Effect of high-pressure treatment on microbiology,proteolysis, lipolysis and levels of flavour compounds in mature blue-veined cheese

The effect of high-pressure (HP) treatment (400 MPa, 600 MPa) on ripening of mature 42-day-old Irish blue-veined cheese was studied. Counts of non-starter lactic acid bacteria, lactococci, yeasts, moulds, enterococci and total aerobic bacteria significantly decreased due to HP, with moulds being most sensitive and 600 MPa the most effective treatment. The levels of pH 4.6-soluble nitrogen and (12%) trichloroacetic acid-soluble nitrogen increased immediately after both HP treatments; however, after 28 days of storage, values were lower in HP-treated cheeses than in the control cheese. Urea-polyacrylamide gel electrophoresis showed increased breakdown of β-casein due to HP treatment at both 400 MPa and 600 MPa. Levels of free fatty acids were lower in HP-treated cheese than in the control, but not significantly so, and no significant changes could be observed in the level of flavour compounds of blue-veined cheese. Overall, HP treatment of blue-veined cheese reduced microbiological activity and decelerated proteolysis, with no statistically significant effects on development of flavour compounds.Industrial relevanceHigh-pressure treatment has been studied for the past 100 years; nevertheless, it was not applied in dairy industry, until recently, for a cheese spread. In this study, HP-induced inactivation of microbes and enzymes, which could arrest the ripening of high-quality mature (i.e., ripened) Irish farmhouse blue-veined cheese and thus extend shelf-life at optimal quality, was examined.     

Application of high-pressure treatment on ovine brined cheese: Effect on composition and microflora throughout ripening

Ovine brined cheese was high-pressure (HP) treated at 200 or 500 MPa for 15 min at 20 °C on the 15th day of ripening. Compared to control cheese, HP treatment did not affect significantly (P > 0.05) the pH values, moisture, fat in dry matter, protein in dry matter and salt in moisture contents of cheeses at 90 days. The counts of total aerobic mesophilic bacteria, thermophilic lactococci, thermophilic lactobacilli and non starter lactic acid bacteria (NSLAB) were not affected by HP treatment of cheese at 200 MPa throughout ripening. After 90 days of ripening, the same microbial groups in cheese treated at 500 MPa were about 1.2, 3.6, 2.1 and 4 log units lower than in control cheese respectively. Coliforms were reduced faster at non detectable levels in HP treated cheeses than in control cheese. Regarding the bacterial enzymatic activities in cheese, aminopeptidase activity (Apep) was marginally favoured by both HP treatments. However, its activity was decreased at 90 days due probably to loss in brine. In contrast, lactate dehydrogenase (LDH) activity, following the bacteria cell lysis, was negatively affected by HP treatment at 500 MPa throughout ripening.Industrial relevanceThe data obtained from this work suggest that application of HP treatment under optimized conditions on ovine cheese in brine can be used to reduce effectively the undesirable microbial load in it and to cause moderate enhancement of aminopeptidase activity, without modifying its composition.     

Evolution of lipolysis during the ripening of traditional Feta cheese

Lipolysis was studied during ripening of traditional Feta cheese produced in two small dairies, A and B. The cheeses were made from a thermized mixture of ewes’/goats’ milk by using yoghurt as starter and artisanal rennet from lambs’ and kids’ abomasa (cheese A) or mixed artisanal rennet with calf rennet (cheese B).The acid degree value and the free fatty acids (FFA) contents in both cheeses increased sharply up to 18 d (pre-ripening period at 15 °C) and continued to increase throughout ripening. In both mature cheeses, acetic acid was found at high levels (13–18% of the total FFAs). However, except for this, all FFA contents differed significantly (P < 0.05) between the two cheeses throughout ripening. The levels of individual and total C2:0–C8:0, C10:0–C14:0 and C16:0–C18:2 fatty acids were significantly higher (P < 0.05) in cheese A than in cheese B. Presumably the difference, especially in the C2:0–C8:0 content, was due mainly to the type of the rennet used. Butyric acid was the dominant FFA in cheese A (20% of the total FFAs at 120 d), while the most abundant FFAs in cheese B were capric (18%) and lauric acid (18%). In general, the lipolysis degree of the two cheeses was higher than those reported for the industrially-made Feta cheese.In organoleptic evaluation, cheese A had a piquant taste that was attributed to its high content of butyric acid and showed a significantly (P < 0.05) higher total score than cheese B.     

Influence of high-pressure processing (HPP) on physico-chemical properties of fresh cheese

Freshly prepared rennet-coagulated soft cheese was high-pressure (HP) treated at up to 291 MPa and 29 min and using a full 2-factor central composite design of experiment, its physico-chemical properties (colour, fat, lipid oxidation, moisture and protein content, pH, and texture) were examined. HP treatment influenced significantly (p < 0.05) the colour, fat, moisture, lipid oxidation, hardness and adhesiveness of the fresh cheese. Fat content increased apparently as moisture decreased significantly after HP treatment of above 100 MPa. Increased pressures reduced lipid oxidation but increased yellowness although the latter showed more effect over redness in the HP-treated fresh cheese. Also, increased pressures increased hardness, decreased acidity and adhesiveness in HP-treated fresh cheese although increased exposure was found to increase acidity.Industrial relevanceHigh isostatic pressure for processing fresh cheese is yet to be adopted on an industrial scale. There is a need for research to provide evidence that improved properties of fresh cheese can be realized. The effects of HPP on rennet-coagulated soft Scottish cheese are investigated and the data from this study have provided points where optimized characteristic properties of HPP fresh cheese can be attained, which can serve as a lead for HPP users on fresh cheese.     

Study of the chemical composition,proteolysis, lipolysis and volatile compounds profile of commercial Reggianito Argentino cheese: Characterization of Reggianito Argentino cheese

Reggianito is a typical variety of grana-type hard cheese produced in Argentina. It is the most exported and due to its organoleptic characteristics is very appreciated by the consumers. The objective of this study was to characterise the global composition, lipolysis, proteolysis and volatile compound profiles of commercial Reggianito cheeses from different dairy plants.Statistical differences (P ? 0.05) in some physicochemical parameters, nitrogen fractions and FFA levels among commercial brands were detected. The volatile profiles were studied by SPME–GC–MS/FID. A total of 53 compounds were identified, the majority belonging to the groups of ketones, alcohols, acids, esters and aldehydes. All these compounds have been reported in Italian grana-type cheeses. Visualization of the analytical results was performed by principal component analysis. This analysis clustered cheese samples according to dairy plants. This fact could be, among other factors, consequences of differences in technologies and ripening time of different manufacturers.     

A preliminary study on the role of alkaline phosphatase in cheese ripening

A preparation of exogenous alkaline phosphatase (ALP), containing 17,500 mU L⁻¹, was added to pasteurized milk (PM) to study its role in cheese ripening. Three miniature Cheddar-type cheeses were made from PM containing no added ALP (control), PM plus 23 μL ALP (T1), to give ALP concentration similar to that in raw milk, and PM plus 46 μL ALP (T2). Milk, after addition of ALP, was held at 6 °C for 12 h before cheese manufacture and the experiment was replicated three times. The control, T1 and T2 milks contained ALP activity of 415, 2391 and 4705 mU L⁻¹, respectively. The addition of ALP to PM caused significant (P<0.05) changes in moisture content of miniature cheeses but did not cause any changes in protein content. Levels of water-soluble N during ripening of the cheeses were similar for control, T1 and T2 cheeses. The concentration of amino acids was not affected by the level of ALP present in milk. However, reversed-phase HPLC showed differences in the peptide patterns of control, T1 and T2 cheeses, suggesting a role of ALP in cheese ripening. The results suggest that ALP may play a role in cheese ripening, but further studies are needed to confirm this.     

Effect of combinations of high-pressure treatment and bacteriocin-producing lactic acid bacteria on the survival of Listeria monocytogenes in raw milk cheese

The combined effect of high-pressure (HP) treatment and bacteriocin-producing lactic acid bacteria (BP-LAB) on the survival of Listeria monocytogenes Scott A in cheeses made from raw milk that was inoculated with the pathogen at 4.80 log cfu mL⁻¹, a commercial starter and one of seven strains of BP-LAB was investigated. On day 3, the counts of L. monocytogenes were 7.03 log cfu g⁻¹ in a control cheese (without BP-LAB, not HP treated), 6.06–6.74 log cfu g⁻¹ in cheeses with BP-LAB, 6.13 log cfu g⁻¹ in a cheese without BP-LAB and treated on day 2 at 300 MPa, 2.01 log cfu g⁻¹ in a cheese without BP-LAB and treated on day 2 at 500 MPa, 3.83–5.43 log cfu g⁻¹ in cheeses with BP-LAB and treated on day 2 at 300 MPa, and 1.81 log cfu g⁻¹ or less in cheeses with BP-LAB and treated on day 2 at 500 MPa. HP treatment was more effective on day 51 than on day 2.     

Manufacture of Cheddar cheese from high-pressure-treated whole milk

The cheese-making characteristics of high-pressure (HP)-treated milk were examined. The rennet coagulation time of pasteurised milk decreased after HP treatment at 400 MPa but increased after treatment at 600 MPa. The L-value (whiteness) of milk decreased directly after HP treatment but, over the course of coagulation, whiteness of HP-treated milk increased to the same level as in the control. Cheddar cheese was then manufactured from raw whole milk or whole milk treated by high-pressure (HP) at 400 MPa (HP400) or 600 MPa (HP600) for 10 min at 20 °C. HP treatment of raw milk at 600 MPa resulted in a 3.66 log reduction in the initial counts of non-starter lactic acid bacteria (NSLAB), decreased protein and fat content, as well as a lower pH compared to the control. Furthermore, higher treatment pressures resulted in increased incorporation of β-lactoglobulin into the cheese curd, with parallel increases in yield by 1.23% and 7.78% for HP400 and HP600 cheeses, respectively. Overall, this study showed that the effects of HP treatment on milk proteins increased rennet coagulation times and changes in cheese composition at day 1.Industrial relevanceHigh-pressure treatment is a novel technology which has been applied to a number of commercial food products. In this study, HP-induced changes in milk proteins resulted in increased cheese yields and increased cheese whiteness. In addition, HP treatment significantly reduced the microflora of raw milk cheese. Those attributes could be of interest for both industry and consumer.     

Influence of reuterin-producing Lactobacillus reuteri coupled with glycerol on biochemical,physical and sensory properties of semi-hard ewe milk cheese

The biochemical, physical and sensory characteristics of ewe milk cheeses made with reuterin-producing Lactobacillus reuteri and glycerol (substrate for reuterin production) were assessed. Cheese made with lactococci starter (CTRL), cheese made with starter and L. reuteri (SLR), and cheese made with starter, L. reuteri and 30 mM glycerol (SLR-G) were manufactured. L. reuteri reached counts above 7 log cfu/g on day 1. Lactococci survival was enhanced in SLR cheese without affecting cheese pH, dry matter, proteolysis, concentration of most free amino acids (FAA), textural and most color parameters, or sensory characteristics. In situ production of reuterin by L. reuteri was only detected in SLR-G cheese, decreasing LAB counts although acidification remained unaffected. SLR-G cheese showed higher values of cell free aminopeptidase activity, overall proteolysis and FAA, particularly glutamic acid, than CTRL and SLR cheeses. The addition of L. reuteri-glycerol resulted in lower hardness and elasticity values in SLR-G cheese and influenced its L*, a* and b* color parameters. However, these changes, which were detected by instrumental analysis, did not affect the sensory scores for texture and color quality of SLR-G cheese, and it received the highest scores for taste quality. Our results suggest that L. reuteri-glycerol may provide a suitable system to release the antimicrobial reuterin in cheese without affecting negatively its sensory characteristics.     

Applicability of extracts from Centaurea calcitrapa in ripening of bovine cheese

Aqueous extracts obtained from cell suspension cultures of Centaurea calcitrapa were used as proteolytic additive in the manufacture of a commercial bovine cheese, coagulated with animal rennet and typically ripened for 28 d. The cheese was assessed in comparison to standard cheese for two levels of addition of said extract, viz. 0.61 and 1.22 mg of total protein mL⁻¹. The qualitative and quantitative evolutions of the nitrogen fractions were monitored in the experimental cheeses throughout the whole ripening period. In general, the chemical compositions of the cheeses were different depending on the amount of extract used, but no significant differences could be detected in the ripening index. With regard to electrophoretic profiles, the two types of cheese could be distinguished until up to ca. 7 d of ripening, but differences did essentially vanish by 28 d.     

Effect of commercial adjunct cultures on proteolysis in low-fat Kefalograviera-type cheese

The effect of two commercially available adjunct cultures, LBC 80 (Lactobacillus casei subsp. rhamnosus) and CR-213 (containing Lactococcus lactis subsp. cremoris and Lc. lactis subsp. lactis) on the proteolysis in low-fat hard ewes’ milk cheese of Kefalograviera-type was investigated. Two controls, a full-fat cheese (306 g kg⁻¹ fat, 378 g kg⁻¹ moisture) and a low-fat cheese (97 g kg⁻¹ fat, 486 g kg⁻¹ moisture, made using a modified procedure), were also prepared. The effect of adjunct culture on proteolysis, as examined by polyacrylamide gel electrophoresis of cheese and water soluble cheese extracts, was marginal. The reverse-phase HPLC peptide profiles of the water soluble extracts from low-fat cheeses were similar although some quantitative differences were observed between low-fat control cheese and experimental cheeses. The fat content as reflected by the differences in peptide profiles affected the pattern of proteolysis. Proteolysis, as measured by the percentage of total nitrogen soluble in water or in 120 g L⁻¹ trichloroacetic acid, was significantly (P<0.05) affected by the addition of adjunct cultures. Furthermore, the adjunct cultures enhanced the production of low molecular mass nitrogenous compounds; the levels of total nitrogen, soluble in 50 g L⁻¹ phosphotungstic acid, and of free amino acids were significantly (P<0.05) higher in the low-fat experimental cheeses than in the low-fat control cheese.     

Preliminary observations on proteolysis in Manchego cheese made with a defined-strain starter culture and adjunct starter (Lactobacillus plantarum) or a commercial starter

Different authors have demonstrated the potential of adding lactobacilli as adjunct cultures to pasteurized milk used in cheese manufacture. The aim of this work was to observe the effect of the use of a defined-strain starter culture and the addition of an adjunct culture (Lactobacillus plantarum) to cheesemilk in order to determine their effect on the ripening of Manchego cheese. Manchego cheeses were manufactured using one of the following starter culture systems: a defined starter consisting of Lactococcus lactis ssp. lactis and Leuconostoc mesenteroides ssp. dextranicum; a defined starter, as described above, and Lb. plantarum, which were isolated from a good quality Manchego cheese made from raw milk, or a commercial starter comprised of two strains of Lc. lactis. The cheeses were sampled at 15, 45, 90 and 150 d of ripening. Principal component analysis of peak heights of reversed-phase HPLC chromatograms of 70% (v/v) ethanol-insoluble and -soluble fractions distributed the samples according to the starter used in their manufacture. Quantitative differences in several peptides were evident between the three cheeses. Cheeses made with the defined-strain starters received higher scores for the flavour quality and intensity and for overall impression than the cheeses made with the commercial starter.     

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.     

Functional goat milk cheese with feruloyl esterase activity

The aim of this paper was to evaluate the effect of the intake of goat milk cheese manufactured with Lactobacillus fermentum CRL1446 on intestinal feruloyl esterase (FE) activity and oxidative status in Swiss albino mice. This strain was used as single-strain culture (CRL cheese) and in combination with starter culture (Mix cheese). In both cheeses, L. fermentum reached 8–9 log cfu/g and FE activity increased during ripening. Highest activity level was observed in Mix cheese. In vivo studies showed that total intestinal FE activity in mice fed with CRL and Mix cheeses increased 1.5 and 2-fold compared to non-treated mice, respectively. Administration of Mix cheese produced a 2-fold increase in FE activity in small and large intestine mucosa. Mice receiving this cheese also showed an approx. 2-fold decrease in plasmatic thiobarbituric acid-reactive substances (TBARS) levels and an approx. 3-fold increase in glutathione reductase (GR) activity.Goat milk cheese elaborated with FE-producing strain L. fermentum CRL1446 could represent a novel functional food with FE activity, responsible for increasing intestinal FE activity and consequently the bioavailability of antioxidant ferulic acid in the gut, thus enhancing the oxidative status and providing protection against oxidative stress-related disorders.     

Influence of bacteria used as adjunct culture and sunflower oil addition on conjugated linoleic acid content in buffalo cheese

The influence of bacteria and sunflower oil addition on conjugated linoleic acid content (CLA) in buffalo cheese was determined. Fresh and short-ripened cheeses were manufactured using the same starter culture and four different adjunct strains previously selected by their CLA production rate. Lactobacillus casei, Lactobacillus rhamnosus, Bifidobacterium bifidum and Streptococcus thermophilus were individually used as adjunct culture. Sunflower oil (SO) was added to obtain a final concentration of 200 μg/ml of linoleic acid. CLA levels in cheese were higher than raw milk, especially after ripening time. SO supplementation increase CLA concentrations in fresh cheeses, except in those manufactured with S. thermophilus as adjunct culture. Both, ripening and SO supplementation showed a positive influence on CLA concentration. Similar texture, acidity and colour were determined in cheeses with or without SO supplementation. Buffalo cheeses manufactured with appropriate adjunct cultures may be a natural source of CLA for human consumption.     

Production of biogenic amines during the ripening of Pecorino Abruzzese cheese

The production of biogenic amines (BA) during the manufacturing and ripening of sheep milk Pecorino Abruzzese cheeses prepared from raw milk without starter culture (A) and from pasteurized milk with added starter (B) were compared. At the end of ripening (60 days), the total BA contents of cheeses of batches A and B were 697 and 1086 mg kg⁻¹, respectively; the dominant BA were different. Single isolates of enterococci, pseudomonads and Enterobacteriaceae were screened for their potential to produce BA. Qualitative tests indicated a large spread of BA-forming cultures among the members of the Enterobacteriaceae and lactic acid bacteria (LAB). Differences among the levels of BA produced in UHT milk by representative isolates of coliforms, Pseudomonas and LAB were observed in relation to the microbial group or the isolate. The results emphasize the need to improve the general hygienic conditions of Pecorino Abruzzese cheese manufacture and control the indigenous bacterial population.     

Contribution of coagulant and native microflora to the volatile-free fatty acid profile of an artisanal cheese

The contributions of the coagulant Cynara cardunculus and of the microflora of raw milk to the volatile-free fatty acid profile of Serra da Estrela cheese were evaluated. The experimental design included both a model system and, dual cheeses. The study in the model system showed that isovaleric acid was the predominant volatile compound after 7 d of ripening. The systems inoculated with Enterococcus faecium produced the highest amount of this volatile (ca. 135.8 mg kg⁻¹ curd), while those inoculated with Lactobacillus plantarum produced the least (21.4 mg kg⁻¹ curd); Lactococcus lactis produced moderate amounts (ca. 34.2 mg kg⁻¹ curd) but a total amount of volatile-free fatty acids similar to those found in control samples. This is considered advantageous since this volatile fatty acid confers a harsh, piquant, mature flavour to cheese, coupled with the realisation that excess volatiles may result in off-flavours. The addition of cultures in experimental cheeses helped reduce ripening time to about one half. Inclusion of Lb. plantarum led to cheeses containing the highest amounts of volatiles, and exhibiting an aroma closest to that of typical Serra da Estrela cheese.     

Influence of probiotic bacteria on the proteolysis profile of a semi-hard cheese

Two probiotic strains, Lactobacillus acidophilus and Lactobacillus paracasei subsp. paracasei, were used as adjunct cultures in semi-hard cheesemaking experiments, in order to study their influence on proteolysis during ripening. Cheeses with and without probiotic bacteria were manufactured. The population of probiotics remained above 10⁷ cfu g⁻¹ during all ripening, and they did not influence primary proteolysis. However, L. acidophilus produced a significant increase in the level of low molecular weight nitrogen compounds and individual free amino acids; the amino acid profiles were also different. Multivariate analysis of peptide profiles showed that samples were grouped mainly by ripening time, although the impact of probiotics was also noticeable. L. acidophilus showed a clear influence on secondary proteolysis, while a minor effect of L. paracasei was evidenced at the end of the ripening. These results showed that the tested strains influenced distinctly proteolysis of cheeses, probably as a consequence of their different proteolytic systems and their activity via the alimentary matrix (cheese).