Micro-needle patch for quantitatively detecting blood sugar concentration of human body as well as preparation method and application of micro-needle patch

Abstract

The invention discloses a microneedle patch for quantitatively detecting the blood sugar concentration of a human body as well as a preparation method and application of the microneedle patch, and belongs to the technical field of medicine detection. Mixing the PVA solution and the chitosan solution, adding the silk fibroin solution, and mixing to obtain a mixed solution; adding the mixed solution into a microneedle patch mold, enabling the mixed solution to enter the tip part of a microneedle, and performing vacuum filling on a cavity; adding a glucose oxidase solution and a potassium iodide solution into the mixed solution, mixing, continuously pouring the obtained mixed solution into a microneedle patch mold, filling a backing part at the upper end of a needle tip in the microneedle patch mold with the mixed solution, filling a cavity with the mixed solution in a vacuum manner, and repeatedly freezing, thawing and drying to obtain the microneedle patch for quantitatively detecting the blood glucose concentration of the human body. The microneedle patch for quantitatively detecting the blood sugar concentration of the human body can quantitatively detect the blood sugar of the human body, can visually reflect a detection result, is rapid and painless, low in cost, high in mechanical property and good in stability, and provides guiding significance for early diagnosis of diabetes mellitus.

Description

Technical Field

[0001] The present invention belongs to the technical field of medical detection, and in particular relates to a microneedle patch for quantitatively detecting blood sugar concentration in a human body, and a preparation method and application thereof. Background Art

[0002] Microneedle technology for component detection is a new type of detection technology. It is a technology that can detect the properties or content of specific components in a mixture by extracting them into a microarray. This technology uses a minimally invasive method. After breaking through the surface layer of food / skin, it uses the swelling properties of the matrix components of the microneedle tip to mix the detected components with the detection object in the mechanism, thereby realizing the detection of components. At present, this technology has been combined with a variety of technologies for detection, such as microfluidic chips. Modifying nanomaterials, enzymes, antibodies and other substances on microneedles can greatly improve sensitivity and selectivity. At present, the ability to qualitatively and quantitatively detect components is a major development direction of microneedle technology.

[0003] Usually, the detection of human blood sugar is based on electronic detection equipment with high pain, and the detection of in vitro glucose mostly uses high performance liquid chromatography to detect its content. CN117686488 A discloses a microneedle patch for colorimetric detection of glucose concentration and its preparation method, wherein the needle tip layer includes a high biocompatibility matrix, PVA-modified gold nanoparticles and a phosphate buffer salt providing a neutral environment, and the backing layer includes a high biocompatibility matrix, horseradish peroxidase, 3,3',5,5'-tetramethylbenzidine and an acetate buffer salt providing an acidic environment; it can be used for visual detection of glucose concentration. Compared with the detection method of the traditional blood glucose meter, this means of detecting glucose is simple, fast, intuitive and accurate, and cumbersome operation process is avoided. However, the microneedle detection patch needle tip prepared by this method has insufficient mechanical properties and poor stability during use, which reduces its application in detecting blood sugar.

[0004] Therefore, there is an urgent need for a simple, portable, fast and sensitive detection method for human blood glucose concentration. Summary of the invention

[0005] In view of the problems existing in the prior art, the present invention provides a microneedle patch for quantitatively detecting human blood glucose concentration and a preparation method and application thereof, which can extract glucose components from interstitial fluid and perform visual detection, perform qualitative and quantitative detection, have improved mechanical properties and good stability, and can be widely used in real-time detection of human blood glucose.

[0006] The present invention is achieved through the following technical solutions: A method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body comprises the following steps: (1) mixing a polyvinyl alcohol (PVA) solution and a chitosan solution, and adding the silk fibroin solution to obtain a mixed solution; (2) The mixed solution in step (1) is added to the microneedle patch mold, and the mixed solution enters the microneedle tip, and the cavity is filled with vacuum; (3) Glucose oxidase solution and potassium iodide solution are added to the mixed solution of step (1), and the mixture is evenly mixed. The obtained mixed solution is continuously poured into the microneedle patch mold of step (2), and the mixed solution is filled into the backing part at the upper end of the needle tip in the microneedle patch mold, and the cavity is filled with vacuum, and the mixture is repeatedly frozen and thawed, and dried to obtain a microneedle patch for quantitatively detecting blood glucose concentration in the human body.

[0007] Furthermore, in step (1), the mass percentage concentration of the PVA solution is 10-20%, the mass percentage concentration of the chitosan solution is 1.5-3%, the mass percentage concentration of the silk fibroin solution is 2-4wt%, the mass ratio of the PVA solution to the chitosan solution is 1:1, and the silk fibroin solution accounts for 8-15% of the mass of the mixed solution.

[0008] Furthermore, in step (3), the concentration of the glucose oxidase solution is 0.06-0.08 mg / mL, and the concentration of the potassium iodide solution is 0.6-0.8 mg / mL.

[0009] Furthermore, in step (3), the mass ratio of the glucose oxidase solution to the potassium iodide solution is 1:1-5; and the mass ratio of the total amount of the glucose oxidase solution and the potassium iodide solution to the PVA solution is 1:1-5.

[0010] Furthermore, in step (3), the mass ratio of the glucose oxidase solution to the potassium iodide solution is 1:3; and the mass ratio of the total amount of the glucose oxidase solution and the potassium iodide solution to the PVA solution is 1:3.

[0011] Furthermore, the number of freeze-thaw cycles in step (2) and step (3) is 4 to 6, with freezing at -20°C for 10 h and thawing at room temperature for 4 h. Furthermore, the drying condition in step (3) is drying at 60° C. for 6 hours.

[0012] Furthermore, the needle tip length of the microneedle patch for quantitatively detecting human blood glucose concentration is 600-800 μm, and the backing thickness is 200-400 μm.

[0013] In the present invention, the microneedle patch for quantitatively detecting the blood glucose concentration of the human body prepared by the method described above can detect the blood glucose by detecting the subcutaneous interstitial fluid because the glucose concentration in the interstitial fluid of the human body is similar to that of blood glucose.

[0014] In the present invention, the microneedle patch for quantitatively detecting blood sugar concentration in human body is used in detecting blood sugar.

[0015] In the present invention, the color development system is carried on the microneedle, the tip of the microneedle is immersed in a gradient glucose solution, the glucose solution is absorbed after the tip swells, and then the glucose reacts with the glucose oxidase in the microneedle lining to generate hydrogen peroxide; hydrogen peroxide can oxidize the elemental iodine produced by iodine ions, and then react with PVA to form a gradient red-brown color.

[0016] The beneficial effects achieved by the present invention are: The microneedle patch for quantitatively detecting the blood sugar concentration of the human body in the present invention is used to quantitatively detect the blood sugar of the human body, which can visually and intuitively reflect the test results, and judge the blood sugar concentration by the degree of color development. It is rapid, painless and low-cost. Moreover, by adding silk fibroin, the mechanical properties of the microneedle patch are improved, and the stability is good. It is not only easy to store, but also improves the ability to pierce the human skin, providing guidance for the early diagnosis and monitoring of diabetes. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The color development results of the microneedle patch for quantitative detection of human blood glucose concentration under different mass ratios (glucose oxidase solution and potassium iodide solution); Figure 2 The color development results of the microneedle patch for quantitative detection of human blood glucose concentration under different mass ratios (color development system and PVA solution); Figure 3 The color development result diagram and linear relationship diagram of the microneedle patch for quantitatively detecting human blood glucose concentration under different glucose concentrations; Figure 4 These are pictures of the needle tip portion of the microneedle patch for quantitatively detecting human blood glucose concentration prepared in Comparative Example 1, A is newly prepared, and B is after being placed at room temperature for 12 hours; Figure 5 These are pictures of the needle tip portion of the microneedle patch for quantitatively detecting human blood glucose concentration prepared in Example 1, A is newly prepared, and B is after being placed at room temperature for 12 hours. DETAILED DESCRIPTION

[0018] The following is a comprehensive and detailed description of the technical solutions involved in this patent. The examples described are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments without creative adjustments to the design ideas are within the scope of protection of the present invention. Example 1 (1) PVA was dissolved in distilled water at 90°C for 4 hours to obtain a 15 wt% PVA solution; chitosan was dissolved in 0.1 m acetic acid and stirred continuously at room temperature overnight to obtain a 2 wt% chitosan solution; the PVA solution and the chitosan solution were mixed in a solution mass ratio of 1:1, and after the mixture was evenly mixed, a silk fibroin solution (the concentration of the silk fibroin aqueous solution was 3 wt%) was added, and the solution was evenly mixed by ultrasonic stirring to obtain a mixed solution, wherein the amount of the silk fibroin solution added accounted for 10% of the total mass of the mixed solution; (2) The mixed solution in step (1) is added to the microneedle patch mold, and the mixed solution enters the microneedle tip, and the cavity is filled with vacuum. The length of the needle tip is 650 μm, and a PVA / CS hydrogel needle tip is obtained; (3) Add 0.08 mg / mL glucose oxidase solution and 0.6 mg / mL potassium iodide solution to the mixed solution of step (1), mix well, the mass ratio of glucose oxidase solution to potassium iodide solution is 1:3; the mass ratio of the total amount of glucose oxidase solution and potassium iodide solution to the PVA solution is 1:3, and the obtained mixed solution is continuously poured into the microneedle patch mold of step (2), and the mixed solution is filled into the backing part at the upper end of the PVA / CS hydrogel needle tip in the microneedle patch mold, and the cavity is vacuum filled, and the backing thickness is 300 μm. The above mold is placed in a -20°C freezer for 10 h, thawed at room temperature for 4 h, and the freeze-thaw cycle is repeated 4 times, and dried in a 60°C air drying oven for 6 h to obtain a microneedle patch for quantitative detection of human blood glucose concentration.

[0019] Example 2 The mass ratios of the glucose oxidase solution and the potassium iodide solution in Example 1 were changed to 1:1, 1:3, 1:5, 1:7 and 1:9, respectively. The other steps and operating conditions were the same as those in Example 1, and a series of microneedle patches for quantitatively detecting human blood glucose concentration were prepared.

[0020] The prepared microneedle patch for quantitative detection of human blood glucose concentration was placed in a 0.7 mg / mL glucose solution to detect the relationship between the gray value after color development and the mass ratio of the glucose oxidase solution to the potassium iodide solution. The results are as follows: Figure 1 As shown by Figure 1 It can be seen that the optimal solution mass ratio of glucose oxidase and potassium iodide is 1:3.

[0021] Example 3 The mass ratio of the color development system (the total amount of glucose oxidase solution and potassium iodide solution) to the PVA solution in Example 1 was changed to 1:1, 1:3, 1:5, 1:7, and 1:9, respectively. The other steps and operating conditions were the same as in Example 1, and a series of microneedle patches for quantitatively detecting human blood glucose concentration were prepared.

[0022] The prepared microneedle patch for quantitative detection of human blood glucose concentration was placed in a 0.7 mg / mL glucose solution to detect the relationship between the gray value after color development and the mass ratio of the color development system and the PVA solution. The results are as follows: Figure 2 As shown by Figure 2 It can be seen that the optimal mass ratio of the color development system (the total amount of glucose oxidase solution and potassium iodide solution) to the PVA solution is 1:3.

[0023] Comparative Example 1 In step (1) of comparative example 1, no silk fibroin was added to the mixed solution, and the remaining operations were the same as those of example 1 to prepare a microneedle patch for quantitatively detecting blood glucose concentration in the human body.

[0024] Property detection 1. The microneedle patch for quantitatively detecting human blood glucose concentration prepared in Example 1 was placed in a gradient glucose solution (0-16 mmol / L) for 10 min and then taken out. The color change and gradient value change were as follows: Figure 3 As shown by Figure 3 It can be seen that the color change and gray value change of the microneedle patch for quantitatively detecting the blood glucose concentration in the human body are in a linear relationship with the concentration of the glucose solution.

[0025] 2. The microneedle patch for quantitative detection of human blood glucose concentration prepared in Example 1 was used to detect the blood glucose of hypoglycemic, normal and hyperglycemic mice (8 mice with different blood glucose values), and the tails of the mice were cut to collect blood, which was tested with a blood glucose meter. The test data of the microneedle patch and the test data of the blood glucose meter are shown in Table 1 below.

[0026] Table 1 As can be seen from Table 1, the detection error between the microneedle patch and the blood glucose meter for quantitatively detecting human blood glucose concentration is within ±1, which is highly correlated.

[0027] 3. The picture of the needle tip of the microneedle patch for quantitatively detecting the blood glucose concentration in human body prepared in Comparative Example 1 of the present invention is as follows Figure 4 As shown, A is freshly prepared, and B is after being placed at room temperature for 12 hours; the picture of the needle tip part of the microneedle patch for quantitative detection of human blood glucose concentration prepared in Example 1 is shown Figure 5 As shown, A is freshly prepared, and B is after being placed at room temperature for 12 hours; Figure 4 and Figure 5 It can be seen that silk fibroin has good biocompatibility and can be used for human body detection. Adding silk fibroin to the microneedle can not only stabilize the needle tip matrix, but also improve the mechanical properties and make the needle tip more stable.

Claims

1. A method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body, characterized in that: The following steps are involved: (1) mixing a PVA solution and a chitosan solution, and adding the silk fibroin solution to obtain a mixed solution; (2) The mixed solution in step (1) is added to the microneedle patch mold, and the mixed solution enters the microneedle tip, and the cavity is filled with vacuum; (3) Glucose oxidase solution and potassium iodide solution are added to the mixed solution of step (1), and the mixture is evenly mixed. The obtained mixed solution is continuously poured into the microneedle patch mold of step (2), and the mixed solution is filled into the backing part at the upper end of the needle tip in the microneedle patch mold, and the cavity is filled with vacuum, and the mixture is repeatedly frozen and thawed, and dried to obtain a microneedle patch for quantitatively detecting blood glucose concentration in the human body.

2. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: In step (1), the mass percentage concentration of the PVA solution is 10-20%, the mass percentage concentration of the chitosan solution is 1.5-3%, the mass percentage concentration of the silk fibroin solution is 2-4wt%, the mass ratio of the PVA solution to the chitosan solution is 1:1, and the silk fibroin solution accounts for 8-15% of the mass of the mixed solution.

3. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: In step (3), the concentration of the glucose oxidase solution is 0.06-0.08 mg / mL, and the concentration of the potassium iodide solution is 0.6-0.8 mg / mL.

4. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: In step (3), the mass ratio of the glucose oxidase solution to the potassium iodide solution is 1:1-5; the mass ratio of the total amount of the glucose oxidase solution and the potassium iodide solution to the PVA solution is 1:1-5.

5. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 4, characterized in that: In step (3), the mass ratio of the glucose oxidase solution to the potassium iodide solution is 1:3; the mass ratio of the total amount of the glucose oxidase solution and the potassium iodide solution to the PVA solution is 1:

3.

6. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: The number of freeze-thaw cycles in step (2) and step (3) is 4 to 6, with freezing at -20°C for 10 h and thawing at room temperature for 4 h.

7. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: The drying condition in step (3) is drying at 60° C. for 6 h.

8. The method for preparing a microneedle patch for quantitatively detecting blood glucose concentration in a human body according to claim 1, characterized in that: The needle tip length of the microneedle patch for quantitatively detecting the blood sugar concentration of the human body is 600-800 μm, and the backing thickness is 200-400 μm.

9. A microneedle patch for quantitatively detecting blood glucose concentration in a human body prepared by the method according to any one of claims 1 to 8.

10. Use of the microneedle patch for quantitatively detecting blood glucose concentration in human body as claimed in claim 9 in detecting blood glucose.

Images