Casein is the main protein found in milk. It makes up about 80% of the proteins in cow’s milk. When milk is heated, casein proteins undergo changes in their structure and interactions with other milk components. Understanding these changes is important for manufacturing dairy products like cheese and yogurt.
Casein in Milk
Fresh milk contains casein proteins that are suspended in a colloidal dispersion. The casein proteins exist in a micelle structure, which is a spherical aggregate of many individual casein proteins along with calcium and phosphate ions. These casein micelles are stabilized by hydrophobic interactions and calcium phosphate binding.
There are several different types of casein proteins in milk including αs1-casein, αs2-casein, β-casein and κ-casein. Each has a distinct structure and plays a role in micelle formation and stabilization. κ-casein is particularly important and sits on the outer surface of the micelle structure.
Effects of Heating Milk
When milk is heated to temperatures above 70°C, the casein proteins begin to destabilize and the micelle structure starts to break down. This happens because of changes to the hydrophobic and hydrophilic interactions that hold the micelles together.
One major change is that the whey proteins, especially β-lactoglobulin, start to denature and interact with the κ-casein on the micelle surface. The newly exposed regions then cross-link with other casein proteins via disulfide bonds and cause aggregation.
In addition, colloidal calcium phosphate dissociates from the casein micelles as the milk is heated. This also leads to destabilization of the micelle structure. The changes to the casein interactions differ based on the heating temperature and duration.
Effects at Moderate Temperatures (70-100°C)
When milk is heated to temperatures between 70-100°C, the main effect is denaturation of the whey proteins and their interaction with the casein micelles. The micelles start to aggregate together into larger clumps rather than remaining as individual spherical structures.
The casein micelles maintain much of their structure during this moderate heat treatment. There are only minor changes to the internal structure and integrity of the individual micelles. Primarily, the outer κ-casein layer is affected by interactions with denatured whey proteins.
Effects at High Temperatures (>100°C)
At higher temperatures above 100°C, more extensive breakdown of the casein micelle structures occurs. The individual micelles start to lose their internal structure and the casein proteins within them begin to interact.
The hydrophobic and electrostatic interactions holding the casein proteins together in the micelles are disrupted. The individual casein proteins then associate with caseins from other micelles as well as interacting with whey proteins.
This leads to a matrix of casein aggregates linked together with whey proteins. Extensive cross-linking and coagulation takes place, leading to the formation of a more continuous protein network rather than discrete casein micelles.
Effects on Dairy Product Properties
The heat-induced changes to casein micelle structure and protein interactions have significant effects on the properties of milk and dairy products.
Cheese Making
In cheese making, milk is heated to temperatures above 70°C to cause controlled casein aggregation. This facilitates the separation of curds and whey during cheese manufacturing. The curds consist of the aggregated casein proteins.
Different cheese varieties rely on different casein interactions during heating. For example, mozzarella cheese production involves moderate heat treatment at 80-90°C to maintain some micelle structure. In contrast, cheddar cheese is made by extensive casein breakdown and aggregation induced by heating to >100°C.
Yogurt Manufacture
Milk heating is also important in yogurt production. Milk solids are concentrated by heating above 100°C to promote whey protein denaturation and complex formation with caseins. The concentrated micellar casein is then cooled and inoculated with yogurt cultures.
The heat treatment of milk is standardized to ensure proper fermentation and texture characteristics in the final yogurt product. Insufficient heat treatment can lead to poor yogurt gel strength and whey separation.
Milk Processing
For general milk processing, lower heat treatments are used to maintain casein micelle integrity as much as possible. High heat treatments can lead to considerable casein aggregation and gritty, depositing texture defects in processed milk.
Most pasteurization methods heat milk to 72-75°C for 15-20 seconds in order to eliminate pathogens while minimizing casein denaturation. Higher temperatures or longer holding times will cause more casein changes.
Effects on Nutrition
The impacts of heating on casein structure can also influence the nutritional properties of milk. Heat denaturation decreases the solubility of calcium and can make it less bioavailable. Moderate heat of pasteurization has minimal effects on solubility.
Heating also increases the amount of casein and whey proteins that are digestible by proteolytic enzymes. However, excessive temperatures can negatively impact protein quality by causing aggregation via sites other than those normally cleaved by proteases.
Kinetics of Heat-Induced Changes
The effects of heating on casein occur gradually over time depending on the temperature. Researchers have studied the kinetics of the casein changes induced by heating using techniques like particle size analysis and protein electrophoresis.
In general, the aggregation process begins slowly and accelerates over time. Micellar structural changes occur early at lower temperatures while extensive aggregation occurs faster at high temperatures.
One study heating milk at 140°C found that aggregation accelerated after 8 minutes as measured by particle size. Another paper found the rate of casein decrease measured by electrophoresis increased exponentially with heating time at 120°C.
The kinetics also depend on other factors like pH and presence of calcium and whey proteins. Understanding the time-dependent nature of casein changes during heating has helped refine thermal processes for optimal dairy product qualities.
Reversibility of Heat Effects
An important consideration is whether the heat-induced changes to casein are reversible or permanent. Reversibility depends on the extent of protein denaturation and aggregation that occurs.
At lower temperatures from 70-90°C, many of the interactions between caseins and whey proteins are reversible if the milk is cooled. This enables retention of casein micelle structure.
However, when milk is heated to higher temperatures above 100°C, the casein-whey protein complexes and casein aggregation become irreversible upon cooling. The original casein micelle structure cannot be regained.
Heating milk above 140°C leads to almost total irreversible denaturation and coagulation of caseins. Cooling will not dissociate the aggregates back into discrete casein micelles after such severe heat treatment.
Effects of Other Factors
Several other factors can influence the heat-induced changes to casein in milk besides just temperature and time.
pH
The pH of milk impacts the magnitude of casein denaturation and interaction with whey proteins during heating. A lower pH results in greater casein aggregation. For example, at 140°C, aggregation was four-fold higher in milk at pH 6.7 than 7.2.
The pH affects the ionization state and charge of amino acid side chains that are involved in casein-casein and casein-whey protein interactions during heating.
Calcium
Calcium also plays a key role in controlling casein changes during heating. Milk with higher calcium was found to have reduced casein-whey protein complex formation when heated to 90°C.
Calcium promotes micellar stability and acts as a “bridge” between proteins through electrostatic interactions. Chelating calcium ions resulted in almost 10 times greater casein aggregation upon heating to 140°C.
Whey Proteins
As described earlier, whey proteins interact extensively with caseins during heating and are integral to casein denaturation and aggregation.
When whey proteins are removed from milk prior to heating, the casein micelles remain relatively stable even at high temperatures above 100°C. Without whey protein interactions, the casein micelles experience minimal internal disruption.
Non-Thermal Effects on Casein
In addition to thermal effects, casein proteins can also be altered by other non-thermal treatments of milk.
High Pressure Processing
High pressure processing involves applying pressures from 100-800 MPa to milk. This can induce changes to casein micelles similar to heating such as dissociation of micellar calcium phosphate and casein aggregation.
However, the effects are reversible when pressure is decreased to ambient conditions. So high pressure is sometimes used for microbial inactivation while minimizing permanent impacts on casein.
Ultrasound
Ultrasonication of milk can break up casein micelles into smaller substructures. Power ultrasound at 20 kHz leads to cavitation and shear forces that physically disrupt the micelles.
This effect can be useful for reassembling casein micelles with altered functionality. However, excessive ultrasonication can cause irreversible aggregation through hydrophobic interactions between exposed regions of casein.
Other Technologies
Non-thermal technologies like pulsed electric fields, high voltage arcs, irradiation and UV treatment can also influence casein stability in milk. The magnitude of effects depends on the processing parameters used.
In general, these non-thermal treatments do not induce protein denaturation and aggregation to the same extent as heating milk to high temperatures.
Intentional Modification of Casein
In some dairy products, controlled disruption or cross-linking of caseins is intentionally induced to create desirable textures and functionalities.
Disruption of Micelles
Micellar casein can be decoupled into individual casein proteins using high shear or enzymatic hydrolysis. This can improve foaming and emulsifying properties.
The newly exposed regions of the caseins have different surface properties than the original micelles. This approach can modulate texture in Greek-style yogurts and imitation cheeses.
Cross-Linking Casein Proteins
Cross-linking enzymes like transglutaminase can be added to milk or pre-formed dairy products to induce protein polymerization. This creates a fine-stranded gel network that provides thickness, creaminess and heat stability.
Cross-linking improves the texture and mouthfeel of products like yogurt, ice cream and cream cheese where a smooth, thick consistency is desired.
Conclusion
Heating milk causes significant changes to the casein proteins including denaturation, aggregation with whey proteins, and disruption of their colloidal micelle structure. The extent of these changes depends on the heating temperature and duration as well as milk composition and conditions like pH.
At low to moderate temperatures (70-100°C), the caseins remain somewhat intact but associate with whey proteins. More severe heat treatment above 100°C leads to irreversible breakdown of casein micelles and extensive aggregation.
These heating-induced changes can be beneficial for manufacturing dairy products like cheese but need to be minimized for other applications like fluid milk pasteurization. Non-thermal processing methods can also modify casein structure and functionality in different ways.
Overall, understanding how casein proteins are impacted by heating and other treatments enables dairy technologists to design optimal processes for a wide range of products, applications, and desired functionalities.
