High-throughput and multiplexed LC/MS/MRM method for targeted metabolomics.
Target-based metabolomics, focused on a subset of metabolites representative of key pathways, is a valuable tool for assessing metabolic changes resulting from genetic mutation, altered gene expression, and protein dysfunction in a given disease state or as a consequence of an environmental perturbation, such as a pharmaceutical. However, simultaneously analyzing hundreds of endogenous metabolites presents a challenge because of their diverse chemical structures and properties. In this study, we report a high-throughput, sensitive, and reproducible method for target-based metabolomics studies. It combines different separation conditions, optimal ionization polarities, and the most sensitive triple-quadrupole MS-based data acquisition mode (MRM). In 10 min, 205 endogenous metabolites, divided into three subgroups (amino acids, sugar and nucleic acids, and organic acids), are sequentially analyzed on a LC/MS/MRM system. Low picogram sensitivity is achieved for more than half of the metabolites. A 3-4 order of linearity and assay coefficient of variation less than 15% are observed for approximately 80% of the metabolites. In summary, we have established a multiplex LC/MS/MRM method for quantitatively profiling hundreds of known metabolites from complex biological samples. The methodology is generally applicable and easily expandable to include more endogenous or drug metabolites.
Metabolomics/Proteomics, Applied Quantitative Genotherapeutics, Pfizer Inc. 620 Memorial Drive, Cambridge, Massachusetts 02139, USA. ru.wei@pfizer.com
Ru
R
Wei
Guodong
G
Li
Albert B
AB
Seymour
D000596:Amino Acids / Q000097:blood / Q000737:chemistry; D000818:Animals; D002241:Carbohydrates / Q000097:blood / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D057166:High-Throughput Screening Assays; D013058:Mass Spectrometry / Q000379:methods*; D055432:Metabolomics / Q000379:methods*; D009696:Nucleic Acids / Q000097:blood / Q000737:chemistry; D051381:Rats
D000596:Amino Acids; D002241:Carbohydrates; D009696:Nucleic Acids