Preparation method of ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor and application thereof in detection of leather cultural relics

Abstract

This invention relates to the technical field of leather artifact detection, and discloses a method for preparing a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor and its application in leather artifact detection. The method includes the following steps: S1: Dispersing ZnTiO3, thiourea, and bismuth nitrate in a mixture of water and ethylene glycol, adding potassium hydroxide, and continuously stirring until homogeneous; subjecting the resulting solution to a solvothermal reaction to obtain a ZnTiO3 / Bi2O2S heterojunction; S2: Preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor. This invention uses a hydrothermal method to grow a narrow-bandgap Bi2O2S semiconductor on the surface of ZnTiO3 nanomaterials in situ, forming a heterojunction structure. This broadens the photoresponse range of the material and promotes the separation and transmission of photogenerated electron-hole pairs, significantly improving the photoelectric signal intensity of the heterojunction and enhancing detection sensitivity.

Description

Technical Field

[0001] This invention belongs to the technical field of leather artifact testing, and more specifically relates to a method for preparing a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor and its application in leather artifact testing. Background Technology

[0002] Leather artifacts are important historical and cultural relics that bear witness to the development of human civilization. They encompass various types, including ancient leather clothing, leather goods, documents, and armor, recording information about the craftsmanship, social systems, and cultural life of different historical periods. They possess irreplaceable historical and cultural value. However, leather's main component is natural collagen, which, during long-term burial or preservation, is subject to environmental factors such as microbial erosion, temperature and humidity fluctuations, and acid-base oxidation. This leads to degradation and deterioration, resulting in decreased strength, powdering, and cracking, seriously threatening the preservation of leather artifacts. Accurate detection of the degree of deterioration in leather artifacts and understanding their state of degradation are core prerequisites for conducting value assessments and developing scientific protection and restoration plans, and are of paramount importance for the long-term preservation and inheritance of leather artifacts.

[0003] Currently, common methods for testing leather artifacts include sensory evaluation, mechanical property testing, chromatography-mass spectrometry (GC-MS), and traditional enzyme-linked immunosorbent assay (ELISA). These methods all have limitations: sensory evaluation relies heavily on the experience of the testing personnel, is highly subjective, and cannot achieve accurate quantification; mechanical testing has strict requirements on sample size and can cause irreversible secondary damage to precious leather artifacts; GC-MS offers high precision, but its complex procedures, high cost, and long cycle time make it difficult to meet the needs of rapid, batch testing; traditional immunoassay methods lack sensitivity and cannot detect trace markers of leather degradation in low-damage samples, failing to meet the actual needs of leather artifact testing. Therefore, developing a new testing technology that is highly sensitive, requires small sample volumes, has high specificity, and allows for rapid detection is a crucial issue that urgently needs to be addressed in the field of leather artifact conservation. Summary of the Invention

[0004] To address the aforementioned issues, this invention provides a method for fabricating a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor and its application in the detection of leather artifacts. A narrow-bandgap Bi2O2S semiconductor is grown in situ on the surface of ZnTiO3 nanomaterials using a hydrothermal method, forming a pn-type heterojunction structure. This broadens the material's photoresponse range and improves the utilization efficiency of visible light. Furthermore, it promotes the separation and transport of photogenerated electron-hole pairs, significantly enhancing the photoelectric signal intensity of the heterojunction and improving detection sensitivity. In addition, the ZnTiO3 / Bi2O2S heterojunction has a large specific surface area, enabling it to load more immunorecognition molecules, further improving the sensor's detection performance.

[0005] The performance of photoelectroactive materials directly determines the detection sensitivity and stability of photoelectrochemical immunosensors. ZnTiO3, a typical perovskite semiconductor, possesses excellent chemical stability and biocompatibility, but its wide bandgap results in poor visible light response and high photogenerated electron-hole recombination rate, leading to low photoelectric conversion efficiency when used alone, which cannot meet the requirements for high-sensitivity detection. Bi2O2S, a novel layered bismuth-based semiconductor, has a narrow bandgap and excellent visible light absorption, but its stability and carrier separation efficiency are poor when used alone. Combining the two to construct a ZnTiO3 / Bi2O2S heterojunction allows their complementary band structure advantages, extending the composite material's light absorption range from the ultraviolet region to the entire visible light region, significantly improving the utilization efficiency of sunlight. Simultaneously, due to the difference in their Fermi levels, a built-in electric field is formed at the interface. The built-in electric field is the core driving force for the separation of photogenerated electron-hole pairs, effectively promoting the separation and migration of photogenerated carriers and suppressing carrier recombination. Simultaneously, it combines the high stability of ZnTiO3 with the excellent visible light response performance of Bi2O2S, significantly improving the photoelectric conversion efficiency of optoelectronic materials and providing a material basis for constructing high-performance photoelectrochemical immunosensors. Compared with pure-phase Bi2O2S, the heterojunction composite material of this invention significantly improves photocurrent density and photoelectric signal intensity.

[0006] This preparation process utilizes in-situ hydrothermal construction to create a tight Z-shaped heterojunction, eliminating the need for harsh high-temperature conditions and maximizing the preservation of the material's excellent photoelectric properties and biocompatibility. Ti-OS interfacial bonding effectively enhances interfacial charge transport efficiency, suppresses photogenerated carrier recombination, and improves the sensor's photoelectric response sensitivity and signal stability. Simultaneously, the mild bio-crosslinking modification process does not damage the antibody's biological activity, effectively resisting matrix interference from proteins, pigments, and trace salts in leather artifact extracts. This enables non-destructive, trace-level, and precise detection of artifact samples, making it suitable for practical applications such as screening for leather artifact diseases, assessing aging, and monitoring preservation status.

[0007] This invention uses a ZnTiO3 / Bi2O2S heterostructure as the photosensitive material and immobilizes collagen antibodies with polydopamine. The binding mechanism is primarily based on a multi-interface interaction mechanism, with covalent reactions as the main component and non-covalent interactions as secondary components. These interactions are further enhanced by non-covalent interactions such as hydrogen bonding, hydrophobic interactions, and π–π stacking, ensuring the stability and conformational integrity of the antibody. The resulting sensor exhibits high sensitivity, specificity, stability, and repeatability, while causing minimal damage to cultural relics.

[0008] To achieve the above objectives, the present invention is implemented through the following technical solution:

[0009] One of the technical solutions of the present invention: a method for preparing a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor, comprising the following steps:

[0010] S1: Disperse ZnTiO3, thiourea and bismuth nitrate in a mixture of water and ethylene glycol. After vigorous stirring, add potassium hydroxide and stir continuously until homogeneous. The resulting solution undergoes a solvothermal reaction. The product obtained from the reaction is filtered, washed and dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0011] S2: Disperse the ZnTiO3 / Bi2O2S heterojunction in water, drop the resulting dispersion onto the surface of ITO conductive glass, incubate, and wash with PBS; then add PDA solution and collagen antibody solution dropwise, washing with PBS after each incubation; finally block with BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0012] As a preferred embodiment, in S1, the preparation method of ZnTiO3 includes: adding zinc chloride and titanium dioxide to water, then slowly adding sodium hydroxide to the solution and stirring, then carrying out a hydrothermal reaction, centrifuging and drying the product obtained from the reaction, and finally heat-treating the dried powder to obtain ZnTiO3 with a hexagonal structure.

[0013] Preferably, the molar ratio of zinc chloride to titanium dioxide is 1.0-2.0:1; the stirring time after adding sodium hydroxide is 0.5-1.5 h; and zinc chloride and titanium dioxide are added to water to achieve a zinc chloride concentration of 0.2-0.5 mol·L⁻¹. -1 The mass ratio of sodium hydroxide to zinc chloride is 2-2.5:1.

[0014] Preferably, the hydrothermal reaction is carried out at a temperature of 160-200℃ for 12-24 hours; the heat treatment is carried out at a temperature of 600-800℃ for 2-4 hours.

[0015] Preferably, in S1, the molar ratio of thiourea to bismuth nitrate is 0.8-1.5:1; and the molar ratio of ZnTiO3 to bismuth nitrate is 0.3-0.8:1.

[0016] Preferably, in S1, the volume ratio of water to ethylene glycol in the mixture is 1-1.5:1; and the concentration of ZnTiO3 dispersed in the mixture is 4-7 g·L⁻¹. -1 The mass ratio of potassium hydroxide to ZnTiO3 is 2.5-3:1.

[0017] Preferably, in S1, the temperature of the solvothermal reaction is 120-180℃ and the time is 2-6h.

[0018] Preferably, in S2, the concentration of ZnTiO3 / Bi2O2S heterojunctions in the dispersion is 2-4 mg / mL.

[0019] Preferably, in step S2, the incubation temperature of the dispersion is 37°C and the incubation time is 4-6 h.

[0020] Preferably, in step S2, the volume ratio of the dispersion, PDA solution, collagen antibody solution, and BSA solution is 20-50:10-30:10-20:10-30; the concentration of the PDA solution is 2-4 mg / mL; the concentration of the collagen antibody solution is 8-10 μg / mL; and the concentration of the BSA solution is 1-3 wt%.

[0021] The second technical solution of the present invention: the application of a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor in the detection of leather cultural relics.

[0022] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0023] (1) The ZnTiO3 / Bi2O2S heterojunction constructed by the composite has complementary energy bands, which extends the light absorption to the visible light region and significantly improves the light utilization efficiency; the built-in electric field at the interface effectively drives the separation and migration of photogenerated carriers and suppresses recombination; the heterojunction has both high stability and excellent visible light response, and the photoelectric conversion efficiency is greatly improved.

[0024] (2) By constructing a heterojunction in situ through hydrothermal growth, the resulting nanocomposite material has a large specific surface area, which can load more biorecognition molecules such as antibodies, increasing the capture probability of the target. By constructing a heterojunction, the photoresponse range of the material is broadened to the visible light region, and the separation of photogenerated electron-hole pairs is effectively promoted, significantly enhancing the photoelectric signal and further improving the detection sensitivity and selectivity of the sensor.

[0025] (3) The process of this invention is simple, easy to operate, and has mild reaction conditions. The materials used are biocompatible, green and pollution-free. At the same time, it can realize non-destructive, micro-scale and accurate detection of cultural relics samples. It is suitable for the actual application scenarios of screening for diseases of leather cultural relics, assessing the degree of aging and monitoring the preservation status. Compared with traditional detection methods, it has more application prospects. Attached Figure Description

[0026] Figure 1 This is a schematic diagram of the fabrication of a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0027] Figure 2 The images are scanning electron microscope (SEM) images and mapping diagrams of the materials (A: SEM image of ZnTiO3; B: SEM image of Bi2O2S; C: SEM image and mapping diagram of ZnTiO3 / Bi2O2S heterojunction).

[0028] Figure 3 XRD patterns of ZnTiO3, Bi2O2S, and ZnTiO3 / Bi2O2S heterojunctions;

[0029] Figure 4 High-resolution XPS spectra (A: ZnTiO3 / Bi2O2S; B: Zn2P; C: Ti2p; D: O1S; E: Bi4S / S2p).

[0030] Figure 5 Figure A shows the photocurrent response plot and EIS plot (Figure A is the photocurrent response plot, from top to bottom: ITO / ZnTiO3 / Bi2O2S (blue), ITO / ZnTiO3 / Bi2O2S / PDA (dark purple), ITOZnTiO3 / Bi2O2S / PDA / Ab (purple), ITOZnTiO3 / Bi2O2S / PDA / Ab / BSA (green), ITO / ZnTiO3 / Bi2O2S / PDA / Ab / BSA / AG (yellow), ITO (black); Figure B is the EIS plot, from left to right: ITO (a), ITO / ZnTiO3 / Bi2O2S / PDA (b), ITO / ZnTiO3 / Bi2O2S / PDA (c), ITOZnTiO3 / Bi2O2S / PDA / Ab (d), ITOZnTiO3 / Bi2O2S / PDA / Ab / BSA (e)). ITO / ZnTiO3 / Bi2O2S / PDA / Ab / BSA / AG(f));

[0031] Figure 6 The results of the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunoassay sensor for detecting cultural relics. Detailed Implementation

[0032] The following description is merely 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 foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

[0033] Example 1

[0034] S1: 1.36 g of zinc chloride and 0.79 g of titanium dioxide were added to 40 mL of deionized water at a 1:1 molar ratio and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0035] S2: 100 mg ZnTiO3, 0.076 g thiourea and 0.485 g bismuth nitrate were dispersed in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stirred vigorously for 10 min. Then, 0.28 g potassium hydroxide was added and stirred continuously until homogeneous. The resulting solution was transferred to a 50 mL reactor and reacted at 120 °C for 3 h. After cooling to room temperature, the mixture was washed alternately with deionized water and ethanol and then dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0036] S3: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0037] Example 2

[0038] S1: 1.36 g of zinc chloride and 0.531 g of titanium dioxide were added to 40 mL of deionized water at a molar ratio of 1.5:1 and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0039] S2: 100 mg ZnTiO3, 0.076 g thiourea and 0.485 g bismuth nitrate were dispersed in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stirred vigorously for 10 min. Then, 0.28 g potassium hydroxide was added and stirred continuously until homogeneous. The resulting solution was transferred to a 50 mL reactor and reacted at 120 °C for 3 h. After cooling to room temperature, the mixture was washed alternately with deionized water and ethanol and then dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0040] S3: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0041] Example 3

[0042] S1: 1.36 g of zinc chloride and 0.79 g of titanium dioxide were added to 40 mL of deionized water at a 1:1 molar ratio and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0043] S2: 60 mg ZnTiO3, 0.076 g thiourea and 0.485 g bismuth nitrate were dispersed in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stirred vigorously for 10 min. Then, 0.28 g potassium hydroxide was added and stirred continuously until homogeneous. The resulting solution was transferred to a 50 mL reactor and reacted at 120 °C for 3 h. After cooling to room temperature, the mixture was washed alternately with deionized water and ethanol and then dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0044] S3: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0045] Example 4

[0046] S1: 1.36 g of zinc chloride and 0.79 g of titanium dioxide were added to 40 mL of deionized water at a 1:1 molar ratio and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0047] S2: 100 mg ZnTiO3, 0.076 g thiourea and 0.485 g bismuth nitrate were dispersed in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stirred vigorously for 10 min. Then, 0.28 g potassium hydroxide was added and stirred continuously until homogeneous. The resulting solution was transferred to a 50 mL reactor and reacted at 120 °C for 6 h. After cooling to room temperature, the mixture was washed alternately with deionized water and ethanol and then dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0048] S3: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0049] Example 5

[0050] Detection of collagen in leather artifacts:

[0051] The detection employed a three-electrode system on an electrochemical workstation, using Ag / AgCl as the reference electrode, platinum wire as the counter electrode, and the fabricated photoelectrochemical immunosensor as the working electrode. The assay was performed in 15 mL of phosphate buffer solution with a pH of 5.0–8.0. A time-current method was used, with open-circuit voltage as the bias voltage, a light source wavelength of 300–2500 nm, a light intensity of 300 mW / cm², a distance of 15 cm between the light source and the electrode, and a running time of 30 s. After electrode placement, the light was turned on for 10 s every 10 s, and the photocurrent was recorded. A working curve was plotted using a collagen standard solution. The artifact samples were provided by the Xinjiang Uygur Autonomous Region Institute of Cultural Relics and Archaeology. Collagen solutions were extracted from artifact fragments unearthed from the Baiyanghe Tomb in Fukang, dating back approximately 2000 years, and used to replace the standard solutions for detection.

[0052] Comparative Example 1

[0053] The difference between this comparative example and Example 1 is that only the pure phase ZnTiO3 nanopowder obtained in S1 is used as the photosensitive material to construct the ZnTiO3 photoelectrochemical immunosensor.

[0054] Comparative Example 2

[0055] The difference between this comparative example and Example 1 is that only Bi2O2S nanopowder is used as the photosensitive material to construct the Bi2O2S photoelectrochemical immunosensor.

[0056] S1: Disperse 0.076 g thiourea and 0.485 g bismuth nitrate in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stir vigorously for 10 min; then add 0.28 g potassium hydroxide and stir continuously until homogeneous; transfer the resulting solution to a 50 mL reactor and react at 120 °C for 3 h. After cooling to room temperature, wash alternately with deionized water and ethanol, and then dry to obtain Bi2O2S.

[0057] S2: Bi2O2S was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the Bi2O2S photoelectrochemical immunosensor.

[0058] Comparative Example 3

[0059] The difference between this comparative example and Example 1 is that the ZnTiO3 / Bi2O2S heterojunction was prepared by physical mixing rather than in-situ growth.

[0060] S1: 1.36 g of zinc chloride and 0.79 g of titanium dioxide were added to 40 mL of deionized water at a 1:1 molar ratio and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0061] S2: Disperse 0.076 g thiourea and 0.485 g bismuth nitrate in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stir vigorously for 10 min; then add 0.28 g potassium hydroxide and stir continuously until homogeneous; transfer the resulting solution to a 50 mL reactor and react at 120 °C for 3 h. After cooling to room temperature, wash alternately with deionized water and ethanol, and then dry to obtain Bi2O2S.

[0062] S3: Weigh 3 mg ZnTiO3 and 3 mg Bi2O2S and add them to 20 mL of anhydrous ethanol. The system is then sonicated, and after treatment, the solution is stirred for 1 h to ensure homogeneity. The system is then centrifuged and dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0063] S4: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0064] Comparative Example 4

[0065] The difference between this comparative example and Example 1 is that potassium hydroxide, a mineralizing agent, is not added during the hydrothermal synthesis of Bi2O2S.

[0066] S1: 1.36 g of zinc chloride and 0.79 g of titanium dioxide were added to 40 mL of deionized water at a 1:1 molar ratio and stirred for 15 min. Then, 3.19 g of sodium hydroxide was slowly added to the solution, and the mixture was stirred at room temperature for 30 min. The solution was then transferred to a reaction vessel and reacted at 160 °C for 12 h. After the reaction was completed, the white precipitate obtained after cooling was washed with deionized water and dried at 80 °C for 12 h. Finally, the dried powder was heat-treated at 750 °C for 3 h to obtain ZnTiO3.

[0067] S2: 100 mg ZnTiO3, 0.076 g thiourea and 0.485 g bismuth nitrate were dispersed in a mixture of 20 mL deionized water and ethylene glycol (volume ratio 1:1) and stirred vigorously for 10 min. The resulting solution was transferred to a 50 mL reactor and reacted at 120 °C for 3 h. After cooling to room temperature, the mixture was washed alternately with deionized water and ethanol and then dried to obtain a ZnTiO3 / Bi2O2S heterojunction.

[0068] S3: The ZnTiO3 / Bi2O2S heterojunction was dispersed in water to obtain a dispersion; 30 μL of the 3 mg / mL dispersion was added dropwise to the surface of an ITO conductive glass and incubated at 37 °C for 4 h, followed by washing with PBS buffer; then 15 μL of 3 mg / mL PDA solution was added dropwise and incubated at 4 °C for 1 h, followed by washing with PBS buffer; then 20 μL of 8 μg / mL collagen antibody solution was added dropwise and incubated at 37 °C for 1 h, followed by washing with PBS buffer; finally, it was blocked with 20 μL of 1 wt% BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

[0069] Experimental data analysis:

[0070] 1. Characterization of heterojunctions:

[0071] like Figure 1 The diagram shown illustrates the fabrication process of the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor of this invention. Figure 2 , Figure 3 , Figure 4 As shown, this invention has successfully prepared ZnTiO3, Bi2O2S, and ZnTiO3 / Bi2O2S heterojunctions via a hydrothermal method.

[0072] like Figure 2 As shown, the pure ZnTiO3 sample ( Figure 2 In section A), the particles exhibit a densely aggregated, blocky morphology, with tight packing between them, scarce porosity, and limited exposed active surface sites; single Bi2O2S Figure 2While ZnTiO3 / Bi2O2S heterojunction composites possess a two-dimensional lamellar flower cluster structure, the lamellars are severely stacked and adhered together, with a large number of intrinsic surfaces covered, making it difficult to fully utilize surface effects. In contrast, ZnTiO3 / Bi2O2S heterojunction composites ( Figure 2 In section C), a multi-level, loose structure interwoven with particles and ultrathin sheets was constructed: Bi₂O₂S ultrathin nanosheets were uniformly anchored on the surface of ZnTiO₃ particles, effectively preventing the aggregation of ZnTiO₃ particles themselves, while simultaneously expanding the stacked Bi₂O₂S sheets to form abundant pores and open surfaces. Elemental mapping showed that Bi, Zn, Ti, O, and S were uniformly dispersed throughout the material, confirming that the two components achieved a highly uniform composite, further inhibiting large-scale aggregation of the two phases. This loose, multi-level structure significantly increased the specific surface area of ​​the material, providing sufficient active sites for immobilizing loaded immunorecognition molecules, laying the structural foundation for excellent detection performance in photoelectrochemical immunosensing systems.

[0073] like Figure 3 As shown, the crystal phase structure of the materials was characterized using X-ray diffraction (XRD). All diffraction peaks of the single ZnTiO3 sample perfectly matched the standard card JCPDS No. 26-1500, with no impurity peaks, proving the successful preparation of high-purity zinc titanate. The diffraction peaks of pure Bi2O2S corresponded one-to-one with the standard spectral lines of JCPDS No. 97-002-9451, indicating a pure phase free of impurities. For the ZnTiO3 / Bi2O2S composite system, its XRD curve simultaneously contained the characteristic diffraction peaks of both ZnTiO3 and Bi2O2S phases. The characteristic peaks of both components were clearly visible without significant shift, indicating that the composite process did not change the intrinsic crystal structure of the two materials, and no new impurity phases were generated. This confirms the effective composite of ZnTiO3 and Bi2O2S, successfully constructing a ZnTiO3 / Bi2O2S heterojunction.

[0074] like Figure 4 As shown, X-ray photoelectron spectroscopy (XPS) was used to systematically characterize the surface elemental composition and chemical valence state of the material. The full-spectrum result (A) clearly observed characteristic signal peaks for Bi4f, S2p, Ti2p, O1s, and Zn2p, directly proving that the five target elements Zn, Ti, O, Bi, and S coexist on the surface of the composite material, further confirming the successful composite of the two phases. The high-resolution Zn2p spectrum (B) can be separated into two sets of characteristic peaks: 1021.6 eV (Zn2p2) and 1044.71 eV (Zn2P / ), corresponding to Zn... 2+ The standard valence state signal. The Ti 2p fine spectrum (C) shows a Ti 2p1 / 2 peak at 458.31 eV and a Ti 2p3 / 2 peak at 464.41 eV, which belong to Ti within the ZnTiO3 lattice. 4+The 01s spectrum (Figure D) yielded three peaks: 532.78 eV corresponds to hydroxyl oxygen from water molecules adsorbed on the material surface; 531.18 eV represents lattice oxygen defects; and 529.58 eV belongs to the Zn-Ti-0 and Bi-0 metal-oxygen bond lattice oxygen. The abundant oxygen defect sites are beneficial for the separation of photogenerated carriers. The fine spectrum (E) of the Bi4f and S2p composite shows 157.78 eV and 163.08 eV corresponding to Bi4f5 / 2 and Bi4f7 / 2, respectively, representing Bi... 3+ The double peaks at 160.98 eV and 162.38 eV match the S2p1 / 2 and S2p3 / 2 characteristic signals of S. Comparison with the standard binding energies of the single components reveals a slight shift in the characteristic peaks of each element after recombination, indicating a strong electronic interaction between the ZnTiO3 and Bi2O2S interfaces, and demonstrating effective charge transfer at the heterojunction interface.

[0075] like Figure 5 Figure A illustrates the changes in photocurrent response during the layer-by-layer assembly of the sensor. After modification with ZnTiO3 / Bi2O2S (curve a), the photocurrent is significantly enhanced, attributed to the excellent photoelectric conversion capability of this heterojunction. Subsequent modifications with Ab, BSA, and antigen result in a progressively decreasing photocurrent. This is because the introduction of non-conductive biomolecules increases the steric hindrance of the electrode surface, hindering the diffusion of electron donor AA to the electrode surface, thereby suppressing photocurrent generation. EIS technology was also used to characterize the fabrication process of this ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor. Figure 5 As shown in Figure B, the impedance of each layer gradually decreases with the increase of the modification layer, further confirming the successful construction of the sensor.

[0076] 2. Detection of collagen in leather artifacts:

[0077] The detection employed a three-electrode system on an electrochemical workstation, using Ag / AgCl as the reference electrode, platinum wire as the counter electrode, and the fabricated photoelectrochemical immunosensor as the working electrode. The assay was performed in 15 mL of phosphate buffer solution with a pH of 5.0–8.0. A time-current method was used, with open-circuit voltage as the bias voltage, a light source wavelength of 300–2500 nm, a light intensity of 300 mW / cm², a distance of 15 cm between the light source and the electrode, and a running time of 30 s. After electrode placement, the light was turned on for 10 s every 10 s, and the photocurrent was recorded. A working curve was plotted using a collagen standard solution. The artifact samples were provided by the Xinjiang Uygur Autonomous Region Institute of Cultural Relics and Archaeology. Collagen solutions were extracted from artifact fragments unearthed from the Baiyanghe Tomb in Fukang, dating back approximately 2000 years, and used to replace the standard solutions for detection.

[0078] like Figure 6The detection results of the PEC immunosensor on leather artifact samples showed that (a) was the blank control, and (b) and (k) were ten artifact samples, respectively. All leather artifact samples were successfully detected. This result indicates that the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor exhibits excellent recognition performance in different types of artifact samples, effectively distinguishing between leather and non-leather samples, and is particularly suitable for the accurate identification of precious artifact samples with complex matrices.

[0079] 3. Comparative data analysis

[0080] Table 1

[0081] At the same target concentration, the greater the relative change in IT, the stronger the sensor's response to the detected object, i.e., the higher the sensitivity. As shown in Table 1, the data shows that the photoelectrochemical immunosensors in Examples 1-4 have high sensitivity. The ZnTiO3 / Bi2O2S heterojunction can introduce an electron transfer medium to accelerate electron transfer, promote the separation of photogenerated carriers, and thus increase the absorption of visible light and photocurrent.

[0082] Comparative Examples 1 and 2 illustrate the limitations of a single wide-bandgap / narrow-bandgap semiconductor: Pure ZnTiO3 (bandgap approximately 3.04 eV) only exhibits light absorption response in the ultraviolet region, and its photocurrent density under visible light irradiation is extremely low (set as a baseline value of 1.0), directly demonstrating that a single ZnTiO3 cannot meet the requirements of visible light photoelectric detection, thus verifying the necessity of introducing a narrow-bandgap semiconductor to broaden the light response range. This results in a decrease in the photocurrent response value of the composite material. Although pure Bi2O2S has a visible light response, due to the lack of a built-in electric field, photogenerated electron-hole pairs recombine rapidly in the bulk phase, resulting in a photocurrent density that is only 2.5 times the baseline value, far lower than that of the heterojunction of this invention.

[0083] Comparative Example 3 illustrates the irreplaceable nature of the "in-situ growth" interface bonding method: physical mixing can only achieve mechanical contact between ZnTiO3 and Bi2O2S; the interface lacks chemical bonding, making it impossible to form a continuous pn junction barrier region, resulting in an extremely weak built-in electric field. The photocurrent density is only 3.2, far lower than that of Example 1 (9.5). This directly verifies the necessity of the "hydrothermal in-situ growth" technique of this invention—only in-situ growth can enable Bi2O2S to uniformly nucleate on the ZnTiO3 surface and form a tight heterojunction interface through lattice matching.

[0084] Comparative Example 4 illustrates the decisive role of alkaline mineralizers in the pure-phase synthesis of Bi₂O₂S: In an acidic system without added mineralizers, Bi 3+ It cannot undergo directional hydrolysis to form [Bi2O2]. 2+The precursor, characterized by a layered structure, releases sulfur from the sulfur source under acidic conditions. 2- The efficiency was also significantly reduced, resulting in the formation of a large number of impurity phases such as Bi2O3 and Bi2S3 in the final product, making it impossible to form pure phase Bi2O2S. Because the target pn heterojunction could not be effectively constructed, the photocurrent density was only about 1.5 (close to pure ZnTiO3), and obvious impurity peaks appeared in the XRD pattern. This verified the necessity of the key process parameter of "adding sodium hydroxide / potassium hydroxide to adjust the pH to 10~13" in this invention. This alkaline environment is not only the chemical basis for the formation of pure Bi2O2S, but also a prerequisite for its selective growth on the ZnTiO3 surface.

[0085] Unless otherwise specified, the raw materials and equipment used in this invention are all commonly used in the field; unless otherwise specified, the methods used in this invention are all conventional methods in the field.

[0086] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Any simple modifications, alterations, and equivalent transformations made to the above embodiments based on the technical essence of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A method for preparing a ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor, characterized in that, Includes the following steps: S1: Disperse ZnTiO3, thiourea and bismuth nitrate in a mixture of water and ethylene glycol. After vigorous stirring, add potassium hydroxide and stir continuously until homogeneous. The resulting solution undergoes a solvothermal reaction. The product obtained from the reaction is filtered, washed and dried to obtain a ZnTiO3 / Bi2O2S heterojunction. S2: Disperse the ZnTiO3 / Bi2O2S heterojunction in water, drop the resulting dispersion onto the surface of ITO conductive glass, incubate, and then wash with PBS; then add PDA solution and collagen antibody solution dropwise in sequence, and wash with PBS after each incubation. Finally, it was blocked with BSA solution to obtain the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor.

2. The preparation method of the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1, characterized in that, In S1, the preparation method of ZnTiO3 includes: adding zinc chloride and titanium dioxide to water, then slowly adding sodium hydroxide to the solution and stirring, then carrying out a hydrothermal reaction, centrifuging and drying the product obtained from the reaction, and finally heat-treating the dried powder to obtain ZnTiO3 with a hexagonal structure.

3. The preparation method of the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 2, characterized in that, Zinc chloride and titanium dioxide were added to water to make the concentration of zinc chloride 0.2-0.5 mol·L⁻¹. -1 The molar ratio of zinc chloride to titanium dioxide is 1.0-2.0:

1.

4. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 2 or 3, characterized in that, The hydrothermal reaction is carried out at a temperature of 160-200℃ for 12-24 hours; the heat treatment is carried out at a temperature of 600-800℃ for 2-4 hours.

5. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1, characterized in that, In S1, the molar ratio of thiourea to bismuth nitrate is 0.8-1.5:1; the molar ratio of ZnTiO3 to bismuth nitrate is 0.3-0.8:

1.

6. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1, characterized in that, In S1, the volume ratio of water to ethylene glycol in the mixture is 1-1.5:1; the concentration of ZnTiO3 dispersed in the mixture is 4-7 g·L⁻¹. -1 The mass ratio of potassium hydroxide to ZnTiO3 is 2.5-3:

1.

7. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1, 5, or 6, characterized in that, In S1, the temperature of the solvothermal reaction is 120-180℃ and the time is 2-6h.

8. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1, characterized in that, In S2, the concentration of ZnTiO3 / Bi2O2S heterojunction in the dispersion is 2-4 mg / mL.

9. The method for preparing the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in claim 1 or 8, characterized in that, In S2, the incubation temperature of the dispersion is 37°C and the incubation time is 4-6 h.

10. The application of the ZnTiO3 / Bi2O2S heterojunction photoelectrochemical immunosensor as described in any one of claims 1-9 in the detection of leather artifacts.

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