Mycotoxins in Food: Occurrence, Health Implications, and Control Strategies

Toxicon 248 (2024) 108038 Contents lists available at ScienceDirect Toxicon journal homep...

Toxicon 248 (2024) 108038 Contents lists available at ScienceDirect Toxicon journal homepage: www.elsevier.com/locate/toxicon Mycotoxins in food: Occurrence, health implications, and control strategies-A comprehensive review Rahim Khan Department of Food Science, Faculty of Food Science and Technology, Universiti Putra Malaysia, Serdang, Selangor, 43300, Malaysia A R T I C L E I N F O A B S T R A C T Handling editor: Denise Tambourgi Mycotoxins are secondary metabolites produced by various filamentous fungi, including Aspergillus, Fusarium, Penicillium, Alternaria, Claviceps, Mucor, Trichoderma, Trichothecium, Myrothecium, Pyrenophora, and Stachybotrys. Keywords: They can contaminate various plants or animal foods, resulting in a significant loss of nutritional and commercial Mycotoxins value. Several factors contribute to mycotoxin production, such as humidity, temperature, oxygen levels, fungal Health implications species, and substrate. When contaminated food is consumed by animals and humans, mycotoxins are rapidly Preventive measures absorbed, affecting the liver, and causing metabolic disorders. The detrimental effects on humans and animals Absorbents include reduced food intake and milk production, reduced fertility, increased risk of abortion, impaired immune response, and increased occurrence of diseases. Therefore, it is imperative to implement strategies for mycotoxin control, broadly classified as preventing fungal contamination and detoxifying their toxic compounds. This re- view aims to discuss various aspects of mycotoxins, including their occurrence, and risk potential. Additionally, it provides an overview of mycotoxin detoxification strategies, including the use of mycotoxin absorbents, as po- tential techniques to eliminate or mitigate the harmful effects of mycotoxins and masked mycotoxins on human and animal health while preserving the nutritional and commercial value of affected food products. 1. Introduction Penicillium. Currently, only ochratoxin A (OTA) and ochratoxin B (OTB) are relevant in ruminant feeding. OTA is the most common contaminant Mycotoxins are secondary metabolites produced by certain fungi, in cereals and is known for its high toxicity. Once absorbed, they cause including Aspergillus, Fusarium, Penicillium, and Alternaria. These fungi nephrotoxicity, hepatotoxicity, and immunosuppression (Aboagye-- can contaminate various crops in the field, during storage and trans- Nuamah et al., 2021). However, ruminal bacterial metabolism mitigates portation, especially under favorable conditions for fungal growth the toxicity of ochratoxins in ruminants, although they still pose po- (Pickova et al., 2021). Currently, more than 300 distinct mycotoxins are tential issues (Xu, 2023). Similarly, FMNs are a group of mycotoxins recognized, among which only 30 mycotoxins are essential for humans synthesized by species of the Fusarium genus. They are classified into and animals. Of these 30, some of the critical mycotoxins are aflatoxins two series: Series A consists of amides (FMNA1, FMNA2), and series B (AFs), ochratoxins (OTs), fumonisins (FMNs), trichothecenes, patulin consists of amines (FMNB1, FMNB2, FMNB3, and FMNB4) (Tomaszewska (PAT), zearalenone (ZEN) and alternariol (AOH) (Agriopoulou et al., et al., 2022). Fumonisins B1 and B2 are the most common in field con- 2020). AFs typically generated by A. flavus and A. parasiticus. There are ditions and affect the functionality of the liver and kidneys (Gao et al., four distinct types of AF, including AFB1, AFB2, AFG1, and AFG2. 2023). Additionally, these mycotoxins are hydroxylated during digestion and Fusarium species produce ZEN that exhibits estrogenic effects like metabolism processes in dairy cows and are excreted in milk as de- those of female estrogens. Therefore, changes in reproductive function rivatives called AFM1 and AFM2 (Sun et al., 2022). AFs are typically can occur, reducing fertility and increasing the risk of abortions (Rai carcinogenic, hepatotoxic, mutagenic, teratogenic, and immunosup- et al., 2020). Similarly, trichothecenes are mycotoxins generated by pressive agents (Ciegler, 2020; Khan et al., 2024). AFs have been species of the genus Fusarium that affect the health and productivity of extensively researched for their detrimental impacts on humans and animals. Although more than 40 derivatives have been documented, T-2 animals. and DON (or vomitoxin) are the most vital ones. These mycotoxins cause OTs are mycotoxins produced by species of the genus Aspergillus and lesions in the intestinal mucosa and have a significant E-mail address: [email protected]. https://doi.org/10.1016/j.toxicon.2024.108038 Received 6 March 2024; Received in revised form 14 June 2024; Accepted 18 July 2024 Available online 22 July 2024 0041-0101/© 2024 Elsevier Ltd. All rights are reserved, including those for text and data mining, AI training, and similar technologies. R. Khan Toxicon 248 (2024) 108038 immunosuppressive effect (Gab-Allah et al., 2023). Furthermore, the prevalence of these mycotoxins varies globally across regions like existence of mycotoxins in foods not only has detrimental effects on Northern Europe, Central Europe, Southern Europe, Eastern Europe, fungal growth but also deteriorates the nutritional value of raw mate- North America, Central America, South America, Middle East, rials. Additionally, these mycotoxins can disrupt the digestion processes, Sub-Saharan Africa, South Africa, Oceania, South Asia, Southeast Asia, reducing the digestibility and absorption of nutrients. These effects can East Asia, and Central Asia (Gruber-Dorninger et al., 2019). Northern decrease the nutritional value of food and cause additional subtle eco- Europe had the highest contamination levels, with 74.2% of samples nomic losses (McIlwaine et al., 2021). Thus, it is of utmost importance to testing positive for DON, followed by FUM (22.4%), ZEN (28.9%), and closely monitor and regulate mycotoxin contamination in the food ABF1 (5.9%). OTA and T-2 toxins were detected in 8.1% and 30.3% of supply chain to maintain food safety standards and protect public samples, respectively. health. Researchers and regulatory bodies often work together to Central Europe had a high contamination profile, with DON (69.8%), develop guidelines and regulations to control mycotoxin levels in food FUM (43.2%), and ZEN (45%). Southern Europe has a high prevalence of products. Efforts to mitigate mycotoxin contamination require the FUM contamination (74.9%), while Eastern Europe presented moderate implementation of effective agricultural practices, appropriate storage to high levels of DON (59.9%), FUM (33.6%), and ZEN (42.5%). North conditions, and regular monitoring throughout the entire food supply America had the most prevalent contaminants, with DON (64.1%, chain. Furthermore, extensive research into the advancement of crop 505 μg/kg), and FUM (47.7%, 652 μg/kg). Central America was highly cultivars that are resistant to various threats and optimizing storage contaminated with FUM, with 81.1% of samples containing it. Similarly, techniques is crucial to addressing the issue of global food safety. This South America has the highest contamination levels, with FUM (75.3%, review aims to discuss various aspects of mycotoxins, including their 1390 μg/kg) and DON (26.9%, 344 μg/kg) being the most. Sub-Saharan occurrence, risk potential, regulation, and detection techniques. More- Africa showed significant contamination with, ABF1, and FUM, while over, it provides an overview of the detoxification of mycotoxins, spe- South Africa has significant contamination levels for FUM (62.6%, cifically mycotoxin absorbents, as a potential technique to eliminate or 266 μg/kg) and DON (63.2%, 363 μg/kg) (Gruber-Dorninger et al., reduce mycotoxins in food. 2019). Oceania has moderate contamination, specifically for FUM (22.2%) and DON (34.5%). Similarly, South Asia has a significant 2. Geographical distribution of mycotoxins prevalence of ABF1 (82.2%), with FUM and ZEN detected in 69.0% and 45.9% of samples, respectively. Southeast Asia has elevated contami- Mycotoxins are found in various food and raw materials worldwide; nation levels for FUM (69.6%) and ZEN (58.2%). On the other hand, East however, their frequency increases at latitudes between 26◦ and 35◦ Asia exhibits a notable level of DON (84.8%, 418 μg/kg), while Central north and south (Sun et al., 2023). Table 1 summarizes the geographical Asia demonstrates moderate levels of FUM (25.0%, 13 μg/kg) and ZEN distribution of mycotoxins, including AFs, FMNs, ZEN, DON, OTA, and (23.1%, 22 μg/kg) (Gruber-Dorninger et al., 2019). T2 toxins, which are identified as the most prevalent mycotoxins. The Overall, the geographical distribution and prevalence of mycotoxins, Table 1 Geographical distribution of mycotoxins by continents (Gruber-Dorninger et al., 2019). Region ABF1 FUM ZEN DON OTA T-2 Northern Europe (n ¼ 1958) 5.9% 22.4% 28.9% 74.2% 8.1% 30.3% 3.1 μg/kg 186 μg/kg 35 μg/kg 504 μg/kg 19 μg/kg 34 μg/kg Central Europe (n ¼ 21,036) 12.7% 43.2% 45% 69.8% 11.9% 30.7% 1.6 μg/kg 187 μg/kg 40 μg/kg 428 μg/kg 2.8 μg/kg 11 μg/kg Southern Europe (n ¼ 3527) 28.9% 74.9% 36.3% 52.9% 21.2% 11.7% 2.1 μg/kg 607 μg/kg 44 μg/kg 324 μg/kg 2.6 μg/kg 25 μg/kg Eastern Europe (n ¼ 2382) 17.0% 33.6% 42.5% 59.9% 36.4% 48.2% 3.4 μg/kg 87 μg/kg 15 μg/kg 153 μg/kg 3.6 μg/kg 21 μg/kg North America (n ¼ 5471) 10.5% 47.7% 31.7% 64.1% 4.3% 3.9% 8.7 μg/kg 652 μg/kg 102 μg/kg 505 μg/kg 2.4 μg/kg 29 μg/kg Central America (n ¼ 367) 8.6% 81.8% 38.2% 70.0% 3.8% 4.1% 3.9 μg/kg 929 μg/kg 60 μg/kg 316 μg/kg 2.5 μg/kg 3.1 μg/kg South America (n ¼ 17,332) 23.5% 75.3% 46.9% 26.9% 4.9% 21.5% 3.2 μg/kg 1390 μg/kg 51 μg/kg 344 μg/kg 17 μg/kg 31 μg/kg Middle East/North Africa (n ¼ 1075) 22.2% 66.6% 44.8% 47.0% 20.3% 0.5% 2.4 μg/kg 347 μg/kg 31 μg/kg 236 μg/kg 3.1 μg/kg 14 μg/kg Sub-Saharan Africa (n ¼ 208) 76.0% 72.6% 52.2% 49.5% 31.9% 3.0% 23 μg/kg 789 μg/kg 38 μg/kg 352 μg/kg 7.2 μg/kg 3.0 μg/kg South Africa (n ¼ 1077) 9.0% 62.6% 41.6% 63.2% 5.6% 1.2% 2.2 μg/kg 266 μg/kg 30 μg/kg 363 μg/kg 2.2 μg/kg 4.4 μg/kg Oceania (n ¼ 1695) 11.3% 22.2% 19.6% 34.5% 7.5% 2.0% 2.0 μg/kg 106 μg/kg 37 μg/kg 158 μg/kg 3.6 μg/kg 16 μg/kg South Asia (n ¼ 1136) 82.2% 69.0% 45.9% 23.1% 50.4% 0.9% 20 μg/kg 288 μg/kg 43 μg/kg 96 μg/kg 4.6 μg/kg 13 μg/kg Southeast Asia (n ¼ 4310) 57.4% 69.6% 58.2% 42.5% 15.2% 2.7% 10 μg/kg 573 μg/kg 90 μg/kg 137 μg/kg 3.0 μg/kg 26 μg/kg East Asia (n ¼ 13,232) 10.4% 60.7% 23.1% 84.8% 14.1% 11.0% 7.0 μg/kg 810 μg/kg 1.5 μg/kg 418 μg/kg 29 μg/kg 16 μg/kg Central Asia (n ¼ 15) 7.7% 25.0% 23.1% 28.7% 13.3% 6.7% 1.4 μg/kg 13 μg/kg 22 μg/kg 28 μg/kg 22 μg/kg 25 μg/kg 2 R. Khan Toxicon 248 (2024) 108038 including AFs, OTA, FMNs, DON, T-2 toxins, ZEN, and PAT, provide Therefore, they can be found in foods even after cooking, sterilization, significant information on global food safety concerns. Regions like Asia and exposure to high pH levels. The lifespan of these mycotoxins in food and South America demonstrate high positivity rates for DON and FMNs, products is longer than that of the fungi that produced them. Currently, highlighting the urgent need for targeted mycotoxin management stra- more than 300 mycotoxins have been documented, but only 30 of them tegies. Understanding these geographical discrepancies is crucial for pose significant health risks to humans or animals (Table 2) (Casu et al., articulating effective prevention and mitigation strategies, improving 2024; Sdogati et al., 2024). In 1960, London poultry farms suffered agricultural practices, optimizing food storage conditions, and upgrad- substantial losses after consuming imported peanut meal from Brazil, ing detection methodologies to ensure the safety of the global food which led to an outbreak of “turkey X disease.” A year later, researchers supply chain. discovered a connection between poisoning and the presence of A. flavus. They successfully managed to isolate the first AF, which turned 2.1. Occurrence of mycotoxins in food out to be the most carcinogenic substance ever identified. Given their potential and toxicity for humans and animals, mycotoxins can be Mycotoxins are natural compounds synthesized by the secondary harmful when ingested or inhaled. Mycotoxins commonly found in metabolism of fungi, which have toxic impacts on human and animal agricultural products are AFs, OTs, T2/HT-2 toxins, FMNs, and PAT health, even when ingested in low concentrations (El-Sayed et al., (Hoteit et al., 2024). According to the Food and Agriculture Organiza- 2022). Mycotoxins are tiny molecules that exhibit low solubility in tion (FAO), mycotoxins can be found in approximately 25% of the global water, are resistant to degradation by microorganisms, and are highly crop yield (Jallow et al., 2021). The harmful effects of these substances stable to heat (up to 250 ◦ C) (Casu et al., 2024; Khan et al., 2024). are being extensively studied and documented. They are carcinogenic, Table 2 Main mycotoxins and molds found in human and animal food. Mycotoxin Fungi Food Health Effects Standard Limit in Risk References Food Assessment References Aflatoxins B1, B2, A. flavus, Peanuts, Carcinogenic: liver (hepatocarcinoma), 20 ppb (total Codex (Dhakal et al., G1, G2, M1 A. parasiticus, cottonseeds, bile duct, bronchopulmonary, and aflatoxins in nuts and Alimentarius 2023; Khan A. nomius, cereals, fruits, bronchonic cancers. Mutagenic: dried fruits), (FAO/WHO) et al., 2021a) A. pseudotamarii, and spices abnormality in the synthesis of DNA 0.05 ppb (aflatoxin A. ochraceoroseus, repair enzymes, immunosuppressive. M1 in milk) A. bombycis Ochratoxin A, B, C A. ochraceus, Cereals, coffee Carcinogenic: associated with kidney 5 ppb (cereals), 3 ppb EFSA (2006) (Ding et al., A. niger, beans, cancer. (processed cereal 2023; Kőszegi A. carbonarius, vegetables, fish, Mutagenic: abnormality products) and Poór, P. nordicum, and meat in DNA repair enzyme synthesis, 2016) P. viridicatum, immunosuppressive. Nephrotoxic: P. verrucosum endemic and chronic interstitial nephropathies. Zearalenone F. graminearum, Corn, barley, Estrogenic: precocious puberty, 200 ppb (cereal JECFA (WHO) Ropejko and F. sporotrichoides, F. and wheat gynecomastia, reprotoxic. grains), 60 ppb Twarużek culmorum, (infant foods) (2021) F. vertillioides Fumonisins B1, B2 F. moniliforme, Corn, cereals Carcinogenic: associated with 2000 ppb (corn), (EFSA, 2018) Kamle et al. F. vertillioides, oesophageal cancer. Toxic to the liver, 1000 ppb (corn (2022) F. proliferatum kidneys, brain (horses), and lungs (pigs). products for direct human consumption) Trichothecenes Fusarium spp., Corn, wheat Mutagenic: abnormality in the synthesis 1750 ppb (EFSA et al., (Gab-Allah (Type B: DON) F. graminearum, and of DNA repair enzymes. (unprocessed 2018) et al., 2023; F. culmorum Immunodepressant: alters phagocytosis cereals), 750 ppb Payros et al., and inhibits protein synthesis. (cereal products) 2016) Emetogenic neuroendocrine-active lesions in the gastrointestinal tract. Patulin Penicillium, Aspergillus, Apples, peaches, Immunosuppressant: lymphopenia in 50 ppb (fruit juices), JECFA (WHO) Bacha et al. and Byssochlamis species plums, and chronic intoxication. 25 ppb (solid apple (2023) apricots Neurotoxic: nervous disorders (anti- products) acetylcholinesterase action). Genotoxic, gastrointestinal, and kidney lesions Citrinin P. rubrum, P. citrinum, Barley, rice, Genotoxic: causes a genetic disorder. No international EFSA (2012b) (Kamle et al., P. viridicatum, wheat, and rye Nephrotoxic: toxic to kidneys standards: limits vary 2022; Sainz A. ochraceoroseus, by country. et al., 2015) A. flavus, and Monascus ruber Alternaria toxins Alternaria spp., Apples, olives, Genotoxic or teratogenic No international (EFSA et al., Bacha et al. A. alternata, A. lycopersici, and tomatoes standards; under 2016a,b) (2023) and A. tenuissima study Ergot alkaloids C. purpurea, and Cereals, rye Ergotism (gangrene) and neurological 1000 ppb (rye and EFSA (2012b) (Borkar, 2023; C. fusiformis symptoms rye products), Doi et al., 500 ppb (other 2023) cereals) NIV (type B Fusarium spp. Barley, wheat, Immunotoxic, and hematotoxic Under study EFSA (2012a) Gab-Allah trichothecene) oats et al. (2023) T-2/HT-2 toxins F. langsethiae, and Wheat, oats, rye, Hematotoxic, immunotoxic, and cytotoxic 100 ppb (cereal EFSA (2011) Meneely et al. (type A F. sporotrichoides and barley grains) (2023) trichothecene) 3 R. Khan Toxicon 248 (2024) 108038 teratogenic, tremorgenic, diabetogenic, hepatotoxic, nephrotoxic, an in-depth approach and rigorous regulatory frameworks to ensure hematotoxic, mutagenic, immunotoxic, allergic, neurotoxic, immuno- public health safety while maximizing its industrial potential. suppressive, and necrotizing (Table 2). In summary, mycotoxins, produced by fungi, pose a global health 2.3. Colonization of foods concern due to their widespread distribution and potent toxicity. These toxins persist in food even after processing methods such as cooking and In an uncontrolled environment, fungal contamination is a constant sterilization due to their chemical stability and resistance to degrada- and inevitable risk. The incidence of fungal growth in crops and the tion. A detailed analysis of the most common mycotoxins, such as AFs, production of mycotoxins is influenced by various environmental fac- OTA, FMNs, DON, T-2 toxins, and ZEN, highlights their potential health tors, including the general health of the crop before harvesting, weather hazards that range from carcinogenic to mutagenic, immunosuppres- conditions (rainfall and atmospheric turbulence), harvesting methods, sion, and reproductive toxicity. However, improving agricultural and timing, and hydrothermal conditions before stabilization for proper food processing techniques can effectively mitigate the risk of myco- preservation (Awuchi et al., 2021). toxins and protect public health. Animals, mainly insects and mites, are secondary factors in the dispersal of conidia. The fungus depends on the surrounding organic 2.2. Mycotoxin-producing fungi materials and their structural properties to grow and reproduce (Fig. 1). Fungi contain a high concentration of depolymerase within their cells, Fungi are eukaryotic microorganisms that typically have a filamen- allowing them to break down complex polymers, such as cellulose, tous body. They can be saprophytic, parasitic, or heterotrophic, lignin, and pectic compounds (Zanne et al., 2020). Similarly, the sap- depending on the source of organic carbon. The fungal colony consists of rophytism of these fungi is based on the availability of free water in their a complex network of hyphae known as mycelium (Ugalde and surroundings. Fungi can colonize a wide variety of foods due to their Rodriguez-Urra, 2014). They produce many spores, providing them with varied enzymatic arsenal, tolerance to acid pH levels, minimal water and considerable contamination capabilities. They can be present in various oxygen requirements, and the ability to thrive at temperatures of environments such as soil (primary habitat), air, water, and various 3–40 ◦ C (Pitt and Hocking, 2022). Due to the growth pattern of the foods, such as cereal (corn, rice, wheat, barley), lentils, fruits, peanuts, hyphae, the fungal colony does not remain at the contaminated site as do almonds, walnuts, pistachios, coffee, cotton seeds, spices (pepper, the bacterial colonies. Hyphae continually progress through the sub- paprika, ginger) and meat (Tolosa et al., 2021). Fungi can be beneficial strate, obtaining new sources of nutrients and occupying the entire in industries such as fermentation, pharmaceuticals, agri-food, and edible surface with the help of the lateral mycelial hyphae. biotechnology. They play a vital role in the transformation of raw food materials and in the production of antibiotics, enzymes, condiments, 2.4. Factors affecting mycotoxin production flavoring agents, and proteins that have potential benefits for human health. 2.4.1. Intrinsic factors However, a strain used by the food industry may not always be The nature and quality of mycotoxins are influenced by the specific nontoxic and can potentially become toxic in specific environments. For species that produce them. They vary according to their genetic makeup example, Aspergillus oryzae is a fungal strain commonly used in the and the surrounding environment (Eltariki et al., 2018; Perrone et al., soybean fermentation process to make soy sauce and miso, a classic 2020). Within the same species known to be toxigenic, not all strains Japanese seasoning. In high humidity and temperature settings, exhibit this characteristic. The prevalence of toxin-producing strains A. oryzae can produce mycotoxins such as AFs due to genetic mutations depends on the fungal species, the geographical location, and the nature or contamination with toxin-producing organisms (Xu et al., 2022). On of the substrate (Greeff-Laubscher et al., 2020). Several species from the other hand, harmful fungi can cause food spoilage and the produc- diverse genera can produce certain mycotoxins. The presence of a tion of toxic metabolites (mycotoxins), mycoses, and allergens such as toxigenic fungus at a given time is necessary to produce mycotoxins. Alternaria alternata, which produce spores that can cause allergic re- However, the occurrence of toxic fungi does not always imply the ex- actions. Additionally, A. fumigatus species can contaminate foods and istence of mycotoxins, and the absence of fungus does not necessarily produce allergens, causing respiratory problems. On the other hand, mean the nonexistence of mycotoxins. Cadosporium species are common airborne allergens associated with food spoilage. Fusarium species are known for their production of my- 2.4.2. Extrinsic factors (ecological conditions) cotoxins and allergenic properties, contaminating cereals and crops, and They are physical, physicochemical, and chemical factors. Temper- leading to allergic reactions when consumed (Al Hallak et al., 2023). ature, pH, humidity, and pests are the main factors that contribute to There are two types of fungi, including hygrophilic and xerophilic biodeterioration, leading to multiple biochemical and organoleptic fungi. Hydrophilic or field fungi thrive in environments with high water activity. Fusarium and Alternaria species are good examples of hydro- philic fungi that contaminate agricultural products before and during harvest. Recently, data revealed that Pyrenophora tritici-repentis (the causative agent of the wheat halo spot) thrives in moist conditions and can produce mycotoxins (Wei, 2020). On the contrary, the Aspergillus and Penicillium species are the most formidable storage or xerotolerant fungi. Moreover, there is a third type, which is quite rare. It belongs to xerophilic fungi and develops when the grain has a lower moisture content. It should be mentioned that several researchers have reported the production of mycotoxins by specific fungal genera, including Aspergillus, Fusarium, Penicillium, Alternaria, Claviceps, Mucor, Tricho- derma, Trichothecium, Myrothecium, Mortierella, Pyrenophora, and Sta- chybotrys (Hoteit et al., 2024; Khan et al., 2021a; Khan et al., 2020; Rovetto et al., 2023; Tribelhorn et al., 2023). Overall, mycotoxin-producing fungi are a complex group of organ- isms with significant implications for human health, agriculture, and Fig. 1. Colonization of food. industry. Optimizing the balance between its benefits and risks requires (Source: self-drawn by author) 4 R. Khan Toxicon 248 (2024) 108038 modifications in an ecosystem (Fig. 2) (Daou et al., 2021). dehydrated or poorly hydrated foods, and rehydrated protected powder milk prevent fungal spoilage. 2.4.2.1. Temperature. Fungi are typically mesophilic and thrive under moderate temperature conditions (Daou et al., 2021). Their hyphal 2.4.2.5. Substrate. The composition of the substrate can influence the growth is optimal at 20–25 ◦ C, while it tends to be slower at 5–35 ◦ C. The expression of mycotoxin secretion function (Kumar et al., 2017). An conidia of mesophilic species remain dormant for an extended period at elevated concentration of sugars or lipids promotes the production of − 20 ◦ C (Bakar et al., 2020). Some toxigenic fungi, such as P. expansum, mycotoxins. Cereals and oilseeds, which are rich in sugar and lipids, P. viridicatum, and P. verrucosum, are psychrotrophic and thrive in cold tend to be more susceptible to mycotoxin contamination compared to temperatures. They can grow slowly at temperatures below 4 ◦ C. These substrates with a high protein content. The production of AF by A. flavus fungi are often responsible for the spoilage of refrigerated foods. Fresh is favored by specific sugars such as glucose, mannose, fructose, and meat carcasses stored for an extended period in a cold room can develop sucrose (Tiwari et al., 2023). Iron, zinc, and copper have been tested for polychrome fungal colonies, among which Cladosporium herbarum is the their potential to produce AFs and OTA. All promote the production of most prevalent species (Daou et al., 2021). AFs and OTA at concentrations below 10 mg/L of medium, and zinc is the most influential in terms of development and AF production. The 2.4.2.2. Food pH. Concerning pH, fungi are more tolerant than bacte- impact of iron and copper may be due to their function as catalysts of ria. Bacteria generally thrive in pH ranges of 7–8, while fungi usually lipid peroxidation (Latunde-Dada, 2017). Recent developments in grow at pHs between 3 and 8, with optimal growth occurring at pH chemotaxonomy have enabled the demonstration of distinct substance levels between 5 and 6. Due to their acidity (pH < 6), certain foods, such profiles among fungi collected from the natural substrate or culture in a as vegetables, fruits, and meat, are much more susceptible to fungal Petri dish. An intriguing example involves Penicillium roqueforti, which is damage than bacterial damage. incapable of producing a toxin (roquefortine) on Roquefort cheese, while in vitro it can produce a highly toxic metabolite. However, the 2.4.2.3. Oxygen pressure. All fungi require oxygen to grow optimally. metabolic profile is different if a complex (natural) or defined (chemical) However, for many of them, development is minimally affected by culture medium is used (Al-Saleem et al., 2022). contents that are ten times lower (2.1%) than those in the atmosphere. Byssochlamys fulva and B. nivea can even grow with 0.27% oxygen. 2.5. Biological factors Foods packaged under low oxygen pressure are not safe from fungus contamination, making them potentially unsafe. 2.5.1. Predators Insects and mites are indirectly involved in mycotoxin production by 2.4.2.4. Water accessibility. The high moisture content promotes the being vectors of fungus spores, allowing them to penetrate seeds through growth of fungi. To sustain its growth, the mycelium needs free (readily the wounds they produce. Therefore, the contamination of peanuts, accessible) water. Fungi use the water vapors present in the spaces be- corn, and cottonseeds by A. flavus and AF before harvest is often linked tween grains, which is influenced by the balance between the water to insect infestations. Grains that harbor weevils during storage typically content of the grain (the water content of the grain) and the water in the show a significant fungal population and mycotoxins (Mohapatra et al., form of vapor surrounding it (Cendoya et al., 2018; Pietsch, 2020). The 2022). Birds and rodents exhibit similar behavior on unprotected grain concentration of pore water is known as the water activity (aw). In the reserves (Awuchi et al., 2021). absence of free water, the diffusion of fungal exoenzymes in the envi- ronment to the substrate is hindered. However, after the depolymer- 2.5.2. Interactions among microorganisms ization of the substrate, simple molecules can be diffused inside the The presence of microbes, including fungi, and bacteria modulate the fungal cell. This parameter can fluctuate between 0 and 1 depending on production of mycotoxins; there is competition between different fungi the water retention or the availability of the substrate. The characteristic (Bartholomew et al., 2021). For example, the presence of A. niger in the values of this factor that allow fungus development are between 0.70 same environment inhibits AF production by A. flavus (Khan et al., and 0.99 (Ezrari et al., 2021). aw below 0.60 does not allow fungi to 2021b, 2021c). The presence of multiple fungal species in the same grow, it will not harm conidia. Theoretically, chocolate, spices, commodity inhibits mycotoxin production. The explanation for this lies Fig. 2. Parameters involved in mycotoxin production. (Source: self-drawn by author). 5 R. Khan Toxicon 248 (2024) 108038 in the competition for the substrate, where certain strains can degrade toxicological assessment and risk management of carcinogenic sub- the toxin produced. stances are crucial to understanding and mitigating their risks (Boobis et al., 2016). Boobis et al. (2016) argued for a multifaceted approach to 2.6. Chemical factors assess the risk of carcinogens, focusing on the identification and quan- tification of these substances along with their molecular and cellular The use of insecticides can effectively reduce mycotoxin production, modes of action. Exploring the interaction between carcinogens and either by direct antifungal action on the fungus or by preventing seeds biological systems can provide valuable insights for developing risk from insect and mite-induced lesions. However, caution should be management strategies. Key aspects of such assessment would involve exercised when using these substances. Some studies have shown that in-depth studies, understanding dose-response relationships, analyzing mycotoxin production is stimulated at sublethal concentrations (Jafar- epidemiological data, and considering regulatory policies and public zadeh et al., 2023). Experiments carried out on grapevine organisms health strategies. have revealed that active compounds, including fludioxonil and cypro- dinil, can effectively hinder the dissemination of Aspergillus and Peni- 3.1. Aflatoxins cillium species. Similarly, fosetyl-aluminum and folpet are antifungals that effectively minimize the OTA in wines (Marín et al., 2008). A recent AFs are mycotoxins produced by Aspergillus species and are study has found that the use of a tridemorph systemic fungicide at commonly found in warm and humid environments. They pose a sig- 6–8 ppm effectively promotes the growth of Fusarium sporotrichoides nificant threat to human health, particularly in regions in Africa and while inhibiting T-2 toxin production (Tleuova et al., 2020). Alterna- Southeast Asia, where they lead to high incidences of liver cancer and tively, concentrations of 30–50 ppm of tridemorph systemic fungicide acute poisoning. AFB1, AFG1, and AFM1 are highly toxic to humans and strongly inhibit fungal growth and considerably enhance mycotoxin animals, excreted through milk and urine (Khan et al., 2021a; Wogan production (Gavahian et al., 2020). Black Aspergilli are stressed when et al., 2012). AFM1 is approximately ten times less potent than AFB1 can exposed to specific concentrations of fungicide, producing more OTA. undergo bioactivation like AFB1 and is carcinogenic in animal experi- Thus, mycotoxin production can serve as a defense mechanism for fungi ments. The IARC classified AFM1 as group 1, indicating its potential to against stress agents (Perincherry et al., 2019). Similarly, citrinin serves cause cancer in humans. The commonly prevalent AF contaminant AFB1 as a sunscreen agent for Penicillium verrucosum (Salah et al., 2019). It is undergoes metabolism after ingestion, leading to the production of crucial to study the influence of other factors, such as climate and cul- highly reactive 8,9-epoxide that bonds covalently to DNA and causes ture, to understand the effects of these fungicides. Furthermore, it gene mutations in the TP53 tumor suppressor gene. However, gluta- should be mentioned that the Aspergillus carbonarius species is a highly thione S-transferases and protein binding can detoxify AFB1 epoxide. aggressive fungus (Linde et al., 2016) that infects healthy grape berries Several metabolites, including serum albumin adduct, AF-N7-guanine and produces OTA without external influences. adduct, and AFM1, serve as reliable biomarkers in epidemiological In general, mycotoxin production is influenced by various factors, studies, defining the relationship between exposure and the risk of including the genetic makeup of the fungus, environmental conditions, illness and demonstrating the effectiveness of strategies to reduce it. and competing microorganisms and pests. Chemical interventions such Studies on AF-induced hepatocellular carcinoma have also revealed the as fungicides can suppress or promote mycotoxin production depending synergistic impacts of hepatitis B virus (HBV) disease (Gramantieri et al., on their application and the fungal response. Similarly, geographical 2022; Ismail et al., 2021). Early exposure, particularly in developing location and specific conditions can affect the prevalence of mycotoxin- countries, is crucial for better education on the hazards of AFs and producing strains. Understanding these interactions is crucial to devel- effective intervention strategies to mitigate these risk factors and pre- oping effective strategies to control and minimize mycotoxin contami- vent the onset of acute and chronic diseases. Furthermore, the reference nation in food and feed products. values for AFB1 are critical for risk assessment and regulatory purposes, often determined by international bodies such as the World Health Or- 3. Most prevalent mycotoxins ganization (WHO) and the Food and Agriculture Organization (FAO). Some of the critical mycotoxins present in food are produced by 3.2. Ochratoxin A Aspergillus, Fusarium, Penicillium, Alternaria, and Claviceps (Frisvad et al., 2006). Mycotoxins can be classified according to the fungi that produce OTA is a common contaminant found in food worldwide, particularly them or their chemical structure. From a toxicological perspective, the in grains and their derived products, including coffee, cocoa, grapes, and focus lies on the pathogenic effects observed in animals, specifically wine (Mejri-Omrani et al., 2016). It is also present in animal foods, such potential hazards, particularly carcinogenicity, nephrotoxicity, muta- as milk, cheese, and sausage from foodstuffs containing OTA. Regula- genicity, hepatotoxicity, neurotoxicity, and immunotoxicity. Table 2 tions have been established on the maximum levels of OTA in food due provides a list of mycotoxins produced by toxigenic fungi. Here are a few to its toxicity concerns (Bui-Klimke and Wu, 2015; Obafemi et al., 2023). points to consider: OTA is known for its potent nephrotoxic effect and documented carci- nogenic potential in studies with rodents. IARC categorized OTA as 1) Fungi can produce multiple mycotoxins; for example, Fusarium Group 2B, suggesting its potential link to cancer in humans. However, strains often produce ZEN along with DON. reliable data on its role in human tumors are not available. The putative 2) Different fungi can produce the same mycotoxins; for example, OTA link between Balkan endemic nephropathy (BEN) and exposure to my- can be synthesized by strains of Aspergillus and Penicillium. cotoxins (OTA and CIT) is currently being investigated due to the con- 3) Complex exposures can occur; however, at low levels, interactions trasting pathologies of BEN and OTA-induced rat kidney tumors between multiple contaminants are of secondary importance. (Mantle, 2016; Zhang et al., 2023). However, recent molecular epide- miological studies provide strong evidence supporting the significant Some of the mycotoxins listed in Table 2 have carcinogenic and contribution of aristolochic acid to BEN development (Stiborová et al., mutagenic potential. Based on results from animal experiments and 2016). epidemiological analysis, the IARC (International Agency for Research The toxicokinetics, toxicodynamics, and mechanism of action of OTA on Cancer) categorizes carcinogenic substances into five groups, helping have been extensively examined (Tao et al., 2018). Key insights are to determine their potential impact on humans (Ostry et al., 2017). For summarized including the kidneys being the main organ affected by the example, mycotoxins assessed by IARC were previously grouped. Such toxic effects of OTA. In pigs, a ’lowest observable effect level’ (LOEL) of classifications based on "hazard" are often subject to debate. A thorough 8 μg/kg bw/day was observed for the first markers of nephrotoxic 6 R. Khan Toxicon 248 (2024) 108038 effects. This LOEL serves as a reference point to determine the TDI of than those causing health concerns. OTA for humans (Tao et al., 2018). OTA is partially hydrolyzed in the gastrointestinal tract to produce the non-toxic metabolite OT-alpha 3.5. Citrinin (OTα), which is more efficient in ruminants than in monogastric spe- cies such as pigs or humans. After absorption, small amounts of hy- Citrinin (CIT) is a metabolite produced by various species of fungi, droxylated metabolites (4-HO-OTA, 10-OH-OTA) and conjugates including Penicillium, Aspergillus, and Monascus (Ajithkumar et al., (glucuronides of OTA and OTα) are also produced. OTA itself causes 2024). It is commonly found as an impurity in fermented products such toxic effects and can accumulate in target tissues, such as the kidneys, as red yeast rice. It is nephrotoxic in various species, including swine, and be excreted at different rates depending on the species(Ciarcia et al., leading to Balkan nephropathy (Flajs and Peraica, 2009). The IARC has 2016; Hashimoto et al., 2016). Despite its potential to cause genotox- classified CIT into Group 3, indicating that it cannot be classified as icity, studies have shown that OTA can cause aneuploid and clastogenic carcinogenic for humans. Similarly, the European Food Safety Authority effects without inducing mutations in bacterial and mammalian cells (EFSA) (2012b) does not find conclusive evidence of the carcinogenic (Föllmann et al., 2014). There is a lack of reliable information on the potential of CIT but highlights the lack of a classic long-term study in bioactivation of DNA-reactive metabolites for OTA, so a practical rodents. Multiple studies have shown the genotoxic effects of CIT, threshold for genotoxic effects can also be considered (Boobis et al., including the aneuploidogenic effect and chromosomal aberrations, 2016). without causing mutations in bacteria and mammalian cells (Föllmann et al., 2014). Mixtures of OTA and CIT show additive effects under in 3.3. Fumonisins vitro conditions when combined at their effective concentrations. Research involving multiple animal species has shown that repeated FMNs are polar mycotoxins produced by F. verticillioides and administration of CIT exhibited a nephrotoxic effect and suggested F. proliferatum found in corn and its derivative products. FB1 is the most variations in their susceptibility (De Oliveira Filho et al., 2017). How- critical contaminant due to its prevalence and toxicity (Hove et al., ever, it remains unclear whether species variations in the kinetics have 2016). Chronic exposure to FB1 has been shown to have carcinogenic an impact in this case, as data on the absorption, distribution, meta- effects on the kidneys and liver in rodents and is considered a potential bolism, and excretion (ADME) of CIT are scarce. Rats excrete primarily cancer risk factor for humans (Ostry et al., 2017). FMN also promotes unchanged CIT, along with a small amount of dihydrocitrinone tumors due to genetic factors. The FMN-contaminated feed can cause (DH-CIT), a metabolite with significantly lower in vitro toxicity toxic effects in livestock, such as horses that develop massive leu- (Föllmann et al., 2014). In contrast, human urine samples show much koencephalopathy and pigs developing pulmonary edema. higher concentrations of DH-CIT metabolite compared to unmetabolized Dose-dependent damage to the liver has been observed in various spe- CIT (Ali et al., 2015), suggesting effective detoxification of CIT than cies, with the liver and kidneys being the primary organs affected (Szabó rodents. EFSA has determined a non-toxic dose of 20 μg/kg bw/day for et al., 2018). Exposure to FB1 in pregnant mice leads to neural tube humans for CIT (de Oliveira Filho et al., 2017). defects in the embryos (Riley et al., 2015). Similarly, prolonged disruption of sphingolipid and glycerolipid metabolism can alter cellular 3.6. Ergot alkaloids signaling pathways crucial for normal cell function. Fluctuations in the sphinganine/sphingosine ratio are currently used as a biochemical A fungus Claviceps purpurea can infect cereals, particularly rye, and marker for FMN exposure in humans (Al-Jaal et al., 2019). FB1 mea- produce sclerotia, also known as ergot. These alkaloids, including surement in urine is not sensitive, as only a tiny portion of the ingested ergotamine, ergometrine, ergosine, α- and β-ergocryptine, ergocrystine, dose (0.5%) is eliminated in the kidneys. Studies have shown that FB1 and ergocornine, can cause severe poisoning (ergotism) when ground absorption is rapid after oral administration and bioavailability is into flour (Awuchi et al., 2021). Two forms, gangrenosum ergotis, and exceedingly low (Al-Jaal et al., 2019; Riley et al., 2015). FB1 is excreted convulsivus ergotis are related to the composition of active ergot alka- primarily in its original form through urine. Considering this further loids. In the EU, a limit below 0.05% ergot in grains is widely accepted research is needed to understand the mechanism of action and potential and modern milling technology is used to effectively remove sclerotia. combined effects of FB1 and AFB1, which are frequently found together Ergot alkaloids and derivatives have significant applications in medi- in corn in Africa. cine, such as treating migraines and postnatal uterine contractions (Weaver et al., 2021). EFSA established a group value for ergot alka- 3.4. Alternaria toxins loids, stating a tolerable daily intake (TDI) of 0.6 μg/kg bw/day and a group acute reference dose (ARfD) of 1 μg/kg bw per meal occasion Alternaria, a common fungus genus, infects various products, (EFSA, 2012a). However, the EFSA acknowledges uncertainties including cereals, apples, citrus fruits, olives, potatoes, tomatoes, and regarding exposure due to the potential contributions of food groups not sunflower seeds (Pinto and Patriarca, 2017). Alternaria produces included in the analysis. potentially hazardous substances, such as alternariol (AOH), alternariol monomethylether (AME), tentoxin (TEN), altenuen (ALT), altertoxin I 3.7. Patulin (ATX-I), altertoxin II (ATX-II), altenuisol (ATL), isoaltenuen (isoALT), and tenuazonic acid (TeA). The toxicity of Alternia toxins has not been PAT is a mycotoxin produced by P. expansum, a fungus that primarily adequately characterized (Hessel-Pras et al., 2019). ALT and TeA are affects fruits like apples, and pears (Mahato et al., 2021a,b; Zheng et al., more acutely toxic than AOH and AME, which are easily absorbed. 2021). It is found in various products, including compotes, juices, grains, Previous rodent studies indicated its potential fetotoxic and teratogenic and vegetables (Egmond et al., 2007). PAT is classified in group 3 by the effects but at doses higher than expected human exposure (Meena et al., IARC due to its genotoxic mechanism (Saleh and Goktepe, 2019; Vidal 2017). AOH and AME exhibit genotoxic effects in bacteria and et al., 2019). PAT reacts with sulfhydryl groups, inhibiting enzymes such mammalian cells, but their mutagenic potential is less pronounced than as membrane-bound ATPases, leading to oxidative stress and a geno- that of ATX-I and ATX-II. However, there are insufficient data to toxic effect. However, no carcinogenic effect was found after oral or determine the cancer-causing properties of Alternaria toxins (Hessel-Pras intraperitoneal administration. High doses of PAT can cause mucous et al., 2019). Recent studies have revealed the presence of Alternaria membrane irritation, nausea, vomiting, diarrhea, bleeding, and neuro- toxins in several foods with concentrations in the 1–8700 ng/kg range toxic symptoms in animals. Furthermore, PAT influences immune sys- (Hickert et al., 2016). However, the potential toxicological relevance of tem parameters and has a reprotoxic potential in rodents. The Joint these exposures is uncertain, as the calculated values are often lower Expert Committee on Food Additives (JECFA) has established a 7 R. Khan Toxicon 248 (2024) 108038 reference point (NOAEL) of 43 μg/kg bw for PAT to ensure safe intake. pathogen that infects grain in the field and causes significant damage to Similarly, a PMTDI of 0.4 μg/kg bw/day was determined for PAT (de crops, such as corn and wheat. ZEN is present primarily in corn and corn Melo et al., 2012). Taking this into account, maximum limits have been products but has also been found in other grains such as barley, oats, established for the presence of PAT in fruit juices and similar products, millet, rye, wheat, and soybeans (Mahato et al., 2021a,b). The levels of with the lowest levels (10 μg/L) applied for infant food. ZEN determined are well below those of DON, a contaminant that is much more common in cereals. ZEN differs significantly from other 3.8. Trichothecenes Fusarium metabolites due to its extremely low acute toxicity. It is commonly known as mycoestrogen due to its potent hormonal (estro- Trichothecenes are cyclic sesquiterpenes with an epoxide ring, pro- gen-like) effect. After ingestion, ZEN is efficiently absorbed and con- duced by fungi of various genera, including Fusarium, Trichoderma, verted into multiple metabolites within the body (Mahato et al., 2021a, Stachybotrys, and Trichothecia (Laraba et al., 2021; Wang et al., 2023a, b). Specific metabolites of ZEN are produced in small amounts through b). These toxins are potent inhibitors of protein biosynthesis, causing the catalytic action of cytochrome P450 enzymes, which involve the damage to cells and attacking the gastrointestinal tract after ingestion hydroxylation of the aromatic ring. On the contrary, the dominant (Gil-Serna et al., 2020; Zhu et al., 2020). They affect blood formation, process involves the conversion of 3α-/3β-steroid dehydrogenases into weaken the immune system, and trigger undesirable neuronal effects α-zearalenol (α-ZEL) and β-zearalenol (β-ZEL). The EFSA has confirmed (Janik et al., 2021). However, trichothecenes are not considered muta- the earlier TDI for ZEN, but it is currently used as a group value for total genic or carcinogenic. ZEN and its altered forms (EFSA et al., 2016a,b). Furthermore, concerns about the precision of the exposure assessment highlight the importance 3.8.1. Deoxynivalenol and nivalenol of further food analysis. Human biomonitoring can serve as a comple- DON is a mycotoxin produced by F. graminearum and F. culmorum mentary approach to determining exposure when converting urine and is widely distributed in cereals such as corn, barley, wheat, rye, oats, values for ZEN and its metabolites into intake amounts. However, the and their derivatives (Mishra et al., 2022; Palumbo et al., 2020). It is the database on kinetics in humans and renal excretion rates must be main contaminant of food, along with its acetyl derivatives (3-ADON, improved (Fromme et al., 2016; Mally et al., 2016). 15-ADON), glucoside conjugates, and nivalenol. Its impacts on the gut, In short, mycotoxins, produced by different fungal species, pose the immune system, and neuroendocrine processes vary depending on substantial risks to human and animal health because of their wide range the time frame and level of exposure (Payros et al., 2016). Consistent of toxicological impacts, such as carcinogenicity, neurotoxicity, and consumption of smaller amounts of DON can cause health problems, immunotoxicity. Key mycotoxins, including AFs, OTA, FMNs, and including lesions in the digestive tract, changes in the immune system, trichothecenes, are prevalent in various foods, particularly grains, and stunted growth of pigs (Alizadeh et al., 2015). Pigs are recognized as highlighting the importance of implementing strict regulatory standards a sensitive species due to the higher bioavailability of DON in their and continuous monitoring to control their levels. Advanced research feeding (Palumbo et al., 2020). In humans, the conversion of DON in and comprehensive risk management strategies are essential to effec- DOM-1 could be more efficient. When consumed, it is transformed into tively reduce the risks of mycotoxin exposure. Furthermore, public DON-3-GlcA and DON-15-GlcA by glucuronidation and excreted awareness and global cooperation plat a crucial role in efficiently through the kidneys. The provisional maximum tolerable daily intake managing and minimizing the adverse health effects associated with (PMTDI) for DON is 1 μg/kg bw has been set by the EFSA. Human urine mycotoxins. analyses show lower concentrations of NIV compared to DON in pigs likely due to efficient metabolism. NIV concentrations in urine reflect 3.10. Masked mycotoxins the combined intake of both contaminants (Fromme et al., 2016). Given safety measures and precautions, its unlikely synergistic effects would In addition to native mycotoxins (DON, ZEN, or FMNs), food can also occur even in cases where DON and NIV coexist, considering the contain masked mycotoxins. These compounds are not detected by circumstances. standard analytical methods, often leading to an underestimation of the overall toxin content in a sample. Masked mycotoxins are synthesized in 3.8.2. T-2 and HT-2 toxins plants via conjugation reactions or chemical processes during food T-2 and HT-2 toxins are produced by Fusarium species such as production (Zhang et al., 2020a,b). Plants protect themselves against F. sporotrichoides and F. langsethiae and are commonly found in cereals xenobiotics, including mycotoxins, through a series of metabolic such as oats, rye, barley, and wheat (Foroud et al., 2019; Gab-Allah detoxification steps, including enzymatic conjugation and compart- et al., 2023). They are more cytotoxic than type B trichothecenes, mentalization, which are particularly important for the inactivation of including DON or NIV, and can harm the epithelium of the gastroin- mycotoxins. During enzymatic conjugation, molecules such as glucose, testinal tract and skin. Additionally, they can systemically alter blood glutathione, or malonic acid are attached to the toxin, resulting in formation due to their hemato and myelotoxicity (Gab-Allah et al., compounds of higher polarity. These conjugates typically exhibit 2023). Prolonged contact with T-2/HT-2 toxins increases susceptibility reduced toxicity compared to the original compound (Berthiller et al., to infections due to their immunosuppressive effect. The T-2 toxin un- 2009). Subsequently, these conjugates are either stored in the vacuole in dergoes rapid conversion into different metabolites, especially HT-2 and a soluble form or bound to cell wall constituents in an insoluble form. T-2 triol, which constitute approximately 20% of the dose found in the This leads to the classification of masked mycotoxins into soluble and urine of exposed pigs (Fromme et al., 2016). Data on toxicology in insoluble (bound) forms. humans are scarce; however, the pattern (HT-2 and T2 triol) is similar. Plant metabolites have been identified and characterized for several In pigs, immunological and hematological effects are particularly sig- mycotoxins, including DON, NIV, T-2 toxin, HT-2 toxin, ZON, OTA, and nificant. Based on these data, a TDI of 20 ng/kg bw/day can be deter- fusaric acid (Berthiller et al., 2013). For example, Lijalem et al. (2023) mined for humans using standard extrapolation factors for grouped detected 17 different metabolites of ZON in the model plant Arabidopsis T-2/HT-2 toxins (Gottschalk et al., 2009). thaliana (A. thaliana), including glucosides, diglucosides, and malonyl glucosides. 3.9. Zearalenone In food production, processes such as malting or heating can lead to the formation of masked mycotoxins. For example, thermal treatment of Zearalenone (ZEN), a hydroxylated resorcylic acid lactone, is pro- FB1 in the presence of reducing sugar results in the formation of N-(1- duced by various strains of the Fusarium genus, including F. culmorum deoxy-D-fructose-1-yl)-FB1 (Zachariasova et al., 2012). Additionally, and F. graminearum (Frisvad et al., 2006). Fusarium is a common enzymatic degradation of polysaccharides during food processing can 8 R. Khan Toxicon 248 (2024) 108038 trigger the release of masked mycotoxins (Zachariasova et al., 2012). and the effect is more pronounced (reduction between 11% and 43%) The potential health hazards posed by masked mycotoxins are an when gamma radiation is used (Zhang et al., 2020a,b). Alternatively, emerging area of mycotoxin research. Generally, there are two potential treatments with ultraviolet (UV) rays reduced the PAT content by 5%– pathways for masked mycotoxins to cause harm when ingested through 73% (Visuthiwan and Assatarakul, 2021). Photodegradation from sun feed or food. First, these toxins possibly be broken down in the digestive exposure also reduces mycotoxin levels by 40% in 3 h and 75% in 30 h. tract, leading to the release and absorption of the native mycotoxin, thus Furthermore, sunlight is more effective than other types of radiation increasing the overall toxin load. Secondly, masked mycotoxins could (Kasun and Vanniarachchy, 2023). have inherent toxic effects, which might differ from those of the native In general, physical treatments, including thermal methods, ionizing mycotoxin. radiation, cold temperatures, wave treatments, and exposure to sun- light, can help reduce mycotoxin contamination. However, their effec- 4. Control strategies for mycotoxins tiveness may vary depending on the specific mycotoxin being targeted. Extremely low temperatures prevent fungal growth but do not entirely Mycotoxin contamination prevention strategies can be pre-harvest or sterilize. Additionally, wave treatments, such as microwaves, gamma post-harvest measures. Preventive measures during the harvesting pro- radiation, UV rays, and sunlight exposure, show promise in reducing cess include ensuring timely harvesting, avoiding harvesting with excess mycotoxin levels. However, these physical interventions should be moisture, and, if necessary, drying crops before storage (Nada et al., evaluated for their effects on the nutritional value of the treated 2022). Today, the agri-food industry strives for a high standard of products. quality and safety, using modern technology such as filtration, air sterilization, overpressure sectors, and disinfection of atmospheres and 4.2. Chemical treatments surfaces to produce food under aseptic conditions (Fumagalli et al., 2021). Currently, in agriculture, preventing fungal contamination is a Chemical treatments provide a variable alternative to effectively challenging task. However, it is essential to avoid superinfection of seeds eliminate mycotoxins, with ammonization being a widely studied and and fruits through contact with soil and contaminated equipment (Greff proven effective method against aflatoxin B1 when combined with high et al., 2023). Additionally, these contacts cause injuries, which facilitate temperatures and pressure (Doğan and Hayırlı, 2022). However, its the penetration of hyphae into the plant. When handling fungal effectiveness is limited to certain mycotoxins. Despite the technical ef- contamination, it is imperative to prevent conidial germination and ficiency of the process, it is costly. During processing, there is a sub- hyphal development. When mycotoxins are already present in food, it is stantial decline in the nutritional content of the food, resulting in the necessary to implement control strategies to minimize their adverse emergence of chemical residues that restrict their use in field conditions effects. Given the complexity of the diagnosis, it is helpful to know the (Liu et al., 2022a,b). Other detoxification methods for food contami- incidence based on geographical area, climatic conditions, and potential nated with mycotoxins include oxidizing agents such as ozone, hydrogen food sources. These factors can guide the decision-making process about peroxide, or sodium bisulfite (Liu et al., 2022a,b). However, certain how to proceed in prevention. Various methods can be employed to antifungal additives, including propionate, calcium sorbate, and nata- process food, including physical, chemical, or biological approaches. mycin, are prohibited for packaging purposes in human food. However, Most of them can partially destroy or inactivate mycotoxins, but they antifungals such as benomyl, ethoxyquin, diphenylamine, and thiaben- rarely eliminate them (Mavrommatis et al., 2021). Furthermore, most of dazole are specifically applicable to fruits and vegetables after har- these methods have side effects on the quality of the raw material, which vesting (Hamad et al., 2023). The use of ionizing radiation along with may limit their application. hydrogen phosphide (PH3) in packaged products may offer promising solutions for the future (Pandiselvam et al., 2019). This method effec- 4.1. Physical treatments tively safeguards against recontamination without leaving residue behind. Although the presence of fungus in food is not always associated In the realm of physical treatments, thermal or radiation methods with the existence of mycotoxin, it is vital to eradicate any food that can be employed to maintain low levels of water activity in the product, appears moldy. Chemical treatments for the elimination of mycotoxins specifically below 0.70. This is because low water activity generates an include ammonization, which is effective against AFB1 but may not be unfavorable environment for fungi to thrive and grow. Heat treatment is suitable for other mycotoxins (Conte et al., 2020). Other methods an effective strategy that necessitates temperatures above 160 ◦ C (Mir include oxidizing agents, including ozone, hydrogen peroxide, and so- et al., 2021). In another study, Deng et al. (2021) noted a reduction of dium bisulfite. Future solutions may involve restrictions on antifungal mycotoxin load of 45%–83%. Gómez-Salazar et al. (2023) reported a additives and ionizing radiation with hydrogen phosphide. It is crucial reduction in the load of aflatoxin B1 after treatment at temperatures of to consider the efficacy and potential limitations of each method, along 160 ◦ C. Heat treatment at high temperatures was also effective against with any regulatory restrictions or consumer preferences. Future ochratoxin (Gu et al., 2021). Similarly, heating dry ground wheat ex- research could focus on improving these techniques to optimize their hibits a decrease in OTA content at high temperatures and a long effectiveness, reduce costs, and mitigate potential negative effects on the exposure time (e.g., 50% reduction at 150 ◦ C for 3 h). When the ground quality of food products. material was moistened, the reduction was faster. However, OTA elim- ination was not achieved even at 200 or 250 ◦ C (Gu et al., 2021). 4.3. Biological treatments Treatment with ionizing radiation completely suppressed A. alutaceus growth, but OTA-contaminated feedstuff irradiation led to a Recently, there has been increasing curiosity in exploring biological reduction of only 36%–47% (Al-Abdalall, 2014). However, most of the detoxification methods as a substitute for physical and chemical mycotoxins are stable to high-temperature treatments. It is worth decontamination techniques to overcome their limitations. Natural mentioning that the inactivation of such mycotoxins is often partial, and substances such as some medicinal plants, spices, herbs, or enzymes treatment conditions can affect the nutritional quality of the treated have the potential to aid in the detoxification of mycotoxins in food, feeds. When using cold as a means of protection against fungal attacks, a although their application under field conditions is still complex, temperature below − 20 ◦ C guarantees a fungistatic effect. Similarly, it is expensive, and only partially deactivates mycotoxins (Mahato et al., crucial to mention that cold temperatures, such as − 80 ◦ C, do not 2022). Alternatively, some microorganisms that can reduce mycotoxin possess a sterilizing effect but only help preserve conidia, viruses, bac- content through fermentation processes have been identified. For teria, and foods. Wave treatments have also shown relative efficacy. example, some lactic acid bacteria detoxify aflatoxin M1 during milk Microwaves applied for 10 min can reduce the mycotoxin load by 32%, fermentation (Sanaldi and Coban, 2023). However, its application as a 9 R. Khan Toxicon 248 (2024) 108038 preventive measure in animal feed is limited. establish absorption immediately before mycotoxin is absorbed into the The advantage of biological detoxification methods is that they gastrointestinal tract. Mycotoxin adsorbents are diverse and challenging operate under mild reaction conditions, eliminating the need to use toxic to classify into a strict framework. A general classification would divide chemicals. Furthermore, these methods have little impact on the nutri- them into inorganic (silicates and activated carbon) and organic (yeast tional value or taste of the product. However, most of these methods cell walls, fibers, and bacteria) adsorbents. have been tested primarily in aqueous solutions with only a limited number of experiments in animals. The degradation of AF and OTA by 4.4.1. Mineral adsorbents the microflora of the gastrointestinal tract has been described in several Minerals, also known as silicates, are formed when silica dioxide is studies, such as the rumen of cows and sheep or the gut of rats (Loh et al., combined with other metal oxides (Bañuelos et al., 2023). Typically, 2020). In an in vitro experiment, it was shown that the human intestinal they are classified according to their structure: phyllosilicates (with a flora is also able to degrade OTA. Mwabulili et al. (2023) reported the lamellar structure) and tectosilicates (with more complex structures). partial adsorption of OTA by yeast during beer production. In an in vitro Clays are a type of phyllosilicate with lamellar or tubular structures that experiment, 1% yeast partially adsorbed OTA (100 g/mL) (binding was are smaller than 2 μm. Clays are classified according to the dominant strongly pH dependent). However, in a study of feeding pigs with a 5% mineral (aluminum, magnesium, silica) and the organization within the yeast supplement in the feed (OTA content 1 mg/kg), a slight reduction sheets (with tetrahedral or octahedral sheets) (Gomes and Rautureau, in toxin levels was achieved in the blood serum (Gavahian et al., 2020). 2021). The interaction between silicates and oxygen produces silicate Xu et al. (2022) tested different yeast strains to see if they could break tetrahedral sheets. Four oxygen atoms surround the center of the down OTA into the non-toxic ochratoxin α. They found a strain of the tetrahedral sheet, where silica is located. As this structure is electrically genus Trichosporon that was able to do this. In a broiler feeding study, decompensated (SiO4), oxygen must bond with a cation(Fig. 3, A) lyophilized Trichosporon cells can mitigate the adverse effects of OTA (Mohana, 2023). Eight hydroxyl atoms surround the cation (aluminum, on weight gain and mortality. Certain lactic acid bacteria (Lactobacillus calcium, or magnesium) that is present in the center of the octahedral rhamnosus) were partially able to bind to OTA in in vitro experiments sheet. The tetrahedral and octahedral sheets stack on top of each other to (Luz et al., 2018). The screening of microorganisms identified the bac- form the mineral. Based on the structural organization of different terium Acinetobacter calcoaceticus as effective in degrading DON and layers, the 2:1 configuration consists of two tetrahedral sheets and one OTA (Al-Nussairawi et al., 2023). A harmless type of A. niger bacteria octahedral sheet in the center. The tetrahedral and octahedral sheets broke down OTA into ochratoxin α. The substance was then broken combine to form the 1:1 structure (Fig. 3, B). The intersheet space in down even further into unknown products (Xiong et al., 2017). Fungi these sheets tends to attract cations due to its free valences, which are isolated from grapes, particularly some species of Aspergillus, have also usually negative (Kihal et al., 2022). The exchangeable cation plays a shown the ability to degrade OTA. A Rhizopus isolate was able to degrade crucial role in determining the clay cation exchange capacity (CEC) more than 95% of OTA in moistened wheat (Mwabulili et al., 2023). (Solly et al., 2020). White rot fungi have long been known to be capable of breaking down Clay absorption mechanisms are related to dipolar ions present in persistent environmental pollutants (Söylemez and Yamaç, 2021). OTA mycotoxin carbonyls and cations such as Na+, Ca+2, K+, and Al2+. and ochratoxin B were broken down to 23% and 3%, respectively, by the Cations are present in various places within clays, including the inter- food-grade white rot fungus Pleurotus ostreatus during a four-week laminar space, on the same sheets, and on the periphery of clays as barley fermentation. Ochratoxin α and probably ochratoxin β were the uncoordinated ions. Under low humidity, mycotoxins bind to cations intermediate products. through carbonyl groups through ion-dipole exchange. In conditions In short, biological detoxification methods, including natural sub- with higher humidity, the interlaminar space tends to expand by 10%, stances, medicinal plants, and microorganisms, provide a promising where the hydrogen molecules of water bind to mycotoxins to form an approach to reducing mycotoxins in food. While these methods preserve H-carbonyl oxygen-cation complex (Barrientos-Velazquez et al., 2022; the nutritional value and taste of the food, their implementation in field Szczerba et al., 2022). The main clays used for their absorption capacity conditions can be challenging. Bacteria, yeast, and fungi can break down include bentonites, montmorillonite, and other clays that need to be specific mycotoxins, indicating the potential for customized biological better characterized. interventions in various food matrices. Bentonite, also known as smectite, is formed by silicate sheets in a 2:1 ratio, while clay is formed by montmorillonite. One of the critical 4.4. Mycotoxin adsorbents characteristics of bentonites is their ability to expand the interlayer space because of their low anionic charge, which leads to a weaker force The alternative to these pre-ingestion treatments is the use of holding the sheets together. Therefore, the interlaminar space may differ mycotoxin adsorbents or sequestrants (Costamagna et al., 2024). These based on the specific cation introduced and the degree of hydration. binders are widely recognized as highly effective in controlling myco- Under dehydrated conditions, interlamellar spaces are small toxin (Kihal et al., 2022). Sequestrants effectively bind to mycotoxins in (0.95–1.0 nm), while under maximum hydration conditions, they can the intestinal tract, preventing their absorption, which is eventually expand by tens of manometers. These conditions allow bentonites to eliminated in the feces. There are two types of hijackers, including sequester mycotoxins (Kihal et al., 2021). High-dose bentonites are simple ones that contain a single component and complex ones that effective against T-2 toxin (Wang et al., 2023a,b). They are also effective consist of other elements. A suitable mycotoxin binder must demon- in sequestering AFB1, especially sodium bentonites, since the interlam- strate the effectiveness of mycotoxin absorption in in vitro and in vivo inar space is ample, unlike calcium bentonites, where the smaller tests (Yiannikouris et al., 2021). Although in vitro studies are helpful in interlaminar space does not allow sequestration of AFB1. On the other the early phases of mycotoxin adsorbent development, potential in- hand, Villarreal-Barajas et al. (2022) demonstrated that bentonites were teractions that may occur in vivo must be considered when assessing less efficient in absorbing ZEN, DON, or vomitoxin (DON). Montmoril- their effectiveness. Similarly, it should have a low level of inclusion in lonite, the main constituent of bentonites, is a hydrated the feed (1–2 kg/ton). It must have stable complexes with mycotoxins in sodium-calcium-aluminium-magnesium hydroxysilicate (Belousov a wide range of pH that represent physiological conditions in a way that et al., 2020). they can maintain adsorbed mycotoxin throughout the gastrointestinal Montmorillonite does not crystallize since van der Waals forces bind tract. It should exhibit a high affinity to allow effective absorption even the sheets. Therefore, the penetration of water and cations into inter- at low concentrations of mycotoxins while avoiding potential in- laminar space is relatively easy, which favors its expansion, which leads teractions with other nutrients. It must possess a high absorption ca- to a high ionic exchange coefficient (80–120 meq/100 g) (Qin et al., pacity that allows the use of low doses of the adsorbents. It should 2021). These characteristics allow for the effective absorption of 10 R. Khan Toxicon 248 (2024) 108038 Fig. 3. Molecular structure of delas sheets (A); tetrahedral and octahedral structures of the sheets (B); structure of the micropores in activated carbon (C); structure of the yeast cell wall (D). mycotoxins. Montmorillonite can absorb AFB1 and decrease AFM1 levels FMNs, and DON. However, the following research has questioned the in milk without affecting its quality. Kumi (2021) found that montmo- ability of activated carbon to sequester AFs. (Çakir et al., 2023) observed rillonites are also effective in adsorbing sterigmatocystin, an AF-like that the adsorption of AF decreased when lower doses of activated mycotoxin. In contrast, its ability to sequester ZEN and DON is lower. charcoal were used. Other studies in chickens and turkeys suggest that Tectosilicates are formed from sepiolite, zeolites, and clinopthiolite. the effectiveness of activated charcoal to sequester AF is considerably Structurally, they are a set of chains organized in a three-dimensional lower than that of clays (Dai et al., 2022; Hamad et al., 2022). manner with tiny pores. Tectosilicates are formed by the union of In conclusion, mineral adsorbents, particularly clays such as ben- tetrahedral sheets of silica oxide (SiO4) and aluminum (AlO4), with tonites montmorillonite, and activated carbon, are promising solutions calcium and sodium ions intercalated. Hydrated sodium calcium alu- to sequester mycotoxins (T-2 toxin and AFB1) due to their high cation minosilicates (HSCAS) acquire the ability to adsorb mycotoxins (Tchio exchange capacities and interlayer spaces. On the other hand, activated et al., 2024). The high cation-exchange capacity of HSCAS allows for a carbon, with its highly porous structure and large surface area, has high sequestration potential. HSCAS are selective for AFs and FMNs but excellent adsorption capacity for ZEN, DON, and AFs. However, its lack not for other mycotoxins. of specificity can lead to nutrient retention, raising concerns about its Activated carbon is an insoluble powder produced by the pyrolysis overall effectiveness. Therefore, more research is needed to optimize the process of organic compounds and a subsequent activation process, use of these mineral adsorbents in the management of mycotoxins, allowing the development of highly porous structures (Fig. 3, C). The considering factors like pore size, interaction area, dosage, and myco- pore structure enables a significant expansion in the total surface area of toxin structure. active carbon surpassing 500 m2/g (Wu et al., 2021). Additionally, there is super-activated carbon that boasts a larger surface area of 3500 m2/g 4.4.2. Organic adsorbents (Basaiah et al., 2024). The features of activated carbon are influenced by There are three types of organic adsorbent: yeast cell walls, ionized the type of raw material and the specific activation method used. The microfibers, and bacteria. The walls of yeasts, such as S. cerevisiae, are adsorption mechanism of activated carbon is based on interactions be- composed of β-D-glucans and glucomannans mannoproteins Fig. 3, D) tween the aromatic groups of mycotoxins and the hydrophobic envi- ((Tapingkae et al., 2022). The yeast cell walls consist of two sheets: the ronment within the pores of activated carbon. The high surface inner one, which provides the yeast with its rigidity and morphology, exchange of activated carbon enables it to have a high adsorption ca- and the outer one, which determines the surface properties of the wall. pacity. However, interactions lack specificity, which can lead to the The inner layer mainly consists of D-glucans complexed with chitin, retention or sequestration of other essential nutrients, such as vitamins, while the outer layer is composed of mannoprotein fibers (Bruinenberg organic acids, and proteins (Wu et al., 2021). and Castex, 2021). β-D-glucans can bind to mycotoxins through various Sequestering competence is influenced by the size of pores, the area mechanisms, including hydrogen bonds and ionic or hydrophobic in- of interaction, the dose, and the structure of the mycotoxin. Activated teractions (Ashry et al., 2022). Its efficacy depends on the glu- charcoal has been extensively investigated mainly for its ability to can/mannan ratio of the yeast strain used. The adsorption mechanism of sequester ZEN, DON, and AFs (Kihal et al., 2022; Liu et al., 2022a,b). the yeast cell wall involves β-D-glucans found in the internal lamina. Its Additionally, activated charcoal can be used to sequester OTs, ZEN, efficacy is influenced by various factors, such as shape, structure, 11 R. Khan Toxicon 248 (2024) 108038 bonding between aromatic groups and glucose units, and the presence of certain bacterial strains show promise in reducing the harmful effects of hydrogen bonds with hydroxyl groups (Takalloo et al., 2023). The yeast mycotoxins, but with varying degrees of adsorption capacity. Yeast cell walls can adsorb different types of mycotoxins. Takalloo et al. (2023) walls have moderate adsorption ability against AFs, OTA, and ZEN. demonstrated that β-D-glucans can effectively bind to ZEN. Nešić et al. Ionized microfibers and dietary fibers from plant by-products show (2021) also observed that yeast walls mitigated the negative impacts of effectiveness against specific mycotoxins, such as ZEN and AFB1. Addi- various mycotoxins, such as AF, OTA, ZEN, and T-2 toxins in contami- tionally, some bacterial strains, particularly lactic acid bacteria, show nated chicken feed. However, the adsorption capacity of mycotoxins is potential in the adsorption of mycotoxins through hydrophobic in- moderate and lower than that of clays. Ionized microfibers are derived teractions. However, concluding the most effective adsorbents is chal- from different plants such as cereals (wheat, barley, and rye), apples, lenging due to variability in studies and methodologies. Therefore, more and bamboo. These fibers contain cellulose, hemicellulose, and lignin. research is needed to optimize the use of organic adsorbents and address Thus, the inclusion of alfalfa fibers at levels of 25% in rats and 15% in uncertainties. pigs effectively reduced the adverse effects of ZEN without affecting productivity (Peivasteh-Roudsari et al., 2022). Similarly, alfalfa fibers 4.4.3. Interactions of mycotoxin adsorbents have shown the ability to mitigate the impacts of T-2 toxin in pigs, and Most of the adsorption mechanisms of various sequestering agents rats. are based on general physicochemical principles. This lack of specificity Dietary fibers extracted from by-products of grapes, artichokes, or raises questions about how these sequestrants can affect the availability almonds can effectively mitigate the harmful impacts of AFB1, ZEN, and of other nutrients (Liu et al., 2022a,b). In 2010, the EFSA established OTA. These fibers exhibit better performance in raw materials rich in guidelines to ensure the safety and efficacy of mycotoxin binders in food non-degradable fibers, such as lignin and cellulose. Some strains of and feed. Key aspects of these guidelines include the definition of Lactobacillus, Streptococcus, Propionibacterium, and Bifidobacterium may mycotoxin binders, the conduct of safety and efficacy evaluations, the also be able to adsorb mycotoxins using peptidoglycans and poly- specification of dosage and concentration levels, the implementation of saccharides (Massoud