Ion Mobility Mass Spectrometry (IMMS) is an emerging analytical technique with the potential capability of detecting, quantifying and identifying thousands of compounds in a complex mixture in fractions of a second. While IMMS has been applied primarily to the proteome, this grant application proposes to develop and evaluate its potential to measure and monitor the metabolome. Using both electro spray ionization and matrix assisted laser desorption ionization to create peptide and protein ions in the gas phase, IMS separates isomeric peptides and protein conformations before mass spectral analysis. This separation mechanism is based on ion-molecule collisions as an ion migrates through a neutral buffer gas in an electric field. The time an ion takes to transverse the buffer gas region is related to its size and shape. In IMS instruments currently used for peptide and protein separations, ion migration through the buffer gas occurs at a reduced pressures (typically 10 torr) with a typical separation efficiency of about 2000 theoretical plates. For application to metabolomics, we propose to use ion mobility separations at atmospheric pressures or greater, increasing the separation efficiency to more than 300,000 theoretical plates and improving significantly the two-dimensional resolving power of IMMS. In addition to separation efficiency, separation range and selectivity will be enhanced. Using IMS parameters of temperature, pressure, buffer gas, and voltage, the separation range will be extended to achieve maximal peak capacity and the separation selectivity will be optimized for maximal resolution of a metabolome. IMMS will also be interfaced to both gas chromatography and liquid chromatography to provide maximal metabolite quantification and provide three dimensional separations. With IMMS, both pattern recognition algorithms to identify modification in an entire metabolome as well as identification and quantification of individual metabolites is possible.