Identification of 4-deoxythreonic acid present in human urine using HPLC and NMR techniques

Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47907, USA.
Journal of pharmaceutical and biomedical analysis (Impact Factor: 2.98). 07/2009; 50(5):878-85. DOI: 10.1016/j.jpba.2009.06.007
Source: PubMed


The 1H NMR spectrum of urine exhibits a large number of detectable and quantifiable metabolites and hence urine metabolite profiling is potentially useful for the study of systems biology and the discovery of biomarkers for drug development or clinical applications. While a number of metabolites (50-100) are readily detectable in urine by NMR, a much larger number is potentially available if lower concentration species can be detected unambiguously. Lower concentration metabolites are thought to be more specific to certain disease states and thus it is important to detect these metabolites with certainty. We report the identification of 4-deoxythreonic acid, a relatively low concentration endogenous metabolite that has not been previously identified in the 1H NMR spectrum of human urine. The use of HPLC and NMR spectroscopy facilitated the unequivocal and non-invasive identification of the molecule in urine which is complicated by extensive peak overlap and multiple, similar resonances from other metabolites such as 3-hydroxybutanoic acid. High-resolution detection and good sensitivity were achieved by the combination of multiple chromatographic fraction collection, sample pre-concentration, and the use of a cryogenically cooled NMR probe.

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Available from: Emmanuel Appiah-Amponsah, Feb 19, 2015
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    • "The use of chromatographic methods to simplify sample matrices by isolating metabolites of interest prior to NMR analysis has high utility for a variety of biological investigations [24,25,26,27,28,29]. This approach has also benefited from the use of sample pre-concentration techniques such as solid phase extraction (SPE) and column trapping to extend NMR detection limits significantly and thus circumvent the issue of sample dilution attributed to solvent mixing in the chromatographic step [26]. "
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    09/2013; 3(3):575-591. DOI:10.3390/metabo3030575
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    • "A one-dimensional 1H selective TOCSY experiment (See additional file 1, Figure S1) observed three related 1H resonances in the same molecule. Further two-dimensional experiments [See additional file 1, Figure S2] allowed the full 1H and 13C characterization [See additional file 1, Table S1] of the two diastereoisomers of 2,3-dihydroxybutanoic acid [See additional file 1, Figure S3]: 4-deoxy-erythronic acid and 4-deoxy-threonic acid, both of which have previously been identified in human urine [41]. "
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