N-15 ENRICHMENTS OF CASEIN AND PLASMA-PROTEIN AMINO-ACIDS IN COWS INGESTING N-15-LABELED AMMONIUM-SULFATE

The aim of this work was to determine by ion-exchange liquid chromatog raphy and isotope ratio mass spectrometry the specific N-15 enrichment of amino acids in casein and plasma proteins in cows receiving three successive daily oral doses (300, 150 and 150 g) of ((NH4)-N-15)(2)SO4 (10 atom per cent isotopic enrichment) and to examine the N-15 enrich ments obtained with regard to nitrogen transport and metabolism in the lactating cow. To investigate the N-15 distribution in amino acids in casein and in plasma proteins, samples of N-15-labelled casein and pl asma proteins were extracted either from a pool of several milkings (3 0-96 h after starting to administer the tracer) or from pooled venous blood (removed on the fourth day after the start of administration) fr om the four lactating cows. N-15 enrichments of the proteins studied, expressed as atoms percent excess, mere 0.2509 for casein and 0.0577 f or plasma protein. Chromatographic fractionation of the amino acid mix ture (protein hydrolysates) resulted in nine groups containing between one and four amino acids: Asp, Ser and Thr; Glu; Pro; Gly, Ala, Val a nd Met; Ileu and Leu; Tyr; Phe; His and Lys; and Arg. High N-15 incorp oration was demonstrated in all individual or groups of amino acids st udied. In both proteins, Glu appeared to be the most enriched amino ac id, Phe and Arg the least enriched. Most aliphatic molecules with a si ngle amino group were highly enriched. The much lower (3.5-7.7-fold) e nrichments in plasma protein compared with casein suggest considerable intracellular dilution at the site of liver protein synthesis. Finall y, the amino acid separation methods are discussed and suggestions for improving them considered.