仿生抗生物黏附表面的设计、制备与性能研究
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摘要
“不良”生物黏附的危害是生物医用材料、生物化工、海洋船舶防污等研究和应用领域中涉及的一个共性的、具有基础性和普遍性的问题。自然界中存在着具有优异抗生物黏附特性的生物表面，如可保持表面清洁的海洋生物鲨鱼的皮肤、“出淤泥而不染”的荷叶表面以及具有生物黏附呈惰性的细胞外层膜等，它们所特有的表面化学组成和特异化学结构，以及表面的微/纳结构为我们构建抗生物黏附表面提供非常完美的仿生模板和仿生途径。从目前国内外学者就抗生物黏附表面而进行的研究思路、方法和研究结果来看，表面仿生无疑是最有效的策略。然而，生物表面在抗生物黏附过程中是以一个完整的生命体系发挥作用，因而单一的形貌结构仿生或者表面化学仿生往往难以构建稳定的抗生物黏附表面，更不可能构建出多功能的仿生表面。正是基于这一分析和认识，受到抗黏附的自然界各种生物表面的非光滑结构以及细胞外层膜的化学组成与化学结构的启迪，我们提出将仿生物非光滑表面的微米级结构与可呈现特殊浸润性的纳米结构相结合，以及将表面的形貌结构仿生与表面化学仿生相结合，构建新型复合仿生的抗生物黏附表面的研究思路。本工作的主要研究内容与研究结果如下所述：
1.以鲨鱼皮和植物(狗尾巴草)叶片为表面仿生的模板，将生物表面微复制法与表面热处理方法相结合，在仿生微结构表面构筑具有“荷叶效应”的纳米结构，制备出具有多尺度微/纳复合结构的仿生表面。研究表明：采用生物微复制法能够较精确地复制出天然模板表面的微米级精细结构，但难以有效地复制出纳米级的精细结构；火焰处理能够在通过微复制法得到的微米级结构表面构筑出纳米结构，由此制备出的微/纳复合结构仿生表面具有超疏水特性，水滴接触角大于150°，滚动角小于1°。
2.受细胞外层膜的性质及其化学结构的启发，设计与合成了几种含甜菜碱型两性离子基团的化合物，如硅烷基化两性离子单体SINNS-2和N,N-二甲基-N-甲基丙烯酰胺基丙基-N-丙烷磺酸内盐DMAPMAPS等，采用分子自组装技术和UV固化有机/无机杂化过渡层黏接技术的方法，在基材表面构筑具有亲水性仿细胞外层膜化学特性的自组装表面和聚合物凝胶层。此外，利用光引发表面接枝聚合法在仿鲨鱼皮微结构表面接枝甜菜碱型两性离子聚合物，借助两性离子与水之间的静电作用形成类似于鲨鱼表皮黏液的水化层，使所制备的表面具有复合仿生的特点。在此基础上，通过现代测试技术表征和分析了所制备的各种仿生表面的化学性质、微观形貌及其润湿性；利用流变仪初步探讨了仿生材料表面流变减阻特性，发现具有粘液特性的两性离子亲水表面存在滑移现象。
3.采用有机/无机杂化技术，用羧酸两性离子硅烷共聚物与纳米SiO2制备了有机/无机纳米复合物。用这一复合物为界面材料，采用雾化喷涂和热处理方法在玻璃基材上构建了一系列表面物化性质不同的SiO2杂化超亲水涂层表面。探讨了表面浸润性与表面微观结构和化学性质之间关系，并测量其防雾效果。实验结果表明，通过对表面化学性质和微观形貌的调控，可获得具有空气中超亲水、水下超疏油的润湿特性的表面，且涂层具有很好的防雾性能。
4.以经典Young氏方程、Wenzel及Cassie方程为理论基础，分析和解释了各种仿生表面的化学成分和粗糙度对表面亲/疏水润湿性能的影响，并探讨了超疏水、超亲水/水下超疏油表面的形成条件。针对两性离子亲水表面呈现出的水下超疏油特性，提出了新的见解，即表面亲水基团的水化作用是含两性离子表面亲水性表面具有水下疏油特性的关键。
5.采用蛋白质吸附、细菌黏附和藻类黏附等生物黏附试验，研究了各种仿生表面的物理/化学特性表面微观结构抗生物黏附性能之间的构效关系。实验发现：
(a)仿鲨鱼皮微结构表面并没有显示出抗生物粘附效果，反而会增加生物黏附量，且黏附更加牢固；
(b)复合仿生结构表面的微/纳结构所吸附的空气薄层作为一种物理屏障，能阻碍各种生物体的粘附，在抗生物黏附中起到关键作用。蛋白质等在微/纳结构表面的吸附，会导致表面空气层逐渐消失，影响材料的抗生物黏附性能的持久性或稳定性；
(c)本身具有低表面能性质的有机硅PDMS材料表面比具有微相分离结构的聚氨酯PU材料表面有着更加优异的抗粘附效果；仿生细胞外层膜的两性离子表面，如自组装分子层、聚合物凝胶层、表面接枝聚合物以及杂化涂层表面由于水化层的作用，均具有较好的抗生物黏附性能；
(d)通过调控SiO2杂化超亲水涂层表面的化学基团，可获得具有抗细菌黏附性能的材料表面。含有季铵盐基团的SiO2-PCM表面具有杀菌和抑制细菌繁殖的作用；而经过碱水解之后得到的SiO2-PCMZ表面，在不具有杀菌灭菌作用基础上，也表现出优异的细菌黏附性能。
The harm of adverse bioadhesion has been a fundamental problem that has commonfeatures and popularly exists in the research and application areas of biomedical materials,biochemical analysis, bological chemical engineering and antifouling of marine vessels. Innature, many biological surfaces can exhibit an excellent anti-bioadhesive performance. Forinstance, a shark skin can keep marine organisms away from it, a lotus leaf unstained by dirtshows an outstanding self-cleaning effect, phosphorylcholine (PC) or headgroups of a cellouter membrane is inert to bioadhesion, and so forth. The chemical compositions, specificchemical structure and micro-nano structures of these biological surfaces provide perfectbiomimetic templates and ways for constructing anti-bioadhesive surfaces. Judging from thecurrent research ideas, methods and findings involving the anti-bioadhesive surfacesconducted by domestic and foreign scholars, constructing biomimetic surfaces is undoubtedlythe most effective strategy. However, most biological surfaces play a role in the course ofanti-bioadhesion as a part of a complete life system, and thus it is often difficult to build astable anti-bioadhesive surface just by a single means of imitating morphology or chemicalcharacteristics of a biological surface, not to mention a multi-functional biomimetic surface.Based on the above analysis and understanding, and inspired by the anti-adhesive non-smoothsurfaces of plants and animals in nature and the chemical composition and molecularstructures of the cell outer membrane, herein we first put forward a research thought ofconstructing compound biomimetic surfaces with resistance to bioadhesion via a novelmethod involving (a) the combination of the micron-scale structures of biological non-smoothsurfaces and their nanoscale structures producing special wettability, and (b) the combinationof the biomimetic strategies of both the surface morphology and the surface chemistry. Themain research work and results are described below.
(1) Using shark skin and grass leaves (Green Bristle grass Herb) as a template,respectively, the biomimetic surfaces with micro-nano hierarchical structures were fabricatedby constructing nanoscale structure producing “lotus effect” on the biomimetic micron-scalestructured surfaces, which was conducted by combining the micro-replication method withthe flame treatment of a surface. The studies indicated that the microstructures on the natural templates can be exactly replicated by means of surface micro-replication, while thenanostructures are difficult to be faithfully replicated. Flame-treatment is an effective methodto construct nanostructures on the biomimetic surfaces prepared via the micro-replication. Theas-prepared hierarchical (micro-nano) surfaces can exhibit super-hydrophobic performance,whose measured water contact angle is greater than150°. And the slip angle for water dropletmerely reaches a limiting value of1°.
(2) Inspired by the chemical composition and molecular structures of a cell outermembrane, a few of betaine zwitterionic compounds, zwitterionic silane (SINNS-2),3-dimethyl(3-(N-methacrylamido) propyl) ammonium propane sulfonate (DMAPMAPS), andso on, were designed and synthesized. Then these functional molecules were used to preparechemically biomimetic self-assembled layer and polymer hydrogel layer by means ofmolecular self-assembly and UV curing of organic-inorganic hybrid transition layer bondingtechnology, respectively. Besides, zwitterionic polymer was grafted onto the surface ofbiomimetic shark skin via photo-induced polymerization. So as to form the mucus-likehydration layer with the aid of the electrostatic action between zwitterions and watermolecules. In this way, the prepared surfaces can possess unique compound biomimeticstructures. On this basis, the chemical parameters, morphology and wettability of the preparedbiomimetic surfaces were characterized and analyzed by modern testing techniques. The dragreduction effect of the biomimetic surfaces was preliminarily investigated based on rheometry,the results show that the slip phenomenon of slip phenomenon can occur on the zwitterionichydrophilic surface.
(3) The organic-inorganic nanocomposites comprising zwitterionic carboxybetaine silanecopolymer and silica nanoparticles was prepared via an organic-inorganic hybrid technology.Using these nanocomposites as interfacial materials and a glass slide as a substrate, a series ofpolymer-SiO2hybrid coatings with different physical and chemical properties were obtainedby spray-deposition and heat treatment. The influence of the chemical and morphologicalfactors of a surface on its wettability was investigated. Furthermore, the fogging behavior ofthe hybrid coating was measured. The results indicate that the biomimetic surface uniquewettability, or superhydrophilicity in air and underwater super-oleophobicity, can beconstructed by regulating the chemical features and morphology of surface. And such a biomimetic surface can possess good antifogging performance.
(4) According to the theories of classical Young's equation, Wenzel's equation andCassie's equations, the effects of the chemical composition and roughness of variousbiomimetic surfaces on their wettability were analyzed and explained, and the conditionsunder which a super-hydrophobic, super-hydrophilic and/or underwater super-oleophobicsurface can be formed were explored and discussed. Owing to the fact that thezwitterion-containing hydrophilic surface can exhibit underwater super-oleophobicity, a newinsight is proposed that the hydration of the hydrophilic groups on the zwitterion-containinghydrophilic surface plays a key role in making it possess underwater oleophobiccharacteristic.
(5) The structure-property relationships between the anti-bioadhesive properites ofbiomimetic surface and their physical, chemical properties and microstructures were studiedby a few of bioadhesion experiments including protein adhesion test, bacteria attachmenttest using S. aerues and E. coli as a test strain and algae adsorption tests in the static anddynamic aqueous environment. The following conclusions can be drawn from theexperimental results.
(a) The surface with biomiemtic shark skin microstructure cannot exhibit resistance tobioadhesion. On the contrary, a large number of organisms will adhere firmly to thesurface.
(b) As a physical barrier, the thin air layer adsorbed on the micro-nano hierarchical structureof the compound biomimetic surface can hinder the attachment of organisms, therebyplaying an important role in resisting bioadhesion. The adsorption of proteins and theother substances on the micro-nano structured surface will drive air away and make the airlayer disappear gradually, thus degrading the durability and stability of itsanti-bioadhesive performance.
(c) The PDMS-based surface with low surface energy can exhibit better anti-bioadhesiveperformance than do the PU-based surface with microphase separation structure. Due tothe existence of hydration shell, the zwitterion-containing surfaces imitating the chemicalfeatures of a cell outer membrane, including self-assembly molecular layer, polymeric gel,polymer surface obtained by surface graft polymerization, and hybrid coating, were a novel mimic of the cell outer membrane, and such surfaces often possessed excellentanti-bioadhesion performance.
(d) The biomimetic surface with anti-bacterial adhesion property using polymer-SiO2hybridnanocomposites as an interfacial material can be achieved through tailoring the chemicalfunctional groups of the hybrid nanocomposites. The surface SiO2-PCM bearingquaternary ammonium salt group can sterilize bacterial cells and inhibit bacterialreproduction; and the surface SiO2-PCMZ obtained by the alkaline hydrolysis can possessgood anti-bacterial adhesion property, but SiO2-PCMZ will lost its function of sterilizingbacterial cells.
1.以鲨鱼皮和植物(狗尾巴草)叶片为表面仿生的模板，将生物表面微复制法与表面热处理方法相结合，在仿生微结构表面构筑具有“荷叶效应”的纳米结构，制备出具有多尺度微/纳复合结构的仿生表面。研究表明：采用生物微复制法能够较精确地复制出天然模板表面的微米级精细结构，但难以有效地复制出纳米级的精细结构；火焰处理能够在通过微复制法得到的微米级结构表面构筑出纳米结构，由此制备出的微/纳复合结构仿生表面具有超疏水特性，水滴接触角大于150°，滚动角小于1°。
2.受细胞外层膜的性质及其化学结构的启发，设计与合成了几种含甜菜碱型两性离子基团的化合物，如硅烷基化两性离子单体SINNS-2和N,N-二甲基-N-甲基丙烯酰胺基丙基-N-丙烷磺酸内盐DMAPMAPS等，采用分子自组装技术和UV固化有机/无机杂化过渡层黏接技术的方法，在基材表面构筑具有亲水性仿细胞外层膜化学特性的自组装表面和聚合物凝胶层。此外，利用光引发表面接枝聚合法在仿鲨鱼皮微结构表面接枝甜菜碱型两性离子聚合物，借助两性离子与水之间的静电作用形成类似于鲨鱼表皮黏液的水化层，使所制备的表面具有复合仿生的特点。在此基础上，通过现代测试技术表征和分析了所制备的各种仿生表面的化学性质、微观形貌及其润湿性；利用流变仪初步探讨了仿生材料表面流变减阻特性，发现具有粘液特性的两性离子亲水表面存在滑移现象。
3.采用有机/无机杂化技术，用羧酸两性离子硅烷共聚物与纳米SiO2制备了有机/无机纳米复合物。用这一复合物为界面材料，采用雾化喷涂和热处理方法在玻璃基材上构建了一系列表面物化性质不同的SiO2杂化超亲水涂层表面。探讨了表面浸润性与表面微观结构和化学性质之间关系，并测量其防雾效果。实验结果表明，通过对表面化学性质和微观形貌的调控，可获得具有空气中超亲水、水下超疏油的润湿特性的表面，且涂层具有很好的防雾性能。
4.以经典Young氏方程、Wenzel及Cassie方程为理论基础，分析和解释了各种仿生表面的化学成分和粗糙度对表面亲/疏水润湿性能的影响，并探讨了超疏水、超亲水/水下超疏油表面的形成条件。针对两性离子亲水表面呈现出的水下超疏油特性，提出了新的见解，即表面亲水基团的水化作用是含两性离子表面亲水性表面具有水下疏油特性的关键。
5.采用蛋白质吸附、细菌黏附和藻类黏附等生物黏附试验，研究了各种仿生表面的物理/化学特性表面微观结构抗生物黏附性能之间的构效关系。实验发现：
(a)仿鲨鱼皮微结构表面并没有显示出抗生物粘附效果，反而会增加生物黏附量，且黏附更加牢固；
(b)复合仿生结构表面的微/纳结构所吸附的空气薄层作为一种物理屏障，能阻碍各种生物体的粘附，在抗生物黏附中起到关键作用。蛋白质等在微/纳结构表面的吸附，会导致表面空气层逐渐消失，影响材料的抗生物黏附性能的持久性或稳定性；
(c)本身具有低表面能性质的有机硅PDMS材料表面比具有微相分离结构的聚氨酯PU材料表面有着更加优异的抗粘附效果；仿生细胞外层膜的两性离子表面，如自组装分子层、聚合物凝胶层、表面接枝聚合物以及杂化涂层表面由于水化层的作用，均具有较好的抗生物黏附性能；
(d)通过调控SiO2杂化超亲水涂层表面的化学基团，可获得具有抗细菌黏附性能的材料表面。含有季铵盐基团的SiO2-PCM表面具有杀菌和抑制细菌繁殖的作用；而经过碱水解之后得到的SiO2-PCMZ表面，在不具有杀菌灭菌作用基础上，也表现出优异的细菌黏附性能。
The harm of adverse bioadhesion has been a fundamental problem that has commonfeatures and popularly exists in the research and application areas of biomedical materials,biochemical analysis, bological chemical engineering and antifouling of marine vessels. Innature, many biological surfaces can exhibit an excellent anti-bioadhesive performance. Forinstance, a shark skin can keep marine organisms away from it, a lotus leaf unstained by dirtshows an outstanding self-cleaning effect, phosphorylcholine (PC) or headgroups of a cellouter membrane is inert to bioadhesion, and so forth. The chemical compositions, specificchemical structure and micro-nano structures of these biological surfaces provide perfectbiomimetic templates and ways for constructing anti-bioadhesive surfaces. Judging from thecurrent research ideas, methods and findings involving the anti-bioadhesive surfacesconducted by domestic and foreign scholars, constructing biomimetic surfaces is undoubtedlythe most effective strategy. However, most biological surfaces play a role in the course ofanti-bioadhesion as a part of a complete life system, and thus it is often difficult to build astable anti-bioadhesive surface just by a single means of imitating morphology or chemicalcharacteristics of a biological surface, not to mention a multi-functional biomimetic surface.Based on the above analysis and understanding, and inspired by the anti-adhesive non-smoothsurfaces of plants and animals in nature and the chemical composition and molecularstructures of the cell outer membrane, herein we first put forward a research thought ofconstructing compound biomimetic surfaces with resistance to bioadhesion via a novelmethod involving (a) the combination of the micron-scale structures of biological non-smoothsurfaces and their nanoscale structures producing special wettability, and (b) the combinationof the biomimetic strategies of both the surface morphology and the surface chemistry. Themain research work and results are described below.
(1) Using shark skin and grass leaves (Green Bristle grass Herb) as a template,respectively, the biomimetic surfaces with micro-nano hierarchical structures were fabricatedby constructing nanoscale structure producing “lotus effect” on the biomimetic micron-scalestructured surfaces, which was conducted by combining the micro-replication method withthe flame treatment of a surface. The studies indicated that the microstructures on the natural templates can be exactly replicated by means of surface micro-replication, while thenanostructures are difficult to be faithfully replicated. Flame-treatment is an effective methodto construct nanostructures on the biomimetic surfaces prepared via the micro-replication. Theas-prepared hierarchical (micro-nano) surfaces can exhibit super-hydrophobic performance,whose measured water contact angle is greater than150°. And the slip angle for water dropletmerely reaches a limiting value of1°.
(2) Inspired by the chemical composition and molecular structures of a cell outermembrane, a few of betaine zwitterionic compounds, zwitterionic silane (SINNS-2),3-dimethyl(3-(N-methacrylamido) propyl) ammonium propane sulfonate (DMAPMAPS), andso on, were designed and synthesized. Then these functional molecules were used to preparechemically biomimetic self-assembled layer and polymer hydrogel layer by means ofmolecular self-assembly and UV curing of organic-inorganic hybrid transition layer bondingtechnology, respectively. Besides, zwitterionic polymer was grafted onto the surface ofbiomimetic shark skin via photo-induced polymerization. So as to form the mucus-likehydration layer with the aid of the electrostatic action between zwitterions and watermolecules. In this way, the prepared surfaces can possess unique compound biomimeticstructures. On this basis, the chemical parameters, morphology and wettability of the preparedbiomimetic surfaces were characterized and analyzed by modern testing techniques. The dragreduction effect of the biomimetic surfaces was preliminarily investigated based on rheometry,the results show that the slip phenomenon of slip phenomenon can occur on the zwitterionichydrophilic surface.
(3) The organic-inorganic nanocomposites comprising zwitterionic carboxybetaine silanecopolymer and silica nanoparticles was prepared via an organic-inorganic hybrid technology.Using these nanocomposites as interfacial materials and a glass slide as a substrate, a series ofpolymer-SiO2hybrid coatings with different physical and chemical properties were obtainedby spray-deposition and heat treatment. The influence of the chemical and morphologicalfactors of a surface on its wettability was investigated. Furthermore, the fogging behavior ofthe hybrid coating was measured. The results indicate that the biomimetic surface uniquewettability, or superhydrophilicity in air and underwater super-oleophobicity, can beconstructed by regulating the chemical features and morphology of surface. And such a biomimetic surface can possess good antifogging performance.
(4) According to the theories of classical Young's equation, Wenzel's equation andCassie's equations, the effects of the chemical composition and roughness of variousbiomimetic surfaces on their wettability were analyzed and explained, and the conditionsunder which a super-hydrophobic, super-hydrophilic and/or underwater super-oleophobicsurface can be formed were explored and discussed. Owing to the fact that thezwitterion-containing hydrophilic surface can exhibit underwater super-oleophobicity, a newinsight is proposed that the hydration of the hydrophilic groups on the zwitterion-containinghydrophilic surface plays a key role in making it possess underwater oleophobiccharacteristic.
(5) The structure-property relationships between the anti-bioadhesive properites ofbiomimetic surface and their physical, chemical properties and microstructures were studiedby a few of bioadhesion experiments including protein adhesion test, bacteria attachmenttest using S. aerues and E. coli as a test strain and algae adsorption tests in the static anddynamic aqueous environment. The following conclusions can be drawn from theexperimental results.
(a) The surface with biomiemtic shark skin microstructure cannot exhibit resistance tobioadhesion. On the contrary, a large number of organisms will adhere firmly to thesurface.
(b) As a physical barrier, the thin air layer adsorbed on the micro-nano hierarchical structureof the compound biomimetic surface can hinder the attachment of organisms, therebyplaying an important role in resisting bioadhesion. The adsorption of proteins and theother substances on the micro-nano structured surface will drive air away and make the airlayer disappear gradually, thus degrading the durability and stability of itsanti-bioadhesive performance.
(c) The PDMS-based surface with low surface energy can exhibit better anti-bioadhesiveperformance than do the PU-based surface with microphase separation structure. Due tothe existence of hydration shell, the zwitterion-containing surfaces imitating the chemicalfeatures of a cell outer membrane, including self-assembly molecular layer, polymeric gel,polymer surface obtained by surface graft polymerization, and hybrid coating, were a novel mimic of the cell outer membrane, and such surfaces often possessed excellentanti-bioadhesion performance.
(d) The biomimetic surface with anti-bacterial adhesion property using polymer-SiO2hybridnanocomposites as an interfacial material can be achieved through tailoring the chemicalfunctional groups of the hybrid nanocomposites. The surface SiO2-PCM bearingquaternary ammonium salt group can sterilize bacterial cells and inhibit bacterialreproduction; and the surface SiO2-PCMZ obtained by the alkaline hydrolysis can possessgood anti-bacterial adhesion property, but SiO2-PCMZ will lost its function of sterilizingbacterial cells.
引文
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