Quantitative profiling of endogenous polar metabolites from low volumes of blood samples

The etiology of many diseases is not yet completely understood. The involved biological processes might be resolved using a metabolomics approach, because metabolomics provides unique challenging opportunities to correlate the metabolome with a physiological or pathophysiological status and provides a vision on the relationships between genes, gene expression, environment and lifestyle.
Here, we present the development of two ultra-high performance liquid chromatography (UHPLC) methods coupled to mass spectrometry (MS) for the separation and quantitation of polar metabolites in blood samples. A reversed-phase UHPLC-MS/MS method has been developed to quantify anionic energetic metabolites, whereas hydrophilic interaction chromatography (HILIC)-MS/MS has been used to determine amino acids. Two sample pretreatment procedures have been developed for an optimal recovery of the respective metabolites from whole blood samples. One method involved the precipitation of proteins with acetonitrile and acids. In addition, volumetric absorptive microsampling has been used for the sample preparation. Small and accurate quantities of biological fluids (10 or 20 µL) can be collected with this sampling technique, which is of great interest for volume-limited samples or serial collection of samples.
The developed methods have been validated and will be applied to determine differences in metabolite concentrations between plasma samples from patients and controls. This can lead to a breakthrough in the understanding of diseases and can open new therapeutic perspectives.