Average N-acetyl neuraminic-acid (NANA) content of milk from cow, sheep, goat, mare, donkey and sow (about 10 samples from each species) was 17·7, 10·1, 6·4, 6·1, 12·0 and 64·0mg/100 ml, of which 14·2, 8·4, 3·2, 3·1, 5·0 and 53·7mg/100 ml respectively was in the casein fraction. The NANA content of casein showed marked species variation ranging from an average of 2·18 % for the sow to 0·09 % for the goat.

About 28 % of the casein NANA of mare's milk was liberated in the fraction soluble in 12 % TCA by the action of rennin at pH 6·6 compared with 75 and around 100 % for donkey's and sow's milk respectively; about half the total NANA in the casein of cow's, sheep's and goat's milk was released under the same conditions.

Large differences in new infection rates occurred with half-udder milking machines in which one half of the cluster operated substantially free from vacuum fluctuations while the other half was subjected to various combinations of different types of fluctuation in vacuum. To increase the rate of new infection all teats were contaminated before and after each milking by immersion in a suspension of Streptococcus agalactiae and Str. dysgalactiae. A series of short-duration trials indicated that neither irregular nor cyclic fluctuations in vacuum acting alone were able to increase new infection rate. However, either the Nyhan and Cowhig irregular fluctuation, or a variant of it in combination with cyclic fluctuations in vacuum, was able to do so under the particular experimental conditions used.

A milk-clotting proteolytic enzyme was isolated and purified from the culture filtrate of Bacillus cereus strain x29 by fractionation with acetone or ammonium sulphate and subsequent column chromatography employing DEAE cellulose and DEAE Sephadex. The purified enzyme was found to be homogeneous by acrylamide gel electrophoresis from pH 3·5 to 8·6, with, a molecular weight of about 50000. The single absorption maximum of the native enzyme was at 277 nm and the value of at 280 nm was 7·79. Purification resulted in a 9-fold enhancement of activity with 24 % yield. The optimum activity of the enzyme was at pH 8·0 at 40 °C with casein as the substrate. The enzyme was found to be most stable at pH 6·0 and was stable to freezing and freeze-drying. Heavy metal ions were found to inactivate the enzyme, but no metal ion activation was found. Enzyme activity was inhibited irreversibly by EDTA and reversibly by 1,10-phenanthroline. The enzyme has been identified as a Zn-containing neutral protease.

The biochemical properties of rennin purified from calf stomach, Mucor pusillus and Endothia parasitica have been compared.

The specificities of the enzymes have been studied by separating by disk-gel electrophoresis the macropeptides released on incubation with acid casein and κ-casein. The products formed on incubation with rennin from calf stomach and M. pusillus showed some similarity although several bands were found which did not correspond to one another. The products formed with rennin from E. parasitica differed considerably from those with the 2 other enzymes.

The kinetic properties of the enzymes have been investigated in stopped-flow experiments on the enzyme-induced aggregation of κ-casein. The aggregation has been characterized by 2 parameters—the lag time prior to the increase in extinction and the maximum rate of change in extinction during aggregation.

The specific activity measured from the inverse value of the lag time was under optimal conditions nearly 2·5 times higher for rennin from calf stomach than for rennin from the 2 micro-organisms. The fact that with increasing enzyme concentration the maximum rate of change of extinction during aggregation was identical for all 3 enzymes indicates that the same product is responsible for the formation of the aggregates. The data suggest that the Km value for the aggregation of κ-casein by the enzymes is about 100 times smaller than Km for the proteolytic activity of calfstomach rennin on κ-casein. The possibility is suggested that the high specificity of the enzymes in milk clotting is due to the very low Km values for the peptide bond modifications involved in the formation of para-κ-casein, the first enzymic step in the clotting of milk.

Published data are available for the fatty-acid compositions and molecular-weight distributions of triglycerides in milk and adipose tissue from the guinea-pig, rat, mouse and rabbit, and in milk from the human, cow, mink and hamster. These data have been used to calculate the extent of non-random associations of fatty acids of different chain lengths in these triglycerides.

The results support, in general, the concept of non-correlative acylation, i.e. that during the biosynthesis of these triglycerides, fatty acids already present on the glycerol backbone do not influence the selective acylation of subsequent fatty acids. However, milk triglycerides from these animals show varying degrees of intermolecular specificity which can be explained in terms of positional specificity of fatty acids on the glycerol backbone.

From a series of short-term experiments under accentuated milking machine conditions predisposing to new mastitis infections, it appears that the cow is at increased risk near the end of a milking. The observations recorded are consistent with the view that bacteria implanted in the teat during milking as a result of the action of the machine are the less likely to be washed out the nearer implantation occurs to the end of milk flow.

The effects of 2, 4, 7 and 10% supplements of tallow to a basal diet low in fat on the composition of volatile fatty acids (VFA) in the rumen and on the secretion in milk of fat and its component fatty acids are reported. There were no significant effects of the tallow on rumen VFA or on the secretion of total milk fat. The yields in milk of stearic (18:0) and oleic (18:1) acids were positively correlated and those of palmitic (16:0) and myristic (14:0) acids negatively correlated with their respective dietary intakes. Apart from butyric acid which increased, the yields of other short-chain acids synthesized within the mammary gland decreased with increased intake of tallow. Possible factors influencing the output of total milk fat through a balance between increased transfer to milk of dietary fatty acids and decreased intramammary synthesis of fatty acids are discussed.