Dataset Information

Globally Optimized Targeted Mass Spectrometry: Reliable Metabolomics Analysis with Broad Coverage.

ABSTRACT: Targeted detection is one of the most important methods in mass spectrometry (MS)-based metabolomics; however, its major limitation is the reduced metabolome coverage that results from the limited set of targeted metabolites typically used in the analysis. In this study we describe a new approach, globally optimized targeted (GOT)-MS, that combines many of the advantages of targeted detection and global profiling in metabolomics analysis, including the capability to detect unknowns, broad metabolite coverage, and excellent quantitation. The key step in GOT-MS is a global search of precursor and product ions using a single liquid chromatography-triple quadrupole (LC-QQQ) mass spectrometer. Here, focused on measuring serum metabolites, we obtained 595 precursor ions and 1?890 multiple reaction monitoring (MRM) transitions, under positive and negative ionization modes in the mass range of 60-600 Da. For many of the MRMs/metabolites under investigation, the analytical performance of GOT-MS is better than or at least comparable to that obtained by global profiling using a quadrupole-time-of-flight (Q-TOF) instrument of similar vintage. Using a study of serum metabolites in colorectal cancer (CRC) as a representative example, GOT-MS significantly outperformed a large targeted MS assay containing ?160 biologically important metabolites and provided a complementary approach to traditional global profiling using Q-TOF-MS. GOT-MS thus expands and optimizes the detection capabilities for QQQ-MS through a novel approach and should have the potential to significantly advance both basic and clinical metabolic research.

SUBMITTER: Gu H

PROVIDER: S-EPMC5437843 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Globally Optimized Targeted Mass Spectrometry: Reliable Metabolomics Analysis with Broad Coverage.

Gu Haiwei H Zhang Ping P Zhu Jiangjiang J Raftery Daniel D

Analytical chemistry 20151202 24

Targeted detection is one of the most important methods in mass spectrometry (MS)-based metabolomics; however, its major limitation is the reduced metabolome coverage that results from the limited set of targeted metabolites typically used in the analysis. In this study we describe a new approach, globally optimized targeted (GOT)-MS, that combines many of the advantages of targeted detection and global profiling in metabolomics analysis, including the capability to detect unknowns, broad metabo ...[more]

PMID: 26579731

Similar Datasets

Project description:Metabolic flux analysis (MFA) is highly relevant to understanding metabolic mechanisms of various biological processes. While the pace of methodology development in MFA has been rapid, a major challenge the field continues to witness is limited metabolite coverage, often restricted to a small to moderate number of well-known compounds. In addition, isotopic peaks from an enriched metabolite tend to have low abundances, which makes liquid chromatography tandem mass spectrometry (LC-MS/MS) highly useful in MFA due to its high sensitivity and specificity. Previously we have built large-scale LC-MS/MS approaches that can be routinely used for measurement of up to ∼1,900 metabolite/feature levels [Gu et al. Anal. Chem. 2015, 87, 12355-12362. Shi et al. Anal. Chem. 2019, 91, 13737-13745.]. In this study, we aim to expand our previous studies focused on metabolite level measurements to flux analysis and establish a novel comprehensive isotopic targeted mass spectrometry (CIT-MS) method for reliable MFA analysis with broad coverage. As a proof-of-principle, we have applied CIT-MS to compare the steady-state enrichment of metabolites between Myc(oncogene)-On and Myc-Off Tet21N human neuroblastoma cells cultured with U-13C6-glucose medium. CIT-MS is operationalized using multiple reaction monitoring (MRM) mode and is able to perform MFA of 310 identified metabolites (142 reliably detected, 46 kinetically profiled) selected from >35 metabolic pathways of strong biological significance. Further, we developed a novel concept of relative flux, which eliminates the requirement of absolute quantitation in traditional MFA and thus enables comparative MFA under the pseudosteady state. As a result, CIT-MS was shown to possess the advantages of broad coverage, easy implementation, fast throughput, and more importantly, high fidelity and accuracy in MFA. In principle, CIT-MS can be easily adapted to track the flux of other labeled tracers (such as 15N-tracers) in any metabolite detectable by LC-MS/MS and in various biological models (such as mice). Therefore, CIT-MS has great potential to bring new insights to both basic and clinical metabolism research.

Project description:The lack of effective screening strategies for high-grade serous carcinoma (HGSC), a subtype of ovarian cancer (OC) responsible for 70-80% of OC related deaths, emphasizes the need for new diagnostic markers and a better understanding of disease pathogenesis. Capillary electrophoresis (CE) coupled with high-resolution mass spectrometry (HRMS) offers high selectivity and sensitivity for ionic compounds, thereby enhancing biomarker discovery. Recent advances in CE-MS include small, chip-based CE systems coupled with nanoelectrospray ionization (nanoESI) to provide rapid, high-resolution analysis of biological specimens. Here, we describe the development of a targeted microchip (µ) CE-HRMS method, with an acquisition time of only 3 min and sample injection volume of 4nL, to analyze 40 target metabolites in serum samples from a triple-mutant (TKO) mouse model of HGSC. Extracted ion electropherograms showed sharp, baseline resolved peak shapes, even for structural isomers such as leucine and isoleucine. All calibration curves of the analytes maintained good linearity with an average R2 of 0.994, while detection limits were in the nM range. Thirty metabolites were detected in mouse serum with recoveries ranging from 78 to 120%, indicating minimal ionization suppression and good accuracy. We applied the µCE-HRMS method to biweekly-collected serum samples from TKO and TKO control mice. A time-resolved analysis revealed characteristic temporal trends for amino acids, nucleosides, and amino acid derivatives. These metabolic alterations are indicative of altered nucleotide biosynthesis and amino acid metabolism in HGSC development and progression. A comparison of the µCE-HRMS dataset with non-targeted ultra-high performance liquid chromatography (UHPLC)-MS results showed identical temporal trends for the five metabolites detected with both platforms, indicating the µCE-HRMS method performed satisfactorily in terms of capturing metabolic reprogramming due to HGSC progression while reducing the total data collection time three-fold.

Dataset Information

Globally Optimized Targeted Mass Spectrometry: Reliable Metabolomics Analysis with Broad Coverage.

Publications

Globally Optimized Targeted Mass Spectrometry: Reliable Metabolomics Analysis with Broad Coverage.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets