Effects of Penicillium roqueforti and whey cheese on gross composition,microbiology and proteolysis of mould‐ripened Civil cheese during ripening

Four different types of mould‐ripened Civil cheese were manufactured. A defined (nontoxigenic) strain of a Penicillium roqueforti (SC 509) was used as the secondary starter with and without addition of the whey cheese (Lor); in parallel, secondary starter‐free counterparts were manufactured. Chemical composition, microbiology and proteolysis were studied during the ripening. The incorporation of whey cheese in the manufacture of mould‐ripened Civil cheese altered the gross composition and adversely affected proteolysis in the cheeses. The inoculated P. roqueforti moulds appeared to grow slowly on those cheeses, and little proteolysis was evident in all cheese treatments during the first 90 days of ripening. However, sharp increases in the soluble nitrogen fractions were observed in all cheeses after 90 days. Microbiological analysis showed that the microbial counts in the cheeses were at high levels at the beginning of ripening, while their counts decreased approximately 1–2 log cfu/g towards the end of ripening.     

Transfer of oxytetracycline from ovine spiked milk to whey and cheese

Cheese-making trials with ovine milk spiked with oxytetracycline (OTC) at maximum residue limit (MRL) level were performed to study the OTC distribution between whey and cheese, and assess its effect in cheese-making process and cheese composition. An in-house validated method, based on liquid chromatography-tandem mass spectrometry, was adopted to detect OTC in milk, whey and cheeses. The MRL OTC spike induced a delay in the pH lowering causing a higher moisture content in 1-day OTC cheese compared with the control cheese. No effects of OTC were observed on the mass balance of the process, on the recovery of fat and protein and, in general, on physico-chemical parameters and gross composition of obtained cheeses. OTC was mainly recovered in the 1-day cheeses (61 ± 5%). Despite a 17 ± 8% OTC reduction observed during cheese ripening, probably due to partial degradation, remaining residues were not negligible and could contribute to reach the acceptable daily intake.     

Evolution of chemico-physical characteristics during manufacture and ripening of Castelmagno PDO cheese in wintertime

Biochemical, volatile and textural profiles during manufacture and ripening were determined in samples of Castelmagno PDO cheese obtained from three different batches in the main artisan cheese plant of Castelmagno PDO production area. At the end of manufacture, samples were characterised by a pH of 6.57% and 52.4% moisture content. The HPLC analysis of organic acids and sugars showed the exhaustion of lactose content, while Urea-PAGE indicated extensive primary proteolysis of both β-casein and α_s1-casein. During ripening, cheeses were characterised by high degradation of β-casein and α_s1-casein, due to bacterial action. RP-HPLC profiles showed a high production of peptides eluted between 20 and 30 min. In total, 92 volatile compounds were identified in cheese headspace. Texture profiles showed an increase in hardness, gumminess, chewiness and adhesiveness values, as well as a decrease in cohesiveness during ripening.     

Effect of somatic cell count on Prato cheese composition

The objective of this research was to evaluate the effect of 2 levels of somatic cell counts (SCC) in raw milk on Prato cheese composition, protein and fat recovery, cheese yield, and ripening. A 2 × 6 factorial design with 3 replications was performed in this study: 2 levels of SCC and 6 levels of storage time. Initially, 2 groups of dairy cows were selected to obtain low (<200,000 cells/ mL) and high (>600,000 cells/mL) SCC in milks that were used to manufacture 2 vats of cheese: 1) low SCC and 2) high SCC. Milk, whey, and cheese compositions were evaluated; clotting time was measured; and cheese yield, protein recovery, and fat recovery were calculated. The cheeses were evaluated after 5, 12, 19, 26, 33, and 40 d of ripening according to pH, moisture, pH 4.6 soluble nitrogen, 12% trichloroacetic acid soluble nitrogen as a percentage of total nitrogen, and firmness. High-SCC milk presented significantly higher total protein and nonprotein nitrogen and lower true protein and casein concentrations than did low-SCC milk, indicating an increased whey protein content and a higher level of proteolysis. Although the pH of the milk was not affected by the somatic cell level, the cheese obtained from high-SCC milk presented significantly higher pH values during manufacture and a higher clotting time. No significant differences in cheese yield and protein recovery were observed for these levels of milk somatic cells. The cheese from high-SCC milk was higher in moisture and had a higher level of proteolysis during ripening, which could compromise the typical sensory quality of the product.     

Effect of lactose standardization of milk using low-concentration factor ultrafiltration: Effect of reducing the lactose-to-casein ratio on the properties of milled-curd Cheddar cheese

The pH of cheese is determined by the amount of lactose fermented and the buffering capacity of the cheese. The buffering capacity of cheese is largely determined by the protein contents of milk and cheese and the amount of insoluble calcium phosphate in the curd, which is related to the rate of acidification. The objective of this study was to standardize both the lactose and casein contents of milk to better control final pH and prevent the development of excessive acidity in Cheddar cheese. This approach involved the use of low-concentration factor ultrafiltration of milk to increase the casein content (～5%), followed by the addition of water, ultrafiltration permeate, or both to the retentate to adjust the lactose content. We evaluated milks with 4 different lactose-to-casein ratios (L:CN): 1.8 (control milk), 1.4, 1.1, and 0.9. All cheesemilks had similar total casein (2.3%) and fat (3.4%) contents. These milks were used to make milled-curd Cheddar cheese, and we evaluated cheese composition, texture, functionality, and sensory properties over 9 mo of ripening. Cheeses made from milks with varying levels of L:CN had similar moisture, protein, fat, and salt contents, due to slight modifications during manufacture (i.e., cutting the gel at a smaller size than control) as well as control of acid development at critical steps (i.e., cutting the gel, whey drainage, salting). As expected, decreasing the L:CN led to cheeses with lower lactic acid, residual lactose, and insoluble Ca contents, as well as a substantial pH increase during cheese ripening in cheeses. The L:CN ratio had no significant effect on the levels of primary and secondary proteolysis. Texture profile analysis showed no significant differences in hardness values during ripening. Maximum loss tangent, an index of cheese meltability, was lower until 45 d for the L:CN 1.4 and 0.9 treatments, but after 45 d, all reduced L:CN cheeses had higher maximum loss tangent values than the control cheese (L:CN 1.8). Sensory analyses showed that cheeses made from milks with reduced L:CN contents had lower acidity, sourness, sulfury notes, and chewdown cohesiveness. Standardization of milk to a specific L:CN ratio, while maintaining a constant casein level in the milk, would allow Cheddar cheese manufacturers to have tighter control of pH and acidity.     

Interaction between sodium chloride and texture in semi-hard Danish cheese as affected by brining time,dl-starter culture,chymosin type and cheese ripening

Reduced NaCl in semi-hard cheeses greatly affects textural and sensory properties. The interaction between cheese NaCl concentration and texture was affected by brining time (0–28 h), dl-starter cultures (C1, C2, and C3), chymosin type (bovine or camel), and ripening time (1–12 weeks). Cheese NaCl levels ranged from <0.15 to 1.90% (w/w). NaCl distribution changed during ripening; migration from cheese edge to core led to a more homogeneous NaCl distribution after 12 weeks. As ripening time increased, cheese firmness decreased. Cheeses with reduced NaCl were less firm and more compressible. Cheeses produced with C2 were significantly firmer than those produced with C1; cheeses produced with C3 had higher firmness and compressibility. In NaCl reduced cheese, use of camel chymosin as coagulant resulted in significantly higher firmness than that given using bovine chymosin. Overall, cheese NaCl content is reducible without significant textural impact using well-defined starter cultures and camel chymosin.     

Residual clotting activity and ripening properties of vegetable rennet from Cynara cardunculus in La Serena cheese

Vegetable rennet extracted from Cynara cardunculus flowers is traditionally used in the manufacture of La Serena cheese. High levels of proteolytic enzymes of the flowers are responsible for its clotting activity and strong proteolytic action. The presence of residual coagulant in cheese and whey was measured by adding known amounts of vegetable rennet as internal standard. We found no differences between the residual coagulant activity of La Serena cheese compared with other types of cheese. The coagulant content detected at the end of four cheesemakings (vat of 830 l) in cheese and whey represented 27 and 78%, respectively, of the total amount added to milk. When measurements were carried out in 16 different cheeses, vegetable rennet appeared to be highly stable during cheese ripening. Cheese composition (moisture, pH, NaCl, fat and protein) was kept relatively constant during ripening, which seems to contribute to stability of residual activity. Electrophoretic analyses of water insoluble fractions from cheeses manufactured with vegetable rennet showed that α_s-casein was less susceptible to proteolysis than β-casein. The water soluble nitrogen/total nitrogen (WSN/TN) exhibited higher levels only during the first 30 days of ripening although non-protein nitrogen/total nitrogen (NPN/TN) ratio and amino acid nitrogen (NH₂-N) increased with ripening time.     

Rheological and calcium equilibrium changes during the ripening of Cheddar cheese

The small deformation rheological properties and the calcium (Ca) equilibrium of Cheddar cheese were investigated as a function of ripening time. The proportion of insoluble Ca as a percentage of the total Ca decreased from ∼72 to 57% between 3 days and 9 months; most changes occurred within the first 4 weeks. During ripening, the storage modulus (G′) of the cheese increased at low temperature, but decreased rapidly at high temperature. At temperatures >40 °C, the loss tangent increased to reach a maximum at a temperature of ∼70 °C in young cheese and there was a steady decline in this temperature during ripening. The maximum loss tangent values increased substantially during the first 4 weeks and then showed little change. Changes in the insoluble Ca content significantly correlated with pH 4.6 soluble nitrogen (pH 4.6 SN). Partial correlation analysis indicated that the insoluble Ca content was more significantly correlated with the rheological properties than was pH 4.6 SN.     

Physicochemical analysis of full-fat, reduced-fat, and low-fat artisan-style goat cheese

The objective of this study was to examine the physicochemical properties of cheese elaborated via traditional artisan methods using goat milk containing 5, 1.5, or 0.4% fat and ripened for 1, 7, 14, or 28 d. Seventy-two cheeses were produced (2 batches × 3 fat levels × 4 ripening times × triplicate). Proximal composition, pH, texture analysis, and color were recorded in each cheese. Protein and moisture were increased in cheese, and fat and fat in DM were decreased with decreasing fat in milk. Internal and external pH was higher in low-fat and reduced-fat cheese, and pH values decreased during the first 2 wk of ripening but increased slightly on d 28. Cheese fracturability, cohesiveness, masticability, and hardness increased with decreasing fat, whereas elasticity and adhesiveness decreased. Cheese lightness and red and yellow indexes decreased with decreasing fat content; during ripening, lightness decreased further but yellow index increased.     

Acceleration of cheese ripening at elevated temperature. An estimation of the optimal ripening time of a traditional Argentinean hard cheese

The effect of elevated ripening temperature on physicochemical, biochemical and sensory characteristics in Reggianito Argentino cheese was evaluated to determine the optimal time for cheese ripening at 18 °C that ensures typical cheese characteristics. Cheeses ripened at 12 or 18 °C and 85% relative humidity were analysed at 2, 4 and 6 months. Seventy-eight variables (as determined by urea-PAGE, RP-HPLC of the water-soluble at pH 4.6 fraction, free amino acids, free fatty acids and sensory analysis) were considered for the principal component analysis. The statistical analysis allowed determination of the optimal time for ripening Reggianito Argentino cheese at 18 °C, which was ranged between 2 and 3 months. In conclusion, the results obtained were not only useful in characterising the ripening of an Argentinean hard cheese, but also in evaluating the effect of an increase of ripening temperature on the main physicochemical, biochemical and sensory changes of Reggianito Argentino cheese.     

The effect of lowering salt on the physicochemical,microbiological and sensory properties of São João cheese of Pico Island

São João cheeses with varying curd dry salting treatments were made with decreasing levels of salt (sodium chloride): 4 (control), 3, 2 and 1% (w/w), along with the salt‐free version. The cheeses were ripened at 11 °C over a 40‐day period, and the effect of lowering salt on the physicochemical, microbiological and sensory properties of the cheese was studied. Reduced salt resulted in a concomitant moisture decrease with protein increase, ash and sodium reduction among experimental cheeses at the same ripening day, but there were no significant differences in pH, acidity and fat, or in the microbiological quality. Triangle tests indicated perceptible differences between test and control cheeses at the level of 2% NaCl (w/w) or less, but not with the cheese salted with 3 g NaCl/100 g. Considering the sensory, the physicochemical and the microbiological results, the cheese formulated at 3% NaCl (w/w) (presenting a reduction of 25% in salt) is feasible on an industrial scale, being indistinguishable by the regular consumer.     

A sensitive HPLC method to detect hen's egg white lysozyme in milk and dairy products

Because of the lytic activity on the cell wall of bacteria like Clostridium tyrobutyricum, hen's egg white lysozyme is used in cheese manufacturing to prevent late blowing. A HPLC method capable to quantify as low as 0.8 ppm lysozyme in milk and cheese is proposed. Lysozyme was extracted with 1 m NaCl at pH 6.0 and the extract was deproteinized at low pH values, reaching a recovery up to 90%. Reversed-phase HPLC was performed on a polymeric column and monitoring lysozyme under fluorescence detection (excitation at 280 nm and emission at 340 nm). The repeatability of this determination tested on a 15-month-aged hard cheese and expressed as relative standard deviation was 1.47 (n=6) and no interference of peptides formed during ripening was observed. Four commercial preparations of egg white lysozyme gave a similar fluorescence response and the partitioning over cheese and whey was studied with one of them. About 80% of the lysozyme added to the cheesemilk at concentrations up to 80 ppm was retained in the cheese and the concentration factor of lysozyme from the cheesemilk to the cheese proved to be 8.2 on average. This HPLC method and the microbiological assay using Micrococcus luteus were compared in cheese analysis, proving the former to be more accurate and reliable than the latter. Eighteen commercial samples including both generic cheeses and cheeses having protected designation of origin, all of them not declared to contain lysozyme, showed concentrations of this enzyme ranging from 0 to 111 ppm.     

The effects of salt and pH stress on the growth rates of persistent strains of Listeria monocytogenes collected from specific ecological niches

This study aimed to investigate the growth responses of persistent strains of Listeria monocytogenes to NaCl (5–10%) and pH (4–8.5) stress. Seven isolates of two persistent strains, collected from cheeses with 7 and 42 days of ripening, and from the shelf of one maturation room, and two reference strains were analysed. The experiments were carried out, in Tryptic Soy Broth (TSB), at 37 °C. Response surface methodology (RSM) was used to compare the effects of the selected variables on the growth rate of the isolates. Persistent strains showed, in general, higher growth rates than reference strains. The factor NaCl was highly significant to the isolate collected from the shelf. There was a different growth response between isolates from cheese with 7 and 42 days of ripening. These results suggest the adaptation of L. monocytogenes to specific environments within the dairies.     

Effect of the use of carbon dioxide on Prato cheese making

The effects of the addition of carbon dioxide (CO₂) under pressure (1.6 × 10⁵ Pa at 8 °C) to pasteurised Prato cheese milk (pH 6.0) was investigated through 120 d of refrigerated storage. The addition of CO₂ decreased the curd formation time (30 min), the total manufacturing time (47 min), and the pH of Prato cheese, thus leading to reduced moisture content. The CO₂ treated cheese showed higher firmness and fracturability due to the greater whey loss. In contrast, the microorganism counts, cheese yield, protein loss, cohesiveness, springiness, and gumminess were not significantly affected by the treatment. For the lactose fermentation, no significant differences were observed. The addition of CO₂ did not change the proteolysis indexes, and no significant differences were observed in the sensory acceptance of the CO₂ treated cheese, which was well accepted by consumers.     

FATE OF AFLATOXIN M1 IN KASHAR CHEESE

The distribution and stability of aflatoxin M₁ (AFM₁) in Kashar cheese were investigated. Raw milk samples were spiked with AFM₁ at the levels of 50, 250 and 750 ng/L. Distribution of toxin in milk, cheese curd, whey, kneading brine and cheese, and its stability during ripening were determined by high‐performance liquid chromatography. Concentrations of AFM₁ in curds for each contamination level were 2.93, 3.19 and 3.37 times higher than those in milk. After syneresis, the percentage distribution of AFM₁ was 40–46% in curds and 53–58% in whey indicating that relatively higher concentration of toxin passed to whey. Moreover, by the kneading process approximately 2–5% of AFM₁ passed to kneading brine. Compared to the initial spiking level, the percentage of toxin in cheeses varied between 35–42%. Over a 60‐day storage period, there was no decrease in the concentration of AFM₁, suggesting that the toxin was stable during ripening.     

Microbiological,compositional, biochemical and textural characterisation of Caciocavallo Pugliese cheese during ripening

A microbiological, compositional, biochemical and textural characterisation of the pasta filata Caciocavallo Pugliese cheese during ripening is reported. Fully ripened cheese contained a total of ca. log 8.0 cfu g⁻¹ mesophilic bacteria and ca. log 6.0 cfu g⁻¹ presumptive staphylococci, while the number of thermophilic and mesophilic rod and coccus lactic acid bacteria varied during ripening. A two-step RAPD-PCR protocol was used to differentiate biotypes. The natural whey starter was composed mainly of Lactobacillus delbrueckii, Lb. fermentum, Lb. gasseri, Lb. helveticus and Streptococcus thermophilus strains. After day 1 of ripening, Lb. delbrueckii became dominant and some strains of Enterococcus durans and E. faecalis appeared. Non-starter lactic acid bacteria, such as Lb. parabuchneri and Lb. paracasei subsp. paracasei formed a large part of the lactic microflora at 42 and 60 d of ripening. The level of pH 4.6-soluble nitrogen increased from the outer to the inner of the cheese and also increased in each section as ripening progressed, attaining values of 18–15%. Urea-PAGE electrophoresis showed that degradation of α_s1-casein was more rapid than that of β-casein throughout ripening and the rates at which both caseins were degraded greatly increased from the outside to the inside of the cheese. Based on the primary proteolysis products, both chymosin and plasmin appeared to be active. RP-HPLC profiles of the 70% ethanol-soluble, pH 4.6-soluble nitrogen, showed a large number of peaks, indicating a heterogeneous mixture of proteolytic products. There were both age- and section-related changes in the area of the different peptide peaks. Butyric (C4:0), caproic (C6:0), palmitic (C16:0) and oleic (C18:1) acids were the free fatty acids found at the highest concentrations. The level of short chain fatty acids (e.g., butyric and caproic) decreased from the middle and inner to outer sections of the cheese. Peptidase activity in the curd was pronounced, increased during ripening and varied with the cheese section. The greatest increase of the peptidase activity coincided with a change in the lactic microflora and with the prevalence of non-starter lactic acid bacteria. Microbial esterases were supposed to be active together with rennet paste. Little change in the firmness and fractures stress during maturation were found by textural analyses of the raw cheese. The flowability was similar to that of typical low-moisture Mozzarella cheese, while stretchability was lower. The heat-induced changes in phase angle of Caciocavallo Pugliese cheese indicated a phase transition from largely elastic rheological characteristics in unheated cheese to a more viscous and fluid character in melted cheese.     

Texture and flavour development in Ras cheese made from raw and pasteurised milk

Texture, proteolysis and flavour development in Ras cheeses made from raw or pasteurised milk with two different thermophilic lactic cultures were monitored during ripening. Results showed that at day 1 of manufacture, the moisture content and pH were lower in raw milk cheese than in pasteurised milk cheeses. Levels of water-soluble nitrogen, casein breakdown, free amino groups and free fatty acids were higher in cheese made from raw milk than in that made from pasteurised milk. Textural characteristics, such as hardness, cohesiveness and chewines, increased in all treatments during the first 60 days of ripening due to the reduction in the moisture level during the second stage of salting (dry salting during the first 60 days of ripening). Cheese made from raw milk received the highest texture and flavour scores by panellists.     

Insoluble calcium content and rheological properties of Colby cheese during ripening

Colby cheese was made using different manufacturing conditions (i.e., varying the lactose content of milk and pH values at critical steps in the cheesemaking process) to alter the extent of acid development and the insoluble and total Ca contents of cheese. Milk was concentrated by reverse osmosis (RO) to increase the lactose content. Extent of acid development was modified by using high (HPM) and low (LPM) pH values at coagulant addition, whey drainage, and curd milling. Total Ca content was determined by atomic absorption spectroscopy, and the insoluble (INSOL) Ca content of cheese was measured by the cheese juice method. The rheological and melting properties of cheese were measured by small amplitude oscillatory rheometry and UW-Melt Profiler, respectively. There was very little change in pH during ripening even in cheese made from milk with high lactose content. The initial (d 1) cheese pH was in the range of 4.9 to 5.1. The INSOL Ca content of cheese decreased during the first 4 wk of ripening. Cheeses made with the LPM had lower INSOL Ca content during ripening compared with cheese made with HPM. There was an increase in melt and maximum loss tangent values during ripening except for LPM cheeses made with RO-concentrated milk, as this cheese had pH <4.9 and exhibited limited melt. Curd washing reduced the levels of lactic acid produced during ripening and resulted in significantly higher INSOL Ca content. The use of curd washing for cheeses made from high lactose milk prevented a large pH decrease during ripening; high rennet and draining pH values also retained more buffering constituents (i.e., INSOL Ca phosphate), which helped prevent a large pH decrease.     

Behaviour of Listeria monocytogenes in Lighvan cheese following artificial contamination during making,ripening and storage in different conditions

This study investigated the behaviour and fate of Listeria monocytogenes at different ripening temperatures and NaCl concentrations in traditional Lighvan cheese. L. monocytogenes was added to raw sheep's milk. After producing the cheese, they were stored in 8%, 12% and 15% NaCl at 4, 9 and 14 °C. Sampling was performed for 150 days. Different temperature and NaCl concentrations had a significant effect on the survival of L. monocytogenes (P < 0.001). The lowest growth and survival rates of L. monocytogenes were in 15% NaCl at 14 °C and 12% NaCl at 14 °C, respectively. Also, the highest growth and survival rates of the bacterium were in 8% NaCl at 4 °C.