Interfacial enzyme-catalyzed hydrogel-encapsulated composite flexible bioelectrode and preparation and application thereof
By preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel, the contradiction between ionic conductivity and mechanical properties of hydrogel bioelectrodes was resolved, achieving efficient biological signal conversion and multiple signal monitoring, which is suitable for real-time and continuous monitoring of physiological metabolites.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- TONGJI UNIV
- Filing Date
- 2023-08-14
- Publication Date
- 2026-07-21
AI Technical Summary
Existing hydrogel bioelectrodes suffer from a trade-off between ionic conductivity and mechanical properties during use, which limits their application in biological detection and human-computer interaction devices.
By processing inorganic materials into flexible fibrous electrode materials, modifying them with conductive nanomaterials and loading enzymes, combined with gel precursor solution coating and vacuum freeze-drying, a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel was prepared.
It achieves efficient conversion of biochemical energy into electrical energy, improves catalytic activity and sensitivity, has good biocompatibility and mechanical strength, is suitable for long-term use, and can simultaneously monitor multiple biomarkers.
Smart Images
Figure CN117133510B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of flexible bioelectrode materials technology, and in particular to a composite flexible bioelectrode encapsulated by an interfacial enzyme-catalyzed hydrogel, its preparation and application. Background Technology
[0002] Metabolic disorders severely impact normal human life and health. With improved living standards, an increasing number of people are suffering from metabolic disorders. Furthermore, the occurrence of one metabolic disorder is often accompanied by the simultaneous occurrence of other metabolic disorders. Real-time, continuous, and synchronous in-situ monitoring of important physiological metabolic markers (such as blood glucose, uric acid, lactic acid, electrolytes, and water) is of significant research value for timely detection and prediction of potential physiological and pathological risks, providing health guidance for clinical practice. Currently, many wearable real-time monitoring devices for physiological metabolites have been researched and developed, most of which rely on electrochemistry or chemical reactions to achieve reliable and stable monitoring. However, precisely because of this, these wearable devices often require rigid electrodes for signal acquisition, greatly limiting their convenience and comfort.
[0003] As an energy conversion and supply device, enzyme biofuel cells (EBFCs) can utilize abundant endogenous substances in the human body, such as lactic acid, vitamin C, and glucose, as fuel to convert these substances into electrical energy. This is an effective way to convert chemical signals into electrical signals, providing a new approach for constructing wearable monitoring devices with "implantable" internal power sources. Considering the elasticity, toughness, and dynamic mechanical properties of biological skin, flexible hydrogel-coated electrodes with good biocompatibility, adjustable hardness, and rich functionality are expected to become the preferred material for constructing a new generation of integrated diagnostic and therapeutic systems for monitoring, diagnosing, and evaluating physiological metabolites.
[0004] Hydrogel bioelectrodes use flexible, modifiable conductive fibers as the matrix material. By modifying the fibers with different functional materials, their electron transfer performance can be regulated to achieve the target precision. By loading different oxidases or reductases, biocathodes (reductases) and anodes (oxidases) can be formed to form EBFC electronic circuits and achieve efficient conversion of "biological signals to electrical signals".
[0005] However, hydrogel bioelectrodes also encounter some problems during use, the most significant of which is the contradiction between the ionic conductivity and mechanical properties of the hydrogel. Hydrogels with mechanical properties that meet the electrode requirements often have low conductivity due to their dense network of pores, especially when combined with conductive fibers, which can also affect the electrical properties of the conductive fibers. Conversely, hydrogels with high ionic conductivity often have weak mechanical properties, particularly after swelling in water, making them prone to breakage and posing biosafety risks. This not only limits the application of hydrogel materials in biosensoring but also hinders the development of flexible electrodes in human-computer interaction devices. Summary of the Invention
[0006] The purpose of this invention is to overcome the contradiction between ionic conductivity and mechanical properties of hydrogel bioelectrodes in the prior art, and to provide a composite flexible bioelectrode encapsulated by an interfacial enzyme-catalyzed hydrogel, as well as its preparation and application.
[0007] The objective of this invention can be achieved through the following technical solutions:
[0008] One of the technical solutions of the present invention is to provide a method for preparing a composite flexible bioelectrode encapsulated by an interfacial enzyme-catalyzed hydrogel, comprising the following steps:
[0009] S1. Processing inorganic materials into flexible fibrous electrode materials;
[0010] S2. Modify the flexible fibrous electrode material obtained in step S1 with conductive nanomaterials to obtain the electrode substrate;
[0011] S3. Load the enzyme onto the electrode substrate obtained in step S2 to obtain an enzyme-loaded electrode, wherein the enzyme is selected from any one of oxidase, peroxidase, and dehydrogenase.
[0012] S4. Mix the monomer, crosslinking agent, electrolyte, initiator and solvent, and stir until completely dissolved into a transparent and homogeneous solution to obtain a gel precursor solution, wherein the monomer is selected from acrylate monomers or acrylic monomers;
[0013] S5. Coat the gel precursor solution obtained in step S4 onto the surface of the enzyme-carrying electrode obtained in step S3, cure it into a gel, and then freeze-dry it under vacuum to obtain a hydrogel-encapsulated composite flexible bioelectrode.
[0014] In some specific embodiments, in step S1, the inorganic material is selected from any one of carbon fiber, carbon nanotube, polyacetylene fiber, stainless steel fiber, and copper fiber; the processing technology is selected from any one or more of spinning, twisting, weaving, and winding.
[0015] In some specific embodiments, in step S2, the conductive nanomaterial is selected from any one or more of metal nanoparticles, single-walled carbon nanotubes, multi-walled carbon nanotubes, carbon nanorods, and graphene; the modification process is selected from any one or more of surface oxidation, surface grafting, electropolymerization, and electrostatic adsorption.
[0016] In some specific embodiments, in step S3, the enzyme loading in the enzyme solution is 3 to 10 mg / mL; the loading process is selected from any one or more of the adsorption method, chemical cross-linking method, and encapsulation method.
[0017] In some specific embodiments, in step S3, the oxidase is selected from any one of glucose oxidase, lactate oxidase, uricate oxidase, and bilirubin oxidase.
[0018] The peroxidase is selected from horseradish peroxidase or catalase;
[0019] The dehydrogenase is selected from glucose dehydrogenase or lactate dehydrogenase.
[0020] In some specific embodiments, in step S4, the acrylate monomer is selected from either sodium acrylate or potassium acrylate, the acrylate monomer is selected from either acrylic acid or acrylamide, and the added mass of the monomer is 30%-60% of the total mass of the gel precursor solution.
[0021] The electrolyte is selected from any one or more of sodium hydroxide, sodium oxalate, sodium chloride, and sodium sulfate, and the added mass of the electrolyte is 1% to 4% of the total mass of the gel precursor solution.
[0022] The crosslinking agent is selected from polyethylene glycol (diol) diacrylate or methylene bisacrylamide, and the added mass of the crosslinking agent is 0.4% to 1% of the total mass of the monomers;
[0023] The initiator is selected from any one or more of acetylacetone, N-hydroxysuccinimide, and glucose, and the added mass of the initiator is 0.5% to 2% of the total mass of the monomers.
[0024] The solvent is selected from any one or more of water, glycerol, and ethylene glycol, and the amount of solvent added is to make up to 100% of the total mass of the gel precursor solution.
[0025] In some specific embodiments, in step S4, the stirring temperature is 25-30°C and the stirring time is 1-5 minutes.
[0026] In some specific embodiments, in step S5, the curing temperature is 32-40°C and the curing time is 5-15 minutes.
[0027] The second technical solution of the present invention is to provide a composite flexible bioelectrode, which is based on the preparation method described in one of the above technical solutions.
[0028] The third technical solution of the present invention is to provide an application of the composite flexible bioelectrode as described in the second technical solution above, wherein the composite flexible bioelectrode is used as a raw material to prepare physiological sensors and / or biofuel cells.
[0029] In step S1 of this invention, processing inorganic materials with a porous and loose morphology and a large specific surface area to form flexible fibrous electrode materials is intended to improve the efficiency of electron conduction pathways. In step S2, the enzyme loading is directly proportional to the subsequent monomer polymerization rate; a higher enzyme loading results in a faster reaction rate, but also shorter polymer chains, affecting hardness and toughness. Stirring in step S4 is to ensure rapid dissolution of monomers, crosslinking agents, electrolytes, initiators, and solvents. The amount of monomer added to the gel precursor solution affects the performance of the final hydrogel coating, especially its mechanical properties. Too low a monomer or crosslinking agent content reduces the mechanical strength of the hydrogel coating and may even lead to gelation failure; too high a monomer or crosslinking agent content results in an overly dense gel network, hindering mass diffusion and catalytic effects. The electrolyte in the gel precursor solution is used to control and regulate the ion channels of the hydrogel; a higher electrolyte content results in higher ion migration efficiency, but also a decrease in mechanical strength.
[0030] This invention involves the reaction of the components in the gel precursor solution with the enzyme on the surface of the enzyme-carrying electrode. The free radicals generated by the redox reaction immediately initiate the formation of a special polymer chain conformation and a three-dimensional network structure by the monomers, thereby improving the material exchange capacity and making the enzyme catalytic efficiency at the interface higher, resulting in a hydrogel coating with excellent mechanical properties and reusability.
[0031] Compared with the prior art, the present invention has the following beneficial effects:
[0032] (1) The method of this invention is simple, highly controllable, and easy to operate. The resulting composite flexible bioelectrode has good inorganic-organic interfacial adhesion, thus enabling efficient and direct conversion of biochemical energy into electrical energy. Unlike chemical modification, this invention uses a mild enzymatic polymerization method. During monomer polymerization, the protein conformation and activity of the enzyme are not affected. Furthermore, the porous structure of the hydrogel provides nanoscale confinement space for the reaction between the enzyme and the substrate, thereby improving the catalytic activity and efficiency of the substance. This results in a composite flexible bioelectrode with high sensitivity and accuracy in signal reading, as well as a long service life of more than 30 days, several times that of a naked enzyme electrode. Due to the good plasticity of the hydrogel, the composite flexible bioelectrode of this invention can be flexibly adjusted in shape and size according to different environmental and system usage requirements without affecting its performance, and can be widely used in electrochemical sensors or biofuel cells.
[0033] (2) Compared with traditional metal electrodes, the hydrogel-encapsulated composite flexible bioelectrode prepared in this invention has higher biocompatibility, greater enzyme loading capacity and more stable and mild enzyme immobilization performance due to the special properties of its structure. It is more sensitive in reading biological signals and longer-lasting and safer in use in the in vivo environment.
[0034] (3) Compared with traditional inorganic electrodes, the hydrogel-encapsulated composite flexible bioelectrode prepared in this invention has higher mechanical strength and can achieve specific identification of target biomarkers, while also meeting the requirements for simultaneous reading and transmission of multiple signals.
[0035] (4) Compared with traditional polymer-inorganic composite electrodes, the hydrogel-encapsulated composite flexible bioelectrode prepared in this invention has more adjustable pore structures, which can achieve efficient exchange of biological fluids and achieve higher catalytic efficiency. In addition, by utilizing the interfacial catalytic ability of enzymes, the material can be prepared in an integrated manner, meeting the stability and reliability requirements during use.
[0036] (5) The hydrogel-encapsulated composite flexible bioelectrode prepared by the present invention has a cytoplasmic matrix-like structure in its hydrogel layer, which enables the flexible bioelectrode to have high environmental tolerance and is suitable for long-term stable and repeated use; while the performance and swelling stability of the electrolyte enable the composite flexible bioelectrode to meet the requirements of efficient ion-electron conduction in the process of signal transduction and energy harvesting.
[0037] (6) The hydrogel-encapsulated composite flexible bioelectrode prepared by this invention has wide applications. The composite flexible bioelectrode can be integrated into an array to achieve simultaneous monitoring of multiple biomarkers, while avoiding interference from other substances in body fluids. The presence of the hydrogel coating on the composite flexible bioelectrode also ensures zero leakage of enzymes during use. Attached Figure Description
[0038] Figure 1 This is a photograph of the hydrogel-encapsulated composite flexible bioelectrode from Example 1.
[0039] Figure 2 This is a scanning electron microscope image of the hydrogel-encapsulated composite flexible bioelectrode in Example 1.
[0040] Figure 3 This is a comparison diagram of the mechanical properties of the hydrogel coating on the hydrogel-encapsulated composite flexible bioelectrode in Example 1;
[0041] Figure 4 These are test diagrams of the interfacial adhesion stability of the composite flexible bioelectrodes in Example 1, Comparative Example 1, and Comparative Example 2.
[0042] Figure 5 These are biocompatibility test diagrams of the composite flexible bioelectrodes in Example 1 and Comparative Example 4;
[0043] Figure 6 This is a diagram showing the specificity of glucose signal reading by the hydrogel-encapsulated composite flexible bioelectrode in Example 1.
[0044] Figure 7 This demonstrates the resistance of the hydrogel-encapsulated composite flexible bioelectrode in Example 1 to interference from non-glucose signals.
[0045] Figure 8 This describes the continuous monitoring of glucose signals by the composite flexible bioelectrode in Example 1 and Comparative Example 3.
[0046] Figure 9 The stability of the hydrogel-encapsulated composite flexible bioelectrode in Example 1 during 30 days of storage. Detailed Implementation
[0047] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments. These embodiments are based on the technical solution of the present invention and provide detailed implementation methods and specific operating procedures. However, the scope of protection of the present invention is not limited to the following embodiments.
[0048] Unless otherwise specified, the raw materials or processing techniques used in the following embodiments and comparative examples are all conventional commercially available raw materials or conventional processing techniques in the art.
[0049] Example 1:
[0050] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0051] The resulting composite flexible bioelectrode has a compressive modulus of 3424 MPa, can specifically recognize glucose, and has a detection limit of 0.02 mM.
[0052] like Figure 1 As shown, the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 has a good uniform morphology.
[0053] like Figure 2 The image shown is a scanning electron microscope image of the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1. It shows that the hydrogel has obvious and relatively uniform pore structure with a pore size of 20-30 μm; and the hydrogel is tightly bonded to the inorganic material.
[0054] like Figure 3 The figure shows a comparison of the mechanical properties of the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 before and after hydrogel coating and before and after freeze-drying, reflecting the excellent mechanical properties of the hydrogel-encapsulated composite flexible bioelectrode prepared after freeze-drying.
[0055] like Figure 6 As shown, the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 exhibits a corresponding and shifted redox peak for the biomarker (glucose as an example), demonstrating specific recognition. Furthermore, it displays different peak intensities for different concentrations of the biomarker (glucose as an example), thus allowing for differentiation based on varying signal intensities.
[0056] like Figure 7As shown, for different concentrations of biomarkers (glucose as an example), the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 is resistant to interference from non-glucose biomarkers and does not produce corresponding redox peaks.
[0057] like Figure 9 As shown in the figure, the stability of the composite flexible bioelectrode after 30 days of storage is tested. It can be seen from the figure that the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 has high stability under long-term storage.
[0058] Example 2:
[0059] Twisted carbon fiber bundles, 7 cm in length and 1 mm in diameter, were washed sequentially with ethanol and distilled water and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C and baked in an oven at 80 °C for 8 h, resulting in modification via electrostatic adsorption. At 25 °C, 15 mg of lactate oxidase (LAx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0060] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 3424 MPa, can specifically recognize lactic acid, and has a detection limit of 0.05 mM.
[0061] Example 3:
[0062] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of uricase (UAx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an inorganic material with the enzyme loaded on its surface through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0063] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 3424 MPa, can specifically identify uric acid, and has a detection limit of 0.01 mM.
[0064] Example 4:
[0065] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.46 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0066] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.08 mM.
[0067] Example 5:
[0068] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 0.8 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0069] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.1 mM.
[0070] Example 6:
[0071] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.46 g of sodium hydroxide were added to 0.8 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0072] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.14 mM.
[0073] Example 7:
[0074] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 0.5 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0075] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.2 mM.
[0076] Example 8:
[0077] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.46 g of sodium hydroxide were added to 0.5 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0078] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.28 mM.
[0079] Example 9:
[0080] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 10 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C, and baked in an oven at 80 °C for 8 h, thus modifying the material via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an enzyme-loaded inorganic material through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 37 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0081] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.01 mM.
[0082] Example 10:
[0083] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 15 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly dropped onto the dried carbon fiber bundles at 25 °C and baked in an oven at 80 °C for 8 h, resulting in modification via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h, resulting in an inorganic material with the enzyme loaded on its surface through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent and homogeneous solution. Then, 0.005 g of polyethylene glycol (diol) diacrylate, 0.0025 g of glucose solution (10 mM), and 0.0025 g of acetylacetone (AcAc) were added and stirred at room temperature for 5 min to obtain a transparent and homogeneous precursor solution. The precursor solution was uniformly coated onto the surface of the enzyme-loaded inorganic material, and after standing at 35 °C for 2 min, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0084] The resulting hydrogel-encapsulated composite flexible bioelectrode has a compressive modulus of 4814 MPa, can specifically recognize glucose, and has a detection limit of 0.007 mM.
[0085] Comparative Example 1:
[0086] Compared with Example 1, most of the steps are the same, except that 0.005g of polyethylene glycol (diol) diacrylate is replaced with 10mg of polyethylene glycol (diol) diacrylate, and 0.0025g of glucose solution (10mM) and 0.0025g of acetylacetone are replaced with 0.05mg of 2,2-diethoxyphenethylcopper. The polymerization is then initiated by UV light. The specific steps are as follows:
[0087] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C and baked in an oven at 80 °C for 8 h, resulting in modification via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a yellow homogeneous solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h to obtain an inorganic material with the enzyme loaded on its surface through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent homogeneous solution. Then, 10 mg of polyethylene glycol (diol) diacrylate and 0.05 mg of 2,2-diethoxyacetophenone were added and stirred at room temperature for 5 min to obtain a transparent homogeneous precursor solution. The precursor solution was uniformly coated on the surface of the enzyme-loaded inorganic material, irradiated with ultraviolet light for 15 minutes to form a gel, and then freeze-dried under vacuum for 3 days to obtain a hydrogel-encapsulated composite flexible bioelectrode.
[0088] Comparative Example 2:
[0089] Compared with Example 1, most of the steps are the same, except that 0.005g of polyethylene glycol (diol) diacrylate is replaced with 10mg of polyethylene glycol (diol) diacrylate, and 0.0025g of glucose solution (10mM) and 0.0025g of acetylacetone are replaced with 0.05mg of potassium peroxide. The polymerization is initiated by heat at 60°C to form a gel. The specific steps are as follows:
[0090] Twisted carbon fiber bundles with a length of 7 cm and a diameter of 1 mm were washed sequentially with ethanol and distilled water, and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C and baked in an oven at 80 °C for 8 h, resulting in modification via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a yellow homogeneous solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h to obtain an inorganic material with the enzyme loaded on its surface through encapsulation. 0.5 mL of acrylic acid and 0.23 g of sodium hydroxide were added to 1 mL of distilled water and stirred at 0 °C for 5 min to form a transparent homogeneous solution. Then, 10 mg of polyethylene glycol (diol) diacrylate and 0.05 mg of potassium persulfate were added and stirred at room temperature for 5 min to obtain a transparent homogeneous precursor solution. The precursor solution was uniformly coated on the surface of the enzyme-loaded inorganic material, and after standing at 60°C for 15 minutes, it formed a gel. After vacuum freeze-drying for 3 days, a hydrogel-encapsulated composite flexible bioelectrode was obtained.
[0091] Comparative Example 3:
[0092] The provided electrode is a bare electrode, which is largely the same as that in Example 1, except that it is not coated with hydrogel. Its preparation method includes the following steps:
[0093] Twisted carbon fiber bundles, 7 cm in length and 1 mm in diameter, were washed sequentially with ethanol and distilled water and dried in an oven at 60 °C for 6 h. 5 mg of multi-walled nanotubes were added to an aqueous solution and ultrasonically dispersed for 5 min to form a homogeneous dispersion. This dispersion was then uniformly added dropwise to the dried carbon fiber bundles at 25 °C and baked in an oven at 80 °C for 8 h, resulting in modification via electrostatic adsorption. At 25 °C, 15 mg of glucose oxidase (GOx) was added to 5 mL of distilled water and stirred for 5 min to form a homogeneous yellow solution. The baked carbon fiber bundles were then immersed in the enzyme solution and stirred at 4 °C for 12 h. This process resulted in the encapsulation of the enzyme-loaded inorganic material, which became the bare electrode.
[0094] Comparative Example 4:
[0095] The two methods are largely the same as in Example 1, except that sodium hydroxide, an electrolyte, is not added to the gel precursor solution.
[0096] like Figure 4 The figure shows the interfacial adhesion stability test diagrams of the composite flexible bioelectrodes prepared in Example 1, Comparative Example 1, and Comparative Example 2. Figure 4 A is a diagram of the testing device. The results show that the hydrogel layer and the inorganic material layer of the composite flexible bioelectrode wrapped in hydrogel prepared in Example 1 are more stably bonded.
[0097] like Figure 8 As shown, compared with the bare electrode prepared in Comparative Example 3, the hydrogel-encapsulated composite flexible bioelectrode prepared in Example 1 can perform long-term continuous monitoring and signal transmission for biomarkers (glucose as an example).
[0098] like Figure 5 As shown, the biocompatibility test diagrams of the composite flexible bioelectrodes prepared in Example 1 (right side) and Comparative Example 4 (left side) are shown. After adding 0.5%, 0.1%, and 0.05% of the mass of cell culture medium to the composite flexible bioelectrodes prepared in Example 1, the cell viability at 12h, 24h, and 48h was greater than 100%.
[0099] The above description of the embodiments is provided to enable those skilled in the art to understand and use the invention. It will be apparent to those skilled in the art that various modifications can be made to these embodiments, and the general principles described herein can be applied to other embodiments without inventive effort. Therefore, the present invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the invention should be within the protection scope of the present invention.
Claims
1. A method for preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel, characterized in that, Includes the following steps: S1. Processing inorganic materials into flexible fibrous electrode materials; S2. Modify the flexible fibrous electrode material obtained in step S1 with conductive nanomaterials to obtain the electrode substrate; S3. Load the enzyme onto the electrode substrate obtained in step S2 to obtain an enzyme-loaded electrode, wherein the enzyme is selected from any one of oxidase, peroxidase, and dehydrogenase. S4. Mix the monomer, crosslinking agent, electrolyte, initiator and solvent, and stir until completely dissolved into a transparent and homogeneous solution to obtain a gel precursor solution, wherein the monomer is selected from acrylate monomers or acrylic monomers; S5. Coat the gel precursor solution obtained in step S4 onto the surface of the enzyme-carrying electrode obtained in step S3, solidify it into a gel, and then freeze-dry it under vacuum to obtain a hydrogel-encapsulated composite flexible bioelectrode. In step S1, the inorganic material is selected from any one of carbon fiber, carbon nanotube, polyacetylene fiber, stainless steel fiber, and copper fiber; the processing technology is selected from any one or more of spinning, twisting, weaving, and winding. In step S2, the conductive nanomaterial is selected from any one or more of metal nanoparticles, single-walled carbon nanotubes, multi-walled carbon nanotubes, carbon nanorods, and graphene; the modification process is selected from any one or more of surface oxidation, surface grafting, electropolymerization, and electrostatic adsorption. In step S4, the acrylate monomer is selected from sodium acrylate and potassium acrylate, and the acrylic monomer is selected from acrylic acid and acrylamide. The added mass of the monomer is 30%-60% of the total mass of the gel precursor solution. The electrolyte is selected from any one or more of sodium hydroxide, sodium oxalate, sodium chloride, and sodium sulfate, and the added mass of the electrolyte is 1% to 4% of the total mass of the gel precursor solution. The crosslinking agent is selected from polyethylene glycol (diol) diacrylate or methylene bisacrylamide, and the mass of the crosslinking agent added is 0.4% to 1% of the total mass of the monomers; The initiator is selected from any one or more of acetylacetone, N-hydroxysuccinimide, and glucose, and the added mass of the initiator is 0.5% to 2% of the total mass of the monomers. The solvent is selected from any one or more of water, glycerol, and ethylene glycol, and the amount of solvent added is to make up to 100% of the total mass of the gel precursor solution.
2. The method for preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel according to claim 1, characterized in that, In step S3, the enzyme loading in the enzyme solution is 3~10 mg / mL; the loading process is selected from any one or more of the adsorption method, chemical cross-linking method, and encapsulation method.
3. The method for preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel according to claim 1, characterized in that, In step S3, the oxidase is selected from any one of glucose oxidase, lactate oxidase, uricate oxidase, and bilirubin oxidase; The peroxidase is selected from horseradish peroxidase or catalase; The dehydrogenase is selected from glucose dehydrogenase or lactate dehydrogenase.
4. The method for preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel according to claim 1, characterized in that, In step S4, the stirring temperature is 25~30℃ and the stirring time is 1~5 min.
5. The method for preparing a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel according to claim 1, characterized in that, In step S5, the curing temperature is 32~40℃ and the curing time is 5~15 min.
6. A composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel, characterized in that, Based on the preparation method according to any one of claims 1-5.
7. An application of a composite flexible bioelectrode encapsulated in an interfacial enzyme-catalyzed hydrogel as described in claim 6, characterized in that, The composite flexible bioelectrode is used as a raw material to prepare physiological sensors and / or biofuel cells.