Metabolism of 15N-labeled amino acids in rat lens

Rat lenses were cultured 4-24 hr at 37 degrees C in balanced salt medium containing 5 mM [15N]-glutamate, [15N]-alanine, or amido-[15N]-glutamine. Free amino acids were extracted with 6% trichloracetic acid containing alpha-aminoisobutyrate as an internal standard, and trifluoroacetyl-n-butyl (TAB) derivatives were prepared. Amino acids were quantified by gas chromatography, and 15N enrichment in amino groups of several free amino acids was determined by mass spectrometry. Culture of lenses with either [15N]-glutamate or [15N]-alanine resulted in [15N]-labeling of the glutamate, aspartate, alanine, proline, serine and glycine pools. No detectable amino-15N (less than 5%) was observed in amino acids when lenses were cultured with amido-labeled [15N]-glutamine or with [15N]-ammonium chloride. It is concluded that the alpha-amino groups of several amino acids are actively involved in lens metabolism. In contrast, although glutamine may serve as the major source of glutamate for lens, the amido nitrogen of glutamine plays a minor role, if any, as a source of alpha-amino groups for amino acid metabolism.