Česká společnost pro hmotnostní spektrometrii

(13. ročník České konference hmotnostní spektrometrie a 11. Neformální proteomické setkání - WeS-01)
Quantification of Oxoacids of the Human Metabolome using Liquid Chromatography-Mass Spectrometry

Philipp Hebesberger ¹, Stanislav Opekar ^2 *, Martin Moos ², Petr Šimek ²

1. University of South Bohemia
2. Czech Academy of Sciences

Abstrakt

Detection and quantification of metabolites serves to understand how various organisms function, as it can unveil new metabolic pathways and provide insight into their regulation. Liquid chromatography with mass detection is one of the leading analytical techniques in metabolomics. However, LC-MS detection is limited when analytes have poor stability or poor ionization efficiency. To overcome these hurdles, analytes can be derivatized to enhance sensitivity by conferring a permanent charge and generating a stable derivative [1]. Quaternary Aminooxy (QAO) reagents provide an opportunity to assess a range of metabolites with oxo groups by generating stable, positively charged oxime derivatives at acidic pH [2].
To explore the potential of QAO reagents, a method for the quantification of oxoacids in the human metabolome using QAO derivatization and LC-MS detection was developed and tested on human urine, serum, and HEK293 cells. 2-(aminooxy)-N,N,N-trimethylethylammonium iodide (2QMA) was the derivatization agent of choice and, combined with HILIC separation on a BEH-Amide column, resulted in good separation. Internal standards were generated using isotope-coded derivatization, and cross-derivatization upon mixing the standard and internal standard solutions was prevented by quenching the reaction with acetone. The method was validated according to US FDA guidelines for LLOQ, ULOQ, linearity, and carryover using deionized water as a surrogate matrix.

* Korespondující autor: opekar@bclab.eu

Literatura

1. Zhao S., Li L.: Trends in Analytical Chemistry, 131: 115988 (2020)
2. Deng P. et al.: Analyst, 143(1): 311-322 (2018)

Poděkování:

This work was supported by the Czech Science Foundation (project GAČR, No. 23-06600S).