KR100740363B1 - Apparatus for measuring bionics property using fixed enzyme - Google Patents

Abstract

The present invention relates to an apparatus for measuring biological properties using enzyme fixation in which an optimum activity temperature of an enzyme is maintained in a biosensor for measuring a recognition substance using enzyme activity. The biometrics measuring device is a biosensor device in which a planar heating element is mounted on a biosensor in which an enzyme is fixed, so that the optimum active temperature of the enzyme is directly corrected in the sensor unit, rather than indirectly correcting the algorithm of the sensor body. In addition, the bio-characteristics device is a strip of a glucose measuring device for extracting glucose by using the redox reaction of the enzyme, the strip is particularly provided with a heating unit for maintaining the optimum active temperature of the enzyme to work with the blood flow . In addition, the biological property measurement device is a patch of a glucose-free blood glucose meter, the patch is particularly provided with a heating unit for maintaining the optimum active temperature of the enzyme contained in the hydrophilic gel disk. According to the present invention, since a predetermined current is applied to the heating part included in the strip or patch, only the enzyme active site portion can be generated in a short time, so that an accurate and direct temperature correction can be expected.

Planar heating element, enzyme, biosensor, fixed, temperature, strip

Description

Apparatus for measuring bionics property using fixed enzyme}

1 is a graph showing the general enzyme activity with temperature.

FIG. 2 is an exploded perspective view showing the strip configuration of the glucose bleeding meter in which the optimum activity temperature of the enzyme is maintained.

3 is an exploded perspective view showing the patch configuration of a glucose-free blood glucose meter in which the optimum activity temperature of the enzyme is maintained.

Figure 4 is an embodiment measuring the concentration of glucose when the temperature correction in the present invention.

FIG. 5 is a diagram illustrating a graph measuring a temperature change and a temperature rising rate with time when temperature correction is performed using a planar heating element in the present invention.

<Explanation of symbols on main parts of the drawings>

200-Patch 215 of the Glucoseless Blood Gage Meter-Planar Heating Element

203-Printed Circuit Board 218-Lead Wires

206a, 206b-Gel Disc 224-Adhesive Film

209-Geltle

212-Gel Disc Receptacle

300-strip of glucose blood meter

303-Top plate 324-Strip recognition electrode

306-Reference Electrode 327-Blood Area Fixed Insulator

309-Air outlet 330-Working electrode

312-Middle 333-Lead Wire

315-Blood insertion path 336-Planar heating element insulator

318-Blood Recognition Electrode 339-Planar Heating Element

321-Bottom

The present invention relates to a biosensor unit in which the optimum enzyme activity temperature of a biosensor is directly corrected. Specifically, the enzyme is maximally active when an analyte reacts with an enzyme by combining a planar heating element on the market with a biosensor. The present invention relates to a biosensor device in which an enzyme optimum active temperature of a sensor to be maintained at a temperature is maintained.

Among the instruments for measuring the amount of analyte in a conventional sample, there is a blood glucose meter for measuring the amount of glucose in the blood, and the measuring device is a blood glucose measurement biosensor for measuring by taking blood and measuring the amount of glucose in the blood without blood sampling There is a blood measurement biosensor.

 As a measuring device for measuring the concentration of the analyte in the blood without blood collection, Korean Patent No. 10-453483 discloses a patch for the glucose extractor and its manufacturing method, Korean Patent Publication No. 2001-0067623 No. A blood glucose extraction method is disclosed. As a measuring device for measuring the concentration of analytes in blood by collecting blood, Republic of Korea Patent No. 10-0490185 discloses a reagent strip for measuring blood glucose content.

 The conventional blood collection or bloodless enzyme biosensor as described above detects a current signal of nano to several tens of nanoamps generated at this time when the enzyme is fixed to a support such as a polymer and a microcurrent is applied using the substrate-specific reaction of the enzyme. To quantify specific substrates. However, these enzyme biosensors have superior advantages in sensitivity and substrate specificity compared to the existing Yurin analysin sensors and spectrometer sensors, but there are technical problems in that they use enzyme reactions. Because enzymes are proteins that catalyze bioreactions, their activity depends on their pH, temperature, etc. In the case of blood sampling biosensors to measure the amount of glucose in the blood, a simple blood collection is performed when This is because there is a problem that an error occurs in accurate blood glucose measurement due to different temperatures when blood in the strip is inserted, and it is difficult to obtain consistent measurement values due to seasonal temperature changes.

In order to compensate for this, in the past, in the case of a blood glucose measurement biosensor, a method of keeping the corresponding meter in an appropriate storage container and combining the blood collection only in order to maintain a constant temperature of the strip part has been proposed. Over time, it was difficult to maintain the temperature of the strip portion of the meter.

The present invention has been made in view of this point, and in a biological property measurement apparatus using enzyme fixation, a fast time is provided through a corresponding exothermic portion provided with a predetermined invention to maintain a temperature at which enzymes such as glucose oxidase have maximum activity. It is aimed to enable accurate and direct temperature correction by heating only the portion of the enzyme active site inside.

 An insulator substrate having a reference electrode for measuring a reaction current between an analyte and an enzyme included in a sample, and a working electrode for applying a predetermined voltage for reaction between the analyte and an enzyme, and the reference electrode and the working electrode. An electrochemical enzyme biosensor having an enzyme fixed to react with the analyte to generate an amount of charge corresponding to the concentration of the substance, a planar heating element for maintaining an optimum active temperature of the enzyme of the biosensor A heat generating unit having a lead wire for applying a voltage to the planar heating element is provided.

In this case, the bio-specific device is a strip of a glucose bleeding meter which extracts glucose by using an redox reaction of an enzyme, and the strip discharges air existing inside when blood is injected with one of the working electrode or the reference electrode. A top plate having an air outlet, a middle plate having a groove-shaped blood insertion flow path formed to accommodate the predetermined amount of blood, another one of the working electrode or the reference electrode and the heat generating unit It consists of a bottom plate provided.

In addition, the strip of the glucose measuring device further includes an enzyme fixing plate for fixing the enzyme by printing the enzyme as long as the length of the blood insertion path, the enzyme fixing plate is preferably attached to the upper or lower plate of the strip.

The heat generating unit includes a lead wire which is divided into a positive electrode and a negative electrode side by side on the lower plate of the strip, and generates a predetermined amount of heat when voltage is applied to the lead wire while being connected to the lead wire to maintain an optimum active temperature of the enzyme. It consists of a heating element.

The planar heating element is characterized in that any one or more of the composition, area, length, thickness of the heating element is determined according to the temperature conditions of the enzyme according to the enzyme activity to be used, the maximum active temperature of the enzyme activity is 35 It is preferable that it is FIGS. 40 degrees, and it is preferable that the said lead wire consists of a conductive electrode containing silver, gold, and carbon.

The lower plate further includes a blood area fixing insulator for fixing a portion of the blood received through the flow path to a predetermined area, and the planar heating element is a silkscreen method, an SMT method, an airbrush method, a vacuum deposition method, a sputter method on the lower plate. It is preferably attached to any one of the electrospinning method, is formed to correspond to the area of the blood area fixed insulator is provided on the bottom of the lower plate.

In addition, the bio-specific device is a patch of a glucose-free blood glucose meter for extracting glucose using the redox reaction of the enzyme, the patch is a printed circuit board printed with an electrical electrode used to extract glucose, An electrode holding an enzyme capable of reacting with glucose to produce hydrogen peroxide, two hydrophilic gel discs containing a hydrophilic gel or enzyme, and a gel frame having two holes for accommodating the two hydrophilic gel discs; And a hole formed in a portion corresponding to the hole of the mold, and having a pressure-sensitive adhesive film attached to the lower side of the mold to support the gel disc in the same shape as the mold, wherein the heat generating portion is the printed circuit and the hydrophilic gel. It is preferable to be mounted between the disks.

The heat generating unit includes two planar heating elements formed in a plane shape so as to correspond to the hydrophilic gel disk, and a lead wire for applying a predetermined voltage to maintain the optimum active temperature of an enzyme to the two planar heating elements. In addition, the lead wire is preferably composed of a conductive electrode containing silver, gold, and carbon.

The planar heating element is preferably one or more of the component composition, area, length, thickness of the heating element is determined to suit the temperature conditions of the enzyme according to the enzyme activity to be used, the optimum activity temperature of the enzyme is from 35 degrees to It is preferable that it is 40 degrees. In addition, the planar heating element is attached to the insulating plate by any one of a silk screen method, SMT method, airbrush method, vacuum deposition method, sputter method, electrospinning method.

Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the accompanying drawings.

1 is a graph showing the general enzyme activity with temperature. First, the measuring method of the biosensor applied to the present invention will be described and the enzyme activity change according to the temperature and the temperature effect in the biosensor will be described with reference to the graph of FIG. 1.

Biosensors that measure the amount of analyte in body fluids include blood sampling meters, and the principle of operation is based on colorimetric or electrochemical methods. In the case of the colorimetric method, the intensity of coloration may change in the transport, pretreatment, amount of sample, reaction time and initiation time of the blood sample, and therefore, the coloration intensity must be precisely controlled, and blood coagulation or uric acid, There is a problem in that it can be interrupted by interference materials such as ascorbic acid and bilirubin. Therefore, the proposed electrochemical method to complement this point.

The electrochemical method has high selectivity and sensitivity, and even if the sample is turbid, the sample can be used without any additional pretreatment and can be accurately measured within a short time. However, these electrochemical methods have technical problems in that they use enzymatic reactions. Because the enzyme is a protein that catalyzes a biological reaction, its activity depends on its pH, temperature, etc. As a result, at other pH and temperature, the measured value is measured differently than the actual value, thereby causing a large error. Therefore, whether or not it is possible to maintain the pH and temperature conditions in which the enzyme is maximally active in the biosensor has emerged as an important fact, the present invention has been proposed for this purpose.

The properties of enzymes in biosensors, such as enzyme-immobilized biosensors, are very effective in trace amounts, with protein as the main component. At this time, any one substrate also has a substrate property that only one enzyme acts. In biosensors, it works best at the optimum temperature of 35 degrees and is denatured at 40 degrees or higher, and the reaction rate is doubled every time the temperature rises by 10 degrees from 5 to 40 degrees. do. The body's biological reaction is made by enzymes, and the chemical reaction proceeds well in the body temperature range (37 degrees), 1 atmosphere, and 1 neutral to 5.1 pH. Similarly, colorimetric, photometric, and electrochemical biosensors are used to catalyze enzymes, where enzymes bind to substrates to form enzyme-substrate complexes, resulting in lowered activation energies and chemical reactions. That is, it will only work at the right conditions in the organism, ie at the right temperature and pH.

Figure 1 shows a graph of the results confirmed through the actual experiment the activity pattern of the enzyme described above. In other words, when the temperature of the analyte solution is about 10 degrees, the measured current value shows a low value of about 3nA because the redox reaction is not performed properly.In the case of 20 degrees, the enzyme activity is slightly improved. As the current value is 11nA, the enzyme activity is better than the temperature of the analyte solution at 10 degrees. In addition, when the analyte solution has a temperature range of 37 degrees similar to the temperature of our blood, the enzyme activity is maximized and the measured current value shows a high value of 21 nA. The analyte solution is 40 degrees when the enzyme is denatured. When the ideal temperature is 45 degrees, the enzyme activity is again sensitive, and it can be seen that the measured current value drops to 15 nA.

That is, it can be seen from the graph of FIG. 1 that the optimum enzyme activity temperature is 35 degrees to 37 degrees, and it is important to maintain a proper temperature and pH in the enzyme active biosensor.

Therefore, the following describes the proposed strip of the glucose collection meter of the present invention and the patch of the glucose collection meter proposed to maintain the optimum activity temperature of the enzyme.

First, the patch configuration of the glucose-free blood glucose meter in which the optimum activity temperature of the enzyme is maintained will be described with reference to the exploded perspective view of FIG. 2.

The patch 200 of the glucose-free blood glucose meter of the present invention is composed of a printed circuit board 203, gel disks 206a and 206b, a gel frame 209, and an adhesive film 224.

The printed circuit board 203 is printed with an electrical electrode used to extract glucose in the blood, and two hydrophilic gel disks 206a and 206b each containing an enzyme and an hydrophilic gel or enzyme are fixed. Each contains an enzyme that can react with glucose to produce hydrogen peroxide.

The heat generating portion is formed at a portion corresponding to the positions of the two gel disks and serves to maintain a temperature at which the enzyme activity is optimal. The heat generating part is connected to the two planar heating elements 215 formed in a planar shape so as to correspond to the hydrophilic gel disks 206a and 206b, and the planar heating elements 215 are connected to the two planar heating elements 215. It may be composed of a lead wire 218 for application. In this case, the lead wire 218 is formed by dividing the anode and the cathode, it is preferably composed of a conductive electrode containing silver, gold, carbon.

In addition, the planar heating element 215 may freely adjust the composition, area, length, thickness, and resistance of the heating element to suit the temperature conditions of the enzyme according to the enzyme activity to be used. That is, the temperature can be maintained for a long time by maintaining a constant area, resistance, etc. of the heating element in order to maintain the optimum activity temperature of the enzyme 35 degrees to 40 degrees.

In this case, each of the planar heating element 215 and the lead wire 218 is any one of the silk screen method, SMT method, air brush method, vacuum deposition method, sputter method, electrospinning method on the lower surface of the printed circuit board 203. Can be attached in a manner.

In addition, although not shown in the drawing, the planar heating element printing pad 221 is an inner portion of the gel frame 209 for accommodating the two hydrophilic gel disks 206a and 206b, that is, each hole of the gel frame 209 and the two holes. It is also possible that the two hydrophilic gel disks 206a and 206b are attached to the abutting portion.

In addition, the strip 200 of the glucose measuring device of the present invention includes a gel frame 209 having two holes 212 for accommodating the two hydrophilic gel disks, and a frame in the same form as the gel frame 209. It is attached to the lower side, a hole is formed in a portion corresponding to the contact hole, and further comprises an adhesive film 224 for supporting the gel disk.

Next, the strip configuration of the glucose bleeding meter which maintains the optimum activity temperature of the enzyme will be described with reference to the exploded perspective view of FIG. 3.

Figure 3 is an exploded perspective view showing the strip configuration of the glucose bleeding meter in which the optimum activity temperature of the enzyme is maintained.

Strip 300 of the glucose measurement device of the present invention is composed of the upper plate 303, the middle plate 312, the lower plate 321 is coupled.

The top plate 303 includes an air outlet 309 and a reference electrode 306 that provides a reference potential for measuring a reaction current between glucose and enzyme contained in the introduced blood. In this case, the reference electrode 306 may be provided on the lower plate 321 in some cases.

The middle plate 312 has a groove-shaped blood insertion passage 315 formed to receive a predetermined amount of blood, which is connected to an air outlet 309 attached to the upper plate. The air outlet 309 serves to discharge the air present in the strip when blood is injected.

The lower plate 321 includes a working electrode 330 for applying a predetermined voltage to the blood in order to measure a reaction current between glucose in the blood and an enzyme, and a heating unit for maintaining an optimum active temperature of the enzyme. In addition, the working electrode 330 may be provided on the upper plate 303. The heat generating unit is composed of a conductive electrode including gold, silver, and carbon. The lead wire 333 is divided into an anode and a cathode and attached to the lower surface of the lower plate of the enzyme biosensor 300, and a voltage is applied to the lead wire 333. As it is composed of a planar heating element (339) for providing a predetermined amount of heat to the enzyme, if the temperature compensation application site is required for insulation may further include a planar heating element insulator 336 to insulate the site. The planar heating element 339 may be printed on any one of a silk screen method, an SMT method, an air brush method, a vacuum deposition method, a sputter method, and an electrospinning method and attached to the lower plate 321.

In this case, the planar heating element 339 has a resistance determined according to an area, a length, and a thickness, and the area, length, and thickness are preferably formed in proportion to the size of the blood area fixing insulator 327. At this time, the resistance value is obtained by calculating the power consumption (W) of the main body of the instrument by using the formula of power (W) = voltage (V) * current (A), and this is Joule's law, that is, RI ² = VI = V ² / It can be obtained by using the formula of R = W.

The lower plate 321 may further include a blood area fixing insulator 327 for fixing a portion of the blood received through the blood insertion channel 315 provided in the middle plate 312 to a predetermined area. The working electrode 330 provided on the lower plate 321 is formed in contact with the blood area fixed insulator 327.

In addition, the lower plate 321 may further include a blood recognition electrode 318 for recognizing blood inflow of the upper plate 303 and a strip recognition electrode 324 for strip recognition, and thus the upper plate 303, When the middle plate 312 and the lower plate 321 are formed, the blood measurement type biosensor is completed through a process of combining an enzyme fixation plate and a sensor immobilizing the enzyme by immobilizing the enzyme and immobilizing the enzyme by a surface immobilization method. At this time, the enzyme fixing plate is preferably provided on the upper plate 303 or the lower plate 321 of the strip 300 of the glucose collection meter by printing the enzyme as long as the blood insertion path 315.

When a voltage is applied to the lead wire 333 of the strip 300 with respect to the strip 300 of the glucose measuring device configured as described above, the temperature of the planar heating element 339 connected to the lead wire 333 becomes high in a short time. Therefore, when blood is injected into the strip, an error in the blood glucose measurement caused by the temperature difference does not occur due to the planar heating element 339 which is raised to an appropriate temperature of 37 degrees in advance. In addition, the planar heating element 339 raised to an appropriate temperature maintains a proper temperature at which the enzyme activity is maximized when a constant voltage is continuously applied to the lead wire 333, thereby improving the performance of the enzyme biosensor. do. In this case, the planar heating element is preferably used in a biosensor in which enzymes such as oxidase, amylase, and maltase have maximum activity in the body at 30 degrees to 40 degrees.

An embodiment in which blood glucose concentration is measured by using the biosensor equipped with the strip or patch of the present invention described above will be described with reference to FIG. 4.

In order to investigate the temperature correction effect in the biosensor equipped with the strip of the present invention, the glucose value was measured at 22 degrees and 32 degrees using a 45.0 mg / dL glucose standard solution. As can be seen in the graph, the measured glucose value was 44.6 ± 0.52 mL / dL at 32 ° C, and the measured glucose value was 39.5 ± 0.90mg / dL at 22 ° C. It was 11.1% lower than the concentration of the standard solution and the standard deviation was wide. That is, it can be seen that the enzyme activity at the temperature of 22 degrees is lower than the enzyme activity at the temperature of 32 degrees, it can be seen that the concentration of glucose measured at the temperature appropriate for the enzyme activity represents a more accurate measurement.

That is, as shown in FIG. 4, the temperature correction is not properly performed in the conventional biosensor, resulting in an error value 11/1% lower than that of the standard solution. On the other hand, the present invention can directly overcome the error limit of applying the algorithm by appropriately correcting the temperature condition and satisfying the pH condition by directly heating the strip itself by using a planar heating element to maximize the enzyme activity. It can be seen that.

Next, the temperature change with time of blood to be measured in the biosensor equipped with the strip or patch of the present invention will be described. For reference, Figure 5 shows a graph measuring the temperature change and the temperature rise rate with time when the temperature correction using the planar heating element in the present invention.

In FIG. 5, the temperature change and the temperature rise rate with time were measured while applying a voltage of 3 V to a planar heating element having a resistance of 29 ohms. When a constant voltage of 3 V is applied to the planar heating element, the planar heating element rapidly rises in temperature as shown in the drawing, and after 80 seconds, the temperature reaches equilibrium at a temperature of about 37 degrees. That is, when the user applies a constant voltage to the planar heating element according to the area, length, thickness, and resistance of the planar heating element, the temperature of the planar heating element increases in a short time according to the amount of the corresponding voltage, and the constant area and resistance of the planar heating element are increased. By keeping the same, it can be seen that the temperature of about 37 degrees at which the enzyme activity is maximized for a long time at the same power can be kept constant.

As a result, in the case of the conventional blood glucose measurement biosensor, the blood glucose measurement was performed by storing the measuring instrument in an appropriate storage container and using only the blood collection measurement method in order to maintain a constant temperature of the strip part. According to the present invention, there is a problem in that the measurement value is difficult to maintain the temperature of the strip part of the measuring device. However, in the strip of the present invention, if the constant area and resistance of the planar heating element are kept the same, the enzyme activity is maximized for a long time with low power. It can be seen that the temperature of 37 degrees can be kept constant.

As mentioned above, although this invention was demonstrated to the several example, this invention is not limited to a specific Example. Those skilled in the art will appreciate that various changes and modifications can be made without departing from the spirit of the invention.

As described above, according to the present invention, in the biological property measuring apparatus using enzyme immobilization, a predetermined heating part is included in order to maintain the temperature at which the enzyme of the analyte has the maximum activity, and a current is applied to the heating part within a short time. The exotherm of only the portion of the enzyme active site enables accurate and direct maintenance of the optimum activity temperature.

Claims (15)

delete delete delete delete delete delete delete delete delete delete An insulator substrate having a reference electrode for measuring a reaction current between an analyte and an enzyme included in a sample, and a working electrode for applying a predetermined voltage for reaction between the analyte and an enzyme, and the reference electrode and the working electrode. An electrochemical enzyme biosensor having an enzyme fixed to react with the analyte to generate an amount of charge corresponding to the concentration of the substance, a planar heating element for maintaining an optimum active temperature of the enzyme of the biosensor, In the living body measurement device having a heat generating unit having a lead wire for applying a voltage to the planar heating element, The biometric device is a patch of a glucose-free blood glucose meter for extracting glucose using an enzyme redox reaction, The patch includes a printed circuit board printed with an electrical electrode used to extract glucose, two hydrophilic gel discs each containing an enzyme capable of reacting with glucose to produce hydrogen peroxide, and the two A gel mold having two holes for accommodating a hydrophilic gel disk, and a hole formed in a portion corresponding to the hole of the mold, and attached to the lower side of the mold in the same shape as the mold to support the gel disk. With film, The heat generating part includes two planar heating elements formed in a plane shape so as to correspond to the hydrophilic gel disk, and a lead wire for applying a predetermined voltage to maintain the optimum active temperature of an enzyme to the two planar heating elements. An apparatus for measuring biological properties using enzyme fixation, which is mounted between a circuit and the hydrophilic gel disk. delete The method of claim 11, The planar heating element is a biological property measurement device using enzyme immobilization, characterized in that any one or more of the component composition, area, length, thickness of the heating element is determined to suit the temperature conditions of the enzyme in accordance with the enzyme activity to be used. delete The method of claim 11, The planar heating element is a bio-characteristic measuring device using enzyme fixation, characterized in that attached to the insulating plate by any one of a silk screen method, SMT method, airbrush method, vacuum deposition method, sputter method, electrospinning method.

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