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Isolation of amino acids by preparative gas chromatography for quantification of carboxyl carbon 13 C enrichment by isotope ratio mass spectrometry
Author(s) -
Smith Kenneth,
Scrimgeour Charles M.,
Bennet William M.,
Rennie Michael J.
Publication year - 1988
Publication title -
biomedical and environmental mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 0887-6134
DOI - 10.1002/bms.1200170407
Subject(s) - chemistry , chromatography , ninhydrin , mass spectrometry , amino acid , isotope ratio mass spectrometry , gas chromatography , isotope , isotopes of carbon , hydrolysate , gas chromatography–mass spectrometry , leucine , derivatization , organic chemistry , hydrolysis , biochemistry , total organic carbon , physics , quantum mechanics
We describe a rapid method for isolating amino acids from tissue hydrolysates as trifluoroacetyl isobutyl esters by preparative gas chromatography on 2% EGA. Determination of the 13 C isotopic enrichment of the isolated amino acid is performed by isotope ratio mass spectrometry following the conversion of amino acid carboxyl carbon to CO 2 by reaction with ninhydrin. Using L ‐(1‐ 13 C)leucine as tracer, the procedure permits the measurement of low‐level enrichments (0.01–0.04 atom% excess) from 10–50 mg of tissue protein. The accuracy and reproducibility of the technique is demonstrated with standards of known enrichment, since the isotope ratio observed is unaltered within experimental error, ±4%, with coefficients of variation of <5%. The applicability of the technique to a clinical study of protein metabolism is also demonstrated.