High-molecular antibacterial material based on borneol, preparation method and application thereof
Technical Field
The invention relates to the field of antibacterial polymers, in particular to a high-molecular antibacterial material based on borneol, a preparation method and application thereof.
Background
In recent years, bacterial infections have become one of the major problems affecting the health of millions of people, which is particularly serious in developing countries. Even more unexpected is that abuse of antibiotics provides an unprecedented environment for the evolution of drug-resistant bacteria, which has greatly diminished our ability to control bacterial infections through drugs. To solve these problems, development of new antibacterial means based on advanced antibacterial principles is required. It is well known that adhesion of bacteria to the surface of a material is the primary process for the formation of a bacterial biofilm. Therefore, the development of a functional interface capable of sterilizing and resisting bacterial adhesion has a very important significance for controlling bacterial infection.
Although there have been many achievements in the development of an antimicrobial interface, one bottleneck in this field is the lack of durability of the antimicrobial functional interface. On the one hand, the loss of the antibacterial ability is caused by the weak bonding mode between the antibacterial coating and the substrate, such as adsorption, layer-by-layer assembly and the like, so that the antibacterial coating is easy to fall off. On the other hand, it is inevitable that still a small number of pathogens pass through a strongly antifouling surface. Therefore, it is an indispensable strategy for an excellent antibacterial interface to quickly kill these escaping bacteria and avoid their colonization on the surface. Therefore, direct anchoring of the antimicrobial coating by covalent bonds is essential to produce permanent antimicrobial activity.
Borneol is a natural broad-spectrum antibacterial agent extracted from various plants, and has been proved to have good biocompatibility and various biomedical functions. The patent application with application number 201410081967.7 relates to synthesizing an acryloyl borneol monomer by taking four types of borneol with different chiral configurations and an acryloyl derivative as raw materials, and further forming a high-molecular antibacterial material through polymerization reaction, and finds that the materials can effectively prevent bacterial adhesion.
However, the above techniques are disadvantageous from the viewpoint of long-lasting antibacterial purpose and cost-effectiveness because they have poor mechanical properties and lack of effective binding force with the substrate, so that these polymers are easily released from the substrate.
Disclosure of Invention
In view of the disadvantages of the prior art, the primary object of the present invention is to prepare a borneol-based polymeric antibacterial material having excellent antibacterial activity, effective sterilization and prevention of bacterial adhesion.
The invention also aims to prepare the antibacterial copolymerization material for firmly bonding the borneol-based macromolecular antibacterial material to a substrate, and the material can be anchored with a substrate material containing hydroxyl through a covalent bond by a simple and environment-friendly hydrothermal method, so that a durable antibacterial and anti-adhesion functional interface is formed, and the antibacterial copolymerization material has wide application value in the fields of medicine, food, sanitation and environment protection.
In order to achieve the purpose, the invention provides a borneol-based macromolecular antibacterial material, which takes borneol and acyl halide derivatives as raw materials to synthesize a borneol micromolecular initiator B-X with a halogen atom at the tail end, and further performs a polymerization reaction with dimethylaminoethyl methacrylate (DMAEMA) to form the borneol-based macromolecular antibacterial material BP, and the structure of the borneol-based macromolecular antibacterial material BP is shown as the following formula:
wherein R represents a hydrogen atom or an alkyl group, X represents a halogen atom, and m.epsilon. [5,5000 ];
wherein the reaction molar ratio of the borneol to the acyl halide derivative is 1:20-2:1, and the reaction molar ratio of the borneol small molecular initiator B-X to the dimethylaminoethyl methacrylate (DMAEMA) is 1-300.
Preferably, the borneol configuration comprises D-borneol, L-borneol and ISO-borneol.
Preferably, the acyl halide derivatives include 2-chloroisobutyryl chloride, 2-bromoisobutyryl bromide and derivatives thereof.
The invention also provides a preparation method of the high molecular antibacterial material based on borneol, which is characterized by comprising the following preparation steps:
s1, adding the borneol into an organic solvent, stirring and dissolving, reacting in an alkaline environment with the molar ratio of 1:20-2:1 between the borneol and the acyl halide derivative for 20-180min in an ice-water bath, heating to room temperature, continuing to react for 2-48h, and purifying after the reaction is finished to obtain the borneol micromolecule initiator B-X;
s2, adding the borneol small molecular initiator B-X, the organic solvent, the dimethylaminoethyl methacrylate (DMAEMA), the ligand and the catalyst in the step S1 into a reaction bottle, and carrying out polymerization reaction in an oxygen-free environment, wherein the polymerization reaction temperature is controlled to be 0-100 ℃, and the reaction time is 10min-24 h;
wherein the molar ratio of dimethylaminoethyl methacrylate (DMAEMA) to the borneol small molecular initiator B-X is 1-300, the preferred range of the molar ratio of the ligand to the borneol small molecular initiator B-X is 0.1-2, the molar ratio of the catalyst to the borneol small molecular initiator B-X is 0.01-2, and the molar ratio of the organic solvent to the borneol small molecular initiator B-X is 50-500;
s3, after the reaction is finished, adding an organic solvent or exposing the mixture to air to inactivate an initiation system and terminate the polymerization; then purifying by neutral alumina column or dialysis to obtain the borneol-based macromolecular antibacterial material BP.
Preferably, the organic solvent in step S1 is dichloromethane, methanol, or the like.
Preferably, the ligand in step S2 is Pentamethyldiethylenetriamine (PMDETA) or 1,1,4,7,10, 10-Hexamethyltriethylenetetramine (HMTETA).
Preferably, the catalyst described in step S2 is cuprous halide (cuprous bromide, cuprous chloride).
The invention also provides a preparation method of the antibacterial copolymer for firmly bonding the borneol-based high polymer material BP to a substrate, which comprises the following steps:
s1, adding the borneol-based high-molecular antibacterial material BP, an organic solvent, a ligand, cuprous halide and two monomers, namely Methyl Methacrylate (MMA) and hydroxyethyl methacrylate (HEMA), into a reverse bottle, and carrying out polymerization reaction under an oxygen-free environment, wherein the reaction temperature is controlled to be 0-100 ℃, and the reaction time is 10min-24 h;
wherein the molar ratio of two monomers HEMA and MMA is 1:3-3:1, the molar ratio of the sum of the two monomers to the borneol-based polymeric antibacterial material BP is 5-200, the preferred range of the molar ratio of a ligand to the borneol-based polymeric antibacterial material BP is 0.1-2, the molar ratio of cuprous halide to the borneol-based polymeric antibacterial material BP is 0.01-2, and the molar ratio of an organic solvent to the borneol-based polymeric antibacterial material BP is 50-500;
s2, after the reaction is completed, adding an organic solvent or exposing to air to inactivate the initiation system and terminate the polymerization. Then purifying by neutral alumina column or dialysis to obtain the borneol-based macromolecular antibacterial copolymer BP-b-HM.
Compared with the prior art, the invention has the beneficial effects that:
(1) according to the invention, a series of borneol-based high-molecular antibacterial materials BP with excellent antibacterial performance are synthesized, and borneol groups are innovatively modified on a molecular chain of a dimethylaminoethyl methacrylate (DMAEMA) polymer, so that the antibacterial performance of the polymer is obviously improved;
(2) the invention synthesizes the borneol-based macromolecular antibacterial material BP-b-HM for the first time, and the copolymerization material has the characteristic of easy covalent anchoring with a substrate, thereby preparing a functional surface with lasting antibacterial and antibacterial adhesion, and having wide application prospect in the fields of medicine, food, sanitation and environmental protection;
(3) the borneol-based high-molecular antibacterial material BP is applied to the field of interfacial antibacterial for the first time, for example, a simple, mild and environment-friendly hydrothermal method is only needed to endow a cotton fabric substrate with excellent sterilizing and antibacterial adhesion effects, so that the added value of the cotton fabric is greatly improved;
(4) the invention greatly improves the water solubility of the borneol, enhances the antibacterial capability of the borneol and expands the application of the borneol in the field of high-molecular antibacterial materials.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to these drawings without creative efforts.
Fig. 1 is a graph of the antibacterial kinetics analysis of three finished cotton fabrics provided by the present invention against gram positive (e.coli) and gram negative (s.aureus) bacteria;
fig. 2 is a plate count chart of the antibacterial rate of three finished cotton fabrics provided by the invention to gram-positive bacteria (e.coli) and gram-negative bacteria (s.aureus);
fig. 3 is a graph of the anti-adhesion effect of BP @ CF on gram-positive bacteria (E.coli) and gram-negative bacteria (S.aureus) of cotton fabric finished with BP-b-HM of different concentrations by a field emission scanning electron microscope.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example one
The invention provides a borneol-based macromolecular antibacterial material, which is prepared by taking borneol and acyl halide derivatives as raw materials, synthesizing a borneol micromolecular initiator B-X with a halogen atom at the tail end, and further carrying out polymerization reaction with dimethylaminoethyl methacrylate (DMAEMA) to form the borneol-based macromolecular antibacterial material BP, wherein the structure of the borneol-based macromolecular antibacterial material BP is shown as the following formula:
wherein R represents a hydrogen atom or an alkyl group, X represents a halogen atom, and m.epsilon. [5,5000 ];
wherein the reaction molar ratio of the borneol to the acyl halide derivative is 1:20-2:1, and the reaction molar ratio of the borneol small molecular initiator B-X to the dimethylaminoethyl methacrylate (DMAEMA) is 20-200.
In addition, the cotton fabric substrate can be endowed with excellent sterilization and antibacterial adhesion efficacy only by a simple mild and environment-friendly hydrothermal method, and the added value of the cotton fabric is greatly improved, so that the material has the characteristic of easy covalent anchoring with the substrate, and can prepare a functional surface capable of lasting antibiosis and antibacterial adhesion, and has wide application prospects in the fields of medicine, food, sanitation and environmental protection.
Example two
The invention provides a preparation method of a high-molecular antibacterial material based on borneol, which comprises the following steps:
s1, weighing 1.018g of dextroborneol and 4.6g of triethylamine into a reaction bottle, dissolving 30mL of dichloromethane, and cooling to 0 ℃ by using an ice water bath. Then, 40mL of freshly distilled dichloromethane and 4.02mL of 2-bromoisobutyryl bromide were added to a constant pressure funnel, and the mixture was reacted at room temperature for 12 hours under a nitrogen atmosphere. The final borneol micromolecule initiator B-Br is obtained through the steps of filtering, rotary evaporation, extraction, concentration, vacuum drying and the like, the purity of the product is 96%, and the yield is 95%.
S2, taking the borneol micromolecule initiator B-Br100mg synthesized in the example 1, adding 1.0894mL of dimethylaminoethyl methacrylate (DMAEMA) and 67.93 muL of Pentamethyldiethylenetriamine (PMDETA) into a reaction bottle, adding 3mL of methanol as a solvent, performing 3 times of cycles of 'quick freezing-vacuumizing-nitrogen filling and unfreezing', adding 51.27mg of catalyst CuBr, performing 2 times of cycles of 'quick freezing-vacuumizing-nitrogen filling and unfreezing', and heating to 45 ℃ for reaction.
And S3, after reacting for 4 hours, stopping the reaction in an ice-water bath, adding tetrahydrofuran to dilute the reaction solution, passing through a neutral alumina column, performing rotary evaporation and concentration, settling the polymer concentrated solution in frozen n-hexane for three times, and performing vacuum drying to obtain the borneol-based macromolecular antibacterial material BP with the product purity of 99% and the yield of 95%.
The structure of the borneol-based macromolecular antibacterial material BP is shown as the following formula (1):
wherein R represents a hydrogen atom or an alkyl group, X represents a halogen atom, and m.epsilon. [5,5000 ].
EXAMPLE III
The invention also provides a preparation method of the antibacterial copolymer for firmly bonding the borneol-based high polymer material BP to a substrate, which comprises the following steps:
s1, sequentially injecting 1.156mL of hydroxyethyl methacrylate (HEMA), 0.890mL of Methyl Methacrylate (MMA), 124 mu L of ligand (PMDETA), 20mL of cyclohexanone and the synthesized borneol-based polymer antibacterial material BP (0.97713g) into a reaction bottle, performing 3 times of cycles of 'quick freezing-vacuumizing-nitrogen filling and unfreezing', adding 85mg of catalyst CuBr, performing 2 times of cycles of 'quick freezing-vacuumizing-nitrogen filling and unfreezing', and heating to 80 ℃ for reaction.
And S2, after reacting for 4 hours, stopping the reaction in an ice-water bath, adding tetrahydrofuran to dilute the reaction solution, passing through a neutral alumina column, performing rotary evaporation and concentration, settling the polymer concentrated solution in frozen n-hexane for three times, and performing vacuum drying to obtain the borneol-based macromolecular antibacterial copolymer BP-b-HM with the product purity of 98% and the yield of 90%.
Taking the synthesized borneol-based macromolecular antibacterial copolymer BP-b-HM, preparing solutions with the concentrations of 0mg/mL (control group), 10mg/mL, 20mg/mL and 40 mg/mL) by using water/ethanol (v/v is 10:1), taking a commercially available cotton fabric according to the bath ratio of 1:10, soaking the cotton fabric into the solution for 20min, taking out the cotton fabric, airing the cotton fabric, taking out the cotton fabric after drying for 4h at 100 ℃ in a drying oven, and finally extracting all samples for 1.0h at 60 ℃ in THF to obtain the cotton fabric BP1@ CF finished by the borneol-based macromolecular antibacterial material.
The same technical means is utilized to prepare MP which has the same structure and similar molecular weight with the borneol-based macromolecular antibacterial material BP but does not contain the borneol group, the same technical means is utilized, MP1 which does not contain the borneol group is used as a raw material to prepare MP-b-HM, the means is utilized to prepare cotton fabric MP1@ CF by using the MP-b-HM as a raw material, then a common commercial cotton fabric finishing agent organosilicon quaternary ammonium salt AEM-5700 is purchased, the cotton fabric AEM @ CF is obtained by finishing the same feeding mass concentration of the former two test fabrics, and the cotton fabrics BP1@ CF, the cotton fabric MP1@ CF and the cotton fabric AEM @ CF are used as comparison experiments.
Experiment one
The antibacterial performance of three finished cotton fabrics was tested by the oscillation method (GB/T20944.3-2007):
the tested bacteria include Escherichia coli (E.coli, ATCC 25922) and Staphylococcus aureus (S.aureus, ATCC 6538), a single colony is picked from a solid plate and inoculated in LB liquid culture medium, the culture is carried out overnight at 37 ℃, then the bacterial liquid is diluted according to a ratio of 1:100 and inoculated in new LB culture medium to be cultured to a logarithmic phase, and the bacterial liquid is diluted to 10 degrees by a sterilized PBS buffer solution (PH is 7.2) in a gradient manner4CFU/mL. Respectively taking 0.8g of blank fabric with the same size and three types of finished fabrics (sterilized) in different 50mL test tubes, respectively adding 5mL of diluted bacterium liquid to immerse the fabrics, sealing, placing the fabrics in a shaking table, fully oscillating at 37 ℃ at 200r/min, taking samples every 2h, performing gradient dilution by 10 times by using PBS liquid, taking 100 mu L of the samples to a nutrient agar culture medium, uniformly smearing the samples to dryness, placing the dried samples in an incubator, culturing the samples for 16h at 37 ℃, taking the samples out, calculating the number of bacterial colonies in a plate, and drawing a curve in a graph 1. The plate count results of the 24h sample were calculated as follows:
note: w1The number of cotton cloth colonies after finishing; w0Number of blank cotton colonies.
Fig. 1 shows the dynamics of the antimicrobial activity of a cotton fabric finished with three antimicrobial agents according to an experimental example of the present invention on gram-positive bacteria (e. Experiments prove that the fabric BP1@ CF finished by the borneol-based antibacterial material has excellent and quick antibacterial effect under the conditions that various finishing conditions are the same and the mass concentrations of the three finishing agents are also the same.
Fig. 2 is a plate counting graph of antibacterial rate of a cotton fabric finished by three antibacterial agents to gram-positive bacteria (e.coli) and gram-negative bacteria (s.aureus) in an experimental example of the invention. Experiments prove that the antibacterial finishing agent has the antibacterial effect equivalent to or even better than that of the antibacterial finishing agent sold in the market.
Experiment two
Observing the anti-adhesion effect of the BP-b-HM finished cotton fabric with different concentrations on gram positive bacteria (E.coli) and gram negative bacteria (S.aureus) by using a field emission scanning electron microscope:
and detecting the condition of bacteria adhered to the surface of the sample by adopting SEM. Diluting the bacterial liquid to OD600Sterilized sample fabrics and blank fabrics (3 cm. times.3 cm) were each soaked in 25mL of the diluted bacterial solution described above (0.1). After incubation for 8h in a 37 ℃ incubator statically, the non-adhering bacteria were removed by washing 3 times with 5mL of sterile PBS solution (PH 7.2). (all vaccination experiments were repeated 4 times). Then solidified with 2.5% glutaraldehyde solution (in 0.1% phosphate buffer, pH 7.2) for 4 hours, and finally washed twice with phosphate buffer and once with distilled water. The textile was then dehydrated with an alcohol gradient (20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% and 100%), dried overnight, and after spraying with gold, the textile surface was observed for bacterial adhesion using FESEM.
FIG. 3 shows the anti-adhesion effect of BP-B-HM finished cotton fabrics with different concentrations on gram-positive bacteria (E.coli) and gram-negative bacteria (S.aureus) observed by a field emission scanning electron microscope, wherein (a) and (e) are control groups (original cotton fabrics), and B1-1, B1-2 and B1-3 respectively represent cotton fabrics finished with 10mg/mL, 20mg/mL and 40mg/mL borneol-based antibacterial copolymers (BP-B-HM). As can be seen, the bacterial adhesion on the fabric surface gradually decreased with increasing concentration of finish. At a finish concentration of 40mg/mL, bacterial adhesion to cotton fabric was essentially not observed. The results show that the borneol-based antibacterial copolymer (BP-b-HM) has excellent antibacterial adhesion effects.
In conclusion, the borneol-based macromolecular antibacterial material BP provided by the invention can be anchored with a substrate material containing hydroxyl through a covalent bond by a simple and environment-friendly hydrothermal method, so that a functional interface with lasting sterilization and anti-adhesion is formed, and the borneol-based macromolecular antibacterial material BP has the advantages of excellent antibacterial activity, effective sterilization and bacterial adhesion prevention.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.