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Poster Presentations
Day 1, June 22（Sun.）
Room P (Maesato East, Foyer, Ocean Wing)
- 1P-PM-16
Development of Hexafluoro-2-propanol-Free LC/MS Method for Oligonucleotide Analysis
(1Univ. Osaka, 2Univ. Osaka Shimadzu Lab., 3Shimadzu Corp.)
oTakashi Miyazaki1,2,3, Natsuyo Asano3, Takao Yamaguchi1, Satoshi Obika1
The characterization of oligonucleotide therapeutics is generally performed by liquid chromatography mass spectrometry (LC/MS) using ion-pair reversed-phase (IP-RP) chromatography with 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) and triethylamine (TEA) due to its efficient electrospray ionization (ESI). However, HFIP has challenges, including environmental concerns as a per- and polyfluoroalkyl substances (PFAS), rapid mobile phase degradation, and high reagent costs. Traditional HFIP-free alternatives, such as acetic acid (AA) and TEA, generally result in lower MS sensitivity. This study aims to develop a HFIP-free analytical method for oligonucleotides and explores the potential of AA/TEA-based mobile phase additives to improve MS sensitivity. As a result, an AA/glycine/TEA-based mobile phase demonstrated improved MS sensitivity compared to an AA/TEA-based mobile phase, achieving sensitivity comparable to that of a HFIP/TEA-based mobile phase. Additionally, the AA/glycine/TEA-based mobile phase maintained MS sensitivity for a longer duration than the HFIP/TEA-based mobile phase. While glycine is typically avoided in LC/MS due to its non-volatile nature, employing it at sufficiently low concentrations did not cause contamination that could impair performance. In conclusion, glycine is a promising additive for enhancing ESI-MS sensitivity in HFIP-free IP-RP chromatography for oligonucleotide analysis.