1. The diurnal nature of nitrogen (N) homoeostasis was investigated in adults fed increasing protein intakes. N balance was estimated during a 48 h period of consecutive 12 h periods of feeding hourly meals and fasting, after 12 days of adaptation to diets containing 0.36 +0.01, 0.77 + 0.03, 1.59 +0.08 and 2.31 +0.65 g of protein day−1 kg−1. N losses were determined from measured urinary N excretion corrected for changes in the body urea pool, and estimated faecal and miscellaneous losses. [13C] Leucine and [2H5]phenylalanine balances were measured during a primed, continuous infusion of the two amino acids during the fasting and feeding phase on the second day.

2. Increasing fasting N losses were observed (47 +7, 60+6, 95+15 and 140+36 mg day−1 kg−1) on the four intakes, with corresponding increasing fed gains of 8.2+3.9, 40.2+7.1, 112+24 and 180+ 56 mg day−1 kg−1.

3. Increasing fed-state amino acid gains with increasing protein intake were observed with both [13C]leucine and [2H5]phenylalanine, whereas increasing fasting amino acid losses were confirmed with [13C]leucine.

4. The N equivalent of the leucine oxidation rate was mostly in the range of 10–50% lower than expected from the N excretion rates. This may reflect the timing of the amino acid balance measurements and non-uniform rates of gain and loss throughout the diurnal cycle.

5. We conclude on the basis of both N and amino acid balances that the amplitude of the diurnal cycling of body protein N in human adults increases with increasing dietary protein intake. Thus one component of the protein requirement for N balance reflects a demand for repletion of fasting losses which increases with increasing habitual protein intake.

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