Mass spectrometry-based untargeted metabolomics: from experimental design to metabolite identification

Metabolomics aims at the comprehensive and quantitative analysis of wide array of metabolites in biological samples. These numerous analytes have very diverse physio-chemical properties and a wide abundance range, presenting significant analytical challenges. Metabolite profiling can be performed with either targeted or untargeted approach. In targeted metabolomics, a defined set of previously identified metabolites is analysed to achieve accurate quantitation. This is effective for testing defined hypotheses. In contrast, untargeted metabolomics aims to detect as many metabolites as possible. This unbiased system level assessment of metabolism is highly attractive because it can potentially reveal altered pathways that had not been previously anticipated. Untargeted metabolomics has become a popular tool in metabolite biomarker discovery and in the evaluation of disease-associated changes in metabolic pathways. Since the set of metabolites being profiled is not well-defined and may include “unknown” compounds that have not yet been characterized, it is important to carefully construct an appropriate experimental design. We aim for a protocol that maximizes the number of metabolites detected while still maintaining high quantitative reproducibility using a workflow that will yield high-quality data used for analysis. For those who are interested in combining metabolomics studies in their research, representative research projects conducted in the Metabolomics Center demonstrate key aspects in study design, analytical methodology and data analysis that should be considered when setting-up a successful metabolomics project. With proper design and execution, untargeted metabolomics can be an important tool generating unique data that cannot be obtained otherwise