CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system, preparation method and application

Abstract

The application discloses a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system and a preparation method and application thereof. After sterilization of CdS / C3N4 photocatalytic material, the CdS / C3N4 photocatalytic material is mixed with a denitrifying culture medium to obtain a mixture of the CdS / C3N4 photocatalytic material and the denitrifying culture medium. After centrifugation of LB culture medium containing denitrifying bacteria, wet bacteria bodies are obtained, and the wet bacteria bodies are added into the mixture of the CdS / C3N4 photocatalytic material and the denitrifying culture medium. The coupling system can promote the growth and cell activity of Micrococcus candidus, and the photocatalytic effect of the material reduces the degree of light damage of the bacteria. In addition, the electrons of the photocatalytic material can provide additional electron donors for the bacteria after being excited under light, promote the reactions involving electron transfer in the bacterial metabolism, and are suitable for treatment of wastewater containing nitrate ions.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of sewage treatment, and relates to a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system, a preparation method and application. BACKGROUND

[0002] With the rapid growth of industrial activities and population, the over-consumption of fertilizers and other nitrogen compounds has led to serious environmental problems of nitrogen pollution, such as eutrophication and climate change. The discharge of nitrogen-rich wastewater leads to serious eutrophication and deoxygenation of the receiving water body. NO3 - As a pollutant, N is widely present in surface water and groundwater. If not completely treated, it will lead to a large amount of nitrogen source entering the environment, aggravate the situation of nitrogen source pollution, and even cause harm to human health, such as methemoglobinemia and colon cancer.

[0003] Among the current methods for removing nitrogen pollution, microbial denitrification is one of the important methods for removing nitrogen from water or sewage due to its low operating cost and easy operation and maintenance. The main pathway of biological denitrification is the reduction of nitrate (NO3 - -N) by a number of denitrifying enzymes in microorganisms, so it is widely used in wastewater treatment plants to remove NO3 - -N and (NH4 + -N). However, in actual use, a series of problems need to be solved. Among them, the problem of photosensitivity of microbial denitrifying bacteria, that is, under light conditions, the bacteria have a long start-up time and a reduced denitrification rate.

[0004] Solar energy is the most important clean energy. In recent years, researchers have proposed the idea of using solar energy to drive microbial cell factories. The basic principle is to combine semiconductor materials with microorganisms. The semiconductor material generates photoelectrons under sunlight, and the photoelectrons and photoholes can effectively accelerate the electron transfer between microorganisms and semiconductors. In most cases, the degree of electron transfer and utilization on the semiconductor-microbial interface determines the performance of the pollutant degradation or synthesis process.

[0005] The method of combining semiconductor materials with microorganisms has the technical problem of low denitrification efficiency.

[0006] Therefore, it is of great significance to study a semiconductor and denitrifying bacteria coupling system to solve the photosensitivity of denitrifying bacteria and improve the efficiency of microbial denitrification in the field of pollutant degradation. SUMMARY

[0007] In order to solve the problem of low denitrification efficiency in the prior art, the present application aims to provide a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system, a preparation method and application thereof, and the coupling system prepared by the method has high denitrification efficiency.

[0008] In order to achieve the above-mentioned purpose, the technical scheme adopted by the present application is as follows:

[0009] A preparation method of a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system, comprising the following steps:

[0010] After sterilizing the CdS / C3N4 photocatalytic material, the CdS / C3N4 photocatalytic material is mixed with the denitrifying culture medium according to a mass ratio of 1:(1000-5000) to obtain a mixture of the CdS / C3N4 photocatalytic material and the denitrifying culture medium.

[0011] After centrifuging the LB culture medium containing denitrifying bacteria, wet bacteria bodies are obtained, and the wet bacteria bodies are washed and added to the mixture of the CdS / C3N4 photocatalytic material and the denitrifying culture medium to obtain the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system.

[0012] Further, the CdS / C3N4 photocatalytic material is prepared by heating urea at 500 DEG C for 3 hours to obtain C3N4 material, suspending the C3N4 material in 50 mL ultrapure water, adding Cd(NO3)2.4H2O, ultrasonicating to obtain a suspension, adding Na2S.9H2O to the suspension to obtain a precipitate, and drying to obtain the CdS / C3N4 photocatalytic material.

[0013] Further, the denitrifying culture medium is prepared by the following process: adding KNO3, NH4Cl, Na2HPO4.12H2O, NaH2PO4, CH3COONa and a trace element aqueous solution into sterile water to obtain a mixture, sterilizing the mixture, adding MgSO4.7H2O, adjusting the pH to 7-8, then passing in argon and sealing to obtain the denitrifying culture medium.

[0014] Further, the amount ratio of KNO3, NH4Cl, Na2HPO4.12H2O, NaH2PO4, CH3COONa, the trace element aqueous solution, sterile water and MgSO4.7H2O is 3.14g:0.38g:11.74g:1.46g:3.54g:0.10mL:1L:0.06g.

[0015] Further, the trace element aqueous solution is prepared by the following process: adding Na2-EDTA, FeCl3.6H2O, MnCl2.4H2O, Na2MoO4.2H2O, CuCl2.2H2O and zinc chloride into sterile water.

[0016] Further, the trace element aqueous solution contains 9.6 mmol / L Na2-EDTA, 9.0 mmol / L FeCl3·6H2O, 0.1 mmol / L MnCl2·4H2O, 1.0 mmol / L Na2MoO4·2H2O, 0.8 mmol / L CuCl2·2H2O and 2.5 mmol / L zinc chloride.

[0017] Further, the LB medium containing denitrifying bacteria is prepared by the following process: culture and amplification of denitrifying microorganisms: inoculating the strain Paracoccus denitrificans of denitrifying microorganisms into the LB medium for large-scale culture to obtain the LB medium containing denitrifying bacteria, and the formula of the LB medium is: dissolving 10 g of tryptone, 10 g of NaCl and 5 g of yeast extract in 1 L of distilled water.

[0018] Further, the ratio of the mixture of the LB medium containing denitrifying bacteria and the CdS / C3N4 photocatalytic material and the denitrifying medium is 1 mL:(200-500) mL.

[0019] A CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system.

[0020] Application of a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system in treating wastewater containing nitrate ions.

[0021] Compared with the prior art, the present application has the beneficial effects of:

[0022] (1) Accelerating the denitrification rate: In the present application, the photoelectron conversion of CdS / C3N4 photocatalytic material provides additional energy for Paracoccus denitrificans, enabling it to denitrify more quickly.

[0023] (2) Environmentally friendly: The use of CdS / C3N4 photocatalytic material in the present application avoids the environmental pollution caused by traditional chemical reagents, and has better environmental friendliness and sustainability.

[0024] (3) Simple synthesis method: In the present application, only CdS / C3N4 photocatalytic material is added to the water containing Paracoccus denitrificans, and the bacteria and the material are adsorbed together by electrostatic attraction to form a coupling system. This simple synthesis method reduces the technical requirements for implementing the method and lowers the threshold for implementation.

[0025] (4) Improving application flexibility: The simple synthesis process of the present application makes its application more flexible, and the ratio and amount of photocatalytic material and Paracoccus denitrificans can be flexibly adjusted according to different wastewater treatment needs to achieve the best denitrification effect.

[0026] (5) Good denitrification effect: under light conditions, the coupling system prepared by the application can promote the growth and cell activity of Paracoccus denitrificans, and the photocatalytic effect of the material reduces the degree of light damage to the bacteria. In addition, the electrons of the photocatalytic material can provide additional electron donors to the bacteria after being excited under light, promoting reactions involving electron transfer in bacterial metabolism, such as the denitrification process. Therefore, through the photocatalytic effect and electron transfer of the photocatalytic material under light conditions, the bacterial-semiconductor coupling system can achieve a more efficient denitrification process, and the coupling system is suitable for the treatment of wastewater containing nitrate ions. BRIEF DESCRIPTION OF DRAWINGS

[0027] Figure 1 It is a scanning electron microscope graph of the denitrifying bacteria Paracoccus denitrificans and CdS / C3N4 photocatalytic coupling system prepared in Example 1 of the application; wherein (a) is a scanning electron microscope graph, and (b) is a local enlarged view of Figure (a);

[0028] Figure 2 It is a Fourier transform infrared spectrum graph of the denitrifying bacteria Paracoccus denitrificans and CdS / C3N4 photocatalytic material coupling system prepared in Example 1 of the application;

[0029] Figure 3 It is a denitrification effect graph of Example 1 of the application, i.e. the content of nitrate in the medium of each system at different time periods;

[0030] Figure 4 It is a denitrification effect graph of Example 1 of the application, i.e. the total nitrate nitrogen removal rate of each system at different time periods;

[0031] Figure 5 It is a denitrification effect graph of Example 2 of the application, i.e. the content of nitrate in the medium of each system at different time periods;

[0032] Figure 6 It is a denitrification effect graph of Example 1 of the application, i.e. the total nitrate nitrogen removal rate of each system at different time periods;

[0033] Figure 7 It is the protein content of denitrifying bacteria Paracoccus denitrificans in the dark, denitrifying bacteria Paracoccus denitrificans under light, and denitrifying bacteria Paracoccus denitrificans + CdSC3N4 system in the application;

[0034] Figure 8 It is the extracellular polymer content of denitrifying bacteria Paracoccus denitrificans in the dark, denitrifying bacteria Paracoccus denitrificans under light, and denitrifying bacteria Paracoccus denitrificans + CdSC3N4 system in the application;

[0035] Figure 9 It is a photocurrent response graph of CdSC3N4 and denitrifying bacteria Paracoccus denitrificans + CdSC3N4 in the application;

[0036] Figure 10 NADH content in Paracoccus denitrificans under dark, Paracoccus denitrificans under light, Paracoccus denitrificans+CdSC3N4 system;

[0037] Figure 11 Figure is a diagram of electron transfer system activity of Paracoccus denitrificans under dark, Paracoccus denitrificans under light, Paracoccus denitrificans+CdSC3N4 system in the present application. DETAILED DESCRIPTION

[0038] In order to facilitate the understanding of the present application, the present application will be described more fully below with reference to the accompanying drawings. The preferred embodiments of the present application are shown in the drawings. However, the present application can be implemented in various forms, and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the disclosure of the present application more thorough and comprehensive.

[0039] The preparation method of the CdS / C3N4 photocatalytic material and the denitrifying bacteria coupling system of the present application comprises the following steps:

[0040] S1, culture and amplification of denitrifying microorganisms: inoculate the strain Paracoccus denitrificans (ATCC 19367) of denitrifying microorganisms into LB culture medium, then transfer it to a culture bottle, and place the culture bottle in a constant temperature shaker, set at 30℃ and 120r / min to carry out amplification culture, amplify the denitrifying bacteria for 24 hours, and obtain LB culture medium containing denitrifying bacteria.

[0041] In step S1, the strain of denitrifying microorganisms is the purchased Paracoccus denitrificans (ATCC 19367) strain; the formula of the LB culture medium is: dissolve 10g of tryptone, 10g of NaCl and 5g of yeast extract in 1L of distilled water.

[0042] S2, preparation of CdS / C3N4 photocatalytic material: place urea in a muffle furnace, heat it to 500℃ at a rate of 5℃ / min, and keep it for 3 hours. After the obtained product is naturally cooled to room temperature, C3N4 material is obtained. Take 240mg of prepared C3N4 and suspend it in 50mL of ultrapure water, and add 130mg of Cd(NO3)2·4H2O. After ultrasonic treatment for 30min, a suspension is obtained. Add 0.15g of Na2S·9H2O to the suspension to obtain a precipitate. After washing the precipitate with ultrapure water for several times, place it in a 60℃ vacuum drying oven to dry overnight to obtain CdS / C3N4 material.

[0043] Preparation of trace element aqueous solution: Na2-EDTA, FeCl3·6H2O, MnCl2·4H2O, Na2MoO4·2H2O, CuCl2·2H2O and ZnCl2 were added into sterile water to prepare the trace element aqueous solution, which contained 9.6 mmol / L Na2-EDTA, 9.0 mmol / L FeCl3·6H2O, 0.1 mmol / L MnCl2·4H2O, 1.0 mmol / L Na2MoO4·2H2O, 0.8 mmol / L CuCl2·2H2O and 2.5 mmol / L ZnCl2.

[0044] Preparation of denitrification medium: 3.14 g of KNO3, 0.38 g of NH4Cl, 11.74 g of Na2HPO4·12H2O, 1.46 g of NaH2PO4, 3.54 g of CH3COONa and 0.10 mL of trace element aqueous solution were added into 1 L of sterile water to obtain a mixture, the mixture was autoclaved at 121 ℃ for 20 min, then 0.06 g of MgSO4·7H2O was added into the mixture after being dissolved by passing through a sterilization filter, and then hydrochloric acid or sodium hydroxide solution was used to adjust the pH to be in the range of 7-8, then argon was introduced and the mixture was sealed to isolate air, thereby obtaining the denitrification medium.

[0045] Preparation of denitrification medium containing CdS / C3N4 photocatalytic material: the prepared CdS / C3N4 photocatalytic material was irradiated under a UV lamp for 1 hour for sterilization, then the sterilized CdS / C3N4 photocatalytic material was mixed with the denitrification medium at a mass ratio of 1:(1000-5000) to obtain a mixture. Subsequently, the mixture was placed in an ultrasonic instrument at room temperature for ultrasonic treatment for 10 minutes to ensure that the CdS / C3N4 photocatalytic material was fully dispersed, thereby obtaining the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium.

[0046] S3, inoculation of denitrifying bacteria: the LB medium containing denitrifying bacteria obtained in step S1 was centrifuged at 6000 rpm for 5 min to obtain wet bacteria. The wet bacteria were washed with PBS buffer solution, and then the washed wet bacteria were inoculated (i.e., added) into the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium obtained in step S2, thereby obtaining a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system.

[0047] The ratio of the LB medium containing denitrifying bacteria to the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium was 1 mL:(200-500) mL.

[0048] The denitrification culture was carried out under light intensity of one solar intensity, and the content of nitrate in the denitrification medium was tested periodically.

[0049] Example 1

[0050] S1, culture and expansion of denitrifying microorganisms: inoculate the purchased Paracoccus denitrificans (ATCC 19367) strain of denitrifying bacteria into LB culture medium, then transfer to a culture bottle, place the culture bottle in a constant temperature shaker, set at 30°C and 120 r / min, and carry out expansion culture for 24 hours to obtain LB culture medium containing denitrifying bacteria.

[0051] S2, preparation of CdS / C3N4 photocatalytic material: place urea in a muffle furnace, heat to 500°C at a rate of 5°C / min, and maintain for 3 hours. After the obtained product is naturally lowered to room temperature, C3N4 material is obtained. Take 240 mg of prepared C3N4 and suspend in 50 mL of ultrapure water, and add 130 mg of Cd(NO3)2·4H2O. After ultrasonic treatment for 30 min, a suspension is obtained. Add 0.15 g of Na2S·9H2O to the suspension to obtain a precipitate. After washing the precipitate with ultrapure water for multiple times, place it in a 60°C vacuum drying oven to dry overnight to obtain CdS / C3N4 material.

[0052] Preparation of trace element aqueous solution: Na2-EDTA, FeCl3·6H2O, MnCl2·4H2O, Na2MoO4·2H2O, CuCl2·2H2O and zinc chloride are added to sterile water to obtain a trace element aqueous solution containing 9.6 mmol / L Na2-EDTA, 9.0 mmol / L FeCl3·6H2O, 0.1 mmol / L MnCl2·4H2O, 1.0 mmol / L Na2MoO4·2H2O, 0.8 mmol / L CuCl2·2H2O and 2.5 mmol / L zinc chloride.

[0053] Preparation of denitrification medium: add 3.14 g of KNO3, 0.38 g of NH4Cl, 11.74 g of Na2HPO4·12H2O, 1.46 g of NaH2PO4, 3.54 g of CH3COONa and 0.10 mL of trace element aqueous solution to 1 L of sterile water to obtain a mixture, autoclave the mixture at 121°C for 20 min, then to prevent the formation of precipitate, add 0.06 g of MgSO4·7H2O dissolved by a sterile filter, then use hydrochloric acid or sodium hydroxide solution to adjust the pH to be in the range of 7-8, then pass in argon and seal to isolate air to obtain a denitrification medium.

[0054] Preparation of denitrification medium containing CdS / C3N4 photocatalyst: The prepared CdS / C3N4 photocatalyst was sterilized by irradiating it under a UV lamp for 1 hour. The sterilized CdS / C3N4 photocatalyst was then mixed with the denitrification medium at a mass ratio of 1:2000 to obtain a mixture. The mixture was then placed in an ultrasonic instrument at room temperature for ultrasonic treatment for 10 minutes to ensure that the CdS / C3N4 photocatalyst was fully dispersed, thus obtaining a mixture of CdS / C3N4 photocatalyst and denitrification medium.

[0055] S3, Inoculation with denitrifying bacteria: Take the LB medium containing denitrifying bacteria obtained in step S1, centrifuge at 6000 rpm for 5 min to obtain wet bacterial cells. Wash the wet bacterial cells with PBS buffer solution, and then inoculate them into the mixture of CdS / C3N4 photocatalyst and denitrifying medium obtained in step S2, to obtain the CdS / C3N4 photocatalyst and denitrifying bacteria coupling system. The ratio of LB medium containing denitrifying bacteria to the mixture of CdS / C3N4 photocatalyst and denitrifying medium is 1 mL: 200 mL.

[0056] Take 50 mL of CdS / C3N4 photocatalyst material and denitrifying bacteria coupling system and put them into a multi-channel instrument. Adjust the light intensity to the intensity of sunlight and the rotation speed to 120 rpm / min. Test the nitrate content in the denitrification medium periodically.

[0057] Figures 1-2 The images are scanning electron microscope images and Fourier transform infrared spectra of denitrifying bacteria combined with CdS / C3N4 photocatalytic material, respectively.

[0058] Depend on Figure 1 It can be seen that the CdS / C3N4 photocatalytic material is closely attached to the denitrifying bacteria, and the smaller photocatalytic material is attached to the denitrifying bacteria.

[0059] Depend on Figure 2 It can be seen that denitrifying bacteria thrive at 3290 cm³. -1 The NH stretching vibration peak decreases at 795.7 cm⁻¹, while in CdS-C₃N₄, it is at 795.7 cm⁻¹. -1 At this time, the hydrogen bonds between amino groups weaken, meaning that hydrogen bonds (-C--N·…·HN) are formed. This occurs at 1380–1450 cm⁻¹. -1 and 2870~2960cm -1 The decreased intensity of the CH bond vibration signal in the cell membrane of the region further confirmed the coupling between -C--N and alkyl groups.

[0060] Performance testing: *Paracococcus denitrifyingans* and *Paracococcus denitrifyingans* + CdS / C3N4 were exposed to sunlight at the same intensity. *Paracococcus denitrifyingans* was exposed to darkness, and *Paracococcus denitrifyingans* + CdS / C3N4 were exposed to darkness. Denitrification rates were tested at different time points (3h, 6h, and 9h). Results are shown in [link to results]. Figure 3 and Figure 4 .

[0061] Depend on Figure 3 It is known that light irradiation inhibits the denitrification rate of bacteria. However, the addition of photocatalytic materials significantly increases the denitrification rate of the previously inhibited denitrifying bacteria. Furthermore, in the dark, the addition of photocatalytic materials does not significantly inhibit the denitrification of *Paracococcus denitrifyingus*, indicating the environmental friendliness of this photocatalytic material.

[0062] Depend on Figure 4 It can be seen that under light conditions, the addition of photocatalytic materials increased the total nitrate nitrogen removal efficiency of denitrifying bacteria from 0.92% to 93.88% within 11 hours. This proves that under light, the addition of CdS / C3N4 photocatalytic materials significantly improves the denitrification rate of Paracoccus denitrifying. Therefore, the present invention can treat wastewater containing nitrate ions.

[0063] Example 2

[0064] S1, Cultivation and amplification of denitrifying microorganisms: The purchased denitrifying bacteria, Paracoccus denitrifyingis, was inoculated into LB medium and then transferred to a culture flask. The culture flask was placed in a constant temperature shaker and cultured at 30℃ and 120r / min for 24 hours to obtain LB medium containing denitrifying bacteria.

[0065] Preparation of S2, CdS / C3N4 photocatalytic material: Same as in Example 1;

[0066] Preparation of trace element aqueous solution: Same as in Example 1;

[0067] Preparation of denitrification medium: Same as in Example 1;

[0068] Preparation of denitrification medium containing CdS / C3N4 photocatalysis: The prepared CdS / C3N4 photocatalyst material was sterilized by irradiation under a UV lamp for 1 hour. The sterilized CdS / C3N4 photocatalyst material was then mixed thoroughly with the denitrification medium at a mass ratio of 1:5000 to obtain a mixture, specifically, 0.01 g of CdS / C3N4 photocatalyst material was added to 50 mL of medium. The mixture was then subjected to ultrasonic treatment at room temperature for 10 minutes to ensure complete dispersion of the CdS / C3N4 photocatalyst material, resulting in a mixture of CdS / C3N4 photocatalyst material and denitrification medium.

[0069] S3, Inoculation with denitrifying bacteria: Take the LB medium containing *Paracoccus denitrifyingus* obtained in step S1, centrifuge at 6000 rpm for 5 min to obtain wet bacterial cells. Wash the wet bacterial cells with PBS buffer solution and inoculate them into the denitrification medium containing photocatalytic material treated in step S2, thus obtaining the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system. The ratio of LB medium containing denitrifying bacteria to the mixture of CdS / C3N4 photocatalytic material and denitrification medium is 1 mL: 200 mL.

[0070] Take 50 mL of CdS / C3N4 photocatalyst material and place it into a multichannel instrument coupled with denitrifying bacteria. Adjust the light intensity to the level of sunlight and the rotation speed to 120 rpm / min. Periodically test the nitrate content in the denitrification medium.

[0071] Depend on Figure 6 It can be seen that under light conditions, when the CdS / C3N4 photocatalyst material and denitrification medium are mixed at a mass ratio of 1:5000, the nitrate nitrogen removal rate can be increased from 0.92% to 16.64% in 11 hours. When the CdS / C3N4 photocatalyst material and denitrification medium are mixed at a mass ratio of 1:1600, the nitrate nitrogen removal rate can be increased from 0.92% to 95.10% in 11 hours. This example demonstrates that by reasonably adding CdS / C3N4 photocatalyst material, the denitrification effect can be improved. After simple adjustments, this method can be adapted to denitrification processes with different water qualities.

[0072] Example 3

[0073] Similar to Example 2, except that the sterilized CdS / C3N4 photocatalyst material was mixed evenly with the denitrification culture medium at a mass ratio of 1:1666 to obtain a mixture, that is, 0.03g of CdS / C3N4 photocatalyst material was added to 50mL of culture medium.

[0074] Depend on Figure 5 It is evident that different amounts of photocatalytic material added result in varying effects on increasing the denitrification rate. The more photocatalytic material added, the better the denitrification effect of *Paracoccus denitrifyingans*.

[0075] Example 4

[0076] S1, same as in Example 1;

[0077] Preparation of S2, CdS / C3N4 photocatalytic material: Same as in Example 1;

[0078] Preparation of trace element aqueous solution: Same as in Example 1;

[0079] Preparation of denitrification medium: Same as in Example 1;

[0080] Preparation of the denitrification medium containing CdS / C3N4 photocatalysis: the prepared CdS / C3N4 photocatalytic material was irradiated under a UV lamp for 1 hour, sterilized, and then mixed with the denitrification medium at a mass ratio of 1:3000 to obtain a mixture. Subsequently, the mixture was placed in an ultrasonic instrument at room temperature for ultrasonic treatment for 10 minutes to ensure that the CdS / C3N4 photocatalytic material was fully dispersed, thereby obtaining the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium.

[0081] S3, inoculation of denitrifying bacteria: the LB medium containing the Paracoccus denitrificans obtained in step S1 was centrifuged at 6000 rpm for 5 min to obtain wet bacteria. After the wet bacteria were washed with a PBS buffer solution, they were inoculated into the denitrification medium containing the photocatalytic material treated in step S2 to obtain a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system. The ratio of the LB medium containing the denitrifying bacteria to the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium was 1 mL:200 mL.

[0082] Example 5

[0083] S1, same as example 1;

[0084] S2, preparation of CdS / C3N4 photocatalytic material: same as example 1;

[0085] Preparation of trace element aqueous solution: same as example 1;

[0086] Preparation of denitrification medium: same as example 1;

[0087] Preparation of the denitrification medium containing CdS / C3N4 photocatalysis: the prepared CdS / C3N4 photocatalytic material was irradiated under a UV lamp for 1 hour, sterilized, and then mixed with the denitrification medium at a mass ratio of 1:3000 to obtain a mixture. Subsequently, the mixture was placed in an ultrasonic instrument at room temperature for ultrasonic treatment for 10 minutes to ensure that the CdS / C3N4 photocatalytic material was fully dispersed, thereby obtaining the mixture of the CdS / C3N4 photocatalytic material and the denitrification medium.

[0088] S3, inoculation of denitrifying bacteria: the LB medium containing Paracoccus denitrificans obtained from step S1 was centrifuged at 6000 rpm for 5 min to obtain wet bacteria. The wet bacteria were washed with PBS buffer solution, and then inoculated into the denitrification medium containing the photocatalytic material treated in step S2 to obtain a coupling system of CdS / C3N4 photocatalytic material and denitrifying bacteria. The ratio of the mixture of the LB medium containing denitrifying bacteria and the denitrification medium containing the CdS / C3N4 photocatalytic material was 1 mL: 500 mL.

[0089] Example 6

[0090] S1, same as example 1;

[0091] S2, preparation of CdS / C3N4 photocatalytic material: same as example 1;

[0092] Preparation of trace element aqueous solution: same as example 1;

[0093] Preparation of denitrification medium: same as example 1;

[0094] Preparation of denitrification medium containing CdS / C3N4 photocatalytic material: the prepared CdS / C3N4 photocatalytic material was sterilized under ultraviolet light for 1 hour, and then mixed with the denitrification medium at a mass ratio of 1:1000 to obtain a mixture. The mixture was then placed in an ultrasonic instrument at room temperature for ultrasonic treatment for 10 minutes to ensure that the CdS / C3N4 photocatalytic material was fully dispersed, thereby obtaining a mixture of the CdS / C3N4 photocatalytic material and the denitrification medium.

[0095] S3, inoculation of denitrifying bacteria: the LB medium containing Paracoccus denitrificans obtained from step S1 was centrifuged at 6000 rpm for 5 min to obtain wet bacteria. The wet bacteria were washed with PBS buffer solution, and then inoculated into the denitrification medium containing the photocatalytic material treated in step S2 to obtain a coupling system of CdS / C3N4 photocatalytic material and denitrifying bacteria. The ratio of the mixture of the LB medium containing denitrifying bacteria and the denitrification medium containing the CdS / C3N4 photocatalytic material was 1 mL: 500 mL.

[0096] Microbial denitrification is carried out by denitrifying microorganisms, and the growth and proliferation of these microorganisms have a significant impact on the process. Meanwhile, extracellular polymers (APIs) are biopolymers secreted by microorganisms, providing a protective barrier against adverse external environmental conditions and storing nutrients. Furthermore, the secretion of APIs can reflect bacterial activity. Therefore, under the conditions of Example 1, the protein and API content of each system (i.e., *Paracoccus denitrifyingans* under darkness, *Paracoccus denitrifyingans* under light, and *Paracoccus denitrifyingans* + CdS / C3N4 system) after 11 hours of denitrification were tested to assess bacterial growth.

[0097] After 11 hours of reaction, all denitrification medium from each system was removed. The bacteria in each system were washed three times with PBS buffer and resuspended. Intracellular proteins were extracted using an ice-water bath sonication method, and the intracellular protein content of each system was tested. Figure 7 It can be seen that under light, the protein content of *Paragonimula denitrificans* decreases significantly, to about one-third of that of *Paragonimula monocytogenes* under darkness, indicating that light inhibits the growth of *Paragonimula denitrificans*. However, the addition of the material increases the protein content, proving that the addition of the CdS / C3N4 photocatalytic material further promotes protein growth in the system.

[0098] After 11 hours of reaction, all denitrification culture media from each system were removed, and the OD600 values ​​of each system were adjusted with PBS buffer to ensure consistency. 10 ml of the adjusted OD600 culture media from each system was then used to wash the bacteria three times with PBS buffer to obtain bacterial precipitates. The extracellular polymeric content of the bacterial precipitates was then tested.

[0099] Depend on Figure 8 It was found that under light irradiation, the amount of extracellular polymers secreted by bacteria decreased, but the addition of CdS-C3N4 material under light irradiation could promote the secretion of extracellular polymers by bacteria. This further demonstrates the promoting effect of CdS / C3N4 photocatalytic material on the activity of denitrifying paracocci.

[0100] Microbial denitrification requires the participation of electrons, which mainly come from photoelectrons converted from light energy captured by materials and direct electron donors (NADH). Electrons are transferred through electron transport systems.

[0101] The CdS / C3N4 material and the denitrifying paracoccus + CdS-C3N4 coupling system were tested for photocurrent under simulated sunlight using a 100W LED lamp with an AM1.5 filter and denitrification medium as the electrolyte.

[0102] Depend on Figure 9 It can be seen that the introduction of *Paracococcus denitrifyingans* reduced the peak photocurrent during light irradiation and accelerated the decrease in photocurrent during the dark reaction. The photocurrent indicates that *Paracococcus denitrifyingans* captured photoelectrons.

[0103] The effect of the material on the intracellular direct electron donor of the denitrifying bacteria was studied by measuring the content of NADH in each system after 11 hours of reaction. Figure 10 It can be seen that light inhibits the production of intracellular electron donor, but the content of NADH in the denitrifying bacteria is increased after the addition of CdS / C3N4, which exceeds the content of NADH in the bacteria in the dark, and the denitrification process is accelerated due to the sufficient electron donor.

[0104] The electron transfer efficiency is closely related to the denitrification performance of Paracoccus denitrificans. The activity of the electron transfer system in each system after 11 hours of reaction was tested. Figure 11 As shown in Table 2, compared with the dark condition, light significantly inhibits the electron transfer efficiency of Paracoccus denitrificans, but the addition of CdS / C3N4 under light significantly improves the electron transfer activity of the bacteria, and exceeds the electron transfer efficiency of the bacteria in the dark. It is proved that the CdS / C3N4 material can adjust the operation of the intracellular electron transfer chain, and make it more efficient.

[0105] The present application fully proves that under light conditions, the material can promote the growth and cell activity of Paracoccus denitrificans, and the photocatalytic effect of the material reduces the degree of light damage to the bacteria. In addition, the electrons of the semiconductor material can provide additional electron donors to the bacteria after being excited under light, and promote the reactions involving electron transfer in bacterial metabolism, such as the denitrification process. Therefore, through the photocatalytic effect and electron transfer effect of the semiconductor material under light conditions, the coupling system of bacteria and semiconductor can realize a more efficient denitrification process.

[0106] The above only describes the best embodiments of the present application, but cannot be understood as limiting the claims. The present application is not limited to the above embodiments, and the specific structure allows changes. Any changes made within the protection scope of the independent claims of the present application are within the protection scope of the present application.

[0107] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used in the description of the application herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application. As used herein, the term "and / or" includes any and all combinations of one or more of the associated listed items.

Claims

1. A method for preparing a coupling system of CdS / C3N4 photocatalytic material and denitrifying bacteria, characterized in that, Includes the following steps: After sterilizing the CdS / C3N4 photocatalyst material, it was mixed with the denitrification medium at a mass ratio of 1:(1000-5000) to obtain a mixture of CdS / C3N4 photocatalyst material and denitrification medium. After centrifuging the LB medium containing denitrifying bacteria, wet bacterial cells were obtained. The wet bacterial cells were washed and added to a mixture of CdS / C3N4 photocatalyst and denitrification medium to obtain a CdS / C3N4 photocatalyst and denitrifying bacteria coupling system. Preparation of CdS / C3N4 photocatalytic material: Urea was heated at 500℃ for 3 hours to obtain C3N4 material. The C3N4 material was suspended in 50mL of ultrapure water, and Cd(NO3)2·4H2O was added. After sonication, a suspension was obtained. Na2S·9H2O was added to the suspension to obtain a precipitate. After drying, CdS / C3N4 photocatalytic material was obtained. The denitrification medium is prepared by the following process: KNO3, NH4Cl, Na2HPO4·12H2O, NaH2PO4, CH3COONa and trace element aqueous solution are added to sterile water to obtain a mixture. After sterilizing the mixture, MgSO4·7H2O is added and the pH is adjusted to 7-8. Then argon gas is introduced and the mixture is sealed to obtain the denitrification medium. LB medium containing denitrifying bacteria is prepared through the following process: Cultivation and amplification of denitrifying microorganisms: The strain of denitrifying microorganism Paracoccus denitrificans is inoculated into LB medium and amplified to obtain LB medium containing denitrifying bacteria. The LB medium formula is: 10g tryptone, 10g NaCl and 5g yeast extract are dissolved in 1L of distilled water.

2. The preparation method of the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system according to claim 1, characterized in that, The ratio of KNO3, NH4Cl, Na2HPO4·12H2O, NaH2PO4, CH3COONa, trace element aqueous solution, sterile water and MgSO4·7H2O is 3.14g:0.38g:11.74g:1.46g:3.54g:0.10mL:1L:0.06g.

3. The preparation method of the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system according to claim 1, characterized in that, The trace element aqueous solution is prepared by adding Na2-EDTA, FeCl3·6H2O, MnCl2·4H2O, Na2MoO4·2H2O, CuCl2·2H2O and zinc chloride to sterile water.

4. The preparation method of the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system according to claim 3, characterized in that, The trace element aqueous solution contains 9.6 mmol / L Na2-EDTA, 9.0 mmol / L FeCl3·6H2O, 0.1 mmol / L MnCl2·4H2O, 1.0 mmol / L Na2MoO4·2H2O, 0.8 mmol / L CuCl2·2H2O, and 2.5 mmol / L zinc chloride.

5. The preparation method of the CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system according to claim 1, characterized in that, The ratio of LB medium containing denitrifying bacteria to the mixture of CdS / C3N4 photocatalyst and denitrifying medium is 1 mL:(200-500) mL.

6. A coupling system of CdS / C3N4 photocatalytic material and denitrifying bacteria prepared by any one of claims 1-5.

7. The application of a CdS / C3N4 photocatalytic material and denitrifying bacteria coupling system prepared according to any one of claims 1-5 in the treatment of nitrate ion-containing wastewater.

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