**3. Main mycotoxins**

The worldwide contamination of foods and feeds with mycotoxins is a significant problem. It was estimated that 25% of the world's crops may be contaminated with these metabolites. Mycotoxigenic fungi involved with the human food chain belong mainly to three genera *As‐ pergillus*, *Penicillium* and *Fusarium*. The toxins produced by *Alternaria* have recently been of particular interest. The biochemistry, physiology and genetics of mycotoxigenic fungi have been discussed in several review articles [28, 41, 42].

Mycotoxins diffuse into grain and can be found in all grind fractions and, due to their ther‐ mo-resistant properties, also in products subjected to thermal processing [43].

The characteristics of major toxins that contaminate foods and feeds in the EU, described from the economic and toxicological point of view, are presented below.

#### **3.1. Aflatoxins (AFs)**

Aflatoxins are difuranocumarin derivatives. The main naturally produced aflatoxins based on their natural fluorescence (blue or green) are called B1, B2, G1, and G2. Aflatoxin M1 is a monohy‐ droxylated derivative of AFB1 which is formed and excreted in the milk of lactating animals. AFs are very slightly soluble in water (10–30 μg/mL); insoluble in non-polar solvents; freely soluble in moderately polar organic solvents (e.g. chloroform and methanol) and extremely soluble in dimethyl sulfoxide. They are unstable under the influence of ultraviolet light in the presence of oxygen, to extremes of pH (< 3, > 10) and to oxidizing agents [44].

Aflatoxins are produced only by a closely related group of aspergilli: *Aspergillus flavus*, *A.parasiticus,* and *A.nomius* strains [45]. These species are very widespread in the tropical and subtropical regions of the world. Other species such as *A.bombycis*, *A.ochraceoroseus*, and *A.pseudotamari* are also aflatoxin-producing species, but they are found less frequently [46, 47]. Aflatoxins constitute a problem concerning many commodities (nuts, spices), however, in terms of grain they are primarily problematic in case of maize. This is because only maize can be colonised by *A.flavus* and related species in the field. Out of the other grains, rice is an important dietary source of aflatoxins in tropical and subtropical areas. In regions with moderate climate, the problem is connected with imported commodities or the local crops that are wet or stored in improper conditions [45]. The carcinogenicity, mutagenicity and acute toxicology of AFB1 have been well documented. The IARC determined it to be a hu‐ man carcinogen (group 1A).

#### **3.2. Ochratoxin A (OTA)**

At 30o

192 Soybean - Pest Resistance

glyceollin [40].

**3. Main mycotoxins**

**3.1. Aflatoxins (AFs)**

*A.flavus*, and *A.niger* in 0.995 aw. At 18o

the production of six types of A trichothecenes in cereals [38].

been discussed in several review articles [28, 41, 42].

C, the ochratoxin production by *Aspergillus ochraceus* was inhibited by *A.candidus*,

*ochraceus* and *Alternaria alternata* resulted in a significant stimulation of ochratoxin A pro‐ duction [36]. Therefore, several microorganisms were reported as effective biocontrol agents against several fungal plant pathogens [37]. It was determined that *Trichoderma harzianum* produces a lytic enzyme, chitinase, which manifests antifungal activity against a wide range of fungal strains. It was found that non-toxigenic *T.harzianum* isolates significantly reduce

According to [39], soybean is not a favourable medium for ZEA production since it possess‐ es some features that limit the production of this toxin by *Fusarium* isolates. Similarly, the production of aflatoxin B1 by *Aspergillus flavus* was suppressed by soybean phytoalexin –

The worldwide contamination of foods and feeds with mycotoxins is a significant problem. It was estimated that 25% of the world's crops may be contaminated with these metabolites. Mycotoxigenic fungi involved with the human food chain belong mainly to three genera *As‐ pergillus*, *Penicillium* and *Fusarium*. The toxins produced by *Alternaria* have recently been of particular interest. The biochemistry, physiology and genetics of mycotoxigenic fungi have

Mycotoxins diffuse into grain and can be found in all grind fractions and, due to their ther‐

The characteristics of major toxins that contaminate foods and feeds in the EU, described

Aflatoxins are difuranocumarin derivatives. The main naturally produced aflatoxins based on their natural fluorescence (blue or green) are called B1, B2, G1, and G2. Aflatoxin M1 is a monohy‐ droxylated derivative of AFB1 which is formed and excreted in the milk of lactating animals. AFs are very slightly soluble in water (10–30 μg/mL); insoluble in non-polar solvents; freely soluble in moderately polar organic solvents (e.g. chloroform and methanol) and extremely soluble in dimethyl sulfoxide. They are unstable under the influence of ultraviolet light in the

Aflatoxins are produced only by a closely related group of aspergilli: *Aspergillus flavus*, *A.parasiticus,* and *A.nomius* strains [45]. These species are very widespread in the tropical and subtropical regions of the world. Other species such as *A.bombycis*, *A.ochraceoroseus*, and *A.pseudotamari* are also aflatoxin-producing species, but they are found less frequently [46, 47]. Aflatoxins constitute a problem concerning many commodities (nuts, spices), however, in terms of grain they are primarily problematic in case of maize. This is because only maize

mo-resistant properties, also in products subjected to thermal processing [43].

from the economic and toxicological point of view, are presented below.

presence of oxygen, to extremes of pH (< 3, > 10) and to oxidizing agents [44].

C and 0.995 aw, the interaction between *Aspergillus*

Ochratoxin A is a chlorinated isocumarin derivative, which contains a chlorinated isocoumar‐ in moiety linked through a carboxyl group to L-phenylalanine via an amide bond. It is colour‐ less, crystalline, and soluble in polar organic solvents compounds. This toxin is more stable in the environment than AFs. The studies of [45] reported that thermal destruction of OTA oc‐ curs after exceeding 250o C. OTA is produced by *Penicillium* species such as *P.verrucosum*, *P.au‐ riantiogriseum*, *P.nordicum*, *P.palitans*, *P. commune*, *P.variabile* and by *Aspergillus* species e.g. *A.ochraceus*, *A.melleus, A.ostanius*, as well as the aspergilli species of section *Nigri*. In moderate climates, the main producers of OTA are *Penicillium* species, while *Aspergillus* species domi‐ nate in tropical and subtropical climates. Ochratoxin A is often found with citrinin produced by *Penicillium aurantiogriseum*, *P.citrinum*, and *P.expansum* [48]. Significant human exposure comes from the consumption of grape juice, wine, coffee, spices, dried fruits and cereal-based products, e.g. whole-grain breads, and in addition to this from products of animal origin, e.g. pork and pig blood-based products. The Scientific Panel on Contaminants in the Food Chain of the European Food Safety Authority (EFSA) has derived an OTA tolerable weekly intake (TWI) on the level of 120ng/kg b.w. The IARC [49] determined it to be a possible human carci‐ nogen (group 2B). Ochratoxins are the cause of urinary tract cancers and kidney damage. In ru‐ minants, ochratoxin A is divided to non-toxic ochratoxin alfa and phenylalanine [44].

#### **3.3. Citrinin**

Citrinin is a polyketide nephrotoxin produced by several species of the genera *Aspergillus*, *Penicillium* and *Monascus*. Some of the citrinin-producing fungi are also able to produce ochratoxin A or patulin. Citrinin is insoluble in cold water, but soluble in aqueous sodium hydroxide, sodium carbonate, or sodium acetate; in methanol, acetonitrile, ethanol, and most other polar organic solvents. Thermal decomposition of citrinin occurs at >175 °C un‐ der dry conditions, and at > 100 °C in the presence of water. The known decomposition products include citrinin H2 which did not show significant cytotoxicity, whereas the de‐ composition product citrinin H1 showed an increase in cytotoxicity as compared to the pa‐ rent compound [50].The most commonly contaminated commodities are barley, oats, and corn, but contamination can also occur in case of other products of plant origin e.g. beans, fruits, fruit and vegetable juices, herbs and spices, and also in spoiled dairy products [50].

#### **3.4. Fumonisins (Fs)**

Fumonisins are a group of diester compounds with different tricarboxylic acids and polyhy‐ dric alcohols and primary amine moiety. There are several fumonisins, but only fumonisins B1 (FB1) and B2 (FB2) have been found in significant amounts. Some technological processes hydrolyze the tricarboxylic acid chain in fumonisin B1. The product of this reaction is more toxic than fumonisin [51].

(ALT). They belong to three structural classes: dibenzopyrone derivatives, perylene deriva‐ tives, and tetramic acid derivatives. Alternariol and related metabolites (AME and ALT) are produced by *Alternaria alternate, A.brassicae, A.citri, A.cucumerina, A.dauci, A.kikuchiana, A.sol‐ ani, A.tenuissima*, and *A.tomato*. These strains are known as plant, especially fruit and vegeta‐ ble pathogens. In cereals, soybean and oilseeds, AOH, AME and ALT are produced mainly by *Alternaria alternata, A.tennuisima,* and *A.infectoria.* AOH has been reported to possess cyto‐ toxic, genotoxic, mutagenic, carcinogenic, and oestrogenic properties [27]. Tenuazonic acid (TeA) is a mycotoxin and phytotoxin produced primarily by *Alternaria alternata* and other phytopatogenic *Alternaria* species. The overview of the chemical characterisation, producers,

Mycotoxins in Cereal and Soybean-Based Food and Feed

http://dx.doi.org/10.5772/54470

195

Sterigmatocystin (STC) is a precursor of the aflatoxins produced mainly by many *Aspergillus* species such as *A.versicolor, A.chevalieri, A.ruber, A.aureolatus, A.quadrilineatus, A.sydowi*, *Euro‐ tium amstelodami,* and less often by *Penicillium*, *Bipolaris*, *Chaetomium*, and *Emericella* genera [30]. Sterigmatocystin was reported as a fungal metabolite in mouldy wheat, rice, barley, ra‐ peseed, peanut, corn, and cheeses or salami. The STC producers, occurrence and toxic prop‐

**4. Contamination level in cereal and soybean-based food and feed**

Food security strategy in the European Union (EU) includes the Rapid Alert System for Food and Feed. The RASFF was established by the European Parliament and Council Regu‐ lation No. 178/2002 laying down the general principles and requirements of food law, estab‐ lishing the European Food Safety Authority and specifying the procedures in matters

In 2002 – 2011, the number of notifications to the RASFF system due to mycotoxin contami‐ nation of food was respectively: 302, 803, 880, 996, 878, 760, 933, 669, 688, 631 notifications identifying the presence of aflatoxin B1 (AFB1) and the amount of AFB1, B2, G1, G2, AFM1, ochratoxin A (OTA), fumonisins B1 and B2 (FB1, FB2), patulin, deoxynivalenol (DON) and zearalenone (ZEA) in such groups of foods, as nuts and milk, oilseeds, cereal, dried fruit, fruit, cocoa, coffee, herbs and spices, wine, milk, products for children. Approximately 95% of the notifications concerned foodstuffs contaminated with aflatoxins. During this period, the number of notifications regarding mycotoxin contamination of grains did not exceed 15% of the total number of notifications. The data in Figure 1 show that in 2002-2011 aflatox‐ ins, ochratoxin A and fumonisins were the main contaminants isolated from cereals [59].

In the research of [60], ninety-fife cereal samples from retail shops and local markets of dif‐ ferent locations in Pakistan were examined in terms of the presence of aflatoxins. The results showed the percentage of aflatoxin contamination samples in the commodities such as in:

toxicity, analysis and occurrence in foodstuffs was summarised by [27].

**3.8. Sterigmatocystin**

**products**

erties were reviewed by [30, 57].

concerning food safety [58].

FB1 is produced by fungi from *Fusarium* genera, especially by *F.moniliforme* and *F.prolifera‐ tum.* The study of [11] suggests that the risk of contamination with *Fusarium* toxins is higher for maize and wheat than for soybean and pea. High concentrations of fumonisins are asso‐ ciated with hot and dry weather, followed by the periods of high humidity. Studies on fu‐ monisin residues in milk, meat and eggs are incomplete [52, 53]. Human exposure assessments on fumonisin B1 have rarely been reported. The mean daily intake in Switzer‐ land is estimated to be 0.03 μg/kg bw/day. In the Netherlands the exposure estimates ranged from 0.006 to 7.1 μg/kg bw/day. In South Africa, the estimates ranged from 14 to 440 μg/kg bw/day, showing that the exposure to FB1 is considerably higher than in the other countries in which exposure assessments were performed [54]. It was concluded that for Fs there was inadequate evidence in humans for carcinogenicity. Therefore, the IARC classified *Fusarium monilliforme* toxins, including fumonisins, as potential carcinogens to humans (group 2B).

#### **3.5. Zearalenone**

Zearalenone is a macrocyclic lactone with high binding affinity to oestrogen receptors. ZEA is produced mainly by *Fusarium graminearum* and *F.sporotrichoides* in the field and during storage of commodities such as maize, barley, sorghum, and soybean. The IARC has evalu‐ ated the carcinogenicity of zearalenone and found it to be a possible human carcinogen (group 2B). Residues of zearalenone in meat, milk and eggs do not appear to be a practical problem [53, 54].

#### **3.6. Trichotecenes**

Trichothecenes constitute a group of 50 mycotoxins produced by *Fusarium*, *Cephalosporium* and *Stachybotrys* genera in different commodities. There are including T-2 toxins, deoxyniva‐ lenol, nivalenol, and diacetoxyscirpenol. Beside trochothecenes, deoxynivalenol (DON, wo‐ mitoxin) is probably the most widely distributed in cereal and soybean foods and feeds. In contaminated cereals, DON derivatives such as 3-acetyl DON and 15-acetyl DON can occur in significant amounts (10 – 20%) with DON. DON is produced by closely related *Fusarium graminearum, F.culmorum* and *F.crokwellense* species [55].

T-2 toxin produced mainly by *F.sporotrichoides* and *F.poae* is primarily associated with mould millet, wheat, rye, oats, and buckwheat. This toxin can be transmitted from dairy cattle feed to milk [56].

#### **3.7. Alternaria toxins**

*Alternaria* species, besides *Fusarium,* is the most isolated fungi from soybean and other cere‐ als. Several species are known producers of toxic metabolites called *Alternaria* mycotoxins. The most important *Alternaria* mycotoxins include alternariol (AOH), alternariol monometh‐ yl ether (AME), altertoxins I, II, and III (ATX-I, -II, III), tenuazonic acid (TeA), and altenuene (ALT). They belong to three structural classes: dibenzopyrone derivatives, perylene deriva‐ tives, and tetramic acid derivatives. Alternariol and related metabolites (AME and ALT) are produced by *Alternaria alternate, A.brassicae, A.citri, A.cucumerina, A.dauci, A.kikuchiana, A.sol‐ ani, A.tenuissima*, and *A.tomato*. These strains are known as plant, especially fruit and vegeta‐ ble pathogens. In cereals, soybean and oilseeds, AOH, AME and ALT are produced mainly by *Alternaria alternata, A.tennuisima,* and *A.infectoria.* AOH has been reported to possess cyto‐ toxic, genotoxic, mutagenic, carcinogenic, and oestrogenic properties [27]. Tenuazonic acid (TeA) is a mycotoxin and phytotoxin produced primarily by *Alternaria alternata* and other phytopatogenic *Alternaria* species. The overview of the chemical characterisation, producers, toxicity, analysis and occurrence in foodstuffs was summarised by [27].

### **3.8. Sterigmatocystin**

B1 (FB1) and B2 (FB2) have been found in significant amounts. Some technological processes hydrolyze the tricarboxylic acid chain in fumonisin B1. The product of this reaction is more

FB1 is produced by fungi from *Fusarium* genera, especially by *F.moniliforme* and *F.prolifera‐ tum.* The study of [11] suggests that the risk of contamination with *Fusarium* toxins is higher for maize and wheat than for soybean and pea. High concentrations of fumonisins are asso‐ ciated with hot and dry weather, followed by the periods of high humidity. Studies on fu‐ monisin residues in milk, meat and eggs are incomplete [52, 53]. Human exposure assessments on fumonisin B1 have rarely been reported. The mean daily intake in Switzer‐ land is estimated to be 0.03 μg/kg bw/day. In the Netherlands the exposure estimates ranged from 0.006 to 7.1 μg/kg bw/day. In South Africa, the estimates ranged from 14 to 440 μg/kg bw/day, showing that the exposure to FB1 is considerably higher than in the other countries in which exposure assessments were performed [54]. It was concluded that for Fs there was inadequate evidence in humans for carcinogenicity. Therefore, the IARC classified *Fusarium monilliforme* toxins, including fumonisins, as potential carcinogens to humans (group 2B).

Zearalenone is a macrocyclic lactone with high binding affinity to oestrogen receptors. ZEA is produced mainly by *Fusarium graminearum* and *F.sporotrichoides* in the field and during storage of commodities such as maize, barley, sorghum, and soybean. The IARC has evalu‐ ated the carcinogenicity of zearalenone and found it to be a possible human carcinogen (group 2B). Residues of zearalenone in meat, milk and eggs do not appear to be a practical

Trichothecenes constitute a group of 50 mycotoxins produced by *Fusarium*, *Cephalosporium* and *Stachybotrys* genera in different commodities. There are including T-2 toxins, deoxyniva‐ lenol, nivalenol, and diacetoxyscirpenol. Beside trochothecenes, deoxynivalenol (DON, wo‐ mitoxin) is probably the most widely distributed in cereal and soybean foods and feeds. In contaminated cereals, DON derivatives such as 3-acetyl DON and 15-acetyl DON can occur in significant amounts (10 – 20%) with DON. DON is produced by closely related *Fusarium*

T-2 toxin produced mainly by *F.sporotrichoides* and *F.poae* is primarily associated with mould millet, wheat, rye, oats, and buckwheat. This toxin can be transmitted from dairy cattle feed

*Alternaria* species, besides *Fusarium,* is the most isolated fungi from soybean and other cere‐ als. Several species are known producers of toxic metabolites called *Alternaria* mycotoxins. The most important *Alternaria* mycotoxins include alternariol (AOH), alternariol monometh‐ yl ether (AME), altertoxins I, II, and III (ATX-I, -II, III), tenuazonic acid (TeA), and altenuene

*graminearum, F.culmorum* and *F.crokwellense* species [55].

toxic than fumonisin [51].

194 Soybean - Pest Resistance

**3.5. Zearalenone**

problem [53, 54].

**3.6. Trichotecenes**

to milk [56].

**3.7. Alternaria toxins**

Sterigmatocystin (STC) is a precursor of the aflatoxins produced mainly by many *Aspergillus* species such as *A.versicolor, A.chevalieri, A.ruber, A.aureolatus, A.quadrilineatus, A.sydowi*, *Euro‐ tium amstelodami,* and less often by *Penicillium*, *Bipolaris*, *Chaetomium*, and *Emericella* genera [30]. Sterigmatocystin was reported as a fungal metabolite in mouldy wheat, rice, barley, ra‐ peseed, peanut, corn, and cheeses or salami. The STC producers, occurrence and toxic prop‐ erties were reviewed by [30, 57].
