Isolation of amino acids by preparative gas chromatography for quantification of carboxyl carbon 13 C enrichment by isotope ratio mass spectrometry

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.