Photoelectrochemical sensor for gentamicin detection

Abstract

The invention relates to the technical field of photoelectrochemical sensors, in particular to a photoelectrochemical sensor for gentamicin detection. The preparation method of the photoelectrochemical sensor for gentamicin detection comprises the following steps: performing first hydrothermal reaction on copper salt, Bi2S3 and 5, 10, 15, 20-tetra (4-pyridine) porphyrin, and then centrifuging, washing and drying to obtain a Bi2S3 coated Cu-TPyP-MOF heterojunction; the photoelectrochemical sensor is obtained by coating an electrode with a heterojunction and then adsorbing an antibody. An n-n type heterojunction is prepared by taking TPyP as a building block network unit, copper salt as a metal node and Bi2S3 as a template, so that the separation of photo-induced electrons and holes can be promoted, the light current response capability is improved, and the constructed sensor has low detection limit (0.041 fg / mL), wide detection range, high selectivity, good reproducibility and good stability.

Description

Technical Field

[0001] The invention relates to the technical field of photoelectrochemical sensors, and in particular to a photoelectrochemical sensor for detecting gentamicin. Background Art

[0002] Gentamycin (GEN) is an aminoglycoside antibiotic that mainly acts on Gram-negative bacteria. GEN is commonly used in veterinary clinics and animal feed additives because of its broad antibacterial spectrum, good efficacy and low price. However, the effective concentration of GEN in serum is close to the toxic concentration, resulting in low safety. It is prone to toxic side effects during application, especially when large doses are used irregularly, which may lead to excessive gene residues in animal food, posing a threat to human health and causing nephrotoxicity and ototoxicity. Therefore, establishing a detection method for GEN residues in animal-derived foods is of great significance to protecting human health and the ecological environment.

[0003] To monitor the therapeutic drug gentamicin, there are two conventional detection methods, namely particle-enhanced turbidity inhibition immunoassay (PETINIA) and competition-based high-throughput fluorescence polarization immunoassay (FPIA). However, these techniques consume a lot of medical resources and require the use of skilled technicians, milliliter-scale sample volumes, and long turnaround times. So far, various advanced analytical methods have been used for high-sensitivity detection of gentamicin, such as microcantilever sensors, immunochromatographic assays, fluorescence, chemiluminescent enzyme immunoassays, optical strategies, integration of fluorescence and colorimetric assays, electrochemical methods, surface plasmon resonance, and photoelectrochemistry (PEC), etc. Among them, PEC immunoassay technology has been widely used in antibiotic determination due to its unique advantages such as low background signal, high sensitivity, low cost, and miniaturized instrumentation. The establishment of PEC sensors is based on rapid photoelectric conversion and efficient and specific recognition of biomolecules. In contrast to the traditional electrochemical analysis process, in the PEC sensing system, light is used as the excitation source and the measured electrical signal is used as the detection signal.

[0004] In the GEN detection process, the selection of electrode materials is crucial. Finding excellent electrode materials that not only have enhanced photoelectric conversion and electron transfer capabilities but also have high bioaffinity for GEN antibodies can create advanced PEC immunosensors for GEN detection.

[0005] Metal-organic frameworks (MOFs), as an emerging class of organic porous frameworks, are formed by linking metal ions with organic linkers, and exhibit highly ordered porosity and large specific surface area, easy functionalization, feasible preparation, tunable structure, and universal designability. Given that organic linkers can effectively absorb light energy and excite electrons into metal-oxygen clusters, resulting in charge transfer to the clusters and the generation of electron-hole pairs, this property endows MOFs with an extraordinary semiconductor-like property. Therefore, a variety of MOFs and MOF-based composites have been used as biological platforms for PEC sensors, which have broad application prospects in the analysis of antibiotics, such as tetracycline, n-1-methyladenine, ofloxacin, and GEN sulfate.

[0006] However, how to construct a highly sensitive and selective photoelectrochemical sensor for gentamicin detection remains an urgent problem to be solved. Summary of the invention

[0007] The invention provides a photoelectrochemical sensor for detecting gentamicin, which solves the problems of low sensitivity and poor selectivity of the existing photoelectrochemical sensor when detecting gentamicin.

[0008] In order to solve the above technical problems, the technical solution of the photoelectrochemical sensor for gentamicin detection of the present invention is:

[0009] A photoelectrochemical sensor for gentamicin detection, the preparation method of the photoelectrochemical sensor comprising the following steps:

[0010] 1) copper salt, Bi2S3 and 5,10,15,20-tetra(4-pyridine)porphyrin are subjected to a first hydrothermal reaction in a solvent, and the reaction is completed by solid-liquid separation to obtain a Bi2S3@Cu-TPyP-MOF heterojunction;

[0011] 2) After coating the Bi2S3@Cu-TPyP-MOF heterojunction obtained in step 1) on the electrode, gentamicin antibody is adsorbed to obtain a photoelectrochemical sensor.

[0012] The present invention improves the prior art and provides a photoelectrochemical sensor for gentamicin detection. A hydrothermal method is adopted to prepare a nn-type Bi2S3@Cu-TPyP-MOF heterojunction with 5,10,15,20-tetra(4-pyridine)porphyrin (TPyP) as a building block network unit, copper salt as a metal node, and Bi2S3 as a template. With the in-situ synthesis of Bi2S3 on the surface of Cu-TPyP-MOF, the band gap of the heterojunction becomes narrower, the light absorption range becomes wider, and an enhanced light adsorption capacity is exhibited, which promotes the separation of photogenerated electrons and holes. In addition, the effective interface generated between Cu-TPyP and Bi2S3 is conducive to enhancing its electrochemical activity and light absorption capacity, and expanding the photocurrent response capacity.

[0013] The photoelectrochemical sensor for gentamicin detection provided by the present invention has an ultra-low detection limit of 0.041 fg / mL, which is significantly lower than the reported GEN biosensor. In addition, the constructed photoelectrochemical sensor also has a wide detection range from 1 fg / mL to 1 ng / mL, and also has high selectivity, good reproducibility and good stability, and has wide practicality in the analysis of GEN in raw milk and sewage.

[0014] In order to further improve the electron separation / transfer ability of the heterojunction, preferably, the molar ratio of copper ions, Bi2S3 and 5,10,15,20-tetra(4-pyridine)porphyrin in the copper salt in step 1) is (1-1.5):(0.5-1):(0.1-1).

[0015] In order to further regulate the three-dimensional structure of the heterojunction, preferably, the temperature of the first hydrothermal reaction in step 1) is 120-150° C. and the time is 24-30 hours.

[0016] In order to further improve the structure of bismuth sulfide and regulate its in-situ growth in the heterojunction, preferably, the preparation method of Bi2S3 in step 1) includes: dissolving bismuth salt and organic sulfide in water and then performing a second hydrothermal reaction, and after the reaction is completed, solid-liquid separation to obtain Bi2S3. This method is used to obtain nanorod-shaped bismuth sulfide.

[0017] In order to further regulate the nanorod-like structure of bismuth sulfide, preferably, the temperature of the second hydrothermal reaction is 180-200° C. and the time is 24-30 h.

[0018] In order to further regulate the nanorod-like structure of bismuth sulfide, preferably, the molar ratio of the bismuth salt to the organic sulfide is (1-1.5): (6-6.5).

[0019] In order to further improve the detection capability of the sensor, preferably, the surface density of the Bi2S3@Cu-TPyP-MOF heterojunction coated on the electrode in step 2) is 0.3-1.2 mg / cm 2 .

[0020] In order to further improve the distribution uniformity of the heterojunction on the electrode of the photoelectrochemical sensor, preferably, the coating includes: dispersing the Bi2S3@Cu-TPyP-MOF heterojunction in water to form a heterojunction suspension, drop-coating the heterojunction suspension on the electrode, and drying after the drop-coating is completed.

[0021] In order to further improve the detection capability of the sensor, preferably, the concentration of the heterojunction suspension is 1-2 mg / mL, and the drop coating volume is 10-15 μL.

[0022] In order to further improve the specific adsorption of gentamicin by the sensor, preferably, the adsorption of gentamicin antibody in step 2) is performed by immersing in a gentamicin antibody solution, the concentration of the gentamicin antibody solution is 1 fg / mL to 1 ng / mL, and the immersion time is 40 to 60 min. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The morphology of Bi2S3@Cu-TPyP-MOF heterojunction;

[0024] Figure 2 PXRD patterns and full XPS spectra of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF;

[0025] Figure 3 XPS spectra of Cu 2p and Bi 4f of Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF;

[0026] Figure 4 The photoelectric performance test results of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF are shown in the figure;

[0027] Figure 5 This is a graph showing the GEN quantitative detection performance test results of the photoelectrochemical sensor of Example 1 of the present invention. DETAILED DESCRIPTION

[0028] The technical concept of the photoelectrochemical sensor for gentamicin detection provided by the present invention is as follows:

[0029] Although MOFs-based composites have the potential to be used in the construction of photoelectrochemical sensors, the rapid recombination of photogenerated electrons and holes on the original MOF electrode greatly limits its application as a PEC biosensor platform. In order to overcome this shortcoming of MOFs, various semiconductors such as CdS, TiO2 and Ag / AgBr are compounded with intrinsic MOFs to form a heterojunction structure. Among them, Bi2S3 is a direct bandgap semiconductor material (1.2-1.7eV) and an inorganic semiconductor material that has received extensive attention from researchers. It has the advantages of high cost-effectiveness, non-toxicity, high chemical stability, and convenient commercial application. It is considered to be a promising PEC electrode material.

[0030] In particular, porphyrin-based MOFs (pMOFs) prepared with porphyrin or metalloporphyrin ligands as linkers have found interesting applications in PEC biosensing due to their high bioaffinity, biochemical and photochemical functionalities. However, the application of pMOFs in PEC bioplatforms is still in its early stages due to the limited development of pMOFs-based bioplatforms, such as PCN-224, PCN-222 or Ce-pMOF. According to previous reports, three-dimensional interpenetrating pMOFs can be synthesized using 5, 10, 15, 20-tetra(4-pyridyl)porphyrin (TPyP) as building blocks, and the porphyrin groups have superior electron separation / transfer capabilities due to their abundant π-π interactions.

[0031] The present invention integrates porphyrin-based Cu-TPyP-MOF and Bi2S to form a nn-type Bi2S3@Cu-TPyP-MOF heterojunction in which Bi2S3 is in situ grown on the surface of Cu-TPyP-MOF. The invention combines the enhanced electron transport and light absorption capacity, large specific surface area and pore size, high biological affinity for GEN antibodies of Cu-TPyP-MOF, and the narrow band gap and visible light absorption capacity of Bi2S3, and establishes an effective interface between Cu-MOF and Bi2S3. The obtained Bi2S3@Cu-TPyP-MOF heterojunction has enhanced separation capacity of photogenerated electrons and holes, and improves the photocurrent signal. The photoelectrochemical sensor constructed by the invention has an ultra-low detection limit (LOD) of 0.041 fg / mL and a wide detection range of 1 fg / mL to 1 ng / mL. It also has high selectivity, good reproducibility and good stability, and has wide practicality in GEN analysis in raw milk and sewage.

[0032] The preparation method of the photoelectrochemical sensor of the present invention is specifically as follows: after dissolving bismuth salt and organic sulfide in water, a second hydrothermal reaction is carried out, and after the reaction is completed, centrifugation, washing and drying are performed to obtain Bi2S3; copper salt, Bi2S3 and 5,10,15,20-tetra(4-pyridine)porphyrin are subjected to a first hydrothermal reaction in a solvent, and after the reaction is completed, centrifugation, washing and drying are performed to obtain a Bi2S3@Cu-TPyP-MOF heterojunction; after coating the Bi2S3@Cu-TPyP-MOF heterojunction on the electrode, it is immersed in a gentamicin antibody solution to obtain a photoelectrochemical sensor.

[0033] In a specific embodiment, the bismuth salt is Bi(NO3)3·5H2O, the organic sulfide is thioacetamide; and the copper salt is Cu(NO3)2.

[0034] In a specific embodiment, the solvent is a mixed solvent of n-butanol and N,N-dimethylformamide, and the volume ratio of n-butanol to N,N-dimethylformamide is (1-2):(1-2).

[0035] In a specific embodiment, after the gentamicin antibody is adsorbed, the incubation is carried out in a BSA solution, and the concentration of the BSA solution is 1-2 wt.%.

[0036] The embodiments of the present invention are further described below in conjunction with specific examples. The chemical reagents involved in the following examples are all commercially available conventional products unless otherwise specified.

[0037] 1. Specific embodiments of the photoelectrochemical sensor for gentamicin detection of the present invention

[0038] Example 1

[0039] The photoelectrochemical sensor for gentamicin detection in this embodiment is prepared as follows:

[0040] 1) Preparation of Bi2S3: 485 mg Bi(NO3)3·5H2O and 451 mg thioacetamide were weighed and dissolved in 60 mL deionized water, and then the solution was ultrasonically dissolved for 10 min using an ultrasonic instrument. The solution was then loaded into a reactor and placed in a high-temperature oven for a second hydrothermal reaction, heated to 180°C, and reacted for 24 h. After the reaction was completed, the product was collected by centrifugation and rinsed with deionized water 3-5 times. Finally, Bi2S3 was obtained by vacuum drying at 60°C overnight.

[0041] 2) Preparation of Bi2S3@Cu-TPyP-MOF heterojunction: Weigh 20mg Cu(NO3)2, 40mgBi2S3 synthesized in step 1) and 10mg TPyP, dissolve in a mixed solvent formed by 10mL n-butanol and 10mL N, N-dimethylformamide (DMF) under ultrasound to form a homogeneous solution, then stir for 30 minutes, and then put the homogeneous solution into a reactor and put it into a high-temperature oven for the first hydrothermal reaction at 120°C for 24h. After the reaction is completed, the product is collected by centrifugation and rinsed 3-5 times with a mixed solvent formed by DMF and ethanol. Finally, vacuum dry overnight at 60°C to obtain the Bi2S3@Cu-TPyP-MOF heterojunction.

[0042] 3) Preparation of photoelectrochemical sensor: First, the glassy carbon electrode was polished with alumina powder. The Bi2S3@Cu-TPyP-MOF heterojunction obtained in step 2) was prepared into a 1 mg / mL heterojunction suspension by deionized water. Then, 10 μL of the heterojunction suspension was dropped on the glassy carbon electrode, so that the surface density of the Bi2S3@Cu-TPyP-MOF heterojunction coated on the glassy carbon electrode was 0.57 mg / cm 2After being dried in an oven, the glassy carbon electrode was immersed in gentamicin (GEN) antibody solution (1fg / mL~1ng / mL, rabbit anti-GEN1 monoclonal antibody, D290574, purchased from Shanghai Shenggong Bioengineering Technology Service Co., Ltd.) for 40min, and the gentamicin antibody was adsorbed on the electrode by van der Waals force. Then, the unadsorbed GEN antibody on the surface of the glassy carbon electrode was washed with PBS and stored at 4°C for further use.

[0043] 2. Experimental Examples

[0044] Experimental Example 1

[0045] In this experimental example, the morphology and components of the Bi2S3@Cu-TPyP-MOF heterojunction were tested and analyzed.

[0046] The morphology of Bi2S3@Cu-TPyP-MOF is shown in Figure 1 As shown, Figure 1 a and b are low-magnification and high-magnification SEM images, respectively. Figure 1 c and d are low-magnification and high-magnification TEM images, respectively. Figure 1 e and f are EDS spectrum and element mapping diagram respectively. Figure 1 From a and b, we can see that the Bi2S3@Cu-TPyP-MOF heterojunction is an irregular spherical structure. Figure 1 c It can be further observed that the structure of the heterojunction is that Bi2S3 nanorods are in-situ grown on stacked Cu-TPyP two-dimensional nanosheets; Figure 1 From the HR-TEM image of (d), it can be seen that the interplanar distance of the lattice fringes of the heterojunction is 0.18 nm, which is caused by the (610) crystal plane of Cu-TPyP-MOF; Figure 1 e shows that the atomic percentages of C, N, O, S, Cu, and Bi are 83.48%, 9.63%, 3.02%, 0.39%, and 0.31%, respectively; Figure 1 f, it can be seen that the C, N, O, S, Cu and Bi elements in Bi2S3@Cu-TPyP-MOF are uniformly distributed in the entire area.

[0047] Powder X-ray diffraction (PXRD) patterns and XPS full spectra of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF are shown in Figure 2 As shown. Among them, the PXRD spectrum is as follows Figure 2As shown in a, Cu-TPyP-MOF has obvious diffraction peaks at 2θ of 4.8, 9.5 and 22.5°, while the diffraction peaks of Bi2S3@Cu-TPyP-MOF at 9.1 and 24.0° can be attributed to Cu-TPyP-MOF, and the diffraction peaks at 32.4, 33.3 and 46.6° are consistent with the diffraction peaks of Bi2S3. The chemical structure and composition of Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF were further studied using XPS technology. Figure 2 The XPS full spectrum of b shows obvious Cu 2p (284.1 eV), C 1s (284.1 eV), N 1s (398.7 eV), O 1s (531.3 eV), Bi 4f (531.3 eV) and S2p (531.3 eV) signals.

[0048] The high-resolution XPS spectra of Cu 2p and Bi 4f of Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF composites are shown in Figure 2. Figure 3 The XPS high-resolution spectrum of Cu 2p includes Cu 2p 3 / 2 (934.5 eV) and Cu 2p 1 / 2 (954.4 eV). It is worth noting that the Cu 2p 3 / 2 The two peak binding energies of the XPS high-resolution spectrum are located at 933.2 eV (Cu 0 ) and 934.8eV(Cu + ), while the Cu 2p 3 / 2 The binding energies are 932.0 eV and 934.5 eV, corresponding to Cu + and Cu 0 . Obviously, compared with Cu-TPyP-MOF, the binding energy of Cu in the Bi2S3@Cu-TPyP-MOF composite material shifts toward the direction of low binding energy by about 0.7 eV. Compared with Bi2S3, the binding energy of Bi 4f in the Bi2S3@Cu-TPyP-MOF heterojunction shifts toward the direction of high binding energy by about 1.0 eV. XPS results show that there is obvious electron transfer from Bi2S3 to Cu-TPyP-MOF at the Bi2S3@Cu-TPyP-MOF heterojunction interface, which is beneficial to enhance its electrochemical activity and light absorption ability.

[0049] Experimental Example 2

[0050] This experimental example tests and analyzes the optical and electrochemical properties of the Bi2S3@Cu-TPyP-MOF heterojunction. The test results of the photoelectric performance of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF are shown in the figure. Figure 4 shown.

[0051] The optical absorption properties of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF were characterized by ultraviolet diffuse reflectance (UV-DRS) technique. Figure 4 As shown in a, it can be observed that several materials have broad absorption from the ultraviolet to the near-infrared region (200-1000nm), which can be attributed to the light-harvesting properties caused by the large cyclic structure of porphyrin. Compared with the single Cu-TPyP-MOF, the absorption intensity of Bi2S3@Cu-TPyP-MOF from the ultraviolet to the near-infrared region is enhanced, which can be attributed to the formation of MOF, which expands its absorption. By fitting the Tauc function with the Kubelka-Munk absorbance, the band gaps of the three materials can be derived.

[0052] The band gap diagrams of the three materials are shown in Figure 2. Figure 4 As shown in b, the band gaps of Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF are 1.35eV, 1.87eV and 1.69eV respectively. From the band gap results, as Bi2S3 is in situ synthesized on the surface of Cu-TPyP-MOF, a heterogeneous structure is generated, which makes the band gap of Bi2S3@Cu-TPyP-MOF narrower, the light absorption range wider, and the light absorption capacity stronger. From the Mott-Schottky image ( Figure 4 c) It can be seen that the slopes of the three materials are all positive, indicating that they are all n-type semiconductors, that is, a nn-type heterojunction is formed on the surface of Bi2S3@Cu-TPyP-MOF. Figure 4 From the fluorescence spectrum of d, it can be seen that the fluorescence intensities of Bi2S3 and Cu-TPyP-MOF are higher than that of the composite material Bi2S3@Cu-TPyP-MOF, indicating that the recombination of photogenerated electron-hole pairs is inhibited in the composite material, so the fluorescence intensity decreases, further proving the successful synthesis of the composite material nn-type heterojunction.

[0053] In order to further explore the electrochemical properties of the materials and verify the formation of the heterojunction, EIS tests and CV scanning tests were performed on Bi2S3, Cu-TPyP-MOF and Bi2S3@Cu-TPyP-MOF using an electrochemical workstation. Figure 4 e is the EISNyquist diagram, from Figure 4 e It can be seen that the electrochemical impedance fitting R of Cu-TPyP-MOF alone ctThe value is about 962Ω, while the electrochemical impedance fitting R ct The value is significantly smaller (438.3Ω), indicating that the nn-type heterojunction formed between Bi2S3 and Cu-TPyP-MOF enhances the electron transfer efficiency and reduces the electrochemical impedance. Figure 4 f is the CV scanning curve, Figure 4 f further illustrates the above phenomenon. The redox peaks of the Bi2S3@Cu-TPyP-MOF composite material are better than those of the single Cu-TPyP-MOF, proving that with the formation of the nn-type heterojunction, the binding of electron-hole pairs inside the material is suppressed, thereby enhancing the electrochemical conductivity of the composite material and increasing the redox ability.

[0054] Experimental Example 3

[0055] This experiment evaluates the GEN quantitative detection performance of the photoelectrochemical sensor. The test results are as follows: Figure 5 shown.

[0056] 1) Photoelectric response capability

[0057] First, the photocurrent response ability of the photoelectrochemical sensor during construction was tested. The glassy carbon electrode (GCE) coated with the Bi2S3@Cu-TPyP-MOF heterojunction was recorded as Bi2S3@Cu-TPyP-MOF / GCE, and then impregnated with GEN antibody and recorded as Antibody / Bi2S3@Cu-TPyP-MOF / GCE. Antibody / Bi2S3@Cu-TPyP-MOF / GCE was incubated with BSA solution (1wt%) to eliminate the nonspecific adsorption between the antibody and the Bi2S3@Cu-TPyP-MOF matrix. After incubation, it was recorded as BSA / Antibody / Bi2S3@Cu-TPyP-MOF / GCE. BSA / Antibody / Bi2S3@Cu-TPyP-MOF / GCE was used to detect GEN, which was recorded as GEN / BSA / Antibody / Bi2S3@Cu-TPyP-MOF / GCE. The photocurrent response curves of each stage in the above process are shown in the figure. Figure 5 As shown in a, from Figure 5a It can be seen that Bi2S3@Cu-TPyP-MOF / GCE has a large PEC response, and the response photocurrent intensity can reach about 0.78μA; with the adsorption of antibody molecules, the photocurrent response dropped from 0.82μA to about 0.58μA. This is because a large number of antibody molecules are anchored on the porous Bi2S3@Cu-TPyP-MOF surface through van der Waals forces, electrostatic adsorption and hydrophobic effects. The anchored antibody layer hinders the photoelectron transfer and electron transport at the electrode and electrolyte interface, thus resulting in a decrease in the photocurrent signal; after BSA incubation, the photocurrent response dropped from 0.58μA to about 0.35μA. This result shows that there are certain nonspecific interactions in the sensing system; when detecting GEN (1fg / mL), the photocurrent is further reduced to about 0.29μA. This is because the specific recognition reaction between the antibody and the GEN antigen, their combination can further inhibit the photoelectric conversion and prevent the transfer of electrons. Therefore, the developed Bi2S3@Cu-TPyP-MOF-based PEC biosensor has great potential for quantitative detection of GENs in complex environments.

[0058] Due to the enhanced biosensing ability of the developed Bi2S3@Cu-TPyP-MOF-based biosensor, it can be better used for the quantitative detection of GEN. The detection limit (LOD) of the constructed biosensor was evaluated by measuring a series of GEN concentrations, as follows: the constructed Bi2S3@Cu-TPyP-based PEC biosensor electrode was immersed in different concentrations of GEN target solutions (1fg / mL, 10fg / mL, 100fg / mL, 1.0pg / mL, 10pg / mL, 100pg / mL and 1ng / mL) and the corresponding photocurrent was recorded.

[0059] The photocurrent response curves of different concentrations of GEN are shown in Figure 2. Figure 5 As shown in b, from Figure 5 b It can be seen that as the concentration of GEN increases from 1fg / mL to 1ng / mL, the photocurrent response caused by GEN detection becomes smaller and smaller. This is due to the gradual increase in the binding interaction between GEN and the antibody.

[0060] The curve of the photocurrent response change △I caused by different concentrations of GEN detection and GEN concentration is shown in the figure Figure 5 c shows that as the GEN level increases from 1 fg / mL to 10 pg / mL, the obtained ΔI (ΔI = I BSA -I GEN) also increased significantly. When the GEN concentration exceeded 100 pg / mL, the value approached a platform (△I=0.3μA). Taking ΔI as the logarithmic function of GEN concentration, the linear relationship obtained is ΔI(μA)=0.033+0.61logCon GEN (fg / mL) linear regression equation (correlation coefficient R 2 =0.9907, n=3). According to LOD=3SD / k (SD is the standard deviation of the blank sample, k is the slope of the standard curve), the obtained LOD is 0.041 fg / mL.

[0061] By comparing the performance of some reported biosensors for detecting GEN (as shown in Table 1), the PEC biosensor based on Bi2S3@Cu-TPyP-MOF heterojunction assembly of the present invention has an ultra-low LOD in a wide linear detection range of 1fg / mL to 1ng / mL. Based on the characteristics of Bi2S3@Cu-TPyP-MOF heterojunction, the improved biosensing performance of the obtained PEC biosensor can be attributed to the following aspects: (i) Cu in Cu-TPyP-MOF 2+ / Cu + The redox pairs of substances such as Cu2S3 and porphyrin can accelerate electron transfer and light adsorption; (ii) Due to the large specific surface area and pore size, high functionalization and regular distribution of copper nodes of Cu-TPyP-MOF, abundant antibodies can be anchored not only on the MOF surface but also inside the skeleton. Therefore, this structural advantage can improve the detection sensitivity of GEN and a wide linear detection range; (iii) The formation of Bi2S3 and Cu-TPyP-MOF heterojunction can promote the separation of photogenerated electrons and holes, thereby improving the efficiency of photoelectric conversion, which can further amplify the photoelectric response and help improve the adaptive ability.

[0062] Table 1 Performance comparison of Bi2S3@Cu-TPyP-MOF-based biosensor and other sensors for detecting GEN.

[0063] Material Detection Methods Detection range Detection limit References Lys-AuNPs Colorimetric 5-60nM 1.22nM [1] <![CDATA[MoS2@MWCNTs]]> DPV <![CDATA[1×10 -6 -80μg / mL]]> <![CDATA[1×10 -6 μg / mL]]> [2] GNPs Colorimetric <![CDATA[2.67-33.93ng mL -1 ]]> <![CDATA[0.354ng mL -1 ]]> [3] Ag / PDDA SPR <![CDATA[5×10 -8 M-1×10 -4 M]]> <![CDATA[1×10 -9 M]]> [4] QD fluorometric 1.03-16.55mM 0.493mM [5] <![CDATA[Bi2S3@Cu-TPyP-MOF]]> PEC 1fg / mL-1ng / mL 0.041fg / mL The present invention

[0064] The references in Table 1 are as follows:

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[0070] 2) Selectivity, stability and reproducibility of photoelectrochemical sensors

[0071] The selectivity of the constructed PEC biosensor is another important indicator for the effective detection of GEN in the pollution system. To detect the selectivity, different antibiotics such as midecamycin (MY), tetracycline (TET), kanamycin (KAN), cefotetan (CTE), enrofloxacin (ENR), ciprofloxacin (CIP) and norfloxacin (NFX) were used as interferents, where the levels of these interferents were set to 100 fg / mL, which is 100 times higher than that of GEN (1 fg / mL). The selectivity was analyzed by comparing the photocurrent responses caused by the detection of GEN and interferents.

[0072] Selective test results such as Figure 5 As shown in Figure d, the Bi2S3@Cu-TPyP-MOF-based biosensor was used to determine other interferents, including midecamycin (MY), tetracycline (TET), kanamycin (KAN), cefotetan (CTE), enrofloxacin (ENR), ciprofloxacin (CIP) and norfloxacin (NFX), as well as a mixture (MIX) of the seven interferents and GEN. The concentration of each of these interferents (100 fg / mL) was set to 100 times the detection concentration of GEN (1 fg / mL), but it could only cause a slight change in the photocurrent response, which was significantly smaller than the detection of GEN, and the photoelectric response of the detection mixture was almost the same as the response intensity when GEN was detected alone. These findings indicate that the developed PEC biosensor still has superior selectivity even when detecting GEN in a complex environment containing multiple interferents.

[0073] Six Bi2S3@Cu-TPyP-MOF-based biosensors were assembled in parallel to evaluate the reproducibility of the biosensor. The reproducibility test results of the six photoelectrochemical sensors are shown in Figure 2. Figure 5 As shown in Figure e, the photocurrent signals of the six photoelectrochemical sensors used for GEN analysis (1 fg / mL) are very close, and their relative standard deviation (RSD) is low, at 1.51%. This finding indicates that the biosensor developed in the present invention has excellent reproducibility.

[0074] In order to further explore the stability of the constructed photoelectrochemical sensor, the photoelectrochemical sensor was used to test the changes in the photoelectric response intensity caused by GEN (1 fg / mL) in the same period of time for 15 consecutive days. The stability test results are shown in Figure 2. Figure 5 As shown in Figure 5, the PEC intensity tested every day was found to be comparable, which was mainly due to the high photoelectric conversion performance of the Bi2S3@Cu-TPyP-MOF heterojunction, resulting in good and stable sensing performance.

[0075] In summary, the developed Bi2S3@Cu-TPyP-MOF-based PEC biosensor has excellent selectivity, good reproducibility and outstanding stability.

[0076] Experimental Example 4

[0077] This experimental example evaluates the practicality of the photoelectrochemical sensor of the present invention. In order to further explore the real sample detection capability of the developed biosensor, the constructed photoelectrochemical sensor was used to detect GEN in real samples, and the spiked method was used for recovery experiments. Milk and sewage were selected as actual samples, and GEN of different standard concentrations was added. The linear fitting curve obtained above was combined with the PEC response curve obtained by detecting the actual sample to infer the theoretical GEN content in the actual sample solution, and compared with the actual sample addition amount, and the recovery rate was calculated to verify its practicality.

[0078] The test results of the spiked recovery of GEN in sewage and milk are shown in Table 2. As can be seen from Table 2, when the photoelectrochemical sensor of the present invention detects GEN in sewage, the recovery rate is 86.7% to 126.13%, and the relative standard deviation RSD is 1.73% to 4.89%; when detecting GEN in milk, the recovery rate is 92.53% to 111.58%, and the relative standard deviation RSD is 1.07% to 4.91%. These findings reveal the excellent practicality of the constructed PEC biosensor.

[0079] Table 2 Recovery of GEN in sewage and milk determined by Bi2S3@Cu-TPyP-MOF-based biosensor

[0080]

[0081] Finally, it should be noted that the above is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A photoelectrochemical sensor for detecting gentamicin, characterized in that: The preparation method of the photoelectrochemical sensor comprises the following steps: 1) The copper salt, Bi2S3 and 5,10,15,20-tetra(4-pyridine)porphyrin are subjected to a first hydrothermal reaction in a solvent, and the reaction is completed by solid-liquid separation to obtain a Bi2S3@Cu-TPyP-MOF heterojunction; 2) After coating the Bi2S3@Cu-TPyP-MOF heterojunction obtained in step 1) on the electrode, gentamicin antibody was adsorbed to obtain a photoelectrochemical sensor.

2. The photoelectrochemical sensor for detecting gentamicin according to claim 1, characterized in that: In the step 1), the molar ratio of copper ions, Bi2S3 and 5,10,15,20-tetrakis(4-pyridine)porphyrin in the copper salt is (1-1.5):(0.5-1):(0.1-1).

3. The photoelectrochemical sensor for detecting gentamicin according to claim 1, characterized in that: The temperature of the first hydrothermal reaction in step 1) is 120-150° C. and the time is 24-30 hours.

4. The photoelectrochemical sensor for detecting gentamicin according to claim 1, characterized in that: The preparation method of Bi2S3 in step 1) comprises: dissolving bismuth salt and organic sulfide in water and then performing a second hydrothermal reaction, and after the reaction is completed, solid-liquid separation is performed to obtain Bi2S3.

5. The photoelectrochemical sensor for detecting gentamicin according to claim 4, characterized in that: The temperature of the second hydrothermal reaction is 180-200° C., and the time is 24-30 hours.

6. The photoelectrochemical sensor for detecting gentamicin according to claim 4, characterized in that: The molar ratio of the bismuth salt to the organic sulfide is (1-1.5): (6-6.5).

7. The photoelectrochemical sensor for detecting gentamicin according to claim 1, characterized in that: In step 2), the surface density of the Bi2S3@Cu-TPyP-MOF heterojunction coated on the electrode is 0.3-1.2 mg / cm 2 .

8. The photoelectrochemical sensor for detecting gentamicin according to claim 1 or 7, characterized in that: The coating comprises: dispersing the Bi2S3@Cu-TPyP-MOF heterojunction in water to form a heterojunction suspension, dripping the heterojunction suspension on the electrode, and drying after the dripping is completed.

9. The photoelectrochemical sensor for detecting gentamicin according to claim 8, characterized in that: The concentration of the heterojunction suspension is 1-2 mg / mL, and the drop coating volume is 10-15 μL.

10. The photoelectrochemical sensor for detecting gentamicin according to claim 1, characterized in that: In the step 2), the gentamicin antibody is adsorbed by immersing in a gentamicin antibody solution, the concentration of the gentamicin antibody solution is 1 fg / mL to 1 ng / mL, and the immersion time is 40 to 60 minutes.

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