JPH07104314B2 - Carbon sensor electrode and method of manufacturing the same - Google Patents

Description

Detailed Description of the Invention

[0001]

This invention relates to an electrochemical detector,
The present invention relates to a carbon sensor electrode used for a sensor for environmental analysis, a sensor for pathological examination, a probe electrode for detecting a physiologically active substance or the like in a biological system, and a method for producing the same.

More specifically, pure carbon fine particles, short carbon fibers, etc. are used as an electrode material, and these are dispersed and mixed with an electron accepting / donating substance such as an enzyme, a metal complex, an organic substance or a metabolite in a liquid for insulation. The present invention relates to a carbon sensor electrode, which is held in a flexible tube, and a manufacturing method thereof. The electron-accepting / donating substance means a substance that reacts chemically or electrochemically with a compound to be measured directly or through a film to accept an electron from the compound to be measured or donate an electron to the compound to be measured. To do. Hereinafter, in order to simplify the description of the electron accepting / donating substance, it will be referred to as a reactant or a reactant.

[0003]

2. Description of the Related Art As a major field of electrochemical measurement,
In recent years, the simple rapid analysis method using a sensor that has been rapidly developed as a detection method has extremely high selectivity and enables highly sensitive measurement. Therefore, the measurement target component is extremely small and many compounds coexist. It is beginning to be actively used for analysis and evaluation of clinical biological samples and environmental samples.

Further, in recent years, by using a sensor electrode capable of detecting such a specific substance with high sensitivity and speed, in a living body system or the like, in a living body (in vivo) or in situ (in situ), Getting information has become very important. For this purpose, it is required to dispose an electrode that can be easily manufactured or to have a structure in which a new electrode surface can be easily obtained, thereby enabling selective measurement of a specific substance in a living body.

Conventionally, as such measuring electrodes, a pH meter for measuring hydrogen ions, an ion sensor for detecting inorganic ions such as sodium ions, carbon fiber (CF)
Various types of sensors, such as a chemically modified complex or organic substance on the surface of glass or glassy carbon (GC), a glucose sensor in which an enzyme such as glucose oxidase is mixed with a paste of a carbon paste (CP) electrode and covered with an ion permeable membrane The enzyme sensor, the biosensor that retains the immune metabolites, etc. have been studied and are being put to practical use.

However, except for the pH meter and the inorganic ion sensor, most of the chemically modified sensor, the enzyme sensor and the biosensor cannot withstand long-term use and have a short life.

[0007]

In order to solve this life problem, it is necessary to make a disposable sensor electrode or to construct a sensor having a mechanism capable of constantly supplying a new reactant to the electrode surface.

A commercially available carbon paste (CP) electrode prepared by mixing carbon fine particles with a liquid has electrode carbon particles mixed and held in a liquid at the tip of the electrode because of its constitution.
Some reactants were mixed simultaneously and some were used as disposable sensor electrodes. However, this CP electrode has to be handmade every time, and since it is handmade, the mixed state and the electrode surface are often slightly different each time it is made.

[0009]

[Means for Solving the Problems] A necessary condition for a sensor electrode is that the potential window is large and the blank current is small.

It is possible to react with a specific substance with good reproducibility.

Electrode reaction activity.

There is no solid difference in electrode characteristics.

The content of impurities is small and the electrode reaction is not hindered.

Easy handling and pretreatment.

[0015] These characteristics are obtained by filling the carbon paste (CP) prepared by mixing the carbon fine particles and the reaction substance with the liquid in a thin insulating tube and removing the used portion. Can also be achieved. Further, if the carbon paste containing a reactant is pushed out by a piston or a low speed pump, it can be used as a sensor electrode having a new electrode surface at all times.

The biological system to be measured includes proteins,
Since a large amount of adsorptive substances such as lipids coexist, and these organic substances once adsorbed on the carbon surface are difficult to remove unless they are removed by oxidation, the electrodes should be either disposable or the electrodes that have already been used should be removed. Consideration is required. Further, re-use of the same sensor used for another person's sample in a clinical test causes a detection error. Therefore, the inventors have invented a structure having a disposable property or an extrusion property. The conditions supplied to such a sensor electrode are listed as follows: (1) Friendly to biological systems.

(2) High activity as an electrode.

(3) Reactants such as enzymes can be kept active in the sensor.

(4) It is easy and sufficient to keep a reaction product such as an enzyme in the sensor active.

(5) It is possible to fold (cut) or extrude and remove the used electrode part.

(6) It is inexpensive enough to be disposable.

The sensor electrode intended for disposable use must satisfy at least the conditions of the above six items. For this purpose, it is desirable to have a sensor electrode that allows desired reactants such as an enzyme, a metal complex, an organic substance, and a metabolite to be always present on the surface of the electrode and can be produced economically.

The carbon sensor electrode of the present invention is characterized in that an insulating outer layer is filled with a carbon paste, and the carbon paste comprises at least a carbon fine material and a carrier.

Further, in the method for manufacturing a carbon sensor electrode of the present invention, at least a carbon fine material and a carrier are uniformly mixed with a dispersion medium to prepare a carbon paste, and the carbon paste is incorporated into a cylindrical outer layer. It is a method characterized by that.

Here, the carbon fine material is preferably selected from activated carbon particles, graphite particles, amorphous carbon particles, short carbon fibers, and a mixture thereof.

As the carrier, at least one of petrolatum, glycerin, oil, paraffin, agar and gelatin is preferably used.

The carbon paste preferably contains a reactive substance.

The reactive substances include (active enzymes, nitrogen metabolites such as uric acid, amino acid metabolites, neurotransmitters such as dopamine, vitamins, organic acids such as acetic acid and citric acid, amines, alcohols. , Organic substances (including anesthetics and drugs such as drugs), complex ions, inorganic acids (such as hydrochloric acid and sulfuric acid), alkalis (such as ammonia and hydroxide), salts of the inorganic acids and alkalis (oxygen) And any soluble gas, such as chlorine, chlorine, nitric oxides, etc., and mixtures thereof.

Further, as the dispersion medium for mixing the reactant and the carbon fine material, preferably, an aqueous solution, an organic solvent, or a mixture thereof is used.

As the insulating outer layer, an insulating tube or an insulating sheet is preferably used.

Further, the tip opening of the insulating outer layer is
It may be sealed with an ion permeable membrane or a dialysis membrane.

Furthermore, a carbon paste supply device capable of continuously supplying the carbon paste is further connected to the base end portion of the outer layer so that a fresh sensor end can always be obtained. May be.

The present invention will be further described below with reference to the drawings.

In the carbon sensor electrode of the present invention, the outer layer 1 is an insulating tube, as shown in the schematic structure of FIG. 1, and as shown in FIG. And the holding substance (carrier) are mixed together to form a carbon paste 4
And this is filled. This electrode is used with the sensor part facing up or in a hanging type.

As the reaction substance 2, (active enzyme,
Organic compounds, including nitrogen metabolites such as uric acid, amino acid metabolites, neurotransmitters such as dopamine, vitamins, organic acids such as acetic acid and citric acid, amines, alcohols, pharmaceuticals such as anesthetics and drugs) Ions, inorganic acids (such as hydrochloric acid and sulfuric acid), alkalis (such as ammonia and hydroxide), salts of said inorganic acids and alkalis, soluble gases (such as oxygen, chlorine and nitric oxides), and mixtures thereof. Any of these, or those containing these are used. In order to prevent these reactants 2 contained in the tube from being immediately washed out into the bulk solution during the reaction, vaseline,
Coexist with a holding substance such as glycerin, oil, paraffin, agar, and gelatin.

The tip of the tube may be covered with an ion exchange membrane having a low electric resistance, a conductive polymer membrane, a dialysis membrane or the like to prevent the liquid from flowing out.

As a method for producing this electrode, first, petroleum jelly, glycerin, oil, paraffin, agar, and so on, for holding the reactant 2 so as not to immediately flow into the solution,
A carbon material 3 made of carbon fine particles or short carbon fibers is sufficiently dispersed and mixed with water and an organic solvent together with gelatin or the like to prepare a carbon paste 4. Next, insulate this carbon paste 4 with a piston or the like (outer layer 1)
It is pushed into the inside or wrapped with an insulating film (outer layer 1) in a so-called "paste-like" manner. This creates the sensor electrode.

In the electrode of the present invention, it is possible to construct a sensor in which the reaction substance is not washed away due to the viscous holding substance.

In the electrode of the present invention, the sensor portion (electrode tip) that has been used once should be cut (cut) to always use a new electrode surface, or the paste of the used tip portion should be extruded to remove it. Reproducible results are obtained by providing an electrode surface.

Further, even when the holding substance is rarely made to coexist, the paste can be held in the pipe and used as a sensor by making the pipe slightly negative pressure by a piston or the like.

Further, in the electrode of the present invention, as shown in FIG. 1, a connection port 5 is provided at one end (base end) of the pipe 1, and a piston (syringe) 6 is provided in the connection port 5 via a tube (not shown).
Alternatively, the pump 7 may be connected to supply new carbon paste.

In the above constitution, nothing is mixed with the enzyme and the like, and a mixture prepared by kneading in a paste form with a solvent is extruded so as not to disperse in a solution, and a new carbon paste surface is produced. Can be used as a carbon paste suspension electrode and a carbon paste disc electrode.

[0043]

The carbon sensor electrode of the present invention having the above-described structure contains a carbon fine material and preferably contains a carbon paste containing a reactant such as an enzyme in an insulative outer layer, and easily pushes out a deteriorated tip portion for use, It is a structure that can be removed, so that a fresh sensor electrode surface can be easily provided over a long period of time, which is extremely practical.

[0044]

EXAMPLES Examples of the present invention will be described below, but the present invention is not limited to these examples.

(Example 1) 35% of liquid paraffin was added to the amount of graphite particles (mechanical pencil core powder), and the particles were well dispersed and kneaded, and then poured into a syringe without a needle to leave both ends empty. The paste was injected with the piston of the syringe from one end of a 1.5 mm diameter silicone tube.

After cutting off one end of the tube in which the paste was sufficiently introduced and flattening the electrode surface with a smooth sheet of paraffin paper or the like, a dialysis membrane (manufactured by Union Carbide), which was slightly cut into large circles, was attached to the tube. It was put on the tip and fixed with an O-ring.

Uric acid in urine was detected using this electrode. Since the pH of human urine is generally around 6.5, the measurement system used was a fixed amount of human urine added to a known amount of Ringer's solution whose pH was adjusted to 6.5. The lactam type of uric acid has three quinone groups -C = 0, one of which is a lactim type hydroxyl group-O due to the redox reaction.
Since a rapid electrode reaction occurs with H, it shows a redox wave that is easy to observe. Its redox potential is about 0.5 V vs. this pH. SCE, the reduction wave is at a much negative potential. The results are shown in FIG. No wave is observed in the blank Ringer's solution as in (a) in the figure, but 1
The case where 50 ml of uric acid is contained in the ml solution is (b),
The result (c) due to uric acid in human urine is slightly less than that in (b). If you have created a calibration curve, you can easily quantify. However, if left soaked for about 5 minutes or longer, other components in urine seemed to be adsorbed, and the wave height gradually decreased. Since this adsorbed substance cannot be removed without strong oxidizing treatment, it is desirable to cut off or extrude the paste electrode at the tip of the electrode once it has been used, and perform the next quantitative determination on a new electrode surface. Also, when a large amount of uric acid is added to human urine, precipitation occurs, which hinders quantification.

(Example 2) In most natural enzymatic reactions, hydrogen peroxide is produced after the reaction is completed. Therefore, the enzymatic reaction progresses by measuring the content of hydrogen peroxide in the solution. Can be guessed. The final target is a sensor electrode with a built-in peroxidase in native horseradish, but instead, iron EDTA is used to detect hydrogen peroxide due to the electrochemical catalytic reaction shown in FIG. Can be done easily.

Therefore, Fe-EDTA is added to glycerin 80.
%, A saturated pseudo enzyme was kneaded into the solution to try to detect hydrogen peroxide in the solution. Graphite particles (mechanical pencil core powder) were added to an 80% aqueous solution of glycerin (manufactured by Wako Pure Chemical Industries), and the mixture was well kneaded, then poured into a syringe without a needle, and the ends of the tube with a diameter of 0.5 mm were vacant. The paste was injected with the piston of a syringe to create a carbon sensor electrode with a built-in pseudo enzyme.

Using this electrode, an attempt was made to detect 0.03% amount of hydrogen peroxide present in an acetic acid buffer solution having a pH of 5.5. The result is shown in FIG. Curve (a) is a voltammogram of iron EDTA with a sensor electrode containing iron EDTA, and curve (b) is 0.03 using this sensor electrode.
3 is a voltammogram obtained in a solution containing% hydrogen peroxide. It can be clearly observed that the wave height is increased by hydrogen peroxide.

Also, an attempt was made to detect 0.03% hydrogen peroxide by using a paste electrode without iron EDTA. The result is the curve (c).

[0052]

As described above, the implementation of the reactant-embedded carbon sensor electrode according to the present invention replaces a sensor or a sensor electrode which has not been used for a long period of time with a specific one of many biological samples. It is possible to provide a disposable carbon sensor electrode for in situ orientation for substance detection.

That is, the present carbon sensor electrode is a sensor electrode having a structure that can be used for a long period mainly composed of carbon paste in which a reaction substance such as an enzyme is contained in an insulating tube.
It becomes a practical sensor electrode that can easily provide a new sensor electrode surface by folding (cutting) the tip to dispose of it or extruding it.

[Brief description of drawings]

FIG. 1 is a schematic configuration diagram of a sensor electrode of the present invention.

FIG. 2 is an enlarged configuration diagram of a portion II in FIG.

FIG. 3 is a voltammogram of uric acid at a sensor electrode of the present invention that does not contain a reactant.

FIG. 4 is a conceptual diagram of an enzyme-like reaction with iron EDTA.

FIG. 5 is a graph showing a cyclic voltammogram of iron EDTA by the sensor electrode of the present invention and a detection result of hydrogen peroxide by the sensor electrode.

[Explanation of symbols]

 1 Outer layer 2 Reactive substance 3 Carbon fine material 4 Carbon paste 5 Connection port 6 Piston (syringe) 7 Pump

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI technical display location G01N 27/327 27/38 27/404

Claims (18)

[Claims] 1. An insulating outer layer is continuously filled with a carbon paste from the end to the tip of the outer layer, and the carbon paste is composed of at least a carbon fine material, a carrier, and an electron accepting / donating substance. The carbon sensor electrode is characterized by being uniformly mixed with each other. 2. The carbon sensor electrode according to claim 1, wherein the carbon fine material is selected from activated carbon particles, graphite particles, amorphous carbon particles, short carbon fibers, and a mixture thereof. 3. The carbon sensor electrode according to claim 1, wherein the carrier is at least one of aqueous solution, petrolatum, glycerin, oil, paraffin, agar, and gelatin. 4. The electron-accepting / donating substance is an active enzyme, nitrogen metabolites such as uric acid, amino acid metabolites, neurotransmitters such as dopamine, vitamins, organic acids such as acetic acid and citric acid, and amines. , Organic substances including alcohols, anesthetics and drugs, complex ions, inorganic acids such as hydrochloric acid and sulfuric acid, alkalis such as ammonia and hydroxides, salts of the inorganic acids and alkalis, oxygen, chlorine and oxidation A soluble gas such as nitrogen, and any of these mixtures,
Alternatively, the carbon sensor electrode according to any one of claims 1 to 3, which contains the above. 5. A dispersion medium is used to mix the electron accepting / donating substance and the carbon fine material, and the dispersion medium is any one of an aqueous solution, an organic solvent, and a mixture thereof. The carbon sensor electrode according to any one of claims 1 to 4. 6. The carbon sensor electrode according to claim 1, wherein the insulating outer layer is an insulating tube. 7. The carbon sensor electrode according to claim 1, wherein the insulating outer layer is an insulating sheet. 8. The carbon sensor electrode according to claim 1, wherein the tip end opening of the insulating outer layer is sealed with an ion permeable membrane or a dialysis membrane. 9. The carbon paste supply device capable of continuously supplying the carbon paste is connected to a base end portion of the outer layer.
The carbon sensor electrode according to any one of 1. 10. A carbon paste is prepared by uniformly mixing at least a carbon fine material, a carrier, and an electron accepting / donating substance with a dispersion medium, and the carbon paste is applied to a cylindrical outer layer from the end of the outer layer to the tip. A method of manufacturing a carbon sensor electrode, which is characterized in that the carbon sensor electrode is continuously built in over a period of time. 11. The activated carbon particles as the carbon fine material,
The method for producing a carbon sensor electrode according to claim 10, wherein the carbon sensor electrode is selected from graphite particles, amorphous carbon particles, short carbon fibers, and a mixture thereof. 12. An aqueous solution, petrolatum, as the carrier
At least one of glycerin, oil, paraffin, agar, and gelatin is used, The manufacturing method of the carbon sensor electrode of Claim 10 or 11 characterized by the above-mentioned. 13. As said electron accepting / donating substance, active enzymes, nitrogen metabolites such as uric acid, amino acid metabolites, neurotransmitters such as dopamine, vitamins, organic acids such as acetic acid and citric acid, amines. , Organic substances including alcohols, anesthetics and drugs, complex ions, inorganic acids such as hydrochloric acid and sulfuric acid, alkalis such as ammonia and hydroxides, salts of the inorganic acids and alkalis, oxygen, chlorine and oxidation 13. The method for producing a carbon sensor electrode according to claim 10, wherein any one of a soluble gas such as nitrogens and a mixture thereof, or one containing them is used. 14. An aqueous solution, an organic solvent, as a dispersion medium for mixing the electron accepting / donating substance and the carbon fine material,
14. The method for manufacturing a carbon sensor electrode according to claim 10, wherein any one of these and a mixture thereof is used. 15. The insulating tube is used as the insulating outer layer, according to claim 10.
5. The method for manufacturing a carbon sensor electrode according to any one of 1. 16. The method for manufacturing a carbon sensor electrode according to claim 10, wherein an insulating sheet is used as the insulating outer layer. 17. The method for manufacturing a carbon sensor electrode according to claim 10, wherein the tip end opening of the insulating outer layer is sealed with an ion permeable membrane or a dialysis membrane. 18. The carbon paste supply device capable of continuously supplying the carbon paste is further connected to the base end portion of the outer layer, and the carbon paste supply device is connected to the carbon paste supply device. Manufacturing method of carbon sensor electrode of.