Porcine and bovine pepsins were very stable in 5·0 M-urea at pH 2·0 or 2·5 at 30 °C while chymosin and Mucor miehei and M. pusillus proteases were rapidly denatured under these conditions. These differences may provide the basis of a quantitative assay of the complement of individual milk clotting enzymes in a coagulant and a possible scheme is suggested.

Confirmation was obtained for the occurrence of an interaction between high and low molecular weight fractions of milk during its pasteurization, resulting in the formation of germinant(s) for Bacillus cereus. Various milk fractions were added to supernatants or dialysates of raw skim-milk, pasteurized and assayed for germinant. Micellar casein obtained by high speed centrifugation of raw milk stimulated germinant formation, but whole casein obtained by acid precipitation was less stimulatory and individual casein fractions showed no stimulation. A membrane rich ‘fluff’ layer obtained by high speed centrifugation of raw milk was highly stimulatory and suggested the possibility that the somatic cell count of milk may be related to germinant production.

Using milk from endotoxin infused and infected quarters of individual cows, a correlation between cell count and germinant formation was clearly demonstrated. Adjustment of the cell counts of individual milk samples was generally accompanied by corresponding changes in germinant formation, though removal of cells by filtration did not have this effect, possibly since germinant precursor had leached from the cells and remained in the milk. Pasteurization was necessary, not simply to express germinant from cells but to effect an interaction of germinant precursors. It is not clear whether the cell associated germinant precursor is derived from the cells themselves or from a related component. Addition of blood serum proteins was without stimulatory effect.

Since low cell count milks sometimes supported appreciable germination after pasteurization, but showed still higher levels when the cell count was raised, germinant may be formed via 2 pasteurization dependent processes, one between milk constituents alone and the other involving both milk constituents and somatic cells.

Several factors which influence the shape of activation curves relating the degree of agitation-induced lipolysis in raw milk to the temperature of agitation were investigated. Curves of the type previously reported (Fitz-Gerald, 1974) containing a maximum at 12–15 °C resulted from low speed agitation with incorporation of air, with fresh (< 12-h old) milk, with milk of fat content ≥ 3·0 % approx. and by low temperature incubation of activated milks. Activation curves containing no distinct maximum at 12–15 °C were obtained from high speed agitation and homogenization, with milks of low fat contents and with aged (≥ 24-h old) milk. Agitation at low temperatures resulted in a maximum degree of activation in fresh milk after 2–4 h cold storage.

During agitation of raw milk redistribution of lipase activity between the cream and skim-milk phases occurred. The extent of redistribution depended on the time and temperature of agitation. Cream prepared from milk agitated at low temperatures contained lipase activity several times greater than cream from unactivated milk. The practical significance of this effect is discussed.

Levels of Cu and Fe in fresh milk and in bulk pasteurized milk were measured. The endogenous levels varied during the lactation period. The level of Cu in bulk pasteurized milk was greater than the endogenous level, suggesting some contamination. This apparent Cu contamination was greater in winter than in summer. No similar contamination was observed with Fe. The levels of Cu and Fe in butter manufactured each month and in retail butter were measured. Higher levels of Cu were detected in ripened cream butter produced in winter compared to that produced in summer. No similar trend was evident for Fe in ripened cream butter. Very low levels of Cu, Fe and lipid oxidation were detected in Irish sweet cream butter and consequently Cu and Fe contamination and resultant oxidation are not a problem in this product.

Porcine pepsin was rapidly denatured in phosphate buffers, pH 6·4–6·7, in the temperature range 31–39 °C and was only slightly more stable in milk under similar conditions. However, the enzyme was considerably more stable in Cheddar cheese curd in which the extent of denaturation was very markedly influenced by the pH of the milk at setting. Under normal cheese-making conditions, porcine pepsin was about equally stable with chymosin. Two modifications of the cheesemanufacturing procedure were developed which permit the manufacture of cheese almost free of coagulant and suitable for the assessment of the contribution of starter proteinases to proteolysis during cheese ripening.