Formation of biogenic amines in raw milk Hispánico cheese manufactured with proteinases and different levels of starter culture

Two proteinases, a neutral proteinase from Bacillus subtilis and a cysteine proteinase from Micrococcus sp., were used to accelerate the ripening process of raw cow's milk Hispánico cheese, a semihard variety. Two levels (0.1% and 1%) of a commercial starter culture containing Lactococcus lactis subsp. lactis and L. lactis subsp. cremoris were added for cheese manufacture. The influence of both factors, proteinase addition and level of starter culture, on the growth of amino acid-decarboxylating microorganisms and on the formation of biogenic amines during cheese ripening was investigated in duplicate experiments. The population of tyrosine decarboxylase-positive bacteria, which represented less than 1% of the total bacterial population in most cheese samples, and tyrosine decarboxylase-positive lactobacilli was not influenced by proteinase addition or level of starter culture. Tyramine was detected in all batches of cheese from day 30. Its concentration was significantly (P < 0.05) influenced by proteinase addition but not by the level of starter culture and increased with cheese age. After 90 days of ripening, 103 to 191 mg/kg of tyramine was found in the different cheese batches. Histamine was not detected until day 60 in cheese with neutral proteinase and 1% starter culture and until day 90 in the rest of the cheeses. The concentration of this amine did not exceed 20 mg/kg in any of the batches investigated. Phenylethylamine and tryptamine were not found in any of the samples.     

Effect of high-pressure treatment of ewe raw milk curd at 200 and 300 MPa on characteristics of Hispánico cheese

Hispánico cheese is a semihard variety made from a mixture of cow and ewe milks. Production of ewe milk declines in summer and autumn. To surmount the seasonal shortage of ewe milk and prevent the inactivation of milk enzymes by pasteurization, curd made in spring from ewe raw milk was pressurized at 200 and 300 MPa and stored frozen for 4 mo. Thawed ewe milk curds were added to fresh curd made from pasteurized cow milk for the manufacture of experimental Hispánico cheeses. Control cheese was made from a mixture of pasteurized cow and ewe milk in the same proportions as those used for experimental cheeses. Experimental cheeses exhibited lower dry matter content, higher aminopeptidase activity and total free amino acid concentration, and higher levels of acetic and propionic acids, aldehydes, alcohols, and esters compared with control cheese. In contrast, the concentration of total free fatty acids and ketones and the levels of textural parameters were significantly higher in control cheese. The use of ewe raw milk curd pressurized at 200 and 300 MPa, stored frozen and thawed for Hispánico cheese manufacture, was generally beneficial for cheese characteristics and increased cheese yield because of the lower dry matter content of experimental cheeses.     

Proteolysis, volatile compounds, and sensory evaluation in Hispánico cheese manufactured with the addition of a thermophilic adjunct culture, nisin, and calcium alginate-nisin microparticles

Nisin, free or incorporated in calcium alginate microparticles, was added to pasteurized milk (80% cows' and 20% ewes' milk) used for the manufacture of Hispánico cheese with a mesophilic starter and a thermophilic adjunct culture of high aminopeptidase activity. Addition of nisin incorporated in microparticles promoted early lysis of thermophilic adjunct culture bacteria. Extracellular aminopeptidase activity in 1-d-old cheese made with both thermophilic adjunct culture and nisin incorporated in microparticles was 1.8-fold higher than in cheese made with thermophilic adjunct culture and free nisin and 2.0-fold higher than in cheese made only with thermophilic adjunct culture without any addition of nisin. Addition of nisin, free or incorporated in microparticles, did not influence cheese proteolysis measured using hydrophilic or hydrophobic peptides, o-phthaldialdehyde ripening index, or free amino acids. Moreover, a total of 37 volatile compounds were identified in the volatile fraction of Hispánico cheese, using a dynamic headspace technique with a purge and trap system followed by a gas chromatography mass spectrometry analysis. The volatile compound profile was not influenced by nisin addition, either free or incorporated in microparticles, but addition of thermophilic adjunct culture enhanced the formation of 2-butanone, diacetyl, 2,3-pentanedione and acetoin and improved the flavor quality (sensory panel) of cheese.     

Microbiological,chemical and biochemical characteristics of ‘Tetilla’ raw cows-milk cheese

The microbiological, chemical and biochemical characteristics of Tetilla raw cow's-milk cheese produced in Galicia (NW Spain) were studied. The identification of the main microbial groups of technological interest was carried out. Mean log mesophilic counts, lactic acid bacteria on M17, citrate-utilizing bacteria and enterococci in 24 cheese samples were 10·60, 10·34, 9·35 and 7·30, respectively. High mean log counts of total coliforms (6·09),Micrococcaceae (5·68) and yeasts (4·44) were also measured. On the other hand, moulds (<2·60),Staphylococcus aureus (<1·79) and Escherichia coli (<1·72) counts were low. Listeria monocytogenes was detected in two samples. None of the samples yielded Salmonella spp. Isolates of technological-interest bacteria were characterized as enterococci (39·8%), lactococci (19·0%), lactobacilli (12·3%) and leuconostocs (7·6%). Micrococci (8·0%) were also isolated.Enterococcus faecalis and Lactococcus lactis subsp. lactis were the species most frequently found. In accordance with the results of pH and chemical composition, only nine samples fulfil the specifications of the Appellation of Origin. β -casein content was higher than α_{s 1}-casein content for all cheeses analysed and a low peptide α_{s 1}-I/α_{s 1}-casein ratio and nitrogen soluble fraction content were found. High volatile free fatty acid and long chain free fatty acid contents were measured, which can be attributed to the action of native lipase. A high level of diacetyl-acetoin was also determined.     

Effect of high-pressure treatment and a bacteriocin-producing lactic culture on the proteolysis, texture, and taste of Hispánico cheese

The effects of high-pressure treatment, by itself or in combination with a bacteriocin-producing culture added to milk, on the proteolysis, texture, and taste of Hispánico cheese were investigated. Two vats of cheese were manufactured from a mixture of cow and ewe milk. Milk in one vat was inoculated with 0.5% Lactococcus lactis ssp. lactis INIA 415, a nisin Z and lacticin 481 producer; 0.5% L. lactis ssp. lactis INIA 415-2, a bacteriocin-nonproducing mutant; and 2% of a commercial Streptococcus thermophilus culture. Milk in the other vat was inoculated with 1% L. lactis ssp. lactis INIA 415-2 and 2% S. thermophilus culture. After ripening for 15 d at 12°C, half of the cheeses from each vat were treated at 400 MPa for 5 min at 10°C. Ripening of high-pressure-treated and untreated cheeses continued at 12°C until d 50. High-pressure treatment of cheese made from milk without the bacteriocin producer accelerated casein degradation and increased the free AA content, but it did not significantly influence the taste quality or taste intensity of the cheese. Addition of the bacteriocin producer to milk lowered the ratio of hydrophobic peptides to hydrophilic peptides, increased the free AA content, and enhanced the taste intensity. The combination of milk inoculation with the bacteriocin producer and high-pressure treatment of the cheese resulted in higher levels of both hydrophobic and hydrophilic peptides but had no significant effect on the free AA content, taste quality, or taste intensity.     

Microstructural, Textural and Colour Characteristics During Ripening of Hispánico Cheese Made Using High-Pressure-Treated Ovine Milk Curd

High-pressure (HP) treatment of curd at low temperatures, on top of efficiently reducing microbial loads while maintaining most of the enzymatic activity, can be useful to overcome the seasonal shortage in ovine milk production. In this work, raw ovine milk curds were subjected to a wide range of HP treatments and, after frozen storage, used in the manufacture of Hispánico cheese. Confocal scanning laser microscopy of HP-treated curds showed increasing denser and more compact protein networks with the pressure applied. Although no differences in overall porosity were recorded between HP-treated and non-pressurised curds, HP treatment lowered pore number and modified pore size and shape. Whereas confocal scanning laser microscopy of control cheese showed open structures with numerous and irregular cavities, additional highly compact protein network areas could be observed in experimental cheeses micrographs. These differences trailed off with time and were no longer observed after 60 days of ripening. Experimental cheeses had significantly lower dry matter values than control cheese, and higher proteolytic indexes. Their texture was less firm than that of control cheese throughout the whole ripening period, requiring less force to be broken and deformed. Experimental cheeses showed lower lightness values than control cheese after 30 days of ripening, but no differences were found after 60 days. Our results indicate that frozen storage of HP-treated curds from ovine milk is a viable option for the production of Hispánico cheese.     

Effect of κ-casein B relative content in bulk milk κ-casein on Montasio, Asiago, and Caciotta cheese yield using milk of similar protein composition

The aim of this study was to investigate the effect exerted by the relative content of κ-casein (κ-CN) B in bulk milk κ-CN on coagulation properties and cheese yield of 3 Italian cheese varieties (Montasio, Asiago, and Caciotta). Twenty-four cheese-making experiments were carried out in 2 industrial and 1 small-scale dairy plant. Detailed protein composition of bulk milk of 380 herds providing milk to these dairies was analyzed by reversed-phase HPLC. To obtain 2 experimental milks differing in the relative content of κ-CN B in κ-CN, herds were selected on the basis of bulk milk protein composition and relative content of κ-CN genetic variants. Milk was collected and processed separately for the 2 groups of selected herds. A difference of 20% in the relative content of κ-CN B in κ-CN was obtained for the 2 experimental milks for Montasio and a difference of 15% for Asiago and Caciotta. The 2 experimental milks were of similar protein and CN content, casein number, pH, CN composition, and β-CN genetic composition. For each cheese-making trial, amounts of milk, ranging from 2,000 to 6,000 kg, were manufactured. Each vat contained milk collected at least from 4 dairy herds. Cheese yield after brining and at the end of the aging was recorded. Milk with a greater proportion of κ-CN B in κ-CN (HIGHB) exhibited similar coagulation properties and greater cheese yield compared with milk with a lower proportion of κ-CN B in κ-CN (LOWB). The increased cheese yield observed for HIGHB when manufacturing Montasio cheese was ascribed to a greater fat content compared with LOWB. The probability of HIGHB giving a cheese yield 5% greater than that of LOWB ranged from 51 to 67% for Montasio cheese, but was less than 21% for Asiago and Caciotta cheeses. Variation in relative content of κ-CN B in κ-CN content did not relevantly affect industrial cheese yield when milks of similar CN composition were processed. An indirect effect due to the increased κ-CN content of κ-CN B milk is thought to explain the favorable effects of κ-CN B on cheese yield reported in the literature.     

Structure of beef Longissimus dorsi muscle frozen at various temperatures: Part 2-ultrastructure of muscles frozen at -10, -22, -33, -78 and -115°C

The changes in electron micrographs of muscles frozen at -10, -22, -33, -78 and -115°C were analyzed. The ultrastructure of muscle successively changed with decreasing freezing temperature whereas light microscopy indicated anomalous behaviour at -22°C. It appeared that, in muscles frozen at -10°C, there was no freezing of water intracellularly; in those frozen at -22°C, water was frozen intracellularly (but only in the I-band region); whereas, in muscles frozen at -33°C, water was frozen inside the fibres, both in the I- and the A-bands. In muscles frozen at -78 and -115°C, water is frozen intracellularly. These findings can be explained on the basis that, in the I-band region, the major protein is actin, which has a relatively high proportion of non-polar residues and holds water weakly, whereas the predominant protein in the A-band is myosin, which contains many polar residues and has a high water-holding capacity.     

Stability of α-tocopherol, γ-tocopherol and β-carotene during ripening of pasta-filata cheese made from raw and pasteurised milk with different vitamin contents

Ripening stability of α-tocopherol, γ-tocopherol and β-carotene in cheese was evaluated in relation to different milk vitamin content and to the use of either pasteurised or raw milk. Milk from two farms with different management systems was used to obtain different vitamin content. Milk was divided into two parts, of which only one was pasteurised. Four blocks of cheese were made from each batch and ripened for 0, 15, 30, or 60 days at 14–16 °C. There was a notable variation in cheese vitamin levels, with the differences in milk vitamin content due to farm management having the highest impact. Pasteurisation had no effect on cheese vitamin content. Cheese γ-tocopherol and β-carotene content decreased after 30 and 60 days, respectively, whereas α-tocopherol content remained stable. γ-Tocopherol appeared to be the most efficient antioxidant in cheese, followed by β-carotene. Vitamin stability was not influenced by milk vitamin content or pasteurisation.     

Microbiological,biochemical and compositional changes during ripening of São Jorge – a raw milk cheese from the Azores (Portugal)

The microbial, compositional and biochemical profiles of São Jorge cheese (PDO) obtained from three distinct cheese plants, throughout the ripening period were determined. Fully ripened cheeses (i.e. by 130 days) contained a total of 3.1 × 10⁷ CFU g⁻¹ mesophilic bacteria, and a decrease in moisture content, concomitantly with an increase in salt content, was observed throughout the same time frame. The pH decreased until 30 days of ripening; thereafter, a slight increase was reported, up to 5.6 by the end of ripening. Urea-PAGE results showed extensive primary proteolysis, of both β-casein and α_s1-casein − degraded at essentially similar rates; plasmin and chymosin accordingly appear to be active in the cheese curd. RP-HPLC profiles of water-soluble fractions showed minor differences between 1 and 130 day old cheeses, whereas equivalent profiles of 7% (v/v) ethanol-soluble fractions contained several peaks, indicative of a heterogeneous mixture of products of proteolysis, that evolved with time.     

Effects of ozonation on microbiological,chemical and sensory attributes of vacuum-packaged rainbow trout stored at 4±0.5°C

The effects of ozone in aqueous solution on the shelf life of whole, vacuum-packaged rainbow trout, stored under refrigeration (4±0.5°C) were studied by monitoring the microbiological, chemical and sensory changes for a period of 15 days. Vacuum-packaged non-ozonated trout served as the control sample. Ozonation affected populations of bacteria namely, mesophilic aerobic bacteria, Pseudomonas spp. and H₂S-producing bacteria until day 11 of storage, Brochothrix thermosphacta, lactic acid bacteria and Enterobacteriaceae until day 8 of storage. Trimethylamine (TMA) values of all rainbow trout samples remained low (<3 mg N/100 g) until day 11 of storage, and then increased to 12.2, 8.9 and 4.7 mg N/100 g for the control and the samples ozonated for 60 and 90 min, respectively on day 15 of storage. Total volatile basic nitrogen (TVB-N) values remained relatively constant (20–25 mg N/100 g) until day 11 of storage, but increased to 61.1, 37.6 and 39.4 mg N/100 g flesh for the control and ozonated specimen for 60 and 90 min, respectively on day 15 of storage. Thiobarbituric acid (TBA) values remained relatively constant (1–3 mg MA/kg flesh) until day 12 of storage but increased to 8.4, 6.4 and 3.8 mg MA/kg flesh on day 15 of storage for the control and the ozonated trout for 60 and 90 min, respectively. Sensory evaluation (odor, taste and texture) of cooked rainbow trout showed a very good correlation with bacterial populations. On the basis of both sensory and microbiological data, a shelf life of 10 and 12 days was obtained for ozonated, vacuum-packaged and refrigerated rainbow trout at 60 and 90 min, respectively.     

Structure of beef Longissimus dorsi muscle frozen at various temperatures: Part 1-histological changes in muscle frozen at -10, -22, -33, -78, -115 and -196°C

To elucidate the relationship between the lowering of freezing temperature and the distribution of ice crystals in frozen muscle, samples of beef Longissimus dorsi muscle were frozen at -10, -22, -33, -78, -115 and -196°C. Histological preparations of these samples indicated that ice crystals were formed in muscles frozen at -10°C intercellularly, at -22°C inter- and intracellularly, at -33°C intercellularly and at -78, -115 and -196°C only intracellularly. In muscles frozen at -78°C, ice crystals in fibres were large; in those frozen at -115°C they were relatively smaller; and in those frozen at -196°C they were evenly distributed throughout the muscles. The greatest damage was caused at -22°C due to the simultaneous formation of the ice intra- and intercellularly. The results indicated that the pattern of distribution of ice crystals at the latter temperature deviates from that predicted from a linear progression of change with temperature of freezing.     

Milk with different somatic cells counts and the physicochemical,microbiological characteristics and fatty acid profile of pasteurised milk cream: is there an association?

In this study, raw cow milk containing somatic cells counts (SCC) at mean levels of 39 000 cells mL^?1 (low), 349 000 cells mL^?1 (intermediate) and 1 297 000 cells mL^?1 (high) was used for the production of pasteurised cream. Physicochemical (pH, fat and fatty acid profile) and microbiological analyses (mesophilic and psychrotrophs) were performed in the obtained creams during 30 days of refrigerated storage at 5 °C ± 2. No interactions (P > 0.05) were found between SCC, storage time and the physicochemical and microbiological variables studied. Fatty acid profile was similar among the SCC creams, except for oleic acid (C18:1), which decreased (P < 0.05) in intermediate and high SCC creams. Considering the technological aspect, our findings suggest that milk cream manufacture can be an interesting option for the use of high SCC milk.     

Characterization of the differences in the aroma of cherry wines from different price segments using gas chromatography–mass spectrometry,odor activity values,sensory analysis,and aroma reconstitution

The aroma composition, aroma-active compounds, and sensory attributions of cherry wines from three different price segments were investigated. A total of 48 aroma compounds were identified and quantified using headspace solid-phase microextraction (HS-SPME)/gas chromatography–mass spectrometry (GC–MS), among which 23 aroma compounds were further screened out as important odorants based on their odor activity values (OAVs); then, the previously mentioned 23 volatiles were correlated to sensory attributes using partial least-squares regression (PLSR). The result indicates that the differently priced wines were associated with different compounds and aroma attributions. Finally, aroma reconstitution was performed by mixing odor-active aroma compounds (OAVs>1) on the basis of their measured concentrations in the original sample. The result indicated that the aroma profile of the reconstituted sample was similar to that of the original sample.