Seasonal variation of aflatoxin M1 contamination in industrial and traditional Iranian dairy products

This study aimed to determine the occurrence of aflatoxin M₁ (AFM₁) contamination in 682 dairy product samples consisting of raw milk of cow, goat and sheep; Lighvan cheese; and industrial and traditional yoghurt, Kashk and Doogh samples collected from popular markets and dairy ranches in four large Iranian cities. Thin layer chromatography (TLC) technique was used for analysis of the samples. Results showed that the incidence and levels of AFM₁ contamination in raw cow milk and industrial products (manufactured from cow milk) were higher than raw goat or sheep milk, and traditional products (made from goat and sheep milk), respectively. Moreover, seasonal variations influenced the concentration of AFM₁ in most of the analyzed dairy products. Owing to the abundance and popularity of the industrial products, contamination of these products in such a level could be a potential hazard for public health.     

Effect of seasonal variations and lactation times on aflatoxin M1 contamination in milk of different species from Punjab, Pakistan

During October 2009 to September 2010, aflatoxin M₁ (AFM₁) levels were analyzed by HPLC-FLD in 356 milk samples of different lactating species (buffalo, cow, goat, sheep and camel) from Punjab (Pakistan). Recoveries of AFM₁ ranged from 92 to 97% and the limit of detection (LOD) was 0.004 μg/L. For all lactating species the mean concentration of AFM₁ was significantly higher in winter season than in summer (p < 0.05). The results showed that 55, 56, 32, 58 and 27% of winter milk samples of buffalo, cow, goat, sheep and camel exceeded the EU maximum limit (0.05 μg/kg), compared with 38, 33, 21, 36 and 14% of summer milk samples, respectively. For all lactating species the mean concentration of AFM₁ was significantly higher in morning milks than in evening milks (p < 0.05). The percentage of morning milk samples exceeding the EU maximum limit was 72, 67, 69, 71 and 44% for buffalo, cow, goat, sheep and camel, while for evening milks percent non compliant rates were 39, 30, 18, 33 and 25%, respectively. The level of AFM₁ tended to be higher in animal species fed mainly on concentrate mixtures (buffalo and cow) than in other species grazing on fresh greens.     

Aflatoxin M1 in milk and traditional dairy products from west part of Iran: Occurrence and seasonal variation with an emphasis on risk assessment of human exposure

In the present study, a total of 358 samples consisting of raw milk of cow (n = 64), goat (n = 56) and sheep (n = 52); traditional cheese (n = 40), yoghurt (n = 42), Kashk (n = 40), Doogh (n = 44) and Tarkhineh (n = 20) were analyzed for aflatoxin M₁ (AFM₁) by using an enzyme linked immunosorbent assay (ELISA). Frequency of AFM₁ and its concentration ranges in the ELISA positive samples were determined by high performance liquid chromatography with fluorescence detection (HPLC-FD). AFM₁ contamination was 84.3%, 44.6% and 65.3% for cow, goat and sheep raw milks, respectively. Moreover, AFM₁ was in 65.5%, 23.8%, 14%, 13.6% and 35.0% of cheese, yoghurt, Kashk, Doogh and Tarkhineh samples, respectively. Percentages of cow milk, goat milk, sheep milk and cheese samples exceeding the EU limit were 35.9%, 11.1%, 26.9% and 10%, respectively. HPLC analyses confirmed the ELISA results although the percentages of AFM₁ contamination in raw milk and dairy products were lower than that of ELISA. There were significant differences (P < 0.05) between the mean AFM₁ contents of raw milk, cheese and yoghurt samples during winter and summer seasons. Our study demonstrated that there is a potential risk for liver cancer due to the consumption of milk and dairy products in Iranian consumers.     

Aflatoxin M1 contamination in milk from five dairy species in Pakistan

Aflatoxin M₁ (AFM₁) in milk samples of five different species of mammals from the area of Faisalabad district of the Punjab province of Pakistan was determined by using high-performance liquid chromatography (HPLC) with fluorescent detection. Immunoaffinity columns were used for clean-up purposes which are based on the principle of affinity chromatography. The present study has been designed to find the milk which is safe from aflatoxin contamination point of view. Total 169 milk samples were analyzed and these were taken from five species made up of 55, 40, 30, 24, and 20 samples of buffaloes, cows, goats, sheep, and camels respectively. The percentage of AFM₁ contamination in buffalo milk, cow milk, goat milk, and sheep milk has been found to be 34.5%, 37.5%, 20%, and 16.7%, respectively. AFM₁ contamination has not been detected in the camel milk.     

Variation of levels of aflatoxin M1 in raw milk from different localities in the central areas of Punjab, Pakistan

The occurrence of aflatoxin M₁ (AFM₁) in samples of raw milk of buffaloes and cows from different localities in the central areas of Punjab, the province of Pakistan, was determined by using high-performance liquid chromatography (HPLC) with prior clean-up step applying immunoaffinity columns. The present study has been designed to find out the variation of levels of aflatoxin M₁ in raw milk of different localities. Feed has major role for prevalence of aflatoxin M₁ in milk and different feed regimen is being used in different areas. Total 480 milk samples were analyzed, among these 360 were of buffalo milk and 120 were of cow milk. The percentage of AFM₁ contamination in buffalo milk and cow milk was 42.5% and 52.5%, respectively. The mean value of AFM₁ was 0.027 μg L⁻¹ in buffaloes’ milk and was 0.044 μg L⁻¹ in cows’ milk. In both types of milk, level of AFM₁ concentration was higher in milk samples obtained from urban and semi-urban areas and it was minimal in milk from rural areas.     

Occurrence and factors associated with the presence of aflatoxin M1 in breast milk samples of nursing mothers in central Mexico

Aflatoxin M₁ (AFM₁) is the main mycotoxin frequently found in milk and dairy products. Exposure of infants to AFM₁ is of great health public concern because of its toxic and carcinogenic properties. The aim of this study was to determine the occurrence, levels and factors associated with the presence of aflatoxin M₁ in breast milk samples of nursing mothers in central Mexico. A total of 112 samples were collected and analyzed by a competitive-ELISA assay during January–August 2014. A Semi-Quantitative Food Frequency Questionnaire was used to determine weekly intake of food groups. Results showed that 89% of breast milk samples contained AFM₁ in a range of 3.01–34.24 ng/L. AFM₁ mean level was significantly higher (P < 0.05) in winter (12.78 ng/L) and spring (12.09 ng/L) than summer (7.91 ng/L). It was also detected that 7% of samples exceeded the European Community regulation (25 ng/L) for milk infant formula. The presence of aflatoxin M₁ was significantly associated (P < 0.05) to consumption of egg, cola drink and sunflower oil. The high occurrence and noticeable levels of AFM₁ detected, as well as the mean AFM₁ estimated daily intake (2.35 ng/kg b.w./day) suggested, for the first time, that breast-fed infants in the central region of Mexico, may be exposed to significant levels of this toxin through mother's breast milk.     

Screening of aflatoxin M1 occurrence in selected milk and dairy products in Terengganu,Malaysia

The study was conducted to screen the occurrence of aflatoxin M₁ (AFM₁) in 53 selected milk and dairy product samples (11 liquid milk, 12 powdered milk, 8 3-in-1 beverages, 6 condensed sweetened milk, 2 evaporated milk, 7 cultured milk drink, 5 yogurt and 2 cheese samples). These samples were purchased from selected markets in Terengganu, Malaysia in January 2014 based on a questionnaire survey among 212 respondents on the types and brands of milk and dairy products that were frequently consumed. Based on the responses, 53 milk and dairy products were purchased and the competitive enzyme-linked immune-absorbent assay (ELISA) method was used to determine the level of AFM₁ in the samples. Of 53 samples, 19 samples were positive with AFM₁ (35.8%) ranging from 3.5 to 100.5 ng/L. Although 4/53 (7.5%) of the tested samples had the contamination level greater than the European Commission (EC) limit (>50 ng/L), the contamination levels were still below the Malaysia Food Regulation 1985 limit (less than 500 ng/L). This study provided a pioneering data on the occurrence of AFM₁ in milk and dairy products in Malaysia.     

Assessment of aflatoxin M1 contamination in the milk of four dairy species in Croatia

Aflatoxin M1 (AFM1) concentrations were measured in bulk cow milk samples from eastern Croatia, and in cow, goat, sheep and donkey bulk milk samples from other parts of Croatia during the period July–September 2013. AFM1 levels in milk were measured in the ranges (ng/L): cow 3.65–162.3 (eastern Croatia) and 2.69–44.9 (other regions of Croatia); goat 2.78–40.8; sheep 2.11–5.87; donkey 3.43–10.4. The concentration of AM1 exceeded the EU MRL in 6.7% of cow milk samples from eastern Croatia. The highest level measured was 162.3 ng/L. AFM1 levels exceeded the LOQ value (23.2 ng/L) in only 59 samples of cow milk and two samples of goat milk of the total 402 samples analysed. A significant difference was found between the mean AFM1 concentrations of cow milk from eastern and other regions of Croatia (P < 0.05). The elevated AFM1 levels in cow milk from eastern Croatia indicate the use of contaminated supplementary feedstuff in some farms during the study period.     

Seasonal variation of aflatoxin M1 in raw milk from the Yangtze River Delta region of China

The objective of this study was to evaluate the occurrence of aflatoxin M₁ (AFM₁) in raw milk samples from 18 dairy farms in the Yangtze River Delta region during four different seasons. A total of 72 tank milk samples was collected with 18 samples for each season. Milk AFM₁ was detected using LC-MS/MS. The AFM₁ was detected in 43 milk samples (59.7%) ranging in concentration from 10 to 420 ng/L. The concentration of AFM₁ in raw milk was significantly higher during the winter (123 ng/L) than during other seasons (P < 0.05). There was no significant difference between the spring (29.1 ng/L), summer (31.9 ng/L), and autumn (31.6 ng/L) (P > 0.05) seasons. This indicates that raw milk collected during the winter is at high risk for AFM₁ and that seasonal factors should be considered for the management of aflatoxins in both the feed and milk.     

The seasonal variation of aflatoxin M1 in milk and dairy products and assessment of dietary intake in Punjab,Pakistan

Total 372 samples (169 sample from summer and 203 samples from winter) of milk and dairy products were collected from major cities of Punjab, Pakistan, during 2014–2015. The aflatoxin M₁ (AFM₁) levels were analyzed using HPLC, equipped with floresceⁿce detector. The results have showed that AFM₁ was detected in 77 (45.5%) samples of milk and dairy products from summer at the range of LOD to 229.6 ng/L, and 114 (56.1%) samples from winter at a range of LOD to 345.8 ng/L. The highest mean of AFM₁ i.e. 94.9 ± 5.4 ng/L from summer and 129.6 ± 8.4 ng/L from winter was detected in raw milk samples. The results have shown that 37.5% of milk, 32% of UHT milk, 12.5% of powdered milk, 20% of flavored milk, 20% of yogurt and 16% of flavored milk samples from summer were found exceeded the recommended levels of AFM₁ i.e 50 ng/L (European Union permissible limit), as compared to 38.1%, 37.1%, 15.6%, 21.4%, 27.7% and 40% samples of milk, UHT milk, powdered milk, flavored milk, yogurt and flavored yogurt from winter, respectively. The mean level of AFM₁ in samples of milk, UHT milk and powdered milk samples from winter season has significantly (p˂ 0.05) higher as compared to the mean levels in samples from summer season. The dietary exposures of AFM₁ in milk and dairy products were calculated in five different age groups and the highest level of 3.42 and 3.33 ng/L/day in summer and 4.7 and 4.6 ng/L/day in winter were found in 1–5 years of male and female infants, respectively. The results of present study have shown significant levels of AFM₁ in milk and dairy products from Punjab, Pakistan and could pose considerable thereat to the health of local population.     

Occurrence of aflatoxin M1 in white cheese samples from Tehran, Iran

This study was undertaken to determine the occurrence of aflatoxin M₁ (AFM₁) in 50 white cheese samples from 2 dairy factories in summer 2008 and winter 2009. Enzyme-linked immunosorbent assay (ELISA) method was used for analysis of the samples. Aflatoxin M₁ was found in 60% of the cheese samples, ranging from 40.9 to 374 ng/kg. Toxin levels in 6% of the samples exceeded the Iranian national standard limit i.e. 200 ng/kg. Considering seasonal variability, mean concentration of AFM₁ in the samples collected in winter was significantly (P < 0.03) higher than those collected in summer. Therefore, high occurrence of AFM₁ in cheese samples could be a potential hazard for public health.     

Occurrence of aflatoxin M1 from rural subsistence and commercial farms from selected areas of South Africa

Aflatoxin M₁ (AFM₁) is a hydroxylated metabolite of aflatoxin B₁ and its presence in milk is considered to be a potential health risk for humans. Due to the important role of milk in humans, especially in infant nutrition, this study is intended to evaluate the quality of milk consumed on daily basis in South Africa by both rural and urban population in regard to AFM₁ contamination. To achieve this, samples were collected from rural subsistence (RSFs) and commercial dairy farms (CDFs) in selected areas of South Africa and samples were extracted using two clean-up procedures, C₁₈ cartridges and immunoaffinity columns (IAC) and analysed using thin layer chromatography (TLC) or by fluorometry (VICAM) (VF) and high pressure liquid chromatography (HPLC) coupled with a fluorescence detector and a coring cell (CoBrA cell) for AFM₁ derivatisation. Results obtained showed a frequency of contamination with AFM₁ of RSFs milk samples at 22.8% by TLC, 93.9% by VF 86% by HPLC and in CDFs of 17.8% (TLC) 96.5% (VF) and 100% (HPLC). No significant differences were obtained between milk from rural subsistence and commercial farms with mean varying between 0.15 and 0.17 μg kg⁻¹     

Aflatoxin M1 in UHT milk consumed in Turkey and first assessment of its bioaccessibility using an in vitro digestion model

Contamination of milk and dairy products with aflatoxin M₁ (AFM₁) continues to receive increased attention because of its potential health hazard to humans. The first aim of this study was to know the occurrence and levels of AFM₁ in whole UHT milk from main processors in Turkey in order to make a preliminary exposure assessment. A total of 40 milk samples were analysed for AFM₁ using high performance liquid chromatography with fluorescence detection (HPLC-FD) after immunoaffinity column clean-up. Aflatoxin M₁ was detected in 20% of samples at levels ranging from <0.004 to 0.076 μg l⁻¹. Only two samples contained AFM₁ above the EU limit of  0.05 μg l⁻¹.The second aim of this study was to determine the bioaccessibility of AFM₁ from milk using an in vitro digestion model. The bioaccessibility of AFM₁ in spiked and naturally contaminated milk samples ranged from 80.5 to 83.8% and from 81.7 to 86.3%, respectively. No difference (P > 0.05) in AFM₁ bioaccessibility was found between spiked and naturally contaminated milk samples. This study also assessed the binding of AFM₁ by six probiotic bacteria under simulated gastrointestinal conditions. A 15.5–31.6% reduction in AFM₁ bioaccessibility was observed in the presence of probiotic bacteria. Based on the results obtained in the present study, the mean daily intake of AFM₁ through milk consumption was estimated as 0.008 ng kg⁻¹ b.w. day⁻¹ for Turkish adults.     

Analysis of aflatoxin M1 and M2 in commercial dairy products using high-performance liquid chromatography with a fluorescence detector

Aflatoxin M₁ (AFM₁) and M₂ (AFM₂) in commercial dairy products were analyzed by high-performance liquid chromatography (HPLC) with a fluorescence detector (FLD). To ensure an accurate analysis, two derivatization methods, bromination and aflatoxin–trifluoroacetic acid derivatization (ATD), were compared. The limits of detection (LODs) of the bromination method were 124.42–151.73 ng/kg, and the recovery rates were between 64 and 102%. The detection rates and concentration levels of AFM₁ were 6–74% and 14.48–270.94 ng/kg, respectively. AFM₁ was detected in 74% of milk powder samples and 36% of ice cream samples. The mean values of AFM₁ in milk powder and ice cream samples were 270.94 and 33.16 ng/kg, respectively. In the case of AFM₂, the detection rates were 2–10%, and the concentration levels were 20.62–55.67 ng/kg in milk and milk powder. Among milk and milk powder samples, ultra heat-treated (UHT) milk had lower AFM₁ contamination levels than pasteurized milk.     

Aflatoxin M1 in raw,pasteurized and powdered milk available in the Syrian market

The incidence of contamination of aflatoxin M1 (AFM1) in milk samples collected from the Syrian market was investigated by using the competitive enzyme linked immunosorbent assay (ELISA) technique. A total of 126 samples composed of raw cow milk (74 samples), raw sheep milk (23), raw goat milk (11), pasteurized cow milk (10) and powdered milk (8) showed that 80% of tested samples were contaminated with various levels of AFM1 ranging from >20 to 765 ng/l. Percentages of AFM1-contaminated samples exceeding the American, Syrian and European tolerance limits were 22%, 38% and 52%, respectively.The range of contamination was relatively higher in pasteurized milk than in raw cow and sheep milk. 80% of AFM1-contaminated pasteurized cow milk samples exceeded the European tolerance limit with a range of contamination between 89 and 765 ng/l. Percentages of contaminated raw cow, sheep and goat milk exceeding the European tolerance limit were 59%, 24% and 14%, respectively.Milk powder was almost free of AFM1 contamination with only one sample containing a concentration lower than the European tolerance limit (12 ng/l).Extrapolation of aflatoxin B1 (AFB1) from AFM1 levels of contamination in milk samples indicates that contamination in dairy cattle feeds may range from 0.5 to 47.8 μg/kg.     

Occurrence of aflatoxin M1 in raw milk in South Korea using an immunoaffinity column and liquid chromatography

The level of aflatoxin M₁ (AFM₁) contamination in raw milk produced in South Korea was investigated using immunoaffinity column chromatography and high performance liquid chromatography with fluorescence detector. A total of 100 raw milk samples were collected from 100 cattle ranches located in three different provinces of South Korea. Forty eight out of 100 raw milk samples contained AFM₁ at low level (0.002–0.08 μg/L) with mean value of 0.026 μg/L. Among the AFM₁ contaminated samples, 29 raw milk samples contained only traceable amount of AFM₁ below the limit of LOQ, 0.02 μg/L. None of samples exceeded the maximum level (0.5 μg/L) of Korean regulation for AFM₁ in milk. The limit of detection was 0.002 μg/L. The result of recovery test with 0.5 μg/L AFM₁ in raw milk sample was 96.3% (SD 3.6, n = 5). This is the first pioneering study to investigate the level of AFM₁ in raw milk used in dairy industries in South Korea.     

Contamination of UHT milk by aflatoxin M1 in Iran

In this study the levels of aflatoxin M₁ (AFM₁) in UHT milk samples were determined in May, August and November and February. Two hundred and ten UHT milk samples were obtained from supermarkets in Tehran, Iran. The occurrence and concentration range of AFM₁ in the samples were investigated by competitive enzyme-linked immunoabsorbent assay (ELISA) method. AFM₁ was found in 116 (55.2%) of 210 UHT milk samples examined. The levels of AFM₁ in 70 (33.3%) samples were higher than the maximum tolerance limit (0.05 μg/l) accepted by some European countries while none of the samples exceeded the prescribed limit of US regulations. The highest mean concentration of AFM₁ was recorded in February (0.087 μg/l). The lowest mean concentration of AFM₁ was recorded in August (0.021 μg/l). Statistical evaluation showed that there were significant difference (P < 0.01) between the mean concentrations of AFM₁ of UHT milk samples taken in February with May and August. AFM₁ contents of milk samples taken in February were not higher than UHT milk samples taken in November (P < 0.01). The AFM₁ incidence of exceeding legal limit in UHT milk samples (33.3%) was relatively much higher than some other countries. It was therefore concluded that, the levels of AFM₁ in UHT milk samples consumed in Iran were high and seemed to pose a threat to public health.     

Occurrence of aflatoxin M1 in dairy products in Brazil

The objective of this study was to investigate the incidence and occurrence of aflatoxin M₁ (AFM₁) in dairy products produced in Brazil. A total of 123 samples of three different groups of dairy products (cheese, yoghurt, and dairy drinks) consumed by Brazilians were collected during 2010. All samples including 58 cheese samples, 53 samples of yoghurt and 12 dairy drinks were purchased from grocery stores in the Ribeirão Preto-SP area. Cheese samples were classified into three categories depending on their moisture and fat contents: Minas Frescal cheese, Minas Frescal light cheese and Minas Padrão cheese. Samples were analyzed for AFM₁ by a published method. The method comprised aqueous methanol extraction, immunoaffinity column purification and isolation, reversed phase liquid chromatography separation and fluorescence detection. AFM₁ was detected in 84% of the analyzed cheese samples (>3 ng/kg) with levels ranging from 10 to 304 ng/kg in 67% of the samples. AFM₁ was detected in 95% of the yoghurt and dairy drink samples with levels ranging from 10 to 529 ng/kg in 72% of the samples. Despite the lack of a Brazilian regulatory limit for AFM₁ in yoghurt and dairy drinks the survey data of this study may offer information useful in the determination of whether the occurrence of AFM₁ in Brazilian dairy products may be considered as a possible risk for consumer health and whether Brazilian regulatory guidelines for AFM₁ in dairy products are needed.     

Occurrence of aflatoxin M1 in pasteurised,UHT milk and milk powder from goat origin

In the present study, 36 samples of pasteurised, ultra-high-temperature (UHT) treated and goat milk powder traded in the city of Campinas, Brazil, were analysed for aflatoxin M₁ (AFM₁), from October to December 2004 and March to May 2005. Results showed 25 (69.4%) positive samples for AFM₁ at levels of 0.011–0.161 μg L⁻¹ of milk, which were below the tolerance limit of 0.500 μg L⁻¹ as adopted for AFM₁ in milk by Brazilian regulations. Mean levels of AFM₁ in pasteurised, UHT and goat milk powder were 0.072 ± 0.048, 0.058 ± 0.044 and 0.056 ± 0.031 μg L⁻¹, respectively. It is concluded that the incidence of AFM₁ in goat milk traded in Campinas is high, but at levels that probably leads to a non-significant human exposure to AFM₁ by consumption of goat milks.     

Aflatoxin M1 in milk and distribution and stability of aflatoxin M1 during production and storage of yoghurt and cheese

The objective of this study was to investigate the incidence and occurrence of aflatoxin M₁ (AFM₁) in Brazilian milk and infant formula. The distribution and stability of AFM₁ in cheese and yoghurt were also determined. Milk samples and infant formula samples were purchased in Ribeirão Preto-SP, Brazil and were analyzed for AFM₁ using immunoaffinity column purification, liquid chromatography separation and fluorescence detection. AFM₁ was detected in 83% of the milk samples (>3 ng/kg) with levels ranging from 8 to 437 ng/kg for fluid milk, and 20–760 ng/kg for powdered milk. No AFM₁ was found in infant formula. Processing and storage was shown to have little effect on AFM₁ content in milk and milk products. Total AFM₁ mass in milk was reduced by 3.2% in cheese and by 6% in yoghurt (pH 4.4). The mean concentration of AFM₁ in curds was 1.9-fold higher and whey was 0.6-fold lower than in unprocessed milk.