Method for inactivating Aspergillus niger spores in water

Abstract

The present invention provides a method for inactivating Aspergillus niger spores in a water body, relates to the technical field of inactivation of pathogenic microorganisms in water bodies, and aims to solve or improve the technical problem of poor inactivation effect of Aspergillus niger spores in the prior art. The method for inactivating Aspergillus niger spores in a water body comprises: preparing a silver single atom-graphite phase carbon nitride composite material by a cryo-photoreduction method; adding ClO2 to a water body to be tested containing Aspergillus niger spores under lightless conditions; and placing the silver single atom-graphite phase carbon nitride composite material into the water body to be tested under visible light irradiation conditions to inactivate the Aspergillus niger spores. The method for inactivating Aspergillus niger spores in a water body provided by the present invention, wherein ClO2 can destroy the melanin protective layer of the Aspergillus niger spores, so that the spore cells are exposed to the outside, and the silver single atom-graphite phase carbon nitride composite material can be in close contact with the exposed spore cells through adsorption, thereby acting as an efficient photocatalyst to achieve inactivation of the spores at the interface of the spores.

Description

Technical Field

[0001] The invention relates to the technical field of inactivation of pathogenic microorganisms in water bodies, and in particular to a method for inactivating Aspergillus niger spores in water bodies. Background Art

[0002] Due to the dual pressures of population growth and climate change, drinking water shortages are expected to persist in the future. The United Nations has established a Sustainable Development Goal to ensure public access to safe and affordable drinking water. However, the presence of pathogenic microorganisms in aquatic environments poses a significant threat to drinking water safety. Fungi, among the most resistant pathogens, have been shown to be widespread in surface water, tap water, and drinking water systems. Fungal metabolites can deteriorate water quality, and mycotoxins can cause illnesses such as skin infections, asthma, and pneumonia. An estimated 1.7 million people died from fungal diseases in 2020, and fungal contamination caused economic losses of hundreds of billions of dollars. Water utilities worldwide have established regulations to limit the number of fungi in drinking water. For example, one water utility has proposed a regulation limiting the number of fungi in drinking water supplies to no more than 100 CFU (colony-forming units) per 100 mL. Therefore, effective control of fungi in water is crucial to ensuring drinking water safety.

[0003] Aspergillus niger belongs to the genus Aspergillus and is frequently detected in drinking water. Aspergillus niger reproduces through spores, making it more environmentally tolerant than bacteria and viruses. The cell wall of Aspergillus niger spores is covered with melanin, a negatively charged hydrophobic polymer composed of phenol or indole compounds. Melanin has a high reactivity to oxidative disinfectants (such as free chlorine and chlorine dioxide), so the disinfectants will preferentially react with melanin and be consumed, thereby losing their inactivation effect on the spores. According to literature reports, 4.0 mg·L -1 Free chlorine cannot effectively inactivate 10 6 CFU·mL -1 of Aspergillus niger spores, while only 1mgL -1 Free chlorine can inactivate nearly 6.0 log 10 The high resistance of Aspergillus niger spores to chlorine-based disinfectants requires increased disinfectant dosage to achieve the desired inactivation effect, which not only increases costs but also may increase the risk of disinfectant residues and the formation of carcinogenic disinfection by-products.

[0004] To address these issues, researchers have developed a number of new methods to inactivate Aspergillus niger spores in water. For example, combining ultraviolet irradiation with low-dose chlorine can effectively inactivate about 2.0 log of Aspergillus niger spores within 1 hour. 10(99%). Ultraviolet light primarily destroys the melanin protective layer of cells, thereby facilitating the entry of reactive oxygen species (ROS) generated by UV-induced chlorine photolysis. However, UV / chlorine disinfection technology faces challenges in the production of carcinogenic byproducts and the high energy consumption of UV light sources. To address these challenges, researchers have explored clean and sustainable photocatalytic disinfection technologies driven by visible light to inactivate Aspergillus niger spores. For example, palladium-carbon-doped titanium dioxide photocatalysts effectively inhibit the activity of Aspergillus niger. However, the main drawback of this method is its low inactivation rate, typically requiring up to 100 hours to completely inactivate Aspergillus niger spores. This is because the melanin on the surface of Aspergillus niger acts as a protective barrier, preventing the photocatalyst from directly contacting the spore membrane, thereby weakening the interaction between the cell and the photocatalyst. Therefore, developing efficient and clean disinfection processes to remove Aspergillus niger from water bodies has become a pressing task and is of great significance for improving water quality and hygiene. Summary of the Invention

[0005] The present invention aims to solve or improve the technical problem of poor inactivation effect of Aspergillus niger spores in the prior art.

[0006] The invention provides a method for inactivating Aspergillus niger spores in water.

[0007] The present invention provides a method for inactivating Aspergillus niger spores in water, comprising: obtaining a water body to be tested containing Aspergillus niger spores; preparing a silver single atom-graphite phase carbon nitride composite material by a cryo-photoreduction method; adding ClO2 to the water body to be tested under light-freezing conditions; and placing the silver single atom-graphite phase carbon nitride composite material into the water body to be tested under visible light irradiation conditions to inactivate the Aspergillus niger spores in the water body to be tested.

[0008] The present invention provides a method for inactivating Aspergillus niger spores in water. ClO2 destroys the melanin protective layer of the Aspergillus niger spores, exposing the spore cells. The silver single atom-graphite carbon nitride composite material then closely contacts the exposed spore cells through adsorption, acting as a highly efficient photocatalyst to inactivate the spores at the interface. Furthermore, because the low concentration of ClO2 is completely consumed by melanin, no chlorinated disinfectant remains in the water, and no carcinogenic byproducts such as chlorite are generated in the water.

[0009] The sterilization mechanism of the silver single atom-graphite carbon nitride composite material is that when the composite is exposed to visible light, g-C3N4 (graphite carbon nitride) absorbs photons and excites them to generate electron-hole pairs. The Ag single atom acts as a highly efficient co-catalyst, capturing these electrons and reducing recombination, thereby enhancing photocatalytic efficiency. Simultaneously, the Ag single atom injects the captured electrons into the cell membrane of the spores in close contact with it, disrupting the membrane structure and inhibiting their metabolic activity, ultimately achieving a highly effective inactivation effect.

[0010] In some technical solutions, optionally, in the silver single atom-graphite phase carbon nitride composite material, the mass percentage of silver in the silver single atom-graphite phase carbon nitride composite material is greater than or equal to 1% and less than or equal to 5%.

[0011] In this technical solution, in the silver single atom-graphite phase carbon nitride composite material, the mass percentage of silver in the silver single atom-graphite phase carbon nitride composite material is greater than or equal to 1% and less than or equal to 5%, for example, it can be 1%, 3% or 5%. Among them, when the mass of silver is too high, it will aggregate to form larger nanoparticles instead of maintaining a single atomic dispersion state. This will reduce the number of effective active sites and reduce the photocatalytic efficiency. If the Ag single atom loading is too low, it is difficult to provide enough co-catalytic sites to capture photogenerated electrons, and the photocatalytic efficiency cannot be significantly improved.

[0012] In some technical solutions, optionally, the wavelength of visible light is greater than or equal to 400 nm and less than or equal to 760 nm.

[0013] In this technical solution, the wavelength of visible light is greater than or equal to 400 nm and less than or equal to 760 nm. If it is less than 400 nm, it belongs to the ultraviolet light range, or if it is greater than 760 nm, the photocatalytic efficiency will be significantly reduced.

[0014] In some technical solutions, optionally, the illumination intensity of visible light is greater than or equal to 50 mW·cm -2 , and less than or equal to 200mW·cm -2 .

[0015] In this technical solution, the illumination intensity of visible light is greater than or equal to 50mW·cm -2 , and less than or equal to 200mW·cm -2 For example, it can be 50mW·cm -2 、100mW·cm -2 or 200 mW·cm -2The rate of photocatalytic reaction usually follows the saturation effect of light intensity. After exceeding a certain light intensity, the increase in reaction rate gradually becomes limited or even tends to be stable. Therefore, excessively high light intensity does not significantly increase the reaction rate, but instead causes the extra light energy to be wasted. Lower light intensity is not enough to fully excite the electrons in the photocatalyst, resulting in a limited number of electron-hole pairs generated, which in turn affects the efficiency of the entire photocatalytic sterilization process.

[0016] In some technical solutions, optionally, before the step of adding ClO2 to the water body to be tested, the step also includes: calculating the actual concentration of Aspergillus niger spores in the water body to be tested; wherein the amount of ClO2 and silver single atom-graphite phase carbon nitride composite material added is related to the actual concentration of Aspergillus niger spores.

[0017] In this technical solution, since the actual concentration of Aspergillus niger spores in different water bodies is different, it is necessary to measure the actual concentration of Aspergillus niger spores in the water body to be tested in advance, and then determine the amount of ClO2 and silver single atom-graphite phase carbon nitride composite material to be added based on the actual concentration of Aspergillus niger spores in the water body to be tested, so as to ensure the inactivation effect.

[0018] In some technical solutions, optionally, the amount of ClO2 added = the volume of the water to be tested × the standard concentration of ClO2 × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, wherein the standard concentration of ClO2 is greater than or equal to 8.0 mg·L -1 , and less than or equal to 10.0 mg·L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 .

[0019] In some technical solutions, optionally, the amount of the silver single atom-graphite carbon nitride composite material added = the volume of the water to be tested × the standard concentration of the silver single atom-graphite carbon nitride composite material × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, wherein the standard concentration of the silver single atom-graphite carbon nitride composite material is equal to 200 mg·L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 .

[0020] In these technical solutions, the inactivation effect is ensured by constructing the relationship between the added amount of ClO2 and silver single atom-graphite phase carbon nitride composite material and the actual concentration of Aspergillus niger spores.

[0021] In some technical solutions, optionally, the steps of preparing a silver single atom-graphite phase carbon nitride composite material by a cryophotoreduction method specifically include: adding graphite phase carbon nitride sheets and silver nitrate solution to deionized water to obtain a solid-liquid mixture, and then quickly freezing the solid-liquid mixture in liquid nitrogen to form an ice crystalline solid, irradiating the ice crystalline solid with an ultraviolet lamp, and maintaining the ice crystalline solid at a first preset temperature to obtain a silver single atom-graphite phase carbon nitride composite material.

[0022] In some technical solutions, optionally, the first preset temperature is greater than or equal to -5°C and less than or equal to 0°C. For example, it can be -5°C, -2°C or 0°C.

[0023] In some technical solutions, optionally, the wavelength of the ultraviolet light emitted by the ultraviolet lamp is greater than or equal to 360 nm and less than or equal to 370 nm; for example, it can be 360 ​​nm, 365 nm or 370 nm.

[0024] In some technical solutions, optionally, the intensity of the ultraviolet light emitted by the ultraviolet lamp is greater than or equal to 50mW·cm -2 , and less than or equal to 60mW·cm -2 ; For example, it can be 50mW·cm -2 、52.8mW·cm -2 or 60 mW·cm -2 .

[0025] In this technical solution, in the process of preparing silver single atom-graphite phase carbon nitride composite materials, 10g of urea was loaded into a crucible and placed in a muffle furnace, heated in an air atmosphere at 550°C for 4h with a heating rate of 2°C / min to obtain bulk g-C3N4. 0.4g of bulk g-C3N4 was ground into powder, evenly spread on aluminum foil, and heated again in an air atmosphere at 500°C for 2h with a heating rate of 5°C / min to obtain exfoliated g-C3N4 nanosheets. 20mg of g-C3N4 nanosheets and 24.3μL of silver nitrate solution (concentration of 67.9mg·mL -1 ) was added to 10 mL of deionized water and stirred for 12 h to allow g-C3N4 to fully adsorb Ag ions. The mixture was quickly frozen in liquid nitrogen to form ice crystals and irradiated under a UV lamp for 10 min. The UV wavelength was 365 nm and the light intensity on the surface of the reaction system was 52.8 mW·cm -2 , the ambient temperature is kept below 0°C to keep the ice crystals from melting, thereby preventing the nucleation of Ag and ultimately forming a silver single atom-graphite phase carbon nitride composite material.

[0026] In some technical solutions, optionally, the volume of the water body to be tested is 50 ml, and the concentration of Aspergillus niger spores in the water body to be tested is 10 6 cells·mL-1 The method for inactivating Aspergillus niger spores in water also includes: transferring the inactivated water to be tested into a centrifuge tube and centrifuging to obtain a spore precipitate; placing the spore precipitate in sterile physiological saline to form a spore suspension; adding a fluorescent dye to the spore suspension to stain the Aspergillus niger spores in the spore suspension, wherein the fluorescent dye includes a first fluorescent dye for staining living spores and a second fluorescent dye for staining dead spores; introducing the dyed spore suspension into a flow cytometer, and determining the inactivation rate of the Aspergillus niger spores by the flow cytometer.

[0027] In this technical solution, the present invention can not only inactivate Aspergillus niger spores in water, but also detect the inactivation rate. Since the low concentration of ClO2 is completely consumed by melanin, no chlorinated disinfectant will remain in the water, and no carcinogenic byproducts such as chlorite will be generated in the water, thereby improving safety.

[0028] In some technical solutions, optionally, the excitation light wavelength of the first fluorescent dye is greater than or equal to 480 nm and less than or equal to 500 nm, for example, it can be 488 nm, the emission light wavelength of the first fluorescent dye is greater than or equal to 500 nm and less than or equal to 530 nm, for example, it can be 530 nm, the excitation light wavelength of the second fluorescent dye is greater than or equal to 488 nm and less than or equal to 546 nm, for example, it can be 488 nm, and the emission light wavelength of the second fluorescent dye is greater than or equal to 600 nm and less than or equal to 650 nm, for example, it can be 630 nm.

[0029] In some technical solutions, the first fluorescent dye optionally includes one or a combination of the following: SYTO 9 dye (a dye from the SYTO dye series), SYBR Green I dye (SYBR Green I Nucleic Acid Gel Stain), and DAPI (4',6-diamidino-2-phenylindole) dye. Specifically, the fluorescent dye for staining viable spores can be SYTO 9 dye, a dibenzimidazole dye with an excitation wavelength of approximately 480 nm, an emission wavelength of approximately 500 nm, and can emit green fluorescence. Alternatively, SYBR Green I dye can be a cyanine dye with two ring structures, an excitation wavelength of approximately 497 nm, an emission wavelength of approximately 520 nm, and can emit green fluorescence.

[0030] In some technical solutions, the second fluorescent dye optionally includes one or a combination of the following: propidium iodide, 7-AAD (7-Aminoactinomycin D), and ethidium bromide. Specifically, the fluorescent dye used to stain dead spores can be propidium iodide, which has an excitation wavelength of approximately 490 nm to 535 nm and an emission wavelength of approximately 617 nm (red fluorescence). Alternatively, it can be 7-AAD, which has an excitation wavelength of approximately 546 nm and an emission wavelength of approximately 647 nm (red fluorescence). Alternatively, it can be ethidium bromide, which has an excitation wavelength of approximately 500 nm to 540 nm and an emission wavelength of approximately 605 nm (orange-red fluorescence).

[0031] It should be noted that, in the actual inactivation rate test process, the existing water body can be directly used, and of course a spore suspension can also be prepared. The steps of preparing the spore suspension specifically include: applying the Aspergillus niger powder suspension on a potato dextrose agar plate and placing it for a preset time to activate and form an Aspergillus niger spore group; washing the Aspergillus niger spore group on the potato dextrose agar plate with sterile saline and resuspending it to obtain a spore suspension; filtering the spore suspension with sterile gauze to obtain a first spore suspension.

[0032] In this technical solution, when preparing the first spore suspension, 200 μL of Aspergillus niger powder suspension can be evenly spread on a potato dextrose agar plate and cultured at 25°C for 5 days to activate the strain; the Aspergillus niger spores on the plate are rinsed with 10.0 mL of sterile saline (0.85% NaCl) and resuspended, the obtained spore suspension is filtered twice with three layers of sterile gauze to remove mycelia, and then centrifuged and washed three times at a speed of 8000 rpm to obtain a spore suspension with higher purity, and the high-concentration spore suspension is diluted to a standard concentration of 10 using the counting function of a flow cytometer. 6 cells·mL -1 .

[0033] Additional aspects and advantages of the present invention will become apparent from the following description or may be learned by practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] The above and / or additional aspects and advantages of the embodiments of the present invention will become apparent and readily understood from the description of the embodiments in conjunction with the following drawings, in which:

[0035] Figure 1a is a scanning electron microscope image of untreated Aspergillus niger spores;

[0036] Figure 1b This is a scanning electron microscope image of Aspergillus niger spores treated with ClO2 and Ag1 / CN;

[0037] Figure 2a The dot plot shows the relationship between SSC-H and FSC-H of the stained spore samples;

[0038] Figure 2b This is the dot matrix of the sample containing 40% dead spores;

[0039] Figure 3a This is one of the transmission electron microscope images of Ag1 / CN;

[0040] Figure 3b This is the second transmission electron microscope image of Ag1 / CN;

[0041] Figure 3c AC-HAADF-STEM image of Ag1 / CN;

[0042] Figure 4a Schematic diagram of spore suspension, ClO2-treated spore suspension and ClO2 solution;

[0043] Figure 4b is the relationship between ClO2 dose and optical density (275nm);

[0044] Figure 4c FT-IR spectra of Aspergillus niger spores in the ClO2 treated and untreated groups;

[0045] Figure 5a Ag1 / CN under light conditions and 8.0 mg·L under dark conditions -1 ClO2 combined with Ag1 / CN and light conditions using 8.0 mg·L -1 Bar graph showing the inactivation rate of ClO2 combined with Ag1 / CN on Aspergillus niger spores;

[0046] Figure 5b The bar graph shows the inactivation rate of different concentrations of ClO2 on different concentrations of Aspergillus niger spores;

[0047] Figure 6 20.0 mg·L -1 ClO2 disinfection system and 8.0mg·L -1 Relationship between ClO2 residual amount and time after treatment of Aspergillus niger spores by ClO2 combined with Ag1 / CN photocatalytic disinfection system;

[0048] Figure 7 Schematic diagram of the process for inactivating Aspergillus niger spores in water. DETAILED DESCRIPTION

[0049] In the following description, many specific details are set forth to facilitate a full understanding of the embodiments according to the present invention. However, the embodiments according to the present invention may also be implemented in other ways different from those described herein. Therefore, the scope of protection according to the embodiments of the present invention is not limited to the specific embodiments disclosed below.

[0050] like Figure 7 As shown, this embodiment provides a method for inactivating Aspergillus niger spores in water, comprising the following steps:

[0051] S702: Obtaining a water body to be tested containing Aspergillus niger spores;

[0052] S704: Preparation of silver single atom-graphite carbon nitride composites by cryo-photoreduction method;

[0053] S706: Adding ClO2 to the water to be tested under light-free conditions;

[0054] S708: Under the condition of visible light irradiation, the silver single atom-graphite carbon nitride composite material is placed into the water body to be tested to inactivate the Aspergillus niger spores in the water body to be tested.

[0055] The present invention provides a method for inactivating Aspergillus niger spores in water. ClO2 destroys the melanin protective layer of the Aspergillus niger spores, exposing the spore cells. The silver single atom-graphite carbon nitride composite material then closely contacts the exposed spore cells through adsorption, acting as a highly efficient photocatalyst to inactivate the spores at the interface. Furthermore, because the low concentration of ClO2 is completely consumed by melanin, no chlorinated disinfectant remains in the water, and no carcinogenic byproducts such as chlorite are generated in the water.

[0056] The sterilization mechanism of the silver single atom-graphite carbon nitride composite material is that when the composite is exposed to visible light, g-C3N4 (graphite carbon nitride) absorbs photons and excites them to generate electron-hole pairs. The Ag single atom acts as a highly efficient co-catalyst, capturing these electrons and reducing recombination, thereby enhancing photocatalytic efficiency. Simultaneously, the Ag single atom injects the captured electrons into the cell membrane of the spores in close contact with it, disrupting the membrane structure and inhibiting their metabolic activity, ultimately achieving a highly effective inactivation effect.

[0057] In some embodiments, optionally, in the silver single atom-graphite carbon nitride composite material, the mass percentage of silver in the silver single atom-graphite carbon nitride composite material is greater than or equal to 1% and less than or equal to 5%.

[0058] In this embodiment, in the silver single atom-graphite phase carbon nitride composite material, the mass percentage of silver in the silver single atom-graphite phase carbon nitride composite material is greater than or equal to 1% and less than or equal to 5%, for example, it can be 1%, 3% or 5%. Among them, when the mass of silver is too high, it will aggregate to form larger nanoparticles instead of maintaining a single atomic dispersion state. This will reduce the number of effective active sites and reduce the photocatalytic efficiency. If the Ag single atom loading is too low, it is difficult to provide enough co-catalytic sites to capture photogenerated electrons, and the photocatalytic efficiency cannot be significantly improved.

[0059] In some embodiments, optionally, the wavelength of the visible light is greater than or equal to 400 nm and less than or equal to 760 nm.

[0060] In this embodiment, the wavelength of visible light is greater than or equal to 400 nm and less than or equal to 760 nm. If it is less than 400 nm, it belongs to the ultraviolet light range, or if it is greater than 760 nm, the photocatalytic efficiency will be significantly reduced.

[0061] In some embodiments, optionally, the illumination intensity of visible light is greater than or equal to 50 mW·cm -2 , and less than or equal to 200mW·cm -2 .

[0062] In this embodiment, the illumination intensity of visible light is greater than or equal to 50 mW·cm -2 , and less than or equal to 200mW·cm -2 For example, it can be 50mW·cm -2 、100mW·cm -2 or 200 mW·cm -2 The rate of photocatalytic reaction usually follows the saturation effect of light intensity. After exceeding a certain light intensity, the increase in reaction rate gradually becomes limited or even tends to be stable. Therefore, excessively high light intensity does not significantly increase the reaction rate, but instead causes the extra light energy to be wasted. Lower light intensity makes it difficult to fully excite the electrons in the photocatalyst, resulting in a limited number of electron-hole pairs generated, which in turn affects the efficiency of the entire photocatalytic sterilization process.

[0063] In some embodiments, optionally, before the step of adding ClO2 to the water body to be tested, the method further includes: calculating the actual concentration of Aspergillus niger spores in the water body to be tested; wherein the amount of ClO2 and silver single atom-graphite phase carbon nitride composite material added is related to the actual concentration of Aspergillus niger spores.

[0064] In this embodiment, since the actual concentration of Aspergillus niger spores in different water bodies is different, it is necessary to measure the actual concentration of Aspergillus niger spores in the water body to be tested in advance, and then determine the amount of ClO2 and silver single atom-graphitic carbon nitride composite material added according to the actual concentration of Aspergillus niger spores in the water body to be tested, so as to ensure the inactivation effect.

[0065] In some embodiments, optionally, the amount of ClO2 added = the volume of the water to be tested × the standard concentration of ClO2 × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, wherein the standard concentration of ClO2 is greater than or equal to 8.0 mg·L -1 , and less than or equal to 10.0 mg L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 .

[0066] In some embodiments, optionally, the amount of the silver single atom-graphite carbon nitride composite material added = the volume of the water to be tested × the standard concentration of the silver single atom-graphite carbon nitride composite material × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, wherein the standard concentration of the silver single atom-graphite carbon nitride composite material is equal to 200 mg·L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 .

[0067] In these examples, the inactivation effect was ensured by constructing the relationship between the added amount of ClO2 and the silver single atom-graphitic carbon nitride composite material and the actual concentration of Aspergillus niger spores.

[0068] In some embodiments, optionally, the step of preparing a silver single atom-graphite phase carbon nitride composite material by a cryophotoreduction method specifically includes: adding a graphite phase carbon nitride sheet and a silver nitrate solution to deionized water to obtain a solid-liquid mixture, and then quickly freezing the solid-liquid mixture in liquid nitrogen to form an ice crystalline solid, irradiating the ice crystalline solid with an ultraviolet lamp, and maintaining the ice crystalline solid at a first preset temperature to obtain a silver single atom-graphite phase carbon nitride composite material.

[0069] In some embodiments, optionally, the first preset temperature is greater than or equal to -5°C and less than or equal to 0°C. For example, it can be -5°C, -2°C or 0°C.

[0070] In some embodiments, optionally, the wavelength of the ultraviolet light emitted by the ultraviolet lamp is greater than or equal to 360 nm and less than or equal to 370 nm; for example, it can be 360 ​​nm, 365 nm or 370 nm.

[0071] In some embodiments, the UV light emitted by the UV lamp has an intensity greater than or equal to 50 mW·cm -2 , and less than or equal to 60mW·cm -2 ; For example, it can be 50mW·cm -2 、52.8mW·cm -2 or 60 mW·cm -2 .

[0072] In this embodiment, in the process of preparing silver single atom-graphite phase carbon nitride composite material, 10g of urea was loaded into a crucible and placed in a muffle furnace, and heated in an air atmosphere at 550°C for 4h with a heating rate of 2°C / min to obtain bulk g-C3N4. 0.4g of bulk g-C3N4 was ground into powder, evenly spread on aluminum foil, and heated again in an air atmosphere at 500°C for 2h with a heating rate of 5°C / min to obtain exfoliated g-C3N4 nanosheets. 20mg of g-C3N4 nanosheets and 24.3μL of silver nitrate solution (concentration of 67.9mg·mL) were mixed with 1% urea and 2% urea. -1 ) was added to 10 mL of deionized water and stirred for 12 h to allow g-C3N4 to fully adsorb Ag ions. The mixture was quickly frozen in liquid nitrogen to form ice crystals and irradiated under a UV lamp for 10 min. The UV wavelength was 365 nm and the light intensity on the surface of the reaction system was 52.8 mW·cm -2 , the ambient temperature is kept below 0°C to keep the ice crystals from melting, thereby preventing the nucleation of Ag and ultimately forming a silver single atom-graphite phase carbon nitride composite material.

[0073] In some technical solutions, optionally, the volume of the water body to be tested is 50 ml, and the concentration of Aspergillus niger spores in the water body to be tested is 10 6 cells·mL -1 The method for inactivating Aspergillus niger spores in water also includes: transferring the inactivated water to be tested into a centrifuge tube and centrifuging to obtain a spore precipitate; placing the spore precipitate in sterile physiological saline to form a spore suspension; adding a fluorescent dye to the spore suspension to stain the Aspergillus niger spores in the spore suspension, wherein the fluorescent dye includes a first fluorescent dye for staining living spores and a second fluorescent dye for staining dead spores; introducing the dyed spore suspension into a flow cytometer, and determining the inactivation rate of the Aspergillus niger spores by the flow cytometer.

[0074] In this embodiment, the present invention can not only inactivate Aspergillus niger spores in water, but also detect the inactivation rate. Since the low concentration of ClO2 is completely consumed by melanin, no chlorinated disinfectant will remain in the water, and no carcinogenic byproducts such as chlorite will be generated in the water, thereby improving safety.

[0075] In some technical solutions, optionally, the excitation light wavelength of the first fluorescent dye is greater than or equal to 480 nm and less than or equal to 500 nm, for example, it can be 488 nm, the emission light wavelength of the first fluorescent dye is greater than or equal to 500 nm and less than or equal to 530 nm, for example, it can be 530 nm, the excitation light wavelength of the second fluorescent dye is greater than or equal to 488 nm and less than or equal to 546 nm, for example, it can be 488 nm, and the emission light wavelength of the second fluorescent dye is greater than or equal to 600 nm and less than or equal to 650 nm, for example, it can be 630 nm.

[0076] In some technical solutions, the first fluorescent dye optionally includes one or a combination of the following: SYTO9 dye, SYBR Green I dye, and DAPI dye. Specifically, the fluorescent dye for staining viable spores can be SYTO9 dye, a dibenzimidazole dye with an excitation wavelength of approximately 480 nm, an emission wavelength of approximately 500 nm, and can emit green fluorescence. Alternatively, the first fluorescent dye can be SYBR Green I dye, a cyanine dye with two ring structures, an excitation wavelength of approximately 497 nm, an emission wavelength of approximately 520 nm, and can emit green fluorescence.

[0077] In some technical solutions, the second fluorescent dye optionally includes one or a combination of the following: propidium iodide, 7-AAD, and ethidium bromide. Specifically, the fluorescent dye used to stain dead spores can be propidium iodide, which has an excitation wavelength of approximately 490 nm to 535 nm and an emission wavelength of approximately 617 nm (red fluorescence). Alternatively, it can be 7-AAD, which has an excitation wavelength of approximately 546 nm and an emission wavelength of approximately 647 nm (red fluorescence). Alternatively, it can be ethidium bromide, which has an excitation wavelength of approximately 500 nm to 540 nm and an emission wavelength of approximately 605 nm (orange-red fluorescence).

[0078] It should be noted that, in the actual inactivation rate test process, the existing water body can be directly used, and of course a spore suspension can also be prepared. The steps of preparing the spore suspension specifically include: applying the Aspergillus niger powder suspension on a potato dextrose agar plate and placing it for a preset time to activate and form an Aspergillus niger spore group; washing the Aspergillus niger spore group on the potato dextrose agar plate with sterile saline and resuspending it to obtain a spore suspension; filtering the spore suspension with sterile gauze to obtain a first spore suspension.

[0079] In this embodiment, when preparing the first spore suspension, 200 μL of Aspergillus niger powder suspension can be evenly spread on a potato dextrose agar plate and cultured at 25°C for 5 days to activate the strain; the Aspergillus niger spores on the plate are rinsed and resuspended with 10.0 mL of sterile saline (0.85% NaCl), and the resulting spore suspension is filtered twice with three layers of sterile gauze to remove mycelia, and then centrifuged and washed three times at a speed of 8000 rpm to obtain a spore suspension with higher purity. The high-concentration spore suspension is diluted to a standard concentration of 10 using the counting function of a flow cytometer. 6 cells·mL -1 .

[0080] Another embodiment of the present invention provides a method for efficiently inactivating Aspergillus niger spores in water using low-concentration ClO2 in conjunction with Ag single-atom-loaded graphite-phase carbon nitride (Ag1 / CN) photocatalyst. The method is efficient and rapid, simple to operate, has no disinfectant residue and by-product generation, and is low in cost.

[0081] In order to achieve the above purpose, the technical solution adopted by the present invention is: using 8.0 mg·L -1 The ClO2 destroyed the Aspergillus niger spore suspension in about 10 6 cells·mL -1 The melanin protective layer of Aspergillus niger spores exposes the spore cells. Ag1 / CN can closely contact the exposed spore cells through adsorption, thereby achieving efficient photocatalytic interfacial inactivation of the spores. The scanning electron microscope image of Aspergillus niger spores before treatment is shown in Figure 2. Figure 1a As shown in the scanning electron microscope image of Aspergillus niger spores after ClO2+Ag1 / CN treatment. Figure 1b shown.

[0082] Alternatively, the preparation method of the Aspergillus niger spore suspension is as follows: 200 μL of Aspergillus niger powder suspension is evenly spread on a potato dextrose agar plate and cultured at 25° C. for 5 days to activate the strain; the Aspergillus niger spores on the plate are rinsed with 10.0 mL of sterile saline (0.85% NaCl) and resuspended, and the obtained spore suspension is filtered twice with three layers of sterile gauze to remove mycelia, and then centrifuged and washed three times at a speed of 8000 rpm to obtain a spore suspension with higher purity, and the high-concentration spore suspension is diluted to a standard concentration of 10 using the counting function of a flow cytometer. 6 cells·mL -1 .

[0083] Alternatively, 8.0 mg·L -1 The ClO2 destroyed the melanin for 15 minutes, and the suspension changed from black to light brown.

[0084] Alternatively, the preparation method of Ag1 / CN is as follows: Ag single atoms are loaded onto g-C3N4 nanosheets by freeze-UV reduction. Specifically, 20 mg of g-C3N4 and 24.3 μL of silver nitrate solution (concentration of 67.9 mg mL -1 ) was added to 10 mL of deionized water and stirred for 12 h to allow g-C3N4 to fully adsorb Ag ions. The mixed solution was then quickly frozen in liquid nitrogen to form ice crystals and irradiated under ultraviolet light for 10 min. The ultraviolet wavelength was 365 nm, and the light intensity on the surface of the reaction system was measured using an ultraviolet power meter and was 52.8 mW·cm -2 The ambient temperature was kept below 0°C to prevent the ice crystals from melting, thereby preventing the nucleation of Ag. The obtained Ag1 / CN catalyst was centrifugally washed three times with deionized water at a centrifugal speed of 12000 rpm, and then dried in a vacuum oven at 60°C for 24 h for use.

[0085] Optionally, the loading amount of Ag in Ag1 / CN is 2.81 wt %.

[0086] Optionally, the optimal killing concentration of Ag1 / CN is 200 mg·L -1 , the optimal killing time is 1h.

[0087] In addition, the present invention also provides a method for detecting the efficiency of inactivating Aspergillus niger spores, which is as follows:

[0088] Collected spore samples were stained with LIVE / DEAD dye (LIVE stands for live, DEAD stands for dead) and analyzed by flow cytometry. This kit contains two fluorescent nucleic acid dyes, SYTO9 and propidium iodide (PI). Live spores, whose membrane permeability is intact, are stained only with green fluorescence by SYTO9 (excitation wavelength: 488 nm, emission wavelength: 530 nm), while dead spores, which have lost membrane permeability, are stained with red fluorescence by PI (excitation wavelength: 488 nm, emission wavelength: 630 nm). To prepare the staining stock solution, mix 3µL of SYTO9 and 3µL of PI in 1.0mL of water. 100µL of sample was thoroughly mixed with an equal volume of the staining stock solution, incubated at room temperature for 15 minutes, and then injected into the flow cytometer for inactivation analysis. The flow cytometer was set up as follows: the sample acquisition speed was adjusted to the medium speed mode (50 μL / min), and the FSC-H (Forward Scatter Height) threshold was set to 5 × 10 4The sampling stop condition was set to 20,000 events, and the fluorescence detector was selected in FITC (Fluorescein isothiocyanate) mode and PE (Phycoerythrin) mode. FSC-H is the forward scattered light signal, which represents the size of the sample particles, and SSC-H (Side Scatter Height) is the side scattered light height, which represents the complexity of the sample particles. In the sample dot matrix, the SSC-H axis is located at 10 5 and FSC-H axis 10 6 The clusters at the junction were identified as Aspergillus niger spores, as shown in Figure 2a As shown in Figure 2, the number of these spore clusters (spore cells) in the sample is counted by the volume counting hardware of the flow cytometer. In the dual-parameter dot matrix of PI and SYTO9, two clusters can be distinguished by the defined gates, namely live spores and dead spores, as shown in Figure 2. Figure 2b shown.

[0089] In the present invention, the g-C3N4 nanosheets used can be synthesized by themselves, ClO2 is purchased from commercial tablets, and other reaction devices include glass reactors, photocatalytic devices, etc. All components used can be obtained from relevant equipment suppliers or designed and built by themselves.

[0090] Compared with the prior art, the present invention has the following beneficial effects:

[0091] 1. Significantly reduced the amount of chlorinated disinfectant used to inactivate Aspergillus niger spores;

[0092] 2. The low concentration of ClO2 used reacts completely with the melanin of Aspergillus niger spores and no residue remains in the water;

[0093] 3. No carcinogenic byproducts such as chlorite are generated in the system;

[0094] 4. After low-concentration ClO2 treatment, Aspergillus niger spores are exposed to the outside and can fully contact with the Ag1 / CN photocatalyst, and the inactivation efficiency can reach nearly 100% in 1 hour;

[0095] 5. It is safe and environmentally friendly, low cost, simple to operate, and can effectively inactivate Aspergillus niger spores, and has broad application prospects.

[0096] The applicant will now describe the method for inactivating Aspergillus niger spores and the method for detecting the inactivation rate of the present invention from multiple perspectives, as follows:

[0097] Example 1, preparation of Ag single-atom photocatalyst.

[0098] (1) 10 g of urea was loaded into a crucible and placed in a muffle furnace. It was heated at 550 °C in an air atmosphere for 4 h at a heating rate of 2 °C / min to obtain bulk g-C3N4.

[0099] (2) Grind 0.4 g of bulk g-C3N4 obtained in step (1) into powder, spread it evenly on aluminum foil, and heat it again at 500°C in air atmosphere for 2 h at a heating rate of 5°C / min to obtain exfoliated g-C3N4 nanosheets.

[0100] (3) 20 mg of g-C3N4 nanosheets obtained in step (2) and 24.3 μL of silver nitrate solution (67.9 mg mL -1 ) was added into 10 mL of deionized water and stirred for 12 h to allow g-C3N4 to fully adsorb Ag ions.

[0101] (4) The mixture solution obtained in step (3) was quickly frozen in liquid nitrogen to form ice crystals, and then irradiated under ultraviolet light for 10 minutes. The ultraviolet wavelength was 365 nm. The light intensity on the surface of the reaction system was measured using an ultraviolet power meter and was 52.8 mW·cm -2 , the ambient temperature is kept below 0°C to keep the ice crystals from melting, thereby preventing the nucleation of Ag and obtaining Ag1 / CN.

[0102] (5) The Ag1 / CN obtained in step (4) was washed three times with deionized water by centrifugation at a speed of 12000 rpm, and then dried in a vacuum oven at 60°C for 24 h.

[0103] The Ag1 / CN prepared in Example 1 was tested by transmission electron microscopy and spherical aberration electron microscopy. The transmission electron microscopy images are as follows: Figure 3a and Figure 3b As shown in the AC-HAADF-STEM (Annular Contrast-High Angle Annular Dark Field-Scanning Transmission Electron Microscopy) image (an electron microscope image with atomic-level resolution) Figure 3c As shown, Figure 3c The bright spot in the middle circle is a single Ag atom, indicating that the prepared Ag1 / CN has a nanosheet structure and the single Ag atom is successfully loaded.

[0104] Example 2: Method for inactivating Aspergillus niger spores in water using low concentration ClO2 in conjunction with Ag1 / CN photocatalyst.

[0105] (1) Preparation of Aspergillus niger spore suspension. Aspergillus niger powder was purchased from a biological company. Specifically, freeze-dried Aspergillus niger powder was mixed with 0.5 mL of sterile saline. 200 μL of powder suspension was evenly spread on a potato dextrose agar plate and cultured at 25°C for 5 days to activate the strain. The Aspergillus niger spores on the plate were rinsed and resuspended with 10.0 mL of sterile saline (0.85% NaCl). The obtained spore suspension was filtered twice with three layers of sterile gauze to remove the mycelium, and then centrifuged and washed three times at 8000 rpm to obtain a spore suspension with higher purity. The high-concentration spore suspension was diluted to 10 using the counting function of the flow cytometer. 6 cells·mL -1 concentration for subsequent inactivation experiments.

[0106] (2) In the first step of the ClO2 combined with Ag1 / CN disinfection experiment, 8.0 mg·L -1 The ClO2 pretreatment concentration of 50mL is 10 6 cells·mL -1 The spore suspension was incubated for 15 min in the dark.

[0107] (3) In the second step of the ClO2 combined with Ag1 / CN disinfection experiment, the pretreated spore suspension was used for photocatalytic disinfection. 6 cells·mL -1 Ag1 / CN (0, 50 mg·L) was added to the quartz tube containing the spore suspension. -1 , 100mg·L -1 and 200 mg·L -1 The photocatalytic reaction was carried out in a multi-channel photocatalytic reaction device. The visible light spectrum range was 400nm-760nm. The light intensity on the surface of the spore suspension was 100mW·cm -2 , measured by an optical power meter. The stirring speed and temperature of the photocatalytic reaction device were set to 250 rpm and 25°C, respectively. 3.0 mL samples were collected at the corresponding sampling time to evaluate the inactivation rate of spores. Each experiment was repeated three times.

[0108] like Figure 4a As shown in the figure, the color of the spores is black when they are not treated, and the color of ClO2 is transparent. After the spore suspension is pretreated with ClO2, the color changes from black to brown. The absorbance of the spore suspension is tested, and the results are as follows: Figure 4b As shown, 8.0 mg·L -1 After ClO2 treatment, 10 6 cells·mL -1 OD of spores 275nm(OD is Optical Density, optical density, OD 275nm That is, the optical density measured at a wavelength of 275 nanometers) reaches its minimum. The spore suspension pretreated with ClO2 was tested by Fourier transform infrared spectroscopy. The FT-IR (Fourier Transform Infrared Spectroscopy) spectrum of Aspergillus niger spores is shown in the figure below. Figure 4c As shown, 1641cm -1 、1744cm -1 and 2925cm -1 The peak intensity at 37° is significantly reduced, which corresponds to the stretching vibration of aromatic C=C or C=O groups in melanin, the C=O stretching of ester or carboxylic acid, and the asymmetric stretching of CH2 and CH3, proving that ClO2 can remove the melanin of spores.

[0109] Example 3. The efficiency of low concentration ClO2 synergistically inactivating Aspergillus niger spores in water using Ag1 / CN photocatalyst.

[0110] (1) Stain the collected spore samples with LIVE / DEAD dye. Mix 3 µL of SYTO9 and 3 µL of PI in 1.0 mL of water to prepare a staining solution. Mix 100 µL of sample with an equal volume of the staining solution and incubate at room temperature for 15 min.

[0111] (2) The stained samples in step (1) were analyzed by flow cytometry. The flow cytometer settings were as follows: the sample acquisition speed was adjusted to the medium speed mode (50 μL / min), and the FSC-H threshold was set to 5´10 4 The sampling stop condition was set to 20,000 events, and the fluorescence detector was selected in FITC and PE mode. In the dual-parameter dot matrix of PI and SYTO9, two clusters can be distinguished by the defined gates: live spores (Live) and dead spores (Dead).

[0112] (3) The inactivation rate of spores is calculated according to Formula 1:

[0113] ;

[0114] Among them, Dead is the number of dead spores, and Live is the number of living spores.

[0115] like Figure 5a As shown, 50.0 mg·L -1 and 100.0 mg·L -1 The inactivation rates of Ag1 / CN on spores reached 36.0% and 42.8% within 60 min, respectively, while 200.0 mg·L -1Ag1 / CN had the best sterilization activity, reaching a nearly 100% inactivation rate in 60 min.

[0116] Example 4, ClO2 residual and chlorite detection.

[0117] (1) ClO2 residues were detected by cresol red spectrophotometry. At pH = 3, ClO2 and cresol red undergo an oxidation-reduction reaction, and the remaining cresol red turns purple under alkaline conditions. The minimum detection concentration of ClO2 is 0.02 mg·L-1 under a colorimetric quantification wavelength of 573 nm. -1 .

[0118] (2) Prepare ClO2 standard 0, 0.1 mg·L -1 , 0.2 mg·L -1 , 0.5mg·L -1 , 1.0mg·L -1 and 2.0 mg·L -1 , adjust the pH to 3 with hydrochloric acid.

[0119] (3) Take 5 mL of sample and adjust the pH to 3 with hydrochloric acid.

[0120] (4) Dissolve 0.1 g of cresol red in 20 mL of ethanol, then dilute to 100 mL with deionized water. Take 1 mL of the above solution and dilute to 50 mL with deionized water for later use.

[0121] (5) Take 0.1 mL of the cresol red solution prepared in step (4) and add it to 5 mL of the standard and sample, and let it stand at room temperature for 10 minutes.

[0122] (6) Add 1.0 mL of 8.0 g·L -1 Add the sodium hydroxide solution to the solution in step (5) and shake well.

[0123] (7) Detect the light absorbance of the solution in step (6) at a wavelength of 573 nm and calculate the concentration of ClO2 based on the standard curve.

[0124] (8) Chlorite was detected by ion chromatography.

[0125] Specifically, the concentrations of sodium chlorite standards were 0 and 0.05 mg·L -1 , 0.1mg·L -1 , 0.2 mg·L -1 , 0.5mg·L -1 and 1.0 mg·L -1. Take 10 mL of the sample in a 100 mL volumetric flask, dilute to the mark with ultrapure water, and sonicate for 30 minutes. Take an appropriate amount of sample and centrifuge at 12,000 rpm for 10 minutes. Take the supernatant, add acetonitrile at a ratio of 1:1, vortex for 1 minute, and centrifuge at 12,000 rpm for 10 minutes. Take 10 mL of the supernatant and pass it through a 0.45 μm nylon membrane syringe filter and an OnGuard Ⅱ RP column. Discard the first 3 mL and collect the subsequent filtrate for testing. Use an ion chromatography system to detect chlorite in samples and standards.

[0126] like Figure 6 As shown in the figure, after 15 min of pretreatment in the first stage, the ClO2 residual was below the detection limit (<0.1 mg·L -1 ), so there is no risk of disinfectant residue in the system. In addition, during the second stage of photocatalytic treatment, the harmful byproduct chlorite produced by ClO2 under the action of light was not detected.

[0127] Comparative Example 1, efficiency of Ag1 / CN photocatalyst alone in inactivating Aspergillus niger spores in water.

[0128] (1) The preparation of Ag1 / CN photocatalyst and Aspergillus niger spore suspension was the same as that in Example 1 and Example 2.

[0129] (2) In a 50 mL container containing 10 6 cells·mL -1 Ag1 / CN (0, 50 mg·L) was added to the quartz tube containing the spore suspension. -1 , 100mg·L -1 and 200 mg·L -1 The photocatalytic reaction was carried out in a multi-channel photocatalytic reaction device. The visible light spectrum range was 400nm-760nm. The light intensity on the surface of the spore suspension was 100mW·cm -2 , measured by an optical power meter. The stirring speed and temperature of the photocatalytic reaction apparatus were set at 250 rpm and 25°C, respectively. 3.0 mL samples were collected at the corresponding sampling times to evaluate the spore inactivation rate. Each experiment was repeated three times.

[0130] (3) The sample in step (2) was tested using the method for testing the inactivation rate of Aspergillus niger spores in Example 3.

[0131] like Figure 5a As shown in the results, without ClO2 pretreatment, Ag1 / CN alone had almost no inactivation effect on spores under light irradiation.

[0132] Comparative Example 2, efficiency of ClO2 combined with Ag1 / CN photocatalyst in inactivating Aspergillus niger spores in water under dark conditions.

[0133] (1) The preparation of Ag1 / CN photocatalyst and Aspergillus niger spore suspension was the same as that in Example 1 and Example 2.

[0134] (2) In the first step of the ClO2 combined with Ag1 / CN disinfection experiment under dark conditions, 8.0 mg·L -1 50 mL of ClO2 pre-treated 10 6 cells·mL -1 The spore suspension was incubated for 15 min and the reaction was carried out in the dark.

[0135] (3) In the second step, 50 mL of 10 6 cells·mL -1 Ag1 / CN (0 mg·L -1 , 50mg·L -1 , 100mg·L -1 and 200 mg·L -1 The reaction was carried out in the dark for 60 min. 3.0 mL samples were collected at the corresponding sampling times to assess spore inactivation. Each experiment was repeated three times.

[0136] (4) The sample in step (3) was tested using the method for testing the inactivation rate of Aspergillus niger spores in Example 3.

[0137] like Figure 5a As shown in the figure, under dark conditions, ClO2 combined with Ag1 / CN only inactivated about 10% of the spores, which was mainly attributed to the effect of ClO2.

[0138] Comparative Example 3, efficiency of ClO2 inactivating Aspergillus niger spores in water and detection of ClO2 residues.

[0139] (1) The preparation of Aspergillus niger spore suspension was the same as in Example 2.

[0140] (2) Use 0.5 mg·L -1 -20mg·L -1 Different concentrations of ClO2 were used to treat 50.0 mL of spore suspension, where the spore concentrations were 10 4 cells·mL -1 , 10 5 cells·mL -1 and 10 6 cells·mL -1 Note that the reactions were performed in the dark because ClO2 decomposes in the presence of light. 3.0 mL samples were collected at the corresponding sampling times to determine the spore inactivation rate. Each experiment was repeated three times.

[0141] (3) The sample in step (2) was tested using the method for testing the inactivation rate of Aspergillus niger spores in Example 3.

[0142] (4) The method in Example 4 was used to detect the ClO2 residue in the sample of step (2).

[0143] like Figure 5b As shown, 10 6 cells·mL -1 Spores in extremely high concentrations of ClO2 (20.0 mg·L -1 ) to achieve 100% inactivation rate. -1 ClO2 disinfection system and 8.0mg·L -1 ClO2 combined with Ag1 / CN disinfection system can effectively inactivate Aspergillus niger spores, but 20.0 mg·L -1 The residual ClO2 in the ClO2 disinfection system is approximately 2.7 mg·L -1 , specifically Figure 6 As shown, it far exceeds the drinking water disinfectant standard (<0.8mg·L -1 ,GB5749-2022).

[0144] In the embodiments according to the present invention, the terms "first", "second", and "third" are used only for descriptive purposes and are not to be understood as indicating or implying relative importance; the term "plurality" refers to two or more, unless otherwise expressly defined. Terms such as "installed", "connected", "connected", and "fixed" should be understood in a broad sense. For example, "connected" can be a fixed connection, a detachable connection, or an integral connection; "connected" can be a direct connection or an indirect connection through an intermediate medium. For those skilled in the art, the specific meanings of the above terms in the embodiments according to the present invention can be understood according to specific circumstances.

[0145] In addition, although adopting specific order to describe each operation, this should be understood as requiring such operation to be carried out in the specific order or in sequential order, or requiring that all illustrated operations should be carried out to obtain desired result.Under certain environment, multitasking and parallel processing may be advantageous.Similarly, although comprising some specific implementation details in the above discussion, these should not be construed as limiting the scope of the present invention.Some features described in the context of independent embodiment can also be implemented in a single implementation in combination.On the contrary, the various features described in the context of independent implementation also can be implemented in a plurality of implementations individually or in the mode of any suitable subcombination.

[0146] Although the subject matter has been described in the language of specific structural features and / or methodological logical actions, it should be understood that the subject matter defined in the appended claims is not necessarily limited to the specific features or actions described above. On the contrary, the specific features and actions described above are merely example forms of implementing the claims.

[0147] The above are merely examples of embodiments according to the present invention and are not intended to limit the embodiments according to the present invention. It will be apparent to those skilled in the art that various modifications and variations may be made to the embodiments according to the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the embodiments according to the present invention shall be included within the scope of protection of the embodiments according to the present invention.

Claims

1. A method for inactivating Aspergillus niger spores in water, characterized in that: include: Obtaining a water body to be tested containing Aspergillus niger spores; Silver single atom-graphite carbon nitride composites were prepared by cryo-photoreduction method; Under light-free conditions, adding ClO2 to the water to be tested; placing the silver single atom-graphite carbon nitride composite material into the water to be tested under visible light irradiation conditions to inactivate Aspergillus niger spores in the water to be tested; The step of preparing the silver single atom-graphite phase carbon nitride composite material by the cryo-photoreduction method specifically includes: Graphite phase carbon nitride sheets and silver nitrate solution are added to deionized water to obtain a solid-liquid mixture, and then the solid-liquid mixture is quickly frozen in liquid nitrogen to form an ice crystalline solid. The ice crystalline solid is irradiated by an ultraviolet lamp and maintained at a first preset temperature to obtain a silver single atom-graphite phase carbon nitride composite material.

2. The method for inactivating Aspergillus niger spores in water according to claim 1, characterized in that: In the silver single atom-graphite carbon nitride composite material, the mass percentage of silver in the silver single atom-graphite carbon nitride composite material is greater than or equal to 1% and less than or equal to 5%; and / or The wavelength of the visible light is greater than or equal to 400nm and less than or equal to 760nm, and the illumination intensity of the visible light is greater than or equal to 50mW·cm -2 , and less than or equal to 200mW·cm -2 .

3. The method for inactivating Aspergillus niger spores in water according to claim 1, characterized in that: Before the step of adding ClO2 to the water to be tested, the method further comprises: Calculating the actual concentration of Aspergillus niger spores in the water to be tested; The added amounts of ClO2 and the silver single atom-graphite phase carbon nitride composite material are related to the actual concentration of the Aspergillus niger spores.

4. The method for inactivating Aspergillus niger spores in water according to claim 3, characterized in that: The amount of ClO2 added = the volume of the water to be tested × the standard concentration of ClO2 × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, where the standard concentration of ClO2 is greater than or equal to 8.0 mg·L -1 , and less than or equal to 10.0 mg·L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 and / or The amount of silver single atom-graphite carbon nitride composite material added = the volume of the water to be tested × the standard concentration of the silver single atom-graphite carbon nitride composite material × the actual concentration of Aspergillus niger spores / the standard concentration of Aspergillus niger spores, where the standard concentration of the silver single atom-graphite carbon nitride composite material is equal to 200 mg·L -1 The standard concentration of Aspergillus niger spores is 10 6 cells·mL -1 .

5. The method for inactivating Aspergillus niger spores in water according to claim 1, characterized in that: The first preset temperature is greater than or equal to -5°C and less than or equal to 0°C; and / or The wavelength of the ultraviolet light emitted by the ultraviolet lamp is greater than or equal to 360nm and less than or equal to 370nm; and / or The intensity of the ultraviolet light emitted by the ultraviolet lamp is greater than or equal to 50mW·cm -2 , and less than or equal to 60mW·cm -2 .

6. The method for inactivating Aspergillus niger spores in water according to claim 1, characterized in that: The volume of the water to be tested is 50 mL, and the concentration of Aspergillus niger spores in the water to be tested is 10 6 cells·mL -1 The method for inactivating Aspergillus niger spores in the water body further comprises: Transferring the inactivated water to be tested into a centrifuge tube and centrifuging it to obtain a spore precipitate; placing the spore pellet into sterile physiological saline to form a spore suspension; adding a fluorescent dye to the spore suspension to stain the Aspergillus niger spores in the spore suspension, wherein the fluorescent dye comprises a first fluorescent dye for staining living spores and a second fluorescent dye for staining dead spores; The stained spore suspension is introduced into a flow cytometer, and the inactivation rate of the Aspergillus niger spores is determined by the flow cytometer.

7. The method for inactivating Aspergillus niger spores in water according to claim 6, characterized in that: The excitation light wavelength of the first fluorescent dye is greater than or equal to 480 nm and less than or equal to 500 nm, the emission light wavelength of the first fluorescent dye is greater than or equal to 500 nm and less than or equal to 530 nm, the excitation light wavelength of the second fluorescent dye is greater than or equal to 488 nm and less than or equal to 546 nm, and the emission light wavelength of the second fluorescent dye is greater than or equal to 600 nm and less than or equal to 650 nm.

8. The method for inactivating Aspergillus niger spores in water according to claim 7, characterized in that: The excitation light wavelength of the first fluorescent dye is equal to 488 nm, the emission light wavelength of the first fluorescent dye is equal to 530 nm, the excitation light wavelength of the second fluorescent dye is equal to 488 nm, and the emission light wavelength of the second fluorescent dye is equal to 630 nm.

9. The method for inactivating Aspergillus niger spores in water according to claim 6, wherein: The first fluorescent dye comprises one or a combination of the following: SYTO9 dye, SYBR Green I dye and DAPI dye; and / or The second fluorescent dye includes one or a combination of the following: propidium iodide dye, 7-AAD dye and ethidium bromide dye.

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