Abstract An investigation was conducted to ascertain whether gastric lipase has a primary role in the hydrolysis of milk fat in the abomasum of the calf. The treatments of rumen-fistulated calves were: (a) fresh whole milk given orally from a nipple-feeder; (b) fresh whole milk passed through a tube directly into the abomasum; (c) same as (b) except that the calf was sham-fed immediately before abomasal feeding; and (d) same as (b) except that sham-fed milk from another calf was given in lieu of fresh milk. Abomasal contents withdrawn at various intervals after feeding were analyzed for total free fatty acids. The level of free fatty acids was high only in those treatments involving the passage of milk through the oral cavity, the site of pregastric esterase secretion. This observation indicated that gastric lipase, if present at all, contributed little to the abomasal hydrolysis of fat. Electrophoretic analyses of both abomasal and intestinal digesta collected after the oral feeding of milk revealed a pattern of esterases similar to that reported previously for pregastric esterase. These observations substantiate the belief that pregastric esterase is primarily responsible for the digestion of milk fat in the abomasum.