A novel method for simultaneous measurement of concentration and enrichment of NO synthesis-specific amino acids in human plasma using stable isotopes and LC/MS ion trap analysis.

Autor: Oosterink JE; Department of Pediatrics, Emma Children's Hospital, Academic Medical Center, Amsterdam, The Netherlands., Buijs N; Department of Surgery, VU University Medical Center, Amsterdam, The Netherlands; Department of Surgery, Medical Centre Alkmaar, Trial Centre Holland Health, Alkmaar, The Netherlands., van Goudoever JB; Department of Pediatrics, Emma Children's Hospital, Academic Medical Center, Amsterdam, The Netherlands; Department of Pediatrics, VU University Medical Center, Amsterdam, The Netherlands., Schierbeek H; Department of Pediatrics, Emma Children's Hospital, Academic Medical Center, Amsterdam, The Netherlands; Department of Pediatrics, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: h.schierbeek@amc.nl.
Jazyk: angličtina
Zdroj: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences [J Chromatogr B Analyt Technol Biomed Life Sci] 2014 May 01; Vol. 958, pp. 10-5. Date of Electronic Publication: 2014 Mar 15.
DOI: 10.1016/j.jchromb.2014.03.005
Abstrakt: Stable isotope studies offer the opportunity to study the in-depth metabolic pathway of glutamine, citrulline, and arginine amino acids involved in NO synthesis. The use of multiple stable isotopes can be used to elucidate the exact transformation of glutamine to citrulline and arginine de novo synthesis. This novel method provides a purification step using cation exchange resin in combination with a rapid and easy derivatization procedure for a precise and robust measurement of the concentration and isotopic enrichments of NO synthesis-specific amino acids using a liquid chromatography mass spectrometry (LC/MS) ion trap system with high sensitivity and selectivity. The ethyl chloroformate derivatization procedure is beneficial in terms of robustness, velocity, simplicity, and derivative stability. In addition, the ethyl chloroformate derivatization can be performed at room temperature in an aqueous environment without incubation and the isolation of the derivatives from the reaction mixture also serves as a purification step. The concentration and enrichment of NO synthesis-specific amino acids as well as phenylalanine and tyrosine to determine protein turnover, were measured with good inter-day precision for the concentration (<7.4%) and enrichment (<12.7%) in plasma samples at low and high levels. The low limit of quantification was 0.2μmol/L for most of the amino acids and the purification method showed to have good recoveries between 78% and 98%. No ion-suppression was observed by post-column infusion experiments. In order to develop new nutritional strategies, this novel method can be used to support the elucidation of the effect of administration of specific supplements on the glutamine-citrulline-arginine pathway by using stable isotope studies.
(Copyright © 2014 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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