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An analysis of quenching and extraction techniques for performing metabolomics in Acidithiobacillus ferrooxidans.
(2016-10-07) Doran, Marney Lynn
I present here a protocol for accurate liquid chromatography – mass spectrometry based determination of the metabolome of Acidithiobacillus ferrooxidans. At. ferrooxidans metabolic processes contribute to the environmentally destructive process of acid mine drainage (AMD), and metabolomic profiling can offer insight into how At. ferrooxidans contributes to these processes. Metabolism must be stopped (quenched) instantaneously while limiting the amount of cell damage. I tested a wide variety of quenching solutions and found that ammonium formate was the most effective quenching solution. An accurate metabolomics analysis also requires extraction of as large a number of metabolites as possible, with as little chemical bias (in extraction or degradation) as possible. I tested a wide variety of extraction solutions and found that isopropanol:methanol:water was the best extraction solution. Standardizing a protocol to quench and extract metabolites in At. ferrooxidans will contribute to understanding how At. ferrooxidans contributes to the AMD phenomenon.