Use of porous support materials in microbial immobilization

Abstract

The application discloses application of a porous carrier material in microbial immobilization and belongs to the technical field of microbial immobilization. a The porous carrier material has a positive surface (pK a 8) and moderate hydrophilicity (contact angle of 20-60 degrees), is used for immobilization of microorganisms, especially white rot fungi, and obtains a porous carrier-microorganism composite material, which realizes efficient organic matter degradation and stable activity of the immobilized white rot fungi in actual wastewater without pH adjustment, solves the technical problem that the white rot fungi can only be used under acidic conditions in the prior art, and provides a new idea for application of functional microorganisms and degradation of emerging pollutants.

Description

Technical Field

[0001] This application belongs to the field of microbial immobilization technology, specifically relating to the application of porous carrier materials in microbial immobilization. Background Technology

[0002] Emerging pollutants generally possess complex structures such as highly branched chains, polycyclic structures, and electron-withdrawing functional groups, making them difficult to degrade. Furthermore, they are widely distributed and remain a risk even at environmental concentrations. Wastewater containing emerging pollutants is characterized by large volumes, low concentrations, and high risks. Conventional wastewater treatment facilities cannot remove these recalcitrant compounds, thus necessitating cost-effective methods for their reduction and removal.

[0003] Therefore, a large number of functional microorganisms with special degradation capabilities have been extensively studied, such as Bacillus thuringiensis B1 and Klebsiella sp. KSC. These functional microorganisms can directly metabolize specific PPCPs as the sole carbon source. Besides degrading bacteria, white-rot fungi are a class of microorganisms capable of degrading lignin, possessing a degradation enzyme system represented by laccase, and are currently the only microorganisms capable of completely degrading dozens of PPCPs. Among them, Trametes versicolor (T. versicolor) has been extensively studied due to its high degradation efficiency.

[0004] However, these functional microorganisms still suffer from insufficient degradation efficiency and difficulty in surviving in actual wastewater. For example, degrading bacteria can generally only remove certain types of fixed-structure compounds and have poor removal effects on mixed pollutants. White-rot fungi face the problems of slow degradation rate and bacterial competition in actual wastewater. Therefore, almost all studies have adjusted the wastewater to a pH of 4.5, which is favorable for fungal growth.

[0005] Microbial (carrier) immobilization is a method of fixing microbial cells onto a carrier using physical and chemical means. Based on the immobilization mechanism, it can be divided into adsorption, covalent bonding, encapsulation, and other methods. This technology offers advantages such as providing a safe and stable growth environment, high mechanical strength, and barrier against toxic substances. Commonly used commercial carriers include biochar, polyethylene, and polyurethane, which have been widely applied in wastewater treatment. For example, biofilms achieved on carriers offer advantages such as large biomass and long sludge age. Polyurethane has also been used for the immobilization of functional microorganisms, and biofilms of degrading bacteria and fungi have been successfully implemented. However, these carrier materials are generally non-porous or have a low specific surface area (<100 m²). 2While existing immobilization carriers (e.g., *[g]*) offer little adsorption for emerging pollutants, they cannot further enhance the degradation efficiency of functional microorganisms. Furthermore, they suffer from biomass erosion; the binding between immobilized functional microorganisms and these carriers is not tight, leading to contamination in the environmental media. Overall, existing immobilization carriers suffer from low removal efficiency and biomass erosion, making it difficult to apply immobilized functional microorganisms in practical wastewater treatment. Further research is needed on novel carrier materials for microbial immobilization. Summary of the Invention

[0006] 1. The problem to be solved

[0007] This invention addresses the problems of low removal efficiency, difficulty in survival in actual wastewater, or competition from environmental microorganisms in existing functional microorganisms, especially white-rot fungi. It provides a porous carrier material for microbial immobilization, wherein the surface of this porous carrier material carries a positive charge (pK). a >8) It is moderately hydrophilic (contact angle is 20°~60°) and can be used for the immobilization of microorganisms, especially white-rot fungi, to obtain a porous carrier-microorganism composite material. When this composite material is used for wastewater treatment, it can simultaneously improve the organic matter degradation effect and biological activity of functional microorganisms.

[0008] 2. Technical Solution

[0009] To solve the above problems, the technical solution adopted in this application is as follows:

[0010] Firstly, this application provides a porous support material whose BET specific surface area, measured by the nitrogen isotherm method, is >600 m². 2 g -1 pK a >8 and the contact angle is 20° to 60°.

[0011] Furthermore, specific surface area > 650 m² 2 g -1 .

[0012] Furthermore, specific surface area >700m² 2 g -1 .

[0013] Furthermore, pK a >9.

[0014] Furthermore, the contact angle is 30° to 50°.

[0015] Furthermore, the contact angle is 30° to 40°.

[0016] Furthermore, the aforementioned porous carrier material includes any one of porous thiophene-based cyclodextrin polymers, porous thiophene-based chitosan polymers, and porous thiophene-based columnar aromatic polymers. Microbial immobilization involves complex processes such as carrier adsorption, microbial degradation, and enzyme degradation. It is necessary to avoid using carriers that are harmful to microorganisms. Chitosan, cyclodextrin, and the like are all bio-friendly materials and have a large number of functional groups on their surfaces. They can be surface modified to achieve immobilization for specific microorganisms.

[0017] Furthermore, the aforementioned porous support material includes a porous thiophene-based cyclodextrin polymer; cyclodextrin polymers have the advantage of fast adsorption rate and can combine with microbial degradation to further improve degradation performance.

[0018] Furthermore, the aforementioned cyclodextrin is any one of α, β, or γ cyclodextrin.

[0019] Secondly, this application provides a method for preparing the aforementioned porous support material. This method uses cyclodextrin, chitosan, or columnar aromatic hydrocarbons as monomers, grafts thiophene rings to form thiophene monomers, and then uses an alkylating agent containing an aromatic ring or methoxy group as a crosslinking agent to carry out a Friedel-Crafts alkylation reaction under the catalysis of a Lewis acid ionic liquid catalyst. Under the catalysis of a Lewis acid, the alkylating agent forms an alkyl carbocation, which then undergoes an electrophilic reaction with an electrophilic compound. Since this reaction can be continuously substituted, a porous material with a high degree of polymerization can be obtained.

[0020] Furthermore, the aforementioned cyclodextrin is any one of α, β, or γ cyclodextrin.

[0021] Furthermore, the above-mentioned grafted thiophene ring comprises: replacing the hydroxyl group of the monomer with sodium alkoxide using anhydrous NaH, and then adding a halothiophene ring for grafting.

[0022] Furthermore, the aforementioned halothiophene ring includes thiophene bromide or thiophene chloride.

[0023] Furthermore, the Lewis acid ionic liquid catalysts mentioned above include: imidazole-type or quaternary ammonium-type ionic liquids, and composite ionic liquids formed with metal chlorides.

[0024] Furthermore, the aforementioned metal chlorides include ferric chloride and / or aluminum chloride.

[0025] Furthermore, the above-mentioned ionic liquid is a 1-ethyl-3-methylimidazolium chloride ionic liquid.

[0026] Further, the crosslinking agent includes any one or more of biphenyl dichlorobenzyl, p-dichlorobenzyl, biphenyl dibromobenzyl, p-dibromobenzyl, dimethoxymethane, trimethyl orthoformate, and 1,4-dimethoxybenzene. Even further, the crosslinking agent includes trimethyl orthoformate.

[0027] Furthermore, the reaction temperature conditions for the above-mentioned Friedel-Crafts alkylation reaction include: reacting at 45°C for 4–6 h, followed by heating to 80–150°C for 18–20 h. Even further, the reaction temperature conditions for the above-mentioned Friedel-Crafts alkylation reaction include: reacting at 45°C for 5 h, followed by heating to 80–150°C for 19 h. If the reaction is not within the above range, the synthesized polymer will exhibit incomplete reaction, lower specific surface area, and lower yield.

[0028] Furthermore, the above reaction temperature conditions include: reacting at 45°C for 5 hours, and then heating to 120°C for 19 hours.

[0029] Furthermore, the aforementioned porous support material is a porous thiophene-based cyclodextrin polymer, and its preparation method includes the following steps:

[0030] Cyclodextrin was dissolved in anhydrous DMF; then anhydrous NaH was slowly added at 0°C and stirred for 15 minutes; 3-bromomethylthiophene (3-Bm-Th) was slowly added dropwise, and then stirred at room temperature for 24 hours; after the reaction was completed, the reaction was quenched with methanol; then the product was mixed with water and concentrated to near dryness under vacuum at 84°C; the obtained product was mixed with water and extracted with dichloromethane; the organic phase was dried with anhydrous sodium sulfate, then filtered and concentrated under vacuum at 35°C, and finally obtained by silica gel chromatography as a light yellow compound thiophene cyclodextrin (ThCD);

[0031] At 0 °C, ThCD was dissolved in 1-ethyl-3-methylimidazolium chloride / ferric chloride ([EMIm]Cl / FeCl3) ionic liquid; under nitrogen protection, anhydrous trimethyl orthoformate (Tmof) was slowly added dropwise at room temperature, the reaction mixture was stirred and heated to 45 °C for 5 h, and then heated to 120 °C for 19 h; after cooling, the brown precipitate was washed with water and methanol until the filtrate was colorless, then extracted with methanol by Soxhlet for 24 h, and dried under vacuum at 60 °C for 24 h to obtain porous thiophene cyclodextrin polymer (Th-CDP).

[0032] Furthermore, the molar ratio of 3-bromomethylthiophene to cyclodextrin is 14 to 21:1. By adjusting the ratio of thiophene ring grafted onto cyclodextrin, some hydroxyl groups on the cyclodextrin can be selectively retained, thereby controlling the hydrophilicity and hydrophobicity of the carrier surface. β-cyclodextrin has a total of 21 hydroxyl groups. When the ratio of grafted thiophene to cyclodextrin is 14:1 and 21:1, the contact angle of the prepared carrier surface increases, and the carrier surface changes from superhydrophilic to moderately hydrophilic, thus having a good ability to maintain the activity of specific functional microorganisms.

[0033] Furthermore, the molar ratio of the alkylating agent to the thiophene cyclodextrin is 5–100:1.

[0034] Thirdly, this application provides the application of the aforementioned porous carrier material in microbial immobilization, wherein the microorganisms include one or more of white-rot fungi, nitrifying bacteria, denitrifying bacteria, Bacillus, Pseudomonas, and activated sludge; the BET specific surface area of ​​the porous carrier material, measured by the nitrogen isotherm method, is >600 m². 2 g -1 pK a >8 and the contact angle is 20° to 60°.

[0035] Furthermore, the aforementioned microorganisms are white-rot fungi.

[0036] Furthermore, the aforementioned white-rot fungi are one or more of Phanerochaetechrysosporium, Trametesversicolor, Corilus versicolor, Pleurotus sajor-caju, and Pleurotus ostreatus.

[0037] Furthermore, the aforementioned white-rot fungus is *Trametes versicolor*.

[0038] Furthermore, the application of the above-mentioned porous carrier material in microbial immobilization involves first adding the porous carrier material to the culture medium, then adding (functional) microorganisms through mycelial inoculation, and finally immobilizing the microorganisms inside the porous carrier material through physical adsorption by shaking culture.

[0039] Furthermore, the application of the aforementioned porous carrier material in the immobilization of microorganisms, specifically white-rot fungi, includes:

[0040] Preparation of mycelial solution:

[0041] T. versicolor agar blocks were inoculated into potato dextrose (PD) medium and cultured with shaking (135 rpm) at 25°C for 4–5 days. The obtained mycelial balls were separated from the medium, the mycelial balls were collected and 0.85% sterile physiological saline was added, and the mixture was stirred with a high-speed homogenizer for 15 min to obtain mycelial solution, which was stored at 4°C.

[0042] Immobilized microorganisms:

[0043] The porous carrier material was added to PD medium and autoclaved. 1.5 mL of mycelial solution of white-rot fungus T. versicolor was inoculated into the medium and cultured at 25°C with shaking at 135 rpm for 5 days to prepare immobilized white-rot fungus.

[0044] Fourthly, this application provides a porous carrier-microbial composite material, wherein: the microorganisms include one or more of white-rot fungi, nitrifying bacteria, denitrifying bacteria, Bacillus, Pseudomonas, and activated sludge; and the BET specific surface area of ​​the porous carrier material, measured by the nitrogen isotherm method, is >600 m². 2 g -1 pK a >8 and the contact angle is 20° to 60°.

[0045] Furthermore, the aforementioned microorganisms are white-rot fungi.

[0046] Furthermore, the aforementioned white-rot fungi are one or more of Phanerochaetechrysosporium, Trametesversicolor, Corilus versicolor, Pleurotus sajor-caju, and Pleurotus ostreatus.

[0047] Furthermore, the aforementioned white-rot fungus is *Trametes versicolor*.

[0048] Fifthly, this application provides a method for preparing the aforementioned porous carrier-microorganism composite material. The method includes: adding the porous carrier material to a culture medium, then adding functional microorganisms by bacterial inoculation, and then culturing by shaking to fix the microorganisms into the porous carrier material through physical adsorption.

[0049] Furthermore, in the above-mentioned method for preparing a porous carrier-microorganism composite material, the microorganism is a white-rot fungus, specifically including:

[0050] Preparation of mycelial solution:

[0051] T. versicolor agar blocks were inoculated into PD medium and cultured with shaking (135 rpm) at 25°C for 4–5 days. The obtained mycelial balls were separated from the medium, the mycelial balls were collected and 0.85% sterile physiological saline was added, and the mixture was stirred with a high-speed homogenizer for 15 min to obtain mycelial solution, which was stored at 4°C.

[0052] Preparation of porous carrier-microorganism composite materials:

[0053] The porous carrier material was added to PD medium and autoclaved; the mycelial solution of white-rot fungus T. versicolor was inoculated into the medium and cultured at 25°C with shaking at 135 rpm for 5 days to prepare the porous carrier-microorganism composite material (immobilized white-rot fungus).

[0054] Sixthly, this application provides the application of the aforementioned porous carrier-microorganism composite material in wastewater treatment.

[0055] Furthermore, the microorganisms in the aforementioned porous carrier-microorganism composite material are white-rot fungi, which are used to degrade PPCPs in wastewater.

[0056] Furthermore, the aforementioned PPCPs include one or more of diclofenac (DCF), ibuprofen (IBU), sulfamethoxazole (SMX), and carbamazepine (CBZ).

[0057] 3. Beneficial effects

[0058] Compared with the prior art, the advantages of this application are as follows:

[0059] (1) The application of a porous carrier material in microbial immobilization provided in this application, wherein the porous carrier material has a BET specific surface area >600m² as measured by the nitrogen isotherm method. 2 g -1 pK a With a contact angle of >8° and 20°–60°, this study investigated the effects of porous structure, surface charge, and hydrophilicity / hydrophobicity on the growth, metabolism, and community function of immobilized microorganisms. It was found that a positively charged, moderately hydrophilic porous carrier material achieved efficient immobilization of the white-rot fungus *T. versicolor* and its laccase. This material was used to treat actual wastewater from a sewage treatment plant without pH adjustment, while simultaneously achieving efficient degradation of PPCPs and stable activity of *T. versicolor*. This provides new insights for the application of functional microorganisms and the degradation of emerging pollutants.

[0060] (2) The application of a porous carrier material provided in this application in microbial immobilization degrades emerging pollutants in wastewater under high concentration load conditions. Compared with unimmobilized T. versicolor and PUF, the porous cyclodextrin polymer-immobilized T. versicolor significantly improves the removal rate of PPCPs, with an average of 81.3%. Meanwhile, the material control reactors B-I (PBCD-BD) and B-II (Th-CDP) became saturated in the later stage of the reaction, and the effluent concentration increased significantly. However, the porous cyclodextrin-immobilized T. versicolor maintained a low effluent concentration throughout, especially Tv@Th-CDP, whose effluent concentration was only 6.2% of the material control after 18 days. The coupling of porous carrier and biodegradation extended the service life of the reactor.

[0061] (3) The application of a porous carrier material provided in this application in microbial immobilization, which degrades emerging pollutants in wastewater at environmental concentrations, has been found to contain high levels of laccase and bioactivity within the porous carrier. The degradation of PPCPs is related to the pK of the immobilized carrier. aPositive correlation: positively charged Tv@Th-CDP retains T. versicolor and its laccase through electrostatic interaction, forming an adsorption-biodegradation synergistic system. The average removal rate of PPCPs reached 90.3%, which is 17.2% higher than that of unfixed T. versicolor. The degradation efficiency of negatively charged Tv@PBCD-BD and Tv@GAC was affected.

[0062] (4) The application of a porous carrier material provided in this application in microbial immobilization was studied using metagenomic sequencing to investigate the effect of the hydrophilicity of the porous carrier on biological activity. After continuous treatment in the reactor for 10 days, the abundance of unimmobilized *T. versicolor* decreased by 97.1%, while the moderately hydrophilic *Tv@Th-CDP* still had 31.2% *T. versicolor* abundance, showing almost no loss, and also exhibited high cytochrome P450 functional abundance. Other biomaterials were negatively affected by *Proteobacteria*, and the superhydrophilic surface of *Tv@PBCD-BD* even caused bacterial overgrowth. Attached Figure Description

[0063] Figure 1 This is a schematic diagram of carrier materials immobilizing the white-rot fungus T. versicolor, including Tv@PBCD-BD, Tv@Th-CDP, Tv@GAC, and Tv@PUF.

[0064] Figure 2 These are scanning electron microscope images of immobilized white-rot fungus T. versicolor, including: (a) T. versicolor; (b) Tv@PBCD-BD; (c) Tv@Th-CDP; (d) Tv@GAC; (e) Tv@PUF.

[0065] Figure 3 This is a schematic diagram of the synergistic degradation of immobilized white-rot fungus Tv@Th-CDP in actual wastewater. The positively charged Th-CDP achieves simultaneous immobilization of T. versicolor and its laccase through electrostatic interaction.

[0066] Figure 4 The results show the removal of high concentrations of PPCPs by immobilized white-rot fungi reactors under rapid saturation conditions. The reactors used were: A1 (T. versicolor); C1 (Tv@PBCD-BD); D1 (Tv@Th-CDP); E1 (Tv@GAC); F1 (Tv@PUF); Diclofenac (DCF); Ibuprofen (IBU); Sulfamethoxazole (SMX); and Carbamazepine (CBZ).

[0067] Figure 5This data represents the results of removing environmental concentrations of PPCPs from a porous cyclodextrin polymer-immobilized white-rot fungus *T. versicolor* and its control material in a reactor. Influent1 represents the influent to reactors B-I, C1, and D1, and Influent2 represents the influent to reactor B-II. Reactors B-I (PBCD-BD), B-II (Th-CDP), C1 (Tv@PBCD-BD), and D1 (Tv@Th-CDP) were used. The reactors used contained diclofenac (DCF), ibuprofen (IBU), sulfamethoxazole (SMX), and carbamazepine (CBZ).

[0068] Figure 6 The results of removing PPCPs by immobilized white-rot fungus T. versicolor in reactors under environmental concentration equilibrium conditions were obtained from reactors A2 (T. versicolor), C2 (Tv@PBCD-BD), D2 (Tv@Th-CDP), E2 (Tv@GAC), and F2 (Tv@PUF); diclofenac (DCF); ibuprofen (IBU); sulfamethoxazole (SMX); and carbamazepine (CBZ).

[0069] Figure 7 The concentrations of adenosine triphosphate (ATP), laccase activity, and glucose in T. versicolor immobilized in the suspension (sus) and inside the material (ex) under environmental concentration equilibrium conditions are: (a) ATP concentration during the immobilization process; (b) ATP concentration in the reactor; (c) laccase activity; and (d) glucose concentration.

[0070] Figure 8 These are the parameters for the reactor suspension of the immobilized white-rot fungus *T. versicolor*. Environmental concentration equilibrium conditions: (a) laccase activity and adenosine triphosphate (ATP) concentration; (b) glucose concentration and pH. Rapid saturation pre-running conditions: (c) laccase activity and ATP concentration; (d) glucose concentration and pH. Reactor A (*T. versicolor*); Reactor BI (PBCD-BD); Reactor B-II (Th-CDP); Reactor C (*Tv@PBCD-BD*); Reactor D (*Tv@Th-CDP*); Reactor E (*Tv@GAC*); Reactor F (*Tv@PUF*).

[0071] Figure 9The species distribution in the immobilized white-rot fungus *T. versicolor* reactor is shown in the following conditions: (a) rapid saturation pre-operation; (b) environmental concentration equilibrium. Wastewater from the secondary sedimentation tank of a wastewater treatment plant (GT_WWTP); Reactor A (*T. versicolor*); Reactor C (*Tv@PBCD-BD*); Reactor D (*Tv@Th-CDP*); Reactor E (*Tv@GAC*); Reactor F (*Tv@PUF*). Suspension (s); *T. versicolor* particles (p).

[0072] Figure 10 The COG functional distribution at the levels of secondary metabolite biosynthesis, transport, and decomposition in the immobilized white-rot fungus *T. versicolor* reactors; (a) rapid saturation pre-operation conditions; (b) environmental concentration equilibrium conditions; wastewater treatment plant secondary sedimentation tank effluent (GT_WWTP); reactor A (*T. versicolor*); reactor C (*Tv@PBCD-BD*); reactor D (*Tv@Th-CDP*); reactor E (*Tv@GAC*); reactor F (*Tv@PUF*). Suspension (s); *T. versicolor* particles (p). Detailed Implementation

[0073] The present application will be further described below with reference to specific embodiments.

[0074] It should be noted that terms such as "upper", "lower", "left", "right", and "middle" used in this specification are only for clarity of description and are not intended to limit the scope of implementation. Changes or adjustments to their relative relationships, without substantially altering the technical content, should also be considered as within the scope of this application.

[0075] 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 pertains; the term “and / or” as used herein includes any and all combinations of one or more of the associated listed items.

[0076] Unless otherwise specified in the examples, the procedures should be performed under standard conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available products.

[0077] As used herein, the term “about” is used to provide for the flexibility and imprecision associated with a given term, measure, or value. Those skilled in the art can readily determine the degree of flexibility for a particular variable.

[0078] As used herein, the term “at least one of…” is intended to be synonymous with “one or more of…”. For example, “at least one of A, B, and C” explicitly includes only A, only B, only C, and combinations thereof.

[0079] Concentration, amount, and other numerical data may be presented in range format herein. It should be understood that such range format is used solely for convenience and brevity and should be flexibly interpreted to include not only the values ​​explicitly stated as the limits of the range, but also all individual values ​​or subranges encompassed within the range, as if each value and subrange were explicitly stated. For example, a range of values ​​from about 1 to about 4.5 should be interpreted to include not only the explicitly stated limits of 1 to 4.5, but also individual numbers (such as 2, 3, 4) and subranges (such as 1 to 3, 2 to 4, etc.). The same principle applies to ranges that describe only a single value, such as “less than about 4.5,” which should be interpreted to include all the aforementioned values ​​and ranges. Furthermore, this interpretation should apply regardless of the breadth of the range or characteristic described.

[0080] In this application, the instruments and methods used for characterizing and evaluating the performance of the composite materials are as follows:

[0081] Physicochemical adsorption instrument: ASAP 2460 (Micromeritics, USA), used to determine the specific surface area of ​​carrier materials by nitrogen adsorption.

[0082] Water contact angle measuring instrument: OCA20 (Dataphysics, Germany).

[0083] Zeta potentiometer: Zetasizer Nano ZS90 (Malvern, UK).

[0084] Scanning electron microscope: QUANTA 250FEG (FEI, USA). After washing with PBS, the samples were fixed with glutaraldehyde, then dehydrated and freeze-dried, and sputter-coated with gold for microscopic examination.

[0085] Liquid chromatography (LC) and liquid chromatography-mass spectrometry (LC-MS): Agilent 1260 (Agilent, USA), API4000 (AB SCIEX, USA), for the concentration detection of PPCPs.

[0086] UV-Vis spectrophotometer: Specord 210 (Analytik Jena, Germany), used DMP method to determine laccase activity and DNS method to detect glucose concentration.

[0087] Microplate reader: Synergy H1 (BioTek, USA), ATP concentration was detected using the BacTiter-Glo™ Microbial Cell Viability Assay Kit.

[0088] Metagenomic sequencing: used for species and functional analysis of microorganisms in reactors.

[0089] Example 1

[0090] This embodiment provides a porous carrier material and its preparation method.

[0091] In this embodiment, the porous support material is a porous thienylcyclodextrin polymer (Th-CDP), and its preparation method is referenced in the literature: Tu, Y.; Wang, H.; Yang, P.; Xu, G.; Hu, X.; Li, A.; Xie, XA porous thienylcyclodextrin polymer synthesized in a homogeneous ionic liquid catalytic system for the rapid removal of pharmaceuticals and personal care products (PPCPs). Green Chemistry, 2022, 24(1):227-237. Specifically, it includes:

[0092] Preparation of 1-ethyl-3-methylimidazolium chloride / ferric chloride ([EMIm]Cl / FeCl3) ionic liquid:

[0093] 29.92 g of [EMIm]Cl and 66.20 g of FeCl3 were dried in a vacuum at 60 °C for 24 h, and then immediately transferred to a glove box. The reaction was carried out in a ceramic bead cooling bath (pre-cooled). FeCl3 was slowly added to a beaker containing [EMIm]Cl and continuously stirred with a glass rod. The reaction was exothermic, and the two components melted into a liquid. During the stirring process, the temperature of the solution rose to about 20-40 °C. After stirring at room temperature for 24 h, the Lewis acid ionic liquid [EMIm]Cl / FeCl3 was obtained.

[0094] Synthesis of thiophene cyclodextrin (ThCD):

[0095] 2 g of β-cyclodextrin (β-CD) was dissolved in anhydrous N,N-dimethylformamide (DMF); then 2.22 g of 60% anhydrous NaH was slowly added at 0 °C and stirred for 15 min; 10.34 g of 3-bromomethylthiophene (3-Bm-Th) was slowly added dropwise, and then stirred at room temperature for 24 h; the reaction was quenched with 5 mL of methanol; then the product was mixed with 200 mL of water and concentrated to near dryness under vacuum at 84 °C; the obtained product was mixed with 100 mL of water and extracted three times with 150 mL of dichloromethane, 50 mL each time; the organic phase was dried with anhydrous sodium sulfate, then filtered and concentrated under vacuum at 35 °C, and finally purified by silica gel chromatography to obtain the pale yellow compound ThCD;

[0096] Synthesis of porous thiophene-based cyclodextrin polymer (Th-CDP):

[0097] At 0℃, 0.8 g of ThCD was dissolved in [EMIm]Cl / FeCl3; after the solution was replaced with N2 three times, 2.68 g of anhydrous trimethyl orthoformate (Tmof) was slowly added dropwise at room temperature under N2 protection. The reaction mixture was stirred and heated to 45℃ for 5 h, and then heated to 120℃ for 19 h. After cooling, the brown precipitate was washed with water and methanol until the filtrate was colorless. Then, it was extracted with methanol by Soxhlet extraction for 24 h and dried under vacuum at 60℃ for 24 h to obtain porous thiophene cyclodextrin polymer (Th-CDP).

[0098] Example 2

[0099] This embodiment provides the effect of different grafting reagents and catalyst systems on the surface charge of porous cyclodextrin polymers.

[0100] Grafting different functional groups onto cyclodextrins alters the surface properties of the resulting polymers. Polypyrrole, polythiophene, and polyfuran are commonly used conductive materials; therefore, different aromatic heterocyclic reagents were grafted onto cyclodextrins to evaluate the effects of different grafting reagents on the surface charge of porous cyclodextrin polymers.

[0101] The preparation method is the same as in Example 1, except that the different grafting reagents include: 3-bromomethylthiophene (3-Bm-Th, 10.34 g), 3-bromomethylpyrrole (3-Bm-Py, 11.43 g), 3-bromomethylfuran (3-Bm-FF, 8.93 g); benzyl bromide (BnBr, 9.48 g); the catalyst system includes: [EMIm]Cl / FeCl3, FeCl3; the alkylating agent is: BCMBP+DCX (biphenyl dichlorobenzyl, 10.44 g; p-dichlorobenzyl, 2.43 g).

[0102] The surface charge of the prepared porous cyclodextrin polymer was measured, and the results are shown in Table 1. This indicates that, compared to the benzene ring structure, positively charged porous cyclodextrin polymers can be prepared by grafting aromatic heterocyclic compounds under a Lewis acid ionic liquid catalytic system. Furthermore, when the thiophene ring is used as the graft monomer in the aromatic heterocyclic compound, the resulting polymer has a higher pK value. a The value is higher (>8), because after grafting, the S atoms on the thiophene cyclodextrin and the α-H atoms on the thiophene ring carry a certain amount of positive charge, and the ionic liquid catalytic system may charge the surface properties of the polymer. Therefore, 3-Bm-Th is preferred as the grafting agent, and [EMIm]Cl / FeCl3 ionic liquid is preferred as the catalytic system.

[0103] Table 1

[0104] serial number Cyclodextrin Grafting reagent Alkylating reagents catalyst Surface charge 1 β 3-Bm-Th BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> Positive charge 2 β 3-Bm-Py BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> neutral 3 β 3-Bm-FF BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> neutral 4 β BnBr BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> neutral 5 β BnBr BCMBP+DCX <![CDATA[FeCl3]]> negative charge

[0105] Note: Positive surface charge: pK a >8; Neutral surface charge: 6≤pK a ≤8; Negative surface charge: pK a <6.

[0106] Example 3

[0107] This embodiment provides the effect of different alkylating agents on the specific surface area of ​​porous cyclodextrin polymers.

[0108] The preparation method is the same as in Example 1, except that different alkylating agents are used, including: trimethyl orthoformate (Tmof, 2.68 g), dimethoxymethane (FDA, 1.92 g), biphenyl dichlorobenzyl + p-dichlorobenzyl (BCMBP, 4.75 g; DCX, 1.10 g), biphenyl dichlorobenzyl (BCMBP, 6.33 g), p-dichlorobenzyl (DCX, 4.41 g), and 1,4-dimethoxybenzene (3.48 g).

[0109] The specific surface area of ​​the prepared porous cyclodextrin polymers was measured, and the results are shown in Table 2. Tmof and BCMBP+DCX, as alkylating agents, yielded porous cyclodextrin polymers with high specific surface areas; while other alkylating agents yielded relatively low specific surface areas.

[0110] Table 2

[0111] serial number Cyclodextrin Grafting reagent Alkylating reagents catalyst Specific surface area 6 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> high 7 β 3-Bm-Th FDA <![CDATA[[EMIm]Cl / FeCl3]]> medium 8 β 3-Bm-Th BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> high 9 β 3-Bm-Th BCMBP <![CDATA[[EMIm]Cl / FeCl3]]> medium 10 β 3-Bm-Th DCX <![CDATA[[EMIm]Cl / FeCl3]]> medium 11 β 3-Bm-Th 1,4-Dimethoxybenzene <![CDATA[[EMIm]Cl / FeCl3]]> Low

[0112] Note: Specific surface area > 500m² 2 g -1 Medium specific surface area: 200m² 2 g -1 -500m2 g -1 Low specific surface area: <200m² 2 g -1 .

[0113] Example 4

[0114] This embodiment demonstrates the effect of different reaction temperatures on the specific surface area of ​​porous cyclodextrin polymers.

[0115] The preparation method is the same as in Example 1 (number 12 is Example 1), except that the final reaction temperatures are 60°C, 80°C, 100°C, 120°C and 150°C, and the reaction is carried out at a constant temperature of 80°C for 24 hours.

[0116] The specific surface area of ​​the prepared porous cyclodextrin polymer was measured, and the results are shown in Table 3. The reaction temperature has a significant impact on the polymerization reaction of thiophene cyclodextrin, and programmed temperature increase can be beneficial to obtaining a higher specific surface area.

[0117] Table 3

[0118] serial number Cyclodextrin Grafting reagent Alkylating reagents catalyst Temperature / °C <![CDATA[Specific surface area / m 2 g -1 > 12 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 730 13 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→60 56 14 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→80 650 15 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→100 695 16 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→150 736 17 β 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 80 527

[0119] Note:

[0120] 45℃→120℃: The temperature is programmed, first reacting at 45℃ for 5 hours, then increasing to 120℃ and reacting for 19 hours;

[0121] 80℃: Reaction at a constant temperature of 80℃ for 24 hours.

[0122] Example 5

[0123] This embodiment demonstrates the effect of different degrees of substitution in grafting reactions on the hydrophilicity of porous cyclodextrin polymers.

[0124] The preparation method is the same as in Example 1 (number 12 is Example 1), except that the grafting ratio is 21:1, 14:1, 7:1, or 3:1.

[0125] The contact angle of the prepared porous cyclodextrin polymer was measured, and the results are shown in Table 4. Controlling the number of hydroxyl groups on the cyclodextrin can achieve the regulation of the hydrophilicity and hydrophobicity of the polymer material. When the grafting ratio is greater than 14, the surface of the porous cyclodextrin polymer is moderately hydrophilic.

[0126] Table 4

[0127] serial number Cyclodextrin Grafting reagent Grafting ratio Alkylating reagents catalyst Temperature / °C Contact angle / ° 12 β 3-Bm-Th 21 Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 38 18 β 3-Bm-Th 14 Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 26 19 β 3-Bm-Th 7 Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 9 20 β 3-Bm-Th 3 Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 5

[0128] Note:

[0129] Grafting ratio: molar ratio of thiophene reagent to cyclodextrin;

[0130] Hydrophilic surface: contact angle < 20°; moderately hydrophilic surface: contact angle 20°~60°; hydrophobic surface: contact angle > 60°.

[0131] Example 6

[0132] This embodiment provides the surface charge and hydrophilicity of porous cyclodextrin polymers prepared from different supramolecular compounds.

[0133] The preparation method is the same as in Example 1 (number 12 is Example 1), except that the supramolecular compounds include: β-cyclodextrin (2g), α-cyclodextrin (1g), γ-cyclodextrin (3g), chitosan (3g), and columnar aromatic hydrocarbons (4g).

[0134] The surface charge and contact angle of the prepared porous cyclodextrin polymers were measured, and the results are shown in Table 5. The surface charge of the porous cyclodextrin polymers prepared by different supramolecular compounds was positive, and the surface was moderately hydrophilic.

[0135] Table 5

[0136] serial number supramolecular compounds Grafting reagent Alkylating reagents catalyst Temperature / °C Surface charge Contact angle / ° 12 β-Cyclodextrin 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 38 21 α-Cyclodextrin 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 36 22 γ-Cyclodextrin 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 45 23 Chitosan 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 33 24 Columnar aromatics 3-Bm-Th Tmof <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 59 25 β-Cyclodextrin 3-Bm-Th Tmof <![CDATA[FeCl3]]> 45→120 Positive charge 55 26 β-Cyclodextrin 3-Bm-Th BCMBP+DCX <![CDATA[[EMIm]Cl / FeCl3]]> 45→120 Positive charge 52

[0137] Note:

[0138] Surface charge is positive: pK a >8; Neutral surface charge: 6≤pK a ≤8; Negative surface charge: pK a <6;

[0139] Hydrophilic surface: contact angle < 20°; moderately hydrophilic surface: 20° ≤ contact angle ≤ 60°; hydrophobic surface: contact angle > 60°.

[0140] Comparative Example 1

[0141] This comparative example provides a porous cyclodextrin polymer (PBCD-BD) with a superhydrophilic surface.

[0142] The preparation method of the surface-superhydrophilic porous cyclodextrin polymer (PBCD-BD) is referenced from Tu, Y.; Xu, G.; Jiang, L.; Hu, X.; Xu, J.; Xie, X.; Li, A. Amphiphilic hyper-crosslinked porous cyclodextrin polymer with high specific surface area for rapid removal of organic micropollutants. Chemical Engineering Journal 2020, 382, ​​123015. β-cyclodextrin was grafted with a certain amount of benzyl groups, and crosslinked with biphenyl dichlorobenzyl and p-dichlorobenzyl to obtain a polymer with a specific surface area of ​​1445 m². 2 g -1 The PBCD-BD polymer retains most of the hydroxyl groups of the cyclodextrin, and the polymer surface is superhydrophilic.

[0143] Comparative Example 2

[0144] This comparative example provides commercially available immobilized carrier material, granular activated carbon.

[0145] In this comparative example, the granular activated carbon (GAC, DARCO) was purchased from Sigma-Aldrich and has a microporous structure with a specific surface area of ​​612 m². 2 g -1 .

[0146] Comparative Example 3

[0147] This comparative example provides a commercially available immobilized carrier material, polyurethane foam (PUF).

[0148] In this comparative example, polyurethane foam (PUF) was purchased from Henan Tianyi Environmental Protection Technology Co., Ltd. It has a non-porous structure and is commonly used for loading biofilms.

[0149] Example 7

[0150] This embodiment provides the application of porous thiophene-based cyclodextrin polymer (Th-CDP) (No. 12), porous cyclodextrin polymer (PBCD-BD), granular activated carbon (GAC), and polyurethane foam (PUF) as carrier materials in microbial immobilization.

[0151] In this embodiment, the surface properties of the carrier material are shown in Table 6.

[0152] Table 6

[0153] carrier material <![CDATA[Specific surface area m 2 g -1 > Contact angle ° <![CDATA[pK a ]]> Th-CDP No. 12 730 38 9.5 PBCD-BD 1445 9 5.5 GAC 612 49 4.4 PUF 0.6 123 5.9

[0154] In this embodiment, the microorganism is the white-rot fungus *Trametes versicolor*.

[0155] Immobilizing microorganisms on a carrier material involves first adding the carrier material to the culture medium, then adding functional microorganisms via mycelial inoculation, and finally, through shaking culture, immobilizing the microorganisms inside the carrier through physical adsorption. A schematic diagram is shown below. Figure 1 As shown, it specifically includes:

[0156] Preparation of mycelial solution:

[0157] Four 1cm 2 T. versicolor agar blocks of the desired size were inoculated into 100 mL of PD medium and cultured with shaking (135 rpm) at 25 °C for 4–5 days. The obtained mycelial balls were separated from the medium, and the mycelial balls were collected and 8 mL of 0.85% sterile physiological saline was added. The mixture was stirred with a high-speed homogenizer for 15 min to obtain mycelial solution, which was stored at 4 °C.

[0158] Immobilized microorganisms (preparation of porous carrier-microbial composite materials):

[0159] Add 1g of the above carrier material to 500mL of PD medium and autoclave; inoculate 1.5mL of mycelial solution of white-rot fungus T. versicolor into the medium and culture at 25℃ with shaking at 135rpm for 5 days to prepare immobilized white-rot fungus.

[0160] Scanning electron microscope image of immobilized white-rot fungi as shown below Figure 2 As shown, (a) is T.versicolor; (b) is Tv@PBCD-BD; (c) is Tv@Th-CDP; (d) is Tv@GAC; and (e) is Tv@PUF. From Figure 2 As can be seen, *T. versicolor* hyphae are banded, approximately 1 μm wide (hyphae ball diameter 6–7 mm). Immobilizing *T. versicolor* with the superhydrophilic PBCD-BD resulted in smaller biomaterial particles (3 mm in diameter) with better dispersibility and tighter binding. In contrast, *T. versicolor* immobilized with the moderately hydrophilic Th-CDP and GAC produced larger particles (Tv@Th-CDP, 6–7 mm in diameter) and Tv@GAC, 8–9 mm in diameter, with material aggregation within them. Due to its macroporous structure, *T. versicolor* immobilized with PUF adhered and grew within it. It can be seen that porous granular carriers and bulk PUFs exhibit different morphologies after immobilizing *T. versicolor*. Regarding biomaterial particles, due to dispersibility, carriers with stronger hydrophobicity produce larger particles.

[0161] Example 8

[0162] This embodiment provides the application of the immobilized white-rot fungus obtained in Example 7 in actual wastewater for the degradation of PPCPs in wastewater.

[0163] In this embodiment, four recalcitrant PPCPs—diclofenac (DCF), ibuprofen (IBU), sulfamethoxazole (SMX), and carbamazepine (CBZ)—were selected as target pollutants. A 0.5L continuous upflow reactor was constructed with an aeration rate of 200 L / min. -1 To study the degradation effect and bioactivity of immobilized white-rot fungi on recalcitrant PPCPs in the effluent from the secondary sedimentation tank of a wastewater treatment plant.

[0164] A schematic diagram illustrating the synergistic degradation of immobilized white-rot fungi Tv@Th-CDP in actual wastewater is shown below. Figure 3 As shown, the positively charged, moderately hydrophilic porous thiophene cyclodextrin polymer (Th-CDP) retains T. versicolor and its laccase (negatively charged) through electrostatic interaction, forming an adsorption-biodegradation synergistic system.

[0165] (1) Removal of high concentrations of PPCPs under rapid saturation conditions

[0166] Experimental method: 10 mg L was added to the reactor for the first 3 days. -1 The hydraulic retention time (HRT) of the above four types of PPCPs is 12h.

[0167] Degradation efficiency of four recalcitrant PPCPs as follows Figure 4 As shown, reactor A1 is T.versicolor, reactor C1 is Tv@PBCD-BD, reactor D1 is Tv@Th-CDP, reactor E1 is Tv@GAC, and reactor F1 is Tv@PUF. From Figure 4 As can be seen, compared with unfixed T.versicolor and PUF, porous carriers significantly improved the degradation performance of T.versicolor, especially T.versicolor fixed with porous cyclodextrin polymers (Tv@PBCD-BD and Tv@Th-CDP), which removed an average of 81.3% of PPCPs.

[0168] (2) Removal of PPCPs at ambient concentrations

[0169] Experimental method: After running at high concentration for 3 days, the addition amount of the four PPCPs was reduced to 10 μg / L. -1 HRT is 1d.

[0170] Results of the removal of environmental concentrations of PPCPs by porous cyclodextrin polymer-immobilized white-rot fungus *T. versicolor* and porous cyclodextrin polymer in a reactor are as follows: Figure 5 As shown, Influent1 is the influent to reactors B-I, C1, and D1; Influent2 is the influent to reactor B-II; B-I is PBCD-BD, B-II is Th-CDP, C1 is Tv@PBCD-BD, and D1 is Tv@Th-CDP. From Figure 5 As can be seen, in the later stage of the reaction, the effluent concentration of PPCPs in the porous cyclodextrin polymer-immobilized T. versicolor reactors (reactors C1 and D1) was much lower than that in their material control groups (reactors B-I and B-II). This coupling of adsorption and biodegradation extended the performance of the T. versicolor reactors, especially Tv@Th-CDP, where the average effluent concentration of PPCPs on day 18 was only 6.2% of that in its material control, achieving higher PPCP biodegradation.

[0171] (3) Removal of PPCPs under environmental concentration equilibrium conditions

[0172] Experimental method: The concentration of each of the four PPCPs was 10 μg / L. -1 HRT is always 1d.

[0173] Degradation efficiency of four recalcitrant PPCPs as follows Figure 6 As shown, reactor A2 is T. versicolor, reactor C2 is Tv@PBCD-BD, reactor D2 is Tv@Th-CDP, reactor E2 is Tv@GAC, and reactor F2 is Tv@PUF. Correlation analysis showed that the degradation efficiency of PPCPs was related to the pK of the carrier material. a A positive correlation was found, with positively charged Tv@Th-CDP achieving an average removal rate of 90.3% for PPCPs, a 17.2% improvement compared to unfixed T.versicolor; however, the removal efficiency of negatively charged Tv@PBCD-BD and Tv@GAC was affected.

[0174] Example 9

[0175] This embodiment provides a study on the parameter changes of immobilized white-rot fungi during the degradation of PPCPs in Example 8.

[0176] (1) ATP concentration, laccase activity, and glucose concentration in the suspension (sus) and inside the material (ex).

[0177] Experimental methods: During the immobilization process and reactor studies, water samples were periodically collected from the culture medium or reactor. Simultaneously, biomaterial particles or material samples were removed, weighed, and subjected to ultrasonic extraction twice, with 7.5 mL of 0.01 mol / L PBS added each time. -1 Extract the internal extract by ultrasonic extraction for 30 minutes at pH 7.2–7.4.

[0178] Figure 7 The concentrations of adenosine triphosphate (ATP), laccase activity, and glucose concentration of immobilized *T. versicolor* in the suspension (sus) and inside the material (ex) under environmental concentration equilibrium conditions are shown in the figures. (a) ATP concentration during immobilization; (b) ATP concentration in the reactor; (c) laccase activity; and (d) glucose concentration. Figure 7 As can be seen, the porous carrier material contains high levels of laccase and bioactivity. The positively charged Tv@Th-CDP retains the negatively charged T. versicolor laccase through electrostatic interaction, forming an adsorption-biodegradation synergistic system.

[0179] (2) Experimental methods for laccase, ATP, glucose concentration and pH value in suspension of immobilized white-rot fungus T. versicolor reactor: In the reactor study, water samples were collected from the reactor periodically to determine biological activity indicators and water quality parameters.

[0180] Figure 8 The concentrations of laccase, ATP, glucose, and pH in the suspension of the immobilized white-rot fungus *T. versicolor* in a reactor were investigated. Correlation analysis was used to study the factors influencing the carrier-biosynthetic degradation. The pH of the reactor influent was relatively stable, averaging 7.6, achieving efficient degradation and stable activity without pH adjustment. pH and ATP were key factors, while electrostatic interactions and the activity of *T. versicolor* significantly influenced the synergistic degradation of the carrier-biocomposite material.

[0181] (3) Species distribution in the immobilized white-rot fungus T. versicolor reactor

[0182] Experimental methods: The suspension and biomaterial particles (mycelial balls) in the reactor were extracted for metagenomic sequencing. The solution samples were filtered through MCE (0.2 μm) and the filter membrane was extracted. The distribution of the top 20 abundant communities in the reactor was analyzed at the species level.

[0183] Species distribution in immobilized white-rot fungi reactor as follows Figure 9 As shown, under rapid saturation pre-operation conditions ( Figure 9a) Compared to the bulk composite biomaterial Tv@PUF, the abundance of T. versicolor immobilized on porous particle carriers was significantly higher. At day 17, the abundance of T. versicolor in the particles of reactors C1 and D1 (Tv@PBCD-BD and Tv@Th-CDP) decreased but remained dominant, while the particles in reactor A1 (unimmobilized T. versicolor) had already fragmented, indicating that immobilization promoted the maintenance of T. versicolor activity in the reactors. Under environmental concentration equilibrium conditions ( Figure 9 b) The hydrophilicity or hydrophobicity of the carrier affected the abundance of T. versicolor. After 10 days of treatment, the abundance of T. versicolor in reactor A2 (unfixed) decreased by 97.1%. The superhydrophilic Tv@PBCD-BD even caused an increase in the abundance of Proteobacteria, while the moderately hydrophilic Tv@Th-CDP still had 31.2% T. versicolor abundance, with almost no loss. The dominant species in the reactor did not change significantly, providing a more stable environment for the growth of T. versicolor.

[0184] (4) Functional distribution of COG at the levels of secondary metabolite biosynthesis, transport, and decomposition in an immobilized white-rot fungus T. versicolor reactor

[0185] Experimental methods: The functions of the top 20 most abundant COGs (biosynthesis, transport, and degradation of secondary metabolites) in the reactor were analyzed by metagenomic sequencing.

[0186] The functional distribution of COG at the levels of secondary metabolite biosynthesis, transport, and degradation in an immobilized white-rot fungus *T. versicolor* reactor is as follows: Figure 10 As shown, cytochrome P450 (COG2124) plays a key degradation role in *T. versicolor* and is generally considered to participate in the degradation of pollutants in conjunction with laccase. Under rapid saturation pre-run conditions ( Figure 10 a) Regarding the abundance of COG2124, the abundance of T. versicolor immobilized on porous supports was significantly increased compared to both unimmobilized T. versicolor and PUF. Under environmental concentration equilibrium conditions ( Figure 10 b) After 10 days of treatment, the activity of Tv@Th-CDP was maintained, allowing it to retain high levels of cytochrome P450 function.

[0187] Example 10

[0188] This embodiment provides the application of porous thiophene-based cyclodextrin polymer (Th-CDP) (No. 12) as a carrier material in microbial immobilization.

[0189] In this embodiment, the microorganisms include nitrifying bacteria, denitrifying bacteria, Bacillus, Pseudomonas, and activated sludge.

[0190] Immobilizing microorganisms on carrier materials involves first adding the carrier material to the culture medium, then adding functional microorganisms through bacterial inoculation, and finally immobilizing the microorganisms inside the carrier through physical adsorption by shaking culture.

[0191] The morphology, degradation efficiency, and bioactivity of the prepared microbial composite materials are shown in Table 7.

[0192] Table 7

[0193] name carrier microorganism Biomaterial morphology Degradation efficiency Activity maintenance ability NB@Th-CDP Th-CDP Nitrifying bacteria Attachment growth higher better DB@Th-CDP Th-CDP Denitrifying bacteria Attachment growth higher better Bt@Th-CDP Th-CDP Bacillus Attachment growth Very high very good P@Th-CDP Th-CDP Pseudomonas Attachment growth Very high very good AS@Th-CDP Th-CDP Activated sludge Attachment growth Very high very good

[0194] Note:

[0195] Degradation efficiency: Very high: >80%; High: 60%–80%; Poor: 40%–60%; Very poor: <40%;

[0196] Excellent activity maintenance: after 10 days of immobilized microorganisms being added to the reactor, the abundance loss was <10%; Good: abundance loss was 10%–30%; Poor: abundance loss was 30%–60%; Very poor: abundance loss was >60%.

Claims

1. A porous carrier-microorganism composite material, characterized in that, The microorganism is a white-rot fungus; the porous carrier material is a porous thiophene-based cyclodextrin polymer; the BET specific surface area of ​​the porous carrier material is >600 m². 2 g -1 pK a >8 and the contact angle is 20°~60°; The method for preparing the porous support material includes the following steps: Cyclodextrin was dissolved in anhydrous DMF; then anhydrous NaH was slowly added at 0°C and stirred for 15 min; 3-bromomethylthiophene was slowly added dropwise and stirred at room temperature for 24 h; after the reaction was completed, the reaction was quenched with methanol; then the product was mixed with water and concentrated to near dryness under vacuum at 84°C; the obtained product was mixed with water and extracted with dichloromethane; the organic phase was dried with anhydrous sodium sulfate, then filtered and concentrated under vacuum at 35°C, and finally obtained by silica gel chromatography as a light yellow compound, thiophene-based cyclodextrin. Thiophene cyclodextrin was dissolved in 1-ethyl-3-methylimidazolium chloride / ferric chloride ionic liquid at 0°C. Anhydrous trimethyl orthoformate was slowly added dropwise at room temperature under nitrogen protection. The reaction mixture was stirred and heated to 45°C for 5 h, then heated to 120°C for 19 h. After cooling, the brown precipitate was washed with water and methanol until the filtrate was colorless. Then, it was extracted with methanol by Soxhlet for 24 h and dried under vacuum at 60°C for 24 h to obtain a porous thiophene cyclodextrin polymer.

2. The porous carrier-microorganism composite material according to claim 1, characterized in that, The molar ratio of 3-bromomethylthiophene to cyclodextrin is 14~21:1; the molar ratio of anhydrous trimethyl orthoformate to thiophene cyclodextrin is 5~100:

1.

3. The application of the porous carrier-microorganism composite material according to claim 1 or 2 in wastewater treatment.

4. The application according to claim 3, characterized in that, The microorganisms in the porous carrier-microbial composite material are white-rot fungi, which are used to degrade PPCPs in wastewater.

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