Dataset Information

NMR-Guided Mass Spectrometry for Absolute Quantitation of Human Blood Metabolites.

ABSTRACT: Broad-based, targeted metabolite profiling using mass spectrometry (MS) has become a major platform used in the field of metabolomics for a variety of applications. However, quantitative MS analysis is challenging owing to numerous factors including (1) the need for, ideally, isotope-labeled internal standards for each metabolite, (2) the fact that such standards may be unavailable or prohibitively costly, (3) the need to maintain the standards' concentrations close to those of the target metabolites, and (4) the alternative use of time-consuming calibration curves for each target metabolite. Here, we introduce a new method in which metabolites from a single serum specimen are quantified on the basis of a recently developed NMR method [ Nagana Gowda et al. Anal. Chem. 2015 , 87 , 706 ] and then used as references for absolute metabolite quantitation using MS. The MS concentrations of 30 metabolites thus derived for test serum samples exhibited excellent correlations with the NMR ones (R2 > 0.99) with a median CV of 3.2%. This NMR-guided-MS quantitation approach is simple and easy to implement and offers new avenues for the routine quantification of blood metabolites using MS. The demonstration that NMR and MS data can be compared and correlated when using identical sample preparations allows improved opportunities to exploit their combined strengths for biomarker discovery and unknown-metabolite identification. Intriguingly, however, metabolites including glutamine, pyroglutamic acid, glucose, and sarcosine correlated poorly with NMR data because of stability issues in their MS analyses or weak or overlapping signals. Such information is potentially important for improving biomarker discovery and biological interpretations. Further, the new quantitation method demonstrated here for human blood serum can in principle be extended to a variety of biological mixtures.

SUBMITTER: Nagana Gowda GA

PROVIDER: S-EPMC6260976 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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NMR-Guided Mass Spectrometry for Absolute Quantitation of Human Blood Metabolites.

Nagana Gowda G A GA Djukovic Danijel D Bettcher Lisa Fan LF Gu Haiwei H Raftery Daniel D

Analytical chemistry 20180109 3

Broad-based, targeted metabolite profiling using mass spectrometry (MS) has become a major platform used in the field of metabolomics for a variety of applications. However, quantitative MS analysis is challenging owing to numerous factors including (1) the need for, ideally, isotope-labeled internal standards for each metabolite, (2) the fact that such standards may be unavailable or prohibitively costly, (3) the need to maintain the standards' concentrations close to those of the target metabo ...[more]

PMID: 29293320

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Project description:A current challenge in metabolomics is the reliable quantitation of many metabolites. Limited resolution and sensitivity combined with the challenges associated with unknown metabolite identification have restricted both the number and the quantitative accuracy of blood metabolites. Focused on alleviating this bottleneck in NMR-based metabolomics, investigations of pooled human serum combining an array of 1D/2D NMR experiments at 800 MHz, database searches, and spiking with authentic compounds enabled the identification of 67 blood metabolites. Many of these (?1/3) are new compared with those reported previously as a part of the Human Serum Metabolome Database. In addition, considering both the high reproducibility and quantitative nature of NMR as well as the sensitivity of NMR chemical shifts to altered sample conditions, experimental protocols and comprehensive peak annotations are provided here as a guide for identification and quantitation of the new pool of blood metabolites for routine applications. Further, investigations focused on the evaluation of quantitation using organic solvents revealed a surprisingly poor performance for protein precipitation using acetonitrile. One-third of the detected metabolites were attenuated by 10-67% compared with methanol precipitation at the same solvent-to-serum ratio of 2:1 (v/v). Nearly 2/3 of the metabolites were further attenuated by up to 65% upon increasing the acetonitrile-to-serum ratio to 4:1 (v/v). These results, combined with the newly established identity for many unknown metabolites in the NMR spectrum, offer new avenues for human serum/plasma-based metabolomics. Further, the ability to quantitatively evaluate nearly 70 blood metabolites that represent numerous classes, including amino acids, organic acids, carbohydrates, and heterocyclic compounds, using a simple and highly reproducible analytical method such as NMR may potentially guide the evaluation of samples for analysis using mass spectrometry.

Dataset Information

NMR-Guided Mass Spectrometry for Absolute Quantitation of Human Blood Metabolites.

Publications

NMR-Guided Mass Spectrometry for Absolute Quantitation of Human Blood Metabolites.

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