The HPLC resolution of nucleic-acid bases and analog hypoxanthine on R,S-2-hydroxypropyl derivatized beta-cyclodextrin bonded chiral stationary phaseusing a water-based mobile phase

A HPLC approach using R,S-2-hydroxypropyl derivatized beta -cyclodextrin pa cked column as the stationary phase was developed to resolve five nucleic-a cid bases and analog hypoxanthine in the reversed-phase mode. These bases a re not only similar in structure but also very close in basicity. However, the resolution can be completed in less than ten minutes and is considered to be better carried out on the R,S-2-hydroxypropyl derivatized beta -cyclo dextrin phase than that obtained on the native beta -cyclodextrin phase und er the same chromatographic conditions. The mechanism involved in the resol ution is believed to be inclusion complexation between the analyte and the cavity of cyclodextrin in the reversed-phase mode. The retention time was f ound relevant to the size of the analyte. The number of groups on analyte t hat is available to form hydrogen bonding with hydroxyl groups on CDs also affects the retention scale. Factors of introducing organic acid and base o r organic modifier such as methanol to the water-based mobile phase or incr easing their percentages in the mobile phase decreases the retention time w ithout degrading the resolution significantly.