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Tracing 15 N with chemical reaction interface mass spectrometry: A demonstration using 15 N‐labeled glutamine and asparagine substrates in cell culture
Author(s) -
Kusmierz Jozef J.,
Abramson Fred P.
Publication year - 1994
Publication title -
biological mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1052-9306
DOI - 10.1002/bms.1200231207
Subject(s) - asparagine , glutamine , proline , alanine , amino acid , chemistry , biochemistry , incubation , metabolism , mass spectrometry , glutamate receptor , chromatography , receptor
This research demonstrates how the chemical reaction Interface mass spectrometry (CRIMS) approach works for a study of amino acid metabolism in cell culture. 15 N‐selective chromatograms from both the culture medium and the cytosol of human hepatoma Hep G2 cells that were incubated in the presence of either 12 mM (α‐ 15 N)glutamine or (α‐ 15 N)asparagine have been produced. The time course of the distribution of 15 N among different amino acids, as well as the enrichment for each amino acid, were observed over a 144 h period. Labeled glutamine was quickly converted into glutamate. After 144 h of incubation, the total amount of 15 N was distributed primarily among alanine (50%), proline (28%) and glutamate (21%). The 15 N enrichment of alanine and proline reached 44% and 41% respectively. Asparagine was only slowly metabolized by the cells. In addition to the 82% that was retained in asparagine, the remaining 15 N in the media at 144 h was found primarily in alanine (8%), glutamate (6.8%) and proline (2.2%). Their enrichments were 20%, 36% and 19% respectively. The minimum detectable amount was 17 pg of 15 N entering the CRI. CRIMS appears to be a powerful, facile approach for 15 N‐tracer experiments.