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. 2013 Sep 17;85(18):8715-21.
doi: 10.1021/ac401712a. Epub 2013 Aug 26.

15N-cholamine--a smart isotope tag for combining NMR- and MS-based metabolite profiling

Fariba Tayyari  1 , G A Nagana Gowda, Haiwei Gu, Daniel Raftery
Affiliations

15N-cholamine--a smart isotope tag for combining NMR- and MS-based metabolite profiling

Fariba Tayyari et al. Anal Chem. .

Erratum in

Abstract

Recently, the enhanced resolution and sensitivity offered by chemoselective isotope tags have enabled new and enhanced methods for detecting hundreds of quantifiable metabolites in biofluids using nuclear magnetic resonance (NMR) spectroscopy or mass spectrometry. However, the inability to effectively detect the same metabolites using both complementary analytical techniques has hindered the correlation of data derived from the two powerful platforms and thereby the maximization of their combined strengths for applications such as biomarker discovery and the identification of unknown metabolites. With the goal of alleviating this bottleneck, we describe a smart isotope tag, (15)N-cholamine, which possesses two important properties: an NMR sensitive isotope and a permanent charge for MS sensitivity. Using this tag, we demonstrate the detection of carboxyl group containing metabolites in both human serum and urine. By combining the individual strengths of the (15)N label and permanent charge, the smart isotope tag facilitates effective detection of the carboxyl-containing metabolome by both analytical methods. This study demonstrates a unique approach to exploit the combined strength of MS and NMR in the field of metabolomics.

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Figures

Figure 1
Figure 1
Schematic figure illustrating the “smart isotope tag” approach used to detect the same metabolites using NMR and MS with high sensitivity. Tagging carboxyl-containing metabolites with 15N-cholamine enables their detection by both NMR and MS with high sensitivity.
Figure 2
Figure 2
Figure 2a. A portion of the 1H-15N HSQC spectrum of human serum tagged with 15N-cholamine. 1: aconitic acid; 2: adipic acid; 3: alanine; 7: aspartic acid; 8: betaine; 9: citric acid; 11: cystine; 12: formic acid; 15: glutamic acid; 17: glycine; 20; histidine; 21: 3-hydroxybutyric acid; 24: isocitric acid; 28: lactic acid; 29: leucine; 32: malic acid; 37: phenylalanine; 40: pyroglutamic acid; 45: threonine; 46: tryptophan; 47: tyrosine; 48: valine. Figure 2b. A portion of the 1H-15N HSQC spectrum of a mixture of standard compounds at various concentrations obtained after tagging with 15N-cholamine. The peak numbers correspond to the compounds shown in Table I.
Figure 2
Figure 2
Figure 2a. A portion of the 1H-15N HSQC spectrum of human serum tagged with 15N-cholamine. 1: aconitic acid; 2: adipic acid; 3: alanine; 7: aspartic acid; 8: betaine; 9: citric acid; 11: cystine; 12: formic acid; 15: glutamic acid; 17: glycine; 20; histidine; 21: 3-hydroxybutyric acid; 24: isocitric acid; 28: lactic acid; 29: leucine; 32: malic acid; 37: phenylalanine; 40: pyroglutamic acid; 45: threonine; 46: tryptophan; 47: tyrosine; 48: valine. Figure 2b. A portion of the 1H-15N HSQC spectrum of a mixture of standard compounds at various concentrations obtained after tagging with 15N-cholamine. The peak numbers correspond to the compounds shown in Table I.
Figure 3
Figure 3
A portion of the 1H-15N HSQC spectrum of human urine tagged with 15N-cholamine. 1: aconitic acid; 2: adipic acid; 3: alanine; 5: arginine; 6: asparagine; 7: aspartic acid; 9: citric acid; 12: formic acid; 15: glutamic acid; 18: glycolic acid; 19: hippuric acid; 24: isocitric acid; 28: lactic acid; 33: malonic acid; 39: propionic acid; 40: pyroglutamic acid; 43: succinic acid; 45: threonine; 46: tryptophan.
Figure 4
Figure 4
Typical LC-QTOF-MS spectra for formic acid and pyruvic acid obtained after tagging with the smart isotope tag, 15N-cholamine. The permanent charge on the tagged metabolites enables their sensitive detection; the observed peak is from the intact tagged metabolite.
Scheme 1
Scheme 1
Synthesis of 15N substituted phthalimide.
Scheme 2
Scheme 2
Alkaline and acid hydrolyses of the 15N substituted phthalimide to yield 15N-cholamine.
Scheme 3
Scheme 3
General reaction for tagging carboxyl-containing metabolites with the smart isotope tag 15N-cholamine.
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