CN111982989A - Preparation method and application of a SiO2-MWCNTs enzyme-catalyzed glucose electrochemical sensor - Google Patents

Abstract

_The invention discloses a preparation method and application of a SiO ₂ MWCNTs enzyme _- catalyzed glucose electrochemical sensor, and belongs to the technical field of electrochemical sensors. The milled _SiO2 nanopowder was added to the carbon nanotube NMP dispersion; 1 ml of deionized water was added to the carbon nanotube NMP dispersion, and the milled _SiO2 nanopowder was mixed with carbon The nanotube NMP dispersion was mixed evenly, and the SiO ₂ ‑MWCNTs composite material was fully obtained after dispersion; 0.0700 g of thionine was weighed into a beaker, and deionized water at 40°C was added to the beaker to dissolve; the dissolved thionine solution was poured into Dilute to a 25mL brown volumetric flask to obtain a 0.01M thionine solution; take 0.001g of chloroauric acid in a beaker, add deionized water to the beaker and stir well with a glass rod.

Description

Preparation method and application of a SiO2-MWCNTs enzyme-catalyzed glucose electrochemical sensor

technical field

The invention relates to an electrochemical sensor, in particular to a preparation method and application of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor, and belongs to the technical field of electrochemical sensors.

Background technique

The accurate detection of glucose is of great value for biomedicine. In modern medicine, due to the sharp increase in the number of diabetics, it is a growing threat to human beings, but the lack of its diagnosis and treatment technology has always plagued modern medicine. By detecting the glucose content in the blood or saliva of diabetic patients to quickly and accurately determine the blood sugar concentration, the blood sugar of diabetic patients can be effectively monitored and treated.

At present, there are mainly the following methods for detecting glucose concentration: kinetic spectrophotometry, laser Raman spectroscopy, internal standard method, and biosensor measurement. Among them, electrochemical glucose sensors with the advantages of low cost, high reliability, and easy operation are widely used. Since the first glucose oxidase biosensor produced by Updike et al. in 1967, it has been widely used by scientists all over the world due to its simple operation, low production cost, easy miniaturization, high sensitivity, short response time, good selectivity and stability. Developed, electrochemical glucose sensors are divided into enzymatic electrochemical glucose sensors and non-enzymatic electrochemical glucose sensors according to different catalytic methods. Among them, non-enzymatic glucose electrochemical sensors usually have the characteristics of high sensitivity and high specificity, but their activity It is susceptible to interference from external environments such as humidity, temperature, and pH, which leads to the shortcomings of poor reproducibility and stability in the detection of glucose. Compared with the enzyme electrochemical glucose sensor, the enzyme electrochemical glucose sensor The sensor has the advantages of low detection limit, wide detection range, good stability and low cost, which makes it widely concerned.

In the prior art, the electrode material of the enzyme-catalyzed glucose electrochemical sensor has the disadvantage that it is easily affected by other interfering substances such as uric acid and chloride ions in the process of detecting glucose, which makes the final detection result biased;

The existing enzymatic glucose electrochemical sensor has a narrow linear range for detecting glucose, which is smaller than the fasting range of normal blood glucose concentration in the human body, which greatly limits the detection of diabetes.

Therefore, it is urgent to research new electrode materials that are not easily interfered by other substances and have a wide detection range. For this reason, a preparation method and application of SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor are designed to optimize the above problems.

SUMMARY OF THE INVENTION

The main purpose of the present invention is to provide a preparation method and application of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor, the composite material is used for the construction of the sensor sensing interface, so as to achieve the effect of improving the sensitivity of the sensor, and then use glucose to oxidize The principle of enzyme-catalyzed glucose reaction is tested to realize the quantitative detection of glucose. The prepared composite material probe not only has strong catalytic conductivity and strong adsorption capacity, but also has the advantages of simple detection method, convenient carrying and low cost.

Object of the present invention can be achieved by adopting the following technical solutions:

A preparation method of SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor, comprising the following steps:

Step 1: take 0.01-0.05g SiO ₂ nano-powder, fully grind the SiO ₂ nano-powder, and add the milled SiO ₂ nano-powder into the carbon nanotube NMP dispersion;

Step 2: Add 1 ml of deionized water to the carbon nanotube NMP dispersion, continuously vibrate with an ultrasonic cleaner for 2 d, mix the milled _SiO2 nanopowder with the carbon nanotube NMP dispersion evenly, and fully obtain _SiO2 after dispersion -MWCNTs composite material;

Step 3: Weigh 0.0700g of thionine into the beaker, add deionized water at 40°C to the beaker to dissolve;

Step 4: Pour the dissolved thionine solution into a 25mL brown volumetric flask to constant volume to obtain a 0.01M thionine solution;

Step 5: Take 0.001g of chloroauric acid in a beaker, then add deionized water to the beaker and stir well with a glass rod;

Step 6: The chloroauric acid solution in the beaker is heated to boiling and then dripped with 0.01% sodium citrate. When the color does not change, heat for another 15 minutes to obtain a nano-gold solution;

Step 7: Take the glassy carbon electrode and polish it to a mirror surface with alumina powder with a diameter of 0.3 μm and a diameter of 0.05 μm in turn;

Step 8: Rinse the polished glassy carbon electrode with deionized water, place it in absolute ethanol for ultrasonic cleaning for 15 minutes, and then place it in deionized water for ultrasonic cleaning again for 15 minutes;

Step 9: Place the cleaned glassy carbon electrode in a sulfuric acid solution, and activate the electrode by scanning 15-25 circles at a potential range of -0.6-1.0V by cyclic voltammetry;

Step 10: Rinse with deionized water after activation, and put it in deionized water for later use;

Step 11: Take 4 μl of the SiO ₂ -MWCNTs composite material prepared in Step 2, drop it on the surface of the activated glassy carbon electrode, and store it in a refrigerator at 4-8 °C for about 4 hours to form a uniform solid film ;

Step 12: Apply 4 μL of thionine, 4 μL of nano-gold, and 4 μL of glucose oxidase respectively. After each layer of modification, store it in a refrigerator at 4-8 °C for about 4 hours. After it is fixed, continue to modify the next layer to make it A uniform solid film is formed.

Preferably, in step 2, both the chloroauric acid and the sodium citrate solution are of analytical grade.

Preferably, in step 9, the concentration of the sulfuric acid solution is 0.1M.

A kind of SiO ₂ -MWCNTs has the application of enzyme-catalyzed glucose electrochemical sensor, comprising the following steps:

Step 1: Weigh a certain amount of dipotassium hydrogen phosphate into a beaker, add deionized water to the beaker to dissolve it to a constant volume to prepare a PBS buffer solution A with a concentration of 0.1mol/L;

Step 2: Weigh a certain amount of sodium dihydrogen phosphate into another beaker, and add deionized water to the beaker to dissolve to constant volume to prepare a PBS buffer solution B with a concentration of 0.1 mol/L;

Step 3: Mix the A and B solutions in Step 1 and Step 2 in a certain proportion to form a PBS buffer solution with pH=7.00;

Step 4: Weigh a certain amount of glucose to dissolve in PBS buffer solution, make up to a constant volume to prepare a glucose solution with a concentration of 0.1M, and use ultrapure water to gradient dilution to prepare a standard solution for detection;

Step 5: Weigh a certain amount of potassium ferricyanide, dissolve it in ultrapure water, and prepare a 0.1M potassium ferricyanide solution in a constant volume;

Step 6: Take 4 μL of saliva and dilute it 10,000 times;

Step 7: Use the PBS buffer solution with pH=7.00 obtained in step 3 and the potassium ferricyanide solution obtained in step 5 as the bottom liquid, use a three-electrode system for detection, and set working conditions;

Step 8: In 0.05M PBS (pH=7), the electrochemical properties of the electrode modification process were characterized by cyclic voltammetry;

Step 9: Under the working conditions of Step 7, pipette 5mL of PBS into the electrolytic cell, and take the glucose standard solution in sequence from the lowest concentration, and take 1 μL, 2 μL, 4 μL, 5 μL, 7 μL, 8 μL, 9 μL of each concentration and take it once. Measure once and add it to the bottom liquid; stir the bottom liquid to be tested for 1-2min with a magnetic stirrer to make it evenly mixed, and draw a standard curve after the test is completed;

Step 10: Under the working conditions of Step 7, pipette 5 mL of PBS into the electrolytic cell, stir 1 μL of the sample prepared in Step 6 for 1-2 min, and mix it evenly for detection, and calculate the sample concentration from the standard curve obtained in Step 9 above. .

Preferably, the three-electrode system in step 7 is: a glassy carbon electrode modified with a silica/carbon nanotube composite material as a working electrode, a platinum electrode as a counter electrode, and a saturated calomel electrode as a reference electrode.

Preferably, the working conditions set in step 7 are: the static time of CV detection is 2min, the initial voltage is 1V, the vertex voltage is 0.8V, the sampling interval is 0.02V, the scanning speed is 0.05V/s, and the sensitivity is 1μA/V.

Beneficial technical effects of the present invention:

The invention provides a preparation method and application of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor,

1. The composite material is used for the construction of the sensor sensing interface to achieve the effect of improving the sensitivity of the sensor, and then the principle of glucose oxidase catalyzing the glucose reaction is used to conduct experiments to realize the quantitative detection of glucose. The prepared composite material probe not only It has strong catalytic conductivity and strong adsorption capacity, and has the advantages of simple detection method, convenient carrying and low cost;

2. The lowest detection limit of the present invention for glucose can reach 1.960×10-37M, and it exhibits good linearity within 1.960×10-37-1.916×10-34M and 1.915×10-34-1,873×10-31M relation;

3. The saliva sample processing method is simple and easy to operate.

Description of drawings

1 is a schematic diagram of the operation of preparing a modified electrode according to a preferred embodiment of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor preparation method and application of the present invention;

2 is a CV characterization diagram of the electrochemical properties of the electrode modification process according to a preferred embodiment of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor preparation method and application of the present invention;

Fig. 3 is a working principle diagram of a three-electrode detection system according to a preferred embodiment of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor preparation method and application of the present invention;

4 is a standard curve diagram drawn after detecting a standard sample according to a preferred embodiment of the preparation method and application of a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor of the present invention.

Detailed ways

In order to make the technical solution of the present invention clearer and clearer to those skilled in the art, the present invention will be described in further detail below with reference to the embodiments and accompanying drawings, but the embodiments of the present invention are not limited thereto.

As shown in FIG. 1 to FIG. 4 , a method for preparing a SiO ₂ -MWCNTs enzyme-catalyzed electrochemical glucose sensor provided in this embodiment includes the following steps:

Step 1: take 0.01-0.05g SiO ₂ nano-powder, fully grind the SiO ₂ nano-powder, and add the milled SiO ₂ nano-powder into the carbon nanotube NMP dispersion;

Step 2: Add 1 ml of deionized water to the carbon nanotube NMP dispersion, continuously vibrate with an ultrasonic cleaner for 2 d, mix the milled _SiO2 nanopowder with the carbon nanotube NMP dispersion evenly, and fully obtain _SiO2 after dispersion -MWCNTs composite material;

Step 3: Weigh 0.0700g of thionine into the beaker, add deionized water at 40°C to the beaker to dissolve;

Step 4: Pour the dissolved thionine solution into a 25mL brown volumetric flask to constant volume to obtain a 0.01M thionine solution;

Step 5: Take 0.001g of chloroauric acid in a beaker, then add deionized water to the beaker and stir well with a glass rod;

Step 6: The chloroauric acid solution in the beaker is heated to boiling and then dripped with 0.01% sodium citrate. When the color does not change, heat for another 15 minutes to obtain a nano-gold solution;

Step 7: Take the glassy carbon electrode and polish it to a mirror surface with alumina powder with a diameter of 0.3 μm and a diameter of 0.05 μm in turn;

Step 8: Rinse the polished glassy carbon electrode with deionized water, place it in absolute ethanol for ultrasonic cleaning for 15 minutes, and then place it in deionized water for ultrasonic cleaning again for 15 minutes;

Step 9: Place the cleaned glassy carbon electrode in a sulfuric acid solution, and activate the electrode by scanning 15-25 circles at a potential range of -0.6-1.0V by cyclic voltammetry;

Step 10: Rinse with deionized water after activation, and put it in deionized water for later use;

Step 11: Take 4 μl of the SiO ₂ -MWCNTs composite material prepared in Step 2, drop it on the surface of the activated glassy carbon electrode, and store it in a refrigerator at 4-8 °C for about 4 hours to form a uniform solid film ;

Step 12: Apply 4 μL of thionine, 4 μL of nano-gold, and 4 μL of glucose oxidase respectively. After each layer of modification, store it in a refrigerator at 4-8 °C for about 4 hours. After it is fixed, continue to modify the next layer to make it A uniform solid film is formed.

In this example, in step 2, both chloroauric acid and sodium citrate solution are of analytical grade.

In this embodiment, the concentration of sulfuric acid solution in step 9 is 0.1M.

As shown in Figures 1-4, a SiO ₂ -MWCNTs has an enzyme-catalyzed glucose electrochemical sensor application, including the following steps:

Step 1: Weigh a certain amount of dipotassium hydrogen phosphate into a beaker, add deionized water to the beaker to dissolve it to a constant volume to prepare a PBS buffer solution A with a concentration of 0.1mol/L;

Step 2: Weigh a certain amount of sodium dihydrogen phosphate into another beaker, and add deionized water to the beaker to dissolve to constant volume to prepare a PBS buffer solution B with a concentration of 0.1 mol/L;

Step 3: Mix the A and B solutions in Step 1 and Step 2 into a PBS buffer solution with pH=7.00 in a certain proportion;

Step 4: Weigh a certain amount of glucose to dissolve in PBS buffer solution, make up to a constant volume to prepare a glucose solution with a concentration of 0.1M, and use ultrapure water to gradient dilution to prepare a standard solution for detection;

Step 5: Weigh a certain amount of potassium ferricyanide, dissolve it in ultrapure water, and prepare a 0.1M potassium ferricyanide solution in a constant volume;

Step 6: Take 4 μL of saliva and dilute it 10,000 times;

Step 7: Use the PBS buffer solution with pH=7.00 obtained in step 3 and the potassium ferricyanide solution obtained in step 5 as the bottom liquid, use a three-electrode system for detection, and set working conditions;

Step 8: In 0.05M PBS (pH=7), the electrochemical properties of the electrode modification process were characterized by cyclic voltammetry;

Step 9: Under the working conditions of Step 7, pipette 5mL of PBS into the electrolytic cell, and take the glucose standard solution in sequence from the lowest concentration, and take 1 μL, 2 μL, 4 μL, 5 μL, 7 μL, 8 μL, 9 μL of each concentration and take it once. Measure once and add to the bottom liquid; stir the bottom liquid to be tested for 1-2min with a magnetic stirrer to make it evenly mixed, and draw a standard curve after the test is completed, and the drawn standard curve is shown in Figure 4;

Step 10: Under the working conditions of Step 7, pipette 5 mL of PBS into the electrolytic cell, stir 1 μL of the sample prepared in Step 6 for 1-2 min and mix it evenly for detection, and calculate the sample concentration from the standard curve obtained in Step 9 above. .

As shown in Figure 3 in this example, the three-electrode system in step 7 is: a glassy carbon electrode modified with silica/carbon nanotube composite material as a working electrode, a platinum electrode as a counter electrode, and a saturated calomel electrode as a reference electrode.

In this embodiment, the working conditions set in step 7 are: the static time of CV detection is 2min, the initial voltage is 1V, the vertex voltage is 0.8V, the sampling interval is 0.02V, and the scanning speed is 0.05V/s , the sensitivity is 1μA/V.

During the test, glucose molecules diffused from the solution to the surface of the composite electrode material, and glucose oxidase catalyzed the reaction of glucose under a certain voltage, and then electron transfer occurred to generate a current signal, which was finally presented as a cyclic voltammetry image in the electrochemical workstation.

The above are only further embodiments of the present invention, but the protection scope of the present invention is not limited thereto. Equivalent replacements or changes to the concept all belong to the protection scope of the present invention.

Claims (6)

1. a kind of SiO ₂ -MWCNTs has an enzyme-catalyzed glucose electrochemical sensor preparation method, it is characterized in that: comprise the steps: Step 1: take 0.01-0.05g SiO ₂ nano-powder, fully grind the SiO ₂ nano-powder, and add the milled SiO ₂ nano-powder into the carbon nanotube NMP dispersion; Step 2: Add 1 ml of deionized water to the carbon nanotube NMP dispersion, continuously vibrate with an ultrasonic cleaner for 2 d, mix the milled _SiO2 nanopowder with the carbon nanotube NMP dispersion evenly, and fully obtain _SiO2 after dispersion -MWCNTs composite material; Step 3: Weigh 0.0700g of thionine into the beaker, add deionized water at 40°C to the beaker to dissolve; Step 4: Pour the dissolved thionine solution into a 25mL brown volumetric flask to constant volume to obtain a 0.01M thionine solution; Step 5: Take 0.001g of chloroauric acid in a beaker, then add deionized water to the beaker and stir well with a glass rod; Step 6: The chloroauric acid solution in the beaker is heated to boiling and then dripped with 0.01% sodium citrate. When the color does not change, heat for another 15 minutes to obtain a nano-gold solution; Step 7: Take the glassy carbon electrode and polish it to a mirror surface with alumina powder with a diameter of 0.3 μm and a diameter of 0.05 μm in turn; Step 8: Rinse the polished glassy carbon electrode with deionized water, place it in absolute ethanol for ultrasonic cleaning for 15 minutes, and then place it in deionized water for ultrasonic cleaning again for 15 minutes; Step 9: Place the cleaned glassy carbon electrode in a sulfuric acid solution, and activate the electrode by scanning 15-25 circles at a potential range of -0.6-1.0V by cyclic voltammetry; Step 10: Rinse with deionized water after activation, and put it in deionized water for later use; Step 11: Take 4 μl of the SiO ₂ -MWCNTs composite material prepared in Step 2, drop it on the surface of the activated glassy carbon electrode, and store it in a refrigerator at 4-8 °C for about 4 hours to form a uniform solid film ; Step 12: Apply 4 μL of thionine, 4 μL of nano-gold, and 4 μL of glucose oxidase respectively. After each layer of modification, store it in a refrigerator at 4-8 °C for about 4 hours. After it is fixed, continue to modify the next layer to make it A uniform solid film is formed. 2 . The method for preparing a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor according to claim 1 , wherein in step 2, both chloroauric acid and sodium citrate solution are of analytical grade. 3 . 3 . The method for preparing a SiO ₂ -MWCNTs enzyme-catalyzed glucose electrochemical sensor according to claim 2 , wherein the concentration of the sulfuric acid solution in step 9 is 0.1M. 4 . 4. a kind of SiO ₂ -MWCNTs has enzyme catalysis glucose electrochemical sensor application, it is characterized in that: comprise the steps: Step 1: Weigh a certain amount of dipotassium hydrogen phosphate into a beaker, add deionized water to the beaker to dissolve it to a constant volume to prepare a PBS buffer solution A with a concentration of 0.1mol/L; Step 2: Weigh a certain amount of sodium dihydrogen phosphate into another beaker, and add deionized water to the beaker to dissolve to constant volume to prepare a PBS buffer solution B with a concentration of 0.1 mol/L; Step 3: Mix the A and B solutions in Step 1 and Step 2 in a certain proportion to form a PBS buffer solution with pH=7.00; Step 4: Weigh a certain amount of glucose to dissolve in PBS buffer solution, make up to a constant volume to prepare a glucose solution with a concentration of 0.1M, and use ultrapure water to gradient dilution to prepare a standard solution for detection; Step 5: Weigh a certain amount of potassium ferricyanide, dissolve it in ultrapure water, and prepare a 0.1M potassium ferricyanide solution in a constant volume; Step 6: Take 4 μL of saliva and dilute it 10,000 times; Step 7: Use the PBS buffer solution with pH=7.00 obtained in step 3 and the potassium ferricyanide solution obtained in step 5 as the bottom liquid, use a three-electrode system for detection, and set working conditions; Step 8: In 0.05M PBS (pH=7), the electrochemical properties of the electrode modification process were characterized by cyclic voltammetry; Step 9: Under the working conditions of Step 7, pipette 5mL of PBS into the electrolytic cell, and take the glucose standard solution in sequence from the lowest concentration, and take 1 μL, 2 μL, 4 μL, 5 μL, 7 μL, 8 μL, 9 μL of each concentration and take it once. Measure once and add it to the bottom liquid; stir the bottom liquid to be tested for 1-2min with a magnetic stirrer to make it evenly mixed, and draw a standard curve after the test is completed; Step 10: Under the working conditions of Step 7, pipette 5 mL of PBS into the electrolytic cell, stir 1 μL of the sample prepared in Step 6 for 1-2 min, and mix it evenly for detection, and calculate the sample concentration from the standard curve obtained in Step 9 above. . 5. a kind of _SiO2 -MWCNTs according to claim 4 has the application of enzyme-catalyzed glucose electrochemical sensor, it is characterized in that: wherein in step 7, the three-electrode system is: the glass with silica/carbon nanotube composite material modified The carbon electrode was used as the working electrode, the platinum electrode was used as the counter electrode, and the saturated calomel electrode was used as the reference electrode. 6. a kind of SiO ₂ -MWCNTs according to claim 4 has enzyme catalysis glucose electrochemical sensor application, it is characterized in that: wherein the working condition set in step 7 is: the rest time of CV detection is 2min, the initial voltage is 1V, the vertex voltage is 0.8V, the sampling interval is 0.02V, the scanning speed is 0.05V/s, and the sensitivity is 1μA/V.

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