Preparation method and application of copper formate nanomaterial

Abstract

The invention belongs to the field of nanomaterials, and specifically relates to a preparation method and application of a copper formate nanomaterial. The preparation method comprises the following steps: 1) adding hydrogen peroxide, a copper formate solution and an amine solution to a buffer solution in sequence and mixing them evenly, and waiting for the reaction to be completed to obtain a copper formate nanomaterial; 2) adding a stabilizer to the colloidal copper formate nanomaterial obtained in step 1) to solidify it into a hydrogel; the copper formate-containing nanohydrogel material can be used as a glucose sensor for detecting glucose in human urine in vitro; the glucose content is detected by using the urine of a diabetic patient, which, on the one hand, reduces the harm caused to the patient during blood sampling, and on the other hand, does not require additional clinical test samples to be provided, and only requires the patient to provide urine normally discharged when fasting for diagnosis.

Description

Technical Field

[0001] The invention belongs to the field of nano materials, and specifically relates to a preparation method and application of a copper formate nano material. Background Art

[0002] Diabetes is a chronic metabolic disease. Continuous high blood sugar may lead to other metabolic disorders in patients. In more serious cases, it may lead to damage to tissues and organs throughout the body and their dysfunction and failure. It is well known that glucose in serum is an important indicator in clinical biochemical tests. It can provide an objective basis for the diagnosis, treatment, monitoring and prevention of diseases such as diabetes, hypertension and cardiovascular and cerebrovascular systems. However, routine testing for diabetes requires puncture and blood sampling from patients. The blood sampling process may lead to bacterial infection and may also cause physical discomfort to diabetic patients. In addition to routine blood tests for diabetes diagnosis, the glucose content in the body fluids and urine of diabetic patients is also higher than that of normal people; therefore, in order to facilitate diabetic patients to effectively detect the glucose content in the body as early as possible, while reducing the risk of patient diagnosis.

[0003] At present, glucose-sensitive hydrogels are mainly made of network-cross-linked polymers, which are characterized by easy response and high detection sensitivity; however, the preparation of polymer hydrogels is complex, costly, and difficult to prepare in large quantities. Summary of the invention

[0004] In view of the above problems, the present invention prepares a hydrogel containing copper formate nanomaterials for glucose detection in vitro. The hydrogel is easy to prepare, has high sensitivity in urine detection, and has adjustable particle size, thus providing a convenient and fast urine detection method for diabetic patients.

[0005] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0006] A method for preparing a copper formate nanomaterial comprises the following steps:

[0007] 1) Add hydrogen peroxide, copper formate solution and amine solution to the buffer solution in sequence and mix them evenly, and wait for the reaction to be completed to obtain copper formate nanomaterials;

[0008] 2) Adding a stabilizer to the colloidal copper formate nanomaterial obtained in step 1) to solidify it into a hydrogel.

[0009] Furthermore, the buffer is selected from at least one of phosphate buffer, acetate buffer, citrate buffer and carbonate buffer.

[0010] Among them, the phosphate buffer can be selected from at least one of sodium phosphate buffer, disodium hydrogen phosphate-potassium dihydrogen phosphate buffer, potassium dihydrogen phosphate-sodium hydroxide buffer, disodium hydrogen phosphate-sodium hydroxide, etc.; the acetate buffer can be selected from sodium acetate buffer; the citrate buffer can be selected from at least one of citric acid-sodium hydroxide-hydrochloric acid buffer, citric acid-sodium citrate buffer, etc.; the carbonate buffer can be selected from at least one of sodium carbonate-sodium bicarbonate buffer, sodium carbonate-sodium hydroxide buffer, etc.

[0011] The buffer solution can adjust the reaction rate by adjusting the pH of the system. That is, when the solution is acidic, the reaction rate of the system is slower; otherwise, the reaction rate is faster.

[0012] Furthermore, the amine solution may be at least one of benzidine and tetramethylbenzidine solution.

[0013] Furthermore, in step 1), the reaction temperature is -5 to 45°C, preferably 25°C, wherein 25°C has the fastest reaction.

[0014] Furthermore, the concentration of the buffer solution is 10±1mM; the concentration of hydrogen peroxide is 100±10mM; the concentration of the copper formate solution is 0.05~20mM; the concentration of the amine solution is 2±0.5mM; and the volume ratio of the buffer solution, hydrogen peroxide, copper formate and amine solution is 3:1:1:1.

[0015] The concentration of the copper formate solution can adjust the particle size of the copper formate nanomaterial, which is the product of the reaction.

[0016] Furthermore, the stabilizer is selected from at least one of cellulose, graphene, sodium alginate, polyvinyl alcohol, and agarose. The volume ratio of the stabilizer to the colloidal substance obtained in step 1) can be 1-5:8.

[0017] The present invention further provides an application of the copper formate-containing nano-hydrogel material prepared by the above-mentioned preparation method. The copper formate-containing nano-hydrogel material can be used as a glucose sensor for detecting glucose in human urine in vitro.

[0018] The method for detecting glucose using a nano-hydrogel material containing copper formate comprises the following steps:

[0019] S1: Incubate the urine of diabetic patients with glucose oxidase aqueous solution at 37°C for 10 min, add the reaction product into the copper formate nano-hydrogel, and make the reaction product evenly dispersed in the hydrogel;

[0020] S2: Add amine solution to detect absorbance and observe the color development effect.

[0021] Furthermore, the concentration of the glucose oxidase aqueous solution is 10±1 mg / mL, the volume ratio of the diabetic patient's urine to the glucose oxidase aqueous solution is 9:1; and the concentration of the amine solution is 2±0.5 mM.

[0022] The copper formate nanomaterial provided by the present invention can provide a simple and quick detection method for diabetic patients, and the glucose content is detected by using the urine of diabetic patients, which can reduce the damage caused to patients during blood sampling on the one hand, and on the other hand, there is no need to provide additional clinical test samples, and the diagnosis can be made only by the patient providing normal urine discharged when fasting.

[0023] The present invention has the following advantages:

[0024] 1) The raw materials used in the preparation of the copper formate nano-hydrogel of the present invention are low in price and environmentally friendly.

[0025] 2) The preparation method of the copper formate nano-hydrogel provided by the present invention is simple and quick and can be prepared in large quantities.

[0026] 3) The present invention is efficient and rapid in detecting urine sugar content in diabetic patients, and has clear color difference and obvious absorbance difference for different urine glucose contents in a short time.

[0027] 4) The present invention is simple to operate and has rapid and accurate detection. It only requires dropping urine onto the hydrogel and observing its color change.

[0028] 5) The present invention adopts an in vitro urine testing method, which avoids the harm caused to patients by traditional blood tests and is safe and reliable. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is a flow chart for the preparation of copper formate nanomaterials.

[0030] Figure 2 This is an optical photograph of copper formate nanomaterials.

[0031] Figure 3 This is the TEM image of copper formate nanomaterials.

[0032] Figure 4 This is the DLS graph of copper formate nanomaterials at pH = 7. The hydrated particle size of the colloid is 68~78 nm.

[0033] Figure 5 This is an optical picture of the synthesis ratio of copper formate nanomaterial hydrogel. Among them, 9:1, 8:2, 6:4, 4:6, 2:8, and 1:9 represent the volume ratio of stabilizer: copper formate nanomaterial.

[0034] Figure 6 Flow chart of the steps for glucose detection using copper formate nanomaterials.

[0035] Figure 7 The fitted straight line is the absorbance of copper formate nanohydrogel detecting hydrogen peroxide. As can be seen from the figure, the higher the concentration of hydrogen peroxide, the greater the absorbance.

[0036] Figure 8 The absorbance fitting curve of copper formate nanohydrogel detecting different concentrations of glucose. As can be seen from the figure, the higher the glucose concentration, the greater the absorbance.

[0037] Fig. 9 This is an optical photo of using copper formate nanoparticles to detect urine from a diabetic patient and urine from a normal person. In the photo, the urine from a diabetic patient appears clear blue, while the urine from a normal person appears the base color of the test object.

[0038] Fig.10 This is a comparison of the absorbance of urine from diabetic patients and normal people detected by copper formate nanohydrogel. As can be seen from the figure, the urine of diabetic patients has a higher absorbance due to the high glucose content, which is significantly different from the absorbance of normal people's urine.

[0039] Fig.11 This is a bar graph showing the selectivity of copper formate nanohydrogel for detecting glucose. As can be seen from the figure, the hydrogel has good selectivity for glucose detection. DETAILED DESCRIPTION

[0040] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0041] Refer to the attached Figure 1 As shown, the present invention provides a method for preparing a copper formate nanomaterial, comprising the following steps:

[0042] 1) Add hydrogen peroxide (~100mM), copper formate solution (0.05~20mM), and amine solution (~2mmol) to a buffer solution in sequence, mix well, and allow to react at -5~45°C until the reaction is complete to obtain a copper formate nanomaterial; the buffer solution can be at least one of phosphate buffer, acetate buffer, citrate buffer, and carbonate buffer; the amine solution can be at least one of benzidine and tetramethylbenzidine solution.

[0043] 2) Add a stabilizer to the colloid obtained in step 1) to solidify it into a hydrogel. The stabilizer can be selected from at least one of cellulose, graphene, sodium alginate, polyethylene ester, agarose, etc. Figure 5 The volume ratio of the stabilizer to the colloidal substance is preferably 1-5:8.

[0044] Refer to the attached Figure 6 As shown, this embodiment also provides a method for detecting glucose using copper formate nanomaterials, comprising the following steps:

[0045] S1: 450 μL of diabetic urine or different concentrations of glucose aqueous solution were incubated with 50 μL of glucose oxidase aqueous solution (~10 mg / mL) at 37°C for 10 min, and the reaction product was added to the copper formate nanohydrogel to make the reaction product evenly dispersed in the hydrogel;

[0046] S2: Add amine solution (~2 mM) for absorbance detection and observe the color development effect.

[0047] Embodiment 1:

[0048] To prepare the copper formate hydrogel for detection, first weigh 0.87 sodium phosphate and dissolve it in 30 mL deionized water and stir it evenly, and use acetic acid (~1 mM) and sodium hydroxide (~1 mM) to adjust the pH of the solution to 7; weigh 0.01 g copper formate and dissolve it in 10 mL deionized water and stir it evenly; weigh 0.04 mg benzidine (~2mmol) and dissolve it in 10 mL anhydrous ethanol and stir it evenly. Add 10 mL hydrogen peroxide (~100mM), 10 mL copper formate solution and 10 mL benzidine ethanol solution to the sodium phosphate solution with pH = 7 in sequence, and let it stand at 35 °C for 3 h until the reaction is completed to obtain the colloid. Weigh 5 g sodium alginate and dissolve it in 50 mL deionized water and stir it evenly, then mix it with the colloid in a ratio of 5:8 and stir it evenly, let it stand for 30 min, and solidify it into a hydrogel containing copper formate nanomaterials.

[0049] Embodiment 2:

[0050] For the copper formate hydrogel used for detection, first weigh 0.64 g of sodium acetate and dissolve it in 30 mL of deionized water and stir it evenly, and use acetic acid (~1 mM) and sodium hydroxide (~1 mM) to adjust the pH of the solution to 7; weigh 0.01 g of copper formate and dissolve it in 10 mL of deionized water and stir it evenly; weigh 0.04 mg of benzidine (~2mmol) and dissolve it in 10 mL of anhydrous ethanol and stir it evenly. Add 10 mL of hydrogen peroxide (~300mM), 10 mL of copper formate solution and 10 mL of benzidine ethanol solution to the sodium acetate solution with pH = 7 in sequence, and let it stand at 45 °C for 2 h until the reaction is completed, and a copper formate nanomaterial is obtained. Weigh 50 mL of polyvinyl alcohol, then mix it with the colloid in a ratio of 3:8 and stir it evenly, let it stand for 10 min, and solidify into a hydrogel containing copper formate nanomaterials.

[0051] Experimental Example 3:

[0052] For the copper formate hydrogel used for detection, first weigh 0.64 g of sodium acetate and dissolve it in 30 mL of deionized water and stir evenly, and use acetic acid (~1 mM) and sodium hydroxide (~1 mM) to adjust the pH of the solution to 7; weigh 0.01 g of copper formate and dissolve it in 10 mL of deionized water and stir evenly; weigh 0.04 g of tetramethylbenzidine (~2mmol) and dissolve it in 10 mL of anhydrous ethanol and stir evenly. Add 10 mL of hydrogen peroxide (~100mM), 10 mL of copper formate solution and 10 mL of tetramethylbenzidine ethanol solution to the sodium acetate solution with pH = 7 in sequence, and let it stand at 25 °C until the reaction is completed. Weigh 5 g of agarose and dissolve it in 50 mL of deionized water and stir evenly, then mix it thoroughly with the colloid in a ratio of 2:8, heat it at 100 °C and stir evenly, let it stand for 20 min, and solidify it into a hydrogel containing copper formate nanomaterials. The product of the prepared copper formate nanomaterial is as follows Figure 2 As shown, it is yellow-brown colloid with obvious Tyndall effect. Its hydrated particle size is as follows Figure 3-4 As shown, it is approximately 68 nm.

[0053] Experimental Example 4:

[0054] For the copper formate hydrogel used for detection, first weigh 0.64 g sodium acetate and dissolve it in 30 mL deionized water and stir evenly, and use acetic acid (~1 mM) and sodium hydroxide (~1 mM) to adjust the pH of the solution to 7; weigh 0.01 g copper formate and dissolve it in 10 mL deionized water and stir evenly; weigh 0.04 g tetramethylbenzidine (~2mmol) and dissolve it in 10 mL anhydrous ethanol and stir evenly. Add 10 mL hydrogen peroxide (~100mM), 10 mL copper formate solution and 10 mL tetramethylbenzidine ethanol solution to the sodium acetate solution with pH = 7 in sequence, and let it stand at 25 °C until the reaction is completed. Weigh 5 g agarose and dissolve it in 50 mL deionized water and stir evenly, then mix it with the colloid in a ratio of 2:8, heat it at 100 °C and stir evenly, let it stand for 20 min, and solidify into a hydrogel containing copper formate nanomaterials.

[0055] The following tests were performed on the hydrogel containing copper formate nanomaterials prepared in Experimental Example 4:

[0056] 1) 50 μL of hydrogen peroxide of different concentrations (0 mM, 0.01 mM~0.1 mM, 0.01 mM~1mM, 1 mM~10mM) was added dropwise to the copper formate nanohydrogel prepared in Experimental Example 4. After the hydrogen peroxide was evenly dispersed on the hydrogel, 50 μL of TMB ethanol solution was added dropwise. The catalytic hydrogen peroxide property of the copper formate nanomaterial was used to oxidize TMB, thereby achieving the effect of detecting hydrogen peroxide. The absorbance of hydrogen peroxide of different concentrations was detected as shown in FIG. Figure 7 As shown, with the increase of hydrogen peroxide concentration, the ability to oxidize TMB becomes stronger, the absorbance becomes greater, and the color development effect becomes more obvious.

[0057] 2) Mix 450 μL of glucose aqueous solution (0 mM, 0.01 mM, 0.1 mM, 1 mM~10 mM) with 50 μL of glucose oxidase aqueous solution (10 mg / mL), incubate at 37 °C for 10 min, drop the reaction product onto the copper formate nanohydrogel prepared in Experimental Example 4; drop 50 μL of TMB ethanol solution, let it stand for color development, and detect its absorbance as shown in Figure 2. Figure 8 As shown, the higher the glucose concentration, the greater the absorbance.

[0058] 3) Take 5 450 μL urine samples from diabetic patients and mix them with the glucose oxidase aqueous solution, and 5 450 μL normal human urine and 50 μL glucose oxidase aqueous solution (10 mg / mL), and incubate them at 37 °C for 10 min. Drop the reaction product onto the copper formate nanohydrogel prepared in Experimental Example 4; add 50 μL TMB ethanol solution and let it stand for color development. The color development results are as follows: Fig. 9As shown in the figure, it can be clearly observed that the urine samples of diabetic patients are blue in color, while the urine samples of normal people still maintain the background color of the hydrogel; in addition, the absorbance is detected as follows Fig. 9 As shown, the urine of diabetic patients has a higher absorbance due to the high glucose content, which is significantly different from the absorbance of normal human urine. The results were measured within 50 minutes after the urine was dropped onto the material.

[0059] Urine with different glucose content was tested, and the test results were referred to the attached Fig.10 As shown in the figure, it can be seen that the urine of diabetic patients has a higher absorbance due to the high glucose content, which is significantly different from the absorbance of normal people's urine.

[0060] 4) Take 7 portions of 5 mM 450 μL fructose, lactose, maltose, ascorbic acid, and glucose solution and mix with 50 μL glucose oxidase aqueous solution (10 mg / mL), and incubate at 37 °C for 10 min. Drop the reaction product onto the copper formate nanohydrogel prepared in Experimental Example 4; add 50 μL TMB ethanol solution and let it stand for color development. The absorbance was measured and the result was as follows: Fig.11 As shown, from left to right are the blank control group, fructose, lactose, maltose, ascorbic acid dopamine and glucose solution. At the same concentration of 5 mM, the absorbance of glucose is much higher than that of the control group, indicating that the copper formate nanowater material has good selectivity for glucose detection.

[0061] The above embodiments further illustrate the purpose, technical solutions and advantages of the present invention in detail. It should be understood that the above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

[0062] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing a copper formate nanomaterial, characterized in that: The following steps are involved: 1) Add hydrogen peroxide, copper formate solution and amine solution to the buffer solution in sequence and mix them evenly, and wait for the reaction to be completed to obtain copper formate nanomaterials; 2) adding a stabilizer to the colloidal copper formate nanomaterial obtained in step 1) to solidify it into a hydrogel; The concentration of the buffer solution is 10±2mM; the concentration of hydrogen peroxide is 100±10mM; the concentration of the copper formate solution is 0.5~20mM; the concentration of the amine solution is 2±0.5mM; the volume ratio of the buffer solution, hydrogen peroxide, copper formate and amine solution is 3:1:1:1; the amine solution is at least one of benzidine and tetramethylbenzidine solution; the reaction temperature in step 1) is -5~45°C.

2. The method for preparing a copper formate nanomaterial according to claim 1, characterized in that: The buffer is selected from at least one of a phosphate buffer, an acetate buffer, a citrate buffer and a carbonate buffer.

3. The method for preparing a copper formate nanomaterial according to claim 1, characterized in that: The stabilizer is selected from at least one of cellulose, graphene, sodium alginate, polyvinyl alcohol and agarose.

4. A nano hydrogel material containing copper formate obtained by the method for preparing a copper formate nano material as claimed in any one of claims 1 to 3.

5. Use of the nano hydrogel material containing copper formate obtained by the preparation method according to any one of claims 1 to 3 in preparing a detection kit for human urine glucose in vitro.

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