Method for constructing antibacterial film interface by chemical bond crosslinking antibacterial peptide

Abstract

The invention discloses a method for constructing an antibacterial film interface by chemical bond cross-linked antibacterial peptide, and belongs to the field of drinking water purification and wastewater pollution treatment. The invention aims to solve the problems that the existing microorganisms cause serious pollution to the membrane, the requirement on the operation environment in the membrane modification process is high, and antibacterial active substances on the surface of the membrane are released. An ultrathin antibacterial peptide modified layer is constructed on the surface of the membrane through alkali-activated air oxidation-Michael addition / Schiff base reaction, the stability of antibacterial active substances on the surface of the membrane is maintained through chemical crosslinking of plant polyphenol, and long-acting biological pollution resistance is achieved. The invention provides a method for constructing an anti-pollution membrane interface by chemical bond crosslinking antibacterial peptide. According to the method, the structure of the base membrane is not damaged, the method is simple, adjustable, short in period and low in cost, the thickness of the coating layer is nanoscale, the ultrathin coating layer relieves reduction of modification on the water passing capacity of the membrane, the hydrophilicity and the antibacterial property are remarkably improved, extra pressurization energy consumption is not needed, and filtering can be achieved through gravity water head driving.

Description

Technical Field

[0001] The present invention belongs to the field of drinking water purification and wastewater pollution treatment, and particularly relates to a method for constructing an antibacterial membrane interface by chemically cross-linking antibacterial peptides. Background Art

[0002] Membrane separation technology is an efficient physical treatment method with low energy consumption, small floor area, simple operation, and easy coupling with other processes. It has been studied and applied in the fields of water treatment, food and beverage processing, industrial gas treatment, and pharmaceutical and biomedical applications.

[0003] During the filtration process, pollutants will deposit on the membrane surface or block the membrane pores, causing membrane fouling, resulting in reduced membrane flux, deteriorated effluent quality, and increased operating energy consumption. Membrane biofouling is a major research hotspot in membrane fouling problems. Microorganisms will not only deposit, adhere, and grow on the membrane surface, but also produce pollutants such as extracellular polymers during metabolism, thereby forming biofilms, leading to a significant reduction in membrane flux and a significant decrease in separation performance. In gravity-driven membrane processes, biofouling will cause bio-leakage of macroporous membranes, affecting the effluent quality.

[0004] Conventional hydraulic cleaning methods are ineffective when biofouling is severe. To avoid severe biofouling, membrane modules need to be cleaned and maintained frequently, resulting in increased operating costs. Currently, people prepare anti-biofouling membrane materials by modifying or blending the membrane surface with antibacterial active materials such as metals or metal nanoparticles, metal oxides, guanidine, quaternary ammonium compounds, and antibiotics. The preparation of common nanomaterials is difficult and costly. During use, the release of active materials not only reduces the antibacterial effect, but also enters the effluent, affecting the effluent quality. It may also diffuse on the membrane surface and affect the composition of the microbial community in gravity-driven membranes. In addition, some materials will affect the hydrophilicity, hydrophobicity, roughness, and other properties of the membrane itself, leading to the adsorption of other pollutants such as humic acid and proteins, and the control effect of membrane fouling is not ideal from the perspective of practical applications. Therefore, how to prepare stable, non-toxic, and anti-biofouling membrane materials through mild and convenient modification methods is the research focus in the field of membrane separation. Summary of the Invention

[0005] The purpose of the present invention is to solve the problems of serious membrane fouling caused by existing microorganisms, high requirements for the operating environment during membrane modification, and the release of antibacterial active substances on the membrane surface, and to provide a method for constructing an antibacterial membrane interface by chemically cross-linking antibacterial peptides.

[0006] The present invention constructs an ultrathin antibacterial peptide modified layer on the membrane surface through alkali-activated air oxidation-Michael addition / Schiff base reaction, and maintains the stability of antibacterial active substances on the membrane surface and realizes long-term anti-biofouling performance through chemical cross-linking of plant polyphenols, providing a method for constructing an anti-fouling membrane interface by chemically cross-linking antibacterial peptides.

[0007] A method for constructing an antibacterial film interface with chemically bonded cross-linked antibacterial peptides is completed according to the following steps:

[0008] I. Pretreatment of the base film:

[0009] Clean and activate the base film to remove contaminants, and obtain the pretreated base film;

[0010] II. Preparation of the prefabricated solution:

[0011] ①. Adjust the pH value of deionized water to alkaline, then add polyphenols and stir until completely dissolved to obtain a polyphenol prefabricated solution;

[0012] ②. Adjust the pH value of deionized water to alkaline, then add antibacterial peptides and stir until completely dissolved to obtain an antibacterial peptide prefabricated solution;

[0013] III. Immerse the pretreated base film in the polyphenol prefabricated solution and shake for a period of time. After taking it out, dry the surface of the base film, and then immerse the base film in the antibacterial peptide prefabricated solution and shake for a period of time;

[0014] IV. Repeat step III for 0 to 5 times, then wash with deionized water, and finally store in deionized water to obtain a chemically bonded cross-linked antibacterial peptide modified film, thus completing a method for constructing an antibacterial film interface with chemically bonded cross-linked antibacterial peptides.

[0015] Principle of the present invention:

[0016] The present invention innovatively induces the air oxidation of polyphenols by alkali to generate quinone free radicals and undergoes Michael addition / Schiff base reaction with the amino groups of antibacterial peptides, and then forms a stable chemically bonded cross-linked antibacterial peptide antibacterial modified layer by layer-by-layer self-assembly; changing the coating layer number can flexibly change the thickness of the modified layer. Plant polyphenols are a type of commonly used green antibacterial material. After chemical covalent cross-linking with antibacterial peptides, a coating with a nanoparticle structure is formed on the film surface, significantly improving the hydrophilicity and antibacterial ability of the modified film, and having good application prospects.

[0017] Advantages of the present invention:

[0018] 1. The present invention has low requirements for the material of the base film during the preparation of the film, and has a large applicable range of film modification;

[0019] 2. The preparation method of the present invention does not damage the structure of the base film, is simple, adjustable, has a short cycle, and low cost;

[0020] 3. The antibacterial peptide modified film prepared by the present invention improves the hydrophilicity and antibacterial property of the modified film;

[0021] 4. The coating layer thickness of the modified film prepared by the present invention is nanoscale, and the ultra-thin coating layer alleviates the reduction of the water passing ability of the film caused by the modification;

[0022] 5. In the present invention, the prepared modified membrane does not require additional pressure energy consumption and can achieve filtration by utilizing the gravity head. Description of the Drawings

[0023] Figure 1 Scanning electron microscope surface image of the chemically crosslinked antibacterial peptide modified membrane prepared in Example 1;

[0024] Figure 2 Scanning electron microscope cross-sectional image of the chemically crosslinked antibacterial peptide modified membrane prepared in Example 2;

[0025] Figure 3 Scanning electron microscope cross-sectional image of the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3;

[0026] Figure 4 Contact antibacterial effects of the PVDF base membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 against Escherichia coli and Staphylococcus aureus. In the figure, A is the PVDF base membrane and B is the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3;

[0027] Figure 5 Anti-pollution effects of the PVDF base membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 against Escherichia coli and Staphylococcus aureus during the filtration process;

[0028] Figure 6 Anti-biofouling effects of the PVDF base membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 during the river water filtration process;

[0029] Figure 7 Change trend of the membrane flux of the chemically crosslinked antibacterial peptide modified membrane and the base membrane prepared in Example 3 during the pollution-cleaning cycle test in the Escherichia coli dispersion. Detailed Embodiments

[0030] Detailed Embodiment 1: A method for constructing an antibacterial membrane interface with chemically crosslinked antibacterial peptides is completed according to the following steps:

[0031] I. Pretreatment of the base membrane:

[0032] Clean and activate the base membrane to remove contaminants, and obtain the pretreated base membrane;

[0033] II. Preparation of the prefabricated solution:

[0034] ①. Adjust the pH value of deionized water to alkaline, then add polyphenol and stir until completely dissolved to obtain a polyphenol prefabricated solution;

[0035] ②. Adjust the pH value of deionized water to alkaline, then add antibacterial peptide and stir until completely dissolved to obtain an antibacterial peptide prefabricated solution;

[0036] III. Immerse the pretreated base membrane in the polyphenol pre-prepared solution and shake for a period of time. After taking it out, dry the surface of the base membrane, and then immerse the base membrane in the antimicrobial peptide pre-prepared solution and shake for a period of time;

[0037] IV. Repeat step III 0 - 5 times, then wash with deionized water, and finally store in deionized water to obtain a chemically cross-linked antimicrobial peptide modified membrane, thus completing a method for constructing an antimicrobial membrane interface with chemically cross-linked antimicrobial peptides.

[0038] Specific Embodiment 2: The difference between this embodiment and Specific Embodiment 1 is that the method for cleaning and activating in step I is as follows: First, immerse the base membrane in isopropanol for 1 h - 2 h, then take it out, and then rinse the isopropanol on the surface of the base membrane with deionized water. Finally, place it in deionized water and soak for 24 h to obtain the pretreated base membrane. Other steps are the same as those in Specific Embodiment 1.

[0039] Specific Embodiment 3: The difference between this embodiment and one of Specific Embodiments 1 or 2 is that the base membrane described in step I is a polysulfone membrane, a polyethersulfone membrane, a polyvinyl chloride membrane, or a polyvinylidene fluoride membrane. Other steps are the same as those in Specific Embodiment 1 or 2.

[0040] Specific Embodiment 4: The difference between this embodiment and one of Specific Embodiments 1 to 3 is that in step II ①, the pH value of deionized water is adjusted to 8 - 8.5 with a 1 mol / L sodium hydroxide solution. Other steps are the same as those in Specific Embodiments 1 to 3.

[0041] Specific Embodiment 5: The difference between this embodiment and one of Specific Embodiments 1 to 4 is that the polyphenol described in step II ① is one or several of epigallocatechin gallate, epicatechin gallate, epigallocatechin, epicatechin, tannic acid, gallic acid, and pyrogallol. Other steps are the same as those in Specific Embodiments 1 to 4.

[0042] Specific Embodiment 6: The difference between this embodiment and one of Specific Embodiments 1 to 5 is that the concentration of the polyphenol pre-prepared solution described in step II ① is 0.05 mg / mL - 0.2 mg / mL. Other steps are the same as those in Specific Embodiments 1 to 5.

[0043] Specific Embodiment 7: The difference between this embodiment and one of Specific Embodiments 1 to 6 is that in step II ②, the pH value of deionized water is adjusted to 8 - 8.5 with a 1 mol / L sodium hydroxide solution. Other steps are the same as those in Specific Embodiments 1 to 6.

[0044] Embodiment VIII: The difference between this embodiment and any one of Embodiments I - VII is that: the antimicrobial peptide described in Step 2② is polylysine; the concentration of the antimicrobial peptide pre - solution described in Step 2② is 0.05 mg / mL to 0.2 mg / mL. Other steps are the same as those in Embodiments I - VII.

[0045] Embodiment IX: The difference between this embodiment and any one of Embodiments I - VIII is that: the oscillation time described in Step 3 is 10 min to 60 min; the oscillation speed is 70 r / min to 200 r / min. Other steps are the same as those in Embodiments I - VIII.

[0046] Embodiment X: The difference between this embodiment and any one of Embodiments I - IX is that: the number of cleaning times described in Step 4 is 3 to 5 times; the thickness of the chemically - crosslinked antimicrobial peptide on the chemically - crosslinked antimicrobial peptide - modified membrane described in Step 4 is 5 nm to 100 nm. Other steps are the same as those in Embodiments I - IX.

[0047] The following examples are used to verify the beneficial effects of the present invention:

[0048] Example 1: A method for constructing an antimicrobial membrane interface by chemically cross - linking antimicrobial peptides is completed according to the following steps:

[0049] I. Pretreatment of the base membrane:

[0050] First, immerse the base membrane in isopropanol for 1 h, then take it out, and then rinse the isopropanol on the surface of the base membrane with deionized water. Finally, place it in deionized water and soak for 24 h to obtain the pretreated base membrane;

[0051] The base membrane described in Step I is a polyvinylidene fluoride membrane;

[0052] II. Preparation of the pre - solution:

[0053] ①. Use a 1 mol / L sodium hydroxide solution to adjust the pH value of deionized water to 8.5, and then add polyphenol and stir until completely dissolved to obtain a polyphenol pre - solution;

[0054] The polyphenol described in Step 2① is tannic acid;

[0055] The concentration of the polyphenol pre - solution described in Step 2① is 0.1 mg / mL;

[0056] ②. Use a 1 mol / L sodium hydroxide solution to adjust the pH value of deionized water to 8.5, and then add the antimicrobial peptide and stir until completely dissolved to obtain an antimicrobial peptide pre - solution;

[0057] The antimicrobial peptide described in Step 2② is polylysine;

[0058] The concentration of the antimicrobial peptide prefabricated solution described in Step 2② is 0.1 mg / mL.

[0059] III. Immerse the pretreated base membrane in the polyphenol prefabricated solution and shake for 30 min. After taking it out, dry the surface of the base membrane, and then immerse the base membrane in the antimicrobial peptide prefabricated solution and shake for 30 min.

[0060] IV. Repeat Step III 0 times, then wash it 3 times with deionized water, and finally store it in deionized water to obtain a chemically crosslinked antimicrobial peptide modified membrane, thus completing a method for constructing an antimicrobial membrane interface with chemically crosslinked antimicrobial peptides.

[0061] Example 2: The difference between this example and Example 1 is that in Step IV, Step III is repeated 2 times. Other steps and parameters are the same as those in Example 1.

[0062] Example 3: The difference between this example and Example 1 is that in Step IV, Step III is repeated 4 times. Other steps and parameters are the same as those in Example 1.

[0063] In Example 1, the thickness of the chemically crosslinked antimicrobial peptide on the chemically crosslinked antimicrobial peptide modified membrane described in Step IV is 5.3 nm;

[0064] In Example 2, the thickness of the chemically crosslinked antimicrobial peptide on the chemically crosslinked antimicrobial peptide modified membrane described in Step IV is 13.5 nm;

[0065] In Example 3, the thickness of the chemically crosslinked antimicrobial peptide on the chemically crosslinked antimicrobial peptide modified membrane described in Step IV is 50.2 nm.

[0066] Figure 1 is the scanning electron microscope surface image of the chemically crosslinked antimicrobial peptide modified membrane prepared in Example 1;

[0067] Figure 2 is the scanning electron microscope cross-sectional image of the chemically crosslinked antimicrobial peptide modified membrane prepared in Example 2;

[0068] Figure 3 is the scanning electron microscope cross-sectional image of the chemically crosslinked antimicrobial peptide modified membrane prepared in Example 3;

[0069] From Figure 2 and Figure 3 it can be seen that the chemically crosslinked antimicrobial peptide modified layer with a nanoscale thickness is continuously coated on the surface of the polyvinylidene fluoride membrane and there is no obvious demarcation line with the membrane surface, and the tight combination of the two ensures the stability of the modified membrane.

[0070] Figure 4The contact antibacterial effects of the PVDF-based membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 against Escherichia coli and Staphylococcus aureus. In the figure, A is the PVDF-based membrane, and B is the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3;

[0071] From Figure 4 it can be seen that: the absence of an inhibition zone indicates that the antibacterial active substance has not been released, and the number of colonies in the part in contact with the membrane has decreased significantly, indicating that the modified membrane has played a good contact antibacterial role.

[0072] Figure 5 The anti-pollution effects of the PVDF-based membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 against Escherichia coli and Staphylococcus aureus during the filtration process;

[0073] From Figure 5 it can be seen that: after filtering the bacterial suspension, the bacteria on the surface of the modified membrane have decreased significantly, and the integrity of the bacteria has been damaged, indicating that the modified membrane can prevent bacteria from adhering and sterilize at the same time during the filtration process.

[0074] Figure 6 The anti-biofouling effects of the PVDF-based membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 during the filtration of river water;

[0075] The bacterial coverage rate of the PVDF-based membrane is 16.6%, while that of the modified membrane drops to 1.8%, and the activity decreases significantly, demonstrating that the modified membrane can reduce bacterial adhesion and play an antibacterial role during the actual surface water filtration process.

[0076] A membrane filtration device constructed with a Millipore 8200 ultrafiltration cup produced by Millipore Corporation of the United States as the core was used to evaluate the anti-pollution performance of the membrane. Using an Escherichia coli dispersion as the membrane pollution test solution, a pollution-cleaning cycle test was adopted to characterize the anti-pollution performance and its stability of the membrane through the change trend of the membrane flux. See Figure 7 as shown Figure 7 for the change trend of the membrane flux during the pollution-cleaning cycle test;

[0077] A pollutant filtration experiment was carried out on the original membrane (PVDF-based membrane) and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3; the configuration steps of the pollutant were as follows: newly activated Escherichia coli was dispersed in physiological saline and stirred evenly to prepare an Escherichia coli pollution solution with a concentration of 2×10 6 CFU / mL, and a filtration experiment was carried out with a 10 cm gravity head. The total number of colonies in the effluent of the solution filtered through the original membrane and the chemically crosslinked antibacterial peptide modified membrane prepared in Example 3 were 205 CFU / mL and 0 CFU / mL respectively, and the stable fluxes were 101.7 L·m -2 ·h -1 and 131.8 L·m-2 ·h -1 ; It can be judged therefrom that the ability of the chemically crosslinked antimicrobial peptide modified membrane to resist biological fouling has been enhanced.

Claims

1. A method for constructing an antibacterial membrane interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The method is completed according to the following steps: I. Pretreatment of the base membrane: Clean and activate the base membrane to remove contaminants, obtaining the pretreated base membrane; II. Preparation of the prefabricated solution: ①. Adjust the pH value of deionized water to alkaline, then add polyphenols and stir until completely dissolved to obtain the polyphenol prefabricated solution; ②. Adjust the pH value of deionized water to alkaline, then add antimicrobial peptides and stir until completely dissolved to obtain the antimicrobial peptide prefabricated solution; III. Immerse the pretreated base membrane in the polyphenol prefabricated solution and shake for a period of time, take out and dry the surface of the base membrane, and then immerse the base membrane in the antimicrobial peptide prefabricated solution and shake for a period of time; IV. Repeat step III 0 to 5 times, then wash with deionized water, and finally store in deionized water to obtain the chemically cross-linked antimicrobial peptide modified membrane, thus completing a method for constructing an antimicrobial membrane interface with chemically cross-linked antimicrobial peptides.

2. The method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide according to claim 1, characterized in that The method of cleaning and activation described in step I is as follows: First, immerse the base membrane in isopropanol for 1 h to 2 h, then take it out, and then rinse the isopropanol on the surface of the base membrane with deionized water, and finally place it in deionized water and soak for 24 h to obtain the pretreated base membrane.

3. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The base membrane described in step I is a polysulfone membrane, a polyethersulfone membrane, a polyvinyl chloride membrane or a polyvinylidene fluoride membrane.

4. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that In step II①, the pH value of deionized water is adjusted to 8 to 8.5 using a sodium hydroxide solution with a concentration of 1 mol / L.

5. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The polyphenols described in step II① are one or more of epigallocatechin gallate, epicatechin gallate, epigallocatechin, epicatechin, tannic acid, gallic acid and pyrogallol.

6. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The concentration of the polyphenol prefabricated solution described in step II① is 0.05 mg / mL to 0.2 mg / mL.

7. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that In step II②, the pH value of deionized water is adjusted to 8 to 8.5 using a sodium hydroxide solution with a concentration of 1 mol / L.

8. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The antimicrobial peptide described in step II② is polylysine; the concentration of the antimicrobial peptide prefabricated solution described in step II② is 0.05 mg / mL to 0.2 mg / mL.

9. A method for constructing an antibacterial film interface with a chemically bonded cross-linked antibacterial peptide, characterized in that The shaking time described in step III is 10 min to 60 min; the shaking speed is 70 r / min to 200 r / min.

10. The method for constructing an antibacterial film interface by chemically bonding cross-linked antibacterial peptides according to claim 1, wherein The number of washing times described in step IV is 3 to 5 times; the thickness of the chemically cross-linked antimicrobial peptide on the chemically cross-linked antimicrobial peptide modified membrane described in step IV is 5 nm to 100 nm.

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