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Several toxic metabolites, such as aflatoxin M1 (AFM1), are known to contaminate dairy milk. However, as mentioned in an external EFSA report, there is a knowledge gap regarding the carry-over of certain emerging toxins such as microcystin-LR (MC-LR). Therefore, this work aimed to develop an LC-MS/MS method for MC-LR quantification in dairy milk. Also, the method included AFM1 as a common fungal metabolite and applied to analyze 113 dairy milk samples collected directly after the end of the summer peak. Both toxins were below their LODs, keeping the question on MC-LR carry-over still unanswered. Moreover, an in silico analysis, using a 3D molecular modeling was performed, pointing to a possible interaction between MC-LR and milk proteins, especially β-lactoglobulin. Since AFM1 and MC-LR are hepatotoxic, their interaction in inducing mitochondrial dysfunction in HepG2 cells was investigated at low (subcytotoxic) concentrations. Live cell imaging-based assays showed an inhibition in cell viability, without involvement of caspase-3/7, and a hyperpolarization in the mitochondrial membrane potential after the exposure to a mixture of 100 ng mL -1 AFM1 and 1000 ng mL -1 MC-LR for 48h. Extracellular flux analysis revealed inhibitions of several key parameters of mitochondrial function (basal respiration, ATP-linked respiration, and spare respiratory capacity)., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this study., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Milk is an essential part of the human diet and is a nutrient-rich food that improves nutrition and food security. The aim of this study was to determine the presence and concentration of aflatoxin M1 (AFM1), adulterants, microbial loads, and physicochemical properties of raw cow's milk (CM) in Nekemte City, Ethiopia. A total of 12 samples of fresh CM were purposefully collected from four kebeles in the city (Bake Jama, Burka Jato, Cheleleki, and Bakanisa Kese) based on the potential of each milk production and distributor site. The AFM1 concentration was determined by high-performance liquid chromatography (HPLC) with a Sigma-Aldrich standard (St. Louis, MO, USA). The concentrations of AFM1 in Bake Jama, Burka Jato, Cheleleki, and Bakanisa Kese were found to be 0.01-0.03 g/L, 0.31-0.35 g/L, 0.19-0.21 g/L, and 0.04-0.07 g/L, respectively. The concentrations of AFM1 in the present study varied significantly ( p < 0.05) and ranged from 0.01 g/L to 0.35 g/L. These results show that of the 12 samples tested, all were positive for AFM1 and contaminated to varying degrees. The results of this study also revealed that the concentration of AFM1 in 7 (58%) of the 12 milk samples was above the European Union's (EU) maximum tolerance limit (0.05 g/L). The present study also revealed that of the investigated adulterants, only the addition of water had positive effects on three milk samples, while the remaining adulterants were not detected in any of the milk samples. The total bacterial count (TBC) and total coliform count (TCC) were significantly ( p < 0.05) different and ranged from 5.53 to 6.82 log 10 cfumL -1 and from 4.21 to 4.74 log 10 cfumL -1 , respectively. The physicochemical properties of the milk samples in the present study were significantly ( p < 0.05) different and ranged from 2.8% to 5.75% fat, 7.03% to 9.75% solid-not-fat (SNF), 2.35% to 3.61% protein, 3.33% to 5.15% lactose, 11.54% to 13.69% total solid, 0.16% to 0.18% titratable acid, 26.7 to 32.1°C, 6.35 to 6.55 pH, and 1.027 to 1.030 specific gravity. The physicochemical parameters of the raw milk in the study area met the required quality standards. Hence, further studies are required to determine the extent of the problem and the factors associated with high levels of AFM1 in raw milk in the study areas, including the detection of aflatoxin B1 (AFB1) in animal feed., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 Leila Nasir and Habtamu Fekadu Gemede.)

Infant formula in liquid for childcare can be stored at room temperature for a certain period of time, reducing the burden of childcare and preparing for disasters. Against this background, domestic manufacturing and sales began in March 2019. AFM 1 is a metabolite of aflatoxin B 1 (AFB 1 ), a carcinogenic mycotoxin, and is contained in the milk of livestock fed a diet contaminated with AFB 1 . At present, standard values have not been set for infant formula in liquid as well as prepared infant formula in liquid, and infants consume a large amount of dairy products per body weight, so care must be taken in the intake.In this study, we investigated the actual condition of AFM 1 content in dairy products with high intake of infants. As a result of the investigation, the AFM 1 of the detected dairy products was 0.001 to 0.005 μg/kg, which was extremely small compared to the AFM 1 in the dairy products reported so far. Since infant nutrition depends on dairy products, it is undeniable that they may consume more than adults, so continuous research is needed.

The present study was conducted to estimate the exposure and characterize the risk for the child population of Serbia to Aflatoxin M1 (AFM1) from milk and milk-based food. A total of 3404 samples comprising milk and different milk-based food samples were collected from various regions of Serbia from 2017 to 2019. Evaluation of AFM1 exposure was carried out using the deterministic method, whereas risk characterization was evaluated using the margin of exposure (MOE) and the risk of hepatocellular carcinoma (HCC). Detection rates for AFM1 in milk and milk-based food samples ranged between 2% and 79%, with the highest incidence (79%) and mean level (22.34 ± 0.018 ng kg -1 ) of AFM1 being detected in pasteurized and UHT milk. According to the three consumption estimates, the values of estimated daily intake (EDI) were higher for toddlers as compared with children aged 3-9 years. Children aged 1-3 years had the highest risk of exposure to AFM1 in milk, with an estimated daily intake of 0.164 and 0.193 ng kg -1 bw day -1 using lower bound (LB) and upper bound (UB) exposure scenarios, respectively. Such difference could result from the higher consumption to weight in younger children. Based on the estimated daily intake (EDI) found in this study, the risk of AFM1 exposure due to consumption of milk and milk-based food was low since the MOE values obtained were >10,000. In addition, the risk of HCC cases/year/10 5 individuals of different age groups showed that the value of HCC, using potency estimates of 0.0017 (mean), was maximum (0.00034) in the age group 1-3 years, which indicates no health risk for the evaluated groups. The present study revealed the importance of controlling and preventing AFM1 contamination in milk through continuous monitoring and regular inspection to reduce the risk of AFM1 exposure, especially in children.

Aflatoxins, which commonly contaminate animal feeds and human food, present a major public health challenge in sub-Saharan Africa. After ingestion by cows, aflatoxin B1 is metabolized to aflatoxin M1 (AFM1), some of which is excreted in milk. This study involved smallholder dairy farms in urban and periurban areas of Nairobi and Kisumu, Kenya. The objective was to determine the effectiveness of training and providing farmers with aflatoxin binder (NovaSil ® ) on AFM1 contamination in raw milk. A baseline survey was undertaken and 30 farmers whose milk had AFM1 levels above 20 ppt were randomly selected for inclusion in the study. Of these, 20 farmers were part of the intervention, and were given training on the usage of the NovaSil ® binder, while 10 served as a control group. All farmers were visited biweekly for three months for interviews and milk samples were collected to measure the AFM1 levels. The AFM1 levels were quantified by enzyme linked immunosorbent assay. The NovaSil ® binder significantly reduced AFM1 concentrations in the raw milk produced by the farmers in the intervention group over the duration of the study ( p < 0.01). The control farms were more likely to have milk with AFM1 levels exceeding the regulatory limit of 50 ppt compared to the intervention farms ( p < 0.001) (odds ratio = 6.5). The farmers in the intervention group perceived that there was an improvement in milk yield, and in cow health and appetite. These farmers also felt that the milk they sold, as well as the one they used at home, was safer. In conclusion, the use of binders by dairy farmers can be effective in reducing AFM1 in milk. Further research is needed to understand their effectiveness, especially when used in smallholder settings.

Aflatoxins are the most harmful mycotoxins causing health problems to human and animal. Many acute aflatoxin outbreaks have been reported in Africa, especially in Kenya and Tanzania. When ingested, aflatoxin B1 is converted by hydroxylation in the liver into aflatoxin M1, which is excreted in milk of dairy females and in urine of exposed populations. This review aims to highlight the AFM1 studies carried out in African regions (North Africa, East Africa, West Africa, Central Africa, and Southern Africa), particularly AFM1 occurrence in milk and dairy products, and in human biological fluids (breast milk, serum, and urine) of the populations exposed. Strategies for AFM1 detoxification will be considered, as well as AFM1 regulations as compared to the legislation adopted worldwide and the assessment of AFM1 exposure of some African populations. Egypt, Kenya, and Nigeria have the highest number of investigations on AFM1 in the continent. Indeed, some reports showed that 100% of the samples analyzed exceeded the EU regulations (50 ng/kg), especially in Zimbabwe, Nigeria, Sudan, and Egypt. Furthermore, AFM1 levels up to 8,000, 6,999, 6,900, and 2040 ng/kg have been reported in milk from Egypt, Kenya, Sudan, and Nigeria, respectively. Data on AFM1 occurrence in human biological fluids have also shown that exposure of African populations is mainly due to milk intake and breastfeeding, with 85-100% of children being exposed to high levels. Food fermentation in Africa has been tried for AFM1 detoxification strategies. Few African countries have set regulations for AFM1 in milk and derivatives, generally similar to those of the Codex alimentarius, the US or the EU standards., (© 2021. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Aflatoxin M1 (AFM1) residues in milk pose a major threat to human health, so there is an urgent need for a simple, rapid, and sensitive method for the determination of trace AFM1 in milk. In this study, a competitive immunochromatographic assay (ICA), using visual (PEI/PSS) 4 red silica nanoparticles (SiNPs) as signal amplification probes, was used for the highly sensitive detection of AFM1. The (PEI/PSS) 4 red SiNPs were used to label AFM1 monoclonal antibody (mAb) to prepare ICA for the detection of AFM1. After exploring the optimal conditions of mAb and immunoprobe dosage conditions, the lowest visual detection limit (VDL) of AFM1 in phosphate-buffered saline with Tween 20 (PBST, 10 mM, pH 7.4, containing 1% BSA, 3% sucrose, 1% trehalose, and 0.5% Tween 20) can reach 0.1 pg/mL. The intuitive visually visible value of AFM1 in both PBST and milk was 10 pg/mL. The results showed that the immunochromatographic system based on high chroma color (PEI/PSS) 4 red SiNPs has high sensitivity and broad application prospects for the detection of trace AFM1 residues in milk. The high chroma (PEI/PSS) 4 red SiNPs are expected to be a convenient biomarker for improving the sensitivity of immune chromatography bands. Graphical abstract The schematic diagram shows the detection principle. In this work, in the competitive experiment, (PEI/PSS) 4 red SiNPs were selected as visual labeling materials, and the specific antibody combined with the labeled material was selected as an immune probe. The AFM1-BSA antigen coupled with the macromolecular BSA was fixed on the T line of the nitrocellulose (NC) membrane. The AFM1 in sample solution competes with AFM1-BSA for the specific binding site of immune probe. The detection sensitivity of this method for AFM1 is obtained by judging the change of the red signal intensity produced by the positive sample, compared with the color at the T line of the negative sample.

In this study, we developed an indirect competitive plasmonic immunoassay using glucose oxidase (GOx)-induced redox reaction to etch Au nanorods (AuNRs) for qualitative and quantitative detection of aflatoxin M1 (AFM1) in milk. In this system, streptavidin (SA) was selected as a linker between biotinylated anti-AFM1-monoantibody (bio-mAb) and biotinylated GOx (bio-GOx) to form the immunocomplexes bio-mAb-SA-bio-GOx. After the oxidation of the glucose and I - , the resultant I 2 could etch cetytrimethylammonium bromide (CTAB)-stabilized AuNRs. This resulted in the blue shift of the longitudinal localized surface plasmon resonance (LSPR) extinction peak, with a color change from blue to pink. The linear range and limit of detection (LOD) of the plasmonic immunoassay were 0.25-10 ng mL -1 and 0.11 ng mL -1 , respectively. It displayed quantitative detection with high sensitivity, specificity, recovery, and accuracy, which is promising for qualitative and quantitative detection of AFM1 in food safety., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

A two-analyte immunochromatographic strip (immunostrip) was developed for the simultaneous detection of aflatoxin M1 (AFM1) and chloramphenicol (CAP) in milk. Protein conjugates (AFM1-ovalbumin (OVA) and CAP-OVA) and goat anti-rabbit IgG were respectively drawn on nitrocellulose membrane as two test lines (T 1 and T 2 ) and a control line (C). The immunostrip was dipped into a well that contained a 200 μL milk sample, 5 μL AFM1 antibody-gold conjugates, and 8 μL CAP antibody-gold conjugates; the whole assay was completed in 15 min and the results could be interpreted visually or using a reader. This immunostrip has cut-off levels of 0.1 ng/mL and 0.5 ng/mL for AFM1 and CAP, respectively. Analysis of CAP and AFM1 in milk samples revealed that data from the immunostrip test agreed closely with those obtained from ELISA. The two-analyte immunostrip is a rapid way for on-site simultaneous detection of AFM1 and CAP in milk.

Aflatoxin M1 (AFM1) is a mycotoxin from Aspergillus flavus and A. parasiticus , classified as carcinogenic and hepatotoxic. The objective of the present investigation was to determine its presence in raw milk from north-central Ecuador, constituted by the provinces of Pichincha, Manabí, and Santo Domingo de los Tsáchilas. These areas represent approximately 30% of Ecuadorian milk production. By the end of the investigation, a total of 209 raw milk samples were collected, obtained both during the dry (June and August) and rainy seasons (April and November) of 2019. AFM1 concentrations were measured with lateral flow immunochromatographic assays, and 100% of the samples were positive for this mycotoxin, presenting a mean value of 0.0774 μg/kg with a range of 0.023 to 0.751 μg/kg. These AFM1 levels exceeded the European Union regulatory limit of 0.05 μg/kg in 59.3% (124/209) of samples, while only 1.9% (4/209) exceeded the Ecuadorian legal limit of 0.5 μg/kg. By using non-parametric tests, significant differences were determined ( p ≤ 0.05) between the provinces for months of study, climatic season (being higher in the dry season), and climatic region (greater in the coast region). On the other hand, there were no significant differences ( p ≥ 0.05) between the types of producers or between production systems. Therefore, AFM1 contamination in raw milk does not present a serious public health problem in Ecuador, but a monitoring and surveillance program for this mycotoxin in milk should be developed to prevent consumer health problems.

Since the high incidence of aflatoxin M1 (AFM1) in milk and dairy products poses a serious risk to human health, this work aimed to investigate the complex formation between bovine α-lactalbumin (α-La) and AFM1 using different spectroscopic methods coupled with molecular docking studies. Fluorescence spectroscopy measurements demonstrated the AFM1 addition considerably reduced the α-La fluorescence intensity through a static quenching mechanism. The results indicated on the endothermic character of the reaction, and the hydrophobic interaction played a major role in the binding between AFM1 and α-La. The binding site stoichiometric value (n = 1.32) and a binding constant of 2.12 × 10 3  M -1 were calculated according to the Stern-Volmer equation. The thermodynamic parameters ΔH, ΔS and ΔG b were determined at 93.58 kJ mol -1 , 0.378 kJ mol -1  K -1 and -19.17 ± 0.96 kJ mol -1 , respectively. In addition, far-UV circular dichroism studies revealed alterations in the α-La secondary structures when the α-La-AFM1 complex was formed. An increased content of the α-helix structures (from 35 to 40%) and the β-sheets (from 16 to 19%) were observed. Furthermore, protein-ligand docking modelling demonstrated AFM1 could bind to the hydrophobic regions of α-La protein. Overall, the gathered results confirmed the α-La-AFM1 complex formation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Major databases were searched until January 2019 and 77 eligible studies were included in the meta-analysis to estimate the overall mean of AFM1 in milk in Iran. The mean of AFM1 levels was obtained 55.97 ng/kg (95% CI: 50.09-61.84). However, the pooled estimated mean of AFM1 levels in milk were 94.58 (95% CI: 70.24-118.92), 59.19 (95% CI: 51.84-66.54) and 35.23 ng/kg (95% CI: 31.53-38.92), considering 4, 55 and 18 TLC, ELISA and HPLC-based studies (including 354, 9224 and 2606 samples), respectively. Also, there is a wide variation of AFM1 levels among different geographical regions which were the highest in northern (88.77 ng/kg). The AFM1 contamination of milk taken from the areas with humid climate was higher than the arid climate. AFM1 Levels were the highest in winter (48.70 ng/Kg). The level of AFM1 in pasteurized, raw, and UHT milk were 49.76, 55.08 and 94.81 ng/kg, respectively., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

In the present study, a sensitive label-free electrochemical aptasensor is introduced to measure aflatoxin M1 (AFM1) by using platinum nanoparticles (PtNPs) decorated on a glassy carbon electrode (GCE) modified with Fe-based metal-organic frameworks, MIL-101(Fe). The MIL-101(Fe) and the PtNP/MIL-101(Fe) are synthesized and characterized by Fourier transform infrared spectroscopy, X-ray diffraction, UV-Visible spectroscopy, and field-emission scanning electron microscopy. Cyclic voltammetry and electrochemical impedance spectroscopy (EIS) are done to monitor the fabrication processes of the aptasensor. In optimum conditions, the linear calibration range of 1.0 × 10 -2 to 80.0 ng mL -1 and the detection limit of 2.0 × 10 -3  ng mL -1 are obtained to measure AFM1 concentration using the EIS method. Finally, the fabricated aptasensor is successfully applied to measure AFM1 concentration in powder and pasteurized milk samples., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Aflatoxin B 1 (AFB 1 ) is a secondary metabolite produced by some Aspergillus spp . fungi affecting many crops and feed materials. Aflatoxin M 1 (AFM 1 ), the 4-hydroxylated metabolite of AFB 1, is the main AFB 1 -related compound present in milk, and it is categorized by the International Agency for Research on Cancer (IARC) as a "group 1 human carcinogen". The aim of this work was to evaluate and compare the analytical performances of two commercial immunoassays widely applied for the detection of AFM 1 in milk, namely strip test immunoassay and enzyme linked immunosorbent assay (ELISA). Assay validation included samples at AFM 1 levels of 25, 50, 75 ng/kg and blank samples (AFM 1 < 0.5 ng/kg). With respect to a screening target concentration (STC) of 50 ng/kg the two assays showed cut-off values of 37.7 ng/kg and 47.5 ng/kg for strip test and ELISA, respectively, a false suspect rate for blanks <0.1% (for both assays) and a false negative rate for samples containing AFM 1 at levels higher than STC, of 0.4% (for both assays). The intermediate precision (RSD ip ) was <32% for the strip test and <15% for the ELISA. Method verification through long-term intra-laboratory quality control (QC) measurements confirmed the results from the validation study. Furthermore, a satisfactory correlation of the results obtained with both immunoassays and the AOAC Official Method 2000.08 was obtained for the analysis of cow milk samples naturally contaminated with AFM 1 at levels within "not detected" (< 0.5 ng/kg) and 50 ng/kg. Finally, the extension of the scope of the strip test method to goat and sheep milk was evaluated by applying the experimental design foreseen in the EU regulation.

Aflatoxin M1 (AFM1) contamination in 150 samples of milk, sold in market of Hisar city of Haryana, India, was investigated by using High Pressure Liquid Chromatography (HPLC). Out of these, 40 samples contained AFM1 at a concentration below the limit of detection (LOD) of 0.052 μg/kg. Among the AFM1 contaminated samples, 46 raw milk samples contained a concentration above the LOD but less than the limit of quantitation (LOQ), whereas 64 samples were above the LOQ. Of these samples, 31 exceeded the maximum limit of 0.5 μg/kg prescribed by FSSAI, India. Based on this study, the dietary intake of AFM1 for adults through consumption of milk was estimated. The results indicated that AFM1 contamination can be a food safety issue for raw and pasteurised milk consumed in India. Therefore, there is a need for a national monitoring programme to control the level of mycotoxins in milk.

To determine the relationship between seasons and regions and aflatoxin M 1 (AFM 1 ) contamination of milk distributed in Fukuyama City, we conducted a survey once during the summer and once during the winter between June 2018 and January 2019. We compared the AFM 1 contamination levels in milk drinks available in Fukuyama City during the same period, to provide more about the factors causing AFM 1 contamination. All milk samples examined exhibited AFM 1 contamination levels below the standard AFM 1 contamination level (0.5 μg/kg). For the comparison based on seasons, one milk sample collected in the summer (0.07 μg/kg) exceeded the EU limit (heat-treated milk: 0.050 μg/kg). However, there was no significant difference in the AFM 1 contamination level (p>0.05). For the comparison based on regions, the AFM 1 contamination level in the milk sample from the Chugoku Region was significantly higher in the winter and significantly lower in the summer compared to those from other regions. AFM 1 contamination of milk did not have a direct relationship with seasons or regions, but was instead influenced by the type, amount, and management of feed supplied to dairy cattle. For the comparison between milk and milk drinks, the AFM 1 contamination levels in milk drinks were significantly lower (p<0.01). The highest AFM 1 concentration (0.08 μg/kg) was detected in one sample of milk drink sampled during the summer. The AFM 1 contamination of milk drinks is likely affected by the level of contamination in raw materials, the proportion of such raw materials in the drinks, and the process type. An increase in non-fat milk solids was assumed to be a factor that increases AFM 1 contamination.

The present study was carried out with the objectives to determine the occurrence of aflatoxin M1 (AFM1) in raw milk and identification of associated risk factors from three different types of animal farming systems. A total of 189 pooled raw milk samples were tested for AFM1 with enzyme linked immunosorbant assay (ELISA) as screening and high performance liquid chromatography with fluorescence detection (HPLC-FLD) as confirmatory techniques. AFM1 was found positive in 58% pooled milk samples with the mean level of 0.917 µg/L, while 50.8% and 36.5% samples were found above the legal limits set by European Commission (EC) and Food safety and standard authority of India (FSSAI), respectively for AFM1 in milk. Among all investigated risk factors, feeding of readymade concentrate feed and leftover household cereals, longer feed storage duration and feed storage quality were found significantly associated with presence of AFM1 in farm milk. High prevalence as well as levels of AFM1 in farm milk poses a health risk to the consumers especially children. Therefore, efforts are urgently required to create awareness among dairy farmers about aflatoxins contamination of feed and milk, to improve farm management practices and to control AFM1 level in milk below the legal limits.

A study was conducted to investigate the frequency and levels of AFM 1 and AFM 2 in urine from children who attended the emergency service of a pediatric referral hospital in Bogota, Colombia. A survey on the consumption of foods likely to be a source of aflatoxins and on sociodemographic variables was conducted as well. The frequency of AFM 1 in urine was found to be 41.7% with an average concentration in positive samples of 16 pg mL -1  ± 10.7 pg mL -1 (range > LOD-48.5 pg mL -1 ). The presence of AFM 1 in the urine was related to the consumption of cereals likely to be contaminated with AFB 1 , especially corn and rice. No detectable levels of AFM 2 were found in any sample. The results show that children's exposure to aflatoxins in Colombia is indeed a problem and should be one of the priorities of the health authorities. Continuous monitoring of aflatoxins in foods should be carried out, in compliance with Colombian regulations, using analytical methods that allow determination and quantification of aflatoxins in different biological and non-biological matrices at trace levels.

This study examined the presence of Aflatoxin M 1 (AFM 1 ) in raw milk, cheese, and cheese halva samples collected from the Çanakkale province of Turkey. Raw milk, Ezine cheese, and cheese halva samples (n 120 for each group, total 360) were collected from bazaars, local manufacturers, and supermarkets in the Çanakkale province of Turkey during 2014 and 2015. ELISA was used to detect AFM 1 in these products. AFM 1 mean values were 5.14-78.69, 19.43-158.30, and 50.25-213.50 ng/kg in raw milk, Ezine cheese, and cheese halva samples, respectively. The levels of AFM1 in four raw milk samples (3.3%) were above the legal limits, whereas AFM 1 levels in Ezine cheese and cheese halva samples were within the legal limits according to the requirements of the Turkish Food Codex and the European Commission. Turkey has a wealth of cultures and local delicacies. Over previous decades, gastronomic tourism in Turkey has become popular owing to an increase in visitor demand, and traditional foods are being exported abroad. Our research investigated the presence of AFM 1 in these types of food products. Çanakkale is one of the best representative cities in the Marmara Region in Turkey as a source of these unique products.

Aflatoxins (AF) are naturally occurring mycotoxins, produced by many species of Aspergillus. Among aflatoxins, Aflatoxin M1 (AFM1) is one of the most frequent and dangerous for human health. The acceptable maximum level of AFM1 in milk according to EU regulation is 50 ppt, equivalent to 152 pM, and 25 ppt, equivalent to 76 pM, for adults and infants, respectively. Here, we study a photonic biosensor based on Si 3 N 4 asymmetric Mach-Zehnder Interferometers (aMZI) functionalized with Fab' for AFM1 detection in milk samples (eluates). The minimum concentration of AFM1 detected by our aMZI sensors is 48 pM (16.8 pg/mL) in purified and concentrated milk samples. Moreover, the real-time detection of the ligand-analyte binding enables the study of the kinetics of the reaction. We measured the kinetic rate constants of the Fab'-AFM1 interaction.

Aflatoxins (AF) are carcinogenic metabolites produced by different species of Aspergillus which readily colonize crops. AFM1 is secreted in the milk of lactating mammals through the ingestion of feedstuffs contaminated by aflatoxin B1 (AFB1). Therefore, its presence in milk, even in small amounts, presents a real concern for dairy industries and consumers of dairy products. Different strategies can lead to the reduction of AFM1 contamination levels in milk. They include adopting good agricultural practices, decreasing the AFB1 contamination of animal feeds, or using diverse types of adsorbent materials. One of the most effective types of adsorbents used for AFM1 decontamination are those of microbial origin. This review discusses current issues about AFM1 decontamination methods. These methods are based on the use of different bio-adsorbent agents such as bacteria and yeasts to complex AFM1 in milk. Moreover, this review answers some of the raised concerns about the binding stability of the formed AFM1-microbial complex. Thus, the efficiency of the decontamination methods was addressed, and plausible experimental variants were discussed.

The objective of this research was to perform an exposure assessment of aflatoxin M1 (AFM1) intake through the consumption of milk and yoghurt by the student population in Serbia and Greece. A food consumption survey of milk and yoghurt was performed during the first half of 2018 in the two countries with at least 500 interviewees (aged between 18 and 27 years) per country, covering their dietary habits and body weight based on one-day and seven-day recall methods. Values for the concentration of AFM1 were extracted from published research. Finally, a Monte Carlo analysis of 100,000 iterations was performed to estimate the intake of AFM1 from the consumption of the two dairy products. Results revealed that the estimated average exposure of students to AFM1 was in the range of 1.238⁻2.674 ng kg -1 bw day -1 for Serbia, and 0.350⁻0.499 ng kg -1 bw day -1 for Greece, depending on the dietary recall method employed. High estimations for hepatocellular carcinoma (HCC) cases/year/10⁵ individuals, depending on the prevalence of Hepatitis B virus surface antigen positive individuals (HBsAg+), were 0.0036⁻0.0047 and 0.0007⁻0.0009 for Serbia and Greece, respectively. Presented Margin of Exposure (MOE) and Hazard Index (HI) values indicate increased risk from exposure to AFM1, particularly in Serbia.

Background: Aflatoxin M1 (AFM1) is a mycotoxin found in milk as a result of the ingestion of aflatoxin B1 (AFB1) by dairy cattle. Because of its carcinogenic properties, the control of its occurrence in dairy products is of great importance. We evaluated the occurrence and seasonality of AFM1 in milk from farms with subtropical and temperate climates, where significant milk production occurs. Forty samples of raw milk were collected from bulk tansk milk during the summer (five samples from a subtropical climate and 14 from a temperate climate) and winter (six samples from a subtropical climate and 15 from a temperate climate) months of 2017 and conducted an analysis using an enzyme-linked immunosorbent assay (ELISA) (detection limit 5 ng L -1 ). Data from eight random samples were compared with respect to liquid chromatography with mass spectrometry detection and ELISA. A significant correlation was observed., Results: The presence of AFM1 was detected in 87.50% (n = 35) of the samples analyzed (a mean of 16.66 ng L -1 and a median of 12.42 ng L -1 in positive samples); however, its concentration in all samples was below the maximum limit allowed by European (50 ng L -1 ) and Brazilian (500 ng L -1 ) legislations. There were no significant differences in the levels of AFM1 between the properties located in the two climate zones, in both summer and winter. The estimated daily intake of AFM1 based on the milk analyzed was 0.0107 ng kg -1 day -1 for adolescents, 0.0072 ng kg -1 day -1 for adults and 0.0098 ng kg -1 day -1 for the elderly., Conclusion: The present work demonstrated a low exposure to AFM1. © 2018 Society of Chemical Industry., (© 2018 Society of Chemical Industry.)

Herein, we report the design of a novel label-free aptasensor based on ferrocene and silicon nanoparticles (SiNPs) for ultrasensitive detection of aflatoxin M1 (AFM1) in milk. Given that silicon nanomaterials stand out by their high capacitive power, we used them to develop a novel capacitive transduction system based on electrochemical capacitance spectroscopy (ECS). This strategy relies on the changes of the redox capacitance signal owed to the surface-tethered ferrocene film, by performing electrochemical impedance spectroscopy (EIS) measurements without using an external redox probe. The redox capacitance variation was found to correlate well with the increasing concentrations of AFM1 in the linear range from 10 to 500 fmol⋅L -1 with a sensitivity of 0.46 μF*fM-1*cm - 2. Furthermore, the aptasensor allowed to reach very low limits of detection and quantification equal to 4.53 fM and 14.95 fM, respectively. The platform revealed a high selectivity toward the target analyte, and it was applied to quantify very low concentrations of AFM1 in commercial pasteurized milk. Finally, the results of real sample analysis were successfully gauged against those obtained using commercially available enzyme-linked immunoassay (ELISA) kits., (Copyright © 2018 Elsevier B.V. All rights reserved.)

This study was conducted to determine the effect of Bacillus subtilis ANSB060 biodegradation product (BDP) in reducing the milk aflatoxin M₁ (AFM₁) content of dairy cows fed a diet contaminated with aflatoxin B₁ (AFB₁). Twenty-four Chinese Holstein cows (254 ± 19 d in milk; milk production 19.0 ± 1.2 kg d -1 ) were assigned to three dietary treatments, as follows: (1) control diet (CON), consisting of a basal total mixed ration (TMR); (2) aflatoxin diet (AF), containing CON plus 63 μg of AFB₁ kg -1 of diet dry matter; and (3) aflatoxin diet plus BDP (AF + BDP), containing AF plus BDP at 0.2% of diet dry matter. The experiment lasted 12 days, including an AFB₁-dosing period from days one to eight, followed by a clearance period from days nine to twelve. Milk samples were collected on days 2, 4, 6, and 8⁻12, and the plasma was sampled on day 9, before morning feeding. Short-term AFB₁ exposure did not affect the milk production and composition. The plasma biochemical indices, except for lactic dehydrogenase (LDH), were also not changed by the AFB₁ intake. The plasma LDH level was significantly elevated ( p < 0.05) following dietary treatment with AFB₁, while no significant difference was observed between the AF + BDP and CON treatments. Adding BDP to the AFB₁-contaminaed diet resulted in a significant reduction in AFM₁ concentration (483 vs. 665 ng L -1 ) in the milk, AFM₁ excretion (9.14 vs. 12.71 μg d -1 ), and transfer rate of dietary AFB₁ to milk AFM₁ (0.76 vs. 1.06%). In conclusion, the addition of BDP could be an alternative method for reducing the dietary AFB₁ bioavailability in dairy cows.

In this survey aflatoxin, M 1 was quantified in raw and processed milk from various areas of two big cities of Punjab province, i.e. Lahore and Multan. The results indicated that approximately 90% of the raw milk samples collected from Lahore city was contaminated with aflatoxin M 1 . Similarly, around 92% of the raw milk samples collected from Multan city was contaminated with aflatoxin M 1 . All samples of processed milk and tea whiteners were contaminated and 56% of the contaminated processed milk samples and 66% of the contaminated tea whitener samples were violating the maximum limits. The dietary exposure data of AFM 1 among six different groups was calculated, which indicated that the male children population was the most vulnerable group to AFM 1 , up to 6.68 ng L -1 per day and the least affected one was the female group above 20 years of age with 1.13 ng L -1 per day.

A longitudinal one-year study was conducted to determine aflatoxin M₁ levels in different types of milk marketed in Pakistan. Processed and raw liquid milk from 21 sources, two milk powder and six tea whitener brands were sampled on monthly basis from Islamabad. The aflatoxin M₁ levels in liquid milk were lower ( p < 0.05) in summer (April to July) compared with the levels in winter (January, November and December). The mean aflatoxin M₁ levels were 254.9, 939.5, and 1535.0 ng/L in UHT, pasteurized, and raw milk, respectively (differing at p < 0.001). The mean toxin level in powdered milk after reconstitution was 522.1 ng/L. Overall, 12.9, 41.0, 91.9 and 50.0% of the UHT, pasteurized, raw and powdered milk samples, respectively, exceeded the Codex maximum tolerable limit of 500 ng of aflatoxin M₁/L. It was estimated that consumers of raw and processed milk were exposed to 11.9 and 4.5 ng aflatoxin M₁, respectively, per kg of body weight daily. The study indicates potential aflatoxin M₁ exposure risks for the consumers of raw milk in the country. The levels of the toxin though comparatively lower in milk powder, requires attention as this type of milk is consumed by infants.

A mini-survey of 29 different foods produced by 21 different Indian manufacturers was conducted for the presence of aflatoxins B1, B2, G1 and G2, aflatoxin M1 and deoxynivalenol. The products were purchased from local markets in Kolkata, India and commonly used in the complementary feeding of infants and toddlers in India. Using a previously established direct competitive enzyme-linked immunoassay for this analysis we show that 100% of the samples contained aflatoxin M1 at levels exceeding the recommended European Union levels of 25 ng kg -1 by more than an order of magnitude. Also, several (66%) of them contained detectable concentrations of deoxynivalenol with two samples (6.9%) exceeding European Union guidelines for baby food products (200 μg kg -1 ) and 51.7% samples with DON levels that can lead to dietary intake higher than 1  μg kg -1 recommended by the joint FAO/WHO expert committee on food additives. None of the samples contained aflatoxins B1, B2, G1 and G2. The results, therefore, suggest that complementary feeding can put Indian infants and toddlers at risk of simultaneous exposures to deoxynivalenol and aflatoxin M1 and warrant an urgent in-depth research to track, increase surveillance and reduce mycotoxin contamination of baby foods manufactured in India., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

During 2016-2017, 156 samples of fresh milk samples were collected from local markets of Karachi, Pakistan and analysed for aflatoxin M 1 (AFM 1 ) contamination using ELISA technique. AFM 1 was detected in 143 (91.7%) samples, ranged from 20 to 3090 ng L -1 with a mean level of 346.2 ng L -1 . In 125 (80.1%) samples, the AFM 1 contamination was greater than the maximum limit (ML = 50 ng L -1 ) set by EU. However, in 51 (32.7%) samples, the AFM 1 level was higher than the ML of 500 ng L -1 as assigned by the USA. Statistical analysis showed that the AFM 1 level in milk samples from summer was significantly (p < 0.05) higher than that obtained in winter. It was concluded that the AFM 1 levels in the tested samples appear to be a serious public health problem. Therefore, immediate measures should be taken and re-evaluation done for the procedures for farming, transportation, refrigeration, and storage for the control of AFM 1 level in milk samples.

The occurrence of aflatoxin M1 in 69 milk samples collected in a south region of Italy in 2016 was evaluated. The samples were analysed using an automated method based on online SPE coupled with UHPLC tandem mass spectrometry. After a salt induced liquid-liquid extraction with acetonitrile to remove protein from milk, the extract was diluted with water and analysed using an automated online SPE MS/MS method. Among the analysed samples no one had AFM1 higher than the legally allowable limits whereas 71.4% of the other analysed samples were above the LOD of the method. The highest contamination level of AFM1 was found in pasteurised milk (44.39 ng kg -1 ). The results show the worrying and widespread of AFM1 contamination, highlighting the necessity of monitoring studies in order to evaluate the reduction of the maximum legal limit.

This study aimed to investigate the ability of chitin and heat-treated shrimp shells to bind aflatoxin M1 (AFM1) in liquid matrix. Several concentrations of chitin or shrimp shells (grinded and ungrinded) were incubated in AFM1-contaminated phosphate-buffered saline (PBS) at different incubation times. The stability of the formed adsorbent-AFM1 complex was also tested in milk at different incubation times and temperatures. The unbound AFM1 was quantified by HPLC. Thereby, the percentages of the initial bounded AFM1 varied between 14.29 and 94.74%. Interestingly, in milk, an increase in incubation time coupled with a decrease in temperature affected positively the amount of bounded AFM1 to chitin and negatively those bounded to ungrinded shells. Results also revealed a partial reversibility in the binding of AFM1 to these adsorbents. These findings provided strong evidence on ability of chitin or shrimp shells by-product to bind AFM1 in milk and in PBS.

During February and August 2016 (winter and summer season), 192 samples of raw bulk tank milk have been collected from small dairy farms in five different regions of Kosova (Prishtina, Prizren, Peja, Mitrovica, and Gjilan). These samples were analyzed for aflatoxin M 1 (AFM 1 ) contamination level by ultra-high performance liquid chromatography coupled with electrospray ionization triple quadrupole tandem mass spectrometry (UHPLC-ESI-MS/MS). The limit of quantification for AFM1 was 0.02 μg/L and recovery at 0.1-0.5 μg/L was 90-95% (coefficient of variation 15-25%). A total of 74 milk samples (38%) were found to be contaminated with AFM 1 and 11 samples (5.7%) slightly exceeded the European Union maximum level of 0.05 μg/L. The maximum AFM 1 level was at 0.082 μg/L. There was no relevant difference between summer and winter with regard to AFM 1 frequency and levels. Although the frequency of AFM 1 -positive samples varied between regions, from 7% (Prishtina) to 53% (Prizren), only minor regional differences were observed with regard to average and maximum toxin levels. The high percentage of milk samples which were non-compliant with AFM 1 maximum levels indicates that efforts to reduce the contamination levels of aflatoxin B 1 in cows feed in Kosovo are necessary.

Aflatoxins are a major class of fungal toxins that have food safety importance due to their economic and health impacts. This pilot aflatoxin exposure biomonitoring study on 84 individuals was conducted in a rural (Ilumafon) and a semi-urban community (Ilishan Remo) of Ogun state, Nigeria, to compare aflatoxin exposures among the two population cohorts. First morning urine samples were obtained from the participants, and the urinary aflatoxin M 1 (AFM 1 ) levels were measured by a quantitative Helica Biosystems Inc. ELISA kit assay. About 99% (83 out of 84) of the urine samples had detectable AFM 1 levels in the range of 0.06 to 0.51 ng mL -1 (median: 0.27 ng mL -1 ). The mean urinary AFM 1 levels were significantly (p = 0.001) higher in the semi-urban population (0.31 ± 0.09 ng mL -1 ) compared to the rural population (0.24 ± 0.07 ng mL -1 ). There were, however, no significant differences in mean urinary AFM 1 levels of males and females, and among children, adolescents and adults. This study indicates high aflatoxin exposure to the extent of public health concerns in the studied populations. Thus, more efforts are required for aflatoxin exposure monitoring and control in high-risk regions.

This survey was performed to investigate the occurrence of aflatoxin M1 (AFM1) contamination of raw milk from manufacturers of infant milk powder in China. A total of 1207 raw milk samples were collected overall from four seasons of 2016 in Northeast China, Northwest China, Northern China, and Central China (11 provinces and one municipality). Results showed that 56 of the 1207 raw milk samples (4.64%) were positive for AFM1, which were obtained from Heilongjiang (two samples), Gansu (one sample), Shaanxi (46 samples), Beijing (one sample), and Hunan (six samples) provinces. None of the raw milk samples from manufacturers of infant milk powder exceeded the Chinese limit (62.5 ng/L) in 2016. Only a few raw milk samples were not suitable for use in infant milk according to EU (European Union) or U.S. infant milk limits. Furthermore, based on this survey and previous studies, it is particularly important to avoid AFM1 contamination in raw milk during the winter.

Aflatoxin is a food contaminant and its exposure through the diet is frequent and ubiquitous. A long-term dietary aflatoxin exposure has been linked to the development of liver cancer in populations with high prevalence of aflatoxin contamination in foods. Therefore, this study was conducted to identify the association between urinary aflatoxin M₁ (AFM₁), a biomarker of aflatoxin exposure, with the dietary intake among adults in Hulu Langat district, Selangor, Malaysia. Certain food products have higher potential for aflatoxin contamination and these were listed in a Food Frequency Questionnaire, which was given to all study participants. This allowed us to record consumption rates for each food product listed. Concomitantly, urine samples were collected, from adults in selected areas in Hulu Langat district, for the measurement of AFM₁ levels using an ELISA kit. Of the 444 urine samples collected and tested, 199 were positive for AFM₁, with 37 of them exceeding the limit of detection (LOD) of 0.64 ng/mL. Cereal products showed the highest consumption level among all food groups, with an average intake of 512.54 g per day. Chi-square analysis showed that consumption of eggs ( X ² = 4.77, p = 0.03) and dairy products ( X ² = 19.36, p < 0.01) had significant associations with urinary AFM₁ but both food groups were having a phi and Cramer's V value that less than 0.3, which indicated that the association between these food groups' consumption and AFM₁ level in urine was weak., Competing Interests: The authors declare no conflict of interest.

A highly sensitive aptasensor for aflatoxin M₁ (AFM₁) detection was constructed based on fluorescence resonance energy transfer (FRET) between 5-carboxyfluorescein (FAM) and palladium nanoparticles (PdNPs). PdNPs (33 nm) were synthesized through a seed-mediated growth method and exhibited broad and strong absorption in the whole ultraviolet-visible (UV-Vis) range. The strong coordination interaction between nitrogen functional groups of the AFM₁ aptamer and PdNPs brought FAM and PdNPs in close proximity, which resulted in the fluorescence quenching of FAM to a maximum extent of 95%. The non-specific fluorescence quenching caused by PdNPs towards fluorescein was negligible. After the introduction of AFM₁ into the FAM-AFM₁ aptamer-PdNPs FRET system, the AFM₁ aptamer preferentially combined with AFM₁ accompanied by conformational change, which greatly weakened the coordination interaction between the AFM₁ aptamer and PdNPs. Thus, fluorescence recovery of FAM was observed and a linear relationship between the fluorescence recovery and the concentration of AFM₁ was obtained in the range of 5-150 pg/mL in aqueous buffer with the detection limit of 1.5 pg/mL. AFM₁ detection was also realized in milk samples with a linear detection range from 6 pg/mL to 150 pg/mL. The highly sensitive FRET aptasensor with simple configuration shows promising prospect in detecting a variety of food contaminants., Competing Interests: The authors declare no conflict of interest.

Aflatoxin B 1 (AFB 1 ) is the most potent of the dietary aflatoxins, and its major metabolite, aflatoxin M 1 (AFM 1 ), is frequently found in the breast milk of lactating mothers. The aim of this study was to assess the occurrence and factors associated with AFM 1 contamination of breast milk collected from lactating mothers in Lebanon. A total of 111 breast milk samples were collected according to the guidelines set by the World Health Organization. Samples were analyzed with a competitive enzyme-linked immunosorbent assay between December 2015 and November 2016. A survey was used to determine the demographic and anthropometric characteristics of participating lactating mothers. Dietary habits were assessed using a semiquantitative food frequency questionnaire. Mean (±standard deviation) concentration of AFM 1 in the breast milk samples was 4.31 ± 1.8 ng/L, and 93.8% of samples contained AFM 1 at 0.2 to 7.9 ng/L. The mean concentration of AFM 1 was significantly lower (P < 0.05) in fall and winter (4.1 ± 1.9 ng/L) than in spring and summer (5.0 ± 1.7 ng/L). None of the samples exceeded the European Commission regulation limit (25 ng/L) for infant milk replacement formula. AFM 1 contamination was significantly associated (P < 0.05) with the daily consumption of white cheeses but not with the consumption of meat or cereal products. No significant association (P > 0.05) was observed between AFM 1 concentrations in breast milk and anthropometric sociodemographic factors (age and level of education) or the governorate of residence of the nursing mothers. The mean AFM 1 estimated daily intake was found to be 0.69 ng/day/kg of body weight. Although the incidence of AFM 1 contamination was low, our first-of-its-kind study highlights the importance of conducting investigations on mycotoxin contamination in breast milk and of developing protection strategies to tackle the exposure of infants to this potent chemical hazard.

An inductively coupled plasma mass spectrometry (ICP-MS)-based immunoassay has been developed to quantify aflatoxin M1 (AFM1) at ultra-trace levels in milk samples. AFM1 detection is carried out by means of a competitive immunoassay using secondary biotinylated antibodies and streptavidin-conjugated Au nanoparticles. After acid addition, nanoparticles are decomposed and Au signal is registered by means of ICP-MS. Results demonstrate that, under optimum conditions, the limit of detection of the immunoassay (0.005μgkg -1 ) is low enough to quantify AFM1 according to current international policies (including the more restrictive European one). Method accuracy and precision was checked by analyzing an AFM1 certified reference material and different milk samples spiked with known amounts of AFM1. AFM1 recovery values range from 80% to 102% whereas inter-assay and intra-assay precision are lower than 15%. Finally, this immunoassay methodology affords a higher dynamic working range (0.012-2.5μgkg -1 ) than other immunoassay methodologies described in the literature., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The aim of this systematic review is to provide information regarding the incidence and levels of aflatoxin M 1 (AFM 1 ) in raw and heat processed cow's milk in Serbia during 2015-16 and to compare these with collected data on the occurrence of AFM 1 in raw milk and dairy products during the last decade in our region. Estimation of dietary exposure (EDI) and hazard index (HI) calculations for different age groups of the population were also carried out, based on the AFM 1 content of milk samples and on available food consumption data in Serbia. AFM 1 was detected in 69.9% (984/1408) of raw milk samples in 2015 versus 84.9% (3094/3646) in 2016, while in heat-processed milk, AFM 1 was detected in 77.8% (364/468) in 2015 versus 98.5% (753/765) in 2016. On the basis of the obtained results, 450 (9%) of raw and 14 (1.1%) of heat-processed milk samples were contaminated with AFM 1 levels above the maximum permitted level in Serbia (0.25 μg kg -1 ). However, a large percentage of raw and heat processed milk in Serbia (30.1% and 17.3%, respectively) was contaminated with AFM 1 levels above the maximum permitted level regulated in the European Union (0.05 μg kg -1 ). Therefore, in order to protect consumer health, it is extremely important to further control the level of aflatoxins in milk, and this should be considered as a high priority for risk management actions.

During the last 3 years, high aflatoxin M1 (AFM1) concentrations in milk and dairy products occurred in Serbia. It resulted in periodical change of the official regulations regarding maximum levels (MLs) of AFM1 as set by the Serbian Government. The aim of this study was to compare the occurrence of AFM1 in raw milk and dairy products during 2015 and also to determine whether there were some differences in AFM1 level among seasons. The AFM1 level exceeded the European Union ML in 29.3% of raw milk and 4.2% of milk product samples. The highest level of AFM1 in raw milk was found during the autumn season, while during the rest of the 2015, it was significantly lower. Although the improvement of dairy products safety was evident in 2015 when compared to 2013 and 2014, the cause of high concentrations in raw milk remained unresolved yet. This study indicates that dairy plants introduced control measures and refused reception of too high contaminated raw milk.

The study reports the results of testing the sensitivity of an early warning sampling plan for detecting milk batches with high aflatoxin AFM 1 concentration. The effectiveness of the method was investigated by the analysis of 9017 milk samples collected in Italian milk processing plants that applied control plans with different action limits (AL). For those milk processing plants where 30 ng kg -1 AL has been applied, the AFM 1 contamination was significantly lower at or above the 95th percentile of the milk samples when compared with plants that used 40 ng kg -1 AL. The results show that the control plan can be used effectively for early warning of occurrence of high AFM 1 contamination of milk and to carry out pro-active measures to limit the level of contamination. Estimation of dietary exposure was also carried out, based on the aflatoxin M 1 content of the milk samples and on Italian food consumption data. Estimated Daily Intakes (EDI) and Hazard Indices (HI) were calculated for different age groups of the population. HIs show that no adverse effects are expected for the adult population, but in the case of children under age three, the approximate HI values were considerably higher. This underlines the importance of the careful monitoring and control of aflatoxin M 1 in milk and dairy products.

This study aims to detect aflatoxins (AFs) in dairy cow feed, milk and milk products using a high-performance liquid chromatography coupled with fluorescence detection (HPLC-FLD) method. All the validation parameters met the method performance criteria of the European Union. The samples comprised 76 dairy cow feeds and 205 milk and milk products (including yoghurt and yoghurt-based beverage, ayran). AFs were present in 26.3% of the feed samples. Two feed samples exceeded the maximum limit (ML) of 5 µg kg(-1) for AFB1 as established by the EU. Nineteen milk samples (21.1%) contained aflatoxin M1 (AFM1) of which three exceeded the EU ML of 0.05 µg l(-1). In addition, only two yoghurt samples and one ayran sample contained AFM1, but the levels were lower than the EU ML.

Aflatoxin M1 (AFM1) contamination in raw milk from household cows fed with sunflower seedcakes or sunflower-based seedcake feeds was determined in 37 milk samples collected randomly from different locations in Singida region, Tanzania. Aflatoxin B1 (AFB1) contamination in sunflower-based seedcake feed was determined in 20 feed samples collected from the same household dairy farmers. The samples were analysed by RP-HPLC using fluorescent detection after immunoaffinity column clean-up. Recoveries were 88.0% and 94.5%, while the limits of detection (LOD) were 0.026 ng mL(-1) and 0.364 ng g(-1) for AFM1 and AFB1, respectively. Of the analysed cow's milk samples, 83.8% (31/37) contained AFM1, with levels ranging from LOD to 2.007 ng mL(-1), exceeding both the European Commission (EC) and Tanzania Food and Drug Authority (TFDA) limit of 0.05 ng mL(-1). Of the contaminated samples, 16.1% exceeded the Codex Alimentarius limit of 0.5 ng mL(-1). AFB1 was present in 65% (13/20) of the feed samples with levels ranging from LOD to 20.47 ng g(-1), 61.53% exceeding the TFDA and EC maximum limits of 5 ng g(-1) for complete dairy animal feed. The observed AFM1 and AFB1 contamination necessitates the need to raise awareness to dairy farmers in Tanzania to safeguard the health of the end-users.

The frequency and levels of aflatoxin M1 (AFM1) in pasteurized milk samples in Sari, located in Mazandaran province, Iran, were determined by enzyme immunoassay. Seventy-six samples of pasteurized milk from different retail stores were randomly collected over four seasons during the year 2015. AFM1 contamination was detected in all milk samples. The mean concentration of aflatoxin M1 was 65.8 ng/l, with a range of 11.7-106.6 ng/l. The highest AFM1 level was detected in milk samples collected during spring. Forty-six (60.53 %) samples had AFM1 levels that exceeded the maximum acceptable levels (50 ng/l) recommended by the European Union (EU). Comparison of these results with previously published data for AFM1 in milk in Iran shows that the percentage of samples exceeding the EU maximum level is consistently high over the years, indicating a general problem related to AFB1 contamination in dairy feedingstuff.

Mycotoxins and industrial chemicals, such as aflatoxin M1 and melamine, now commonly exist in milk and cause potential health risks. This study presents an indirect competitive immunoassay through multiplex planar waveguide fluorescence immunosensor (MPWFI) for rapid, sensitive, and simultaneous detection and quantification of aflatoxin M1 and melamine by applying the principle of immunoreaction and total internal reflect fluorescent. Double-channel standard curves with appropriate logistic correlation (R(2)>0.99) were plotted, respectively. The working ranges (0.073-0.400 ng/mL and 26.38-270.00 ng/mL, respectively) were calculated, as well as the limit of detection (0.045 and 13.37 ng/mL, respectively), when two analytes were simultaneously detected. Both results satisfied the requirements for the maximum amount set by the WHO, which illustrated that the current method was better than some other standard methods. The recovery rates in the actual samples ranged from 85% to 103%, with relative standard deviations between 1.3% and 6.5%, which indicated high accuracy and repeatability., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Aflatoxins (AFs) are mycotoxins produced by some species of Aspergillus. In dairy cows, ingested AFB1 is metabolized into carcinogenic AFM1 which is eliminated through milk, thus posing a risk for consumer health. Here we describe the set, validation, and application of screening (ELISA) and confirmatory (HPLC) tests carried out on milk samples collected through official control of mycotoxin levels in northern Italy over a three-year period (2012-2014). The limit of detection (LOD) was set at 5 ppt and 2 ppt for ELISA and HPLC, respectively, and the limit of quantification (LOQ) was 10 ppt for confirmatory HPLC. A total of 1668 milk samples were analyzed: ELISA identified 36 (2.2%) positive milk samples that were subsequently confirmed by HPLC. The level of AFM1 in the positive samples ranged between 18 ± 2 and 208 ± 27 ppt. Of the total samples, only eight (0.5%) were found non-compliant with the EU regulatory limit (50 ppt; range 74 ± 10 to 208 ± 27 ppt). Use of ELISA and HPLC tests in series allows for high-volume analysis of samples, thus saving time and money while guaranteeing high analytical precision and accuracy.

An aptasensor was designed for the determination of aflatoxin M1 (AFM1) in milk based on DNA-aptamer recognition and electrochemical impedance spectroscopy detection. A hexaethyleneglycol-modified 21-mer oligonucleotide was immobilized on a carbon screen-printed electrode through carbodiimide immobilization, after diazonium activation of the sensing surface. Cyclic voltammetry and electrochemical impedance spectroscopy in the presence of ferri/ferrocyanide redox probe were used to characterize each step of the aptasensor development. Aptamer-AFM1 interaction induced an increase in electron-transfer resistance, allowing the determination of AFM1 in buffer in the range 2-150 ng/L (LOD=1.15 ng/L). Application to milk analysis showed that a preliminary treatment was mandatory. A simple filtration through a 0.2 µm PTFE membrane allowed determination of AFM1 in milk for concentrations ranging from 20 to 1000 ng/kg. These performances are compatible with the AFM1 levels set in European Union for milk and dairy products for adults (50 ng/kg) and infants (25 ng/kg)., (Copyright © 2015 Elsevier B.V. All rights reserved.)

An efficient, simple and fast low-density solvent based dispersive liquid-liquid microextraction (LDS-DLLME) followed by vortex-assisted dispersive solid phase extraction (VA-D-SPE) has been developed as a new approach for extraction and preconcentration of aflatoxin M1 in milk samples prior to its micelle enhanced spectrofluorimetic determination. In this LDS-DLLME coupled VA-D-SPE method, milk samples were first treated with methanol/water (80:20, v/v) after removing the fat layer. This solvent was directly used as the dispersing solvent in DLLME along with using 1-heptanol (as a low-density solvent with respect to water) as the extracting solvent. In VA-D-SPE approach, hydrophobic oleic acid modified Fe3O4 nanoparticles were used to retrieve the analyte from the DLLME step. It is considerably that the target of VA-D-SPE was 1-heptanol rather than the aflatoxin M1 directly. The main parameters affecting the efficiency of LDS-DLLME and VA-D-SPE procedures and signal enhancement of aflatoxin M1 were investigated and optimized. Under the optimum conditions, the method was linear in the range from 0.02 to 200 µg L(-1) with the correlation coefficient (R(2)) of 0.9989 and detection limit of 13 ng L(-1). The intra-day precision was 2.9 and 4.3% and the inter-day precision was 2.1 and 3.3% for concentration of 2 and 50 µg L(-1) respectively. The developed method was applied for extraction and preconcentration of AFM1 in three commercially available milk samples and the results were compared with the official AOAC method., (Copyright © 2014 Elsevier B.V. All rights reserved.)

A total of 635 raw milk samples from 45 dairy farms, from three regions of São Paulo state - Brazil, were evaluated during 15 months for aflatoxin M1 (AFM1). AFM1 was determined by high performance liquid chromatograph with fluorescence detection. AFM1 was detected (>0.003 µg kg(-1)) in 72.9%, 56.3% and 27.5% of the samples from Bauru, Araçatuba and Vale do Paraíba regions, respectively. The mean AFM1 contamination considering all the samples was 0.021 µg kg(-1). Furthermore, the concentration of AFM1 was quite different among Bauru (0.038 µg kg(-1)), Araçatuba (0.017 µg kg(-1)) and Vale do Paraíba (<0.01 µg kg(-1)) regions. Only three samples (0.5%) had higher contamination than the tolerated limit in Brazil (0.50 µg kg(-1)) and 64 samples (10.1%) had a higher contamination than the maximum limit as set by the European Union (0.050 µg kg(-1)). The estimated AFM1 daily intake was 0.358 and 0.120 ng kg(-1) body weight per day for children and adults, respectively.

Aflatoxins are toxic fungal metabolites, which can be found in feed. Aflatoxin M1 (AFM1) is excreted into milk when ruminants ingest aflatoxin B1 contaminated feedstuffs. Due to its carcinogenic potential, contamination of milk and dairy products with AFM1 may pose a risk for consumers. Hence, it is considered a public health concern. In this survey, the level of AFM1 contamination of dairy products marketed in Costa Rica was determined by enzyme-assisted extraction, immunoaffinity clean-up and high-performance liquid chromatography coupled with a fluorescent detector (HPLC-FLD) in fluid milk (n = 70), fresh cheese (n = 70) and sour cream (n = 70) collected at local convenience stores and supermarkets. AFM1 concentrations in milk and fresh cheese ranged from 19 to 629 ng/L and from 31 to 276 ng/L, with mean values of 136 ng/L and 74 ng/L, respectively, whereas none of the sour cream samples analysed tested positive for this aflatoxin. In 30 milk samples, and 10 cheese samples, AFM1 concentrations surpassed threshold concentrations as established by the European Commission. Thus, sour cream and - to a lesser extent - cheese manufacturing seems to reduce the amount of AFM1 present in milk, possibly due to fraction redistribution or microbiological degradation. The survey results reveal improper quality control procedures in the Costa Rican dairy industry. Therefore, a surveillance programme for dairy products in our country is recommended.