The field of omics involves comprehensive investigations based on genomics, transcriptomics, proteomics, and metabolomics, and omics studies have developed rapidly. Metabolomics, metabolome analysis, involves technology to analyze the concentrations of low-molecular-weight metabolites comprehensively, and has recently rapidly developed along with improvements in analytical technology. Therefore, metabolome analysis is just beginning to be applied to not only food science and environmental research fields but also medical research. In the medical research field, especially, metabolome analysis plays an important role in novel disease biomarker discovery. The metabolome represents the endpoint of the omics cascade and, therefore, is considered to be closer to the phenotype. In addition, there is also a possibility that the metabolome is affected by exogenous factors such as environmental and food factors, as well as endogenous factors such as DNA/mRNA/protein. Therefore, metabolome analysis can be a useful approach for discovering effective biomarkers. Here, we explain the characteristics of metabolome analysis, and also outline metabolome analysis using a liquid chromatograph mass spectrometer (LC-MS), gas chromatograph mass spectrometer (GC-MS), capillary electrophoresis mass spectrometer (CE-MS), and matrix-assisted laser desorption ionization mass spectrometer (MALDI-MS). Then, we describe the findings of studies that used metabolome analysis in an attempt to discover biomarker candidates for pancreatic cancer, and discuss metabolome analysis-based disease diagnosis.