Short communication: Inactivation of microbial contaminants in raw milk La Serena cheese by high-pressure treatments

La Serena cheese, a Spanish variety made from Merino ewes’ raw milk, has a high pH value, low salt content, and high moisture, conditions that are all favorable for growth and survival of contaminating microorganisms, including pathogens. To improve its microbiological quality and safety, high-pressure treatments at 300 or 400 MPa for 10 min at 10°C were applied to 2 batches of La Serena cheese on d 2 or 50 of ripening. Cheese treated on d 2 at 300 MPa showed viable aerobic counts that were 0.99 log units lower than those for control cheese on d 3 and showed counts of enterococci, coagulase-positive staphylococci, gram-negative bacteria, and coliforms that were 2.05, 0.49, 3.14, and 4.13 log units lower, respectively, than control cheese. For cheese treated on d 2 at 400 MPa, the respective reductions in counts were 2.02, 2.68, 1.45, 3.96, and 5.50 log units. On d 60, viable aerobic counts in cheese treated on d 50 at 300 MPa were 0.50 log units lower than those in control cheese, and counts of enterococci, gram-negative bacteria, and coliforms were 1.37, 2.30, and 4.85 log units lower, respectively. For cheese treated on d 50 at 400 MPa, the respective reductions in counts were 1.29, 1.98, 4.47, and > 5 log units. High-pressure treatments at 300 or 400 MPa on d 2 or 50 reduced significantly the counts of undesirable microorganisms, improving the microbiological quality and safety of La Serena cheese immediately after treatment and at the end of the ripening period.     

High pressure treatment decelerates the lipolysis in a caprine cheese

Garrotxa cheese, a cheese made of goat's milk, typical of the Catalonian region, has been high pressure-treated (400 MPa, 5 min, 14 °C) to accelerate the ripening. The volatile fraction of Garrotxa cheese was studied on treated and untreated samples at two ripening stages by SDE extraction with dichloromethane, and identified with GC–MS. Pressure treatment at 400 MPa for 5 min decelerated the lipolysis, having treated cheese lower amount of free fatty acids, and, in general, less volatile compounds showing a tendency to decrease differences during ripening. No new volatile compounds could be identified after pressure treatment. Humidity and content of non-casein nitrogen and non-protein were higher on pressurised cheese, but bacterial counts were significantly reduced after pressure treatment. High pressure-treated Garrotxa cheese was not substantially equivalent to the regular-ripened cheese, however the treatment presented in this paper could lead to new varieties of cheese.     

Volatile compounds, odor, and aroma of La Serena cheese high-pressure treated at two different stages of ripening

La Serena cheeses made from raw Merino ewe's milk were high-pressure (HP) treated at 300 or 400 MPa for 10 min on d 2 or 50 after manufacture. Ripening of HP-treated and control cheeses proceeded until d 60 at 8°C. Volatile compounds were determined throughout ripening, and analysis of related sensory characteristics was carried out on ripe cheeses. High-pressure treatments on d 2 enhanced the formation of branched-chain aldehydes and of 2-alcohols except 2-butanol, but retarded that of n-aldehydes, 2-methyl ketones, dihydroxy-ketones, n-alcohols, unsaturated alcohols, ethyl esters, propyl esters, and branched-chain esters. Differences between HP-treated and control cheeses in the levels of some volatile compounds tended to disappear during ripening. The odor of ripe cheeses was scarcely affected by HP treatments on d 2, but aroma quality and intensity scores were lowered in comparison with control cheese of the same age. On the other hand, HP treatments on d 50 did not influence either the volatile compound profile or the sensory characteristics of 60-d-old cheese.     

High-pressure processing of Gorgonzola cheese: influence on Listeria monocytogenes inactivation and on sensory characteristics

The presence of Listeria monocytogenes on the rind of Gorgonzola cheese is difficult to avoid. This contamination can easily occur as a consequence of handling during ripening. The aims of this study were to determine the efficiency of high-pressure processing (HPP) for inactivation of L. monocytogenes on cheese rind and to evaluate the influence of HPP treatments on sensory characteristics. Gorgonzola cheese rinds, after removal, were inoculated (about 7.0 log CFU/g) with L. monocytogenes strains previously isolated from other Gorgonzola cheeses. The inoculated cheese rinds were processed with an HPP apparatus under conditions of pressure and time ranging from 400 to 700 MPa for 1 to 15 min. Pressures higher than 600 MPa for 10 min or 700 MPa for 5 min reduced L. monocytogenes more than 99%. A reduction higher than 99.999% was achieved pressurizing cheese rinds at 700 MPa for 15 min. Lower pressure or time treatments were less effective and varied in effectiveness with the cheese sample. Changes in sensory properties possibly induced by the HPP were evaluated on four different Gorgonzola cheeses. A panel of 18 members judged the treated and untreated cheeses in a triangle test. Only one of the four pressurized cheeses was evaluated as different from the untreated sample. HPP was effective in the reduction of L. monocytogenes on Gorgonzola cheese rinds without significantly changing its sensory properties. High-pressure technology is a useful tool to improve the safety of this type of cheese.     

Changes in water binding in high-pressure treated cheese,measured by TGA (thermogravimetrical analysis)

Garrotxa cheese was pressurised at 400 MPa for 5 min to shorten ripening accordingly to previous studies. Moisture content was followed along ripening. Moisture in cheese was divided into free and bound water according to the weight loss rate during drying in a TGA oven. High-pressure treated cheese to shorten ripening time retained more moisture than control cheese. Bound water remained unaffected by the pressure treatment, whereas free water was higher in treated cheese during ripening. Free water content was linked with a_w.     

High-pressure effects on the microstructure, texture, and color of white-brined cheese

Abstract: White‐brined cheeses were subjected to high‐pressure processing (HPP) at 50, 100, 200, and 400 MPa at 22 °C for 5 and 15 min and ripened in brine for 60 d. The effects of pressure treatment on the chemical, textural, microstructural, and color were determined. HPP did not affect moisture, protein, and fat contents of cheeses. Similar microstructures were obtained for unpressurized cheese and pressurized cheeses at 50 and 100 MPa, whereas a denser and continuous structure was obtained for pressurized cheeses at 200 and 400 MPa. These microstructural changes exhibited a good correlation with textural changes. The 200 and 400 MPa treatments resulted in significantly softer, less springy, less gummy, and less chewy cheese. Finally, marked differences were obtained in a* and b* values at higher pressure levels for longer pressure‐holding time and were also supported by ΔE* values. The cheese became more greenish and yellowish with the increase in pressure level. Practical Application: The quality of cheese is the very important to the consumers. This study documented the pressure‐induced changes in selected quality attributes of semisoft and brine‐salted cheese. The results can help the food processors to have knowledge of the process parameters resulting in quality changes and to identify optimal process parameters for preserving pressure‐treated cheeses.     

Proteolysis and biogenic amine buildup in high-pressure treated ovine milk blue-veined cheese

Penicillium roqueforti plays an important role in the ripening of blue-veined cheeses, mostly due to lactic acid consumption and to its extracellular enzymes. The strong activity of P. roqueforti proteinases may bring about cheese over-ripening. Also, free amino acids at high concentrations serve as substrates for biogenic amine formation. Both facts result in shorter product shelf-life. To prevent over-ripening and buildup of biogenic amines, blue-veined cheeses made from pasteurized ovine milk were high-pressure treated at 400 or 600 MPa after 3, 6, or 9 wk of ripening. Primary and secondary proteolysis, biogenic amines, and sensory characteristics of pressurized and control cheeses were monitored for a 90-d ripening period, followed by a 270-d refrigerated storage period. On d 90, treatments at 400 MPa had lowered counts of lactic acid bacteria and P. roqueforti by less than 2 log units, whereas treatments at 600 MPa had reduced lactic acid bacteria counts by more than 4 log units and P. roqueforti counts by more than 6 log units. No residual α-casein (CN) or κ-CN were detected in control cheese on d 90. Concentrations of β-CN, para-κ-CN, and γ-CN were generally higher in 600 MPa cheeses than in the rest. From d 90 onwards, hydrophilic peptides were at similar levels in pressurized and control cheeses, but hydrophobic peptides and the hydrophobic-to-hydrophilic peptide ratio were at higher levels in pressurized cheeses than in control cheese. Aminopeptidase activity, overall proteolysis, and free amino acid contents were generally higher in control cheese than in pressurized cheeses, particularly if treated at 600 MPa. Tyramine concentration was lower in pressurized cheeses, but tryptamine, phenylethylamine, and putrescine contents were higher in some of the pressurized cheeses than in control cheese. Differences in sensory characteristics between pressurized and control cheeses were generally negligible, with the only exception of treatment at high pressure level (600 MPa) at an early ripening stage (3 wk), which affected biochemical changes and sensory characteristics.     

Effect of various high-pressure treatments on the properties of reduced-fat Cheddar cheese

A major problem with reduced-fat cheese is the difficulty in attaining the characteristic flavor and texture of typical full-fat versions. Some previous studies have suggested that high hydrostatic pressure (HHP) can accelerate the ripening of full-fat cheeses. Our objective was to investigate the effect of HHP on reduced-fat (~7.3% fat) Cheddar cheese, with the goal of improving its flavor and texture. We used a central composite rotatable design with response surface methodology to study the effect of pressure and holding time on the rheological, physical, chemical, and microbial characteristics of reduced-fat Cheddar cheese. A 2-level factorial experimental design was chosen to study the effects of the independent variables (pressure and holding time). Pressures were varied from around 50 to 400 MPa and holding times ranged from 2.5 to 19.5 min. High pressure was applied 1 wk after cheese manufacture, and analyses were performed at 2 wk, and 1, 3, and 6 mo. The insoluble calcium content as a percentage of total Ca in cheeses were not affected by pressure treatment. Pressure applications ≥225 MPa resulted in softer cheese texture during ripening. Pressures ≥225 MPa increased melt, and resulted in higher maximum loss tangent values at 2 wk. Pressure treatment had a greater effect on cheese microbial and textural properties than holding time. High-pressure-treated cheeses also had higher pH values than the control. We did not observe any significant difference in rates of proteolysis between treatments. In conclusion, holding times of around 5 min and pressures of ≥225 MPa could potentially be used to improve the excessively firm texture of reduced-fat cheese.     

Effects of high-pressure treatment on free fatty acids release during ripening of ewes' milk cheese

The free fatty acid (FFA) profile of high pressure treated ewes' milk cheeses were studied to assess the effect of pressure treatment on cheese lipolysis. Cheeses were treated at 200, 300, 400 or 500 MPa (2P to 5P) at two stages of ripening (after 1 and 15 days of manufacturing; P1 and P15) and FFA were assayed at 1, 15 and 60 d ripening. On the first day of ripening, 3P1-cheeses showed levels of FFA twice that of the control cheeses. However, no significant differences were found between 3P1 and control cheeses at 60 d ripening. On the contrary, 4P1 and 5P1-cheeses had the lowest total FFA levels. The point at which pressure treatment was applied influenced the FFA profile of cheeses; cheeses pressurized at pressures<400 MPa on the first day of ripening were more similar to untreated cheeses than their homologues treated at 15 d.     

Changes in textural, microstructural, and colour characteristics during ripening of cheeses made from raw, pasteurized or high-pressure-treated goats’ milk

Goats’ milk cheeses were made from raw (RA), pasteurized (PA; 72°C, 15 s) or pressure-treated (PR; 500 MPa, 15 min, 20°C) milk to compare textural, microstructural, and colour characteristics in relation to ripening time. Texture, microstructure and colour were evaluated by uniaxial compression and stress relaxation tests, confocal laser scanning microscopy and Hunter colorimetry, respectively.Raw and PR cheeses were firmer and less fracturable than PA cheese, but differences became less notable toward the end of ripening. PA and PR cheeses were less cohesive than RA cheese. Although cheeses exhibited a loss of elastic characteristics with ageing, PR cheese showed the most elastic behaviour initially. Confocal laser scanning micrographs displayed PR cheese with a regular and compact protein matrix, with small and uniform fat globules resembling the structure of RA cheese. Finally, colour evaluation demonstrated significant differences between cheeses due to milk treatments and ripening time.     

Effect of high pressure combined with mild heat or nisin on inoculated bacteria and mesophiles of goat's milk fresh cheese

The effect of pressures ranging from 400 to 500 MPa combined with mild heat on Staphylococcus carnosus inoculated in fresh cheese and the concurrent use of 500 MPa and nisin to inactivate cheese indigenous populations has been studied. Staphylococcus carnosus counts could not be substantially decreased with treatments at 500 MPa at 10 or 25°C for 30 min, whereas treatments at 50°C for 5 min caused a reduction of 7-log₁₀cfu g⁻¹. Multiple-cycle treatments of 500 MPa and times between 15 and 30 min also improved the inactivation rate. Combination of 500 MPa and nisin was the most effective treatment to inactivate cheese indigenous microbiota. Inactivation of Bacillus subtilis spores inoculated in fresh cheese has also been studied. Germination treatments of 60 MPa at 40°C for 210 min followed by vegetative cells inactivation treatments of 500 MPa at 40°C for 15 min caused a lethality of 4·9-log₁₀ofB. subtilis , whereas the same combination of treatments applied at 25°C only caused a 2·7-log₁₀reduction.     

Effect of high-pressure treatments on proteolysis and texture of ewes’ raw milk La Serena cheese

La Serena cheeses, made from Merino ewes’ raw milk, were high-pressure (HP)-treated at 300 or 400 MPa for 10 min at 10 °C, on days 2 or 50 of ripening. Cheeses treated by HP on day 2 showed higher pH values than control cheese on day 3, but cheeses treated by HP on days 2 or 50 and control cheese had similar pH values on day 60. Breakdown of caseins was delayed by HP treatment of cheeses on day 2. Cheeses treated by HP on day 2 showed higher levels of hydrophilic peptides, lower levels of hydrophobic peptides, lower hydrophobic peptides: hydrophilic peptides ratios, and higher total contents of free amino acids than those of control cheese. HP treatment of cheese on day 50 scarcely affected proteolysis of 60-day-old cheeses. Fracturability, hardness and elasticity values of cheeses treated by HP on day 2 were higher than those of control cheese and of cheeses treated on day 50. Cheeses treated at 400 MPa on day 2 received the lowest scores for quality of taste from panellists, whereas the rest of HP-treated cheeses did not differ from control cheese.     

Rheological properties of high-pressure-treated Gouda cheese

The rheological properties of high-pressure-treated (50–400 MPa, 1 h) and untreated Gouda cheese were compared. Immediately after pressure release, oscillation measurements gave lower storage and loss moduli from 50 MPa onwards. Simultaneously, tan δ was higher, indicating a relatively less solid-like behaviour of the pressurized samples. Creep measurements showed that samples treated at 400 MPa got less rigid, less solid-like, and more viscoelastic; from 50 MPa onwards, the samples had less resistance to flow at longer times. Texture profile analysis revealed that samples treated at 225 and 400 MPa showed no macroscopical breakage. Relaxation measurements gave a higher level of stress decay at long relaxation times and a higher rate at which the stress relaxes. During further ripening after pressure release, differences between pressure-treated and untreated samples became smaller. At 42 days of ripening, any or only a slight difference could still be observed. Dissolution experiments showed that hydrophobic interactions in Gouda cheese were weakened by pressure treatment. This possibly led to structural changes of the paracasein network causing the rheological property changes. These pressure effects on proteins in Gouda cheese are possibly reversible as hydrophobic interactions and rheological properties were restored during ripening.     

Texture, proteolysis and viable lactic acid bacteria in commercial cheddar cheeses treated with high pressure

High pressure processing was investigated for controlling Cheddar cheese ripening. One-month-or 4-month-old Cheddar cheeses were subjected to pressures ranging from 200 to 800 MPa for 5 min at 25 C. The number of viable Lactococcus lactis (starter) and Lactobacillus (nonstarter) cells decreased as pressure increased. Subsequent storage of the control and pressure-treated cheeses at 10 degrees C caused viable cell counts to change in some cases. Free amino acid content was monitored as an indicator of proteolysis. Cheeses treated with pressures > or = 400 MPa evolved free amino acids at significantly lower rates than the control. No acceleration in free amino acid development was observed at lower pressures. Pressure treatment did not accelerate the rate of textural breakdown compared with the non-pressure treated control. On the contrary, pressure treatment at 800 MPa reduced the time-dependent texture changes. Results indicate that high pressure may be useful in arresting Cheddar cheese ripening.     

Changes of composition and free fatty acid contents of Urfa cheeses (a white-brined Turkish cheese) during ripening: Effects of heat treatments and starter cultures

The influences of heat treatments (at 65 °C for 20 min or 72 °C for 5 min) applied to the milk and addition of mesophilic or thermophilic starter cultures, prior to cheese-making, on the composition and free fatty acid contents of Urfa cheeses were evaluated throughout the ripening period. Sensory evaluation of cheese samples was also performed on 90th day. The basic composition of ripened cheese samples was not significantly affected by the heat treatments and starter cultures. Heat treatments adversely affected the lipolysis and sensory properties of Urfa cheeses, particularly at 72 °C. The FFA contents of cheeses made from mesophilic and thermophilic cultures were similar. Cheese made from raw milk had a higher level of lipolysis than the cheeses made from milk inoculated with mesophilic or thermophilic lactic starters (p < 0.05).     

Behavior of Yersinia enterocolitica strains inoculated in model cheese treated with high hydrostatic pressure

The effects of high hydrostatic pressure treatment and the ability for survival, repair, and growth of three human pathogenic serotypes (O:1, O:3, O:8) of Yersinia enterocolitica were investigated in washed-curd model cheese made with pasteurized bovine milk. Samples were treated at 300, 400, and 500 MPa for 10 min at 20 degrees C and analyzed at 0, 1, 7, and 15 days to assess the viability of the Yersinia population. A long-term study (up to 60 days of ripening after high hydrostatic pressure treatment) was also undertaken. Treatments at 400 and 500 MPa caused maximum lethality, and only the treatment at 300 MPa showed significant differences (P < 0.05) between serotypes; the most baroresistant was O:3. Ability to repair and grow was not observed after 15 days of storage at 8 degrees C. Yersinia counts in untreated cheese samples also decreased below the detection limit at day 45 in the long-term study. These results suggest that the cheese environment did not allow recovery of injured cells or growth. A primary contributing factor to this effect seemed to be the low pH resulting from the production of lactic acid during cheese ripening.     

Effect of cheese water activity and carbohydrate content on the barotolerance of Listeria monocytogenes scott A

High-pressure processing is an appropriate technique for improving the microbiological safety of packaged ready-to-eat foods. The effect of high-pressure treatment on Listeria monocytogenes Scott A inoculated into fresh Hispánico-type cheese and ripe Mahón cheese was investigated. A 3.8-log reduction in the counts of L. monocytogenes Scott A in fresh cheese was recorded after 3 min at 400 MPa and 12 degrees C, whereas 18 min under the same conditions was required to obtain a 1-log reduction in ripe cheese. Dry matter values were 48.96% for fresh cheese and 58.79% for ripe cheese, and water activity (aw) values were 0.983 and 0.922, respectively. In dehydrated fresh cheese (58.20% dry matter) in which 5% NaCl was added to achieve a 0.904 aw value, L. monocytogenes Scott A counts were lowered by only 0.4 log after treatment for 10 min at 400 MPa. On the other hand, in a 60:40 mixture of ripe cheese:distilled water with a 0.976 aw value, the reduction under the same conditions was 3.9 log. Within the aw range of 0.945 to 0.965, L. monocytogenes Scott A barotolerance was significantly higher in fresh cheese than in ripe cheese for equivalent aw values. Carbohydrate content was higher in fresh cheese than in ripe cheese. The addition of lactose at a concentration of 5 mg/g to an 85:15 mixture of ripe cheese:distilled water did not influence L. monocytogenes Scott A barotolerance during treatment for 10 min at 400 MPa. Galactose at a concentration of 5 mg/g had a protective effect during high-pressure treatment, and glucose at a concentration of 5 mg/g favored L. monocytogenes Scott A survival during refrigerated storage of pressurized samples at 8 degrees C for 5 days.     

Use of high pressure treatment to attenuate starter bacteria for use as adjuncts for Cheddar cheese manufacture

Attenuated starter bacteria cannot produce acid during cheese manufacture, but contain enzymes that contribute to cheese ripening. The aim of this study was to investigate attenuation of starter bacteria using high pressure treatment, for use in combination with a primary starter for Cheddar cheese manufacture, and to determine the effect of such adjunct cultures on secondary proteolysis during ripening. Lactococcus lactis ssp. cremoris HP and L. lactis ssp. cremoris 303 were attenuated by pressure treatment at 200 MPa for 20 min at 20 °C. Cheddar cheese was manufactured using untreated cultures of both these starter strains, either alone or in combination with their high pressure-treated equivalents. High pressure-treated starters did not produce acid during cheese manufacture and starter counts in cheeses manufactured using high pressure-treated starter did not differ from those of the controls. Higher levels of cell lysis were apparent in cheese manufactured using high pressure-treated strains than in the controls after 26 d of ripening. Small differences were observed in the peptide profiles of cheeses, analysed by reversed-phase HPLC; cheeses manufactured using high pressure-treated starters also had slightly higher levels of amino acids than the relevant controls. Overall, addition of high pressure-treated starter bacteria as a secondary starter culture accelerated secondary proteolysis in Cheddar cheese.

Industrial relevance

Attenuated starters provide extra pool of enzymes, which can influence cheese ripening, without affecting the cheese making schedule. This paper presents an alternative method for attenuation of starter bacteria using high pressure treatment and their subsequent use to accelerate secondary proteolysis in Cheddar cheese during ripening.     

Effect of a bacteriocin-producing Lactococcus lactis strain and high-pressure treatment on the esterase activity and free fatty acids in Hispánico cheese

The effect of milk inoculation with a bacteriocin-producing (BP) culture and of high-pressure (HP) treatment of 15-day-old Hispánico cheeses (400 MPa, 5 min, 10 °C), separately or combined, on the release of intracellular esterases and cheese lipolysis was investigated. Esterase activity and free fatty acids (FFAs) content increased during ripening of Hispánico cheese and palmitic, oleic and stearic acids being the most abundant FFAs. On day 15, the highest esterase activity was recorded for HP-treated BP cheese. The activity for HP-untreated BP cheese was the next highest. No difference in the activities was found between HP-treated and untreated cheeses made without BP culture. Total FFAs on day 15 were at a lower concentration in BP cheeses than in cheeses made without BP culture, probably due to the lower pH values of the former. The rate of total FFA accumulation from day 15 to day 50 was higher in BP cheeses (31.1–32.1% increase) than in cheeses made without BP culture (19.3–21.7% increase). The highest total FFA concentration on day 50 (612 mg kg⁻¹) was found for HP-untreated cheese made without BP culture.