Abstract: Phenylketonuria (PKU) is a metabolic disease in which the untreated patients can show irreversible mental retardation and the use of milk by phenylketonurics patients is forbidden. With the aim of preparing low-phenylalanine milk hydrolysates, to be used by phenylketonurics patients with no restriction, the Activated Carbon (AC) was used as the adsorbent support. Also, the effects of certain parameters were analyzed, such as enzyme type (proteases from Aspergillus sojae and from Bacillus subtilis), temperature (30 and 50°C) and enzyme:substrate (E:S) ratio (1:100 and 2:100). The efficiency of Phenylalanine (Phe) removal was evaluated by second derivative spectrophotometry, measuring the Phe content of milk as well as in its hydrolysates after AC treatment. AC showed to be an efficient Phe adsorbent because a Phe removal from 16.28 to 58.76% was achieved, yielding certain hydrolysates with less than 100 mg Phe 100 mL-1, that could be used in the diet of phenylketonurics. The highest Phe removal (58.26%, in average) was achieved using the protease from B. subtilis at 50°C and E:S ratio of 1:100 or 2:100. At 50oC, the use of B. subtilis protease was more efficient than A. sojae protease whereas at 30°C the trend was inversed. The E:S ratio of 2:100 yielded greater Phe removal than 1:100 for B. subtilis protease at 30°C and for A. sojae at 30 and 50°C. It can be inferred that the conditions used in the study showed to be efficient in obtaining milk with reduced Phe content, appropriate for the diet of phenylketonurics.