Characterization of calcium lactate crystals on cheddar cheese by image analysis

Previous research demonstrated that crystal coverage on the surface of Cheddar cheese can be quantitatively and nondestructively measured using image analysis of digital photographs of the cheese surface. The objective of the present study was to extend image analysis methodology to quantify and characterize additional features of visible crystals on cheese surfaces as they grow over time. A random weight (∼300 g) retail sample of naturally smoked Cheddar cheese exhibiting white surface crystals was obtained from a commercial source. The total area occupied by crystals and total number of discrete crystal regions on one of the surfaces (∼55 × 120 mm) was measured at 3-wk intervals for 30 wk using image analysis. In addition, 5 small (∼0.3 mm radius) individual crystals on that surface were chosen for observation over the 30-wk period. The crystals were evaluated for area, radius, and shape factor (circularity) every third week using image analysis. The total area occupied by crystals increased in a linear manner (R² = 0.95) from about 0.44 to 7.42% of the total cheese surface area over the 30-wk period. The total number of discrete crystal regions also increased but in a nonlinear manner that was best described by a quadratic relationship. Measurement of discrete crystal regions underestimated the true number of crystals present at the cheese surface due to merging of adjacent crystals as they grew and merged into a single crystal region over time. Throughout this period, the shapes of the 5 individual crystals closely approximated perfect circles, except when adjacent crystals merged to form a single irregular crystal region, and the area occupied by each of the 5 crystals increased in a near-linear manner (R² = 0.95). Image analysis approaches may be used to evaluate crystal formation and growth rates and morphology on cheese.     

Nonstarter lactic acid bacteria and aging temperature affect calcium lactate crystallization in cheddar cheese

The occurrence of unappetizing calcium lactate crystals in Cheddar cheese is a challenge and expense to manufacturers, and this research was designed to understand their origin. It was hypothesized that nonstarter lactic acid bacteria (NSLAB) affect calcium lactate crystallization (CLC) by producing D(-)-lactate. This study was designed to understand the effect of NSLAB growth and aging temperature on CLC. Cheeses were made from milk inoculated with Lactococcus lactis starter culture, with or without Lactobacillus curvatus or L. helveticus WSU19 adjunct cultures. Cheeses were aged at 4 or 13 degrees C for 28 d, then half of the cheeses from 4 and 13 degrees C were transferred to 13 and 4 degrees C, respectively, for the remainder of aging. The form of lactate in cheeses without adjunct culture or with L. helveticus WSU19 was predominantly L(+)-lactate (> 95%, wt/wt), and crystals were not observed within 70 d. While initial lactate in cheeses containingL. curvatus was only L(+)-lactate, the concentration of D(-)-lactate increased during aging. After 28 d, a racemic mixture of D/L-lactate was measured in cheeses containing L. curvatus; at the same time, CLC was observed. The earliest and most extensive CLC occurred on cheeses aged at 13 degrees C for 28 d then transferred to 4 degrees C. These results showed that production of D(-)-lactate by NSLAB, and aging temperature affect CLC in maturing Cheddar cheese.     

Compositional factors associated with calcium lactate crystallization in smoked Cheddar cheese

Previous researchers have observed that surface crystals of calcium lactate sometimes develop on some Cheddar cheese samples but not on other samples produced from the same vat of milk. The causes of within-vat variation in crystallization behavior have not been identified. This study compared the compositions of naturally smoked Cheddar cheese samples that contained surface crystals with those of samples originating from the same vat that were crystal-free. Six pairs of retail samples (crystallized and noncrystallized) produced at the same cheese plant on different days were obtained from a commercial source. Cheese samples were 5 to 6 mo old at the time of collection. They were then stored for an additional 5 to 13 mo at 4°C to ensure that the noncrystallized samples remained crystal-free. Then, the crystalline material was removed and collected from the surfaces of crystallized samples, weighed, and analyzed for total lactic acid, l(+) and d(−) lactic acid, Ca, P, NaCl, moisture, and crude protein. Crystallized and noncrystallized samples were then sectioned into 3 concentric subsamples (0 to 5 mm, 6 to 10 mm, and greater than 10 mm depth from the surface) and analyzed for moisture, NaCl, titratable acidity, l(+) and d(−) lactic acid, pH, and total and water-soluble calcium. The data were analyzed by ANOVA according to a repeated measures design with 2 within-subjects variables. The crystalline material contained 52.1% lactate, 8.1% Ca, 0.17% P, 28.5% water, and 8.9% crude protein on average. Both crystallized and noncrystallized cheese samples contained significant gradients of decreasing moisture from center to surface. Compared with noncrystallized samples, crystallized samples possessed significantly higher moisture, titratable acidity, l(+) lactate, and water soluble calcium, and significantly lower pH and NaCl content. The data suggest that formation of calcium lactate crystals may have been influenced by within-vat variation in salting efficacy in the following manner. Lower salt uptake by some of the cheese curd during salting may have created pockets of higher moisture and thus higher lactose within the final cheese. When cut into retail-sized chunks, the lower salt, higher moisture samples contained more lactic acid and thus lower cheese pH, which shifted calcium from the insoluble to the soluble state. Lactate and soluble calcium contents in these samples became further elevated at the cheese surface because of dehydration during smoking, possibly triggering the formation of calcium lactate crystals.     

Chemical changes that predispose smoked Cheddar cheese to calcium lactate crystallization

We have observed a high incidence of calcium lactate surface crystals on naturally smoked Cheddar cheese in the retail marketplace. The objective of this study was to identify chemical changes that may occur during natural smoking that render Cheddar cheese more susceptible to calcium lactate crystal formation. Nine random-weight (approximately 300 g) retail-packaged samples of smoked Cheddar cheese were obtained from a commercial manufacturer immediately after the samples were smoked for about 6 h at 20°C in a commercial smokehouse. Three similarly sized samples that originated from the same 19.1-kg block of cheese and that were not smoked were also obtained. Within 2 d after smoking, 3 smoked and 3 control (not smoked) samples were sectioned into 5 subsamples at different depths representing 0 to 2, 2 to 4, 4 to 6, 6 to 8, and 8 to 10 mm from the cheese surface. Six additional smoked cheese samples were similarly sectioned at 4 wk and again at 10 wk of storage at 5°C. Sample sections were analyzed for moisture, l(+) and d(−) lactate, pH, and water-soluble calcium. The effects of treatment (smoked, control), depth from cheese surface, and their interactions were analyzed by ANOVA according to a repeated measures design with 2 within-subject variables. Smoked samples contained signficantly lower moisture and lower pH, and higher total lactate-in-moisture (TLIM) and water-soluble calcium-in-moisture (WSCIM) than control cheeses. Smoked samples also contained significant gradients of moisture, pH, TLIM, and WSCIM, with lower moisture and pH, and higher TLIM and WSCIM, occurring at the cheese surface. Gradients of moisture were still present in smoked samples at 4 and 10 wk of storage. In contrast, the pH, TLIM, and WSCIM equilibrated and showed no gradients at 4 and 10 wk. The results indicate that calcium and lactate in the serum phase of the cheese were elevated because of smoking, especially at the cheese surface immediately after smoking treatment, which presumably predisposes the smoked cheeses to increased susceptibility to calcium lactate surface crystallization.     

Surface roughness and packaging tightness affect calcium lactate crystallization on Cheddar cheese

Calcium lactate crystals that sometimes form on Cheddar cheese surfaces are a significant expense to manufacturers. Researchers have identified several postmanufacture conditions such as storage temperature and packaging tightness that contribute to crystal formation. Anecdotal reports suggest that physical characteristics at the cheese surface, such as roughness, cracks, and irregularities, may also affect crystallization. The aim of this study was to evaluate the combined effects of surface roughness and packaging tightness on crystal formation in smoked Cheddar cheese. Four 20-mm-thick cross-section slices were cut perpendicular to the long axis of a retail block (~300 g) of smoked Cheddar cheese using a wire cutting device. One cut surface of each slice was lightly etched with a cheese grater to create a rough, grooved surface; the opposite cut surface was left undisturbed (smooth). The 4 slices were vacuum packaged at 1, 10, 50, and 90 kPa (very tight, moderately tight, loose, very loose, respectively) and stored at 1°C. Digital images were taken at 1, 4, and 8 wk following the first appearance of crystals. The area occupied by crystals and number of discrete crystal regions (DCR) were quantified by image analysis. The experiment was conducted in triplicate. Effects of storage time, packaging tightness, surface roughness, and their interactions were evaluated by repeated-measures ANOVA. Surface roughness, packaging tightness, storage time, and their 2-way interactions significantly affected crystal area and DCR number. Extremely heavy crystallization occurred on both rough and smooth surfaces when slices were packaged loosely or very loosely and on rough surfaces with moderately tight packaging. In contrast, the combination of rough surface plus very tight packaging resulted in dramatic decreases in crystal area and DCR number. The combination of smooth surface plus very tight packaging virtually eliminated crystal formation, presumably by eliminating available sites for nucleation. Cut-and-wrap operations may significantly influence the crystallization behavior of Cheddar cheeses that are saturated with respect to calcium lactate and thus predisposed to form crystals.     

Gas-flushed packaging contributes to calcium lactate crystals in Cheddar cheese

Gas-flushed packaging is commonly used for cheese shreds and cubes to prevent aggregation and loss of individual identity. Appearance of a white haze on cubed cheese is unappealing to consumers, who may refrain from buying, resulting in lost revenue to manufacturers. The objective of this study was to determine whether gas flushing of Cheddar cheese contributes to the occurrence of calcium lactate crystals (CLC). Cheddar cheese was manufactured using standard methods, with addition of starter culture, annatto, and chymosin. Two different cheese milk compositions were used: standard (lactose:protein = 1.47, protein:fat = 0.90, lactose = 4.8%) and ultrafiltered (UF; lactose:protein = 1.23, protein:fat = 0.84, lactose = 4.8%), with or without adjunct Lactobacillus curvatus. Curds were milled when whey reached 0.45% titratable acidity, and pressed for 16 h. After aging at 7.2°C for 6 mo, cheeses were cubed (1 × 1 × 4 cm) and either vacuum-packaged or gas-flushed with carbon dioxide, nitrogen, or a 50:50 mixture of carbon dioxide and nitrogen, then aged for an additional 3 mo. Heavy crystals were observed on surfaces of all cubed cheeses that were gas-flushed, but not on cheeses that were vacuum-packaged. Cheeses without Lb. curvatus exhibited l(+)-CLC on surfaces, whereas cheeses with Lb. curvatus exhibited racemic mixtures of l(+)/d(−)-CLC throughout the cheese matrices. The results show that gas flushing (regardless of gas composition), milk composition, and presence of nonstarter lactic acid bacteria, can contribute to the development of CLC on cheese surfaces. These findings stress the importance of packaging to cheese quality.     

Cheese pH, protein concentration, and formation of calcium lactate crystals

The occurrence of calcium lactate crystals (CLC) in hard cheeses is a continual expense to the cheese industry, as consumers fail to purchase cheeses with this quality defect. This research investigates the effects of the protein concentration of cheese milk and the pH of cheese on the occurrence of CLC. Atomic absorption spectroscopy was used to determine total and soluble calcium concentrations in skim milk (SM1, 8.7% total solids), and skim milk supplemented with nonfat dry milk (CSM1, 13.5% total solids). Calcium, phosphorus, lactic acid, and citrate were determined in cheeses made with skim milk (SM2, 3.14% protein), skim milk supplemented with ultrafiltered milk (CSM2, 6.80% protein), and nonfat dry milk (CSM3, 6.80% protein). Supplementation with nonfat dry milk increased the initial total calcium in CSM1 (210 mg/100 g of milk) by 52% compared with the total calcium in SM1 (138 mg/100 g of milk). At pH 5.4, soluble calcium concentrations in CSM1 were 68% greater than soluble calcium in SM1. In cheeses made from CSM2 and CSM3, total calcium was 26% greater than in cheeses made from SM2. As the pH of cheeses made from SM2 decreased from 5.4 to 5.1, the concentration of soluble calcium increased by 61.6%. In cheeses made from CSM2 and CSM3, the concentrations of soluble calcium increased by 41.4 and 45.5%, respectively. Calcium lactate crystals were observed in cheeses made from SM2 at and below pH 5.1, whereas CLC were observed in cheeses from CSM2 and CSM3 at and below pH 5.3. The increased presence of soluble calcium can potentially cause CLC to occur in cheese manufactured with increased concentrations of milk solids, particularly at and below pH 5.1.     

Factors affecting calcium lactate and liquid expulsion defects in Cheddar cheese

This paper summarizes the results of 2 studies designed to investigate the influence of several manufacturing and curing treatments on the appearance of Cheddar cheese defects. Specifically, 2 defects, calcium lactate crystal formation and the expulsion of free liquid (weeping) were monitored in Cheddar cheese. Both studies were conducted at a commercial cheese manufacturing facility that produces Cheddar in 18.14-kg (40-lb) blocks. In the first study we monitored cheese calcium, both total and soluble during manufacture and early curing. In the second study we measured cheese pH from 3 d through 8 mo, as well as some factors that are influenced by cheese pH. Early cheese pH (3 d to 7 d) patterns were used to select vats of cheese for retail packaging. Mild Cheddar packaged at 30 d postmanufacture and sharp Cheddar packaged at 8 mo postmanufacture from the same vats were monitored for the incidence and severity of the defects. Our results indicated that factors measured in early stages of manufacture and curing (less than 7 d) such as cheese pH at mill, lactic acid concentration, nonprotein nitrogen, and calcium (total and soluble) in cheese did not correlate with the appearance of either calcium lactate or expulsion of free liquid in packaged cheeses. Factors including pH, lactic acid concentrations, and soluble calcium measured during curing (greater than 7 d) of cheese were found to be statistically significant in the development of defects and appeared to be associated with use of specific starter culture groups. In the study, 5 different starter culture groups, each consisting of a 4-strain blend of Lactococcus lactis ssp. cremoris and Lactococcus lactis ssp. lactis, were used to manufacture the cheeses. Cheese manufactured with one particular culture group showed no incidence of calcium lactate crystal formation or weeping during curing and shelf-life of cheeses in this study. This starter group also generated the least amount of pH change in cheese during the first month of curing. From these results we conclude that starter culture group, more than any other factor measured, played an important role in the development of calcium lactate and liquid expulsion defects in Cheddar cheese. Starter culture group appeared to strongly influence cheese pH, lactic acid, and soluble calcium concentrations during curing and storage.     

Development and application of image analysis to quantify calcium lactate crystals on the surface of smoked Cheddar cheese

Calcium lactate crystals that form white specks or haze on the surface of cheese constitute a significant quality problem for producers of Cheddar cheese. Subjective methods to evaluate crystal coverage of cheese surfaces have been reported previously, but objective methods are currently lacking. The objectives of this work were to develop and evaluate an objective method to measure the area occupied by calcium lactate crystals on surfaces of naturally smoked Cheddar cheese samples using digital photography and image analysis. Coefficients of variation ranged from 1.29 to 4.68% for 5 replicate analyses of 3 different cheese surfaces that ranged from ∼2 to 49% of total surface area occupied by crystals. Thus, results showed a high degree of repeatability for the 3 cheese surfaces, which ranged from very slight and geometrically simple to very heavy and geometrically complex crystal coverage. The method underestimated total area occupied by crystals on the 3 surfaces by 0.24 to 4.83% unless the fainter crystal regions that went undetected during initial thresholding were manually segmented and quantified. The wet weight of crystal substance collected per unit of surface area from 20 different cheese samples increased exponentially as the percentage of total surface area occupied by crystals increased. These data were consistent with subjective observations that crystal regions appeared to grow vertically as well as horizontally as they expanded to occupy greater surface area. Image analysis was well suited for evaluating changes in crystal coverage during cheese aging because measurements were made nondestructively and with minimal disruption to the cheese. The area occupied by crystals on 6 different surfaces from 3 different cheese samples increased linearly (R² = 0.94 to 0.99) during storage at 4°C for up to 33 wk. However, the rates of increase differed significantly among the 3 cheese samples. Image analysis may serve as a useful tool to quantitatively evaluate the effects of factors such as cheese composition, packaging conditions and storage temperature on rate of crystal growth and time of crystal appearance during storage.     

Influence of calcium and phosphorus, lactose, and salt-to-moisture ratio on cheddar cheese quality: pH changes during ripening

The pH of cheese is an important attribute that influences its quality. Substantial changes in cheese pH are often observed during ripening. A combined effect of calcium, phosphorus, residual lactose, and salt-to-moisture ratio (S/M) of the cheese on the changes in cheese pH during ripening was investigated. Eight cheeses with 2 levels of Ca and P (0.67 and 0.47% vs. 0.53 and 0.39%, respectively), lactose at pressing (2.4 vs. 0.78%), and S/M (6.4 vs. 4.8%) were manufactured. All the cheeses were salted at a pH of 5.4, pressed for 5 h, and then ripened at 6 to 8°C. The pH of the salted curds before pressing and the cheeses during 48 wk of ripening was measured. Also, cheeses were analyzed for water-soluble Ca and P, organic P, and bound inorganic P during ripening. Changes in organic acids’ concentration and shifts in the distribution of Ca and P between different forms were studied in relation to changes in pH. Cheeses with low S/M exhibited a larger increase in acid production during ripening compared with high S/M cheeses. Cheeses with the highest concentration of bound inorganic P exhibited the highest pH, whereas cheeses with the lowest concentration of bound inorganic P exhibited the lowest pH among the 8 treatments. Although conversion of lactose to short-chain, water-soluble organic acids decreased cheese pH, bound inorganic phosphate buffered the changes in cheese pH. Production of acid in excess of the buffering capacity (which was the case in low Ca and P and low S/M treatments) led to a low pH, whereas solubilization of bound inorganic P in excess to acid production (which was the case in high Ca and P and high S/M treatments) led to an increase in pH. However, for cheeses with high Ca and P and low S/M, changes in cheese pH were influenced by the level of residual lactose. Hence, pH changes in Cheddar cheese can be modulated by a concomitant control on the amount and state of Ca and P, level of residual lactose, and S/M of the cheese.     

Nonstarter lactic acid bacteria biofilms and calcium lactate crystals in Cheddar cheese

A sanitized cheese plant was swabbed for the presence of nonstarter lactic acid bacteria (NSLAB) biofilms. Swabs were analyzed to determine the sources and microorganisms responsible for contamination. In pilot plant experiments, cheese vats filled with standard cheese milk (lactose:protein = 1.47) and ultrafiltered cheese milk (lactose:protein = 1.23) were inoculated with Lactococcus lactis ssp. cremoris starter culture (8 log cfu/mL) with or without Lactobacillus curvatus or Pediococci acidilactici as adjunct cultures (2 log cfu/mL). Cheddar cheeses were aged at 7.2 or 10°C for 168 d. The raw milk silo, ultrafiltration unit, cheddaring belt, and cheese tower had NSLAB biofilms ranging from 2 to 4 log cfu/100 cm². The population of Lb. curvatus reached 8 log cfu/g, whereas P. acidilactici reached 7 log cfu/g of experimental Cheddar cheese in 14 d. Higher NSLAB counts were observed in the first 14 d of aging in cheese stored at 10°C compared with that stored at 7.2°C. However, microbial counts decreased more quickly in Cheddar cheeses aged at 10°C compared with 7.2°C after 28 d. In cheeses without specific adjunct cultures (Lb. curvatus or P. acidilactici), calcium lactate crystals were not observed within 168 d. However, crystals were observed after only 56 d in cheeses containing Lb. curvatus, which also had increased concentration of d(−)-lactic acid compared with control cheeses. Our research shows that low levels of contamination with certain NSLAB can result in calcium lactate crystals, regardless of lactose:protein ratio.     

Factors affecting solubility of calcium lactate in aqueous solutions

Calcium lactate (CaL2) crystal formation on the surface of cheese continues to be a widespread problem for the cheese industry despite decades of research. To prevent those crystals from forming, it is necessary to keep the concentration of CaL2 below saturation level. The limited data available on the solubility of CaL2 at conditions appropriate for the storage of cheese are often conflicting. In this work, the solubility of L(+)-CaL2 in water was evaluated at 4, 10, and 24 degrees C, and the effects of salt and pH at those temperatures were investigated. The effects of additional calcium and lactate ions on solubility also were studied. The results suggested that temperature and the concentration of lactate ions are the main factors influencing the solubility of CaL2, with the other parameters having limited effect.     

Influence of calcium and phosphorus, lactose, and salt-to-moisture ratio on Cheddar cheese quality: changes in residual sugars and water-soluble organic acids during ripening

Cheddar cheese ripening involves the conversion of lactose to glucose and galactose or galactose-6-phosphate by starter and nonstarter lactic acid bacteria. Under ideal conditions (i.e., where bacteria grow under no stress of pH, water activity, and salt), these sugars are mainly converted to lactic acid. However, during ripening of cheese, survival and growth of bacteria occurs under the stressed condition of low pH, low water activity, and high salt content. This forces bacteria to use alternate biochemical pathways resulting in production of other organic acids. The objective of this study was to determine if the level and type of organic acids produced during ripening was influenced by calcium (Ca) and phosphorus (P), residual lactose, and salt-to-moisture ratio (S/M) of cheese. Eight cheeses with 2 levels of Ca and P (0.67 and 0.47% vs. 0.53 and 0.39%, respectively), lactose at pressing (2.4 vs. 0.78%), and S/M (6.4 vs. 4.8%) were manufactured. The cheeses were analyzed for organic acids (citric, orotic, pyruvic, lactic, formic, uric, acetic, propanoic, and butyric acids) and residual sugars (lactose, galactose) during 48 wk of ripening using an HPLC-based method. Different factors influenced changes in concentration of residual sugars and organic acids during ripening and are discussed in detail. Our results indicated that the largest decrease in lactose and the largest increase in lactic acid occurred between salting and d 1 of ripening. It was interesting to observe that although the lactose content in cheese was influenced by several factors (Ca and P, residual lactose, and S/M), the concentration of lactic acid was influenced only by S/M. More lactic acid was produced in low S/M treatments compared with high S/M treatments. Although surprising for Cheddar cheese, a substantial amount (0.2 to 0.4%) of galactose was observed throughout ripening in all treatments. Minor changes in the levels of citric, uric, butyric, and propanoic acids were observed during early ripening, whereas during later ripening, a substantial increase was observed. A gradual decrease in orotic acid and a gradual increase in pyruvic acid content of the cheeses were observed during 12 mo of ripening. In contrast, acetic acid did not show a particular trend, indicating its role as an intermediate in a biochemical pathway, rather than a final product.     

Powder X-ray diffraction can differentiate between enantiomeric variants of calcium lactate pentahydrate crystal in cheese

Powder X-ray diffraction has been used for decades to identify crystals of calcium lactate pentahydrate in Cheddar cheese. According to this method, diffraction patterns are generated from a powdered sample of the crystals and compared with reference cards within a database that contains the diffraction patterns of known crystals. During a preliminary study of crystals harvested from various Cheddar cheese samples, we observed 2 slightly different but distinct diffraction patterns that suggested that calcium lactate pentahydrate may be present in 2 different crystalline forms. We hypothesized that the 2 diffraction patterns corresponded to 2 enantiomeric forms of calcium lactate pentahydrate (l- and dl-) that are believed to occur in Cheddar cheese, based on previous studies involving enzymatic analyses of the lactate enantiomers in crystals obtained from Cheddar cheeses. However, the powder X-ray diffraction database currently contains only one reference diffraction card under the title “calcium lactate pentahydrate.” To resolve this apparent gap in the powder X-ray diffraction database, we generated diffraction patterns from reagent-grade calcium l-lactate pentahydrate and laboratory-synthesized calcium dl-lactate pentahydrate. From the resulting diffraction patterns we determined that the existing reference diffraction card corresponds to calcium dl-lactate pentahydrate and that the other form of calcium lactate pentahydrate observed in cheese crystals corresponds to calcium l-lactate pentahydrate. Therefore, this report presents detailed data from the 2 diffraction patterns, which may be used to prepare 2 reference diffraction cards that differentiate calcium l-lactate pentahydrate from calcium dl-lactate pentahydrate. Furthermore, we collected crystals from the exteriors and interiors of Cheddar cheeses to demonstrate the ability of powder X-ray diffraction to differentiate between the 2 forms of calcium lactate pentahydrate crystals in Cheddar cheeses. Powder X-ray diffraction results were validated using enzymatic assays for lactate enantiomers. These results demonstrated that powder X-ray diffraction can be used as a diagnostic tool to quickly identify different forms of calcium lactate pentahydrate that may occur in Cheddar cheese.     

Comparison of effect of vacuum-condensed and ultrafiltered milk on cheddar cheese

The objective of this study was to compare the effects of vacuum-condensed (CM) and ultrafiltered (UF) milk on some compositional and functional properties of Cheddar cheese. Five treatments were designed to have 2 levels of concentration (4.5 and 6.0% protein) from vacuum-condensed milk (CM1 and CM2) and ultrafiltered milk (UF1 and UF2) along with a 3.2% protein control. The samples were analyzed for fat, protein, ash, calcium, and salt contents at 1 wk. Moisture content, soluble protein, meltability, sodium dodecyl sulfate-PAGE, and counts of lactic acid bacteria and nonstarter lactic acid bacteria were performed on samples at 1, 18, and 30 wk. At 1 wk, the moisture content ranged from 39.2 (control) to 36.5% (UF2). Fat content ranged from 31.5 to 32.4% with no significant differences among treatments, and salt content ranged from 1.38 to 1.83% with significant differences. Calcium content was higher in UF cheeses than in CM cheeses followed by control, and it increased with protein content in cheese milk. Ultrafiltered milk produced cheese with higher protein content than CM milk. The soluble protein content of all cheeses increased during 30 wk of ripening. Condensed milk cheeses exhibited a higher level of proteolysis than UF cheeses. Sodium dodecyl sulfate-PAGE showed retarded proteolysis with increase in level of concentration. The breakdown of alphas1- casein and alphas1-I-casein fractions was highest in the control and decreased with increase in protein content of cheese milk, with UF2 being the lowest. There was no significant degradation of beta-casein. Overall increase in proteolytic products was the highest in control, and it decreased with increase in protein content of cheese milk. No significant differences in the counts of lactic starters or nonstarter lactic acid bacteria were observed. Extent as well as method of concentration influenced the melting characteristics of the cheeses. Melting was greatest in the control cheeses and least in cheese made from condensed milk and decreased with increasing level of milk protein concentration. Vacuum condensing and ultrafiltration resulted in Cheddar cheeses of distinctly different quality. Although both methods have their advantages and disadvantages, the selection of the right method would depend upon the objective of the manufacturer and intended use of the cheese.     

Influence of salt-to-moisture ratio on starter culture and calcium lactate crystal formation

The occurrence of l(+)-lactate crystals in hard cheeses continues to be an expense to the cheese industry. Salt tolerance of the starter culture and the salt-to-moisture ratio (S:M) in cheese dictate the final pH of cheese, which influences calcium lactate crystal (CLC) formation. This research investigates these interactions on the occurrence of CLC. A commercial starter was selected based on its sensitivity to salt, less than and greater than 4.0% S:M. Cheddar cheese was made by using either whole milk (3.25% protein, 3.85% fat) or whole milk supplemented with cream and ultrafiltered milk (4.50% protein, 5.30% fat). Calculated amounts of salt were added at milling (pH 5.40 ± 0.02) to obtain cheeses with less than 3.6% and greater than 4.5% S:M. Total and soluble calcium, total lactic acid, and pH were measured and the development of CLC was monitored in cheeses. All cheeses were vacuum packaged and gas flushed with nitrogen gas and aged at 7.2°C for 15 wk. Concentration of total lactic acid in high S:M cheeses ranged from 0.73 to 0.80 g/100 g of cheese, whereas that in low S:M cheeses ranged from 1.86 to 1.97 g/100 g of cheese at the end of 15 wk of aging because of the salt sensitivity of the starter culture. Concentrated milk cheeses with low and high S:M exhibited a 30 to 28% increase in total calcium (1,242 and 1,239 mg/100 g of cheese, respectively) compared with whole milk cheeses with low and high S:M (954 and 967 mg/100 g of cheese, respectively) throughout aging. Soluble calcium was 41 to 35% greater in low S:M cheeses (low-salt whole milk cheese and low-salt concentrated milk cheese; 496 and 524 mg/100 g of cheese, respectively) compared with high S:M cheeses (high-salt whole milk cheese and high-salt concentrated milk cheese; 351 and 387 mg/100 g of cheese, respectively). Because of the lower pH of the low S:M cheeses, CLC were observed in low S:M cheeses. However, the greatest intensity of CLC was observed in gas-flushed cheeses made with milk containing increased protein concentration because of the increased content of calcium available for CLC formation. These results show that the occurrence of CLC is dependent on cheese milk concentration and pH of the cheese, which can be influenced by S:M and cheese microflora.     

Chymosin-mediated proteolysis, calcium solubilization, and texture development during the ripening of cheddar cheese

Full fat, milled-curd Cheddar cheeses (2 kg) were manufactured with 0.0 (control), 0.1, 1.0, or 10.0 μmol of pepstatin (a potent competitive inhibitor of chymosin) added per liter of curds/whey mixture at the start of cooking to obtain residual chymosin levels that were 100, 89, 55, and 16% of the activity in the control cheese, respectively. The cheeses were ripened at 8°C for 180 d. There were no significant differences in the pH values of the cheeses; however, the moisture content of the cheeses decreased with increasing level of pepstatin addition. The levels of pH 4.6-soluble nitrogen in the 3 cheeses with added pepstatin were significantly lower than that of the control cheese at 1 d and throughout ripening. Densitometric analysis of urea-PAGE electro-phoretograms of the pH 4.6-insoluble fractions of the cheese made with 10.0 μmol/L of pepstatin showed complete inhibition of hydrolysis of α_S1-casein (CN) at Phe₂₃-Phe₂₄ at all stages of ripening. The level of insoluble calcium in each of 4 cheeses decreased significantly during the first 21 d of ripening, irrespective of the level of pepstatin addition. Concurrently, there was a significant reduction in hardness in each of the 4 cheeses during the first 21 d of ripening. The softening of texture was more highly correlated with the level of insoluble calcium than with the level of intact α_S1-CN in each of the 4 cheeses early in ripening. It is concluded that hydrolysis of α_S1-CN at Phe₂₃-Phe₂₄ is not a prerequisite for softening of Cheddar cheese during the early stages of ripening. We propose that this softening of texture is principally due to the partial solubilization of colloidal calcium phosphate associated with the para-CN matrix of the curd.     

Influence of calcium and phosphorus, lactose, and salt-to-moisture ratio on Cheddar cheese quality: pH buffering properties of cheese

The pH buffering capacity of cheese is an important determinant of cheese pH. However, the effects of different constituents of cheese on its pH buffering capacity have not been fully clarified. The objective of this study was to characterize the chemical species and chemical equilibria that are responsible for the pH buffering properties of cheese. Eight cheeses with 2 levels of Ca and P (0.67 and 0.47% vs. 0.53 and 0.39%, respectively), residual lactose (2.4 vs. 0.78%), and salt-to-moisture ratio (6.4 vs. 4.8%) were manufactured. The pH-titration curves for these cheeses were obtained by titrating cheese:water (1:39 wt/wt) dispersions with 1 N HCl, and backtitrating with 1 N NaOH. To understand the role of different chemical equilibria and the respective chemical species in controlling the pH of cheese, pH buffering was modeled mathematically. The 36 chemical species that were found to be relevant for modeling can be classified as cations (Na⁺, Ca²⁺, Mg²⁺), anions (phosphate, citrate, lactate), protein-bound amino acids with a side-chain pKa in the range of 3 to 9 (glutamate, histidine, serine phosphate, aspartate), metal ion complexes (phosphate, citrate, and lactate complexes of Na⁺, Ca²⁺, and Mg²⁺), and calcium phosphate precipitates. A set of 36 corresponding equations was solved to give the concentrations of all chemical species as a function of pH, allowing the prediction of buffering curves. Changes in the calculated species concentrations allowed the identification of the chemical species and chemical equilibria that dominate the pH buffering properties of cheese in different pH ranges. The model indicates that pH buffering in the pH range from 4.5 to 5.5 is predominantly due to a precipitate of Ca and phosphate, and the protonation equilibrium involving the side chains of protein-bound glutamate. In the literature, the precipitate is often referred to as amorphous colloidal calcium phosphate. A comparison of experimental data and model predictions shows that the buffering properties of the precipitate can be explained, assuming that it consists of hydroxyapatite [Ca₅(OH)(PO₄)₃] or Ca₃(PO₄)₂. The pH buffering in the region from pH 3.5 to 4.5 is due to protonation of side-chain carboxylates of protein-bound glutamate, aspartate, and lactate, in order of decreasing significance. In addition, pH buffering between pH 5 to 8 in the backtitration results from the reprecipitation of calcium and phosphate either as CaHPO₄ or Ca₄H(PO₄)₃.     

Impact of milk preacidification with CO2 on the aging and proteolysis of cheddar cheese

To determine the influence of milk preacidification with CO(2) on Cheddar cheese aging and proteolysis, cheese was manufactured from milk with and without added CO(2). The experiment was replicated 3 times. Carbon dioxide (approximately 1600 ppm) was added to the cold milk, resulting in a milk pH of 5.9 at 31 degrees C in the cheese vat. The starter and coagulant usage rates were equal for the control and CO(2) treatment cheeses. The calcium content of the CO(2) treatment cheese was lower, but no difference in moisture content was detected. The higher CO(2) content of the treatment cheeses (337 vs. 124 ppm) was maintained throughout 6 mo of aging. In spite of having almost one and a half times the salt-in-moisture, proteolysis as measured by pH 4.6 and 12% trichloroacetic acid soluble nitrogen expressed as percentages of total nitrogen, was higher in the CO(2) treatment cheeses throughout aging. The ratio of alpha(s)-casein (CN) to para-kappa-CN decreased faster in the CO(2) treatment cheeses than in the control cheeses, especially before refrigerated storage. No difference was detected in the ratio of beta-CN to para-kappa-CN between the control and CO(2) treatment cheeses. Intact alpha(s)- and beta-CN were found in the expressible serum (ES) from the CO(2) treatment cheese as well as alpha(s1)-I-CN, but they were not detected in the ES from the control cheese. No CN was detected in the ES from the curd before the salting of either the control or CO(2) treatment cheese. Higher proteolysis in the cheese made from milk preacidified with CO(2) may have been due to increased substrate availability in the water phase or increased chymosin activity or retention in the cheese.