15N enrichment of ammonium, glutamine-amide and urea, measured via mass isotopomer analysis of hexamethylenetetramine.
Ammonium is an important intermediate of protein metabolism and is a key component of acid-base balance. Investigations of the metabolism of NH(4)(+) in vivo using isotopic techniques are difficult because of the low concentration of NH(4)(+) in biological fluids and because of frequent artifactual isotopic dilution of the enrichment of NH(4)(+) during the assay. A new gas chromatographic mass spectrometric method was designed to monitor the (15)N enrichment and concentration of NH(4)(+) in vivo. These are both calculated from the mass isotopomer distribution of hexamethylenetetramine (HMT) formed by reacting NH(4)(+) with formaldehyde. The enrichment of NH(4)(+) is amplified four times since the HMT molecule contains four atoms of nitrogen derived from NH(4)(+). This allows the measurement of low (15)N enrichment of NH(4)(+), down to 0.1%. (15)N enrichment of urea and of the amide N of L-glutamine are measured by enzymatic release of NH(4)(+) and conversion of the latter to HMT. These new techniques facilitate in vivo investigations of the metabolism of NH(4)(+) and related compounds.
Yang
D
D
Department of Nutrition, Case Western Reserve University, D-201 Cleveland, Ohio 44106-4906, USA.
Puchowicz
M A
MA
David
F
F
Powers
L
L
Halperin
M L
ML
Brunengraber
H
H
D000577:Amides / Q000032:analysis; D000643:Ammonium Chloride / Q000378:metabolism; D000818:Animals; D004285:Dogs; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D005973:Glutamine / Q000032:analysis*; D008709:Methenamine / Q000032:analysis*; D009587:Nitrogen Isotopes; D000644:Quaternary Ammonium Compounds / Q000032:analysis*; D014508:Urea / Q000032:analysis
D000577:Amides; D009587:Nitrogen Isotopes; D000644:Quaternary Ammonium Compounds; D000643:Ammonium Chloride; D005973:Glutamine; D014508:Urea; D008709:Methenamine