The bactericidal activities of various concentrations of 3 disinfectants commonly used for teat disinfection were compared in vitro using small discs of pig skin previously contaminated with Staphylococcus aureus. Solutions containing 40 g/1 Na hypochlorite, 25 or 50 g/1 chlorhexidine digluconate or iodophor containing 5 g/1 iodine were found to have equivalent or superior bactericidal activity to a solution containing 10 g/1 Na hypochlorite. This in vitro technique offers a convenient alternative to in vivo tests for teat disinfectants and with minor modifications could be applicable to the testing of skin antiseptics in general.

The hygienic condition of 6 milking installations, 3 sanitized by circulation cleaning (CC) with chlorine-based chemicals and 3 by flushing with acidified boiling water (ABW), was tested using rinses of quarter strength Ringer's solution. The bacterial content of the rinses was determined using both colony counts and the direct epifluorescent filter technique (DEFT). A comparison of testing methods gave correlation coefficients between colony count and DEFT of 0·82 for plants using CC and 0·46 for plants using ABW.

Five strains of bacteria belonging to different genera and commonly found on milking equipment were exposed to various degrees of heat and to various concentrations of chlorine. The effects of such treatments on the staining characteristics of the organisms were studied. It was observed that Staphylococcus aureus and Streptococcus lactis, although killed by heat treatment, stained a bright orange when treated with acridine orange dye. Pseudomonas fluorescens, Escherichia coli and vegetative cells of Bacillus cereus did not take up the orange stain after heat treatment, nor did any of the 5 strains stain orange after treatment with NaOCl. It is suggested that the DEFT is a useful and rapid means of counting bacteria in rinses of equipment where sterilization is due primarily to chlorination, but in the absence of a stain which can differentiate more accurately between dead and living organisms its use is limited where sterilization is carried out solely by heat.

Possible mechanisms for protective roles of milk components on footand-mouth disease virus present in the milk of infected cows were examined. Light scattering bands collected from Ficoll-sucrose gradient fractions of skim-milk contained membrane-limited structures but these were non-infectious for bovine kidney cells, lnfectivity titres in buttermilk higher than those of the original cream or butter suggested association of virus with milk fat globules. Increased infectivity titres in skim-milk after treatment with SDS suggested release of virus particles from dissociated casein micelle subunits. Chelating agents, de-emulsifying agents and trypsin, which alter the structure of the individual milk components casein, lipid and milk fat globule membrane were without effect on infectivity titres.

The in vivo action of gastric proteinases on bovine milk proteins was studied in rats fed with skim-milk, by analysing gastric contents after 30, 60 and 240 min of digestion. Gastric proteolysis was already marked after 30 min of digestion and liberated a large number of peptides with different molecular weights. αs1-Casein and β-casein were still the main components of the stomach contents together with α-lactalbumin and β-lactoglobulin, which were little degraded, even after 240 min of digestion. A statistical analysis (multivariate method), made on several parameters (such as pH, N, and NPN) showed changes in the stomach contents during the digestion. The amino acid composition of the protein fraction was close to that of the diet, whilst that of the non-protein fraction was very different, being between the amino acid composition of the endogenous proteins and that of the diet.

Bovine caseins from skim-milk (micellar casein and acid precipitated casein) were separated into 4 discrete components by high performance gel chromatography on a TSK-GEL G4000SW columm. The resolution was better than that obtained by conventional gel chromatography which resulted in separation into 2 components at best. Our results showed that the second and third components consisted of αs1- and κ-caseins and αs1- and β-caseins respectively. They are presumed to be submicellar fractions. Samples prepared by removal of Ca phosphate from large and small micelles were rich in the third and the second components respectively. It is suggested that casein micelles consist of αs1- and κ-casein complexes and complexes and polymers of αs1- and β-casein which are situated preferentially on the periphery and in the interior of the micelles respectively.

Native proteolytic enzymes in good quality normal bovine milk readily hydrolyscd the caseins during incubation or storage, producing the γ-caseins, proteosc-peptone components 5 (PP5) and 8-fast (PP8F) and a considerable number of other unidentified fragments, many of which were also subsequently found in the proteose-pcptone fraction. The rate of casein hydrolysis was greater in pasteurized than in raw milk, with β-casein being slightly more susceptible to attack than αs1-easein. Measurements of γ-cascin and proteose-peptone formation have been made and it was found that PP5 was an intermediate product that was subject to further proteolysis while PP8F was a stable end-product. With the exception of component 3 (PP3), virtually all constituents of the proteose-peptone fraction increased during storage and appeared to be products of the action of proteolytic enzymes. Further evidence was obtained from the effects of various inhibitors that the principal proteinase of normal milk is plasmin, but slight differences were apparent between the protein breakdown patterns induced by storage and by added plasmin, which was consistent with the presence of more than one proteinase. Incubations in the presence of soya bean trypsin inhibitor to prevent plasmin action clearly revealed that another enzyme(s) was also involved.

Milk obtained from cows which were either infected by clinical mastitis or had been subjected to intramammary infusion of Escherichia coli endotoxin possessed high counts of somatic cells and very high levels of proteinase activity which hydrolysed the caseins almost completely in a few hours at 37 °C. The rate of hydrolysis of β-casein was slightly greater than that of αs1-casein, but in both cases hydrolysis was enhanced by 6 cycles of freezing and thawing to disrupt somatic cell membranes. A study of the relationship between proteinase activity and cell count suggested that only some of the proteinase activity originated in the somatic cells and also that the identity of the cells making up the total cellular population was important. Maximum proteolysis occurred at 50–60 °C, but the temperature-activity curve was a broad peak. Likewise the pH versus activity plot was very broad and was almost flat over the pH range 6–9. Experiments with a number of inhibitors of proteinases failed to give a clear cut pattern of inhibition. All evidence obtained was consistent with the view that several different enzymes with different pH and temperature optima and different specificities contributed to the overall hydrolysis of caseins in these milks. From electrophoretic band patterns one of these enzymes was clearly plasmin, but in high cell count milks other proteinases also became significant.

The caseinate complex in bovine milk was partitioned by differential centrifugation at both 20 and 4 °C into 4 micellar fractions and a fraction representing serum casein, and the protein composition of the fractions determined. At both temperatures the relative amount of κ-casein in the micellar caseins increased markedly and that of β-casein decreased appreciably with decreasing micelle size. The relative amount of αs2-casein also tended to decrease with decreasing micelle size, but the relative amounts of αs1- and γ-caseins, and an unidentified casein fraction, showed little systematic variation. The serum casein differed appreciably in composition from the micellar caseins, being very rich in β-casein and comparatively poor in αsl- and αs2-caseins, and the amount present at 4 °C was considerably greater than at 20 °C, with the increase being due almost entirely to β-casein, but with γ-casein also making a significant contribution. The changes in the composition and distribution of micellar and serum caseins induced by cooling milk at 4 °C were completely reversible when the milk was re-equilibrated at 20 °C for 18 h.

Aeration increased the growth and lipase production in milk by Pseudomonas fluorescens strain AFT 36, isolated from refrigerated bulk milk. A heat-stable lipase was isolated from a shaken milk culture of this microorganism by DEAE-chromatography and gel filtration in Sepharose 6B. The lipase-rich fraction from DEAE cellulose contained 3 lipases that were separated by gel filtration; only the principal lipase, which represented ∼ 71 % of total lipolytic activity, was characterized. The purified enzyme showed maximum activity on tributyrin at pH 8·0 and 35 °C; it had a Km on tributyrin of 3·65 mM. and was inhibited by concentrations of substrate > ∼ 17 mM. The enzyme was very stable over the pH range 6–9; it was relatively heat-labile in phosphate buffer in the temperature range 60–80 °C, where it was stabilized significantly by Ca2+. It was, however, very stable at 100–150 °C: the D values at 150 °C were ∼ 22 s and 28 s in phosphate buffer and synthetic milk serum respectively; the corresponding Z values in the temperature range 100–150 °C were ∼ 40 and ∼ 42 °C and the Ea for inactivation were 7·65 × 104 J mol-1 and 6·97 × 104 J mol-1 respectively.

A study has been made of the interdependence of the volume fraction and the degree of dispersion of the fat in creams subjected to single-pass homogenization. The effects of fat volume fraction φ could be incorporated into a semi-empirical correlation of the homogenizing variables established previously. In general, as φ increased from a low value the mean fat globule diameter at first decreased, reached a minimum and subsequently increased. The total area of the fat interface/m.l. volume when the globule diameter was a minimum proved to be independent of the homogenizing pressure and had a value of approximately 1·1 m2; the interface of maximum area was greater than this and, in contrast, its magnitude was pressure-dependent.

The overall results have been interpreted in relation to the variability of the disruptive energy within the homogenizing valve, competing effects of globule coalescence and hindered break-up, and the surface-active properties of the milk proteins.