Cellulose-DNA hydrogel, preparation method of cellulose-DNA hydrogel, enzyme assembled hydrogel for removing organic pollutants and preparation method of enzyme assembled hydrogel

Abstract

The invention relates to the technical field of water treatment, in particular to cellulose-DNA hydrogel, a preparation method of the cellulose-DNA hydrogel, enzyme assembled hydrogel for removing organic pollutants and a preparation method of the enzyme assembled hydrogel. The preparation method of the cellulose-DNA hydrogel comprises the following steps: adding DNA and 1, 4-butanediol diglycidyl ether into a cellulose solution, and fully stirring to obtain a doped mixed solution; centrifugally degassing the mixed solution, and then pouring the mixed solution into a mold for standing and molding to obtain preformed hydrogel; the preformed hydrogel is put into an ethanol solution to be subjected to soaking treatment, then deionized water is used for cleaning, and the cellulose-DNA hydrogel is obtained. The preparation method is simple to operate, and the prepared hydrogel is good in mechanical strength, large in specific surface area and high in POPs removal rate.

Description

Technical Field

[0001] The invention relates to the technical field of water treatment, in particular to a cellulose-DNA hydrogel and a preparation method thereof, and an enzyme-assembled hydrogel for removing organic pollutants and a preparation method thereof. Background Art

[0002] Persistent organic pollutants (POPs) are a class of chemicals with high toxicity, bioaccumulation and long-range migration, which pose a serious threat to the environment and human health. Since POPs can be frequently detected in various water bodies, their effective removal has become a key challenge in the field of environmental remediation.

[0003] At present, the environmental remediation technologies of POPs mainly include adsorption, flocculation, advanced oxidation and biodegradation. Among these methods, biodegradation, especially enzyme-catalyzed degradation, shows great application potential due to its high efficiency, low cost, less secondary pollution and ability to overcome the complex culture process and harsh operating conditions of microbial degradation. However, the industrial application of enzymes is limited by their structural instability and easy inactivation during use, which seriously affects their industrial applicability and stability.

[0004] In order to solve this problem, the relevant technology has developed enzyme immobilization technology, which not only protects the degradation activity of the enzyme but also realizes the recycling of the enzyme by confining the enzyme in a certain space. Among the numerous enzyme carrier materials, hydrogel is considered to be one of the ideal three-dimensional polymer carriers for immobilizing or assembling enzymes because of its similarity to the natural living environment of the enzyme (extracellular matrix).

[0005] As a natural polymer, cellulose is considered to be one of the best polymers for synthesizing hydrogels due to its advantages such as non-toxicity, environmental protection and low cost. Cellulose-based hydrogels provide sufficient fixing sites for enzyme molecules due to their abundant hydroxyl groups on the surface, which helps to further remove POPs in water. However, the cellulose-based hydrogels synthesized by existing technologies have some shortcomings, mainly including:

[0006] 1. The dense hydrogen bonds between cellulose molecules result in a low specific surface area of ​​the synthesized hydrogel, which limits the amount of enzyme immobilization and thus reduces the removal efficiency of POPs;

[0007] 2. Salt ions in the water environment may weaken the hydrogen bonding between cellulose chains, resulting in insufficient mechanical strength of the hydrogel, reducing its reusability in environmental remediation and increasing the risk of pollutant leakage and secondary pollution;

[0008] 3. The matrix in real wastewater is complex and contains a variety of interfering substances, such as heavy metals, other organic pollutants and dissolved organic matter. Existing technologies are difficult to efficiently remove POPs from it, which seriously limits its widespread use in industrial applications.

[0009] Therefore, how to improve the mechanical properties of cellulose hydrogel and the removal rate of POPs has attracted more and more attention from technicians in this field. Summary of the invention

[0010] In order to solve or partially solve the problems existing in the related art, the present invention provides a cellulose-DNA hydrogel and a preparation method thereof, and an enzyme-assembled hydrogel for removing organic pollutants and a preparation method thereof.

[0011] The present invention provides a method for preparing a cellulose-DNA hydrogel, which is characterized by comprising:

[0012] Add DNA and 1,4-butanediol diglycidyl ether into the cellulose solution and stir thoroughly to obtain a doped mixed solution;

[0013] The mixed solution is centrifuged to degas, and then poured into a mold to be formed to obtain a preformed hydrogel;

[0014] The preformed hydrogel is immersed in an ethanol solution and then washed with deionized water to obtain a cellulose-DNA hydrogel.

[0015] Furthermore, the cellulose solution is prepared according to the following method:

[0016] Under low temperature conditions, cotton linters are added into an alkaline solution containing urea and stirred to dissolve to obtain a cellulose solution.

[0017] Furthermore, the DNA is salmon sperm DNA.

[0018] Furthermore, the concentration of the ethanol solution used in the soaking treatment is 75%, and the soaking treatment time is 6 to 8 hours.

[0019] Furthermore, the rotation speed of the centrifugal degassing treatment is 4000-6000 rpm, and the time is 20-25 min; and / or, the temperature of the static molding is 60-65° C., and the time is 4-6 h.

[0020] Furthermore, the usage ratio of the DNA, 1,4-butanediol diglycidyl ether and cellulose is (3-4): (0.6-1.2): (3-4).

[0021] The present invention also provides a cellulose-DNA hydrogel, which is prepared according to any one of the methods described above.

[0022] The present invention also provides a method for preparing an enzyme-assembled hydrogel for removing organic pollutants, which comprises:

[0023] Prepare cellulose-DNA hydrogel according to any of the above methods;

[0024] A buffer containing an enzyme is added to the cellulose-DNA hydrogel for a shaking treatment, and then washed with a buffer to obtain an enzyme-assembled hydrogel for removing organic pollutants.

[0025] Furthermore, the enzyme is laccase.

[0026] The present invention also provides an enzyme-assembled hydrogel for removing organic pollutants, which is prepared according to the above method.

[0027] The preparation method of the cellulose-DNA hydrogel provided by the present invention has the following beneficial effects:

[0028] The cellulose-DNA hydrogel prepared by this method uses 1,4-butanediol diglycidyl ether as a cross-linking agent to link DNA to the cellulose molecular chain to form a more stable three-dimensional network structure. On the one hand, the covalent bond produced by the etherification reaction between 1,4-butanediol diglycidyl ether and DNA / cellulose is stronger and more stable than the hydrogen bond between cellulose, thereby increasing the mechanical strength of the prepared cellulose-DNA hydrogel; on the other hand, the long chain structure of 1,4-butanediol diglycidyl ether can greatly stretch the cellulose chain, which is conducive to increasing the specific surface area and resulting in an increase in exposed sites, so that more enzymes can be fixed for efficient degradation and removal of POPs in real wastewater. Finally, the connected DNA, on the one hand, helps to further increase the specific surface area, and on the other hand, it has a good capture effect on POPs, which is conducive to the efficient removal and degradation of POPs on the subsequent enzyme-fixed cellulose-based hydrogel, and improves the removal rate of POPs. In addition, the preparation method is simple to operate and easy to realize industrial production and application.

[0029] It is to be understood that the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] The above and other objects, features and advantages of the present invention will become more apparent through a more detailed description of exemplary embodiments of the present invention in conjunction with the accompanying drawings, wherein like reference numerals generally represent like components throughout the exemplary embodiments of the present invention.

[0031] Figure 1 is a scanning electron microscope image of the cellulose-DNA hydrogel prepared in Example 3 of the present invention;

[0032] Figure 2 is a scanning electron microscope image of the cellulose-DNA hydrogel prepared in Comparative Example 1 of the present invention;

[0033] Figure 3 is a scanning electron microscope image of the cellulose-DNA hydrogel prepared in Comparative Example 2 of the present invention;

[0034] Figure 4 This is a Michaelis-Menten kinetic diagram of the enzyme-assembled hydrogel and free laccase obtained after immobilizing laccase in Example 3 of the present invention. DETAILED DESCRIPTION

[0035] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0036] The terms used in the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The singular forms "a", "the" and "the" used in the present invention and the appended claims are also intended to include plural forms unless the context clearly indicates other meanings. It should also be understood that the term "and / or" used herein refers to and includes any or all possible combinations of one or more associated listed items.

[0037] It should be understood that although the terms "first", "second", "third", etc. may be used to describe various information in the present invention, such information should not be limited to these terms. These terms are only used to distinguish the same type of information from each other. For example, without departing from the scope of the present invention, the first information may also be referred to as the second information, and similarly, the second information may also be referred to as the first information. Thus, the features defined as "first" and "second" may explicitly or implicitly include one or more of the features. In the description of the present invention, the meaning of "multiple" is two or more, unless otherwise clearly and specifically defined.

[0038] The present invention provides a method for preparing a cellulose-DNA hydrogel, which comprises the following steps:

[0039] Add DNA and 1,4-butanediol diglycidyl ether into the cellulose solution and stir thoroughly to obtain a doped mixed solution;

[0040] The mixed solution is centrifuged to degas, and then poured into a mold to be formed to obtain a preformed hydrogel;

[0041] The preformed hydrogel is immersed in an ethanol solution and then washed with deionized water to obtain a cellulose-DNA hydrogel.

[0042] The preparation method provided in this embodiment uses DNA, 1,4-butanediol diglycidyl ether and cellulose solution as raw materials, wherein:

[0043] Cellulose solution is the basic material, which is the main component of preparing hydrogel and provides the basic skeleton structure of hydrogel. In addition, cellulose molecules contain a large number of hydroxyl groups, which can interact with DNA and cross-linker 1,4-butanediol diglycidyl ether to help form a stable hydrogel network.

[0044] DNA is mainly used for the capture of POPs, which is beneficial for the efficient removal and degradation of POPs on the subsequent enzyme-fixed cellulose-based hydrogel. Specifically: DNA molecules have a long chain structure, which can provide a large amount of surface area in the hydrogel, increasing the chance of contact with POPs, thereby improving the capture efficiency. The base pairs in the NA molecule can form hydrogen bonds with the POPs molecules; the phosphate backbone of the DNA molecule carries a negative charge, while many POPs molecules such as polycyclic aromatic hydrocarbons (PAHs) carry a positive charge or functional groups that can form a positive charge. This interaction between charges helps DNA molecules capture positively charged POPs. The base pairs in the DNA molecule (such as adenine and thymine) can interact with the aromatic rings in the POPs molecule through π-π stacking, enhancing the adsorption capacity of POPs. The double helix structure of the DNA molecule can provide a spatial containment effect for the POPs molecules, especially for smaller pollutant molecules, this three-dimensional structure helps to improve the capture efficiency.

[0045] The role of 1,4-butanediol diglycidyl ether is: first, as a cross-linking agent, it connects DNA to the cellulose molecular chain to form a more stable three-dimensional network structure, allowing DNA to play the above role. Secondly, an etherification reaction occurs between 1,4-butanediol diglycidyl ether and DNA / cellulose to produce covalent bonds, which are stronger and more stable than the hydrogen bonds between cellulose, thereby increasing the mechanical strength of the prepared cellulose-DNA hydrogel; on the other hand, the chain length of 1,4-butanediol diglycidyl ether is significantly longer than the hydrogen bonds between cellulose, which can greatly stretch the cellulose chain, which is conducive to increasing the specific surface area, resulting in an increase in exposed sites, so that more enzymes can be fixed for efficient degradation and removal of POPs in real wastewater.

[0046] In summary, cellulose solution is used as the skeleton material of the hydrogel; 1,4-butanediol diglycidyl ether connects DNA to cellulose to form a stable three-dimensional network structure, so that DNA can stably and efficiently capture POPs, thereby improving the removal effect of POPs; 1,4-butanediol diglycidyl ether works together with DNA and cellulose to improve the mechanical strength and specific surface area of ​​the hydrogel system. As a preferred embodiment of this embodiment, the dosage ratio of DNA, 1,4-butanediol diglycidyl ether and cellulose is (3-4): (0.6-1.2): (3-4), more preferably (3-4): (0.8-1.0): (3-4), and most preferably 3:0.8:4.

[0047] The preparation methods of the above cellulose solution include but are not limited to: acid hydrolysis method, alkali solution treatment method, organic solvent method, enzyme treatment method, ultrasonic treatment method, ionic liquid method, etc. In this embodiment, the preparation method is as follows: under low temperature environment, cotton linters are added to an alkali solution containing urea and stirred to dissolve to obtain a cellulose solution. More preferably, the above cellulose solution can be prepared according to the following method:

[0048] A mixed aqueous solution containing urea and NaOH is prepared, wherein the content of urea in the mixed aqueous solution is 11-13 wt % and the content of NaOH is 6-8 wt %; the mixed aqueous solution is precooled at -12 DEG C for 3-5 hours; cotton linters are added to the precooled mixed aqueous solution, and the mixture is stirred until the cotton linters are dissolved to obtain a cellulose solution; and the obtained cellulose solution is stored at -12 DEG C.

[0049] Urea and NaOH work together to promote the dissolution of cellulose: Urea, as a co-solvent, can help cellulose dissolve more easily under alkaline conditions. NaOH is used to provide an alkaline environment, which helps to break the hydrogen bonds inside the cellulose, making its chains looser and easier to dissolve. Precooling the mixed aqueous solution is to lower the temperature of the solution, which helps to reduce the degradation of cellulose during the dissolution process and also helps to increase the solubility of cellulose. Cotton linters are the source of cellulose, which is composed of a large number of cellulose molecules. Cotton linters are added to the precooled mixed solution and stirred thoroughly at low temperature to make the cellulose evenly dispersed in the solution and begin to dissolve. After sufficient stirring and dissolution process, the cellulose molecules will be dispersed in the solution to form a uniform cellulose solution. Low temperature helps to maintain the uniformity of the solution and the dissolved state of the cellulose molecules. Storing the prepared cellulose solution at -12°C is conducive to maintaining its stability and preventing the cellulose from re-solidifying or precipitating.

[0050] The DNA is preferably salmon sperm DNA, which is widely available and low-cost, and has good biocompatibility and functionality.

[0051] In the above preparation method, DNA and 1,4-butanediol diglycidyl ether are first added to a cellulose solution and stirred thoroughly to obtain a doped mixed solution. The purpose of this step is to evenly disperse DNA and 1,4-butanediol diglycidyl ether in the cellulose solution, laying a foundation for subsequent grafting and cross-linking. In this step, the stirring speed is preferably 1000-1200 rpm / min.

[0052] After obtaining the mixed solution after doping, centrifugal degassing is required. The purpose of centrifugal degassing is to remove bubbles in the reaction solution and prevent uneven pores or defects in the hydrogel. The degassed solution is more conducive to forming a uniform and dense hydrogel structure. As a preferred embodiment of this embodiment, the speed of the centrifugal degassing treatment is 4000-6000rpm, and the time is 20-25min. More preferably, the speed of the centrifugal degassing treatment is 5000rpm, and the time is 20min.

[0053] The mixed solution after centrifugal degassing is transferred to a mold and kept for static molding. This step is used to provide a constant and favorable environment for the cross-linking reaction, promote the completion of the cross-linking reaction, and form a preformed hydrogel. In this step, the static molding temperature is 60-65°C and the time is 4-6 hours. More preferably, the static molding temperature is 45°C and the time is 6 hours.

[0054] After obtaining the preformed hydrogel, it is placed in an ethanol solution for soaking treatment. The purpose of the soaking treatment in this step is: on the one hand, to remove soluble impurities and residual solvents in the hydrogel; on the other hand, the soaking process further physically crosslinks it and enhances its structural stability. Preferably, the concentration of the ethanol solution used in this step is 75%, and the soaking treatment time is 6 to 8 hours.

[0055] After the soaking treatment, the step of washing with deionized water is continued to remove ethanol and other possible residual impurities, ensure the purity of the hydrogel, and balance its water content to achieve the final state before use. After washing with deionized water, the cellulose-DNA hydrogel can be obtained.

[0056] From the above content, it can be seen that the preparation method of cellulose-DNA hydrogel provided by the embodiment of the present invention has the following advantages:

[0057] The cellulose-DNA hydrogel prepared by this method uses 1,4-butanediol diglycidyl ether as a cross-linking agent to link DNA to the cellulose molecular chain to form a more stable three-dimensional network structure. On the one hand, the covalent bond produced by the etherification reaction between 1,4-butanediol diglycidyl ether and DNA / cellulose is stronger and more stable than the hydrogen bond between cellulose, thereby increasing the mechanical strength of the prepared cellulose-DNA hydrogel; on the other hand, the long chain structure of 1,4-butanediol diglycidyl ether can greatly stretch the cellulose chain, which is conducive to increasing the specific surface area and resulting in an increase in exposed sites, so that more enzymes can be fixed for efficient degradation and removal of POPs in real wastewater. Finally, the connected DNA, on the one hand, helps to further increase the specific surface area, and on the other hand, it has a good capture effect on POPs, which is conducive to the efficient removal and degradation of POPs on the subsequent enzyme-fixed cellulose-based hydrogel, and improves the removal rate of POPs. In addition, the preparation method is simple to operate and easy to realize industrial production and application.

[0058] Another embodiment of the present invention also provides the above-mentioned cellulose-DNA hydrogel, which is prepared according to the method of the above-mentioned embodiment. After the enzyme is assembled with the cellulose-DNA hydrogel as a carrier, the capture performance of POPs can be improved, thereby improving the contact efficiency between POPs and enzymes, and improving the removal effect of POPs in sewage. In addition, the cellulose-based hydrogel has good mechanical strength, which is conducive to its recycling. The cellulose-DNA hydrogel has the advantages of good removal effect of POPs and high mechanical strength.

[0059] Another embodiment of the present invention also provides a method for preparing an enzyme-assembled hydrogel for removing organic pollutants, which comprises the following steps:

[0060] Prepare cellulose-DNA hydrogel according to the method of the above example;

[0061] A buffer containing an enzyme is added to the cellulose-DNA hydrogel for a shaking treatment, and then washed with a buffer to obtain an enzyme-assembled hydrogel for removing organic pollutants.

[0062] The above-mentioned shaking treatment is used to shake and help the enzyme to be evenly distributed throughout the structure of the hydrogel, thereby improving the efficiency and uniformity of immobilization. Subsequent buffer washing is used to remove unfixed laccase and impurities to ensure the purity of the final product and the activity of the enzyme. The conditions for the shaking treatment are preferably as follows: a rotation speed of 160 to 200 rpm, a temperature of 22 to 26° C., and a time of 20 to 28 hours.

[0063] The above enzyme is preferably laccase, which can catalyze and oxidize a variety of POPs, including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides, etc. These compounds are usually difficult to remove by traditional biodegradation methods; in addition, laccase also has the advantages of high catalytic activity, environmental friendliness and strong adaptability; most importantly, laccase can be mass-produced by microbial fermentation and other methods, with relatively low cost. The buffer of the above enzyme is a phosphate buffer of laccase.

[0064] Correspondingly, another embodiment of the present invention also provides an enzyme-assembled hydrogel of organic pollutants, which is prepared according to the above method.

[0065] The preparation methods and corresponding products provided in the above embodiments also have the advantages of simple operation, good product mechanical properties, and high POPs removal efficiency.

[0066] The technical solution of the present invention is further described below in conjunction with specific embodiments:

[0067] The cellulose content of the cotton linters in the following examples is greater than 99%.

[0068] Example 1

[0069] (1) Preparation of cellulose solution: First, prepare 96 g of a mixed aqueous solution containing urea (12 wt %) and NaOH (7 wt %), and precool it at -12°C for 4 hours; then add 4 g of cotton linter, stir and dissolve it thoroughly to obtain a cellulose solution, which is stored at -12°C.

[0070] (2) Uniform doping of DNA and cross-linking agent (1,4-butanediol diglycidyl ether): 3 g of DNA from salmon sperm and 0.9 g of cross-linking agent (1,4-butanediol diglycidyl ether) were added to the cellulose solution of step 1 and stirred thoroughly to obtain a doped mixed solution.

[0071] (3) Synthesis of cellulose-DNA hydrogel: The mixed solution obtained in step 2 was degassed by centrifugation in a centrifuge (5000 rpm) for 20 minutes, then poured into a mold and kept at 60°C for 6 hours to form a hydrogel; after being taken out of the mold, it was immersed in a 75% ethanol solution for 12 hours, and finally washed with deionized water for 3 days to remove excess impurities to obtain a cellulose-DNA hydrogel.

[0072] Example 2

[0073] (1) Preparation of cellulose solution: the same as in Example 1.

[0074] (2) Uniform doping of DNA and cross-linking agent (1,4-butanediol diglycidyl ether): 3 g of DNA from salmon sperm and 1.0 g of cross-linking agent (1,4-butanediol diglycidyl ether) were added to the cellulose solution of step 1 and stirred thoroughly to obtain a doped mixed solution.

[0075] (3) Synthesis of cellulose-DNA hydrogel: the same as in Example 1.

[0076] Example 3

[0077] (1) Preparation of cellulose solution: the same as in Example 1.

[0078] (2) Uniform doping of DNA and cross-linking agent (1,4-butanediol diglycidyl ether): 3 g of DNA from salmon sperm and 0.8 g of cross-linking agent (1,4-butanediol diglycidyl ether) were added to the cellulose solution of step 1 and stirred thoroughly to obtain a doped mixed solution.

[0079] (3) Synthesis of cellulose-DNA hydrogel: The mixed solution obtained in step 2 was degassed by centrifugation in a centrifuge (5000 rpm) for 15 minutes, then poured into a mold and kept at 60°C for 5 hours to form a hydrogel; after being taken out of the mold, it was immersed in a 75% ethanol solution for 6 hours, and finally washed with deionized water for 3 days to remove excess impurities to obtain a cellulose-DNA hydrogel.

[0080] Comparative Example 1 (Example 4, without crosslinking agent)

[0081] The only difference from Example 3 is that no cross-linking agent (1,4-butanediol diglycidyl ether) is added in step 2, and the rest is the same as Example 3.

[0082] Comparative Example 2 (Example 5, more crosslinking agent)

[0083] The only difference from Example 3 is that the amount of the cross-linking agent (1,4-butanediol diglycidyl ether) in step 2 is changed to 1.6 g, and the rest is the same as Example 3.

[0084] Comparative Example 3 (Example 6, without DNA)

[0085] The only difference from Example 3 is that no DNA is added in step 2, and the rest is the same as Example 3.

[0086] The cellulose-DNA hydrogel samples prepared in Examples 1-3 and Comparative Examples 1-3 were used to conduct the following experiment.

[0087]

Specific surface area measurement

[0088] The N2 adsorption-desorption isotherms of cellulose-DNA hydrogel were obtained using a surface area and porosity analyzer, and the specific surface area (m 2 The measurement results are listed in Table 1.

[0089]

Compression strength test

[0090] A rectangular cellulose-DNA hydrogel with a length, width and height of 10 mm, 10 mm and 20 mm respectively was placed on an electronic universal testing machine to measure its compressive strength (Mpa). The test results are listed in Table 1.

[0091] Table 1 Specific surface area and compressive strength test results of cellulose-DNA hydrogel

[0092]

[0093]

[0094] The cellulose-DNA hydrogel samples prepared in Examples 1-3 and Comparative Examples 1-3 were immobilized with laccase according to the following method to obtain the corresponding enzyme-assembled hydrogels:

[0095] Laccase was dissolved in 40 mL of KH2PO4-prepared phosphate buffer (pH 6) to prepare a 2.0 mg / mL laccase solution, and then 40 mg of cellulose-DNA hydrogel sample was added and shaken at 180 rpm and 25°C for 24 h to fully fix the laccase, followed by thorough washing with phosphate buffer to obtain the enzyme-assembled hydrogel.

[0096] The prepared enzyme-assembled hydrogel was tested as follows:

[0097]

Determination of laccase immobilization amount

[0098] The enzyme fixation amount (Q, mg / g) is calculated according to the formula Q = [(C0-C e )×V-C1×V1] / m, where C0 and C e are the concentrations of laccase in the solution before and after fixation (mg / L), respectively; V and m represent the volume of the solution (L) and the mass of the cellulose-DNA hydrogel (g), respectively; C1 and V1 represent the concentration of laccase in the cleaning solution (mg / L) and the volume (L), respectively. The results are listed in Table 2.

[0099]

Determination of total removal efficiency (adsorption and degradation) of ∑16PAHs in real wastewater

[0100] Polycyclic aromatic hydrocarbons (PAHs) are a typical type of POPs that are frequently detected in large quantities in water environments. Therefore, PAHs were selected to represent POPs for removal efficiency determination. Real wastewater was taken from the workshop wastewater of a coal chemical plant in Ningxia. Laccase-fixed cellulose-DNA hydrogel was used to remove 16 PAHs that are prioritized by the US Environmental Protection Agency in real wastewater. The 16 PAHs include naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, benzo[a]anthracene, Benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenzo[a,h]anthracene, benzo[g,h,i]perylene and indeno[1,2,3-c,d]pyrene. Take 15 mL of wastewater sample, then add laccase immobilized cellulose-DNA hydrogel in experiment 1, shake at 180 rpm and 25 °C for 48 hours, and then use the formula R = (C0-C e ) / C0×100%to calculate the total removal efficiency (adsorption and degradation) of ∑16PAHs, where C0 and C e are the sum of the concentrations of the 16 PAHs in the solution before and after removal (mg / L). The total removal rate of ∑16PAHs includes the adsorption and degradation of ∑16PAHs by the enzyme-assembled hydrogel, because it has both adsorption and capture of ∑16PAHs and enzyme degradation. The measurement results are listed in Table 3.

[0101]

Determination of degradation efficiency of ∑16PAHs in real wastewater

[0102] The inactivated enzyme-assembled hydrogel was used as a control to calculate the degradation efficiency of ∑16PAHs. Since the inactivated laccase-immobilized cellulose-DNA hydrogel only adsorbed ∑16PAHs, the degradation efficiency of ∑16PAHs (D, %) can be calculated according to the formula D = R - (C 01 -C e1 ) / C 01 × 100%, where R represents the total removal rate (adsorption and degradation) of ∑16PAHs by laccase-immobilized cellulose-DNA hydrogel; C 01 and C e1 The sum of the concentrations of 16 PAHs in the solution before and after adsorption by inactivated laccase-immobilized cellulose-DNA hydrogel (mg / L) is respectively. The experimental procedure of adsorption and removal of ∑16PAHs by inactivated laccase-immobilized cellulose-DNA hydrogel is consistent with that in “Determination of the total removal efficiency (adsorption and degradation) of ∑16PAHs in real wastewater”. The results are listed in Table 3.

[0103]

Reusability determination

[0104] The enzyme-assembled hydrogel was added to 15 mL of wastewater sample, shaken at 180 rpm and 25 °C for 48 hours, washed with phosphate buffer for 12 hours, and added to 15 mL of wastewater sample again for removal. After this process was repeated 7 times, the total removal efficiency and degradation efficiency of ∑16PAHs were calculated. The measurement results are listed in Table 3.

[0105] Table 2 Test results of laccase immobilization by enzyme-assembled hydrogel

[0106] Experimental Project Laccase immobilization amount (Q, mg / g) Example 1 922.9 Example 2 920.5 Example 3 937.3 Comparative Example 1 208.7 Comparative Example 2 805.8 Comparative Example 3 849.4

[0107] Table 3 Test results of enzyme-assembled hydrogel for POPs removal effect and reusability

[0108]

[0109] From the above test results, we can see that:

[0110] 1. The cellulose-DNA hydrogel prepared in the embodiment of the present invention has a high specific surface area, a large amount of laccase immobilization, and high mechanical strength. The corresponding enzyme-assembled hydrogel has excellent performance in the removal and degradation of POPs in real wastewater, high reusability, and is very practical in the field of green water treatment.

[0111] 2. According to the test results of Comparative Example 1, the crosslinking agent 1,4-butanediol diglycidyl ether can stretch the cellulose chain and increase its specific surface area on the one hand, and the etherification reaction with cellulose / DNA can produce strong and stable covalent bonds on the other hand, thereby increasing its mechanical strength; after the doping of the crosslinking agent is removed, the specific surface area, laccase fixation amount, and compression strength of the cellulose-DNA hydrogel are significantly reduced. Therefore, in this example, after the laccase is fixed, the total removal efficiency and degradation efficiency of ∑16PAHs in real wastewater are only 19.45% and 11.04%, respectively; after repeated use for 7 times, the total removal efficiency of ∑16PAHs and the degradation efficiency of ∑16PAHs are only 10.44% and 4.83%, respectively, and the reusability is greatly reduced.

[0112] 2. According to the test results of Comparative Example 2, the doping of excessive (1.6 g) cross-linking agent causes the cross-linking between cellulose and DNA to produce agglomeration, resulting in the specific surface area of ​​cellulose-DNA hydrogel and the amount of laccase fixed to decrease to 118.4 m2 / g and 805.8 mg / g; therefore, in this example, after the laccase was fixed, the total removal efficiency and degradation efficiency of ∑16PAHs in real wastewater were reduced to 75.64% and 65.09%, respectively.

[0113] 3. According to the test results of Comparative Example 3, since DNA is beneficial to the adsorption and pre-capture of POPs (PAHs) and is used for subsequent degradation on the enzyme-immobilized hydrogel, after removing the doped DNA, the total removal efficiency and degradation efficiency of ∑16PAHs in real wastewater after laccase fixation in this example were significantly reduced to 61.73% and 58.62%, respectively.

[0114] Figures 1 to 3 The scanning electron micrographs of the cellulose-DNA hydrogels prepared in Example 3, Comparative Example 1 and Comparative Example 2 are shown respectively. Figures 1 to 3 It can be seen that:

[0115] Since no crosslinking agent was added in Comparative Example 1 ( Figure 2 ), which cannot stretch the cellulose chain, making the pore structure on the surface of the cellulose-DNA hydrogel not rich enough, resulting in a decrease in its specific surface area and enzyme loading; at the same time, due to the addition of excessive cross-linking agent ( Figure 3 ), although the pore structure of cellulose-DNA hydrogel is richer than that of Comparative Example 1 without adding crosslinking agent, it is still not as good as that of Example 3 with adding appropriate amount of crosslinking agent ( Figure 1 ) have rich pore structures; this is consistent with the results of their specific surface areas and laccase immobilization amounts.

[0116] Figure 4 3 is the Michaelis-Menten kinetics of the enzyme-assembled hydrogel and free laccase obtained after immobilizing laccase in Example 3 of the present invention. It can be seen from the figure that the maximum action rate (V max ) is significantly larger than that of free laccase. In addition, the Michaelis constant (K m ) is smaller than free laccase, indicating a greater substrate affinity. These advantages make the enzyme-assembled hydrogel prepared in the embodiment of the present invention have excellent removal and degradation performance for POPs (PAHs) in real wastewater.

[0117] The embodiments of the present invention have been described above, and the above description is exemplary, not exhaustive, and is not limited to the disclosed embodiments. Many modifications and changes will be apparent to those of ordinary skill in the art without departing from the scope and spirit of the described embodiments. The selection of terms used herein is intended to best explain the principles of the embodiments, practical applications, or improvements to the technology in the market, or to enable other persons of ordinary skill in the art to understand the embodiments disclosed herein.

Claims

1. A method for preparing a cellulose-DNA hydrogel, characterized in that: include: Add DNA and 1,4-butanediol diglycidyl ether into the cellulose solution and stir thoroughly to obtain a doped mixed solution; The mixed solution is centrifuged to degas, and then poured into a mold to be formed to obtain a preformed hydrogel; The preformed hydrogel is immersed in an ethanol solution and then washed with deionized water to obtain a cellulose-DNA hydrogel.

2. The preparation method according to claim 1, characterized in that: The cellulose solution is prepared according to the following method: Under low temperature conditions, cotton linters are added into an alkaline solution containing urea and stirred to dissolve to obtain a cellulose solution.

3. The preparation method according to claim 1, characterized in that: The DNA is salmon sperm DNA.

4. The preparation method according to claim 1, characterized in that: The concentration of the ethanol solution used in the soaking treatment is 75%, and the soaking treatment time is 6 to 8 hours.

5. The preparation method according to claim 1, characterized in that: The rotation speed of the centrifugal degassing treatment is 4000-6000 rpm, and the time is 20-25 min; and / or, the temperature of the static molding is 60-65° C., and the time is 4-6 h.

6. The preparation method according to any one of claims 1 to 5, characterized in that: The dosage ratio of the DNA, 1,4-butanediol diglycidyl ether and cellulose is (3-4): (0.6-1.2): (3-4).

7. A cellulose-DNA hydrogel, characterized in that: It is prepared according to the method according to any one of claims 1 to 6.

8. A method for preparing an enzyme-assembled hydrogel for removing organic pollutants, characterized in that: include: Prepare cellulose-DNA hydrogel according to the method of any one of claims 1 to 6; A buffer containing an enzyme is added to the cellulose-DNA hydrogel for a shaking treatment, and then washed with a buffer to obtain an enzyme-assembled hydrogel for removing organic pollutants.

9. The preparation method according to claim 8, characterized in that: The enzyme is laccase.

10. An enzyme-assembled hydrogel for removing organic pollutants, characterized in that: It is prepared according to the method of claim 8 or 9.

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