Anti-interference electrocatalytic dopamine sensor based on nitrogen-doped graphyne and its construction method and application

Abstract

The present invention belongs to the field of nanofunctional materials and electrochemical sensors, and specifically relates to an anti-interference electrocatalytic dopamine sensor based on specific binding sites on the surface of nitrogen-doped graphyne, and its construction method and application. The construction method first prepares a HEE pyridine solution, and then obtains nitrogen-doped graphyne through coupling reaction and calcination; then the electrode is modified with nitrogen-doped graphyne to prepare an anti-interference electrocatalytic dopamine sensor. The sensor prepared by the present invention has excellent anti-interference performance; density functional theory directly calculates that the most matching binding site between dopamine and the surface of N-doped graphyne is sp-N; the effective binding of dopamine and sp-N directly eliminates the binding of other coexisting interfering components to the interface, thereby improving the anti-interference performance and selectivity of the sensor prepared by the present invention. The conductive properties of nitrogen-doped graphyne improve the sensitivity of the sensor to target detection.

Description

Technical Field

[0001] The present invention belongs to the field of nano-functional materials and electrochemical sensors, and particularly relates to an anti-interference electrocatalytic dopamine sensor based on nitrogen-doped graphyne, and a construction method and application thereof. Background Art

[0002] Dopamine is the most abundant catecholamine neurotransmitter in the brain. As a neurotransmitter, it regulates numerous physiological functions of the central nervous system. Therefore, research on dopamine is of crucial importance in neurophysiology, clinical medicine, and pharmaceutics. In recent years, highly sensitive and selective methods for dopamine detection have become a hot topic. Electrochemical analysis, with its advantages of high selectivity, sensitivity, and stability, has become a promising method for dopamine detection.

[0003] In the existing technology, since the test sample contains multiple coexisting interfering components and the interfering substances are all at very high concentrations, they can easily interfere with the detection. Therefore, it is difficult to directly detect the target substance dopamine in real and complex physiological media such as serum solutions, and the detection sensitivity needs to be improved. Summary of the Invention

[0004] In response to the problems existing in the existing technology, the present invention provides a method for constructing an interference-resistant electrocatalytic dopamine sensor based on nitrogen-doped graphyne. The sensor constructed by this method can be directly applied to the highly selective and sensitive detection of dopamine, a human disease marker, in complex physiological media (artificial sweat, fetal bovine serum, etc.).

[0005] The present invention also provides an interference-resistant electrocatalytic dopamine sensor obtained by using the above-mentioned construction method.

[0006] The present invention also provides a use of the above-mentioned anti-interference electrocatalytic dopamine sensor in detecting dopamine in a biological sample.

[0007] The technical solutions adopted by the present invention to achieve the above-mentioned purpose are as follows:

[0008] The present invention provides a method for constructing an interference-resistant electrocatalytic dopamine sensor based on nitrogen-doped graphyne, comprising the following steps:

[0009] (1) Monomer deprotection: Hexa(trimethylsilyl)ethynylbenzene was added to tetrahydrofuran, and nitrogen was passed through the reaction system to remove oxygen. A THF solution of tetrabutylammonium fluoride was added to carry out the reaction. The mixture was then extracted with a saturated NaCl solution, dried over anhydrous MgSO4, and the solvent was removed. The resulting HEB was dissolved in pyridine to obtain a HEB pyridine solution for later use.

[0010] (2) Coupling reaction: Pyridine was added to the pretreated copper foil, oxygen was removed from the reaction system, and HEB pyridine solution was slowly added to the reaction system under oil bath conditions to carry out coupling reaction; after the reaction was completed, ultrasonication was performed, and the copper foil was rinsed with pyridine, N,N-dimethylformamide and deionized water, and the suspension was collected and freeze-dried;

[0011] (3) Preparation of nitrogen-doped graphyne: graphyne and melamine are mixed to obtain a mixture, the mixture is dispersed in water to form a uniform solution, heated to react, and then freeze-dried; finally, the resultant is transferred to a tube furnace under an argon atmosphere to obtain nitrogen-doped graphyne;

[0012] (4) Electrode modification:

[0013] ① The glassy carbon electrode is polished in an alumina suspension, rinsed with ethanol and water, ultrasonicated, and placed aside for later use; ② Nitrogen-doped graphyne is dispersed in the ink and ultrasonicated to form a uniform mixed solution;

[0014] ③ Quickly drop the mixed solution onto the surface of the pretreated glassy carbon electrode and leave it for a while to form a uniform electrocatalytic sensing interface.

[0015] Furthermore, in step (1), the ratio of hexa(trimethylsilyl)ethynylbenzene to tetrahydrofuran is 5 mg:3 mL; the volume ratio of tetrahydrofuran to the THF solution of tetrabutylammonium fluoride is 60:1; the concentration of the THF solution of tetrabutylammonium fluoride is 1 M; and the volume ratio of tetrahydrofuran to pyridine is 3:1.

[0016] Furthermore, in step (1), the reaction is carried out in an ice bath at 0°C for 20 minutes.

[0017] Furthermore, in step (2), 100 mL of pyridine is added to 6-8 pieces of pretreated copper foil; the volume ratio of the pyridine to the HEB pyridine solution is 1:5.

[0018] Furthermore, in step (2), the temperature of the oil bath is 80° C., and the coupling reaction time is 72 h.

[0019] Furthermore, in step (3), the mass ratio of graphyne to melamine is 1:2; and the concentration of the mixture in water is 2 mg / mL.

[0020] Furthermore, in step (3), the heating reaction is carried out at 150° C. for 15 h; the heating temperature in the tubular furnace is 800° C. for calcination for 2 h; and the flow rate of the argon gas is 150 mL / min.

[0021] Furthermore, in step ②, the ink is composed of 0.3 mL of ethanol, 0.15 mL of water, and 60 µL of naphthol; the ratio of nitrogen-doped graphyne to ink is 3 mg:0.51 mL; and in step ③, the placement time is 24 h.

[0022] The present invention also provides an interference-resistant electrocatalytic dopamine sensor prepared by the above-mentioned construction method.

[0023] The present invention also provides an application of the above-mentioned anti-interference electrocatalytic dopamine sensor in directly detecting dopamine in a biological sample.

[0024] The copper foil used in the present invention is in three sections of I-shape. The copper foil is ultrasonically treated with 0.1 M dilute hydrochloric acid and ethanol for 20 minutes respectively, and then quickly blown dry with a large flow of nitrogen gas to obtain the pretreated copper foil. The diameter of the glassy carbon electrode used in the present invention is 3 mm.

[0025] The beneficial effects of the present invention are:

[0026] 1) The sensor fabricated in this invention exhibits excellent anti-interference performance. Density functional theory calculations directly indicate that the optimal binding site for dopamine on the surface of N-doped graphyne is sp-N. The effective binding of dopamine to sp-N eliminates the binding of other coexisting interfering components to the interface, thereby enhancing the anti-interference performance and selectivity of the sensor fabricated in this invention. The conductive properties of nitrogen-doped graphyne enhance the sensor's sensitivity to target detection.

[0027] 2) Thanks to its ultra-high selectivity and sensitivity, the sensor enables direct detection of dopamine in complex physiological media. Compared to detection results in phosphate buffer (0.2 M, pH 7.4), the sensor's detection range remains unchanged in complex physiological media, from 1.0 to 550 µM, with a minimum detection limit of 0.46 µM. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 Four different N-doped graphyne structures: a. N1-GDY (sp-N), b. N2-GDY (sp-N), c. Pyrrolic nitrogen-GDY, and d. Graphene-N-GDY.

[0029] Figure 2 It is a complex structure formed by the combination of sp-N and dopamine which has lost two protons;

[0030] Figure 3 This is a graph showing the results of the electrochemical sensor prepared in the present invention detecting dopamine in a phosphate buffer solution;

[0031] Figure 4Graphs showing the results of direct detection of dopamine by the electrochemical sensor prepared for the present invention in complex physiological media artificial sweat (a and b) and 50% (v / v) fetal bovine serum (c and d). DETAILED DESCRIPTION

[0032] The technical solution of the present invention is further explained and illustrated by means of specific embodiments below.

[0033] Example 1

[0034] (1) Preparation of nitrogen-doped graphyne nanomaterials:

[0035] (1) Synthesis of Graphdiyne:

[0036] ① Copper foil pretreatment: ultrasonically treat the copper foil with dilute hydrochloric acid and ethanol respectively, and quickly dry it with a large flow of nitrogen.

[0037] ② Monomer Deprotection: The monomer hexaethynylbenzene (HEB) should be prepared and used immediately. Before the coupling reaction, place 100 mg of HEB-TMS hexa(trimethylsilyl)ethynylbenzene in a 100 mL three-necked round-bottom flask. Add 60 mL of THF (tetrahydrofuran) and purge with nitrogen for 5 minutes to remove oxygen from the reaction system. Add 1 mL of a 1 M solution of tetrabutylammonium fluoride (TBAF) in THF. Incubate at 0°C for 20 minutes (on ice). Extract with saturated NaCl three times, dry over anhydrous MgSO₄, and remove the solvent on a rotary evaporator (below 40°C). Dissolve the resulting HEB in 20 mL of pyridine. Note: The entire operation must be performed in the dark and as quickly as possible to prevent HEB decomposition.

[0038] ③ Coupling Reaction: Add pretreated copper foil (6-8 pieces, I-shaped, three sections) and 100 mL of pyridine to a 250 mL three-necked round-bottom flask. Place the flask in an oil bath and purge nitrogen through a gas tube for 15 minutes to remove oxygen. Raise the oil temperature to 80°C. Slowly add the pyridine from HEB to the reaction system via a constant pressure funnel (allowing half an hour to complete the addition and wrapping the mixture in a black plastic bag). Allow to react for 72 hours. After the reaction, sonicate the flask and rinse with pyridine, N,N-dimethylformamide, and deionized water. Collect the suspension in centrifuge tubes (approximately 10 mL per tube) and freeze-dry.

[0039] (2) Preparation of nitrogen-doped graphyne:

[0040] 20 mg of gydnyle (GDY) was mixed with 40 mg of melamine and dispersed in 30 mL of water to form a homogeneous solution. The solution was then heated at 150°C in a Teflon-coated stainless steel autoclave for 15 hours. The solution was then dried overnight in a freeze-vacuum dryer. Finally, the resulting solution was transferred to a tube furnace and heated at 800°C for 2 hours under an argon atmosphere at 150 mL / min. This yielded nitrogen-doped GDY.

[0041] (2) Electrode modification:

[0042] ① Polish a glassy carbon electrode (3 mm diameter) in 1.0, 0.5, and 0.03 µm alumina suspensions, rinse with ethanol and water, and ultrasonicate. Set aside for later use.

[0043] ② Prepare ink: Add 60.0 µL of naphthol (5.0 wt.%) to 0.30 mL of ethanol and 0.15 mL of water.

[0044] ③ 3.0 mg of nitrogen-doped graphyne was dispersed in the ink prepared above and ultrasonically treated to form a uniform mixed solution.

[0045] ④ Take 7 μL of the mixed solution and quickly drop-coat it on the surface of the pretreated glassy carbon electrode and leave it for 24 h to form a uniform electrocatalytic sensing interface.

[0046] Effect embodiment

[0047] (I) Density functional theory calculations to find the best-matched binding site for dopamine and its mechanism of action:

[0048] After the N-doping process, four N-doped structures will appear on the surface of N-doped graphyne, such as Figure 1 As shown, they are a. N1-GDY (sp-N), b. N2-GDY (sp-N), c. Pyrrolic nitrogen-GDY and graphite-N-GDY:

[0049] from Figure 1 As can be seen in Figures ad, compared to the originally highly positively charged surface of graphyne, the surface charge of nitrogen-doped graphyne has been redistributed and become more uniform. The doped N atoms exhibit a greater net charge, which will facilitate the binding of dopamine, which has lost two protons, to the highly matching N-doped sites.

[0050] The inventors used density functional theory to obtain the optimal binding site of dopamine and nitrogen-doped graphyne interface as sp-N ( Figure 1 This is because sp-N (a) has the largest net charge and is suitable for the adsorption of dopamine that has lost two protons.

[0051] Density functional theory was also used to simulate the composite structure formed after sp-N and dopamine combined, as shown below Figure 2 This composite structure effectively eliminates the adsorption of other components and sites. This composite structure prevents other interfering components coexisting in complex physiological media from binding to the nitrogen-doped graphyne interface, thus ensuring the strong anti-interference performance of the sensing interface. This in turn improves the selectivity of dopamine detection and provides support for the detection of dopamine in complex media.

[0052] (II) Detection of dopamine in phosphate buffer solution:

[0053] First, the ability of the constructed N-doped graphyne sensor to directly detect the target substance dopamine in phosphate buffer solution (0.2 M, pH 7.4) was tested. The detection performance of dopamine in phosphate buffer solution (0.2 M, pH 7.4) was tested using electrochemical it curve testing technology (electrochemical workstation, 760E, Shanghai Chenhua). The applied voltage for the test was +0.45 V. A magnetic stirrer was used to keep the phosphate buffer solution stirred at a constant speed. The interval time for dopamine addition was 50 s. A 10 µL syringe was used for addition. The concentration of dopamine added showed a gradient increase. The test results are as follows Figure 3 As shown in a and b, the linear detection range of the sensor for dopamine in phosphate buffer solution is 1 to 550 μM, with a minimum detection limit of 0.46 μM, indicating a wide detection range and a low detection limit.

[0054] Detection of dopamine in complex physiological media:

[0055] The electrochemical sensor based on nitrogen-doped graphyne realizes the direct detection of dopamine in complex physiological media, as shown below Figure 4 1) Direct detection of dopamine in artificial sweat, the operation method is the same as the detection in phosphate buffer solution. Figure 4 As shown in a and b, the linear detection range in artificial sweat is still 1 to 550 μM, and the lowest detection limit is 0.46 μM. 2) Dopamine was directly detected in 50% (V / V) fetal bovine serum, and the operation method was the same as the detection in phosphate buffer above. Figure 4 As shown in c and d, the linear detection range in 50% (v / v) fetal bovine serum is still 1 to 550 µM, and the minimum detection limit is 0.46 µM.

Claims

1. Application of an anti-interference electrocatalytic dopamine sensor based on nitrogen-doped graphyne for direct detection of dopamine in biological samples, characterized in that: The biological sample is artificial sweat or fetal bovine serum; The construction of an interference-resistant electrocatalytic dopamine sensor based on nitrogen-doped graphyne includes the following steps: (1) Monomer deprotection: Hexa(trimethylsilyl)ethynylbenzene was added to tetrahydrofuran, and nitrogen was passed through the reaction system to remove oxygen. A THF solution of tetrabutylammonium fluoride was added to carry out the reaction. The mixture was then extracted with a saturated NaCl solution, dried over anhydrous MgSO4, and the solvent was removed. The resulting HEB was dissolved in pyridine to obtain a HEB pyridine solution for later use. (2) Coupling reaction: Pyridine was added to the pretreated copper foil, oxygen was removed from the reaction system, and HEB pyridine solution was slowly added to the reaction system under oil bath conditions to carry out coupling reaction; after the reaction was completed, ultrasonication was performed, and the system was rinsed with pyridine, N,N-dimethylformamide and deionized water, and the suspension was collected and freeze-dried; (3) Preparation of nitrogen-doped graphyne: graphyne and melamine are mixed to obtain a mixture, the mixture is dispersed in water to form a uniform solution, heated to react, and then freeze-dried; finally, the resultant is transferred to a tube furnace under an argon atmosphere to obtain nitrogen-doped graphyne; The heating reaction is carried out at 150° C. for 15 h; the heating temperature in the tube furnace is calcined at 800° C. for 2 h; the flow rate of the argon gas is 150 mL / min; (4) Electrode modification: ① Polish the glassy carbon electrode in alumina suspension, rinse with ethanol and water, ultrasonicate, and place it aside for later use; ② Dispersing nitrogen-doped graphyne in ink and ultrasonically treating it to form a uniform mixed solution; ③ Rapidly drop the mixed solution onto the pre-treated glassy carbon electrode surface and leave it for a while to form a uniform electrocatalytic sensing interface; In step ②, the ink is composed of 0.3 mL of ethanol, 0.15 mL of water, and 60 µL of naphthol; the ratio of nitrogen-doped graphyne to ink is 3 mg:0.51 mL; in step ③, the placement time is 24 h.

2. The use according to claim 1, characterized in that In step (1), the ratio of hexa(trimethylsilyl)ethynylbenzene to tetrahydrofuran is 5 mg:3 mL; the volume ratio of tetrahydrofuran to the THF solution of tetrabutylammonium fluoride is 60:1; the concentration of the THF solution of tetrabutylammonium fluoride is 1 M; and the volume ratio of tetrahydrofuran to pyridine is 3:

1.

3. The use according to claim 1 or 2, characterized in that In step (1), the reaction is carried out in an ice bath at 0°C for 20 min.

4. The use according to claim 1, characterized in that In step (2), 100 mL of pyridine is added to 6-8 pieces of pretreated copper foil; the volume ratio of the pyridine to the HEB pyridine solution is 1:

5.

5. The use according to claim 1 or 4, characterized in that In step (2), the temperature of the oil bath is 80° C., and the coupling reaction time is 72 h.

6. The use according to claim 1, characterized in that In step (3), the mass ratio of graphyne to melamine is 1:2; and the concentration of the mixture in water is 2 mg / mL.

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