The effects of microfluidization on the physical, microbial, chemical, and coagulation properties of milk.

This work examines the use of mild heat treatments in conjunction with 2-pass microfluidization to generate cheese milk for potential use in soft cheeses, such as Queso Fresco. Raw, thermized, and high temperature, short time pasteurized milk samples, standardized to the 3% (wt/wt) fat content used in cheesemaking, were processed at 4 inlet temperature and pressure conditions: 42°C/75 MPa, 42°C/125 MPa, 54°C/125 MPa, and 54°C/170 MPa. Processing-induced changes in the physical, chemical, and microbial properties resulting from the intense pressure, shear, and cavitation that milk experiences as it is microfluidized were compared with nonmicrofluidized controls. A pressure-dependent increase in exit temperature was observed for all microfluidized samples, with inactivation of alkaline phosphatase in raw and thermized samples at 125 and 170 MPa. Microfluidization of all samples under the 4 inlet temperature and processing pressure conditions resulted in a stable emulsion of fat droplets ranging from 0.390 to 0.501 µm, compared with 7.921 (control) and 4.127 (homogenized control) µm. Confocal imaging showed coalescence of scattered fat agglomerates 1 to 3 µm in size during the first 24 h. We found no changes in fat, lactose, ash content or pH, indicating the major components of milk remained unaffected by microfluidization. However, the apparent protein content was reduced from 3.1 to 2.2%, likely a result of near infrared spectroscopy improperly identifying the micellar fragments embedded into the fat droplets. Microbiology results indicated a decrease in mesophilic aerobic and psychrophilic milk microflora with increasing temperature and pressure, suggesting that microfluidization may eliminate bacteria. The viscosities of milk samples were similar but tended to be higher after treatment at 54°C and 125 or 170 MPa. These samples exhibited the longest coagulation times and the weakest gel firmness, indicating that formation of the casein matrix, a critical step in the production of cheese, was affected. Low temperature and pressure (42°C/75 MPa) exhibited similar coagulation properties to controls. The results suggest that microfluidization at lower pressures may be used to manufacture high-moisture cheese with altered texture whereas higher pressures may result in novel dairy ingredients.

Comparing the effect of homogenization and heat processing on the properties and in vitro digestion of milk from organic and conventional dairy herds.

We compared the effects of homogenization and heat processing on the chemical and in vitro digestion traits of milk from organic and conventional herds. Raw milk from organic (>50% of dry matter intake from pasture) and conventional (no access to pasture) farms were adjusted to commercial whole and nonfat milk fat standards, and processed with or without homogenization, and with high-temperature-short-time or UHT pasteurization. The milk then underwent in vitro gastrointestinal digestion. Comparison of milk from organic and conventional herds showed that the milks responded to processing in similar ways. General composition was the same among the whole milk samples and among the nonfat milk samples. Protein profiles were similar, with intact caseins and whey proteins predominant and only minor amounts of peptides. Whole milk samples from grazing cows contained higher levels of α-linolenic (C18:3), vaccenic (C18:1 trans), and conjugated linoleic acids, and lower levels of palmitic (C16:0) and stearic (C18:0) acids than samples from nongrazing cows. Processing had no effect on conjugated linoleic acid and linolenic acid levels in milk, although homogenization resulted in higher levels of C8 to C14 saturated fatty acids. Of the 9 volatile compounds evaluated, milk from grazing cows contained lower levels of 2-butanone than milk from nongrazing cows, and milk from both farms showed spikes for heptanal in UHT samples and spikes for butanoic, octanoic, nonanoic, and N-decanoic acids in homogenized samples. At the start of in vitro digestion, nonfat raw and pasteurized milk samples formed the largest acid clots, and organic milk clots were larger than conventional milk clots; UHT whole milk formed the smallest clots. Milk digests from grazing cows had lower levels of free fatty acids than digests from nongrazing cows. In vitro proteolysis was similar in milk from both farms and resulted in 85 to 95% digestibility. Overall, milk from organic/grass-fed and conventional herds responded in similar ways to typical homogenization and heat processing used in United States dairy plants and showed only minor differences in chemical traits and in vitro digestion. Findings from this research enhance our knowledge of the effect of processing on the quality traits and digestibility of milk from organic/pasture-fed and confined conventional herds and will help health-conscious consumers make informed decisions about dairy selections.

Characterization of starter-free Queso Fresco made with sodium-potassium salt blends over 12 weeks of 4°C storage.

Development of reduced-sodium cheese to meet the demands of consumers concerned about sodium levels in their diet is challenging when a high-moisture, higher pH, fresh cheese, such as Queso Fresco (QF), depends on its NaCl salt content to obtain its signature flavor and quality traits. This study evaluated the effects of different Na-K salt blends on the compositional, sensorial, microbial, functional, and rheological properties of QF stored for up to 12 wk at 4°C. Queso Fresco curd from each vat was divided into 6 portions and salted with different blends of NaCl-KCl (Na-K, %): 0.75-0.75, 1.0-0.5, 1.0-1.0, 1.0-1.3, 1.0-1.5, and 2.0-0 (control). Within this narrow salt range (1.5 to 2.5% total salt), the moisture, protein, fat, and lactose levels; water activity; pH; and the textural and rheological properties were not affected by salt treatment or aging. The total salt, sodium, potassium, and ash contents reflected the different Na-K ratios added to the QF. Total aerobic microbial count, overall proteolysis, the release of casein phosphopeptides, and the level of volatile compounds were affected by aging but not by the salt treatment. Only the 1.0-1.3 and 1.0-1.5 Na-K cheeses had sensory saltiness scores similar to that of the 2.0-0 Na-K control QF. Loss of free serum from the cheese matrix increased steadily over the 12 wk, with higher losses found in QF containing 1.5% total salt compared with the higher Na-K blends. In conclusion, KCl substitution is a viable means for reduction of sodium in QF resulting in only minor differences in the quality traits, and levels of 1.0-1.3 and 1.0-1.5 Na-K are recommended to match the saltiness intensity of the 2.0-0 Na-K control. The findings from this study will aid cheese producers in creating reduced-sodium QF for health-conscious consumers.

Effect of high-pressure processing on reduction of Listeria monocytogenes in packaged Queso Fresco.

The effect of high-hydrostatic-pressure processing (HPP) on the survival of a 5-strain rifampicin-resistant cocktail of Listeria monocytogenes in Queso Fresco (QF) was evaluated as a postpackaging intervention. Queso Fresco was made using pasteurized, homogenized milk, and was starter-free and not pressed. In phase 1, QF slices (12.7 × 7.6 × 1 cm), weighing from 52 to 66 g, were surface inoculated with L. monocytogenes (ca. 5.0 log10 cfu/g) and individually double vacuum packaged. The slices were then warmed to either 20 or 40°C and HPP treated at 200, 400, and 600 MPa for hold times of 5, 10, 15, or 20 min. Treatment at 600 MPa was most effective in reducing L. monocytogenes to below the detection level of 0.91 log10 cfu/g at all hold times and temperatures. High-hydrostatic-pressure processing at 40°C, 400 MPa, and hold time ≥ 15 min was effective but resulted in wheying-off and textural changes. In phase 2, L. monocytogenes was inoculated either on the slices (ca. 5.0 log10 cfu/g; ON) or in the curds (ca. 7.0 log10 cfu/g; IN) before the cheese block was formed and sliced. The slices were treated at 20°C and 600 MPa at hold times of 3, 10, and 20 min, and then stored at 4 and 10°C for 60 d. For both treatments, L. monocytogenes became less resistant to pressure as hold time increased, with greater percentages of injured cells at 3 and 10 min than at 20 min, at which the lethality of the process increased. For the IN treatment, with hold times of 3 and 10 min, growth of L. monocytogenes increased the first week of storage, but was delayed for 1 wk, with a hold time of 20 min. Longer lag times in growth of L. monocytogenes during storage at 4°C were observed for the ON treatment at hold times of 10 and 20 min, indicating that the IN treatment may have provided a more protective environment with less injury to the cells than the ON treatment. Similarly, HPP treatment for 10 min followed by storage at 4°C was the best method for suppressing the growth of the endogenous microflora with bacterial counts remaining below the level of detection for 2 out of the 3 QF samples for up to 84 d. Lag times in growth were not observed during storage of QF at 10°C. Although HPP reduced L. monocytogenes immediately after processing, a second preservation technique is necessary to control growth of L. monocytogenes during cold storage. However, the results also showed that HPP would be effective for slowing the growth of microorganisms that can shorten the shelf life of QF.

Effect of hydrostatic high-pressure processing on the chemical, functional, and rheological properties of starter-free Queso Fresco.

Queso Fresco (QF), a popular high-moisture, high-pH Hispanic-style cheese sold in the United States, underwent high-pressure processing (HPP), which has the potential to improve the safety of cheese, to determine the effects of this process on quality traits of the cheese. Starter-free, rennet-set QF (manufactured from pasteurized, homogenized milk, milled before hooping, and not pressed) was cut into 4.5- × 4.5- × 15-cm blocks and double vacuum packaged. Phase 1 of the research examined the effects of hydrostatic HPP on the quality traits of fresh QF that had been warmed to a core temperature of 20 or 40 °C; processed at 200, 400, or 600 MPa for 5, 10, or 20 min; and stored at 4 °C for 6 to 8d. Phase 2 examined the long-term effects of HPP on quality traits when QF was treated at 600 MPa for 3 or 10 min, and stored at 4 or 10 °C for up to 12 wk. Warming the QF to 40 °C before packaging and exposure to high pressure resulted in loss of free whey from the cheese into the package, lower moisture content, and harder cheese. In phase 2, the control QF, regardless of aging temperature, was significantly softer than HPP cheeses over the 12 wk of storage. Hardness, fracture stress, and fracture rigidity increased with length of exposure time and storage temperature, with minor changes in the other properties. Queso Fresco remained a bright white, weak-bodied cheese that crumbled and did not melt upon heating. Although high pressures or long processing times may be required for the elimination of pathogens, cheese producers must be aware that HPP altered the rheological properties of QF and caused wheying-off in cheeses not pressed before packaging.

Impact of curd milling on the chemical, functional, and rheological properties of starter-free Queso Fresco.

The manufacture of Queso Fresco (QF), a high-moisture fresh Mexican cheese that is popular in the Americas, varies from country to country, with many manufacturers milling the curd before forming the cheese block to disrupt the protein matrix and ensure the crumbly nature of the QF. Because this traditional milling step does take time and may be an unnecessary point of microbial contamination, this study was undertaken to determine whether the curd-milling step could be omitted without altering the chemical, functional, and textural properties of the QF. Starter culture-free, rennet-set QF was prepared from pasteurized, homogenized milk. Curds were cooked at 39°C for 30 min, wet salted at 1.45 g of NaCl/100 g of milk, chilled, and divided into 4 portions. Curds were not milled or were subjected to coarse, medium, or fine milling and hand-packed into molds. After 12h at 4°C, the cheese was divided, vacuum packaged, and stored at 4°C for up to 8 wk. Fresh QF contained 57.3 ± 1.2% moisture, 20.9±0.8% fat, 16.0 ± 1.3% protein, 2.61 ± 0.15% lactose, and 2.25 ± 0.22% salt and had a pH of 6.36 ± 0.03%. Moisture decreased over the 8 wk of storage, whereas the fat level tended to increase. All cheeses lost 1.3 to 1.7% of their weight in whey during the first week after manufacture, and the weight gradually increased to 2.1% (nonmilled) to 3.2% (milled) by wk 8. Milling did result in QF that were softer, less chewy, and less rigid and with lower viscoelastic properties than nonmilled cheeses. Sensory panelists differentiate the finely milled QF from the other treatments, but they detected no significant differences among the nonmilled, coarsely milled, and medium-milled QF. Milling of the curd did not affect the ability of Listeria monocytogenes to grow on the cheese surface. Results from this study indicate that the milling step, which lengthens the manufacturing time, does increase wheying off during storage and results in a more fragile protein matrix. Cheese manufacturers can use this information to produce a QF that meets the demands of their customers.

Mexican Queso Chihuahua: functional properties of aging cheese.

Queso Chihuahua, a semi-hard cheese manufactured from raw milk (RM) in northern Mexico, is being replaced by pasteurized milk (PM) versions because of food safety concerns and the desire for longer shelf life. In this study, the functional traits of authentic Mexican Queso Chihuahua made from RM or PM were characterized to identify sources of variation and to determine if pasteurization of the cheese milk resulted in changes to the functional properties. Two brands of RM cheese and 2 brands of PM cheese obtained in 3 seasons of the year from 4 manufacturers in Chihuahua, Mexico, were analyzed after 0, 4, 8, 12, and 16 wk of storage at 4°C. A color measurement spectrophotometer was used to collect color data before and after heating at 232°C for 5 min or 130°C for 75 min. Meltability was measured using the Schreiber Melt Test on samples heated to 232°C for 5 min. Sliceability (the force required to cut through a sample) was measured using a texture analyzer fitted with a wire cutter attachment. Proteolysis was tracked using sodium dodecyl sulfate-PAGE. Compared with PM cheeses, RM cheeses showed less browning upon heating, melted more at 232°C, and initially required a greater cutting force. With aging, cheeses increased in meltability, decreased in whiteness when measured before heating, and required less cutting force to slice. Seasonal variations in the cheesemilk had minimal or no effect on the functional properties. The differences in the functional properties can be attributed, in part, to the mixed microflora present in the RM cheeses compared with the more homogeneous microflora added during the manufacture of PM cheeses. The degree of proteolysis and subsequent integrity of the cheese matrix contribute to melt, slice, and color properties of the RM and PM cheeses. Understanding the functional properties of the authentic RM cheeses will help researchers and cheesemakers develop pasteurized versions that maintain the traditional traits desired in the cheeses.

Fate of lysostaphin in milk from individual cows through pasteurization and cheesemaking.

Transgenic cows secreting over 3 microg of lysostaphin/ mL of milk are protected against mastitis caused by Staphylococcus aureus, but it is unknown if active lysostaphin persists through dairy processing procedures or affects the production of fermented dairy foods. The objective of this study was to determine the fate of lysostaphin as milk was pasteurized and then processed into cheese. Raw milk from transgenic cows was heat treated at 63 degrees C for 30 min, 72 degrees C for 15 s (high temperature, short time), or 140 degrees C for 2 s (UHT). Portions of the high temperature, short-time milk were manufactured into semi-hard cheeses. Aliquots taken at each processing step were assayed to determine the quantity (ELISA) and activity (ability to inhibit S. aureus growth) of lysostaphin. Results indicated that most of the lysostaphin was present in the aqueous portion of the milk and was not affected by pasteurization, although UHT treatment reduced enzyme concentration by 60%. The quantity and activity of the lysostaphin decreased during cheesemaking. Based on the amount of lysostaphin present in the starting cheesemilk, 10 to 15% of the lysostaphin was recovered in the whey, 21 to 55% in the cheese curd at d 1, and 21 to 36% in cheese stored at 4 degrees C for 90 d. Enough of the lysostaphin secreted into milk by transgenic cows survived typical dairy processing conditions to impart potential value as a bioprotective agent against staphylococci in dairy foods.

Proteolytic and rheological properties of aging cheddar-like caprine milk cheeses manufactured at different times during lactation.

The effects of 24 wk of aging on the proteolytic and rheological properties of cheddar-like cheese made from caprine milk collected at different lactation periods were evaluated. Cheddar cheese was made weekly using whole milk from Alpine goats and cheeses manufactured at weeks 4, 5, 12, 14, 15, 21, 22, and 23 of lactation were evaluated for proteolytic and rheological properties at 5 d after manufacture and after 8, 16, and 24 wk of aging at 4 degrees C. Rheology results indicated that a minimum of 8 wk of aging was needed to stabilize the texture of the cheese and that the most uniform cheeses were made from mid lactation milk. Cheeses manufactured at weeks 12 to 15 of lactation were the firmest, had the least flexible protein matrix (highest values for hardness, chewiness, and shear stress and rigidity at point of fracture), and the lowest degree of proteolysis. Understanding the factors that impact the texture of cheese, such as aging and the period of lactation that cheesemilk is obtained, will help develop guidance for maintaining the production of high quality and uniform caprine milk cheeses.

Effect of frozen storage on the proteolytic and rheological properties of soft caprine milk cheese.

Freezing and long-term frozen storage had minimal impact on the rheology and proteolysis of soft cheese made from caprine milk. Plain soft cheeses were obtained from a grade A goat dairy in Georgia and received 4 storage treatments: fresh refrigerated control (C), aged at 4 degrees C for 28 d; frozen control (FC), stored at -20 degrees C for 2 d before being thawed and aged in the same way as C cheese; and 3-mo frozen (3MF), or 6-mo frozen (6MF), stored at -20 degrees C for 3 or 6 mo before being thawed and aged. Soft cheeses had fragile textures that showed minimal change after freezing or over 28 d of aging at 4 degrees C. The only exceptions were the FC cheeses, which, after frozen storage and aging for 1 d at 4 degrees C, were significantly softer than the other cheeses, and less chewy than the other frozen cheeses. Moreover, after 28 d of aging at 4 degrees C, the FC cheeses tended to have the lowest viscoelastic values. Slight variation was noted in protein distribution among the storage treatment, although no significant proteolysis occurred during refrigerated aging. The creation and removal of ice crystals in the cheese matrix and the limited proteolysis of the caseins showed only slight impact on cheese texture, suggesting that frozen storage of soft cheeses may be possible for year-round supply with minimal loss of textural quality.

Torsion gelometry of cheese.

Torsion gelometry, a fundamental rheological test in which specimens are twisted until they fracture, was applied to several different cheese varieties to determine its suitability for measuring their textural properties. Fresh and aged Brick, Cheddar, Colby, Gouda, Havarti, Mozzarella, and Romano cheeses were subjected to torsion analysis, and the results were compared with those from small amplitude oscillatory shear (SAOS) tests and texture profile analysis (TPA). Strong relationships (correlation coefficients > 0.8) were found between torsion shear stress and TPA hardness, and between torsion shear strain and TPA cohesiveness. SAOS, which measures rheological properties of intact samples, did not correlate well with torsion or TPA. A map showing trends during aging toward brittle, mushy, rubbery, and tough texture was drawn using the torsion data. The findings show that torsion gelometry provides fundamental rheological data on cheese at the fracture point. The information can be used to compare textural qualities of cheese samples as they are being cut.

Molten globule structures in milk proteins: implications for potential new structure-function relationships.

Recent advances in the field of protein chemistry have significantly enhanced our understanding of the possible intermediates that may occur during protein folding and unfolding. In particular, studies on alpha-lactalbumin have led to the theory that the molten globule state may be a possible intermediate in the folding of many proteins. The molten globule state is characterized by a somewhat compact structure, a higher degree of hydration and side chain flexibility, a significant amount of native secondary structure but little tertiary folds, and the ability to react with chaperones. Purified alpha(s1)- and kappa-caseins share many of these same properties; these caseins may thus occur naturally in a molten globule-like state with defined, persistent structures. The caseins appear to have defined secondary structures and to proceed to quaternary structures without tertiary folds. This process may be explained, in part, by comparison with the architectural concepts of tensegrity. By taking advantage of this "new view" of protein folding, and applying these concepts to dairy proteins, it may be possible to generate new and useful forms of proteins for the food ingredient market.

Heat treatment adaptations in Clostridium perfringens vegetative cells.

Vegetative cells of Clostridium perfringens enterotoxigenic strains NCTC 8679, NCTC 8238. and H6 were grown at 37 degrees C followed by a 60-min exposure to 28 degrees C or 46 degrees C. D10-values, as a measure of thermal resistance at 60 degrees C, were significantly lower for 28 degrees C exposures as compared with cultures given 37 and 46 degrees C exposures. Following refrigeration at 4 degrees C for 24 h, D10-values for the 37 and 46 degrees C samples could not be differentiated from 28 degrees C samples. Western immunoblot analyses of lysates from heat-adapted cells also detected the increased expression of proteins reacting with antiserum directed against the molecular chaperonins from Escherichia coli; GroEL, DnaJ, and the small acid soluble protein from Bacillus subtilis, SspC. Differential scanning calorimetry (DSC) identified thermal transitions corresponding to ribosomal protein denaturations at 72.1 +/- 0.5 degrees C. Any cellular heat adaptations in the DSC profiles were lost following refrigeration for several days to simulate minimally processed food storage conditions. Further analyses of high-speed pellets from crude cell extract fractions using two-dimensional gel electrophoresis detected the differential gene expression of at least four major proteins in heat-adapted vegetative cells of C. perfringens. N-terminal amino acid analyses identified two of the proteins as glyceraldehyde 3-phosphate dehydrogenase and rubrerythrin. Both appear to have roles in this anaerobe under stressful conditions.

Rheology of dairy foods that gel, stretch, and fracture.

Casein gels are found in hard and soft dairy foods such as cheese and yogurt, and their rheological properties can be analyzed with various instrumental techniques. Empirical tests of cheese have been used since cheese making began, and imitative tests have been conducted in laboratories since the 1930s. Texture profile analysis is now the most frequently used imitative test. Fundamental tests provide a better representation of cheese and yogurt rheology because of the systematic mathematical treatment involved. Uniaxial compression and stress and strain relaxation tests are among the fundamental tests performed on cheese, and viscosity data are often used to investigate the physical characteristics of yogurt. Small amplitude oscillatory shear measurements are applicable to both products. Food scientists can acquire a clearer picture of the structure of dairy foods by performing appropriate rheological studies of the casein matrix.

Cold shock and its effect on ribosomes and thermal tolerance in Listeria monocytogenes.

Differential scanning calorimetry (DSC) and fatty acid analysis were used to determine how cold shocking reduces the thermal stability of Listeria monocytogenes. Additionally, antibiotics that can elicit production of cold or heat shock proteins were used to determine the effect of translation blockage on ribosome thermal stability. Fatty acid profiles showed no significant variations as a result of cold shock, indicating that changes in membrane fatty acids were not responsible for the cold shock-induced reduction in thermal tolerance. Following a 3-h cold shock from 37 to 0 degrees C, the maximum denaturation temperature of the 50S ribosomal subunit and 70S ribosomal particle peak was reduced from 73.4 +/- 0.1 degrees C (mean +/- standard deviation) to 72.1 +/- 0.5 degrees C (P < or = 0.05), indicating that cold shock induced instability in the associated ribosome structure. The maximum denaturation temperature of the 30S ribosomal subunit peak did not show a significant shift in temperature (from 67.5 +/- 0.4 degrees C to 66.8 +/- 0.5 degrees C) as a result of cold shock, suggesting that either 50S subunit or 70S particle sensitivity was responsible for the intact ribosome fragility. Antibiotics that elicited changes in maximum denaturation temperature in ribosomal components also elicited reductions in thermotolerance. Together, these data suggest that ribosomal changes resulting from cold shock may be responsible for the decrease in D value observed when L. monocytogenes is cold shocked.

Effect of carbon dioxide under high pressure on the survival of cheese starter cultures.

A new processing method that rapidly forms curds and whey from milk has the potential to improve cheesemaking procedures if cheese starter cultures can tolerate the processing conditions. The survival of Lactobacillus delbrueckii ssp. bulgaricus, Lactococcus lactis ssp. lactis, or Streptococcus thermophilus through this new process was evaluated. Inoculated milk containing 0, 1, or 3.25% fat or Lactobacillus MRS broth or tryptone yeast lactose broth (depending on microorganism used) was sparged with CO2 to a pressure of 5.52 MPa and held for 5 min at 38 degrees C. Broth contained 7.93 to 8.78 log CFU/ ml before processing and 7.84 to 8.66 log CFU/ml afterward. Before processing, milk inoculated with L bulgaricus, L. lactis, or S. thermophilus contained 6.81, 7.35, or 6.75 log CFU/ml, respectively. After processing, the curds contained 5.68, 7.32, or 6.50 log CFU/g, and the whey had 5.05, 6.43, or 6.14 log CFU/ml, respectively. After processing, the pHs of control samples were lower by 0.41 units in broth, 0.53 units in whey, and 0.89 units in curd. The pH of the processed inoculated samples decreased by 0.3 to 0.53 units in broth, 0.32 to 0.37 units in whey, and 0.93 to 0.98 units in the curd. Storing curds containing L. lactis at 30 degrees C or control curds and curds with L. bulgaricus or S. thermophilus at 37 degrees C for an additional 48 h resulted in pHs of 5.22, 5.41, 4.53, or 4.99, respectively. This study showed that milk inoculated with cheese starter cultures and treated with CO2 under high pressure to precipitate casein-produced curds that contained sufficient numbers of viable starter culture to produce lactic acid, thereby decreasing the pH.