Catalyst and detection system for luminol electrochemiluminescence reaction and preparation method thereof

Abstract

The present invention discloses a catalyst and detection system for luminol electrochemiluminescence reaction and a preparation method thereof. The detection system adopts a Fe-N-C single-atom catalyst modified electrode. In an alkaline electrolyte, H2O2 is used as a co-reactant, and an electrochemiluminescence test can be performed at an ultra-low potential (≤±0.2V), which can achieve a strong electrochemiluminescence reaction of the luminol-H2O2 system. Wherein, the preparation method of the Fe-N-C single-atom catalyst includes: dissolving Zn(NO3)2·6H2O and Fe(NO3)3·9H2O in methanol to obtain a first mixed solution; adding a methanol solution of 2-methylimidazole to the first mixed solution to obtain a second mixed solution; allowing the second mixed solution to undergo a mixed reaction at 55-65°C, and calcining the solid obtained by the reaction at 850-950°C under an inert atmosphere to obtain the Fe-N-C SACs single-atom catalyst.

Description

Technical Field

[0001] The invention belongs to the technical field of electrochemiluminescence reaction, and in particular to the technical field of catalysts for driving luminol electrochemiluminescence reaction at low potential. Background Art

[0002] Early diagnosis and early treatment are the most effective ways to prevent and treat tumors and reduce cancer mortality. Therefore, for a long time, finding a detection method that can sensitively and accurately diagnose tumor markers has become the focus of attention. Among them, the electrochemiluminescence (ECL) detection method has the advantages of simple instrumentation, convenient operation, and easy automation, and has been clinically applied to trace tumor marker detection.

[0003] According to the different ways of generating the excited state of the luminophore or the positive and negative ion free radicals, the ECL luminescence pathways can be divided into two types, namely the annihilation pathway and the co-reactant pathway. Compared with the annihilation pathway, the co-reactant introduced in the co-reactant pathway can overcome the limitation of the solvent phase potential and produce stronger ECL emission than the annihilation reaction. For example, the mature electrochemiluminescence immunoassay system Elecsys, which is based on ruthenium trispyridine (Ru(bpy) 3 ) 2+ ) is an electrochemiluminescent probe and TPA is a co-reactant. Through the co-reactant pathway, the detection of trace targets is achieved.

[0004] However, due to Ru(bpy 3 ) 2+ The slow kinetics of TPA on the electrode surface usually require a large voltage to generate enough free radical intermediates to produce ECL signals. Applying a large voltage will inevitably destroy the biological activity of the target, thereby reducing the sensitivity and specificity of the detection. 3 ) 2+ Luminol has become one of the most commonly used organic light emitters due to its high ECL quantum yield and low excitation potential, and is expected to be used in the clinical detection of tumor markers by ECL.

[0005] However, for the luminol ECL system, small molecule compounds are usually required as co-reactants to promote the oxidation of luminol by decomposing small molecules to produce reactive oxygen species (ROS), thereby enhancing the ECL performance of luminol. The reaction process is cumbersome and the reaction efficiency is low. Summary of the invention

[0006] In view of the defects of the prior art, the purpose of the present invention is to provide a method that can drive the luminol electrochemiluminescence reaction at an ultra-low potential (≤±0.2V). The present invention also provides a preparation method of a luminol electrochemiluminescence reaction catalyst and a detection system. The detection system can realize the luminol luminescence reaction at an ultra-low potential, and is expected to accurately, sensitively and efficiently detect tumor markers, and can effectively protect the biological activity of the target during the detection process.

[0007] The technical solution of the present invention is as follows:

[0008] A method for preparing a catalyst for luminol electrochemiluminescence reaction, comprising:

[0009] Will Zn(NO 3 ) 2 6H 2 O and Fe(NO 3 ) 3 9H 2 O is dissolved in methanol to obtain a first mixed solution;

[0010] adding a methanol solution of 2-methylimidazole to the first mixed solution to obtain a second mixed solution;

[0011] Allowing the second mixed solution to react at 55-65° C., and separating the resulting solid precipitate;

[0012] The solid precipitate is washed and dried, and then calcined in an inert atmosphere at a temperature of 850-950° C. for a time of 0.8-1.2 h to obtain the catalyst.

[0013] According to some preferred embodiments of the present invention, the Zn(NO 3 ) 2 6H 2 O and the Fe(NO 3 ) 3 9H 2 The mass ratio of O is (20~40):1.

[0014] According to some preferred embodiments of the present invention, in the first mixed solution, the Zn(NO 3 ) 2 6H 2 The concentration of O is 5-6 mg / mL.

[0015] According to some preferred embodiments of the present invention, the Zn(NO 3 ) 2 6H 2 The mass ratio of O to the 2-methylimidazole is 1:(1-1.2).

[0016] According to some preferred embodiments of the present invention, in the second mixed solution, the concentration of 2-methylimidazole is 6-7 mg / mL.

[0017] According to some preferred embodiments of the present invention, the drying temperature is 55-65°C.

[0018] The present invention further provides a catalyst Fe-NC SACs single-atom catalyst for luminol electrochemiluminescence reaction prepared by the above preparation method, wherein the catalyst does not contain nanoparticles and clusters and is a single-atom structure.

[0019] The present invention further provides an electrochemical detection system containing the above-mentioned catalyst for luminol electrochemiluminescence reaction, which can realize electrochemical luminescence (ECL) of luminol at an ultra-low potential (≤±0.2V), and the resulting luminescence signal is relatively strong.

[0020] According to some preferred embodiments of the present invention, the electrochemical detection system comprises: a glassy carbon electrode to which the luminol electrochemical detection catalyst is added, an electrolyte containing an acid-base buffer solution of luminol, and a small molecule co-reactant H 2 O 2 .

[0021] According to some preferred embodiments of the present invention, the method for preparing the detection system comprises:

[0022] Dispersing the luminol electrochemical detection catalyst in a mixed solution of ethanol, deionized water and perfluorosulfonic acid with a concentration of 4 to 6 wt % to obtain a catalyst dispersion;

[0023] Adding the catalyst dispersion dropwise onto the glassy carbon electrode to form a uniform film, thereby obtaining the glassy carbon electrode to which the luminol electrochemical detection catalyst is added;

[0024] The glassy carbon electrode is added to an electrolyte to obtain a reaction system; wherein the electrolyte is a BR buffer solution containing luminol, the pH of which is 10, and the concentration of luminol is 0.1 to 0.15 mM;

[0025] Add the coreactant H to the reaction system 2 O 2 , and obtain the detection system.

[0026] The present invention has the following beneficial effects:

[0027] The present invention can obtain a Fe-NC single-atom catalyst, which exhibits significant H 2 O 2Reducing ability can produce a large amount of reactive oxygen species (ROS) in the reaction, accompanied by a strong ECL signal, which is suitable for high-precision detection of markers.

[0028] The detection system of the present invention can use the obtained catalyst as a highly efficient co-reaction catalyst to react with a small molecule compound H 2 O 2 As a co-reactant, by decomposing H 2 O 2 The generation of ROS such as hydroxyl radicals, superoxide anion radicals and singlet oxygen promotes the oxidation of luminol, thereby enhancing the ECL performance of luminol.

[0029] In the detection system of the present invention, the catalyst can reduce H at an ultra-low potential of 0 to -0.2 V due to its unique electronic structure. 2 O 2 A large amount of reactive oxygen species is generated, which then oxidizes the luminol anion into luminol anion radicals. This process avoids the cumbersome electrochemical oxidation process of luminol. The luminol anion radicals formed in situ will then directly react with the reactive oxygen species to produce a strong ECL signal. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is the spherical aberration scanning transmission electron microscope image of the Fe-NC SACs single atom catalyst in Example 1.

[0031] Figure 2 The Fourier transform EXAFS spectra of Fe in the Fe-NC SACs single atom catalyst, iron foil (Fe foil) and iron porphyrin (FePc) in Example 1.

[0032] Figure 3 The Fe-NC SACs single atom catalyst, NC modified glassy carbon electrode and bare glassy carbon electrode in Example 1 were subjected to luminol / H 2 O 2 ECL intensity in the ECL system.

[0033] Figure 4 The Fe-NC SACs single atom catalyst, NC-modified GCE and bare GCE in Example 1 were prepared under luminol-H 2 O 2 Cyclic voltammetry curves in the ECL system.

[0034] Figure 5 The Fe-NC SACs / GCE electrode in Example 1 is 2 With / without H in the ECL system 2 O 2CV curve of.

[0035] Figure 6 The Fe-NC SACs-luminol-H 2 O 2 The stability of the system. DETAILED DESCRIPTION

[0036] The present invention is described in detail below in conjunction with the embodiments and drawings, but it should be understood that the embodiments and drawings are only used to exemplify the present invention and do not constitute any limitation on the protection scope of the present invention. All reasonable changes and combinations within the scope of the inventive concept of the present invention fall within the protection scope of the present invention.

[0037] According to the technical solution of the present invention, some specific methods for preparing luminol electrochemical detection catalysts include the following process:

[0038] Will Zn(NO 3 ) 2 6H 2 O and Fe(NO 3 ) 3 9H 2 O is dissolved in methanol to obtain a first mixed solution;

[0039] adding a methanol solution of 2-methylimidazole to the first mixed solution to obtain a second mixed solution;

[0040] stirring the second mixed solution at 55-65° C., and collecting the resulting solid precipitate;

[0041] After the solid precipitate is washed and dried, it is calcined in an inert atmosphere at a calcination temperature of 850-950° C. for a calcination time of 0.8-1.2 h to obtain the catalyst Fe-NC SACs.

[0042] In some specific embodiments, Zn(NO 3 ) 2 6H 2 O and Fe(NO 3 ) 3 9H 2 The mass ratio of O is (20~40):1.

[0043] In some preferred embodiments, in the first mixed solution, Zn(NO 3 ) 2 6H 2 The concentration of O is 5-6 mg / mL.

[0044] In some preferred embodiments, Zn(NO 3 )2 6H 2 The mass ratio of O to 2-methylimidazole is 1:(1-1.2).

[0045] In some preferred embodiments, in the second mixed solution, the concentration of 2-methylimidazole is 6-7 mg / mL.

[0046] In some preferred embodiments, the stirring time of the second mixed solution is 20 to 28 hours.

[0047] In some preferred embodiments, the washing uses ethanol.

[0048] In some preferred embodiments, the drying is vacuum drying at 55-65°C.

[0049] In some preferred embodiments, the inert atmosphere is provided by nitrogen.

[0050] According to the technical solution of the present invention, some specific detection systems for detecting tumor markers using the obtained luminol electrochemical detection catalyst include:

[0051] A glassy carbon electrode containing the luminol electrochemical detection catalyst, an electrolyte containing luminol and a BR buffer solution, and a small molecule co-reactant H 2 O 2 .

[0052] The BR buffer is composed of NaH 2 PO 4 , H 3 BO 3 and CH 3 COONa was prepared and the buffer was adjusted to pH 10.

[0053] In some preferred embodiments, the preparation of the detection system comprises:

[0054] The prepared Fe-NC SACs catalyst was ultrasonically dispersed in a mixed solution of ethanol, deionized water and perfluorosulfonic acid (nafion) to obtain a catalyst dispersion;

[0055] Adding the catalyst dispersion dropwise onto the glassy carbon electrode to form a uniform film, thereby obtaining the glassy carbon electrode to which the luminol electrochemical detection catalyst is added;

[0056] The glassy carbon electrode is transferred into an electrolytic cell, and an electrolyte is added, wherein the electrolyte is a BR buffer solution containing luminol, to obtain a reaction system;

[0057] Add the coreactant H to the reaction system 2 O 2 , and obtain the detection system.

[0058] In some preferred embodiments, the concentration of perfluorosulfonic acid (nafion) is 4 to 6 wt.%.

[0059] In some preferred embodiments, the concentration of luminol in the electrolyte is 0.1-0.15 mM.

[0060] In some preferred embodiments, the BR buffer is composed of 0.035-0.045 M NaH 2 PO 4 , H 3 BO 3 and CH 3 It was prepared by preparing COONa and adjusting the pH to 10 with NaOH solution.

[0061] The technical solution of the present invention is demonstrated below in conjunction with specific embodiments:

[0062] Example 1

[0063] The Fe-NC SACs catalyst was prepared by the following process:

[0064] 3.39 g Zn(NO 3 ) 2 6H 2 O and 100 mg Fe(NO 3 ) 3 9H 2 O was dissolved in 600 mL of methanol to obtain a mixed solution, to which 600 mL of a methanol solution containing 3.94 g of 2-methylimidazole was added, and the mixture was continuously stirred at 60 °C for 24 h. The precipitate was collected and washed with ethanol, dried in a vacuum oven at 60 °C, and then calcined in a tube furnace at 900 °C for 1 h under a nitrogen atmosphere to obtain a catalyst, which was marked as Fe-NC SACs.

[0065] The obtained catalyst was characterized by spherical aberration scanning transmission electron microscopy. Figure 1 As shown, it can be seen that there are no nanoparticles or clusters in the Fe-NC SACs, and there are atomic-scale light spots, indicating that there are atomically distributed Fe species in the Fe-NC SACs.

[0066] The obtained Fe-NC SACs single atom catalyst was characterized and compared with iron foil (Fe foil) and iron porphyrin (FePc) by EXAFS spectroscopy. The Fourier transform EXAFS spectrum of Fe is shown in the attached figure. Figure 2 As shown in Figure 2, it can be seen that the Fe k-edge EXAFS curves of Fe-NC SACs are There is no peak of Fe-Fe bond at , while the Fe k-edge EXAFS curve of FePc is about There is only one main peak at , which can be attributed to the first Fe-N shell. This indicates that there are Fe-N bonds but no Fe-Fe bonds in Fe-NC SACs, and Fe in Fe-NC SACs is atomically dispersed, which is consistent with the spherical aberration scanning transmission electron microscopy image data.

[0067] The electrochemiluminescence performance of the above catalysts was tested:

[0068] Preparation of catalyst solution: 2 mg of the prepared Fe-NC SACs catalyst was dispersed in 1 mL of a mixed solution of ethanol, deionized water and 5 wt.% nafion (volume ratio of 79:20:1), and ultrasonicated for 30 minutes to obtain a catalyst solution.

[0069] 3 μL of the obtained 2 mg / mL catalyst solution was evenly dropped onto the polished glassy carbon electrode to form a uniform film.

[0070] The prepared electrode was transferred to an electrolytic cell, and the electrolyte was a BR buffer solution containing 0.12 mM luminol, which was composed of 0.04 M NaH 2 PO 4 , H 3 BO 3 and CH 3 COONa was prepared and the pH was adjusted to pH 10 with 1 M NaOH solution.

[0071] The electrolytic cell was subjected to an electrochemiluminescence test. During the test: the electrochemiluminescence scanning rate was 100 mV / s, and the potential scanning range was 0 to -0.2 V. The voltage of the photomultiplier tube (PMT) was set to 400 V during the detection process.

[0072] According to the above test, the following Figure 3-6 The test curve is shown.

[0073] in,

[0074] As attached Figure 3 As shown, Fe-NC SACs act as luminol / H 2 O 2 The ECL signal was significantly enhanced when the co-reaction catalyst of the system was used. 2 O 2 The ECL effect of the system is much smaller than that of Fe-NC SACs. Moreover, when the scanning potential is only -0.2 V, Fe-NC SACs can achieve luminol / H2 O 2 The significant electrochemiluminescence signal of the ECL system indicates that Fe-NC SACs are efficient co-reaction catalysts.

[0075] As attached Figure 4 As shown in the cyclic voltammetry electrochemical test, the change trend of the current density of Fe-NC SACs / GCE, NC / GCE and bare GCE is consistent with the change of their ECL intensity, indicating that Fe-NC SACs have significant H 2 O 2 Activation ability.

[0076] As attached Figure 5 As shown, the Fe-NC SACs / GCE electrode was placed in N 2 Saturated electrolyte contains or does not contain H 2 O 2 By comparing the cyclic voltammetry curves of 2 O 2 N 2 In saturated electrolyte, there is an obvious reduction peak at about -0.05 V, indicating that Fe-NC SACs can significantly reduce H at this ultra-small potential. 2 O 2 Therefore, Fe-NC SACs can act as co-reaction catalysts to activate H 2 O 2 , luminol / H is achieved at a very low potential 2 O 2 Cathodoluminescence of the ECL system.

[0077] As attached Figure 6 As shown, Fe-NC SACs were used as co-catalysts to drive luminol-H 2 O 2 It exhibits excellent stability in electrochemiluminescence system.

[0078] The above embodiments are only preferred implementations of the present invention, and the protection scope of the present invention is not limited to the above embodiments. All technical solutions under the concept of the present invention belong to the protection scope of the present invention. It should be pointed out that for ordinary technicians in this technical field, improvements and modifications without departing from the principle of the present invention should also be regarded as the protection scope of the present invention.

Claims

1. An electrochemical detection system containing a catalyst for luminol electrochemiluminescence reaction, wherein include: A glassy carbon electrode containing a luminol catalyst for electrochemical detection, an electrolyte containing a luminol acid-base buffer solution, and a small molecule co-reactant H 2 O 2 The electrochemical detection system detects the marker to be detected by a luminol electrochemical luminescence reaction, wherein the preparation of the luminol electrochemical detection catalyst comprises: Will Zn(NO 3 ) 2 6H 2 O and Fe(NO 3 ) 3 9H 2 O is dissolved in methanol to obtain a first mixed solution; adding a methanol solution of 2-methylimidazole to the first mixed solution to obtain a second mixed solution; Allowing the second mixed solution to react at 55-65° C., and separating the resulting solid precipitate; The solid precipitate is washed and dried, and then calcined in an inert atmosphere at a temperature of 850-950° C. for a time of 0.8-1.2 h to obtain the catalyst; The Zn(NO 3 ) 2 6H 2 O and the Fe(NO 3 ) 3 9H 2 The mass ratio of O is (20-40):1; In the first mixed solution, the Zn(NO 3 ) 2 6H 2 The concentration of O is 5-6 mg / mL; The Zn(NO 3 ) 2 6H 2 The mass ratio of O to the 2-methylimidazole is 1:(1-1.2); In the second mixed solution, the concentration of 2-methylimidazole is 6-7 mg / mL.

2. The electrochemical detection system according to claim 1, It is characterized in that The drying temperature is 55-65°C.

3. A method for preparing the electrochemical detection system according to claim 1, It is characterized in that It includes: Dispersing the luminol electrochemical detection catalyst in a mixed solution of ethanol, deionized water and perfluorosulfonic acid with a concentration of 4 to 6 wt % to obtain a catalyst dispersion; Adding the catalyst dispersion dropwise onto the glassy carbon electrode to form a uniform film, thereby obtaining the glassy carbon electrode to which the luminol electrochemical detection catalyst is added; The glassy carbon electrode is added to an electrolyte to obtain a reaction system; wherein the electrolyte is a BR buffer solution containing luminol, the pH of which is 10, and the concentration of luminol is 0.1 to 0.15 mM; Add the coreactant H to the reaction system 2 O 2 , and obtain the electrochemical detection system.

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