Electrochemical biosensor electrode for selectively detect dopamine, electrochemical biosensor including the same and manufacturing method thereof

Abstract

The present invention relates to an electrode for an electrochemical biosensor for selectively detecting dopamine, an electrochemical biosensor comprising the same, and a method for manufacturing the same. The electrode for an electrochemical biosensor for selectively detecting dopamine has a detection limit of trace amounts compared to existing dopamine detection sensors, and possesses excellent stability, reproducibility, and selectivity. The invention also relates to an electrochemical biosensor comprising the same and a method for manufacturing the same.

Description

Technology Field

[0001] The present invention relates to an electrode for an electrochemical biosensor for selectively detecting dopamine, an electrochemical biosensor comprising the same, and a method for manufacturing the same. The electrode for an electrochemical biosensor for selectively detecting dopamine has a detection limit of trace amounts compared to existing dopamine detection sensors, and possesses excellent stability, reproducibility, and selectivity. The invention also relates to an electrochemical biosensor comprising the same and a method for manufacturing the same. Background Technology

[0003] Dopamine is a substance belonging to the catecholamine and phenylethylamine families that plays an important role in the central nervous system, cardiovascular system, kidneys, and hormonal system, and serves as an indicator of human metabolism. Abnormal dopamine levels in the human brain (too low or too high) can cause neurodegenerative diseases such as Alzheimer's and Parkinson's disease.

[0004] Therefore, there is a need to develop a simple, selective, and sensitive method to check dopamine levels in the human body.

[0005] Pain is a common symptom in patients with Parkinson's disease, and paracetamol has been frequently used to alleviate this pain. Both paracetamol and dopamine are electrochemically active substances and are essential for human metabolism because they are often found together in the extracellular fluid of the central nervous system. Additionally, tyrosine, a non-essential amino acid, is reported to be a precursor for many neurotransmitters such as dopamine, thyroxine, and epinephrine. Tyrosine plays a significant role in Alzheimer's and Parkinson's diseases, and studies indicate that tyrosine supplementation can increase norepinephrine levels and minimize orthostatic hypotension in patients with Parkinson's disease. Since drug treatment for Parkinson's disease often requires biomolecules such as dopamine, paracetamol, and tyrosine, it is also important to establish a simple and sensitive electrochemical method capable of measuring these substances simultaneously.

[0006] Meanwhile, electrochemical sensors are widely used for the detection and analysis of chemical substances, playing a significant role in biological and medical applications, particularly due to their high sensitivity and selectivity. The demand for electrochemical sensors is continuously increasing in various fields, such as environmental monitoring, food safety assessment, drug development, and biosensing.

[0007] As is known, the electrochemical properties of electrodes are influenced by electronic conductivity and a large active surface area. Therefore, to improve the electrochemical properties of sensors, designing composite materials by combining highly electrically conductive electrocatalytic materials with highly electrically conductive materials that modify the electrode surface is emerging as an effective strategy.

[0008] Due to the rapid growth of nanotechnology, various nanomaterials are currently being used to fabricate electrodes for electrochemical sensing applications. Among these diverse nanomaterials, transition metal nanoparticles such as platinum (Pt), gold (Au), and palladium (Pd) are receiving particular attention due to their excellent electrocatalytic activity and surface area. While platinum-based nanoparticles are currently the most widely used, they have the disadvantages of low scarcity and high cost.

[0009] In this regard, recent research suggests that palladium-based nanoparticles can be a viable alternative to platinum and gold for developing electrochemical applications due to their highly heterogeneous catalytic activity, low cost, and non-toxicity. To enhance the performance of palladium-based nanoparticles and reduce palladium usage, it is necessary to select a suitable support matrix that is stable, inexpensive, and synergistic; the most efficient method is to add metal oxides to the catalyst to improve electrocatalytic activity through a dual-function mechanism. Prior art literature

[0011] Korean Published Patent No. 10-2011-0000647 (Publication Date: 2011.01.04) The problem to be solved

[0012] The present invention was devised to solve the above-mentioned problems and aims to provide an electrode for an electrochemical biosensor for selectively detecting dopamine, which exhibits high performance in detecting biochemical substances such as dopamine and can be utilized in medical diagnosis, such as measuring the concentration of specific drugs in the blood, and which can play an important role in disease diagnosis and treatment through rapid and sensitive detection, an electrochemical biosensor including the same, and a method for manufacturing the same. means of solving the problem

[0014] To solve the above-mentioned problem, the electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention may include an electrode and reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on a surface formed on one side of the electrode.

[0015] In a preferred embodiment of the present invention, the palladium-cerium oxide nanocomposite may have an average particle size of 7 to 10 nm.

[0016] In a preferred embodiment of the present invention, the palladium-cerium oxide nanocomposite may have an average crystalline size of 6.6 to 10.6 nm.

[0017] In a preferred embodiment of the present invention, the electrode for the electrochemical biosensor of the present invention is 0.15 to 0.25 cm 2 It can have an electrochemical active surface area (EASA).

[0018] Meanwhile, the electrochemical biosensor for selectively detecting dopamine according to the present invention may include an electrode for the electrochemical biosensor for selectively detecting dopamine according to the present invention.

[0019] Furthermore, the method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention may include a first step of preparing an electrode and an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface, respectively, and a second step of applying the aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface to one side of the electrode and then drying to form reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface on one side of the electrode.

[0020] In a preferred embodiment of the present invention, an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface comprises: Step 1-1, preparing a graphene oxide dispersion, a solution containing a cerium oxide (CeO2) precursor, and a solution containing a palladium (Pd) precursor, respectively; Step 1-2, preparing a mixture by mixing the graphene oxide dispersion, the solution containing the cerium oxide (CeO2) precursor, and the solution containing the palladium (Pd) precursor, and adding ascorbic acid and a pH adjuster to the mixture to prepare a reaction mixture having a pH of 10 to 14; Step 1-3, preparing a reaction product by stirring and reacting the reaction mixture at a temperature of 120 to 160°C for 3 to 7 hours; Step 1-4, preparing reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface by drying the reaction product; and Step 1-4, preparing the reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface in methanol. It can be prepared by including steps 1-5, which involve dissolving and then ultrasonically treating to prepare an aqueous solution of reduced graphene oxide in which a palladium-cerium oxide nanocomposite is formed on the surface.

[0021] In a preferred embodiment of the present invention, in step 1-1, graphene oxide, cerium oxide (CeO2) precursor, and palladium (Pd) precursor may have a weight ratio of 1 : 0.47 to 0.72 : 0.58 to 0.88.

[0022] In a preferred embodiment of the present invention, a graphene oxide dispersion may be prepared by comprising the steps of: adding graphene powder, sodium nitrate (NaNO3), and potassium permanganate (KMnO4) to sulfuric acid (H2SO4) and stirring and reacting to produce a reaction product; adding hydrogen peroxide (H2O2) to the reaction product and filtering it, then washing the supernatant of the filter with ultrapure water to obtain a supernatant having a pH of 5 to 9; centrifuging the precipitate contained in the supernatant and drying it to produce graphene oxide; and adding the graphene oxide to ethylene glycol and ultrasonically treating it to produce a graphene oxide dispersion. Effects of the invention

[0024] The electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention, the electrochemical biosensor including the same, and the method for manufacturing the same provided high detection sensitivity and selectivity, and the high detection capability of the sensor can be usefully utilized in various fields such as detecting harmful substances in the environment, measuring the concentration of specific drugs in the blood, and detecting toxic substances in the food industry.

[0025] Furthermore, the electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention, the electrochemical biosensor including the same, and the method for manufacturing the same present the possibility of new sensor technology, which plays a role in technological advancement in the relevant field and can make a significant contribution to the technological development of related industries.

[0026] In addition, the electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention, the electrochemical biosensor including the same, and the method for manufacturing the same can be used for the detection of specific substances and the detection of biological activity in biomaterials because high sensing ability and selectivity can be equally applied to biological samples. Brief explanation of the drawing

[0028] Figure 1(A) shows the PXRD spectrum of reduced graphene oxide, b shows the PXRD spectrum of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Preparation Example 1, c shows the PXRD spectrum of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Preparation Example 2, and d shows the PXRD spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2. FIG. 1(B) shows the XPS full survey spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(C) shows the high-resolution XPS spectrum of C 1s in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(D) shows the high-resolution XPS spectrum of O 1s in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(E) shows the high-resolution XPS spectrum of Pd 3d in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, and FIG. 1(F) shows the high-resolution XPS spectrum of Ce 3d in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2. It shows the spectrum (high-resolution XPS spectrum). FIGS. 2(A) and FIGS. 2(B) are FESEM images of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 2(C) is an EDX spectrum of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 2(D) is a mapping image of C atoms, Pd atoms, and O atoms mixed in reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, a mapping image of C atoms, a mapping image of Pd atoms, and a mapping image of O atoms. Figures 2(E) and 2(F) are FESEM images of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, Figure 2(G) is an EDX spectrum of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, and Figure 2(H) is a mapping image of mixed C atoms, Ce atoms, and O atoms of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, a mapping image of C atoms, a mapping image of Ce atoms, and a mapping image of O atoms. Figures 2(I) and 2(J) are FESEM images of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, Figure 2(K) is an EDX spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, and Figure 2(L) is a mapping image of C atoms, O atoms, Pd atoms, and Ce atoms mixed in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, a mapping image of C atoms, a mapping image of Ce atoms, a mapping image of O atoms, a mapping image of Pd atoms. FIGS. 3(A) to FIGS. 3(B) are TEM images of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 3(C) is a TEM and HRTEM image of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, and FIG. 3(D) is a particle histogram of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1. FIGS. 3(E) to 3(F) are TEM images of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 3(G) is a TEM and HRTEM image of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, and FIG. 3(H) is a particle histogram of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2. Figure 4(A) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution. 3- / 4- In this graph, the scanning rate CV curves at 100 mV were measured for each of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE). Figure 4(B) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution. 3- / 4- In this graph, the scanning rate range CV curve of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 is measured and shown at 10 to 100 mV. Figure 4(C) is a graph showing a Tafel plot plotted between the peak potential and the logarithm of the current for each of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE), based on the results of a CV investigation. Figure 4(D) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution via electrochemical impedance spectroscopy (EIS). 3- / 4- This is a graph showing the Nyquist plots of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE). Figures 5(A) and 5(B) are graphs showing the results of cyclic voltammetry (CV) performed at 100 mV / s in 0.1 M PBS (pH 7) containing 1 mM dopamine to evaluate the electro-oxidation of dopamine in each of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE). Figure 5(C) is a graph showing the effect of pH on the electrochemical behavior of 1 mM dopamine in the dopamine detection electrode prepared in Example 1 through CV using 0.1 M PBS (pH 5–9) of various pH at 100 mV / s. Figure 5(D) is a plot showing the peak current and potential of the dopamine detection electrode prepared in Example 1 for dopamine oxidation in relation to various pH levels. Figure 6(A) is a graph showing the CV curves for various scan rates of 10 to 200 mV / s in 1 mM dopamine dissolved in 0.1 M sterile phosphate buffer (PBS) (pH 7) for the dopamine detection electrode prepared in Example 1. Figure 6(B) is a graph showing a plot between the positive and negative peak currents of dopamine and various sweep speeds for the dopamine detection electrode prepared in Example 1. FIG. 6(C) is a graph showing a plot between the log of the peak current and the log of the scan rate for the dopamine detection electrode prepared in Example 1, and FIG. 6(D) is a graph showing a plot between the log of the scan rate and the log of the peak transition for the dopamine detection electrode prepared in Example 1. Figure 7(A) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 at various concentrations of dopamine dissolved in 0.1M sterile phosphate buffer solution (PBS) (pH 7). Figure 7(B) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 in various concentrations of dopamine dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7), 5 μM paracetamol (PC), and 10 μM tyrosine (TY). FIG. 7(C) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 at various concentrations of paracetamol (PC), 5 μM dopamine (DA), and 10 μM tyrosine (TY) dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7). FIG. 7(D) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 in various concentrations of tyrosine (TY), 5 μM dopamine (DA), and 5 μM paracetamol (PC) dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7). Figures 7(E) and 7(F) are DPV curves showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 at various concentrations of dopamine (DA), tyrosine (TY), and paracetamol (PC) dissolved in 0.1M sterile phosphate buffer solution (PBS) (pH 7). FIGS. 8(A) and 8(B) show the results of evaluating the selectivity of the dopamine detection electrode prepared in Example 1, using 30 μM dopamine (DA) as well as Na as an interfering substance in 0.1 M sterile phosphate buffer (PBS) (pH 7). + , Al 3+ , AA (ascorbic acid), glucose, Zn 2+ and NO 3- This is a graph showing the results of performing DPV in this dissolved solution. FIG. 8(C) is a graph showing the DPV of each of the six dopamine detection electrodes, prepared by manufacturing six electrodes as in Example 1 to evaluate the reproducibility of the dopamine detection electrode. FIG. 8(D) is a graph showing the DPV of the dopamine detection electrode prepared in Example 1 before and after storage to evaluate the storage stability of the dopamine detection electrode. Figure 9(A) is a graph showing the results of performing DPV on pharmaceutical injection samples and dopamine (DA) added at different concentrations, and Figure 9(C) is a graph showing the results of performing DPV on human serum samples and dopamine (DA) added at different concentrations. Figure 9(B) shows the peak current (I) according to the amount of pharmaceutical injection sample added. pa Figure 9(D) is a graph showing a linear plot of peak current versus dopamine (DA), and Figure 9(D) is a graph showing a linear plot of peak current versus dopamine (DA) according to the amount of human serum sample added. Specific details for implementing the invention

[0029] Hereinafter, embodiments of the present invention are described in detail with reference to the attached drawings so that those skilled in the art can easily implement the present invention. The present invention may be embodied in various different forms and is not limited to the embodiments described herein. In the drawings, parts unrelated to the explanation have been omitted to clearly explain the present invention, and the same reference numerals are assigned to identical or similar components throughout the specification.

[0031] The electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention may comprise an electrode and reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on a surface formed on one side of the electrode.

[0032] At this time, the electrode may include various electrodes used in the industry as a working electrode, preferably a glassy carbon electrode, a rotating disk electrode (RDE), a rotating ring-disk electrode (RRDE), a screen-printed carbon electrode, a carbon nanotube film, or an ITO electrode, and more preferably a glassy carbon electrode.

[0033] Meanwhile, the palladium-cerium oxide nanocomposite of the present invention may have an average particle size of 7 to 10 nm, preferably an average particle size of 8 to 9 nm.

[0034] In addition, the palladium-cerium oxide nanocomposite may have an average crystal size of 6.6 to 10.6 nm, preferably 7.6 to 9.6 nm, and more preferably 8.1 to 9.1 nm.

[0035] Meanwhile, the electrode for the electrochemical biosensor for selectively detecting dopamine according to the present invention is 0.15 to 0.25 cm 2 The electrochemical active surface area (EASA) of, preferably 0.18 to 0.21 cm 2 It can have an electrochemical active surface area (EASA).

[0037] Furthermore, the electrochemical biosensor for selectively detecting dopamine according to the present invention may include the electrode for the electrochemical biosensor for selectively detecting dopamine described above.

[0039] Meanwhile, the method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention includes a first step and a second step.

[0040] The first step of the method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention may involve preparing an electrode and an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface, respectively. At this time, the electrode is as described above.

[0041] In addition, an aqueous solution of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface can be prepared by including steps 1-1 to 1-5.

[0042] Step 1-1 of the method for preparing an aqueous solution of reduced graphene oxide in which a palladium-cerium oxide nanocomposite is formed on the surface may involve preparing a graphene oxide dispersion, a solution containing a cerium oxide (CeO2) precursor, and a solution containing a palladium (Pd) precursor, respectively. At this time, the graphene oxide, the cerium oxide (CeO2) precursor, and the palladium (Pd) precursor may have a weight ratio of 1:0.47 to 0.72:0.58 to 0.88, preferably 1:0.53 to 0.66:0.65 to 0.81. If the weight ratio of the graphene oxide and the cerium oxide (CeO2) precursor is less than 1:0.47, there may be a problem with difficulty in manufacturing the desired palladium-cerium oxide nanocomposite, and if it exceeds 1:0.72, there may be an issue with economic feasibility. In addition, if the weight ratio of graphene oxide and palladium (Pd) precursor is less than 1:0.58, there may be a problem with the yield of the palladium-cerium oxide nanocomposite being produced decreasing, and if it exceeds 1:0.88, there may be an economic problem.

[0043] In addition, the cerium oxide (CeO2) precursor may include one or more selected from ceric ammonium nitrate, cerium(III) carbonate, and cerium(III) nitrate hexahydrate, and preferably may include ceric ammonium nitrate.

[0044] In addition, the palladium (Pd) precursor may include one or more selected from palladium (II) acetate, tris(dibenzylideneacetone)dipalladium (O), palladium dipropionate, and palladium (II) acetylacetonate, and preferably may include palladium (Pd(CH3COO)2) acetate.

[0045] Meanwhile, a graphene oxide dispersion can be prepared by including the steps of: adding graphene powder, sodium nitrate (NaNO3), and potassium permanganate (KMnO4) to sulfuric acid (H2SO4) and stirring and reacting to produce a reaction product; adding hydrogen peroxide (H2O2) to the reaction product and filtering it, then washing the supernatant of the filter with ultrapure water to obtain a supernatant having a pH of 5 to 9, preferably 6 to 7; centrifuging the precipitate contained in the supernatant and drying it to produce graphene oxide; and adding the graphene oxide to ethylene glycol and ultrasonically treating it to produce a graphene oxide dispersion. At this time, if the pH of the supernatant is less than 5, there may be a problem with defects occurring in the reduced graphene oxide, and if it exceeds 9, there may be a problem with difficulty in utilizing it as an electrochemical biosensor.

[0046] In addition, specifically, the reactants can be prepared by carrying out a first reaction by stirring at a temperature of 0 to 5°C for 4 to 6 hours, and a second reaction by stirring at a temperature of 50 to 70°C, preferably 55 to 65°C, for 3 to 5 hours. If the temperature of the first reaction is below 0°C, there may be a problem of incomplete reaction, and if it exceeds 5°C, there may be a problem of significantly increased manufacturing time. Furthermore, by setting the temperature of the second reaction higher than that of the first reaction, the problem of incomplete reaction can be resolved.

[0047] In addition, when adding hydrogen peroxide (H2O2) to the reactants, the temperature of the reactants may be lowered to 18 to 26°C, preferably 20 to 24°C, before adding, and if the temperature is not lowered in this way, there may be a problem with unreacted substances.

[0048] In addition, drying can be performed at a temperature of 60 to 80°C.

[0049] Furthermore, a solution containing a cerium oxide (CeO2) precursor can be prepared by adding a cerium oxide precursor to ethylene glycol, and a solution containing a palladium (Pd) precursor can be prepared by adding a palladium precursor to ethylene glycol.

[0051] Step 1-2 of the method for preparing an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface involves mixing the graphene oxide dispersion prepared in Step 1-1, a solution containing a cerium oxide (CeO2) precursor, and a solution containing a palladium (Pd) precursor to prepare a mixture, and adding ascorbic acid and a pH adjuster to the prepared mixture to prepare a reaction mixture having a pH of 10 to 14, preferably 11 to 13. At this time, if the pH of the reaction mixture is less than 10, there may be a problem in that the rate of the reduction reaction slows down. In addition, the pH adjuster may include one or more selected from sodium hydroxide, potassium hydroxide, and calcium hydroxide.

[0053] Step 1-3 of the method for preparing an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface may produce a reaction product by stirring and reacting the reaction mixture prepared in Step 1-2 at a temperature of 120 to 160°C, preferably 130 to 140°C, for 3 to 7 hours, preferably 4 to 6 hours. If these manufacturing conditions are not met, there may be a problem in that the desired compound cannot be produced.

[0055] Step 1-4 of the method for preparing an aqueous solution of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface may produce reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface by drying the reaction product prepared in Step 1-3. At this time, drying may be performed at a temperature of 70 to 90°C, preferably 75 to 85°C, for 6 to 10 hours, preferably 7 to 9 hours.

[0057] Step 1-5 of the method for preparing an aqueous solution of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface involves dissolving the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface, prepared in Step 1-4, in methanol, and then performing ultrasonic treatment to prepare an aqueous solution of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface.

[0059] The second step of the method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine according to the present invention may involve applying an aqueous solution of reduced graphene oxide, prepared in the first step and having a palladium-cerium oxide nanocomposite formed on its surface, to one side of the electrode, and then drying to form reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface on one side of the electrode. At this time, drying may be performed in an oven at 40 to 60°C, preferably 45 to 55°C, for 2 to 20 minutes, preferably 7 to 13 minutes.

[0061] The present invention will be explained in more detail below through examples, but the following examples are not intended to limit the scope of the invention and should be interpreted as being for the purpose of aiding understanding of the invention.

[0063] Preparation Example 1: Preparation of graphene oxide

[0064] (1) 2g of graphite powder was added to 100ml of concentrated sulfuric acid (H2SO4) and sonicated for 1 hour. Then, sodium nitrate (NaNO3) and potassium permanganate (KMnO4) were added, and the first reaction was carried out by stirring at a temperature of 0 to 5℃ for 5 hours, and the second reaction was carried out by stirring at a temperature of 60℃ for 4 hours to prepare the reaction product.

[0065] (2) After lowering the temperature of the prepared reaction mixture to room temperature (22℃), a 30% w / v hydrogen peroxide (H2O2) solution was added and filtered, and the supernatant of the filtrate was washed with ultrapure water to obtain a supernatant with a pH of 7.

[0066] (3) The precipitate contained in the obtained supernatant was centrifuged at 8,000 rpm and dried at a temperature of 70°C to produce graphene oxide.

[0068] Preparation Example 2: Preparation of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface

[0069] (1) 70 mg of graphene oxide prepared in Preparation Example 1 was added to 40 ml of ethylene glycol and sonicated for 30 minutes to prepare a graphene oxide dispersion. Additionally, 41.65 mg of ceric ammonium nitrate was added to 30 ml of ethylene glycol and sonicated for 1 hour to prepare a solution containing 10 wt% of a cerium oxide (CeO2) precursor. Additionally, 51.25 mg of palladium acetate (Pd(CH3COO)2) was added to 30 ml of ethylene glycol and sonicated for 1 hour to prepare a solution containing 20 wt% of a palladium (Pd) precursor.

[0070] (2) A mixture was prepared by mixing a graphene oxide dispersion, a solution containing 10 wt% of a cerium oxide (CeO2) precursor, and a solution containing 20 wt% of a palladium (Pd) precursor, and 50 mg of ascorbic acid and a small amount of sodium hydroxide were slowly added to the mixture to maintain the pH, thereby preparing a reaction mixture with a pH of 12.

[0071] (3) The reaction mixture was stirred at room temperature (22℃) for 24 hours, and then refluxed and stirred at 140℃ for 5 hours to produce the reaction product.

[0072] (4) The reaction product was washed three times with an ethanol-water mixture (50% : 50%) and then dried at a temperature of 80°C for 8 hours to produce reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface.

[0074] Comparative Preparation Example 1: Preparation of reduced graphene oxide with palladium nanoparticles formed on the surface

[0075] (1) 70 mg of graphene oxide prepared in Preparation Example 1 was added to 40 ml of ethylene glycol and sonicated for 30 minutes to prepare a graphene oxide dispersion. In addition, 51.25 mg of palladium acetate (Pd(CH3COO)2) was added to 30 ml of ethylene glycol and sonicated for 1 hour to prepare a solution containing 20 wt% of palladium (Pd) precursor.

[0076] (2) A mixture was prepared by mixing a graphene oxide dispersion and a solution containing 20% ​​by weight of a palladium (Pd) precursor, and 50 mg of ascorbic acid and a small amount of sodium hydroxide were slowly added to the mixture to maintain the pH, thereby preparing a reaction mixture with a pH of 12.

[0077] (3) The reaction mixture was stirred at room temperature (22℃) for 24 hours, and then refluxed and stirred at 140℃ for 5 hours to produce the reaction product.

[0078] (4) The reaction product was washed three times with an ethanol-water mixture (50% : 50%) and then dried at a temperature of 80°C for 8 hours to produce reduced graphene oxide with palladium nanoparticles formed on the surface.

[0080] Comparative Preparation Example 2: Preparation of reduced graphene oxide with cerium oxide nanoparticles formed on the surface

[0081] (1) 70 mg of graphene oxide prepared in Preparation Example 1 was added to 40 ml of ethylene glycol and sonicated for 30 minutes to prepare a graphene oxide dispersion. Additionally, 41.65 mg of ceric ammonium nitrate was added to 30 ml of ethylene glycol and sonicated for 1 hour to prepare a solution containing 10 wt% of a cerium oxide (CeO2) precursor.

[0082] (2) A mixture was prepared by mixing a graphene oxide dispersion and a solution containing 10% by weight of a cerium oxide (CeO2) precursor, and 50 mg of ascorbic acid and a small amount of sodium hydroxide were slowly added to the mixture to maintain the pH, thereby preparing a reaction mixture with a pH of 12.

[0083] (3) The reaction mixture was stirred at room temperature (22℃) for 24 hours, and then refluxed and stirred at 140℃ for 5 hours to produce the reaction product.

[0084] (4) The reaction product was washed three times with an ethanol-water mixture (50% : 50%) and then dried at a temperature of 80°C for 8 hours to produce reduced graphene oxide with cerium oxide nanoparticles formed on the surface.

[0086] Experimental Example 1: Structural Analysis 1

[0087] To confirm the crystal phase and chemical phase composition of the reduced graphene oxide prepared in Preparation Example 1, the reduced graphene oxide prepared in Preparation Example 2 with a palladium-cerium oxide nanocomposite formed on its surface, the reduced graphene oxide prepared in Comparative Preparation Example 1 with palladium nanoparticles formed on its surface, and the reduced graphene oxide prepared in Comparative Preparation Example 2 with cerium oxide nanoparticles formed on its surface, powder X-ray diffraction (PXRD) analysis was performed. The PXRD analysis was performed using an X-ray diffraction analyzer (3003 TT) with Cu Kα radiation (λ = 1.54106 Å), and the results are shown in Figure 1(A) below.

[0088] Figure 1(A) shows the PXRD spectrum of reduced graphene oxide, b shows the PXRD spectrum of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Preparation Example 1, c shows the PXRD spectrum of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Preparation Example 2, and d shows the PXRD spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2.

[0089] As can be seen in a of Fig. 1(A), the PXRD plot shows a broad diffraction peak of 2θ with a value of about 24.6° for the (002) crystal plane, which is a peak representing reduced graphene oxide.

[0090] As can be seen in Fig. 1(A)b, the PXRD pattern corresponds to the face-centred cubic phase of palladium (Pd), and the diffraction peaks at 2θ values ​​of 39.77°, 46.20°, and 67.56° can be indexed to the (111), (200), and (220) planes, respectively.

[0091] As can be seen in c of Fig. 1(A), the PXRD pattern corresponds to the cubic fluorite structure of cerium oxide (CeO2), and the diffraction peaks at 2θ values ​​of 28.69°, 33.45°, 47.64°, 59.09°, and 69.40° can be indexed to the (111), (200), (220), (222), and (400) planes, respectively.

[0092] As can be seen in d of Fig. 1(A), the PXRD pattern shows diffraction peaks at 2θ values ​​of 39.37°, 46.20°, and 67.12° due to palladium (Pd) (indexed to the (111), (200), and (220) planes, respectively) and diffraction peaks at 2θ values ​​of 28.96°, 33.42°, and 59.10° due to cerium oxide (CeO2) (indexed to the (111), (200), and (222) planes, respectively).

[0093] Meanwhile, when calculating the average crystal size, the palladium-cerium oxide nanocomposite on the surface was calculated to be 8.6 nm, the palladium nanoparticles 11.2 nm, and the cerium oxide nanoparticles 11.9 nm.

[0095] Experimental Example 2: Structural Analysis 2

[0096] XPS analysis was performed to confirm the chemical properties of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface, prepared in Preparation Example 2. XPS analysis was performed using an X-ray photoelectron microscope (XPS; Kratos, AXIS Supra, Manchester, UK) with a monochromatic Al-Kα X-ray source (15 Kv, 375 W), and the results are shown in Figures 1(B) to 1(F).

[0097] FIG. 1(B) shows the XPS full survey spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(C) shows the high-resolution XPS spectrum of C 1s in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(D) shows the high-resolution XPS spectrum of O 1s in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 1(E) shows the high-resolution XPS spectrum of Pd 3d in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, and FIG. 1(F) shows the high-resolution XPS spectrum of Ce 3d in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2. It shows the spectrum (high-resolution XPS spectrum).

[0098] As can be seen in Figure 1(B), the peaks appearing at binding energies of 283 eV, 530 eV, 330 eV, 560 eV, 900 eV, and 115.1 eV represent C 1s, O 1s, Pd 3d, Pd 3p, Ce 3d, and Ce 4d, respectively.

[0099] As can be seen in Fig. 1(C), a strong peak was observed for the binding energy of C 1s at 284.8 eV. This indicates that the CC / C=C of non-oxygenated carbon is dominant sp² in the reduced graphene oxide support. 2It exhibits a hybrid graphene structure and indicates a significant reduction in the oxygen-rich functional group. Additionally, peaks appeared at 286.2 eV and 288.8 eV, which are attributed to the epoxy and carbonyl groups, respectively.

[0100] As can be seen in Fig. 1(D), the binding energy of O 1s shows a peak at 532.1 eV, which represents the CO group. Additionally, the peak at 529.3 eV represents the Ce-O bond, which confirms that an interaction is occurring between graphene oxide and cerium oxide.

[0101] As can be seen in Fig. 1(E), in the Pd 3d spectrum, Pd 3d 5 / 2 and Pd 3d 3 / 2 Subpeaks of were observed, and such peaks at 338.1 eV and 343.2 eV can be attributed to Pd(II), and peaks at 335.8 eV and 341.2 eV indicate the presence of Pd(0).

[0102] As can be seen in Fig. 1(F), in the Ce 3d spectrum, Ce 3d 5 / 2 and Ce 3d 3 / 2 A peak group appeared, and the binding energy peaks at 883.8 eV, 886.4 eV, and 905.4 eV were Ce 3+ 3D 5 / 2 and Ce 3+ 3D 3 / 2 It is attributed to. Also, Ce 4+ 3D 3 / 2 corresponds to the binding energy peak at 902.2 eV. Therefore, Ce 4+ From Ce 3+The change in the valence state of the cerium oxide structure is attributed to oxygen vacancies, and this result shows that charge transfer from mobile holes in the graphene layer to localized electrons in the cerium oxide structure favorably affects the electrocatalytic properties of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2.

[0104] Experimental Example 3: Structural Analysis 3

[0105] Field emission scanning electron microscope (FESEM, S-4800) and mapping using energy dispersive X-rays (EDX) were performed on each of the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, the reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Preparation Example 1, and the reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Preparation Example 2, and are shown in Fig. 2.

[0106] FIGS. 2(A) and FIGS. 2(B) are FESEM images of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 2(C) is an EDX spectrum of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 2(D) is a mapping image of C atoms, Pd atoms, and O atoms mixed in reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, a mapping image of C atoms, a mapping image of Pd atoms, and a mapping image of O atoms.

[0107] Figures 2(E) and 2(F) are FESEM images of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, Figure 2(G) is an EDX spectrum of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, and Figure 2(H) is a mapping image of mixed C atoms, Ce atoms, and O atoms of reduced graphene oxide with cerium oxide nanoparticles formed on the surface prepared in Comparative Example 2, a mapping image of C atoms, a mapping image of Ce atoms, and a mapping image of O atoms.

[0108] Figures 2(I) and 2(J) are FESEM images of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, Figure 2(K) is an EDX spectrum of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, and Figure 2(L) is a mapping image of C atoms, O atoms, Pd atoms, and Ce atoms mixed in the reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on the surface prepared in Preparation Example 2, a mapping image of C atoms, a mapping image of Ce atoms, a mapping image of O atoms, a mapping image of Pd atoms.

[0109] As can be seen in Figs. 2(A) and 2(B), some spherical particles, which are palladium nanoparticles, were captured as bright colors on the surface of three-dimensional interconnected wrinkled reduced graphene oxide flakes. Additionally, as can be seen in Fig. 2(C), reduced graphene oxide with palladium nanoparticles formed on its surface, prepared in Comparative Example 1, was primarily observed. Furthermore, as can be seen in Figs. 2(C) and 2(D), the observed elements were carbon (C), oxygen (O), and palladium (Pd), and no other impurities were observed.

[0110] As can be seen in Figures 2(E) and 2(F), it was confirmed that cerium oxide nanoparticles were attached to the surface of the reduced graphene oxide flakes. Additionally, as can be seen in Figure 2(G), it was confirmed that reduced graphene oxide with cerium oxide nanoparticles formed on its surface was successfully prepared. Furthermore, as can be seen in Figures 2(G) and 2(H), the observed elements were carbon (C), oxygen (O), and cerium (Ce), and no other impurities were observed.

[0111] As can be seen in Figures 2(I) and 2(J), palladium-cerium oxide nanocomposites were observed on the surface of reduced graphene oxide. Additionally, as can be seen in Figures 2(K) and 2(L), the reduced graphene oxide prepared in Preparation Example 2, with palladium-cerium oxide nanocomposites formed on its surface, was found to contain carbon (C), oxygen (O), palladium (Pd), and cerium (Ce) without other impurities.

[0113] Experimental Example 4: Structural Analysis 4

[0114] Transmission electron microscopy (TEM, JEM-F200) was performed on reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2 and reduced graphene oxide with palladium nanoparticles formed on its surface prepared in Comparative Preparation Example 1, respectively, and is shown in Fig. 3.

[0115] FIGS. 3(A) to FIGS. 3(B) are TEM images of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, FIG. 3(C) is a TEM and HRTEM image of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1, and FIG. 3(D) is a particle histogram of reduced graphene oxide with palladium nanoparticles formed on the surface prepared in Comparative Example 1.

[0116] FIGS. 3(E) to 3(F) are TEM images of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, FIG. 3(G) is a TEM and HRTEM image of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2, and FIG. 3(H) is a particle histogram of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface prepared in Preparation Example 2.

[0117] As can be seen in Figures 3(A) to 3(C), it was confirmed that palladium nanoparticles were well immobilized on the reduced graphene oxide support. In addition, as can be seen in Figure 3(D), it was confirmed that the palladium nanoparticles had an average particle size of 10 to 12 nm.

[0118] As can be seen in FIGS. 3(E) to 3(G), it was confirmed that spherical palladium-cerium oxide nanocomposites were well immobilized on the reduced graphene oxide support. Additionally, as can be seen in the HRTEM image of FIG. 3(G), the reduced graphene oxide prepared in Preparation Example 2, with palladium-cerium oxide nanocomposites formed on its surface, exhibited lattice fringes of 0.28 nm and 0.34 nm, which were attributed to the palladium and cesium oxide nanocomposites, respectively. Furthermore, as can be seen in FIG. 3(H), it was confirmed that the palladium-cerium oxide nanocomposites had an average particle size of 8 to 9 nm.

[0120] Example 1: Preparation of a dopamine detection electrode

[0121] (1) A glassy carbon electrode was prepared as a working electrode. In addition, 1 mg of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface, prepared in Preparation Example 2, was dissolved in 1.0 ml of methanol and sonicated for 20 minutes to prepare an aqueous solution of reduced graphene oxide with a palladium-cerium oxide nanocomposite formed on its surface.

[0122] (2) A dopamine detection electrode was prepared by dropping 5.0 μl of an aqueous solution of reduced graphene oxide, on which a palladium-cerium oxide nanocomposite was formed on the surface of the prepared working electrode, and then drying it in an oven at 50°C for 10 minutes to form reduced graphene oxide, on which a palladium-cerium oxide nanocomposite was formed on the surface of the working electrode.

[0124] Comparative Example 1: Preparation of a dopamine detection electrode

[0125] (1) A glassy carbon electrode was prepared as a working electrode. In addition, 1 mg of reduced graphene oxide with palladium nanoparticles formed on its surface, prepared in Comparative Preparation Example 1, was dissolved in 1.0 ml of methanol and sonicated for 20 minutes to prepare an aqueous solution of reduced graphene oxide with palladium nanoparticles formed on its surface.

[0126] (2) A 5.0 μl aqueous solution of reduced graphene oxide with palladium nanoparticles formed on its surface was dropped onto one side of the working electrode, and then dried in an oven at 50°C for 10 minutes to form reduced graphene oxide with palladium nanoparticles formed on its surface on one side of the working electrode, thereby producing a dopamine detection electrode.

[0128] Comparative Example 2: Preparation of a dopamine detection electrode

[0129] (1) A glassy carbon electrode was prepared as a working electrode. In addition, 1 mg of reduced graphene oxide with cerium oxide nanoparticles formed on its surface, prepared in Comparative Preparation Example 2, was dissolved in 1.0 ml of methanol and sonicated for 20 minutes to prepare an aqueous solution of reduced graphene oxide with cerium oxide nanoparticles formed on its surface.

[0130] (2) A 5.0 μl aqueous solution of reduced graphene oxide with cerium oxide nanoparticles formed on the surface of the prepared working electrode was dropped onto one side of the working electrode and dried in an oven at 50°C for 10 minutes to form reduced graphene oxide with cerium oxide nanoparticles formed on the surface of the working electrode, thereby preparing a dopamine detection electrode.

[0132] Experimental Example 5: Electrochemical Analysis

[0133] A three-electrode system was prepared to perform electrochemical analysis on the dopamine detection electrodes prepared in Example 1 and Comparative Examples 1 and 2. Specifically, the dopamine detection electrode prepared in Example 1, the dopamine detection electrode prepared in Comparative Example 1, the dopamine detection electrode prepared in Comparative Example 2, and a glassy carbon electrode (GCE) were each used as working electrodes, a platinum wire (Pt wire) was used as a counter electrode, and a saturated Ag / AgCl electrode was used as a reference electrode.

[0135] (1) CV curve

[0136] Figure 4(A) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution. 3- / 4-In this graph, the scanning rate CV curves at 100 mV were measured for each of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE).

[0137] As can be seen in Fig. 4(A), the glassy carbon electrode (=bare GCE) exhibited a minimum response to peak current, and the maximum separation of the peak potential (△Ep) was approximately 115 mV, which was found to be insufficient to meet the requirements of a high-performance electrochemical sensor. Additionally, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2 was measured to have a maximum separation of the peak potential (△Ep) of approximately 105 mV, indicating superior electron transferability compared to the glassy carbon electrode (=bare GCE). Furthermore, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1 was measured to have a maximum separation of the peak potential (△Ep) of approximately 96 mV, which was determined to be a result of its excellent electronic conductivity. Finally, the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 was measured to have a maximum separation of peak potential (△Ep) of approximately 88 mV, confirming that it possesses the best electron transportability. In other words, it was confirmed that the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 exhibited the maximum augmentation of the redox peak current response.

[0138] Meanwhile, as a result of calculating the electrochemical active surface area (EASA), the dopamine detection electrode prepared in Example 1 was 0.194 cm² 2 , the dopamine detection electrode prepared in Comparative Example 1 is 0.097 cm 2 , the dopamine detection electrode prepared in Comparative Example 2 is 0.074 cm 2 , the glassy carbon electrode is 0.068 cm 2 It was calculated as follows. In conclusion, it was confirmed that the dopamine detection electrode prepared in Example 1 has a large electrochemical active surface area and excellent conductivity compared to other detection electrodes, thereby promoting electrochemical sensing performance for dopamine.

[0140] Figure 4(B) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution. 3- / 4- In this graph, the CV curve of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 is measured and shown in the scanning rate range of 10 to 100 mV. As can be seen in Fig. 4(B), as the scanning rate increases from 10 mV to 100 mV, the redox peak currents (I pa / I pc It was possible to confirm that ) increased linearly.

[0142] Figure 4(C) is a graph showing a Tafel plot plotted between the peak potential and the logarithm of the current for each of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1, the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and the glassy carbon electrode (=bare GCE), based on the results of a CV investigation. As can be seen in Fig. 4(C), the glassy carbon electrode (=bare GCE) exhibits very small peak separation, while the dopamine detection electrode prepared in Example 1 (=Pd-CeO2 / rGO / GCE) exhibits large peak separation, indicating that the absorption properties and electrocatalytic activity of the dopamine detection electrode prepared in Example 1 (=Pd-CeO2 / rGO / GCE) are superior to those of other dopamine detection electrodes. Furthermore, the high potential shift of the dopamine detection electrode prepared in Example 1 (=Pd-CeO2 / rGO / GCE) indicates excellent electrical conductivity.

[0144] (2) Electrochemical Impedance Spectroscopy (EIS)

[0145] Figure 4(D) shows 1.0 mM [Fe(CN)6] in a 0.1 M KCl solution via electrochemical impedance spectroscopy (EIS). 3- / 4- This graph shows the Nyquist plots of the dopamine detection electrode prepared in Example 1 (=Pd-CeO2 / rGO / GCE), the dopamine detection electrode prepared in Comparative Example 1 (=Pd / rGO / GCE), the dopamine detection electrode prepared in Comparative Example 2 (=CeO2 / rGO / GCE), and the glassy carbon electrode (=bare GCE), respectively. A Nyquist plot represents the charge transfer resistance (R ct; charge transfer resistance), double-layer capacitance (C dI ; double layer capacitance), Warburg resistance (Z W ; Warburg resistance), solution resistance (R s A Randles equivalent circuit composed of ; solution resistance) was used. As can be seen in Fig. 4(D), the glassy carbon electrode (=bare GCE) has a charge transfer resistance (R ct The value was measured to be 235 Ω, indicating a larger semicircular diameter compared to other detection electrodes. This result demonstrates that the glassy carbon electrode (=bare GCE) has the lowest active site, indicating poor electron mobility. Meanwhile, the charge transfer resistance (R) of the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 ct The value of ) is 88 Ω, and the charge transfer resistance (R) of the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1 is 88 Ω. ct The value of ) is 191 Ω, and the charge transfer resistance (R) of the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2. ct The values ​​were measured as 205 Ω, respectively. Therefore, the lowest charge transfer resistance (R ct It was confirmed that the dopamine detection electrode prepared in Example 1, having a value of ), had the best electron transfer kinetics.

[0146] In conclusion, since the dopamine detection electrode prepared in Example 1 has low impedance, a high electrochemical active surface area (EASA), and excellent conductivity, it was confirmed that the biosensor containing the dopamine detection electrode prepared in Example 1 is highly suitable for electrochemical applications.

[0148] (3) Electrochemical behavior of dopamine

[0149] To evaluate the electro-oxidation of dopamine in each of the dopamine detection electrode prepared in Example 1 (=Pd-CeO2 / rGO / GCE), the dopamine detection electrode prepared in Comparative Example 1 (=Pd / rGO / GCE), the dopamine detection electrode prepared in Comparative Example 2 (=CeO2 / rGO / GCE), and the glassy carbon electrode (=bare GCE), cyclic voltammetry (CV) was performed at 100 mV / s in 0.1 M PBS (pH 7) containing 1 mM dopamine, and the results are shown in Figures 5(A) and 5(B). As can be seen in Figure 5(A), no peak current response was observed in the glassy carbon electrode without dopamine (=GCE without DA), which indicates that a Faraday reaction did not occur on the surface of the glassy carbon electrode. A reversible redox peak current response was observed in the glassy carbon electrode (=bare GCE). However, compared to other detection electrodes, the glassy carbon electrode (=bare GCE) showed the lowest peak current response, indicating that the conductivity was reduced due to the lowest active site. The reason the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2 showed a higher peak current response than the glassy carbon electrode (=bare GCE) is that oxygen vacancies were present in the cerium oxide nanoparticles and reduced graphene oxide was introduced. On the other hand, the reason the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1 showed a higher peak current response than the glassy carbon electrode (=bare GCE) is due to the synergistic effect between the palladium nanoparticles and the reduced graphene oxide. Meanwhile, the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 was found to have a superior peak current response compared to other detection electrodes.Specifically, as can be seen in Fig. 5(B), it was confirmed that the dopamine detection electrode (=Pd-CeO2 / rGO / GCE) prepared in Example 1 showed a peak current response 2.5 times higher than the dopamine detection electrode (=Pd / rGO / GCE) prepared in Comparative Example 1, 3.6 times higher than the dopamine detection electrode (=CeO2 / rGO / GCE) prepared in Comparative Example 2, and 7 times higher than the glassy carbon electrode (=bare GCE).

[0150] Meanwhile, the effect of pH on the electrochemical behavior of 1 mM dopamine in the dopamine detection electrode prepared in Example 1 was measured by using CV with 0.1 M PBS (pH 5–9) of various pH values ​​at 100 mV / s, and the results are shown in Fig. 5(C). As can be seen in Fig. 5(C), as the pH gradually increased, the oxidation peak potential shifted to a less positive potential. When the pH dropped below 7, both the anode and cathode peak potentials shifted toward the anode due to strong electrostatic repulsion between the anode surface and the positively charged dopamine. Additionally, when the pH increased above 7, both the anode and cathode peak potentials shifted toward the cathode due to strong electrostatic repulsion between the cathode surface and the positively charged dopamine. In conclusion, it was confirmed that the dopamine detection electrode prepared in Example 1 is affected by the pH of the electrolyte during the oxidation of dopamine.

[0151] FIG. 5(D) is a plot showing the peak current and potential of the dopamine detection electrode prepared in Example 1 for dopamine oxidation in relation to various pH levels. As can be seen in FIG. 5(D), the oxidation peak (I pa In the plot of pH versus pH, the maximum peak current of dopamine was observed at pH 7.0, confirming that pH 7.0 is the optimal pH.

[0152] Meanwhile, for the dopamine detection electrode prepared in Example 1, CV curves for various scan rates of 10 to 200 mV / s in 1 mM dopamine dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7) are shown in Fig. 6(A). As can be seen in Fig. 6(A), it was observed that the peak current response of dopamine gradually increased as the scan rate increased. In addition, Fig. 6(B) is a graph showing a plot between the anode and cathode peak currents of dopamine and various sweep rates for the dopamine detection electrode prepared in Example 1, and as can be seen in Fig. 6(B), it was observed that the electrooxidation of dopamine using the dopamine detection electrode prepared in Example 1 was controlled by the adsorption process. Furthermore, FIG. 6(C) is a graph showing a plot between the logarithm of the peak current and the logarithm of the scan rate for the dopamine detection electrode prepared in Example 1, and FIG. 6(D) is a graph showing a plot between the logarithm of the scan rate and the logarithm of the peak transition for the dopamine detection electrode prepared in Example 1. As can be seen in FIG. 6(C) and FIG. 6(D), it was confirmed that dopamine exhibits rapid electron transfer kinetics at the electrode interface through the dopamine detection electrode prepared in Example 1. Additionally, the amount of dopamine adsorbed on the surface of the dopamine detection electrode prepared in Example 1 is 1.34 mol / cm². 2 It is calculated as such, and through this, it was confirmed that the dopamine detection electrode prepared in Example 1 has excellent absorption.

[0154] (4) Electrochemical measurement of dopamine

[0155] The electrochemical characteristics of dopamine in the dopamine detection electrode prepared in Example 1 were measured using the DPV method (differential pulse voltammetry method) and are shown in Figure 7.

[0156] Figure 7(A) is a DPV curve showing the degree of dopamine detection by the dopamine detection electrode prepared in Example 1 at various concentrations of dopamine dissolved in 0.1M sterile phosphate buffered saline (PBS) (pH 7). As can be seen in Figure 7(A), it was observed that the dopamine peak current gradually increased as the concentration of dopamine (DA) increased. Meanwhile, based on these results, the limit of detection (LOD) of the dopamine detection electrode prepared in Example 1 was 0.69 μM, the limit of quantification (LOQ) was 2.31 μM, and the sensitivity was 0.894 μA / μM. -1 cm -2 It was calculated as such. This confirmed that it has a low detection limit and excellent sensitivity compared to existing dopamine detection biosensors.

[0157] Figure 7(B) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 in various concentrations of dopamine dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7), 5 μM paracetamol (PC), and 10 μM tyrosine (TY). As can be seen in Figure 7(B), the oxidation peak currents of paracetamol (PC) and tyrosine (TY) remained almost constant, confirming that paracetamol (PC) and tyrosine (TY) did not interfere with the detection of dopamine (DA).

[0158] Figure 7(C) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 in various concentrations of paracetamol (PC), 5 μM dopamine (DA), and 10 μM tyrosine (TY) dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7). As can be seen in Figure 7(C), the peak current response of paracetamol (PC) increased linearly with increasing concentration, and the oxidation peak currents of dopamine (DA) and tyrosine (TY) remained almost constant, confirming that the presence of dopamine (DA) and tyrosine (TY) did not interfere with the oxidation of paracetamol (PC).

[0159] Figure 7(D) is a DPV curve showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 in various concentrations of tyrosine (TY), 5 μM dopamine (DA), and 5 μM paracetamol (PC) dissolved in 0.1 M sterile phosphate buffer solution (PBS) (pH 7). As can be seen in Figure 7(D), the peak current response of tyrosine (TY) increased linearly with increasing concentration, and the oxidation peak currents of dopamine (DA) and paracetamol (PC) remained almost constant, confirming that the presence of dopamine (DA) and paracetamol (PC) did not interfere with the oxidation of tyrosine (TY).

[0160] From these results, it was confirmed that dopamine (DA), paracetamol (PC), and tyrosine (TY) could be detected individually using the dopamine detection electrode prepared in Example 1.

[0161] Figures 7(E) and 7(F) are DPV curves showing the degree of dopamine detection at the dopamine detection electrode prepared in Example 1 at various concentrations of dopamine (DA), tyrosine (TY), and paracetamol (PC) dissolved in 0.1M sterile phosphate buffer solution (PBS) (pH 7). As can be seen in Figures 7(E) and 7(F), when the concentrations of all three substances increased simultaneously, it was confirmed that the DPV oxidation peak currents increased linearly and separately for each.

[0162] FIGS. 8(A) and 8(B) show the results of evaluating the selectivity of the dopamine detection electrode prepared in Example 1, using 30 μM dopamine (DA) as well as Na as an interfering substance in 0.1 M sterile phosphate buffer (PBS) (pH 7). + , Al 3+ , AA (ascorbic acid), glucose, Zn 2+ and NO 3- This is a graph showing the results of performing DPV in this dissolved solution. As can be seen in Figures 8(A) and 8(B), it was confirmed that even if 100 times the amount of interfering material is included, it does not have a significant effect on the detection of dopamine (DA). In other words, the relative error of the signal is less than 6%, confirming that the dopamine detection electrode prepared in Example 1 has excellent selectivity for dopamine detection.

[0163] Furthermore, to evaluate the reproducibility of the dopamine detection electrode, six dopamine detection electrodes were prepared as in Example 1, and DPV was performed on each of the six dopamine detection electrodes, as shown in Fig. 8(C). As can be seen in Fig. 8(C), it was confirmed that the reproducibility was excellent with a relative standard deviation of 6.2%.

[0164] In addition, to evaluate the storage stability of the dopamine detection electrode, DPV of the dopamine detection electrode prepared in Example 1 before and after storage was performed and is shown in Fig. 8(D). As can be seen in Fig. 8(D), it was confirmed that it has high storage stability by maintaining 94.6% of the initial value even after 10 days.

[0166] (5) Detection of dopamine in biological samples

[0167] Figure 9 is a graph showing the results of performing DPV on pharmaceutical injection samples and human serum samples to evaluate the applicability of the dopamine detection electrode prepared in Example 1 to a biological environment. A commercial dopamine (DA) injection sample was diluted with an appropriate amount of 0.1 M PBS (pH 7.0), and a standard solution was also prepared as a dopamine injection solution of the same concentration. To ensure a concentration range suitable for dopamine detection prior to analysis, the human serum sample was diluted 100-fold with 0.1 M PBS (pH 7.0). Figure 9(A) is a graph showing the results of performing DPV on pharmaceutical injection samples and samples with dopamine (DA) added at different concentrations, and Figure 9(C) is a graph showing the results of performing DPV on human serum samples and samples with dopamine (DA) added at different concentrations. As can be seen in Figs. 9(A) and 9(C), the recovery rates in the pharmaceutical injection sample and human serum sample were measured to be 98.3–100.15%. Fig. 9(B) shows the peak current (I) according to the amount of pharmaceutical injection sample added. paFigure 9(B) is a graph showing a linear plot of peak current versus dopamine (DA), and Figure 9(D) is a graph showing a linear plot of peak current versus dopamine (DA) according to the amount of human serum sample added. As can be seen in Figures 9(B) and 9(D), it was confirmed that the dopamine detection electrode prepared in Example 1 is more sensitive and reliable in clinical and diagnostic applications.

[0169] Simple variations or modifications of the present invention can be easily implemented by those skilled in the art, and all such variations or modifications are considered to be included within the scope of the present invention.

Claims

Claim 1 Comprising an electrode; and reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on a surface formed on one side of the electrode; and 0.15 to 0.25 cm 2 Electrode for an electrochemical biosensor for selectively detecting dopamine, characterized by having an electrochemical active surface area (EASA). Claim 2 An electrode for an electrochemical biosensor for selectively detecting dopamine, characterized in that, in claim 1, the palladium-cerium oxide nanocomposite has an average particle size of 7 to 10 nm. Claim 3 An electrode for an electrochemical biosensor for selectively detecting dopamine, characterized in that, in claim 1, the palladium-cerium oxide nanocomposite has an average crystalline size of 6.6 to 10.6 nm. Claim 4 delete Claim 5 An electrochemical biosensor for selectively detecting dopamine comprising an electrode for the electrochemical biosensor of claim 1. Claim 6 A first step of preparing an electrode and an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface, respectively; and a second step of applying the aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface to one side of the electrode and then drying to form reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface on one side of the electrode; wherein a dopamine detection electrode is prepared, and the prepared dopamine detection electrode is 0.15 to 0.25 cm 2 A method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine, characterized by having an electrochemical active surface area (EASA). Claim 7 In claim 6, the aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface comprises: Step 1-1, preparing a graphene oxide dispersion, a solution containing a cerium oxide (CeO2) precursor, and a solution containing a palladium (Pd) precursor, respectively; Step 1-2, preparing a mixture by mixing the graphene oxide dispersion, the solution containing the cerium oxide (CeO2) precursor, and the solution containing the palladium (Pd) precursor, and adding ascorbic acid and a pH adjuster to the mixture to prepare a reaction mixture having a pH of 10 to 14; Step 1-3, preparing a reaction product by stirring and reacting the reaction mixture at a temperature of 120 to 160°C for 3 to 7 hours; and Step 1-4, preparing reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface by drying the reaction product. A method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine, characterized by comprising the first-fifth step of preparing an aqueous solution of reduced graphene oxide having a palladium-cerium oxide nanocomposite formed on its surface by dissolving the reduced graphene oxide, on which a palladium-cerium oxide nanocomposite is formed on its surface, in methanol, and then ultrasonically treating the solution. Claim 8 A method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine, characterized in that, in step 1-1 above, the graphene oxide, cerium oxide (CeO2) precursor, and palladium (Pd) precursor have a weight ratio of 1 : 0.47 ~ 0.72 : 0.58 ~ 0.

88. Claim 9 A method for manufacturing an electrode for an electrochemical biosensor for selectively detecting dopamine according to claim 7, wherein the graphene oxide dispersion comprises the steps of: adding graphene powder, sodium nitrate (NaNO3), and potassium permanganate (KMnO4) to sulfuric acid (H2SO4), stirring, and reacting to produce a reaction product; adding hydrogen peroxide (H2O2) to the reaction product, filtering, and then washing the supernatant of the filtered product with ultrapure water to obtain a supernatant having a pH of 5 to 9; centrifuging the precipitate contained in the supernatant and drying it to produce graphene oxide; and adding the graphene oxide to ethylene glycol and ultrasonically treating it to produce a graphene oxide dispersion.

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