We have conducted optimizaiton studies to develop a superior ¹³C-phenylalanine breath test for the diagnosis of liver disease. First, we examined the optimum ¹³C-labeling position in phenylalanine for use in a breath test based on infrared spectroscopic detection of ¹³CO₂ in exhaled air. L-[1-¹³C]Phenylalanine gave the best result. Next, a suitable dosage to give a short peak time (the time expressed in minutes as which ¹³CO₂ excretion is maximal) after administration was determined. The ¹³CO₂/¹²CO₂ ratio in exhaled air after administration of 100 mg/body of L-[1-¹³C]phenylalanine peaked sharply at 15 min. We also examined the effect of food on the hepatic metabolism of L-[1-¹³C]phenylalanine. We found that a fasting period of over 7 h before the test resulted in a higher ¹³CO₂ peak excretion. The peak appeared sooner than that in the ¹³C-phenacetin breath test and, therefore, the ¹³C-phenylalanine breath test appears preferable for the rapid evaluation of hepatic function.