To determine tryptophan requirements with use of the indicator amino acid oxidation technique, we studied 5 healthy, premenopausal women who each consumed 6 different intakes of tryptophan. The energy-sufficient diet contained 1.0 g x kg(-1) x d(-1) of an L-amino acid mixture patterned after egg protein, with adequate phenylalanine (14 mg x kg(-1) x d(-1)) and excess tyrosine (40 mg x kg(-1) x d(-1)). The effect of graded alterations in dietary tryptophan on phenylalanine flux and oxidation was studied by using L-[1-(13)C]phenylalanine as the indicator amino acid. Graded increases in tryptophan had no effect on phenylalanine flux. Determined by two-phase linear regression crossover analysis, the mean tryptophan requirement based on indicator (phenylalanine) oxidation was 4.01 mg x kg(-1) x d(-1), with a safe intake at 5.02 mg x kg(-1) x d(-1). These results show that the indicator amino acid oxidation technique can be used to determine the requirement for amino acids such as tryptophan that cannot be successfully studied by direct oxidation because of their characteristic metabolism. Our value of safe intake for 95% of the population was found to be 43% higher than that reported previously for adults.