Towards an unbiased metabolic profiling of protozoan parasites: optimisation of a Leishmania sampling protocol for HILIC-orbitrap analysis

Anal Bioanal Chem. 2010 Nov;398(5):2059-69. doi: 10.1007/s00216-010-4139-0. Epub 2010 Sep 8.


Comparative metabolomics of Leishmania species requires the simultaneous identification and quantification of a large number of intracellular metabolites. Here, we describe the optimisation of a comprehensive metabolite extraction protocol for Leishmania parasites and the subsequent optimisation of the analytical approach, consisting of hydrophilic interaction liquid chromatography coupled to LTQ-orbitrap mass spectrometry. The final optimised protocol starts with a rapid quenching of parasite cells to 0 °C, followed by a triplicate washing step in phosphate-buffered saline. The intracellular metabolome of 4 × 10(7) parasites is then extracted in cold chloroform/methanol/water 20/60/20 (v/v/v) for 1 h at 4 °C, resulting in both cell disruption and comprehensive metabolite dissolution. Our developed metabolomics platform can detect approximately 20% of the predicted Leishmania metabolome in a single experiment in positive and negative ionisation mode.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chromatography, Liquid / methods
  • Hydrophobic and Hydrophilic Interactions
  • Leishmania / chemistry*
  • Leishmania / growth & development
  • Metabolome*