The metabolome of an organism at a given time is a snapshot of the phenotypical expression of biomolecules in response to environmental and biological events. Recent metabolomics studies identified key pathways and biomarkers for a wide array of human diseases. Despite these promising studies we still lack a thorough understanding of the metabolome. Enhancing and developing novel techniques enables us to expand metabolome coverage. Imaging mass spectrometry allows us to gain the understanding of dynamic changes occurring spatially within specific cellular populations within a tissue which is lost in traditional platforms. Electrophoretic separations coupled to mass spectrometry provide us with the ability to identify biologically relevant expression changes with higher mass accuracy and confidence than imaging alone. In this work, I present the development of CZE-ESI-MS and MALDI-TOFMS methods to investigate the spatial and temporal alterations in metabolism in complex biological samples: breast cancer models and during key stages during early embryonic development.