JPH06174680A - Glucose measuring device - Google Patents

Abstract

PURPOSE:To improve contamination resistance and stability by performing graft polymerization of hydrophilic monomer with a specific constituent onto the surface covered with a film of a glucose sensor. CONSTITUTION:Graft polymerization of hydrophilic monomer of a constituent similar to a living organism containing a compound expressed by an expression I (R; hydrogen, carboxyl group, or carboxylic ester group, R1; hydrogen or 1-4C alkyl group, R2; 2-6C amine group or choline group, n; an integer 1-5) is performed on the surface covered with a film of a glucose sensor, thus reducing cell adhesion property and indicating an effect which is superb in adhesion suppression etc., of fibrin. Therefore, stability is high and measurement can be repeated. One or more compounds which are selected from acrylamide, N,N'- dimethylacyrylamide, N-isopropylacrylamine, 2-hydroxyethylmethacrylate, 2- hydroxyethylacrylate, acrylic acid, N-vinylpyrrolidone other than compounds expressed by the expression I are suited.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a medical diagnostic apparatus and method, and more particularly to a glucose detecting apparatus for diabetes.

[0002]

2. Description of the Related Art It is said that about 10% of Japanese people are naturally prone to diabetes, and diabetes is a major illness of adult diseases, which is one of modern diseases, and has become a social problem. In addition, about 8% of adult diseases diagnosed over 40 years old are diagnosed with diabetes, and the rate tends to increase year by year. It is rare to be cured once it occurs, and diabetic retina cannot be recovered if insufficient treatment is continued. , Diabetic renal failure, myocardial infarction, cerebral thrombosis, and other complications are dangerous. In order to prevent such complications, in the case of diabetes mellitus, drug treatment with insulin and dietary restriction are mainly performed, and in the case of non-insulin dependence, dietary restriction is mainly performed. In addition, it is necessary to perform treatment while controlling the blood glucose concentration of oneself, and the blood glucose concentration is usually measured in a hospital. However, recently, simple measuring instruments that can measure at home have become widespread and convenient.

[0003]

Since the measurement of blood glucose concentration in a hospital is limited in time, it is difficult to carry out daily examinations, but nowadays simple measuring instruments have become popular and use these. Now you can easily measure every day. Examples of simple measuring instruments include Toeko Super, trade name, manufactured by Kodama Co., Ltd., and Glutest E, trade name, manufactured by Sanwa Chemical Co., Ltd.

However, the glucose sensor used in these simple measuring instruments has a drawback that it cannot be repeatedly used because proteins, lipids, etc. in blood adhere to the sensor, and when the measurement fails, a new sensor is required. It must be replaced and is not economical. In addition, since dirt easily adheres to the sensor, there is concern about the stability and reliability of the sensor itself.

Therefore, the present invention has been accomplished by earnest research to solve these problems. That is, an object of the present invention is to provide a glucose measuring device having excellent stain resistance and high stability by graft-polymerizing a hydrophilic monomer that is a biomimetic component on the surface of an envelope of a glucose sensor. .

[0006]

The present invention has found that this object can be achieved by graft-polymerizing a hydrophilic monomer as a specific component on the surface of an envelope of a glucose sensor. That is, by graft-polymerizing a hydrophilic monomer as a biomimetic component alone or together with another hydrophilic monomer or a cross-linking agent to the encapsulating membrane, it was possible to obtain a glucose measuring device having excellent stain resistance and high stability. .

The present invention has been completed based on such findings, and the present invention is as follows. At least the general formula on the surface of the envelope,

[0008]

[Chemical 2]

(Wherein R is hydrogen, a carboxyl group or a carboxylic acid ester group, R ₁ is hydrogen or an alkyl group having 1 to 4 carbon atoms, R ₂ is an amine group having 2 to 6 carbon atoms, and n is 1 to 1).
1. A glucose measuring device, wherein a hydrophilic monomer containing a compound represented by the formula 5) is graft-polymerized. _2. A glucose measuring device characterized in that R ₂ in the general formula is a choline group; In the hydrophilic monomer, in addition to the compound represented by the general formula, at least acrylamide, N, N′-dimethylacrylamide, N-isopropylacrylamide, 2-hydroxyethyl methacrylate, 2-hydroxyethyl acrylate, acrylic acid, N-vinylpyrrolidone A glucose measuring device comprising one or more of the following.

The present invention will be described in detail below. In order to achieve this object, the glucose measuring device of the present invention needs to graft-polymerize a specific hydrophilic monomer on the surface of the envelope film. That is, it must be a biomimetic component in order to ensure biocompatibility, and it is also necessary that stain components such as proteins and fibrin do not easily adhere. Based on such knowledge, it was found that a component similar to phospholipid which is a biomembrane component is most suitable.

The hydrophilic monomer which is graft-polymerized on the surface of the envelope film of the present invention has a general formula:

[0012]

[Chemical 3]

(Wherein R is hydrogen, a carboxyl group or a carboxylic acid ester group, R ₁ is hydrogen or an alkyl group having 1 to 4 carbon atoms, R ₂ is an amine group having 2 to 6 carbon atoms, and n is 1 to 1).
5 represents an integer of 5), for example, 2-
Examples thereof include acryloyloxyethylphosphorylcholine, 2-methacryloyloxybutylphosphorylcholine, 2-methacryloyloxyethylphosphorylcholine, and 2-fumaroyloxyethylphosphorylcholine.

The hydrophilic monomer represented by the general formula used in the present invention may be used alone or in combination with other hydrophilic monomers. As the other hydrophilic monomer, acrylamide, N, N'-dimethyl, which are conventionally known, may be used. Acrylamide, 2-hydroxyethyl methacrylate, N-isopropyl acrylamide, 2-hydroxyethyl acrylate, acrylic acid, N-vinylpyrrolidone, polyethylene glycol monomethacrylate, 2,3-dihydroxypropyl methacrylate, vinyl acetate and the like can be mentioned.

The glucose oxidase-immobilized membrane used in the present invention can be obtained by a general method such as an adsorption / dry immobilization method, a covalent bond method using glutaraldehyde or the like, or an encapsulation method utilizing structural transition of the membrane. As the immobilization membrane, general membranes such as nitrocellulose, cellulose, polyhydroxyethyl methacrylate, silk are used. Alternatively, a method in which a binder such as glutaraldehyde is vapor-deposited on the electrode surface and glucose oxidase is bonded thereon may be used.

The encapsulating membrane used in the present invention includes, but is not limited to, dialysis membranes of cellulose, cellulose derivatives, EVA and the like.

The hydrophilic monomer used in the present invention can be graft-polymerized to the envelope film by a general method. That is, the encapsulating membrane is subjected to plasma treatment or corona discharge treatment in air or oxygen atmosphere to generate active species, and the treated encapsulating membrane is immersed in a hydrophilic monomer solution to perform graft polymerization. As the initiator at this time, for example, iron (II) sulfate ammonium (Mor salt), ammonium persulfate, or the like may be used. After the graft polymerization, the unreacted hydrophilic monomer is extracted with water or the like to finish. The encapsulating membrane thus obtained has good hydrophilicity because the surface is graft-polymerized with a hydrophilic biomimetic component, and also has a graft chain containing a compound represented by the general formula and similar to a cell membrane. Due to the fluctuation of, the stain components such as proteins are hardly attached and the stain resistance is excellent. Therefore,
It is possible to obtain a glucose measuring device having good stain resistance and excellent biocompatibility.

[0018]

The glucose measuring device according to the present invention is graft-polymerized on the surface of the encapsulating membrane with a hydrophilic monomer represented by the general formula, which is a bio-similar component, as a main component, so that the glucose measuring device has no stain resistance to stains such as proteins. Since the hydrophilic monomer represented by the general formula is remarkably improved, it is excellent in biocompatibility, in particular, reduction in cell adhesion and suppression of fibrin adhesion, since it is a component similar to a biomembrane. Therefore, it is possible to provide a glucose measuring device which has high stability and can be repeatedly measured.

[0019]

EXAMPLES The present invention will be specifically described below with reference to examples, but the present invention is not limited thereto. (Example 1) Cellulose membrane for dialysis was separated by 3 cm between electrodes,
It was installed between the electrodes of a corona discharge treatment device having an inter-electrode voltage of 20 kilovolts and a frequency of 2000 hertz for discharge treatment. The treated contact lens was placed in an 8% by weight aqueous solution of 2-methacryloyloxyethylphosphorylcholine (containing 0.2% by weight of iron iron sulfate), and after substituting with nitrogen gas, it was sealed and graft-polymerized at 50 ° C. for 1 hour. Graft-polymerized cellulose membrane for dialysis was added to warm pure water at 60 ° C for 10
By immersing for a period of time to remove the by-product polymer, a cellulose membrane for dialysis with graft treatment was obtained. Glucose oxidase (200
0 IU) 2 mg dissolved in 3 ml of 0.1 M phosphate buffer to a nitrocellulose membrane (pore size 0.45 μm)
Was immersed in the solution for 60 minutes, immobilized by adsorption, washed with a phosphate buffer and dried to prepare an immobilized enzyme membrane. This enzyme membrane was attached to a Clark-type oxygen electrode to prepare a glucose measuring device.

<Platelet Adsorption Test> Platelet-rich plasma prepared from fresh rabbit blood was brought into contact with the graft-polymerized encapsulation membrane for 24 hours. After rinsing with physiological saline after 24 hours, the surface of the encapsulating membrane was observed with a scanning electron microscope. As a result, fibrin produced by adsorbing and activating platelets was hardly observed, showing good biocompatibility and contamination resistance. Showed sex.

<Glucose measuring range> 1 to 100 mg
When a glucose solution having a concentration of 1 / l was prepared and its potential decrease value (mV) was measured, a good linear relationship was observed in the range of 8 to 800 mg / l. <Stability test> Glucose concentration in human urine is about 200 mg
Repeated measurements were carried out using urine prepared to have a concentration of 1 / l. As a result, no significant decrease in sensitivity was observed even after 100 repeated tests, indicating good stability. The results are shown in Fig. 1.

(Example 2) P-type silicon having a thickness of about 250 μm, silicon dioxide having a thickness of about 0.7 μm, thickness of about 0.1
A 450 × 4700 μm-sized hydrogen peroxide microelectrode having a gold lead wire with a width of about 70 μm was used on an electrode substrate made of silicon nitride of μm (the detecting portion is about 1 m at the tip).
m is also a site and the rest is insulated with tartar oxide.) About 80 μl of γ-aminopropyltriethoxysilane was vapor-deposited on this electrode, and further about 90 μl of glutaraldehyde was vapor-deposited and reacted for 20 hours. Glucose oxidase (2000 IU) 2 mg was dissolved in 1000 μl of 0.01 M phosphate buffer pH 7.0, and 10 μl of glutaraldehyde was further added to the solution, and the solution was applied on the surface of the vapor-deposited electrode, followed by reaction immobilization at 5 ° C. for 20 hours. hand,
A glucose oxidase-immobilized hydrogen peroxide electrode was obtained.

A polyhydroxyethyl methacrylate film was placed between the electrodes of a corona discharge treatment device having a distance between electrodes of 3 cm, a voltage between electrodes of 15 kilovolts, and a frequency of 1000 hertz to perform discharge treatment. The treated polyhydroxyethylmethacrylate film was put into an aqueous solution of 7% by weight of 2-methacryloyloxyethylphosphorylcholine and 1% by weight of N, N'-methylenebisacrylamide (containing 0.2% by weight of ammonium iron sulfate) and filled with nitrogen gas and then sealed. Then 5
Graft polymerization was carried out at 5 ° C. for 1 hour. The graft-polymerized polyhydroxyethyl methacrylate film was immersed in warm pure water at 60 ° C. for 10 hours to remove the by-produced polymer to obtain a graft-treated polyhydroxyethyl methacrylate film.

A glucose measuring device was prepared by covering a glucose oxidase-immobilized hydrogen peroxide electrode with a graft-treated polyhydroxyethyl methacrylate film.

<Platelet adsorption test> When evaluated in the same manner as in Example 1, almost no fibrin produced by adsorption and activation of platelets was observed, and good biocompatibility and stain resistance were exhibited.

<Glucose measurement range> 0.1 to 100
A glucose solution having a concentration of mg / l was prepared, the potential of the electrode was set to 1.0 V, and the potential increase value (mV) was measured. As a result, a good linear relationship was observed in the range of 0.5 to 600 mg / l. Was given.

<Stability test> Repeated measurement was carried out using urine prepared so that the glucose concentration in human urine was about 100 mg / l. No significant decrease in sensitivity was observed even after 100 repeated tests. , Showed good stability.
The results are shown in Figure 2.

(Comparative Example 1) The oxygen electrode used in Example 1 was used without an encapsulating membrane.

<Platelet adsorption test> When evaluated in the same manner as in Example 1, fibrin produced by adsorption and activation of platelets was found on the surface of the glucose oxidase-immobilized membrane, and platelet activation was good. It showed no significant biocompatibility and stain resistance.

<Glucose measuring range> 1 to 100 mg
When a glucose solution having a concentration of 1 / l was prepared and the potential decrease value (mV) was measured, a good linear relationship was observed in the range of 10 to 800 mg / l. <Stability test> Repeated measurement was carried out using urine prepared so that the glucose concentration in human urine was about 200 mg / l. As a result, sensitivity was reduced in eight repeated tests. The results are shown in Fig. 1.

(Comparative Example 2) The hydrogen peroxide electrode used in Example 2 was used without attaching an encapsulating membrane.

<Platelet adsorption test> When evaluated in the same manner as in Example 1, fibrin produced by adsorption and activation of platelets was observed on the surface of the glucose oxidase-immobilized electrode, and platelet activation was favorable. It showed no significant biocompatibility and stain resistance.

<Glucose measurement range> 0.1 to 100
When a glucose solution having a concentration of mg / l was prepared and the potential increase value (mV) was measured, it was 0.5 to 600 mg / l.
A good linear relationship was observed in the range of.

<Stability test> Repeated measurement was carried out using urine prepared so that the glucose concentration in human urine was about 200 mg / l. A decrease in sensitivity was observed after 10 repeated tests. The results are shown in Figure 2.

[0035]

INDUSTRIAL APPLICABILITY Since the hydrophilic monomer, which is a biomimetic component, is graft-polymerized on the surface of the encapsulating membrane, the present invention can provide a glucose measuring device excellent in stain resistance and stability.

[Brief description of drawings]

FIG. 1 is a graph showing potential changes due to repeated measurement in Example 1 and Comparative Example 1 of the present invention.

FIG. 2 is a graph showing a potential change due to repeated measurement in Example 2 and Comparative Example 2 of the present invention.

Claims (3)

[Claims] 1. A glucose measuring device having a glucose sensor comprising a detection electrode, a glucose oxidase-immobilized membrane and an encapsulation membrane, wherein at least the general formula: (In the formula, R is hydrogen, a carboxyl group or a carboxylic acid ester group, R ₁ is hydrogen or an alkyl group having 1 to 4 carbon atoms,
R ₂ is an amine group having 2 to 6 carbon atoms, n is an integer of 1 to 5), and a hydrophilic monomer containing a compound represented by the formula is graft-polymerized. 2. The glucose measuring device according to claim 1, wherein R ₂ in the general formula is a choline group. 3. The hydrophilic monomer may be acrylamide, N, N′-dimethylacrylamide, N-isopropylacrylamide, 2 in addition to the compound represented by the general formula.
The glucose measuring device according to claim 1, comprising one or more compounds selected from -hydroxyethyl methacrylate, 2-hydroxyethyl acrylate, acrylic acid, and N-vinylpyrrolidone.