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A novel method has been developed to determine protein requirements, which is called indicator amino acid oxidation (IAAO). This technique has been validated by comparison with the "gold standard" nitrogen balance. Using IAAO we have shown that minimum protein requirements have been underestimated by 30%-50%. The National Academy of Sciences has for macro-nutrients proposed "Acceptable Macronutrient Distribution Ranges", which for protein is 10% to 35% of total energy. In practice, we suggest 1.5-2.2 g/(kg·day) of a variety of high-quality proteins.

Our studies have focused on the regulation of whole body and skeletal muscle protein metabolism in premature infants. Net deposition of protein is the result of a positive balance between protein synthesis and breakdown. To measure protein metabolism we have employed end-product studies with [15N]glycine and 13[C]leucine. Myofibrillar protein degradation was estimated by measuring the excretion of N-t-methylhistidine in urine. Energy expenditure and substrate utilization were also measured. Premature infants have high rates of protein synthesis (12 g.kg-1.d-1), twice those measured in children and four times those found in adults. Intrauterine malnourished babies have increased rates of protein turnover. Very low birth weight infants (less than 1500 g) have higher myofibrillar protein turnover than larger babies. Intravenous feeding decreases whole body protein turnover, and we estimate visceral protein synthesis to be approximately 4 g.kg-1.d-1. Suboptimal energy intake worsens nitrogen utilization by reducing the reutilization of endogenous amino acids for protein synthesis. We have also examined the effects of varying the source of nonprotein energy (i.e., glucose only versus glucose plus lipid) at requirement levels and have shown there is no effect on protein metabolism. Recent improvements in technology have opened the way to detailed study of individual amino acid metabolism in neonates in the future.

Rates of whole body amino nitrogen flux were measured in 16 obese adolescents undergoing weight reduction with a high protein low energy diet. The subjects received approximately 2.5 g of animal protein per day per kilogram ideal body weight and maintained nitrogen balance throughout the 18 days on the diet. Flux rates were calculated separately from the cumulative excretion of 15N in urinary ammonia and urea following the administration of a single dose of [15N]glycine. The pattern of 15N label appearance in urinary ammonia and urea nitrogen was followed for 72 h after the administration of [15N]glycine. Significant amounts of label continued to be excreted in both urinary ammonia and nitrogen for 36-48 h after label administration. The weight-reducing diet accelerated 15N cumulative excretion in urinary urea, but not in ammonia nitrogen compared with the control diet. Whole body nitrogen flux rates increased rapidly and significantly on the diet. Using the urea end product, this increase was evident on the 4th diet day, but not by the 7th or subsequent days. On the other hand, using the ammonia end product, flux rate increased markedly (p less than 0.0001) and remained elevated throughout the whole study. Our results demonstrate adaptive changes in whole body amino-nitrogen metabolism in response to the reducing diet. Different patterns of change are seen depending upon whether an ammonia or a urea end product is used. Our data thus add to the evidence for compartmentation of the body's amino-nitrogen pools.

We investigated the effect of human milk feeding on the nitrogen metabolism of appropriate-for-gestational age infants of birth weight 1.5-2.0 kg. Eight infants received pooled mature human milk. The remaining 20 were divided into two equal groups, who received one of two low-protein, milk-based formulae. The formulae were identical in composition except for the protein source, which was either casein- or whey-predominant. The three diet groups received similar total nitrogen (390 mg N.kg-1.d-1) and energy (500 kJ.kg-1.d-1) intakes. The human-milk-fed group, however, received a significantly higher intake of nonprotein and urea nitrogen and a significantly lower true protein nitrogen. Nitrogen metabolism was studied using a modified constant infusion of [15N]glycine, mixed with the feeding every 2-3 h. Urine was collected in approximately 3-h aliquots and analysed for total ammonia and urea nitrogen. Excretion of the 15N label was measured in urinary urea and ammonia. No differences were seen between the three diet groups in total [15N]urea or [15N]ammonia urinary excretion. However, the concentration of 15N in urinary urea in the human-milk-fed group was lower than in the two formula-fed groups. This reduction in concentration appeared due to a higher dietary intake of urea among the human-milk-fed group, and the consequent dilution of the label in the urine. As a result, protein turnover rates calculated from the [15N]urea end product were artificially raised in the milk-fed group, and were significantly higher than those in the formula groups.(ABSTRACT TRUNCATED AT 250 WORDS)

The pattern of urinary urea excretion and labelling with 15N was examined in eight meal-fed 6 to 9 year old children, over a 3-day period using a simulated constant infusion of the label. The children had cystic fibrosis but were healthy and in a good nutritional status at the time of the study. Reciprocal diurnal patterns of urea excretion and [15N]urea enrichment were noted and found to be suitable for mathematical description. Urea excretion was maximal in the evening at approximately 2000 and minimal at 0800, whereas the [15N]urea enrichment was maximal at about 0800 and minimal at 2000. In addition to the diurnal variation the [15N]urea enrichment increased exponentially to a plateau or isotopic steady state. The diurnal variation in [15N]urea enrichment resulted in large diurnal changes in the calculated rates of whole body amino nitrogen flux, synthesis, and breakdown. Flux rates were approximately 44% higher in the evening than in the morning. Synthesis rates were 19% higher in the evening, whereas breakdown rates were 27% greater in the morning. Mean amino nitrogen flux rates were 1.28 (SD 0.13) . g N . kg-1 . day-1. Isotope recycling was estimated from the slope of the [15N]urea enrichment curve between 30 and 54 h from the start of the study. There was a wide range in recycling, 2.9 - 19.4% (mean 11.4, SD 5.4). Some of the biological and pharmacological importance of the diurnal variation in the protein metabolism is discussed.