A glucose bioelectrochemical sensor

Abstract

The application discloses a glucose bioelectrochemical sensor and relates to the technical field of glucose detection.The glucose bioelectrochemical sensor comprises the following steps: (1) preparing composite silver nanoparticles; (2) preparing electrode modification material; (3) electrode modification treatment; and (4) obtaining the glucose bioelectrochemical sensor.The glucose bioelectrochemical sensor prepared by the application has high sensitivity and a wide linear range; after the electrode is modified by the electrode modification material prepared by the application, the stability of the sensor can be greatly improved; and the biological activity of glucose oxidase can be maintained for a long time, so that the detection sensitivity of the sensor is effectively ensured.

Description

Technical Field

[0001] This invention belongs to the field of glucose detection technology, specifically a glucose bioelectrochemical sensor. Background Technology

[0002] Diabetes mellitus is a chronic metabolic disease characterized primarily by hyperglycemia, and the number of people with diabetes worldwide has been rising in recent years. Therefore, early prevention and care of diabetes should be given high priority. The components of human blood are important indicators for assessing health, and blood glucose levels are a key indicator for measuring metabolic capacity and clinical diagnosis of diabetes. Therefore, in order to manage and control the condition of diabetic patients, it is particularly important to regulate daily diet and exercise through real-time and continuous monitoring of the condition.

[0003] Blood glucose levels are a key indicator for assessing metabolic capacity and for the clinical diagnosis of diabetes. Current methods for detecting blood glucose mainly include liquid chromatography, fluorescence spectroscopy, capillary electrophoresis, and electrochemical methods. Most of these methods are slow or expensive. Among these, electrochemical sensors containing glucose oxidase (GOD) have attracted significant attention for blood glucose detection due to their advantages such as low detection limit, good selectivity, and high sensitivity.

[0004] High-purity glucose oxidase (GOD) is a pale yellow crystal, readily soluble in water, and can be formulated into liquid preparations. GOD is completely insoluble in organic solvents such as ether, chloroform, butanol, pyridine, glycerol, and ethylene glycol. Glucose oxidase has an effective pH range of 3.5–6.5, with an optimum pH of around 5.0. It will rapidly inactivate at pH > 8.0 or pH < 3.0 without the presence of a preservative.

[0005] Therefore, ensuring the activity of glucose oxidase is crucial, as it directly affects the sensitivity of glucose detection. Thus, to improve the detection sensitivity of the sensor, it is necessary to ensure high activity of glucose oxidase.

[0006] Therefore, further improvements to existing technologies are needed. Summary of the Invention

[0007] The purpose of this invention is to provide a glucose bioelectrochemical sensor to overcome the shortcomings of the prior art.

[0008] The technical solution adopted in this invention is as follows:

[0009] A glucose bioelectrochemical sensor includes the following steps:

[0010] (1) Preparation of composite silver nanoparticles;

[0011] (2) Preparation of electrode modification material:

[0012] First, prepare the ammonium persulfate solution: specifically, add 0.35-0.4g of ammonium persulfate to 10-15mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution;

[0013] Add glacial acetic acid to distilled water to prepare an acetic acid solution;

[0014] Add 1-2g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain chitosan solution;

[0015] Add 0.1-0.15g of composite silver nanoparticles to 100mL of chitosan solution and disperse by ultrasonication for 10min to obtain a composite silver nanoparticle dispersion.

[0016] Add 3-5g of aniline monomer and 5-7g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution.

[0017] Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base liquid and stirred for 3-5 minutes. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was completed, the mixture was ultrasonically dispersed for 1 minute to obtain the electrode modification material.

[0018] (3) Electrode modification treatment:

[0019] In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again.

[0020] (4) Insert the modified electrode into the glucose oxidase solution and immerse it for 8-10 hours. Then take it out and wait for the water to evaporate. Then coat the electrode surface with a 5% Nafion solution and dry it to obtain the final product.

[0021] As a further technical solution: the method for preparing composite silver nanoparticles in step (1) includes:

[0022] Disperse 0.12-0.18g of nano-silver particles uniformly into 50mL of deionized water, then add 3-5g of polyvinylpyrrolidone, stir at 5000r / min for 20min, filter, and then uniformly disperse into 50mL of ethanol solution to obtain nano-silver particle ethanol dispersion.

[0023] Add 3-4 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.2-0.3 g of TEOS, stir for 1 hour, then add 0.05-0.08 g of graphene, continue stirring for 8-10 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles.

[0024] As a further technical solution: the ethanol solution has a mass fraction of 75%;

[0025] The ammonia water has a mass fraction of 30%.

[0026] As a further technical solution: the mass fraction of the peracetic acid solution in step (2) is 1%.

[0027] As a further technical solution: the concentration of the hydrochloric acid solution in step (2) is 0.5 mol / L.

[0028] As a further technical solution: the inert atmosphere in step (2) is either nitrogen or neon.

[0029] As a further technical solution: the ultrasonic redispersion treatment time in step (3) is 2-3 minutes;

[0030] The drying and setting temperature is 38℃.

[0031] As a further technical solution: (4) The glucose oxidase solution is a 2000 U / mL glucose oxidase solution.

[0032] As a further technical solution: the immersion treatment temperature in step (4) is room temperature.

[0033] As a further technical solution: in step (4), a 5% Nafion solution is coated on the electrode surface, and the coating amount is 1:30-32 according to the mass ratio of electrode to Nafion solution.

[0034] The electrode modifier prepared by this invention has a complex three-dimensional network structure and carries a variety of functional groups, especially a large number of hydroxyl and amino groups. The composite silver nanoparticles are tightly bound to the electrode modifier through a large number of functional groups on their surface. Glucose oxidase can be bound to the electrode modifier through chemical cross-linking. The various active functional groups on the electrode modifier provide a large number of chemical cross-linking points for the immobilization of glucose oxidase. At the same time, the complex porous structure of the prepared electrode modifier provides a suitable environment and a large amount of immobilization space for glucose oxidase. Furthermore, the distributed composite silver nanoparticles can provide a bridge for the rapid transfer of electrons.

[0035] Beneficial effects:

[0036] This invention provides a glucose bioelectrochemical sensor. The glucose bioelectrochemical sensor prepared by this invention has high sensitivity and a wide linear range. After modifying the electrode with the electrode modifier prepared by this invention, the stability of the sensor can be greatly improved, and the bioactivity of glucose oxidase can be maintained for a long time, thereby effectively ensuring the detection sensitivity of the sensor.

[0037] The electrode modifier prepared by this invention not only has excellent conductivity, ensuring rapid electron transfer, but also good biocompatibility and good immobilization capacity for glucose oxidase. Attached Figure Description

[0038] Figure 1 This is a standard curve of anolyte current density corresponding to different scan rates. Detailed Implementation

[0039] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0040] Example 1

[0041] A glucose bioelectrochemical sensor includes the following steps:

[0042] (1) Preparation of composite silver nanoparticles; the methods for preparing composite silver nanoparticles include:

[0043] 0.12g of nano-silver particles were uniformly dispersed in 50mL of deionized water, and then 3g of polyvinylpyrrolidone was added. The mixture was stirred at 5000r / min for 20min, then filtered, and then uniformly dispersed in 50mL of ethanol solution to obtain an ethanol dispersion of nano-silver particles.

[0044] Add 3 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.2 g of TEOS, stir for 1 hour, then add 0.05 g of graphene, continue stirring for 8 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles; the ethanol solution has a mass fraction of 75%.

[0045] The ammonia solution has a mass fraction of 30%.

[0046] (2) Preparation of electrode modification material:

[0047] First, prepare the ammonium persulfate solution: specifically, add 0.35g of ammonium persulfate to 10mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution;

[0048] Add glacial acetic acid to distilled water to prepare an acetic acid solution;

[0049] Add 1g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain a chitosan solution.

[0050] 0.1g of composite silver nanoparticles were added to 100mL of chitosan solution and ultrasonically dispersed for 10min to obtain a composite silver nanoparticle dispersion.

[0051] Add 3g of aniline monomer and 5g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution.

[0052] Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base solution and stirred for 3 minutes. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was complete, the mixture was ultrasonically dispersed for 1 minute to obtain the electrode modification material. The mass fraction of the peracetic acid solution was 1%.

[0053] The concentration of the hydrochloric acid solution was 0.5 mol / L.

[0054] The inert atmosphere is nitrogen.

[0055] (3) Electrode modification treatment:

[0056] In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again.

[0057] (4) The modified electrode is inserted into the glucose oxidase solution and immersed for 8 hours at room temperature. Then it is taken out and the water evaporates. Then the electrode surface is coated with a 5% Nafion solution and dried. The glucose oxidase solution is a 2000 U / mL glucose oxidase solution.

[0058] Specifically, a 5% Nafion solution is coated onto the electrode surface, with the coating amount calculated at a mass ratio of 1:30 between the electrode and the Nafion solution.

[0059] Example 2

[0060] A glucose bioelectrochemical sensor includes the following steps:

[0061] (1) Preparation of composite silver nanoparticles; the methods for preparing composite silver nanoparticles include:

[0062] 0.14 g of silver nanoparticles were uniformly dispersed in 50 mL of deionized water, and then 3.5 g of polyvinylpyrrolidone was added. The mixture was stirred at 5000 r / min for 20 min, then filtered, and then uniformly dispersed in 50 mL of ethanol solution to obtain an ethanol dispersion of silver nanoparticles.

[0063] Add 3.6 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.24 g of TEOS, stir for 1 hour, then add 0.06 g of graphene, continue stirring for 9 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles; the ethanol solution has a mass fraction of 75%.

[0064] The ammonia solution has a mass fraction of 30%.

[0065] (2) Preparation of electrode modification material:

[0066] First, prepare the ammonium persulfate solution: specifically, add 0.38g of ammonium persulfate to 12mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution;

[0067] Add glacial acetic acid to distilled water to prepare an acetic acid solution;

[0068] Add 1.2g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain a chitosan solution.

[0069] 0.12 g of composite silver nanoparticles were added to 100 mL of chitosan solution and ultrasonically dispersed for 10 min to obtain a composite silver nanoparticle dispersion.

[0070] Add 3.5g of aniline monomer and 6g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution.

[0071] Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base solution and stirred for 3.5 min. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was complete, the mixture was ultrasonically dispersed for 1 min to obtain the electrode modification material. The mass fraction of the peracetic acid solution was 1%.

[0072] The concentration of the hydrochloric acid solution was 0.5 mol / L.

[0073] The inert atmosphere is nitrogen.

[0074] (3) Electrode modification treatment:

[0075] In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again.

[0076] (4) The modified electrode is inserted into the glucose oxidase solution and immersed for 9 hours at room temperature. Then it is taken out and the water evaporates. Then the electrode surface is coated with a 5% Nafion solution and dried. The glucose oxidase solution is a 2000 U / mL glucose oxidase solution.

[0077] Specifically, a 5% Nafion solution was coated onto the electrode surface, with the coating amount calculated at a mass ratio of 1:31 between the electrode and the Nafion solution.

[0078] Example 3

[0079] A glucose bioelectrochemical sensor includes the following steps:

[0080] (1) Preparation of composite silver nanoparticles; the methods for preparing composite silver nanoparticles include:

[0081] 0.15g of nano-silver particles were uniformly dispersed in 50mL of deionized water, and then 4g of polyvinylpyrrolidone was added. The mixture was stirred at 5000r / min for 20min, then filtered, and then uniformly dispersed in 50mL of ethanol solution to obtain an ethanol dispersion of nano-silver particles.

[0082] Add 3.5 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.28 g of TEOS, stir for 1 hour, then add 0.07 g of graphene, continue stirring for 9 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles; the ethanol solution has a mass fraction of 75%.

[0083] The ammonia solution has a mass fraction of 30%.

[0084] (2) Preparation of electrode modification material:

[0085] First, prepare the ammonium persulfate solution: specifically, add 0.38g of ammonium persulfate to 13mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution;

[0086] Add glacial acetic acid to distilled water to prepare an acetic acid solution;

[0087] Add 1.6g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain chitosan solution;

[0088] 0.14 g of composite silver nanoparticles were added to 100 mL of chitosan solution and ultrasonically dispersed for 10 min to obtain a composite silver nanoparticle dispersion.

[0089] Add 4g of aniline monomer and 6g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution.

[0090] Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base solution and stirred for 4 minutes. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was complete, the mixture was ultrasonically dispersed for 1 minute to obtain the electrode modification material. The mass fraction of the peracetic acid solution was 1%.

[0091] The concentration of the hydrochloric acid solution was 0.5 mol / L.

[0092] The inert atmosphere is nitrogen.

[0093] (3) Electrode modification treatment:

[0094] In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again.

[0095] (4) The modified electrode is inserted into the glucose oxidase solution and immersed for 9 hours at room temperature. Then it is taken out and the water evaporates. Then the electrode surface is coated with a 5% Nafion solution and dried. The glucose oxidase solution is a 2000 U / mL glucose oxidase solution.

[0096] Specifically, a 5% Nafion solution was coated onto the electrode surface, with the coating amount calculated at a mass ratio of 1:31 between the electrode and the Nafion solution.

[0097] Example 4

[0098] A glucose bioelectrochemical sensor includes the following steps:

[0099] (1) Preparation of composite silver nanoparticles; the methods for preparing composite silver nanoparticles include:

[0100] 0.18g of nano-silver particles were uniformly dispersed in 50mL of deionized water, and then 5g of polyvinylpyrrolidone was added. The mixture was stirred at 5000r / min for 20min, then filtered, and then uniformly dispersed in 50mL of ethanol solution to obtain an ethanol dispersion of nano-silver particles.

[0101] Add 4 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.3 g of TEOS, stir for 1 hour, then add 0.08 g of graphene, continue stirring for 10 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles; the ethanol solution has a mass fraction of 75%.

[0102] The ammonia solution has a mass fraction of 30%.

[0103] (2) Preparation of electrode modification material:

[0104] First, prepare the ammonium persulfate solution: specifically, add 0.4g of ammonium persulfate to 15mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution;

[0105] Add glacial acetic acid to distilled water to prepare an acetic acid solution;

[0106] Add 2g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain a chitosan solution.

[0107] 0.15g of composite silver nanoparticles were added to 100mL of chitosan solution and ultrasonically dispersed for 10min to obtain a composite silver nanoparticle dispersion.

[0108] Add 5g of aniline monomer and 7g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution.

[0109] Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base solution and stirred for 5 minutes. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was complete, ultrasonic dispersion was performed for 1 minute to obtain the electrode modification material. The mass fraction of the peracetic acid solution was 1%.

[0110] The concentration of the hydrochloric acid solution was 0.5 mol / L.

[0111] The inert atmosphere is nitrogen.

[0112] (3) Electrode modification treatment:

[0113] In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again.

[0114] (4) The modified electrode is inserted into the glucose oxidase solution and immersed for 10 hours at room temperature. Then it is taken out and the water evaporates. Then the electrode surface is coated with Nafion solution with a mass fraction of 5%. After drying, the electrode is obtained. The glucose oxidase solution is a glucose oxidase solution with a mass fraction of 2000 U / mL.

[0115] Specifically, a 5% Nafion solution was coated onto the electrode surface, with the coating amount calculated at a mass ratio of 1:32 between the electrode and the Nafion solution.

[0116] Comparative Example 1:

[0117] Based on Example 1, the composite silver nanoparticles in step (1) are replaced with pure silver nanoparticles, and the remaining steps are the same as the technical solution of Example 1.

[0118] test

[0119] The experiment was conducted using platinum sheets as electrodes:

[0120] The linear response range of the sensor under low glucose concentration was tested for both the example and comparative examples, with an operating voltage of +0.65V, as shown in Table 1.

[0121] Table 1

[0122] Example 1 0-10.5 Example 2 0-11.0 Example 3 0-10.8 Example 4 0-10.1 Comparative Example 1 0-7.3

[0123] As can be seen from Table 1, the sensor prepared by this invention has a wide linear response range.

[0124] Using Example 1 as the base sample, the effects of different operating voltages on the linear response range of the sensor under low glucose concentrations were compared:

[0125] Table 2

[0126]

[0127]

[0128] As can be seen from Table 2, as the operating voltage increases, the linear response range of the sensor first increases and then decreases.

[0129] The glucose detection sensitivity of the sensor in this embodiment of the invention is >172.6 μA / mMcm 2 .

[0130] A standard curve of anolyte current density corresponding to different scan rates was tested in a 0.2M phosphate buffer solution with a glucose concentration of 10mM and a pH of 7.0. Figure 1 .

[0131] The above description is only a preferred embodiment of the present invention, but the present invention is not limited to the scope of implementation shown. Any changes made in accordance with the concept of the present invention, or equivalent embodiments modified to have equivalent changes, shall be within the protection scope of the present invention as long as they do not exceed the spirit covered by the specification.

Claims

1. A glucose bioelectrochemical sensor, characterized in that: Includes the following steps: (1) Preparation of composite silver nanoparticles; (2) Preparation of electrode modification materials: First, prepare the ammonium persulfate solution: specifically, add 0.35-0.4g of ammonium persulfate to 10-15mL of distilled water, stir for 10min at room temperature until well mixed, and obtain the ammonium persulfate solution; Add glacial acetic acid to distilled water to prepare an acetic acid solution; Add 1-2g of chitosan to 100mL of acetic acid solution and stir to mix evenly to obtain chitosan solution; Add 0.1-0.15g of composite silver nanoparticles to 100mL of chitosan solution and disperse by ultrasonication for 10min to obtain a composite silver nanoparticle dispersion. Add 3-5g of aniline monomer and 5-7g of 3-aminophenylboronic acid to 100mL of distilled water in sequence, stir and mix well, then add 30mL of hydrochloric acid solution, adjust the temperature to 40℃, and continue stirring for 30min to obtain the base solution. Under an inert atmosphere, the composite silver nanoparticle dispersion was added to the base liquid and stirred for 3-5 minutes. Then, ammonium persulfate solution was added dropwise while stirring. After the addition was completed, the mixture was ultrasonically dispersed for 1 minute to obtain the electrode modification material. (3) Electrode modification treatment: In the preparation of electrode modifiers, after ultrasonic dispersion for 1 minute, the electrode is immediately inserted into the electrode modifier and ultrasonic redispersion is continued. After removal, drying and shaping, it is cleaned and dried again. (4) Insert the modified electrode into the glucose oxidase solution and immerse it for 8-10 hours. Then take it out and wait for the water to evaporate. Then coat the electrode surface with a 5% Nafion solution and dry it to obtain the product. The method for preparing composite silver nanoparticles described in step (1) includes: Disperse 0.12-0.18g of nano-silver particles uniformly into 50mL of deionized water, then add 3-5g of polyvinylpyrrolidone, stir at 5000r / min for 20min, filter, and then uniformly disperse into 50mL of ethanol solution to obtain nano-silver particle ethanol dispersion. Add 3-4 mL of ammonia to the ethanol dispersion of silver nanoparticles, stir and mix, then add 0.2-0.3 g of TEOS, stir for 1 hour, then add 0.05-0.08 g of graphene, continue stirring for 8-10 hours, then centrifuge, wash and dry to obtain composite silver nanoparticles.

2. The glucose bioelectrochemical sensor according to claim 1, characterized in that: The ethanol solution has a mass fraction of 75%. The ammonia water has a mass fraction of 30%.

3. The glucose bioelectrochemical sensor according to claim 1, characterized in that: The acetic acid solution in step (2) has a mass fraction of 1%.

4. The glucose bioelectrochemical sensor according to claim 1, characterized in that: The concentration of the hydrochloric acid solution in step (2) is 0.5 mol / L.

5. A glucose bioelectrochemical sensor according to claim 1, characterized in that: The inert atmosphere mentioned in step (2) is either nitrogen or neon.

6. A glucose bioelectrochemical sensor according to claim 1, characterized in that: The ultrasonic redispersion treatment in step (3) takes 2-3 minutes; The drying and setting temperature is 38℃.

7. A glucose bioelectrochemical sensor according to claim 1, characterized in that: (4) The glucose oxidase solution mentioned is a glucose oxidase solution with a concentration of 2000 U / mL.

8. A glucose bioelectrochemical sensor according to claim 1, characterized in that: The immersion treatment temperature in step (4) is room temperature.

9. A glucose bioelectrochemical sensor according to claim 1, characterized in that: In step (4), a 5% Nafion solution is coated on the electrode surface, with the coating amount calculated as an electrode to Nafion solution mass ratio of 1:30-32.

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