A novel fibrous mesoporous silica nanoparticles (fSiO2) stationary phase grafted with polymer (Poly (2-(dimethylamino) ethyl methacrylate) (PDMAEMA) was developed for open tubular capillary electrochromatography (OT-CEC). The preparation procedure included synthesizing fSiO2 through biphase stratification approach, removing the surfactants, silanization and in situ graft polymerization with monomers via atom transfer radical polymerization (ATRP). Subsequently, PDMAEMA-modified mesoporous silica nanoparticles (P-fSiO2)/ethanol solution was immobilized onto the inner surface of the pretreated capillary and functionalized with octadecylsilane to fabricate the open-tubular column. Separation of polycyclic aromatic hydrocarbons (PAHs) and proteins were carried out to evaluate the performance of the column in CEC. The run-to-run, day-to-day and column-to-column reproducibility in terms retention time of naphthalene was 1.9%, 2.2%, and 3.7%, respectively. The effects of solvent concentration and pH on the separation were evaluated. The method was also used for the separation of real bio-sample, egg white proteins.