AbstractThe characterization of industrial yeast strains by examining their metabolic footprints (exometabolomes) was investigated and compared to genome‐based discriminatory methods. A group of nine industrial brewing yeasts was studied by comparing their metabolic footprints, genetic fingerprints and comparative genomic hybridization profiles. Metabolic footprinting was carried out by both direct injection mass spectrometry (DIMS) and gas chromatography time‐of‐flight mass spectrometry (GC–TOF–MS), with data analysed by principal components analysis (PCA) and canonical variates analysis (CVA). The genomic profiles of the nine yeasts were compared by PCR–restriction fragment length polymorphism (PCR–RFLP) analysis, genetic fingerprinting using amplified fragment length polymorphism (AFLP) analysis and microarray comparative genome hybridizations (CGH). Metabolomic and genomic analysis comparison of the nine brewing yeasts identified metabolomics as a powerful tool in separating genotypically and phenotypically similar strains. For some strains discrimination not achieved genomically was observed metabolomically. Copyright © 2007 John Wiley & Sons, Ltd.