Ion Chromatography Coupled with Mass Spectrometry for Metabolomics

The fields of metabolomics and metabonomics attempt to phenotype and quantify the vast array of metabolites present in biological samples. Reversed-phase liquid chromatography (LC) coupled with mass spectrometry (MS) is a valuable tool in the separation and identification of these compounds. Reversed-phase HPLC techniques cover a wide range of compounds. However, ionic and polar compounds such as organic acids, carbohydrates, nucleotides, and amino acids are difficult to separate or even retain on traditional reversed-phase columns. Ion-exchange chromatography offers a far better separation process for these compounds. The problem with the separation mode in this application relates to the salt eluents employed in ion exchange being incompatible with mass spectrometers. Here, the author describes an ion-exchange system that provides good separation of these polar compounds with on-line desalting to allow MS detection. This technique will allow studies of key metabolites that do not separate well on traditional reversed-phase columns. Some ionic metabolites may be positional isomers adjacent to each other in a synthetic pathway and as such, are isobaric. In such cases, MS is not sufficient to differentiate the two compounds. There is clearly a need for adequate separation techniques for isomeric metabolites.