The Research On Modification Of Cellulose And Application In Separation Of Biomacromolecule

Capillary electrophoresis (CE) has been proven to be a powerful method for DNA and protein analysis. It offers great advantages over conventional slab gel electrophoresis in terms of higher resolution, shorter analysis time, minimal sample requirement, negligible waste of toxic chemicals, high detection sensitivity, and ease of automation. Like the other chromatography technique, the capillary electrophoresis column is the most important component. Although some biomacromolecules can be analyzed in the bare fused-silica capillaries, the separation medium has played an important role in high performance CE for DNA and protein separations. There are many reasons for modification of the capillary wall in electrophoresis. Goals may include reduction or elimination of analyte-wall interactions, alteration of electroosmotic flow (EOF) for rapid separation, improved reproducibility, and better resolution of particularly difficult separation problems. Base on these findings, we designed work on the following aspects:1. HEC and LPA composite matrices for DNA sequencingA network was formed by matrix of HEC and LPA for DNA sequencing, trying to combine the high sieving ability of LPA and the dynamic coating ability of HEC. The results showed that under the appropriate ratio of HEC to LPA, the networks yielded a read length of up to 900 bases of contiguous sequence in 73 min without optimized base-calling software.2. Novel hydroxyethylcellulose graft copolymer for separation of double-stranded DNA fragmentsAn ideal separation medium for dsDNA separations using CE should possess the following properties: high sieving ability, dynamic coating ability, and relatively low viscosity. In this work, hydroxyethylcellulose-graft-polyacrylamide (HEC-g-PAM) copolymer was synthesized using atom transfer radical polymerziation (ATRP). The reason we chosen HEC as the backbone is that HEC has both good sieving ability and dynamic coating ability for DNA separation. HEC-g-PAM with well-defined structures was applied for the first time as a medium for the separation of dsDNA in the bare fused-silica capillaries. The effectiveness of HEC-g-PAM for dsDNA fragment analysis was evaluated in comparison with that of HEC conventionally used for this purpose. Furthermore, the influences of PAM graft length on the separation performances of dsDNA were also discussed.3. Cationized hydroxyethylcellulose as a novel, adsorbed coating for basic protein separationProtein is another important biomacromolecules. In practice, the efficiencies typically achieved in protein separations by CE are considerably lower because of different types of interactions, including hydrogen-bonding, electrostatic, and hydrophobic interactions, between the proteins and the silica wall of the capillary. In order to minimize protein adsorption and stabilize EOF, a large amount of polymers have been explored to form stable dynamic coatings on capillary wall and show high efficiency in reducing protein adsorption. In previous studies, the cationic polymer derivatives were shown to be capable of suppressing the adsorption of fluorescently labeled amino acids, peptides, and proteins. Therefore, we take account of introducing cationized groups into HEC, then using cationized hydroxyethylcellulose (cat-HEC) as novel, hydrophilic, adsorbed capillary coating for electrophoretic protein analysis. The reason we chosen cat-HEC is that the HEC backbone has good sieving ability and the cat-HEC polymer contains cationic groups as substituents which can form ionic bond with the negative silanol groups on the silica surface. The results showed that highly efficient and rapid protein separation has been obtained over a wide pH range. These coatings demonstrated the excellent reproducibility of migration times of the three proteins, with relative standard deviation (RSD) values less than 1.0%.4. Novel hydroxyethylcellulose graft copolymer for protein separationNew separation medium, hydroxyethylcellulose-graft-poly(4-vinylpyridine) (HEC-g-P4VP), synthesized by using ceric ammonium nitrate (CAN) initiator in aqueous nitric acid solution, used for protein separation by CE is presented. The properties of HEC-g-P4VP as coating of capillary wall for basic, acidic and neutral protein separation were studied. The polymer efficiently coated the capillary inner surface by a dynamic process, thereby leaded to high efficiency, reproducibility and recovery of proteins analyzed by CE.