High-pressure treatment of milk: effects on casein micelle structure 
and on enzymic coagulation

High isostatic pressures up to 600 MPa were applied to samples of skim milk before addition of rennet and preparation of cheese curds. Electron microscopy revealed the structure of rennet gels produced from pressure-treated milks. These contained dense networks of fine strands, which were continuous over much bigger distances than in gels produced from untreated milk, where the strands were coarser with large interstitial spaces. Alterations in gel network structure gave rise to differences in rheology with much higher values for the storage moduli in the pressure-treated milk gels. The rate of gel formation and the water retention within the gel matrix were also affected by the processing of the milk. Casein micelles were disrupted by pressure and disruption appeared to be complete at treatments of 400 MPa and above. Whey proteins, particularly β-lactoglobulin, were progressively denatured as increasing pressure was applied, and the denatured β-lactoglobulin was incorporated into the rennet gels. Pressure-treated micelles were coagulated rapidly by rennet, but the presence of denatured β-lactoglobulin interfered with the secondary aggregation phase and reduced the overall rate of coagulation. Syneresis from the curds was significantly reduced following treatment of the milk at 600 MPa, probably owing to the effects of a finer gel network and increased inclusion of whey protein. Levels of syneresis were more similar to control samples when the milk was treated at 400 MPa or less.

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Heat stability of milk: pH-dependent dissociation of micellar κ-casein on heating milk at ultra high temperatures

Journal of Dairy Research ◽

10.1017/s0022029900024481 ◽

1985 ◽

Vol 52 (4) ◽

pp. 529-538 ◽

Cited By ~ 82

Author(s):

Harjinder Singh ◽

Partick F. Fox

Keyword(s):

Whey Protein ◽

Whey Proteins ◽

Heat Stability ◽

Casein Micelle ◽

Casein Micelles ◽

Ph Profile ◽

Ph Values ◽

Maximum Stability ◽

Β Lactoglobulin ◽

Maximum Minimum

SUMMARYPreheating milk at 140 °C for 1 min at pH 6·6, 6·8, 7·0 or 7·2 shifted the heat coagulation time (HCT)/pH profile to acidic values without significantly affecting the maximum stability. Whey proteins (both β-lactoglobulin and α-lactalbumin) co-sedimented with the casein micelles after heating milk at pH < 6·9 and the whey protein-coated micelles, dispersed in milk ultrafiltrate, showed characteristic maxima–minima in their HCT/pH profile. Heating milk at higher pH values (> 6·9) resulted in the dissociation of whey proteins and κ-casein-rich protein from the micelles and the residual micelles were unstable, without a maximum–minimum in the HCT/pH profile. Preformed whey protein–casein micelle complexes formed by preheating (140 °C for 1 min) milk at pH 6·7 dissociated from the micelles on reheating (140 °C for 1 min) at pH > 6·9. The dissociation of micellar-κ-casein, perhaps complexed with whey proteins, may reduce the micellar zeta potential at pH ≃ 6·9 sufficiently to cause a minimum in the HCT/pH profile of milk.

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Comparison of heat and pressure treatments of skim milk, fortified with whey protein concentrate, for set yogurt preparation: effects on milk proteins and gel structure

Journal of Dairy Research ◽

10.1017/s0022029900004301 ◽

2000 ◽

Vol 67 (3) ◽

pp. 329-348 ◽

Cited By ~ 102

Author(s):

ERIC C. NEEDS ◽

MARTA CAPELLAS ◽

A. PATRICIA BLAND ◽

PRETIMA MANOJ ◽

DOUGLAS MACDOUGAL ◽

...

Keyword(s):

Heat Treatment ◽

Whey Protein ◽

Skim Milk ◽

Milk Proteins ◽

Whey Protein Concentrate ◽

Protein Concentrate ◽

Casein Micelles ◽

Micelle Structure ◽

Dynamic Structures ◽

Β Lactoglobulin

Heat (85 °C for 20 min) and pressure (600 MPa for 15 min) treatments were applied to skim milk fortified by addition of whey protein concentrate. Both treatments caused > 90% denaturation of β-lactoglobulin. During heat treatment this denaturation took place in the presence of intact casein micelles; during pressure treatment it occurred while the micelles were in a highly dissociated state. As a result micelle structure and the distribution of β-lactoglobulin were different in the two milks. Electron microscopy and immunolabelling techniques were used to examine the milks after processing and during their transition to yogurt gels. The disruption of micelles by high pressure caused a significant change in the appearance of the milk which was quantified by measurement of the colour values L*, a* and b*. Heat treatment also affected these characteristics. Casein micelles are dynamic structures, influenced by changes to their environment. This was clearly demonstrated by the transition from the clusters of small irregularly shaped micelle fragments present in cold pressure-treated milk to round, separate and compact micelles formed on warming the milk to 43 °C. The effect of this transition was observed as significant changes in the colour indicators. During yogurt gel formation, further changes in micelle structure, occurring in both pressure and heat-treated samples, resulted in a convergence of colour values. However, the microstructure of the gels and their rheological properties were very different. Pressure-treated milk yogurt had a much higher storage modulus but yielded more readily to large deformation than the heated milk yogurt. These changes in micelle structure during processing and yogurt preparation are discussed in terms of a recently published micelle model.

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High pressure treatment of bovine milk: effects on casein micelles and whey proteins

Journal of Dairy Research ◽

10.1017/s002202990300640x ◽

2004 ◽

Vol 71 (1) ◽

pp. 97-106 ◽

Cited By ~ 142

Author(s):

Thom Huppertz ◽

Patrick F Fox ◽

Alan L Kelly

Keyword(s):

High Pressure ◽

Treatment Time ◽

Whey Proteins ◽

Bovine Milk ◽

Casein Micelle ◽

Pressure Treatment ◽

Casein Micelles ◽

High Pressure Treatment ◽

Micelle Size ◽

Β Lactoglobulin

Effects of high pressure (HP) on average casein micelle size and denaturation of α-lactalbumin (α-la) and β-lactoglobulin (β-lg) in raw skim bovine milk were studied over a range of conditions. Micelle size was not influenced by treatment at pressures <200 MPa, but treatment at 250 MPa increased micelle size by ∼25%, while treatment at [ges ]300 MPa irreversibly reduced it to ∼50% of that in untreated milk. The increase in micelle size after treatment at 250 MPa was greater with increasing treatment time and temperature and milk pH. Treatment times [ges ]2 min at 400 MPa resulted in similar levels of micelle disruption, but increasing milk pH to 7·0 partially stabilised micelles against HP-induced disruption. Denaturation of α-la did not occur [les ]400 MPa, whereas β-lg was denatured at pressures >100 MPa. Denaturation of α-la and β-lg increased with increasing pressure, treatment time and temperature and milk pH. The majority of denatured β-lg was apparently associated with casein micelles. These effects of HP on casein micelles and whey proteins in milk may have significant implications for properties of products made from HP-treated milk.

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Disruption and reassociation of casein micelles during high pressure treatment: influence of whey proteins

Journal of Dairy Research ◽

10.1017/s0022029906002263 ◽

2007 ◽

Vol 74 (2) ◽

pp. 194-197 ◽

Cited By ~ 21

Author(s):

Thom Huppertz ◽

Cornelis G de Kruif

Keyword(s):

High Pressure ◽

Treatment Time ◽

Light Transmission ◽

Whey Proteins ◽

Casein Micelle ◽

Casein Micelles ◽

Β Lactoglobulin ◽

Steric Stabilisation ◽

Prolonged Holding

In the study presented in this article, the influence of added α-lactalbumin and β-lactoglobulin on the changes that occur in casein micelles at 250 and 300 MPa were investigated by in-situ measurement of light transmission. Light transmission of a serum protein-free casein micelle suspension initially increased with increasing treatment time, indicating disruption of micelles, but prolonged holding of micelles at high pressure partially reversed HP-induced increases in light transmission, suggesting reformation of micellar particles of colloidal dimensions. The presence of α-la and/or β-lg did not influence the rate and extent of micellar disruption and the rate and extent of reformation of casein particles. These data indicate that reformation of casein particles during prolonged HP treatment occurs as a result of a solvent-mediated association of the micellar fragments. During the final stages of reformation, κ-casein, with or without denatured whey proteins attached, associates on the surface of the reformed particle to provide steric stabilisation.

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Influence of aggregation on the susceptibility of casein to proteolysis

Journal of Dairy Research ◽

10.1017/s0022029900013200 ◽

1970 ◽

Vol 37 (2) ◽

pp. 173-180 ◽

Cited By ~ 37

Author(s):

P. F. Fox

Keyword(s):

Calcium Phosphate ◽

Equilibrium Dialysis ◽

Limited Proteolysis ◽

Skim Milk ◽

Sodium Caseinate ◽

Casein Micelle ◽

Casein Micelles ◽

Micelle Structure ◽

Colloidal Calcium Phosphate

SummaryThe susceptibility of the casein in milk to proteolysis was shown to be greatly influenced by its state of aggregation. In normal milk, where the casein is largely in micellar form, the αs1- and β-caseins are almost inaccessible to proteolysis. On removal of the colloidal phosphate, the casein micelles disintegrate, rendering the components, especially the αs1-casein, accessible to proteolysis. The role of colloidal calcium phosphate in the casein micelle is believed to be that of a non-specific aggregating agent which can be effectively replaced by calcium. Dissolved colloidal phosphate can be effectively reformed by elevation of the pH of colloidal phosphate-free (CPF) milk before equilibrium dialysis. Addition of κ-casein to CPF milk also causes aggregation of the component caseins but the micelles formed are smaller than those of normal milk.The behaviour of micellar β-casein differs considerably from that of micellar αs1-casein. The evidence suggests that part of the β-casein freely dissociates either outside or within the micelle when the temperature is reduced. The temperature dependence of the susceptibility of β-casein to proteolysis was similar in skim-milk and in solutions of sodium caseinate, and increased as the temperature was reduced. αs1-Casein was quite resistant to proteolysis in normal milk but became susceptible when the micelle structure was disrupted on removal of colloidal phosphate.It is concluded that limited proteolysis may prove a valuable technique in the study of casein micelle structure.

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Electrophoretic mobility of casein micelles

Journal of Dairy Research ◽

10.1017/s0022029900017477 ◽

1979 ◽

Vol 46 (3) ◽

pp. 441-451 ◽

Cited By ~ 29

Author(s):

Donald F. Darling ◽

John Dickson

Keyword(s):

Electrophoretic Mobility ◽

Moving Boundary ◽

Ionic Composition ◽

Skim Milk ◽

Casein Micelle ◽

Casein Micelles ◽

Zeta Potentials ◽

Micelle Structure ◽

Increase With Increase ◽

Micelle Size

SummaryA simplified moving boundary electrophoresis technique has been developed for the measurement of the electrophoretic mobility of casein micelles. The zeta potentials of casein micelles from different skim-milk samples were calculated using Henry's equation and shown to decrease with decrease in pH between pH 6.9 and 5.3 and to increase with increase in temperature between 10 and 45 °C. Neither severe heat treatment (up to 135 °C for 51 min) nor centrifugal fractionation of micelles into different micelle size ranges had any significant effect on zeta potential. The ionic composition of the serum phase has been shown to be extremely important in determining the electrophoretic mobility. Casein micelles electrophoresed through milk ultrafiltrate consistently gave a lower mobilities than the same micelles centrifuged through milk centrifugate. The results are discussed in relation to present theories of casein micelle structure; these theories do not accommodate all of the observations.

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Association of denatured whey proteins with casein micelles in heated reconstituted skim milk and its effect on casein micelle size

Journal of Dairy Research ◽

10.1017/s0022029902005903 ◽

2003 ◽

Vol 70 (1) ◽

pp. 73-83 ◽

Cited By ~ 194

Author(s):

Skelte G Anema ◽

Yuming Li

Keyword(s):

Heat Treatment ◽

Particle Size ◽

Whey Protein ◽

Volume Fraction ◽

Whey Proteins ◽

Skim Milk ◽

Casein Micelle ◽

Casein Micelles ◽

Prolonged Heating ◽

Micelle Size

When skim milk at pH 6·55 was heated (75 to 100 °C for up to 60 min), the casein micelle size, as monitored by photon correlation spectroscopy, was found to increase during the initial stages of heating and tended to plateau on prolonged heating. At any particular temperature, the casein micelle size increased with longer holding times, and, at any particular holding time, the casein micelle size increased with increasing temperature. The maximum increase in casein micelle size was about 30–35 nm. The changes in casein micelle size were poorly correlated with the level of whey protein denaturation. However, the changes in casein micelle size were highly correlated with the levels of denatured whey proteins that were associated with the casein micelles. The rate of association of the denatured whey proteins with the casein micelles was considerably slower than the rate of denaturation of the whey proteins. Removal of the whey proteins from the skim milk resulted in only small changes in casein micelle size during heating. Re-addition of β-lactoglobulin to the whey-protein-depleted milk caused the casein micelle size to increase markedly on heat treatment. The changes in casein micelle size induced by the heat treatment of skim milk may be a consequence of the whey proteins associating with the casein micelles. However, these associated whey proteins would need to occlude a large amount of serum to account for the particle size changes. Separate experiments showed that the viscosity changes of heated milk and the estimated volume fraction changes were consistent with the particle size changes observed. Further studies are needed to determine whether the changes in size are due to the specific association of whey proteins with the micelles or whether a low level of aggregation of the casein micelles accompanies this association behaviour. Preliminary studies indicated lower levels of denatured whey proteins associated with the casein micelles and smaller changes in casein micelle size occurred as the pH of the milk was increased from pH 6·5 to pH 6·7.

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Effect of high hydrostatic pressure and whey proteins on the disruption of casein micelle isolates

Journal of Dairy Research ◽

10.1017/s0022029907002762 ◽

2007 ◽

Vol 74 (4) ◽

pp. 452-458 ◽

Cited By ~ 8

Author(s):

Federico M Harte ◽

Subba Rao Gurram ◽

Lloyd O Luedecke ◽

Barry G Swanson ◽

Gustavo V Barbosa-Cánovas

Keyword(s):

Hydrostatic Pressure ◽

Whey Protein ◽

High Hydrostatic Pressure ◽

Analytical Ultracentrifugation ◽

Whey Proteins ◽

Skim Milk ◽

Casein Micelle ◽

Casein Micelles ◽

Transmission Electron ◽

Thermally Processed

High hydrostatic pressure disruption of casein micelle isolates was studied by analytical ultracentrifugation and transmission electron microscopy. Casein micelles were isolated from skim milk and subjected to combinations of thermal treatment (85°C, 20 min) and high hydrostatic pressure (up to 676 MPa) with and without whey protein added. High hydrostatic pressure promoted extensive disruption of the casein micelles in the 250 to 310 MPa pressure range. At pressures greater than 310 MPa no further disruption was observed. The addition of whey protein to casein micelle isolates protected the micelles from high hydrostatic pressure induced disruption only when the mix was thermally processed before pressure treatment. The more whey protein was added (up to 5 g/l) the more the protection against high hydrostatic pressure induced micelle disruption was observed in thermally treated samples subjected to 310 MPa.

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CPG-chromatography studies on the stability of casein micelles

Journal of Dairy Research ◽

10.1017/s0022029900017222 ◽

1979 ◽

Vol 46 (2) ◽

pp. 313-316 ◽

Cited By ~ 6

Author(s):

Märtha Larsson-Raźnikiewicz ◽

Elisabeth Almlöf ◽

Bo Ekstrand

Keyword(s):

Freeze Drying ◽

Whey Proteins ◽

Skim Milk ◽

Casein Micelles ◽

Controlled Pore Glass ◽

Milk Serum ◽

Colloidal Calcium Phosphate ◽

Pore Glass ◽

The Stability ◽

Β Lactoglobulin

SUMMARYCasein micelles fractionated on controlled pore glass (CPG-10/3000) were shown to be stable by recycling experiments. Only minor effects on the size distribution of the casein micelles were found after heating skim-milk to 100 °C for 10 min, or freeze-drying skim-milk at – 70 °C followed by resuspension in the synthetic milk serum of Jenness & Koops (1962). The heating caused some whey proteins (β-lactoglobulin) to enter the micelle fractions while the freeze-drying caused some of the largest micelles to disrupt. In colloidal calcium phosphate-free skim-milk prepared according to Pyne & McGann (1960) all the micelles appeared to dissociate into monomeric caseins.

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