【作者】 成会玲；

【导师】 丁中涛；

【作者基本信息】 云南大学 ， 有机化学， 2012， 博士

【摘要】 分子印迹技术(Molecular Imprinting Technology, MIT)是在仿生科学和模拟自然界中酶-底物及受体-抗体作用的基础之上发展起来的一项技术。其基本原理是通过模拟自然界存在的分子识别作用,在聚合物材料中引入印迹分子识别位点,制备在空间结构和结合位点上与目标分子(印迹分子)完全匹配的聚合物／膜,即分子印迹聚合物/膜(Molecularly Imprinted Polymers/Membranes, MIP/MIM),以实现对目标分子的特异性选择。随着MIT研究的不断深入和应用领域的不断扩展,MIT已经在手性拆分、固相萃取、传感器、催化和有机合成等领域得到了应用,但MIP/MIM的研究仍然存在许多问题和不足,其中之一就是制备MIP/MIM所能使用的功能单体种类还很有限。目前关于新型功能单体设计和合成的研究报道还较少,制备MIP/MIM一般都是采用十来种常规的功能单体,而可供选择的模板分子种类相对较多且结构多样,因此制备的MIP/MIM还不能很好满足分子识别的要求。基于上述原因,本论文以非共价印迹法为基础,针对特定的目标分子(鬼臼毒素和靛玉红等)设计合成了一些可适用于分子印迹体系的新型功能单体,并把它们用来制备了一系列鬼臼毒素、靛玉红的MIP/MIM,通过MIP/MIM制备条件的优化及其性能测试,筛选出合适的功能单体。最后把优选出的MIP/MIM用于实际样品的富集分离,探讨了MIP/MIM的识别机理,为所合成的新功能单体在功能材料制备中的应用及所制备的MIP/MIM在复杂样品中的应用奠定了基础。其具体研究工作如下:针对特定的目标分子(鬼臼毒素和靛玉红等),设计合成了14个应用于共价或非共价印迹体系的新型功能单体(分别是酰胺类功能单体、手性功能单体及具有β-酮酸酯结构和p-氨基酯单元的功能单体)并取得良好的收率,采用1HNMR及13CNMR谱等分别对其结构进行了鉴定。以鬼臼毒素(PPT)为模板分子,自主设计合成的化合物(1-12)为新型功能单体,对鬼臼毒素分子印迹聚合物的合成条件及制备方法进行了优化。优化得到合成聚合物的最佳条件,即选用本体聚合法,以化合物2(L(-)-N-烯丙基吡咯烷基-2-羰基酰胺)为功能单体,氯仿为致孔溶剂,印迹分子、功能单体及交联剂比例为1：4:20。平衡结合实验表明,在氯仿中制备的分子印迹聚合物MIP14具有较大的吸附量和较高的印迹因子。接着实验结合超高效液相色谱(UPLC)分析方法,以MIP14为吸附剂成功用于混合样品溶液[含PPT结构类似物4’-去甲表鬼臼毒素(DMEP)]及桃儿七样品溶液中的PPT的固相萃取。在此基础上,采用表面接枝的方法(表面印迹法),考察了功能单体及交联剂用量、基膜种类、浸泡时间等因素对鬼臼毒素吸附性能的影响,制备了20个PPT分子印迹复合膜(MICMs)及相应的非印迹复合膜(NICMs)。平衡结合实验表明,以功能单体1-苯基-3-甲基-4-甲基丙烯酰基-5-吡啉酮(简称PMMP)制备的MICM对PPT具有较高的吸附容量和印迹因子。渗透实验表明,在其结构类似干扰物DMEP存在时,最佳膜MICM2对模板分子PPT表现出良好的渗透选择性。并在不同的pH条件下,研究了MICM2的渗透性能,实验研究表明MICM2具有良好的pH响应性(pH“色质”响应性)。通过扫描电镜(SEM)及氮气吸附等方法对MICM2进行了结构表征,并结合UPLC方法考察了MICM2对PPT和DMEP的甲醇混合标准样品及实际样品桃儿七甲醇提取液中鬼臼毒素的分离富集能力。以靛玉红为模板分子,自主设计合成的化合物(1-12)为新型功能单体,通过MIP制备方法的筛选及制备条件的优化,确定了合成MIP最佳的实验条件是：采用牺牲硅胶法,以化合物4(L(-)-N-烯丙基-2-羟基酰胺)为功能单体,THF为致孔溶剂,印迹分子、功能单体及交联剂比例为1：4:30。作为对照,采用上述实验条件,以常用的甲基丙烯酸(MAA)、丙烯酰胺(AM)以及4-乙烯基吡啶(4-VP)为功能单体,合成了3个MIPs。平衡吸附实验研究表明,与实验室常用的功能单体MAA、AM及4-VP相比,以化合物4为功能单体制备的MIP20对靛玉红具有更大的印迹因子和吸附容量,表明化合物4对于模板分子靛玉红来说是合适的功能单体。以MIP20为固相萃取微柱(MISPE)的吸附剂成功实现了板蓝根样品溶液中靛玉红的分离富集。在此基础上,以靛玉红为模板分子,考察了功能单体及交联剂用量、基膜种类、浸泡时间等因素对靛玉红吸附性能的影响,制备了一系列靛玉红分子印迹复合膜(MICMs)。探讨了优化得到的分子印迹复合膜(MICM20)对靛玉红和靛蓝甲醇溶液混合标准样品及实际样品板蓝根甲醇提取液的渗透选择性,膜渗透实验结果表明,MICM20对印迹分子靛玉红具有一定的富集能力。更多还原

【Abstract】 Molecular imprinting technique (MIT) is a technology developed on the basis of the bionic science, as well as the simulation of enzyme-substrate and antigen-antibody interactions in the nature. By introducing imprinting molecular recognition sites analogous to the natural effects of molecular recognition, a molecularly imprinted polymers/membranes (MIP/MIM) with spatial binding sites is artificially synthesized to achieve the specific selectivity to the target molecule (template molecule). With the development of the research and application concerning MIT, this technique has been widely used in many fields, such as the chiral separation, solid phase extraction, sensors, catalysis, organic synthesis, and so on. However, there are a number of unresolved problems in the MIP/MIM researches waiting for further efforts. One of them is that, while the template molecules are relatively diverse in terms of the types and structures, the number of available functional monomers is limited in the preparation of MIP/MIM. And there are few reports on the design and synthesis of new functional monomers up to now. Consequently, it is unable for the MIPs or MIMs to meet the requirements of recognition for template molecules with a wide range of molecular types and chemical structures.The goal of this paper is to design and synthesize a number of novel monomers applicable to molecularly imprinted systems by the non-covalent imprinting method, and use them to prepare MIPs/MIMs specific to target molecules including podophyllotoxin and indirubin. After optimizing the conditions of preparation and testing the performance, the optimum MIP or MIM with suitable functional monomer was finally selected and used in the enrichment and separation of active components in complex samples. The recognition mechanism of MIP/MIM was further speculated. This study laid a solid foundation for the applications of these new functional monomers in preparation of functional materials, as well as the applications of this group of MIP/MIM for the separation of complex samples. The details of this research work are as follows: Fourteen novel functional monomers belonging to three types, i.e. lactam functional monomer, chiral functional monomer, and functional monomer with β-keto ester structure or β-amino ester unit, were designed and synthesized for specific target molecules including podophyllotoxino, indirubin, etc. Their structures were identified by1HNMR and13CNMR spectra.The self-designed compounds (1-12) were synthesized and used as functional monomers for the preparation of MIPs for podophyllotoxin (PPT). The preparation methods and conditions of MIPs for podophyllotoxin were optimized. The optimal conditions for preparation of polymers are as follow: the compound2used as the functional monomer, chloroform as the porogenic solvent, ratio of template molecule to functional monomer to crosslinker as1:4:20. The equilibrium binding experiments show that the molecularly imprinted polymer prepared in chloroform (MIP14) has a larger adsorption capacity and higher imprinting factor. As a result, MIP14was successfully used as an adsorbent in the solid-phase extraction for PPT in the complex sample solution. In addition, a series of molecularly imprinted composite membrane (MICMs) for PPT were also prepared by the surface grafting method (surface imprinting method). The effects of the amounts of functional monomer and crosslinker, the type of base membrane, soaking-time and other factors on the adsorption properties of PPT were investigated. The equilibrium binding experiments indicated that the MICM2prepared using the functional monomer1-phenyl-3-methyl-4-methyl propylene acyl-5-Pyrazoline ketone (PMMP) has a higher adsorption capacity and imprinting factor. Permeation experiments showed that, in the presence of its analogue DMEP, the optimum membrane MICM2exhibited a good permeating selectivity to the template molecule PPT. Especially, it was found that MICM2has good pH sensitivity (pH-responsibility) in the permeating tests. The structural characteristic of MICM2was investigated by scanning electron microscopy (SEM) and nitrogen adsorption method. Finally, the separation and enrichment capacity of MICM2in PPT-and DMEP-methanol mixture and actual samples of methanol extracts of Himalayan mayapple was also examined by ultra-high performance liquid chromatography (UPLC) method. Compounds (1-12) were also used as the functional monomers for the preparation of MIPs for indirubin. The optimal experimental conditions of preparation of MIPs were determined as follow:Silica gel as sacrificial material, compound4as the functional monomer, THF as the porogenic solvent, ratio of template molecule to functional monomer to crosslinker as1:4:30. Moreover, methacrylic acid (MAA), acrylamide (AM) and4-vinylpyridine (4-VP) were used as functional monomer to prepareMIPs for indirubin, respectively. The equilibrium adsorption experimental studiesshowed that, compared with the general functional monomer MAA AM and4-VP, the MIP20prepared with the compound4as the functional monomer had a larger imprinting factor and adsorption capacity for indirubin. This implied that compound4is a suitable functional monomer for the template molecule indirubin. Using MIP20as sorbent in solid phase extraction micro-column (MISPE), indirubin was successfully separated and enriched from real sample solution. Amounts of functional monomer and crosslinker, the types of base membrane, soaking time duration and other factors were taken into account, a series of indirubin imprinted composite membrane (MICMs) and the control NICMs were also prepared. The selectivity of the optimum membrane (MICM20) in the standard mixture of indirubin and indigo, and in the sample solution of methanol extract of Radix Isatidis was investigated. The results of permeating experiments showed that MICM20is capable of enriching indirubin in complex solution.更多还原