N-15 LABELING AND PREPARATION OF MILK, CASEIN AND WHEY PROTEINS

One of the main problems of in vivo protein digestion studies is the c ontribution of endogenous protein secretion. Nutritional studies have shown that the use ot stable isotopes in this context is an appropriat e technique to perform certain metabolic experiments with proteins. Th us the purpose of this work was to determine the optimum conditions fo r the production of stable isotope N-15-labelled milk and for the subs equent partition of two crude fractions of milk proteins: casein and w hey proteins. N-15-labelled milk was prepared with milk from two lacta ting cows: one received daily 25 g ((NH4)2SO4)-N-15 and the second rec eived 50 g. Native phosphocaseinate (NPPC) and whey protein concentrat e (WPC) were separated from raw pooled N-15-milk (RPM) by membrane mic rofiltration and then purified through water diafiltration. The N-15-e nrichment of milk reached 0.4213 atom-%. excess (APE) and 0.5177 APE f or the cows receiving 25 g and 50 g ((NH4)2SO4)-N-15, respectively. Th e microfiltration technique used allowed to obtain from 47 kg RPM both WPC (1.3 kg) and NPPC (9.76 kg) with yields ot 34.4% and 82.5%, respe ctively. NPPC was 0.5070 APE N-15-enriched and consisted of 99.9% prot eic nitrogen. WPC was 0.4999 APE N-15-enriched and consisted of 96.8% proteic nitrogen. The N-15 enrichments of skim milk, NPPC and WPC were not significantly different (P < 0.05) and were high enough to perfor m in vivo metabolic experiments.