A standardized protocol for comparable analysis of GSH/GSSG by UHPLC-ESI-MSMS for human plasma

J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Jan 1;1104:67-72. doi: 10.1016/j.jchromb.2018.11.007. Epub 2018 Nov 10.


Variability in the levels of GSH and GSSG in plasma is suggested to derive from inadequate pre-processing methods. The aim of this study was to develop a protocol for comparable and reliable measurements of GSH/GSSG. Venous blood from 8 healthy individuals were collected and divided into 7 different pre-processing procedures. For three of the samples an extraction mixture was added after 0 (baseline), 4 and 8 min and for three of the samples the extraction mixture was added at different times during defrost. A worst case scenario where a sample was left in a cool box during 6 h was also included. The samples were analyzed with UHPLC-ESI-MSMS. A large difference in the levels of GSH and GSSG were identified and it was clearly associated with the sample handling procedures. A sample left untreated for 4 min will have significantly reduced amount of GSH. Stability tests showed that the level of GSH was reduced after 3 months in -80 °C.

Keywords: GSH; GSSG; Glutathione; Plasma; Sample pre-processing; UHPLC-ESI-MS.

MeSH terms

  • Chromatography, High Pressure Liquid / methods*
  • Drug Stability
  • Glutathione / blood*
  • Glutathione / chemistry*
  • Glutathione Disulfide / blood*
  • Glutathione Disulfide / chemistry*
  • Humans
  • Linear Models
  • Reproducibility of Results
  • Spectrometry, Mass, Electrospray Ionization
  • Tandem Mass Spectrometry / methods


  • Glutathione
  • Glutathione Disulfide