A series of hydrolysates from commercial sodium caseinate and rennet casein were prepared using different commercial proteolytic enzymes. Increasing degree of hydrolysis, DH, resulted in markedly changed solubility of the products. The level of bitterness varied with the enzyme used. Non-bitter hydrolysates of sodium caseinate were obtained with a DH of 10–12 and protein solubilities of 99%. The rennet casein hydrolysates exhibited low levels of bitterness and high solubility (90%), even at high DH values (DH = 55).

On renneting, the electrical conductivity of milk decreased as viscosity increased. The sigmoidal time course of the decrease resembled the time course of shear modulus, but was more rapid. The total amount of change was independent of the amount of rennet and proportional to milk conductivity and its casein content. The conductivity change was interpreted as a change in the way casein micelles obstructed the path of the charge-carrying ions.

The surface energies of highly hydrated casein micelle layers isolated from variously pretreated skim milks have been determined by means of contact angle measurements. The long range Lifshitz-Van der Waals (LW) and the short range hydrogen bonding (SR) components of surface energy were determined using α-bromonaphthalene and water for contact angle measurements. Casein micelles isolated from untreated and heat treated milks showed similar surface energy values of about 63·5 mJ.m-2 with an LW component of 19·2 mJ.m-2 and an SR component of 44·3 mJ.m-2. The calculated attraction potential energy was − 0·7 mJ.m-2. Casein micelles isolated from renneted milk showed a surface energy of 33·0 mJ.m-2 with an LW component of 30·7 mJ.m-2 and an SR component of 2·3 mJ.m-2. The attraction potential energy of renneted micelles was nearly two orders of magnitude higher than those of micelles from other milks ( − 63·3 mJ.m-2). The SR component of interfacial energy accounted for 98% of this attraction potential. The importance of attractive forces in relation to casein micelle stability is discussed.

The extent of cream feathering was observed in samples of six brands of instant coffee at 80 °C and classified on a scale of 0 (non-existent)–3 + (very severe). The hypothesis that it is caused by the amount of chlorogenic acid in the samples was tested. Ultraviolet spectra were recorded between 230 and 486 nm, and the amount of chlorogenic acid was measured from the absorbance at 340 nm. It was found that: (i) u.v. spectra of a given brand of coffee prepared at different concentrations could all be superimposed when concentration and dilution were taken into account, and the spectrum did not change between room temperature and 80 °C; (ii) for a given coffee a minimum amount of chlorogenic acid was needed to cause feathering, but that this amount differed by up to a factor of 10 depending upon the coffee; (iii) only two fundamental shapes of spectra were observed and coffees could be classified by a ratio, R, which lay between two ranges: 0·82 < R < 0·90 or 1·10 < R < 1·27; (iv) a knowledge of the chlorogenic acid content of a sample and the value of R appropriate to that coffee could be used to predict the occurrence of feathering.

Two slime-forming strains (T5 and MLS96) and a non-slime-forming strain (HA) of Lactococcus lactis subsp. cremoris were investigated for autolysis and optimal conditions for the autolytic process were developed. The rate of autolysis was maximal in exponential phase cells in 0·01 M-Na phosphate buffer, pH 6·5–7·5, at 30–45 °C. Autolysis of the native exponential phase walls was activated by trypsin and inhibited by lipoteichoic acid and cardiolipin. Decreased trypsin activation was found in intact exponential phase cells and no activation in stationary phase cells. N-acetylmuramylhydrolase action in the autolytic system of Lc. lactis subsp. cremoris strains was indicated by the progressive release of reducing groups. No amidase or endopeptidase action was found. Great variation in the rate of autolysis of whole cells was observed between the two slime-forming strains. Intact cells of strain T5 exhibited decreased autolytic activity, but a higher rate of autolysis of isolated exponential phase walls. Autolysins from strain T5 walls exhibited lower hydrolytic activity against sodium dodecyl sulphate-treated walls of the other two strains as compared with strains MLS96 and HA. Quantitative analysis revealed higher protein and phosphorus and lower hexosamine and rhamnose in cell wall preparations of strain T5 compared with the other two strains. Results suggest that the decreased rate of autolysis of strain T5 cells may at least in part be caused by cell surface components other than cell-wall peptidoglycan. It is proposed that the difference in autolytic characteristics should be used as a criterion when selecting starter strains of lactococci (streptococci).

Eight commercially available yogurt starter cultures were examined for their ability to synthesize water-soluble vitamins. In order to yield comparable assaying conditions, 12·5% reconstituted whole milk was used for yogurt production throughout the experiments. When applying the traditional method of yogurt manufacture (short-time incubating at 42 °C/3–4 h) with all cultures the following vitamins were enriched during fermentation by more than 20%: thiamin (two cultures), pyridoxine (four cultures), folic acid (one culture), and biotin (two cultures). Two starter cultures were selected and used to compare vitamin profiles during the two different methods of fermentation. In contrast to short-time incubation, long-time yogurt production (30 °C/14–16 h) led to a lesser production of folic acid, but to increased concentrations of thiamin and of nicotinic acid.

Ultraviolet spectra of solutions of instant and filter coffees have been analysed as a linear combination of component Gaussian bands. We show that the ratio, R′, of two of these bands, one at 329 nm due almost entirely to chlorogenic acid, and the other at 272 nm due to a coffee component not appearing in the chlorogenic acid spectrum, is analogous to the ratio R (Hamboyan et al. 1989). The use of R which is easier to measure than R′ has therefore been justified on physical grounds, based on the existence of component spectral bands. Filter coffees appeared to exhibit behaviour similar to that of instant coffees.

To improve acid clotting properties, enzymic dephosphorylation of caseins with calf intestinal alkaline phosphatase (CAP) or potato acid phosphatase (PAP) was investigated. Greater dephosphorylation was achieved using αs1- or β-casein as substrates, compared to whole casein or skim milk. Electrophoresis of PAP-modified caseins revealed bands with lower mobility and a multibanded pattern in the β-casein region which was similar to that of human β-casein. On the other hand, CAP modification produced electrophoretic bands having lower mobility of the β-casein component, but with higher mobility in the αs1-casein component as well as increased net negative charge in the CAP-casein. PAP-casein formed a fine dispersion upon acidification to pH 4. with a microstructure similar to that of acidified human casein. Greater initial rates of hydrolysis by pepsin at pH 4 were observed for both CAP- and PAP-modified caseins, compared to bovine and human caseins. The rate and extent of hydrolysis remained high for CAP-casein but tended to level off with PAP-casein during sequential digestion with pepsin and pancreatin. There may be advantages in the use of partial dephosphorylation to improve acid clotting and digestibility properties of bovine casein for infant feeding.

A mathematical model for chymosin action on casein micelles is presented in a two-stage equation which results in a single curve demonstrating the lag time from enzyme addition to the end of coagulum firming. The model uses the Michaelis–Menten enzyme kinetics equation for the first reaction followed by an nth order reaction for the casein micelles agglomeration stage. The computer output using these equations shows that lag time is elongated as enzyme concentration is lowered. Regression analysis of time of gelation against l/E0 shows good correlation. Viscosity of the milk drops at the beginning of the κ-casein hydrolysis and increases thereafter, when the coagulum is being formed.

Zn binding by whole bovine and human casein and by purified bovine caseins and whey proteins was investigated by equilibrium dialysis. Bovine αs1 casein had the greatest Zn-binding capacity (˜ 11 atoms Zn/mol). Protein aggregation was observed as Zn concentration was increased and- the protein precipitated at a free Zn concentration of 1·7 mM. Zn binding increased with increasing pH in the range 5·4–7·0 and decreased with increasing ionic strength. Competition between Zn and Ca was observed for binding to αs1-casein indicating common binding sites for these two metals. Bovine β-casein bound up to 8 atoms Zn/ mol and precipitated at a free Zn concentration of ˜ 2·5 mM, while K-casein bound 1–2 atoms Zn/mol. Whole bovine and human casein bound 5–8 atoms Zn/mol and precipitated at a free Zn concentration of ˜ 2·0 mM. Scatchard plots for Zn binding to caseins showed upward convexity, possibly due to Zn-induced association of caseins. Apparent average association constants (K¯app) for all caseins were similar (log K¯app 3·0–3·2). Enzymic dephosphorylation of αs1- or whole bovine casein markedly reduced, but did not eliminate, Zn binding. Thus, phosphoserine residues appeared to be the primary Zn-binding sites in caseins. With the exception of bovine serum albumin. which bound over 8 atoms Zn/mol, the bovine whey proteins, β-lactoglobulin, α-lactalbumin and lactotransferrin, had little capacity for Zn binding.

The rapid isolation of major bovine caseins in gram quantities was investigated. Whole casein was precipitated from individual cow's milk by adjusting the pH to 4·6 and the precipitated casein was suspended in 4·5 M urea (pH 8·0) containing 0·02 M imidazole and 0·03 M β-mercaptoethanol, and bound on a QAE Zeta Prep 250 cartridge. Stepwise elution with the urea/imidazole β-mercaptoethanol buffer and varying amounts of NaCl gave five well resolved peaks, which were identified by polyacrylamide gel electrophoresis and fast protein liquid chromatography to be pure γ-casein, κ-casein. β-casein, β-casein and αs-casein, respectively. The ion exchange cartridge was regenerated by flushing with buffer containing 0·50 Μ-NaCl followed by equilibration with starting buffer before separation of next sample. The time required to run each sample including cartridge regeneration and equilibration was 4 hours.

About 90% of the Zn in bovine skim milk was sedimented by ultracentrifugation at 100000 g for 1 h. About half of the non-sedimentable Zn was non-dialysable, indicating that it was associated with protein, probably non-sedimented casein micelles. Casein micelles incorporated considerable amounts of Zn added to skim milk as ZnCl2, and at Zn concentrations ≥ 16 mM coagulation of casein micelles occurred. Ca was displaced from casein micelles by increasing ZnCl2 concentration and ˜ 40% of micellar Ca was displaced by 16 mM-ZnCl2. Micellar Zn, Ca and P1 were gradually rendered soluble as the pH of milk was lowered and at pH 4·6 > 95% of the Zn, Ca and P1 were non-sedimentable. These changes were largely reversible by readjustment of the pH to 6·7. About 40% of the total Zn in skim milk was non-sedimentable at 0·2 mM-EDTA and most of the remainder was gradually rendered soluble by EDTA over the concentration range 1–50 mM. This indicates that there are two distinct micellar Zn fractions. No micellar Ca or P1 was solubilized at EDTA concentrations up to 1·0 mM, indicating that both colloidal calcium phosphate (CCP) and casein micelles remained intact under conditions where the more loosely bound micellar Zn fraction dissolved. Depletion of casein micelles of colloidal Ca and P1 by acidification and equilibrium dialysis resulted in removal of Zn, and in colloidal Pi-free milk non-dialysable Zn was reduced to ·-2 mg/1 (˜ 32% of the original Zn). Thus, ˜ 32% of the Zn in skim milk is directly bound to caseins, while ˜ 63% is associated with CCP. Over 80% of the Zn in colloidal Pi-free milk was rendered soluble by 0·2 mM-EDTA, indicating that the casein-bound Zn is the loosely bound Zn fraction in casein micelles. A considerable fraction of the Zn in acid whey (pH 4·6) co-precipitated with Ca and Pi on raising the pH to 6·7 and heating for 2 h at 40 °C, indicating that insoluble Zn phosphate complexes form readily under these conditions. Studies on dialysis of milk against water, or dilution of milk or casein micelles with water, showed that CCP and its associated Zn is very stable and dissolves only very slowly at pH 6·6. The nature of Zn binding in casein micelles may help to explain the lower nutritional bioavailability of Zn in bovine milk and infant formulae compared with human milk.

Effects of the reaction variables pH, temperature, time, enzyme concentration and solids concentration on the hydrolysis and solubilization of lactalbumin slurries by Alcalase have been determined. Reaction progress curves, solubilization v. time, were very unusual with a maximum solubilization of 96% occurring at a short reaction time corresponding to a degree of hydrolysis of 11–12%. Further hydrolysis beyond this point produced a rapid decrease in solubilization to about 67%, followed by a further gradual increase in solubilization with prolonged hydrolysis. The unusual solubilization profile was also produced by increasing enzyme concentration at fixed reaction time. The effects of pH, temperature and solids concentration were similar to those found in the hydrolysis and solubilization of other insoluble proteins. The implications of the results for the design and operation of an enzymic hydrolysis process are discussed.

The cheesemaking properties of recombinant chymosin from Kluyveromyces lactis (Gist-Brocades Chymosin 610) were Compared with those of standard rennet in parallel trials with the same milk and mixed culture starter. For each pair of cheeses the cheesemaking characteristics, mass balance results, composition of the cheeses at 6 weeks and maturation rates were similar. A taste panel was not able to differentiate between the cheeses at 3, 6 or 12 months.

A new method for measuring the activity of mesophilic and thermophilic lactic acid bacteria is proposed based on measuring the pH of cultures at extremely short intervals (30–90 s) during growth and calculating several kinetic parameters, i.e. the maximum acidification rate (Vm), the time and pH at which Vm occurred, the time or pH range during which the observed rates were greater than Vm/2. These were easily calculated by coupling the pH meter to a microcomputer.

The phosphorylation of different caseins by a number of tyrosine-specific kinases has been studied. The different kinases showed a range of specificities, suggesting differences in recognition mechanisms. The site of phosphorylation of αs2-casein by one phosphorylase has been identified, and compared with analogous sites in other proteins.

An intracellular proteinase was purified from Lactococcus lactis subsp. lactis NCDO 763 after spheroplast formation from cell wall proteinase-deficient variants. The proteinase was active at pH 7·5 and 45 °C and affected by metalloenzyme inhibitors. Its specificity, determined on B-chain of insulin, was thermolysin-like. The B-chain of insulin was hydrolysed rapidly while hydrolysis of β-casein was slower. This enzyme has a Mr of ∽ 93000.

By linear regression analysis, a highly significant negative correlation (r = −0·96) was found between the mean ash concentration values (g/100 g moisture) and water activity (aw) of six types of processed cheeses (low-fat, semi-fat, fat, extra-fat, double fat and special). The regression equation aw = 0·9951 − 0·0032* (ash), applied to 40 cheese samples, yielded aw values which differed by < 0·005 aw units from those measured experimentally in 75% of the samples. The maximum differences between the calculated and experimental aw values (found in only two samples) were ±0·01 aw units.

Hydrogenated milk fats have been subjected to a process of fractional melting. In five of the six samples, the procedure yielded two fractions, defined as oil/upper solid and lower solid fractions, whilst in the remaining sample, three fractions (oil, upper solid and lower solid) were obtained. The oil and oil/upper solid fractions were rich in short-chain fatty acids (FA) (4:0 and 6:0), whereas the lower solid fractions contained large amounts of long-chain FA (16:0 and 18:0). Triglyceride distributions reflected these differences in FA composition, the oil and oil/upper solid fractions being dominated by triglycerides with carbon numbers (CN) 36–38; the dominant CN for the lower solid fraction was 48–54. Melting spectra, obtained using differential scanning calorimetry, showed that the ability of the fractionation procedure to separate on the basis of melting properties ranged from excellent to relatively poor, depending on the type of milk fat.

Dietary manipulation was used to produce a similar series of milks from both Friesian and Jersey cows. The gross compositions of the milks, the fatty acid (FA) composition of the milk fats, the distribution of molecular sizes in the triglycerides of the milk fat, the melting properties of the milk fats, and the whipping properties of creams containing 360 and 400 g fat/kg were measured. Changes in gross composition and FA composition were as expected from the use of dietary oil supplements, but it was established that the mathematical relation between 18:0 and 18:1 differed between breeds, the Jersey yielding a milk fat with a lower proportion of 18:1 for a given value of 18:0. Control diets free from added fat produced milk fats with essentially unimodal triglyceride distributions, whereas fatrich diets produced bimodal distributions. The slight differences in these distributions between breeds were merely a reflection of variations in FA composition rather than in synthetic procedures. Differences in the whipping properties of creams containing 360 and 400 g fat/kg were consistent with literature observations. Dietary manipulation had little effect on the whipping properties of creams derived from Friesian cows, but caused considerable changes in the corresponding properties of the creams from Jersey cows. The only property that behaved similarly in the creams from the two breeds was the butter time, i.e. the time taken for butter granules to form on prolonged whipping of the cream. A major determinant of the butter time appeared to be the proportion of the fat that was molten at the temperature at which the whipping experiments were carried out.