Development, validation and application ofan ion-pair reversed-phase liquidchromatography-tandem massspectrometry method for the quantificationof nusinersen: supplementary materials

Background: The fully phosphorothioate-modified oligonucleotide (OGN) nusinersen has low ionization
efficiency in the negative ion mode, resulting in a low mass spectrometry response. There have
been no relevant reports on developing a LC–MS method for the determination of nusinersen by
optimizing mobile phase composition. Materials & methods: Mobile phase additives comprised of 15 mM
triethylamine/25 mM 1,1,1,3,3,3-hexafluoro-2-propanol with a pH of 9.6. Nusinersen was extracted from
plasma using Oasis R ? HLB solid-phase extraction (Waters, MA, USA). Results & conclusion: By adjusting
the pH of the mobile phase to 9.6 by optimizing the type and concentration of ion-pair reagents, a high
mass spectrometry response was obtained. The developed method was applied to nusinersen and met the
requirements for the pharmacokinetic study of nusinersen in rabbits.