A Yeast Metabolite Extraction Protocol Optimised for Time-Series Analyses

Kalesh Sasidharan, Tomoyoshi Soga, Masaru Tomita, Douglas B. Murray

Research output: Contribution to journalArticlepeer-review

21 Citations (Scopus)

Abstract

There is an increasing call for the absolute quantification of time-resolved metabolite data. However, a number of technical issues exist, such as metabolites being modified/degraded either chemically or enzymatically during the extraction process. Additionally, capillary electrophoresis mass spectrometry (CE-MS) is incompatible with high salt concentrations often used in extraction protocols. In microbial systems, metabolite yield is influenced by the extraction protocol used and the cell disruption rate. Here we present a method that rapidly quenches metabolism using dry-ice ethanol bath and methanol N-ethylmaleimide solution (thus stabilising thiols), disrupts cells efficiently using bead-beating and avoids artefacts created by live-cell pelleting. Rapid sample processing minimised metabolite leaching. Cell weight, number and size distribution was used to calculate metabolites to an attomol/cell level. We apply this method to samples obtained from the respiratory oscillation that occurs when yeast are grown continuously.

Original languageEnglish
Article numbere44283
JournalPloS one
Volume7
Issue number8
DOIs
Publication statusPublished - 2012 Aug 29

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Fingerprint Dive into the research topics of 'A Yeast Metabolite Extraction Protocol Optimised for Time-Series Analyses'. Together they form a unique fingerprint.

Cite this