Effect of addition of CO₂ to raw milk on quality of UHT-treated milk

The objective of this study was to evaluate the effect of addition of CO(2) to raw milk on UHT milk quality during storage. Control milk (without CO(2) addition) and treated milk (with CO(2) addition up to pH 6.2) were stored in bulk tanks at 4°C for 6d. After storage, both samples were UHT processed using indirect heating (140°C for 5s). Samples were aseptically packed in low-density polyethylene pouches and stored in the dark at room temperature. Raw milk was evaluated upon receipt for physicochemical composition, proteolysis, lipolysis, standard plate count, psychrotrophic bacteria, and Pseudomonas spp. counts, and after 6d of storage for proteolysis, lipolysis, and microbial counts. After processing, UHT milk samples were evaluated for physicochemical composition, proteolysis, and lipolysis. Samples were evaluated for proteolysis and lipolysis twice a month until 120d. Peptides from pH 4.6-soluble N filtrates were performed by reversed-phase HPLC after 1 and 120d of storage. A split-plot design was used and the complete experiment was carried out in triplicate. The results were evaluated by ANOVA and Tukey's test. After 6d of storage, CO(2)-treated raw milk kept its physicochemical and microbiological quality, whereas the untreated milk showed significant quality losses. A significant increase in proteolysis occurred during 120d of storage in both treatments, but the increase occurred 1.4 times faster in untreated UHT milk than in CO(2)-treated UHT milk. In both UHT milks, the proteolysis was a consequence of the action of plasmin and microbial proteases. However, the untreated UHT milk showed higher microbial protease activity than the treated UHT milk. The addition of CO(2) to the raw milk maintained the quality during storage, resulting in UHT milk with less proteolysis and possibly longer shelf life, which is usually limited by age gelation of UHT milk.     

IMPROVED SHELF LIFE ESTIMATION OF UHT MILK BY PREDICTION OF PROTEOLYSIS

ABSTRACT

During growth in raw milk, many psychrotrophic bacteria produce proteases that can retain activity following ultra‐high temperature (UHT) treatment. In this study, casein and skim milk powder assays for detecting very low levels of protease in UHT milk were optimized, and the suitability of azocasein and fluorescein isothiocyanate‐casein (FITC‐casein) as substrates was investigated. The strongest correlations of protease activity with proteolysis in stored UHT milk were observed when FITC‐casein was used as substrate in the assays. Assays using casein and FITC‐casein as substrates yielded the highest activities. To determine sensitivity, crude protease was added at low concentrations to UHT milk, and the milk was assayed for progress of proteolysis over 12 months and for protease activity using the casein and FITC‐casein assays. With long assay incubation times, the FITC‐casein assay was more sensitive than the casein assay and may be suitable for detecting very low levels of protease activity and predicting progress of proteolysis in stored UHT whole milk.

PRACTICAL APPLICATIONS

This study contributes to the development and evaluation of practical assays for the detection of protease activity in the industry to identify potential premature spoilage of contaminated UHT milk before it is distributed for sale. The developed assays are also useful for assessing the quality of milk powder as active protease can persist in milk powder to cause spoilage in reconstituted milk. Although the assays require up to 14 days to complete, this is not an excessive time, compared with the time required for microbiological clearance and total shelf life of the product. High protease activity can be identified with less incubation time. The cost of protease detection assays developed during this work is quite low and, although 20 min analysis time is required per sample, the tests can be very cost efficient when run in batches, as would be expected in a commercial testing facility.

     

Proteolysis,protein distribution and stability of UHT milk during storage at room temperature

Proteolytic degradation and distribution of caseins and whey proteins between the soluble and colloidal phases were studied in six batches of commercial UHT milk (three skim and three whole milks) during storage at 25 ± 2 °C. For that purpose, at 30 day intervals, milk samples were ultracentrifuged and the pellets and supernatants analysed by capillary electrophoresis and SDS‐PAGE. Samples were also visually examined for signs of gelation. Extensive proteolytic degradation of the micellar fractions and severe changes in the electrophoretic pattern of the proteins present in the serum fractions were observed in all the batches. A higher proportion of denatured whey proteins not attached to the micelle surface was found in the skim milk samples as compared with the whole milk samples that could provide less resistance against gelation. In addition to β‐Lg, para‐κ‐casein was also found in the serum fraction. A high proteolytic activity against κ‐casein could be responsible for the hydrolysis of serum‐liberated κ‐casein or could have enhanced the liberation of β‐Lg–para‐κ‐casein complexes through proteolysis of micellar κ‐casein. © 1999 Society of Chemical Industry     

Effects of somatic cell count on quality and shelf-life of pasteurized fluid milk

Milk was collected from eight Holstein cows four times before and four times after intramammary infection with Streptococcus agalactiae. Postinfection milk had significantly higher somatic cell count (SCC) (849,000 cells/ml) than preinfection milk (45,000 cells/ml). High SCC raw milk had more lipolysis and proteolysis than low SCC raw milk. Pasteurized, homogenized, 2% fat milks from pre- and postinfection periods were stored at 5 degrees C and analyzed for lipolysis, proteolysis, microbial quality, and sensory attributes at 1, 7, 14, and 21 d post processing. During refrigerated storage, the average rates of free fatty acid increase (i.e., lipolysis) and casein hydrolysis in high SCC milk were, respectively, three and two times faster than those in low SCC milk. In general, standard plate counts, coliform counts, and psychrotrophic bacterial counts of both the high and low SCC milks remained low (<100,000 cfu/ ml) during 5 degrees C storage. Low SCC milk maintained high organoleptic quality for the entire 21-d shelf-life period. However, for high SCC milk, between 14 and 21 d, sensory defects were detected, which resulted in low overall quality ratings. The sensory defects mainly included rancidity and bitterness and were consistent with higher levels of lipolysis and proteolysis. Hence, mastitis adversely affected the quality of pasteurized fluid milk. It is recommended that the fluid milk industry consider implementation of premium quality payment programs for low SCC milks.     

UHT乳贮藏期间理化特性和感官品质的变化

为探讨超高温瞬时杀菌(ultra-high temperature instantaneous sterilization,UHT)乳的营养价值随贮藏时间的变化,对不同贮藏期UHT乳的理化特性和感官品质进行监测。在6个月的贮藏期内,UHT乳中脂肪、蛋白质、乳糖以及固形物的总量并无明显变化,维生素C和泛酸的含量均损失了约25%,游离氨基酸和钙离子分别增加了5.35%和6.73%。不同贮藏期UHT乳的黏度变化不显著(P>0.05),但粒径大小强烈依赖于贮藏时间,乳脂肪球和酪蛋白胶束的尺寸在贮藏初期分别增加了约38 nm和14 nm,然后分别稳定在约173 nm和612 nm。贮藏1个月的UHT乳样品在风味和口感方面整体表现较好,贮藏6个月后,UHT乳的感官品质大幅下降,出现了许多不良风味。     

Short communication: Relationships between α-lactalbumin and quality traits in bulk milk

The main objective of this study was to investigate whether the α-lactalbumin (α-LA) content of bulk milk is related with some known inflammatory markers and milk quality traits. An additional objective was to study whether combining α-LA, haptoglobin (Hp), and serum amyloid A (SAA) in an acute phase index (API) could be useful as an alternative marker for bulk milk quality. For the dairy industry, it is of great importance to receive high quality bulk milk for manufacture of liquid milk and dairy products. The somatic cell count (SCC) is currently used as an indirect marker for bulk milk quality, but because it is somewhat insensitive and unspecific, interest exists in alternative markers. Bulk milk samples were analyzed for α-LA, SCC, polymorphonuclear leukocyte count, Hp, SAA, fat, lactose, total protein and casein contents, casein number, protein composition, proteolysis, and coagulating properties. No significant differences were found in SCC, polymorphonuclear leukocyte count, Hp, or SAA between milk samples containing low, medium, or high concentrations of α-LA. Differences between α-LA groups were, however, found in some milk quality traits because high α-LA concentration was related to low concentrations of α_S1-, α_S2-, and β-caseins and high concentrations of lactose and β-lactoglobulin. A high API was related to low lactose content and casein number. Samples with high SCC contained less casein and had a lower casein number than milk with a low SCC, and proteolysis was significantly higher in high SCC milk than in low SCC milk. Neither α-LA nor API proved to be a better marker than SCC for the quality traits investigated, and α-LA was not considered to be a useful inflammatory marker in bulk milk.     

Effect of storage and separation of milk at udder quarter level on milk composition, proteolysis, and coagulation properties in relation to somatic cell count

Coagulation properties of milk are altered by elevated somatic cell count (SCC), partly due to increased proteolytic and lipolytic activity in the milk and, thereby, degradation of protein and fat during storage. Milk is commonly stored on the farm at cooling conditions for up to 2 d before transport to the dairy for processing. This study evaluated the effects of storage on milk with altered composition due to high SCC and the effects of exclusion of milk from individual udder quarters with high SCC on milk composition, proteolysis, and coagulation properties. Udder-quarter milk and cow-composite milk samples from 13 cows having at least 1 quarter with SCC above 100,000 cells/mL were collected on 1 occasion. In addition, commingled milk from only healthy quarters (<100,000 cells/mL) of each cow was collected, representing a cow sample where milk with elevated SCC was excluded. The milk samples were analyzed for total protein content; protein content in the whey fraction; casein, fat, and lactose contents; SCC; proteolysis; curd yield; coagulation time; and total bacterial count, on the day of sampling and after 2 and 5 d of storage at +4°C. In addition to SCC, duration of storage and total bacterial count had an effect on milk quality. The content of total protein, fat and protein contents in the whey fraction, and curd yield were found to have different storage characteristics depending on the level of SCC at udder-quarter level. The exclusion of milk from udder quarters with elevated SCC decreased the content of total protein and protein content in the whey fraction and increased the content of lactose at cow level. However, the effect of separating milk at udder-quarter level needs to be further studied at bulk tank level to evaluate the effect on overall total milk quality.     

Effects of somatic cell counts in milk on physical and chemical characteristics of yoghurt

The quality of plain stirred yoghurt produced from whole milk with somatic cell counts (SCC) at low (147,000 cells mL⁻¹), intermediate (434,000 cells mL⁻¹) and high (1,943,000 cells mL⁻¹) levels was examined. Each milk treatment was obtained from selected cows, according to its SCC status and milk composition. Yoghurt samples were analysed on days 1, 10, 20 and 30 after production. Analyses included pH, acidity, fat, lipolysis (expressed as free fatty acids, FFA), proteolysis and apparent viscosity. Viscosity of high SCC yoghurt was higher (P<0.05) than the low SCC yoghurt on days 10, 20 and 30 of storage. High SCC yoghurt also had higher FFA content (P<0.05). SCC did not affect pH, acidity, fat content and proteolysis of the yoghurt (P>0.05). Results indicate that SCC in milk increases the lipolysis in the resulting yoghurt during storage for 30 d.     

原料乳离心除菌对UHT乳品质的改善作用

离心除菌可以改善原料乳的质量,本文研究了常规UHT(N-UHT)乳和离心除菌UHT乳(C-UHT)在贮藏过程中理化性质、蛋白分布变化.结果表明:UHT乳贮藏期颜色变深、粒度增加、Zeta电位绝对值降低、黏度增加、酸度增加和纤溶蛋白酶活性增加,乳胶体稳定性降低;离心除菌UHT乳(C-UHT)的物理化学稳定性优于普通UHT...     

Proteolysis and storage stability of UHT milk produced in Turkey

The effect of raw milk quality (total and psychrotrophic bacterial and somatic cell counts, proteinase and plasmin activity) and UHT temperature (145 or 150 °C for 4 s) on proteolysis in UHT milk processed by a direct (steam-injection) system was investigated during storage at 25 °C for 180 d. High proteinase activity was measured in low-quality raw milk, which had high somatic cell count, bacterial count and plasmin activity. The levels of 12% trichloroacetic acid–soluble and pH 4.6-soluble nitrogen in all milk samples increased during storage, and samples produced from low-quality milk at the lower UHT temperature (145 °C) showed the highest values. Bitterness in UHT milk processed from low-quality milk at 145 °C increased during storage; gelation occurred in that milk after 150 d. The RP-HPLC profiles of pH 4.6-soluble fraction of the UHT milk samples produced at 150 °C showed quite small number of peaks after 180 d of storage. Sterilization at 150 °C extended the shelf-life of the UHT milk by reducing proteolysis, gelation and bitterness.     

Characterization of a new isolate of Lactobacillus fermentum IFO 3956 from Egyptian Ras cheese with proteolytic activity

More than 200 isolates were obtained from 15 Egyptian traditional dairy products (Domiatti cheese, Ras cheese and Rayeb milk) collected from local markets of Alexandria, Tanta and Kafr El-Sheikh. Examination with optical microscope of these dairy samples allowed to classify 92 bacilli, 64 of which were identified as lactobacilli. The proteolytic activity of lactobacilli isolates was tested on skim milk agar. Eight isolates showing a high proteolytic activity were further tested on UHT skim milk. The strain showing the highest proteolytic activity was purified and identified as Lactobacillus fermentum IFO 3656. The specific proteolytic activity of this strain and the factors affecting it (pH, temperature and presence of inhibitors) were studied. The proteolysis targeted mainly caseins (73% of whole casein), especially β-casein (85%). Smaller portions of whey proteins were proteolyzed (20%) essentially β-lactoglobulin. The proteolysis process gave rise to medium-sized peptide populations. The optimum conditions for the proteolysis activity of the studied strains were pH 6.5 and 37 °C. Proteolytic activities were very slightly affected by the increase of the temperature to 42 °C or the pH to 8.2. The protease system of Lactobacillus fermentum IFO 3956 is most probably composed from a high amount of metalloproteases and small amount of cysteine and serine proteases.     

UHT Milk Processing and Effect of Plasmin Activity on Shelf Life: A Review

Abstract: The demand for ultra‐high‐temperature (UHT) processed and aseptically packaged milk is increasing worldwide. A rise of 47% from 187 billion in 2008 to 265 billon in 2013 in pack numbers is expected. Selection of UHT and aseptic packaging systems reflect customer preferences and the processes are designed to ensure commercial sterility and acceptable sensory attributes throughout shelf life. Advantages of UHT processing include extended shelf life, lower energy costs, and the elimination of required refrigeration during storage and distribution. Desirable changes taking place during UHT processing of milk such as destruction of microorganisms and inactivation of enzymes occur, while undesirable effects such as browning, loss of nutrients, sedimentation, fat separation, cooked flavor also take place. Gelation of UHT milk during storage (age gelation) is a major factor limiting its shelf life. Significant factors that influence the onset of gelation include the nature of the heat treatment, proteolysis during storage, milk composition and quality, seasonal milk production factors, and storage temperature. This review is focused on the types of age gelation and the effect of plasmin activity on enzymatic gelation in UHT milk during a prolonged storage period. Measuring enzyme activity is a major concern to commercial producers, and many techniques, such as enzyme‐linked immunosorbent assay, spectrophotometery, high‐performance liquid chromatography, and so on, are available. Extension of shelf life of UHT milk can be achieved by deactivation of enzymes, by deploying low‐temperature inactivation at 55 °C for 60 min, innovative steam injection heating, membrane processing, and high‐pressure treatments.     

贮藏期UHT奶中蛋白质组成结构及其糖基化的变化

通过测定UHT奶在贮藏过程中蛋白水解,蛋白在胶束相和乳清相的分布,乳蛋白糖基化位点变化,探究乳蛋白在贮藏过程中性质的变化.结果表明贮藏过程中UHT奶中菌落总数增大,纤溶酶含量增大.UHT奶在内外源酶的作用下乳蛋白发生不同程度的水解.热处理导致从胶束表面脱落的κ-CN和乳清蛋白以热诱导聚合物的形式,在贮藏过程中重新结合到...     

Effect of somatic cell count on proteolysis and lipolysis in pasteurized fluid milk during shelf-life storage

The general goal of this research was to provide fluid milk processors with data to enable them to estimate the economic benefits they might derive from longer fluid milk shelf-life or new marketing opportunities due to a reduction in raw milk SCC. The study objectives were: 1) to measure the time in days for pasteurized homogenized 2% milk to achieve a level of lipolysis and proteolysis caused by native milk enzymes present in milks of different somatic cell count (SCC) at 0.5 and 6 degrees C that would be sufficient to produce an off-flavor, 2) to determine whether milk fat content (i.e., 1, 2, and 3.25%) influences the level of proteolysis or lipolysis caused by native milk enzymes at 6 degrees C, and 3) to determine the time in days for milks containing 2% fat with different SCC to undergo sufficient lipolysis or proteolysis to produce an off-flavor due to the combination of the action of native milk enzymes and microbial growth at 0.5 and 6 degrees C. In experiment 1, pasteurized, homogenized milks, containing 2% fat were prepared from raw milk containing four different SCC levels from < 100,000 to > 1,000,000 cells/ml. Each of the four milks was stored at 0.5 and 6 degrees C for 61 d. In experiment 2, pasteurized, homogenized milks containing 1, 2, and 3.25% fat were prepared starting from two raw milks containing two different SCC levels, one < 100,000 and the other > 1,000,000 cells/ml. In experiment 3, pasteurized, homogenized 2% fat milks were prepared starting from raw milks containing two different SCC levels, one < 100,000 and the other > 1,000,000 cells/ml. For experiments 1 and 2, all milks were preserved with potassium dichromate to prevent microbial growth but to allow the activity of native milk proteases and lipases during storage. For experiment 3, one set of milk was preserved with potassium dichromate to prevent microbial growth but to allow the activity of native milk proteases and lipases, and a second set of milk was unpreserved during storage at 0.5 and 6 degrees C for 29 d. Based on previous work, an off-flavor due to proteolysis was detected by 50% of panelists when the decrease in casein as a percentage of true protein (CN/TP) was > 4.76%. Our data indicated (assuming 50% of consumers would detect an off-flavor when CN/TP decreases 5%) that pasteurized milk containing 2% fat would develop an off-flavor at a time long after 61 and at 54 d for the low SCC milk, and at about 54 and 19 d for the high SCC milk, at 0.5 and 6 degrees C, respectively. Previous research reported that 34% of panelists could detect an off-flavor in milk containing 2% fat due to lipolysis at a (free fatty acid) FFA concentration of 0.25 meq/kg of milk. Based on these results, it was estimated in the present study that 34% of panelists would detect an off-flavor in a 2% fat pasteurized milk with low SCC at a time long after 61 and just after 61 d at 0.5 and 6 degrees C, respectively, while for milk with high SCC, an off-flavor would be detected by 34% of panelists at slightly longer than 61 and 35 d at 0.5 and 6 degrees C, respectively. The combination of low SCC milk and low storage temperature when coupled with processing technology to produce very low initial bacteria count in fluid milk could produce fluid milk that will maintain flavor quality for more than 61 d of storage at temperatures < 6 degrees C.     

Interactions between bacteria type,proteolysis of casein and physico-chemical properties of bovine milk

The effects of separate infection with four major pathogens frequently associated with the occurrence of subclinical mastitis in cows (Staphylococcus aureus, S. chromogenes, Escherichia coli and Streptococcus dysgalactiae) on milk quality for cheese production were studied for quarters of the same animal. Infection increased somatic cell count (SCC), modified leucocyte distribution, decreased lactose concentration and increased proteolysis of casein. Regardless of bacteria type, the plasmin activity in milk from the infected glands increased ∼2 fold compared with uninfected quarters. These changes were associated with increased rennet clotting time and decreased curd firmness for milk from infected glands, indicating that milk quality for cheese production was negatively affected by infection. Although the general pattern of bacterial invasion was similar, each type of bacteria elicited the above-described responses in a specific manner. SCC, commonly used by the dairy industry as a measure of milk hygienic quality, provided the poorest prediction of milk quality for cheese production in comparison to indices of proteolysis of casein.     

Detection of rennet whey solids in UHT milk by capillary electrophoresis

The fraudulent presence of rennet whey solids in UHT milk was studied by capillary electrophoresis (CE). Commercial UHT samples of different origins, genuine milk samples and milk samples adulterated with rennet whey were analysed. Linear discriminant functions using ratios of peak areas of caseinmacropeptide (CMP) and two other CMP-like degradation products were defined. The interference of proteolysis in the detection was estimated in samples adulterated on purpose with rennet whey, UHT treated in a pilot plant, and stored at 10, 20 and 30°C for up to 150 days. The application of the classification functions obtained allowed the detection of rennet whey solids added to milk. Only interferences due to very severe proteolysis, occurring after very long storage periods and/or at storage temperatures above room temperature, were observed.     

On the influence of non‐enzymatic crosslinking of caseins on the gel strength of yoghurt

After storage of UHT milk at 37°C resp. 50°C, yoghurt was prepared. For a storage temperature of 37°C, breaking strength of the yoghurt samples increased from 2.7 to 5.8 N with increasing storage duration of the UHT milk. A plateau is reached after 17 days of storage. This increase in breaking strength correlates with a significant increase in non‐reducible casein oligomerization from 14% for fresh UHT milk to 25% measured using size exclusion chromatography under reducing and denaturing conditions and calculated as sum of predominantly formed dimers and trimers at the total casein fraction. At a storage temperature of 50°C, a less increase in breaking strength from 2.7 to 4.6 N with a plateau after 17 days was observed while casein oligomerization increased to 63%. After acid hydrolysis, only lysinoalanine and histidinoalanine were detected in the caseinate samples via amino acid analysis. The quantified concentration of lysinoalanine and histidinoalanine could not explain the observed casein oligomerization. Thus, unknown crosslinked amino acids must have been formed during storage, inducing significant changes in the functional properties of milk proteins.     

Survival analysis,consumer perception and physico-chemical analysis of low fat UHT milk stored for different time periods

Survival analysis based on consumers' acceptance or rejection of milk of different storage ages, was used to validate the shelf-life of low fat ultra-high temperature treated (UHT) milk in high density polyethylene bottles, as previously determined by a multivariate accelerated shelf-life test (MASLT). UHT milk between 120 and 290 d of storage were evaluated. Based on 50% of consumers rejecting the product, the shelf-life was estimated to be 214 d, validating the shelf-life of 211 d estimated by the MASLT. In addition, consumers completed check-all-that-apply attribute questions and rated the acceptability of the milk. The consumers noted positive sensory attributes more frequently in fresher milk samples with an increase in negative attributes with storage. Along with this, hedonic scores for the milk decreased and physicochemical and enzymatic reactions associated with the deterioration of UHT milk increased with storage.     

Gelation Profiles of Yogurt as Affected by Heat Treatment of Milk

Milk was heated at 140°C for 6 s (UHT), 98°C for 1.87 min (HTST) and 85°C for 10 min (vat-pasteurized), homogenized, inoculated with yogurt culture, and incubated at 45°C. During incubation, pH, ionic Ca, and viscosity changes were continuously monitored by interfacing to a microcomputer. Samples for transmission electron microscopy were taken at selected pH intervals.Acid fermentation of milk was a multistage process comprising 1) an initial lag period of low viscosity; 2) a period of rapid viscosity change, and 3) a stage of high constant viscosity. The shapes of the viscosity versus incubation time curves were the same for all heat treatments differing only in the extent of viscosity change in stage 2. A microstructural study of UHT milk indicated that at pH 5.1, casein micelles appeared to dissociate into subparticles of approximately 30 to 40 nm diameter. By pH 4.8, subparticles reassociated into large casein aggregates of nonspecific shape and dimensions. Micellar dissociation was thought to be influenced by conversion of colloidal Ca to ionic Ca.     

Proteolysis and texture of hard ewes' milk cheese during ripening as affected by somatic cell counts

Ewes' milk samples with low (<500,000 ml(-1)), medium (1,000,000-1,500,000 ml(-1)) and high (> 2,500,000 ml(-1)) somatic cell counts (SCC) were used to manufacture hard ewes' cheese using the Zamorano cheese manufacturing protocol. Cheeses that had been ripened for 1, 2 and 3 months were used to obtain isoelectric ovine casein that was analysed by capillary electrophoresis. The texture of the cheeses during ripening was determined instrumentally using the Warner-Bratzler maximum shear force and assessed for sensory qualities by consumers using hedonic tests. The study revealed that the pH value and the lactose content of the milk were affected by high SCC and that the coagulation properties were dependent on the somatic cell content. The protein and moisture contents of the cheeses were unaffected by SCC but a significant increase of pH with ripening time were observed in high-SCC cheeses. The results also pointed to a significant increase in proteolysis related to SCC levels, showing that intact casein, both alphas1 and beta-casein, decreased as the SCC of milk increased, and that the proteolytic fragments, mainly I-alphas1, increased with SCC levels. Significant differences in texture were found among the samples, the cheeses made with high levels of SCC being significantly less compact at each ripening time. The differences in texture were detected by the consumers, who reported defects in cheeses made with high levels of SCC. Indeed, high SCC cheeses were significantly less well accepted.