Influence of storage, heat treatment, and solids composition on the bleaching of whey with hydrogen peroxide

The residual annatto colorant in liquid whey is bleached to provide a desired neutral color in dried whey ingredients. This study evaluated the influence of starter culture, whey solids and composition, and spray drying on bleaching efficacy. Cheddar cheese whey with annatto was manufactured with starter culture or by addition of lactic acid and rennet. Pasteurized fat-separated whey was ultrafiltered (retentate) and spray dried to 34% whey protein concentrate (WPC34). Aliquots were bleached at 60 °C for 1 h (hydrogen peroxide, 250 ppm), before pasteurization, after pasteurization, after storage at 3 °C and after freezing at -20 °C. Aliquots of retentate were bleached analogously immediately and after storage at 3 or -20 °C. Freshly spray dried WPC34 was rehydrated to 9% (w/w) solids and bleached. In a final experiment, pasteurized fat-separated whey was ultrafiltered and spray dried to WPC34 and WPC80. The WPC34 and WPC80 retentates were diluted to 7 or 9% solids (w/w) and bleached at 50 °C for 1 h. Freshly spray-dried WPC34 and WPC80 were rehydrated to 9 or 12% solids and bleached. Bleaching efficacy was measured by extraction and quantification of norbixin. Each experiment was replicated 3 times. Starter culture, fat separation, or pasteurization did not impact bleaching efficacy (P > 0.05) while cold or frozen storage decreased bleaching efficacy (P < 0.05). Bleaching efficacy of 80% (w/w) protein liquid retentate was higher than liquid whey or 34% (w/w) protein liquid retentate (P < 0.05). Processing steps, particularly holding times and solids composition, influence bleaching efficacy of whey. PRACTICAL APPLICATION: Optimization of whey bleaching conditions is important to reduce the negative effects of bleaching on the flavor of dried whey ingredients. This study established that liquid storage and whey composition are critical processing points that influence bleaching efficacy.     

Effect of temperature and bleaching agent on bleaching of liquid Cheddar whey

The use of whey protein as an ingredient in foods and beverages is increasing, and thus demand for colorless and mild-tasting whey protein is rising. Bleaching is commonly applied to fluid colored cheese whey to decrease color, and different temperatures and bleach concentrations are used. The objectives of this study were to compare the effects of hot and cold bleaching, the point of bleaching (before or after fat separation), and bleaching agent on bleaching efficacy and volatile components of liquid colored and uncolored Cheddar whey. First, Cheddar whey was manufactured, pasteurized, fat-separated, and subjected to one of a number of hot (68°C) or cold (4°C) bleaching applications [hydrogen peroxide (HP) 50 to 500 mg/kg; benzoyl peroxide (BP) 25 to 100 mg/kg] followed by measurement of residual norbixin and color by reflectance. Bleaching agent concentrations were then selected for the second trial. Liquid colored Cheddar whey was manufactured in triplicate and pasteurized. Part of the whey was collected (no separation, NSE) and the rest was subjected to fat separation (FSE). The NSE and FSE wheys were then subdivided and bleaching treatments (BP 50 or 100 mg/kg and HP 250 or 500 mg/kg) at 68°C for 30 min or 4°C for 16 h were applied. Control NSE and FSE with no added bleach were also subjected to each time-temperature combination. Volatile compounds from wheys were evaluated by gas chromatography-mass spectrometry, and norbixin (annatto) was extracted and quantified to compare bleaching efficacy. Proximate analysis, including total solids, protein, and fat contents, was also conducted. Liquid whey subjected to hot bleaching at both concentrations of HP or at 100mg/kg BP had greater lipid oxidation products (aldehydes) compared with unbleached wheys, 50mg/kg BP hot-bleached whey, or cold-bleached wheys. No effect was detected between NSE and FSE liquid Cheddar whey on the relative abundance of volatile lipid oxidation products. Wheys bleached with BP had lower norbixin content compared with wheys bleached with HP. Bleaching efficacy of HP was decreased at 4°C compared with 68°C, whereas that of BP was not affected by temperature. These results suggest that fat separation of liquid Cheddar whey has no effect on bleaching efficacy or lipid oxidation and that hot bleaching may result in increased lipid oxidation in fluid whey.     

The Effect of Feed Solids Concentration and Inlet Temperature on the Flavor of Spray Dried Whey Protein Concentrate

Previous research has demonstrated that unit operations in whey protein manufacture promote off‐flavor production in whey protein. The objective of this study was to determine the effects of feed solids concentration in liquid retentate and spray drier inlet temperature on the flavor of dried whey protein concentrate (WPC). Cheddar cheese whey was manufactured, fat‐separated, pasteurized, bleached (250 ppm hydrogen peroxide), and ultrafiltered (UF) to obtain WPC80 retentate (25% solids, wt/wt). The liquid retentate was then diluted with deionized water to the following solids concentrations: 25%, 18%, and 10%. Each of the treatments was then spray dried at the following temperatures: 180 °C, 200 °C, and 220 °C. The experiment was replicated 3 times. Flavor of the WPC80 was evaluated by sensory and instrumental analyses. Particle size and surface free fat were also analyzed. Both main effects (solids concentration and inlet temperature) and interactions were investigated. WPC80 spray dried at 10% feed solids concentration had increased surface free fat, increased intensities of overall aroma, cabbage and cardboard flavors and increased concentrations of pentanal, hexanal, heptanal, decanal, (E)2‐decenal, DMTS, DMDS, and 2,4‐decadienal (P < 0.05) compared to WPC80 spray dried at 25% feed solids. Product spray dried at lower inlet temperature also had increased surface free fat and increased intensity of cardboard flavor and increased concentrations of pentanal, (Z)4‐heptenal, nonanal, decanal, 2,4‐nonadienal, 2,4‐decadienal, and 2‐ and 3‐methyl butanal (P < 0.05) compared to product spray dried at higher inlet temperature. Particle size was higher for powders from increased feed solids concentration and increased inlet temperature (P < 0.05). An increase in feed solids concentration in the liquid retentate and inlet temperature within the parameters evaluated decreased off‐flavor intensity in the resulting WPC80.     

Characterizing endogenous and exogenous peroxidase activity for bleaching of fluid whey and retentate

The lactoperoxidase (LP) system may be used to achieve the desired bleaching of fluid whey with the addition of low concentrations (<50 mg/kg) of hydrogen peroxide. The addition of an exogenous peroxidase (EP) to whey may also be used to aid in whey bleaching when the LP system is not fully active. The objectives of this study were to monitor LP activity in previously refrigerated or frozen milk, fluid whey, and whey retentate (10% solids) and to evaluate peroxidase activity in fluid whey and whey retentate (10% solids), with and without additional EP (2, 1, or 0.5 dairy bleaching units), over a range of pH (5.5–6.5) and temperatures (4–60°C). Subsequent experiments were conducted to determine the relationship between enzyme activity and bleaching efficacy. Raw and pasteurized milk, fat-separated pasteurized whey, and whey retentate (10% solids) were evaluated for LP activity following storage at 4 or −20°C, using an established colorimetric method. A response surface model was applied to evaluate both endogenous and EP activity at various temperatures and pH in freshly manufactured whey and retentate. Refrigerated or frozen storage at the parameters evaluated did not affect LP activity in milk, whey, or retentate. In fluid whey, with and without added EP, as pH decreased (to 5.5) and temperature increased (to 60°C), peroxidase activity increased. Retentate with EP exhibited behavior similar to that of fluid whey: as pH decreased and temperature increased, activity increased. However, in retentate without EP, as pH increased and temperature increased, activity increased. Enzyme activity was negatively correlated to bleaching time (time for >80% norbixin destruction) in fluid whey but a linear relationship was not evident in retentate. When fluid whey is bleached enzymatically, if pH is decreased and temperature is increased, the rate of reaction increases (e.g., bleaching occurs in less time). When bleaching in retentate, a higher pH (pH 6.5 vs. pH 5.5) is desired for optimal bleaching by the LP system. Due to processing restraints, this may not be possible for all dairy producers to achieve and, thus, addition of EP could be beneficial to improve bleaching efficacy.     

The effect of bleaching agents on the degradation of vitamins and carotenoids in spray-dried whey protein concentrate

Previous research has shown that bleaching affects flavor and functionality of whey proteins. The role of different bleaching agents on vitamin and carotenoid degradation is unknown. The objective of this study was to determine the effects of bleaching whey with traditional annatto (norbixin) by hydrogen peroxide (HP), benzoyl peroxide (BP), or native lactoperoxidase (LP) on vitamin and carotenoid degradation in spray-dried whey protein concentrate 80% protein (WPC80). An alternative colorant was also evaluated. Cheddar whey colored with annatto (15 mL/454 L of milk) was manufactured, pasteurized, and fat separated and then assigned to bleaching treatments of 250 mg/kg HP, 50 mg/kg BP, or 20 mg/kg HP (LP system) at 50°C for 1 h. In addition to a control (whey with norbixin, whey from cheese milk with an alternative colorant (AltC) was evaluated. The control and AltC wheys were also heated to 50°C for 1 h. Wheys were concentrated to 80% protein by ultrafiltration and spray dried. The experiment was replicated in triplicate. Samples were taken after initial milk pasteurization, initial whey formation, after fat separation, after whey pasteurization, after bleaching, and after spray drying for vitamin and carotenoid analyses. Concentrations of retinol, a-tocopherol, water-soluble vitamins, norbixin, and other carotenoids were determined by HPLC, and volatile compounds were measured by gas chromatography-mass spectrometry. Sensory attributes of the rehydrated WPC80 were documented by a trained panel. After chemical or enzymatic bleaching, WPC80 displayed 7.0 to 33.3% reductions in retinol, β-carotene, ascorbic acid, thiamin, α-carotene, and α-tocopherol. The WPC80 bleached with BP contained significantly less of these compounds than the HP- or LP-bleached WPC80. Riboflavin, pantothenic acid, pyridoxine, nicotinic acid, and cobalamin concentrations in fluid whey were not affected by bleaching. Fat-soluble vitamins were reduced in all wheys by more than 90% following curd formation and fat separation. With the exception of cobalamin and ascorbic acid, water-soluble vitamins were reduced by less than 20% throughout processing. Norbixin destruction, volatile compound, and sensory results were consistent with previous studies on bleached WPC80. The WPC80 colored with AltC had a similar sensory profile, volatile compound profile, and vitamin concentration as the control WPC80.     

Effect of bleaching whey on sensory and functional properties of 80% whey protein concentrate

Whey is a highly functional food that has found widespread use in a variety of food and beverage applications. A large amount of the whey proteins produced in the United States is derived from annatto-colored Cheddar cheese. Color from annatto is undesirable in whey and must be bleached. The objective of this study was to compare 2 commercially approved bleaching agents, benzoyl peroxide (BP) and hydrogen peroxide (HP), and their effects on the flavor and functionality of 80% whey protein concentrate (WPC80). Colored and uncolored liquid wheys were bleached with BP or HP, and then ultrafiltered, diafiltered, and spray-dried; WPC80 from unbleached colored and uncolored Cheddar whey were manufactured as controls. All treatments were manufactured in triplicate. The WPC80 were then assessed by sensory, instrumental, functionality, color, and proximate analysis techniques. The HP-bleached WPC80 were higher in lipid oxidation compounds (specifically hexanal, heptanal, octanal, nonanal, decanal, dimethyl disulfide, and 1-octen-3-one) and had higher fatty and cardboard flavors compared with the other unbleached and BP-bleached WPC80. The WPC80 bleached with BP had lower norbixin concentrations compared with WPC80 bleached with HP. The WPC powders differed in Hunter color values (L, a, b), with bleached powders being more white, less red, and less yellow than unbleached powders. Bleaching with BP under the conditions used in this study resulted in larger reductions in yellowness of the powders made from whey with annatto color than did bleaching with HP. Functionality testing demonstrated that whey bleached with HP treatments had more soluble protein after 10 min of heating at 90°C at pH 4.6 and pH 7 than the no-bleach and BP treatments, regardless of additional color. Overall, HP bleaching caused more lipid oxidation products and subsequent off-flavors compared with BP bleaching. However, heat stability of WPC80 was enhanced by HP bleaching compared with control or BP-bleached WPC80.     

The Impact of Iron on the Bleaching Efficacy of Hydrogen Peroxide in Liquid Whey Systems

Whey is a value‐added product that is utilized in many food and beverage applications for its nutritional and functional properties. Whey and whey products are generally utilized in dried ingredient applications. One of the primary sources of whey is from colored Cheddar cheese manufacture that contains the pigment annatto resulting in a characteristic yellow colored Cheddar cheese. The colorant is also present in the liquid cheese whey and must be bleached so that it can be used in ingredient applications without imparting a color. Hydrogen peroxide and benzoyl peroxide are 2 commercially approved chemical bleaching agents for liquid whey. Concerns regarding bleaching efficacy, off‐flavor development, and functionality changes have been previously reported for whey bleached with hydrogen peroxide and benzoyl peroxide. It is very important for the dairy industry to understand how bleaching can impact flavor and functionality of dried ingredients. Currently, the precise mechanisms of off‐flavor development and functionality changes are not entirely understood. Iron reactions in a bleached liquid whey system may play a key role. Reactions between iron and hydrogen peroxide have been widely studied since the reaction between these 2 relatively stable species can cause great destruction in biological and chemical systems. The actual mechanism of the reaction of iron with hydrogen peroxide has been a controversy in the chemistry and biological community. The precise mechanism for a given reaction can vary greatly based upon the concentration of reactants, temperature, pH, and addition of biological material. In this review, some hypotheses for the mechanisms of iron reactions that may occur in fluid whey that may impact bleaching efficacy, off‐flavor development, and changes in functionality are presented. Practical Application: Cheese whey is bleached to remove residual carotenoid cheese colorant. Concerns regarding bleaching efficacy, off‐flavor development, and functionality changes have been reported for whey proteins bleached with hydrogen peroxide and benzoyl peroxide. It is very important for the dairy industry to understand how whey bleaching can impact flavor and functionality of dried ingredients. Proposed mechanisms of off‐flavor development and functionality changes are discussed in this hypothesis paper.     

Optimization of Bleaching Parameters in Refining Process of Kenaf Seed Oil with a Central Composite Design Model

Kenaf seed oil has been suggested to be used as nutritious edible oil due to its unique fatty acid composition and nutritional value. The objective of this study was to optimize the bleaching parameters of the chemical refining process for kenaf seed oil, namely concentration of bleaching earth (0.5 to 2.5% w/w), temperature (30 to 110 °C) and time (5 to 65 min) based on the responses of total oxidation value (TOTOX) and color reduction using response surface methodology. The results indicated that the corresponding response surface models were highly statistical significant (P < 0.0001) and sufficient to describe and predict TOTOX value and color reduction with R² of 0.9713 and 0.9388, respectively. The optimal parameters in the bleaching stage of kenaf seed oil were: 1.5% w/w of the concentration of bleaching earth, temperature of 70 °C, and time of 40 min. These optimum parameters produced bleached kenaf seed oil with TOTOX value of 8.09 and color reduction of 32.95%. There were no significant differences (P > 0.05) between experimental and predicted values, indicating the adequacy of the fitted models.     

Changes in the Hydrophobicity and Functionality of Whey during the Processing of Whey Protein Concentrates

Cheddar cheese whey was ultrafiltered to yield whey protein concentrates (80% WPC). The retentates were heated at 64 or 72°C for 1.5 set or received no heat treatment. Changes in composition and hydrophobicity during processing were related to WPC functionality. Heating at 72°C decreased retentate hydrophobic@ and had a detrimental affect of WPC functionality, while heating at 64°C did not. Day to day variation in the milk supply and processing conditions did not affect hydrophobicity; but the unit operations did have an effect. Ultrafiltration increased the alkane binding values of the retentate compared to the whey. Spray drying the retentate increased surface hydrophobicity and decreased alkane binding values of the WPC.     

Surface Tension of Whey and Whey Derivatives

The surface tension of various whole wheys, solutions of component whey proteins, UF fractions, and the effect of heating on the surface tensions of these solutions were determined using the Wilhemy plate method. The mean surface tension of three commercial cottage cheese wheys, a commercial cheddar cheese whey, and a laboratory rennet whey was found to be 41.7 ± 1.2 dyne/cm (25°C) and did not vary significantly with the type of whey despite differences in both pH and protein content. The surface tensions of aqueous solutions of individual pure protein fractions of whey (serum albumin, β-lactoglobulin, α-lactalbumin, and gammaglobulins), in concentrations approximating normal whey contents, were significantly different and greater than for the whole wheys.Heating of individual protein solutions at 80°C for 50 min produced insignificant changes in measured surface tension despite producing protein precipitation in some of the solutions. Similar heating of the whole whey solutions resulted in a significant decrease in surface tension and marked precipitation in most cases.The fractionation of the wheys into UF permeates and retentates resulted in a retentate fraction of significantly lower surface tension than for UF permeates. Heating increased the surface tension of retentate fractions while the permeate fractions showed a decrease.     

Effect of Whey Pretreatment on Composition and Functional Properties of Whey Protein Concentrate

The effect of pretreatment upon the composition and physicochemical and functional properties of whey, ultrafiltration (UF) retentate and freeze-dried and spray-dried whey protein concentrates (WPC) was investigated. Pretreatment was by cooling cheese whey to 0-5°C, adding calcium chloride, adjusting to pH 7.3, warming to 50°C, and removing the insoluble precipitate that formed by centrifugation or decantation. UF permeation flux rate of pretreated whey was about double that for control whey. Pretreated whey was essentially turbidity free, contained 85% less milkfat, 37% more calcium and 40% less phosphorus than whey. Pretreated whey WPC proteins were slightly more soluble at pH 3, but less functional for emulsification than whey WPC proteins. Neither whey WPC proteins nor pretreated whey WPC proteins was functional for foaming at 6% protein concentration.     

Effect of HTST Pasteurization of Milk, Cheese Whey and Cheese Whey UF Retentate upon the Composition, Physicochemical and Functional Properties of Whey Protein Concentrates

The effect of high temperature-short time (HTST) pasteurization of milk, Cheddar cheese whey and cheddar cheese whey ultrafiltration (UF) retentate upon the composition, physicochemical and functional properties of whey protein concentrates (WPC) was investigated. HTST pasteurization (72°C-15 sec) of milk, whey and UF retentate caused no significant differences in chemical composition of resulting WPCs. HTST pasteurization of milk and whey had no significant effect upon WPC solubility, whereas, heating UF retentate caused significant loss of WPC solubility. HTST pasteurization of milk caused a significant lowering (P<0.10) of maximum foam expansion of WPC dispersions, but HTST pasteurization of whey and UF retentate had no significant effect upon this latter parameter.     

Viscoelastic properties of caseinmacropeptide isolated from cow,ewe and goat cheese whey

Caseinmacropeptide (CMP) is a C‐terminal glycopeptide released from κ‐casein by the action of chymosin during cheese‐making. It is recognised as a bioactive peptide and is thought to be an ingredient with a potential use in functional foods. CMP occurs in sweet cheese whey and whey protein concentrate (WPC). Its composition is variable and depends on the particular whey source and the fractionation technology employed in the isolation. There were no significant (P < 0.05) differences in the relative apparent viscosities between species of CMPs (cow, ewe and goat). Analyses at different pH (2, 4, 7, 10), ionic strength (0, 0.2, 0.4 and 0.7 as NaCl molarity) and protein concentration (50, 100 and 200 g kg^?1) at temperatures from 10 to 90 °C carried out found pH 7 and high protein concentration (200 g kg^?1) conditions to be the best for CMP solutions to keep low and constant relative viscosity values with increasing temperature up to 75 °C. The viscoelastic properties–storage modulus, loss modulus and phase angle–of the different CMPs and WPC solutions were determined. Heat‐induced rheological changes in CMP solutions occurred at moderate temperatures (40–50 °C) with no appreciable differences in viscosity. Gelation took place significantly (P < 0.05) earlier in goat CMP (41 °C), followed by cow CMP (44 °C), ewe CMP (47 °C) and WPC (56 °C). Heating at 90 °C showed that WPC required significantly (P < 0.05) longer times to form gels (>5 min) than the CMPs (<5 min). WPC gels had higher (>20°) phase angle than CMP (<20°), which could be associated with untidy structures, limiting elastic properties of the gel. Copyright © 2006 Society of Chemical Industry     

The use of lactoperoxidase for the bleaching of fluid whey

Lactoperoxidase (LP) is the second most abundant enzyme in bovine milk and has been used in conjunction with hydrogen peroxide (H₂O₂) and thiocyanate (SCN^–) to work as an antimicrobial in raw milk where pasteurization is not feasible. Thiocyanate is naturally present and the lactoperoxidase system purportedly can be used to bleach dairy products, such as whey, with the addition of very little H₂O₂ to the system. This study had 3 objectives: 1) to quantify the amount of H₂O₂ necessary for bleaching of fluid whey using the LP system, 2) to monitor LP activity from raw milk through manufacture of liquid whey, and 3) to compare the flavor of whey protein concentrate 80% (WPC80) bleached by the LP system to that bleached by traditional H₂O₂ bleaching. Cheddar cheese whey with annatto (15 mL of annatto/454 kg of milk, annatto with 3% wt/vol norbixin content) was manufactured using a standard Cheddar cheesemaking procedure. Various levels of H₂O₂ (5–100 mg/kg) were added to fluid whey to determine the optimum concentration of H₂O₂ for LP activity, which was measured using an established colorimetric method. In subsequent experiments, fat-separated whey was bleached for 1 h with 250 mg of H₂O₂/kg (traditional) or 20 mg of H₂O₂/kg (LP system). The WPC80 was manufactured from whey bleached with 250 mg of H₂O₂/kg or 20 mg of H₂O₂/kg. All samples were subjected to color analysis (Hunter color values and norbixin extraction) and proximate analysis (fat, protein, and moisture). Sensory and instrumental volatile analyses were conducted on WPC80. Optimal LP bleaching in fluid whey occurred with the addition of 20 mg of H₂O₂/kg. Bleaching of fluid whey at either 35 or 50°C for 1 h with LP resulted in >99% norbixin destruction compared with 32 or 47% destruction from bleaching with 250 mg of H₂O₂/kg, at 35 or 50°C for 1 h, respectively. Higher aroma intensity and increased lipid oxidation compounds were documented in WPC80 from bleached whey compared with WPC80 from unbleached whey. Monitoring of LP activity throughout cheese and whey manufacture showed that LP activity sharply decreased after 30 min of bleaching (17.01 ± 1.4 to <1 U/mL), suggesting that sufficient bleaching takes place in a very short amount of time. Lactoperoxidase averaged 13.01 ± 0.7 U/mL in unpasteurized, fat-separated liquid whey and 138.6 ± 11.9 U/mL in concentrated retentate (11% solids). Lactoperoxidase may be a viable alternative for chemical whey bleaching.     

Hydroxyl Radical-Stressed Whey Protein Isolate: Chemical and Structural Properties

Many whey protein-containing foods are prepared in the presence of reactive oxygen species generated during processing. The objective of the present study was to determine chemical and structural changes, including carbonyls, sulfhydryls (SHs), dityrosine, surface hydrophobicity, turbidity, and cross-linking, in whey protein isolate (WPI) exposed to FeCl₃/H₂O₂ hydroxyl radical-generating systems (HRGS) at room temperature (20 °C). Protein carbonyl content in WPI increased (P < 0.05) with increasing concentrations of H₂O₂ when incubated for up to 10 h; total SH groups decreased (P < 0.05) in a similar fashion. The HRGS-oxidized WPI also showed a higher dityrosine content, surface hydrophobicity, and turbidity (P < 0.05) than nonoxidized WPI. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed substantial losses of β-lactoglobulin, α-lactalbumin, and bovine serum albumin, and a concomitant formation of protein polymers in oxidized WPI. The protein-oxidation indexes were all significantly correlated (P < 0.01). These oxidation-induced changes demonstrate high susceptibility of WPI to oxidative stress at room temperature and may explain variations in functionality of whey proteins often observed in formulated foods.