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Mobile Phase Optimization for the Analysis of Synthetic Oligonucleotides by Ion-Pair Reversed Phase Liquid Chromatography

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by Brian Rivera and Dr. Bryan Tackett, Phenomenex, Inc., 411 Madrid Ave., Torrance, CA 90501 USA

Therapeutic oligonucleotides represent a recent breakthrough in the pharmaceutical industry. Improved reliability to synthetic routes coupled with new drug delivery technologies has given a heightened level of interest in these clinically interesting targets. However, characterization of oligos, specifically by ion-pair reversed phase liquid chromatography (IP-RPLC), can be quite challenging.

Most oligos are manufactured by solid phase synthesis, where nucleotides are added in a step-wise phase, leading to characterization of the commonly associated n-1 and n+1 impurities. However, even with ion-pair to facilitate the analytical separation, for full characterization of process and product related impurities for synthetic oligonucleotides, extensive method development and mobile phase optimization is required.

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