Structural changes induced by high-pressure processing in micellar casein and milk protein concentrates

Reconstituted micellar casein concentrates and milk protein concentrates of 2.5 and 10% (wt/vol) protein concentration were subjected to high-pressure processing at pressures from 150 to 450 MPa, for 15 min, at ambient temperature. The structural changes induced in milk proteins by high-pressure processing were investigated using a range of physical, physicochemical, and chemical methods, including dynamic light scattering, rheology, mid-infrared spectroscopy, scanning electron microscopy, proteomics, and soluble mineral analyses. The experimental data clearly indicate pressure-induced changes of casein micelles, as well as denaturation of serum proteins. Calcium-binding α_S1- and α_S2-casein levels increased in the soluble phase after all pressure treatments. Pressurization up to 350 MPa also increased levels of soluble calcium and phosphorus, in all samples and concentrations, whereas treatment at 450 MPa reduced the levels of soluble Ca and P. Experimental data suggest dissociation of calcium phosphate and subsequent casein micelle destabilization as a result of pressure treatment. Treatment of 10% micellar casein concentrate and 10% milk protein concentrate samples at 450 MPa resulted in weak, physical gels, which featured aggregates of uniformly distributed, casein substructures of 15 to 20 nm in diameter. Serum proteins were significantly denatured by pressures above 250 MPa. These results provide information on pressure-induced changes in high-concentration protein systems, and may inform the development on new milk protein-based foods with novel textures and potentially high nutritional quality, of particular interest being the soft gel structures formed at high pressure levels.     

Use of micellar casein concentrate and milk protein concentrate treated with transglutaminase in imitation cheese products—Unmelted texture

The amount of intact casein provided by dairy ingredients is a critical parameter in dairy-based imitation mozzarella cheese (IMC) formulation because it has a significant effect on unmelted textural parameters such as hardness. From a functionality perspective, rennet casein (RCN) is the preferred ingredient. Milk protein concentrate (MPC) and micellar casein concentrate (MCC) cannot provide the required functionality due to the higher steric stability of casein micelle. However, the use of transglutaminase (TGase) has the potential to modify the surface properties of MPC and MCC and may improve their functionality in IMC. The objective of this study was to determine the effect of TGase-treated MPC and MCC powders on the unmelted textural properties of IMC and compare them with IMC made using commercially available RCN. Additionally, we studied the degree of crosslinking by TGase in MPC and MCC retentates using capillary gel electrophoresis. Three lots of MCC and MPC retentate were produced from pasteurized skim milk via microfiltration and ultrafiltration, respectively, and randomly assigned to 1 of 3 treatments: no TGase (control); low TGase: 0.3 units/g of protein; and high TGase: 3.0 units/g of protein, followed by inactivation of enzyme (72°C for 10 min), and spray drying. Each MCC, MPC, and RCN was then used to formulate IMC that was standardized to 21% fat, 1% salt, 48% moisture, and 20% protein. The IMC were manufactured by blending, mixing, and heating ingredients (4.0 kg) in a twin-screw cooker. The capillary gel electrophoresis analysis showed extensive inter- and intramolecular crosslinking. The IMC formulation using the highest TGase level in MCC or MPC did not form an emulsion because of extensive crosslinking. In MPC with a high level of TGase, whey protein and casein crosslinking were observed. In contrast, crosslinking and hydrolysis of proteins were observed in MCC. The IMC made from MCC powder had significantly higher texture profile analysis hardness compared with the corresponding MPC powder. Further, many-to-one (multiple) comparisons using the Dunnett test showed no significant differences between IMC made using RCN and treatment powders in hardness. Our results demonstrated that TGase treatment causes crosslinking hydrolysis of MCC and MPC at higher TGase levels, and MPC and MCC have the potential to be used as ingredients in IMC applications.     

Invited review: Microfiltration-derived casein and whey proteins from milk

Milk, a rich source of nutrients, can be fractionated into a wide range of components for use in foods and beverages. With advancements in filtration technologies, micellar caseins and milk-derived whey proteins are now produced from skim milk using microfiltration. Microfiltered ingredients offer unique functional and nutritional benefits that can be exploited in new product development. Microfiltration offers promise in cheesemaking, where microfiltered milk can be used for protein standardization to improve the yield and consistency of cheese and help with operation throughputs. Micellar casein concentrates and milk whey proteins could offer unique functional and flavor properties in various food applications. Consumer desires for safe, nutritious, and clean-label foods could be potential growth opportunities for these new ingredients. The application of micellar casein concentrates in protein standardization could offer a window of opportunity to US cheese makers by improving yields and throughputs in manufacturing plants.     

Processing factors that influence casein and serum protein separation by microfiltration

Our objective was to demonstrate the effect of various processing factors on the performance of a microfiltration system designed to process skim milk and separate casein (CN) from serum proteins (SP). A mathematical model of a skim milk microfiltration process was developed with 3 stages plus an additional fourth finishing stage to standardize the retentate to 9% true protein (TP) and allow calculation of yield of a liquid 9% TP micellar CN concentrate (MCC) and milk SP isolate (MSPI; 90% SP on a dry basis). The model was used to predict the effect of 5 factors: 1) skim milk composition, 2) heat treatment of skim milk, 3) concentration factor (CF) and diafiltration factor (DF), 4) control of CF and DF, and 5) SP rejection by the membrane on retentate and permeate composition, SP removal, and MCC and MSPI yield. When skim milk TP concentration increased from 3.2 to 3.8%, the TP concentration in the third stage retentate increased from 7.92 to 9.40%, the yield of MCC from 1,000 kg of skim milk increased from 293 to 348 kg, and the yield of MSPI increased from 6.24 to 7.38 kg. Increased heat treatment (72.9 to 85.2°C) of skim milk caused the apparent CN as a percentage of TP content of skim milk as measured by Kjeldahl analysis to increase from 81.97 to 85.94% and the yield of MSPI decreased from 6.24 to 4.86 kg, whereas the third stage cumulative percentage SP removal decreased from 96.96 to 70.08%. A CF and DF of 2× gave a third stage retentate TP concentration of 5.38% compared with 13.13% for a CF and DF of 5×, with the third stage cumulative SP removal increasing from 88.66 to 99.47%. Variation in control of the balance between CF and DF (instead of an equal CF and DF) caused either a progressive increase or decrease in TP concentration in the retentate across stages depending on whether CF was greater than DF (increasing TP in retentate) or CF was less than DF (decreasing TP in retentate). An increased rejection of SP by the membrane from an SP removal factor of 1 to 0.6 caused a reduction in MSPI yield from 6.24 to 5.19 kg/1,000 kg of skim milk, and third stage cumulative SP removal decreased from 96.96 to 79.74%. Within the ranges of the 5 factors studied, the TP content of the third stage retentate was most strongly affected by the target CF and DF and variation in skim milk composition. Cumulative percentage SP removal was most strongly affected by the heat treatment of skim milk, the SP removal factor, and the target CF and DF. The MCC yield was most strongly affected by initial skim milk composition. Yield of MSPI was strongly affected by skim milk composition, whereas the heat treatment of milk and SP removal factor also had a large effect.     

浓缩酪蛋白胶束成分的影响因素及其在奶酪生产中的应用（网络首发、推荐阅读）

目前,采用膜过滤技术可从脱脂奶中分离酪蛋白,随后通过浓缩、杀菌、干燥等工艺获得浓缩酪蛋白胶束。对浓缩酪蛋白胶束成分的影响因素及其在奶酪生产中的应用进行综述,发现膜过滤期间的pH值、温度和洗滤条件均会影响浓缩酪蛋白胶束的成分,使其具有不同浓度的酪蛋白、乳清蛋白、乳糖以及钙。而且可以利用浓缩酪蛋白胶束标准化原奶,从而制备成分和品质一致的奶酪;也可以利用不同成分的浓缩酪蛋白胶束获得不同的原奶组合物,从而制备所需品质和功能的奶酪。总之,在奶酪生产过程中添加浓缩酪蛋白胶束能够影响奶酪的成分、质地以及风味等,但通过调整膜过滤和奶酪生产的工艺参数可以解决这些问题。未来还需获得一种经济有效的方式来保存浓缩酪蛋白胶束,赋予其更长的保质期,良好的凝乳酶凝乳特性,从而保证奶酪的品质和产量。     

Use of micellar casein concentrate and milk protein concentrate treated with transglutaminase in imitation cheese products—Melt and stretch properties

Melt and stretch properties in dairy-based imitation mozzarella cheese (IMC) are affected by the amount of intact casein provided by dairy ingredients in the formulation. Rennet casein (RCN) is the preferred ingredient to provide intact casein in a formulation. Ingredients produced using membrane technology, such as milk protein concentrate (MPC) and micellar casein concentrate (MCC), are unable to provide the required functionality. However, the use of transglutaminase (TGase) has potential to modify the physical properties of MPC or MCC and may improve their functionality in IMC. The objective of this study was to determine the effect of TGase-treated MPC and MCC retentates on melt and stretch properties when they are used in IMC and to compare them with IMC made using RCN. The MCC and MPC retentates were produced using 3 different lots of pasteurized skim milk and treated with 3 levels of TGase enzyme: no TGase (control), low TGase: 0.3 units/g of protein, and high TGase: 3.0 units/g of protein. Each of the MCC and MPC treatments was heated to 72°C for 10 min to inactivate TGase and then spray dried. Each MCC, MPC, and RCN powder was then used in an IMC formulation that was standardized to 48% moisture, 21% fat, 20% protein, and 1% salt. The IMC were manufactured in a twin-screw cooker by blending, mixing, and heating various ingredients (4.0 kg). Due to extensive crosslinking, the IMC formulation with the highest TGase level (MCC or MPC) did not form an emulsion. The IMC made from MCC treatments had significantly higher stretchability on pizza compared with their respective MPC treatments. The IMC made from TGase-treated MCC and MPC had significantly lower melt area and significantly higher transition temperature (TT) and stretchability compared with their respective controls. Comparison of IMC made using TGase-treated MCC and MPC to the RCN IMC indicated no difference in TT or texture profile analysis-stretchability; however, the Schreiber melt test area was significantly lower. Our results demonstrated that TGase treatment modifies the melt and stretch characteristics of MCC and MPC in IMC applications, and TGase-treated MPC and MCC can be used to replace RCN in IMC formulations.     

Use of micellar casein concentrate for Greek-style yogurt manufacturing: Effects on processing and product properties

The objective of this work was to develop and optimize an alternative make process for Greek-style yogurt (GSY), in which the desired level of protein was reached by fortification with micellar casein concentrate (MCC) obtained from milk by microfiltration. Two MCC preparations with 58 and 88% total protein (MCC-58 and MCC-88) were used to fortify yogurt milk to 9.80% (wt/wt) protein. Strained GSY of similar protein content was used as the control. Yogurt milk bases were inoculated with 0.02% (wt/wt) or 0.04% (wt/wt) direct vat set starter culture and fermented until pH 4.5. The acidification rate was faster for the MCC-fortified GSY than for the control, regardless of the inoculation level, which was attributed to the higher nonprotein nitrogen content in the MCC-fortified milk. Steady shear rate rheological analysis indicated a shear-thinning behavior for all GSY samples, which fitted well with the power law model. Dynamic rheological analysis at 5°C showed a weak frequency dependency of the elastic modulus (G′) and viscous modulus (G″) for all GSY samples, with G′ > G″, indicating a weak gel structure. Differences in the magnitude of viscoelastic parameters between the 2 types of GSY were found, with G′ of MCC-fortified GSY < G′ of control, indicating a different extent ofprotein interactionsin the 2 types of yogurt. Differences were also noticed in water-holding capacity, which was lower for the MCC-fortified GSY compared with the control, attributed to lower serum protein content in the former. Despite some differences in the physicochemical characteristics of the final product compared with GSY manufactured by straining, the alternative process developed here is a feasible alternative to the traditional GSY make process, with environmental and possibly financial benefits to the dairy industry.     

Protein,casein, and micellar salts in milk: Current content and historical perspectives

The protein and fat content of Dutch bulk milk has been monitored since the 1950s and has increased considerably, by 11 and 20%, respectively, whereas milk yield has more than doubled. The change in protein and fat content of milk is advantageous for the dairy industry, as these are the 2 most economically valuable constituents of milk. Increases in protein and fat content of milk have allowed increases in the yield of various products such as cheese and butter. However, for cheese and other applications where casein micelles play a crucial role in structure and stability, it is not only casein content, but also the properties of the casein micelles that determine processability. Of particular importance herein is the salt partition in milk, but it is unknown whether increased protein content has affected the milk salts and their distribution between casein micelles and milk serum. It was, therefore, the objective of this research to determine the salt composition and protein content for individual cow milk and bulk milk over a period of 1 yr and to compare these data to results obtained during the 1930s, 1950s, and 1960s in the last century. Calcium, magnesium, sodium, potassium, and phosphorus content were determined by inductively coupled plasma atomic emission spectrometry and inorganic phosphate, citrate, chloride, and sulfate content by anion-exchange chromatography in bulk milk and milk ultracentrifugate. In addition, ionic calcium and ionic magnesium concentration were determined by the Donnan membrane technique. We concluded that historical increase in milk yield and protein content in milk have resulted in correlated changes in casein content and the micellar salt fraction of milk. In addition, the essential nutrients, calcium, magnesium, and phosphorus in milk have increased the past 75 yr; therefore, the nutritional value of milk has improved.     

Rheological properties of rennet gels containing milk protein concentrates

Different milk protein concentrates (MPC), with protein concentrations of 56, 70, and 90%, were dispersed in water under different treatments (hydration, shear, heat, and overnight storage at 4°C), as well as in a combination of all the treatments in a factorial design. The particle size distribution of the dispersions was then measured to determine the optimal conditions for the dispersion. Heating at 60°C for 30 min with 5 min of shear was chosen as the best condition to dissolve MPC powders. The samples were also characterized for composition, presence of protein aggregates, and ratio of calcium to protein. The total calcium present in MPC increased with increasing concentration of protein; however, the total calcium-to-protein ratio was lower in MPC90 than in MPC56 and MPC70. The level of whey protein denaturation, the presence of κ-casein-whey protein aggregates in the supernatant after centrifugation, and the amount of caseins dissociated from the micelle increased as the protein concentration in the powder increased. The total amount of casein macropeptide released was lower in samples from powders with a higher protein concentration than for MPC56 or the skim milk control. The gelation behavior of reconstituted MPC was tested in systems dispersed in water (5% protein) as well as in systems dispersed in skim milk (6% protein). The gelation time of MPC dispersions was considerably lower and the gel modulus was higher than those of reconstituted skim milk with the same protein concentration. When MPC dispersions were dialyzed against skim milk, a significant decrease in the gelation time and modulus were shown, with a complete loss of gelling functionality in MPC90 dispersed in water. This demonstrated that the ionic equilibrium was key to the functionality of MPC.     

Stability of casein micelles cross-linked by transglutaminase

In this study, caseins micelles were internally cross-linked using the enzyme transglutaminase (TGase). The integrity of the micelles was examined on solubilization of micellar calcium phosphate (MCP) or on disruption of hydrophobic interactions and breakage of hydrogen bonds. The level of monomeric caseins, determined electrophoretically, decreased with increasing time of incubation with TGase at 30°C; after incubation for 24 h, no monomeric β- or κ-caseins were detected, whereas only a small level of monomeric α_S1-casein remained, suggesting near complete intramicellar cross-linking. The ability of casein micelles to maintain structural integrity on disruption of hydrophobic interactions (using urea, sodium dodecyl sulfate, or heating in the presence of ethanol), solubilization of MCP (using the calcium-chelating agent trisodium citrate) or high-pressure treatment was estimated by measurement of the L*-value of milk; i.e., the amount of back-scattered light. The amount of light scattered by casein micelles in noncross-linked milk was reduced by >95% on complete disruption of hydrophobic interactions or complete solubilization of MCP; treatment of milk with TGase increased the stability of casein micelles against disruption by all methods studied and stability increased progressively with incubation time. After 24 h of cross-linking, reductions in the extent of light scattering were still apparent in the presence of high levels of dissociating agents, possibly through citrate-induced removal of MCP nanoclusters from the micelles, or urea- or sodium dodecyl sulfate-induced increases in solvent refractive index, which reduce the extent of light-scattering.     

超声处理对酪蛋白胶束粉加工特性的影响

研究超声预处理对酪蛋白胶束粉(micellar casein concentrate,MCC)加工特性的影响。通过微滤生产的酪蛋白截留液经超声处理(超声功率600 W,超声时间0.0、0.5、1.0、2.0、5.0 min)后进行喷雾干燥,得到超声改性的MCC,并进行了加工特性的研究。结果表明,随着超声时间的延长,MCC溶液的电导率、溶解性、乳化性、凝胶性显著增加(P0.05),pH值变化不显著(P0.05)。超声对MCC粉体性质也有影响,随着超声时间的延长,MCC压缩性、流动指数明显减小(P0.05);基本流动能在超声处理0.5 min达到最小值;特殊流动能在超声1 min达到最大值。超声处理可以改善MCC的加工特性,将会促进MCC在食品工业中的应用。     

Invited review: Understanding the behavior of caseins in milk concentrates

The colloidal properties of the casein micelles play a major role in the structural properties of milk protein concentrates. Because of their great technological importance, the structural-functional relationships of casein micelles have been studied for decades in skim milk; however, novel ingredients are now available with higher protein concentrations and varying in composition. The colloidal behavior of caseins in these systems is not fully understood. Concentrates prepared with membrane technologies, and subjected to pre- or post-modifications that affect their technological functionality, have become increasingly widespread. This has created large opportunities for innovation and generation of value-added ingredients. The manner in which caseins interact with themselves and the other components in these concentrates will affect the structure of the final matrix. During concentration by filtration, the interparticle distance between the micelles decreases considerably, increasing their spatial correlation and decreasing their diffusivity. Rearrangements occur due to changes in environmental conditions, such as ionic composition, osmotic stress, shear, pH, or heating temperature. This will have important consequences on bulk viscosity of the concentrates, as well as on the mode of formation of structures' building blocks. This paper aims at highlighting some of the important factors affecting the colloidal structure of casein micelles, their destabilization and network formation, namely, processing history, volume fraction, composition of the serum phase, and ionic equilibrium. Understanding these factors will lead to a better quality control of dairy ingredients and to the development of a new generation of ingredients with targeted functionality.     

Manufacture of process cheese products without emulsifying salts using acid curd and micellar casein concentrate

Process cheese products (PCP) are dairy foods prepared by blending dairy ingredients (such as natural cheese, protein concentrates, butter, nonfat dry milk, whey powder, and permeate) with nondairy ingredients [such as sodium chloride, water, emulsifying salts (ES), color, and flavors] and then heating the mixture to obtain a homogeneous product with an extended shelf life. The ES, such as sodium citrate and disodium phosphate, are critical for the unique microstructure and functional properties of PCP because they improve the emulsification characteristics of casein by displacing the calcium phosphate complexes that are present in the insoluble calcium-paracaseinate-phosphate network in natural cheese. The objectives of this study were to determine the optimum protein content (3, 6, and 9% protein) in micellar casein concentrate (MCC) to produce acid curd and to manufacture PCP using a combination of acid curd cheese and MCC that would provide the desired improvement in the emulsification capacity of caseins without the use of ES. To produce acid curd, MCC was acidified using lactic acid to get a pH of 4.6. In the experimental formulation, the acid curd was blended with MCC to have a 2:1 ratio of protein from acid curd relative to MCC. The PCP was manufactured by blending all ingredients in a KitchenAid blender (Professional 5 Plus, KitchenAid) to produce a homogeneous paste. A 25-g sample of the paste was cooked in the rapid visco analyzer (RVA) for 3 min at 95°C at 1,000 rpm stirring speed during the first 2 min and 160 rpm for the last min. The cooked PCP was then transferred into molds and refrigerated until further analysis. This trial was repeated 3 times using different batches of acid curd. MCC with 9% protein resulted in acid curd with more adjusted yield. The end apparent viscosity (402.0–483.0 cP), hardness (354.0–384.0 g), melting temperature (48.0–51.0°C), and melting diameter (30.0–31.4 mm) of PCP made from different acid curds were slightly different from the characteristics of typical PCP produced with conventional ingredients and ES (576.6 cP end apparent viscosity, 119.0 g hardness, 59.8°C melting temperature, and 41.2 mm melting diameter) due to the differences in pH of final PCP (5.8 in ES PCP compared with 5.4 in no ES PCP). We concluded that acid curd can be produced from MCC with different protein content. Also, we found that PCP can be made with no ES when the formulation uses a 2:1 ratio of acid curd relative to MCC (on a protein basis).     

Technical note: Fourier transform infrared spectral analysis in tandem with 31P nuclear magnetic resonance spectroscopy elaborates detailed insights into phosphate partitioning during skimmed milk microfiltration and diafiltration

Our previous study identified peaks in the ³¹P nuclear magnetic resonance (³¹P NMR) spectra of skim milk, denoting the interaction of different phosphate species such as inorganic and casein-associated phosphate during the separation of colloidal and serum phases of skim milk by microfiltration (MF) and diafiltration (DF). In the current study, we investigated the same samples generated by the aforementioned separation using attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopy analysis. The results confirmed that the technique was not only capable of differentiating between the mineral equilibrium of the casein phosphate nanocluster (CPN) and milk serum, but also complemented the application of ³¹P NMR. An ATR-FTIR broad band in the region of 1,055 to 1,036 cm^?1 and a specific band at 1,076 cm^?1 were identified as sensitive to the repartitioning of different phosphate species in milk in accordance with the ³¹P NMR signals representing casein-associated phosphate and inorganic phosphate in the serum. A third ATR-FTIR signal at 1,034 cm^?1 in milk, representing precipitated inorganic calcium phosphate, had not previously been detected by ³¹P NMR. Thus, the results indicate that a combination of ATR-FTIR and ³¹P NMR spectroscopies may be optimally used to follow mineral and protein phase changes in milk during membrane processing.     

牛羊乳蛋白组分比较研究

羊乳被国际营养学界誉为"奶中之王",逐渐被人们列为日常生活的营养保健佳品。该文对羊乳和牛乳的蛋白组分(主要是酪蛋白和乳清蛋白)含量、氨基酸组成及变异体等方面的差异进行了综述,并且对两者酪蛋白胶束的差异进行了比较,为羊乳检验和加工提供理论依据。