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Metabolites extraction from Staphylococcus aureus

Instrument: Precellys® 24 Dual

Sample:  Staphylococcus aureus

Protocol:

  • Precellys kit: 03961-1-005 (0.1mm glass beads).
  • Competitor kit: 7mm bead (tungsten carbide).
  • Add Sample: 1 mL of cell suspension from exponential growth phase.
  • Fast vacuum dependent filtration to separate cells and medium.
  • Run the Precellys at 6800 rpm, 1-3 cycles x 30s vs competitor: 2 min at maximum speed.
  • Extraction of metabolites using water or 60% (w/v) ethanol.
  • Analysis of intracellular metabolites by GC/MS and LC/MS using standardized protocols.

Results:

The comparison of both mechanical disruption, using water as an extraction solution, showed that Precellys is more efficient. In addition the Precellys method after metabolite extraction by 60% (w/v) ethanol was optimized by the addition of several cycles. Two cycles at 6800 rpm for 30s were adopted for the cell disruption part of the protocol for S.aureus metabolome analysis, since a higher number of cycles could not obtain higher metabolite concentrations.

Percentage of relative metabolite concentration is shown for no cycle (defined as 100%) (X-axis) and compared to 1,2,3 cycles for each metabolite (Y-axis). Significant difference by two-tailed t test, P<0.1. Cells were harvested in exponential growth phase and metabolites were extracted with 60% (w/v) ethanol