This thesis is an investigation of binding of casein micelles to polyphenols such as epigallocatechin gallate (EGCG), resveratrol, and curcumin. The incorporation of the bioactive molecules in casein micelles seems to be beneficial. The results from this study clearly demonstrated that casein micelles can incorporate polyphenols. There is a difference in the binding behaviour between curcumin and resveratrol. Curcumin is able to penetrate the core of the micelles, but resveratrol has less affinity for the hydrophobic sites, instead, it can be bind in the core of the micelles through the water channels. The processing of milk (heating, cooling, static high pressure and microfluidization) alters the surface or the internal structure of the casein micelles resulting in increased incorporation. The release of β-casein caused alteration to the core of the casein micelles, without any effect on the colloidal calcium phosphate composition and any changes in the surface properties of the micelles. These internal rearrangements lead to an increase in the affinity of the hydrophobic sites for curcumin and resveratrol in the inner core of the micelles. This work clearly confirmed that β -casein plays a role in stabilizing internal structures in the casein micelle and its release causes an increase in the micellar size by increasing the hydration and repulsion occurring within the water pockets present in the inhomogeneous inner structure of the casein micelles. Inner core of casein micelles undergoes rearrangements during application of static high pressure and microfluidization. In the case of static high pressure, the results indicated that the persistent rearrangement of the amino acid residues induced by high-pressure treatment result in an increase in the amount of curcumin association with milk proteins. It was shown that in both untreated and microfluidized milk, the presence of polyphenol molecules significantly affects rennet induced gelation, by delaying the gelation. However, the behavior of casein micelles incorporated with resveratrol is different comparing to curcumin, as resveratrol strongly affected the surface interactions during rennet-induced gelation. Further research is needed to explain the practical aspect ( functionalities such as acid gelation, emulsification and foaming) of application of casein micelles as natural nanocarrier of bioactive compounds. / The National Sciences and Engineering Research Council (NSERC),The Ontario Dairy Council.

Pasteurized skim milk was acidified using glucono-δ-lactone (GDL) at 10, 20, 30, and 40°C or with 1.2% freeze-dried yogurt starter culture at 40°C. Milk coagulation was followed by measuring turbidity, curd firmness, particle size, and casein micelle microstructural changes using transmission electron microscopy . The pH of milk was gradually lowered during acidification with GDL or starter culture. Acidification rate showed greater influence on turbidity change at 10°C than at 20, 30, or 40°C. Average casein micelle size increased with decreasing temperature. The patterns of average micelle size versus pH were not affected by temperature. No great variation of average micelle size was observed above pH 5.2. Below pH 5.0 the size increased exponentially as the milk gelled. Acidification rate did not influence average micelle size at 10°C. Acidification rate, types of acidifying agents, and temperature had no effect on the Formagraph gelation pH and the rate at which curd firmness developed. Casein micelles became less compact and less distinct with decreasing temperature before acidification. As pH was lowered, protein was dissociated from and then reassociated with casein micelles. Acidification rate had no effect on microstructure change of casein micelles at 10°C.

The purpose of this study was to determine if size exclusion chromatography could be used to separate casein micelles from whey proteins for a rapid, direct test to measure percent casein in milk. A size exclusion chromatography column was developed for the separation having dimensions 100 by .4 cm. Packing material selected was glycophase coated porous glass supports. A Beckman DU-8B spectrophotometer monitored the casein and whey protein peaks as they eluted and a Tektronix 4052 computer accepted data points every 4 sec, storing these on tape. Absorbances and areas of each peak were used in the evaluation of samples. Treatments of temperature, pH and calcium addition were performed on a commingled milk sample from Utah State University Dairy Laboratory. It was determined that addition of calcium and pre-warming to 40 C before injection is important for good separation. Several samples of milk from individual cows were run through the column and parameters obtained. For each sample, percent casein was measured using the standard method of acid precipitation and Kjeldahl nitrogen determination. Percent casein was then estimated using area and absorbance of each casein peak from the elution plots of milk from individual cows. A regression line of predicted vs actual percent casein resulted in a correlation coefficient (r) of .92.

Master of Science / Department of Food Science / Karen A. Schmidt / People who suffer from swallowing disorders are diagnosed with dyphasgia. The beverage for the dyphagia patients should have the apparent viscosity in the range of nectar-like (51 to 350 mPa•s) or honey-like (351 to 1750 mPa•s). Due to the swallowing problems, dysphagia patients usually consume beverages slowly. Thus, the apparent viscosity of beverage for such patients should be high enough to be in the suitable range during the entire time of consumption. Three ratios of α-lactalbumin (α-la)/β-lactoglobulin (β-lg) (3:8, 1:1 and 8:3) were used to prepare the milk systems. These ratio adjusted milk systems were either processed at 70, 80, and 90ºC for 30 min or at 25ºC, and cooled to 25 ± 1ºC. After the process was completed, the milk systems were set quiescently 120 min at 25 ±1ºC. Physical and chemical properties were assessed at various time. For the milk systems at 0 min, the apparent viscosity increased in all 90°C processed-samples, and the increase was in the order of 8:3 (15.96%), 1:1 (6.38%) and 3:8 (2.11%) compared with the 25ºC samples at each ratio. When the milk systems set for 120 min, apparent viscosity increased slightly by 3.7%. The maximum apparent viscosity was 2.18 mPa•s, which was less than nectar-like. Therefore, xanthan gum was added at 0.15 w/w % to enhance rheological properties of the milk systems. α-La/β-lg ratio adjusted milk systems either with or without xanthan gum were prepared, and processed at 90ºC or 25ºC, and cooled to 25 ± 1ºC. Apparent viscosity increased by 48.61 and 89.61% in 3:8 and 8:3 milk systems, respectively for those at 0.15% xanthan gum concentration and processed at 90ºC compared with at 25ºC. Apparent viscosity of 8:3 milk systems at xanthan gum concentration of 0.15% processed at 90°C was 58.7 ± 2.12 mPa•s which was within the nectar-like range. When the samples were set for 120 min, no changes were found in the apparent viscosity of the milk systems. If the rheological properties of the milk systems can be controlled by ingredients interactions, this can be used to develop nutritious products with different forms for dysphagia patients.

Structure function changes of casein micelles were studied as a function of concentration using a non invasive concentration method, osmotic stressing. A combination of serum analysis, light scattering and rheological measurements were used to characterize the physico-chemical properties of casein micelles. In heated and unheated milk, rheological studies indicated that casein micelles behave as hard spheres of similar volume fractions, if the viscosity changes in the serum phase and the particle particle interactions are taken into account. The differences in the distribution of the heat induced complexes between colloidal and soluble phase affected the colloidal properties of casein micelles. Above 70 g L-1 protein, the protein particles were no longer free diffusing. Re-dilution of the suspensions showed no irreversible aggregation. The data suggested that in the range of concentration studied casein micelles behave as hard spheres. Age gelation was also investigated on heated and unheated concentrated milk. In unheated concentrated milk proteolysis played an important role in imparting an increase in viscosity by causing aggregation of the casein micelles. On the other hand, in heated milk, there was a significant effect of the whey protein aggregates, which increased their interaction with the casein micelles over time. This effect, together with proteolysis caused age gelation in heated concentrated milk. The method of concentration used in this research, osmotic stressing, was then compared to ultrafiltration. It was demostrated that these two methods are not equivalent, as shear and mixing during ultrafiltration cause rearrangements to the casein micells. The differences were clearly demonstrated by adding soluble caseins to the milk before or after concentration. This project brings a better understanding on the effects of concentration on the structure-function of casein micelles and the interactions occurring in milk proteins during concentration.

The effect of ultra-high temperature processing by direct steam injection and room temperature storage of pH-adjusted and unadjusted 3X (volume reduction) skim milk retentate and the effect of storage at various temperatures of 3X skim milk retentate without pH adjustment on their ζ-potential, viscosity, and pH were determined. Pasteurized skim milk was concentrated to 3X by ultrafiltration. In the pH study portions of skim milk retentate were adjusted to pH 6.38 ± .02 with HCl and 6.85 ± .01 and 7.32 ± .02 with NaOH between ultrafiltration and ultra-high temperature processing. In the storage temperature study, storage temperatures used for pH-unadjusted retentate samples were 11°C, 23°C (room temperature), and 37°C. Although pH 6.38 samples had the lowest viscosity in the pH study before ultra-high temperature processing, these samples precipitated during ultra-high temperature processing. For non-acidified samples, increasing pH of retentate resulted in higher viscosities and quicker age gelation times. Destabilization occurred more rapidly at 37°C than at 23°C or 11°C. The pH drop tended to be greater for samples stored at a higher temperature or adjusted to a higher pH. During 28 weeks of 37°C storage the pH decreased from 6.54 to 6.06. During 32 weeks of 23°C storage pH of samples initially adjusted to pH 7.32 dropped to 7.06. ζ-Potentials of casein micelles were calculated from electrophoretic mobility obtained by measuring Doppler frequency shifts of scattered laser light in samples that had been diluted 300 fold with their own ultra.filtrate. Absolute values of ζ-potential of samples stored at 37°C decreased from -23.4 millivolts immediately after ultra-high temperature processing to -18.5 millivolts after 28 weeks of storage. For samples stored at 11°C and 23°C in the storage temperature study and control samples in the pH study, absolute values of ζ-potential decreased approximately 1.5 to 2.0 millivolts during 28 or 32 weeks of storage.

The objective of this study was to investigate the effects of preheat treatments on fouling by fresh whole milk (FWM), recombined whole milk (RCB) and reconstituted whole milk (Recon) in the high-temperature heater of indirect UHT plants. Various preheat treatments prior to evaporation during milk powder manufacture were applied to skim milk powder (SMP, 75 °C 2 s, 85 °C, 155 s and 95 °C, 155 s) and whole milk powder (WMP, 95 °C, 33 s). These preheat treatments were so-called “evaporator preheat treatments”. Skim milk powder (SMP) and whole milk powder (WMP) were derived from the same original batch of pasteurised FWM to remove the effects of the variation in milk composition between different milk batches. These SMPs were recombined with anhydrous milk fat and water to prepare RCB, and WMPs were reconstituted with water to prepare Recon. Then, (homogenized) FWM, RCB and Recon were subjected to various preheat treatments (75 °C, 11 s, 85 °C, 147 s and 95 °C, 147 s) prior to UHT processing. These preheat treatments were so-called “UHT preheat treatments”. Temperature difference (hot water inlet temperature – milk outlet temperature) was taken as a measure of the extent of fouling in the high-temperature heater. The slope of the linear regression of temperature difference versus time (for two hours of UHT processing) was taken as fouling rate (°C/h). Increasing both evaporator and UHT preheat treatments resulted in increasing fouling rate and total deposit weight for all three whole milk types for several milk batches. In the case of FWM, there was no reduction in fouling rate with increasing UHT preheat treatment whether FWM was homogenized then preheated, preheated then homogenized or not homogenized at all. These findings, which are wholly consistent and well replicated, are in apparent conflict with the results of most previous comparable studies. Possible reasons for this are explained. Further investigations of the effects of homogenization relating to the role of whey protein on the surface of the fat globules showed that whey protein associated with the membrane covering the surface of fat globules for homogenized then preheated FWM, RCB and Recon and that association increased with increasing heating process stage. The increasing association of whey protein with the milk fat globules membrane with increasing severity of heating process stage became faster when preheat treatment was more severe: the association of whey protein plateaued on intermediate temperature heating when the milks were preheated at 75°C, 11 s and on preheating when the milks were preheated at 95°C, 147 s. In the case of FWM, the thickness of the membrane covering the surface of fat globules for homogenized then preheated FWM, which increased with the severity of heating process stage, was greater than the thickness of the membrane in preheated then homogenized FWM. Preheating then homogenization resulted in the greater interfacial spreading of small molecules on the surface of fat globules, i.e. whey protein or small molecules from the disintegration of casein micelles during preheating. Possible basic mechanisms for UHT fouling in the high-temperature heater include: the reduction in the solubility of calcium phosphate and the deposition of protein as fat-bound protein and non-fat-bound protein. When non-fat-bound protein in milk plasma deposited, it could be a carrier for the deposition of mineral, such as, the precipitate of calcium phosphate in the casein micelles or the deposition of complexes between whey protein and casein micelles.

The objective of this study was to investigate the effects of preheat treatments on fouling by fresh whole milk (FWM), recombined whole milk (RCB) and reconstituted whole milk (Recon) in the high-temperature heater of indirect UHT plants. Various preheat treatments prior to evaporation during milk powder manufacture were applied to skim milk powder (SMP, 75 °C 2 s, 85 °C, 155 s and 95 °C, 155 s) and whole milk powder (WMP, 95 °C, 33 s). These preheat treatments were so-called “evaporator preheat treatments”. Skim milk powder (SMP) and whole milk powder (WMP) were derived from the same original batch of pasteurised FWM to remove the effects of the variation in milk composition between different milk batches. These SMPs were recombined with anhydrous milk fat and water to prepare RCB, and WMPs were reconstituted with water to prepare Recon. Then, (homogenized) FWM, RCB and Recon were subjected to various preheat treatments (75 °C, 11 s, 85 °C, 147 s and 95 °C, 147 s) prior to UHT processing. These preheat treatments were so-called “UHT preheat treatments”. Temperature difference (hot water inlet temperature – milk outlet temperature) was taken as a measure of the extent of fouling in the high-temperature heater. The slope of the linear regression of temperature difference versus time (for two hours of UHT processing) was taken as fouling rate (°C/h). Increasing both evaporator and UHT preheat treatments resulted in increasing fouling rate and total deposit weight for all three whole milk types for several milk batches. In the case of FWM, there was no reduction in fouling rate with increasing UHT preheat treatment whether FWM was homogenized then preheated, preheated then homogenized or not homogenized at all. These findings, which are wholly consistent and well replicated, are in apparent conflict with the results of most previous comparable studies. Possible reasons for this are explained. Further investigations of the effects of homogenization relating to the role of whey protein on the surface of the fat globules showed that whey protein associated with the membrane covering the surface of fat globules for homogenized then preheated FWM, RCB and Recon and that association increased with increasing heating process stage. The increasing association of whey protein with the milk fat globules membrane with increasing severity of heating process stage became faster when preheat treatment was more severe: the association of whey protein plateaued on intermediate temperature heating when the milks were preheated at 75°C, 11 s and on preheating when the milks were preheated at 95°C, 147 s. In the case of FWM, the thickness of the membrane covering the surface of fat globules for homogenized then preheated FWM, which increased with the severity of heating process stage, was greater than the thickness of the membrane in preheated then homogenized FWM. Preheating then homogenization resulted in the greater interfacial spreading of small molecules on the surface of fat globules, i.e. whey protein or small molecules from the disintegration of casein micelles during preheating. Possible basic mechanisms for UHT fouling in the high-temperature heater include: the reduction in the solubility of calcium phosphate and the deposition of protein as fat-bound protein and non-fat-bound protein. When non-fat-bound protein in milk plasma deposited, it could be a carrier for the deposition of mineral, such as, the precipitate of calcium phosphate in the casein micelles or the deposition of complexes between whey protein and casein micelles.

The objective of this study was to investigate the effects of preheat treatments on fouling by fresh whole milk (FWM), recombined whole milk (RCB) and reconstituted whole milk (Recon) in the high-temperature heater of indirect UHT plants. Various preheat treatments prior to evaporation during milk powder manufacture were applied to skim milk powder (SMP, 75 °C 2 s, 85 °C, 155 s and 95 °C, 155 s) and whole milk powder (WMP, 95 °C, 33 s). These preheat treatments were so-called “evaporator preheat treatments”. Skim milk powder (SMP) and whole milk powder (WMP) were derived from the same original batch of pasteurised FWM to remove the effects of the variation in milk composition between different milk batches. These SMPs were recombined with anhydrous milk fat and water to prepare RCB, and WMPs were reconstituted with water to prepare Recon. Then, (homogenized) FWM, RCB and Recon were subjected to various preheat treatments (75 °C, 11 s, 85 °C, 147 s and 95 °C, 147 s) prior to UHT processing. These preheat treatments were so-called “UHT preheat treatments”. Temperature difference (hot water inlet temperature – milk outlet temperature) was taken as a measure of the extent of fouling in the high-temperature heater. The slope of the linear regression of temperature difference versus time (for two hours of UHT processing) was taken as fouling rate (°C/h). Increasing both evaporator and UHT preheat treatments resulted in increasing fouling rate and total deposit weight for all three whole milk types for several milk batches. In the case of FWM, there was no reduction in fouling rate with increasing UHT preheat treatment whether FWM was homogenized then preheated, preheated then homogenized or not homogenized at all. These findings, which are wholly consistent and well replicated, are in apparent conflict with the results of most previous comparable studies. Possible reasons for this are explained. Further investigations of the effects of homogenization relating to the role of whey protein on the surface of the fat globules showed that whey protein associated with the membrane covering the surface of fat globules for homogenized then preheated FWM, RCB and Recon and that association increased with increasing heating process stage. The increasing association of whey protein with the milk fat globules membrane with increasing severity of heating process stage became faster when preheat treatment was more severe: the association of whey protein plateaued on intermediate temperature heating when the milks were preheated at 75°C, 11 s and on preheating when the milks were preheated at 95°C, 147 s. In the case of FWM, the thickness of the membrane covering the surface of fat globules for homogenized then preheated FWM, which increased with the severity of heating process stage, was greater than the thickness of the membrane in preheated then homogenized FWM. Preheating then homogenization resulted in the greater interfacial spreading of small molecules on the surface of fat globules, i.e. whey protein or small molecules from the disintegration of casein micelles during preheating. Possible basic mechanisms for UHT fouling in the high-temperature heater include: the reduction in the solubility of calcium phosphate and the deposition of protein as fat-bound protein and non-fat-bound protein. When non-fat-bound protein in milk plasma deposited, it could be a carrier for the deposition of mineral, such as, the precipitate of calcium phosphate in the casein micelles or the deposition of complexes between whey protein and casein micelles.

Dans le lait on peut distinguer les protéines sériques et les caséines. Les protéines sériques sont des protéines globulaires qui se trouvent dans le sérum du lait et elles sont connues pour leurs propriétés fonctionnelles exceptionnelles. Quand une solution de protéines sériques est chauffée, elles perdent leur structure native et peuvent s'agréger. Elles forment des agrégats de différentes formes, tailles et densités : des cylindres, des agrégats fractals, des microgels et des agrégats fibrillaires. De l'autre côté, les caséines sont organisées dans des micelles de caséine d'un rayon environ 100-200 nm stabilisées par du phosphate de calcium colloïdal.Au cours de ce travail, nous avons cherché à comprendre comment les agrégats de protéines sériques pouvaient être utilisés en mélange avec les micelles de caséine pour obtenir et contrôler la texture de produits laitiers. Dans un premier temps, nous avons étudié le processus de « cold gelation » induit par ajout de calcium et/ou acidification d'agrégats et de microgels de protéines sériques seuls. Dans une deuxième partie, nous nous sommes intéressés à la fonctionnalité des agrégats dans les mélanges plus complexes avec les autres protéines laitières et en présence de minéraux. L'addition de petites quantités d'agrégats fractals dans des suspensions de micelles diminuait leur température critique de gélification, augmentait le module élastique et diminuait la synérèse des gels.Les agrégats de protéines sériques peuvent être utilisés pour modifier la viscosité des mélanges, comme gélifiant ou pour enrichir la teneur en protéine du milieu sans en augmenter la viscosité. / The proteins of milk can be divided into whey proteins and caseins. Whey proteins are compact globular proteins that are found in the aqueous phase of milk. They are well-known for their exceptional functional properties. Upon heating, individual whey proteins denature and aggregate, forming aggregates of different morphologies and sizes, such as strands, fractal aggregates, microgels and fibrillar aggregates, depending on the heating conditions. On the other hand, the caseins in milk are organized in complex protein units with a diameter of 100-200 nm called casein micelles stabilized by colloidal calcium phosphate (CCP).The current work is an endeavor to understand how whey protein aggregates might be used in mixtures with other dairy proteins, such as casein micelles, in order to get a particular texture in a dairy product. We first extended the understanding of so-called “cold gelation” of pure WPI aggregates induced by calcium and acidification and then studied how the aggregates work in more complex mixtures of proteins and minerals. Interestingly, addition of small amounts of fractal aggregates to suspensions of casein micelles has been demonstrated to decrease the critical gelation temperature, increase the elastic modulus and decrease the syneresis of the gels.The aggregates are to be used to modify the viscosity of dairy products, as a gelling agent and for protein enrichment. The properties of strands, fractal aggregates and microgels have been studied and compared. WPI aggregates might be considered as “clean label” texturizing ingredients that do not require approval from the European Food Safety Authority (EFSA).