A method for removing organic pollutants by immobilizing laccase on an ionic liquid-modified metal-organic framework

Abstract

The present invention provides a method for removing organic pollutants by an ionic liquid-modified metal organic framework immobilized laccase. First, a metal organic framework is synthesized by a hydrothermal method, and the metal organic framework is surface-modified with a carboxyl-functionalized ionic liquid, and then the laccase is immobilized by activation with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide. The enzyme loading and activity recovery of the immobilized laccase prepared by this method are respectively as high as 61.3 mg / g and 83.3%, and its stability and pollutant removal efficiency are significantly improved. The immobilized laccase has good oxidative removal ability for common organic pollutants such as 2,4-DCP, bisphenol A, indole, anthracene, etc., has good application value in removing pollutants, and is conducive to the further application of immobilized laccase in organic wastewater treatment.

Description

Technical Field

[0001] The invention belongs to the technical field of water pollution treatment, and particularly relates to a method for removing organic pollutants by immobilizing laccase on a metal organic framework modified with ionic liquid. Background Art

[0002] In recent years, with the accelerated growth of the global economy, water pollution has become a growing concern due to the large amount of industrial waste being discharged into water bodies without any systematic management. In particular, the pollution of dyes, phenols and polycyclic aromatic hydrocarbons (PAHs) has attracted widespread attention due to their toxicity, carcinogenicity, mutagenicity and poor biodegradability. The treatment strategies for wastewater pollutants mainly include physical and chemical treatment and biodegradation treatment. Although physical and chemical methods have the advantage of high removal efficiency, their disadvantages such as high energy consumption, high cost and secondary pollution limit their further application. Biodegradation is considered to be an attractive and promising alternative method for removing a wide range of water pollution.

[0003] Laccase is a type of multi-copper oxidase. Its excellent oxidative degradation activity and specificity for a variety of substrates make it show good application prospects in the field of organic industrial wastewater treatment. However, its low operational stability and difficulty in separation and reuse limit its practical application. Seyed Mehdi (Bioresour Technol. 2020, 306, 123169) et al. used aminated magnetic iron oxide to immobilize laccase by glutaraldehyde cross-linking. Although the stability was improved, the activity recovery rate was only 27%. Enhui Wu (Chemosphere. 2019, 233, 327-335) et al. used metal-organic frameworks for adsorption and covalent immobilization of laccase. Although the immobilized enzyme of the sample had a high activity recovery rate, the removal efficiency of 2,4-dichlorophenol was only 87%. The development of a new technology for removing organic pollutants by immobilized laccase with good wastewater treatment effect and strong applicability has high scientific research and potential application value. Summary of the Invention

[0004] The present invention provides an ionic liquid-modified metal-organic framework-immobilized laccase composite biocatalyst. Using a metal-organic framework as a carrier, the metal-organic framework is modified with an ionic liquid. Laccase is then immobilized on the carrier under the action of an activator and applied to the removal of organic pollutants in water. The immobilized laccase prepared by this method achieves enzyme loading and activity recovery rates as high as 61.3 mg / g and 83.3%, respectively. Furthermore, its stability, pollutant removal efficiency, and reusability are significantly improved.

[0005] The technical solution adopted by the present invention is as follows: a method for removing organic pollutants by immobilizing laccase using an ionic liquid-modified metal-organic framework, comprising the following steps:

[0006] (1) Hydrothermal synthesis of the metal-organic framework NH2-MIL-101;

[0007] (2) Preparation of carboxyl functionalized ionic liquid modified IL-NH2-MIL-101;

[0008] (3) activating the carboxyl-functionalized ionic liquid-modified IL-NH2-MIL-101 obtained in step (2); (4) preparing magnetic polyethyleneimine-immobilized laccase: adding the activated ionic liquid-modified metal organic framework obtained in step (3) to a laccase solution having a concentration of 0.2-0.8 mL laccase solution / mL, a temperature of 25-50° C., a pH of 2.5-5.5, and shaking for 1-10 hours to perform a laccase immobilization reaction; obtaining Lac-IL-NH2-MIL-101;

[0009] (5) The immobilized laccase prepared in step (4) is applied to the removal of various organic pollutants.

[0010] Preferably, the concentration of the laccase solution in step (4) is 0.6 mL laccase solution / mL, the temperature is 30° C., the pH is 3.0, and the reaction time is 4 hours.

[0011] Preferably, the preparation method of the metal organic framework NH2-MIL-101 in step (1) is as follows: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate and 0.4 g of sodium hydroxide are dissolved in 30 ml of deionized water and transferred to a hydrothermal reactor, and heated at 150°C in a drying oven for 12 hours; the resulting product is washed multiple times with N,N-dimethylformamide, and the washed product is dispersed in ethanol, placed in an oven at 90°C for 6 hours, and the reaction is repeated twice.

[0012] Preferably, the preparation method of the metal-organic framework modified with carboxyl functionalized ionic liquid in step (2) is as follows: The preparation method of the metal-organic framework modified with carboxyl functionalized ionic liquid is as follows: take 3 mg of 1,8-diazabicycloundec-7-ene as a catalyst and add it to 20 ml of toluene, add 1.0 g of the metal-organic framework material synthesized in step (1), stir in a low-temperature stirrer for 1.5, add 2.2 mmol N,N'-carbonyldiimidazole, slowly warm to room temperature, stir at room temperature for 12 hours, wash and dry; then divide the obtained solid into 50 ml of dimethyl sulfoxide, slowly add 1-(3-aminopropyl)-imidazole (0.3-0.8 g) to the reaction system and stir at room temperature for 12 hours. Then wash and dry the solid. The dried sample was then alkylated with excess chloroacetic acid (0.3-0.8 g) in acetonitrile at 80°C under nitrogen for 24 hours, washed several times, and dried. Finally, the intermediate product was ion-exchanged with excess sodium tetrafluoroborate (NaBF4) in water for a specified period of time, washed, and dried. The final product was named IL-NH2-MIL-101.

[0013] Preferably, the activation method of the carboxyl functionalized ionic liquid modified NH2-MIL-101 in step (3) is as follows: 0.1 mol N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), 0.1 mol N-hydroxysuccinimide (NHS) and 0.5 g NH2-MIL-101 are dispersed in 50 mL of citrate buffer solution pH 6.0 and stirred in a low-temperature stirrer for 1.5 h;

[0014] Preferably, the laccase immobilization process in step (4) is as follows: adding the activated ionic liquid-modified metal organic framework obtained in step (3) to the laccase solution to carry out a laccase immobilization reaction; and then obtaining the ionic liquid-modified metal organic framework immobilized laccase by centrifugation, washing, and freeze-drying.

[0015] Preferably, the immobilized laccase is applied to the removal of various organic pollutants such as 2,4-dichlorophenol, bisphenol A, indole, and anthracene.

[0016] Laccase was immobilized on a metal-organic framework modified with a carboxyl-functionalized ionic liquid (LIF) by activation with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). The specific steps are as follows:

[0017] 1) Metal-organic frameworks were synthesized by hydrothermal synthesis: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate, and 0.4 g of sodium hydroxide were dissolved in 30 ml of deionized water and transferred to a hydrothermal reactor. The mixture was heated at 150°C in a drying oven for 12 h. The resulting product was washed multiple times with N,N-dimethylformamide, dispersed in ethanol, and placed in an oven at 90°C for 6 h. The reaction was repeated twice.

[0018] 2) Take 3.0 mg of 1,8-diazabicycloundec-7-ene as a catalyst and add it to 20 ml of toluene. Add 1.0 g of the metal organic framework material synthesized in step (1) and stir in a low temperature stirrer for 1.5 seconds. Then add 2.2 mmol of N,N'-carbonyldiimidazole and slowly warm to room temperature. Stir at room temperature for 12 hours and wash and dry. Then, divide the obtained solid into 50 ml of dimethyl sulfoxide and slowly add 0.5 g of 1-(3-aminopropyl)-imidazole to the reaction system and stir at room temperature for 12 hours. Then wash and dry the solid. Then, alkylate the dried sample with chloroacetic acid (0.5-1.0 g) in acetonitrile at 80°C under nitrogen for 24 hours, wash several times and dry. Finally, ion exchange the intermediate product with excess sodium tetrafluoroborate in water for a certain period of time and wash and dry. The final product is named IL-NH2-MIL-101.

[0019] The excess 1-(3-aminopropyl)-imidazole 0.3g-0.8g, preferably 0.5g.

[0020] The excess chloroacetic acid is 0.3-0.8 g, preferably 0.5 g.

[0021] The anion exchange time is 0-24 hours, preferably 12 hours.

[0022] 3) NH2-MIL-101 and IL-NH2-MIL-101 were added to laccase solutions of varying concentrations and shaken in a water bath for a specified period of time under different temperature and pH conditions to immobilize the laccase. Immobilized laccases were then separated using an external magnetic field, washed, and freeze-dried to obtain the immobilized laccases. The immobilized laccases were designated Lac-NH2-MIL-101 and Lac-IL-NH2-MIL-101, respectively.

[0023] The concentration of the laccase solution in the immobilization reaction is 0.3-0.7 mL laccase solution / mL, preferably 0.6 mL laccase solution / mL.

[0024] The immobilization reaction time is 4-8 hours, preferably 4 hours.

[0025] The temperature of the immobilization reaction is 20-50°C, preferably 30°C.

[0026] The pH of the immobilization reaction is 3.0-5.0, preferably 3.0.

[0027] The method for determining the laccase activity in the method is as follows: a 1 mmol / L 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt (ABTS) solution is prepared with a citric acid buffer solution (pH 3.0), an appropriate amount of immobilized laccase is added to 5 mL of the ABTS solution, and the reaction is carried out at 30°C for 5 minutes. The absorbance of the supernatant at 420 nm is then measured using a UV-visible spectrophotometer. The enzyme activity unit (1U) is defined as the amount of laccase required to catalyze the oxidation of 1 μmol of ABTS in one minute under certain conditions. The experiment is repeated three times and the average value is taken. The activity calculation formula is as follows:

[0028] Apparent activity (U / g biocatalyst) = A × 10 6 ×V t / (36000×t×m1)

[0029] Specific activity (U / g protein) = A × 10 6 ×V t / (36000×t×m2)

[0030] A–ABTS ·+ absorbance at 420 nm;

[0031] V t –Total volume of reaction solution (L);

[0032] 36000–ABTS ·+ The molar extinction coefficient (M -1 cm -1 );

[0033] t – reaction time (min);

[0034] m1 – mass of immobilized enzyme (g);

[0035] m2 – the amount of protein contained in the immobilized enzyme (g).

[0036] The method for removing organic pollutants by using the ionic liquid modified metal organic framework immobilized laccase is as follows: adding the immobilized laccase to an aqueous solution of various organic pollutants to react for a certain time, and after the reaction is completed, determining the removal efficiency of various pollutants by high performance liquid chromatography.

[0037] RE(%)=(C0-C t ) / C0×100

[0038] Among them, C0 and C t Represent the concentrations of pollutants before and after treatment, respectively.

[0039] The beneficial effects of the present invention are:

[0040] (1) The loading capacity and activity recovery rate of the metal organic framework immobilized laccase modified by the ionic liquid of the present invention are as high as 61.3 mg / g and 83.3%, respectively. The metal organic framework immobilized laccase of Example 1 has the best loading capacity and activity recovery rate compared with other examples. It removes 2,4-DCP aqueous solution, and the removal effect is 100% after 6 hours. Moreover, the immobilized laccase can still maintain a removal efficiency of more than 90% after being reused 8 times. At the same time, its thermal stability has also been greatly improved. Compared with other immobilized laccases, it has a strong competitive advantage. It is significantly higher than the degradation effect of laccase on 2,4-DCP by adsorption and covalent means of metal organic framework reported in the literature (Chemosphere.2019,233,327-335) (87% degradation in 12 hours, 40% degradation efficiency retained after repeated use 3 times).

[0041] (2) The modification of carboxyl functionalized ionic liquid can enhance the carrier loading capacity and the activity of immobilized laccase. The activity recovery rate of Lac-IL-NH2-MIL-101 was increased by 25.2% compared with Lac-NH2-MIL-101.

[0042] (3) The ionic liquid-modified metal organic framework-immobilized laccase of the present invention not only has a good removal efficiency for common phenolic organic pollutants, but also has a removal efficiency of more than 85% for organic pollutants such as indole and anthracene that are more difficult to remove.

[0043] (4) As shown in Table 1, the laccase solution concentration in step (4) is 0.6 mL laccase solution / mL, the temperature is 30°C, the pH is 3.0, and the reaction time is 4 hours. The ionic liquid-modified metal organic framework immobilized laccase has the best loading capacity and activity recovery rate. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] The present invention will be further described below with reference to the accompanying drawings.

[0045] Figure 1 1 and 2 are XRD spectra of the products NH2-MIL-101 and IL-NH2-MIL-101 of each part in the synthesis process of the metal organic framework and the ionic liquid modified organic framework prepared in Example 1.

[0046] Figure 2 It is the XPS spectra of NH2-MIL-101 and IL-NH2-MIL-101 prepared in Example 1.

[0047] Figure 3 This is a scanning electron microscope image of NH2-MIL-101 prepared in Example 1.

[0048] Figure 4 This is a scanning electron microscope image of IL-NH2-MIL-101 prepared in Example 1. DETAILED DESCRIPTION

[0049] The present invention will be further described below with reference to the following examples, which are provided for illustration only and to illustrate that the spirit and scope of the present invention are not limited to the details and modifications herein.

[0050] Example 1:

[0051] (1) Hydrothermal synthesis of metal-organic frameworks: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate, and 0.4 g of sodium hydroxide were dissolved in 30 ml of deionized water and transferred to a hydrothermal reactor. The mixture was heated at 150°C in a drying oven for 12 h. The resulting product was washed several times with N,N-dimethylformamide, dispersed in ethanol, and placed in an oven at 90°C for 6 h. The reaction was repeated twice. The final product was dried in a vacuum oven.

[0052] (2) Take 3 mg of 1,8-diazabicycloundec-7-ene as a catalyst and add it to 20 ml of toluene. Add 1.0 g of the metal organic framework material synthesized in step (1) and stir it in a low temperature stirrer for 1.5 seconds. Then add 2.2 mmol of N,N'-carbonyldiimidazole and slowly warm it to room temperature. Stir it at room temperature for 12 hours and wash and dry it. Then, divide the obtained solid into 50 ml of dimethyl sulfoxide and slowly add 0.5 g of 1-(3-aminopropyl)-imidazole to the reaction system and stir it at room temperature for 12 hours. Then wash and dry the solid. Then, alkylate the dried sample with 0.5 g of chloroacetic acid in acetonitrile under nitrogen at 80 ° C for 24 hours, wash several times and dry it. Finally, ion exchange the intermediate product with 0.8 g of sodium tetrafluoroborate in water for 12 hours and wash and dry it. The final product is named IL-NH2-MIL-101.

[0053] (3) The activation method of carboxyl functionalized ionic liquid modified IL-NH2-MIL-101 is as follows: 0.1 mol N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), 0.1 mol N-hydroxysuccinimide (NHS) and 1.0 g IL-NH2-MIL-101 are ultrasonically dispersed in 50 mL citrate buffer solution pH 6.0 and stirred in a low-temperature stirrer for 1.5 h;

[0054] (4) adding the activated ionic liquid-modified metal organic framework obtained in step (3) to the laccase solution to carry out a laccase immobilization reaction; and then obtaining the ionic liquid-modified metal organic framework-immobilized laccase by centrifugation, washing, and freeze-drying.

[0055] (5) 0.15 g of the activated ionic liquid-modified metal organic framework was added to the laccase solution (0.6 mL of laccase solution / mL), and the mixture was shaken in a water bath shaker at 30°C and pH 3.0 for 4 hours to perform the laccase immobilization reaction; then, the ionic liquid-modified metal organic framework immobilized laccase was obtained by centrifugation, washing, and freeze-drying.

[0056] (6) To measure the thermal stability of immobilized laccase, the thermal stability parameters and thermodynamic parameters of laccase from Aspergillus oryzae (Ningxia Xiasheng Industrial Group Co., Ltd.) and immobilized laccase were determined. The activation energies of free laccase and immobilized laccase were 77.59 and 93.20 kJ·mol -1 The half-lives of free laccase and immobilized laccase at 50°C were 52.21 and 101.93 min, respectively. The enzyme loading and activity recovery of the immobilized laccase prepared by this method were as high as 61.3 mg / g and 83.3%, respectively.

[0057] To measure the storage stability of immobilized laccase, free and immobilized laccases were stored at 4°C for one month, and their residual enzyme activity was measured every five days. The results showed that after 30 days of storage, only 10.97% of the free laccase activity remained, while 65.15% of the immobilized enzyme activity remained.

[0058] To further enhance the advantages of ionic liquids, laccase was immobilized on NH2-MIL-101 under optimized conditions (enzyme concentration: 0.6 mL / mL; immobilization time: 2 h; pH: 3.0; immobilization temperature: 30°C). The enzyme loading and activity recovery were 39.1 mg / g and 58.1%, respectively. Lac-IL-NH2-MIL-101 exhibited a 22.2 mg / g increase in loading compared to Lac-NH2-MIL-101, while activity recovery increased by 25.2%.

[0059] Example 2:

[0060] (1) Hydrothermal synthesis of metal-organic frameworks: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate, and 0.4 g of sodium hydroxide were dissolved in 30 ml of deionized water and transferred to a hydrothermal reactor. The mixture was heated at 150°C in a drying oven for 12 h. The resulting product was washed several times with N,N-dimethylformamide, dispersed in ethanol, and placed in an oven at 90°C for 6 h. The reaction was repeated twice. The final product was dried in a vacuum oven.

[0061] (2) Take 3 mg of 1,8-diazabicycloundec-7-ene as a catalyst and add it to 20 ml of toluene, add 1.0 g of the metal organic framework material synthesized in step (1), stir in a low-temperature stirrer for 1.5, add 2.2 mmol of N,N'-carbonyldiimidazole, slowly warm to room temperature, stir at room temperature for 12 hours, wash and dry; then divide the obtained solid into 50 ml of dimethyl sulfoxide, slowly add 0.3 g of 1-(3-aminopropyl)-imidazole to the reaction system and stir at room temperature for 12 hours. Then wash and dry the solid. Then alkylate the dried sample with chloroacetic acid in acetonitrile under nitrogen at 80 ° C for 24 hours, wash several times and dry; finally, ion exchange the intermediate product with excess sodium tetrafluoroborate in water for 12 hours, wash and dry. The final product is named IL-NH2-MIL-101

[0062] (4) adding the activated ionic liquid-modified metal organic framework obtained in step (3) to the laccase solution to carry out a laccase immobilization reaction; and then obtaining the ionic liquid-modified metal organic framework-immobilized laccase by centrifugation, washing, and freeze-drying.

[0063] (5) The activated ionic liquid-modified metal organic framework was added to the laccase solution (0.6 mL laccase solution / mL), and the mixture was shaken in a water bath shaker at 30°C and pH 3.0 for 4 hours to perform the laccase immobilization reaction; the ionic liquid-modified metal organic framework immobilized laccase was then obtained by centrifugation, washing, and freeze-drying. The metal organic framework immobilized laccase was then obtained by washing and freeze-drying.

[0064] When the amount of 1-(3-aminopropyl)-imidazole in step (2) was reduced to 0.3 g, the loading amount and activity recovery rate of the ionic liquid-modified metal organic framework immobilized enzyme were 45.6 mg / g and 60.2%, respectively. The loading amount was reduced compared to that obtained in Example 1.

[0065] Example 3: Synthesis of immobilized enzymes was performed as in Example 1, except that in step (2), the amount of 1-(3-aminopropyl)-imidazole was 0.8 g. The resulting ionic liquid-modified metal organic framework immobilized enzyme had a loading capacity of 58.8 mg / g and an activity recovery rate of 79.8%, respectively. Considering economic benefits, 0.5 g is preferred.

[0066] Example 4: Synthesis of immobilized enzymes Referring to Example 1, the difference is that the amount of chloroacetic acid in step (2) is reduced to 0.3 g, the loading amount of the obtained ionic liquid modified metal organic framework immobilized enzyme is 48.7 mg / g, and the activity recovery rate is 75.3%.

[0067] Example 5: The synthesis process of the immobilized enzyme refers to Example 1, except that the amount of chloroacetic acid in step (2) is reduced to 0.8 g. The loading amount of the immobilized enzyme on the obtained ionic liquid-modified metal-organic framework is 55.7 mg / g, and the activity recovery rate is 77.5%. Considering the economic benefit, 0.5 g is preferred.

[0068] Example 6: Synthesis of the immobilized enzyme was performed as in Example 1, except that the anion exchange time in step (2) was reduced by 10 h. The immobilized laccase loading was 53.6 mg / g, and the enzyme activity recovery was 67.8%. Compared to Example 1, the enzyme loading and activity recovery were slightly lower.

[0069] Example 7: Synthesis of immobilized laccase The synthesis process was similar to that of Example 1, except that the laccase immobilization reaction conditions in step (2) were different. The loading amount and activity recovery rate of immobilized laccase Lac-IL-NH2-MIL-101 under different reaction conditions are shown in Table 1:

[0070] Table 1 Effects of different immobilization conditions on the immobilization results of IL-NH2-MIL-101

[0071]

[0072]

[0073] Example 8: Synthesis of Immobilized Laccase Reference Example 1. Add 10 mg of the immobilized enzyme to 10 ml of a 10 mg / L 2,4-DCP aqueous solution. Stir the reaction at 25°C, and sample the solution every two hours to monitor the removal efficiency. After six hours, the immobilized enzyme achieved 100% 2,4-DCP removal. This is significantly higher than the reported degradation of 2,4-DCP by laccase using metal-organic frameworks for adsorption and covalent degradation (87% degradation in 12 hours, with 40% retention after three reuses) (Chemosphere. 2019, 233, 327-335).

[0074] Moreover, the immobilized laccase can still maintain a removal efficiency of more than 90% after being reused 8 times.

[0075] Example 9: Synthesis of Immobilized Laccase Reference Example 1. 10 mg of the immobilized enzyme was added to 10 ml of a 10 mg / L aqueous solution of bisphenol A. The reaction was stirred at 25°C, and samples were taken every two hours to monitor the removal efficiency. After 10 hours, the immobilized enzyme achieved 100% removal. After eight repetitions, the degradation efficiency remained above 80%.

[0076] Example 10: Synthesis of Immobilized Laccase Referring to Example 1, 10 mg of Lac-IL-NH2-MIL-101 from Example 1 was added to 10 mL of an anthracene aqueous solution (10 mg / L). The reaction was stirred at 25°C, and samples were taken every two hours to monitor the removal efficiency. After 12 hours, the anthracene removal efficiency reached a maximum of 88.3%.

[0077] Example 11: Synthesis of Immobilized Laccase: Refer to Example 1. Lac-IL-NH2-MIL-101 (10 mg) from Example 1 was added to 10 mL of an aqueous indole solution (10 mg / L). The reaction was stirred at 25°C, and samples were taken every two hours to monitor the removal efficiency. After 12 hours, the indole removal efficiency reached a maximum of 85.3%.

Claims

1. A method for removing organic pollutants by immobilizing laccase on an ionic liquid-modified metal-organic framework, characterized in that: Here are the steps: (1) Hydrothermal synthesis of the metal-organic framework NH2-MIL-101; (2) Preparation of IL-NH2-MIL-101 modified with carboxyl functionalized ionic liquid; The preparation method of the metal organic framework modified with carboxyl functionalized ionic liquid in step (2) is as follows: 3 mg of 1,8-diazabicycloundec-7-ene was taken as a catalyst and added to 20 ml of toluene, 1.0 g of the metal organic framework material synthesized in step (1) was added, and stirred in a low temperature stirrer at 0-3 ° C for 1.5 h, and then 2.2 mmol of N,N'-carbonyldiimidazole was added, and the mixture was slowly heated to room temperature, stirred at room temperature for 12 h, and washed and dried; then the obtained solid was dispersed in 50 ml of dimethyl sulfoxide, and 0.5 g of 1-(3-aminopropyl)-imidazole was slowly added dropwise to the reaction system and stirred at room temperature for 12 h; then the solid was washed and dried, and the dried sample was alkylated with 0.5 g of chloroacetic acid in acetonitrile at 80 ° C under nitrogen for 24 h, washed several times and dried; finally, the intermediate product was mixed with 0.8 g sodium tetrafluoroborate was ion exchanged in water for 12 h, washed and dried, and the final product was named IL-NH2-MIL-101; (3) activating the carboxyl-functionalized ionic liquid-modified IL-NH2-MIL-101 obtained in step (2); (4) Preparation of ionic liquid-modified metal organic framework immobilized laccase: adding the activated ionic liquid-modified metal organic framework obtained in step (3) to a laccase solution derived from Aspergillus oryzae, wherein the concentration of the laccase solution is 0.2-0.8 mL laccase solution / mL, the temperature is 25-50 °C, the pH is 2.5-5.5, and the mixture is shaken for 1-10 hours to perform a laccase immobilization reaction; thereby obtaining Lac-IL-NH2-MIL-101; (5) The immobilized laccase prepared in step (4) is applied to the removal of organic pollutants such as 2,4-dichlorophenol, bisphenol A, indole, and anthracene.

2. The method for removing organic pollutants by using laccase immobilized on an ionic liquid-modified metal-organic framework according to claim 1, wherein: The concentration of the laccase solution in step (4) is 0.6 mL laccase solution / mL, the temperature is 30 °C, the pH is 3.0, and the reaction time is 4 hours.

3. The method for removing organic pollutants by using laccase immobilized on an ionic liquid-modified metal-organic framework according to claim 2, characterized in that: The preparation method of the metal organic framework in step (1) is as follows: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate and 0.4 g of sodium hydroxide are dissolved in 30 ml of deionized water, stirred at room temperature for 30 min, transferred to a hydrothermal reactor, and heated at 150°C in a forced air drying oven for 12 h; the obtained product is washed with N,N-dimethylformamide several times, the washed product is dispersed in ethanol, placed in an oven at 90°C for 6 h, and the reaction is repeated twice; and the final product is dried in a vacuum oven to obtain the final product; named NH2-MIL-101.

4. The method for removing organic pollutants by using laccase immobilized on an ionic liquid-modified metal-organic framework according to claim 2, wherein: The activation method of the carboxyl functionalized ionic liquid modified IL-NH2-MIL-101 in step (3) is as follows: 0.1 mol N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride EDC, 0.1 mol N-hydroxysuccinimide NHS and 1.0 g IL-NH2-MIL-101 are ultrasonically dispersed in 50 mL citrate buffer solution pH 6.0 and stirred in a low-temperature stirrer for 1.5 h.

5. The method for removing organic pollutants by using laccase immobilized on an ionic liquid-modified metal organic framework according to claim 2, characterized in that: The laccase immobilization process in step (4) is as follows: adding the activated ionic liquid-modified metal organic framework obtained in step (3) to the laccase solution to carry out a laccase immobilization reaction; then, centrifuging, washing, and freeze-drying to obtain an ionic liquid-modified metal organic framework immobilized laccase, named Lac-IL-NH2-MIL-101.

6. The method for removing organic pollutants by using laccase immobilized on an ionic liquid-modified metal-organic framework according to claim 2, characterized in that: The following steps are involved: (1) Hydrothermal synthesis of metal-organic frameworks: 0.72 g of 2-aminoterephthalic acid, 1.6 g of chromium nitrate nonahydrate, and 0.4 g of sodium hydroxide were dissolved in 30 ml of deionized water and transferred to a hydrothermal reactor. The mixture was heated at 150°C in a drying oven for 12 h. The resulting product was washed several times with N,N-dimethylformamide, dispersed in ethanol, and placed in an oven at 90°C for 6 h. The reaction was repeated twice. The final product was dried in a vacuum oven and named NH2-MIL-101. (2) Take 3 mg of 1,8-diazabicycloundec-7-ene as a catalyst and add it to 20 ml of toluene, add 1.0 g of the metal organic framework material synthesized in step (1), stir it in a low-temperature stirrer at 0-3 ° C for 1.5 h, add 2.2 mmol N,N'-carbonyldiimidazole, slowly warm it to room temperature, stir it at room temperature for 12 h, wash and dry it; then disperse the obtained solid in 50 ml of dimethyl sulfoxide, slowly add 0.5 g of 1-(3-aminopropyl)-imidazole to the reaction system and stir it at room temperature for 12 h, then wash and dry the solid, and then alkylate the dried sample with 0.5 g of chloroacetic acid in acetonitrile at 80 ° C under nitrogen for 24 h, wash several times and dry it; finally, the intermediate product is ion exchanged with 0.8 g of sodium tetrafluoroborate in water for 12 h, washed and dried; the final product is named IL-NH2-MIL-101; (3) The activation method of carboxyl functionalized ionic liquid modified IL-NH2-MIL-101 is as follows: 0.1 mol N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride EDC, 0.1 mol N-hydroxysuccinimide NHS and 1.0 g IL-NH2-MIL-101 are ultrasonically dispersed in 50 mL citrate buffer solution pH 6.0 and stirred in a low-temperature stirrer for 1.5 h; (4) 0.15 g of the activated ionic liquid-modified metal organic framework was added to a laccase solution of 0.6 mL laccase solution / mL, and the solution was shaken in a water bath shaker at 30 °C and pH 3.0 for 4 hours to perform a laccase immobilization reaction; the ionic liquid-modified metal organic framework-immobilized laccase was then obtained by centrifugation, washing, and freeze-drying. The metal organic framework-immobilized laccase was then washed and freeze-dried to remove organic pollutants.

Images