PMID: 9174264Apr 11, 1997Paper

Isolation and quantitation of isotopically labeled amino acids from biological samples

Journal of Chromatography. B, Biomedical Sciences and Applications
H M van EijkN E Deutz

Abstract

To face the problem of simultaneous isolation and quantitation of isotopically labeled amino acids in biological samples, two semi-preparative chromatographic methods were developed. One method was especially designed to isolate radioactively labeled amino acids for which we used derivatization with the fluorophore o-phthaldialdehyde (OPA), which is known to be easy and reliable. Isolation of amino acids labeled with stable isotopes required another approach as we wanted to use isotope ratio mass spectroscopy (IRMS), which can only be performed on pure, non-derivatized amino acids. Because the OPA probe cannot be removed after isolation of the derivative, we used 9-fluorenylmethylchloroformate (FMOC) instead. This probe is linked to an amino acid via a peptide bond which can easily be broken by gas-phase acid hydrolysis (103% recovery after 5 h at 150 degrees C: S.D=3.5%, n=14). Run time (injection to injection) was 60 min for the OPA method and 75 min for the FMOC method. Both fluorescence and UV absorbance detection can be employed. The coefficient of variation (C.V.) for peak area measurement was below 2% for most OPA amino acids and below 3% for most FMOC amino acids. At maximum, a total of 1000 microl could be injected, re...Continue Reading

Citations

May 19, 2000·The Proceedings of the Nutrition Society·A J Wagenmakers
Aug 6, 2004·Current Opinion in Clinical Nutrition and Metabolic Care·Hans M H van Eijk, Nicolaas E P Deutz
Sep 14, 2007·Mass Spectrometry Reviews·Jean-Philippe GodinGérard Hopfgartner
Jun 12, 2002·Clinical Nutrition : Official Journal of the European Society of Parenteral and Enteral Nutrition·M M HallemeeschN E P Deutz
May 2, 2003·American Journal of Physiology. Endocrinology and Metabolism·Marcella M HallemeeschNicolaas E P Deutz

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