Impurity Profiling of GalNAc-conjugated Oligonucleotides by LC-MS-UV

This application note details the use of ion-pairing reversed-phase liquid chromatography (IP-RPLC) coupled with MS and UV detection for characterizing a GalNAc-conjugated siRNA duplex. Initial attempts with a Biozen Oligo C18 100 A column under denaturing conditions failed to fully resolve the strands, but switching to a Biozen WidePore C4 400 A column, which features a larger pore size and lower carbon load, achieved successful separation and impurity profiling of the duplex at elevated temperatures. This method effectively identifies and quantifies impurities, making it ideal for metabolite profiling and characterization of siRNA therapeutics.