Lactoperoxidase (LP), thiocyanate (SCN-), pH and somatic cell counts (SCC) were measured in mammary secretions from 20 cows collected 14 d before drying-off, 7 and 21 d after drying-off, and 3–18 d postcalving. The inhibitory activity of the secretions on Streptococcus uberis was determined and the susceptibility of the udder to infection by this organism was tested by intramammary infusion of 250 colony forming units at the above stages. LP, SCN-, pH and SCO increased during involution and fell postcalving. The secretions collected before drying-off, 7 d after drying-off and postcalving inhibited growth of Str. uberis.; those collected 21 d after drying-off did not. Inhibitory activity in pre-drying-off secretions was destroyed by heating and restored by addition of LP, glucose and glucose oxidase, but addition of these substances to secretion 21 d after drying-off did not provide a full inhibitory system. The growth of Str. uberis in the secretions was correlated with intramammary susceptibility, since challenges with Str. uberis at 14 d before drying-off, at 7 and 21 d after drying-off and postcalving led to 43·8, 25·0, 81·3 and 37·5% of quarters becoming infected. It is suggested that the LP/SCN-/H2O2 system plays a role in protecting the lactating mammary gland from infection with Str. uberis but becomes ineffective as involution progresses.

The effect of subclinical mastitis on levels of plasminogen and plasmin in milk from cows in a high-yielding herd was investigated. Comparisons were made with levels of milk Na, antitrypsin and N-acetyl-β-D-glucosaminidase (NAGase). In samples from mastitic quarters plasminogen activity, as measured after activation to plasmin, increased by only 21% and plasmin by 82%, while NAGase increased by 307 %. Plasminogen was the only component that was normally distributed, all other components showed more or less skewed distributions. Plasmin and plasminogen were significantly related to the other components. However, plasminogen plateaued when the other components continued to increase. There was thus no further increase in plasminogen with the severity of inflammation as with the other components. Plasmin showed a similar although less pronounced tendency. Results of treatment of mastitic whey samples with acid suggested that the non-linear increase in plasmin activity was due to interaction with acid-labile proteinase inhibitors. Mastitis led to dissociation of plasminogen and plasmin from the casein micelles. The degree of activation of plasminogen was higher with casein-associated than with soluble plasminogen in both healthy and mastitic milks. Plasmin was very closely related to milk Na, which is a sensitive indicator of epithelial integrity. It is suggested that plasmin contributes to Na leakage into milk by degrading membrane proteins of the epithelial lining. Plasminogen and antitrypsin, which are both plasma proteins, were not identically affected by stage of lactation, indicating nonidentical modes of transport from plasma to milk.

The free fatty acid (FFA) levels after milking (initial FFA) and after 22 h storage at 4°C (FFA-22) were determined in milk from 57 cows every 2 weeks for one year. FFA levels which were higher in summer than in winter were explained by later stages of lactation occurring in summer. Initial FFA content increased regularly during lactation. FFA-22 content increased after 12 weeks, and further after 32 weeks of lactation. The first increase was due to stage of lactation. The second increase, which appeared in late lactation, was essentially due to the stage of pregnancy combined with the effect of milk yield. FFA-22 content increased in the second part of pregnancy, chiefly after 24 weeks. A low milk yield (≼ 5 kg) further increased lipolysis, but only after 16 weeks of pregnancy. The effect of stage of pregnancy may have been enhanced by management factors such as feeding conditions. In this investigation, the seasonal FFA variation in bulk tank milk could be explained entirely by physiological factors.

Milk serum, whey and milk ultrafiltrate were examined by 31P nuclear magnetic resonance (31P-NMR). About 20 phosphorylated milk constituents gave rise to resonances in the spectra. Most of these have been assigned to such well-known milk constituents as inorganic phosphate, N-acetylglucosamine-1-phosphate and glycerophosphorylcholine. Resonances from previously unknown constituents such as phosphocreatine were also observed. When the pH-dependence of inorganic phosphate, N-acetylglucosamine-1 -phosphate, glycerophosphorylcholine and gly-cerophosphorylethanolamine was examined it was observed that the resonance of inorganic phosphate overlapped that of N-acetylglucosamine-1-phosphate around neutral pH. This is the most probable explanation as to why this constituent was not observed in earlier 31P-NMR studies on milk.

Using either Sephacryl S-300 or TSK-G4000SW gel chromatography, the larger of the two major components of bovine casein micelles (F2) was eluted at the same retention time irrespective of sample concentration and column temperature. Its mol. wt was estimated to be 500K by examination of the eluate from a TSK-gel G4000SW column using a low-angle laser light-scattering photometer and a precision differential refractometer. With the second component (F3), retention time in Sephacryl gel chromatography was significantly affected by both sample concentra-tion and volume applied. Limiting retention times for F3 could be obtained by extrapolation of these two parameters to higher and lower values, and corresponded to those of water-soluble protein with mol. wt 200K and 100K respectively. Approach to sedimentation equilibrium technique at the above limiting sample concentrations gave values of F3 consistent with the above molecular weight. Sephacryl gel chromatography at either low sample concentration or low column temperature resulted in a new component being eluted at a position of mol. wt 25K

The strength of binding of the individual caseins and the nature of the bonding within bovine casein micelles were examined through dissociation of the micelles by dialysis of skim milk either against phosphate-free buffers containing 3 or 6 mm-CaCl2, or against buffers that were nearly saturated with respect to micellar calcium phosphate, but which had a free Ca2+ concentration in the range 0·4–5·9 mm. Dissociation was followed by ultracentrifuging the dialysed milks and determining the partition of the total and the individual caseins between the pellet and serum. During dialysis against the phosphate-free buffers both colloidal Ca and Pi in the milks decreased and about 30 % of the Pi could be removed without significant casein dissociation. With further loss of Pi, however, increasing dissociation occurred and the proportions of the individual caseins retained in the casein pellet were in the order αs2- > αs1- > β- ≈ κ-casein. Dialysis against the calcium phosphate buffers resulted in no loss of colloidal Pi but colloidal Ca increased with the free Ca2+ concentration of the buffer. Little change in the casein partition occurred in the presence of more than 1 mm free Ca2+, but serum casein increased markedly at lower levels, and the strength of binding of the individual caseins in the pelleted casein was in the order αs2-> αs1- > β- > κ-casein. In both types of buffer, dissociation is considered to occur through the breaking of linkages between the caseins and inorganic constituents. Analysis of the amino acids in a calcium phosphate-rich material obtained after exhaustive proteolytic digestion of casein micelles suggests that these linkages involve the phosphate centres of the caseins.

Casein micelles in a reconstituted bovine skim milk were dissociated by dialysing them against various concentrations of EDTA. Gel chromatography showed a gradual but limited dissociation when micellar Ca was chelated and two main fractions were isolated. Monolayer technique and gel electrophoresis were used to characterize the interfacial properties and composition of the fractions. The first fraction contained highly cohesive complexes that did not unfold at the air-water interface. From π-A isotherms the close-packing area (A0) was 0·08 m2 mg-1 and was related to an average particle diameter of 9·1 nm. κ-Casein was concentrated in this fraction in an equimolar ratio with β-caseins and a variable ratio with αs-casein (0·55–1·10). These particles are thought to represent submicelles. The complexes of the second fraction were easily disrupted and spread out at the interface like casein monomers, with a close-packing area of 1·05 m2 mg-1. Two different complexes were identified in this last fraction. One of them contained an approximately equimolar ratio of αs- and β-caseins, and the other was composed exclusively of αs-casein. The results are discussed in relation to casein interactions leading to micelle formation.

The effect of heat treatment on the profiles of casein micelles obtained at 340 and 280 nm from controlled pore-glass chromatography of milk was studied in a series of unhomogenized whole milk samples which had been heated in a pilot plant at 75, 90, 105, 120, 135 and 150 °C for holding times of 4, 16, 32, 72, 150 and 300 s. A second experiment was carried out in which whole milk was preheated at temperatures between 70 and 110 °C for 13, 60 and 180 s before being treated at 120 °C for 4 s or 150 °C for 16 s. The average micelle size in some of the samples was determined by photon correlation spectroscopy. Profiles were divided into four fractions in which micelle size decreased from fraction 1 to fraction 4. The dimensions of these fractions were expressed in terms of size relative to that of the total profile. Fraction 4 was not affected by any of the heat treatments. Fractions 1–3 were significantly affected only by temperatures above 120 °C. Between 120 and 150 °C fraction 1 increased significantly while fractions 2 and 3 showed a corresponding decrease. Holding time affected fraction 1 only above 120 °C. For fraction 2 there was no effect in < 32 s at temperatures less than 150 °C. Fraction 3 was the most sensitive to changes in holding time, but no effect was observed below 105 °C. Average micelle size measurements indicated that a substantial size increase only occurred in those milks heated at 135 °C for longer than 16 s and in all the samples at 150 °C. To assess whether this increase in micelle size was reversible, samples were dialysed against a 70 mm-NaCl solution containing 10 mm-EDTA and then separated by column chromatography on Sephacryl S-300. The proportion of materials which was undissociated by dialysis remained unchanged for temperatures below 120 °C, but progressively increased with treatments more severe than 120 °C for 16 s. Column profiles were not influenced by variations in the conditions of preheating

Taking as an example a tryptic hydrolysate of bovine β-casein, it was shown that the method used for the determination of optimal elution conditions for isocratic systems also applied to the separation of complex peptide mixtures by reversed-phase high-performance liquid chromatography with linear gradients. Using a C18 column and the same solvents, successive studies of the influence of flow rate, pH and temperature allowed a satisfactory separation of the sample in less than 30 min. Valuable information on the specificity of the action of trypsin on β-casein was deduced from the yield of the eluted peptides.

The report that addition of KI03 (0·1 mm) to milk before ultra high temperature (UHT) treatment induces extensive proteolysis during subsequent storage at 37 °C was confirmed. None was produced by addition of H202 KMn04 or K2Cr207. The pH optimum for KI03-induced proteolysis was between 7·0 and 8·0 and the temperature optimum 37—45 °C. β-Casein was particularly susceptible and the proteolysis pattern was similar to that caused by indigenous alkaline milk proteinase (MPA, plasmin). Addition of plasmin to milk before UHT treatment (140 °C/10 s) caused slight proteolysis during subsequent storage but addition of 0·1 mm-KI03 and plasmin caused extensive proteolysis which was prevented by addition of soyabean trypsin inhibitor, indicating the probable involvement of plasmin in KI03-induced proteolysis in UHT-treated milk. Equally extensive proteolysis occurred in serum protein-free casein micelle systems (SPFCM), with or without KI03, during storage at 37 °C following UHT treatment, indicating a role for whey proteins in KI03-induced proteolysis. Addition of β-lactoglobulin (β-lg) to a SPFCM system inhibited proteolysis, but extensive proteolysis occurred in a SPFCM system containing both β-lg and KI03. MPA-free Na caseinate (prepared by heating at 140 °C for 7 min) underwent extensive proteolysis when treated with plasmin before UHT treatment; proteolysis was inhibited by addition of °-lg to this system and KI03 reversed the inhibitory effect of β-lg. Plasmin proteolysis of isolated αs1-casein was inhibited by denatured β-lg (90 °C/30 min) at a level of 4 mg/ml but not by native β-lg. When denatured in the presence of KI03, β-lg had a lower free SH content than the control and was less inhibitory for plasmin in proteolysis of isolated αsl-casein. The results show that denatured β-lg inhibits plasmin proteolysis of caseins in UHT milk and that inhibition is prevented by KI03. This inhibition may occur via thiol–disulphide interchange, which is prevented if the SH group of ²-lg is oxidized by KI03, thus permitting the stimulatory effect of KI03 on proteolysis in UHT-treated milk.

The rates of change in light reflectance and in CIELAB tri-stimulus colour values were compared for direct and indirect ultra heat treated (UHT) and sterilized milk in glass and polyethene bottles stored under different conditions. The rate of change of milk reflectance was higher at shorter wavelengths and the milk colour changed more rapidly at 30 and 37 °C than at room temperature. Sterilized milk in polyethene bottles was bleached when stored in an illuminated cabinet. The colour of skimmed milk changed more rapidly than that of whole milk. The rate of change in reflectance of direct UHT milk above 590 nm was found to be higher than in other milks. A statistical interaction was found between the fat content and the storage temperature for the CIELAB values. The homogenization pressure used in processing UHT milk samples did not affect the rate of change of the milk colour. The main implication of these findings may be that milk colour cannot be used to assess the heat treatment of a milk when the age or storage conditions of the milk are unknown.

Incubation of bacteria with casein at 30 °C (pH 7·2) resulted in the formation of a number of peptide fragments. The peptides were separable when subjected to electrophoresis. A proteinase-positive strain of Streptococcus lactis preferentially hydrolysed the β-casein moiety of whole casein, and after incubation for 5 min produced a characteristic pattern of four peptides on sodium dodecyl sulphate (SDS) polyacrylamide gels. After 30 min incubation only one peptide remained. This single polypeptide was then further hydrolysed to produce one (60 min) and eventually two (120 min) lower molecular weight peptides. Hydrolysis was generally complete after about 480 min. Two proteinase-negative variants of this strain did not hydrolyse casein. Crude proteinase preparations gave the same characteristic peptide patterns. SDS-polyacrylamide gel electrophoresis was used to screen more proteinase-positive strains of Str. Lactis for casein hydrolysing activity. Of those so far tested all produced the same peptide patterns from β-casein. Proteinase negative variants of all these strains did not hydrolyse casein.

The effect of milk containing increased somatic cells on the starter organisms Streptococcus thermophilus, Lactobacillus bulgaricus and L. acidophilus was examined. Increased somatic cell count resulted in stimulation of Str. thermophilus owing to increased proteolysis, but inhibition of L. acidophilus as a consequence of increased phagocytic activity of the polymorphonuclear leucocytes

Wateractivity (aw), pH, and several chemical components were determined in 20 samples of Blue cheese belonging to the varieties Bleu de Bresse, Cabrales, Danablu, Edelpilzkäse, Gorgonzola and Roquefort. The aw in these cheeses could be calculated from chemical data on a water content basis (g/100 g H2O) by two simple equations:

where SN is the soluble nitrogen.

Differences between calculated and measured aw using these relationships were similar to those obtained with a more complex equation described by Rüegg & Blanc (1983).

Cheddar cheeses were prepared from low somatic cell count (SCC) milks and from milks containing 10, 20 and 30% added high SCC milk to give modestly elevated SCC. Increased SCO in the milk were associated with a reduction in the coagulum strength and an increase in moisture content of the cheeses. Cheese texture was affected significantly as judged both by the findings of an assessment panel and by instrumental physical measurement. The panel scores for flavour were also slightly altered. It was concluded that even a small increase in SCC is detrimental to cheese composition and quality, although no significant effects on yield were detected. Relationships between cheese yield, composition and properties, coagulation properties and composition of the milk were also studied. Yield was related to the total solids and fat in the milk but not to the coagulation properties or the flavour or texture of the cheeses. Coagulum strength, cheese composition and cheese texture were strongly interrelated.