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Study of Muti-separaction Characteristic Silica Hybrid Monolithic Column Based on Alkenyl Connection Terminations

Author: ChenYingZhuang
Tutor: YaoShouZhuo
School: Hunan University
Course: Analytical Chemistry
Keywords: Capillary electrophoresis Capillary electrochromatography Nano-Liquidchromatography Monolithic column Protein Electroosmotic flow Proteomicresearch Polymer monolith TMOS monolith VTMS monolith Enzyme Biological sample Enzyme microreactor On-line hydrolysis Biphase hybridmonolith On-line precocentration
CLC: O658
Type: PhD thesis
Year: 2012
Downloads: 39
Quote: 0
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Abstract


As one of the core technologies in nano/micro-separation field,monolithic columntechnology is increasingly attracting the attention of researchers, due to its high mass transferand excellent separation performance. Up to date, it has been successfully applied in almostall field of life sciences, such as proteomics, metabolomics etc. Monolithic columns aremainly divided into two types, organic polymer-based monoliths and silica-based monoliths.To the best of our knowledge, the organic monolithic column, characterized by a richdiversity of surface groups, is easy to swell when exposed to organic solvents. Compared tothe organic polymer monolith, the silica monolithic column has a more stable column bed, butthe monolith types are very limited, due to the small number of silanes and cumbersomederivation. So more wide application of monolithic column is greatly limited. As an emergingtechnology, the silica matrix hybrid monolithic column, which takes advantage of bothorganic monolith and silica monolith, can resolve the problem to some extent. However,related researches and applications are limited. Given this situation, we focus on thediversification of the silica-based monolith based on hybrid technology. The main content isas follows:1. effects of the porogen (PEG, urea) and temperature on pore structure formation ofTMOS monolithic column were investigated in detail. Further investigation was carried outusing vinyltrimethoxysilane (VTMS) and tetramethoxysilane (TMOS) as precursor forpreparation of VTMS hybrid monolithic column. The effects of porogen (PEG, urea) andtemperature on pore structure formation of VTMS monolith were also examined. A novelγ-MAPS-TMOS hybrid monolith was synthesized by using γ-methacryloxypropyltrimeth-oxysilane (γ-MAPS) instead of VTMS. Highly reactive vinyl groups were formed on thesurface, and further modification via thiol-ene click reaction and radical polymer reactionunder mild condition was achieved.2. Based on the hybrid organic-inorganic monolithic capillary column, a novel thiol-ene“click” strategy for the preparation of monolithic trypsin microreactor was proposed. It wasfound that the thio-ene click strategy is feasible and simple. In contrast to the conventionalazide-alkyne click reaction, the microreactor can give high catalytic efficiency and stability.The proposed thiol-ene click chemistry strategy is proved to be a promising method for thehighly selective immobilization of proteins under mild conditions, especially for enzymeswith free thiol radicals. 3. A novel method to synthesize sulfo/vinyl biphasic silica hybrid monolithic column byone step was developed for on-column preconcentration. In this method, sulfo-based segmentis located at the inlet of capillary column, which acts as preconcentration column. Close to thepreconcentration column, a vinyl functionalized segment is formed and serves as separationcolumn. Vinyl groups on vinyl functionalized segment are modified with ligand containingsulfhydryl group, such as octadecanethiol and6-mercapto-1-hexanol, via thiol-ene clickreaction. The applicability of this system is demonstrated by successful separation of closelyrelated amines. Good separation and enrichment are obtained. The proposed system is alsosuccessfully applied to complex biological samples, such as peptide, diluted bovine serumalbumin (BSA) hydrolysate, and the results indicate that the system has a capability forpreconcentration of low abundance peptides.4. A novel strategy for surface modification of silica-based monolith via one-pot methodwas developed in recent year. With this method, a polymer organic silica hybrid monolith canbe fabricated by adding polymer monomer into pre-condensation solution of VTMS monolith.The polymer integrated in monolith could provide better interaction between analytes andstationary phase. However, the adding monomer will lead to a change of composition inpre-condensation solution, and this change usually has greatly impact on pore structureformation of monolith. So the effect of adding monomer on the pore structure is veryimportant for the fabrication of monolith. In this work, the effect of monomer on the porestructure of monolith was investigated in detail. For stronger acidic monomer, the resultindicates that the adding amount of monomer would be depressed by the low pH value ofpre-condensation solution. In contrast, the adding amount of quaternary ammonium monomerwould be promoted under the strong acid conditions (pH <2). For weak acidic monomer,good pore structure of monolith can be obtained in a wide range of monomer concentrations.For weak alkaline monomer, adding amount was affected by many factors, such as porogen(PEG, urea) and temperature.5. A novel sulfoalkylbetaine-based zwitterionic organic-silica hybrid monolith wassynthesized by using N, N-dimethyl-N-methacrylamidopropyl-N-(3-sulfopropyl) ammoniumbetaine (DMMPPS, netural sulfoalkyl-betaine monomer). The key factors for effect of addingamount of zwitterionic monomer were proved to be pH value of monomer and temperature.The adding amount of zwitterionic monomer was significantly increased when DMMPPS wasused instead of N, N-dimethyl-N-methacryloxyethyl-N-(3-sulfopropyl) ammonium betaine(SPE), and this led to a significantly improved hydrophilicity of the monolith. Within thelinear velocity ranging from1to7mm/s, the DMMPPS-based organic-silica hybrid monolithexhibited good mechanical stability and excellent separation performance. The proposed monolithic column was successfully applied to separate purines/pyrimidines, benzoic acidderivatives and nucleotides, as well as tryptic digest of bovine hemoglobin (BHb) in a nano-hydrophilic chromatography (HILIC) mode, and the result demonstrated that such monolithhas the potential for separation of a variety of hydrophilic substances.

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