An antibacterial polymer selectively eliminating gram-negative bacteria, and a preparation method and application thereof

Abstract

The application provides an antibacterial polymer selectively removing gram-negative bacteria and a preparation method and application thereof, and solves the problem that the existing antibacterial polymer with multiple advantages still cannot effectively remove gram-negative bacteria in a specific manner. m HF n The chemical formula of the antibacterial polymer is PD m HF n , an amine-containing monomer 2-(dimethylamino)ethyl methacrylate (DMAEMA) is used as a cation block unit, and a fluorine-containing monomer 2,2,3,3,4,4,4-heptafluorobutyl methacrylate (HFBMA) is used as a hydrophobic group block unit, wherein the ratio of n to m represents the molar ratio of the monomers HFBMA and DMAEMA, and 0.1≤n / m≤0.2.

Description

Technical Field

[0001] This invention belongs to the field of biomedical engineering materials technology, specifically relating to an antibacterial polymer that selectively eliminates Gram-negative bacteria, its preparation method, and its application. Background Technology

[0002] The overuse and inappropriate use of antibiotics has accelerated the development of antibiotic resistance in bacteria, and the rapid spread of multidrug-resistant (MDR) bacteria in recent years has evolved into a global medical crisis. A group comprising two Gram-positive bacteria (E. faecalis, S. aureus) and four Gram-negative bacteria (K. pneumoniae, A. baumannii, P. aeruginosa, and Enterobacter species) is known as the infamous ESKAPE pathogen, particularly concerning due to their ability to evade antibiotic treatment and exhibit high rates of resistance. However, the development of new antibiotics lags far behind the rate at which bacteria develop resistance. Currently, some new antibiotics are undergoing clinical trials, showing potential in eradicating Gram-positive bacteria; however, no new class of antibiotics has been approved for the treatment of Gram-negative bacterial infections in over 50 years.

[0003] Carbapenem-resistant Gram-negative ESKAPE bacteria, along with extended-spectrum β-lactamase (ESBL) species such as *K. pneumonia* and *Enterobacter*, have been listed by the World Health Organization as a priority pathogen requiring urgent treatment in 2023. These drug-resistant Gram-negative ESKAPE pathogens can cause serious nosocomial infections, such as pneumonia, surgical site infections, urinary tract infections, and bloodstream infections. In the United States, these Gram-negative bacterial infections account for 70% of infections in intensive care units, and similar reports have emerged in other parts of the world. Therefore, there is an urgent need to develop novel antimicrobial agents that are less likely to induce resistance in order to combat MDR Gram-negative ESKAPE bacterial infections.

[0004] In recent years, antimicrobial peptides, leveraging their cationic amphiphilic structural properties, exert bactericidal activity through membrane perturbation, electrostatics, and hydrophobic interactions. As a novel antimicrobial agent that is less likely to induce bacterial resistance, they have gradually attracted increasing attention from researchers. However, antimicrobial peptides are limited in widespread application due to drawbacks such as complex production and purification processes, high production costs, difficulty in mass production, and easy decomposition and loss of activity. Therefore, antimicrobial polymers, with their multiple advantages including low cost, large-scale production capability, ease of synthesis, and controllable structure, have emerged and are expected to replace antimicrobial peptides as the next generation of antimicrobial agents. However, currently developed antimicrobial polymers still primarily target broad-spectrum bacteria. While killing pathogenic bacteria, they indiscriminately kill other bacteria, leading to widespread bacterial resistance, significantly weakening the therapeutic effect of drugs, and seriously threatening human health. Compared to Gram-positive bacterial infections, Gram-negative bacteria, due to their complex cell wall structure and highly active efflux pump system, are more difficult to treat. Therefore, developing novel antibacterial drugs that can selectively kill Gram-negative bacteria can effectively prevent the overuse of antibiotics, slow down the development of bacterial resistance, and is of great significance for the treatment of Gram-negative bacterial infections.

[0005] To this end, the research team of this invention has developed an antibacterial polymer that can selectively eliminate Gram-negative bacteria. Summary of the Invention

[0006] The purpose of this invention is to address the shortcomings of existing antimicrobial polymers with multiple advantages that still cannot specifically and effectively eliminate Gram-negative bacteria, and to provide an antimicrobial polymer that selectively eliminates Gram-negative bacteria, its preparation method, and its application; this antimicrobial polymer can not only selectively eliminate Gram-negative bacteria, but also delay the development of drug resistance.

[0007] To achieve the above objectives, the technical solution provided by this invention is:

[0008] An antibacterial polymer, characterized by its chemical formula PD m HF n The structural formula is as follows:

[0009]

[0010] The polymer uses amine-containing monomer 2-(dimethylamino)ethyl methacrylate (DMAEMA) as a cationic block unit and fluorine-containing monomer 2,2,3,3,4,4,4-heptafluorobutyl methacrylate (HFBMA) as a hydrophobic block unit. The ratio of n to m represents the molar ratio of monomers HFBMA and DMAEMA, and 0.1 ≤ n / m ≤ 0.2.

[0011] Furthermore, its chemical formula is PD.45 HF5, at this ratio, the antimicrobial polymer is more effective at eliminating Gram-negative bacteria.

[0012] This invention also provides a method for preparing the above-mentioned antibacterial polymer, characterized by comprising the following steps:

[0013] 1) Dissolve the initiator, chain transfer reagent, DMAEMA and HFBMA in an organic solvent and add them into a Schlenk reaction tube, stirring until completely dissolved;

[0014] 2) After the reaction system in step 1) undergoes at least three double-row tube freeze-pump-charge cycles, the atmosphere is replaced with an inert gas.

[0015] 3) Place the reaction system from step 2) in a constant-temperature stirring environment to carry out the reaction;

[0016] 4) After the reaction in step 3) is completed, the reaction product is allowed to settle and then vacuum dried to obtain the target antibacterial polymer, namely the fluorinated amphiphilic block copolymer PD. m HF n .

[0017] In short, the above preparation method involves synthesizing monomers DMAEMA and HFBMA using a reversible addition-fragmentation chain transfer (RAFT) polymerization method, followed by precipitation and vacuum drying of the reaction product to obtain the target product PD. m HF n .

[0018] Further, in step 1), the initiator is azobisisobutyronitrile, dimethyl azobisisobutyrate, benzoyl peroxide, or tert-butyl peroxide.

[0019] The chain transfer agent is 4-cyanopentanoic acid dithiobenzoic acid, dithiobenzoic acid cyanoisopropyl ester, or 2-[dodecylthio(thiocarbonyl)thio]-2-methylpropionic acid;

[0020] The organic solvent is dimethyl sulfoxide, N,N-dimethylformamide, acetonitrile, or dioxane.

[0021] Further, in step 1), the molar ratio of the initiator, chain transfer reagent, DMAEMA and HFBMA is 1:10:400:100-1:10:450:50, wherein the total concentration of the two monomers is 1 mmol / L.

[0022] Further, in step 2), the inert gas is nitrogen or argon.

[0023] Furthermore, in step 3), a constant temperature stirrer is used, the reaction temperature is 50-70℃, and the reaction time is 18-24h.

[0024] Furthermore, in step 4), the reaction product is precipitated by a poor solvent, which is a solvent that cannot dissolve the antibacterial polymer, such as n-hexane, petroleum ether, n-pentane, etc.

[0025] Furthermore, the present invention also provides the application of the above-mentioned antimicrobial polymer in the selective elimination of Gram-negative bacteria, and an antimicrobial agent based on this application for eliminating Gram-negative bacteria, the active ingredient of which is the aforementioned antimicrobial polymer PD. m HF n .

[0026] The concept and principle of this invention:

[0027] To address the lack of Gram selectivity in current antimicrobial polymers, this invention investigates the structure-activity relationship (SOR) of antimicrobial polymers. It reveals that this can be achieved by optimizing parameters such as polymer molecular weight, monomer arrangement, polymer charge, side groups, and hydrophilic / hydrophobic ratio. Furthermore, it demonstrates that balancing the ratio of cationic to hydrophobic units in the antimicrobial polymer structure is crucial for achieving high antimicrobial activity and low toxicity (hemolysis and cytotoxicity). This indicates that positive charge and hydrophilicity / hydrophobicity are important factors influencing the biomedical properties of antimicrobial polymers. Therefore, the key to designing novel antimicrobial polymers lies in the rational selection of cationic and hydrophobic monomers and their appropriate ratio. Unlike Gram-positive bacteria, lipopolysaccharide (LPS), as a unique component of the outer membrane of Gram-negative bacteria, can serve as a target for Gram selection. Thus, the key to developing antimicrobial polymers capable of killing Gram-negative bacteria also lies in the selection of cationic and hydrophobic monomers and their ratio.

[0028] Therefore, this invention screened cationic and hydrophobic monomers. Currently, the hydrophobic portion of antibacterial polymers is typically composed of various types of aliphatic (cyclic, linear, and branched) and aromatic groups. However, during the research process, this invention discovered that fluorinated groups exhibit superior hydrophobicity and oleophobicity, which is crucial for cell membrane penetration and enables a more efficient membrane disruption antibacterial mechanism. Simultaneously, fluorinated polymers show lower cytotoxicity and hemolytic activity, but no literature has yet demonstrated the antibacterial effects of fluorinated amphiphilic polymers. Therefore, considering the bacterial membrane disruption mechanism and leveraging their superior hydrophobicity and oleophobicity, the research team of this invention intends to use fluorinated groups as hydrophobic monomers to enhance antibacterial activity. Furthermore, during the screening process, this invention discovered that the amine-containing cationic monomer 2-(dimethylamino)ethyl methacrylate (DMAEMA) has a specific affinity for LPS from Gram-negative bacteria. Therefore, the research team of this invention intends to use DMAEMA as a cationic monomer. Fluorinated amphiphilic polymers were synthesized using DMAEMA and HFBMA as cationic and hydrophobic monomers, respectively, and their antibacterial activity was verified.

[0029] The advantages of this invention are:

[0030] 1. Based on the synergistic effect of the efficient membrane disruption mechanism of HFBMA and the LPS-specific affinity of DMAEMA, this invention designs a novel antibacterial polymer—the fluorinated amphiphilic polymer PD. m HF n These antibacterial polymers exert their antibacterial effect through a membrane disruption mechanism, achieving a minimum inhibitory concentration (MIC) of 4 μg / mL against multidrug-resistant Gram-negative pathogens in ESKAPE, demonstrating superior efficacy compared to current first-line antibiotics. Even after continuous treatment of E. coli ESBL 110 generations, no resistance was observed. Therefore, this fluorinated amphiphilic polymer can efficiently and selectively kill multidrug-resistant Gram-negative bacteria while reducing the probability of resistance development. The fluorinated amphiphilic polymer designed in this invention will fill the gap in current drugs for treating Gram-negative bacterial infections.

[0031] 2. This invention employs the RAFT polymerization method to polymerize monomers DMAEMA and HFBMA into a fluorinated amphiphilic block copolymer. The preparation process is simple, low-cost, suitable for large-scale production, and the structure is controllable. The introduced fluorinated monomers exhibit superior hydrophobicity and oleophobicity, enhancing the polymer's antibacterial ability. The introduced amine monomers show specific affinity for Gram-negative bacteria LPS. Furthermore, it is demonstrated that the regulation of positive charge and hydrophobicity can endow the antibacterial polymer with Gram selectivity. This provides a new approach for developing novel antibacterial drugs with low resistance and strong killing effects to combat Gram-negative bacterial infections, effectively broadening the drug spectrum for targeted elimination of Gram-negative bacteria. Attached Figure Description

[0032] Figure 1 PD is a fluorinated amphiphilic polymer m HF n The molecular structure diagram.

[0033] Figure 2 PA is a fluorinated amphiphilic polymer. m HF n Molecular structure diagram.

[0034] Figure 3 The graph shows the cytotoxicity results for Example 1 and Comparative Example 5.

[0035] Figure 4 The images show the morphology of bacteria after treatment in Example 1 and Comparative Examples 2, 3, and 5.

[0036] Figure 5 The graph shows the drug resistance results for Example 1 and Comparative Example 5.

[0037] Figure 6The image shows the LPS inhibition test results for Example 1. Detailed Implementation

[0038] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments:

[0039] Example 1:

[0040] like Figure 1 As shown, the method for preparing the antibacterial polymer includes the following steps:

[0041] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid, DMAEMA and HFBMA in 4 mL of dioxane at a molar ratio of 1:10:450:50, where the total concentration of the two monomers is 1 mmol / L. Add the solution to a Schlenk reaction tube and stir until completely dissolved.

[0042] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0043] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 60°C for 24 hours;

[0044] 4) The reaction product from step 3) was subjected to precipitation with n-hexane and vacuum drying to obtain a light pink solid, which is the fluorinated amphiphilic polymer PD. 45 HF5.

[0045] The antimicrobial properties of the polymer against multidrug-resistant ESKAPE were characterized by a minimum inhibitory concentration (MIC) test. The obtained PD 45 The MICs of HF5 against multidrug-resistant ESKAPE were >512, >512, 64, 4, 4, and 4 μg / mL, respectively, which are the PD values. 45 HF5 showed almost no antibacterial activity against Gram-positive bacteria, but had a strong killing effect on Gram-negative bacteria, indicating that PD... 45 HF5 exhibits significant selective antibacterial activity against Gram-negative bacteria.

[0046] Example 2:

[0047] like Figure 1 As shown, the method for preparing the antibacterial polymer includes the following steps:

[0048] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid, DMAEMA and HFBMA in 4 mL of dioxane at a molar ratio of 1:10:400:100, where the total concentration of the two monomers is 1 mmol / L. Add the solution to a Schlenk reaction tube and stir until completely dissolved.

[0049] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0050] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 60°C for 24 hours;

[0051] 4) The reaction product from step 3) is precipitated with n-hexane and dried under vacuum. The resulting pale pink solid is the fluorinated amphiphilic polymer PD. 40 HF 10 .

[0052] The antibacterial properties of the polymer against multidrug-resistant ESKAPE were characterized by MIC testing. The obtained PD 40 HF 10 The MICs for multidrug-resistant ESKAPE were >512, >512, 256, 16, 8, and 8 μg / mL, respectively, indicating PD. 40 HF 10 It showed almost no antibacterial activity against Gram-positive bacteria, but at this ratio, its antibacterial effect against Gram-negative bacteria gradually weakened compared to Example 1, but it still showed Gram-selective antibacterial ability.

[0053] As can be seen from the MICs of Examples 1 and 2, the fluorinated amphiphilic polymer PD... m HF n It can selectively eliminate Gram-negative bacteria, but shows almost no antibacterial activity against Gram-positive bacteria; when the fluoride content is too high, its antibacterial effect against Gram-negative bacteria is weakened.

[0054] Comparative Example 1:

[0055] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid, DMAEMA and HFBMA in 4 mL of dioxane at a molar ratio of 1:10:350:150, where the total concentration of the two monomers is 1 mmol / L. Add the solution to a Schlenk reaction tube and stir until completely dissolved.

[0056] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0057] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 60°C for 24 hours;

[0058] 4) The reaction product from step 3) is precipitated with n-hexane and dried under vacuum. The resulting pale pink solid is the fluorinated amphiphilic polymer PD. 35 HF 15 .

[0059] PD obtained 35 HF 15 It exhibits poor water solubility at room temperature and pH around 7.4, thus preventing further testing of its MIC for multidrug-resistant ESKAPE; this demonstrates that the regulation of the ratio of cationic monomers to hydrophobic monomers is particularly important.

[0060] Comparative Example 2:

[0061] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid, dithiobenzoic acid and DMAEMA in 4 mL of dioxane at a molar ratio of 1:10:500, wherein the monomer concentration is 1 mmol / L, and add it to a Schlenk reaction tube and stir until completely dissolved.

[0062] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0063] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 60°C for 24 hours;

[0064] 4) The reaction product from step 3) was precipitated with n-hexane and dried under vacuum to obtain a light pink solid amphiphilic polymer PD. 50 .

[0065] The antibacterial properties of the polymer against multidrug-resistant ESKAPE were characterized by MIC testing. The obtained PD 50 The MICs for multidrug-resistant ESKAPE were >512, >512, 256, 8, 64, and 16 μg / mL, respectively, which correspond to PD. 50 It showed almost no antibacterial activity against Gram-positive bacteria, but had a certain clearance effect on Gram-negative bacteria.

[0066] Compared with Examples 1-2, it can be seen that PD 50 It also exhibits the ability to selectively eliminate Gram-negative bacteria, but its elimination effect is weaker than that of Examples 1 and 2, which is insufficient to support its practical application. This further highlights the importance of fluorinated monomers. Fluorinated monomers have superior hydrophobicity and oleophobicity, play an important role in cell membrane penetration, and can improve antibacterial ability to a certain extent, which in turn confirms the necessity of the fluorinated amphiphilicity of the present invention.

[0067] Comparative Example 3:

[0068] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid and BocAEMA in 4 mL of dioxane at a molar ratio of 1:10:500, wherein the monomer concentration is 1 mmol / L, and add it to a Schlenk reaction tube and stir until completely dissolved.

[0069] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0070] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 65°C for 24 hours;

[0071] 4) The reaction product from step 3) is precipitated with n-hexane and then dried under vacuum;

[0072] 5) The reaction product from step 4) was dissolved in 2 mL of a 4 mol HCl / dioxane system and reacted for 12 h to deprotect the product, yielding a pale pink solid amphiphilic polymer PA. 50 .

[0073] The antibacterial properties of the polymer against multidrug-resistant ESKAPE were characterized by MIC testing. The obtained PA 50 The MICs for multidrug-resistant ESKAPE were 32, 32, >512, 256, 64, and 256 μg / mL, respectively.

[0074] Compared with Examples 1-2 and Comparative Example 2, it can be seen that PA 50 It exhibits broad-spectrum activity, capable of simultaneously eliminating Gram-positive and some Gram-negative bacteria, with a significantly stronger ability to eliminate Gram-positive bacteria than Gram-negative bacteria. Therefore, it lacks the ability to selectively eliminate Gram-negative bacteria. Thus, the selective elimination of Gram-negative bacteria primarily stems from the contribution of the monomer DMAEMA; using it as a cationic block unit endows the antimicrobial polymer with the function of selectively eliminating Gram-negative bacteria.

[0075] Comparative Example 4:

[0076] like Figure 2 As shown, the method for preparing the antibacterial polymer includes the following steps:

[0077] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid, BocAEMA and HFBMA in 4 mL of dioxane at a molar ratio of 1:10:450:50, where the total concentration of the two monomers is 1 mmol / L. Add the solution to a Schlenk reaction tube and stir until completely dissolved.

[0078] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0079] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 65°C for 24 hours;

[0080] 4) The reaction product from step 3) is precipitated with n-hexane and then dried under vacuum;

[0081] 5) The reaction product from step 4) was dissolved in 2 mL of a 4 mol HCl / dioxane system and reacted for 12 h to deprotect the product, yielding a pale pink solid amphiphilic polymer PA. 45 HF5.

[0082] The antibacterial properties of the polymer against multidrug-resistant ESKAPE were characterized by MIC testing. The obtained PA 45 The MICs of HF5 against multidrug-resistant ESKAPE were 32, 16, 256, 256, 32, and 256 μg / mL, respectively.

[0083] Compared with Examples 1-2 and Comparative Example 3, it can be seen that PA 45 HF5 also lacks the ability to selectively eliminate Gram-negative bacteria, but with the addition of fluorinated hydrophobic monomers in its hydrophobic portion, its effectiveness in eliminating Gram-positive bacteria is significantly improved. This further confirms that fluorinated hydrophobic monomers have the ability to enhance antibacterial effects.

[0084] Comparative Example 5:

[0085] like Figure 1 As shown, the method for preparing the antibacterial polymer includes the following steps:

[0086] 1) Dissolve azobisisobutyronitrile, 4-cyanopentanoic acid dithiobenzoic acid, BocAEMA and HFBMA in 4 mL of dioxane at a molar ratio of 1:10:400:100, where the total concentration of the two monomers is 1 mmol / L. Add the solution to a Schlenk reaction tube and stir until completely dissolved.

[0087] 2) After the reaction system in step 1) undergoes at least three freeze-pump-charge cycles in double-row tubes, the atmosphere is replaced with an argon atmosphere;

[0088] 3) Place the reaction system from step 2) in a constant temperature stirrer and slowly heat it to 65°C for 24 hours;

[0089] 4) The reaction product from step 3) is precipitated with n-hexane and then dried under vacuum;

[0090] 5) The reaction product from step 4) was dissolved in 2 mL of a 4 mol HCl / dioxane system and reacted for 12 h to deprotect the product, yielding a pale pink solid amphiphilic polymer PA. 40 HF 10 .

[0091] The antibacterial properties of the polymer against multidrug-resistant ESKAPE were characterized by MIC testing. The obtained PA 40 HF10 The MICs for multidrug-resistant ESKAPE were 32, 16, 64, 32, 32, and 64 μg / mL, respectively.

[0092] Compared with Comparative Example 4, it can be seen that as the fluorine content increases, the antibacterial ability of the antimicrobial polymer can be significantly improved, but its ability to eliminate Gram-negative bacteria is still limited, and it has no selectivity for Gram-negative bacteria.

[0093] In summary, comparing Examples 1-2, Comparative Example 2, and Comparative Example 3-4, it was found that the polymers of DMAEMA monomer introduced in Examples 1-2 and Comparative Example 2 selectively kill Gram-negative bacteria, while Comparative Example 3-4 exhibits a broad-spectrum antibacterial effect but has no ability to eliminate Gram-negative bacteria.

[0094] Figure 3 Based on the cell compatibility results of Example 1 and Comparative Example 5, Comparative Example 5 showed no cytotoxicity at 64 and 128 μg / mL in a short period of time, but showed slight cytotoxicity after long-term co-culture; Example 1 not only showed no cytotoxicity at 64 and 128 μg / mL, but also had a slight effect of promoting cell proliferation. Thus, it can be shown that the antibacterial polymer prepared by the present invention has good biocompatibility.

[0095] Figure 4 The images show scanning electron microscope (SEM) images of bacteria after co-incubation with the polymers in Examples 1, 2-3, and 5. After polymer treatment, varying degrees of wrinkling, depressions, and even perforations appeared on the surface of each bacterium. Compared to Comparative Examples 2 and 3, the polymers in Example 1 and Comparative Example 5, which introduced fluorinated hydrophobic monomers, exhibited stronger membrane-disrupting capabilities. The surfaces of almost all bacteria treated with these polymers showed severe perforation; some bacteria were so fragmented that their morphology was no longer observable, confirming a significant membrane-disrupting antibacterial mechanism.

[0096] Figure 5 The results are from the drug resistance test of Example 1 and Comparative Example 5. After 110 generations of continuous treatment with levofloxacin (Lev) at 0.5×MIC, the MIC of E. coli ESBL treated with Lev increased 16 times, while Example 1 and Comparative Example 5, although showing slight fluctuations, did not cause a significant increase in MIC. Therefore, Example 1 and Comparative Example 5 have an extremely low probability of inducing bacterial drug resistance, and can be considered to be unlikely to induce bacterial resistance, attributed to their significant membrane disruption antibacterial mechanism.

[0097] Figure 6The results of the LPS inhibition test for Example 1 are shown. Co-incubation of LPS with Example 1 significantly affected the antibacterial effect of the polymer in a concentration-dependent manner. Even at concentrations below 1 μg / mL, LPS doubled the MIC of E. coli ESBL in Example 1. As the LPS concentration was gradually increased to 25 μg / mL, the antibacterial effect of E. coli ESBL in Example 1 gradually weakened to 8 × MIC. Therefore, it can be inferred that Example 1 has a certain affinity for LPS on the surface of Gram-negative bacteria.

[0098] Of course, during the research process, within the aforementioned limitations, adjustments were made to the initiator, chain transfer reagent, organic solvent, unsuitable solvent, and even the reaction temperature and reaction time. All these adjustments resulted in the acquisition of the target antimicrobial polymer that met the objectives of the invention, and it also efficiently and selectively eliminates Gram-negative bacteria while reducing the probability of drug resistance development, without affecting the performance of the target product. To facilitate core comparisons and minimize influencing factors, the same initiator, chain transfer reagent, organic solvent, unsuitable solvent, and even the reaction temperature and reaction time were used in all the aforementioned embodiments and comparative examples.

[0099] This invention proposes that, based on the same antibacterial mechanism, if the hydrophobic monomer 2,2,3,3,4,4,4-heptafluorobutyl methacrylate is replaced with other fluorinated groups having the same methacrylate structure, such as 2,2,2-trifluoroethyl methacrylate, 2,2,3,3,3-pentafluoropropyl methacrylate, etc., it is also possible to synthesize antibacterial polymers with the same selective ability to eliminate Gram-negative bacteria.

[0100] In summary, the fluorinated amphiphilic polymer prepared by this invention based on the specific affinity and efficient membrane-breaking mechanism of LPS has precise Gram-selective antibacterial ability, low probability of drug resistance development, and good biocompatibility, thus having broad application prospects.

[0101] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any person skilled in the art can easily conceive of various equivalent modifications or substitutions within the scope of the technology disclosed in the present invention, and such modifications or substitutions should all be covered within the scope of protection of the present invention.

Claims

1. An antimicrobial polymer that selectively eliminates Gram-negative bacteria, characterized in that, The chemical formula is PD m HF n The structural formula is as follows: ； Wherein, the ratio of n to m represents the molar ratio of monomers 2,2,3,3,4,4,4-heptafluorobutyl methacrylate and 2-(dimethylamino)ethyl methacrylate, and 0.1≤n / m≤0.

2.

2. The antimicrobial polymer for selectively eliminating Gram-negative bacteria according to claim 1, characterized in that: Its chemical formula is PD 45 HF5.

3. The method for preparing the antimicrobial polymer for selectively eliminating Gram-negative bacteria as described in claim 1, characterized in that, Includes the following steps: 1) Dissolve the initiator, chain transfer reagent, 2-(dimethylamino)ethyl methacrylate and 2,2,3,3,4,4,4-heptafluorobutyl methacrylate in an organic solvent and add them into a Schlenk reaction tube, stirring until completely dissolved. 2) After the reaction system in step 1) undergoes at least three double-row tube freeze-pump-charge cycles, the atmosphere is replaced with an inert gas; 3) Place the reaction system from step 2) in a constant-temperature stirring environment to carry out the reaction; 4) After the reaction in step 3) is completed, the reaction product is precipitated and vacuum dried to obtain the target antibacterial polymer.

4. The method for preparing the antimicrobial polymer for selectively eliminating Gram-negative bacteria according to claim 3, characterized in that: In step 1), the initiator is azobisisobutyronitrile, dimethyl azobisisobutyrate, benzoyl peroxide, or tert-butyl peroxide. The chain transfer agent is 4-cyanopentanoic acid dithiobenzoic acid; The organic solvent is dimethyl sulfoxide, N,N-dimethylformamide, acetonitrile, or dioxane.

5. The method for preparing the antimicrobial polymer for selectively eliminating Gram-negative bacteria according to claim 3, characterized in that: In step 1), the molar ratio of the initiator, chain transfer reagent, 2-(dimethylamino)ethyl methacrylate and 2,2,3,3,4,4,4-heptafluorobutyl methacrylate is 1:10:400:100-1:10:450:50, wherein the total molar concentration of the two monomers is 1 mmol / L.

6. The method for preparing the antimicrobial polymer selectively scavenging Gram-negative bacteria according to any one of claims 3-5, characterized in that: In step 2), the inert gas is nitrogen or argon.

7. The method for preparing the antimicrobial polymer for selectively eliminating Gram-negative bacteria according to claim 6, characterized in that: In step 3), a constant temperature stirrer is used, the reaction temperature is 50-70℃, and the reaction time is 18-24h.

8. The method for preparing the antibacterial polymer according to claim 7, characterized in that: In step 4), the reaction products are precipitated using a poor solvent.

9. An antibacterial agent for eliminating Gram-negative bacteria, characterized in that: Its active ingredient is the antimicrobial polymer that selectively eliminates Gram-negative bacteria as described in claim 1.

Images