Determination of 4-aminobutyric acid, aspartate, glutamate and glutamine and their 13C stable-isotopic enrichment in brain tissue by gas chromatography-mass spectrometry.
A selected-ion monitoring method was developed for measuring 4-aminobutyric acid, aspartate, glutamate, and glutamine in brain tissue. Natural isotopes of these amino acids and their stable-isotopic enrichment following intravenous infusion of a precursor, [13C]glucose, were quantitated. Frozen mouse brain tissue was homogenized in cold 80% ethanol, and the supernatant, equivalent to 1 mg of wet weight brain tissue, was extracted using solid-phase bonded silica ion-exchange columns. Aspartate and glutamate (dicarboxylic acids) were isolated from strong anion-exchange columns, whereas 4-aminobutyric acid and glutamine (neutral amino acids) were isolated from strong-cation exchange columns. n-Butyl ester pentafluoropropionyl amide derivatives of these amino acids were analyzed by gas chromatography-mass spectrometry using a methane positive chemical ionization mode after gas chromatographic separation on a wide-bore, fused-silica capillary column. The method is applicable to determination of brain concentrations of these amino acids as well as their fluxes following administration of a stable-isotopic tracer.
D000596:Amino Acids / Q000032:analysis*; D000818:Animals; D001224:Aspartic Acid / Q000032:analysis; D001923:Brain Chemistry; D002247:Carbon Isotopes; D008401:Gas Chromatography-Mass Spectrometry; D005971:Glutamates / Q000032:analysis; D005973:Glutamine / Q000032:analysis; D008297:Male; D051379:Mice; D005680:gamma-Aminobutyric Acid / Q000032:analysis
D000596:Amino Acids; D002247:Carbon Isotopes; D005971:Glutamates; D005973:Glutamine; D001224:Aspartic Acid; D005680:gamma-Aminobutyric Acid