CN110849813A - CuO-Cu2Preparation method and application of O/CM nanowire array heterostructure - Google Patents

Abstract

The invention belongs to the technical field of new nano materials, and particularly relates to CuO-Cu₂A preparation method and application of an O/CM nanowire array heterostructure. The preparation method comprises the following steps: mixing and stirring NaOH solution and ammonium persulfate solution to obtain clarified solution, placing the pretreated copper mesh in the mixed solution for soaking reaction, washing the product with distilled water, and air-drying to obtain Cu (OH)₂A precursor. Subsequently, the obtained Cu (OH)₂The precursor is placed in a tubular furnace to be calcined to obtain CuO-Cu₂An O/CM nanowire array. The nanowire array prepared by the invention has the advantages of large surface area, high active site density, good stability, high catalytic efficiency and the like, and is beneficial to generating a PEC signal.

Description

CuO-Cu2Preparation method and application of O/CM nanowire array heterostructure

Technical Field

The invention belongs to the technical field of new nano materials, and particularly relates to CuO-Cu₂A preparation method and application of an O/CM nanowire array heterostructure.

Background

The detection of tumor markers is of great significance for early clinical diagnosis and subsequent cancer recurrence. Prostate Specific Antigen (PSA) secreted by prostate epithelial cells is considered a classical biomarker for screening for prostate cancer, one of the most common malignancies in men. It is recognized that while a threshold level of PSA of 4 ng/mL is typically found in cancer patients, it is typically in the range of the threshold to 10 ng/mL. Therefore, there is a great need for an efficient method to achieve sensitive detection of PSA. To date, a number of techniques have been used to quantitatively detect PSA, such as fluoroimmunoassay, electrochemiluminescence, and colorimetric immunoassay. And electrochemical methods. Despite the enormous efforts to accurately and sensitively determine PSA, expensive equipment or time-consuming procedures remain disadvantages of these methods. Therefore, there is a need to develop a simple, rapid, low cost and specific method for efficiently detecting PSA.

Photoelectrochemical (PEC) bioanalysis has attracted considerable research interest as an emerging analytical method with low cost, simple instrumentation, short response times, simple preparation and good portability. More importantly, it has the inherent advantage of high sensitivity compared to conventional techniques, since the unique signal conversion scheme converts excitation light into electrical energy output, thereby significantly reducing background noise signals. The cathode photoelectrochemical biosensor based on the p-type semiconductor has high anti-interference capability, can eliminate false positive signals, and has high sensitivity. Recently, there has been a growing interest in self-powered PEC biosensors that operate without additional voltage using semiconductors, a unique technology that eliminates the general need for an external power source and has become a promising sensing strategy.

In general, the performance of PEC sensors depends on the material that is photoactive sensitive to light irradiation. Therefore, the materials used to construct the sensor are critical to achieving excellent PEC response. Cu₂O and CuO are two attractive p-type semiconductors, CuO and Cu due to their p-type semiconductor properties and appropriate band gap to enable good visible light harvesting, and their advantages of high abundance, low cost and non-toxic properties₂O is a promising material for use as a photocathode in PEC analysis. Even so, due to photo-corrosion of the light absorbing layer in the electrolyte, it is generally subjected to Cu in practice due to its poor stability₂O use, and the need to improve the CuO and Cu₂Photocatalytic activity of O. To overcome these limitations, heterostructures were synthesized that had better photocatalytic activity and stability than the individual phases.

Disclosure of Invention

The invention aims to provide CuO-Cu₂The preparation method of the O/CM nanowire array heterostructure has the advantages of large surface area, high active site density, good stability, high catalytic efficiency and the like, and is beneficial to generating a PEC signal; the invention also provides the CuO-Cu₂The application of the O/CM nanowire array heterostructure is used for a photoelectrochemical biosensor for rapidly detecting PSA, and has good detection stability and low detection limit on PSA.

In order to achieve the purpose, the invention adopts the technical scheme that:

CuO-Cu₂The preparation method of the O/CM nanowire array heterostructure is characterized by comprising the following steps of:

(1) mixing and stirring a NaOH solution and an ammonium persulfate solution to obtain a clear solution, placing a pretreated copper net in the clear solution for soaking reaction, washing and drying an obtained sample to obtain Cu (OH)₂A precursor;

(2) mixing the Cu (OH) prepared in the step (1)₂Calcining the precursor to obtain CuO-Cu₂An O/CM nanowire array.

Further, the mass ratio of NaOH to ammonium persulfate in the step (1) is 3.2-3.5: 0.91-1.0; the mass-volume ratio of NaOH to water is (3.2-3.5) g: (20-25) mL; the mass volume ratio of the ammonium persulfate to the water is (0.91-1.0) g: (20-25) mL;

further, the water in the step (1) is deionized water; the washing step in the step (1) is washing with deionized water and ethanol respectively;

further, the drying temperature in the step (1) is 60 ℃; the calcining temperature in the step (2) is 350 ℃, and the calcining time is 15-20 minutes.

Further, the stirring time in the step (1) is 5-10 minutes; the soaking reaction time is 20-25 minutes.

A photoelectrochemical biosensor comprises a working electrode, a reference electrode and a counter electrode which are connected with an electrochemical workstation, wherein the CuO-Cu prepared by the method is modified on the working electrode₂An O/CM nanowire array.

The application of the photoelectrochemical biosensor is to sensitive detection of PSA.

Further, the application adopts the following steps to detect:

(1) dropping 0.1% chitosan on the surface of the photoelectrode, drying in air at 60 ℃, and fixing the aptamer CuO-Cu₂O/CM on electrode, incubated at 4 ℃ for 16h, washed with Tris-HCl buffer solution and then N₂Drying;

(2) the prepared photoelectrochemical biosensor was incubated with 1% BSA at room temperature for 1h, and the BSA/aptamer/CuO-Cu buffer solution was thoroughly washed with Tris-HCl buffer solution₂Storing the O/CM electrode for later use;

(3) and (3) incubating the sensor and PSA at 37 ℃ for 1h, irradiating under a xenon lamp, and detecting the PSA according to the change of photoelectric signals of the photoelectrochemical biosensor.

The aptamer is a DNA single chain, the single chain has a special sequence and can specifically capture a target, and the aptamer in the experiment is an aptamer of PSA (prostate specific antigen), namely the DNA single chain for capturing the PSA can be specifically identified. Heterojunction CuO-Cu in the invention₂The nanostructure of O is a nanowire, which has a large surface to volume ratio compared to nanoparticles, nanocubes or nanospheres, can provide a fast channel for carrier transfer, thereby inhibiting carrier recombination and improving the activity of the PEC.

Advantageous effects

(1) CuO-C prepared by the inventionu₂The O/CM nanowire array has the advantages of large surface area, high active site density, good stability, high catalytic efficiency and the like, and is beneficial to generating a PEC signal.

(2) The novel photoelectrochemical biosensor for quickly detecting PSA prepared by the invention has good detection stability on PSA and lower detection limit of 0.33 ng.mL^-1。

In general, based on the use of CuO-Cu₂The O/CM nanowire array designed a simple PEC bioanalytical platform for monitoring PSA activity; experiments have demonstrated that the constructed photoelectrochemical biosensor platform is simple and economical and has high sensitivity, selectivity and reliability for PSA detection, a new universal PEC aptamer sensor that can be extended to detect other biological interactions of interest.

Drawings

FIG. 1 CuO-Cu prepared in example 1 of the present invention₂Schematic diagram of a photoelectrochemical biosensor with an O/CM nanowire array for detection of PSA;

FIG. 2, CuO-Cu₂Schematic diagram of O/CM nanowire array electron transfer process;

FIG. 3, (A) CuO-Cu prepared in example 1₂X-ray diffraction Spectroscopy (XRD) of O/CM; (B) CuO-Cu prepared in example 1₂Scanning Electron Micrographs (SEM) of O/CM; (C) CuO-Cu prepared in example 1₂Transmission Electron Microscopy (TEM) of O nanocomposites; (D) CuO-Cu prepared in example 1₂High Resolution Transmission Electron Microscopy (HRTEM) of O nanocomposites. (E, F) CuO-Cu prepared in example 1₂An energy dispersive X-ray spectroscopy (EDS) mapping analysis plot of the O nanocomposite;

FIG. 4, CuO-Cu prepared in example 1₂X-ray photoelectron spectroscopy (XPS) of O/CM, (A) CuO-Cu₂An O/CM total spectrum; (B) high resolution XPS spectra of Cu 2p region;

FIG. 5 shows a photoelectrochemical biosensor in which (A) is 5.0mM [ Fe (CN)₆]^3-/4-Electrochemical Impedance Spectroscopy (EIS) of (1), the following are the respective working electrodes: (a) CuO-Cu₂O/CM, (b) BSA/aptamer/CuO-Cu₂O/CM and (c) PSA/BSA/aptamer/CuO-Cu₂And (B) a photocurrent response of the PEC aptamer sensor after stepwise modification at an applied potential of 0V under visible light irradiation in 0.01M Tris-HCl buffer. (a) CuO-Cu₂O/CM, ((b) BSA/aptamer/CuO-Cu₂O/CM and (c) PSA/BSA/aptamer/CuO-Cu₂O/CM;

FIG. 6, the photoelectrochemical biosensor prepared in example 1, was used to detect the current response (A) of PSA at different concentrations, and the corresponding calibration curve (B);

FIG. 7, (A) a control chart of the photoelectrochemical biosensor prepared in example 1 for detection of PSA selectivity; (B) measurement of aptamer/CuO-Cu with optical on and off₂Stability and reproducibility of O/CM electrodes.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

Example 1

(1) Respectively dissolving 3.2g of NaOH and 0.91g of ammonium persulfate in 25mL of deionized water, mixing the NaOH solution and the ammonium persulfate solution, and stirring for 5 minutes to obtain a clear solution; placing the pretreated copper mesh in the clear solution for soaking reaction for 20 minutes; the sample was taken out, washed with deionized water and ethanol, respectively, and air-dried at 60 ℃ to obtain Cu (OH)₂A precursor.

(2) Mixing the Cu (OH) prepared in the step (1)₂The precursor was calcined and calcined in air at 350 ℃ for 15 minutes. The copper mesh was taken out, and the obtained sample (CuO-Cu) was finally used₂O/CM) was washed several times with deionized water and dried in air.

Example 2

(1) Respectively dissolving 3.5g of NaOH and 1.0g of ammonium persulfate in 20mL of deionized water, mixing a NaOH solution and an ammonium persulfate solution, and stirring for 10 minutes to obtain a clear solution; will be provided withPlacing the pretreated copper mesh in the clear solution for soaking reaction for 25 minutes; the sample was taken out, washed with deionized water and ethanol, respectively, and air-dried at 60 ℃ to obtain Cu (OH)₂A precursor.

(2) Mixing the Cu (OH) prepared in the step (1)₂The precursor was calcined and calcined in air at 350 ℃ for 20 minutes. The copper mesh was taken out, and the obtained sample (CuO-Cu) was finally used₂O/CM) was washed several times with deionized water and dried in air.

Example 3

(1) Respectively dissolving 3.3g of NaOH and 0.95g of ammonium persulfate in 23mL of deionized water, mixing the NaOH solution and the ammonium persulfate solution, and stirring for 8 minutes to obtain a clear solution; placing the pretreated copper mesh in the clear solution for soaking reaction for 23 minutes; the sample was taken out, washed with deionized water and ethanol, respectively, and air-dried at 60 ℃ to obtain Cu (OH)₂A precursor.

(2) Mixing the Cu (OH) prepared in the step (1)₂The precursor was calcined and calcined in air at 350 ℃ for 18 minutes. The copper mesh was taken out, and the obtained sample (CuO-Cu) was finally used₂O/CM) was washed several times with deionized water and dried in air.

Photoelectrochemical biosensor

The photoelectrochemical biosensor comprises a working electrode and a reference electrode (Ag | AgCl | Cl) connected with an electrochemical workstation^-) A counter electrode (platinum electrode), a xenon lamp irradiation is adopted as a simulation light source, the working electrode is a Copper Mesh (CM), and the CuO-Cu prepared in the embodiment 1 is modified on the Copper Mesh (CM)₂Performing ultrasonic cleaning on the O/CM nanowire array in diluted hydrochloric acid for 2 min before modification, wherein the area of a CM electrode is 0.5 x 0.5 CM²。

Direct immersion reaction of the photoelectrochemical biosensor described in example 1 on the CM working electrode to Cu (OH)₂Precursor, then finally obtaining CuO-Cu by calcining₂And (3) an O nanowire array.

When the photoelectrochemical biosensor described in example 1 detects PSA, 20. mu.L of 0.1% chitosan was dropped on the surface of the photoelectrode,air-dried at 60 ℃ and then immobilized with an aptamer (available from Shanghai Biotechnology Ltd.) CuO-Cu₂O/CM on electrode, incubated at 4 ℃ for 16h, washed with Tris-HCl buffer solution and then N₂Blow-drying, after which the prepared aptamer sensor was incubated with 20 μ L of 1% BSA at room temperature for 1 hour to block non-specific sites. The BSA/aptamer/CuO-Cu was then rinsed thoroughly with buffer solution₂O/CM electrode to wash away excess BSA and store at 4 ℃ for later use. Finally, the prepared sensor was incubated with 20 μ L of PSA at 37 ℃ for 1 h. The PSA is detected according to the change of photoelectric signals of the photoelectrochemical biosensor by irradiating under a 300W xenon lamp.

As shown in FIG. 1, based on BSA/aptamer/CuO-Cu₂The O/CM photoelectrode establishes a new platform for the ultra-sensitive detection of PSA;

as shown in fig. 2, a schematic diagram of the electron transfer process of the proposed PEC biosensor is shown;

as shown in FIG. 3, Panel A shows the CuO-Cu prepared₂XRD of O nanowire samples. Diffraction peaks of CuO at 67.7 °, 38.1 °, 48.1 °, 53.5 °, 57.3 °, 32.5 ° point to the (-110), (111), (-202), (020), (202), (113) (311) plane (JCPDS numbering 48-1548). The peaks at 36.4 ° and 61.4 ° are labeled as Cu₂O (JCPDS number 48-1548), matches well with the (111) and (220) planes and shows a very prominent and clear diffraction peak at 36.4 deg.. FIG. B shows CuO-Cu₂Scanning Electron Microscope (SEM) images of O/CM. As observed, CuO-Cu₂The O composite material is in the form of nano wire, and the smooth surface of the copper net is completely covered by CuO-Cu₂And covering the O nanowire. In addition, the morphology of the nanowires was examined by Transmission Electron Microscopy (TEM) (fig. C). Single CuO-Cu₂High Resolution Transmission Electron Microscope (HRTEM) images of O nanowires showed good crystallization. 0.243 nm lattice spacing in HRTEM images assigned to Cu₂The (111) plane of O, and the lattice spacing of 0.252 nm corresponds to the (-111) plane of CuO (Panel D). Energy dispersive X-ray spectroscopy (EDS) mapping analysis confirmed the elemental composition of Cu and O and their compositionIn the presence of CuO-Cu₂Even distribution over O/CM (FIGS. E and F).

To further confirm the chemical composition of the prepared samples, XPS analysis was performed. CuO-Cu₂The measured spectrum of the O composite material is shown in fig. 4A, showing that only Cu, O and C (from the atmosphere) elements were detected, and no other impurities were present. The high resolution XPS spectrum of Cu 2p is shown in FIG. 4B. The peak positions of Cu 2p1/2 and Cu 2p3/2 are 953.9 eV and 933.7 eV, respectively. The fitted peaks at 934.3 eV and 954.2 eV are attributed to CuO. Peaks of 933.2 eV and 952.4 eV correspond to Cu₂And O. In addition, two detectable rocking satellite peaks further indicate the presence of CuO. The results agree well with those of XRD, which further confirms that CuO-Cu₂Successful fabrication of O heterojunctions.

As shown in FIG. 5A, the use of [ Fe (CN)₆]^{3- / 4-}Nyquist plots for electrodes of different fabrication processes as redox probes, the inset of which is the corresponding equivalent circuit. CuO-Cu₂The O/CM had the lowest value of Rct (curve a). However, after anchoring the aptamer and incubation in blocking agent (BSA) to block the remaining active sites, the resistance gradually increases (curve b), probably due to the formation of a less conductive complex, which greatly hinders the transfer interface of electrons from bulk solution to the electrode. When PSA is introduced, the aptamer sensor can react specifically with PSA, PSA/aptamer-CuO-Cu₂The impedance of O/CM (curve c) is less than that of BSA-aptamer-CuO-Cu₂Impedance of O/CM (curve b). This may be attributed to the fact that PSA can interact with aptamers to form PSA-aptamer complexes with CuO-Cu in solution₂The O/CM electrodes are separated, thereby reducing steric hindrance. All these results provide supporting evidence that the proposed PEC sensing platform has been successfully manufactured and can be used for PSA assays.

PEC technology is also used to monitor the construction process of PEC-adapted sensors and their feasibility in PSA detection. Fig. 5B depicts the photocurrent response of the PEC aptamer sensor after stepwise modification at an applied potential of 0V under visible light illumination. As shown in the curve a, CuO-Cu₂The O/CM is sensitive to radiation of visible light and exhibits good cathode photoelectric propertyFlow, indicating CuO-Cu₂The O nanowire heterostructure has good photocatalytic activity. However, in immobilizing PSA binding aptamers and BSA to CuO-Cu₂After O/CM, the photocurrent intensity dropped significantly, resulting in a lower PEC background signal and no further detection of PSA (curve b). The steric hindrance experienced by the biomolecules hinders the transfer of photo-generated charges, thereby accelerating their recombination with holes. However, the introduction of PSA resulted in a recovered cathode photocurrent signal (curve c), which may be due to the specific capture of PSA by the aptamer sensor, with release of the PSA-aptamer complex from the sensing interface. All results obtained were consistent with EIS results, indicating successful construction of a stepwise modified PEC aptamer sensor for PSA detection.

As shown in FIG. 6, by configuring PSA aqueous solutions with different concentrations and testing the current response curves of the photoelectrochemical biosensor to the PSA with different concentrations, it can be seen that the PSA concentrations have better correlation, and the linear regression equation is A =159.1+0.95C_PSA(R²=0.9931), and the lower detection limit is 0.33 ng · mL when S/N =3^-1。

Selectivity is another important criterion for designing PEC sensor analysis applications, and was detected by replacing the PSA standard with several representative interfering proteins (including CA125, IgG and AFP). As shown in fig. 7A, the photocurrent response of the photocathode was not affected by possible interfering proteins, while PSA provided a sensitive and significant photocurrent signal, indicating an increase in signal from specific binding. In addition, 2 ng/mL PSA solutions containing 20 ng/mL interfering substances were also measured. The change in signal caused by the addition of the control protein was negligible, indicating that the selectivity of the aptamer sensor was completely acceptable. The stability of the proposed immunoassay was also investigated (fig. 7B). Clearly, no significant change was found in the photocurrent signal when the illumination was repeatedly turned on/off within 2400 s. Good photocatalytic stability should result from the formation of a CuO-Cu2O heterostructure that can improve photocatalytic stability.

The above description is only exemplary of the present invention and should not be taken as limiting the invention, as any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims (8)

1. CuO-Cu₂The preparation method of the O/CM nanowire array heterostructure is characterized by comprising the following steps of:

(1) mixing and stirring a NaOH solution and an ammonium persulfate solution to obtain a clear solution, placing a pretreated copper net in the clear solution for soaking reaction, washing and drying an obtained sample to obtain Cu (OH)₂A precursor;

(2) mixing the Cu (OH) prepared in the step (1)₂Calcining the precursor to obtain CuO-Cu₂An O/CM nanowire array.

2. The preparation method according to claim 1, wherein the mass ratio of NaOH to ammonium persulfate in the step (1) is 3.2-3.5: 0.91-1.0; the mass-volume ratio of NaOH to water is (3.2-3.5) g: (20-25) mL; the mass volume ratio of the ammonium persulfate to the water is (0.91-1.0) g: (20-25) mL.

3. The method according to claim 1, wherein the water in step (1) is deionized water; the washing step in the step (1) is washing with deionized water and ethanol respectively.

4. The method according to claim 1, wherein the drying temperature in the step (1) is 60 ℃; the calcining temperature in the step (2) is 350 ℃, and the calcining time is 15-20 minutes.

5. The production method according to claim 1, wherein the stirring time in the step (1) is 5 to 10 minutes; the soaking reaction time is 20-25 minutes.

6. The photoelectrochemistry biosensor comprises a working electrode, a reference electrode and a counter electrode which are connected with an electrochemical workstation, and is characterized in that the working electrode is arranged on the working electrodeModified with CuO-Cu obtained according to any one of claims 1 to 5₂An O/CM nanowire array.

7. Use of the photoelectrochemical biosensor of claim 6 for sensitive detection of PSA.

8. Use of the photoelectrochemical biosensor according to claim 7, wherein the detection is performed by the following steps:

(1) dropping 0.1% chitosan on the surface of the photoelectrode, drying in air at 60 ℃, and fixing the aptamer CuO-Cu₂O/CM on electrode, incubated at 4 ℃ for 16h, washed with Tris-HCl buffer solution and then N₂Drying;

(2) the prepared photoelectrochemical biosensor was incubated with 1% BSA at room temperature for 1h, and the BSA/aptamer/CuO-Cu buffer solution was thoroughly washed with Tris-HCl buffer solution₂Storing the O/CM electrode for later use;

(3) and (3) incubating the sensor and PSA at 37 ℃ for 1h, irradiating under a xenon lamp, and detecting the PSA according to the change of photoelectric signals of the photoelectrochemical biosensor.

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