Device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing

Abstract

The invention belongs to the technical field of biomedical instruments, and particularly relates to a device for detecting the glucose concentration of tear based on non-enzymatic colorimetric sensing, wireless and non-invasive detection of the glucose concentration in the tear is realized by introducing a paper-based sensing unit into a corneal contact lens, and in the paper-based sensing unit, the glucose concentration in the tear is detected by the paper-based sensing unit. Glucose is catalyzed by glucose oxidase to generate gluconic acid, and meanwhile hydrogen peroxide is released; the hydrogen oxide and TMB are subjected to oxidation reaction under the physiological pH condition under the catalytic action of the ferroferric oxide / graphene oxide enzyme-free compound to form a conjugated structure and present blue, so that the RGB color of the paper-based sensing unit is changed. Due to the use of the ferriferrous oxide / graphene oxide enzyme-free compound, the problem that the traditional enzyme catalysis is sensitive to the environment when being used is solved, so that the colorimetric sensing has quick reaction and stability at the same time. Compared with the prior art, better biocompatibility and stability are obtained, the structure is simple, and large-scale production is easy.

Description

Technical Field

[0001] This invention belongs to the field of biomedical device technology, specifically relating to a device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing, which is used for wireless and non-invasive bio-detection of corneal contact lenses. Background Art

[0002] In the field of wearable device technology, flexible wearable physical sensing electronics have developed rapidly in recent years. These devices, integrated into wearable products, enable real-time tracking of physiological signals from areas such as the wrist, chest, and eyes. For example, flexible contact lenses with built-in microsensors, as a new generation of wearable devices, have demonstrated significant application value in medical diagnosis and treatment. Contact lenses integrating capacitive coils can monitor intraocular pressure changes in glaucoma patients in real time, while contact lenses integrating drug delivery units can treat eye diseases by precisely controlling drug release. This wireless and non-invasive detection and treatment method provides a new solution for the diagnosis and treatment of eye diseases.

[0003] However, traditional electrochemical detection methods in the field of blood glucose monitoring technology still have many shortcomings. Currently, this method requires an electrochemical workstation to analyze and process the collected samples, which has the following limitations: First, it is easily affected by interfering substances in the blood, leading to inaccurate test results; second, it requires collecting a large amount of blood and demands a high level of professional expertise, causing discomfort to users and affecting the user experience; third, it is highly dependent on instruments, with poor device portability, making it difficult to meet the needs of real-time blood glucose monitoring; in addition, some patients may experience skin irritation or allergic reactions. Therefore, there is an urgent need to develop a more accurate, convenient, non-invasive blood glucose monitoring method suitable for daily monitoring. Summary of the Invention

[0004] The purpose of this invention is to provide a device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing, so as to solve the problems existing in the above-mentioned blood glucose monitoring.

[0005] To achieve the above objectives, this experiment adopts the following technical solution:

[0006] A device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing includes a first silicone hydrogel encapsulation layer, a second silicone hydrogel encapsulation layer, a paper-based sensing unit, and a colorimetric analysis unit.

[0007] Both the first and second silicone hydrogel encapsulation layers are made by modifying the hydrogel by introducing hydrophilic monomers and silicone monomers.

[0008] The paper-based sensing unit is disposed between the first and second silicone hydrogel encapsulation layers, and together with the first and second silicone hydrogel encapsulation layers, constitutes a corneal contact lens; it is used to change the RGB color of the paper-based substrate under physiological pH conditions, including the paper-based substrate, a colorimetric sensing unit attached to the paper-based substrate, and the colorimetric sensing unit is prepared by TMB colorimetric object, glucose oxidase GOD, and Fe3O4 / GO NPs, an enzyme-free complex of iron oxide / graphene oxide, in a mass ratio of 8:1:1.

[0009] The colorimetric analysis unit is used to capture the RGB color change values ​​of the paper substrate. The colorimetric method is used to compare the RGB color change values ​​with the actual values ​​to detect the glucose concentration in the tear duct.

[0010] Furthermore, the actual value was obtained by experimentally correlating the RGB color channel values ​​with glucose concentration.

[0011] Furthermore, the enzyme-free Fe3O4 / graphene oxide composite is prepared by mixing Fe3O4 nanoparticles and graphene oxide powder at a mass ratio of 1:1 in a buffer solution with a pH of 7; and using ultrasonic-assisted dispersion to ensure sufficient diffusion and binding of graphene oxide and Fe3O4. The Fe3O4 particles are attached to the graphene oxide, and the graphene oxide (GO) provides a high specific surface area, enhances the dispersibility and electron transfer capacity of Fe3O4, improves catalytic efficiency, thereby improving the reaction of glucose molecules on the mixture and enhancing the performance of the enzyme-free sensor.

[0012] Furthermore, the iron oxide nanoparticles are synthesized from FeCl2·4H2O, FeCl3·6H2O, and NH3·H2O; wherein the mass ratio of FeCl2·4H2O to FeCl3·6H2O is 1:2.

[0013] Furthermore, the paper substrate is a patterned paper substrate prepared using laser cutting technology. Laser cutting technology allows the paper substrate to be flexibly cut into any desired shape, thus adapting to the needs of different working environments and occasions.

[0014] Furthermore, the preparation process of the silicone hydrogel encapsulation layer is as follows:

[0015] NVP, KH-570, and HEMA were mixed in a mass ratio of 35%:15%:50%, and a thermal initiator and binder were added. The mixture was then thoroughly mixed to obtain a silicone hydrogel solution. The hydrophilic monomers were provided by hydroxyethyl methacrylate (HEMA) and N-vinylpyrrolidone (NVP), and the silicone monomers were provided by γ-propyltrimethoxysilane (KH-570).

[0016] Furthermore, the thermal initiator is azobisisobutyronitrile (AIBN), and the binder is N,N-methylenebisacrylamide (NMBA); the ratio of the thermal initiator (AIBN) to the crosslinking agent (NMBA) is 0.7%:0.3%.

[0017] This invention achieves wireless and non-invasive detection of glucose concentration in tears by introducing a paper-based sensing unit into a contact lens. In the paper-based sensing unit, glucose is catalyzed by glucose oxidase to produce gluconic acid, simultaneously releasing hydrogen peroxide. Under physiological pH conditions, the hydrogen peroxide reacts with TMB (tumor monoxide) under the catalysis of an enzyme-free iron(III) oxide / graphene oxide complex, forming a conjugated structure that exhibits a blue color, thus causing a change in the RGB color of the paper-based sensing unit. The use of the iron(III) oxide / graphene oxide enzyme-free complex solves the environmental sensitivity of traditional enzyme-catalyzed methods, enabling colorimetric sensing to achieve both rapid response and stability. In the encapsulation section, a modifier prepared from hydrophilic and silicon monomers is introduced to modify the silicone hydrogel encapsulation layer. This modifies the material's hydrophilicity and water permeability while maintaining oxygen permeability, ensuring that glucose molecules can pass through the silicone hydrogel encapsulation layer and the paper-based sensing unit. The corneal contact lens of the present invention overcomes the drawback of traditional PDMS as a hydrophobic encapsulation layer by introducing hydrophilic monomers to improve the hydrophilicity of the material and by introducing silicon monomers to improve the oxygen permeability and gas permeability of the material.

[0018] Compared with existing technologies, the device of the present invention uses general-purpose materials, which achieves better biocompatibility and stability, and has a simple structure, simple design, and is easy to mass-produce. Attached Figure Description

[0019] Figure 1 This is a schematic diagram of the device structure for detecting tear glucose concentration based on non-enzymatic colorimetric sensing in Example 1.

[0020] Figure 2 The preparation process of the silicone hydrogel encapsulation layer in Example 1 is as follows;

[0021] Figure 3 This is a flowchart illustrating the specific preparation process of the corneal contact lens in Example 1;

[0022] Figure 4 The ultraviolet absorption spectrum provided for the apparatus of Example 1 in Experiment 1 illustrates the principle and results of the colorimetric reaction in the experimental examples.

[0023] Figure 5 The figure shows the colorimetric reaction results of the device in Example 1 in Experiment 2. This figure proves that the device can identify glucose solutions of different concentrations, demonstrating the feasibility of the device in Example 1 in distinguishing tear glucose concentration. DETAILED DESCRIPTION

[0024] The technical solution of the present invention will be described in detail below with reference to the accompanying drawings and embodiments.

[0025] Example 1: As Figure 1 As shown, this embodiment provides a device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing, including a first silicone hydrogel encapsulation layer, a second silicone hydrogel encapsulation layer, a paper-based sensing unit, and a colorimetric analysis unit.

[0026] Both the first and second silicone hydrogel encapsulation layers are made by modifying the hydrogel by introducing hydrophilic monomers and silicone monomers.

[0027] The paper-based sensing unit is disposed between the first and second silicone hydrogel encapsulation layers, and together with the first and second silicone hydrogel encapsulation layers, constitutes a corneal contact lens; it is used to change the RGB color of the paper-based substrate under physiological pH conditions. The paper-based sensing unit includes a paper-based substrate and a colorimetric sensing unit attached to the paper-based substrate. The paper-based substrate is laser-cut into the required shape on a piece of paper. The colorimetric sensing unit is prepared from TMB colorimetric material, glucose oxidase (GOD), and an enzyme-free complex of iron(III) oxide / graphene oxide (Fe3O4 / GO NPs) in a mass ratio of 8:1:1.

[0028] The colorimetric analysis unit is implemented through analysis software installed on a user terminal, such as a smartphone. It captures the RGB color change values ​​of a paper-based substrate and compares these values ​​with actual values ​​using a colorimetric method, enabling wireless, non-invasive, real-time, and convenient detection of glucose concentration in tear ducts. The advantage of this embodiment using a colorimetric method is that the color change of glucose in the sample after reacting with a specific reagent can be identified by the naked eye or simple optical equipment. The actual values ​​shown are obtained by experimentally correlating RGB color channel values ​​with glucose concentration.

[0029] The preparation method of the above-mentioned device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing is as follows:

[0030] S1. Prepare silica hydrogel solution and iron(III) oxide / graphene oxide enzyme-free complex respectively:

[0031] The preparation method of silica hydrogel solution is as follows: Figure 2As shown, hydrophilic monomers were prepared by uniformly mixing hydroxyethyl methacrylate (HEMA) and N-vinylpyrrolidone (NVP) at a mass ratio of 50%:35%. Then, 15% γ-propyltrimethoxysilane (KH-570) was added to provide silica monomers for the hydrogel. After stirring evenly, a thermal initiator was added, followed by a binder. The mass ratio of thermal initiator (AIBN) to crosslinking agent (NMBA) was 0.7%:0.3%. The mixture was then magnetically stirred for 30 minutes to obtain the final product. In this embodiment, the thermal initiator added was azobisisobutyronitrile (AIBN), and the binder was N,-methylene N-bisacrylamide (NMBA).

[0032] The preparation method of the iron oxide / graphene oxide enzyme-free complex is as follows:

[0033] 0.1 mol FeCl2·4H2O and 0.2 mol FeCl3·6H2O were uniformly dissolved in 1.2 L of deionized water, and 0.6 L of NH3·H2O was added while stirring until the suspension turned black. Then, the mixture was stirred continuously in a 50 °C water bath for 30 minutes, and the precipitated powder was collected after cooling. Finally, the powder was washed several times with deionized water to obtain iron oxide nanoparticles.

[0034] The graphene oxide and iron oxide were mixed in a buffer solution with a pH of 7 at a mass ratio of 1:1, and ultrasonic-assisted dispersion was used to ensure that the graphene oxide and iron oxide were fully diffused and combined to obtain an enzyme-free iron oxide / graphene oxide complex.

[0035] S2. Add 10 mL of 10 mg / mL glucose oxidase, 10 mL of 20 mg / mL Fe3O4 / GO NPs (10 mg Fe3O4 and 10 mg graphene oxide were added to 1 mL of pH=7 buffer solution and dispersed by sonication) and 80 mL of TMB mixture to a brown light-proof bottle. Add 2 mL of the mixture to a paper substrate and then dry it in an oven at 37 °C to obtain a paper-based sensing unit.

[0036] S3.1 Preparation of corneal contact lenses, the preparation process is as follows: Figure 3 As shown, the following steps are included:

[0037] The paper-based sensing unit made of S2 was cut into the desired shape using a laser cutting instrument. This shape is a polygonal ring structure with a ring width of 2mm and an inner ring radius of 5mm. Compared with a circular ring structure, the polygonal ring structure can increase and improve the flexibility of the corneal contact lens. Setting the inner ring radius to 5mm can reduce light obstruction, thereby reducing the impact of the corneal contact lens on vision.

[0038] S3.2. Take 1 ml of the prepared silicone hydrogel solution and drop it onto the center of the glass slide. Then, use a spin coater to evenly spin coat the silicone hydrogel solution onto the glass slide. The spin coater speed is 800 rpm and the spin coat time is 30 s. Place the spin-coated glass slide into an oven at 75-80 degrees Celsius and dry it for 2 hours to obtain a cured silicone hydrogel film, which is the first silicone hydrogel encapsulation layer.

[0039] S3.3 Place the product obtained in S3.1 on the cured first silicone hydrogel film, then drop 1 ml of silicone hydrogel solution onto the surface of the product obtained in S3.1, and coat it again using a spin coater at a speed of 800 rpm for a spin coating time of 45 s. Then place it in an oven to dry for 2 hours to form a second silicone hydrogel encapsulation layer. Finally, mechanically peel the film off the glass slide to obtain the silicone hydrogel encapsulated colorimetric unit film.

[0040] S3.4. Add 0.2 ml of silicone hydrogel to the corneal contact lens mold, and then place the silicone hydrogel film containing the colorimetric unit as described in 3.3 at the center of the corneal contact lens mold. Apply mechanical pressure to the structure obtained in S3.3 through the corneal contact lens mold to compress it and ensure that each layer is tightly bonded. Place the compressed sample in an oven for drying and shaping. After drying and shaping, demold the corneal contact lens to obtain a corneal contact lens with a built-in colorimetric sensing unit.

[0041] To better illustrate the effectiveness of the device in this embodiment, experiments 1 and 2 are conducted below. Experiment 1 is used to verify the principle of color development by the colorimetric unit in this embodiment, and experiment 2 is used to verify the feasibility of the device provided in embodiment 1 for glucose concentration detection.

[0042] Experiment 1: This experiment verifies the catalytic effect of Fe3O4 / GO NPs and explains the principle of the colorimetric reaction. Colorimetry, a classic method for glucose detection, relies on specific chemical reactions that cause color changes in the solution. A common method is the glucose oxidase-peroxidase colorimetric method, where glucose oxidase oxidizes glucose, producing a colored compound proportional to the glucose concentration, which is then analyzed colorimetrically to determine the glucose content. In this experiment, using TMB as the chromogenic substrate, glucose oxidase specifically and efficiently catalyzes the oxidation of glucose molecules to gluconic acid, simultaneously generating hydrogen peroxide. At physiological pH, TMB reacts with H2O2, catalyzed by Fe3O4 / GO NPs. The amino group in TMB loses an electron to become a cationic radical, which exists in the system as a dimer. This dimer exhibits maximum absorption at 370 nm and 652 nm, appearing blue.

[0043] The catalytic activity of Fe3O4 / GO NPs in Example 1 was verified by ultraviolet absorption spectroscopy. The specific experimental procedure is as follows:

[0044] First, prepare three portions each of TMB solution, H2O2 solution, and Fe3O4 / GO NPs solution, 10 ml each. Then, thoroughly mix the different solutions and divide them into three groups: Group 1, a mixture of TMB solution and H2O2 solution (control group 1); Group 2, a mixture of TMB solution and Fe3O4 / GO NPs solution (control group 2); and Group 3, a mixture of TMB solution, H2O2 solution, and Fe3O4 / GO NPs solution (experimental group). The mixed solutions were thoroughly mixed at 37 degrees Celsius and incubated for 30 minutes to allow for complete reaction. Afterward, the mixed liquid was placed in a UV absorption instrument to detect its UV absorption spectrum. The results showed that... Figure 4 As shown, the experimental group exhibits strong absorption around 650 nm, which confirms the experimental principle that the copolymer has maximum absorption at 652 nm. The control group, however, did not show a significant absorption peak, indicating that Fe3O4 / GO NPs have a catalytic effect on TMB and H2O2, verifying the feasibility of the enzyme-free catalytic design. The blue colorimetric result confirms the feasibility of the device for glucose colorimetry.

[0045] Experiment 2 verified the resolution of the device in Example 1 by using glucose solutions of different concentrations. Furthermore, it used a smartphone and data processing analysis to fit the color of the device in Example 1 with the glucose concentration, visually linking color and concentration. The specific experimental design is as follows:

[0046] Different concentrations of glucose solutions were prepared to simulate the glucose concentration in tears (0.01 mM–6 mM). These solutions were then added dropwise to a prepared colorimetric sensing unit, and the color changes were observed after allowing the reaction to proceed at 37 degrees Celsius. Next, RGB pigment capture software on a smartphone was used to analyze the color changes of the colorimetric unit at different concentrations. Specifically, the phone's camera was used to capture images of the color changes, thus quantifying the changes in the three primary colors (red, green, and blue). Finally, data processing was used to correlate the color changes with the concentration of the analyte. The experimental results are as follows: Figure 5 As shown, when the glucose concentration ranges from 0.01 mM to 6 mM, a good linear relationship is observed between (R+B) / 2R and the glucose concentration: y = 0.257x + 1.11,R 2 =0.9869. This indicates that Experiment 2 can effectively distinguish the color change of the colorimetric unit, thus demonstrating that the device in Example 1 can effectively distinguish glucose solutions of different concentrations.

[0047] The above embodiments are merely illustrative of the principles and effects of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in the present invention should still be covered by the claims of the present invention.

Claims

1. A device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing, characterized in that, It includes a first silicone hydrogel encapsulation layer, a second silicone hydrogel encapsulation layer, a paper-based sensing unit, and a colorimetric analysis unit; Both the first and second silicone hydrogel encapsulation layers are made by modifying the hydrogel by introducing hydrophilic monomers and silicone monomers. The paper-based sensing unit is disposed between the first silicone hydrogel encapsulation layer and the second silicone hydrogel encapsulation layer, and together with the first silicone hydrogel encapsulation layer and the second silicone hydrogel encapsulation layer, it constitutes a corneal contact lens. It is used to change the RGB color of a paper substrate under physiological pH conditions. It includes a paper substrate and a colorimetric sensing unit attached to the paper substrate. The colorimetric sensing unit is prepared by TMB colorimetric object, glucose oxidase GOD and Fe3O4 / GO NPs, which are enzyme-free complexes of iron oxide / graphene oxide, in a mass ratio of 8:1:

1. The colorimetric analysis unit is used to capture the RGB color change values ​​of the paper substrate. The colorimetric method is used to compare the RGB color change values ​​with the actual values ​​to detect the glucose concentration in the tear duct.

2. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 1, characterized in that, The actual values ​​were obtained by experimentally correlating RGB color channel values ​​with glucose concentration.

3. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 1, characterized in that, The enzyme-free iron oxide / graphene oxide composite is prepared by mixing iron oxide nanoparticles and graphene oxide powder at a mass ratio of 1:1 in a buffer solution with a pH of 7, and then using ultrasonic-assisted dispersion to ensure that the graphene oxide and iron oxide are fully diffused and combined.

4. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 3, characterized in that, The iron tetroxide nanoparticles were synthesized from FeCl2·4H2O, FeCl3·6H2O, and NH3·H2O; wherein the mass ratio of FeCl2·4H2O to FeCl3·6H2O was 1:

2.

5. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 1, characterized in that, The paper substrate is a patterned paper substrate prepared using laser cutting technology.

6. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 1, characterized in that, The method for preparing the silicone hydrogel encapsulation layer is as follows: NVP, KH-570, and HEMA are mixed in a mass ratio of 35:15:50, and a thermal initiator and binder are added. After mixing, a silicone hydrogel solution is obtained. The hydrophilic monomers are provided by hydroxyethyl methacrylate and N-vinylpyrrolidone, and the silicone monomers are provided by γ-propyltrimethoxysilane.

7. The device for detecting tear glucose concentration based on non-enzymatic colorimetric sensing according to claim 5, characterized in that, The thermal initiator is azobisisobutyronitrile, and the binder is N,N-methylenebisacrylamide; the ratio of the thermal initiator to the crosslinking agent is 0.7%:0.3%.

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