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Graphene oxide (GO) nanosheets were immobilized onto the capillary wall using 3-aminopropyldiethoxymethyl silane as coupling agent. Graphene coated column (G@column) was fabricated by hydrazine reduction of GO modified column. Scanning electron microscopy (SEM) images provided visible evidence of the GO grafted on the capillary wall. Energy dispersive X-ray spectrometry (EDS) indicated the high coverage of the GO on the capillary wall. The G@column exhibited a pH-dependent electroosmotic flow (EOF) from anode to cathode in the pH range of 3-9 while the graphene oxide coated column (GO@column) showed a constant EOF. Both GO@column and G@column were evaluated for open-tubular capillary electrochromatography (OT-CEC). The GO@column was also evaluated for open-tubular capillary liquid chromatography (OT-CLC). Good separation of the tested neutral analytes on the GO@column was achieved on the basis of a typical reversed-phase behavior. On the contrary, G@column showed poor separation performance because of the strong π-π stacking and hydrophobic interactions between graphene and polyaromatic hydrocarbons. The high coverage of GO improved the column phase ratio which makes the GO@column promising for OT-CLC separation. Five of the major known proteins including three glycoisoforms of ovalbumin in chicken egg white were identified in a single run on the GO@column with phosphate buffer (5 mM, pH 7.0) and an applied voltage of 20 kV. The run-to-run, day-to-day, and column-to-column reproducibilities are evaluated by calculating the relative standard deviations (RSDs) of the EOF in OT-CEC and retention time of naphthalene in OT-CLC, respectively. These RSD values were found to be less than 3%.

The paper presents a novel on-line transient moving chemical reaction boundary method (tMCRBM) for simply but efficiently stacking ionizable analytes in high-salt matrix in capillary zone electrophoresis (CZE). The powerful function and stability of the tMCRBM are elucidated with the ionizable test analytes of L-phenylalanine (Phe) and L-tryptophan (Trp) in the matrix with 85.6-165.6 mM sodium ion and further compared with the normal CZE of Phe and Trp samples dissolved in running buffer. The results verify that (1) the on-line tMCRBM mode can evidently increase separation efficiency, peak height, and resolution, (2) with the mode, the analytes in a 28-cm high-salt matrix plug can be stacked successfully and further separated well, (3) the values of relative standard deviation of peak height, peak area, and migrating time range from 3.9% to 6.1%; the results indicate the high stability of the technique of tMCRBM-CZE. The techniques implies obvious potential significance for those ionizable analytes, e.g., protein, peptide, and weak alkaline or acidic compound, in such matrixes as serum, urine, seawater, and wastewater, with high salt, which has a deleterious effect on isotachophoresis (ITP) and especially on electrostacking and field-amplified sample injection (FASI). The mechanism of stacking of zwitterionic analytes in a high-salt matrix by the tMCRBM relies on non-steady-state isoelectric focusing (IEF) but not on transient ITP, electrostacking, and FASI.

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