Abstract
Anion-exchange chromatography is an established method for the separation of nucleotides, nucleotide sugars, oligonucleotides, DNA, and RNA and their closely related synthetic impurities and metabolites. It is particularly useful for the separation of failure sequences (shortmers and longmers), impurities in phosphorothioated oligonucleotides, and the separation of diastereomers of phosphorothioated oligonucleotides. Purification of synthetic oligonucleotides by anion-exchange chromatography at the laboratory scale through process applications is described. Multidimensional separations using automated multiple heart cutting allow an improved separation for complex mixtures and sequential desalting of target fractions for mass spectrometric detection. Practical protocols are described for the analysis and purification of oligonucleotides as synthetic raw materials, therapeutic agents, and extracts of biological samples.
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