Establishing 18 O-Labeled Inositol Phosphates for Quantitative Capillary Electrophoresis-Mass Spectrometry: Fragmentation Pathways and Comparison with 13 C-Labeled Analogs
Year: 2025
Type: article
Abstract: High Resolution Image Download MS PowerPoint Slide Capillary electrophoresis mass spectrometry (CE-MS) allows for the rapid and accurate quantitative analysis of inositol phosphates (InsPs) and inositol pyrophosphates (PP-InsPs). The recent discovery of new InsPs and PP-InsPs isomers in plants and mammals necessitates new heavy isotope references for quantitative analysis of complex cellular extracts. Here, we evaluate 18 O-labeled InsPs and PP-InsPs as alternatives to 13 C labeled internal standards for quantitation by CE-MS. In contrast to 13 C labels, the 18 O labels are introduced at the end of a synthetic campaign and not at the beginning, rendering 18 O much more accessible and affordable as a label. A series of 18 O-labeled InsPs and PP-InsPs with different numbers and positions of 18 O atoms were synthesized, enabling systematic investigation of MS2 fragmentation pathways. We propose two major dissociation pathways to elucidate the 18 O redistribution of the dominant product ion (the loss of H 3 PO 4 ). Based on these insights, we identified the loss of HPO 3 as a suitable transition for minimizing isotope redistribution in MS2 analysis. The ratios of this alternative product ion and dominant product ion were reproducible across replicates, concentration, and measurement days, supporting the use of this alternative product ion as a reliable product ion for quantitative analysis. Application to Saccharomyces cerevisiae, HCT116 cells, and Arabidopsis thaliana extracts confirmed accurate quantitation and precision comparable to 13 C-based methods.
Source: Analytical Chemistry
Authors: Guizhen Liu, Tobias Dürr-Mayer, Mengsi Lu, Henning J. Jessen
Institution: University of Freiburg
Language: English
Subfield: Spectroscopy
Field: Chemistry
Domain: Physical Sciences
Open Access status: hybrid
Estimated APC paid: $4,500
OpenAlex ID: W4415929682