Distribution of aflatoxin M1 during Grana Padano cheese production from naturally contaminated milk

The distribution of aflatoxin M₁ (AFM₁) has been investigated in samples of whey, curd and a typical hard and long maturing cheese like Grana Padano (ripened for twelve months), produced with naturally contaminated milk in a range of 30–98 ng AFM₁/kg. AFM₁ determinations were carried out on 25 samples of each product by reverse-phase HPLC and fluorescence detection with post-column derivatisation, after a preliminary C18-SPE clean-up. Experimental results show that, in comparison to milk, AFM₁ concentration levels increased both in curd (3-fold) and in long maturing cheese (4.5-fold), while AFM₁ occurrence in whey decreased by 40%.     

Distribution and stability of aflatoxin M1 during processing, ripening and storage of Telemes cheese

Telemes cheeses were produced using milk that was artificially-contaminated with aflatoxin M1 at the levels of 0.050 and 0.100 microg/l. The cheeses produced in the two cheese-making trials were allowed to ripen for 2 months and stored for an additional 4 months to simulate commercial production of Telemes cheese. Concentrations of aflatoxin M1 in whey, curd, brine, and the produced cheeses were determined at intervals by liquid chromatography and fluorometric detection coupled with immunoaffinity column extraction. Concentrations of aflatoxin M1 in the produced curds were found to be 3.9 and 4.4 times higher than those in milk, whereas concentrations in whey were lower than those in curd and milk. Aflatoxin M1 was present in cheese at higher concentrations at the beginning than at the end of the ripening/storage period, and it declined to concentrations 2.7 and 3.4 times higher than those initially present in milk by the end of the sixth month of storage. Concentrations of aflatoxin M1 in brine started low and increased by the end of the ripening/storage period but only a portion of the amounts of aflatoxin M1 lost from cheese was found in the brine. Results showed that Telemes cheeses produced from milk containing aflatoxin M1 at a concentration close to either the maximum acceptable level of 0.05 microg/l set by the European union (EU) or at double this value, will contain the toxin at a level that is much lower or slightly higher, respectively, than the maximum acceptable level of 0.250 microg of aflatoxin M1/kg cheese set by some countries.     

Fate of aflatoxin M1 in cheese whey processing

Aflatoxin M₁ (AFM₁) is an important mycotoxin frequently found in milk and in dairy products. It is a minor metabolic product of Aspergillus flavus and A parasiticus. However, it occurs in dairy products as a metabolite formed in cows from aflatoxin B₁ contained in animal feeds. In cheese production, AFM₁ distributes between curd and whey, being present in products derived from cheese whey processing. In this study, cheese whey from dairy processing was artificially contaminated with the mycotoxin at about 0.1 µg l⁻¹. Ultra‐filtration experiments of whey were carried out in order to determinate AFM₁ distribution between retentate (protein‐rich fraction) and permeate (lactose‐rich fraction). Recoveries of AFM₁ in retentate were 72.6–86.4% while, in permeate, recoveries were in the range 2.4–14.7%. Partition coefficients of AFM₁, lactose and protein were calculated to determine whether there was an interaction between AFM₁ and protein. In all experiments, AFM₁ partition coefficient was lower than 1, whilst for lactose coefficients close to 1 were determined, showing an affinity of aflatoxin M₁ to the protein‐rich fraction (retentate). Copyright © 2005 Society of Chemical Industry     

Sterigmatocystin: Incidence,fate and production by Aspergillus versicolor in Ras cheese

The occurrence, distribution, and stability of sterigmatocystin (STG) in Ras cheese were investigated. An incidence value for STG in market samples of Ras cheese was 35% with a mean value of 22.2 μg/kg. In experimental Ras cheese from milk contaminated with STG, 80% of the toxin was retained in the curd while 20% was found in the whey. The temperature for cheese ripening affected the toxin content. At 6 °C the toxin concentration was hardly affected, but at 20 °C the concentration was reduced by 16% after 90 days. In Ras cheese contaminated with spores of Aspergillus versicolor, toxin production started after 45 days of the ripening, reached a maximum after 90 days, and declined thereafter. Cow's milk favoured toxin formation in comparison with buffaloe's milk. Aged cheese (more than 6 months) inhibited toxin production.     

Prediction and Characterization of Normal and Vat-Failed Cottage Cheese

An acidification-heat-coagulation test has been developed for predicting cottage cheese vat-failure potential of milk. Milk is fist acidified to pH 5.06 at 10°C and then heated at a slow rate (1°C increment per min). Poor quality acidified milk (> 10⁴ CFU/ml) forms small curds at 37°C and below. Good quality acidified milk (< 10⁴ CFU/ml) will form small curds at higher temperatures. By this procedure cottage cheese vat-failure potential of milk containing different levels of psychrotrophs can be predicted. Normal and vat-failed cottage cheese curds are characterized by % of grit in cottage cheese and amount of curd fines in whey.     

Role of salt in Iranian ultrafiltered Feta cheese: Some textural and physicochemical changes during ripening

The effect of NaCl (0–4%) on the ripening of Iranian ultrafiltered (UF)‐Feta cheese was assessed over 120 days of ripening at 4 °C. Whey percentage, whey salt, whey pH, cheese pH and textural properties of hardness and cohesiveness were monitored, and experimental modelling performed using response surface methodology. Texture, pH and whey percentage were significantly affected by NaCl and ripening. The maximum whey of 22% was recorded at the end of ripening period. Texture of this cheese becomes harder during ripening confirming cheese pH and whey percentage being the major determining factors. Cheese samples were more elastic than viscous with cohesiveness values of 0.6–0.9.     

The influence of milk pasteurization temperature and pH at curd milling on the composition, texture and maturation of reduced fat cheddar cheese

Reduced fat milks were pasteurized, for 15 s, at temperatures ranging from 72 to 88°C to give levels of whey protein denaturation varying from ˜ 3 to 35%. The milks were converted into reduced fat cheddar cheese (16–18% fat) in 500 litre cheese vats; the resultant cheese curds were milled at pH values of 5.75 and 5.35. Raising the milk pasteurization temperature resulted in impaired rennet coagulation properties, longer set-to-cut times during cheese manufacture, higher cheese moisture and moisture in the non-fat cheese substance, lower levels of protein and calcium and lower cheese firmness. Increasing the pH at curd milling from 5.35 to 5.75 affected cheese composition and firmness, during ripening, in a manner similar to that of increasing milk pasteurization temperature. Despite their effects on cheese composition and rheology, pasteurization temperature and pH at curd milling had little influence on proteolysis or on the grading scores awarded by commercial graders during ripening over 303 days .     

益生性植物乳杆菌对切达干酪挥发性风味形成的影响

将分离自西藏灵菇的益生性植物乳杆菌1-2通过在杀菌乳中添加活菌数8.0、9.0（lg（CFU/mL））和在排乳清后添加于凝乳块8.0（lg（CFU/g））的方式分别加入到切达干酪中,考察植物乳杆菌活菌数量、添加方式和成熟时间对干酪挥发性风味物质组成的影响。利用固相微萃取和气相色谱-质谱联用技术检测出对照组干酪的风味物质26种,益生菌干酪组风味物质30种,添加植物乳杆菌1-2可产生乙苯、十二烷、己醇和丙酮4种挥发性风味物质。成熟时间对干酪风味的影响最大,随成熟时间的延长,益生菌干酪组中苯含量显著增加,而对照组干酪在成熟12周时才检测到苯。益生菌添加量和添加方式对干酪挥发性风味的影响相似,丁酸受益生菌活菌数和添加方式的影响最大,益生菌干酪组成熟12周时,丁酸含量最高达对照组的3.96倍（P＜0.05）。在杀菌乳中添加益生菌活菌数8.0（lg（CFU/mL））组和9.0（lg（CFU/mL））组干酪中挥发性风味物质含量有显著差异,但在杀菌乳中添加高活菌数9.0（lg（CFU/mL））和在排乳清后添加低活菌数8.0（lg（CFU/g））于凝乳块中对干酪挥发性风味的形成具有相似的影响。本研究结果为改进益生菌干酪的加工工艺和风味品质提供了实验依据。     

Survey of aflatoxin M1 in milk and dairy products consumed in Burdur,Turkey

The aim of this study was to investigate the occurrence of aflatoxin M₁ (AFM₁) in dairy product samples in Burdur city. A total of 315 samples of dairy products were collected during 2008. Of the 315 samples analysed, AFM₁ in 246 samples (78.1%) was found to range from 5.5 to 800 ng/kg. In addition, AFM₁ levels of 16 raw milk, two pasteurised milk, only one milk powder and three white cheese samples were above the Turkish Food Codex. It is concluded that the occurrence of AFM₁ in dairy products may be considered as a possible hazard for public health.     

Recent aflatoxin survey data in milk and milk products: A review

Aflatoxin M₁ (AFM₁) occurrence in human and animal milk, infant formula, powdered milk, cheese and yoghurt represents a risk for health. The last four years (2010–2014) of data, as well as the most frequently and updated analytical methods applied for AFM₁ quantification, are evaluated. Aflatoxin B₁, considered the most potent toxic aflatoxin, is metabolised to form the monohydroxy derivative AFM₁. This metabolized, expressed in the milk, is relatively stable, and it is not eliminated by heat treatments or pasteurisation, and thus represents a serious health concern.     

Assessment of aflatoxin M1 levels in selected dairy products in north‐western Iran

The purpose of this survey was to evaluate the natural occurrence and content of aflatoxin M_1, AFM_1, in dairy products marketed in Urmia. During September 2007, 40 samples of pasteurised milk, 40 samples of ultra high temperature‐treated (UHT) milk, 40 samples of creamy cheese and 40 samples of Iranian Feta cheese were collected from different supermarkets in Urmia city. AFM₁ contents were determined by the competitive enzyme‐linked imunosorbent assay (ELISA) technique. All milk samples analysed showed a mean of AFM₁ concentrations lower than the permissible level of 50 ng/kg in Iran (23.22 and 19.53 ng/kg in pasteurised milk and UHT milk respectively). The mean levels of AFM₁ contamination were 43.31 ng/kg in Feta cheeses and 21.96 ng/kg in creamy cheeses. The potential risk of human exposure to aflatoxin M₁ via consumption of milk and milk products is well known. Dairy products must therefore be evaluated for aflatoxin and kept free from fungal contamination as much as possible.     

Reconstituted skim milk concentration affects partitioning of some individual caseins between rennet curd and whey

Milk composition is a major factor determining cheesemaking efficiency where caseins (CNs) play key roles, but little is known about individual partitioning during coagulation. This study evaluated the impact of reconstituted skim milk concentration (8%–25% solids) on the partitioning of individual CNs between rennet curd and whey produced with or without slight acidification. α_s2-CN and intact κ-CN fractions partitioned in rennet whey were less than 10% and decreased with rise in milk concentration, whereas α_s1-CN and β-CN appeared 100% retained into the curds. Loss of some α_s2- and κ-CN into rennet whey is attributed to their soluble complexes with whey proteins.     

Aflatoxin M1 determination and stability study in milk samples using a screen-printed 96-well electrochemical microplate

A highly sensitive method for the determination of aflatoxin M₁ (AFM₁) is described that involves the use of a disposable multichannel microplate coupled to intermittent pulse amperometry (IPA) for immunosensor development based on a direct competitive assay. Immunoassay parameters were evaluated and optimized to establish an electrochemical enzyme-linked immunosorbent assay (ELISA) procedure for AFM₁ in milk samples. The suitability of the immunosensors for the direct analysis of the toxin in milk was assessed. AFM₁ was measured with a working range of 5–250 pg mL⁻¹ and a detection limit of 1 pg mL⁻¹. Good recovery values (90–105%) were obtained. The method was used to study the recovery of this toxin in frozen (−30 °C) or lyophilized (4 °C) milk samples stored for up to 3 months. A significant, but variable, decrease (>50%) of the measured AFM₁ concentration with respect to the initial toxin contamination levels was found.     

Association of aflatoxin M1 with casein

Summary Equilibrium dialysis was used to determine whether aflatoxin M1 (AFM₁) binds to casein. Simulated milk ultrafiltrate (SMUF) containing 10 or 20 ng/ml of AFM₁ was dialyzed against SMUF containing casein micelles. After 24 h at 7°C, the casein suspension contained 2.5- or 2.9-fold, respectively, more toxin than that found in SMUF. An average of 17.9 or 55.3 g of AFM₁, respectively, per gram of casein was bound. In a separate experiment, milk naturally contaminated with AFM₁ was treated with a proteolytic enzyme. An average of 30.7% more toxin was found in treated than in untreated milk. This result also suggests binding of AFM₁ by milk protein.

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Verbindung zwischen Aflatoxin M₁ und Casein

Zusammenfassung Die Gleichgewichtsdialyse wurde benutzt, um festzustellen, ob es eine Bindung zwischen Aflatoxin M₁ (AFM₁) and Casein gibt. Simuliertes Milchultrafiltrat (SM) mit entweder 10 oder 20 ng von AFM₁/ml wurde gegen SM mit Caseinmicellen dialysiert. Nach 24 Std bei 7°C enthielt die Caseinsuspension 2,5- bzw. 2,9-mal so viel AFM₁ als SM. Durchschmittlich wurden 17,9 bzw. 55,3 g AFM₁/g Casein gebunden. In einem Einzelexperiment wurde Milch, die natürlich mit AFM, kontaminiert war, mit einem eiweißabbauenden Enzym behandelt. Im Durchschnitt fand man etwa 30,7% mehr Toxin in behandelter als in unbehandelter Milch. Dieses Ergebnis weist auf eine Verbindung zwischen AFM₁ and Casein hin.

     

Efficiency of Lactic Acid Bacteria Strains for Decontamination of Aflatoxin M1 in Phosphate Buffer Saline Solution and in Skimmed Milk

The aim of this study was to evaluate the ability of seven lactic acid bacteria (LAB) strains to remove aflatoxin M₁ (AFM₁) in phosphate buffer saline (PBS) and in skimmed milk samples. The mean AFM₁ removal by LAB in PBS ranged from 5.60?±?0.45% to 45.67?±?1.65% (n?=?3). Heat-killed cells showed AFM₁ removal percentages significantly higher than viable cells in contact times of 15 min or 24 h, although there were no significant differences between those times. AFM₁/LAB complex resulted from tests with PBS was unstable, as 40.57?±?4.66% to 87.37?±?1.82% of AFM₁ retained by bacteria were recovered in solution after washing with PBS. Heat-killed cells of Lactobacillus rhamnosus, Lactobacillus delbrueckii spp. bulgaricus, and Bifidobacterium lactis had the highest percentage (>33%) of AFM₁ removal in PBS tests. In ultrahigh temperature (UHT) skimmed milk spiked with AFM₁, the three selected LAB strains showed no significant differences in removing AFM₁ at 37 °C, and only B. lactis had greater ability to remove AFM₁ at 4 °C. Results demonstrated that AFM₁ removal by LAB has a potential application to reduce toxin concentrations until safe levels in milk.     

Fate of Aflatoxin M1 during Manufacture and Storage of Feta Cheese

ABSTRACT:  The effect of feta cheese manufacture on aflatoxin M₁ (AFM₁) content was studied using an enzyme immunoassay technique. Feta cheese was made from milk spiked with 1 and 2 μg AFM₁ per kilogram milk. Pasteurization at 63 °C for 30 min caused <10% destruction of AFM₁. During cheese making, the remaining AFM₁ in milk was partitioned between curd and whey with two-thirds retained in the curd and one-third going into the whey. Cheeses were then stored for 2 mo in 8%, 10%, and 12% brine solutions at 6 and 18 °C. There was a 22% to 27% reduction of AFM₁ during the first 10 d of storage, with slightly more loss as salt concentration increased and when the cheese was stored at 18 °C. Further storage caused only slight decrease in AFM₁ and after 30 d of brining there was no difference in AFM₁ content of the cheese based upon salt concentration of the brine. At 18 °C, no further losses of AFM₁ occurred after 30 d, and at 6 °C, there was continued slight decrease in AFM₁ levels until 50 d. After 60 d of brining, there was a total loss of 25% and 29% of the AFM₁ originally present for cheese brined at 6 and 18 °C, respectively. Thus, the combination of pasteurization, conversion of milk into feta cheese, and at least 50 d storage of cheese in brine caused a total loss of about 50% of the AFM₁ originally present in the raw milk.     

Determination of microbial contamination sources for use in quality management of cheese industry: “Dil” cheese as an example

The microbiological quality, safety and shelf-life of cheeses depend on manufacture and handling in an environment that meets basic standards for hygiene and the management of hygiene in the process. In this research contamination sources of “Dil” cheese during production in a local dairy plant in Bursa, Turkey were determined. Eighteen different control points (raw milk, pasteurized milk, heated curd, molded cheese before kneading, kneaded cheese, brine solution for kneading, thermophilic culture, rennet, calcium chloride solution, brine solution for cheese, cheese vat, workers hands, production room air, production room floor, production room wall, packaging material and packaged cheese) have been examined for the enumeration of total aerobic mesophilic bacteria, Staphylococci, Enterobacteriaceae, Salmonella spp., Escherichia coli, lactic acid bacteria, Pseudomonas spp. and yeast-moulds. It was determined that viability of lactic acid bacteria in thermophilic culture was not in high numbers and some contaminations to “Dil” cheese were detected from the starter culture. Brine solutions and rennet were contaminated with Staphylococci. Yeast and moulds in production room air were the major sources of contamination. Pasteurization and kneading in hot brine solution can eliminate some of the microorganisms but that was not sufficient in the production of Dil cheese. Finish cheese should meet specific hygienic standards, with respect to regulations post-contaminations to the cheese must be inhibited and a HACCP plan should be established during production.     

Distribution and stability of Aflatoxin M1 during processing and ripening of traditional white pickled cheese

The distribution of aflatoxin M₁ (AFM₁) has been studied between curd, whey, cheese and pickle samples of Turkish white pickled cheese produced according to traditional techniques and its stability studied during the ripening period. Cheeses were produced in three cheese-making trials using raw milk that was artificially contaminated with AFM₁ at the levels of 50, 250 and 750 ng/l and allowed to ripen for three months. AFM₁ determinations were carried out at intervals by LC with fluorescence detection after immunoaffinity column clean-up. During the syneresis of the cheese a proportionately high concentration of AFM₁ remained in curd and for each trial the level was 3.6, 3.8 and 4.0 times higher than levels in milk. At the end of the ripening, the distribution of AFM₁ for cheese/whey + brine samples was 0.9, 1.0 and 1.3 for first, second and third spiking respectively indicating that nearly half of the AFM₁ remained in cheese. It has been found that only 2-4% of the initial spiking of AFM₁ transferred into the brine solution. During the ripening period AFM₁ levels remained constant suggesting that AFM₁ was quite stable during manufacturing and ripening.     

Effects of somatic cell count and stage of lactation on the plasmin activity and cheese-making properties of ewe milk

The experiment was conducted from March to July 2002 using 5 intensively managed flocks of Southern Italy. In each flock, 2 groups of 50 ewes were created. The groups were designated LSCC (low somatic cell count [SCC]) when their milk SCC was lower than 500,000/mL and HSCC (high SCC) when their milk SCC was higher than 1,000,000/mL. Bulk milk and whey samples were analyzed for fat, total protein, lactose, casein, and whey protein contents. Renneting properties of milk were also determined. Moisture, NaCl, and nitrogen fractions were determined in fresh cheese curds. In addition, plasmin (PL) and plasminogen (PG) activities in milk and cheese were monitored. The proteolytic activity of plasmin by urea-polyacrylamide gel electrophoresis and the white blood cell (WBC) differentials were determined. The HSCC resulted in higher pH values in milk and in higher moisture and lower fat contents in fresh cheese curds. Moreover, a lower recovery of fat and whey proteins was obtained from the HSCC than from the LSCC raw milk. The crude protein and casein contents were higher in the HSCC than in the LSCC curds during early and midlactation; an opposite trend was observed in late lactation. Plasmin and PG activities underwent more marked fluctuations in the LSCC than in the HSCC curds through lactation. The results of this experiment demonstrate that the PL activity in ewe milk is markedly influenced by the SCC, although SCC is not the only parameter for predicting PL and PG evolution in ewe milk. The LSCC milk resulted in a higher proteolytic potential of Canestrato pugliese cheese curds.