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Microorganisms are the only entities in the biosphere with an incomparable ability to employ diverse organic and inorganic compounds for growth and convert it to simple form that is no longer harmful to human health and environment. Food grade microorganisms such as lactic acid bacteria, bifidobacteria, propionibacteria as well as several yeast species are associated with food fermentation processes as well as have gained probiotic status owing to their noteworthy offerings in health stimulation as a natural gut microbiota in animals and humans. However, as biological agents little is known about their application for bioremediation and biotransformation aptitude. In context to this, aflatoxin M1 is a class of mycotoxins often associated with milk through consumption of fungus contaminated feed & fodders by cattle and well documented for their adverse health effects. Therefore, current review summarizes significance of aflatoxins present in milk and dairy products in human life, their source, types & health implications; food grade bacteria including probiotic strains and their mechanism of action involved in the removal of aflatoxin; and last section discusses the outcome of major studies showing aflatoxin reduction potential of food grade bacteria in milk and milk based products., 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 © 2024 Elsevier Ltd. All rights reserved.)

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.)

In the present study, encapsulation of Lactiplantibacillus plantarum (L.p) was performed using chitosan and casein through calcium phosphate intercrossing. Chitosan and casein both considered as non-toxic and biocompatible food derived components with intrinsic antioxidant properties. Layer by layer strategy was performed for deposition of modified cross-linked chitosan along with casein as the novel protective layers on the surface of probiotics. After confirmation of successful encapsulation, the viability and antioxidant activity of encapsulated L.p was evaluated. The results showed enhanced survival and antioxidant activity of encapsulated L.p compared to free bacteria in simulated digestive conditions. The survival of free and encapsulated L.p was respectively 1.38 ± 0.29 log cfu/ml and 6.99 ± 0.12 log cfu/ml in SGF and 8.54 ± 0.05 log cfu/ml and 7.25 ± 0.23 log cfu/ml in SIJ after 2 h of incubation. HPLC analysis was also used to investigate the detoxification activity of probiotics toward Aflatoxin M1 and obtained results showed encapsulated bacteria could significantly reduce aflatoxin M1 (68.44 ± 0.5 %) compared to free bacteria (43.76 ± 0.54 %). The results of this research suggest that the chitosan/casein mediated encapsulation of L.p with layer-by-layer technology is an effective method to improve the survival and antioxidant properties of probiotics with enhanced detoxification of AFM1., 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 © 2024 Elsevier B.V. All rights reserved.)

Aflatoxin M1 (AFM1) is a metabolite of the more toxic aflatoxin B1 (AFB1) a mycotoxin produced by the fungi Aspergillus flavus and A. parasiticus; which contaminates maize, peanuts, tree nuts, and oilseeds, among other crops in warm regions of the world. When mammals consume AFB1 in these foods, they secrete AFM1 in milk. A recent analysis indicated negligible cancer risk associated with AFM1 exposure, but whether AFM1 impairs children's growth is important to understand because children consume relatively more milk than adults worldwide. Our paper reviews the evidence on the link between AFM1 exposure and child growth impairment. We find that the existing studies are contradictory and necessitates further studies on this question; in particular, those that control for potentially confounding factors such as household socioeconomic status, children's overall diets, hygienic factors, and agroecological zone. Though many nations have policies for maximum AFM1 limits in dairy foods, they are not based on an explicit health risk analysis of AFM1 but on conversion rates of AFB1 levels to AFM1 in dairy products. Future studies on AFM1's potential harmful effects on child growth will help to better inform policies on maximum allowable AFM1 in milk and other dairy products., Competing Interests: Declaration of competing interest The authors declare that they have no financial interests that could have influenced the work reported in this paper., (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.)

The numerous strategies have been conducted worldwide to mitigate the presence of these hazardous toxins. In this systematic study, these researches are summarized. The search of this study was done with keywords aflatoxin M1, AFM1, reduce, decrease, mitigation, prevent, prevention, and milk in databases without a time limit. A total of 49 manuscripts were carefully reviewed, and their data were extracted. Some interventions focused on modifying animal rations, aiming to reduce AFM1 in milk. Some were applied directly to the animals. In this method, which was done more than other research interventions, some toxin binders are used as feed additives. The third type of intervention consisted of measures that were taken directly on the milk itself. Among the three types of interventions, the use of toxin binders in animal feed was more practical and effective., Competing Interests: 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., (© 2024 The Authors.)

Aflatoxin B1 (AFB1), a common mycotoxin, can occur in agricultural products. As a metabolite of AFB1, aflatoxin M1 (AFM1) mainly exist in dairy products. These two mycotoxins threaten human health, although it is unclear how they affect the function of the intestinal barrier. In this study, mice were exposed to AFB1 (0.3 mg/kg body b.w.) and AFM1(3.0 mg/kg b.w.) either individually or in combination for 28 days to explore the main differentially expressed proteins (DEPs) and the associated enriched pathways. These findings were preliminarily verified by the transcriptomic and proteomic analyses in differentiated Caco-2 cells. The results revealed that AFB1 and AFM1 exposure in mice disrupted the function of the intestinal barrier, and the combined toxicity was greater than that of each toxin alone. Further proteomic analysis in mice demonstrated that the mechanisms underlying these differences could be explained as follows: (i) lipid metabolism was enriched by AFB1-induced DEPs. (ii) protein export pathway was stimulated by AFM1-induced DEPs. (iii) cell metabolic ability was inhibited (as evidenced by changes in UDP-GT1, UDP-GT2, and Gatm6), apoptosis was induced (MAP4K3), and epithelial cell integrity was disrupted (Claudin7 and IQGAP2), resulting in more extensive intestinal damage after combined treatment. In conclusion, the hazardous impact of co-exposure to AFB1 and AFM1 from proteomic perspectives was demonstrated in the present study., 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 © 2024. Published by Elsevier Inc.)

The detection of the amount of aflatoxin M1 (AFM1) in milk is crucial for food safety. Here, we utilize a fiber optic (FO) localized surface plasmon resonance (LSPR) biosensor by constructing gold nanoparticle (AuNP) multimers, in which the nanogaps amplified the LSPR signal by the hot spot effect, and achieved a highly sensitive detection of f AFM1. Through the optimization of parameter conditions for the fabrication of the sensor and detection system, a high performance result from the FO LSPR biosensor was obtained, and the method for AFM1 detection was established, with a wide detection range of 0.05-100 ng/mL and a low limit of detection (LOD) of 0.04 ng/mL, and it has been successfully validated with the actual sample milk. Therefore, it is a good strategy to fabricate highly sensitive FO LSPR sensors for detecting AFM1 by constructing AuNP multimers, and this approach is suitable for developing other biosensors.

This paper presents the first assessment of dietary exposure to aflatoxin M1 (AFM1) and associated health risks through milk and dairy product consumption in Armenia. Data on AFM1 in raw milk were obtained from an annual residue monitoring program. Additionally, commonly consumed dairy products (pasteurized milk, cheese, sour cream, curd cheese) were sampled, considering the sources of raw milk used by dairy companies. Per capita consumption of raw milk was sourced from national food balance databases, while individual consumption data for dairy products was collected via a 24 h recall survey with 1400 adult respondents. Detectable levels of AFM1 were observed in 7.14% of raw milk samples (up to 0.334 μg/kg) and, albeit at lower amounts (up to 0.009 µg/kg), in 30% and 40% of sour cream and curd cheese, respectively. The AFM1 levels were lower than the national maximum permitted level (0.5 μg/kg); however, levels in raw milk exceeded the EU ML (0.05 μg/kg). The estimated margin of exposure values for dairy products indicated no significant risk, whereas a reasonable worst-case estimate, using the measurable levels of AFM1 in raw milk consumption indicated a potential public health concern. This study provides a scientific basis for evaluating aflatoxin issues in the Caucasus area.

Given the highly mutagenic and carcinogenic nature of Aflatoxin M1 (AFM1), the quantity assessment of AFM1 residues in milk and dairy products is necessary to maintain consumer health and food safety. Herein, CRISPR-Cas12a-based colorimetric aptasensor was developed using the catalytic activity of flower-like nanozymes of MnO 2 and trans-cleavage property of CRISPR-Cas12a system to quantitatively detect AFM1. The basis of the developed colorimetric aptasensor relies on whether or not the CRISPR-Cas12a system is activated, as well as the contrast in oxidase-mimicking capability exhibited by flower-like MnO 2 nanozymes when AFM1 is absent or present. When AFM1 is not present in the sample, single-stranded DNA (ssDNA) is degraded by the activated CRISPR-Cas12a, and the solution turns into yellow due to the catalytic activity of the nanozymes. While, in the attendance of AFM1, ssDNA degradation does not occur due to the inactivation of the CRISPR-Cas12a. Therefore, with the adsorption of the ssDNA on the MnO 2 nanozymes, their catalytic activity decreases, and the solution color becomes pale yellow due to less oxidation of the chromogenic substrate. In this aptasensor, the relative absorbance changes increased linearly from 6 to 160 ng L -1 , and the detection limit was 2.1 ng L -1 . The developed aptasensor displays a selective detection performance and a practical application for quantitative analysis of AFM1 in milk samples. The results of the introduced aptasensor open up the way to design other selective and sensitive aptasensors for the detection of other mycotoxins by substitution of the used sequences., 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 © 2024 Elsevier B.V. All rights reserved.)

Breast milk (BM) is considered as the best source of nutrition which could have prevention effects on various diseases in the first years of a child. Along with nutritive compounds, presence of contaminants such as mycotoxins in BM could be transmitted into neonate. The aim of this study was to determine the occurrence, levels, and factors associated with the presence of aflatoxin M 1 (AFM 1 ) and ocratoxin a (OTA) in BM samples of nursing mothers in rural centers of Yazd, Iran. The presence and average AFM 1 and OTA concentration in 72 BM samples was measured by competitive ELISA. The demographic and diet parameters of nursing mothers were collected by a questionnaire and were analyzed using SPSS 18 software. AFM 1 and OTA were detected in 63 (87.5%) and 47 (65.2%) samples with the mean concentration levels of 19.46 ± 13.26 ng/L (ranges from 5.1 to 53.9) and 200 ± 160 ng/L (ranges from 100 to 2460), respectively. Of these, 32 samples (50.7%) for AFM 1 and 23 samples (48.9%) for OTA had values exceeding the limit set by the European Union regulation for infant foods (25 ng/L for AFM 1 and 500 ng/L for OTA). It was also found that the risk of AFM 1 and OTA occurrence in BM increased significantly with the consumption of beans, bread, cereals, fruit juice and crackers, and cream, respectively. This study showed that the estimated daily intake for AFM 1 and OTA by 1 month of age infants was 2.7 and 28.5 ng/kg bw/day, respectively, while, as the age of the infant increased, the values were lower and close to 0.9 and 9.9 ng/kg bw/day for AFM1 and OTA in 12 months of age infants, respectively. The high occurrence and noticeable levels of AFM 1 and OTA detected in this study indicated that some infants receive undesirable exposures to AFM 1 and OTA with breast milk. Therefore, it is recommended that mothers are advised to avoid certain foods during pregnancy and breastfeeding that are likely sources of mycotoxins., (© 2024. The Author(s) under exclusive licence to Society for Mycotoxin (Research Gesellschaft für Mykotoxinforschung e.V.) and Springer-Verlag GmbH Germany, part of Springer Nature.)

A novel ratiometric electrochemical aptasensor based on split aptamer and Au-reduced graphene oxide (Au-rGO) nanomaterials was proposed to detect aflatoxin M1 (AFM1). In this work, Au-rGO nanomaterials were coated on the electrode through the electrodeposition method to increase the aptamer enrichment. We split the aptamer of AFM1 into 2 sequences (S1 and S2), where S1 was immobilized on the electrode due to the Au-S bond, and S2 was tagged with methylene blue (MB) and acted as a response signal. A complementary strand to S1 (CS1) labeled with ferrocene (Fc) was introduced as another reporter. In the presence of AFM1, CS1 was released from the electrode surface due to the formation of the S1-AFM1-S2 complex, leading to a decrease in Fc and an increase in the MB signal. The developed ratiometric aptasensor exhibited a linear range of 0.03 μg L -1 to 2.00 μg L -1 , with a detection limit of 0.015 μg L -1 for AFM1 detection. The ratiometric aptasensor also showed a linear relationship from 0.2 μg L -1 to 1.00 μg L -1 , with a detection limit of 0.05 μg L -1 in natural milk after sample pretreatment, indicating the successful application of the developed ratiometric aptasensor. Our proposed strategy provides a new way to construct aptasensors with high sensitivity and selectivity., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Context: Aflatoxins are the most harmful mycotoxins that cause human and animal health concerns. Aflatoxin M 1 (AFM 1 ) is the primary hydroxylated metabolite of aflatoxin B 1 and is linked to the development of hepatocellular carcinoma and immunotoxicity in humans and animals. Because of the important role of dairy products in human life, especially children, AFM 1 is such a major concern to humans because of its frequent occurrence in dairy products at concentrations high enough to cause adverse effects to human and animal health. Reduced its bioavailability becomes a high priority in order to protect human and animal health., Objectives: This study aimed to investigate, in vivo , the ability of lactic acid bacteria ( lactobacillus rhamnosus GAF01, LR) and clay mineral (bentonite, BT) mixture to mitigate/reduce AFM 1 -induced immunotoxicity, hepatotoxicity, nephrotoxicity and oxidative stress in exposed Balb/c mice., Materials and Methods: The in vivo study was conducted using male Balb/c mice that treated, orally, by AFM 1 alone or in combination with LR and/or BT, daily for 10 days as follows: group 1 control received 200 µl of PBS, group 2 treated with LR alone (2.10 8 CFU/mL), group 3 treated with BT alone (1 g/kg bw), group 4 treated with AFM 1 alone (100 μg/kg), group 5 co-treated with LR + AFM 1 , group 6 co-treated with BT + AFM 1 , group 7 co-treated with BT + LR + AFM 1 . Forty-eight h after the end of the treatment, the mice were sacrificed and the blood, spleen, thymus, liver and kidney were collected. The blood was used for biochemical and immunological study. Spleen and thymus samples were used to thymocytes and splenocytes assessments. Liver and kidney samples were the target for evaluation of oxidative stress enzymes status and for histological assays., Results: The results showed that AFM 1 caused toxicities in male Blab/c mice at different levels. Treatment with AFM 1 resulted in severe stress of liver and kidney organs indicated by a significant change in the biochemical and immunological parameters, histopathology as well as a disorder in the profile of oxidative stress enzymes levels. Also, it was demonstrated that AFM 1 caused toxicities in thymus and spleen organs. The co-treatment with LR and/or BT significantly improved the hepatic and renal tissues, regulated antioxidant enzyme activities, spleen and thymus viability and biochemical and immunological parameters. LR and BT alone showed to be safe during the treatment., Conclusion: In summary, the LR and/or BT was able to reduce the biochemical, histopathological and immunological damages induced by AFM 1 and indeed it could be exploited as one of the biological strategies for food and feedstuffs detoxification.

Mycotoxins have been linked to adverse health impacts, including liver cancer and kidney diseases. The objectives of the current study were to evaluate the dietary exposure of Lebanese adults to multi-mycotoxins (aflatoxin B1 (AFB1), aflatoxin M1 (AFM1), ochratoxin A (OTA), ochratoxin B (OTB), deoxynivalenol (DON), T-2 and HT-2) and to assess their associated health risks. Hence, a nationally representative sample of 449 participants aged 18-64 years old were interviewed to obtain their socio-demographic characteristics, food consumption data and exposure estimates. A food frequency questionnaire and 24 h-recall were used to collect data. The concentration of mycotoxins in all foods consumed by the participants was collected from previous national published studies. The estimated daily intake (EDI), the hazard quotient (HQ) and the margin of exposure (MOE) were calculated. The total exposure to AFB1, AFM1, OTA and DON was 1.26, 0.39, 4.10 and 411.18 ng/kg bw/day, respectively. The MOE to AFB1, AFM1, OTA and DON in the Lebanese food basket was 316, 1454, 3539 and 510, respectively, indicating high health-related risks. Per food items, the MOE to AFB1 was below 10,000 in cereals (466.5), mainly in rice (827.9) and Burgul (4868.5). Similarly, the MOE to OTA in cereals was 1439, in which bread (4022), rice (7589) and bulgur (7628) were considered unsafe. Moreover, the MOE to DON in cereals (605) is alarming, especially in bread (632) and manakesh (6879). The MOE to AFM1 in dairy products was 1454, indicating health-related risks with a focus on yogurt (9788) and labneh (8153). As for the herbs/spices group and traditional dishes, the MOE to AFB1 was relatively lower than 10,000 (3690 and 1625, respectively), with a focus on thyme (2624) and kishik (3297), respectively. It is noteworthy that the MOE to DON and the MOE to OTA in traditional foods and coffee were lower than 10,000 (8047 and 8867, respectively). All hazard quotient (HQ) values were below 1, except the HQ value of milk and dairy products (1.96). The intake of some food groups varied between age categories, corresponding to differences in EDI between them. Thus, it is essential to put control measures in place to decrease the contamination and exposure to mycotoxins by Lebanese consumers.

Background: The assessment of risks related to food safety is becoming a challenge in developing countries with its consequent health hazards. Chemical risk assessment in dairy products is important to maintain consumer health locally and internationally. Since milk and dairy products are essential foods for a wide range of customers, mostly children, patients, and pregnant women, it is very important to estimate the risks of some chemical residues, such as pesticides, some heavy metals, and aflatoxins., Aim: This work aims to determine the levels of chemical contamination in milk and traditional Egyptian cheese., Methods: Heavy metals were determined in samples by atomic absorption spectrometry. GC-mass spectrometry (MS)/MS and LC-MS/MS were also used for measuring pesticide residues. The Aflatoxin M1 was determined by enzyme-linked immune-sorbent assay., Results: Raw milk samples were tested and showed elevated concentrations of lead and cadmium, (46% and 4%, respectively). The heavy metals detected in the Egyptian cheese samples were variable depending on the type of cheese. Moreover, p.p.-DDE phenofose was present in 45% and 29% of raw milk and Ras cheese samples, respectively. For Aflatoxin M1, only 7% of milk samples and 2.9% of Ras cheese samples exceeded the acceptable limits., Conclusion: More surveying and risk assessment of chemical residues in milk and milk products are essential for controlling health risks to consumers., Competing Interests: The authors declare that there is no conflict of interest.

Aflatoxin M1 (AFM1) and its precursor, Aflatoxin B1 (AFB1), are highly pathogenic and mutagenic substances, making the detection and sensing of AFB1/M1 a long-standing focus of researchers. Among various detection techniques, surface-enhanced Raman spectroscopy (SERS) is considered an ideal method for AFB1/M1 detection due to its ability not only to enhance characteristic frequencies but also to detect shifts in these frequencies with high repeatability. Therefore, we employed density functional theory in conjunction with surface-enhanced Raman spectroscopy to investigate the interaction between AFB1/M1 and a Au substrate in the context of the SERS effect for the first time. To predict the potential binding sites of AFB1/M1 and Au within the SERS effect, we performed calculations on the molecular electrostatic potential of AFB1/M1. Considering the crucial role of the binding energy in molecular docking studies, we computed the binding energy between two molecules interacting with Au at different binding sites. The molecular frontier orbitals and related chemical parameters of AFB1/M1 and "molecular-Au" complexes were computed to elucidate the alterations in AFB1/M1 molecules under the SERS effect. Subsequently, the theoretical Raman spectra of AFB1/M1 and the complexes were compared and analyzed, enabling determination of the adsorption conformation of AFB1/M1 on the gold surface based on SERS surface selection rules. These findings not only provide a deeper understanding of the interaction mechanism between molecules and substrates in the SERS effect but also offer theoretical support for developing novel aflatoxin SERS sensors.

Wagashi is a West African type cottage cheese locally prepared from cow milk. Wagashi like other milk products, is prone to microbial contamination, particularly by fungi. Many of these fungal species produce mycotoxins which are of serious public health concern. This work aimed to update the mycoflora profile and determine the concentrations of aflatoxin M1 and its health risk characterization due to the consumption of wagashi. Culturing the wagashi on mycological media (Oxytetracycline Glucose Yeast Extract OGYE, Dichloran Rose Bengal Chloramphenicol DRBC) caused a de-novo growth of the quiescent spores at 28-30 °C for 5-7 days. The analysis of AFM1 levels in the samples was done using High-Performance Liquid Chromatography connected to a Fluorescence detector (HPLC-FLD). The exposure and risk assessment to the AFMI levels were determined using deterministic models prescribed by the European Food Safety Authority (EFSA). The fungal counts ranged between 2.36-4.30 log10 CFU/g. In total, thirteen (13) fungal species from eight (8) genera were isolated from all wagashi samples. They are; Fusarium oxysporum, Aspergillus flavus, Aspergillus niger, Fusarium verticillioides, Penicillium digitatum, Trichoderma harzianum, Aspergillus terreus, Rhodotorula mucilaginosa, Rhizopus stolonifer, Aspergillus fumigatus, Yeast sp., Mucor racemosus and Fusarium oligosporum belonging to the genera Fusarium, Aspergillus, Penicillium, Trichoderma, Rhodotorula, Rhizopus, Yeast, and Mucor . The AFM1 observed in the wagashi samples' analysis was low, ranging from 0.00 (Not Detected) ± 0.00 - 0.06 ± 0.002 µg/Kg. Risk assessments of AFM1 using deterministic models produced outcomes that ranged between 5.92 × 10 -3 - 0.14 ng/kg bw/day, 1.42 -44.35, 0-0.0323 ng aflatoxins/kg bw/day, and 1.51 × 10 -3 - 9.69 × 10 -4 cases/100,000 person/yr for estimated daily intake (EDI), margin of exposure (MOE), average potency, and cancer risks, respectively, for the age categories investigated. Fungal counts were interpreted as medium to high. It was also established that the consumption of wagashi may pose adverse health effects on all age categories in the selected zones of the study since all calculated MOE values were less than 100,000., Competing Interests: 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., (© 2024 The Authors.)

Presence of aflatoxin B 1 (AFB 1 ) in food and feed is a serious problem, especially in developing countries. Human exposure to this carcinogenic mycotoxin can occur through dietary intake, but also through inhalation or dermal contact when handling and processing AFB 1 -contaminated crops. A suitable biomarker of AFB 1 exposure by all routes is the occurrence of its hydroxylated metabolite aflatoxin M 1 (AFM 1 ) in urine. To assess mycotoxin exposure in mill workers in Bangladesh, we analyzed AFM 1 levels in urine samples of this population group who may encounter both dietary and occupational AFB 1 exposure. In this pilot study, a total of 76 participants (51 mill workers and 25 controls) were enrolled from the Sylhet region of Bangladesh. Urine samples were collected from people who worked in rice, wheat, maize and spice mills and from controls with no occupational contact to these materials. A questionnaire was used to collect information on basic characteristics and normal food habits of all participants. Levels of AFM 1 in the urine samples were determined by a competitive enzyme linked immunosorbent assay. AFM 1 was detected in 96.1% of mill workers' urine samples with a range of LOD (40) of 217.7 pg/mL and also in 92% of control subject's urine samples with a range of LOD of 307.0 pg/mL). The mean level of AFM 1 in mill workers' urine (106.5 ± 35.0 pg/mL) was slightly lower than that of the control group (123.3 ± 52.4 pg/mL), whilst the mean AFM 1 urinary level adjusted for creatinine was higher in mill workers (142.1 ± 126.1 pg/mg crea) than in the control group (98.5 ± 71.2 pg/mg crea). Yet, these differences in biomarker levels were not statistically significant. Slightly different mean urinary AFM 1 levels were observed between maize mill, spice mill, rice mill, and wheat mill workers, yet biomarker values are based on a small number of individuals in these subgroups. No significant correlations were found between the study subjects' urine AFM 1 levels and their consumption of some staple food items, except for a significant correlation observed between urinary biomarker levels and consumption of groundnuts. In conclusion, this pilot study revealed the frequent presence of AFM 1 in the urine of mill workers in Bangladesh and those of concurrent controls with dietary AFB 1 exposure only. The absence of a statistical difference in mean biomarker levels for workers and controls suggests that in the specific setting, no extra occupational exposure occurred. Yet, the high prevalence of non-negligible AFM 1 levels in the collected urines encourage further studies in Bangladesh regarding aflatoxin exposure.

Background: Fungal metabolites known as aflatoxin M 1 (AFM 1 ) are linked to contaminated milk and milk products. Consuming food contaminated with AFM 1 poses major health risks and may even be fatal., Methods: The retrieved publications were categorized in this bibliometric study using the Web of Science (WoS) database Jan 1, 1970 to Nov 30, 2022 based on a variety of factors, including the time of publication of articles, citation totals, languages, research areas, countries, affiliations, funding agencies, journals, and keywords analysis to identify any hot and developing subjects. Additionally, VOSviewer software version 1.6.18 provided the bibliometric analysis of the global collaboration network and hot research themes., Results: Overall, 679 published documents were detected. Food Control was the top-line journal in publications on AFM 1 research with 540 published articles, while the USA was the best productive country in AFM 1 publications as well as the major country with the maximum co-authorship collaboration. This study ensures quantitative and qualitative analyses of the top 25 journals, most cited published articles, most relevant authors and title word occurrences in published documents on AFM 1 publications. Over the past two decades, there has been an enormous rise for research conducted on global AFM 1 ., Conclusion: The assessment of the historical state and development trend in AFM 1 scientific research can serve as a roadmap for future research and eventually, serve as a foundation for bettering management practices for territorial decisions, healthcare, and dairy industries., (Copyright© 2024 Altun et al. Published by Tehran University of Medical Sciences.)

Aim and Background: Aflatoxins, produced by Aspergillus flavus and Aspergillus parasiticus, are among the most toxic mycotoxins. Aflatoxin M1 (AFM1) is a hydroxylated metabolite of aflatoxin B1 (AFB1), found in milk and dairy products from animals fed AFB1-contaminated feed. Consumption of AFM1 has related adverse effects on human health. Breast milk can be a source of contamination for infants due to the presence of AFM. AFM1 can also contaminate powdered milk, a significant product of the milk industry. Consequently, monitoring dairy products for these toxins is imperative., Study Method: A total of 50 samples (25 samples of breast milk and 25 samples of powdered infant milk formula) were collected in Tehran from December 2021 to February 2022. HPLC method was used for the determination of AFM1 in samples., Results: and Discussion: AFM1 was detected in 72% of breast milk samples and 96% of powdered milk samples. AFM1 levels varied significantly between the two sample types (p < 0.05). The average amount of AFM1 in breast milk samples was 25.82 ± 4.72 ng/kg, while the average amount in powdered milk samples was 40.59 ± 7.76 ng/kg. Moreover, 44% of the breast and 68% of powdered milk samples exceeded the AFM1 content limit of the European Union and the Iranian national standard. This study concludes that given the importance of breast milk and formula to maternal and infant health, monitoring and regulating the toxin levels in these products in Tehran is crucial., 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.Authors declare no conflict of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Aflatoxin M1 (AFM1) is highly toxic, widely distributed, and difficult to monitor, posing a serious threat to human health. Therefore, a highly sensitive, rapid, convenient, and low-cost detection method must be urgently established. In this study, a triple strategy-enhanced immunochromatographic assay (ICA) was developed to satisfy these detection requirements. First, a turn-on signal output mode of the fluorescence quenching ICA substituted the turn-off mode of the traditional ICA for sensitive response to trace AFM1, with the limit of detection (LOD) reduced by approximately 4.9-fold. Then, a novel Au and polydopamine (PDA) cogrowth chrysanthemum-like blackbody was prepared as the quenching probe to reduce the background signal. This probe combined the excellent properties of Au nanoparticles with PDA. Thus, its fluorescence quenching constant was higher than that of single Au and PDA nanoparticles by 25.8- and 4.9-fold, respectively. Furthermore, an aggregation-induced emission fluorescence microsphere with a 5.7-fold higher relative quantum yield than a commercial fluorescence microsphere was selected as the signal output carrier to improve the signal-to-noise ratio. The integration of the above triple strategies established a 53.4-fold sensitivity-enhanced fluorescence quenching ICA (LOD = 0.9 pg/mL) for detecting AFM1 in milk, providing a strong technical guarantee for the safety monitoring of milk products., 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 © 2023 Elsevier B.V. All rights reserved.)

The aim of this work was to evaluate interaction between aflatoxin M1 (AFM1) and structural models of β-lactoglobulin (β-LG) at pH 4.0 and 6.5. This information would provide an explanation of the variability in AFM1 during cheese production. Once β-LG models were optimized using molecular dynamic (MD) simulation, it was found that a region of the Calyx cavity underwent conformational changes, at the E-F loop, from the closed conformation at pH 6.5 to the open at pH 4.0. No differences in Site C conformation were observed at both pH. The binding free energy ( ΔG b ) of the β-LG-AFM1 complexes at the different pHs were determined by molecular docking. The ΔG b values obtained for the Calyx cavity showed that at pH 4.0 there is a more stable complex formation compared to pH 6.5 with values of -42.6 and -32.0 kJ mol -1 , respectively. On the contrary, in the complexes formed in Site C at both pH´s there were no differences. Likewise, the ΔG b in the dimer interface was evaluated, obtaining a value of -29.3 kJ mol -1 , like those obtained at Site C. In addition, by the MD simulations of the β-LG-AFM1 complexes, it was observed that at acidic pH the binding of AFM1 with β-LG is more stable. In conclusion, the computational tools showed that the most stable complex was formed at the Calyx cavity at pH 4.0. This suggests that during cheese production using acidic coagulation, the whey proteins show higher affinity toward AFM1 which may explain the observed variability of mycotoxin.Communicated by Ramaswamy H. Sarma.

Food safety can be seriously threatened by the existence of both aflatoxin M1 (AFM1) and ochratoxin A (OTA) in milk and corresponding products. The importance of intestine integrity in preserving human health is widely understood in vitro, but the fundamental processes by which AFM1 and OTA cause disruption of the intestinal barrier are as yet unknown, especially in vivo. Based on the analysis of the whole transcriptome of BALB/c mice, the competing endogenous RNA (ceRNA) regulation network was obtained in the current study. Each of 12 mice were separated into five treatments: saline solution treatment, 1.0% DMSO vehicle control treatment, 3.0 mg/kg b.w. individual AFM1 treatment (AFM1), 3.0 mg/kg b.w. individual OTA treatment (OTA), and combined mycotoxins treatment (AFM1 +OTA). The study period lasted 28 days. The jejunum tissue was collected for the histological assessment and whole transcriptome analysis, and the whole blood was collected, and determination of serum biochemical indicators. The phenotypic results demonstrated that AFM1 and OTA caused intestinal barrier disruption via an increased apoptosis level and decreased expression of tight junction (TJ) proteins. The ceRNA network demonstrated that AFM1 and OTA induced cell apoptosis through activating the expression of DUSP9 and suppressing the expression of PLA2G2D, which were regulated by differentially expressed microRNAs (DEmiRNAs) (miR-124-y, miR-194-z, miR-224-x, and miR-452-x) and differentially expressed long non-coding RNAs (DElncRNAs) (FUT8 and GPR31C). And AFM1 and OTA decreased TJ proteins via inhibiting the expression of PAK6, which was regulated by several important DEmiRNAs and DElncRNAs. These DE RNAs in intestinal integrity were involved in MAPK and Ras signaling pathway. Overall, our findings expand the current knowledge regarding the potential mechanisms of intestinal integrity disruption brought on by AFM1 and OTA in vivo., 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. Declaration of competing interest All the authors declare no conflict of interest., (Copyright © 2023. Published by Elsevier Inc.)

Herein, we synthesized a novel porphyrinic covalent organic polymer (TPAPP-PTCA PCOP) for constructing a polarity-switchable dual-wavelength photoelectrochemical (PEC) biosensor with ferrocene (Fc) and hydrogen peroxide (H 2 O 2 ) as regulator and amplifier simultaneously. Interestingly, this new PCOP possessed both n-type and p-type semiconductor characteristics, which thus enabled the appearance of a dual-polarity photocurrent at two different excitation wavelengths. Furthermore, Fc and H 2 O 2 could readily switch the photocurrent of PCOP to the cathode and anode stemming from its efficient electron collection and donation function, respectively. Based on these, a PCOP-based PEC biosensor skillfully integrating dual wavelengths with reliable accuracy and polarity switch with high sensitivity was instituted. As a result, the developed PEC biosensor exhibited a low detection limit down to 0.089 pg mL -1 for the most powerful natural carcinogen aflatoxin M1 (AFM1) assay. Impressively, the target exhibited a completely opposite photocurrent difference to the interfering substances, and the linear correlation coefficient of the assay was improved compared to single-wavelength detection. The PEC sensing platform not only provided a basis for exploring multicharacteristic photoactive material but also innovatively developed the detection mode of the PEC biosensor.

The aim of this study, research the potential use of probiotics in reducing the toxic effect of Aflatoxin M1 in cow milk, goat milk, sheep milk, and Phosphate-buffered saline (PBS). Milk and Phosphate-buffered saline were contaminated with Aflatoxin M1 at a concentration of 100 ppt. Then, various study groups were formed by adding Lactobacillus acidophilus DSMZ 20079, Lactobacillus rhamnosus GG, and Bifidobacterium bifidum DSMZ 20456 probiotic bacteria at a density of 108 CFU/ml. Then, working groups were stored for 1 day and Aflatoxin M1 levels were analyzed by an Enzyme-Linked Immunosorbent Assay kit. The binding level of Aflatoxin M1 by probiotic bacteria varies between 2.32-12.52% in Phosphate-buffered saline, 9.08-40.14% in cow milk, 15.01-38.01% in goat milk, and 32.49-42.90% in sheep milk. The highest binding level of Aflatoxin M1 was detected in sheep milk and the lowest in Phosphate-buffered saline. The binding ability of Aflatoxin M1 is ranked from highest to lowest in sheep milk, cow milk, and goat milk. The data obtained from this study is important because it is the first study to show that if sheep and goat milk is enriched with probiotics, it can reduce AFM1 exposure.

Aflatoxins are considered as reproductive toxins for mammalian species. Here, we studied the effect of aflatoxin B1 (AFB1) and its metabolite aflatoxin M1 (AFM1) on the development and morphokinetics of bovine embryos. Cumulus oocyte complexes (COCs) were matured with AFB1 (0.032, 0.32, 3.2, 32 µM) or AFM1 (0.015, 0.15, 1.5, 15, 60 nM), then fertilized and the putative zygotes were cultured in an incubator equipped with a time-lapse system. Exposing COCs to 32 µM AFB1 or 60 nM AFM1 reduced the cleavage rate, whereas exposing them to 3.2 or 32 µM AFB1 further reduced the blastocyst formation. A delay was recorded for the first and second cleavages in a dose-dependent manner for both AFB1- and AFM1-treated oocytes. A delay was recorded in the third cleavage in the AFM1-treated group. To explore potential mechanisms, subgroups of COCs were examined for nuclear and cytoplasmic maturation (n = 225; DAPI and FITC-PNA, respectively), and mitochondrial function was examined in a stage-dependent manner. COCs were examined for their oxygen consumption rates (n = 875; Seahorse XFp analyzer) at the end of maturation, MII-stage oocytes were examined for their mitochondrial membrane potential (n = 407; JC1), and putative zygotes were examined using a fluorescent time-lapse system (n = 279; IncuCyte). Exposing COCs to AFB1 (3.2 or 32 µM) impaired oocyte nuclear and cytoplasmic maturation and increased mitochondrial membrane potential in the putative zygotes. These alterations were associated with changes in the expression of mt-ND2 (32 µM AFB1) and STAT3 (all AFM1 concentrations) genes in the blastocyst stage, suggesting a carryover effect from the oocyte to the developing embryos., 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 © 2023 Elsevier Inc. All rights reserved.)

Cheese is popular in Iran because of its high nutritional value; therefore, it is necessary to control this product regarding health risk factors, particularly aflatoxin M1 (AFM1). This research reviewed AFM1 in various varieties of cheese in Iran to assess the potential health risks associated with consuming these products for different age groups. In this regard, all accessible papers from different databases were screened between June 27, 2000 and October 10, 2022 b y systematic research and then considering the selection criteria of the studies; finally, 22 articles were selected for the current review. The amount and prevalence of AFM1 were calculated and separated based on the cheese variety, and the sampling location; health risk assessment (HRA), statistical, uncertainty, and sensitivity analysis for AFM1 of cheese for different age groups were performed. The study results for 2143 samples showed that the overall average AFM1 for cheese is 160 ± 175 ng/kg, below the European Commission (EC) regulation (250 ng/kg). AFM1 contaminated 72.42% of all cheese samples, and 13% of these contaminated samples had a higher AFM1 than the EC regulation. Cheese varieties were ranked based on average levels of AFM1 as white pasteurized > traditional > creamy > probiotic > Lighvan, and this ranking was obtained based on sampling locations as market > dairy factories > livestock farms. Based on the HRAs, from the perspective of the liver cancer risk (LCR), the margin of exposure (MOE), and the hazard index (HI) approach, it can be concluded that cheese produced in Iran, in terms of AFM1, particularly for children, poses serious health risks. Accordingly, it is imperative to carefully consider implementing suitable management methods to inhibit the growth of aflatoxin B1 (AFB1) in livestock fodder, and training in sanitary production and processing of dairy products according to world standards is suggested for industrial and traditional cheese producers across Iran., Competing Interests: 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., (© 2023 The Authors.)

Milk is a staple food that is essential for human nutrition because of its high nutrient content and health benefits. However, it is susceptible to being contaminated by Aflatoxin M1 (AFM1), which is a toxic metabolite of Aflatoxin B1 (AFB1) presented in cow feeds. This research investigated AFM1 in Tunisian raw cow milk samples. A total of 122 samples were collected at random from two different regions in 2022 (Beja and Mahdia). AFM1 was extracted from milk using the QuEChERS method, and contamination amounts were determined using liquid chromatography (HPLC) coupled with fluorescence detection (FD). Good recoveries were shown with intra-day and inter-day precisions of 97 and 103%, respectively, and detection and quantification levels of 0.003 and 0.01 µg/L, respectively. AFM1 was found in 97.54% of the samples, with amounts varying from values below the LOQ to 197.37 µg/L. Lower AFM1 was observed in Mahdia (mean: 39.37 µg/L), respectively. In positive samples, all AFM1 concentrations exceeded the EU maximum permitted level (0.050 µg/L) for AFM1 in milk. In Tunisia, a maximum permitted level for AFM1 in milk and milk products has not been established. The risk assessment of AFM1 was also determined. Briefly, the estimated intake amount of AFM1 by Tunisian adults through raw cow milk consumption was 0.032 µg/kg body weight/day. The Margin of Exposure (MOE) values obtained were lower than 10,000. According to the findings, controls as well as the establishment of regulations for AFM1 in milk are required in Tunisia.

Milk contaminated with aflatoxin can lead to liver cancer. Aflatoxin B1 (AFB1), a serious animal feed contaminant, is transformed into Aflatoxin M1 (AFM1) and secreted in milk. In this study, a biological method using probiotic bacteria, Lactobacillus rhamnosus ( L. rhamnosus ) in combination with Saccharomyces cerevisiae ( S. cerevisiae ), was used to assess their antiaflatoxigenic effect in animal milk. A Box-Behnken design was used to establish the optimal ratio of L. rhamnosus and S. cerevisiae , incubation time, and temperature for efficient AFM1 detoxification from milk. To achieve this, the primary, interaction, and quadratic effects of the chosen factors were investigated. To investigate the quadratic response surfaces, a second-order polynomial model was built using a three-factor, three-level Box-Behnken design. The quantity of AFM1 was detected by the ELISA technique. The results of these experiments obtained an optimum condition in AFM1 detoxification of the three tested factors in order to maximize their effect on AFM1 detoxification in milk. The model was tested in three highly contaminated milk samples to assure the efficacy of the model. AFM1 detoxification was up to 98.4% in contaminated milk samples. These promising results provide a safe, low-cost, and low-time-consuming solution to get rid of the problem of milk contamination with AFM1.

The increasing incidence of diseases caused by highly carcinogenic aflatoxin M 1 (AFM 1 ) in food demands a simple, fast, and cost-effective detection technique capable of sensitively monitoring AFM 1 . Recent works predominantly focus on the electrochemical aptamer-based biosensor, which still faces challenges and high costs in experimentally identifying an efficient candidate aptamer. However, the direct electrochemical detection of AFM 1 has been scarcely reported thus far. In this study, we observed a significant influence on the electrochemical signals of ferric ions at a gold nanoparticle-modified glassy carbon electrode (AuNPs/GCE) by adding varying amounts of AFM 1 . Utilizing ferricyanide as a sensitive indicator of AFM 1 , we have introduced a novel approach for detecting AFM 1 , achieving an unprecedentedly low detection limit of 1.6 × 10 -21 g/L. Through monitoring the fluorescence quenching of AFM 1 with Fe 3+ addition, the interaction between them has been identified at a ratio of 1:936. Transient fluorescence analysis reveals that the fluorescence quenching process is predominantly static. It is interesting that the application of iron chelator diethylenetriaminepentaacetic acid (DTPA) cannot prevent the interaction between AFM 1 and Fe 3+ . With a particle size distribution analysis, it is suggested that a combination of AFM 1 and Fe 3+ occurs and forms a polymer-like aggregate. Nonetheless, the mutual reaction mechanism between AFM 1 and Fe 3+ remains unexplained and urgently necessitates unveiling. Finally, the developed sensor is successfully applied for the AFM 1 test in real samples, fully meeting the detection requirements for milk.

Aflatoxin M1 (AFM1) is detected in the milk of animals after ingestion of aflatoxin B1-contaminated food; since 2002, it has been categorized as a group I carcinogen. In this work, a silicon-based optoelectronic immunosensor for the detection of AFM1 in milk, chocolate milk, and yogurt has been developed. The immunosensor consists of ten Mach-Zehnder silicon nitride waveguide interferometers (MZIs) integrated on the same chip with the respective light sources, and an external spectrophotometer for transmission spectra collection. The sensing arm windows of MZIs are bio-functionalized after chip activation with aminosilane by spotting an AFM1 conjugate with bovine serum albumin. For AFM1 detection, a three-step competitive immunoassay is employed, including the primary reaction with a rabbit polyclonal anti-AFM1 antibody, followed by biotinylated donkey polyclonal anti-rabbit IgG antibody and streptavidin. The assay duration was 15 min with limits of detection of 0.005 ng/mL in both full-fat and chocolate milk, and 0.01 ng/mL in yogurt, which are lower than the maximum allowable concentration of 0.05 ng/mL set by the European Union. The assay is accurate (% recovery values 86.7-115) and repeatable (inter- and intra-assay variation coefficients <8%). The excellent analytical performance of the proposed immunosensor paves the way for accurate on-site AFM1 determination in milk.

Camel milk is known as a source of nutritional and health supplements. It is known to be rich in peptides and functional proteins. One main issue facing it is related to its contamination, mainly with aflatoxins. The present study aimed to evaluate camel milk samples from different regions while trying to reduce its toxicity using safe approaches based on probiotic bacteria. Collected samples of camel milk were sourced from two main regions: the Arabic peninsula and North Africa. Samples were tested for their contents of aflatoxins (B 1 and M 1 ) using two techniques to ensure desired contamination levels. Additionally, feed materials used in camel foods were evaluated. Applied techniques were also tested for their validation. The antioxidant activity of camel milk samples was determined through total phenolic content and antioxidant activity assays. Two strains of probiotic bacteria ( Lactobacillus acidophilus NRC06 and Lactobacillus plantarum NRC21) were investigated for their activity against toxigenic fungi. The result revealed high contamination of aflatoxin M 1 for all samples investigated. Furthermore, cross-contamination with aflatoxin B 1 was recorded. Investigated bacteria were recorded according to their significant inhibition zones against fungal growth (11 to 40 mm). The antagonistic impacts were between 40% and 70% against toxigenic fungi. Anti-aflatoxigenic properties of bacterial strains in liquid media were recorded according to mycelia inhibition levels between 41 to 52.83% against Aspergillus parasiticus ITEM11 with an ability to reduce aflatoxin production between 84.39% ± 2.59 and 90.4% ± 1.32 from media. Bacteria removed aflatoxins from the spiked camel milk in cases involving individual toxin contamination.

Aflatoxin B1 (AFB1) and aflatoxin M1 (AFM1) are universally found as environmental pollutants. AFB1 and AFM1 are group 1 human carcinogens. Previous sufficient toxicological data show that they pose a health risk. The intestine is vital for resistance to foreign pollutants. The enterotoxic mechanisms of AFB1 and AFM1 have not been clarified at the metabolism levels. In the present study, cytotoxicity evaluations of AFB1 and AFM1 were conducted in NCM 460 cells by obtaining their half-maximal inhibitory concentration (IC50). The toxic effects of 2.5 μM AFB1 and AFM1 were determined by comprehensive metabolomics and lipidomics analyses on NCM460 cells. A combination of AFB1 and AFM1 induced more extensive metabolic disturbances in NCM460 cells than either aflatoxin alone. AFB1 exerted a greater effect in the combination group. Metabolomics pathway analysis showed that glycerophospholipid metabolism, fatty acid degradation, and propanoate metabolism were dominant pathways that were interfered with by AFB1, AFM1, and AFB1+AFM1. Those results suggest that attention should be paid to lipid metabolism after AFB1 and AFM1 exposure. Further, lipidomics was used to explore the fluctuation of AFB1 and AFM1 in lipid metabolism. The 34 specific lipids that were differentially induced by AFB1 were mainly attributed to 14 species, of which cardiolipin (CL) and triacylglycerol (TAG) accounted for 41%. AFM1 mainly affected CL and phosphatidylglycerol, approximately 70% based on 11 specific lipids, while 30 specific lipids were found in AFB1+AFM1, mainly reflected in TAG up to 77%. This research found for the first time that the lipid metabolism disorder caused by AFB1 and AFM1 was one of the main causes contributing to enterotoxicity, which could provide new insights into the toxic mechanisms of AFB1 and AFM1 in animals and humans.

Monitoring aflatoxin M1 (AFM1) in dairy products and milk-based foods is very important. The main purpose of this research was to investigate and determine the amount and human health risk assessment of aflatoxin M1 (AFM1) in the most famous and widely used brands of infant dried powder milk (IDPM) consumed in Iran. For this study, 45 imported IDPM (IM-IDPM) samples and 45 domestically produced IDPM (DO-IDPM) samples (a total of 90 samples) were selected randomly. All samples were analyzed for AFM1 using a competitive enzyme-linked immunosorbent assay (ELISA) technique. The mean level and the percentage of positive samples for AFM1 in DO-IDPM were 9.2 ± 5.4 ng/kg and 73.3%, and for IM-IDPM, they were 5.1 ± 3.8 ng/kg and 33.3%, respectively. The average level of AFM1 in all samples was lower than the EU and Iranian national standards (25 ng/kg). AFM1 intake through IDPM consumption by Iranian infants less than one-year-old was lower than the allowable level, but the hazard quotient for infants less than 6 months was higher than the allowable level. Although the concentration of AFM1 in IDPM consumed in Iran is not hazardous, since Iranian infants may be exposed to AFM1 through other sources, including baby food, breast milk, raw and pasteurized milk, continuous monitoring of IDMP quality in Iranian markets should be taken., Competing Interests: Declaration of Competing Interests 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 © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Aflatoxin M 1 (milk toxin) found in milk is formed from the hepatic biotransformation of AFB 1 (aflatoxin B 1 ) and poses a risk to human health when consumed. The risk assessment of AFM 1 exposure due to milk consumption is a valuable way to assess health risk. The objective of the present work was to determine an exposure and risk assessment of AFM 1 in raw milk and cheese, and it is the first of its kind in Ethiopia. Determination of AFM 1 was conducted using an enzyme-linked immunosorbent assay (ELISA). The results indicated that AFM 1 was positive in all samples of milk products. The risk assessment was determined using margin of exposure (MOE), estimated daily intake (EDI), hazard index (HI), and cancer risk. The mean EDIs for raw milk and cheese consumers were 0.70 and 0.16 ng/kg bw/day, respectively. Our results showed that almost all mean MOE values were <10,000, which suggests a potential health issue. The mean HI values obtained were 3.50 and 0.79 for raw milk and cheese consumers, respectively, which indicates adverse health effects for large consumers of raw milk. For milk and cheese consumers, the mean cancer risk was 1.29 × 10 -6 and 2.9 × 10 -6 cases/100,000 person/year, respectively, which indicates a low risk for cancer. Therefore, a risk assessment of AFM 1 in children should be investigated further as they consume more milk than adults.

The aim of this study was to investigate the in vivo transfer of aflatoxin B1 (AFB1) to Assaf ewes' milk (aflatoxin M1, AFM1) and its effect on animal performance and health. Thirty Assaf ewes were allocated to three groups (C, L, H), and received a different individual daily dose of AFB1 (0, 40 and 80 μg) for 13 days. Milk (days 1, 2, 3, 4, 7, 14, 16 and 18) and blood (days 1, 7, 14 and 18) samples were collected. Milk yield, composition (except protein) and somatic cell counts (SCC) were not affected by AFB1 intake ( p > 0.05). Haemoglobin concentration increased ( p < 0.05) and haematocrit and alanine aminotransferase levels tended to increase ( p < 0.10) in group H on day 14. AFM1 excretion was highly variable and detected in L and H animals from days 1 to 16 (3 days increase, 10 days steady-state, 3 days clearance). Carry-over rate (0.23%) was significantly higher in L (0.22-0.34%) than in H (0.16-0.19%) animals ( p < 0.05). AFB1 daily doses of 40 to 80 µg do not impair milk yield; however, it may start affecting animals' health. Milk AFM1 depends mainly on the AFB1 intake whereas carryover rate is positively influenced by the level of milk production.

Aflatoxin M1 (AFM1) and ochratoxin A (OTA) are common mycotoxins in cereal foods and milk products, and may cause serious negative impacts on human health. The intestine is crucial for immune regulation as it protects host homeostatic health from external contaminants; however, the underlying mechanisms of AFM1 and OTA mediated intestinal immunotoxicity remain unclear. In this study, whole transcriptome analysis was used to characterize BALB/c mouse intestines exposed to individual and combined AFM1 and OTA [3.0 mg/kg body weight (BW)] for 28 days to screen for key intestinal immunotoxicity-related differentially expressed mRNAs (DEmRNAs), differentially expressed microRNAs (DEmiRNAs), differentially expressed long non-coding RNAs (DElncRNAs), and associated enriched signaling pathways. Functional validation was then conducted in intestinal differentiated Caco-2 cells using different inhibitor assays to verify the accuracy of transcriptome and the importance of the key screened regulatory factors. In vivo data revealed that AFM1 and OTA exposure disrupted the intestines and exerted intestinal immunosuppression effects. When compared with AFM1, OTA had stronger intestinal toxicity in combined treatments. Further analyses of competitive endogenous RNA (ceRNA) regulatory networks in mice showed that AFM1 and OTA mediated-intestinal immunosuppression was putatively explained as follows: (i) toxins affected DEmRNAs regarding transfer and transduction mechanisms between cells (Csf1, Csf1r, Cxcl10, Cx3cr1, and Irf1), which were regulated by key DEmiRNAs (miR-106-x, miR-107-y, and miR-124-y) and the DElncRNA Rian, and (ii) toxins inhibited transforming growth factor-β-activated kinase 1 (TAK1)/I-kappaB kinase (IKK)/inhibitor of kappa Bα (IκBα)/p65 nuclear factor-κB (NF-κB) signaling phosphorylation levels, which was validated in differentiated Caco-2 cells using the TAK1 inhibitor (5Z-7-oxozeaenol). In conclusion, we evaluated the risk of co-exposure to AFM1 and OTA and associated health hazards from a whole transcriptome perspective., 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 © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Aflatoxins are natural toxicants produced mainly by species of the Aspergillus genus, which contaminate virtually all feeds and foods. Apart from their deleterious health effects on humans and animals, they can be secreted unmodified or carried over into the milk of lactating females, thereby posing health risks to suckling babies. Aflatoxin M1 (AFM1) is the major and most toxic aflatoxin type after aflatoxin B1 (AFB1). It contaminates human breast milk upon direct ingestion from dairy products or by carry-over from the parent molecule (AFB1), which is hydroxylated in the liver and possibly in the mammary glands by cytochrome oxidase enzymes and then excreted into breast milk as AFM1 during lactation via the mammary alveolar epithelial cells. This puts suckling infants and children fed on this milk at a high risk, especially that their detoxifying activities are still weak at this age essentially due to immature liver as the main organ responsible for the detoxification of xenobiotics. The occurrence of AFM1 at toxic levels in human breast milk and associated health conditions in nursing children is well documented, with developing countries being the most affected. Different studies have demonstrated that contamination of human breast milk with AFM1 represents a real public health issue, which should be promptly and properly addressed to reduce its incidence. To this end, different actions have been suggested, including a wider and proper implementation of regulatory measures, not only for breast milk but also for foods and feeds as the upstream sources for breast milk contamination with AFM1. The promotion of awareness of lactating mothers through the organization of training sessions and mass media disclosures before and after parturition is of a paramount importance for the success of any action. This is especially relevant that there are no possible control measures to ensure compliance of lactating mothers to specific regulatory measures, which can yet be appropriate for the expansion of breast milk banks in industrialized countries and emergence of breast milk sellers. This review attempted to revisit the public health issues raised by mother milk contamination with AFM1, which remains undermined despite the numerous relevant publications highlighting the needs to tackle its incidence as a protective measure for the children physical and mental health.

In this present study, 195 cow milk, 100 goat milk, 50 buffalo milk, 50 camel milk, and 50 yak milk samples were collected in China in May and October 2016. The presence of aflatoxin M1 (AFM1) was determined using enzyme-linked immunosorbent assay method. For all cow milk samples, 128 samples (65.7%) contained AFM1 in concentrations ranging from 0.005 to 0.191 µg/L, and 6 samples (3.1%) from Sichuan province in October were contaminated with AFM1 above 0.05 µg/L (EU limit). For all goat milk samples, 76.0% of samples contained AFM1 in concentrations ranging from 0.005 to 0.135 µg/L, and 9 samples (9.0%) from Shanxi province in October were contaminated with AFM1 above 0.05 µg/L. For all buffalo milk samples, 24 samples (48.0%) contained AFM1 in concentrations ranging from 0.005 to 0.089 µg/L, and 2 samples collected in October were contaminated with AFM1 above 0.05 µg/L. Furthermore, 28.0% of samples contained AFM1 in concentrations ranging from 0.005 to 0.007 µg/L in camel milk samples, and 18.0% of samples contained AFM1 in concentrations ranging from 0.005 to 0.007 µg/L in yak milk samples. Our survey study has expanded the current knowledge of the occurrence of AFM1 in milk from five dairy species in China, in particular the minor dairy species.

Aflatoxin M1 (AFM1), a group 1 carcinogen, is a risk factor to be monitored in milk. This study aimed to investigate the occurrence of AFM1 in milk in Xinjiang, China, and to assess the risk of exposure for milk consumers in different age-sex groups. A total of 259 milk samples including pasteurized milk (93 samples), extended-shelf-life (ESL) milk (96), and raw donkey milk (70) were collected in Xinjiang from January to March in 2022. The AFM1 content of the milk samples was detected using a validated ELISA method. Of the 259 total samples analyzed for AFM1, 84 (32.4%) samples were contaminated at levels greater than the detection limit of 5 ng/L, with the maximum level of 16.5 ng/L. The positive rates of AFM1 in pasteurized milk and ESL milk were 43.0% ( n = 40) and 45.8% ( n = 44), respectively, and AFM1 was undetectable in donkey milk. The estimated daily intakes of AFM1 in each age group were lower than the hazard limits and were similar between male and female milk consumers. Therefore, the AFM1 contamination of milk in Xinjiang is low but still needs to be continuously monitored considering that children are susceptible to AFM1.

A new green competitive ELISA for aflatoxin M1 quantification in raw milk was developed. This diagnostic tool is based on an anti AFM1 mAb produced by plant molecular farming in alternative to classical systems. Our assay, showing an IC 50 below 25 ng/L, fits with the requirements of EU legislation limits for AFM1 (50 ng/L). Optimal accuracy was achieved in correspondence of the decision levels (25 and 50 ng/L), and the assay enabled AFM1 quantification in the range 5-110 ng/L, with limit of detection 3 ng/L. Moreover, to evaluate a real applicability in diagnostics, raw milk-spiked samples were analysed, achieving satisfactory recovery rates of AFM1. In conclusion, an efficient and ready-to-use diagnostic assay for the quantification of aflatoxin M1 in milk, based on a plant-produced recombinant mAb, has been successfully developed.

Cold plasma technology is a novel non-thermal technology that has shown promising results for food decontamination and improving food safety. This study investigates the efficacy of high voltage atmospheric cold plasma (HVACP) system to reduce Aflatoxin M 1 (AFM 1 ) in skim and whole milk. A dielectric barrier discharge HVACP was employed at 90 kV using modified air (MA65: 65 % O 2 , 30 % CO 2 , 5 % N 2 ) fill gas for 1,3, and 5 min. Skim and whole milk was spiked with 1.0 μg/L AFM 1 and exposed to HVACP treatment in both direct or indirect mode with no post-treatment storage or 4.0 h post storage at room temperature. Optimum condition of toxin degradation was chosen as for quality assessment including color, conductivity, total dissolved solid (TDS), pH, viscosity, peroxide value (PV), Thiobarbituric acid reactive substance (TBARS) assay, Fourier-transform infrared spectroscopy (FTIR), as well as nutrient composition. A one-minute HVACP treatment degrades 41.9 % and 37.8 % of AFM 1 in skim milk and whole milk, respectively. However, much greater reductions were seen after a short treatment and then post-treatment storage. A greater than 87 % reduction in AFM 1 was observed for all samples after a 3 min HVACP treatment with 4.0 h of post treatment. These results suggest that a few minutes of HVACP treatment is sufficient to generate significant reactive plasma species in the milk. Quality changes were less significant with shorter post treatment time and indirect mode of exposure. Overall, HVACP is an effective solution for decontamination of milk from AFM 1 ., 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 © 2022 Elsevier Ltd. All rights reserved.)

Currently, there is a great demand for simple, sensitive, and accurate sensors for aflatoxin M1 (AFM1) in dairy products. In the present research, a novel fluorescent immunosensor based on nitrogen-doped carbon quantum dots (CQDs) has been developed for AFM1 analysis. The N-doped CQDs were synthesized through the hydrothermal approach using citric acid and polyethyleneimine as precursors. The CQDs showed bright blue emission under ultraviolet light irradiation and a maximum emission was observed at 450 nm upon excitation at 350 nm. The anti-AFM1 antibody (Ab) was immobilized on the as-obtained amine-functionalized CQDs and the obtained CQDs/Ab probe was then directly used for developing the immunoassays for AFM1. The fluorescence of the CQDs/Ab solution was effectively quenched in the presence of increasing AFM1 concentrations. Under the optimized conditions, the fluorescent nanosensor exhibited high sensitivity towards AFM1 in the range of 0.2-0.8 ng/mL with low limit of detection i.e., 0.07 ng/mL in standard buffer. Furthermore, the CQDs/Ab immunosensor was developed as a lateral flow design for detecting the aflatoxin residues in milk. This strategy can be used for the development of low-cost, rapid, and highly sensitive sensor strips for the detection of AFM1 in dairy products., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The simultaneous detection of three kinds of small-molecule contaminants (antibiotics, mycotoxins, and hormones) in milk was realized by using an 8-17 DNAzyme-based fluorescent enzyme-linked immunosorbent assay (ELISA), in which 8-17 DNAzyme was utilized as the catalytic enzyme for amplifying the signal. Compared with the conventional ELISA in which horseradish peroxidase is used as the catalyzing factor, this 8-17 DNAzyme-based ELISA could achieve multicolor signal output with lower detection limits. The linearities for chloramphenicol, 17β-estradiol, and aflatoxin M 1 were in the range of 0.3 ng/mL-3 μg/mL, 3 ng/mL-3 μg/mL, and 3 pg/mL-3 ng/mL with quantitation limits of 0.3, 3, and 0.003 ng/mL, respectively. This proposed scheme demonstrated that the 8-17 DNAzyme might be an effective substitute for horseradish peroxidase in ELISA for the development of ultrasensitive and multicolor fluorescence immunoassay, which would stimulate the development of ELISA in a new orientation.

This study investigates the reduction of aflatoxin M1 (AFM1) in skim milk by using ultraviolet light at 254 nm and the effects of influencing factors on the efficacy including treatment time (min), depth of samples (mm), contamination level (μg L -1 ), stirring, temperature, and fat content in milk. The colour and pH of milk samples were measured to evaluate the influence of the treatment on these values. It was found that short-wave ultraviolet radiation (UVC) reduced up to 50% of AFM1 in milk after 20 min of treatment regardless of the initial AFM1 contamination level. Treatment time, depth of samples, and stirring were all found to significantly (P < 0.05) enhance the reduction of AFM1. The milk colour was affected but there was no influence on the pH of milk samples at any duration of UV exposure. It is concluded that UVC light treatment has the potential to reduce AFM1 in milk., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Cold plasma has potential for the degradation of aflatoxins in corn and hazelnuts; however, this has not been demonstrated for aflatoxin in milk. In this study, the efficacy of high voltage atmospheric cold plasma (HVACP) on the reduction of aflatoxin M1 (AFM1) in skim milk improved with increasing treatment times (1-20 min), using gas containing 65% oxygen (MA65) rather than air, increasing voltage (60-80 kV) and reducing sample volume (30 mL-10 mL). Direct treatment was more effective than indirect treatment. AFM1 in milk was degraded by 65.0 % and 78.9 % by air and MA65 respectively in 20 min with no change in milk colour. The toxicity of AFM1 after treatment was assessed using a brine shrimp model. A five-minute HVACP treatment reduced the toxicity of AFM1 by 83.9 % based on the increase in brine shrimp survival. HVACP is a promising method to reduce AFM1 in milk., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The analysis of food samples is a challenging task. The high complexity of food matrices hinders the extraction and detection of analytes from them. Therefore, the correct preparation of food samples is a crucial step for their subsequent analysis, as it achieves the proper isolation and preconcentration of analytes and removes the interfering proportion of the food matrix before instrumental analysis. We aimed to develop a method that not only satisfies the requirement of detecting trace compounds in complex matrices but also achieves a "greener" approach by reducing the use of organic solvents and non-degradable materials to minimize the health hazards posed to the operators as well as pollution to the environment. In this study, we prepared egg white as a concentrated gel and used this material for the biological purification of milk samples. After the milk protein was removed by acidification and salting, the residual amount of aflatoxin M1 in milk samples was quantitatively determined by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The results showed that the novel egg white purification method possessed advantages over the immunoaffinity technique used as the reference method in extraction recovery, sensitivity, repeatability, and operability. The limit of detection (LOD) was 0.001 μg/kg. In spiked samples containing 0.01 μg/kg to 2 μg/kg of AFM1, the average recovery was 88.3-94.7%, with a precision of 6.1-11.0%. Improved repeatability was obtained by significantly reducing the operation time and resource requirements compared with the immunoaffinity technique currently used internationally. This study provides a reference for the further improvement of the relevant international standards in place for the detection of aflatoxin M1 in milk.

We assessed the occurrence of aflatoxin M 1 (AFM 1 ) in urine in a sample of the MAUCO population-based cohort (n = 120) using an enzyme-linked immune sorbent assay (ELISA) kit specially designed for the analysis of AFM 1 in urine. We found AFM 1 in the urine of 59% of the participants (> limit of detection), with 12% of the samples being over the limit of quantification. The mean of the quantifiable samples was 0.66 (± 0.35) ng/mg adjusted creatinine, ranging from 0.31 to 1.39 ng/mg creatinine. The mean probable daily intake (PDI) of AFB 1 was 0.23 (± 0.37) ng/kg bw according to the upper bound (UB), being significantly higher in women and 0.14 (± 0.23) ng/kg bw in the modified lower bound (mLB) approach, ranging from 0.01 to 1.98 ng/kg bw. The risk of AFB 1 was assessed with the margin of exposure (MOE) approach estimated at 2800 in the mean mLB and 1733 in the mean UB. According to the MOE values obtained in this study, aflatoxin B 1 exposure must be considered a public health concern and must be taken as a priority for food risk management., (© 2022. The Author(s) under exclusive licence to Society for Mycotoxin (Research Gesellschaft für Mykotoxinforschung e.V.) and Springer-Verlag GmbH Germany, part of Springer Nature.)

This study's objective was to estimate the seasonal occurrence of aflatoxin M 1 (AFM 1 ) in cow's milk between winter 2016 and winter 2022 and to assess dietary exposure and risk assessment for the adult Croatian population. In total, 5817 cow milk samples were screened for AFM 1 concentrations using the enzyme immunoassay assay (ELISA). For confirmation purposes of AFM 1 concentration above the European Union maximum permitted level (MRL), ultra high-performance liquid chromatography with tandem mass spectrometry was performed. In 94.7% of milk samples, AFM 1 levels were below the detection limit (LOD) of the ELISA test. For 3.47% of samples, the AFM 1 was between the LOD and MRL values. Only 1.87% of all samples exceeded the MRL. The mean value of elevated AFM 1 in different seasons ranged between 59.2 ng/kg (autumn 2017) and 387.8 ng/kg (autumn 2021). The highest incidences of positive AFM 1 were determined in autumn and winter and the maximum (6.4%) was in winter 2019/2020. The largest percentage of positive samples (69.7%) was found in central Croatia. The estimated daily intakes for positive samples ranged between 0.17 and 2.82 ng/kg body weight/day. Risk assessment indicated a high level of concern during autumn and winter, especially for consumers of large amounts of milk.

Due to the climatic change, an increase in aflatoxin B1 (AFB1) maize contamination has been reported in Europe. As an alternative to mineral binders, natural phytogenic compounds are increasingly used to counteract the negative effects of AFB1 in farm animals. In cows, even low dietary AFB1 concentrations may result in the milk excretion of the genotoxic carcinogen metabolite aflatoxin M1 (AFM1). In this study, we tested the ability of dietary turmeric powder (TP), an extract from Curcuma longa (CL) rich in curcumin and curcuminoids, in reducing AFM1 mammary excretion in Holstein-Friesian cows. Both active principles are reported to inhibit AFM1 hepatic synthesis and interact with drug transporters involved in AFB1 absorption and excretion. A crossover design was applied to two groups of cows ( n = 4 each) with a 4-day washout. Animals received a diet contaminated with low AFB1 levels (5 ± 1 µg/kg) for 10 days ± TP supplementation (20 g/head/day). TP treatment had no impact on milk yield, milk composition or somatic cell count. Despite a tendency toward a lower average AFM1 milk content in the last four days of the treatment (below EU limits), no statistically significant differences with the AFB1 group occurred. Since the bioavailability of TP active principles may be a major issue, further investigations with different CL preparations are warranted.