Capillary Electrochromatography with Novel Stationary Phases. 3. Retention Behavior of Small and Large Nucleic Acids on Octadecyl-Sulfonated-Silica

Abstract

In this investigation, the potentials of porous and nonporous octadecyl-sulfonated-silica (ODSS) microparticles were demonstrated in the capillary electrochromatography (CEC) of small (e.g., nucleotides and dinucleotides) and large (e.g., transfer ribonucleic acids (t-RNAs)) nucleic acids. The ODSS stationary phase comprised two layers: a hydrophilic sulfonated (permanently charged) sublayer and an octadecyl top layer. While the sublayer is to provide a relatively strong electroosmotic flow, the octadecyl top layer is to ensure the retentivity and selectivity required for the separation of the analytes. Mono-, di-, and triphosphate nucleotides were best separated when a small amount of tetrabutylammonium bromide was added to the mobile phase. The tetrabutylammonium bromide functioned as an ion-pairing agent and consequently allowed the rapid separation of 12 different nucleotides. It is believed that the dynamic complex exchange model explains the basis of retention in ion pair reversed-phase CEC. Eight different dinucleotides, which have similar mass-to-charge ratios, separated very well by CEC. These solutes exhibited similar migration times (i.e., little or no separation) in capillary zone electrophoresis (CZE). Similarly, t-RNAs that did not separate by CZE were well resolved in CEC with nonporous ODSS. This demonstrates that CEC is very suitable for the separation of solutes that have similar mass-to-charge ratios but differ in their hydrophobicity.