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• Three structurally similar stationary phases were prepared and evaluated. • Differences in retention behavior among the three columns were studied. • The retention mechanisms were explored by linear solvation energy relationship. • The retention mechanisms were explored by Van't Hoff plots. • The column efficiency for uridine on Sil-Phe was up to 7.3 × 104 plates/m. Three structurally similar silane reagents with different terminal groups were prepared and bonded to silica to obtain three structurally similar stationary phases (Sil-Ph-COOH, Sil-Phe and Sil-Ph-NH 2). The prepared stationary phases were characterized through elemental analysis (EA) and Fourier Transform Infrared Spectroscopy (FT-IR). These three stationary phases provided acceptable retention repeatability (relative standard deviations between 0.08% and 0.13%) and high column efficiency (7.3 × 104 plates/m for uridine on Sil-Phe). The retention behavior of the three columns was investigated under different chromatographic conditions including different mobile phase ratio, salt concentration, pH etc. The retention mechanisms were explored by linear solvation energy relationships and Van't Hoff plots. Applications in separation under reversed phase liquid chromatography (RPLC), hydrophilic interaction liquid chromatography (HILIC) and ion exchange chromatography (IEC) mode were investigated. The results showed that the retention capacity of the stationary phases with different terminal groups to the analytes is very different, especially for carboxylic acids, because the surface charges of amino groups and carboxyl groups under weakly acidic conditions produce different electrostatic effects with dissociated carboxylic acids. Finally, the Sil-Phe column was employed to detect ibuprofen extracted from pharmaceutical ibuprofen capsules and vitamins extracted from vitamin tablets. [ABSTRACT FROM AUTHOR]

Peptide Nucleic Acid (PNA) is a nucleic acid analogue, whose neutrally charged and hydrophobic backbone makes it more stable in vivo, so it might act as a potentially better protein probe as compared to aptamer. Currently the investigation of PNA and protein interaction is scarce. In this research, multiple modes of capillary electrophoresis were established and applied for PNA characterization and its interaction with ssDNA and protein. A 15-mer PNA having the same nucleobase sequence as 15-mer anti-thrombin DNA aptamer was chosen as PNA model for this study, its pI (7.71) was estimated by capillary isoelectric focusing (cIEF). Due to its neutral charge and strong hydrophobicity, three micellar electrokinetic chromatography (MEKC) modes containing (a) SDS, (b) Triton X-100 and (c) CTAB were compared for PNA related analysis. CTAB was not applicable for PNA analysis, while in 4 mM SDS or 2 mM Triton X-100, PNA and PNA-ssDNA complex can be identified directly. The significant peak of PNA-ssDNA complex helped in validating the two MEKC modes for PNA and target interaction study. Furthermore, the effect of SDS and Triton X-100 concentrations in the two MEKC modes on the protein target thrombin analysis was investigated by capillary zone electrophoresis (CZE). 4 mM SDS caused thrombin denaturation. So in 2 mM Triton X-100, interactions of PNA with thrombin, PNA with RNase A and a non-aptameric PNA ( n -PNA) with thrombin were compared. PNA with thrombin exhibited strongest binding. In summary, cIEF mode for PNA pI determination, MECK mode for direct PNA, PNA-ssDNA and PNA-protein complex identification and CZE mode for the effect of surfactant in MEKC modes on protein target thrombin analysis were applied. Above results showed that multiple modes of CE provide rapid and very low-sample cost methods for PNA related studies. [ABSTRACT FROM AUTHOR]