| Capillary electrophoresis is refer to the charged particle or ion depends on the high voltage electric field drive, directional movement behavior at different speeds in the capillary. Due to its high separation efficiency, less sample consumption, rapid separation time, easy operation and other characteristics, it is applied in various kinds of separation and analysis fields, such as applied to bio-macromolecules separation such as amino acid, nucleic acid, peptide and protein, etc. Fused-silica is a kind of material which is used to manufacture the capillary owing to its good thermal conductivity and ultraviolet transparency. However, under the effect of buffer solution, the capillary inner wall would be negatively charged due to the dissociation of silanol groups. When the capillary was used to separate protein, the interaction between capillary inner wall and proteins would result in protein adsorption. It would lead to the decrease of the separation efficiency and reproducibility of migration time, peak trailing, deformation, and even unable to achieve effective separation and so on.Numerous approaches have been applied to minimize this adsorption, such as extreme pH, zwitterionic additives, high ionic strength, but by far the most common and effective way is using covalent bonding or physical adsorption to form polymer coating on the capillary wall. Silanol groups on the capillary inner wall can be masked by the polymer coating, the electroosmotic flow is suppressed, and the interaction of protein-wall would be decreased, etc. From the aspects of stability of the coating, coating which covalently bonded onto the inner wall of capillary is more stability than the physically adsorbed coating, however, process of the preparation of covalently linked coating is cumbersome and difficult to control, and coating reproducibility is poor. Compared to the covalently bonded coating, process of the preparation of physically adsorbed coating is simple, easy to control, strong regeneration, and therefore in recent years the physically adsorbed coating is more and more get the favour of scientists.Around the preparation of physically adsorbed polymer coatings, the main results are as follows:1. The graft copolymers of hydroxyethylcellulose-graft-poly(2-(dimethylamino)ethyl methacrylate) (HEC-g-PDMAEMA) with different graft ratio were synthesized by using ceric ammonium nitrate initiator in aqueous nitric acid solution. Electroosmotic flow measurement results showed that the synthesized HEC-g-PDMAEMA graft copolymer coated capillary could suppress EOF effectively compared to the bare fused-silica capillary, and efficient separations of basic proteins were also achieved. The electrical charge of the coated capillary wall could be modulated by varying not only the pH of the running buffer, but also the grafting ratio of poly(2-(dimethylamino)ethyl methacrylate) grafts, which makes it is possible to analyze the basic and acidic proteins in the same capillary. The effects of poly(2-(dimethylamino)ethyl methacrylate) grafting ratio in HEC-g-PDMAEMA and buffer pH on the separation of basic proteins for capillary electrophoresis were investigated in detail. Furthermore, egg white proteins and milk powder samples were separated by the HEC-g-PDMAEMA coated capillary, respectively.2. A novel noncovalent adsorbed coating for capillary electrophoresis has been prepared and explored. This coating was based on quaternized poly(2-(dimethylamino)ethyl methacrylate)-block-poly(ethylene oxide)-block-poly(2-(dimethylamino)ethyl methacrylate)(QDED) triblock copolymer which was synthesized by atomic transfer radical polymerization(ATRP) in our laboratory. The polycationic polymer and the negatively charged fused-silica surface attracted each other through electrostatic interactions and hydrogen bonds. It was demonstrated that the coated capillaries provided an electroosmotic flow with reverse direction, and the magnitude of the electroosmotic flow can be modulated by varying the molecular mass of poly(2-(dimethylamino)ethyl methacrylate)(PDMAEMA) block and pH value of the buffer, respectively. The effects of the molecular mass of PDMAEMA block in QDED triblock copolymer and pH value of the buffer on the separation of basic proteins were investigated in detail. The triblock copolymer coatings showed higher separation efficiency, better migration time repeatability and would apply to wider range of pH than bare fused-silica capillary when used in separating proteins. Proteins from egg white were also separated respectively through this QDED triblock copolymer coated capillary. These results demonstrated that the QDED triblock copolymer coatings are suitable for analyzing biosample.3. Firstly, the poly(DMA-co-SBMA)s with different feed ratio (SBMA/DMA) were synthesized via the reversible addition fragmentation chain transfer (RAFT) polymerization. And then, X-ray photoelectron spectroscopy (XPS) and water contact angel (CA) were used to investigate the composition and hydrophilicity of poly(DMA-co-SBMA) coating formed on the glass slide surfaces. CA measurements revealed that the poly(DMA-co-SBMA) coating became more hydrophilic with the increment of feed ratio (SBMA/DMA), and at the same time the XPS results showed that the coating ability was also increased with the increment of feed ratio. Followed, the copolymer was applied to coat the fused-silica capillary inner wall and the coated capillary was used to separate the mixture of proteins (lysozyme, cytochrome c, ribonuclease A and α-chymotrypsinogen A) in a pH range from3.0to5.0. Under the optimum conditions, an excellent separation of basic proteins with peak efficiencies ranging from551000to1509000N/m had been accomplished within10min. Furthermore, the compare of separation efficiency among the bare, PSBMA and poly(DMA-co-SBMA) coated capillary was also investigated. |
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