The main objective of a metabolomics experiment is to determine the composition of the set of small molecules in one or more biological samples, both with respect to quantity and chemical characteristics. The focus is on concepts that are of particular importance for subsequent analysis and interpretation of the data. Gas chromatography is a technique that can be used for the separation and analysis of gaseous, liquid solutions and volatile solid materials that are thermally stable. Several stationary phases are used for the separation of small molecules in LC-MS such as reversed phase, ion exchange and hydrophilic interaction liquid chromatography. The basic principle of mass spectrometry is to determine the exact mass of a number of molecules by converting them to a beam of charged gas phase ions and measure their mass to charge ratio. Mass spectrometry is an almost universal detection method offering very high sensitivity and specificity.