Structural mass spectrometry (MS) is a chemical structure-revealing analytical technique that measures mass/charge ratio of ions. Biochemical analyses in the context of metabolomics-which seeks chemical descriptions of cellular, organismal, and even community biology-pose challenges distinct from past applications of MS. The scientific need for both far greater depth and breadth of metabolic analytes is manifested analytically in two general approaches: the "Biomarker" approach and the "Pathway" approach. In this chapter, the basic principles of mass spectrometry relevant to these two approaches to metabolomics are reviewed. The emphasis is on practical aspects of how experimental design interacts with instrument designs and capabilities. Both simulated theoretical and real-life analyses are used to illustrate the key concepts and their ramifications.