Quantitation of Diastereomer Content in PS-Modified Synthetic Oligonucleotides using cIMS-MS

) containing PS linkages. To assess whether fragment diastereomer relative abundances reflect those of the intact oligonucleotide, 5-mer analogs (shortmers) matching the intact termini were synthesized and analyzed via nESI-cIMS-MS, ion pairing reversed phase liquid chromatography with UV detection (IPRP-LC-UV), and IPRP-LC with IMS detection (IPRP-LC-IMS-MS). Comparison of the fragment ion IMS distributions with their shortmer counterparts showed very similar characteristics, indicating these fragments could be directly compared to the shortmers. By aligning LC retention time with cIMS-MS mobility distributions, the relative abundances of the diastereomers from the fragment ions to those of the shortmers were compared. Our results demonstrate that CID-IMS-MS enables relative quantitation of the PS diastereomers, establishing a framework for PS modified oligonucleotide characterization.