Seed dormancy and germination are regulated by several plant hormones, such as abscisic acid, gibberellin, auxin (indole-3-acetic acid), ethylene, and brassinosteroid. Endogenous concentrations of a hormone are determined by the balance between biosynthesis and deactivation, and contribute to the regulation of physiological responses. Therefore, profiling of all hormones and their metabolites (hormonome) is a powerful approach to elucidate the regulatory networks of hormone metabolism. The methods involved in the use of liquid chromatography-electrospray ionization-tandem mass spectrometry to develop a high-sensitive and high-throughput hormonome platform are described in this chapter.