Preparation Of Hydrophilic Bicontinuous Polystyrene Monliths By ATRP And Their Separation Application

With the continuous development of the bioengineering industry,biological macromolecules such as proteins and polysaccharides are widely used in the field of biotechnology.However,the structures of these biological macromolecules are complex,and it is difficult to separate and purify them.The organic polymer monolithic column has become a research hotspot in the field of biomacromolecule separation due to its simple preparation method,easy derivatization,good chemical stability,and high mechanical strength.However,it is difficult for the current organic polymer monolithic columns to balance mechanical strength,biocompatibility and separation efficiency.In this study,an amphiphilic diblock polymer(ADG)was firstly synthesized by the method of atom transfer radical polymerization(ATRP).Then the ADG and Pluronic 127 were used as composite surfactants to prepare the bicontinuous medium internal phase emulsion(MIPE)Finally,the hydrophilic bicontinuous polystyrene monoliths(HBPMs)were fabricated by using MIPEs as templates via electron activated regeneration ATRP(AGET ATRP)reaction in one step.The chromatographic performance of the HBPM in the field of rapid protein separation was preliminarily verified.This dissertation is divided into four parts.The first part is the synthesis of ADG.According to the synthetic route developed in our lab,diacetone-ADG was synthesized through esterification reaction,ATRP reaction and acid hydrolysis by using diacetone-D-glucose as the raw material.The hydrophilic-lipophilic balance(HLB)and the molecular weight of ADG were controlled by the reaction conditions of ATRP.The chemical composition of ADG was characterized by infrared absorption spectrum and hydrogen nuclear magnetic spectrum.The static contact angle of ADG was measured by a contact angle meter to verify its hydrophilicity and hydrophobicity.Finally,a surface tension meter was used to measure the critical micelle concentration(CMC value)of ADG in the oil phase.The rersult reveals that ADG can form reverse micelles at a lower concentration,which provides a basis for the subsequent preparation of MIPE.The second part is the preparation of bicontinuous MIPEs by the synergistic assembly of ADG and PF127 at the oil/water interface.By examining various conditions,it is concluded that the molecular weight of ADG affects the amount of ADG added,and the amount of water added between 45% and 70% can form a stable uniform emulsion structure.On this basis,the synergistic effect between ADG with different HLB values and PF127 with different concentrations on the structural properties of the emulsion was investigated.The structures and stability of the emulsion were observed by laser confocal,inverted fluorescence microscope and nanoparticle size analyzer.The third part is to prepare the HBPM in one step by using ADG as the ATRP macroinitiator via the AGET ATRP reaction of the bicontinuous MIPE.The factors that affecting the pore structures of the monolithic column were systemically investigated.The results indicate that the HBPM presented an ideal pore structure,uniform skeleton and a polysaccharide gel-like surface.The skeleton sizes of the monoliths were different depending on the molecular weights of ADG.The addition of water phase is controlled between 45%-60%to prepare the HBPMs with good permeability and high mechanical strength.In addition,there is a certain correspondence between the HLB value of ADG and the amount of PF127 added.Mercury intrusion analysis shows that the prepared HBPM has a bimodal pore size distribution.Through the measurement of the water contact angle,it was found that the participation of ADG can effectively mask the hydrophobic skeleton of polystyrene monolithic column,which is also beneficial to the post derivatization of HBPM.The fourth part is to study the chromatographic performance of the HBPM.Firstly,HBPM was used to directly separate glycoproteins in hydrophilic interaction chromatography mode.By changing the flow rate of the mobile phase,the separation speed of protein can be accelerated without reducing the resolution.Secondly,the DEAE group was introduced on HBPM to obtain a DEAE-HBPM anion exchange medium,which can effectively separate proteins with different isoelectric points(p I).The purifity of the eluted proteins was verified by SDS electrophoresis.It was preliminarily confirmed that the monolithic column has great potentials in the field of biomacromolecules separation.