JPH02165046A - Enzyme function electrode - Google Patents

Abstract

PURPOSE:To obtain an enzyme function electrode which is flexible and easy to work with and reaction controllable from outside by plating a cloth foil comprising non-metallic fiber with metal to form an electrode, on which an enzyme is immobilized directly. CONSTITUTION:A cloth foil comprising plant fiber, animal fiber, artificial fiber or inorganic fibers as non-metallic fiber is plated with metal to form an electrode and an enzyme is solidified directly thereon. The cloth foil herein used can be any of cotton, hemp and wool with titer being about 1-100 denier and bent and mixed fiber may be used. Metal of the electrode herein used is nickel, copper or the like. The cloth foil is plated with nickel, copper or the like and then with gold or silver to form an electrode. The solidification of the enzyme may employ a proper immobilization method but normally, the enzyme is contained in a film forming resin liquid such as photosetting resin to coat. Any enzyme is applicable only if electrons are given and taken in reaction. Being an cloth-like electrode, the electrode thus obtained is large in flexibility, light and easy to cut and the immobilization of the enzyme on the surface of the electrode allows the controlling of a reaction such as that for changing voltage from outside.

Description

[Detailed Description of the Invention] (Industrial Application Field) The present invention provides an enzyme function for biosensors and bioreactors that is flexible, has a large surface area, is lightweight, easy to cut, and has a wide range of applications! Regarding poles.

(Prior art) Enzyme-functional electrodes for biosensors are used as a means of measuring the concentration of biochemical substances in a liquid to be measured, by using an enzyme to cause a reaction between the biochemical substances to be measured and measuring the concentration of the product using electricity. It is used to quantify substrates that indirectly contribute to enzymatic reactions.

Conventionally, enzyme electrodes used in the above-mentioned biosensors are known to consist of an active electrode and a reference electrode. This electrolyte uses a polymer membrane in which an enzyme whose substrate is the biochemical substance to be measured is immobilized on a base electrode made of platinum, and one of the products of the enzymatic reaction by this enzyme is selected and permeated. It consists of a film that

(Problems to be Solved by the Invention) As mentioned above, a base electrode made of platinum is normally used for enzyme electrodes.
No. 57 forms a polymer film on the very surface of the base, and in this polymer film, an enzyme that is contained in the sample to be measured and uses the biochemical substance to be measured as a substrate is oxidized or reduced as a result of the reaction by this enzyme. This invention discloses an enzyme electrode consisting of a working electrode formed by fixing an acid (arsenic reducing substance) and a reference electrode which is immersed together with the working electrode in a sample to be measured.
The platinum plate itself is used. The Pond Special Publication Showa 62-
Various enzyme functional electrodes are disclosed in No. 215861, JP-A No. 62-144C161, etc., but the enzyme electrodes disclosed in these documents use a platinum plate itself as a cathode, an enzyme-permeable membrane, an enzyme-immobilized membrane, and This is an enzyme electrode formed by laminating semipermeable membranes.

The above-mentioned electrodes, platinum electrodes, etc. are inconvenient when it is desired to change the shape of the electrode depending on the shape of the sample container to be measured, since they are flat and lack flexibility, so they cannot be bent freely. It also has problems in that it cannot be cut easily and has poor workability, and the surfaces of platinum electrodes and the like are flat and have a small surface area, resulting in low reactivity and low sensitivity.

Recently, it has become possible to use enzymes as polar bioreactors by immobilizing enzymes directly on the surface of the pole, and by changing the voltage applied to the pole to trigger the reaction from the outside. . At this time, reactivity is important in bioreactors. In this case, there are problems in that the platinum electrode has a small surface area and therefore has low reactivity and low sensitivity.

It is required that bioreactor enzyme electrodes can be packed into a reaction tower in a desired shape, and that they be lightweight as the amount packed increases, but platinum electrodes and the like have many problems in this respect as well.

(Means for Solving the Problems) An object of the present invention is to solve the drawbacks of the above-mentioned prior art.

The enzyme functional electrode of the present invention includes vegetable fibers that are non-metallic fibers;
A fabric made of animal fiber, artificial fiber, or inorganic fiber is plated with metal and an enzyme is directly fixed to it.
Surface unevenness, lightness, ease of cutting, and tFf of metal
1 performance is very effectively combined, and the drawbacks of the prior art are eliminated.

The flexibility of the enzyme function of the present invention is JIS109
6 According to the rigidity A method (the rigidity force method Chilever method), 10m
It is a soft material with a diameter of 130 mm to 130 mm. It should be noted that it is virtually impossible to measure with a normal platinum electrode at a length of 200 mm or more.

The cloth foil used in the present invention includes vegetable fibers such as cotton and linen, animal fibers such as wool and silk, man-made fibers such as cupro, acetate, polyester, polyamide, polyacrylonitrile, polyurethane, and polypropylene. Whichever fiber you prefer, each of these alone or a blend of two or more,
A mixed fiber form is also acceptable. The form of cloth foil is woven fabric, knitted fabric, lace, net, non-woven fabric, paper, etc. The preferred fineness is 1
Denier to 100 denier, but not limited thereto. Thin threads of 1 denier or less are acceptable, but production of thin threads of 1 denier or less requires advanced technology and increases costs. Moreover, if it is more than 1000 deniers, it will become hard and it will be difficult to obtain an electrode with flexibility, which is the object of the present invention.

The metal used for the electrode in the present invention includes gold, silver, platinum, etc., and gold and silver are particularly preferred.

These may be plated directly onto the non-metallic fiber cloth, but it is also preferable to plate them with nickel, copper or the like. As the plating method, electroless plating is preferred. Electrolytic plating may be performed after metal is plated in advance by an electroless plating method.

The plating thickness is preferably 0.05μ, but is not limited thereto. If it is less than 0.05 μm, it tends to cause poor isoelectricity and film strength, and if it is more than 1 μm, it may impair the flexibility, which is the object of the present invention, and the cost will increase, which is not preferable.

In this way, a strong metallization is achieved at low cost.

The enzyme can be immobilized onto the metallized cloth by an appropriate immobilization method.9 Usually, the enzyme or enzyme solution is contained in a solution or dispersion of a film-forming resin such as a photocuring resin. This is done by a method such as coating. One preferred method is to use a polyurethane polymer dispersion.

The enzyme used in the present invention may be any enzyme as long as it transfers electrons in the reaction, and includes oxidases such as glucose oxidase, galactose oxidase, alcohol oxidase, and cholesterol oxidase, glucose dehydrogenase, and alcohol dehydrogenase. Examples include dehydrogenase enzymes, diaphorase, catalase, and the like.

(Example) Examples of the present invention will be described below.

(A) Preparation of tWi The next one is for glucose sensor! This is an example in which the pole and the gluconolactone reactor IS are made of Fe.

Preparation of O-plated cloth foil Taffeta made of polyester filament yarn with a fineness of 5o denier (24 filaments) according to the following recipe.
First, nickel plating was performed on the cloth (warp density: 170 lines/inch, weft density: 85 lines/inch) to obtain a nickel-plated cloth with a plating thickness of 0.3 μm.

・Catalyst Fft treatment formula Palladium chloride 0.3gy'L Chloride-M
15, Og, / I.

Salt M 200 cc/L liquid Temperature 30°C Treatment time Thoroughly washed with water for 10 minutes and dried with hot air at 80°C
Minute treatment/electroless nickel plating method recipe Nickel sulfate 25 g/L Sodium hypophosphite 25 g/L Sodium birophosphate 50 g/L pH 10-11 Solution temperature 50°C Time 10 minutes The above nickel-plated cloth was immersed in a 15-fold diluted Gerhardt solution and plated with gold at 90° C. for 110 minutes to obtain a gold-plated cloth with a plating thickness of 05 μm.

○Preparation of immobilization solution Immobilization was carried out using the method disclosed in JP-A No. 62-263668.

Izerax 1020 (Hodogaya Chemical) 500mg,
A mixture of 12.5 mg of Izerax O ([! Tsuchiya (Higaku)) and 10 mg of Izerax P (Hodogaya Chemical) was added and mixed thoroughly. Next, glucose oxidase ■ (Toyobo > 1200/mg) was added and mixed thoroughly. 30 mg was added and thoroughly mixed.

0 Preparation of enzyme-immobilized electrode The gold-plated cloth was fixed to < (112 mm2) with rhi solution (3
0mg> was applied, dried at 25°C for 24 hours, and fixed by forming a film.

(B) Cyclic portammetry performance was checked by portammetry similar to the triode method. The measurement conditions are as follows.

Equipment Potentiostat (Nippon thickness measurement NPGFZ2501A
) Recorder (GRAPHTECWXIloo) Reference electrode (
Silver/salt fII control electrode (platinum plate) Electrolyte 0, IM citrate buffer (p) 15.4) 0.1
M Salt 1 Hypotassium 0.5mM Ferrocenecarboxylic acid (Tokyo Kasei) Comparison
-1'' Sensing property The enzyme functional electrode of the present invention was immersed in the above electrolyte, and the
Cyclic portamograms were recorded between sweep potentials of -0.1 V and 0.5 V at a potential insertion rate of 2 V/S. Figure 1
As shown in Figure 2, there was a difference between the curves with no addition of glucose (A curve) and with the addition of 100ml of glucose (Curve B), indicating responsiveness.

FIG. 2 shows the difference between the current value at 0.5 V and the value without the addition of glucose by varying the amount of glucose added and plotted. Linearity was observed, and the electrode of the present invention was found to be useful as a biosensor for measuring glucose concentration.

■ Ability to hold cloth-like materials horizontally The enzyme functional electrode of the present invention has a large surface area, as shown in the surface photograph (Figure 3), and since the electrodes are cloth-like, it has great flexibility, and is lightweight and easy to cut. was gotten. Flexibility was measured using JIS1096 stiffness A method (
A piece of 80 mm was obtained when measured by a bending force method (Chilebar method).

Figure 4 shows the measurement of cyclic portamendoli by changing the electrode surface area. (1) has a surface area of 56 m■
2, and (2) is twice that, 112 mm", 0
．． When the current value at 5V was measured, the area (2) was twice as large as the area (1).

As is clear from the above results, when the enzyme functional electrode of the present invention is used in a bioreactor or a biosensor, high productivity and high sensitivity can be obtained due to its large surface area.

■ Direct Immobilization Characteristics Since the enzyme functional electrode of the present invention immobilizes the enzyme on the electrode area, the movement of electrons at the electrode can be controlled by changing the voltage applied to the electrode.

It became possible to control the reaction externally.

(Effects of the Invention) Since the enzyme function ti of the present invention has a large surface area, it provides a bioreactor biosensor with high productivity and high sensitivity. In addition, it has flexibility and is easy to process such as cutting, allowing the shape of the enzyme functional electrode to be changed freely, giving it the advantage of a wide range of applications. Furthermore, since the enzyme is directly immobilized (as a resin), the reaction can be controlled externally.

This is a diagram showing the results.

Claims (4)

[Claims] (1) An enzyme-functional electrode that has flexibility, has a large surface area, is light and easy to cut, and is made by directly immobilizing a fabric made of plant fiber, animal fiber, artificial fiber, or inorganic fiber on a metal-plated electrode. (2) The enzyme functional electrode according to claim 1, wherein the metal is gold or silver. (3) The enzyme functional electrode according to claim 1, wherein the metal is nickel or copper as the lower layer and gold or silver as the upper layer. (4) The enzyme functional electrode according to claim 1, wherein the plating method comprises an electroless plating method or a combination of an electroless plating method and a subsequent electrolytic plating method.