Study On Antibacterial Biofunctional Coatings Made Of Self-Assembled Colloidal Particles

Implant infection has become a huge obstacle to the clinical application of biomedical metal implants.It is necessary and urgent to develop and research anti-implant infection materials.The essence of implant infection is bacterial contamination.Bacterial colonization and subsequent biofilm formation on the surfaces of medical materials has become one of the key factors to determine the success rate of implantation.Polymeric coatings with multiple antibacterial mechanisms is an effective strategy for solving the problems of bacterial colonization and biofilm formation.Although antibacterial materials and antibacterial mechanisms based on polymeric have received extensive attention in recent years,there are still many problems should be considered,including the accumulation of dead bacteria on the surface of bactericidal coatings,bacterial adhesion-resistant coatings which lacks the balance and regulation of corresponding biological activities,and unsatisfactory coating preparation process.Self-assembly of amphiphilic copolymer has attracted much attention during the past decade.Over the years,scientists have paid more attention to exploring the functionalization and application of polymeric self-assembled aggregates through multi-component and hierarchical assembly.These polymeric aggregates have been used in many promising fields,such as drug delivery,biological sensing,and interfacial catalysis.Amphiphilic block copolymers with regular structure are still the preferred for most of the application researches.However,the practical application of block copolymer-based self-assembled aggregates maybe limited by their complicated and time-consuming synthesis processes.In contrast,aggregates self-assembled from amphiphilic random copolymers with simple synthesis procedures and low costs have been developed in recent years.Random copolymer self-assemble colloidal particles can also be used as a container for loading bioactive component such as antibacterial,anchoring component,and drugs.Based on this feature,random copolymer colloid particles have great potential applications in functional coatings.Therefore,using random copolymer colloidal particles as building blocks may provide a simple and convenient method for constructing structure-controllable multifunctional coating materials.The practical application of self-assembled aggregates has very important theoretical value and practical significance.Considering the above aspects,antibacterial coatings with multifunctional and structure-controllable properties were prepared by using self-assembled colloid particles of amphiphilic random copolymers as building blocks,and the content of the thesis are described as follow.Firstly,the contact-based antibacterial coating were prepared on the surface of NiTi alloy through direct electrodeposition of the self-assembly colloidal particles from synthetic quaternary ammonium copolymers,and the accumulation of dead bacteria on the surface of antibacterial coatings was verified.Secondly,in order to solve the accumulation of dead bacteria on materials,we proposed an amphiphilic bacterial adhesion-resistant coating based on molecular design which could promote cell adhesion and proliferation,the mechanism of inhibiting bacterial adhesion and promoting cell adhesion were explored.Furthermore,based on the structure-activity relationship,the antibacterial coating materials with multiple mechanisms were constructed to further improve the antibacterial effect,the cell adhesion behavior was regulated by adjusting the copolymer structural parameters,the regulation mechanism were investigated in detail.Finally,the multi-function antibacterial coatings with better antibacterial effect were prepared through the copolymer structure and synthetic path optimization,and the mechanism of regulating cell adhesion based on hydrophobicity had also been verified.The work in this thesis includes the following parts:1.Quaternary ammonium antibacterial functional coatings on NiTi alloyThe contact-based antibacterial functional coating material were prepared on the surface of NiTi alloy.The ability of inhibition toxic Ni²⁺release and sterilization of the coatings was investigated.Firstly,the photo-sensitive quaternized copolymers Poly（7-hydroxy-4-methylcoumarin methacrylate-co-2-Hydroxyethyl methacrylate-co-2-（Dimethylamino）ethyl methacrylate）（PCHD-Q）with many hydroxyl and tertiary amine groups were synthesized,and self-assembled into colloidal particles in selective solvent dioxane/water.The paiticles size,morphology and surface charges and their responsive on pH,salt concentration,and UV irradiation were successfully characterized by transmission electron microscopy（TEM）,scanning electron microscopy（SEM）,ZetaPALS particle size and zeta potential analyzer.The antibacterial functional coatings were formed on the NiTi alloy by direct electrodeposition of these colloidal particles and subsequent photo-cross-linking process under deposition conditions of 15 V,5 min and UV irradiation conditions of 10 min.The formation mechanism of coating is the classical electrochemical particle agglomeration theory.SEM images revealed that photo-cross-linking process enhanced the compactness and uniformity of the coating materials.The coatings showed good protective properties for NiTi alloy in vitro environment and could significantly decrease the release of Ni²⁺through physical barriers and physical adsorption based on the-OH and-N（CH₃）₂ groups,which could obviously reduce cytotoxicity inducing by Ni²⁺.Meanwhile,the quaternized coating materials not only showed the low cytotoxicity against NIH-3T3 cells,but also obtained excellent antimicrobial activity against S.aureus and E.coli.This antibacterial coating with toxic ion adsorption ability have the potential for improving the safety and success rate of NiTi-based implants.In addition,the antibacterial results showed that large amounts of dead bacteria adhered to the sterilization surface.2.Hydrophobic zwitterionic-based coating materials with bacterial resistance but cell adhesion propertiesIn order to solve the accumulation of dead bacteria on materials,we proposed an amphiphilic bacterial adhesion-resistant coating based on molecular design which could promote cell adhesion and proliferation.The hydrophobized zwitterionic-based copolymer poly（sulfobetaine methacrylate-co-2-（dimethylamino）-ethyl methacrylate-co-isobornyl acrylate）（P（SBMA-co-DMA-co-ISA）,PSDI）containing a chiral structure（ISA）were successfully synthesized by free radical polymerization and subsequent sulfonation reactions.Together with tannic acid（TA）,PSDI colloidal particles（CP）were anchored onto the Ti alloy surface by the co-electrodeposition forming coating materials.The composition,morphology and surface properties of the CP-TA composite coatings were characterized by Fourier transform infrared spectroscopy（FTIR）,SEM,X-ray photoelectron spectroscopy（XPS）,Optical contact angle（OCA）and other methods.The results show that the introduced of TA improve the stability of coatings by participating into the noncovalent interactions including ionic pairing and hydrogen bonding.According to the in vitro bacterial adhesion test,the composite coatings could effectively inhibit the initial adhesion of bacteria through the synergistic of chiral surface,electrostatic shielding surface and a certain degree of hydration surface from chiral content ISA and zwitterionic content SBMA.Furthermore,the coatings are favorable to cell adhesion and growth.A key feature is that the introduced ISA increased the hydrophobicity of the coating and provided a moderately hydrophilic surface for cell adhesion and proliferation,which is of great significance for zwitterionic-based surfaces.This work provides a simple and practical strategy for preparing zwitterionic-based multifunctional coating.3.Multi-function antibacterial coating material with controllable cell adhesion behaviorOn the basis of improving the antibacterial activity of the coating,designing specific polymers and studying their structure on regulation of cell adhesion behavior is the focus of this chapter.Firstly,a series of amphiphilic zwitterionic-based copolymers P（SBMA-co-DMA-co-ISA）and P（DMA-co-ISA）were designed and synthesized with different monomer ratios and sulfonation degrees（marked as PDI-S and PDI）.By using TA,a natural polyphenol which can anchor to the surface of any substrate and is a highly-effective antibacterial agent,as the functional assembly unit,colloidal particles were obtained by hybrid self-assembly of copolymers and TA in ethanol.Then,these hybrid particles were deposited on Ti substrate with relatively optimized deposition conditions to construct multi-function antibacterial coating.The research results show that the hybrid coating has uniform and complete morphology,and shows good solution stability.The physicochemical properties of the coating surface are affected by polymer composition,sulfonation degree and TA assembly content.Furthermore,the hybrid coatings have excellent antibacterial properties and adjustable cell adhesion properties.The antibacterial mechanism is described as follows:the initial adhesion of bacteria was inhibited by increasing the ISA or SBMA functional components,and the already attached bacteria were killed by TA component,which significantly reduced the amount of dead/live bacteria on the coating surface.The regulation mechanism of cell adhesion was explained as that the hydrophilic and hydrophobic properties of hybrid coating surfaces could be regulated by changing the structural parameters of the copolymers,so as to realize the regulation of cell adhesion behavior.Wherein,the threshold of ISA content in copolymers to achieve antibacterial properties is 30%,and 50%of ISA content is the optimal value of the coating from promoting cell adhesion to inhibiting cell adhesion.Finally,animal experiments showed that hybrid coatings have potential applications in the field of implantable materials.4.Polycarboxybetaine zwitterionic-based multi-function antibacterial coating materialsIn order to verify the universality of regulation mechanism of cell adhesion behavior meanwhile not compromise the antibacterial properties,a more convenient control mode of bacterial and cell adhesion behavior was investigated.Polycarboxybetaine zwitterionic-based copolymers P（CB-tBu-co-EHA-co-DOPA）were successfully synthesized by free radical polymerization.The colloidal particles loaded with Cu²⁺were prepared in ethanol/isopropanol selective solvent and deposited on Ti substrate for constructing multi-function antibacterial coating.The composition,morphology and surface properties of the composite coatings were characterized by FTIR,SEM,XPS,OCA and other methods.The research results show that the coatings can be converted from quaternized surfaces to zwitterionic-based surfaces by the treatment of trifluoroacetic acid solution to remove the protective groups of carboxyl group.The hydrophilicity of the coatings increases obviously without affecting the solution stability.Therefore,the coating excellently controls cell adhesion behavior through this simple surface treatment process,which verify the feasibility and universality of the regulation mechanism of cell adhesion behavior based on the hydrophilicity of coating.Furthermore,the coatings have excellent antibacterial properties,and the resistance is derived from the bacterial resistance ability,contact-and release-bactericidal ability of the coatings.Indirect cytotoxicity assay showed that the cumulative released of Cu²⁺from the coatings with 60 days did not induce obvious cytotoxicity,suggesting that Cu²⁺loaded into the coating is biosafety.Animal experiments showed that the coating materials had good biocompatibility in vivo.Our coating system provide a simple and practical strategy for regulating cell adhesion and solving bacterial infections,which has potential applications in both orthopedics and cardiovascular fields.In summary,we used amphiphilic random copolymers and functional elements as the multi-component units of assembly to prepare colloidal particles.The obtained particles were then electrodeposited on the surface of Ti and its alloys to obtain antibacterial coating materials with multi-functional and structure-controllable properties.The results imply that the coatings have ideal antibacterial activity and controllable cell adhesion ability.Firstly,the obtained coatings with multiple antibacterial mechanisms were utilized for solving the problems of initial bacteria adhesion,biofilm formation,and dead bacteria adhesion on material surfaces.Meanwhile,we propose a specific mechanism for regulating cell adhesion behavior through the control of copolymer structures and assembly elements,and confirm its universality.This research provides a simple and convenient strategy for constructing multi-function coating materials to solve many problems faced by antibacterial coating.