Separation of nucleosides using capillary electrochromatography

The analysis of nucleosides and nucleotides have in most cases been performed by HPLC using either reversed-phase HPLC with gradient elution or using reversed-phase ion-pair chromatography. In this paper we have explored the possibility of using capillary electrochromatography (CEC) in order to avoid the use of gradients or ion-pairing reagents. CEC is in many ways comparable to HPLC, but CEC is theoretically able to provide better separations due to the higher efficiency caused by the flowfront being more plug-like as also is the case in CE, which is to be compared to the more parabolic flow observed in HPLC. The separation of six nucleosides (adenosine, cytidine, guanosine, inosine, thymidine and uridine) was investigated with respect to concentration of buffers, pH, amount of acetonitrile, temperature and voltage in order to optimise the separation. Baseline separation was achieved for the six nucleosides in less than 13 min using a background electrolyte consisting of (5 mM acetic acid, 3 mM triethylamine, pH 5.0)-acetonitrile (92:8, v/v).