【作者】 王洪；

【导师】 谭业邦；

【作者基本信息】 山东大学 ， 高分子化学与物理， 2012， 博士

【摘要】 把生物多胺小分子引入到聚合物(特别是结构规整的嵌段共聚物)中得到的多胺聚合物,与生物多胺小分子一样具有一系列的生物学性能,在生物体的生理过程中如细胞新陈代谢,DNA复制等起着非常重要的作用,而且聚合物的链结构有利于胺基的稳定和功能基团的发挥。本文系统介绍了胺基的性质,常见多胺聚合物的结构、合成方法以及在工业生产中的应用；并且研究了多胺聚合物的特征和在水溶液中的自组装结构和在生物材料方面的初步应用。研究了多胺聚合物均聚物或嵌段共聚物的合成方法,在水溶液中形成的聚集体形态,通过调控水溶液的pH值,水溶液的温度或者水溶液的盐浓度,来调节水溶液中聚合物自组装形成的聚集体的聚集形态。(1)制备了含胺结构的单体5-甲基丙烯酰胺基戊胺盐酸盐(MAAPA),以大分子引发剂单甲基聚乙二醇偶氮二异氰基戊酸(mPEG-ACVA)为引发剂在水溶液中通过自由基聚合的方法合成了pH敏感的多胺嵌段共聚聚合物。通过1H-NMR, GPC-MALLS和元素分析表征了聚合物的结构和分子量。此外,通过动态光散射(DLS),透射电镜(TEM),共振光散射研究了在水溶液中pH值,盐浓度与聚合物的自组装的聚集体尺寸大小。从DLS和TEM的实验结果可以得出,随着水溶液pH值的升高,聚合物中的亲水基团质子化的铵基逐渐去质子化,聚集体的疏水基团增加,而且去质子化后的氨基形成的氢键相互作用力增大,使得聚集体聚集,引起聚集体的尺寸变大,当聚集体中的质子化的NH3+越来越少时,聚集体就会沉淀出来；而当水溶液中盐的浓度增加时,溶液中的离子浓度变大,聚集体表面电荷作用力增加,使得聚集体尺寸变小。而共振光散射的强度和聚集体的大小有很大的关系,所以随着pH值的降低,盐浓度的增加,共振光散射强度减小(2)制备了多胺单体N’-(4-乙烯苄基)-1,5-戊二胺盐酸盐(VBPDA),以偶氮二异氰基戊酸(ACVA)为引发剂在水溶液中通过自由基聚合的方法合成了pH敏感的多胺聚合物。通过FTIR,1H-NMR和GPC-MALLS表征了聚合物的一级结构和分子量。此外,通过动态光散射(DLS),透射电镜(TEM),紫外光谱,共振光散射和荧光光谱计研究了在水溶液中pH值,盐浓度与聚合物的自组装的聚集体尺寸大小,光学性质的关系。从DLS和TEM的实验结果可以得出,水溶液的pH变化对水溶液中聚集体的尺寸影响比水溶液的盐浓度的影响要大：随着水溶液pH值的升高,聚合物中的亲水基团质子化的铵基逐渐去质子化,聚集体的疏水基团增加,而且去质子化后的氨基形成的氢键相互作用力增大,使得聚集体聚集,引起聚集体的尺寸变大,当聚集体中的质子化的铵基越来越少时,聚集体就会沉淀出来；而当水溶液中盐的浓度增加时,溶液中的离子浓度变大,电荷间作用力增强,使得聚集体尺寸变小。而共振光散射的强度和聚集体的大小有很大的关系,所以随着pH值的降低,盐浓度的增加,共振光散射强度减小。对于光学性质的影响,主要是由于发射基团芳香环周围的电荷变化,使得紫外吸收和发射荧光强度发生了改变,随着pH值的降低或者盐浓度的增加,荧光强度变大。(3)合成含有侧链正戊二胺的PVBPDA直链和亲水的mPEG直链组成的两嵌段多胺共聚物mPEG-b-PVBPDA。通过’H-NMR和GPC-MALLS表征了聚合物的结构和分子量。双亲水嵌段聚合物mPEG-b-PVBPDA在水溶液中能够自组装,改变水溶液的pH或者水溶液的温度来调节嵌段共聚合物的自组装行为。使用态光散射(DLS),紫外光谱,共振光散射和荧光光谱研究了聚合物在水溶液中自组装的聚集体尺寸大小,光学性质的关系与水溶液的pH值,温度的关系。随着水溶液的pH值的升高,嵌段共聚物中的质子化的氨基去质子化后,疏水链增多,水溶液中的小聚集体由于氨基的氢键的相互作用发生了聚集,而且随着pH值的升高,形成了以PVBPDA为核,mPEG为壳的核壳结构,所以随着pH值的升高,水溶液中聚集体的尺寸变大,DLS测得的流体力学半径随着pH值的升高而变大,共振光散射的强度也会升高。而随着pH值的升高,紫外吸收强度,荧光的强度由于发射官能团芳香环周围电子密度降低,所以强度都会减弱。而水溶液的温度变化,使得分子的动能增加,聚集体的尺寸变小,但是氨基形成的氢键作用力比较大,所以聚集体的大小随温度变化较小。同时初步研究了多胺嵌段共聚物分别与DNA, BSA形成的生物大分子在水溶液中的自组装行为与构象的变化。在对DNA转染后,形成了生物大分子,通过凝胶电泳研究了合成的三种嵌段共聚物与DNA的相互作用形成的聚电解质生物大分子的电泳,可以得出嵌段共聚物中氨基的含量越高,对DNA的转染效率越高；通过共振光散射研究了含有不同聚合物的聚电解质在水溶液中聚集体的尺寸变化,随着多胺聚合物的加入,聚集体尺寸变大,当达到1：1后,聚集体的尺寸变化比较小。也用圆二色光谱仪(CD光谱仪)来测量了DNA与不同量的多胺聚合物相互作用后生成的聚电解质的构象变化,发现直线型的多胺聚合物使得DNA的构象发生了改变,手性变低。而多胺聚合物与BSA形成的聚电解质在水溶液中的聚集体通过DLS,功能荧光来研究,随着多胺聚合物的加入,BSA和多胺聚合物中的氨基相互作用,生成大的生物大分子,形成的聚集体逐渐变大；用CD光谱仪测定了形成的生物大分子的圆二色谱光谱,通过CD Pro软件计算,其中生物大分子中的α-螺旋结构随着多胺聚合物的加入逐渐变少,而生物大分子中的p-螺旋结构随着多胺聚合物的加入逐渐增加,表明了多胺加入后,诱发蛋白质BSA的多肽键断裂,断裂后的多肽键重新组合后,形成了新的手性构象的多肽键。(4)水溶性的嵌段准聚轮烷大分子通过含有侧链戊二胺基团的嵌段聚合物mPEG-b-PVBPDA和葫芦脲CB[6]在水溶液中相互作用来制备。嵌段准聚轮烷可以通过调节聚合物与CB[6]不同的摩尔比来调配准聚轮烷中CB[6]的含量。含有不同量CB[6]的准聚轮烷的性质通过元素分析,核磁来表征并且推算出CB[6]的准确含量。这些准聚轮烷在水溶液中的自组装现象通过动态光散射(DLS),透射电镜(TEM),紫外光谱,共振光散射和荧光光谱来研究。同时,研究了的含有不同含量的CB[6]的嵌段准聚轮烷水溶液的聚集形态与水溶液温度的关系。DLS, TEM和共振光散射的实验数据表明随着随着CB[6]含量的增加,水溶液中准聚轮烷形成的聚集体的尺寸变大,可能是随着CB[6]的加入,与嵌段共聚物中的亲水的正戊二胺基团形成轮烷结构后,亲水性链端变为疏水链端,聚集体发生了膨胀；另外一个可能的原因是分子本身的体积随着CB[6]的加入也逐渐变大,这两种原因使得聚集体尺寸变大；由双亲水嵌段共聚物变为两亲性嵌段准聚轮烷,在水溶液中聚集成“核壳”结构,由于中间的核太大所以在TEM中外面的PEG链就没法显示出来。所以随着CB[6]含量的增加,聚集体尺寸逐渐变大,而且越来越紧密。也研究了水溶液的温度对于不同含量CB[6]的嵌段准聚轮烷的聚集体的聚集形态的影响。从DLS的数据可以看出,随着温度的升高,聚集体的尺寸逐渐变小,但是从35到45℃温度变化时,聚集体的尺寸变化不大。而且当温度为0℃时,由CB[6]与嵌段共聚物mPEG-b-PVBPDA按1：1摩尔比形成的嵌段准聚轮烷,在水溶液中形成的聚集体在水溶液中沉淀出来,嵌段准聚轮烷疏水链太多,形成的聚集体太大,由于低温下,分子运动动能小,所以产生了沉淀现象。共振光散射强度随温度变化的趋势与DLS测得的水溶液中聚集体随温度变化的趋势一致。更多还原

【Abstract】 In the field of polymer science it has been of great interest to combine properties of useful functional groups and polymers, especially well-defined block copolymers. The application of this approach provides a prospect of modifying polymer constitution to discovery new type of polymer which can be used as a pharmacological tool. Aminated polyelectrolytes have caused a great deal of attention because of their applications in drug delivery, immobilization matrices for enzymes and cells, and tissue engineering. As a matter of fact, the incorporation of primary amino groups onto the polymer backbone can tune both stabilization of polymer fragment and the useful biological activity of amino groups, which are among the most important classes of polyelectrolytes.Amino functionalized polymers aggregate to spherical, cylindrical, tubular and vesicular phases in selective solvent spontaneously. Such aggregations can be used as templates for making nano-materials, immobilization matrices for enzymes and cells, and tissue engineering, and applicated in drug delivery such as vesicles possessing nano-sized hollow cubages.We summarize the amino group properties; structures, synthesis and industrial applications of cationic polyelectrolyte; characteristics and aggregation of polyelectrolyte solution. In this paper, we study on the synthesis methods, conformation and self-assembly behavior of amino functional cationic homo-or block polyelectrolytes. We try to find out the relationship between the aggregation structures and self-assembly condition, by which to analyses the self-assembly mechanism and control the aggregation.(1) A water-soluble monomer methacrylamido-pentylamine hydrochloride (MAAPA) was synthesized. The block copolymers were prepared by free radical polymerization using macroinitiator bis[methoxy poly(ethylene glycol) ethyl]4,4’-(diazene-1,2-diyl)bis(4-cyano-pentanoate)(mPEG-ACVA) as the initiator. The structure and molecular weight of polymers were characterized by1H-NMR and GPC-MALLS. Aggregation behavior of the polymer in aqueous solution was investigated by dynamic light scattering (DLS), TEM and Resonance Light Scattering (RLS) spectra. The experimental results show that the fluorescence intensity of the aggregates reduces and the size of the aggregates increases due to the amino groups with increasing solution pH. It was demonstrated that polyelectrolyte having a pH-responsive polyamine segment in water in prompt response to pH. Also the NaCl concentration could effect the size of the aggregates.(2) A water-soluble monomer N1-(4-vinylbenzyl)-pentane-1,5-diamine dihydrochloride (VBPDA)(with cadaverine salts) was synthesized. The polymers with cadaverine side groups were prepared by free radical polymerization using4’,4’-azobis(4-cyanovaleric acid)(ACVA) as the initiator. The structure and molecular weight of polymers were characterized by FTIR,1H-NMR and GPC-MALLS. Aggregation behavior of the polymer in aqueous solution was investigated by dynamic light scattering (DLS), UV-spectrophotometer, RLS and fluorescence spectra. The experimental results show that the fluorescence intensity of the aggregates reduces and the size of the aggregates increases due to the cadaverine side groups with increasing solution pH. It was demonstrated that polyelectrolyte having a pH-responsive polyamine segment in water in prompt response to pH and the sizes of aggregates are in the range of23to406nm, increasing with the increase of pH.(3) A new pH-responsive diblock copolymer, methoxy poly (ethylene glycol)-b-poly[N1-(4-vinylbenzyl) pentane-1,5-diamine dihydrochloride](mPEG-b-PVBPDA). The monomer with cadaverine side group (N1-(4-vinylbenzyl) pentane-1,5-diamine dihydrochloride, VBPDA) and macro-initiator (mPEG-ACVA) were synthesized, respectively, and mPEG-b-PVBPDA was then obtained by free radical polymerization. The synthesis of mPEG-b-PVBPDA was confirmed by FTIR,1H NMR and GPC-MALLS measurements. At low pH, it is hydrophilic due to the protonation of the amine groups. With increasing pH, deprotonation occurs and the hydrophobicity of PVBPDA block increases. This molecular feature leads to interesting aggregation behavior of mPEG-b-PVBPDA in aqueous solutions at different pH as revealed by DLS measurements, TEM observations; RLS and fluorescence spectrometry. This polymer was further subjected to gene delivery evalutions and promising DNA transfection capacity has been found.In comparison, comb-like copolymers, consisting of the same hydrophobic PVBPDA backbone at high-pH, albeit with non-interaction hydrophilic mPEG polymer side chain, could self-assemble only into hollow nanoparticles of single-wall in aqueous media. Moreover, the copolymer aggregates exhibit a reversible change in fluorescence intensity in aqueous media within a pH range of2.6to10.8. Finally, the transfection activity of mPEG-b-PVBPDA as novel carrier for synthetic DNA in gene therapy was evaluated.(4) A novel water-soluble block polypseudorotaxanes is synthesized in water from cucurbituril (CB[6]) and a diblock copolymer, methoxy poly (ethylene-glycol)-b-poly[N1-(4-vinylbenzyl) pentane-1,5-diaminedihydrochloride](mPEG-b-PVBPDA), by simple stirring at room temperature. Driven by hydrophobic/hydrophobic and charge/dipole interactions, CB[6] beads are localized on pentamethylene units in side chains of3as found by NMR studies. The degree of threading, i.e., the average number of CB[6] beads per repeat recognition unit of mPEG-b-PVBPDA (denoted as q/n hereafter), can be controlled from0.2to1.0by varying the amount of CB[6] added. This molecular feature leads to interesting aggregation behavior of the polypseudorotaxanes in aqueous solutions at different q/n as revealed by DLS measurements, TEM observations, UV-vis and fluorescence spectrometry. The average hydrodynamic radius (Rh), the intensity of UV-vis absorption band,RLS and the fluorescence intensity (If) of the block polypseudorotaxanes in solution increase with the increasing of CB[6] threaded.The degree of CB[6] threaded can be controlled from0.25to1.0. The polypseudorotaxanes in aqueous solution have more rigid chain conformation because of the threaded CB[6]. The Rh, the intensity of UV-vis absorption band and If of the polypseudorotaxanes increases with the increasing amount of CB[6] threaded.更多还原