electrode is rigidity slightly, can use scissor cut, and the surface is very hydrophobic.Conductivity is purchased GDL material～50% in the plane.

Embodiment 26: biological-cathode catalyst carrier 2

This biological-cathode catalyst carrier electrode is similar to biological-cathode catalyst carrier 1 on physical property, but different be the use of carbon black conductibility component and PVDF and the ratio of PTFE.This carrier material is sneaked into the conducting polymer of the doping of chemical graft on the carbon black materials.For improving the direct electron rate of transform greatly, electron transfer takes place to be formed for the careful grafting of mode with the interactional nano whisker structure of the metal center of enzyme in this center in the conducting polymer of this doping.When using this modification carbon, do not carry out among the embodiment 25 sintering step under 300 ℃ of temperature, to prevent the softening or fusing of polymer nanocomposite structure; And the ratio that changes adhesive reduces PTFE to increase PVDF, because the sintering temperature that PTFE has relatively high expectations.This electrode material is hydrophobic and have the part hydrophily, has high surface area, helps the direct electron transfer reaction between enzyme and the electrode.The electrode of the type be purchased GDL similar fashion utilization, and be used in the MEA preparation.This material that is used for this electrode is a preferred material, but carbon black, pore former and adhesive can be replaced (table 1 and 3) with other.

The dried component of this GDL thickener is DKDX graphite fibre (Cytec Carbon Fibers), 3.00g poly-(vinylidene fluoride) powder (Sigma), 1.00g polytetrafluorethylepowder powder (Sigma) and the 2.00g ammonium carbonate (Sigma) that 4.00g is grafted with 150 microns long of Monarch 1400 carbon blacks, the 4.00g of poly-(pyrroles) nano whisker that hydroxide 1-(3-sulfopropyl) pyridinium inner salt mixes.These dried components were mixed in food processing equipment 1 minute.Solvent composition used in the GDL thickener is～4.00mL ethanol, and its increment is added, until reaching the expectation denseness.

With spatula these dried components are mixed in small beaker, reach the denseness of toothpaste until this mixture.Then this thickener is coated onto in frame structure and the PTFE coated fibres sheet glass structure by above-mentioned, and only once 200 ℃ of following sintering 20 minutes.With the electrode cool to room temperature, and prepare as biological-cathode catalyst carrier and electrode.The gained electrode is rigidity slightly, can use scissor cut, and the surface is very hydrophobic.Conductivity is purchased GDL material～50% in the plane.

Embodiment 27: biological-cathode catalyst carrier 3

Preparation has air from breathing side and enzyme side and having the double-deck biological-cathode catalyst carrier electrode of porous metals silk screen carrier as collector body.Because the low ratio (for example 1: 10) of electrode enzyme side polymer adhesive and carbon solid and owing to the combination of electrode layer, so porous metals silk screen carrier is necessary for said preparation.Preferred arrange be enzyme side at electrode have micropore and slightly hydrophilic-structure and provide fine and close ventilative and hydrophobic structure with control water and keep electrolyte in battery increasing the interaction between enzyme and the electrolyte in air side.This realizes by using double-deck electrode.This two-layer electrode needs multistep preparation and sintering process.This material that is used for this electrode is a preferred material, but carbon black and adhesive can be replaced (table 1 and 3) with other.

The air that is used for this electrode is the Milled XN-I00 graphite fibre (CytecCarbon Fibers) and the 2.50g polytetrafluorethylepowder powder (Sigma) of 150 microns long of 4.00g Monarch1400 carbon blacks (Cabot), 1.00g from the dried component of breathing the GDL thickener of side.These dried components were mixed in food processing equipment 1 minute.Solvent used in the GDL thickener is～4.00mL 1-propyl alcohol, and its increment is added, until reaching the expectation denseness.

The dried component of GDL thickener that is used for the enzyme side of this electrode is Milled XN-100 graphite fibre (Cytec Carbon Fibers), 0.25g polytetrafluorethylepowder powder (Sigma) and the 1.00g ammonium carbonate (Sigma) of 150 microns long of Monarch 1400 carbon blacks (Cabot), the 0.50g of the modification of 2.00g conducting polymer.These dried components were mixed in food processing equipment 1 minute.Solvent used in the GDL thickener is～4.00mL 1-propyl alcohol, and its increment is added, until reaching the expectation denseness.

To be used for air with spatula and mix at small beaker with solvent, reach the denseness of toothpaste until this mixture from the dried component of breathing the side thickener.Then this thickener is coated onto a side of porous metals silk screen and frame structure by " biological anode " part is described, and 300 ℃ of following sintering 20 minutes.Electrode is cooled off.And mix at small beaker with solvent with the dried component that spatula will be used for enzyme side thickener, reach the denseness of toothpaste until this mixture.Then side is the same to be coated onto the opposite side of porous metals silk screen and frame structure with this thickener from breathing with air, and for the second time 200 ℃ of following sintering 20 minutes.With the electrode cool to room temperature, and prepare as biological-cathode catalyst carrier and electrode.One side of gained electrode is hydrophobic, and the opposite side that is coated with the enzyme layer is hydrophilic.

Embodiment 28: with the comparison that is purchased the GDL material

Figure 54 has provided biological-cathode catalyst carrier 1 and two-sided Elat (BASF) and has been purchased comparison between the negative electrode GDL, wherein uses with oxygen as the laccase base biological-cathode of oxidant with make platinum black anode on the Elat of fuel with hydrogen.Nafion 115 is as the polymer dielectric film among this hot pressing MEA.The activity of this biological-cathode is based on DET, and do not have the electron transfer amboceptor in the catalyst layer.These two systems compare approaching in whole polarization scope.The biological-cathode carrier layer is uniform biological-cathode carrier layer and does not optimize for the mass transfer of product or reactant.In addition, do not use pore former, this causes surface area to reduce.

Figure 55 has provided biological anode catalyst carrier 3 and two-sided Elat and has been purchased comparison between the anode GDL, wherein uses with 5.0% methyl alcohol and makes the platinum ruthenium black anode of fuel and go up the platinum black negative electrode with oxygen as the Elat of oxidant.Nafion 115 is as the polymer dielectric film among this MEA.Except the open circuit voltage zone, prepared GDL ratio is purchased material and all does very well on whole polarization curve.This may be because the hydrophilic nanoporous structure of prepared GDL causes increases that contact of fuel solution and film, cause since fuel more leap cause the cathodic depolarization increase.

Embodiment 29: the carbon black particle of the polyaniline nano-line grafting that 1-(3-sulfopropyl) hydroxide pyridine mixes

For the carbon black particle of preparation modification, the 300mL deionized water is transferred in the beaker of one liter of size, and beaker is placed ice bath.With glacial acetic acid (10mL, 99%, Sigma) add in the beaker, and continuous stirring 20 minutes is to reduce the temperature of liquid in the beaker, preferably near 5 ℃.Temperature at acid flux material reaches after 5 ℃, and 40 gram Monarch 1400 (Cabot) carbon blacks are added in the acid flux material.This carbon was starched continuous stirring 10 minutes.Then, add the mixture that is dissolved in gram 1-(3-sulfopropyl) the hydroxide pyridinium inner salt of 8 in the 50mL deionized water (Sigma).This mixture was stirred 20 minutes.(10mL Sigma), and stirred whole mixture 20 minutes more dropwise to add aniline monomer in above mixture.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram ammonium persulfate of 18 in the 60mL deionized water (Sigma) through 2 hours time-histories.Preferably not rising in this elementary reaction medium temperature is higher than 5 ℃, otherwise obtains macropore fiber rather than nano wire.After in case all ammonium persulfate solutions all add, with slurry continuous stirring 24 hours in ice bath.After first 24 hours stir, add the mixture that is dissolved in the gram of 8 in 50mL deionized water 1-(3-sulfopropyl) hydroxide pyridinium inner salt.This mixture was stirred 20 minutes.In above mixture, dropwise add the 10mL aniline monomer, and whole mixture was stirred 20 minutes again.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram of 18 in 60mL deionized water ammonium persulfate through 2 hours time-histories.Preferably should not rise in this elementary reaction medium temperature is higher than 5 ℃, otherwise obtains macropore fiber rather than nano wire.After in case all ammonium persulfate solutions all add, with slurry continuous stirring 24 hours in ice bath.Stir in ice bath 24 hours when finishing at this second, carbon starched vacuum filtration, and with a large amount of water washings with except that disacidify.Then, with this modification carbon 100 ℃ of following vacuumizes 10 hours.

Embodiment 30: 1, the carbon black particle of the polyaniline nano-line grafting that the 5-naphthalenedisulfonic acid mixes

(300mL) transfers in the beaker of one liter of size with deionized water, and beaker is placed ice bath.Glacial acetic acid (10mL, 99%) is added in the beaker, and 20 minutes temperature with liquid in the reduction beaker of continuous stirring, preferably near 5 ℃.Temperature at acid flux material reaches after 5 ℃, and 40 gram Monarch 1400 (Cabot) carbon blacks are added in the acid flux material.This carbon was starched continuous stirring 10 minutes.Then, adding is dissolved in the gram of 8 in the 50mL deionized water 1, and 5-naphthalenedisulfonic acid tetrahydrate (97%, mixture Sigma).This mixture was stirred 20 minutes.In above mixture, dropwise add aniline monomer (10mL), and whole mixture was stirred 20 minutes again.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram of 18 in 60mL deionized water ammonium persulfate through 2 hours time-histories.Preferably should not rise in this elementary reaction medium temperature is higher than 5 ℃, otherwise obtains macropore fiber rather than nano wire.After in case all ammonium persulfate solutions all add, with slurry continuous stirring 2 hours in ice bath.Carbon is starched vacuum filtration, and with a large amount of water washings to remove disacidify.Then, with this modification carbon 100 ℃ of following vacuumizes 10 hours.

Embodiment 31: the carbon black particle of the polyaniline nano-line grafting that the 2-naphthalene sulfonic acids mixes

(300mL) transfers in the beaker of one liter of size with deionized water, and beaker is placed ice bath.Glacial acetic acid (10mL, 99%) is added in the beaker, and 20 minutes temperature with liquid in the reduction beaker of continuous stirring, preferably near 5 ℃.Temperature at acid flux material reaches after 5 ℃, and 40 gram Monarch 1400 (Cabot) carbon blacks are added in the acid flux material.This carbon was starched continuous stirring 10 minutes.Then, add be dissolved in the gram of 8 in 50mL deionized water 2-naphthalene sulfonic acids (70%, mixture Sigma).This mixture was stirred 20 minutes.In above mixture, dropwise add aniline monomer (10mL), and whole mixture was stirred 20 minutes again.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram of 18 in 60mL deionized water ammonium persulfate through 2 hours time-histories.Preferably should not rise in this elementary reaction medium temperature is higher than 5 ℃, otherwise obtains macropore fiber rather than nano wire.After in case all ammonium persulfate solutions all add, with slurry continuous stirring 2 hours in ice bath.Carbon is starched vacuum filtration, and with a large amount of water washings to remove disacidify.Then, with this modification carbon 100 ℃ of following vacuumizes 10 hours.

Embodiment 32: the carbon black particle of the polyaniline nano-line grafting that 1-(3-sulfopropyl) hydroxide pyridine mixes

The 300mL deionized water is transferred in the beaker of one liter of size, and beaker is placed on the agitating plate.Glacial acetic acid (10mL, 99%) is added in the beaker, and descended continuous stirring 20 minutes in room temperature (roughly 24 ℃).(20 grams Cabot) add in this acid flux material with Black Pearls 2000 carbon blacks.This carbon was starched continuous stirring 30 minutes.Then, add the mixture that is dissolved in gram 1-(3-sulfopropyl) the hydroxide pyridinium inner salt of 8 in the 50mL deionized water (Sigma).This mixture was stirred 20 minutes, in above mixture, dropwise add the 10mL aniline monomer, and whole mixture was stirred 20 minutes again.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram iron chloride of 10 in the 60mL deionized water (III) through 2 hours time-histories.After in case all ferric chloride solutions all add, with slurry continuous stirring 2 hours at room temperature.Carbon is starched vacuum filtration, and with a large amount of water washings to remove disacidify.Then, with this modification carbon 100 ℃ of following vacuumizes 10 hours.

Embodiment 33: the carbon black particle of the polyaniline nano-line grafting that sulfonation 1-(3-sulfopropyl) hydroxide pyridine mixes, it is used for the biological-cathode of high power output

(300mL) transfers in the beaker of one liter of size with deionized water, and beaker is placed ice bath.With glacial acetic acid (10mL, 99%, Sigma) add in the beaker, and continuous stirring 20 minutes is to reduce the temperature of liquid in the beaker, preferably near 5 ℃.Temperature at this acid flux material reaches after 5 ℃, and 40 gram Monarch 1400 (Cabot) carbon blacks are added in the acid flux material.This carbon was starched continuous stirring 10 minutes.Then, add the mixture that is dissolved in gram 1-(3-sulfopropyl) the hydroxide pyridinium inner salt of 8 in the 50mL deionized water (Sigma).This mixture was stirred 20 minutes, and (10mL Sigma), and stirred whole mixture 20 minutes more dropwise to add aniline monomer in above mixture.For making aniline begin polymerization, dropwise add the mixture that is dissolved in the gram ammonium persulfate of 18 in the 60mL deionized water (Sigma) through 2 hours time-histories.Preferably not rising in this elementary reaction medium temperature is higher than 5 ℃, otherwise obtains macropore fiber rather than nano wire.After in case all ammonium persulfate solutions all add, with slurry continuous stirring 24 hours in ice bath.After stirring in 24 hours this first, dropwise add the mixture of the 6mL concentrated sulfuric acid (97%) and 12mL glacial acetic acid (97%), the while, beaker was still in ice bath.Slurry was stirred 1 hour, then vacuum filtration, and with a large amount of water washings.At last, with this modification carbon 100 ℃ of following vacuumizes 10 hours.

Embodiment 34:Coat enzyme/carbon diffusion electrode 1:ADH immobilization anodic half-cell

Alcohol dehydrogenase is coated on the hydrocarbon black powder, drying, and be coated onto on the electrode through said procedure.PVDF is as adhesive, and the DKDX graphite fibre is used to provide rigid electrode, and Printex XE 2 is as filler, and polyethylene glycol is a hydrophilizing agent.This paste formulation is carbon, 0.45gPrintex XE 2,0.25g DKDX, 0.25g PVDF and the 0.5g polyethylene glycol that 0.5g ADH coats.The methyl alcohol of q.s is sneaked in this thickener giving its high-consistency, and prepare electrode according to embodiment 21 described programs.Then test this half-cell the enzymatic of methyl alcohol has been replied to examine enzymatic activity, shown positive response.The results are shown in Figure 58.

Carry out the half-cell test existing and do not exist under the methyl alcohol in 1M pH is 7.2 phosphate buffer.Shown in Figure 58, observe the about 5mA that shifts through direct electron and reply.

Embodiment 35: coat enzyme/carbon diffusion electrode 2: the ADH immobilization anodic half-cell of amboceptor mediation

Alcohol dehydrogenase is coated on the hydrocarbon black powder, drying, and be coated onto on the electrode through said procedure.Described identical among these components and the embodiment 34 at coating enzyme/carbon diffusion electrode 1.This paste formulation is 1.09g ADH carbon coated, 0.5g DKDX, 0.5g PVDF and 0.5g polyethylene glycol.The methyl alcohol of q.s is sneaked in this thickener giving its high-consistency, and prepare electrode according to said procedure.In preparation, do not have under the situation of Printex XE 2 carbon fillers, must use amboceptor to observe enzymatic and reply.For this embodiment, in solution, use hexamine ruthenic chloride (III) (Sigma).Also can coat the solubility amboceptor and observe and reply with enzyme.The results are shown in Figure 59.

Shown in Figure 59, this electrode can show catalysis at the place, amboceptor peak of about 0.65V (to NHE) and reply.

Embodiment 36: biological-cathode ink formulations 1: enzyme immobilization is to ELAT GDL carrier

Carbon with enzyme coats is coated onto electrode on the ELAT carrier structure.In a kind of ink formulations, the ordinary carbon black particle is as the filler material in the ink, and another preparation only has carbon and Nafion solution that enzyme coats.This carbon filler base ink formulations comprises carbon, 320mgMonarch 1400 and the 2.0mL 5%Nafion solution that the 160mg laccase coats.The ink formulations of carbon containing filler does not comprise carbon and the 0.667mL 5%Nafion solution that the 160mg laccase coats.

In the ink preparation process, at first enzyme is coated solution and add in the plastic jar.Next step added carbon filler and vortex mixed about 30 seconds.Add Nafion solution, and stir slurry with spatula and all wet until all particles.Before being applied on the electrode holder, this ink is mixed into the time of making the necessary weak point of trying one's best of even denseness ink with Fisher ScientificSonicating Dismembrator.This ink is applied on the two-sided ELAT, then they was pressed onto on Nafion 115 films in following 35 seconds and is positioned at the platinum black electrode opposite side at 125 ℃.

The data of Figure 60 are to use with 0.500mg/cm ²Carrying capacity is loaded into that 24Unit/mg laccase on the electrode gathers.The electrode that does not contain any carbon filler shows direct electron and shifts (DET), does not need electron mediator to obtain power, still uses the carbon of high conductance high surface can increase overall power density and the current density that battery provides greatly in ink formulations.Favourable carbon filler for example is Monarch 1400, Black Pearls 1300, Printex XE 2 and Vulcan XC-72.Other acceptable carbon black is listed among Figure 60.In catalyst ink, use carbon black to make filler, obtain greater than 85mA/cm ²Current density.The key parameter that influences the performance of this cathode electrode is the thickness of immobilized polymer layer of specific activity, coating of the amount of carbon black, operating temperature, immobilization laccase and the amount of 5%Nafion solution.

Embodiment 37: biological-cathode ink formulations 2: the amount that changes carbon filler in the ink formulations

Shown in Figure 61, the amount that increases carbon black has strengthened electrode performance, and until reaching the catalyst based layer thickness of critical enzyme, this moment, performance began to descend because reactant and proton transfer limit.Select to use 3 kinds of electrode preparations of 320mg, 240mg and 400mg Monarch 1400 carbon to help support direct electron to transfer to enzyme.320mg carbon filler ink formulations is made by carbon, 320mgMonarch 1400 and 2.0mL 5%Nafion solution that the 160mg laccase coats.240mg carbon filler ink formulations is made by carbon, 240mg Monarch 1400 and 2.0mL 5%Nafion solution that the 160mg laccase coats.400mg carbon filler ink formulations is made by carbon, 400mgMonarch 1400 and 2.0mL 5%Nafion solution that the 160mg laccase coats.

As preparation ink formulations as described in the embodiment 36, wherein before adding Nafion solution with the carbon vortex mixed of dried carbon with the enzyme coating.Then with the ink sonicated and be applied on the ELAT.125 ℃ following 35 seconds they be pressed onto on Nafion 115 films and be positioned at the platinum black electrode offside.

When using Monarch 1400 carbon blacks, observing critical thickness is 480mg total carbon weight.Reach the required total carbon weight of critical bed thickness and depend on the carbon that is used for enzyme immobilization and is used as filler.For example, if use closeer carbon, then before performance descended owing to ink thickness, acceptable total carbon weight was higher.Shown in Figure 61, when with the 80mg increment with content of carbon black when 240mg is increased to 400mg, the peak value carrying capacity appears at 320mg (～13mg/cm ²).For these tests, use 0.500mg/cm ²The 24Unit/mg laccase of enzyme carrying capacity.

Embodiment 38: biological-cathode ink formulations 3: change types of carbon black as filler

Select 3 kinds of carbon blacks to illustrate that the carbon type variation can be to the influence of battery overall performance.The carbon black that has compared Monarch 1400, Pure Black 115 and doped polypyrrole among Figure 62.Ink formulations provides below.

Monarch 1400 carbon filler ink formulations are made by carbon, 80mgMonarch 1400 and 1.0mL 5%Nafion solution that the 80mg laccase coats.The carbon that the carbon black carbon filler ink formulations of doped polypyrrole is coated by the 80mg laccase, the carbon black and the 1.0mL 5%Nafion solution of 80mg doped polypyrrole are made.Pure Black 115 ink formulations are made by carbon, 80mg Pure Black 115 and 1.0mL 5%Nafion solution that the 80mg laccase coats.

As described above, before adding Nafion solution with the carbon vortex mixed of dried carbon with the enzyme coating.Then with the ink sonicated and be applied on the ELAT.They were pressed onto on the Nafion115 film in the platinum black electrode offside in following 35 seconds at 125 ℃.

When considering carbon black as enzyme during the filler in the catalyst based ink, the existence of polymer-doped thing grafting, conductibility and surface area can influence the overall performance of electrode.Several carbon blacks have carried out test to improve battery performance, preferred Monarch 1400 as filler.But any carbon black can substitute the interaction that Monarch 1400 improves enzyme and collector body shown in Figure 62.All carbon blacks all do very well than independent enzyme, but every kind of carbon black has the Different Optimization parameter that consider.In the various carbon that tried 3 types have been shown among Figure 62, and except being used as the carbon type of filler, all parameters are all identical.The enzyme carrying capacity of figure power curve D.11 is 0.250mg/cm ²Other carbon that is tried provides in table 1.

Embodiment 39: biological-cathode ink formulations 4: immobilization layer thickness evaluation and test

For of the influence of test immobilization bed thickness, carry out following program to the direct electron rate of transform (it can also be directly related with battery performance).In this program, carbon filler total content is held constant at 480mg, and increase is put into the immobilised enzymes/carbon amount of ink to keeping constant enzyme carrying capacity.The key parameter that this experiment changed is the amount that is coated onto the modification Nafion enzyme solutions on the carbon in the encapsulation steps.At first, the enzyme solutions with following surface compositions is fixed on the carbon.17% immobilised enzymes prescription by the carbon of 1g nano wire grafting, 200mg in 0.5M pH 7.2 phosphate buffers laccase and the Nafion of 4mL 5% Tetrabutylammonium bromide modification make.8.5% immobilised enzymes prescription by the carbon of 1g nano wire grafting, 100mg in 0.5MpH 7.2 phosphate buffers laccase and the Nafion of 2mL 5% Tetrabutylammonium bromide modification make.4.2% immobilised enzymes prescription by the carbon of 1g nano wire grafting, 50mg in 0.5M pH 7.2 phosphate buffers laccase and the Nafion of 1mL 5% Tetrabutylammonium bromide modification make.

With laccase be dissolved in phosphate buffer (pH 7.2,0.5M) in, mix with the carbon sample of 1g nano wire grafting (for example: embodiment 29, the polyaniline grafted carbon black particle that 1-(3-sulfopropyl) hydroxide pyridine mixes).This enzyme/carbon was starched vortex mixed 5 minutes.Then, add the Nafion solution (5wt% in ethanol) of Tetrabutylammonium bromide modification, and every addings 1mL is afterwards with its vortex mixed 1 minute with the 1mL increment.With this mixture spray drying (open) at room temperature to surrounding air.This obtains loading on the immobilised enzymes on the carbon of nano wire grafting.Following to every kind of ink formulations that the immobilised enzymes formulation is independent.The carbon ink water formulation that 17% enzyme coats is made by 20mg 17% immobilised enzymes prescription, 460mg Monarch 1400 and 1.6mL 5%Nafion solution.The carbon ink water formulation that 8.5% enzyme coats is made by 40mg 8.5% immobilised enzymes prescription, 440mg Monarch 1400 and 1.6mL 5%Nafion solution.The carbon ink water formulation that 4.2% enzyme coats is made by 80mg 17% immobilised enzymes prescription, 400mg Monarch 1400 and 1.6mL 5%Nafion solution.

Discuss as in the previous examples, ink formulations has immobilised enzymes carbon and the carbon filler that passed through vortex mixed before adding Nafion solution.In case add Nafion solution, use the spatula mixed slurry, then carry out sonicated with ultrasonic dismembrator, be applied to subsequently on the ELAT carrier material.In case pole drying was pressed onto it on Nafion 115 films and the corresponding platinum black anode at 125 ℃ in following 35 seconds.The results are shown in Figure 63.

When using less modification Nafion to fix enzyme/carbon,, higher by the electrons spread speed of this layer because the thickness of separator reduces.Figure 63 shows to the enzyme increment is held constant at 0.136mg/cm ²And with the relevant adjustment (tradeoff) of amount that reduces the carbon filler.The activity of used enzyme is 120U/mg in the coating process.

Embodiment 40: biological-cathode ink formulations 5: be purchased the comparison of GCL (ELAT) and inner homemade GDL

For comparing biological-cathode catalyst carrier 1 and the carrier (being two-sided ELAT in this case) that is purchased, use the same ink preparation to carry out following program, with H ₂/ O ₂PEM biological-cathode fuel cell is tested under the same conditions.Ink formulations used in this program is made by carbon, 440mg Monarch 1400 and the 1.6mL 5%Nafion solution that the 40mg enzyme coats.ELAT GDL ink formulations is made by carbon, 440mg Monarch 1400 and the 1.6mL 5%Nafion solution that the 40mg enzyme coats.

For ink formulations, before adding Nafion solution, pass through vortex mixed immobilised enzymes carbon and carbon filler.Add after the Nafion solution, use the spatula mixed slurry, then carry out sonicated, be applied to subsequently on the ELAT carrier material with ultrasonic dismembrator.In case pole drying was pressed onto it on Nafion 115 films and the corresponding platinum black anode at 125 ℃ in following 35 seconds.Use the enzyme immobilization carbon of same batch to smear ink, the gained data are shown in Figure 64.

Use the enzyme of 120Units/mg activity to prepare the electrode ink.The inner homemade GDL ELAT GDL that compares has more high-performance under the same conditions.

Embodiment41: ink formulations 6: chitin immobilized enzyme is applied on the ELAT

Test different fixing formed material.Use the Nafion of the chitosan to replace Tetrabutylammonium bromide modification of hydrophobically modified.Any immobilization material can be used in the ink formulations, as long as they can sufficiently keep active and stable in solvent environment.In the spray drying step, sneak into modification of chitosan.In case the carbon drying that enzyme coats is sneaked into following ink formulations with it.The chitin immobilized enzyme ink formulations is made by carbon, 400mg Monarch 1400 and the 1.6mL 5%Nafion solution that the 80mg chitin immobilized enzyme coats.The ink smearing step is described identical with front embodiment, but changes pressing step.Shitosan has lower fusing and decomposition temperature, thus it be higher than under 85 ℃ the temperature unstable.Therefore, the shitosan electrode was pressed 35 seconds at 85 ℃.

The laccase that the Nafion of Tetrabutylammonium bromide modification of comparing seals, the shitosan of hydrophobically modified demonstrate performance slightly descend (seeing Figure 65).

The present invention allows various modifications and replacement form, and this paper describes its specific embodiments in detail by example.But should be understood that not to be to be intended to limit the invention to particular forms disclosed, all modifications, equivalents and the replacement that drops in the spirit and scope that claims limit contained in opposite the present invention.

Can see that from the front description some purposes of the present invention realize, and obtain other favourable outcome.Because above without departing from the present invention method can be carried out various variations, so above description comprises or all the elements shown in the drawings all are intended to be interpreted as exemplary and nonrestrictive.

When introducing the key element of the present invention or its preferred embodiment, modifier " ", " a kind of ", " described " all are intended to expression and have one or more these key elements.Term " comprises ", " comprising " and " having " be intended to represent to exist other key element outside the listed key element.

Claims (113)

1. biological fuel cell equipment that is used to produce electric current, it comprises:

The fuel chamber of confluxing, it has face, defines at least one chamber in fuel storage pond in face, is communicated with storage pond fluid and is used for fuel fluid to flow into the chamber of confluxing filling the import in storage pond, and be communicated with and be used for the conflux outlet in chamber of fuel fluid outflow with storage pond fluid;

Anode assemblies, it comprises that setting is to store up at least one biological anode that the fuel fluid in the pond contacts with described fuel;

Cathode assembly, it comprises that setting is to be used for fuel fluid flows to negative electrode by biological anode at least one negative electrode; And

Controller, it is connected with the cathode assembly operability with anode assemblies, is used for the output of Control current from described biological fuel cell equipment.

2. according to the biological fuel cell equipment of claim 1, described at least one chamber in the chamber of wherein confluxing has entrance point surface and port of export surface, and described port of export surface is positioned at the height that is higher than described entrance point surface.

3. according to the biological fuel cell equipment of claim 1, wherein said face comprises the front in the chamber of confluxing, the chamber that has first series in the described front, each chamber of first series defines fuel storage pond, described front also has the runner that connects described chamber, is used for fuel fluid flows to first series from a storage pond of first series next one storage pond.

4. according to the biological fuel cell equipment of claim 3, the wherein said chamber of confluxing also comprises the back side, and the described back side has at least one chamber that defines fuel storage pond.

5. according to the biological fuel cell equipment of claim 4, wherein the conflux described back side in chamber has the chamber of second series, each chamber of second series defines fuel storage pond, the described back side also has the runner in the described chamber that connects second series, is used for fuel fluid flows to second series from a storage pond of second series next one storage pond.

6. according to the biological fuel cell equipment of claim 5, also comprise being used for fuel fluid flows to the storage pond of second series from the storage pond of first series runner.

7. according to the biological fuel cell equipment of claim 1, wherein:

The described chamber of confluxing has a plurality of chambeies therein, and each in described a plurality of chambeies defines fuel storage pond;

Described anode assemblies, it comprises a plurality of biological anodes, each is provided with to contact with fuel fluid in the corresponding fuel storage pond in the described biological anode;

Described cathode assembly, it comprises a plurality of negative electrodes, each setting flows to respective cathode to be used for fuel fluid by biological anode in the described negative electrode; And

At least one runner is used for fuel fluid and flows to next fuel storage pond from each fuel storage pond.

8. according to the biological fuel cell equipment of claim 7, also comprise the check-valves that is connected with each runner operability, be used for allowing only to flow through described pipeline with a direction.

9. according to the biological fuel cell equipment of claim 1, wherein:

Each anode assemblies comprises the anode framework, is used to keep described at least one biological anode;

Each cathode assembly comprises the negative electrode framework, is used to keep described at least one negative electrode; And

Described anode framework and negative electrode framework are fixed on position fixing for the chamber of confluxing, and described at least one biological anode and at least one negative electrode registration each other, and with the chamber of confluxing in described at least one chamber registration.

10. according to the biological fuel cell equipment of claim 1, comprise that also its inner surface has the housing of projection, described projection is used for the chamber of confluxing, anode assemblies and cathode assembly are remained on fixed position for housing.

11. according to the biological fuel cell equipment of claim 1, wherein:

The described chamber of confluxing comprises the integral type body that is formed with a plurality of chambeies, and each chamber defines fuel storage pond;

Described anode assemblies comprises a plurality of biological anodes, and each is provided with to contact with fuel fluid in the corresponding fuel storage pond in the described biological anode; And

Described cathode assembly comprises a plurality of negative electrodes, and each is provided with to be used to receive from corresponding biological anode and flows the fuel fluid of coming in the described negative electrode.

12., wherein on relative two faces of described body, form at least two described chambeies according to the biological fuel cell equipment of claim 11.

13., wherein on same of described body, form at least two described chambeies according to the biological fuel cell equipment of claim 11.

14. according to the biological fuel cell equipment of claim 1, wherein:

Described anode assemblies comprises the anode framework with coupling frame part; And

Described biological anode is fixed between the described frame part.

15. according to the biological fuel cell equipment of claim 1, wherein said at least one biological anode comprises:

Collector body;

Gas diffusion layers; With

The catalyst layer that comprises enzyme and enzyme immobilization material.

16. according to the biological fuel cell equipment of claim 1, wherein:

Described cathode assembly comprises the negative electrode framework with coupling frame part; And

Described negative electrode is fixed between the described frame part.

17. according to the biological fuel cell equipment of claim 1, wherein said negative electrode comprises collector body, gas diffusion layers and catalyst.

18. according to the biological fuel cell equipment of claim 1, wherein said controller comprises:

The controller that is used for operational mode control fuel cell output according to the rules; And

The switching circuit that operability is connected between fuel cell and load, described switching circuit response controller are used for alternately connecting and disconnecting fuel cell and load according to operational mode.

19. a biological fuel cell equipment that is used for to load supply electric power, described equipment comprises:

Fuel cell;

Operability is connected to the controller of fuel cell, is used for the output of operational mode control fuel cell according to the rules; And

Switching circuit between fuel cell and load, described switching circuit response controller are used for alternately connecting and disconnecting fuel cell and load according to operational mode.

20. according to the biological fuel cell equipment of claim 19, wherein fuel cell comprises a plurality of fuel cells that are electrically connected in series.

21. according to the biological fuel cell equipment of claim 20, wherein operational mode comprises according to predetermined duty ratio and alternately at least one fuel cell being connected with load and fuel cell and the load disconnection that at least one is other.

22. according to the biological fuel cell equipment of claim 20, wherein operational mode comprises according to predetermined duty ratio and alternately all fuel cells is disconnected with load together and be connected.

23. according to the biological fuel cell equipment of claim 20, wherein operational mode comprises alternately one in the fuel cell is disconnected with load, and other fuel cell is connected with load.

24. the biological fuel cell equipment according to claim 19 also comprises additional power circuit, its response controller is connected with the output of fuel cell optionally to make additional power supply, replenishes thus by the electric power of described biological fuel cell equipment to the load supply.

25. biological fuel cell equipment according to claim 24, its middle controller comprises the output that is used for the comparison fuel cell and the comparator of threshold value, and described additional power circuit responds this comparison and is connected with the output of fuel cell will replenish power supply in the output of fuel cell during less than threshold value.

26., but wherein replenish the battery that power supply comprises recharge according to the biological fuel cell equipment of claim 24.

27. biological fuel cell equipment according to claim 26, its middle controller is configured to the output according to the operational mode of another regulation control fuel cell, and the output of fuel cell is connected with the load disconnection and with battery and is used for battery recharge in described operational mode.

28. a biological fuel cell equipment that is used for to load supply electric power, described equipment comprises:

Fuel cell;

Operability is connected to the controller of fuel cell, is used for the output of operational mode control fuel cell according to the rules; And

Replenish power circuit, its response controller is connected with the output of fuel cell optionally to make additional power supply, replenishes thus by the electric power of described biological fuel cell equipment to the load supply.

29. a biological fuel cell equipment that is used for to load supply electric power, described equipment comprises:

The a plurality of fuel cells that are electrically connected in series;

Operability is connected to the controller of each fuel cell, and at least one that is used for according to the operational mode of a plurality of regulations controlled the output of each fuel cell; And

Operability is connected the switching circuit between fuel cell and the load, described switching circuit response controller be used for according to operational mode optionally makes fuel cell at least one be connected with load.

30. the method that electricity is adjusted is carried out in the output to one or more fuel cells of biological fuel cell equipment, described biological fuel cell equipment is adapted to thus to load supply electric power, and described method comprises:

Between fuel cell and load, be electrically connected switching circuit; And

Switching switch circuit is connected with load optionally to make in the fuel cell at least one according in a plurality of regulation operational modes at least one.

31. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises biological anode and negative electrode; And

Described biological anode comprises gas diffusion layers, can produce at least a anode enzyme of the fuel fluid of oxidised form with the fuel fluid reaction, and the enzyme immobilization material that can fix and stablize described enzyme; Described anode endonuclease capable is discharged into gas diffusion layers with electronics, and described immobilization material is permeable to fuel fluid.

32. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises biological anode and negative electrode; And

Described biological anode comprises the anode enzyme of the electron mediator of gas diffusion layers, the fuel fluid that can produce oxidised form with the electron mediator and the fuel fluid reaction of oxidised form and reduction form, and fuel fluid and the permeable enzyme immobilization material of electron mediator, the electron mediator of described reduction form can be discharged into gas diffusion layers with electronics.

33. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises biological anode and negative electrode; And

Described biological anode comprises gas diffusion layers, the anode enzyme of the fuel fluid that can produce oxidised form and the electron mediator of reduction form with the electron mediator and the fuel fluid reaction of oxidised form, the permeable enzyme immobilization material of fuel fluid and electron mediator, and adjacent the eelctro-catalyst of electronic conductor, the described eelctro-catalyst of oxidised form can produce the electron mediator of oxidised form and the eelctro-catalyst of reduction form with the electron mediator reaction of reduction form, and the eelctro-catalyst of reduction form can be discharged into gas diffusion layers with electronics.

34. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises anode and biological-cathode;

Described biological-cathode comprises gas diffusion layers, can generate at least a cathode enzyme of water with oxidant reaction, and can fix and stablize this enzyme and the permeable enzyme immobilization material of this oxidant; Described cathode enzyme can obtain electronics from gas diffusion layers.

35. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises anode and biological-cathode;

Described biological-cathode comprises gas diffusion layers, can generate the electron mediator of oxidised form and at least a cathode enzyme of water with the electron mediator and the oxidant reaction of reduction form, and can fix and stablize this enzyme and the permeable enzyme immobilization material of this oxidant; The electron mediator of described oxidised form can obtain the electron mediator that electronics forms the reduction form from gas diffusion layers.

36. according to each biological fuel cell equipment or method among the claim 1-30, wherein:

Biological fuel cell equipment comprises anode and biological-cathode;

Described biological-cathode comprises gas diffusion layers, can generate the electron mediator of oxidised form and at least a cathode enzyme of water with the electron mediator and the oxidant reaction of reduction form, the eelctro-catalyst of oxidised form, and can fix and stablize this enzyme and the permeable enzyme immobilization material of this oxidant; Wherein, the eelctro-catalyst of described oxidised form can obtain electron production reduction form eelctro-catalyst from gas diffusion layers, described reduction form eelctro-catalyst can produce reduction form electron mediator and oxidised form eelctro-catalyst with the electron mediator reaction of oxidised form, and the eelctro-catalyst of this oxidised form can obtain the eelctro-catalyst of electron production reduction form from gas diffusion layers.

37. according to each biological fuel cell equipment among the claim 31-36, wherein biological anode or biological-cathode also comprise collector body.

38. according to claim 32,33 and 35-37 in each biological fuel cell equipment, wherein the enzyme immobilization material comprises electron mediator, eelctro-catalyst or electron mediator and eelctro-catalyst.

39. according to the biological fuel cell equipment of claim 37 or 38, the collector body material that comprises conduction electron and provide grid to support for gas diffusion layers and catalyst layer wherein.

40. according to each biological fuel cell equipment among the claim 31-39, wherein gas diffusion layers comprises the graphite worm of carbon cloth, carbon paper, carbon filament reticulated printing electrode, carbon black, carbon dust, carbon fiber, Single Walled Carbon Nanotube, double-walled carbon nano-tube, multi-walled carbon nano-tubes, carbon nano pipe array, coated with CVD (chemical vapor deposition) diamond conductor, vitreous carbon, mesoporous carbon, graphite, uncompressed, purifying flake graphite, high-performance graphite, carbon dust, pyrolytic graphite, polycrystalline graphite, epoxy resin, polytetrafluoroethylene or its combination of tearing layer open.

41. according to each biological fuel cell equipment among the claim 31-39, wherein gas diffusion layers be by have high surface be used for metastatic electron the first electrical conductivity material, be used to support second electrical conductivity material of electronic conductor and the individual layer that adhesive constitutes, wherein the weight ratio of the second electrical conductivity material and the first electrical conductivity material is at least 0.5: 1, thinks that electronic conductor provides the rigidity that enough makes its self-supporting.

42. according to each biological fuel cell equipment among the claim 31-41, wherein the anode enzyme is alcohol dehydrogenase, aldehyde dehydrogenase, hydrogenlyase, formaldehyde dehydrogenase, glucose dehydrogenase, glucose oxidase, lactic dehydrogenase, lactose dehydrogenase, pyruvic dehydrogenase, lipoxygenase, PQQ dependent form alcohol dehydrogenase or its combination.

43. according to the biological fuel cell equipment of claim 42, wherein the anode enzyme is the PQQ dependent form alcohol dehydrogenase with the PQQ that combines with enzyme.

44. according to the biological fuel cell equipment of claim 42 or 43, wherein fuel fluid comprises ammonia; methyl alcohol; ethanol; propyl alcohol; isobutanol; butanols; isopropyl alcohol; allyl alcohol; aryl alcohol; glycerine; propylene glycol; sweet mellow wine; glucuronic acid; aldehyde; carbohydrate; glucose; glucose-1; D-glucose; L-glucose; G-6-P; lactic acid; 9--phosphoric acid; D-lactic acid; L-lactic acid; fructose; galactolipin-1; galactolipin; aldose; sorbose; mannose; glyceric acid; coacetylase; acetyl group Co-A; malic acid; isocitric acid; formaldehyde; acetaldehyde; acetate; citric acid; the L-gluconic acid; beta-hydroxysteroid; alpha-hydroxysteroid; lactic aldehyde (lactaldehyde); testosterone; gluconic acid; aliphatic acid; lipid; phosphoglyceric acid; retinene; estradiol; cyclopentanol; hexadecanol; long-chain alcohol; coniferyl alcohol; cinnamyl alcohol; formic acid; long-chain aldehyde; pyruvic acid; butyraldehyde; acyl group Co-A; steroids; amino acid; riboflavin; NADH; NADH ₂, NADPH, NADPH ₂, hydrogen or its combination.

45. according to the biological fuel cell equipment of claim 44, wherein fuel fluid comprises methyl alcohol or ethanol.

46. a utilization according to the method for each biological fuel cell equipment generating among claim 1-29 or the 31-45, is included in biological anodic oxidation fuel fluid and in negative electrode or biological-cathode reduction-oxidation agent.

47. an electrode, it comprises:

Air can see through and first district of the electrical conductivity material of fuel fluid impermeable;

Fuel fluid and air be second district of permeable conductive of material all; And

Can contact the non-precious metal catalyst of fuel fluid and air,

Comprising the air self-respiration type half-cell of described electrode at room temperature, 0.4V electrode potential and 10mg/cm ²Catalyst loading produces during operation down at least about 16,20,25,30,35,40,45,50,55,60,65,70,75,80 or bigger mA/cm ²Current density.

48. according to the electrode of claim 47, wherein the permeability in first district and second district is controlled by the porosity in each district of control.

49. according to the electrode of claim 47 or 48, wherein the catalyst neighbour second district.

50. according to each electrode among the claim 47-49, wherein catalyst comprises transition metal, transition metal macrocyclic compound or its combination.

51. according to the electrode of claim 50, wherein transition metal macrocyclic compound is transition metal phthalocyanine, transition metal porphyrin, its derivative or analog or their combination.

52. according to the electrode of claim 51, wherein transition metal macrocyclic compound is FePC, phthalocyanine cobalt, PORPHYRIN IRON, Cobalt Porphyrin, its derivative or analog or their combination.

53. according to the electrode of claim 51, wherein transition metal macrocyclic compound comprises 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II), or derivatives thereof or analog.

54. one kind through heat treated electrode, it comprises:

Electronic conductor; And

At least a can optionally be the non-precious metal catalyst of water with hydrogen reduction;

Comprising the air self-respiration type half-cell of described electrode at room temperature, 0.4V electrode potential and 10mg/cm ²Catalyst loading produces during operation down at least about 16,20,25,30,35,40,45,50,55,60,65,70,75,80 or bigger mA/cm ²Current density.

55. according to the electrode of claim 54, wherein electrode also comprises carbon load polyamine, and described electrode is through the interaction of heat treatment with increase metallic atom and polyamine.

56. according to the electrode of claim 54 or 55, wherein current density is at least about 50mA/cm ²

57. according to the electrode of claim 54 or 55, wherein current density is at least about 80mA/cm ²

58. according to each electrode among the claim 54-57, wherein electronic conductor comprises at least a carbon-based material, and described carbon-based material is the graphite worm of carbon cloth, carbon paper, carbon filament reticulated printing electrode, carbon black, carbon dust, carbon fiber, Single Walled Carbon Nanotube, double-walled carbon nano-tube, multi-walled carbon nano-tubes, carbon nano pipe array, coated with CVD (chemical vapor deposition) diamond conductor, vitreous carbon, mesoporous carbon, graphite, uncompressed, purifying flake graphite, high-performance graphite, high order pyrolytic graphite, pyrolytic graphite, polycrystalline graphite or its combination of tearing layer open.

59. according to each electrode among the claim 54-58, wherein non-precious metal catalyst is the tolerance fuel fluid, particularly tolerance alcohol.

60. according to the electrode of claim 59, wherein non-precious metal catalyst is acidproof.61. according to each electrode among the claim 54-60, wherein non-precious metal catalyst comprises 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II), or derivatives thereof or analog.

62. a catalyst comprises 1,2,3,4,8,9,10,11,15,16,17,18,22,23,24,25-ten hexafluoros-29H, 31H-phthalocyanine cobalt (II) (CoPcF) and polypyrrole, wherein CoPcF and polypyrrole through Overheating Treatment to increase the interaction between cobalt metallic atom and the polypyrrole nitrogen-atoms.

63. according to the catalyst of claim 62, wherein polypyrrole loads on the carbon.

64. an electrode, it comprises:

Air can see through and first district of the electrical conductivity material of fuel fluid impermeable;

Second district of the permeable substantially conductive of material of fuel fluid and air; And

Can contact the catalyst of fuel fluid and air.

65. a biological-cathode, it comprises:

(a) electronic conductor;

(b) can generate at least a cathode enzyme of water with oxidant reaction; And

(c) can fix and stablize the permeable enzyme immobilization material of this enzyme and this oxidant, wherein electronic conductor comprise functionalized multi-walled carbon nano-tubes, based on material or its combination of activated carbon.

66. according to the biological-cathode of claim 65, wherein the material based on activated carbon comprises carbon black, wherein said carbon black once was heated to about 600 ℃-Yue 900 ℃ and also then soaked in water.

67. according to the biological-cathode of claim 65, wherein functionalized multi-walled carbon nano-tubes is functionalized by at least one hydroxyl or carboxylic group or its combination.

68. according to each biological-cathode among the claim 65-67, wherein enzyme is bilirubin oxidase, laccase, superoxide dismutase, peroxidase or its combination.

69. particle that contains the nuclear core that is coated with immobilised enzymes, described enzyme be fixed in the immobilization material also (i) have with respect to its in immobilization and the initial activity before coating at least about 0.65,0.7,0.75,0.8,0.85,0.9,0.95 or bigger activity, or (ii) when described enzyme continuous catalysis chemical conversion, keep it at least about 75% initial catalytic activity at least 1,2,3,4,5,6 or 7 days.

70. according to the particle of claim 69, wherein said enzyme keeps its initial catalytic activity at least 5 of at least 75%, 10,15,20,25,30,45,60,75,90,105,120,150,180,210,240,270,300,330,365,400,450,500,550,600,650,700,730 or more days.

71. according to the particle of claim 69 or 70, wherein said enzyme is fixed in the enzyme immobilization material, this material is the described enzyme of covalent bond not.

72. according to each particle among the claim 69-71, wherein said nuclear core is a kernel.

73. according to each particle among the claim 69-72, wherein said nuclear core is not a polymer.

74. according to each particle among the claim 69-73, the enzyme that wherein is immobilized is cambium layer on the nuclear core.

75. according to the particle of claim 74, wherein said layer is continuous.

76. according to each particle among the claim 69-75, wherein said enzyme keeps it at least about 75% initial catalytic activity at least 30 days when the continuous catalysis chemical conversion.

77. according to each particle among the claim 69-76, wherein said enzyme have with respect to its in immobilization and the activity before coating at least about 0.7,0.75,0.8,0.85,0.9,0.95 or higher activity.

78. a particle that contains the nuclear core that is coated with the immobilized cell device, described organelle is fixed in the immobilization material.

79. according to the particle of claim 78, wherein organelle is glyoxysome, peroxisome, mitochondria, mitochondria matter, thylakoid, chloroplaset, hydrogenosome or its combination.

80. method for preparing the particle that is coated with immobilised enzymes or organelle, described method comprises: mix solution that contains enzyme or organelle and the suspension that contains at least a nuclear core particle, immobilization material and liquid medium with the formation mixture, and with described mixture spray drying.

81. 0 method is wherein used spray gun to carry out spray drying, and described mixture is spray dried on the surface according to Claim 8.

82. 0 or 81 method according to Claim 8, wherein liquid medium is solvent, buffer solution, ionic liquid or its combination.

83. according to each particle or method among the claim 69-82, its center core particle is polymer particle, carbon particle, zeolite particles, metallic, ceramic particle, metal oxide particle, the insoluble organic metal particle of reaction medium or its combination.

84. according to each particle or method among claim 69-77 and the 80-83, wherein said enzyme comprises lipase; glucose isomerase; nitrilase; glucose oxidase; protease; pepsin; amylase; fungal amylase; produce maltogenic amylase; cellulase; lactase; esterase; carbohydrase; hemicellulase; pentosanase; zytase; amylopectase; 1,4 beta-glucanase; acetolactate decarboxylase; β-Pu Tangganmei; glutaminase; PA ase; chloroperoxidase; aspartic acid β-decarboxylase; cyclodextrin glycosyl transferases; subtilopeptidase A; amino-acylase; alcohol dehydrogenase; amino acid oxidase; phosphatidase; urase; cholesterase; desulfinase; lignin peroxidase; pectase; oxidoreducing enzyme; dextranase; glucosidase; galactosidase; glucoamylase; maltose; invertase; invertase; naringinase (naringanase); bromelain; ficin; papain; pepsin; peptase; renin; thermolysin; trypsase; the triacylglycerol esterase; glandular stomach (pregastric) esterase; phosphatase; phytase; amidase; glutaminase; lysozyme; catalase; dehydrogenase; peroxidase; lyase; fumarase; histidase (histadase); aminopherase; ligase; cyclase; racemase; mutase; oxidizing ferment; reductase; lignoenzyme or its combination.

85. according to each particle or method among the claim 69-84, wherein said particle comprises about 90wt% nuclear core of about 25wt%-and the about 75wt% coating of about 10wt%-.

86. 5 particle according to Claim 8, wherein coating comprises the about 29wt% enzyme of about 0.1wt%-, about 0.1wt%-about 43wt% enzyme immobilization material and about at the most 29wt% electron mediator.

87. each particle or method among the 0-84 according to Claim 8, wherein said solution comprises about 15wt% enzyme of about 0.1wt%-and the about 99.9wt% solvent of about 85wt%-, and described suspension comprises the about 28.7wt% nuclear of about 0.1wt%-core particle, about 4wt%-about 10wt% enzyme immobilization material and the about 75wt% liquid medium of about 50wt%-.

88. a self-bearing type electronic conductor comprises:

By have high surface be used for metastatic electron the first electrical conductivity material, be used to support second electrical conductivity material of electronic conductor and the individual layer that adhesive is formed, wherein the weight ratio of the second electrical conductivity material and the first electrical conductivity material is at least 0.5: 1, thinks that described electronic conductor provides the rigidity that enough makes its self-supporting.

89. 8 electronic conductor according to Claim 8, wherein the weight ratio of the second electrical conductivity material and the first electrical conductivity material is at least 0.6,0.7,0.8,0.9 or 1: 1.

90. 8 or 89 electronic conductor according to Claim 8, wherein electron mediator be grafted to the first electrical conductivity material to small part.

91. each electronic conductor among the 8-90 also comprises the collector body that contacts with individual layer according to Claim 8, wherein this electronic conductor does not comprise the structural support except that the structural support that collector body and the second electrical conductivity material provide.

92. according to the electronic conductor of claim 91, wherein collector body has along the near-end and the far-end of the longitudinal axis and limits certain-length betwixt, and described individual layer contacts with collector body and from the coaxial far-end that extends to collector body of the near-end of collector body.

93. each electronic conductor among the 8-92 according to Claim 8, wherein the weight ratio of the adhesive and the second electrical conductivity material is for being at least 0.8,0.9 or 1: 1.

94. each electronic conductor among the 8-93 according to Claim 8, wherein the first electrical conductivity material comprises carbon black, and the second electrical conductivity material comprises carbon fiber, and adhesive comprises polyvinylidene fluoride or polytetrafluoroethylene.

95. each electronic conductor among the 8-93 according to Claim 8, wherein individual layer also comprise claim 69-77,83 or 85-87 in each particle.

96. each electronic conductor among the 8-94 according to Claim 8, wherein the enzyme layer contacts with the surface of described individual layer, and described enzyme layer comprises and is fixed in the enzyme immobilization material and the enzyme that contacts with the nanostructure that is made of the nano wire that is grafted on the carbon black particle.

97. according to each biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or technology among claim 15,31-47, the 65-87 or 96, wherein the enzyme immobilization material comprises micella or reverse micelle structure.

98. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 97, wherein the enzyme immobilization material comprises the perfluorinated sulfonic acid-PTFE copolymer of modification or the alginates of modification.

99. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 98, wherein enzyme immobilization material quilt is greater than NH ₄ ⁺The dewatering cationic modification.

100. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 99, wherein dewatering cationic comprises ammonium cation, quaternary ammonium cation, alkyl trimethyl ammonium cation, organic cation, phosphonium cation, triphenyl phosphonium, pyridylium, glyoxaline cation, cetyl pyridinium, second ingot, viologen, methyl viologen, benzyl viologen, two (triphenylphosphine) imonium, metal complex, Bipyridine metal complexes, phenanthrolines Base Metal complex compound, [Ru (bipyridine) ₃] ²⁺Or [Fe (phenanthrolines) ₃] ³⁺

101. biological fuel cell equipment according to claim 99, method, biological-cathode, particle, electronic conductor or technology, wherein dewatering cationic comprises tetrapropyl ammonium (T3A), four pentyl ammonium (T5A), tetrahexyl ammonium (T6A), four heptyl ammoniums (T7A), trimethyl icosyl ammonium (TMICA), trimethyl octadecyl ammonium (TMODA), trimethyl cetyltrimethyl ammonium (TMHDA), trimethyl myristyl ammonium (TMTDA), trimethyl octyl group ammonium (TMOA), trimethyldodecane base ammonium (TMDDA), trimethyl decyl ammonium (TMDA), trimethyl hexyl ammonium (TMHA), TBuA (TBA), triethyl group hexyl ammonium (TEHA) and combination thereof.

102. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 99, wherein dewatering cationic comprises the quaternary ammonium ion by following formula 4 expressions:

R wherein ₁, R ₂, R ₃And R ₄Be the alkyl or the heterocycle of hydrogen, alkyl, replacement, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.

103. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 102, wherein R ₁, R ₂, R ₃And R ₄Be hydrogen, methyl, ethyl, propyl group, butyl, amyl group, hexyl, heptyl, octyl group, nonyl or decyl, wherein R independently ₁, R ₂, R ₃And R ₄In at least one is not a hydrogen.

104. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 102, wherein R ₁, R ₂, R ₃And R ₄Identical and be methyl, ethyl, propyl group, butyl, amyl group or hexyl.

105. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 102, wherein R ₁, R ₂, R ₃And R ₄In one be hexyl, octyl group, decyl, dodecyl or myristyl, and other is methyl, ethyl or propyl group independently.

106. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 102, wherein R ₁, R ₂, R ₃And R ₄It is butyl.

107. according to each biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or technology among claim 15,31-47, the 65-87 or 96, wherein immobilization material is the polysaccharide of micellelike hydrophobically modified.

108. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 107, wherein polysaccharide comprises shitosan.

109. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 107 or 108, wherein the polysaccharide of micellelike hydrophobically modified is corresponding to formula 2:

Wherein n is an integer; R ₁₀Be the alkyl or the hydrophobic redox mediators of hydrogen, alkyl, replacement independently; And R ₁₁Be the alkyl or the hydrophobic redox mediators of hydrogen, alkyl, replacement independently.

110. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 109, wherein R ₁₀Be hydrogen or alkyl independently, R ₁₁Be hydrogen or alkyl independently.

111. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 109, wherein R ₁₀Be hydrogen or hexyl independently, R ₁₁Be hydrogen or hexyl independently.

112. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 109, wherein R ₁₀Be hydrogen or octyl group independently, R ₁₁Be hydrogen or octyl group independently.

113. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 109, wherein R ₁₀Be hydrogen or butyl independently, R ₁₁Be hydrogen or butyl independently.

114. according to biological fuel cell equipment, method, biological-cathode, particle, electronic conductor or the technology of claim 109, wherein R ₁₀Be hydrogen or hydrophobic redox mediators independently, R ₁₁Be hydrogen or hydrophobic redox mediators independently.

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