JPH05188037A - Concentration measuring device - Google Patents

Abstract

PURPOSE:To improve the concentration measuring accuracy of measuring object substance by excluding the quantity of electricity based on the biochemical reaction between bio-substance and interfering substance, from the quantity of electricity converted by a sensor element part, and obtaining concentration on the basis of the quantity of electricity reflecting only the concentration of the measuring object substance. CONSTITUTION:A biosensor 1 receives the impression of measuring weak voltage through a connector 35 so as to output the current value outputted from sensor element parts 5-11, to an electronic control device 40. Upon decision to execut measuring of a solution of unknown concentration to be measured, for instance, glucose concentration of urine, the sensor output (current value) of the respective sensor element parts 5-11 is read, and each sensor output read from the interfering substance measuring sensor element part among the read sensor output is converted into the sensor output of the glucose measuring sensor element part 5. After conversion, the sensor output of the element part 5 is corrected, and the concentration of glucose in the urine based on the sensor output after correction is computed. A control signal corresponding to the numeric value of the computed glucose concentration is then outputted to display equipment 50 from the device 40.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a concentration measuring device for measuring the concentration of a substance to be measured based on a biochemical reaction between a substance to be measured and a biological substance in a solution to be measured.

[0002]

2. Description of the Related Art A biosensor measures a substance to be measured in a solution to be measured such as urine by converting the progress state of a biochemical reaction that progresses between a biological substance such as an enzyme or a microorganism and the substance to be measured into an electric quantity. The various things are known. For example, the potential value is measured based on the ion concentration change of various substances involved in biochemical reaction, or the current value obtained by the electrode reaction due to the chemical substance (electrode active substance) produced or consumed in this reaction is measured. In addition to electrode biosensors of the type, biosensors that measure thermal changes due to biochemical reactions with a thermal measurement device, and biosensors that guide biochemical reactions to chemiluminescence and measure the amount of light emitted by a photocounter. and so on.

By using various enzymes or microorganisms as the biological substance to be used, it is possible to selectively detect the substance to be measured that reacts with it. For example, when glucose oxidase is used as the biological substance, it becomes a biosensor for detecting glucose in urine. Further, when ascorbate oxidase is used as the biological substance, it becomes a biosensor for detecting ascorbic acid in urine. That is, the biological substance to be used is appropriately selected according to the substance to be measured.

To obtain the concentration of the substance to be measured using the biosensor, a concentration measuring device that processes the sensor output from the biosensor is used. This concentration measuring device includes a biosensor and a microcomputer including a CPU, a ROM, a RAM, and the like. Then, if the biosensor is brought into contact with the solution to be measured whose concentration is unknown, this concentration measuring device converts the amount of electricity (sensor output) input from the sensor into the concentration of the substance to be measured. The substance concentration can be obtained. Note that a calibration curve stored in advance in the ROM is generally used to convert the amount of electricity into the concentration.

When the concentration of the substance to be measured is measured by the concentration measuring device, there are various methods for bringing the solution to be measured into contact with the biosensor, and the method is appropriately selected. For example, the biosensor is immersed in the solution to be measured, or the solution to be measured is poured into the biosensor.

[0006]

However, when the concentration of the substance to be measured in the solution to be measured is obtained, the concentration may not be measured correctly. This is for the following reason. Since the solution to be measured is urine as described above, various substances other than the substance to be measured are present in the solution. And, although a certain substance is not a measurement target substance, it causes a biochemical reaction with a biological substance used to convert the concentration of the measurement target substance into an electric quantity, and converts the concentration of the measurement target substance into an electric quantity. It becomes an interfering substance that inhibits.
For example, when glucose concentration in urine is measured using glucose oxidase, ascorbic acid, uric acid, bilirubin, etc. in urine serve as interfering substances, so glucose oxidase starts glucose and some of this ascorbic acid etc. Will cause the biochemical reaction of. Therefore, the converted amount of electricity does not depend on the concentration of glucose alone, and the glucose concentration cannot be accurately measured.

Therefore, there has been proposed a technique for preventing such an interfering substance from being oxidized or kept close to glucose oxidase (JP-A-2-245650, JP-A-58-5642). However, since the chemical structure of perchloric acid such as NaClO ₄ used for the oxidation of the interfering substance is unstable, the reliability of the oxidation of the interfering substance is low, and as a result, the detection accuracy of the substance to be measured (glucose) is low. I cannot hope for improvement. On the other hand, if the interfering substance is not close to the biological substance,
Since ascorbic acid or the like has a negative charge in a solution, a negative charge is applied to the periphery of the electrode so that the ascorbic acid or the like is electrically repelled so as not to come close. However, it is not preferable that the negative charges are constantly charged around the electrodes, because the electric configuration becomes complicated.

The present invention has been made to solve the above problems, and an object of the present invention is to eliminate the influence of interfering substances with a simple structure to improve the measurement accuracy of the concentration of the substance to be measured.

[0009]

Means for Solving the Problems In order to achieve such an object, the means adopted by the present invention is to determine the concentration of a substance to be measured based on a biochemical reaction between the substance to be measured and a biological substance in a solution to be measured. In a concentration measuring device for measuring, an object to be measured is provided with an identification layer carrying a biological substance that causes a biochemical reaction with the object substance to be measured, and the state of progress of the biochemical reaction proceeding in the identification layer is converted into an electric quantity and output. A substance sensor element part, and an identification layer carrying a biological substance that causes a biochemical reaction with an interfering substance that inhibits the conversion of electricity in the measurement target substance sensor element part, and the progress state of the biochemical reaction that proceeds in the identification layer. To correct the sensor output from the measurement target substance sensor element unit based on the sensor output from the interfering substance sensor element unit and the sensor substance output from the interfering substance sensor element unit. Alternatively, the sensor output from the interfering substance sensor element unit is converted into a sensor output when the biological substance carried on the discrimination layer in the measurement target substance sensor element unit causes a biochemical reaction with the interfering substance. The gist of the present invention is to include means and a concentration calculation means for calculating the concentration of the substance to be measured based on the sensor output obtained by subtracting the converted sensor output from the sensor output of the substance to be measured sensor element unit.

[0010]

The concentration measuring device having the above-described structure is provided with the measurement target substance sensor element section and the interfering substance sensor element section, each sensor element section converts the next electric quantity, and outputs the converted electric quantity as a sensor output. .. In the measurement target substance sensor element section, the biological substance carried on the discrimination layer causes a biochemical reaction between the measurement target substance and the interfering substance in the discrimination layer. The state of progress of this dependent biochemical reaction is converted into an electric quantity. On the other hand, in each interfering substance sensor element part, the biological substance carried on the discrimination layer of each interfering substance sensor element part causes a biochemical reaction with the interfering substance in this discriminating layer, so that it depends only on the interfering substance concentration. The state of progress of this biochemical reaction is converted into an electric quantity.

Since the biological substance carried on the discrimination layer in the sensor element part of the measurement object causes a slight biochemical reaction with the interfering substance, the electric quantity (sensor output) converted by the sensor element part of the measurement object shows this interference. It includes the amount of electricity (sensor output) that is based on biochemical reactions with substances. By the way, in the concentration measuring device of the present invention, the sensor output conversion means causes the sensor output from the interfering substance sensor element part to cause a biological substance carried on the discrimination layer in the target substance sensor element part to cause a biochemical reaction with the interfering substance. Converted to the sensor output when Further, the concentration calculation means calculates the concentration of the substance to be measured based on the sensor output obtained by subtracting the converted sensor output from the sensor output of the substance to be measured sensor element section. As a result, the electric quantity based on the biochemical reaction with the interfering substance is excluded from the electric quantity converted by the sensor element part to be measured to calculate the concentration. The concentration of the substance to be measured can be obtained based on the amount.

[0012]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Next, a concentration measuring device according to an embodiment of the present invention will be described with reference to the drawings. FIG. 1 is a schematic perspective view of a biosensor 1 which is a main component of the concentration measuring device of the embodiment, and FIG. 2 is a concentration measuring device for obtaining the concentration of a substance to be measured based on the sensor output from the biosensor 1. 20 is a block diagram showing an electrical configuration of 20. FIG. The concentration measuring device 20 is used when measuring glucose in urine containing a solution to be measured, for example, glucose and ascorbic acid or the like that interferes with glucose measurement.

First, the biosensor 1 will be described.
As shown in FIG. 2, the biosensor 1 includes a device body 30.
While being used by being attached to and detached from the attached connector 35,
As shown in FIG. 1, it is a plate-type electrode biosensor and has the following configuration.

As shown in FIG. 1, the biosensor 1 comprises an insulating substrate 3 having a plate thickness of 0.7 mm produced by sintering alumina.
On (20 mm × 30 mm), the glucose measuring sensor element portion 5, the ascorbic acid measuring sensor element portion 7,
The sensor element unit 9 for measuring uric acid and the sensor element unit 11 for measuring bilirubin are provided.

The glucose measuring sensor element portion 5 includes a working electrode 5a and a reference electrode 5b formed on the insulating substrate 3.
The counter electrode 5c, an identification layer 5d in which glucose oxidase (GOD) is carried and immobilized on the working electrode 5a, and terminal portions 5e, 5f, 5g of each electrode such as the working electrode 5a are provided. Similarly, the ascorbic acid measurement sensor element unit 7, the uric acid measurement sensor element unit 9, and the bilirubin measurement sensor element unit 11 also include a working electrode, a reference electrode, a counter electrode, and the like. In addition,
The working electrodes and the like in each sensor element portion are denoted by reference numerals a to g as in the case of the glucose measuring sensor element portion 5.

The ascorbic acid measuring sensor element portion 7 is provided with an identification layer 7d which is immobilized by carrying ascorbic acid oxidase on the working electrode 7a, and the uric acid measuring sensor element portion 9 is mounted on the working electrode 9a. Is provided with an identification layer 9d that is immobilized by carrying uricase that is a urate oxidase, and the bilirubin-measuring sensor element portion 11 is provided with an identification layer 11d that is immobilized by carrying bilirubin oxidase on the working electrode 11a. ..

Each electrode such as the working electrode in each of the sensor element portions is covered with an insulating layer 13 so that the side on which the identification layer is formed becomes the sensitive portion 15.

When the solution to be measured comes into contact with the sensitive portion 15, a biochemical reaction occurs between the above-mentioned biological substances carried in the discrimination layer on the upper surface of each working electrode and the substance to be measured in the solution to be measured, and the biosensor 1 Outputs a current value reflecting this biochemical reaction to the device body 30 via the connector 35. The device body 30 processes this current value as a sensor output value, and obtains the density as described later.

Next, the manufacturing process of the biosensor 1 will be described. First, the electrode in each sensor element part is produced. That is, the working electrodes 5a to 11a and the respective terminal portions 5e to 11e following the working electrodes 5a to 11a are formed through screen printing of a platinum paste on the upper surface of the insulating substrate 3 and drying treatment at 50 ° C. for 1 hour. At this time, as the platinum paste, a dispersion paste obtained by kneading 90 wt% of platinum fine powder and 10 wt% of butyl cellulose was used. Next, each reference electrode 5
b to 11b and counter electrodes 5c to 11c and the following terminal portions 5f to 11f and 5g to 11g are formed through screen printing of graphite paste on the upper surface of the insulating substrate 3 and drying treatment at 50 ° C. for 1 hour. .. At this time, as the graphite paste, graphite fine powder 60 wt% and liquid paraffin 40 wt%
The dispersion paste obtained by kneading and was used. In addition,
Each of the electrodes is formed to have a thickness of 10 μm.
In addition, the insulating substrate 3 is cut into the above-described dimensions by using the above-mentioned four sensor element portions including the glucose-measuring sensor element portion 5 as one set after a plurality of sensor element portions are formed.

After that, the insulating layer 13 is formed on each working electrode, reference electrode and counter electrode by printing and drying an appropriate insulating agent such as an epoxy resin.

Then, glucose oxidase (GO
The discriminating layer supporting the above-mentioned biomaterials such as D) is immobilized on the upper surface of each working electrode as described below. Collagen: 94 wt%, GOD: 5 wt%, and ferrocene, which is an electron acceptor (mediator): 1 wt%, are kneaded to prepare a gel-like GOD solution. Then, this GOD solution was applied to the upper surface of the working electrode 5a on the side of the sensitive part 15 with a thickness of about 20 μm by a micropipette, and then the identification layer 5d was fixed through natural drying for 2 hours at room temperature.

5 wt% in place of the above 5 wt% GOD
Ascorbate oxidase, 5 wt% uricase, and 5 wt% bilirubin oxidase are used, and each of the biomaterials is mixed with the collagen and ferrocene to separately prepare a gel-like biomaterial solution. Then, each biomaterial solution is GOed to the upper surface of the corresponding working electrode 7a to 11a.
Coating is performed in the same manner as in the case of the D solution, and each of the identification layers 7d to 11
d was fixed. In this way, the biosensor 1 in which the four sensor element portions are integrally provided on the insulating substrate 3 is completed.

Next, the device body 30 will be described. As shown in FIG. 2, the concentration measuring device 20 includes the biosensor 1 described above and a device body 30. Device body 30
Is an electronic control unit 40 that calculates the concentration as described later.
A display device 50 for displaying the converted concentration, an operation panel 60 having a power-on switch and the like, and a connector 35 for attaching / detaching the biosensor 1.

The electronic control unit 40 includes a CPU 41, R
The OM 42, the RAM 43, and the timer 44 are mainly configured as a logical operation circuit, and input / output with the outside is performed by an input / output port 46 interconnected with these via a common bus 45. The display device 50, the operation panel 60, and the connector 35 are connected to the input / output port 46 of the electronic control unit 40. The display device 50 displays the concentration and the like based on the control signal from the electronic control unit 40, and displays the biosensor 1
Receives the weak voltage for measurement via the connector 35 and outputs the current value output from each of the sensor element units 5 to 11 to the electronic control unit 40. The operation panel 60 also outputs on / off signals of various switches to the electronic control unit 40.

Next, the concentration measurement control (routine) performed by the concentration measuring device 20 of the present embodiment having the above-mentioned structure will be described with reference to the flowchart of FIG. The flowchart shown in FIG. 3 shows a concentration measurement routine that is repeatedly executed from when the power switch (not shown) of the operation panel 60 is pressed to turn on the power of the concentration measuring device 20 until the power is turned off. .. As shown in FIG. 3, in this routine, an initial process executed only when the power is turned on, that is, an internal register of the CPU is cleared and the like, and then sequentially executed. Following the initial processing, first, it is determined whether or not the measurement of the glucose concentration is started from the operation state of a measurement start switch (not shown) of the operation panel 60 (step 200, hereinafter, this step is simply referred to as S). ),
Wait until the measurement start switch is turned on.

If it is determined in S200 that the solution to be measured whose concentration is unknown, for example, the glucose concentration of urine, is to be measured, the sensor output (current value) of each of the sensor element units 5 to 11 is read (S210). Of the read sensor outputs, the sensor outputs read from the interfering substance measuring sensor element units such as the ascorbic acid measuring sensor element unit 7, the uric acid measuring sensor element unit 9 and the bilirubin measuring sensor element unit 11 are used for glucose measurement. The sensor output is converted into a sensor output based on a biochemical reaction between glucose oxidase and each interfering substance in the discrimination layer 5d of the sensor element unit 5 (S220). The reason why the sensor output of each interfering substance measuring sensor element unit is converted into the sensor output of the glucose measuring sensor element unit 5 is as follows.

The glucose oxidase in the discriminating layer 5d of the glucose measuring sensor element 5 slightly reacts with each interfering substance present in urine. Therefore, the sensor output from the sensor element unit 5 for glucose measurement is based on the biochemical reaction between glucose and glucose oxidase in the discrimination layer 5d, and the biochemical reaction between each interfering substance and glucose oxidase in the discrimination layer 5d. Based on. Therefore, if the sensor output based on the biochemical reaction between each interfering substance and glucose oxidase is found among the sensor outputs from the glucose measurement sensor element unit 5, the sensor output based on only the biochemical reaction between glucose and glucose oxidase. Is obtained. However, the reactivity of glucose oxidase with respect to each interfering substance is naturally inferior to that with glucose, and each biological substance with respect to each interfering substance (ascorbic acid oxidase for ascorbic acid, uricase for uric acid,
The reactivity of bilirubin oxidase) to bilirubin is also poor. Therefore, taking into account the reactivity of glucose oxidase with respect to each interfering substance, each sensor output of each interfering substance measuring sensor element unit is compared with each interfering substance and glucose oxidase in the identification layer 5d of the glucose measuring sensor element unit 5. It is converted into the sensor output based on the biochemical reaction of.

This conversion was carried out by multiplying each sensor output read from each of the above-mentioned interfering substance measuring sensor element parts by a conversion coefficient determined according to each interfering substance. It should be noted that this conversion coefficient is set in advance as described below.
It is stored in the OM 42 and is read out for conversion. For example, if the sensor output (current value) from the ascorbic acid measuring sensor element unit 7 is Ia, Ka · Ia obtained by multiplying the sensor output Ia by the conversion factor Ka for ascorbic acid is used as the glucose measuring sensor element unit. The sensor output based on the biochemical reaction between ascorbic acid and glucose oxidase in the identification layer 5d of No. 5 was used. Similarly, Ku · Iu obtained by multiplying the sensor output Iu from the uric acid measurement sensor element unit 9 by the conversion coefficient Ku for uric acid is used as the biochemistry of uric acid and glucose oxidase in the discrimination layer 5d of the glucose measurement sensor element unit 5. The sensor output was based on the reaction. Further, Kb · Ib obtained by multiplying the sensor output Ib from the bilirubin measurement sensor element unit 11 by the conversion coefficient Kb for bilirubin is used as a biochemical reaction between bilirubin and glucose oxidase in the discrimination layer 5d of the glucose measurement sensor element unit 5. Based on the sensor output.

The respective conversion factors Ka, Ku, Kb are stored in advance in the ROM 42 in association with at least the respective interfering substance concentrations (sensor output from each interfering substance measuring sensor element section). Further, the above conversion factors were determined through the experiments described below.

The test biosensor consisting only of the glucose measuring sensor element portion 5 in the present embodiment is immersed in a glucose reagent adjusted to a predetermined concentration, and first, the sensor in the case where only the adjusted concentration of glucose is present in the solution to be measured. Calculate the initial output value IGO. Next, ascorbic acid is gradually added to this glucose reagent, and the transition of the sensor output of the test biosensor is observed. Then, the relationship between the ascorbic acid addition amount, that is, the ascorbic acid concentration and the change amount ΔIG of the sensor output (deviation from the sensor output initial value IG0) is obtained. The amount of change in the sensor output is based on the biochemical reaction between ascorbic acid and glucose oxidase. On the other hand, the ascorbic acid concentration can be converted into a sensor output IA of a biosensor using ascorbate oxidase by a calibration curve. Therefore,
The conversion coefficient Ka for ascorbic acid is the value of the ratio (ΔIG / sensor) of the change amount ΔIG of the sensor output and the sensor output IA.
IA), and the obtained conversion coefficient Ka is stored in the ROM 42 in association with the ascorbic acid concentration (sensor output IA). Similarly, conversion factors Ku and Kb for uric acid and bilirubin are obtained and stored.

The above experiment was repeated for glucose reagents having different adjusted concentrations to obtain the above conversion factors Ka.
, Ku, Kb can be stored in the ROM 42 in association with the glucose concentration (sensor output) and each interfering substance concentration (sensor output). In this way, the sensor output conversion of S220 can be performed by the conversion coefficient that also reflects the glucose concentration (sensor output) in the measured solution, which is preferable because the conversion accuracy is improved.

After the sensor output of each sensor element unit for measuring interfering substances is converted into the sensor output of the sensor element unit for glucose measurement 5 in S220, the sensor output of the sensor element unit for glucose measurement 5 read in S210 is corrected. Then, the glucose concentration in urine is calculated based on the corrected sensor output (S230). That is, from the read sensor output of the glucose measuring sensor element unit 5 to the above-mentioned converted sensor output (Ka) based on the biochemical reaction between glucose oxidase and an interfering substance such as ascorbic acid in the discrimination layer 5d of the glucose measuring sensor element unit 5.・ Ia, K
u * Iu, Kb * Ib) is subtracted to perform the above correction.
As a result, the sensor output of the glucose measuring sensor element unit 5 based on only the biochemical reaction between glucose and glucose oxidase is obtained. Then, the glucose concentration of the glucose reagent is calculated from the obtained sensor output (corrected sensor output) and the glucose calibration curve Kgcal shown in FIG. The calibration curve Kgcal is stored in the ROM 42 in advance as calibration curve map data in which the sensor output and the glucose concentration are associated with each other.

Thereafter, a control signal corresponding to the calculated numerical value of the glucose concentration is sent from the electronic control unit 40 to the display device 50.
To display the glucose concentration value (S240),
The processing of this routine is once ended.

The glucose concentration in the glucose reagent was measured using the concentration measuring device 20 of this embodiment having such a structure. The prepared concentrations of glucose, ascorbic acid, uric acid and bilirubin in the glucose reagent used for the measurement are as shown in Table 1 and are prepared in advance before the measurement. In addition, the element parts 5 to 11 in Table 1
Are abbreviations of the glucose measuring sensor element section 5, the ascorbic acid measuring sensor element section 7, the uric acid measuring sensor element section 9, and the bilirubin measuring sensor element section 11, respectively.

The measured glucose concentration in Table 1 was determined as follows according to the flow chart described. In addition,
This explanation is given for the case where the prepared glucose concentration is 17 mg / dl. That is, of the sensor outputs read from the sensor element units 5 to 11, the sensor output Ia (10 × 10 ⁻⁶ A) of the ascorbic acid measuring sensor element unit 7,
Sensor output Iu of the sensor element unit 9 for measuring uric acid (15 × 1
0 ^-6 A), the sensor output Ib (8 × 10 ^-6 A) of the bilirubin measurement sensor element unit 11 is converted into a conversion factor Ka in the table.
By multiplying (0.2), Ku (0.01), and Kb (0.5), a sensor based on a biochemical reaction between glucose oxidase and each interfering substance in the discriminating layer 5d of the glucose measuring sensor element unit 5 Convert to output (S22
0). As a result, the converted sensor output is 2 × 10 ^-6
A (Ka ・ Ia), 0.15 × 10 ^-6 A (Ku ・ Iu)
), 4 × 10 ⁻⁶ A (Kb · Ib) is obtained.

Then, the glucose measuring sensor element 5
The sensor output Ig of the sensor element unit 5 is corrected by subtracting each converted sensor output from the sensor output Ig (19 × 10 ⁻⁶ A) read from the sensor output Ig and the corrected sensor output I
gh (12.85 × 10 ^-6 A = 19 × 10 ^-6 A-2 × 1
The measured glucose concentration of 16.06 mg / dl is calculated from 0 ⁻⁶ A−0.15 × 10 ⁻⁶ A−4 × 10 ⁻⁶ A) and the glucose calibration curve Kgcal shown in FIG. 4 (S230).

On the other hand, the sensor output Ig (19 × 10 9) read from the glucose measuring sensor element unit 5
^-6 A) was used to calculate the glucose concentration, the glucose concentration was 23.25 m as shown in FIG.
It becomes g / dl.

That is, as is clear from Table 1, the concentration measuring device 20 of the present embodiment can measure the concentration close to each adjusted glucose concentration without the influence of the interfering substance. The glucose concentration measurement accuracy can be improved. Therefore, when measuring the glucose concentration in urine that necessarily contains ascorbic acid, uric acid, and bilirubin, which are interfering substances with respect to the measurement of glucose concentration, glucose is accurately measured by the concentration measuring device 20 of the present embodiment. The concentration can be measured.

Further, in the concentration measuring device 20 of this embodiment,
Since ferrocene is carried as an electron acceptor (mediator) together with the above-mentioned biological substance on the discrimination layer in each sensor element portion, the glucose concentration can be measured more accurately even if the solution to be measured is urine with a small amount of dissolved oxygen.

Next, a concentration measuring device used for measuring lactic acid in sweat containing lactic acid and urea which is an interfering substance for lactic acid measurement will be described. Since the biosensor used in this concentration measuring apparatus for measuring lactic acid is different from that of the concentration measuring apparatus 20 of the above-mentioned embodiment, the description of the apparatus main body will be omitted.

As shown in FIG. 5, the biosensor 70 used in the concentration measuring device for lactic acid measurement is a flat plate electrode type biosensor, like the biosensor 1 described above, and has the following configuration. .. Since the biosensor 1 has the same configuration as that of the biosensor 1, the description of the biosensor 70 will be simplified with respect to the same configuration and manufacturing process.

The biosensor 70 includes a lactate measuring sensor element portion 73 and a urea measuring sensor element portion 75 on an insulating substrate 71 (10 mm × 30 mm) made of polyethylene having a plate thickness of 1.0 mm.

The lactic acid measuring sensor element portion 73 has a working electrode 73a and a reference electrode 73 formed on the insulating substrate 71.
b, the counter electrode 73c, and the identification layer 7 in which lactate oxidase, which is a lactate dehydrogenase, is carried and immobilized on the working electrode 73a
3d and terminal portions 73e, 73 of each electrode such as the working electrode 73a
f, 73g. Urea measurement sensor element part 75
In addition to the working electrode, the reference electrode, the counter electrode, and the like, the working electrode 75a is provided with an identification layer 75d on which urease, which is a urea oxidase, is carried and immobilized.

Each electrode such as the working electrode in each sensor element section is covered with an insulating layer 77, and the side on which each identification layer is formed serves as a sensitive section 79.

When the solution to be measured comes into contact with the sensitive portion 79, a biochemical reaction takes place between each of the biological substances carried on the discrimination layer on the upper surface of each working electrode and the substance to be measured in the solution to be measured, and the biosensor 70. Outputs a current value reflecting this biochemical reaction to the device body 30 via the connector 35. The device body 30 processes the current value as a sensor output value to obtain the density.

The electrodes such as each working electrode and each terminal portion of the biosensor 70 are formed by screen printing after plasma etching the surface of the insulating substrate 71 to make the surface sparse. The paste used was a dispersion paste obtained by kneading 90 wt% of graphite fine powder and 10 wt% of butyl cellulose, and after screen printing, it was dried at 50 ° C. for 1 hour. The thickness of the electrodes was adjusted to 20 μm in all. After that, the insulating layer 77 was formed.

The discriminating layer 73d on the working electrode 73a is a gel-like solution obtained by kneading 10 wt% of lactate oxidase and 90 wt% of albumin with a microsyringe of about 20 μm.
Was dripped and applied at a thickness of 10 .mu.m, and then left standing in glutaraldehyde-saturated air for 30 minutes to be immobilized. The discrimination layer 75d on the working electrode 75a was similarly immobilized by using a gel-like solution in which 10 wt% urease and 90 wt% albumin were kneaded. The immobilization of both discriminating layers is performed simultaneously in glutaraldehyde-saturated air.

Next, an evaluation test relating to lactic acid measurement by the concentration measuring device 20 using the biosensor 70 will be described. The lactic acid reagent used for the measurement was prepared as follows, assuming the case of measuring lactic acid in sweat. Lactic acid reagent A: Reagent that assumes perspiration sweat in daily life Prepared lactic acid concentration → 80 mg / dl Prepared urea concentration → 13 mg / dl Lactate reagent B: Reagent that assumes sweat sweated during continuous exercise Prepared lactic acid concentration → 182 mg / Dl Prepared urea concentration → 28 mg / dl

The measured lactic acid concentration for each of the above reagents was determined as follows according to the flowchart described above. The sensor output and the like at that time are as follows. Lactic acid reagent A Sensor output from the lactic acid measuring sensor element unit 73 → 5.
75 × 10 ⁻⁶ A Sensor output Iu from the sensor element unit for urea measurement 75 → 6.
5 × 10 ⁻⁶ A Used conversion coefficient Ku → 0.25 Converted sensor output Ku · Iu → 1.63 × 10 ⁻⁶ A Corrected sensor output Ilh → 4.12 × 10 ⁻⁶ A (= 5.75 × 10 ⁻⁶ A −1.63 × 10) ^-6 A) This corrected sensor output Ilh and the lactate calibration curve K shown in FIG.
From lcal, 72 mg / dl was calculated as the measured lactic acid concentration. When the lactic acid concentration is calculated using the sensor output Il (5.75 × 10 ⁻⁶ A) from the lactic acid measuring sensor element unit 73 as it is, the lactic acid concentration is 100 m as shown in FIG.
It becomes g / dl.

Lactic acid reagent B Sensor output from the lactic acid measuring sensor element unit 73 →
3.85 × 10 ⁻⁶ A Sensor output from the urea measuring sensor element unit 75 Iu → 14
× 10 ^-6 A conversion factor used Ku → 0.25 scaled sensor output ^{Ku · Iu → 3.5 × 10 -6} A corrected sensor output ^{Ilh → 10.35 × 10 -6 A (} = 13.85 × 10 -6 A-3.5 × 10 - ⁶ A) This corrected sensor output Ilh and the lactate calibration curve K shown in FIG.
From lcal, 180 mg / dl was calculated as the measured lactic acid concentration. When the lactic acid concentration is calculated using the sensor output Il (13.85 × 10 ⁻⁶ A) from the lactic acid measuring sensor element unit 73 as it is, the lactic acid concentration is 24 as shown in FIG.
It becomes 1 mg / dl.

Thus, the concentration measuring device 20 of this embodiment
According to this, even if the substance to be measured is lactic acid, it is possible to accurately measure the lactic acid concentration by eliminating the influence of urea, which is the interfering substance, and to improve the measurement accuracy of the lactic acid concentration. Therefore, the concentration of lactic acid in sweat can be accurately measured.

The present invention is not limited to the above-described embodiments, but can be carried out in various modes without departing from the scope of the invention, and the following modifications are possible. For example, the biosensor 1 is provided on each side of the insulating substrate 3 for each sensor element portion, but two biosensors 1 may be provided on each side of the insulating substrate.
This is preferable because the sensor is compact. Further, since the sensor output obtained by detecting ascorbic acid or the like is obtained, the concentrations of interfering substances such as ascorbic acid and uric acid may be displayed using a calibration curve.

Further, instead of glucose oxidase in the biosensor for measuring glucose, an enzyme such as pyranose oxidase or mutarotase, or Pseudomo
Microorganisms such as nasfluorescens may be used. The substances to be measured are not limited to the above-mentioned glucose and lactic acid, and in addition to sugars such as fructose and galactose, vitamins such as ascorbic acid, metabolites such as urobilinogen, bilirubin, creatine and creatinine, albumin, globulin, Proteins such as hemoglobin, hormones such as human chorionic gonadotropin (hCG), steroids such as 17KS and 17OHCS, amino acids, hippuric acid, glucuronic acid, uric acid, pyruvic acid, ketoglutaric acid, oxalic acid, β-oxybutyric acid, and other acids It may be. It is needless to say that a biological substance may be appropriately selected according to the measurement target substance, and a biological substance that causes a biochemical reaction with an interfering substance with respect to the measurement of each measurement target substance may be appropriately selected.

The biosensor to be used is not limited to the flat plate electrode type biosensor, but a so-called potentiometric sensor for measuring the potential value based on the ion concentration change of various substances involved in the biochemical reaction, Also, a biosensor of a type that measures a thermal change associated with a biochemical reaction with a heat measuring device, or a sensor of a type that guides a biochemical reaction to chemiluminescence and measures the amount of the emitted light with a photocounter may be used.

[0055]

As described in detail above, according to the concentration measuring apparatus of the present invention, the amount of electricity converted by the sensor element portion for measuring the substance to be measured is used to determine the difference between the biological substance and the interfering substance in this sensor element. It is possible to obtain the concentration of the substance to be measured based on the amount of electricity that reflects only the concentration of the substance to be measured, excluding the amount of electricity based on the biochemical reaction of. As a result, the influence of the interfering substance can be eliminated, the concentration of the substance to be measured can be accurately determined, and the measurement accuracy can be improved.

[Brief description of drawings]

FIG. 1 is a schematic perspective view of a biosensor 1 which is a main component of a concentration measuring device 20 according to an embodiment.

FIG. 2 is a block diagram showing an electrical configuration of a concentration measuring device 20 according to an embodiment.

FIG. 3 is a flowchart showing a concentration measurement routine performed by the concentration measuring device 20.

FIG. 4 is a graph showing a glucose calibration curve Kgcal in which glucose concentrations are associated with sensor output current values.

FIG. 5 is a schematic perspective view of a biosensor 70 used when measuring a lactic acid concentration.

FIG. 6 is a graph showing a lactate calibration curve Klcal in which a lactate concentration and a sensor output current value are associated with each other.

[Explanation of symbols]

 1,70 Biosensor 5 Glucose measurement sensor element part 7 Ascorbic acid measurement sensor element part 9 Uric acid measurement sensor element part 11 Bilirubin measurement sensor element part 20 Concentration measuring device 30 Device body 35 Connector 40 Electronic control device 5a to 11a , 73a to 75a Working electrode 5b to 11b, 73b to 75b Reference electrode 5c to 11c, 73c to 75c Counter electrode 5d to 11d, 73d to 75d Identification layer 70 Biosensor 73 Lactate measurement sensor element part 75 Urea measurement sensor element part

[Table 1]

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁵ Identification code Internal reference number FI Technical display location G01N 27/26 371 A 7235-2J 27/28 331 Z 7235-2J 27/27 7235-2J G01N 27 / 46 336 G 7235-2J 338

Claims (1)

[Claims] 1. A concentration measuring device for measuring the concentration of a substance to be measured based on a biochemical reaction between the substance to be measured and a biological substance in a solution to be measured, the biological substance causing a biochemical reaction with the substance to be measured. And a measurement target substance sensor element section for converting the progress state of the biochemical reaction proceeding in the discrimination layer into an electric quantity and outputting the quantity, and inhibiting the conversion of the electric quantity in the measurement target substance sensor element section. And an interfering substance sensor element part for converting the progress state of the biochemical reaction progressing in the discriminating layer into an electric quantity and outputting the electric amount, and the interfering substance. In correcting the sensor output from the measurement target substance sensor element unit based on the sensor output from the sensor element unit, the sensor output from the interfering substance sensor element unit is used as the measurement target. A sensor output conversion means for converting the biological material carried on the discrimination layer in the substance sensor element unit into a sensor output when a biochemical reaction occurs with the interfering substance, and the conversion is performed from the sensor output of the measurement target substance sensor element unit. A concentration measuring device comprising: a concentration calculating unit that calculates the concentration of the substance to be measured based on the sensor output obtained by subtracting the sensor output.