Abstract
In this study, graphene oxide‐octadecylsilane incorporated monolithic nano‐columns were developed for protein analysis by nano liquid chromatography (nano LC). The monolithic column with 100 μm id was first prepared by an in situ polymerization using ethylene dimethacrylate (EDMA), 3‐chloro‐2‐hydroxypropylmethacrylate (HPMA‐Cl), and methacryloyl graphene oxide nanoparticles (MGONPs). MGONPs were synthesized by the treatment of 3‐(trimethoxysilyl)propylmethacrylate (TMSPM) and GO. Tetrahydrofuran (THF) and dodecanol were used as the porogenic solvent. The resulting column was functionalized by dimethyloctadecylch lorosilane (DODCS) for the enhancement of hydrophobicity. The functionalization greatly improved the baseline separation of hydrophobic compounds such as polyaromatic hydrocarbons (PAHs). The optimized monolith with respect to total polymerization mixture was characterized by using Fourier‐transform infrared spectroscopy (FT‐IR), scanning electron microscopy (SEM) X‐ray diffraction (XRD) and chromatographic analyses. The blank monoliths without functionalization exhibited poor separation while a good separation performance of MGONPs functionalized monoliths was achieved. The monolith with 100 μm id was evaluated in protein separation in nano LC using RNase A, Cytochrome C, Lysozyme, Trypsin, and Ca isozyme II as the test proteins. It was shown that protein separation mechanism was based on large π‐system of GO and hydrophobicity of the monolithic structure. Theoretical plates number up to 57 600 plates were achieved. The nano‐column with 50 μm id was also prepared using the same polymerization mixture under the same chemical conditions. These nano‐columns were employed for protein separation by nano LC, and the dependence of both nano‐column performance on the internal diameter was also discussed.