Mutant ddY/DAO- mice lacking D-amino-acid oxidase were examined for their ability to utilize D-phenylalanine in place of its L-isomer. Chemically defined milk powder devoid of phenylalanine was used as a basal diet. Adult ddY/DAO- mice and normal ddY/DAO+ mice having D-amino-acid oxidase lost weight every day when fed this diet. However, they maintained their weight when fed the milk powder diet supplemented with 0.33% L-phenylalanine. Although adult ddY/DAO+ mice maintained their weight when fed the milk powder diet supplemented with 0.33% D-phenylalanine, ddY/DAO- mice lost weight while feeding on this diet, which indicated that the ddY/DAO- mice could not utilize D-phenylalanine in place of its L-isomer. Both ddY/DAO- and ddY/DAO+ mice maintained their weight when feeding on the milk powder diet enriched with 0.41% sodium phenylpyruvate, which suggested to us that the ddY/DAO- mice had the ability to invert this alpha-keto acid to L-phenylalanine. Therefore, these results indicate that D-amino-acid oxidase is indispensable for D-amino acid utilization. This is consistent with the concept that D-amino acids are oxidatively deaminated to the corresponding alpha-keto acids by D-amino-acid oxidase and that these keto intermediates are then asymmetrically reaminated to their L-amino acids.