【作者】 付金红；

【导师】 沈家骢； 计剑；

【作者基本信息】 浙江大学 ， 材料学， 2007， 博士

【摘要】 生物材料界面是决定材料生物相容性和生物功能性的关键科学问题。本论文针对生物医用材料的感染问题，将层层自组装的方法引入到具有抗茵功能的生物医用材料的界面设计中，依据弱聚电解质的pH敏感性和层间穿插或者层间扩散的特点，深入研究了组装条件影响多层膜组份、表面形貌和特殊润湿性的基本规律，进行了生物大分子协同抗菌多层膜、生物大分子负载纳米银多层膜及特殊润湿性多层膜三种不同的抗菌多层膜的功能界面设计。研究利用多层膜的生物大分子固定化作用，采用具有抗菌性能的壳聚糖(chitosan)作为聚阳离子，具有抗细菌黏附的肝素(heparin)作为聚阴离子层层自组装，依据弱聚电解质的pH敏感性和层间穿插的特点，调节多层膜最外层中壳聚糖和肝素的含量，构建杀菌、抗细菌黏附协同作用的功能表面。接触角、紫外光谱数据表明壳聚糖和肝素交替组装到基材表面。吸附到多层膜的聚电解质的量增长缓慢，呈现线性增长的方式。XPS和接触角数据表明，通过改变聚电解质的pH值可以控制多层膜组份的穿插程度，从而可以调节多层膜的组份和润湿性。抗菌试验结果表明，改变聚电解质溶液的pH值可以调节多层膜的杀菌性和抗细菌黏附性。以弱聚电解质生物大分子多层膜为模板，原位合成了有机(天然抗菌分子)-无机(纳米银)复合抗菌多层膜，实现了超强抗菌效果与长效抗菌的结合。研究中利用chitosan对金属银离子的缔合作用，形成chitosan／Ag~+复合，再与肝素层层自组装，将银离子装载进超薄膜内，并用抗坏血酸将银离子原位还原为纳米银。UV结果显示多层膜在414 nm左右出现纳米银的特征吸收峰。通过透射电镜发现，纳米银的尺寸取决于溶液中壳聚糖分子和银离子的比例以及还原剂抗坏血酸的pH。抗菌实验结果表明装载有纳米银的多层膜不仅显著地提高了多层膜的抗菌效果，而且表现出长效的抗菌性。这种抗菌、抗凝血及具有较好生物相容性的复合多层膜在医用抗感染领域具有较大的潜在应用价值。将弱聚电解质的pH敏感性和聚电解质在多层膜中的扩散特性相结合，首先报道了指数增长多层膜的pH增强特性。采用不对称pH的交替组装，即低pH的聚丙烯酸(poly(acrylic acid)，PAA)和高pH的聚乙烯亚胺(polyethylenimine，PEI)溶液交替组装，实现了多层膜从弱指数增长到超强的指数增长方式的转变，在五个双层内，多层膜的厚度增加到一个多微米。这种pH增强的指数增长行为为多层膜的快速制备提供了一种新的选择。进一步对这种pH增强指数增长的机理研究显示pH增强指数增长行为与弱聚电解质的pH敏感性以及PEI在多层膜中扩散相关，两种聚电解质溶液的pH值的交替变换会引起多层膜内弱聚电解质官能团的离子化程度的交替变化，这一变化会增加多层膜的固定电荷，放大PEI扩散出和扩散进多层膜的程度，从而形成了pH增强的多层膜快速指数增长行为。研究进一步发现，pH增强的指数增长多层膜可以自发地形成具有微纳复合二元结构的性貌，这种微纳二元结构的形成与PEI的扩散、相分离及聚电解质发生吸附时所采取的卷曲构象有关。在这种具有微纳复合结构的多层膜上化学气相沉积氟烷基硅烷薄层，可以获得超疏水表面，并可以通过调节聚电解质溶液的pH控制超疏水状态从Wenzel状态转变到Cassie状态。多层膜表面的微纳复合结构是形成超疏水的关键因素。如果将这种多层膜构建在疏水的基材上，多层膜的生长相对缓慢，但最终仍能得到微纳复合结构，经热交联和疏水处理后，多层膜可以从疏水的基材上取下，获得了具有超疏水特征的自支撑多层膜。以pH增强的指数增长多层膜为平台，构建了特征形貌尺寸从500 nm到5 um变化的多层膜表面，为系统研究具有微纳二元结构的多层膜的形貌尺寸和润湿性对细菌黏附的影响奠定了物质基础。细菌黏附实验结果表明，超疏水表面的特殊的自清洁性能够减少细菌的黏附。除超疏水表面外，其余多层膜表面的细菌黏附规律与它们表面的接触角变化趋势完全一致。研究中还发现多层膜表面的形貌对细菌黏附有很大的影响。特征尺寸小于或者等于细菌的尺寸时，细菌黏附较少；而大于细菌的尺寸时，细菌黏附的就较多。更多还原

【Abstract】 Infections arising from the use of biomaterials, such as vascular and urinary catheters, vascular implants, heart valves etc, have become a major concern in the biomedical industry. As only the biomaterials surface is in direct contact with the biological environment, the infections first of all depends on the biomaterials surface properties involved in reactions occurring at the biomaterials-biosystem interface. In the present work, the layer-by-layer (LbL) self-assembly technique was introduced to the surface modification of the biomaterials to improve their anti-bacterial properties. The effects of the pH of dipping solution on the composition, wettability and morphology of the weak polyelectrolyte multilayer films were systemtatically investigated, and two kinds of multilayer films with anti-fouling properties were designed, which will be introduced in the following section.Chitosan/heparin multilayer films and their anti-infection properties—Chitosan as an antibacterial agent and heparin as an anti-adhesive agent were alternatively deposited onto aminolyzed poly (ethylene terephthalate) (PET) to construct anti-infection multilayer films, and the pH of dipping solution was adjusted to change the composition of the multilayer films, and harmonize the anti-bacterial and anti-adhesive properties of the multilayer films to construct more powerful anti-infection coating. The contact angle and UV data verified the progressive buildup of the multilayer film by alternate deposition of the polyelectrolytes. The results of initial adhesion of Escherichia coli (E. coli) showed that the number of adhesive bacteria decreased with decreasing the assembly pH. The in vitro antibacterial test indicated that the multilayer of chitosan/heparin could kill the bacteria effectively and assembly pH has remarkable effect on the antibacterial property of the multilayer. Chitosan/heparin multilayer films were also used as a template to load nano-silver for more powerfull anti-infection coating. Chitosan-silver nitrate complex and heparin were alternately deposited onto an aminolyzed PET film surface, and subsequently, the silver ions within the multilayer films were reduced with ascorbic acid to form silver nanoparticles. UV-visible spectroscopy and transmission electron microscopyconfirmed the formation of well-dispersed nanosilver particles with sizes (10-40 nm) that depended on the initial concentration of silver ions in chitosan solution and the pH of ascorbic acid solution. The chitosan/heparin multilayer films were possessed of bactericidal effect on Escherichia coli, and this antibacterial effect could be significantly enhanced by the incorporation of silver nanoparticles into the multilayer films. The multilayer films containing nanosilver were not only effective as antibacterial but also as anticoagulant coating. And cell toxicity evaluation suggested that the multilayer films containing nanosilver did not show any cytotoxicity. The multilayer films containing nanosilver may have good potentials for surface modification of medical devices, especially for cardiovascular implants.pH amplified exponential growth multilayer films and their anti-fouling function—A facile method to amplify the growth of the exponential multilayers by the alternate deposition of PEI solution at high pH and PAA solution at low pH was demonstrated. A model was given to explain why pH can amplify the growth of multilayer films based on the mechanism of polyelectrolyte "diffusion in and out" of multilayers. Since both PEI and PAA are weak polyelectrolyte, the ionization degree of their functional groups change with the alternate deposition steps, which will facilitate the diffusion of the PEI molecules "in and out" of the whole film and amplify the amount of the polyelectrolytes deposited onto the surface during each adsorption step. The method of pH amplified exopential growth multilayers may provide a novel and convenient pathway to produce multilayer films more efficiently. A hierarchal micro and nano structures on the multilayers can be achieved simultaneously by deposition of PEI solution at high pH and PAA solution at low pH. When the multilayers were constructed with PAA solutions at low pH and PEI solution at high pH, such as PAA2.95/PEI9.00, PAA chains and PEI chains are adsorbed onto the surface with globular conformation due to its low ionization degree, at the same time, a pH-induced ionic-bond-breaking and morphological reorganization process happens in the pH alternative build-up steps. Larger aggregates from more loopy PAA chains and bigger pores by pH-induced morphological reorganization lead to a rougher surface with micro- and nano- structures on themultilayers. A superhydrophobic surface was developed from pH amplified exponential growth multilayers with micro- and nano- structures after being modified by a chemical vapor deposition of (tridecafluoroctyl) triethoxysilane. The pH amplified exponential multilayers with micro-nano structures can be constructed on the PEI-modified hydrophobic substrates such as Teflon, followed by depositing a thin layer of fluorinated silane onto the multilayers. Free-standing superhydrophobic multilayers could be prepared sucessfully after the multilayers were taken away from the hydrophobic subatrates. The amplified exponential growth multilayer films with characteristic size changing from several hundred nanometers to about five micrometers were constructed by adjusting the pH of dipping solution. The effects of the morphology and wettability of the surfaces of pH amplified exponential growth multilayer films on their anti-fouling properties were investigated. The biological tests suggested that the superhydrophobic surfaces having self-cleaning ability can resist the adhesion of the bacteria, and other surfaces increase their anti-fouling properties with increasing the hydrophilicity. The multilayers which structure size is less than or equal to the size of bacteria can more effectively resist the adhesion of bacteria, which is caused by the few contact area between bacteria and the surface.更多还原