Profiling thiol metabolites and quantification of cellular glutathione using FT-ICR-MS spectrometry

Sadakatali S. Gori, Pawel Lorkiewicz, Daniel S. Ehringer, Alex C. Belshoff, Richard M. Higashi, Teresa W M Fan, Michael H. Nantz

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

We describe preparation and use of the quaternary ammonium-based α-iodoacetamide QDE and its isotopologue*QDE as reagents for chemoselective derivatization of cellular thiols. Direct addition of the reagents to live cells followed by adduct extraction into n-butanol and analysis by FT-ICR-MS provided a registry of matched isotope peaks from which molecular formulae of thiol metabolites were derived. Acidification to pH 4 during cell lysis and adduct formation further improves the chemoselectivity for thiol derivatization. Examination of A549 human lung adenocarcinoma cells using this approach revealed cysteine, cysteinylglycine, glutathione, and homocysteine as principal thiol metabolites as well as the sulfinic acid hypotaurine. The method is also readily applied to quantify the thiol metabolites, as demonstrated here by the quantification of both glutathione and glutathione disulfide in A549 cells at concentrations of 34.4±11.5 and 10.1±4.0 nmol/mg protein, respectively.

Original languageEnglish (US)
Pages (from-to)4371-4379
Number of pages9
JournalAnalytical and Bioanalytical Chemistry
Volume406
Issue number18
DOIs
StatePublished - 2014
Externally publishedYes

Keywords

  • Chemoselective
  • Cysteine
  • Hypotaurine
  • Iodoacetamide
  • Metabolomics
  • Oxidative stress

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry

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    Gori, S. S., Lorkiewicz, P., Ehringer, D. S., Belshoff, A. C., Higashi, R. M., Fan, T. W. M., & Nantz, M. H. (2014). Profiling thiol metabolites and quantification of cellular glutathione using FT-ICR-MS spectrometry. Analytical and Bioanalytical Chemistry, 406(18), 4371-4379. https://doi.org/10.1007/s00216-014-7810-z