An electrochemical biosensor, a preparation method thereof and application thereof in dopamine detection

Abstract

The application relates to an electrochemical biosensor and a preparation method thereof and application thereof in dopamine detection, wherein the photoelectrochemical biosensor adopts a three-electrode system, a glassy carbon electrode modified by a single-atom catalyst with Ni-N4 coordination is used as a working electrode, an Ag / AgCl electrode is used as a reference electrode, and a platinum wire electrode is used as a counter electrode. The electrochemical biosensor provided by the application uses the glassy carbon electrode modified by the single-atom catalyst with Ni-N4 coordination as the working electrode, the single-atom catalyst has excellent electron conductivity and extremely high catalytic activity, a large specific surface area, high dispersity, more active sites, high atom utilization rate and the like, the electrochemical biosensor prepared based on the single-atom catalyst has high sensitivity in dopamine detection, can clearly distinguish common interferents such as ascorbic acid, dopamine and uric acid, and has good selectivity.

Description

Technical Field

[0001] This invention belongs to the field of materials testing or analysis technology by measuring the chemical or physical properties of materials, specifically relating to an electrochemical biosensor based on a single-atom catalyst, its preparation method, and its application in dopamine detection. Background Technology

[0002] Organisms contain various electrochemically active small molecules, such as ascorbic acid (AA), dopamine (DA), and uric acid (UA), all of which play important roles in normal life activities. Dopamine, also known as catechol ethylamine, plays a crucial role in the central nervous system, involved in vital activities such as movement, emotion, cognition, and learning and memory. A deficiency in dopamine may lead to neurodegenerative diseases of the motor system, such as Parkinson's disease. Decreased dopamine levels in the central nervous system can also cause brain aging. Therefore, measuring dopamine levels is of great significance for monitoring human health.

[0003] To date, numerous detection methods have been applied to the detection of small biomolecules, such as high-performance liquid chromatography (HPLC), gas chromatography-mass spectrometry (GC-MS), ultraviolet-visible spectrophotometry (UV-Vis spectrophotometry), and fluorescence spectroscopy. However, most of these methods suffer from drawbacks such as high cost, complex sample preparation, and long analysis and detection times. In contrast, electrochemical detection methods offer advantages such as ease of operation, short response time, and low cost, and are therefore widely used for the detection of small biomolecules.

[0004] In electrochemical detection, amplifying the electrochemical signal to improve detection sensitivity and selectivity is a key scientific problem that urgently needs to be solved. This invention modifies the electrode surface using an appropriate catalyst, giving the modified electrode a large number of structurally uniform catalytic active sites, thereby achieving highly sensitive and selective detection of dopamine. Summary of the Invention

[0005] In view of the above-mentioned shortcomings in the prior art, the technical problem to be solved by the present invention is to provide an electrochemical biosensor based on a single-atom catalyst, its preparation method and its application in dopamine detection, which can achieve highly sensitive and selective detection of dopamine.

[0006] To solve the above-mentioned technical problems, the technical solution provided by the present invention is as follows:

[0007] An electrochemical biosensor, wherein the photoelectrochemical biosensor employs a three-electrode system, with a glassy carbon electrode modified by a Ni-N4 coordinated single-atom catalyst as the working electrode, an Ag / AgCl electrode as the reference electrode, and a platinum wire electrode as the counter electrode.

[0008] The present invention also includes a method for preparing the above-mentioned electrochemical biosensor, wherein the steps of the method for preparing the working electrode are as follows:

[0009] 1) Preparation of Ni-N4 coordinated single-atom catalyst: First, nickel acetate and Schiff base ligand are added to a solvent and mixed to obtain Ni(II) Schiff base ligand. Then, carbon black is added, mixed evenly, and heated to evaporate and remove the solvent, so that Ni(II) Schiff base ligand is fully adsorbed on the carbon black support to obtain a complex. Then, the obtained complex is placed in a tube furnace, and sufficient argon gas is introduced to remove air, and then pyrolyzed to obtain Ni-N4 coordinated single-atom catalyst.

[0010] 2) Preparation of electrochemical biosensor: The single-atom catalyst prepared in step 1) is dispersed in a mixture of double-distilled water, ethanol and Nafion solution, and ultrasonically dispersed to obtain a uniform dispersion. The obtained dispersion is uniformly coated on the surface of a glassy carbon electrode, and after drying, a working electrode is obtained, and an electrochemical biosensor is further prepared.

[0011] According to the above scheme, the molar ratio of nickel acetate to Schiff base ligand in step 1) is 1:1 to 2.

[0012] According to the above scheme, the Schiff base ligand in step 1) is 1,10-phenanthroline.

[0013] According to the above scheme, the solvent in step 1) is ethanol or acetone, and the mass-volume ratio of nickel acetate to solvent is 1.0 to 2.5 mg / mL.

[0014] According to the above scheme, the mass ratio of carbon black to nickel acetate in step 1) is 1:10-20.

[0015] According to the above scheme, the heating and evaporation temperature in step 1) is 60-80℃, and the heating and evaporation time is 4-6h.

[0016] According to the above scheme, the pyrolysis process conditions in step 1) are: pyrolysis at 300-600℃ for 2-4 hours.

[0017] Preferably, in step 2), the volume ratio of the double-distilled water, ethanol, and Nafion solution in the mixture is 79:20:1, and the Nafion solution concentration is 5 wt%.

[0018] According to the above scheme, the mass-to-volume ratio of the single-atom catalyst in step 2) to the mixture of double-distilled water, ethanol and Nafion solution is 2-4 mg / mL.

[0019] This invention also includes the application of the aforementioned electrochemical biosensor in dopamine detection. Dopamine can be detected by performing an electrochemical scan in a test solution containing dopamine and detecting the oxidation peak of dopamine.

[0020] The beneficial effects of this invention are as follows: 1. The electrochemical biosensor provided by this invention uses a glassy carbon electrode modified with a Ni-N4 coordinated single-atom catalyst as the working electrode. This single-atom catalyst has excellent electronic conductivity and extremely high catalytic activity, large specific surface area, high dispersion, many active sites, and high atomic utilization. Based on this, the electrochemical biosensor prepared has high sensitivity in detecting dopamine and can clearly distinguish common interfering substances such as ascorbic acid, dopamine, and uric acid, with good selectivity; 2. The preparation method of the electrochemical biosensor of this invention has simple synthesis steps, low cost, and is suitable for commercial application. Attached Figure Description

[0021] Figure 1 This is a scanning electron microscope image of the surface of the working electrode prepared in Example 1 of the present invention;

[0022] Figure 2 The CV diagrams of the working electrode prepared in Example 1 and the unmodified bare glassy carbon electrode in 0.1M PBS buffer solution containing ascorbic acid, dopamine, and uric acid are shown.

[0023] Figure 3 CV diagrams of the working electrode prepared in Example 1 and the electrodes modified with C nanomaterials, N-doped carbon materials and Ni-containing nanoparticle materials in 0.1M PBS buffer solution containing ascorbic acid, dopamine and uric acid;

[0024] Figure 4 DPV diagrams of the working electrode prepared in Example 1 and the electrodes modified with C nanomaterials, N-doped carbon materials and Ni nanoparticles in 0.1M PBS buffer solution containing ascorbic acid, dopamine and uric acid;

[0025] Figure 5 The CV curve of the working electrode prepared in Example 1 in PBS buffer solution with varying dopamine concentration;

[0026] Figure 6 for Figure 5 Linear curves showing the peak current versus concentration of dopamine. Detailed Implementation

[0027] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be further described in detail below with reference to the accompanying drawings.

[0028] Example 1

[0029] An electrochemical biosensor employs a three-electrode system, using a glassy carbon electrode modified with a Ni-N4 coordinated single-atom catalyst as the working electrode, an Ag / AgCl electrode as the reference electrode, and a platinum wire electrode as the counter electrode. The preparation method of the working electrode is as follows:

[0030] 1) First, nickel acetate (12.4 mg) and Schiff base ligand 1,10-phenanthroline (29.7 mg) were added to 5 mL of ethanol and mixed evenly to obtain NiNⅡN Schiff base ligand. Then, carbon black (70 mg) was added and stirred and evaporated at 60 °C for 4 hours to allow the NiNⅡN Schiff base ligand to be fully adsorbed on the carbon black support. Next, the resulting mixture was placed in a tube furnace, and sufficient argon gas was introduced to purge the air. Then, it was pyrolyzed at 300 °C for 2 hours to finally obtain a single-atom catalyst coordinated with Ni-N4 (X-ray absorption near-edge spectroscopy and X-ray absorption fine structure spectroscopy showed that Ni element was distributed in single-atom form, and the fitting results showed that the metal active center was Ni-N4 coordinated structure).

[0031] 2) Disperse 4 mg of the single-atom catalyst prepared in step 1) in a mixture containing 0.79 mL of deionized water, 0.2 mL of ethanol and 0.01 mL of 5 wt% Nafion, and sonicate for 10 minutes. After uniform dispersion, take 3 μL of the dispersion and spin coat it evenly onto the polished glassy carbon electrode surface. After drying, obtain the working electrode and further prepare an electrochemical biosensor.

[0032] Figure 1 The image shows a scanning electron microscope image of the working electrode surface prepared in this embodiment. The image shows that the substrate of the single-atom catalyst is a carbon-based material with a loose and porous structure, which is conducive to the full contact between the single-atom catalyst and the electrochemically active material and the electrolyte used.

[0033] The electrochemical biosensor prepared in this embodiment was transferred to a buffer solution containing 0.1M PBS (pH=7.4) for electrochemical performance testing. Cyclic voltammetry (CV) was used with a test voltage range of -0.80V to 0.80V and a scan rate of 0.05V / s. Differential pulse voltammetry (DPV) was used with a test voltage range of -0.50V to 0.50V and a scan rate of 0.05V / s.

[0034] Figure 2The figure shows the CV curves of the working electrode prepared in this embodiment and the unmodified bare glassy carbon electrode (Bare GCE) in 0.1M PBS buffer solution containing ascorbic acid (AA, 200 μM), dopamine (DA, 200 μM), and uric acid (UA, 200 μM). The scanning voltage range is -0.8V to 0.8V, and the scanning rate is 50mV / s. The glassy carbon electrode modified with a Ni-N4 coordinated single-atom catalyst has good selectivity and exhibits three different oxidation peaks, which can effectively catalyze the oxidation of DA and is not affected by the interfering substances AA and UA. In contrast, the bare glassy carbon electrode only shows one oxidation peak, which cannot specifically quantify a certain substance and is subject to mutual interference.

[0035] N-doped carbon materials were prepared according to step 1) of this embodiment, without the addition of nickel acetate, and the remaining conditions were the same as in step 1) of this embodiment. Ni-containing nanoparticle materials were prepared according to step 1) of this embodiment, without the addition of 1,10-phenanthroline, and the remaining conditions were the same as in step 1) of this embodiment. The method for modifying the electrode was the same as in step 2) of this embodiment. Figure 3 The figure shows the CV curves of the working electrode prepared in this embodiment and the electrodes modified with C nanomaterials (commercial carbon black, average particle size 30 nm), N-doped carbon materials, and Ni-containing nanoparticle materials in 0.1 M PBS buffer solution containing ascorbic acid (AA, 200 μM), dopamine (DA, 200 μM), and uric acid (UA, 200 μM). The scanning voltage range is -0.8 V to 0.8 V, and the scanning rate is 50 mV / s. As can be seen from the figure, the working electrode prepared in this embodiment exhibits the best detection sensitivity. Three independent and interference-free oxidation peaks can be clearly observed in the figure, corresponding to the oxidation of AA, DA, and UA, respectively. Figure 4 The image shows the DPV of the working electrode prepared in this embodiment, as well as the electrodes modified with C nanomaterials, N-doped carbon materials, and Ni nanoparticles, in a 0.1M PBS buffer solution containing ascorbic acid (AA, 200 μM), dopamine (DA, 200 μM), and uric acid (UA, 200 μM). The scanning voltage range is -0.5V to 0.5V, and the scanning rate is 5mV / s. It can be seen that when performing differential potential voltammetry testing, the working electrode prepared in this embodiment exhibits excellent catalytic performance in the voltage range of -0.5V to 0.5V, and can effectively distinguish DA from interfering substances AA and UA by different potentials.

[0036] A 0.1M PBS buffer solution containing ascorbic acid (AA), dopamine (DA), and uric acid (UA) was prepared, maintaining the concentration of AA at 100 μM, UA at 300 μM, and DA concentrations ranging from 2 to 800 μM (concentrations of 2 μM, 5 μM, 10 μM, 20 μM, 50 μM, 100 μM, 200 μM, 500 μM, and 800 μM, respectively). The CV curve of the working electrode prepared in this embodiment was tested, with a scan voltage range of -0.5 to 0.6 V and a scan rate of 5 mV / s. The CV curve is shown below. Figure 5 As shown, the oxidation peak current of DA changes with the concentration, and increases linearly. The linear function corresponding to the peak current of DA and its concentration, obtained through fitting, is I0. P (DA) = 0.1852C DA +12.7992, correlation coefficient R 2 =0.97( Figure 6 The detection limit is 0.6 μM.

[0037] In summary, the electrochemical biosensor using a glassy carbon electrode modified with a Ni-N4 coordinated single-atom catalyst as the working electrode exhibits characteristics such as a wide linear range, low detection limit, high sensitivity, and good selectivity in the detection of dopamine, and has promising application prospects.

Claims

1. Use of an electrochemical biosensor for the detection of dopamine, characterized in that, The electrochemical biosensor adopts a three-electrode system, and a glassy carbon electrode modified by a single-atom catalyst with Ni-N4 coordination is used as a working electrode, an Ag / AgCl electrode is used as a reference electrode, and a platinum wire electrode is used as a counter electrode. The preparation method of the working electrode comprises the following steps: 1) Preparation of a single-atom catalyst with Ni-N4 coordination: first, nickel acetate and a Schiff base ligand are mixed in a solvent to obtain a Ni(II) Schiff base ligand, then carbon black is added, and the mixture is uniformly mixed and heated to evaporate the solvent, so that the Ni(II) Schiff base ligand is fully adsorbed on the carbon black carrier to obtain a composite, then the obtained composite is placed in a tube furnace, sufficient argon is introduced to remove air, and pyrolysis is performed to obtain a single-atom catalyst with Ni-N4 coordination; 2) Preparation of an electrochemical biosensor: the single-atom catalyst prepared in step 1) is dispersed in a mixture of double-distilled water, ethanol and Nafion solution, and ultrasonic dispersion is performed to obtain a dispersion liquid, the obtained dispersion liquid is uniformly coated on the surface of a glassy carbon electrode, and after drying, a working electrode is obtained, and an electrochemical biosensor is further prepared.

2. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 1), the molar ratio of the nickel acetate to the Schiff base ligand is 1:1-2; and the Schiff base ligand is 1,10-phenanthroline.

3. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 1), the solvent is ethanol or acetone, and the mass-volume ratio of the nickel acetate to the solvent is 1.0-2.5 mg / mL.

4. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 1), the mass ratio of the carbon black to the nickel acetate is 1:10-20.

5. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, Step 1) the heating evaporation temperature is 60~80 o C, heating evaporation time is 4~6h.

6. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 1), the pyrolysis process conditions are as follows: pyrolysis at 300-600℃ for 2-4h.

7. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 2), the volume ratio of double-distilled water, ethanol and Nafion solution in the mixture is 79:20:1, wherein the concentration of the Nafion solution is 5wt%.

8. Use of the electrochemical biosensor according to claim 1 for the detection of dopamine, characterized in that, In step 2), the mass-volume ratio of the single-atom catalyst to the mixture of double-distilled water, ethanol and Nafion solution is 2-4 mg / mL.

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