SummaryAseptically drawn milk was made into small cheeses (100 g) under aseptic conditions. The influence of the cow's feeding régime on the composition of the cheese fat, and the hydrolysis of the cheese fat and protein by bacterial and native milk enzymes, was studied.In the absence of a strongly lipolytic bacterial cheese flora, the free fatty acid (FFA) content of cheese appeared to depend on several variables:1. The initial FFA content of the milk varied according to the mode of milking (the lowest was obtained by cannulation of the udder) and the feeding régime of the cow. When one of a pair of monozygotic twin cows was maintained on pasture and the other on winter feed (hay and concentrates), the cheese made from the milk of the latter contained less FFA than did that from the former.2. The major changes in the composition of the milk fat of the cows on winter regime were an increase of myristic and palmitic acids from 8 to 11% and 21 to 28%, respectively, and a decrease in stearic and oleic acid from 11 to 7·5% and 38 to 28%; minor changes in the other fatty acids were also observed.3. The milk lipase in raw milk cheese liberated substantially more FFA than did the weakly lipolytic lactic acid streptococci. Milk lipase was active at the low pH value of cheese, but was inactivated when the milk was pasteurized at 63°C for 30 min.4. Although rennet did not produce any amino acids in the mature cheeses it hydrolysed some of the cheese protein to nitrogen soluble in water (WS), trichloroacetic acid (TCA-S) or phosphotungstic acid (PTA-S). Some of the WS-N appeared to be utilized by the lactic acid streptococci. The native milk protease resisted the heat treatment used and liberated amino acids at the low pH value of cheese, but its contribution to the proteolysis was relatively less important than the contribution of the milk lipase to the fat breakdown in cheese.5. The free amino acids of the milk, which appeared in the whey, were not recovered in the cheese. The cheese made from unheated milk with δ-gluconic acid lactone contained low amounts of free amino acids apparently due to the action of milk protease. The proteolytic activity of the lactic acid bacteria increased the amino acid content appreciably.