Construction, Evaluation And Applications Of Packed Capillary Electrochromatography With Magnetically Immobilized Frits

Capillary electrochromatography (CEC) is a separation technique that combines the high separation efficiency of capillary electrophoresis with the superior selectivity of high-performance liquid chromatograph, which has a wide applied future in the biomedicine field. The problem of column preparation, especially frits fabrication, has hampered the development of CEC. Conventional techniques of frits fabrication, such as thermal sintering, pulling and in situ formation, have many disadvantages including time consuming, easy to generate bubble and bringing the fragility at the frit sections. In view of the strong responsibility and easy manipulation in the magnetic field of magnetic particles, packed CEC with magnetically immobilized frits was constructed in this paper. The details were summarized as follows:First of all, the magnetic microspheres and the frits were evaluated. The solvothermal reduction method was adapted to prepare magnetic microspheres with uniform particle sizes and strong magnetic responsibility, which met the demand of preparing the frits. The frits composed of magnetic microspheres demonstrated good penetrability and strong mechanical property.Secondarily, packed CEC with magnetically immobilized frits was prepared and the feasibility was verified. The packing procedure was summarized as follows: The frits were prepared by hydrodynamically introducing the magnetic microspheres which were immobilized by the magnets outside the capillary. Then the packing materials were packed into the capillary with magnetically immobilized frits to form the column. The feasibility was verified by reversed phase chromatography including C18 and phenyl column, which were used to separate some neutral compounds. As a result, the column efficiency was high enough, and the RSD of retention time, peak area and column efficiency was lower respectively than that of column with sinter frits. Based on changing the packing time, the columns with different column length were prepared, and the effect on electroosmotic mobilities and resolution by column length was investigated.At the last, the chromatography mode of packed CEC with magnetically immobilized frits was used in preparative chromatography and polymer packed CEC. (a) Thanks to the homogeneity of magnetically immobilized frits, large inside micrometer capillary can be used in packed column. In this paper, C18 packing materials were packed into 250μm i.d. capillary to investigate the preparative separation. The linearity of the voltage and current was 0.9986, which can indicate the stability of the preparative separation system. (b) Considering that there was no need to heating the capillary in the process of preparing frits, the novel CEC mode, polymer packed CEC, was proposed for the first time. In this work, polystyrene (PS) microspheres were packed into the capillary to form the column to separate some alkali compound.In summary, seeing that the magnetic microspheres can respond to external magnetic field, the packed CEC with magnetically immobilized frits was constructed. After the feasibility was verified successfully, preparative chromatography and polymer packed CEC were further investigated. In the future, polymer packed CEC can be used in protein analysis, and packed CEC with magnetically immobilized frits is promising for construction of chip-based CEC.