Improving rennet coagulation and cheesemaking properties of reverse osmosis skim milk concentrates by pH adjustment

The use of reverse osmosis (RO) for cheese milk concentration has advantages including obtaining reusable low pollutant permeates and reducing milk transportation costs. However, high levels of lactose and salts in RO concentrates impair their cheesemaking abilities. The objective of this work was to optimise the use of RO concentrates (5–11% protein content) for cheesemaking by pH adjustment. Rennet coagulation kinetics, salt partitioning and cheesemaking properties were studied in comparison with ultrafiltration concentrates. Results showed that concentration by RO induced an increase regarding the coagulation time and the gel maximal firming rate that reached a plateau at 9% protein content. Increases in calcium mineralisation of casein micelles as well as in yield, moisture and lactose content in model cheese were observed. Lowering renneting pH was found to improve the cheesemaking properties of RO concentrates by promoting partial demineralisation of casein micelles, accelerating coagulation kinetics and increasing curd drainage.     

Influence of denaturation and aggregation of β-lactoglobulin on rennet coagulation properties of skim milk and ultrafiltered milk

Skim milk and ultrafiltered milk were heated at temperatures in the range 80–140 °C for 4s in a UHT plate heat exchanger. The extent of denaturation of β-lactoglobulin and its association with the casein micelles increased with increase in heating temperature; the extent of association was markedly less than the amount that denatured. There was no noticeable difference in the extent of denaturation and association of β-lactoglobulin between skim milk and the corresponding ultrafiltered milk. The coagulation properties of renneted heated milks were measured by dynamic oscillation rheometry. Gelation times and storage modulus (G′) could be related to the extent of denaturation of β-lactoglobulin and its association with the casein micelles. Denaturation of β-lactoglobulin up to 60% had no effect on the gelation time, but additional denaturation markedly increased the gelation times. In contrast, G' decreased gradually with increase in the extent of denaturation of β-lactoglobulin and its association with casein micelles. The effects of β-lactoglobulin denaturation and association on coagulation properties were less pronounced in ultrafiltered milk. G' values for ultrafiltered milk were markedly higher than the corresponding skim milks under all heating conditions.     

Rheological properties of concentrated skim milk: importance of soluble minerals in the changes in viscosity during storage

Properties of condensed milks prior to spray drying dictate to a large extent the functionality of the resulting milk powder. Rheological properties of concentrated skim milk, with total solids content of 45% but different mineral content, were studied as a function of shear rate and storage time at 50 degrees C. These milks are proposed as a model to study the effects of minerals on rheology and age gellation of condensed milk prior to drying. During storage of the concentrated milk, the apparent viscosity, particularly after 4 h, increased markedly at all shear rates studied. The yield stress also increased steeply after 4 h of storage at 50 degrees C. The changes in apparent viscosity of concentrated milk stored for up to 4 h were largely reversible under high shear, but irreversible in samples stored for longer time. The appearance of yield stress suggested the presence of reversible flocculation arising from weak attraction between casein micelles, with a transition from reversible to irreversible aggregation during storage. Particle size analysis confirmed irreversible aggregation and fusion of casein micelles during storage. Gradual reduction of mineral content of concentrated milks resulted in a marked decrease in the apparent viscosity and casein micelle aggregation during storage, while addition of minerals to milk had the opposite effect. The results demonstrated that the soluble mineral content is very important in controlling the storage-induced changes in the rheology of concentrated milks.     

Assessment of the rennet coagulation of skim milk: A comparison of methods

Monitoring of rennet-induced coagulation of milk reconstituted from low-heat or medium-heat skim milk powders with or without calcium enrichment (6.25 mmol kg⁻¹) was carried out by five analytical methods. The four types of milk samples allowed evaluation of these methods on substrates characterized by very different rennetability. Thirteen parameters describing the coagulation process were extracted from the various measurements for each sample and compared. The visual flocculation and gelation times evaluated by the Berridge test and dynamic small amplitude oscillatory rheometer (DSAOR), respectively, were not significantly different from specific parameters identified from near-infrared (NIR) spectra analysis. DSAOR, piezoelectric rheometer, and NIR spectrometer allowed and evaluation of the beginning of gel firming. It clearly appeared that NIR spectroscopy, an on-line sensor, could best assess rennet-induced coagulation of studied samples.     

Influence of pH and NaCl on rheological properties of rennet-induced casein gels made from UF concentrated skim milk

Milk concentrates are used in the production of cast cheese. The effects of pH (5.19–6.21) and NaCl concentration (0, 1.75% and 3.50%, w/w) on the rheological and microstructural properties of rennet-induced casein gels made from ultra filtered skim milk (19.8%, w/w casein) were investigated. Low pH and high NaCl concentration reduced the development rate of the gel elasticity after coagulation of the ultra filtrated skim milk. Strain at fracture and stress at fracture from uniaxial compression of casein gels 48 h after coagulation showed maximum and minimum values at pH ∼5.8 and 5.29, respectively. Young's modulus from uniaxial compression of the same gels was almost constant between pH 5.52 and 6.21 but much lower at pH 5.28. Addition of NaCl resulted in increased Young's modulus in the interval pH 6.21–5.52. As pH decreased, the level of colloidal calcium phosphate decreased concomitantly, giving less cross-links in the casein network and partly explaining the changes in the rheological properties. Increased ionic strength by adjusting pH and addition of NaCl also influenced rheological results. The microstructure examined with confocal laser-scanning microscopy was unaffected by the changes in pH and concentrations of NaCl in the range studied as revealed by image analysis and calculations of two- and three-dimensional data from micrographs.     

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.     

Heat-induced coagulation of concentrated skim milk heated by direct steam injection

Direct steam injection heat treatment on pilot scale was applied to investigate heat stability of concentrated skim milk across a broad range of temperatures from 117 °C to 153 °C and from 0.5 to 13 s holding time, assessing options for heat treat treatment without a significant amount of protein sediment formation. The relationship between total solids content of concentrated skim milk and temperature–time combinations of heat treatment could be established using minimal heat-induced coagulation as a criterion. Coagulation of destabilised casein micelles was shown to proceed non-linear over heating temperature. Transition of critical temperature–time combinations resulted in a marked increase in sediment formation indicating that preceding reactions, noticeable as the formation of dissociated material, need to take place to some extent to induce coagulum formation. UHT pre-heat treatment of skim milk prior to concentration was shown to increase heat stability in terms of possible temperature–time combinations without coagulation.     

Effect of heat treatment of rennet skim milk induced coagulation on the rheological properties and molecular structure determined by synchronous fluorescence spectroscopy and turbiscan

Heat treatment applied to milk induces denaturation of whey proteins, leading to a complex mixture of whey protein and whey protein coated casein micelles. The present paper investigates the effects of heat treatment (60 and 80°C during 20min) and rennet-induced coagulation temperature (30 and 40°C) determined by rheology, synchronous fluorescence spectroscopy (SFS) and turbiscan measurements. The gelation times determined by rheology and SFS increased with the increase of heat treatment applied to milk. The rise in temperature induced a decrease in the maximum curd firming rate and an increase in the viscosity of the investigated milk samples. The principal component analysis (PCA) applied, separately, to the SF and turbiscan spectra showed a clear discrimination between: (i) raw milks and heated milks; and (ii) milks renneted at 30°C from those renneted at 40°C. The results showed the ability of SFS as a rapid and non-destructive technique for the: (i) monitoring network structure and molecular interaction during the coagulation process; and (ii) determination of gelation time of rennet-induced coagulation of studied milk samples.     

Chemical interactions among caseins during rennet coagulation of milk

Rennet milk curds were prepared under 4 different temperature and acidity conditions. The development of different types of inter-protein chemical bonds (disulfide, hydrophobic, electrostatic, hydrogen, and calcium bridges) was monitored for 60 min after curd cutting. Hydrophobic inter-protein interactions originally present in casein micelles in milk were substituted by electrostatic, hydrogen, and calcium bonds throughout the curd curing period. Disulfide bonds were not disturbed by the experimental conditions employed in the study, remaining at a constant level in all studied treatments. Acidification of curds increased the availability of soluble ionic calcium, increasing the relative proportion of calcium bridges at the expense of electrostatic-hydrogen bonds. Although pH defined the nature of the interactions established among proteins in curd, temperature modified the rate at which such bonds were formed.     

Viscosity changes during rennet coagulation of Murciano-Granadina goat milk

The viscosity changes during coagulation of raw and pasteurized Murciano-Granadina goat milk with 10 rennets and coagulant enzymes were studied using a cylindrical rotational viscometer. The effect of three shear rates (7.32, 14.68, 73.42 s^-1) was analysed in reconstituted milk using 60% chymosin rennet and different enzyme concentrations. The shear rate selected (14.68 s^-1) corresponded to that in which the coagulation time was not modified. At a sheer rate of 73.42 s^-1 there was a significant difference (p < .05) in the milk clotting time determined compared with the shear rates of 7.32 and 14.68 s^-1. At the beginning of the coagulation process there was a decrease in the viscosity value, which was higher in raw milk (22%) than in the two heat treated samples (17% in milk pasteurized at 65°C for 30 min, low temperature long time, and 19% in milk pasteurized at 75°C for 15 s, high temperature short time). After this period the viscosity rose rapidly and reached a value 2.89 times greater than at time 0. A highly significant Pearson correlation (0.952-0.994) was found between the coagulation time determined by the Berridge method and viscometric measurement in raw and pasteurized milk.     

High-pressure-induced changes in the rennet coagulation properties of bovine milk

The mechanism of high-pressure (HP)-induced changes in rennet coagulation properties of milk, particularly the role of whey protein-casein micelle associations, was studied. Treatment at 100 or 250 MPa reduced the rennet coagulation time (RCT) of raw skimmed bovine milk, compared with untreated milk. Treatment at 400 MPa had little effect, but at 600 MPa, RCT increased considerably. HP-induced increases in RCT did not occur in serum protein-free milk or milk treated with the sulphydryl-oxidising agent KIO₃, which prevents association of denatured β-lactoglobulin with casein micelles. Treatment at 5 or 10 °C at 250–600 MPa resulted in shorter RCT than treatment at 20 °C. In milk without KIO₃, coagulum strength was highest after treatment at 250 or 400 MPa, whereas in milk with KIO₃ it was highest after treatment at 400 MPa. These results indicate the significance of HP-induced association of whey proteins with casein micelles for rennet coagulation properties of milk.     

Acid induced gelation behavior of skim milk concentrated by membrane filtration

This work reports a detailed study of the effect of ultrafiltration (UF) and diafiltration (DF) on the acid-induced gelation behavior of fresh milk retentates (2× and 4×). Concentrates were heated at 80°C for 15 min, and compared to unheated samples. The use of extensive DF caused a significantly greater amount of protein (both caseins and whey proteins) in the supernatant fraction, compared to UF retentates at the same concentration, both in unheated and heated samples. DF retentates showed higher pH of gelation, compared to the corresponding UF retentates. The development of tan δ is reported for the first time as a function of colloidal calcium release, and the protein gelation behavior discussed in light differences in composition of the soluble fraction. The results demonstrate how processing history can affect compositional changes and the gelation behavior of fresh milk retentates. Membrane filtration is a widespread unit operation in the dairy industry, employed either to prepare fresh concentrates for further processing, or ingredients with specific functional properties. This work describes in detail the effect of processing history during membrane filtration on the rheological properties of acid induced gels and will help in optimizing formulations and prepare the right ingredients for the right application. It will also be possible to determine new ways to define processing quality of the milk protein concentrates, as it relates to their ability to form texture in fermented dairy products.     

Initial study on high pressure jet processing using a modified waterjet on physicochemical and rennet coagulation properties of pasteurized skim milk

Developments in material science and engineering have enabled high pressure jet (HPJ) processing at >500 MPa. The objective of this study was to characterize the physicochemical properties of pasteurized skim milk processed at pressures from 0 to 500 MPa using waterjet technology, to explore novel uses of this technology for milk. The apparent particle size of casein micelles increased from approximately 180–220 nm in milk processed from 0 to 200 MPa to approximately 280 nm in milk treated at 500 MPa. All milk samples were shelf-stable up to 14 days at 4 °C. The viscosity of skim milk processed at 400 and 500 MPa almost doubled (4.2 mPa s) when compared with control unprocessed milk (2.2 mPa s). Increasing HPJ processing pressure changed the structure–function properties of the casein micelles and no rennet-induced coagulation was observed for milk processed at 500 MPa.     

Effect of insoluble calcium concentration on rennet coagulation properties of milk

Rennet-induced gels were made from milk acidified to various pH values or milk at pH 6.0 that had added EDTA. The objective was to examine the effect of removing insoluble Ca (INS Ca) from casein micelles (CM) on rennet gelation properties. For the pH trial, diluted lactic acid was added to reconstituted skim milk to decrease the pH to 6.4, 6.0, 5.8, 5.6, and 5.4. For the EDTA trial, EDTA was slowly added (0, 2, 4, and 6 mM) to reconstituted skim milk, and the final pH values were subsequently adjusted to pH 6.0. Dynamic low amplitude oscillatory rheology was used to monitor gel development. The Ca content of CM and rennet wheys made from these milks was measured using inductively coupled plasma spectroscopy. The INS Ca content of milk was altered by the acidification pH values or level of EDTA added. In all samples, the storage modulus (G′) exhibited a maximum (GM), with a decrease in G′ during longer aging times. Gels made at pH 6.4 had higher GM compared with gels made at pH 6.7 probably due to the reduction in electrostatic repulsion, whereas the INS Ca content only slightly decreased. The highest GM value of gels was observed at pH 6.4 and the GM value decreased with decreasing pH from 6.4 to 5.4. This was due to an excessive loss of INS Ca from CM. There was a decrease in GM with the increase in the concentration of added EDTA, which was probably due to the loss of colloidal calcium phosphate, which weakens the integrity of CM. Loss tangent (LT) values at GM increased with a reduction in milk pH and the addition of EDTA to milk. Rennet gels at the point of the GM were subjected to constant low shearing to fracture the gels. With a reduction in INS Ca content, the yield stress decreased, whereas LT values increased indicating a weaker, more flexible casein network. Microstructure of rennet-induced gels near the GM point and 2 to 10 h after this point was studied using fluorescence microscopy. At GM, gels made from milk acidified to pH 6.4 exhibited more branched, interconnected networks, whereas strands and clusters became larger with a reduction in milk pH to 5.4. Gels made from milk with EDTA added had more finely dispersed protein clusters compared with gels made from milk with no EDTA added. These microscopic observations supported the effect of loss of INS Ca on GM and LT. There was a decrease in apparent interconnectivity between strands in gel microstructure during aging, which agreed with the decrease in G′ after GM. It can be concluded that low levels of solubilization of INS Ca and the decrease in milk pH resulted in an increase in GM. With greater losses of INS Ca there was excessive reduction in cross-linking within CM, which resulted in weaker, more flexible rennet gels. This complex behavior cannot be explained by adhesive hard sphere models for CM or rennet gels made from these CM.     

Rheological properties of milk gels formed by a combination of rennet and glucono-delta-lactone

The effects of heat treatment of milk, and a range of rennet and glucono-delta-lactone (GDL) concentrations on the rheological properties, at small and large deformation, of milk gels were investigated. Gels were made from reconstituted skim milk at 30 degrees C, with two levels each of rennet and GDL. Together with controls this gave a total of sixteen gelation conditions, eight for unheated and eight for heated milk. Acid gels made from unheated milks had low storage moduli (G') of < 20 Pa. Heating milks at 80 degrees C for 30 min resulted in a large increase in the G' value of acid gels. Rennet-induced gels made from unheated milk had G' values in the range approximately 80-190 Pa. However, heat treatment severely impaired rennet coagulation: no gel was formed at low rennet levels and only a very weak gel was formed at high levels. In gels made with a combination of rennet and GDL unusual rheological behaviour was observed. After gelation, G' initially increased rapidly but then remained steady or even decreased, and at long ageing times G' values increased moderately or remained low. The loss tangent (tan delta) of acid gels made from heated milk increased after gelation to attain a maximum at pH approximately 5.1 but no maximum was observed in gels made from unheated milk. Gels made by a combination of rennet and GDL also exhibited a maximum in tan delta, indicating increased relaxation behaviour of the protein-protein bonds. We suggest that this maximum in tan delta was caused by a loosening of the intermolecular forces in casein particles caused by solubilization of colloidal calcium phosphate. We also suggest that in combination gels made from unheated milk a low value for the fracture stress and a high tan delta during gelation indicated an increased susceptibility of the network to excessive large scale rearrangements. In contrast. combination gels made from heated milk formed firmer gels crosslinked by denatured whey proteins and underwent fewer large scale rearrangements.     

Structural changes of milk proteins during heating of concentrated skim milk determined using FTIR

This work aimed to establish the impact of total solids (TS) concentration (9, 17 and 25%) of raw skim milk on the conformational properties of proteins during heating up to 121 °C. Conformational changes were determined using Fourier transform infrared spectroscopy and correlated to physicochemical properties of the system. The results showed high total solids dependence in rearrangement of β-sheet of β-lactoglobulin. Heat denaturation of whey proteins was concentration and temperature dependant. The micelle appeared to undergo temperature dependant dissociation that induced redistribution of helical and loop structures. The conformational changes were further evaluated with principal component analysis and normalising for a concentration effect for all samples. The most affected structures were located in the region 1620–1655 cm⁻¹ including intra- and inter-molecular β-sheets, α-helix/loop structures and randomly distributed structures. These findings may assist in predicting the heat stability of concentrates.     

Grazing season and forage type influence goat milk composition and rennet coagulation properties

Two different types of pasture (cultivated and rangeland) and 2 different hay qualities (high and low quality) were examined for their effects on goat milk composition and rennet coagulation properties. Furthermore, the effect of dietary treatments in both the early and late grazing season was studied. As lactation stage is known to influence milk composition, the goats in the early and late grazing season were in the same lactation stage at the start of the experiment. The milk composition was influenced both by dietary treatment and season. Milk from goats on pasture was superior to those on hay by containing a higher content of protein and casein, and the goats on cultivated pasture had the highest milk yield. Casein composition was significantly influenced by forage treatment. Goats grazing on cultivated pasture had higher contents of α_s1-casein and also of κ-casein compared with the other treatments, whereas goats grazing on rangeland had the highest content of β-casein. Factors such as milk yield, casein micelle size, α_s2-casein, and calcium content were reduced in late compared with early season. More favorable rennet coagulation properties were achieved in milk from the early grazing season, with shorter firming time and higher curd firmness compared with milk from the late grazing season, but the firming time and curd firmness were not prominently influenced by forage treatment. The content of α_s2-casein and calcium in the milk affected the firming time and the curd firmness positively. The influence of season and forage treatment on especially milk yield, casein content, and rennet coagulation properties is of economic importance for both the dairy industry and goat milk farmers.     

Modeling rennet coagulation time and curd firmness of milk

Milk coagulation properties (MCP) are traditionally expressed using rennet coagulation time (RCT), time to curd firmness (CF) of 20 mm (k₂₀), and CF 30 min after enzyme addition (a₃₀) values, all of which are single-point measures taken from the output of computerized renneting meters, such as the Formagraph. Thus, traditional MCP use only some of the available information. Moreover, because of the worldwide spreading of breeds such as the Holstein-Friesian, characterized by late-coagulating milk, it happens often that some samples do not coagulate at all, that a₃₀ is strongly and negatively related to RCT, and that k₂₀ is not measurable. The aim of the present work was to model CF as a function of time (CF_t, mm) over a 30-min interval. The model tested was CFt=CFP×(1−e−kCF×(t−RCT)), where CF_P (mm) is the potential asymptotical CF at an infinite time, k_CF (min⁻¹) is the curd firming rate constant, and RCT is measured in minutes. The CF_t model was initially applied to data of milk of each of 105 Brown Swiss cows from 7 herds, each sampled once (trial 1). Four samples did not coagulate within 30 min. Eighty-seven of the 101 individual equations obtained fit the CF data of milk samples very well, even though the samples differed in composition, and were produced by cows of different ages and days in milk, reared on different farms (coefficient of determination >0.99; average residual standard deviation = 0.21 mm). Samples with a very late RCT (slowly coagulating samples) yielded so few observational data points that curve parameters could not be precisely estimated. The repeatability of CF_t equation parameters was estimated using data obtained from 5 replicates of each of 2 samples of bulk milk from 5 Holstein-Friesian cows analyzed every day for 5 consecutive days (trial 2). Repeatability of RCT was better than that of the other 2 parameters. Moreover, traditional MCP values (RCT, a₃₀, and k₂₀) can be obtained from the individual CF_t equations, using all available information. The MCP estimated from equations were very similar to the single-point measures yielded by the computerized renneting meter (coefficient of determination >0.97), but repeatability was slightly better. The model allowed the estimation of k₂₀ for samples with a very late coagulation or with very slow curd firming. Finally, the 3 novel parameters used to assess different milk samples were less interdependent than are the traditional measures, and their practical and scientific utility requires further study.     

Age gelation in UHT-processed reconstituted concentrated skim milk

UHT reconstituted concentrated skim milks made from high-heat powder had considerably longer gelation times than those made from medium- or low-heat powders. Addition of hexametaphosphate to the concentrated milk before UHT processing markedly delayed the onset of gelation during storage. Sediment formation was greatest in the UHT concentrated skim milk made using high-heat powder followed by samples made using medium- and low-heat powders, respectively. The extent of proteolysis, as measured by 12% TCA-soluble amino groups, increased at a faster rate in the UHT milks stored at 40°C than in those stored at 22°C but decreased with increasing heat treatment of the milk prior to powder manufacture. The electrophoretic patterns of samples stored at 22°C clearly showed the breakdown of β-casein with a corresponding increase in slower moving bands, presumably γ-casein and proteose-peptone components. However, storage of samples at 40°C resulted in diffused 'blurred' protein patterns with some protein material not entering the resolving gel. At 22°C there was some evidence of proteolysis but no evidence of high molecular weight polymer formation, while at 40°C both proteolysis and high molecular weight polymer formation increased with storage time. It appeared that both physico-chemical and proteolytic processes play some part in the mechanism of gelation in UHT reconstituted concentrated skim milk.     

Physicochemical characteristics and rennet coagulation time of ultrafiltered goat milk

Goat skim milk was concentrated by ultrafiltration (UF) to volume concentration ratios (VCR) of 2, 3, 4 and 5. Gross composition, titratable acidity, pH, nitrogen distribution, percentage retention and recovery of components and rennet coagulation time (RCT) of skim milk during UF processing were studied. During UF of goat skim milk, all fat, CN, WPN, 19% of NPN, 78.1% of TS, 78.6% of ash and 3.5% of lactose were retained in 5-VCR retentate. Recovery of these components were 14.7, 53, 48, 17 for NPN, TS, ash, lactose and 100% for fat, WPN or CN, respectively. For TN, TS, ash, NPN and lactose, retention was increased by increasing the VCR. The titratable acidity was increased from an initial value of 0.14 to 0.38% in 5-VCR retentate, whereas pH decreased from 6.58 to 6.50. The RCT decreased as the protein concentration of the milk increased, but the precise influence of protein concentration decreased at higher levels of rennet.