Fabrication of Monodisperse Porous Silica Microspheres with a Tunable Particle Size and Pore Size for Protein Separation

Monodisperse porous silica microspheres with a tunable particle size and pore size were fabricated by utilizing porous polymer microspheres as a novel hard template during the sol-gel process followed by calcination to remove the polymer. The particle size and pore size could be simply tuned by the feature of the polymer template and reaction conditions such as different functionalization of the parent polymer template, particle size of polymer template, and amount of TEOS during the sol-gel process. EDA (ethylenediamine), APTES (3-aminopropyl)triethoxysilane, and TMA (trimethylamine hydrochloride) functionalization of porous poly(GMA-co-EGDMA) microspheres were carried out to study their effect on the synthesized porous silica microspheres. The TMA-functionalized polymer microspheres led to a higher yield, smaller silica nanoparticles, and no self-nucleation of TEOS due to their positive surface charge. Furthermore, no addition of NaOH during TMA functionalization and the amount of TEOS during the sol-gel process played key roles in determining the pore size and particle size of porous silica microspheres. Then, through poly(aspartic acid) coating of the APTES-functionalized monodisperse porous silica microspheres, the modified monodisperse porous silica microspheres were explored as the stationary phase of HPLC for protein separation. The effects of particle size and pore size on the chromatographic behavior were discussed. When the protein mixture composed of transferrin, hemoglobin, ribonuclease A, cytochrome C, and lysozyme was used as the model analytes, the as-prepared silica microspheres exhibited an excellent separation performance with a high protein recovery and good reproducibility.