Application Of P4VP-b-PEG-b-P4VP For Separation Of Basic Proteins By CE

The separation of proteins in complex biological mixtures is a challenging task and an important part in biomedical research field, which requires the most advanced separation technologies and sophisticated instrumentation available to human. Capillary zone electrophoresis (CZE) and its derivative methods are separation technologies very suitable for the analysis and separation of proteins and have great potential for becoming one of the key tools in proteome research, concerning its advantages of high efficiency, short analysis time and minimum consumption of both reagents and samples. In the applications, the analysis of human body fluids, agricultural products, and pharmaceuticals are focused on. Protein adsorption is the biggest problem confronted in the separation process, which causes a series of damages like low protein recovery, unstable baseline, poor repeatability and difficulties in qualitative and quantitative analysis, and eventually leads to a greatly decreased lifetime of the capillary column. Numerous approaches have been explored, in which physical adsorbed coatings are favored because of its easy to make and its pretty good stability, in order to minimize protein adsorption and to improve separation effiency.In this paper, we design and synthesis a cationic polymer coating and use it as a physical adsorbed coating of capillaries to separate basic proteins and to explore its mechanism of screening the adsorption of basic proteins. Major work was laying on the following aspects:1. A series of triblock copolymers of poly (4-vinylpyridine)-block-poly (ethylene glycol)-block-poly (4-vinylpyridine) (P4VP-b-PEG-b-P4VP) with different lengths of P4VP blocks were synthesized by atom transfer radical polymerization (ATRP), which were characterized by 1H NMR and GPC.2. Triblock copolymers of P4VP-b-PEG-b-P4VP were used as physical adsorbed coatings of capillaries for separating basic proteins. The effect on proteins separation of various buffer pH and different lengths of P4VP blocks of triblock copolymers was investigated and compared with diblock copolymers. The triblock copolymer was sensitive to buffer pH, because of which we can modify the EOF by changing the buffer pH in order to improve the resolution and to cut the separation time. 3. Electrostatic interaction between proteins and capillary surface was measured in bare and coated capillaries, in support of the shielding effect of cationic polymer coating towards basic proteins. Furthermore, we calculated the electrophoretic mobility of three basic proteins, in accord with the peak order which was determined by the pI and relative molecular mass of proteins.