Development of High Throughput Global Metabolomics Approaches based upon Mass Spectrometry
EMSL Project ID
12499
Abstract
The term 'metabolome' refers to the entire complement of all small molecular weight metabolites and related species at a given time for a system of interest. Quantitative metabolomic measurements that can include a significant fraction of either, or both, the intracellular and extra-cellular metabolites are the key to understanding metabolism and regulation of cellular systems, and compliment gene expression and proteomic information. This project is directed towards developing approaches and methods for quickly detecting many metabolites in an extremely broad manner that is both amenable to quantitation and is also highly sensitive (e.g., able to detect low level species). In this project, we focus on developing methods based upon the use of capillary liquid chromatography (LC)-Fourier transform ion cyclotron resonance (FTICR) or high mass accuracy time-of flight (TOF) mass spectrometry (MS) to increase the comprehensiveness, quantitation and throughput of metabolomics analyses, and leverage the significant separations and mass spectrometry capabilities initially developed at PNNL for proteomics. Due to the low molecular weights of many metabolites, the accurate mass measurements provided by FTICR or TOF MS can often provide specific elemental compositions. Specific metabolite databases, such as the Ligand component of the Kyoto Encyclopedia of Genes and Genomics (KEGG), can then be searched using either empirical formula or molecular weight information in order to identify candidate metabolites.
Similar to our current proteomics approach, we will use various MS/MS methods and the combination of accurate mass and LC elution time information to initially identify metabolome components and define “tags” so that subsequent measurements can be made without the need for routine MS/MS analyses. Using mass and time (MT) tags, broad metabolome measurements can be made with much higher throughput. Under the three specific aims of the project we will: (1) develop the sample processing and separation conditions suitable for metabolomic studies of microbes, (2) use MS/MS methods for the identification of metabolites and the development of an MT tag database, and (3) evaluate the quantitative application of the approach using microbial syntrophic co-culture samples from two companion BSI projects.
Project Details
Project type
Exploratory Research
Start Date
2005-02-25
End Date
2005-10-24
Status
Closed
Released Data Link
Team
Principal Investigator