Detection and quantitation of phosphorus metabolites in crude tissue extracts by1H and31P NMR: use of gradient assisted1H-31P HMQC experiments, with selective pulses, for the assignment of less abundant metabolites
| Autor: | R. Toffanin, Ove Eriksson, E Geimonen, Sergio Paoletti, Piero Pollesello, Franco Vittur |
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| Rok vydání: | 1995 |
| Předmět: |
Uridine Diphosphate Glucose
Resolution (mass spectrometry) Proton Metabolite Analytical chemistry chemistry.chemical_element 010402 general chemistry 01 natural sciences Phosphorus metabolism Phosphoenolpyruvate 03 medical and health sciences chemistry.chemical_compound Animals Radiology Nuclear Medicine and imaging Perchloric acid Rats Wistar Spectroscopy 030304 developmental biology 0303 health sciences Phosphorus Metabolism Adenosine Monophosphate Rats 0104 chemical sciences Liver chemistry Molecular Medicine Protons Quantitative analysis (chemistry) |
| Zdroj: | Scopus-Elsevier |
| ISSN: | 1099-1492 0952-3480 |
| DOI: | 10.1002/nbm.1940080503 |
| Popis: | The analysis of crude tissue extracts by NMR has proven to be of use in the study of metabolism due to the non-destructive and non-selective character of the technique. Lists of 1H and 31P NMR assignments of phosphorus metabolites in water solution at specified pH and ionic composition are of large general value but their usefulness may be limited when analysing complex mixtures of metabolites at low concentrations. In this work we report on the use of gradient-assisted proton detected multiple quantum 1H and 31P coherence experiments with selective pulses for the rapid and unambiguous assignments of some crowded regions in 1H and 31P spectra of crude extracts from rat liver. The amplitudes of the gradient episodes were calibrated to optimize the coherence transfer pathway between proton and phosphorus, and the delay for the evolution of the long-range coupling was calculated from values of 3JPH and 4JPH ranging from 1.4 to 7.5 Hz. Moreover, a selective 90 degrees Gaussian pulse on the 31P channel was introduced to increase the resolution in the F1-domain and make the method even faster. The procedure was then applied to unambiguously assign the ID 31P and 1H spectra of perchloric acid extracts of rat livers that had been stimulated with phenylephrine, dBcAMP and glucagon and thus detect changes in the concentration of less abundant metabolites such as phosphoenolpyruvate, UDP-glucose and AMP. The fact that the quantification of these metabolites by either 31P and 1H methods lead to different results is discussed, and the use of 1H NMR spectroscopy for the quantification of phosphorus metabolites whose signal are too weak or poorly resolved in a 31P spectrum is proposed. |
| Databáze: | OpenAIRE |
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