We investigated the effects of graded hyperaminoacidemia on protein metabolism in eight healthy, young (25 +/- 2 years), normal weight (BMI = 25 +/- 1 kg/m2), overnight-fasted human subjects. A balanced amino acid solution was infused for 180 min at five different rates: 0.5 (study I), 1.0 (study II), 2.0 (study III), 4.0 (study IV). and 6.0 (study V) mg x kg-1 x min-1 on separate days in random order. Studies were performed with [1-14C]leucine infusion and indirect calorimetry to calculate leucine oxidation (LOX), nonoxidative leucine disposal (NOLD) (an index of protein synthesis), and endogenous leucine flux (ELF) (an index of proteolysis). Basal total plasma amino acid concentrations averaged 1.85 +/- 0.1 mmol/l and increased to 2.27 +/- 0.1, 2.70 +/- 0.2, 3.84 +/- 0.2, 5.87 +/- 0.4, and 7.52 +/- 0.3 mmol/l in studies I-V, respectively. ELF decreased from a basal value of 2.27 +/- 0.2 to 2.12 +/- 0.2, 1.97 +/- 0.1, 1.73 +/- 0.2, 1.67 +/- 0.3, and 1.65 +/- 0.1 micromol x kg-1 x min-1 in studies I-V, respectively (P < 0.05 for study I vs. basal, P < 0.01 for studies II-V vs. basal, and NS for studies IV and V vs. study III). LOX increased from a basal value of 0.31 +/- 0.04 to 0.38 +/- 0.05, 0.41 +/- 0.02, 0.64 +/- 0.04, 1.11 +/- 0.07, and 1.56 +/- 0.05 micromol x kg-1 x min-1 in studies I-V (all P < 0.01 vs. basal; P < 0.05-0.01 for each study vs. preceding study). Basal NOLD averaged 1.96 +/- 0.2 and did not change significantly in studies I and II (2.03 +/- 0.2 and 2.10 +/- 0.1 micromol x kg-1 x min-1). In contrast, a significant increase in NOLD was observed in studies III, IV, and V (to 2.3 +/- 0.15, 2.74 +/- 0.2, and 3.25 +/- 0.7 micromol x kg-1 x min-1, respectively; all P < 0.01 vs. basal; P < 0.05-0.01 for each study vs. preceding study). The net leucine balance (difference between ELF and NOLD)(-0.31 +/- 0.06 micromol x kg-1 x min-1) became less negative in study I (P < 0.01 vs. basal) and positive during studies II-V when the rise in plasma total amino acid levels was >/= 50% above basal level (P < 0.01 vs. each preceding study). In conclusion, NOLD, ELF, and LOX exhibit a differential responsiveness to acute changes in substrate availability: 1) small increments (25-50%) in plasma amino acid levels inhibit ELF and stimulate LOX but have no effect on NOLD; 2) stimulation of NOLD is observed only with increments in plasma amino acid levels >/= 100% above basal values; and 3) increments in plasma amino acid concentrations > 100% above basal values cause a progressive dose-related increase in LOX and NOLD but do not induce any further inhibition of ELF.