Dopamine electrochemical sensor based on carbonized wood natural structure as well as preparation method and application of dopamine electrochemical sensor

Abstract

The invention relates to a dopamine electrochemical sensor based on a natural structure of carbonized wood as well as a preparation method and application of the dopamine electrochemical sensor. A working electrode of the electrochemical sensor is modified with carbonized wood-ZnO nanoline-ZIF-8. According to the electrochemical sensor, the carbonized wood is used as a substrate, and the carbonized wood is integrally cut and then pasted on the working electrode by using the conductive carbon paste, so that the conductivity is ensured, the three-dimensional structure of a nano material is not damaged, the performance of the sensor is ensured, and the electrochemical sensor has a good application prospect.

Description

Technical Field

[0001] The present invention belongs to the field of biosensor technology, and in particular relates to a dopamine electrochemical sensor based on the natural structure of carbonized wood, and a preparation method and application thereof. Background Art

[0002] Dopamine (DA), a key neurotransmitter and hormone, is widely involved in the fine-tuning of various physiological processes in the human body, including motor control, mood regulation, reward mechanisms, and cardiovascular function. Accurate measurement of dopamine concentration is crucial for the clinical diagnosis and course monitoring of neurological diseases such as Parkinson's disease (characterized by degeneration of substantia nigra dopamine neurons) and schizophrenia (hyperactivity of dopamine in the mesolimbic pathway). It also provides key molecular evidence for the study of the mechanisms of psychiatric and behavioral disorders such as depression (weakened dopamine signaling in the nucleus accumbens) and drug addiction (abnormal dopamine release in the reward circuit). It also demonstrates irreplaceable clinical value in the regulation of vasoactive drugs in shock patients, the evaluation of the efficacy of antipsychotic drugs, and the development of individualized dosing regimens.

[0003] Because dopamine's physiological concentration in the central nervous system is extremely low—typically 10-100 nM (nanomolar) in the striatal synaptic cleft and even as low as pM (picomolar) in peripheral body fluids—and often coexists with high-concentration interfering substances such as ascorbic acid and uric acid, detection technology must possess both ultra-high sensitivity and excellent selectivity. Among the current mainstream detection methods, the enzyme-linked immunosorbent assay (ELISA) relies on a sandwich structure with dopamine-specific antibodies, achieving indirect quantification through an enzyme-catalyzed colorimetric reaction. While high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) can directly measure dopamine molecules, they face significant limitations such as cumbersome sample pretreatment procedures, hours-long analysis cycles, and reliance on large, sophisticated instruments and specialized operators. In contrast, electrochemical sensing technology has become a very promising alternative due to its advantages such as fast response (detection in seconds), simple operation and miniaturization of instruments. However, the sensitivity of existing sensors (usually in the μM level) is still difficult to meet the needs of accurate detection of trace dopamine (nM-pM level) in real biological samples. There is an urgent need to achieve technological breakthroughs through means such as nanomaterial modification and signal amplification strategies. Summary of the Invention

[0004] The present invention provides a dopamine electrochemical sensor based on the natural structure of carbonized wood, and a preparation method and application thereof, which solve the problems of insufficient sensitivity and long detection time in dopamine analysis and detection.

[0005] The present invention provides a dopamine electrochemical sensor based on the natural structure of carbonized wood. The working electrode of the electrochemical sensor is modified with carbonized wood-ZnO nanowire-ZIF-8. Polyethyleneimine is used as a morphology directing agent during the preparation of the carbonized wood-ZnO nanowire-ZIF-8.

[0006] The carbonized wood-ZnO nanowire-ZIF-8 system of the present invention has a densely packed antenna-like structure, possessing a large specific surface area and numerous catalytic sites, which promotes sufficient contact and reaction with dopamine (DA) molecules in solution. The specific synergistic effects are as follows: 1) The carbonized wood material has excellent electrical conductivity, which can transfer electrons generated by the electrochemical oxidation reaction of dopamine to the sensor electrode to generate an electrical signal. At the same time, the naturally porous structure of the carbonized wood effectively increases the specific surface area of ​​the sensing interface, increasing the probability of contact with dopamine molecules. 2) The ZnO nanowire arrays grown in situ within the carbonized wood pores further increase the material's specific surface area while providing zinc atomic sites to support the growth of ZIF-8. 3) The ZIF-8 shell outside the nanowires acts as a solid base nanocatalyst, deeply oxidizing DA molecules in solution. The electron transfer during the oxidation process generates an electrical signal that is proportional to the number of dopamine molecules detected.

[0007] The present invention also provides a method for preparing an electrochemical sensor for monitoring exocytosis, comprising the following steps:

[0008] S1. Carbonizing a Finnish pine block to obtain carbonized wood;

[0009] S2. The carbonized wood is cut into thin slices, washed and dried to obtain carbonized wood slices;

[0010] S3. Add zinc acetate dihydrate in ethanol solution dropwise onto the carbonized wood sheet, defining the added surface as the front side and drying;

[0011] S4. The carbonized wood flakes obtained in step S3 are calcined and removed after cooling;

[0012] S5. preparing an aqueous solution containing polyethyleneimine, zinc nitrate hexahydrate and hexamethylenetetramine to obtain a precursor solution;

[0013] S6. The carbonized wood flakes obtained in step S4 are suspended upside down on the surface of the precursor solution and heated;

[0014] S7. Remove the carbonized wood flakes, wash and dry them, perform secondary calcination, cool them to room temperature, remove them, and repeat steps S5 and S6 to obtain carbonized wood-ZnO nanowires;

[0015] S8. Preheating 2-dimethylimidazole at the bottom of a glass container, suspending the carbonized wood-ZnO nanowires above the 2-dimethylimidazole, heating at 100-120°C for 5-10 minutes, removing the carbonized wood-ZnO nanowires, cooling to room temperature, washing, and drying to obtain carbonized wood-ZnO nanowires-ZIF-8;

[0016] S9. Coat the working electrode area of ​​the screen-printed electrode with a conductive carbon slurry, and paste the carbonized wood-ZnO nanowire-ZIF-8, solidify, and soak in deionized water to obtain the electrochemical sensor for exocytosis monitoring.

[0017] Preferably, the size of the Finnish pine wood block in step S1 is 3 cm×1 cm×1 cm.

[0018] Preferably, the carbonization treatment in step S1 is performed at a temperature of 700 to 1000° C. for 1 to 5 hours under the protection of an inert gas.

[0019] Preferably, the concentration of the ethanol solution of zinc acetate dihydrate in step S3 is 1.111 mg / mL.

[0020] Preferably, the calcination temperature in step S4 is 300-400° C. and the calcination time is 10-30 minutes.

[0021] Preferably, in the precursor solution in step S5, the concentration of polyethyleneimine is 0.00387 g / mL, the concentration of zinc nitrate hexahydrate is 0.008 g / mL, and the concentration of hexamethylenetetramine is 0.0035 g / mL.

[0022] Preferably, the heating temperature in step S6 is 80-100° C. and the heating time is 1-5 hours.

[0023] Preferably, the secondary calcination temperature in step S7 is 300-400° C., and the time is 10-30 minutes.

[0024] Preferably, the preheating temperature in step S8 is 100-120° C., and the preheating time is 20-40 minutes.

[0025] Traditional electrochemical sensor fabrication methods require the synthesized material to be thoroughly ground and ultrasonically processed into a slurry, which is then applied to the surface of a screen-printed electrode. This method can disrupt the three-dimensional structure of the nanomaterial, thereby affecting sensor performance. The method disclosed in this paper uses carbonized wood as a substrate. The carbonized wood is cut into pieces and then affixed to the working electrode using a conductive carbon paste. This ensures conductivity while preserving the three-dimensional structure of the nanomaterial, thus guaranteeing sensor performance.

[0026] The present invention also provides an application of an electrochemical sensor for monitoring exocytosis in detecting dopamine.

[0027] The specific steps include:

[0028] The dopamine test solution was dripped into the test area of ​​the dopamine electrochemical sensor based on the natural structure of carbonized wood. The solution droplets simultaneously covered the working electrode, counter electrode and reference electrode, and cyclic voltammetry test was performed.

[0029] Furthermore, the amount of the dopamine test solution added is 5 to 10 μL, preferably 5 μL.

[0030] Beneficial effects

[0031] 1. The dopamine electrochemical sensor based on the natural structure of carbonized wood of the present invention has a simple and quick determination method, which does not require complex and precise instrument operation and pretreatment process. The test can be performed by simply adding 5 to 10 μL of the required test liquid to the working area of ​​the electrochemical sensor.

[0032] 2. The present invention uses electrochemical technology and screen-printed electrodes to prepare a dopamine electrochemical sensor based on the natural structure of carbonized wood. The detection cost is low and the electrodes can be used once.

[0033] 3. The traditional electrode preparation method grinds the prepared nanomaterial into powder, mixes it with a conductive resin, and ultrasonically mixes it to make a slurry, which is then coated on the working electrode area of ​​the screen-printed electrode. The grinding and ultrasonic processes may destroy the three-dimensional structure of the nanomaterial. The present invention uses carbonized wood as the substrate, which can be directly cut and fixed on the working electrode of the screen-printed electrode with a conductive slurry. Not only is the operation simple, but it can also retain the three-dimensional structure of the nanomaterial to the greatest extent, thereby improving sensor performance.

[0034] 4. The dopamine electrochemical sensor based on the natural structure of carbonized wood of the present invention has a fast detection time, and the sample preparation and testing time is less than 200 seconds.

[0035] 5. The dopamine electrochemical sensor based on the natural structure of carbonized wood of the present invention has good selectivity and can eliminate the interference of molecules such as adrenaline, uric acid, ascorbic acid, glucose and insulin that coexist with dopamine in body fluids. It also has good selectivity for its structural analogs tyrosine and catechol. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 This is a flow chart for the production of the dopamine electrochemical sensor based on the natural structure of carbonized wood proposed in the present invention.

[0037] Figure 2These are scanning electron microscope characterization images of the materials of Example 1, where (a) is the carbonized wood obtained after the carbonization treatment in step S4, and (b)-(d) are carbonized wood-ZnO nanowire-ZIF-8 nanomaterials at different magnifications.

[0038] Figure 3 The response curves of different materials to dopamine in Example 1 are compared by cyclic voltammetry; wherein, the carbonized wood is the carbonized wood obtained in step S4 of Example 1, the carbonized wood + ZnO nanowires are the carbonized wood-ZnO nanowires obtained in step S7 of Example 1, and the carbonized wood + ZnO nanowires + ZIF-8 are the carbonized wood-ZnO nanowires-ZIF-8 obtained in step S8 of Example 1.

[0039] Figure 4 Figure 1 shows the current-time curve (ampere method) of the dopamine electrochemical sensor based on the natural structure of carbonized wood in Example 1 in response to different concentrations of dopamine in PBS buffer, and the linear fitting results of dopamine concentration and current value, where (a) is the current-time curve (ampere method) in response to different concentrations of dopamine, and (b) is the linear fitting result of dopamine concentration and current value.

[0040] Figure 5 This is a comparison chart of the current responses of the dopamine electrochemical sensor based on the natural structure of carbonized wood in Example 1 to 10 μM tyrosine, 10 μM catechol, 10 μM epinephrine, 10 μM uric acid, 10 μM ascorbic acid, 10 μM glucose, 10 μM insulin and 100 pM dopamine.

[0041] Figure 6 This is the response signal and relative standard deviation analysis of the dopamine electrochemical sensor based on the natural structure of carbonized wood prepared in the same batch as in Example 1 to a 20 pM dopamine solution.

[0042] Figure 7 This is a scanning electron microscope image of carbonized wood-ZnO nanowire-ZIF-8 as an electrode sensing material prepared using the method (a) of Comparative Example 1 and the method (b) of Example 1.

[0043] Figure 8 These are the voltammetric cycling curves of the electrochemical sensors prepared in Comparative Example 1 and Example 1 tested in 1 mM dopamine solution. DETAILED DESCRIPTION

[0044] Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiments are only used to illustrate the present invention and are not used in limiting the scope of the present invention.In addition, should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms fall equally within the scope limited by the appended claims of the application.

[0045] An embodiment of the present invention provides a dopamine electrochemical sensor based on the natural structure of carbonized wood. The working electrode of the electrochemical sensor is modified with carbonized wood-ZnO nanowire-ZIF-8. Polyethyleneimine is used as a morphology directing agent during the preparation of the carbonized wood-ZnO nanowire-ZIF-8.

[0046] An embodiment of the present invention also provides a method for preparing the above-mentioned electrochemical sensor for monitoring exocytosis, wherein polyethyleneimine is used as a morphology-directing agent, zinc nitrate hexahydrate is used as a zinc source, and hexamethylenetetramine is used as an alkali source and a ligand, and ZnO nanowires are directionally grown on carbonized wood slices through a hydrothermal reaction to obtain carbonized wood-ZnO nanowires; then, 2-dimethylimidazole is used as a ligand to in situ grow a metal organic framework on the carbonized wood-ZnO nanowires to obtain carbonized wood-ZnO nanowire-ZIF-8; and the carbonized wood-ZnO nanowire-ZIF-8 is modified on a working electrode to obtain the above-mentioned dopamine electrochemical sensor based on the natural structure of carbonized wood.

[0047] Traditional electrochemical sensor fabrication methods require the synthesized material to be thoroughly ground and ultrasonically processed into a slurry, which is then applied to the surface of a screen-printed electrode. This method can disrupt the three-dimensional structure of the nanomaterial, thereby affecting sensor performance. The method disclosed in this paper uses carbonized wood as a substrate. The carbonized wood is cut into pieces and then affixed to the working electrode using a conductive carbon paste. This ensures conductivity while preserving the three-dimensional structure of the nanomaterial, thus guaranteeing sensor performance.

[0048] The present invention uses Finnish pine blocks as raw materials to prepare carbonized wood, which has the following advantages compared to other materials:

[0049] Finnish pine wood itself has a unique natural pore structure and fiber arrangement, which is preserved during the carbonization process, forming a highly ordered carbon skeleton. This naturally porous structure provides an ideal substrate for electrochemical sensing, facilitating electrolyte penetration, ion transport, and uniform loading of sensing materials (such as ZnO nanowires and ZIF-8), thereby improving sensor sensitivity and response speed. As a natural biomass material, wood is widely available and low-cost. Finnish pine wood has a relatively short growth cycle and is renewable. Compared with nanocarbon materials such as graphene and carbon nanotubes, or synthetic materials such as metal oxides, using wood as a raw material can significantly reduce the cost of sensor preparation and conform to the concepts of green chemistry and sustainable development. Carbonized wood (carbonized wood) retains its natural fiber structure, giving it excellent mechanical strength and stability. This property enables carbonized wood-based sensors to maintain structural integrity even under repeated use or bending, extending their service life. By adjusting the carbonization temperature, heating rate, and atmosphere, the porosity, conductivity, and surface chemistry of carbonized wood can be precisely controlled. This process controllability makes carbonized wood an ideal substrate material for composite with other functional materials (such as ZnO and ZIF-8) to form a high-performance sensing interface. The wood carbonization process does not require the use of toxic reagents, and the carbonized wood itself is chemically inert and does not easily release harmful substances during the electrochemical sensing process. In addition, discarded carbonized wood-based sensors can be treated to achieve carbon neutrality through incineration and other methods, further reducing the environmental burden. The natural hydrophilicity of wood facilitates the uniform deposition of sensing materials (such as ZIF-8) and the sufficient infiltration of the electrolyte, thereby optimizing the electrode / electrolyte interface contact and improving the sensing performance. Although the hydrophilicity is further enhanced by soaking in deionized water in the embodiment, the hydrophilic properties of the wood itself have provided a favorable basis for this step; through cutting (such as 700μm thin slices in step S2) and subsequent functionalization (such as ZnO nanowire growth and ZIF-8 coating), carbonized wood can be designed into a sensing material with specific morphology and function. This structural designability enables carbonized wood-based sensors to optimize the performance for different target molecules (such as dopamine); in summary, Finnish pine blocks are used as raw materials and converted into carbonized wood through a carbonization process, which not only retains the structural advantages of natural materials, but also combines the characteristics of low cost, renewability, mechanical stability and environmental friendliness, providing an ideal platform for the preparation of high-performance electrochemical sensors.

[0050] The dopamine electrochemical sensor based on the natural structure of carbonized wood provided in the embodiment of the present invention can be used to detect dopamine.

[0051] The method for detecting dopamine using the dopamine electrochemical sensor based on the natural structure of carbonized wood of the present invention comprises the following steps:

[0052] The dopamine test solution was dripped into the test area of ​​the dopamine electrochemical sensor based on the natural structure of carbonized wood. The solution droplets simultaneously covered the working electrode, counter electrode and reference electrode, and cyclic voltammetry test was performed.

[0053] In a preferred embodiment of the present invention, the working electrode and the counter electrode of the screen-printed electrode are both carbon electrodes, and the reference electrode is a silver / silver chloride electrode.

[0054] In a preferred embodiment of the present invention, the amount of the dopamine test solution added is 5 to 10 μL, preferably 5 μL.

[0055] In the following embodiments of the present invention, the specific steps of detecting dopamine using a dopamine electrochemical sensor are as follows:

[0056] S1. Prepare the dopamine test solution using PBS buffer (pH = 7.4);

[0057] S2. Add 5-10 μL of the dopamine test solution to the sensor test area, so that the solution droplets cover the working electrode, counter electrode, and reference electrode at the same time.

[0058] S3. Use the ampere method at a potential of 0.34 V for 200 s and read the current value.

[0059] The present invention provides a multidimensional hybrid nanomaterial (carbonized wood-ZnO nanowire-ZIF-8) that is in situ grown on the natural structure of carbonized wood: an array of one-dimensional nanowires is grown on a three-dimensionally carbonized Finnish pine block, and a porous metal-organic framework compound (ZIF-8) is further grown on the surface of the nanowires. This structure has a synergistic effect and can efficiently electrocatalytically detect dopamine molecules. When the carbonized wood-ZnO nanowire-ZIF-8 nanostructured material is modified onto the working electrode of the screen-printed electrode, a dopamine electrochemical sensor is prepared, and testing can be performed by simply adding the dopamine solution to be tested to the detection area. The dopamine electrochemical sensor based on the natural structure of carbonized wood proposed by the present invention can achieve highly sensitive, highly selective and rapid detection of dopamine due to the unique multidimensional nanostructure and scale of the sensitive material.

[0060] Unless otherwise specified, the room temperature in the present invention is 25±2°C.

[0061] All raw materials used in the examples of the present invention are commercially available.

[0062] It should be pointed out that the matters not described in detail in the present invention are conventional operating methods in the field and are not the focus of the present invention. For example, the specific preparation methods such as the aqueous solution containing 25 mM zinc nitrate hexahydrate and 25 mM hexamethylenetetramine are all completed by conventional methods.

[0063] The technical solution of the present invention is further illustrated by the following examples.

[0064] Example 1

[0065] This embodiment provides a method for preparing a dopamine electrochemical sensor based on the natural structure of carbonized wood. The specific steps are as follows:

[0066] S1. A 3 cm × 1 cm × 1 cm block of Finnish pine wood was placed in a tube furnace and heated at 800°C for 2 h under an Ar atmosphere (flow rate of 0.1 L / min) for 4 h to complete carbonization and obtain carbonized wood.

[0067] S2. The carbonized wood was cut into 700 μm thick slices, ultrasonically cleaned in ethanol, and then dried at 150°C.

[0068] S3. Weigh 22.22 mg of zinc acetate dihydrate and dissolve it in 20 mL of ethanol. Ultrasonicate the solution to completely dissolve it in the ethanol. Then, use a 100 mL pipette to dropwise add 25 μL of this solution onto the carbonized wood slice (the surface to which the solution was added is facing the front). Dry at 150°C. Repeat this process four times.

[0069] S4. The carbonized wood slices treated above were placed in a tube furnace and heated at a constant temperature of 350°C for 20 minutes, then cooled to room temperature and removed;

[0070] S5. Weigh 0.387g polyethyleneimine and 0.8g zinc nitrate hexahydrate, add 10mL of deionized water and stir thoroughly to dissolve, then add 0.35g of hexamethylenetetramine and dilute to 100mL with deionized water to obtain a uniformly mixed precursor solution;

[0071] S6. Pour the mixed precursor solution into a 100 mL wide-mouth container, place the carbonized wood flakes upside down and suspend them on the surface of the precursor solution, and heat at 90°C for 4 h.

[0072] S7. Remove the carbonized wood slice, wash it twice with deionized water, then once with ethanol, dry it at 150°C, place it face up in a tube furnace and heat it at 350°C for 10 minutes, then cool it to room temperature and remove it; then repeat steps S5 and S6, remove the carbonized wood slice, wash it twice with deionized water, then once with ethanol, and dry it at 150°C to obtain carbonized wood-ZnO nanowires;

[0073] S8. Weigh 2 g of 2-dimethylimidazole and place it at the bottom of a glass container. Preheat the container at 110°C for 30 min. Then, suspend the carbonized wood-ZnO nanowires obtained in step S7 8 cm above the 2-dimethylimidazole and heat at 110°C for 5 min. Remove the carbonized wood-ZnO nanowires and cool them to room temperature. Rinse the mixture once with ethanol and remove excess ethanol at 110°C to obtain carbonized wood-ZnO nanowire-ZIF-8.

[0074] S9. Apply 1 μL of conductive carbon slurry (Celanese BQ242) to the working electrode area of ​​the screen-printed electrode (the working electrode and the counter electrode are carbon electrodes, and the reference electrode is a silver / silver chloride electrode), and paste the carbonized wood-ZnO nanowire-ZIF-8 cut into 1 mm × 1 mm pieces on it;

[0075] S10. The material obtained in S9 was placed in a 50°C oven for 30 min for curing;

[0076] To enhance the hydrophilicity of the sensing material and ensure adequate electrolyte / electrode interface contact during electrochemical testing, the cured electrochemical sensor was immersed in deionized water for 24 h to obtain a dopamine electrochemical sensor based on the natural structure of carbonized wood.

[0077] The scanning electron microscope characterization of the material prepared in this embodiment is shown in FIG. Figure 2 As shown, from Figure 2 It can be seen from a that the carbonized wood obtained after carbonization treatment has electrical conductivity while still retaining its natural pore structure; Figure 2 b It can be seen that after the in situ growth of ZnO nanowire-ZIF-8 material, the pore size is about 10μm; Figure 2 c It can be seen that ZnO nanowire-ZIF-8 is evenly distributed on the carbonized wood substrate; Figure 2 d It can be seen that the ZIF-8 shell is evenly distributed on the ZnO nanowires, ensuring efficient catalytic oxidation of dopamine molecules in the solution.

[0078] The electrochemical sensor prepared in this embodiment was used to detect dopamine standard solutions of different concentrations, and the response ability and detection limit of the sensor to dopamine were analyzed. The specific implementation operation was as follows: PBS buffer (pH = 7.4) was used as the background liquid to prepare a 1mM dopamine solution. Under the settings of a scan range of -0.2 to 0.6V and a scan rate of 0.05V / s, carbonized wood, carbonized wood-ZnO nanowires, and carbonized wood-ZnO nanowires-ZIF-8 were used as electrode sensing materials to perform cyclic voltammetry tests. The response curves of different materials to dopamine in this embodiment are compared by cyclic voltammetry. Figure 3As shown, the carbonized wood material produced a weak oxidation signal for 1 mM dopamine in solution. However, after in situ growth of ZnO nanowires on the carbonized wood substrate, the increased specific surface area enhanced the oxidation current signal. Furthermore, after in situ growth of ZIF-8 on the ZnO nanowires, the specific catalytic effect of the ZIF-8 solid base on dopamine further increased the oxidation current intensity, resulting in a strong oxidation current at 0.34 V. Therefore, the subsequent amperometric experiments were conducted at a potential of 0.34 V.

[0079] Using PBS buffer (pH = 7.4) as the background solution, 0.5pM, 1pM, 10pM, 20pM, 40pM, 60pM, and 80pM dopamine standard solutions were prepared respectively; the electrochemical sensor was connected to the electrochemical workstation, and 5μL of the dopamine standard solution of the above concentration was dripped into the working area of ​​the electrochemical sensor. The current was detected using the amperometric method: the detection potential was set to 0.34V, the detection time was 200s, and the final current value was read after the detection was completed; each concentration of dopamine solution was tested three times; the average value and standard deviation of the current were calculated, and the dopamine "current-concentration" standard calibration curve was drawn to obtain the linear formula (1) y = 1.54*x + 49.73, where y is the current (in nanoamperes), x is the dopamine concentration (in pM), and the linear regression coefficient R 2 =0.9973. The current-time curves (amperometric method) of the dopamine electrochemical sensor based on the natural structure of carbonized wood in this embodiment in response to different concentrations of dopamine in PBS buffer and the linear fitting results of dopamine concentration and current value are shown in FIG. Figure 4 As shown, from Figure 4 a It can be seen that the current value increases with the increase of dopamine concentration; Figure 4 b It can be seen that there is a good linear relationship between dopamine concentration and current value in the range of 0.5 pM-80 pM, and the detection limit is 0.5 pM.

[0080] The prepared electrochemical sensor was compared to detect the difference in signal response to interfering molecules and dopamine molecules with a 100-fold difference in concentration. The specific implementation steps are as follows:

[0081] 1) Using PBS buffer (pH = 7.4) as the background solution, prepare 10 μM tyrosine, 10 μM catechol, 10 μM epinephrine, 10 μM uric acid, 10 μM ascorbic acid, 10 μM glucose, 10 μM insulin, and 100 pM dopamine standard solutions, respectively;

[0082] 2) Connect the prepared electrochemical sensor to the electrochemical workstation and drop 5 μL of the test solution into the working area of ​​the electrochemical sensor;

[0083] 3) Detect the response current using the amperometric method: set the detection potential to 0.34 V and the detection time to 200 s, and read the final current value after the detection is completed; each test solution is tested three times;

[0084] 4) Calculate the average value and standard deviation of the current and compare them using a bar graph.

[0085] The current response comparison of the dopamine electrochemical sensor based on the natural structure of carbonized wood in this embodiment to 10 μM tyrosine, 10 μM catechol, 10 μM epinephrine, 10 μM uric acid, 10 μM ascorbic acid, 10 μM glucose, 10 μM insulin and 100 pM dopamine is shown in the figure below. Figure 5 As shown in the figure, the response of the sensor to higher concentrations of dopamine analogs tyrosine, catechol and common interfering substances uric acid, ascorbic acid, glucose, and insulin is close to that of the blank sample, while a strong current signal is obtained for low concentration (100pM) of dopamine, proving that the sensor has good selectivity and specificity for dopamine.

[0086] To verify the consistency of the electrochemical sensor based on the natural structure of carbonized wood prepared in this example, the specific implementation steps are as follows:

[0087] 1) Prepare a 20 μM dopamine standard solution using PBS buffer (pH = 7.4) as the background solution;

[0088] 2) Connect the prepared electrochemical sensor to the electrochemical workstation and drop 5 μL of the test solution into the working area of ​​the electrochemical sensor;

[0089] 3) Randomly select five electrochemical sensors from the same batch and measure the response current using the amperometric method: set the detection potential to 0.34 V and the detection time to 200 s. After the detection is complete, read the final current value.

[0090] 4) Compare the current collected by each electrochemical sensor using a bar graph and calculate the average value and relative standard deviation.

[0091] The response signals and relative standard deviations of the dopamine electrochemical sensors based on the natural structure of carbonized wood prepared in the same batch in this example to 20 pM dopamine solution are shown in the following figure: Figure 6 As shown, it can be seen that the randomly selected electrodes prepared from the same batch have very close response current values ​​for the dopamine standard solution with the same concentration, with a relative standard deviation of 3%, which proves that the electrochemical sensor has good consistency.

[0092] Comparative Example 1

[0093] A dopamine electrochemical sensor based on the natural structure of carbonized wood, the specific steps are as follows:

[0094] S1. A 3 cm × 1 cm × 1 cm Finnish pine block was placed in a tube furnace and heated at 800°C for 2 h under an Ar atmosphere (flow rate 0.1 L / min) for 4 h to complete the carbonization process and obtain carbonized wood material.

[0095] S2. The carbonized wood was cut into 700 μm thick slices, ultrasonically cleaned in ethanol, and then dried at 150°C.

[0096] S3. Weigh 22.22 mg of zinc acetate dihydrate and dissolve it in 20 mL of ethanol. Ultrasonicate the solution to completely dissolve it in the ethanol. Then, use a 100 mL pipette to dropwise add 25 μL of this solution onto the carbonized wood slice (the surface to which the solution was added is facing the front). Dry at 150°C. Repeat this process four times.

[0097] S4. The carbonized wood material was placed in a tube furnace and heated at a constant temperature of 350°C for 20 minutes, then cooled to room temperature and removed;

[0098] S5. Prepare an aqueous solution containing 25 mM zinc nitrate hexahydrate and 25 mM hexamethylenetetramine (precursor solution) and transfer it to a 100 mL wide-mouth container. Suspend the carbonized wood flakes face upside down on the surface of the precursor solution and heat at 90°C for 3 h to obtain carbonized wood-ZnO nanowires.

[0099] S6. Weigh 0.5 g of 2-dimethylimidazole and place it at the bottom of a glass container. Preheat the container at 110°C for 30 minutes. Then, suspend the carbonized wood flakes 5 cm above the 2-dimethylimidazole and heat at 110°C for 5 minutes. Remove the flakes, cool them to room temperature, rinse them with ethanol, and remove excess ethanol at 110°C to obtain carbonized wood-ZnO nanowire-ZIF-8.

[0100] S7. Apply 1 μL of conductive carbon slurry (Celanese BQ242) to the working electrode area of ​​the screen-printed electrode (the working electrode and counter electrode are carbon electrodes, and the reference electrode is a silver / silver chloride electrode). Paste the carbonized wood-ZnO nanowire-ZIF-8 cut into 1 mm × 1 mm pieces on top.

[0101] S8. Place in a 50°C oven for 30 minutes to cure;

[0102] S9. To improve the hydrophilicity of the sensing material and ensure adequate contact between the electrolyte and the electrode during electrochemical testing, the electrochemical sensor was immersed in deionized water for 24 hours.

[0103] S10. Prepare a 1 mM dopamine test solution using PBS buffer (pH = 7.4);

[0104] S11. Take 5 μL of the dopamine test solution and drop it onto the sensor test area. The solution droplets cover the working electrode, counter electrode, and reference electrode at the same time.

[0105] S12. Under the settings of a scan range of -0.2 to 0.6 V and a scan rate of 0.05 V / s, electrochemical sensors were prepared using the carbonized wood-ZnO nanowire-ZIF-8 prepared in Comparative Example 1 and Example 1 as electrode sensing materials, and cyclic voltammetry tests were performed.

[0106] The scanning electron microscope images of carbonized wood-ZnO nanowire-ZIF-8 prepared by the methods of Comparative Example 1 and Example 1 are as follows: Figure 7 As shown, it can be seen that the ZnO nanowires grown using the method of Comparative Example 1 cannot achieve complete coverage of the carbonized wood; the method of Example 1 can be used to grow ZnO nanowires and ZIF-8 on the carbonized wood substrate. By performing two hydrothermal growth methods on the ZnO nanowires, complete coverage of the ZnO nanowires on the carbonized wood can be achieved, and the length of the nanowires is significantly increased.

[0107] The voltammetric cycling curves of the electrochemical sensors prepared in Comparative Example 1 and Example 1 tested in 1 mM dopamine solution are as follows: Figure 8 As shown, the material synthesized by the method of Comparative Example 1 has only one oxidation peak with a peak value of 7.4 microamperes; while the material synthesized by the method of Example 1 can deeply oxidize dopamine, with the maximum oxidation peak value of 12.5 microamperes, which significantly improves the signal intensity.

[0108] The above are merely preferred embodiments of the present invention, but the scope of protection of the present invention is not limited thereto. Any changes or substitutions that can be easily conceived by a person skilled in the art within the technical scope disclosed in the present invention should be included in the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be based on the scope of protection of the claims.

Claims

1. A dopamine electrochemical sensor based on the natural structure of carbonized wood, characterized by: The working electrode of the electrochemical sensor is modified with carbonized wood-ZnO nanowire-ZIF-8; and polyethyleneimine is used as a morphology directing agent during the preparation of the carbonized wood-ZnO nanowire-ZIF-8.

2. A method for preparing a dopamine electrochemical sensor based on the natural structure of carbonized wood, comprising the following steps: S1. Carbonizing a Finnish pine block to obtain carbonized wood; S2. The carbonized wood is cut into thin slices, washed and dried to obtain carbonized wood slices; S3. Add zinc acetate dihydrate in ethanol solution dropwise onto the carbonized wood sheet, defining the added surface as the front side and drying; S4. The carbonized wood flakes obtained in step S3 are calcined and removed after cooling; S5. preparing an aqueous solution containing polyethyleneimine, zinc nitrate hexahydrate and hexamethylenetetramine to obtain a precursor solution; S6. The carbonized wood flakes obtained in step S4 are suspended upside down on the surface of the precursor solution and heated; S7. Remove the carbonized wood flakes, wash and dry them, perform secondary calcination, cool them to room temperature, remove them, and repeat steps S5 and S6 to obtain carbonized wood-ZnO nanowires; S8. Preheating 2-dimethylimidazole at the bottom of a glass container, suspending the carbonized wood-ZnO nanowires above the 2-dimethylimidazole, heating at 100-120°C for 5-10 minutes, removing the carbonized wood-ZnO nanowires, cooling to room temperature, washing, and drying to obtain carbonized wood-ZnO nanowires-ZIF-8; S9. Coat the working electrode area of ​​the screen-printed electrode with a conductive carbon slurry, and paste the carbonized wood-ZnO nanowire-ZIF-8, solidify, and soak in deionized water to obtain the electrochemical sensor for exocytosis monitoring.

3. The preparation method according to claim 2, wherein: The carbonization treatment in step S1 is performed at a temperature of 700 to 1000° C. for 1 to 5 hours under the protection of an inert gas.

4. The preparation method according to claim 2, wherein: The concentration of the ethanol solution of zinc acetate dihydrate in step S3 is 1.111 mg / mL.

5. The preparation method according to claim 2, wherein: In step S4, the calcination temperature is 300-400° C. and the calcination time is 10-30 minutes.

6. The preparation method according to claim 2, wherein: In the step S5, the concentration of polyethyleneimine in the precursor solution is 0.00387 g / mL, the concentration of zinc nitrate hexahydrate is 0.008 g / mL, and the concentration of hexamethylenetetramine is 0.0035 g / mL.

7. Use of the dopamine electrochemical sensor based on the natural structure of carbonized wood as claimed in claim 1 in detecting dopamine.

8. The use according to claim 7, characterized in that: The following steps are involved: The dopamine test solution was dripped into the test area of ​​the dopamine electrochemical sensor based on the natural structure of carbonized wood. The solution droplets simultaneously covered the working electrode, counter electrode and reference electrode, and cyclic voltammetry test was performed.

9. The use according to claim 8, characterized in that: The amount of the dopamine test solution added is 5 to 10 μL.

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