Double-embedded particle, preparation method thereof and application of double-embedded particle in removal of mixed estrogens in sewage

Abstract

The invention provides double-embedded particles for sewage treatment and application of the double-embedded particles in removal of mixed estrogens in sewage, and belongs to the technical field of sewage treatment. Artificial photosynthetic particles embedded with a photocatalyst and white-rot fungi and natural photosynthetic particles embedded with bacteria and green algae are coupled to prepare the double-embedded particles. The double-embedded particles have a good removal effect on binding state estrogen and artificially synthesized free state estrogen in sewage. The double-embedded photosynthetic particle solves the problems of low biological treatment degradation efficiency and poor toxicity reduction effect of advanced oxidation treatment in the prior art, and provides a new means for sewage treatment.

Description

Technical Field

[0001] The invention belongs to the technical field of sewage treatment, and in particular relates to a double-embedded particle and a preparation method thereof and an application of the double-embedded particle in removing mixed estrogens in sewage. Background Art

[0002] The operation of municipal sewage treatment plants faces new challenges and opportunities. Although the traditional secondary biochemical treatment process has achieved remarkable results in reducing conventional pollution indicators and meets basic environmental protection requirements, it is inadequate when facing those difficult-to-degrade pollutants with high toxicity and biological resistance. For a long time, estrogen has been widely studied and applied due to its special role in organisms, but its residue and accumulation in the natural environment have gradually revealed its ecological risks that cannot be ignored.

[0003] In the field of sewage treatment, advanced oxidation technology and bioaugmentation technology are two cutting-edge strategies for the deep purification of refractory estrogens, each showing its unique advantages and limitations. First of all, advanced oxidation technology relies on its rapid reaction speed and excellent degradation efficiency, but the double-edged sword nature of this technology cannot be ignored - its degradation products may carry more serious toxicity challenges than the original pollutants, which to a certain extent limits its prospects for widespread application. Bioaugmentation technology is particularly good at reducing the toxicity of pollutants and contributing to environmental restoration and ecological balance. However, due to the biological inhibition of refractory pollutants, the degradation efficiency of traditional bioaugmentation technology is often difficult to achieve expectations, and the effect is unsatisfactory when used alone, which has become a key factor restricting its effectiveness. Summary of the invention

[0004] In view of this, the object of the present invention is to provide a double-embedded particle for sewage treatment, by mixing artificial photosynthetic particles obtained by embedding photocatalysts and white rot fungi with natural photosynthetic particles formed by the symbiosis of bacteria and green algae to synergistically exert the estrogen degradation effect, thereby achieving the purpose of effectively removing estrogen pollutants in sewage.

[0005] The present invention provides a double-embedded particle, comprising an artificial photosynthetic particle and a natural photosynthetic particle;

[0006] The artificial photosynthetic particles include calcium alginate hydrogel and a photocatalyst and white rot fungi embedded in the calcium alginate hydrogel; the photocatalyst includes the MOF material NH2-MIL-53(Fe);

[0007] The natural photosynthetic particles include calcium alginate hydrogel and green algae and bacillus subtilis embedded in the calcium alginate hydrogel.

[0008] Preferably, the concentration of the photocatalyst is 0.1 to 0.5 mg / mL;

[0009] The concentration of the white rot fungus was 3.2×10 8 CFU / mL~4×10 8 CFU / mL.

[0010] Preferably, the green algae includes Chlorella vulgaris; the concentration of the green algae is 1×10 9 cells / L~2×10 9 cells / L;

[0011] The concentration of Bacillus subtilis is 0.6×10 9 CFU / L~0.8×10 9 CFU / L.

[0012] Preferably, the quantity ratio of the artificial photosynthetic particles to the natural photosynthetic particles is (1-6):(1-6).

[0013] Preferably, the preparation method of NH2-MIL-53(Fe) is to dissolve ferric chloride and 2-aminoterephthalic acid in an organic solvent, perform hydrothermal synthesis at 150-170°C for 18-24 hours under sealed conditions, cool the mixture, wash with an organic solvent, remove the solvent, and obtain NH2-MIL-53(Fe).

[0014] The present invention provides a method for preparing the double-embedded particles, comprising the following steps:

[0015] collecting white rot fungus mycelium and / or spores from the cultured white rot fungus liquid, dispersing the white rot fungus mycelium and / or spores and NH2-MIL-53(Fe) in a sodium alginate solution to obtain a material-fungus mixed liquid;

[0016] Dropping the material-fungus mixture into a calcium chloride solution, and undergoing a solidification reaction, obtaining artificial photosynthetic particles;

[0017] Separating green algae bodies from the cultured green algae liquid, separating Bacillus subtilis cells from the cultured Bacillus subtilis liquid, and dispersing the green algae bodies and Bacillus subtilis cells in a sodium alginate solution to obtain a green algae-bacteria mixed solution;

[0018] The green algae-bacteria mixed solution is dripped into a calcium chloride solution to obtain natural photosynthetic particles through a solidification reaction;

[0019] The artificial photosynthetic particles and the natural photosynthetic particles are mixed to obtain double-embedded particles.

[0020] The present invention provides the use of the double-embedded particles or the double-embedded particles prepared by the preparation method in degrading bound estradiol and / or free estradiol in sewage.

[0021] The present invention provides a method for degrading bound estradiol and / or free estradiol in sewage based on the double-embedded particles, comprising the following steps:

[0022] The double-embedded particles are mixed with the sewage to be treated, and the light culture program is run: the light intensity is 60 to 240 μmol·s -1 ·m -2 , the temperature is 20-25℃, and the photoperiod is 6-18h / day.

[0023] Preferably, the drainage ratio during each photoperiod is 50% to 55%, and the hydraulic retention time is 4 to 6 hours.

[0024] Preferably, the operation cycle of the light culture includes water intake, standing, aeration, sedimentation and drainage in sequence;

[0025] The water inlet time is 3 to 5 minutes;

[0026] The standing time is 5 to 10 minutes;

[0027] The aeration time is 205-220 min; the aeration volume is 0.5-1 L / min;

[0028] The precipitation time is 10 to 15 minutes;

[0029] The drainage time is 3 to 5 minutes.

[0030] The present invention provides a double-embedded particle, comprising an artificial photosynthetic particle and a natural photosynthetic particle; the artificial photosynthetic particle comprises a calcium alginate hydrogel and a photocatalyst and a white rot fungus embedded in the calcium alginate hydrogel; the photocatalyst comprises NH2-MIL-53(Fe); the natural photosynthetic particle comprises a calcium alginate hydrogel and green algae and Bacillus subtilis embedded in the calcium alginate hydrogel. The present invention has the following significant advantages:

[0031] (1) The present invention proposes the double-embedded particle technology for the first time. By immobilizing the microbial strains with estrogen degradation function, the survival ability and degradation efficiency of the microorganisms in the sewage environment can be significantly improved. By utilizing the stability of the immobilized microbial particles, the microorganisms can continue to play a degradation role in a complex water quality environment, thereby effectively removing estrogens in sewage, such as estradiol-3-sulfate (E2-3S) and 17α-ethinyl estradiol (EE2).

[0032] (2) Double-encapsulated particle technology can effectively resist these adverse factors by providing microorganisms with a relatively stable microenvironment, allowing microorganisms to maintain efficient degradation activity under a wider range of environmental conditions.

[0033] (3) The present invention uses double-embedded particle technology. On the one hand, the preparation and use of immobilized microbial particles are relatively simple and do not require complex equipment and a large amount of chemical agents; on the other hand, the degradation of microorganisms can be carried out at room temperature and pressure without the need for additional energy support.

[0034] (4) The present invention makes up for the shortcomings of the current deep purification process for difficult-to-degrade organic pollutants, improves the shortcomings of the existing technology of poor removal effect and unstable operation of difficult-to-degrade organic pollutants, and fills the gap in the economical and efficient removal technology of difficult-to-degrade organic pollutants in the secondary effluent of sewage treatment plants at home and abroad.

[0035] (5) The method provided by the present invention can be adjusted according to the characteristics and treatment requirements of different sewage, has high flexibility and controllability, and is suitable for a variety of different types of sewage treatment.

[0036] (6) The present invention embeds white rot fungi and NH2-MIL-53(Fe) to prepare artificial photosynthetic particles, so that the white rot fungi produce laccase, and NH2-MIL-53(Fe) serves as a fixed carrier of laccase. It is revealed that the active site of laccase (Lac) produced by the white rot fungi is fixed and covalently bound to the surface of NH2-MIL-53(Fe), which not only enhances the catalytic activity of the system, but also significantly shortens the accessibility distance of the active site.

[0037] (7) The present invention co-encapsulates green algae and Bacillus subtilis to make natural photosynthetic particles. The CO2 generated by sewage is absorbed by the microalgae, and the oxygen released by the photosynthesis of the microalgae is metabolized by microbial respiration, so that the energy utilization rate is increased to more than 70%, reducing the carbon emissions of aeration by about 50%. Natural photosynthetic particles can not only achieve self-sufficiency in oxygen and nutrients, but also convert carbon, nitrogen and phosphorus in sewage into high-value-added proteins and oils, turning waste into treasure while reducing carbon emissions, and providing an effective way to achieve near-zero discharge of urban sewage. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Schematic diagram of the structural composition of double-embedded particles;

[0039] Figure 2 Schematic diagram of the preparation process of double-embedded particles;

[0040] Figure 3 The scanning electron micrograph of double-embedded particles. DETAILED DESCRIPTION

[0041] The invention provides a double-embedded particle, comprising an artificial photosynthetic particle and a natural photosynthetic particle; the artificial photosynthetic particle comprises a calcium alginate hydrogel and a photocatalyst and a white rot fungus embedded in the calcium alginate hydrogel; the photocatalyst comprises NH2-MIL-53(Fe); and the natural photosynthetic particle comprises a calcium alginate hydrogel and green algae and Bacillus subtilis embedded in the calcium alginate hydrogel.

[0042] In the present invention, the concentration of the white rot fungi in the artificial photosynthetic particles is 3.2×10 8 CFU / mL~4×10 8 CFU / mL, can be 3.5×10 8 CFU / mL, 3.7×10 8 CFU / mL, 3.8×10 8 CFU / mL. The concentration of the photocatalyst is preferably 0.1-0.5 mg / mL, and can be 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL and 0.4 mg / mL. The preparation method of the NH2-MIL-53(Fe) is preferably to dissolve ferric chloride and 2-aminoterephthalic acid in an organic solvent, perform hydrothermal synthesis at 150-170°C under sealed conditions for 18-24 hours, wash with an organic solvent after cooling, remove the solvent, and obtain NH2-MIL-53(Fe). The molar ratio of ferric chloride and 2-aminoterephthalic acid is preferably 0.8-1.2:0.8-1.2, which can be 1:1. The organic solvent is preferably NN dimethylformamide (DMF). When dissolving, it is preferred to use an ultrasonic or magnetic stirring method to promote dissolution. When dissolving, it is preferred to first dissolve 2-aminoterephthalic acid in an organic solvent and then add ferric chloride. The temperature of the hydrothermal synthesis can be 160°C. The time of the hydrothermal synthesis can be 20 hours and 22 hours. The hydrothermal synthesis is preferably carried out in a polytetrafluoroethylene reactor. During washing, the reaction product is preferably cooled to room temperature (25°C). The washing is preferably performed by alternately washing and centrifuging with DMF and anhydrous ethanol, and repeated three times. After the washing, the obtained product is preferably dried. The drying temperature is preferably 70 to 90°C, and can be 80°C. The drying time is preferably 6 hours.

[0043] In the present invention, the working principle of the artificial photosynthetic particles is that white rot fungi have the characteristic of secreting laccase. In the presence of white rot fungi, laccase (Lac) will be produced in the system, and the photocatalyst captures visible light and generates electrons, which are transferred to T1 Cu (II) of laccase to start the enzymatic reaction. Since an organic covalent system is formed between the photocatalyst and laccase, the active site of laccase and the catalytic activity of laccase are improved, catalyzing the ring-opening and bond-breaking of estrogen, which is beneficial to remove mixed estrogens in pollutants.

[0044] In the present invention, in the natural photosynthetic particles, the green algae preferably includes Chlorella vulgaris. The concentration of the green algae is preferably 1×10 9 cells / L~2×10 9 cells / L, which can be 1.2×10 9 cells / L、1.4×10 9 cells / L、1.6×10 9 cells / L / 1.8×10 9 The concentration of Bacillus subtilis is preferably 0.6×10 9 CFU / L~0.8×10 9 CFU / L, can be 0.7×10 9 CFU / L. The green algae not only has the function of further removing mixed estrogens and reducing the estrogen effect produced by them under photosynthesis, but also produces oxygen for microbial respiration. The Bacillus subtilis further degrades the estrogen intermediates degraded by the photocatalyst, and can also produce CO2 and vitamins such as vitamin B1 and B2 for microalgae to absorb. The results of an embodiment of the present invention show that the green algae and Bacillus subtilis synergistically play a role in reducing the estrogen mixture in sewage.

[0045] In the present invention, the ratio of the number of artificial photosynthetic particles to that of natural photosynthetic particles is preferably 1-6:1-6, can be (1-3):(1-3), or can be 1:1. The working concentration of the artificial photosynthetic particles in the double-embedded particles is preferably 100-600 pcs / L, can be 200 pcs / L, 300 pcs / L, 400 pcs / L, 500 pcs / L; the working concentration of the natural photosynthetic particles is 100-600 pcs / L, can be 200 pcs / L, 300 pcs / L, 400 pcs / L, 500 pcs / L. In one embodiment of the present invention, the results show that the artificial photosynthetic particles and the natural photosynthetic particles work together in degrading the bound estrogen (E2-3S) and the synthetic free estrogen (EE2) in the sewage, and the removal efficiency of the estrogen using only the natural photosynthetic particles will be greatly reduced.

[0046] The present invention provides a method for preparing the double-embedded particles, comprising the following steps:

[0047] collecting white rot fungus mycelium and / or spores from the cultured white rot fungus liquid, dispersing the white rot fungus mycelium and / or spores and NH2-MIL-53(Fe) in a sodium alginate solution to obtain a material-fungus mixed liquid;

[0048] Dropping the material-fungus mixture into a calcium chloride solution, and undergoing a solidification reaction, obtaining artificial photosynthetic particles;

[0049] Separating green algae bodies from the cultured green algae liquid, separating Bacillus subtilis cells from the cultured Bacillus subtilis liquid, and dispersing the green algae bodies and Bacillus subtilis cells in a sodium alginate solution to obtain a green algae-bacteria mixed solution;

[0050] The green algae-bacteria mixed solution is dripped into a calcium chloride solution to obtain natural photosynthetic particles through a solidification reaction;

[0051] The artificial photosynthetic particles and the natural photosynthetic particles are mixed to obtain double-embedded particles.

[0052] In the present invention, the method for preparing the white rot fungus liquid is to inoculate the white rot fungus into a comprehensive potato culture medium (pH 6), culture it at 28-32°C for 7-10 days, and wait until the OD 600 When the value is 0.8 to 1.0, the mycelium or spores are collected. The method for collecting the mycelium and / or spores of white rot fungi is preferably centrifugation. The speed of the centrifugation is preferably 5000 to 7000 rpm, and can be 6000 rpm. The time of the centrifugation is preferably 9 to 11 minutes, and can be 10 minutes.

[0053] In the present invention, the method for preparing the green algae liquid preferably inoculates the green algae species into BG11 medium, cultures them at 24-26°C and 1800-2200 Lux for 1-2 weeks, and collects the algae liquid. The method for preparing the Bacillus subtilis liquid preferably inoculates Bacillus subtilis into a nutrient broth medium (pH 7.0-7.2), cultures them at 28-30°C for 24-48 hours, and waits for the OD value of the liquid to reach 0.01. 600 When the pH value is between 0.8 and 1.0, collect the bacterial solution.

[0054] In the present invention, the mass percentage of the sodium alginate solution is preferably 1% to 2%, and can be 1.5%. The pH value of the sodium alginate solution is preferably 7 to 7.5, and can be 7.2 or 7.4. The sodium alginate solution is preferably sterilized before use. The method of the sterilization treatment is preferably 131°C for 30 minutes. The concentration of the calcium chloride solution is preferably 2% to 5%, and can be 3% or 4%. The pH value of the calcium chloride solution is preferably 7 to 7.5, and can be 7.2 or 7.4. The calcium chloride solution is preferably sterilized before use. The method of the sterilization treatment is preferably 131°C for 30 minutes. The method of dripping the calcium chloride solution is preferably to use a 5mL syringe to hold the solution to be dripped. The time of the curing reaction is preferably 30 to 60 minutes, and can be 40 minutes or 50 minutes.

[0055] The present invention provides the use of the double-embedded particles or the double-embedded particles prepared by the preparation method in degrading bound estradiol and / or free estradiol in sewage.

[0056] In the present invention, the bound estradiol preferably includes estradiol-3-sulfate. The free estradiol preferably includes 17α-ethinyl estradiol. The sewage is preferably secondary influent of a sewage treatment plant.

[0057] The present invention provides a method for degrading bound estradiol and / or free estradiol in sewage based on the double-embedded particles, comprising the following steps:

[0058] The double-embedded particles are mixed with the sewage to be treated, and the light culture program is run: the light intensity is 60 to 240 μmol·s -1 ·m -2 , the temperature is 20-25℃, and the photoperiod is 6-18 / day.

[0059] In the present invention, the illumination culture procedure is preferably: the illumination intensity is 120 to 220 μmol·s -1 ·m -2 , which can be 150 μmol·s -1 ·m -2 , 180 μmol·s -1 ·m -2 , 200 μmol·s -1 ·m -2 , 220 μmol·s -1 ·m -2 The temperature can be 21°C, 22°C, 23°C, 24°C.

[0060] In the present invention, the drainage ratio during each light cycle is preferably 50% to 55%, and can be 52% or 54%. The hydraulic retention time is preferably 4 to 6 hours, and can be 5 hours. The operation cycle of the light culture preferably includes water inlet, standing, aeration, sedimentation and drainage in sequence. The water inlet time is preferably 3 to 5 minutes, and can be 4 minutes. The standing time is preferably 5 to 10 minutes, and can be 6 minutes, 7 minutes, or 8 minutes. The aeration time is preferably 205 to 220 minutes, and can be 205 minutes, 210 minutes, 215 minutes, or 220 minutes. The aeration volume is preferably 0.5 to 1 L / min, and more preferably 0.5 L / min. The sedimentation time is preferably 10 to 15 minutes, and can be 12 minutes, 13 minutes, or 14 minutes. The drainage time is preferably 3 to 5 minutes, and can be 4 minutes.

[0061] In one embodiment of the present invention, the use of double-embedded particles has a strong removal effect on E2-3S and EE2, especially for conjugated estrogen (E2-3S).

[0062] The double-embedded particles for sewage treatment provided by the present invention and their application in removing mixed estrogens in sewage are described in detail below in conjunction with the examples, but they should not be construed as limiting the scope of protection of the present invention.

[0063] Example 1

[0064] Preparation of double-embedded particles for sewage treatment and method for removing mixed estrogens in sewage

[0065] In this embodiment, the artificial photosynthetic particles encapsulating NH2-MIL-53 (Fe) and white rot fungi are recorded as particles 1, and the natural photosynthetic particles encapsulating Bacillus subtilis and green algae are recorded as particles 2. Particles 1 and 2 are coupled to form double-encapsulated particles to treat sewage, wherein the sewage is a simulated secondary influent of a sewage treatment plant, which contains estradiol-3-sulfate (E2-3S) and 17α-ethinyl estradiol (EE2). The specific preparation method and treatment process include the following steps:

[0066] (1) Preparation of artificial photosynthetic particles:

[0067] (I) Preparation of NH2-MIL-53(Fe): 1.08 g FeCl3·6H2O, 0.73 g NH2-H2BDC and 50 mL DMF were placed in a 250 mL conical flask and ultrasonicated at 100 W for 5 min to promote dissolution. The obtained solution was sealed in a 100 mL polytetrafluoroethylene reactor and hydrothermally reacted at 170°C for 24 h. After cooling to room temperature, the solution was washed alternately with DMF and anhydrous ethanol and centrifuged three times. The solution was centrifuged at 4000 rpm for 5 min. The material was collected and dried at 80°C for 6 h to obtain the photocatalyst NH2-MIL-53(Fe).

[0068] (II) Method for expanding the culture of white rot fungi: In a clean bench, a certain amount of fungal cells were picked from the slant test tube of white rot fungi with an inoculation loop and inoculated into 100 mL of liquid comprehensive potato medium with a pH of 6, and cultured at 30°C until the bacterial solution OD 600 The liquid comprehensive potato culture medium includes 1L of 20% potato juice, 20g of glucose, 3g of KH2PO4, 1.5g of MgSO4·7H2O and 1g of thiamine.

[0069] (III) Dissolution of the embedding agent: 4 g of anhydrous CaCl2 was added to 100 mL of ultrapure water, and the pH was adjusted to 7 to obtain a CaCl2 solution; 2 g of sodium alginate (SA) solid powder was added to 100 mL of water under ultrasonic vibration, and the solution was assisted in dissolution in a water bath at 50-60°C, and the pH was adjusted to 7 to obtain a sodium alginate solution; the CaCl2 solution and the sodium alginate solution were placed in an autoclave and sterilized at 121°C for 30 min.

[0070] (IV) Particle embedding: After room temperature, weigh 0.05g of photocatalyst NH2-MIL-53(Fe) for use. Take 50mL of white rot fungus culture and centrifuge at 6000rpm for 10min to harvest the fungus, and set aside. Mix the weighed photocatalyst NH2-MIL-53(Fe) and the harvested fungus with sodium alginate solution, and use a 5mL syringe to drop into 50mL of CaCl2 solution, fix the reaction for 30min, and obtain about 300 artificial photosynthetic particles.

[0071] (2) Preparation of natural photosynthetic particles:

[0072] (I) Enrichment culture of bacteria and algae: Inoculate green algae (Chlorella vulgaris) into BG11 medium and culture at 25°C and 2000 Lux for 2 weeks. Take 50 mL of algae solution and centrifuge at 4000 rpm for 10 min to collect the green algae. The above operations were performed in a clean bench.

[0073] Bacillus subtilis was inoculated into nutrient broth medium (5.0 g peptone, 3.0 g beef extract, 5.0 g NaCl and 1 L distilled water, pH 7.0-7.2) and cultured at 28°C and 120 rpm until the OD of the bacterial solution reached 0. 600 The concentration of the bacterial solution was 0.8-1.0, and 50 mL of the bacterial solution was centrifuged at 6000 rpm for 10 min to collect the bacteria.

[0074] (II) Preparation of embedding agent is the same as the dissolution of embedding agent in step (III) of step (1).

[0075] (III) The green algae and bacterial cells collected in step (I) were suspended in 5 mL of ultrapure water, and then evenly mixed with 50 mL of sodium alginate solution to obtain a bacteria-algae mixed solution. The bacteria-algae mixed solution was dripped dropwise into 50 mL of CaCl2 solution using a 5 mL syringe (equipped with a needle), and the reaction was fixed for 30 minutes to obtain about 300 natural photosynthetic particles.

[0076] (3) Wastewater treatment method using double-encapsulated photosynthetic particles:

[0077] (I) Artificial photosynthetic particles and natural photosynthetic particles were added to a 1 L photo-sequential batch bioreactor (PSBR) at a ratio of 1:1. The working concentration of artificial photosynthetic particles was about 300 particles / L, and the concentration of natural photosynthetic particles was about 300 particles / L. The conditions for light culture were set as follows: pH value was 7-8, aeration intensity was 0.5 mL / min, photoperiod was 12 h / day, and light intensity was 120 μmol·s -1 ·m -2 .

[0078] (II) The operation cycle of the photo-sequential batch bioreactor is 4 hours, and each cycle includes 3 minutes of water inlet, 5 minutes of static, 219 minutes of aeration, 10 minutes of sedimentation and 3 minutes of drainage. The height-to-diameter ratio of the sequencing batch bioreactor is 30:8, and the water inlet, drainage and aeration of the sequencing batch bioreactor are all automatically operated by an automatic controller connected to a peristaltic pump, a solenoid valve and an aeration pump, and the aeration device is arranged at the bottom of the sequencing batch bioreactor to provide sufficient oxygen.

[0079] (III) The wastewater added to the photo-sequential batch bioreactor is artificial simulated wastewater, with a COD concentration of 200 mg / L and ammonium nitrogen (NH4 + -N) concentration is 40mg / L, phosphate (PO4 3- -P) concentration is 3.5mg / L, estradiol-3-sulfate (E2-3S) concentration is 2μg / L, and 17α-ethinyl estradiol (EE2) concentration is 2μg / L. The contents of E2-3S and EE2 in the effluent are quantitatively detected by ultra-high performance liquid chromatography tandem mass spectrometry, and the specific steps are as follows:

[0080] 1. Pretreatment: Samples are pretreated;

[0081] 2. Connect the chromatographic column: Select BEH C18 1.7μm, 2.1×50mm;

[0082] 3. Flushing the chromatographic column: Before using the machine, flush the chromatographic column with mobile phase and organic phase for 5 to 10 minutes;

[0083] 4. Edit the instrument parameters; set the mobile phase to ultrapure water, the organic phase to methanol, the injection flow rate to 0.25mL / min, the injection volume to 20μL, and the running time to 6min;

[0084] 5. Analysis: "Single Analysis" or "Quick Batch Processing" in the software, input sample bottles and bottle racks;

[0085] 6. Analyze data: Use the Lab-solution main project "Processing Tools", "Reanalysis", open the analysis results and view them;

[0086] 7. Finally, flush the column with 100% methanol for 30 min;

[0087] 8. Shut down the computer and turn off “LC” and “MC” above the software.

[0088] Yeast cell bioluminescence method was used to determine the estrogen effect, wherein the estrogen treatment steps are:

[0089] 1. Use HLB solid phase extraction column, activate with 3ml of ethyl acetate, 3ml of methanol and 6ml of ultrapure water, the flow rate is 1-3 drops / s;

[0090] 2. Drop the sample into the activated HLB column at a flow rate of 1 to 2 drops / s;

[0091] 3. After loading, the sample is rinsed with 9 ml of ultrapure water to remove salt and impurities;

[0092] 4. Use a suction pump to drain until the filler is in powder form;

[0093] 5. Elute into a pear-shaped bottle using 6 ml of ethyl acetate and methanol respectively;

[0094] 6. Raise the temperature of the nitrogen blower to 40°C, adjust the needle to a small size and insert it 1 cm into the liquid surface, blow until the sample becomes moist, and dilute it to a chromatographic bottle with methanol to obtain the methanol extract of the sample.

[0095] The test method for estrogenic effects is as follows:

[0096] 1. Take 500 μl of yeast cells and inoculate them into 100 ml of yeast basic culture medium without leucine, uracil and tryptophan;

[0097] 2. Set the conditions to incubate at 28°C and 200 rpm, and shake until OD 600 0.6~0.8;

[0098] 3. Transfer 20ul of estradiol (E2) standard to an opaque 96-well plate for yeast estrogen detection;

[0099] 4. Add 20 μl of methanol to three separate wells as blank controls;

[0100] 5. Dilute 20 μl of the sample methanol extract obtained above and add the dilution into an opaque 96-well plate;

[0101] 6. Add methanol to evaporate until no visible wet spots remain. Add 200 μl of yeast culture to the 96-well plate of E2 for detecting estrogen effects;

[0102] 7. Cover the top with tin foil, incubate at 28°C for 8 hours, and use an ELISA reader to detect the luminescence results. The conditions of the ELISA reader are as follows:

[0103] The bioluminescence was detected at 28°C every 1 h with an integration time of 1 s / well for 8 h, and the bioluminescence was calculated according to Formula I.

[0104] Bioluminescence value = minimum value + (maximum value - minimum value) / [1 + (concentration / EC 50)]Formula I.

[0105] Comparative Example 1

[0106] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that: 0.05g MIL-53 (Fe) material is added to a sequencing batch bioreactor (PSBR) instead of artificial photosynthetic particles and natural photosynthetic particles, and the conditions of light culture are set: pH value is 7-8, aeration intensity is 0.5mL / min, light cycle is 12h / day, light intensity is 120μmol·s -1 ·m -2 .

[0107] Among them, the preparation method of MIL-53 (Fe) is as follows: 0.83g of terephthalic acid is added to a beaker containing 28mL of N-N dimethylformamide (DMF), and after it is completely dissolved under magnetic stirring, an equal molar amount of FeCl3·6H2O (1.35g) is added. After the mixture is stirred at room temperature for 30 minutes, the solution is transferred to a 100mL polytetrafluoroethylene-lined autoclave and reacted at 170°C for 24 hours. Finally, after the autoclave is naturally cooled to room temperature, it is alternately washed and centrifuged three times with anhydrous ethanol and N-N dimethylformamide, and the material is collected and dried at 80°C for 6 hours to obtain the photocatalyst MIL-53 (Fe) material.

[0108] Comparative Example 2

[0109] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that 0.05g NH2-MIL-53 (Fe) is added to a 1L sequencing batch bioreactor (PSBR) instead of artificial photosynthetic particles and natural photosynthetic particles, and the conditions of light culture are set: pH value is 7-8, aeration intensity is 0.5mL / min, light cycle is 12h / day, light intensity is 120μmol·s -1 ·m -2 .

[0110] Comparative Example 3

[0111] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the addition of artificial photosynthetic particles is omitted, only about 300 natural photosynthetic particles are added to a 1L sequencing batch bioreactor (PSBR), and the conditions for light culture are set: pH value is 7-8, aeration intensity is 0.5mL / min, light cycle is 12h / day, and light intensity is 120μmol·s -1 ·m -2 .

[0112] Example 2

[0113] The degradation results of mixed estrogens in sewage of Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 3 were tested and analyzed: 100 mL of water sample was filtered with a 0.45 μm glass fiber filter membrane (water system), and after filtration, it was stored in a 4°C refrigerator for subsequent solid phase extraction operation and subsequent nitrogen blowing, and finally the estrogen concentration and effect were determined. Each experiment was repeated three times, and the average value ± standard deviation was taken for analysis. The E2-3S removal rate or EE2 removal rate was calculated according to formula I.

[0114] E2-3S removal rate or EE2 removal rate = (1-C / C0) × 100% Formula I.

[0115] Where C0 is the initial concentration and C is the concentration after the reaction.

[0116] As can be seen from Table 1, the removal effect of the double-embedded particles on E2-3S and EE2 is higher than that of NH2-MIL-53(Fe) (Comparative Example 2) and natural photosynthetic particles (Comparative Example 3), which are 13.9% and 5.8% and 9.2% and 4.4% respectively. Using the yeast cell luminescence method, E2 is used as the positive standard substance in the estrogen-like test. -13 M~10 -7 The E2 of M was used to obtain the estrogenic effect reduction value. For the estrogenic effect reduction effect, the double-embedded particles were significantly better than the NH2-MIL-53(Fe)MOF material and the natural photosynthetic particles, which were 98.2% and 89.3% respectively.

[0117] The results show that the treatment method of NH2-MIL-53(Fe)MOF material and bacteria-algae symbiotic coupled double-embedded particles has a strong removal effect on E2-3S and EE2, especially for bound estrogen (E2-3S), but NH2-MIL-53(Fe)MOF material does not have a stronger reduction ability for estrogen effect. From the analysis of the results, it can be seen that the method of the present invention can effectively remove bound estrogen and free estrogen in sewage, while reducing the concentration of estrogen active substances and ecological risk level in the effluent.

[0118] Table 1. Removal efficiency and estrogen reduction efficiency of E2-3S / EE2 in different modes

[0119] Coupling type Example 1 Comparative Example 1 Comparative Example 2 Comparative Example 3 E2-3S removal rate 98.6％ 78.2％ 84.7％ 92.8％ EE2 removal rate 83.3％ 66.3％ 74.1％ 78.9％ Total Effect Reduction Value (EEQ) 11.1ng / L 508.6ng / L 618.4ng / L 104.6ng / L

[0120] Comparative Example 4

[0121] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the green algae in the natural photosynthetic particles in step (2) are replaced by blue algae (Anabaena).

[0122] Comparative Example 5

[0123] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the Bacillus subtilis in the natural photosynthetic particles in step (2) is replaced by round brown nitrogen-fixing bacteria. The brown spherical nitrogen-fixing bacteria uses a nitrogen-fixing medium, wherein the composition includes 0.5g yeast extract, 20.0g mannitol, 0.2g KH2PO4, 0.8g K2HPO4, 0.2g MgSO4.7H2O, 0.1g CaSO4·2H2O, 0.036g FeCl3 trace, 0.084g Na2MoO4·2H2O, and 1.0L distilled water.

[0124] The degradation effects of Comparative Examples 4 and 5 were tested and analyzed according to the method in Example 1. The results are shown in Table 2.

[0125] As can be seen from Table 2, when the microalgae is replaced by Anabaena and the bacteria is replaced by round brown nitrogen-fixing bacteria, the removal rates of E2-3S and EE2 are significantly lower than the effects of natural photosynthetic particles formed by Chlorella and Bacillus subtilis, with values ​​of 6.3% and 3.0% respectively. At the same time, there is a significant difference in the reduction of the effect. When the microalgae is Anabaena, its effect is about 58ng / L higher than that of Chlorella. This may be because cyanobacteria release algae toxins under the influence of estrogen. When the bacteria is round brown nitrogen-fixing bacteria, its effect is about 3ng / L higher than that of Bacillus subtilis. This may be because round brown nitrogen-fixing bacteria is a functional bacterium for denitrification. From the analysis of the results, it can be seen that the combination of microalgae as Chlorella and bacteria as Bacillus subtilis set in the present invention has a good sewage treatment effect.

[0126] Table 2. Removal efficiency and estrogen reduction efficiency of E2-3S / EE2 in different modes

[0127] Coupling type Example 1 Comparative Example 4 Comparative Example 5 E2-3S removal rate 98.6％ 92.3％ 95.6％ EE2 removal rate 83.3％ 69.7％ 84.2％ Total Effect Reduction (EEQ) 11.1ng / L 69.1ng / L 14.1ng / L

[0128] Example 3

[0129] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the light cycle time in step (3) is 18h / day.

[0130] Example 4

[0131] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the light cycle time in step (3) is 6h / day.

[0132] The degradation effects of Example 3 and Example 4 were detected and analyzed according to the method described in Example 1. The results are shown in Table 3.

[0133] As shown in Table 3, when the photoperiod is 18h / day, both E2-3S and EE2 have a high removal rate, which are 98.8% and 88.1% respectively, and the total estrogen reduction effect is only 4.6ng / L. When the photoperiod is 6h, the removal rates of E2-3S and EE2 decrease slightly, which are 94.3% and 78.6% respectively, and the total estrogen reduction effect is only 29.8ng / L. From the analysis of the results, it can be seen that the longer the photoperiod is, the more conducive it is to the removal of estrogen, so the photoperiod set by the present invention is 6 to 18h / day, which is reasonable.

[0134] Table 3. Removal efficiency and estrogen reduction efficiency of E2-3S / EE2 in different modes

[0135] Coupling type Example 1 Example 3 Example 4 E2-3S removal rate 98.6％ 98.8％ 94.3％ EE2 removal rate 83.3％ 88.1％ 78.6％ Total Effect Reduction (EEQ) 11.1ng / L 4.6ng / L 29.8ng / L

[0136] Example 5

[0137] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the light intensity in step (3) is 240 μmol m -2 s -1 .

[0138] Example 6

[0139] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the light intensity in step (3) is 60 μmol m -2 s -1 .

[0140] The degradation effects of Example 5 and Example 6 were detected and analyzed according to the method described in Example 1. The results are shown in Table 4.

[0141] As shown in Table 4, the light intensity is 240 μmol m -2 s -1 When the light intensity was 60 μmol m -2 s -1 When the light intensity was increased from 60 μmol m -2 s -1 Increased to 240 μmol m -2 s -1When the light intensity is set to 60 μmol m -2 s -1 ~240 μmol m -2 s -1 It is reasonable.

[0142] Table 4. Removal efficiency and estrogen reduction efficiency of E2-3S / EE2 in different modes

[0143] Coupling type Example 1 Example 5 Example 6 E2-3S removal rate 98.6％ 99.0％ 90.2％ EE2 removal rate 83.3％ 87.4％ 66.4％ Total Effect Reduction (EEQ) 11.1ng / L 3.9ng / L 19.7ng / L

[0144] Example 7

[0145] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, differs in that the concentration of artificial photosynthetic particles in step 3 is about 600 particles / L, and the concentration of natural photosynthetic particles is 600 particles / L.

[0146] Example 8

[0147] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, the difference is that the concentration of artificial photosynthetic particles in step 3 is about 100 particles / L and the concentration of natural photosynthetic particles is 300 particles / L.

[0148] Example 9

[0149] A method for preparing double-embedded particles for sewage treatment and removing mixed estrogens in sewage, compared with Example 1, differs in that the concentration of artificial photosynthetic particles in step 3 is about 300 particles / L, and the concentration of natural photosynthetic particles is 100 particles / L.

[0150] The degradation effects of Example 7 and Comparative Examples 6 and 7 were detected and analyzed according to the method described in Example 1. The results are shown in Table 5.

[0151] It can be seen from Table 5 that when the volume ratio of artificial photosynthetic particles to natural photosynthetic particles is 1:1, and the working concentration is increased by 1 times, the removal rate and effect reduction of E2-3S and EE2 are enhanced, and the removal rates are 99.5% and 89.1%, respectively, and the effect value is 0.9ng / L. When the volume ratio of artificial photosynthetic particles to natural photosynthetic particles is 1:3, there is little change in E2-3S and effect reduction, but a greater impact on the removal rate of EE2, with an E2-3S removal rate of 94.32%, an EE2 removal rate of 68.2%, and a total effect reduction of 10.3ng / L. When the volume ratio of artificial photosynthetic particles to natural photosynthetic particles is 3:1, there is little effect on the removal rates of E2-3S and EE2, with removal rates of 95.4% and 86.6%, respectively, and a total effect reduction of 23.8ng / L. From the analysis of the results, it can be seen that the volume ratio of artificial photosynthetic particles to natural photosynthetic particles set by the present invention (1~3): (1-3) is reasonable.

[0152] Table 5. Removal efficiency and estrogen reduction efficiency of E2-3S / EE2 in different modes

[0153]

[0154]

[0155] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A double-embedded particle, characterized in that: Including artificial photosynthetic particles and natural photosynthetic particles; The artificial photosynthetic particles include calcium alginate hydrogel, a photocatalyst and white rot fungi embedded in the calcium alginate hydrogel; the photocatalyst includes an iron-based metal organic framework NH2-MIL-53 (Fe); The natural photosynthetic particles include calcium alginate hydrogel and green algae and bacillus subtilis embedded in the calcium alginate hydrogel.

2. The double-embedded particle according to claim 1, characterized in that: The concentration of the photocatalyst is 0.1 to 0.5 mg / mL; The concentration of the white rot fungus was 3.2×10 8 CFU / mL~4×10 8 CFU / mL.

3. The double-embedded particle according to claim 1, characterized in that: The green algae includes Chlorella vulgaris; the concentration of the green algae is 1×10 9 cells / L~2×10 9 cells / L; The concentration of Bacillus subtilis is 0.6×10 9 CFU / L~0.8×10 9 CFU / L.

4. The double-embedded particle according to claim 1, characterized in that: The quantity ratio of the artificial photosynthetic particles to the natural photosynthetic particles is (1-6):(1-6).

5. The double-embedded particle according to any one of claims 1 to 4, characterized in that: The preparation method of NH2-MIL-53(Fe) comprises dissolving ferric chloride and 2-aminoterephthalic acid in an organic solvent, performing hydrothermal synthesis at 150-170° C. for 18-24 hours under sealed conditions, washing with an organic solvent after cooling, removing the solvent, and obtaining NH2-MIL-53(Fe).

6. A method for preparing the double-embedded particles according to any one of claims 1 to 5, characterized in that: The following steps are involved: collecting white rot fungus mycelium and / or spores from the cultured white rot fungus liquid, dispersing the white rot fungus mycelium and / or spores and NH2-MIL-53(Fe) in a sodium alginate solution to obtain a material-fungus mixed liquid; Dropping the material-fungus mixture into a calcium chloride solution, and undergoing a solidification reaction, obtaining artificial photosynthetic particles; Separating green algae bodies from the cultured green algae liquid, separating Bacillus subtilis cells from the cultured Bacillus subtilis liquid, and dispersing the green algae bodies and Bacillus subtilis cells in a sodium alginate solution to obtain a green algae-bacteria mixed solution; The green algae-bacteria mixed solution is dripped into a calcium chloride solution to obtain natural photosynthetic particles through a solidification reaction; The artificial photosynthetic particles and the natural photosynthetic particles are mixed to obtain double-embedded particles.

7. Use of the double-embedded particles according to any one of claims 1 to 5 or the double-embedded particles prepared by the preparation method according to claim 6 in degrading bound estradiol and / or free estradiol in wastewater.

8. A method for degrading bound estradiol and / or free estradiol in wastewater based on the double-embedded particles according to any one of claims 1 to 5, characterized in that: The following steps are involved: The double-embedded particles are mixed with the sewage to be treated, and the light culture program is run: the light intensity is 60 to 240 μmol·s -1 ·m -2 , the temperature is 20-25℃, and the photoperiod is 6-18h / day.

9. The method according to claim 8, characterized in that: The drainage ratio during each photoperiod is 50% to 55%, and the hydraulic retention time is 4 to 6 hours.

10. The method according to claim 8, characterized in that: The operation cycle of the light culture includes water intake, static standing, aeration, sedimentation and drainage in sequence; The water inlet time is 3 to 5 minutes; The standing time is 5 to 10 minutes; The aeration time is 205-220 min; the aeration volume is 0.5-1 L / min; The precipitation time is 10 to 15 minutes; The drainage time is 3 to 5 minutes.

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