CN114344571B - Antibacterial material and preparation method and application thereof - Google Patents

Abstract

The application discloses an antibacterial material which is formed by the complex crosslinking of alginic acid and zwitter-ion copolymer and copper ions. The zwitterionic polymer end can prevent bacteria from adhering due to good hydrophilicity, so that an anti-adhesion effect is achieved, and an anti-adhesion layer is formed; meanwhile, copper ions can be intelligently released according to environmental changes, so that the aim of sterilization is fulfilled, and the method is particularly suitable for clinical urinary tract infection.

Description

A kind of antibacterial material and its preparation method and application

technical field

The application relates to an antibacterial material, which belongs to the field of biomedical polymer materials.

Background technique

Implantable medical device infection is a serious problem in current clinical practice. Constructing an antibacterial coating on the surface of the device to inhibit the adhesion, growth and reproduction of bacteria is an effective means of fighting infection, and it is also one of the current research hotspots in the field of biomedical materials.

Traditional antibacterial coatings can be roughly divided into anti-adhesion type and bactericidal type, which achieve antibacterial effect by inhibiting bacterial adhesion or killing bacteria. Clinical applications have found that coatings that rely on a single mechanism to exert antibacterial/bactericidal functions have certain limitations, especially the long-term antibacterial effect is poor. For example, the anti-adhesion coating can inhibit but not completely prevent the deposition of bacteria on the surface, and a small amount of bacteria that reach the surface can still multiply and grow to form biofilms; the bactericidal groups that contact the bactericidal coating are usually positively charged, which are easy to adsorb bacterial residues to kill bacteria. The active substance is covered to cause failure; the release of the biocide will be affected after the surface of the release biocide coating is deposited and covered by other substances. These coatings have a single antibacterial mechanism and poor long-term antibacterial effect, which cannot fundamentally solve the problem of coating failure caused by the reproduction of bacteria on the surface or the coating of residues on the surface.

Constructing a multi-mechanism coating with both anti-adhesion and bactericide release functions can effectively improve the antibacterial ability of the material surface. However, surface-loaded fungicides have problems such as easy consumption and biotoxicity of high concentrations of fungicides. How to design a response mechanism so that the coating can selectively regulate the release of fungicides according to environmental changes, "intelligently" adjust the antibacterial ability of the coating, and reduce the probability of infection and complications, is a very worthy concern in the field of antibacterial materials. It has important clinical application prospects.

SUMMARY OF THE INVENTION

In view of the deficiencies of the prior art, according to one aspect of the present application, an antibacterial material is provided.

An antibacterial material, characterized in that the antibacterial material comprises a substrate, a complex of alginic acid and a zwitterionic copolymer and copper ions.

The antibacterial material is formed by complex cross-linking of a copolymer of alginic acid and zwitterion and copper ions. Due to its good hydrophilicity, the end of the zwitterionic polymer can prevent the adhesion of bacteria, achieve anti-adhesion effect, and form an anti-adhesion layer; at the same time, copper ions will be "smartly" released according to environmental changes to achieve the purpose of sterilization and form sterilization. Floor. Through the synergy of the dual mechanism of the anti-adhesion layer and the bactericidal layer, the antibacterial properties of the surface of the implanted device are improved.

Optionally, the copper ion-alginic acid-zwitterion copolymer has a copper ion content of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, Any value of 15, 16, 17, 18, 19, 20 μg/cm ² or a value in the range between any two values.

Optionally, in the alginic acid-zwitterion copolymer, the zwitterion is selected from at least one of carboxylate betaine type zwitterion, sulfonate betaine type zwitterion, and phosphate betaine type zwitterion.

Optionally, the substrate is selected from at least one of a metal substrate and a polymer material substrate.

The second aspect of the present invention provides a method for preparing the above-mentioned antibacterial material.

A method for preparing antibacterial materials, the steps are as follows:

(1) obtaining an aqueous solution containing alginic acid-zwitterionic copolymer;

(2) obtaining an aqueous solution containing copper ions;

(3) the substrate is first immersed in an aqueous solution containing copper ions to obtain a substrate whose surface is coated by the aqueous solution containing copper ions;

(4) Immerse the substrate coated with the aqueous solution of copper ions on the surface in the aqueous solution of alginic acid-zwitterion copolymer, and react to obtain the antibacterial material.

The preparation method of the copolymer of described alginic acid and zwitterion is as follows:

Alginic acid is dissolved in water at 1-5% (v/v), the solution is heated to 60°C and maintained at this temperature during the reaction, then the initiator ammonium or potassium persulfate is added, prior to passing through argon or nitrogen Degassing to keep the reaction system in an oxygen-free environment, argon or nitrogen gas was bubbled into the mixture for another 30 min, followed by the gradual addition of the aqueous solution of zwitterionic monomer to the mixture over 30 min with a syringe, wherein the zwitterionic monomer was added. The molar ratio of body and alginic acid unit was 1:1, and the reaction time was 6h. After the reaction, dialysis was performed using a cellulose membrane (molecular weight cut-off of 12,000) for three days, and the product was collected after lyophilization.

Optionally, in step (1), the mass concentration of the solution of the copolymer of alginic acid and zwitterion is 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5% , 5.5%, 6%, 6.5%, 7% or any value in the range determined by any two values.

Optionally, the aqueous solution of copper ion in step (2), the molar concentration of copper ion is 1mmol/L, 1.5mmol/L, 2mmol/L, 2.5mmol/L, 3mmol/L, 3.5mmol/L, 4mmol/L. Any value or any two values of L, 4.5 mmol/L, 5 mmol/L, 5.5 mmol/L, 6 mmol/L, 6.5 mmol/L and 7 mmol/L determine the numerical value in the range.

The copper ions come from at least one of a copper chloride solution, a copper sulfate solution, a copper nitrate solution, and a copper acetate solution.

Optionally, the immersion time in step (3) is any value or any two in 3min, 6min, 9min, 12min, 15min, 18min, 21min, 24min, 27min, 30min, 33min, 36min, 39min, 42min, 45min. The value in the determination range is determined by the value, and the immersion time in step (4) is any value in 5min, 10min, 15min, 20min, 25min, 30min, 35min, 40min or any two values determine the value in the range.

The third aspect of the present invention provides an anti-infection application of the above-mentioned antibacterial material as an implantable medical device.

Application of the above-mentioned antibacterial material and the antibacterial material obtained by the above-mentioned preparation method as an implantable medical device in anti-infection.

Preferably, the implantable medical device comprises a urinary tract implantable medical device.

The beneficial effects that this application can produce include:

(1) The antibacterial material prepared in this application has a dual mechanism synergistic antibacterial effect. First, the zwitterionic polymer ends can prevent the adhesion of bacteria and form an anti-adhesion layer. At the same time, the release of copper ions can further kill bacteria and form a dual antibacterial effect.

(2) The antibacterial material provided in this application is formed by cross-linking based on the complexation between copper ions and alginic acid-zwitterion copolymers, without additional initiators, cross-linking agents and catalysts. Residual problems are easy for industrial application; at the same time, the complex and cross-linked network structure endows the antibacterial materials with good self-healing properties.

(3) The copper ions in the antibacterial material provided by this application can be released "intelligently" according to the concentration of free NH ₃ in the surrounding environment. When the concentration of free NH ₃ in the solution increases, the release of copper ions will be accelerated, especially Applicable to clinical urinary tract infection.

Description of drawings

Figure 1: Schematic diagram of the structure of the antibacterial material of the embodiment adopted in this application and its antibacterial effect under different pH conditions.

Figure 2: The response behavior of Example 1 used in this application under different pH (containing different concentrations of NH ₃ ) conditions.

Figure 3: Blank substrate (silicon rubber), Comparative Example 1 and Example 1 of the present application silicone rubber and alginic acid-copper ion-alginic acid and zwitterion-copper ion complexes modified surfaces after bacterial adsorption for 4 hours respectively Counting result graph.

Figure 4: Blank substrate (silicone rubber), Comparative Example 1 and Example 1 of the present application silicone rubber and alginic acid-copper ion and alginic acid and zwitterion and copper ion complexes modified surfaces respectively after bacterial adsorption for 4 hours Scanning electron microscope photo.

Detailed ways

The present application will be described in detail below with reference to the examples, but the present application is not limited to these examples.

Example 1

(1) prepare and obtain alginic acid-zwitterionic copolymer;

Alginic acid was dissolved in water at 3% (v/v), the solution was heated to 60°C and maintained at this temperature during the reaction, then the initiator ammonium persulfate was added, before degassing the reaction system with argon to keep the reaction system free. In an oxygen environment, argon gas was bubbled into the mixture for 30 min, followed by adding the aqueous carboxylate betaine zwitterionic monomer solution to the mixture gradually within 30 min with a syringe, wherein the carboxylate betaine zwitterionic monomer was added to the mixture. The molar ratio of ionic monomer and alginic acid was 1:1, and the reaction time was 6 h. After the reaction, dialysis was performed using a cellulose membrane (molecular weight cut-off of 12,000) for three days, and the product was collected after lyophilization.

(2) adopt copper sulfate to prepare copper ion solution, the solvent is water, and wherein copper ion concentration is 7mmol/L;

(3) prepare the copolymer solution of alginic acid-carboxylate betaine type zwitterion, the solvent is water, and wherein the mass concentration is 8%;

(4) Put the polymer-based material silicone rubber into the copper ion solution to soak for 45min;

(5) Put the polymer-based material silicone rubber into the alginic acid and carboxylate betaine type zwitterionic copolymer solution and soak for 40 minutes; namely, the silicone rubber coated with the coating is obtained.

The above coating materials were placed in a 25 mL centrifuge tube containing 10 mM ammonia solution, and stirred at 37 ° C and 250 rpm for 10 min. It was found that the coating material did not change significantly in the low-concentration ammonia solution, but in the high-concentration ammonia solution. , the gel structure was partially destroyed and spread throughout the centrifuge tube, which was caused by the copper ions in the coating material and the free NH ₃ forming a copper ammine complex, and the copper ions were released from the coating material, see Fig. 2. It is proved that the copper ions in the coating can be released "intelligently" according to the concentration of free _NH3 in the surrounding environment. The release schematic is shown in Figure 1.

The above coating material was placed in 25 mL of an aqueous solution containing 100 mM ammonia. Since the copper ions in the coating material and free _NH3 formed a copper ammine complex, the copper ions were rapidly released from the hydrogel and the gel structure was disintegrated. . The gel structure is stable in low concentration ammonia solution (10 mM).

The gram-negative bacteria Proteus mirabilis and the gram-positive bacteria Staphylococcus epidermidis were selected as the test strains, and the antibacterial experiments were carried out and counted, and the antibacterial properties were characterized by scanning electron microscope.

The results of Proteus mirabilis showed that the number of bacteria on silicone rubber and alginate-copper ion and alginic acid-zwitterion-copper ion modified surface after adsorption for 4 hours was 4.5×10 ⁷ , 2.3×10 ⁷ and 0.18×10 ⁷ , respectively. indivual. The 4 hours of bacterial adsorption is shown in the scanning electron microscope photo in Figure 4. It can be seen that the bacteria adsorbed by alginic acid-zwitterion-copper ion is the least, which proves that the coating has a good antibacterial effect on Proteus mirabilis.

The results of Staphylococcus epidermidis showed that the counts of bacteria on silicone rubber and alginate-copper ion and alginate-zwitterion-copper ion modified surfaces were 2.2×10 ⁷ , 0.8×10 ⁷ and 0.4×10 ⁶ , respectively, after 4 hours of adsorption. indivual. The 4 hours of bacterial adsorption is shown in the scanning electron microscope photo in Figure 4. It can be seen that the bacteria adsorbed by alginic acid-zwitterion-copper ion is the least, which proves that the coating has a good antibacterial effect. It proves that the coating has a good antibacterial effect on Staphylococcus epidermidis. .

Example 2

The experimental steps are the same as those in Example 1, the difference is that

Step (1) the volume concentration of alginic acid is 5% (v/v), and the zwitterionic monomer is a sulfonate betaine type zwitterionic monomer;

In step (2), copper nitrate is adopted, and the copper ion concentration is 1mmol/L;

In step (3), the mass concentration of the copolymer of alginic acid and sulfonate betaine type zwitterion is 1%;

In step (4), the base material is a titanium sheet, and the titanium sheet is soaked in the copper ion solution for 3 minutes;

In step (5), the soaking time is 5min.

The results of Proteus mirabilis showed that the number of bacteria on the titanium sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surface after adsorption for 4 hours was 6.3×10 ⁷ , 4.5×10 ⁷ and 1.4×10 ⁷ , respectively. indivual.

The results of Staphylococcus epidermidis showed that the number of bacteria on the titanium sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surface after adsorption for 4 hours was 4.3×10 ⁷ , 2.2×10 ⁷ and 3.6×10 ⁶ , respectively. indivual.

Example 3

The experimental steps are the same as those in Example 1, the difference is that

Step (1) the volume concentration of alginic acid is 1% (v/v), and the zwitterionic monomer is a phosphate betaine type zwitterionic monomer;

In step (2), copper chloride is adopted, and the copper ion concentration is 3mmol/L;

In step (3), the mass concentration of the copolymer of alginic acid and sulfonate betaine type zwitterion is 2%;

In step (4), the base material is stainless steel sheet, and the stainless steel sheet is soaked in copper ion solution for 5min;

The soaking time in step (5) is 10min.

The results of Proteus mirabilis showed that after 4 hours of adsorption of bacteria on the stainless steel sheet and alginate-copper ion and alginic acid-zwitterion-copper ion modified surfaces, the counts were 8.5×10 ⁷ , 5.2×10 ⁷ and 2.1×10 ⁷ , respectively. indivual.

The results of Staphylococcus epidermidis showed that the counts of bacteria on the stainless steel sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surfaces were 5.1×10 ⁷ , 1.9×10 ⁷ and 1.6×10 ⁶ , respectively, after 4 hours of adsorption. indivual.

Example 4

The experimental steps are the same as those in Example 1, the difference is that

Adopt copper acetate in step (2), copper ion concentration is 4mmol/L;

In step (3), the mass concentration of the copolymer of alginic acid and carboxylate betaine type zwitterion is 3%;

In step (4), the silicone rubber is soaked in the copper ion solution for 10 min;

The soaking time in step (5) is 15min.

The results of Proteus mirabilis showed that the counts of bacteria on silicone rubber and alginate-copper ion and alginate-zwitterion-copper ion modified surfaces were 4.5×10 ⁷ , 3.2×10 ⁷ and 0.83×10 ⁷ , respectively, after 4 hours of adsorption. indivual.

The results of Staphylococcus epidermidis showed that the number of bacteria on the silicone rubber and alginate-copper ion and alginate-zwitterion-copper ion modified surface after adsorption for 4 hours was 2.2×10 ⁷ , 1.3×10 ⁷ and 1.6×10 ⁶ , respectively. indivual.

Example 5

The experimental steps are the same as those in Example 2, the difference is that

In step (2), copper ion concentration is 5mmol/L;

In step (3), the mass concentration of the copolymer of alginic acid and sulfonate betaine type zwitterion is 5%;

In step (4), the base material is a titanium sheet, and the titanium sheet is soaked in a copper ion solution for 20 minutes;

The soaking time in step (5) is 25min.

The results of Proteus mirabilis showed that the number of bacteria on the titanium sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surface after adsorption for 4 hours was 6.3×10 ⁷ , 2.5×10 ⁷ and 0.6×10 ⁷ , respectively. indivual.

The results of Staphylococcus epidermidis showed that the number of bacteria on the titanium sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surface after adsorption for 4 hours was 4.3×10 ⁷ , 1.6×10 ⁷ and 1.3×10 ⁶ , respectively. indivual.

Example 6

The experimental steps are the same as those in Example 3, the difference is that

In step (2), copper ion concentration is 2mmol/L;

In step (3), the mass concentration of the copolymer of alginic acid and sulfonate betaine type zwitterion is 6%;

In step (4), the base material is stainless steel sheet, and the stainless steel sheet is soaked in copper ion solution for 25min;

The soaking time in step (5) is 30min.

The results of Proteus mirabilis showed that the counts of bacteria on the stainless steel sheet and alginate-copper ion and alginic acid-zwitterion-copper ion modified surface after adsorption for 4 hours were 8.5×10 ⁷ , 4.8×10 ⁷ and 1.8×10 ⁷ , respectively. indivual.

The results of Staphylococcus epidermidis showed that after 4 hours of adsorption of bacteria on the stainless steel sheet and alginate-copper ion and alginate-zwitterion-copper ion modified surfaces, the counts were 5.1×10 ⁷ , 2.3×10 ⁷ and 1.6×10 ⁶ , respectively. indivual.

Comparative Example 1

Experimental Procedure Copper Example 1, the difference is

In step (1), no carboxylate betaine type zwitterionic monomer is added.

The final obtained alginic acid-copper ion was subjected to the above-mentioned adsorption experiment of Proteus mirabilis and Staphylococcus epidermidis, and the count results after adsorption for 4 hours were 2.3×10 ⁷ and 0.8×10 ⁷ respectively. Example 1, Comparative Example 1, The technical results obtained for the blank sample (silicon rubber) are shown in FIG. 3 .

The above are only a few embodiments of the present application, and are not intended to limit the present application in any form. Although the present application is disclosed as above with preferred embodiments, it is not intended to limit the present application. Without departing from the scope of the technical solution of the present application, any changes or modifications made by using the technical content disclosed above are equivalent to equivalent implementation cases and fall within the scope of the technical solution.

Claims (7)

1. An antibacterial material, which is characterized by comprising a substrate, alginic acid and a complex compound of a zwitterionic copolymer and copper ions;

the preparation method of the antibacterial material comprises the following steps:

(1) alginic acid was dissolved in 1-5% v/v in water, the solution was heated to 60 ℃ and maintained at this temperature during the reaction, then the initiator ammonium persulfate or potassium persulfate was added, the reaction system was previously kept in an oxygen-free environment by argon or nitrogen degassing, argon or nitrogen was bubbled into the mixture for a further 30min, then an aqueous zwitterionic monomer solution was gradually added to the mixture over a period of 30min using a syringe, wherein the molar ratio of zwitterionic monomer to alginic acid units was 1: 1, reacting for 6 hours, dialyzing for three days by using a cellulose membrane after reaction, and collecting a product after freeze-drying;

(2) obtaining an aqueous solution containing copper ions;

(3) immersing the substrate in an aqueous solution containing copper ions to obtain a substrate with the surface coated by the aqueous solution containing copper ions;

(4) and soaking the substrate with the surface coated by the aqueous solution of copper ions in the aqueous solution of alginic acid-zwitter ion copolymer for reaction to obtain the antibacterial material.

2. The antimicrobial material of claim 1, wherein the complex of alginic acid and the zwitterionic copolymer with copper ions has a copper ion content of 1-20 μ g/cm ² 。

3. The antibacterial material according to claim 1, wherein in the complex of alginic acid and the zwitterionic copolymer with copper ions, the zwitterion thereof is at least one selected from the group consisting of carboxylate betaine zwitterion, sulfonate betaine zwitterion and phosphate betaine zwitterion.

4. The antimicrobial material of claim 1, wherein the substrate is selected from at least one of a metal substrate and a polymeric substrate.

5. The antibacterial material according to claim 1, wherein in the step (1), the mass concentration of the solution of alginic acid and the zwitterionic copolymer is 1-8%.

6. The antibacterial material according to claim 1, wherein in the aqueous solution of copper ions in the step (2), the molar concentration of the copper ions is 1 to 7 mmol/L;

the copper ions are at least one of copper chloride solution, copper sulfate solution, copper nitrate solution and copper acetate solution.

7. The antibacterial material according to claim 1, wherein the immersion time in step (3) is 3 to 45min, and the immersion time in step (4) is 5 to 40 min.

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