Open tubular capillary electrochromatography of isomeric dipeptides using a G-quartet forming DNA aptamer stationary phase

DNA oligonucleotides that form intramolecular G-quartet structures were investigated as stationary phase reagents for separation of mixtures of isomeric dipeptides in open-tubular capillary electrochromatography (OTCEC). The oligonucleotides included a thrombin-binding aptamer (tb-ligand) that forms a biplanar G-quartet structure and an oligonucleotide that forms a 4-plane G-quartet structure. Fluorescence, circular dichroism and UV-visible absorbance spectroscopies were used in batch solution studies to detect interactions between the dipeptides and the biplanar G-quartet structure. Successful separations were achieved by OTCEC for a series of isomeric dipeptides. Results for OTCEC separations were compared with results obtained for capillary zone electrophoresis (CZE) separations on a bare, activated fused silica capillary. OTCEC with the aptamer stationary phase showed significant improvements in the resolution and reproducibility of CZE separations. In many studies, partial destabilization of the G-quartet structures by increased temperatures improved the separations and in some cases, different buffer systems were used in the OTCEC for enhanced separations.; Concentrated solutions of Guanosine (Guo) and Guanosine 5' -Monophosphate (GMP) were developed into gels. Gel formation was facilitated by the addition of certain alkaline metal ions and confirmed by visual inspection for increased viscosity and by CD spectroscopic studies. When the gel is formed individual nucleotides and nucleosides orient themselves into guanine-based coplanar tetrads or G-quartets. Like the G-quartet in tb-ligand, the guanine-based gels produce a circular dichroism spectrum that was used to monitor extent and stability of gel formation.; These gels were then developed into injectable monolithic stationary phases for CEC separations of a range of biomolecules based on their interactions with the gel matrix as well as the G-quartet structures. Separations were achieved for many of the isomeric dipeptides used in the first part of this work as well as for mixtures of homodipeptides, proteins and chiral drugs.