Comprehensive analysis of the metabolome can contribute to mechanism of action studies for small molecules discovered in phenotypic screens. Examples are presented in this overview of the rapidly developing field of "metabolic profiling." These examples include the use of NMR in gene function analysis, GC-based studies on the identification of metabolic pathways affected by PPAR-gamma agonists, applications of Fourier-transform MS and the use of stable isotope-based metabolic profiling (SIDMAP) to investigate metabolic adaptive changes induced by effective anticancer agents.