Fast and High-Resolution Reversed-Phase Separation of Synthetic Oligonucleotides

Synthetic DNAand RNA-based oligonucleotides are among many successful biotherapeutic drugs to treat many different kinds of illnesses. They are synthesized in a multistep process. Although coupling efficiencies are high, the overall yield of oligos decreases as the number of cycles increases, with failure in coupling with single (N-1) and double (N-2) deletions as the major impurities. To ensure drug potency, and to reduce the potential for drug interactions, a high-purity product is required, therefore, analyzing the purity of the products is important. There are many methods used for analyzing oligonucleotides. One of the most common involves anion-exchange chromatography. This can provide high resolution, but the separation often takes a long time. Moreover, due to its solvent systems, eluting oligonucleotides with high salt concentrations make this method incompatible with mass-spectrometry. This makes the identification of oligonucleotides and their impurities very cumbersome.