Preparation method and application of photoelectrochemical aptamer sensor for tetracycline detection

Abstract

The invention relates to a preparation method and application of a photoelectrochemical aptamer sensor for tetracycline detection, and the preparation method comprises the following steps: synthesizing an up-conversion material NaYF4: Yb, Tm through solvothermal reaction, reacting with a composite material (NGTT) formed by compounding g-C3N4, Ti3C2 and TiO2 prepared based on melamine, and modifying the obtained NGTT to the surface of an indium tin oxide glass (I TO) electrode to obtain the photoelectrochemical aptamer sensor for tetracycline detection. And combining with an aptamer to form the photoelectrochemical sensor for tetracycline detection. According to the sensor, NaYF4: Yb, Tm is used as an energy transfer donor, near-infrared light is converted into ultraviolet light and visible light, the ultraviolet light and the visible light are efficiently absorbed through the g-C3N4 / TiO2 / Ti3C2 composite material, and high-sensitivity and high-selectivity detection of tetracycline is achieved. The sensor provided by the invention has high light stability, low biotoxicity, strong penetrability and good absorption capacity to near-infrared light, and is suitable for in vivo detection and accurate detection of tetracycline in a complex environment.

Description

Technical Field

[0001] The invention relates to the technical field of chemical analysis, and in particular to a preparation method of a photoelectrochemical aptamer sensor for tetracycline detection and application thereof. Background Art

[0002] Photoelectrochemical aptamer sensing is a new analytical method that combines bioanalysis with photoelectrochemical sensing. Its core lies in the specific binding of photoelectroactive materials and nucleic acid aptamers (aptamers), and the use of the binding reaction between the target detection object and the aptamer to trigger the change of the photoelectric signal, thereby achieving quantitative analysis. Nucleic acid aptamers are single-stranded DNA or RNA with a specific sequence that can bind to target molecules with high specificity and high affinity, making them ideal recognition units for photoelectrochemical sensors. In this technology, the performance of photoelectroactive materials plays a decisive role in the detection sensitivity and signal strength of the sensor.

[0003] In the prior art, the photoelectrochemical aptamer sensor that is closest to the present invention is a tetracycline detection sensor based on aptamer / g-C3N4 / BiVO4 / FTO electrode. Its photoelectric active material is g-C3N4 / BiVO4, and the excitation light source adopts ultraviolet-visible light (LED light) with a wavelength of 420nm. However, due to the inherent band gap limitation, traditional semiconductor photoelectric materials can usually only absorb ultraviolet light and part of visible light, and cannot make full use of near-infrared light, which accounts for 50% of the solar spectrum. Compared with ultraviolet light and visible light, near-infrared light has low phototoxicity, good biocompatibility and deeper tissue penetration ability. These characteristics make sensors excited by near-infrared light have important application value in the fields of biomolecule detection, solar energy conversion and energy catalysis. However, the current single photoelectric semiconductor material has limitations such as low excitation light utilization and insufficient detection sensitivity.

[0004] In addition, existing studies have not applied photoelectrochemical aptamer sensors based on near-infrared light excitation and their related photoelectric materials to the detection of tetracycline. Therefore, the development of a photoelectrochemical aptamer sensor that can achieve high-sensitivity detection under near-infrared light excitation has important research significance and application prospects. This breakthrough can not only solve the bottleneck problems of traditional sensors in light source utilization and detection performance, but also provide innovative technical solutions for antibiotic detection and other analysis in complex environments.

[0005] In view of the above problems, the present invention is proposed. Summary of the invention

[0006] The invention discloses a method for preparing a photoelectrochemical aptamer sensor for tetracycline detection, aiming to solve the technical problems existing in the prior art.

[0007] According to one aspect of the present invention, a method for preparing a photoelectrochemical aptamer sensor for detecting tetracycline is provided, comprising:

[0008] Dissolving NaOH in ultrapure water, adding ethanol and oleic acid, and stirring to obtain a first mixed solution;

[0009] Mix YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O, add into NH4F solution, and stir to obtain a second mixed solution;

[0010] The first mixed solution and the second mixed solution are mixed and stirred to perform a solvothermal reaction to obtain a third mixed solution, and the third mixed solution is centrifuged, washed and dried to obtain an up-conversion material NaYF4:Yb,Tm;

[0011] Dissolving an up-conversion material NaYF4:Yb,Tm in a polyvinylpyrrolidone (PVP) solution to obtain a fourth mixed solution;

[0012] Prepare a g-C3N4 supernatant using melamine, add Ti3C2 into the g-C3N4 supernatant, then add H2O2 dropwise, and stir to obtain a fifth mixed solution;

[0013] The fourth mixed solution and the fifth mixed solution are mixed and stirred, and heated to react to obtain a sixth mixed solution, and the sixth mixed solution is centrifuged, washed and dried to obtain an NGTT composite material;

[0014] The NGTT composite material was modified onto the surface of an indium tin oxide glass (ITO) electrode and incubated in an aptamer solution to obtain an aptamer / NGTT / ITO electrode, which is a photoelectrochemical aptamer sensor for tetracycline detection.

[0015] As a preferred technical solution, in the preparation of g-C3N4 supernatant using melamine:

[0016] Melamine is used as a raw material to be calcined to obtain g-C3N4, and the g-C3N4 is subjected to water dispersion and ultrasonic centrifugation to obtain a g-C3N4 supernatant.

[0017] As a preferred technical solution, the calcination reaction temperature is 550°C and the reaction time is 2h.

[0018] As a preferred technical solution, the molar ratio of YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O is 79.8:20:0.2.

[0019] As a preferred technical solution, the NGTT composite material is modified onto the surface of an indium tin oxide glass (ITO) electrode, including:

[0020] Pre-treat indium tin oxide glass (ITO) electrodes;

[0021] Ultrasonic dispersion of the NGTT composite material in ultrapure water to obtain an NGTT aqueous dispersion;

[0022] The NGTT aqueous dispersion was droplet coated onto the conductive surface of the indium tin oxide glass (ITO) electrode.

[0023] As a preferred technical solution, the pretreatment of the indium tin oxide glass (ITO) electrode includes:

[0024] The slice of indium tin oxide glass (ITO) was immersed in a detergent solution for ultrasonic treatment, and ultrapure water and ethanol were added for ultrasonic treatment respectively, and then dried with nitrogen gas.

[0025] As a preferred technical solution, the NGTT composite material is ultrasonically dispersed in ultrapure water, and the concentration of the obtained NGTT aqueous dispersion is 2 mg / mL.

[0026] As a preferred technical solution, after the NGTT aqueous dispersion is droplet-coated on the conductive surface of the indium tin oxide glass (ITO) electrode, it is dried at a drying temperature of 70° C. and a drying time of 12 h.

[0027] As a preferred technical solution, the heating temperature for mixing and heating the fourth mixed solution and the fifth mixed solution is 80° C., and the reaction time is 1 hour.

[0028] According to another aspect of the present invention, there is also provided a use of a photoelectrochemical biosensor prepared by the above preparation method for detecting tetracycline, comprising:

[0029] The surface of the photoelectrochemical biosensor was incubated in tetracycline (TET) / PBS solutions of different concentrations to obtain TET / aptamer / NGTT / I TO electrodes of different concentrations;

[0030] A three-electrode system was used: TET / aptamer / NGTT / I TO electrode as the working electrode, platinum wire as the counter electrode, silver / silver chloride as the reference electrode, PBS solution as the electrolyte, and a 980nm laser as the excitation light source. The photoresponse performance of the TET / aptamer / NGTT / I TO electrode was detected on an electrochemical workstation.

[0031] The technical solution adopted by the present invention can achieve at least one of the following beneficial effects:

[0032] 1. The present invention uses NaYF4:Yb,Tm as an energy transfer donor, which can efficiently convert near-infrared light into ultraviolet and visible light. This light energy is absorbed by the g-C3N4 / TiO2 / Ti3C2 composite material (NGTT), solving the problem of weak absorption of near-infrared light by ordinary semiconductor materials. Based on the composite characteristics of NaYF4:Yb,Tm and semiconductor materials, the sensor achieves strong adjustability and significantly improved photoelectric conversion efficiency.

[0033] 2. This invention is the first to apply the photoelectrochemical aptamer sensor based on NaYF4:Yb,Tm / g-C3N4 / TiO2 / Ti3C2 composite material (NGTT) to the detection of tetracycline (TET), showing extremely high sensitivity and selectivity. The sensor performs well in light response detection and can accurately detect TET in complex environments, providing a new and widely adaptable technical path for the detection of other antibiotics.

[0034] 3. The photoelectrochemical aptamer sensor of the present invention has good absorption capacity for near-infrared light, and exhibits high photostability, low biotoxicity and strong penetrability. These characteristics make the sensor suitable for preparing microelectrodes for in vivo detection, providing reliable technical support for the medical and biosensing fields.

[0035] 4. Due to its near-infrared light excitation, high sensitivity, low cost and high reliability, the photoelectrochemical biosensor of the present invention is not only suitable for the detection of tetracycline, but can also be expanded to the detection of other antibiotics, heavy metal ions and organic pollutants, and has broad application prospects and market value. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for describing the embodiments, which constitute a part of the present invention. The exemplary embodiments of the present invention and their descriptions explain the present invention and do not constitute improper limitations on the present invention. In the drawings:

[0037] Figure 1 It is a schematic flow chart of a method for preparing a photoelectrochemical aptamer sensor for tetracycline detection according to the present invention;

[0038] Figure 2 It is a material characterization diagram of the NGTT composite material of the present invention;

[0039] Figure 2 A is a scanning electron microscope image (SEM) of the NGTT composite material of the present invention;

[0040] Figure 2 B is a transmission electron microscope image (TEM) of the NGTT composite material of the present invention;

[0041] Figure 2 C is a high resolution image (HRTEM) of the NGTT composite material of the present invention;

[0042] Figure 2 D is an element mapping image (EDS Mapping) of the NGTT composite material of the present invention;

[0043] Figure 3 The X-ray diffraction characterization diagram (XRD) of the NGTT composite material of the present invention is compared with the PDF of the NaYF4:Yb,Tm standard card (#28-1192);

[0044] Figure 4 is an X-ray photoelectron spectrum (XPS) of the NGTT composite material of the present invention;

[0045] Figure 5 The fluorescence spectra of the NaYF4:Yb,Tm (a) and NGTT composite materials (b) of the present invention;

[0046] Figure 6 UV-visible-near infrared diffuse reflectance images of NaYF4:Yb,Tm (a), g-C3N4 / TiO2 / Ti3C2 (b) and NGTT composite materials (c) of the present invention;

[0047] Figure 7 Electrochemical impedance spectroscopy (EIS) of NaYF4:Yb,Tm (a), g-C3N4 / TiO2 / Ti3C2 (b), NGTT composite material (c), Aptamer / NGTT composite material (d) and TET / Aptamer / NGTT composite material (e) of the present invention;

[0048] Figure 8 The photocurrent response diagrams of the electrodes modified with g-C3N4 / TiO2 / Ti3C2 (a), NGTT composite material (b), Aptamer / NGTT composite material (c), 10nM TET / Aptamer / NGTT composite material (d) and 100nM TET / Aptamer / NGTT composite material (e) of the present invention under near-infrared light excitation at a wavelength of 980nm;

[0049] Fig. 9 This is a test diagram of the anti-interference performance of a photoelectrochemical aptamer sensor for tetracycline detection prepared by the present invention;

[0050] Fig.10 This is a stability test diagram of a photoelectrochemical aptamer sensor for tetracycline detection prepared by the present invention;

[0051] Fig.11A photocurrent response diagram of a photoelectrochemical aptamer sensor for tetracycline detection prepared by the present invention to different concentrations of TET;

[0052] Fig.12 The linear fitting equation of the photoelectrochemical aptamer sensor for tetracycline detection prepared by the present invention for different concentrations of TET. DETAILED DESCRIPTION

[0053] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be clearly and completely described below in conjunction with the specific embodiments of the present invention and the corresponding drawings. In the description of the present invention, it should be noted that the term "or" is usually used in the sense of including "and / or", unless the content clearly indicates otherwise.

[0054] Unless expressly indicated to the contrary, the numerical parameters in this specification and the appended claims may be approximate values ​​and can be changed according to the desired properties obtained by the content of the present invention. Specifically, all numbers used in the specification and claims to express the content of the composition, reaction conditions, etc., should be understood to be modified by the term "about" in all cases. In general, the meaning of the expression is to include a change of ±10% in some embodiments, a change of ±5% in some embodiments, a change of ±1% in some embodiments, and a change of ±0.5% in some embodiments by a specific number.

[0055] Furthermore, the word "comprising" does not exclude the presence of materials or steps not listed in the claims. Ordinal numbers such as "first", "second", "third" and Arabic numerals, letters, etc. used in the specification and claims to modify corresponding elements or steps do not themselves mean the order of the manufacturing method. The use of these ordinals is only used to make the steps clearly distinguishable.

[0056] In addition, unless the steps are specifically described or must occur in sequence, the order of the above steps is not limited to the above list, and can be changed or rearranged according to the required design. And the above embodiments can be mixed and matched with each other or with other embodiments based on design and reliability considerations, that is, the technical features in different embodiments can be freely combined to form more embodiments.

[0057] Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0058] In order to solve the problems existing in the prior art, the embodiment of the present invention provides a method for preparing a photoelectrochemical aptamer sensor for tetracycline detection, comprising the following steps:

[0059] S1: Preparation of upconversion material NaYF4:Yb,Tm by solvothermal method;

[0060] S11: dissolving NaOH in ultrapure water to form a NaOH solution, adding ethanol and oleic acid to the NaOH solution, and stirring to obtain a first mixed solution;

[0061] S12: preparing an NH4F solution, mixing YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O, adding the mixture to the NH4F solution, and stirring to obtain a second mixed solution;

[0062] S13: mixing and stirring the first mixed solution and the second mixed solution, performing a solvothermal reaction, obtaining a third mixed solution, and centrifugally washing and drying the third mixed solution to obtain an up-conversion material NaYF4:Yb,Tm;

[0063] S2: Preparation of NGTT composite material, namely NaYF4:Yb,Tm / g-C3N4 / TiO2 / Ti3C2 composite material, by hydrogen peroxide (H2O2) oxidation method;

[0064] S21: calcining melamine as a raw material to obtain g-C3N4, performing water dispersion ultrasonic centrifugation to obtain a g-C3N4 supernatant for standby use;

[0065] S22: dissolving the up-conversion material NaYF4:Yb,Tm prepared in S1 in a polyvinylpyrrolidone (PVP) solution to obtain a fourth mixed solution;

[0066] S23: adding Ti3C2 to the g-C3N4 supernatant, and then adding H2O2 dropwise, stirring, to obtain a fifth mixed solution;

[0067] S24: mixing and stirring the fourth mixed solution and the fifth mixed solution, performing heating reaction to obtain a sixth mixed solution, and centrifugally washing and drying the sixth mixed solution to obtain an NGTT composite material;

[0068] S3: Modify the NGTT composite material onto the surface of ITO (indium tin oxide glass) electrode;

[0069] S31: Pre-treating the I TO electrode, immersing the I TO slice in a detergent solution for ultrasonic treatment, adding ultrapure water and ethanol for ultrasonic treatment respectively, and then drying with nitrogen;

[0070] S32: ultrasonically dispersing the NGTT composite material in ultrapure water to obtain an NGTT aqueous dispersion;

[0071] S33: applying NGTT aqueous dispersion droplets to the conductive surface of the I TO electrode and drying to obtain the NGTT / I TO electrode;

[0072] S4: Incubate the NGTT / I TO electrode in an aptamer solution to obtain an aptamer / NGTT / I TO electrode, which is a photoelectrochemical aptamer sensor for detecting tetracycline.

[0073] The invention discloses a method for preparing a photoelectrochemical aptamer sensor for tetracycline detection. The method is based on an upconversion material NaYF4:Yb,Tm prepared by a solvothermal method, and a NGTT composite material is prepared by a hydrogen peroxide (H2O2) oxidation method. The NGTT composite material and the aptamer are sequentially modified on an ITO electrode to obtain a photoelectrochemical aptamer sensor (aptamer / NGTT / ITO electrode) for detecting tetracycline. The photoelectrochemical aptamer sensor uses NaYF4:Yb,Tm as an energy transfer donor, can convert near-infrared light into ultraviolet-visible absorption light, can be absorbed by a g-C3N4 / TiO2 / Ti3C2 composite material, and obtains a photoelectrochemical aptamer sensor based on a NaYF4:Yb,Tm / g-C3N4 / TiO2 / Ti3C2 composite material (NGTT). The photoelectrochemical aptamer sensor has good absorption of near-infrared light, high photostability, low biotoxicity and strong penetrability, and effectively solves the problem of weak near-infrared light absorption of common semiconductor materials. Compared with single upconversion materials or semiconductor materials, semiconductor composite materials based on upconversion materials have stronger adjustability and higher photoelectric conversion efficiency. In addition, the photoelectrochemical aptamer sensor has high sensitivity, low cost, and simple operation. It is suitable for highly sensitive and highly selective detection of tetracycline (TET), and also provides a new path for the detection of other antibiotics.

[0074] In some preferred embodiments, when preparing the second mixed solution, the molar ratio of YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O used is 79.8:20:0.2.

[0075] In some preferred embodiments, when preparing the third mixed solution, the temperature of the solvent thermal reaction is 200° C. and the reaction time is 12 hours. Such temperature and time parameters help the solvent thermal reaction to proceed better and to react more fully.

[0076] In some preferred embodiments, when preparing the g-C3N4 supernatant, melamine is used as a raw material for calcination, and the calcination reaction temperature is 550°C and the reaction time is 2 hours.

[0077] In some preferred embodiments, when preparing the sixth mixed solution, the fourth mixed solution and the fifth mixed solution are mixed and heated at a heating temperature of 80°C and a reaction time of 1 hour, so as to facilitate partial oxidation of Ti3C2 and facilitate the preparation of NGTT composite materials.

[0078] In some preferred embodiments, the NGTT composite material is ultrasonically dispersed in ultrapure water, and the concentration of the obtained NGTT aqueous dispersion is 2 mg / mL.

[0079] In some preferred embodiments, the NGTT aqueous dispersion is dropwise coated onto the conductive surface of the I TO electrode, and in the step of drying to obtain the NGTT / I TO electrode, the drying temperature is 70°C and the drying time is 12 hours. The temperature and time parameters are conducive to the curing of the NGTT aqueous dispersion. Too high a temperature will easily cause cracking, and too low a temperature will have a poor curing effect.

[0080] A detection method for detecting tetracycline (TET) using the above-mentioned photoelectrochemical aptamer sensor comprises the following steps:

[0081] The aptamer / NGTT / I TO electrodes were incubated in TET / PBS solutions of different concentrations to obtain TET / aptamer / NGTT / I TO electrodes of different concentrations;

[0082] The photoresponse performance of the above-mentioned electrode was detected by an electrochemical workstation. A three-electrode system was adopted, with the above-mentioned TET / aptamer / NGTT / I TO electrode as the working electrode, platinum wire as the counter electrode, silver / silver chloride as the reference electrode, PBS solution as the electrolyte, and a laser with a wavelength of 980 nm as the excitation light source.

[0083] Embodiment 1:

[0084] This embodiment provides a method for preparing a photoelectrochemical aptamer sensor for tetracycline detection, comprising the following steps:

[0085] S1: Preparation of upconversion material NaYF4:Yb,Tm by solvothermal method;

[0086] Dissolve 10 mM NaOH in 2 mL ultrapure water, add 8 mL ethanol and oleic acid to the NaOH solution, and stir to obtain a first mixed solution;

[0087] 0.1 M YbCl3·6H2O, YCl3·6H2O, and TmCl3·6H2O (molar ratio of 79.8:20:0.2) were mixed and added to 2 mL of 2M NH4F solution, and stirred to obtain a second mixed solution;

[0088] The first mixed solution and the second mixed solution were mixed and stirred for 15 minutes, and then transferred to a 50 mL polytetrafluoroethylene-lined autoclave for a solvothermal reaction at a reaction temperature of 200° C. for 12 hours to obtain a third mixed solution. The third mixed solution was centrifuged and washed three times with ultrapure water and ethanol, and then dried in an oven at 70° C. for 12 hours to obtain an upconversion material NaYF4:Yb,Tm.

[0089] S2: Preparation of NGTT composite material, namely NaYF4:Yb,Tm / g-C3N4 / TiO2 / Ti3C2 composite material, by hydrogen peroxide (H2O2) oxidation method;

[0090] Melamine was used as a raw material for calcination at a reaction temperature of 550°C for a reaction time of 2 hours to obtain g-C3N4. 20 mg of g-C3N4 was weighed and added to 20 mL of ultrapure water for ultrasonic centrifugation. The supernatant was taken for later use.

[0091] Dissolve 0.1 g of up-conversion material NaYF4:Yb,Tm in 4.0 mL of 0.2 g / mL polyvinylpyrrolidone (PVP) solution to obtain a fourth mixed solution;

[0092] 0.1 g of Ti3C2 was added to the g-C3N4 supernatant, and 50 μL of H2O2 was added dropwise, and stirred to obtain a fifth mixed solution;

[0093] The fourth mixed solution and the fifth mixed solution were mixed and stirred, and heated to react at a heating temperature of 80° C. for a reaction time of 1 hour to obtain a sixth mixed solution, and the sixth mixed solution was centrifuged and washed three times with ultrapure water and ethanol, and dried in a vacuum oven at 40° C. for 12 hours to obtain an NGTT composite material;

[0094] S3: Modify the NGTT composite material onto the surface of I TO electrode;

[0095] Pretreatment of ITO electrodes: Immerse the ITO slices in a detergent solution for ultrasonic treatment for 0.5 hours, add ultrapure water and ethanol for ultrasonic treatment for 0.5 hours respectively, and then blow dry with nitrogen;

[0096] The NGTT composite material was ultrasonically dispersed in ultrapure water (2.5 mg / mL), and then drop-coated on the conductive surface of the I TO electrode. After being naturally dried, it was transferred to an oven at 70°C and dried for 12 hours to obtain an NGTT / I TO electrode.

[0097] S4: Incubate the NGTT / I TO electrode in 40 μL of 2 μM aptamer solution for 2 hours to obtain an aptamer / NGTT / I TO electrode.

[0098] A detection method for detecting tetracycline (TET) using the above-mentioned photoelectrochemical aptamer sensor comprises the following steps:

[0099] The aptamer / NGTT / I TO electrodes were incubated in 10 mL 0.01 M PBS solution with different concentrations of TET for 1 hour to obtain TET / Aptamer / NGTT / I TO electrodes with different concentrations;

[0100] The photoresponse performance of the above-mentioned electrode was detected by an electrochemical workstation. A three-electrode system was adopted, with the above-mentioned TET / aptamer / NGTT / I TO electrode as the working electrode, platinum wire as the counter electrode, silver / silver chloride as the reference electrode, PBS solution as the electrolyte, and a laser with a wavelength of 980 nm as the excitation light source.

[0101] Material characterization and performance testing:

[0102] like Figure 2 As shown in Figure 2, the morphology of the NGTT composite material samples was characterized by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Figure 2 As shown in A (SEM), in the prepared NGTT composite material, NaYF4:Yb,Tm is rod-shaped with a length of about 1 μm. The g-C3N4 / TiO2 / Ti3C2 composite material is block-shaped as a whole. The rod-shaped NaYF4:Yb,Tm is evenly distributed between the block-shaped g-C3N4 / TiO2 / Ti3C2 composite materials. Figure 2 B (TEM) is consistent, and it can be seen more clearly that the four materials NaYF4:Yb,Tm, g-C3N4, TiO2, and Ti3C2 are closely combined. Figure 2 C (HRTEM) shows that the lattice fringes of NaYF4:Yb,Tm and TiO2 are clearly visible, indicating a high degree of crystallinity. Figure 2 The elements in D (EDS Mapping) are clearly visible and evenly distributed, further proving the successful synthesis of the NGTT composite material.

[0103] like Figure 3 As shown, the prepared NGTT composite material was characterized and analyzed by X-ray diffraction characterization (XRD). The characteristic peaks of the XRD spectrum of the NGTT composite material are consistent with the NaYF4:Yb,Tm standard card PDF comparison (#28-1192). In addition, the characteristic peak at 27.5° corresponds to the (002) crystal plane of g-C3N4. In addition, the strong peaks at 25.4°, 38.0°, 48.3° and 54.1° are attributed to rutile TiO2, which further verifies the oxidation of Ti3C2.

[0104] like Figure 4As shown, X-ray photoelectron spectroscopy (XPS) was used to further verify the elemental composition of the NGTT composite material. Figure 4 From the full spectrum, we can see that nine elements, including Ti, Y, Tm, Yb, C, N, Na, O, and F, are present, proving the successful synthesis of the NGTT composite material.

[0105] like Figure 5 As shown in Figure 2, the feasibility of NGTT composite materials for near-infrared excitation was verified by fluorescence spectra. Figure 5 As shown in Figure 2, under near-infrared 980nm laser excitation, NaYF4:Yb / Tm has strong upconversion emission. In the ultraviolet-visible region, there are three typical emission peaks at 365nm, 465nm and 480nm, corresponding to Tm 3+ of 1 D2→ 3 H6, 1 D2→ 3 F6, and 1 G4→ 3 H6 transition. After being compounded with g-C3N4 / TiO2 / Ti3C2, the NGTT composite material has obvious fluorescence quenching phenomenon, indicating that the energy of NaYF4:Yb / Tm can be effectively transferred to g-C3N4 / TiO2 / Ti3C2, further improving the utilization efficiency of near-infrared light.

[0106] like Figure 6 As shown, the light absorption analysis of the sample was carried out using UV-visible-near-infrared diffuse reflectance. Figure 6 The UV-Vis-NIR spectra of NaYF4:Yb / Tm, g-C3N4 / TiO2 / Ti3C2, and NGTT composite materials. It can be seen from the figure that single NaYF4:Yb / Tm and g-C3N4 / TiO2 / Ti3C2 have full-band absorption, but the absorbance is weak. After the two are combined, the absorbance of the NGTT composite material increases significantly, proving that the composite material effectively increases light absorption.

[0107] like Figure 7 As shown in Figure 1, the electron migration ability of the material was characterized by electrochemical impedance spectroscopy (EIS). Figure 7As shown, the arc diameter of the NGTT composite material (curve c) is smaller than that of NaYF4:Yb / Tm (curve a) and larger than that of g-C3N4 / TiO2 / Ti3C2 (curve b), indicating that the electron migration ability of the NGTT composite material is between g-C3N4 / TiO2 / Ti3C2 and NaYF4:Yb / Tm, and also indicating that in the NGTT composite material, the electron migration is dominated by g-C3N4 / TiO2 / Ti3C2. After modifying the aptamer, the electron migration ability of Aptamer / NGTT (curve d) is reduced compared with NGTT, because the aptamer is adsorbed on the surface of NGTT, which increases the steric hindrance of interfacial charge migration. When detecting TET, the electron migration ability of TET / Aptamer / NGTT (curve e) is significantly increased, indicating that each step of electrode modification is successful.

[0108] like Figure 8 As shown, the modified electrode was photocurrent detected using an electrochemical workstation. A three-electrode system was used, with TET / Aptamer / NGTT / I TO as the working electrode, platinum wire as the counter electrode, silver / silver chloride as the reference electrode, the electrolyte being 0.1M PBS solution, and the excitation light source being a 980nm wavelength laser. Figure 8 As shown, the photocurrent of NGTT is higher than that of g-C3N4 / TiO2 / Ti3C2, corresponding to light absorption; the photocurrent of Aptamer / NGTT is lower than that of NGTT, corresponding to EIS; the photocurrent of 10nM TET / Aptamer / NGTT is significantly higher than that of Aptamer / NGTT; the photocurrent of 100nM TET / Aptamer / NGTT is also significantly improved, which fully proves that the prepared NGTT composite photoelectrochemical aptamer sensor has a significant photoresponse to TET under near-infrared light excitation.

[0109] like Fig. 9 As shown, the selectivity test of the photoelectrochemical aptamer sensor: tetracycline (TET), oxytetracycline (OXY), kanamycin (KAN), streptomycin (STR), neomycin (NEO), chloramphenicol (CAP), chlortetracycline (CTC), doxycycline (DOX) of the same concentration (10 nM) were prepared and placed in the photoelectrochemical aptamer sensor for testing, as shown in FIG. Fig. 9 As shown, the TET / Aptamer / NGTT electrode exhibits significant selectivity for TET, while the effects of the other antibiotics are negligible.

[0110] like Fig.10 As shown, the stability test of the photoelectrochemical aptamer sensor: Fig.10As shown, the photocurrent of the 10nM TET / Aptamer / NGTT electrode remained basically unchanged after the light source was continuously switched on and off for 400s, indicating that the NGTT composite photoelectrochemical aptamer sensor has significant stability.

[0111] like Fig.11 As shown, the photocurrent response of the photoelectrochemical aptamer sensor to different concentrations of TET is as follows: Fig.11 As shown in the figure, the TET concentration range is 0.5nM-350nM. As the concentration increases, the photocurrent increases from 0.979μA to 4.456μA, and the detection limit is as low as 0.15nM. The results are linearly fitted, as shown in Fig.12 As shown, the linear fitting equations corresponding to the two linear ranges are: (1) y = 0.04698x + 0.02601 (0.5nM-10nM, R 2 =0.99673); (2)y=0.00847x+0.44679(10nM-350nM, R 2 =0.99492).

[0112] Comparative Example 1:

[0113] The difference between this comparative example and Example 1 is that in this comparative example:

[0114] In step S2, the fourth mixed solution containing the up-conversion material NaYF4:Yb,Tm is not added, that is, 0.1 g of Ti3C2 is added to the g-C3N4 supernatant, and then 50 μL of H2O2 is added dropwise, stirred, and the mixed solution is centrifuged and washed three times with ultrapure water and ethanol, and dried in a vacuum oven at 40°C for 12 hours to obtain a g-C3N4 / TiO2 / Ti3C2 composite material;

[0115] In step S3, the g-C3N4 / TiO2 / Ti3C2 composite material is modified on the surface of the ITO electrode to obtain a g-C3N4 / TiO2 / Ti3C2 / ITO electrode.

[0116] Performance Test:

[0117] like Figure 8 As shown, the photocurrent of the g-C3N4 / TiO2 / Ti3C2 / I TO electrode in this comparative example is significantly lower than the photocurrent result of the embodiment of the present invention, and the photoresponse to TET under near-infrared light excitation is significantly poor.

[0118] The embodiments of the present invention are described above in conjunction with the accompanying drawings, but the present invention is not limited to the above-mentioned specific implementation modes, which are merely illustrative rather than restrictive. Under the guidance of the present invention, ordinary technicians in this field can also make many forms without departing from the scope of protection of the present invention and the claims, all of which are within the protection of the present invention.

Claims

1. A method for preparing a photoelectrochemical aptamer sensor for tetracycline detection, characterized in that: include: Dissolving NaOH in ultrapure water, adding ethanol and oleic acid, and stirring to obtain a first mixed solution; Mix YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O, add into NH4F solution, and stir to obtain a second mixed solution; The first mixed solution and the second mixed solution are mixed and stirred to perform a solvothermal reaction to obtain a third mixed solution, and the third mixed solution is centrifuged, washed and dried to obtain an up-conversion material NaYF4:Yb,Tm; The up-conversion material NaYF4:Yb,Tm solution is dissolved in a polyvinylpyrrolidone (PVP) solution to obtain a fourth mixed solution; Prepare a g-C3N4 supernatant by using melamine, add Ti3C2 to the g-C3N4 supernatant, then add H2O2 dropwise, and stir to obtain a fifth mixed solution; The fourth mixed solution and the fifth mixed solution are mixed and stirred, and heated to react to obtain a sixth mixed solution, and the sixth mixed solution is centrifuged, washed and dried to obtain an NGTT composite material; The NGTT composite material is modified onto the surface of an indium tin oxide glass (ITO) electrode and incubated in an aptamer solution to obtain an aptamer / NGTT / ITO electrode, which is the photoelectrochemical aptamer sensor for tetracycline detection.

2. The preparation method according to claim 1, characterized in that: In the g-C3N4 supernatant prepared by melamine: The melamine is used as a raw material to be calcined to obtain g-C3N4, and the g-C3N4 is subjected to water dispersion and ultrasonic centrifugation to obtain the g-C3N4 supernatant.

3. The preparation method according to claim 2, characterized in that: The calcination reaction temperature is 550° C. and the reaction time is 2 h.

4. The preparation method according to claim 1, characterized in that: The molar ratio of YbCl3·6H2O, YCl3·6H2O and TmCl3·6H2O is 79.8:20:0.

2.

5. The preparation method according to claim 1, characterized in that: The step of modifying the NGTT composite material onto the surface of an indium tin oxide glass (ITO) electrode comprises: Pre-treating the indium tin oxide glass (ITO) electrode; Ultrasonic dispersion of the NGTT composite material in ultrapure water to obtain an NGTT aqueous dispersion; The NGTT aqueous dispersion is drop-coated on the conductive surface of the indium tin oxide glass (ITO) electrode.

6. The preparation method according to claim 5, characterized in that: The pretreatment of the indium tin oxide glass (ITO) electrode includes: The slice of indium tin oxide glass (ITO) is immersed in a detergent solution for ultrasonic treatment, ultrapure water and ethanol are added for ultrasonic treatment respectively, and then dried with nitrogen.

7. The preparation method according to claim 5, characterized in that: The NGTT composite material is ultrasonically dispersed in ultrapure water, and the concentration of the obtained NGTT aqueous dispersion is 2 mg / mL.

8. The preparation method according to claim 5, characterized in that: After the NGTT aqueous dispersion is drop-coated on the conductive surface of the indium tin oxide glass (ITO) electrode, it is dried at a drying temperature of 70° C. and a drying time of 12 hours.

9. The preparation method according to claim 1, characterized in that: The heating temperature for mixing and heating the fourth mixed solution and the fifth mixed solution is 80° C., and the reaction time is 1 hour.

10. Use of the photoelectrochemical biosensor prepared according to the preparation method of claims 1-9 for detecting tetracycline, characterized in that: include: The surface of the photoelectrochemical biosensor is incubated in tetracycline (TET) / PBS solutions of different concentrations to obtain TET / aptamer / NGTT / ITO electrodes of different concentrations; A three-electrode system was used: the TET / aptamer / NGTT / ITO electrode was used as the working electrode, the platinum wire was used as the counter electrode, the silver / silver chloride was used as the reference electrode, the electrolyte was a PBS solution, and the excitation light source was a laser with a wavelength of 980 nm. The light response performance of the TET / aptamer / NGTT / ITO electrode was detected on an electrochemical workstation.

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