Retention Behavior of Oligonucleotides on a Glycidyl Methacrylate-Based DEAE-Modified Sorbent

The retention behavior of unmodified phosphodiester mixed-sequence oligonucleotides has been investigated under different conditions on a glycidyl methacrylate-based, DEAE-modified anion-exchange Biospher GMB 1000Q column. It was verified that separation is based not only on an anion-exchange mechanism but also on secondary interactions of the bases with the stationary phase. Notably different retention was observed for oligonucleotides differing only in the position of one base in the sequence. The nature of these secondary interactions and the effect of sequence on retention is discussed. The effect on retention of the type and concentration of organic mobile phase modifier, mobile phase pH, sample molecular weight, and the bases in the sample was investigated. A linear relationship was found between retention and the molecular weight of mixed-sequence oligonucleotides with lengths from 15 to 40. Resolution of N − 1 synthetic by-products from the full-length product was also evaluated.