Cathepsin B and D activities were assayed in homogenates of the mucosa of the proximal, middle, and distal sections of the small intestine of suckling rats of various ages and of adult rats. Cathepsin B activity was determined using N-benzoyl-L-phenylalanyl-L-valyl-L-arginine-p-nitroanilide as substrate in the presence of trypsin inhibitor (pH optimum: 5.0-5.5). Cathepsin D activity was determined by measuring the pepstatin A sensitive action on denatured haemoglobin (pH optimum: 3.0-3.5). In the distal intestine, the activities of both cathepsins increased 6- to 10-fold during the first 2 weeks post partum and then sharply declined to the low values seen in adult rats, but in the proximal intestine the activities were low throughout the suckling period. In vitro digestion of casein by ileal mucosa of 8-day-old rats was almost completely inhibited by the combined actions of leupeptin and pepstatin A. The results show that cathepsin B- and D-like proteinases could play an important role in the digestion of milk proteins by suckling rats.