Details

Autor(en) / Beteiligte

• Bornø, Andreas
• van Hall, Gerrit

Titel

Quantitative amino acid profiling and stable isotopically labeled amino acid tracer enrichment used for in vivo human systemic and tissue kinetics measurements

Ist Teil von

• Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2014-03, Vol.951-952, p.69-77

Ort / Verlag

Netherlands: Elsevier B.V

Erscheinungsjahr

2014

Quelle

MEDLINE

Beschreibungen/Notizen

• •A sensitive LC–MS/MS method was developed for the quantification of 20 amino acids.•Achieved by using stable isotopically labeled internal standard for each amino acid.•Simultaneously amino acid tracer enrichments from human kinetic studies.•Interday accuracy for amino acids in spiked plasma was on average 97.3%.•Suitable for AA in small muscle specimens and differences across a tissue (a–v dif.). An important area within clinical functional metabolomics is in vivo amino acid metabolism and protein turnover measurements for which accurate amino acid concentrations and stable isotopically labeled amino acid enrichments are mandatory not the least when tissue metabolomics is determined. The present study describes a new sensitive liquid chromatography tandem mass-spectrometry method quantifying 20 amino acids and their tracer(s) ([ring-13C6]/D5Phenylalanine) in human plasma and skeletal muscle specimens. Before analysis amino acids were extracted and purified via deprotonization/ion exchange, derivatized using a phenylisothiocyanate reagent and each amino acid was quantitated with its own stable isotopically labeled internal standard (uniformly labeled-13C/15N). The method was validated according to general recommendations for chromatographic analytical methods. The calibration curve correlations for amino acids were on average; r2=0.998. Interday accuracy for amino acids determined in spiked plasma was on average 97.3% and the coefficient of variation (CV) was 2.6%. The ([ring-13C6]/D5Phenylalanine) enrichment CV's for machine reproducibility in muscle tissue fluid and plasma were 4.4 and 0.8%, and the interday variability was 3.4% and the recovery was 90.5%, respectively. In conclusion, we have developed and validated a method for quantitative amino acid profiling that meets the requirements for systemic and tissue human in vivo amino acid and protein turnover kinetics measurements. Moreover, citrulline, ornithine, π-methyl-histidine, τ-methyl-l-histidine, hydroxy-proline and carnitine were analysed but when similar precision and accuray are required an additional stable istopically labeled internal standard for these meatablites should be be added