Metabolism and Nutrition

Experiments were conducted to determine the conditions for assay of hepatic phenylalanine hydroxylase (PAH) activity in the chicken and to determine the developmental pattern of PAH activity in liver 25,000 × g supernatant. PAH activity was detected in liver supernatant and (postnuclear) 25,000 × g particulate fraction. Optimum assay conditions differed for the two cell fractions, the most notable difference being a broad pH optimum of 7.7 to 9.2 for the supernatant and 4.7 and 5.6 for the particulate fraction. The PAH activity in the supernatant increased to a maximum as L-phenylalanine concentration in the assay medium increased from 0.02 to 0.5 mM and 1.0 mM. Activity increased in the particulate fraction as the Phe concentration increased to 0.5 mM. Substrate inhibition of PAH activity occurred at Phe concentrations of 3 to 5 mM in the supernatant but not in the particulate fraction. Concentrations of the cofactor, 6(R)-5,6,7,8tetrahydrobiopterin, ranging from 0.09 to 0.75 mM, resulted in maximal PAH activity. The developmental pattern of PAH in supernatant was determined using a modified assay in which substrate and cofactor concentrations and pH were optimum. The PAH activity in liver supernatant was present at a low level in 11 d chick embryos and increased several fold between Days 15 and 17 to a maximum at Days 17 to 21. Activity declined at hatching to levels that were present in 11 to 15 d embryos and remained at this level in male chicks through 4 wk of age. Mature males had higher PAH activity than mature laying females. (