Author's personal copy

Food and Chemical Toxicology 60 (2013) 218–237

Contents lists available at ScienceDirect

Food and Chemical Toxicology

journal homepage: www.elsevier.com/locate/foodchemtox

Review

Mycotoxins: Occurrence, toxicology, and exposure assessment

S. Marin, A.J. Ramos, G. Cano-Sancho, V. Sanchis ⇑
Food Technology Dept., UTPV-XaRTA, Agrotecnio Center, University of Lleida, Rovira Roure 191, 25198 Lleida, Spain

a r t i c l e i n f o a b s t r a c t

Article history: Mycotoxins are abiotic hazards produced by certain fungi that can grow on a variety of crops.
Received 15 March 2013 Consequently, their prevalence in plant raw materials may be relatively high. The concentration of
Accepted 18 July 2013 mycotoxins in finished products is usually lower than in raw materials. In this review, occurrence and
Available online 29 July 2013
toxicology of the main mycotoxins are summarised. Furthermore, methodological approaches for
exposure assessment are described. Existing exposure assessments, both through contamination and
Keywords: consumption data and biomarkers of exposure, for the main mycotoxins are also discussed.
Mycotoxins
Ó 2013 Elsevier Ltd. All rights reserved.
Food
Occurrence
Toxicology
Exposure

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219
2. Main mycotoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219
2.1. Aflatoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219
2.2. Ochratoxin A . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
2.3. Fusarium toxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
2.3.1. Fumonisins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
2.3.2. Zearalenone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222
2.3.3. Trichothecenes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222
2.3.4. Emerging Fusarium mycotoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222
2.4. Patulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223
2.5. Alternaria toxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223
2.6. Ergot alkaloids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223
3. Mycotoxin occurrence in foodstuffs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223
3.1. Aflatoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223
3.2. Ochratoxin A . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224
3.3. Fusarium toxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224
3.3.1. Fumonisins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224
3.3.2. Zearalenone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224

Abbreviations: AAL-toxins, A. alternata f.sp. lycopersici toxins; 3-Ac-DON, 3-acetyldeoxynivalenol; 15-Ac-DON, 15-acetyldeoxynivalenol; AFs, aflatoxins; AFB1, aflatoxin B1;
AFB1-N7-Gua, 8,9-dihydro-8-(n7-guanyl)-9-hydroxy AFB1; AFB1, FAPY, AFB1-formamidopyrimidine; AFM1, aflatoxin M1; ALT, alernuene; AOH, alternariol; AME, alternariol
monomethyl ether; ATA, alimentary toxic aleukia; ATX, altertoxins; aw, water activity; BEN, Balkan endemic nephropathy; BEA, beauvericin; bw, body weight; CER, enzyme
ceramide; CYP, cytochrome P-450; DON, deoxynivalenol; DNA, deoxyribonucleic acid; EAs, ergot alkaloids; EFSA, European Food Safety Authority; ELEM, equine
leukoencephalomalacia; ENNs, enniatins; EU, European Union; FAO, Food and Agricultural Organization of the United Nations; FBs, fumonisins B1, B2 and B3; FB1, fumonisin
B1; FUS, fusaproliferin; HCC, hepatocellular carcinoma; HT2, HT-2 toxin; IARC, International Agency for Research on Cancer; Iso-TeA, iso-tenuazonic acid; JECFA, Joint FAO/
WHO expert committee on food additives; LC–ESI-MS/MS, liquid chromatography–electrospray ionization tandem mass spectrometry; LD50, lethal dose 50%; LOD, limit of
detection; LOQ, limit of quantification; MON, moniliformin; NOEL, non observed effect level; NTD, neural tube defects; OTA, ochratoxin A; OTa, ochratoxin a; PPE, porcine
pulmonary oedema; PTDIs, provisional tolerable daily intakes; PAT, patulin; PMTDI, provisional maximum tolerable daily intake; Phe, phenylalanine; RASFF, Rapid Alert
System for Food and Feed; RNA, ribonucleic acid; Sa, sphinganine; SCF, scientific committee on food So, sphingosine; TDS, total diet studies; TDI, tolerable daily intake; TeA,
tenuazonic acid; TEN, tentoxin; T2, T-2 toxin; WHO, World Health Organization; ZEN, zearalenone.
⇑ Corresponding author.
E-mail address: vsanchis@tecal.udl.cat (V. Sanchis).

0278-6915/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.fct.2013.07.047
 Author's personal copy

S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237 219

3.3.3. Trichothecenes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224

3.3.4. Emerging Fusarium mycotoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226
3.4. Patulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226
3.5. Alternaria toxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226
4. Toxicological aspects of the main mycotoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226
4.1. Aflatoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226
4.2. Ochratoxin A . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 227
4.3. Fusarium toxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228
4.3.1. Fumonisins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228
4.3.2. Zearalenone. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228
4.3.3. Trichothecenes (T-2/HT-2, DON) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228
4.4. Patulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229
5. Exposure assessment to mycotoxins. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229
5.1. Methodological issues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229
5.2. Exposure to mycotoxins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 231
Conflict of Interest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 234
Acknowledgement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 234
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 234

1. Introduction als/bakery products, and foodstuffs, were the most affected catego-
ries (Table 3). Analysis of these notifications reveals which products
Mycotoxins are secondary metabolites produced by fungi that are more susceptible to be contaminated by these fungal
may be injurious to vertebrates upon ingestion, inhalation, or skin metabolites.
contact. The diseases they cause, known as mycotoxicosis, do not
need to involve the toxin-producing fungus. Thus, they are abiotic 2. Main mycotoxins
hazards but with biotic origin. Diagnostic features characterizing
mycotoxicosis are the following: the disease is not transmissible; Mycotoxins are produced by a number of fungal genera, primarily
drug and antibiotic treatments have little or no effect; outbreaks Aspergillus, Penicillium, Alternaria, Fusarium, and Claviceps genus.
are often seasonal; the outbreaks are usually associated with a spe- The most relevant groups of mycotoxins found in food are produced
cific foodstuff; and examination of the suspected food or foodstuff by the following five fungal genera: aflatoxins, produced by Aspergil-
often reveals signs of fungal activity. Mycotoxins can appear in the lus species; ochratoxin A produced by both Aspergillus and Penicil-
food chain because of fungal infection of crops, either by being con- lium; trichothecenes (type A: HT-2 and T-2 toxin, and type B:
sumed directly by humans or used as livestock feed. The metabo- deoxynivalenol), zearalenone, fumonisins B1 and B2, and the emerg-
lism of ingested mycotoxins could result in mycotoxin ing mycotoxins (fusaproliferin, moniliformin, beauvericin, and enni-
accumulation in different organs or tissues, entering into the food atins) produced mainly by Fusarium species; ergot alkaloids
chain through meat, milk, or eggs. produced by Claviceps; and altenuene, alternariol, alternariol methyl
The Food and Agriculture Organization of the United Nations ether, altertoxin, and tenuazonic acid produced by Alternaria species
(FAO) estimated that approximately 25% of the cereals produced (Barkai-Golan, 2008; Bottalico and Logrieco, 1998). The main pro-
in the world are contaminated by mycotoxins (Rice and Ross, ducing species of these mycotoxins are listed in Table 4.
1994). Other foods, like nuts, spices, fruits, and their by-products
can also be contaminated by these fungal metabolites. Mycotoxin 2.1. Aflatoxins
production in agricultural crops can occur at various points in the
food chain: at pre-harvest, harvest and drying, and storage. Poor Aflatoxins (AFs) are difuranocoumarins mainly produced by two
agricultural and harvesting practices, improper drying, handling, species of Aspergillus from section Flavi, primarily found in hot, hu-
packaging, storage, and transport conditions promote fungal
Table 1
growth, increasing the risk of mycotoxin production. Once the food
2012 Notifications by hazard category in the EU.
has been processed, further mycotoxin production is difficult as long
as food commodities are stored under conditions that prevent fungal Hazard category Alert Border Information Information
rejection for attention for follow-up
contamination and mycotoxin bioproduction, especially if the water
activity of the product is low enough to prevent mould growth and Allergens 64 3 17 1
Biocontaminants 6 9 26 2
mycotoxin production. This is the key element for mycotoxin-free
Food additives and 10 59 23 47
products. However, when water activity of the stored products in- flavourings
creases to levels allowing fungal growth and mycotoxin production, Foreign bodies 24 61 26 47
toxins can also accumulate in processed products. GMO/novel food 2 52 14 22
Many countries have adopted regulations to limit mycotoxin Heavy metals 57 108 79 24
Industrial 16 9 18 14
exposure. Their presence is not only related to the effect they might contaminants
have on consumer health, but may also have an impact on world Mycotoxins 38 425 53 9
trade. According to the annual report of the Rapid Alert System for Parasitic infestation 4 13 13 25
Food and Feed (RASFF), in 2012 mycotoxins were the main hazard Pathogenic 162 159 168 103
microorganisms
in border rejection notifications in the European Union (Table 1).
Pesticides residues 19 320 90 18
The number of alert notifications and information for attention noti- Residues of veterinary 12 18 16 14
fications was also outstanding. In the report, aflatoxins were the pri- medicinal products
mary mycotoxins associated to the notifications (Table 2), and nuts,
Rapid Alert System for Food and Feed (RASFF) Annual Report, 2012.
nut products and seeds, fruits and vegetables, herbs and spices, cere-
 Author's personal copy

220 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237

Table 2 found in foods due to fungal contamination during pre- and post-
Mycotoxin notification in EU during 2008–2012. harvest. The rate and degree of contamination depend on differ-
Mycotoxin 2008 2009 2010 2011 2012 Total ent factors such as temperature, humidity, water activity,
Aflatoxins 902 638 649 585 484 3258 concurrent mycobiota, physical damage, and other storage condi-
Deoxynivalenol (DON) 4 3 2 11 4 24 tions (EFSA, 2007).
Fumonisins 2 1 3 4 4 14 Aflatoxin B1 (AFB1) is considered the most toxic aflatoxin. It is
Ochratoxin A 20 27 34 35 32 148 metabolized mainly by the liver to AFB1-8,9-exo-epoxide and
Patulin 3 3
Zearalenone 2 4 6
8,9-endo-epoxide. The exo-epoxide binds to DNA to form the
predominant 8,9-dihydro-8-(N7-guanyl)-9-hydroxy AFB1 (AFB1-
Total 933 669 688 635 525 3450
N7-Gua) adduct. AFB1-N7-Gua may be converted to two secondary
Rapid Alert System for Food and Feed (RASFF) Annual Report, 2012. lesions, an apurinic site and a more stable ring opened AFB1-for-
mamidopyrimidine (AFB1-FAPY) adduct; the latter is far more
mid climates. The B aflatoxin-producing Aspergillus flavus is of persistent in vivo than AFB1-N7-Gua. The major human cytochrome
ubiquitous occurrence in nature, colonizing mostly the aerial P450 (CYP) enzymes involved in aflatoxin metabolism are CYP3A4,
parts of plants (leaves, flowers). Aspergillus parasiticus produces 3A5, 3A7, and 1A2.
B and G aflatoxins, and is more adapted to a soil environment Aflatoxin M1 (AFM1) (Fig. 2) is the main monohydroxylated
with a more limited distribution (EFSA, 2007). Other species of derivative of AFB1 formed in the liver via cytochrome P450-associ-
this section, such as A. nomius, A. pseudotamarii, and A. bombycis ated enzymes. Mammals that ingest AFB1-contaminated diets ex-
also produce aflatoxins (Peterson et al., 2001). The structures of crete amounts of the main 4-hydroxylated metabolite AFM1 into
the key aflatoxins are shown in Fig. 1. These mycotoxins are milk.

Table 3
Mycotoxin notifications by products in the EU during 2012.

Product category Aflatoxins Deoxynivalenol (DON) Fumonisins Ochratoxin A Zearalenone

Cereals and bakery products 17 4 4 6 3
Feed 79
Fruits and vegetables 137 19 1
Herbs and spices 33 4
Nuts, nut products and seeds 204
Other 14 3
Total 484 4 4 32 4

Rapid Alert System for Food and Feed (RASFF) Annual Report, 2012.

Table 4
Non-exhaustive list of mycotoxins and producing species.

Mycotoxin Acronym Species producing

Aflatoxins B1, B2, G1, G2 AFB1 Aspergillus section Flavi
AFB2
AFG1
AFG2
Alternariol AOH Alternaria alternata
Alternariol monomethyl ether AME Alternaria alternata, A. solani
Tenuazonic acid TeA Alternaria alternata,
Altertoxins ALTs A. tenuissima
Altenuene ALT Alternaria alternata
Alternaria alternata
Beauvericin BEA F. sporotrichioides, F. poae,
F. langsethiae, Fusarium section
Liseola, Fusarium avenaceum
Enniatins ENNs Fusarium avenaceum, F. tricinctum
Fusaproliferin FUS F. poae, F. langsethiae, F. sporotrichioides
F. proliferatum, F. subglutinans
Moniliformin MON Fusarium avenaceum, F. tricinctum, Fusarium section Liseola
Ergot alkaloids EAs Claviceps purpurea, C. fusiformis, C. africana, Neotyphodium spp.
Fumonisins B1, B2 FB1, Fusarium section Liseola
FB2
Ochratoxin A OTA Aspergillus section Circumdati
Aspergillus section Nigri
Penicillium verrucosum
Penicillim nordicum
Patulin PAT Penicillim expansum, Bysochlamis nívea, Aspergillus clavatus
HT-2 and T-2 toxin (type A trichothecenes) HT-2 Fusarium acuminatum, F. poae,
T-2 F. sporotrichioides, F. langsethiae
Deoxynivalenol (type B trichothecenes) DON Fusarium graminearum, F. culmorum, F. cerealis
Zearalenone ZEN Fusarium graminearum (F. roseum), F. culmorum, F. equiseti, F. cerealis, F. verticillioides, F. incarnatum
 Author's personal copy
S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
Fig. 1. Molecular structure of aflatoxins B1, B2, G1, and G2.
AFs are very stable and may resist quite severe processes like
roasting, extrusion, baking, and cooking. For this reason, they can
be a problem in processed foods, such as roasted nuts and bakery
products.
2.2. Ochratoxin A
Ochratoxin A (OTA) is a phenylalanyl derivative of a substituted
isocoumarin (R)-N-[5-chloro-3,4-dihydro-8-hydroxy-3-methyl-
1-oxo-1H-2-benzopyran-7-y1)-carbonyl]-L-phenylalanine. The struc-
ture of ochratoxin A is shown in Fig. 3. Ochratoxin a is a product
of the hydrolysis of OTA as a consequence of the lack of the
phenylalanine molecule. Its name derives from Aspergillus ochrac-
eus, the mould from which it was first isolated (van der Merwe
et al., 1965). OTA is produced by two main genera of fungi, Aspergil-
lus and Penicillium. The main producing species belong to the Asper-
gillus section Circumdati, Aspergillus section Nigri, Penicillium
verrucosum, and Penicillium nordicum (EFSA, 2006a).
OTA is the most toxic member of the ochratoxins. This myco-
toxin is structurally similar to the amino acid phenylalanine
(Phe). Thus, it has an inhibitory effect on a number of enzymes that
use Phe as a substrate, in particular, Phe-tRNA synthetase, which
can result in the inhibition of protein synthesis. It is a mitochon-
drial poison, causing mitochondrial damage, oxidative burst, lipid
peroxidation, and interferes with oxidative phosphorylation. In
addition, OTA increases apoptosis in several cell types (Kuiper-
Goodman and Scott, 1989).
OTA is a stable compound that is not destroyed by common
food preparation procedures. Temperatures above 250 °C for sev-
eral minutes are necessary to reduce the concentration of this toxin
(Boudra et al., 1995).
2.3. Fusarium toxins
The most reported Fusarium toxins in foods are the fumonisins,
trichothecenes, zearalenone (ZEN), and a group of mycotoxins
known as the emerging mycotoxins that include beauvericin, enni-
Fig. 2. Molecular structure of aflatoxin M1.
221
Fig. 3. Molecular structure of ochratoxin A.
atins, fusaproliferin, and moniliformin. These mycotoxins can be
found alone or simultaneously, as well as co-occurring with other
mycotoxins such as aflatoxins, in cereals and in cereal-based foods
(Placinta et al., 1999). The co-occurrence is a usual situation in
cereals, particularly of mycotoxins produced by the same mould.
2.3.1. Fumonisins
Fumonisins are a group of mycotoxins with a strong structural
similarity to sphinganine, the backbone precursor of sphingolipids
(Gelderblom et al., 1992). These mycotoxins are mainly produced
by Fusarium species from the Liseola section. Fusarium verticillioides
(syn. Fusarium moniliforme) and Fusarium proliferatum are the main
producing species. Other fungal species, including F. napiforme, F.
dlamini, and F. nygamai, also produce fumonisins (EFSA, 2005a).
At least 12 fumonisin analogues are known, the most important
being the B series (FBs) (fumonisins B1, B2, and B3). From a toxico-
logical perspective, fumonisin B1 (FB1) is the most important
fumonisin. The chemical name of this mycotoxin is 1,2,3-propane-
tricarboxylic acid, 1,10 -[1-(12-amino-4,9,11-trihydroxy-2-methylt-
ridecyl)-2-(1-methylpentyl)-1,2-ethanediyl]ester (EFSA, 2005a).
The structure of fumonisin B1 is shown in Fig. 4. This group of
mycotoxins, especially FB1, strongly inhibits the enzyme ceramide
(CER) synthase that catalyses the acylation of sphinganine (Sa) and
recycling of sphingosine (So). The inhibition of CER synthase in-
creases intracellular Sa and other sphingoid bases, which are
highly cytotoxic compounds. This imbalance has been proposed
as the main responsible of toxicity, and possibly carcinogenicity,
of FBs, based on mechanistic studies with cell cultures and borne
out by animal studies (Norred et al., 1992). Based on this biological
perturbation, alterations of Sa to So ratio in tissues, urine, and
blood have been proposed as potential biomarkers of FBs exposure
in various animal species, but studies have not allowed an accurate
validation (Solfrizzo et al., 2004).
Fumonisins are the most important mycotoxins found in maize,
particularly when grown in warmer regions. As F. verticillioides and
F. proliferatum grow over a wide range of temperatures, but only at
relatively high water activities (aw > 0.9), FBs are formed in maize
prior to harvest or during the early stage of storage. Except under
extreme conditions, the concentration of FBs does not increase
during storage.
FBs are fairly heat-stable, and toxin content is significantly re-
duced only during processes in which the temperature exceeds
150 °C. There is little degradation of FBs during fermentation
(EFSA, 2005a).
Fig. 4. Molecular structure of fumonisin B1.
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2.3.2. Zearalenone
Zearalenone (ZEN) (6-(10-Hydroxy-6-oxo-trans-1-undecenyl)-
beta-resorcylic acid lactone) (Fig. 5) is a mycotoxin produced by
several Fusarium species, particularly by Fusarium graminearum
and also by F. culmorum, F. cerealis, F. equiseti, F. verticillioides,
and F. incarnatum. Generally, these Fusarium species grow and in-
vade crops in moist cool field conditions during blooming, but
growth and toxin production may also occur post-harvest under
poor storage conditions (EFSA, 2011a). These species can produce
small amounts of several related metabolites, a-zearalenol and b-
zearalenol being the most important derivatives (Richardson
et al., 1985). All zearalenones are estrogenic compounds, although
the estrogenic potential of a-zearalenol is higher than that of ZEN
and b-zearalenol (Hagler et al., 1979), probably due to a greater
binding affinity to estrogen receptors (Fitzpatrick et al., 1989).
ZEN is heat-stable up to 150 °C. Degradation has been observed
only at high temperatures or under alkaline conditions (Ryu
et al., 1999).

2.3.3. Trichothecenes
Trichothecenes are a group of structurally related compounds
with a common tetracyclic sesquiterpenoid 12,13-epoxytrichot-
hec-9-ene ring system. The approximately 170 identified trichot-
hecenes (Grove, 1988, 1993) have been divided in four types
(A-D) according to variations in the functional hydroxyl and acet-
oxy side groups. Type A is represented by HT-2 toxin (HT2) and
T-2 toxin (T2), and type B is most frequently represented by deoxy-
nivalenol (DON). The structures of the key trichothecenes are
shown in Figs. 6 and 7. The trichothecenes are in general very sta-
ble compounds, both during storage/milling and the processing/
cooking of food, and are not degraded by high temperatures (EFSA,
2011c; Eriksen and Alexander, 1998). C and D types include some
trichothecenes of lesser importance.
The simple type A and B trichothecenes are produced by several
Fusarium species (Thrane, 2001), but also by some species of Trich-
oderma (Nielsen et al., 2005). However, the most important T-2-
and HT-2-producing species T are F. sporotrichioides, F. langsethiae,
F. acuminatum, and F. poae. The main producers of DON are F.
graminearum, F. culmorum, and F. cerealis. These fungi are soil fungi
and are important plant pathogens that grow on the crop in the
field (Eriksen and Alexander, 1998). Because these species are na-
tive to areas with temperate climates, their mycotoxins are also
found there.
Different studies (Cundliff et al., 1974; Cundliff and Davies,
1977) have shown that these mycotoxins act by inhibiting eukary-
otic protein synthesis (toxicity) by binding to the 60S ribosomal
subunit and by interacting with the enzyme peptidyltransferase.
This interaction leads to varying degrees of inhibition of peptide
bond formation, depending on the chemical structure of the spe-
cific trichothecene. The T-2 toxin can inhibit DNA and RNA synthe-
sis in vivo and in vitro (WHO, 1990). In vivo, the T-2 toxin is rapidly
metabolised to HT-2 toxin, a major metabolite.
Fig. 6. Molecular structure of T2 and HT2 toxins.
moniliformin. There is limited available data on these metabolites,
not only because their late recognition, but particularly due to the
late understanding of their role as mycotoxins.
Fusaproliferin (FUS) is a sesterterpene toxic to Artemia salina,
L,6,10 IARC/LCL 171 human B lymphocytes, and SF-9 insect cells
and has teratogenic and pathogenic effects on chicken embryos.
This mycotoxin is produced by species of Fusarium section Liseola
such as F. proliferatum and F. subglutinans.
Enniatins (ENNs) and beauvericin (BEA) are a group of structur-
ally related cyclic hexadepsipeptides consisting of three d-2
hydroxycarboxylic acid and N-methylamino acid residues linked
alternately. F. avenaceum can produce at least six enniatins and
small amounts of beauvericin (Logrieco et al., 1998). Other Fusar-
ium species with capacity to produce enniatins are F. trincinctum,
F. poae, F. sporotrichioides, and F. langsethiae (Thrane, 2001),
whereas beauvericin producers also include F. sambucinum, F. poae,
F. langsethiae, F. verticillioides, F. sporotrichioides, F. proliferatum, and
F. subglutinans (Logrieco et al., 1998; Thrane, 2001). These myco-
toxins are cytotoxic (Uhlig et al., 2006), and their apolar nature en-
ables them to insert into cell membranes creating cation selective
channels (Ovchinnikov et al., 1974), thereby disturbing the intra-

2.3.4. Emerging Fusarium mycotoxins

Emerging Fusarium mycotoxins include other toxic secondary
metabolites such as fusaproliferin, enniatins, beauvericin, and
Fig. 7. Molecular structure of deoxynivalenol.

Fig. 5. Molecular structure of zearalenone. Fig. 8. Molecular structure of patulin.

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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237 223

cellular ionic homeostasis (Kamyar et al., 2004). Toxicological data
on these mycotoxins is insufficient.
Moniliformin (MON) (3-hydroxycyclobut-3-ene-1,2-dione) is a
very strong acid produced by several Fusarium species (F. avenace-
um, F. tricinctum, F. proliferatum, F. subglutinans, and F. verticillio-
ides). It is an inhibitor of several thiamine pyrophosphate
dependent enzymes, from which pyruvate dehydrogenase is the
best studied. This inhibition leads to the interruption of gluconeo-
genesis (Pirrung et al., 1996). Furthermore, this mycotoxin has
been reported to inhibit glutathione peroxidase and reductase
(Chen et al., 1990).
2.4. Patulin
Patulin (PAT) is a mycotoxin included in a group of compounds
commonly known as toxic lactones (4-hydroxy-4H-furo[3,2-c]pyr-
an-2(6H)-one). The structure of patulin is shown in Fig. 8. It is pro-
duced by a wide range of fungal species of the Penicillium,
Aspergillus, Byssochlamys, Eupenicillium, and Paecilomyces genera
from which Penicillium expansum, a common contaminant of dam-
aged fruits, is the most important (Morales et al., 2007).
PAT has a strong affinity for sulfhydryl groups, which explains
its inhibitory activity on many enzymes. As described for other
mycotoxins, PAT can alter the immune response of the host. In vitro
studies have shown that PAT inhibits several macrophage func-
tions. In vivo studies with mice reveal variable effects of PAT on
the immune system, including an increasing number of splenic T
lymphocytes and depressed serum immunoglobulin concentra-
tions, a modification of the hypersensitivity responses, and an in-
crease in the number of neutrophilsers (Puel et al., 2010).
toxins). A. alternata is the most important mycotoxin-producing
species (EFSA, 2011b).
2.6. Ergot alkaloids
Ergot alkaloids (EAs) are toxic compounds found in of Claviceps
species (sclerotia) that infect some economically important grain
cereals. These compounds are characterized by a tetracyclic ergo-
line ring system and are classified biosynthetically as trypto-
phan-derived alkaloids. The physiological effects of EAs have
been known since biblical times and are used as drugs in humans.
Claviceps purpurea is the main species in European grains and a
number of EAs, namely ergometrine, ergotamine, ergosine, ergo-
cristine, ergocryptine (which is a mixture of a- and b-isomers),
ergocornine, and the corresponding-inine epimers (EFSA, 2012)
can be found in its sclerotia.
Some studies suggest that EAs are absorbed from the gastroin-
testinal tract and are subjected to oxidative biotransformation, pri-
marily by cytochrome P450 3A4, and some EAs (e.g., ergometrine)
can subsequently be conjugated with glucuronic acid. EAs act on a
number of neurotransmitter receptors, particularly adrenergic,
dopaminergic, and serotonergic receptors. On repeated dosing of
various EAs, their effects on receptors result in ischemia, particu-
larly in the extremities (such as the tails of rats), decreased body
weight gain, and changes in the levels of some hormones (EFSA,
2012).
3. Mycotoxin occurrence in foodstuffs
3.1. Aflatoxins

2.5. Alternaria toxins
Alternaria mycotoxins include a small percentage of the more
than 70 phytotoxins produced by this genus that have been chem-
ically characterised and reported to act as mycotoxins in humans
and animals. They belong to three different structural groups:
the dibenzopyrone derivatives, alternariol (AOH), alternariol
monomethyl ether (AME), and altenuene (ALT); the perylene deriv-
atives altertoxins (ATX-I and II); and the tetramic acid derivatives,
tenuazonic acid (TeA) and iso-tenuazonic acid (iso-TeA). Some tox-
ins such as AOH, AME, TeA, and ATX have been described to induce
toxic effects in animals, including fetotoxic and teratogenic effects.
AOH and AME are both mutagenic and clastogenic in various
in vitro systems. Other known Alternaria mycotoxins are tentoxin
(TEN) and AAL-toxins (abbreviation of A. alternata f.sp. lycopersici
AFs are often detected in cereals and their derivatives, nuts, and
species. Food sample analysis reveals that the amount of AFB1 is of-
ten the highest in the mixture of AFs. According to an EFSA report
(2007) in which 34,326 sample analyses from EU countries were
pooled (including export, import, company, and market control
samples), the highest percentage of positive samples occurred in
Brazil nuts, pistachios, and spices (LOD was 0.1–0.2 lg/kg for
AFB1 and 0.2–0.4 lg/kg for total AFs) (Table 5). Brazil nuts and pis-
tachios presented a much higher mean and maximum values than
the other food groups. Furthermore, figs, peanuts, spices, hazelnuts,
and almonds had 97.5th percentile values above 2 lg/kg for AFB1
and above 4 lg/kg for total AFs. High maximum values were deter-
mined for most food categories except for baby foods and maize.
In most cases, values were at the low concentration end and a
few were high or very high; the median was lower or much lower
than the mean. Of the 168 samples with values above 200 lg/kg,

Table 5
Aflatoxin B1 and total aflatoxin (AFT) distribution (lg/kg) in food samples in the European Union (EFSA, 2007).

Food category N samples N samples > LOD Median AFB1/AFT Meana AFB1/AFT Maximum AFB1/AFT
Almonds 1766 471 (27%) 0.20/0.28 1.46/1.82 575/579
Brazil nuts 622 271 (43%) 0.20/0.40 22.2/39.6 1897/3337
Hazelnuts 3163 940 (30%) 0.16/0.30 0.95/1.70 200/200
Cashews 336 33 (10%) 0.10/0.20 0.42/0.60 36/39
Peanuts 8929 1830 (20%) 0.10–0.20 1.93/2.69 935/985
Pistachios 4069 1783 (44%) 0.20/0.40 16.8/19.4 2625/2680
Other nuts 1131 158 (14%) 0.10/0.20 1.16/1.41 385/402
Figs 2067 618 (30%) 0.15/0.24 1.36/2.22 130/151
Other dried fruits 1396 114 (8%) 0.10/0.24 0.26/0.51 20/90
Maize 943 136 (14%) 0.12/0.24 0.26/0.41 8/9
Other cereals 3010 207 (7%) 0.20/0.40 0.35/0.51 109/117
Spices 4698 1988 (42%) 0.20/0.40 1.46/1.88 96/96
Baby foods 592 23 (4%) 0.02/0.04 0.07/0.14 1/2
Other foodstuffs 1604 303 (19%) 0.10/0.20 0.53/0.75 99/99
a
Samples < LOD were given the LOD value.
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224 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237

Table 6
Ochratoxin A (lg/kg) in food samples in the European Union (SCOOP, 2002a report).

Food category N samples N samples > LOD LOD Meana Maximum

Cereals (raw materials and derived products) 5180 2825 (54%) 0.01–0.5 0.29 33.3
Green coffee 1704 620 (36%) 0.01–1 1.62 200.9
Roasted coffee 1184 549 (46%) 0.01–1 0.72 11.5
Beer 496 192 (39%) 0.0005–0.2 0.03 0.3
Wine 1470 872 (59%) 0.003–1 0.36 15.6
Cocoa and derived products 547 445 (81%) 0.01–0.25 0.24 3.6
Dried fruits 800 582 (73%) 0.01–0.5 2.30 53.6
Meat products 1860 328 (18%) 0.01–1 0.20 9.3
Spices 361 188 (52%) 0.1–0.4 1.15 23.8
Other foodstuffs 4927 2472 (50%) 0.0005–0.3 0.20 15.4
a
Samples < LOD were given the LOD/2 value.

raw material susceptible of OTA contamination. Comparison

Table 7
between green and processed coffee samples (3.6 (3641 lg/kg
Fumonisin B1 (lg/kg) in food samples in the European Union (SCOOP, 2003 report).
vs. 1.1 vs. 1092 lg/kg) indicates a marked reduction of OA, probably
Food category N samples N samples > LOD LOD Meana Maximum attributed to blending and losses during the roasting procedures.
Maize grain 801 534 (67%) 5–50 346.4 10,200 Among cereals, rye showed the highest level of contamination
Wheat grain 110 87 (79%) 10–30 71.2 736 (with 50% positive samples), followed by barley and wheat. The ex-
Wheat flour 256 42 (16%) 12–30 64.4 4343
tent of OTA incidence in contaminated beer was rather low as well
Maize grits 172 98 (57%) 5–30 347.6 4800
Maize flour 110 87 (97%) 5–50 408.5 4766 as the average levels (0.301 lg/kg in barley vs. 0.028 lg/L in beer).
Cornflakes 274 125 (46%) 0.2–500 31.5 1092
Sweet maize 145 13 (9%) 5–500 12.4 81
Polenta 29 29 (100%) 20–50 182.2 752 3.3. Fusarium toxins
a
Samples < LOD were given the LOD/2 value.
3.3.1. Fumonisins
Maize is, by far, the most FB-contaminated food commodity
110 were associated to pistachios, 30 to Brazil nuts, 23 to peanuts,
(Table 7), as well as its resulting milling fractions, such as grits
3 to other nuts, and 2 of almonds. Although a rare occurrence, the
(mean 347 lg/kg) and flour (mean 408 lg/kg) used for further pro-
heterogeneous distribution of AFs with occasionally very high val-
cessing. Sweet corn is less susceptible to Fusarium contamination
ues is a concern.
than other maize varieties. Certain thermal treatments or extru-
Cleaning of cereals or nuts, where mould-damaged kernels,
sion, have been proved to reduce to some extent FB presence in
seeds or nuts are removed from the intact commodity, may result
foods; in general, products for direct human consumption such
in 40–80% reduction of AFs (Park, 2002). Milling of cereals results
as corn flakes have low FB levels.
in a redistribution of AFs and other toxins in the different fractions
obtained, being flour and flaking grits intended for human con-
sumption the less contaminated. Baking, cooking, boiling, frying 3.3.2. Zearalenone
and roasting leads to different levels of AFs inactivation (Bullerman According to EFSA (2011a), among the grains for human con-
and Bianchini, 2007). sumption the frequency of occurrence of ZEN in maize (33%) and
Many feeds may contain AFs, and groundnut meal, cottonseed mean content level (15 lg/kg) are significantly higher. This trend
and maize are the most important AFs carriers in cows. Recent data was maintained in grain mill products, although very high levels
on AFM1 in milk samples in several studies from different EU coun- were determined in wheat bran (33 lg/kg). Thus, both the
tries show that the prevalence of AFM1 contaminated samples prevalence and level of ZEN in the analysed cereal products for hu-
seems to be very low. From the summed data (11,831 samples), man consumption was low except for vegetable oils (mostly maize
the incidence of occurrence of samples above the statutory limit germ oil) with 86% positive samples and a mean level of 72 lg/kg
of 0.05 lg/kg was only 0.06% (EFSA, 2004a). (Table 8).
In general, ZEN is redistributed between the milling fractions.
3.2. Ochratoxin A The by-products from cleaning the raw cereal grains (dust, hulls,
and others) were characterised by 3- to 30-fold higher ZEN concen-
More than 50% of positive samples were reported in the SCOOP trations than the cleaned cereal grains, while bran contained up to
report (2002a) for cacao and derived products (81%), dried fruits 2-fold higher concentrations. Generally, ZEN is not affected by
(73%), wine (59%), cereals (54%), and spices (52%), although cooking. Only under alkaline conditions or during extrusion cook-
positive samples are highly dependent on LOD levels (Table 6). ing (heating under a high degree of pressure) more than 40% reduc-
However, the highest mean levels of contamination were reported tion was observed (EFSA, 2011a).
for dried fruits, green coffee, and spices. Although no data was pro-
vided on OTA contamination in cocoa beans, results on processed
products confirmed cocoa beans as a raw material largely suscep- 3.3.3. Trichothecenes
tible to OTA contamination. Among different spices, nutmeg, papri- Regarding DON, among individual commodities, wheat has
ka, coriander, and pepper powder where the most highly and been investigated more widely in comparison with the other
frequently contaminated. Regarding wines, red and sweet wine grains. Maize shows the highest percentage of positive samples
seemed to be the most contaminated, even based on a rather low (89%) (Table 9), followed by wheat. Weighted mean ranged from
number of samples in the case of sweet wine. Data on dried fruits, 37 lg/kg for barley to 594 lg/kg for maize. Considering the EU
with particular reference to vine fruits derived products, indicate maximum limits, the provided occurrence data showed very few
these commodities to be highly susceptible to OTA contamination levels of contamination higher than the proposed legal limit of
as a result of the drying process. Green coffee is confirmed to be a 500 lg/kg (for cereal products as consumed and other cereal prod-
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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237 225

Table 8
Zearalenone (lg/kg) in food samples in the European Union (EFSA, 2011a).

Food category N samples N samples > LOD/LOQ Meana Median Maximum

Grains for human consumption 2190 372 (17%) 5.7 3.0 140
Wheat milling products 3088 432 (14%) 13 5.0 507
Rye milling products 482 31 (6.4%) 4.1 3.0 50
Maize milling products 838 369 (44%) 14 5.0 509
Bread and rolls 1247 94 (7.5%) 5.2 4.0 70
Pasta 330 13 (3.9%) 5.8 3.0 50
Breakfast cereals 1377 120 (8.7%) 5.7 3.0 172
Fine bakery wares 813 195 (24%) 7.7 5.0 98
Biscuits 541 179 (33%) 9.0 5.0 98
Beer 35 2 (5.7%) 1.0 0.5 10
Sweet corn 94 10 (11%) 4.8 4.0 50
Vegetable oils 221 190 (86%) 72 49 823
Food for infants and small children 420 17 (4%) 7.0 6.7 20

LOQ: 0.02–20 lg/kg.

a
Samples < LOD were given the LOD value.

Table 9
Deoxynivalenol (lg/kg) in food samples in the European Union (SCOOP, 2003 report).

Food category N samples N samples > LOD LOD Meana Maximum

Wheat and wheat flour 6350 3891 (61%) 2–220 205 3600
Barley 781 370 (47%) 2.220 37 550
Oat 595 204 (34%) 2–50 95 5004
Rye and rye flour 271 135 (50%) 2–220 42 595
Maize 520 463 (89%) 10–220 594 8850
Biscuits 95 23 (24%) 50–250 50.6 420
Bread 89 29 (32%) 50–250 88.9 560
Pasta 303 169 (56%) 20–250 141.2 3200
Breakfast cereals 71 40 (56%) 25–200 198.8 426
Baby food 185 133 (72%) 20–100 99.6 1075
a
Samples < LOD were given the LOD/2 value.

ucts at retail stage, 6%) and of 750 lg/kg (for flour used as raw
material in food products) (SCOOP report, 2003).
Although the percentage of positive samples was high in raw
materials and food products (after pooling all samples with differ-
ent LOD values), means were still low for processed products. For
example, biscuits, bread, and pasta samples were less often
contaminated than wheat samples and with less than half the
value in comparison to the raw material. Similarly, breakfast cere-
als and baby food that may contain maize among other cereals
contained DON levels lower than wheat and maize. Accordingly,
baking of bread, cookies and biscuits have led to variable reduc-
tions in DON from 0% to 71% (Bullerman and Bianchini, 2007). Can-
ning of maize led to a 12% reduction (Wolf-Hall et al., 1999). A
detoxification over 95% was achieved by extrusion cooking
(Cazzaniga et al., 2001).
Regarding T2 and HT-2, their mean individual concentrations in
food groups followed generally the same pattern as for the sum of
T-2 and HT-2 toxins. In general, HT-2 toxin concentration repre-
sents two-thirds of the sum of T-2 and HT-2 concentrations. From
the available data, it can be concluded that raw oats can be highly
contaminated with T-2 and HT-2, high incidence and concentra-
tions, followed by barley (Table 10). Maize may occasionally be
contaminated at a moderate concentration, while contamination
of wheat occurs infrequently and at low concentration level
(SCOOP report, 2003).
Processing cereals will substantially reduce T-2 and HT-2 con-
tamination in most food products due to redistribution over in
the various fractions. As for most mycotoxins, because T-2 and
HT-2 toxins are mostly attached to the outer hull of the grain,
cleaning, sorting, sieving and, de-hulling of grains leads to
marked increases in T-2 and HT-2 toxins in cereal by-products,
e.g. bran. Food products generally show low incidence and con-
centration of T-2 and HT-2, however, oat products may contain
some T-2 and HT-2. During baking and cooking, T-2 and HT-2
toxins seem to be relatively stable. Malting leads to substantially
lower levels of T-2 and HT-2 toxins in malt, compared to the ori-
ginal barley, although the ratio varies considerably (EFSA,
2011c).

Table 10
Sum of T-2 and HT-2 toxins (lg/kg) in food samples in the European Union (EFSA, 2011c).

Food category N samples N samples > LOD/LOQ Meana Median Maximum

Grains for human consumption 368 221 (60%) 16 12 124
Grain milling products 2281 821 (36%) 7.3 5 204
Bread and rolls 617 364 (59%) 3.4 4 27
Pasta 513 380 (74%) 2.5 1.5 17
Breakfast cereals 1808 976 (54%) 11 6.5 197
Fine bakery wares 531 329 (62%) 3.7 2.4 66
Snack food 36 20 (56%) 5.6 2.0 20
Vegetable and vegetable products 167 124 (74%) 3.7 1.3 10
Food for infants and small children 423 292 (69%) 3.4 1.4 33

LOQ: 0.02–20 lg/kg.

a
Samples < LOD were given the LOD value.
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226 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237

Table 11
Patulin (lg/kg) in food samples in the European Union (SCOOP, 2002b report).

Food category N samples N samples > LOD/LOQ LOD Meana Maximum

Apple juice 7820 4660 (60%) 0.03–10 15.6 1150
Other fruit juice 551 145 (26%) 0.03–10 11.3 750
Cider 215 33 (15%) 1–5 4.0 101
Apple puree 96 7 (7%) 0.20–10 4.9 86
Baby food 312 43 (14%) 0.31–10 2.8 68
a
Samples < LOD were given the LOD/2 value.

3.3.4. Emerging Fusarium mycotoxins
Thorough surveys to determine FUS, BEA, and ENN contamina-
tion of raw and processed foods are still scarce and are generally
limited to Northern Europe and the Mediterranean (Santini et al.,
2012). The presence of BEA or ENNs has been reported during
the last decade in some European countries (Croatia, Finland, Italy,
Norway, Slovakia, Spain, and Switzerland). Most studies have ana-
lyzed grain cereal samples (barley, corn, oat, wheat, and rice) and
few have included samples from cereal products (bread and baby
food) (Juan et al., 2013). High prevalence of ENNs has been partic-
ularly determined in the surveys. High levels of contamination
have been recently reported (up to 10–500 mg/kg) for the sum of
BEA and ENNs in wheat, barley, and corn in southern, central,
and northern Europe (Jestoi et al., 2009). High concentrations of
ENNs have been reported in Spain and northern Africa (maximum
concentration of enniatin A 814 mg/kg in rice in the Spanish mar-
ket), while concentrations were significantly lower in northern
Europe (maximum concentration of enniatin B 18.3 mg/kg in
wheat from Finland) (Santini et al., 2012). The most frequently
found mycotoxin was enniatin B and it concentrations in pasta
and baby food ranged from <LOQ to 106 and from <LOQ to
1100 lg/kg, respectively. High incidence (70.3%) of these mycotox-
ins was determined in multi-grain cereal baby food (Juan et al.,
2013). BEA appears to be of low significance in grains from cooler
climates, while it has been reported to occur at concentrations of
tens of mg/kg in Southern Europe and Morocco (Santini et al.,
2012). However, a maximum level of BEA of 844 lg/kg in a sample
of wheat pasta from Tunisia was found in a survey of cereal-based
products from Mediterranean countries (Serrano et al., 2012a). FUS
has been less investigated. Several reports from the Mediterranean
show its occasional occurrence up to 19.6 mg/kg in rice from Mor-
occo, while its natural occurrence in cooler climates seems to be
uncommon (Santini et al., 2012). Current literature regarding the
presence on these mycotoxins in grains and foods indicates there
may be a continuous low-level exposure to these toxic metabolites
(Jestoi et al., 2009). More attention should be paid to the
toxicological impact of mycotoxin mixtures as well as other Fusar-
ium mycotoxins, including culmorin compounds, fusarin C, fusaro-
chromanone, chlamydosporol, ENNs, butenolide, and wortmannin,
as there is little available data on their toxicity and modes of action
(Gutleb et al., 2002).
3.4. Patulin
Considering the level of 50 lg/kg of JECFA, the provided occur-
rence data for apple juice and apple products rarely show higher
levels of contamination, leading to the general conclusion that
products circulating in EU are of good sanitary quality with respect
to PAT contamination (SCOOP report 2002b, Table 11). Regarding
juices from fruits other than apple, both the percentage of positive
samples and mean levels are lower than for apple juice.
3.5. Alternaria toxins
The information on occurrence of Alternaria toxins in food sam-
ples is scarce. According to EFSA’s Scientific Opinion (2011b), AOH,
AME, TeA, and TEN are generally found in certain grains and grain-
based products, tomato and tomato products, sunflower seeds and
sunflower oil, fruits and fruit products including fruit juices, and in
beer and wine. The highest concentrations for AOH, AME, TeA, and
TEN were found in sunflower seeds. Mean concentrations of AOH
in this food group were in the range of 22–26 lg/kg with a maxi-
mum value of 1200 lg/kg. For AME, the mean values were in the
range 11–12 lg/kg, with a maximum value of 440 lg/kg. TeA
was present at higher concentrations (mean 333–349 lg/kg; max-
imum 5400 lg/kg). Mean concentrations of TEN ranged between
47 and 50 lg/kg, with a maximum value of 880 lg/kg.
4. Toxicological aspects of the main mycotoxins
4.1. Aflatoxins
AFs have shown to be extremely potent carcinogens in all
animal species investigated, i.e. mice, rats, hamsters, fish, ducks,
and monkeys, and in several organs, the liver being the primary
target. Furthermore, AFs are genotoxic compounds. Toxicological
studies led the International Agency for Research on Cancer (IARC)
to consider there was sufficient evidence in humans for the
carcinogenicity of the main AFs (AFB1, AFB2, AFG1, and AFG2) as
to be classified in Group 1 (Table 12). On the other hand, there is
sufficient evidence in experimental animals regarding the
carcinogenicity of AFM1 (its carcinogenic potency has been
estimated to be approximately 10 times lower than AFB1), being
classified as possibly carcinogenic to humans (Group 2B) (IARC,
1993, 2002).
Toxicity of AFs must be distinguished between acute and
chronic. Currently, there is very low incidence of acute AF toxicity
in humans. Acute poisoning occurs when the food is contaminated
with high concentrations of AFs. This occurs sporadically in devel-
oping countries, as happened during the severe acute human afla-
toxicosis outbreak in Kenya in 2004, where 317 cases and 125
deaths (39.4% mortality) were reported (Lewis et al., 2005; Probst

Table 12
IARC classification of the main mycotoxins.

IARC number Definition Mycotoxins

1 The mycotoxin is carcinogenic to humans AFs
2A The mycotoxin is probably carcinogenic to humans –
2B The mycotoxin is possibly carcinogenic to humans AFM1, FBs, OTA, sterigmatocystin
3 The mycotoxin is not classifiable as to its carcinogenicity to humans DON, NIV, PAT, T-2/HT-2, ZEN, citrinin, fusarenon-X
4 The mycotoxin is probably not carcinogenic to humans –
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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
et al., 2007). In this case, corn for human consumption was con-
taminated with AFs levels ranging from 20 to 8000 lg/kg. Acute
human poisoning is characterized by vomiting, abdominal pain,
pulmonary or cerebral oedema, necrosis, and fatty liver. Other
symptoms include anorexia, depression, jaundice, diarrhoea, and
photosensitivity. Occurrence of acute aflatoxicosis in animals is
more common, as highly contaminated feed is more frequent and
susceptibility of livestock species varies.
Chronic toxicity is the most common form of aflatoxicosis and is
caused by the consumption of relatively small amounts of these
toxic compounds over an extended period. Toxic effects will de-
pend on factors such as age, species, gender, nutritional status,
dose, and length of exposure. In animals, symptoms can be quite
unspecific, which often complicates the diagnosis. Usually the
symptoms are related to the decrease in productive parameters,
such as reduction in weight gain, lower feed conversion, decreased
egg or milk production, and increased susceptibility to infectious
diseases. In general, birds are more sensitive than mammals, and
among the latter, ruminants are the most resistant. Fish are also
quite sensitive to the action of AFs, as shown repeatedly in epi-
sodes of aflatoxicosis in trout.
In human beings, chronic consumption of AF-contaminated
foods has been linked to various diseases:
– Liver cancer. There is evidence that hepatitis B and/or C viruses
and AFs act synergistically in the aetiology of the hepatocellular
carcinoma (HCC) (Palliyaguru and Wu, in press; Wu and
Santella, 2012). Eastern and South-Eastern Asia and Middle and
Western Africa are regions of high HCC incidence. Geographic
variations in HCC incidence might be due to geographic
differences in the prevalence of various etiological factors,
particularly chronic infection with hepatitis viruses, and dietary
exposure to AFs.
– Effects on the reproductive system. AFs exert negative repro-
ductive effects in human males, causing delayed testicular
development, testicular degeneration, decreased reproductive
potential, morphological changes, reduced size and weight of
the testes, meiotic index decrease, decline in the percentage
of live sperm, sperm with increased abnormalities, degenera-
tion of the seminiferous epithelium, and reduced plasma con-
centration of testosterone, among others (CAST, 2003).
– Effects on the immune system. AFs act as immunomodulators,
causing a decrease in resistance to secondary infections by
fungi, bacteria, and parasites. The cellular response is particu-
larly sensitive to AFs, as evidenced by decreased T or B lympho-
cyte activity, impaired macrophage/neutrophil effector
functions, modified synthesis of inflammatory cytokines,
suppressed NK cell-mediated cytolysis, induced reactivation of
chronic infection, decreased immunity to vaccination, and
impaired immune function in developing animals (Jiang et al.,
2008).
– Encephalopathy with fatty degeneration of viscera, resembling
Reye’s syndrome (Dvorácková et al., 1977). The role of AFs in
the development of Reye’s syndrome, which produces fatty
degeneration, pale and enlarged liver and kidneys, oedema
and stroke has never been unequivocally proved, despite the
frequent detection of AFs in the liver of children who have died
of this disease and, therefore, is still under discussion.
– Pulmonary interstitial fibrosis. In this case, a possible occupa-
tional risk of AF exposure via the respiratory tract is suggested
(Dvorácková and Píchová, 1986).
4.2. Ochratoxin A
Regarding OTA, to date there is inadequate evidence of carcino-
genicity in humans, but there is sufficient evidence in experimental
227
animals. Therefore, OTA has been classified as possibly carcino-
genic to humans (Group 2B) (IARC, 1993). The latest provisional
tolerable daily intakes (PTDIs) of this toxin set by the European
Food Safety Authority (EFSA, 2006a) and the Joint FAO/WHO Expert
Committee on Food Additives (JECFA, 2007) were 17 and 14 ng/kg
bw/day, respectively.
The kidney is the major target organ for OTA, recognized as a
potent nephrotoxin. In humans, OTA has been identified as the
causative agent of nephropathies, often related with urothelial
cancer of the upper urinary tract. In animals, OTA intake has also
been correlated with an increase in the incidence of testicular can-
cer. Furthermore, OTA is recognized as teratogenic, genotoxic, car-
cinogenic, and immunotoxic, but its neurotoxic effect remains
unconfirmed.
Acute toxicity studies provide very variable OTA LD50 values,
depending on the species and the administration route. Thus, dogs
and pigs seem to be very sensitive, with oral LD50 of 0.2 and
1.0 mg/kg bw, respectively. For chickens the LD50 is 3.3 mg/kg
bw, and in rats and mouse increases to 20–58 mg/kg bw. These val-
ues are frequently lower with intraperitoneal or intravenous
administration (WHO, 2001). Symptoms such as multifocal haem-
orrhages in many organs and fibrin thrombi in the spleen, the cor-
oid plexus of the brain, liver, kidney, and heart have been observed
in rats after acute intake of OTA, suggesting disseminated intravas-
cular coagulation, probably associated to the activation of intrinsic
and extrinsic coagulation systems. Other changes were hepatic and
lymphoid necrosis, enteritis (which mainly affects the jejunum)
and nephrosis (Albassam et al., 1987).
Chronic effects of OTA are of more concern. In animals, it has
been shown that after a prolonged OTA intake, a nephropathy
linked to the degeneration of the convoluted tubule of the nephron
and renal interstitial fibrosis occurs, followed by a decrease in the
thickness of the basal membrane and glomerular hyalinization. Re-
nal lesions observed in birds, pigs, and rodents are very similar. In
humans, OTA has been associated with a kidney disease typical of
the Balkans, the so-called Balkan Endemic Nephropathy (BEN),
mainly present in rural areas of countries like Croatia, Bosnia and
Herzegovina, Serbia, Romania, and Bulgaria (Pfohl-Leszkowicz
et al., 2002). BEN was first described in the late 1950s and consists
of a bilateral chronic renal failure, often associated with
urothelioma and renal carcinomas. It is characterized by tubular
dysfunction, predominantly affecting the proximal tubule. Symp-
toms of BEN usually include anaemia, proteinuria, jaundice, head-
ache, anorexia, and uraemia. Pathological symptoms are
characterized by progressive tubulointerstitial nephropathy result-
ing in tubular atrophy and periglomerular fibrosis, often followed
by aggressive malignant tumours in the upper urinary tract, a
marked reduction in kidney size, and certain changes in the renal
cortex, such as interstitial fibrosis, glomerular hyalinization, tubu-
lar epithelium degeneration, and loss of the brush border of the
renal tubules. OTA (often in co-occurrence with citrinin) has been
presumably linked to BEN (Fuchs and Peraica, 2005; Pfohl-Les-
zkowicz et al., 2007), mainly due to the similarity between the
symptoms of this disease and those caused by OTA in porcine
nephropathy, and because high levels of OTA in food, human plas-
ma, and urine have been detected in sites where the disease is en-
demic. Therefore, OTA is a primary target of current toxicological
investigations, but although the evidence on OTA exposure is
potentially consistent, currently it is insufficient to make a BEN-
OTA association (Stefanovic et al., 2011).
Besides renal symptoms, OTA may affect other body systems. It
can cross the placenta and has been found to be embryotoxic in
rats and mice. animal studies have shown that OTA is immunotox-
ic; the immunosuppressant activity of OTA in animals is character-
ized by size reduction of the thymus, spleen, and lymph nodes,
depression of antibody responses, changes in immune cells
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228 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
number and function, and modulation of cytokine production. Be-
sides, it has been shown that OTA is associated with cerebellar,
hippocampal, and other adverse neurological effects. Finally, OTA
has been found in breast milk, which could represent a significant
source of exposure for infants (Hope and Hope, 2012).
4.3. Fusarium toxins
4.3.1. Fumonisins
Acute and chronic toxicity by FBs has been largely demon-
strated in several animal species, including carcinogenicity and
cardiovascular toxic effects (Gelderblom et al., 1988, 1991). FB1
is a cancer promoter, but a poor cancer initiator. It is not genotoxic
because FB1 as it does not induce unscheduled DNA synthesis in
primary rat hepatocytes (Norred et al., 1992). Based on toxicolog-
ical evidence, the IARC has classified FB1 as possibly carcinogenic
to humans (group 2B) (IARC, 2002). The JECFA evaluated FBs and
allocated a provisional maximum tolerable daily intake (PMTDI)
of 2 lg/kg bw/day to FBs on the basis of a NOEL of 0.2 mg/kg bw/
day and a safety factor of 100 (WHO, 2001). This PMTDI for FBs
has recently been confirmed by the Scientific Committee on Food
(SCF, 2003).
The available studies indicate that FBs do not possess a high
acute toxicity, although some animals, as sheep, revealed acute
toxicity that primarily affected the hepatic and renal function.
However, the tested FBs concentrations were significantly higher
than those usually found in natural feeds (45 mg total FBs/kg
bw/day, for 4 days) (Edrington et al., 1995).
The liver and kidney are the major target organs, but there is
species-, strain-, and sex-dependent differences in response to
the dose (Voss et al., 2007). It has been demonstrated that chronic
dietary exposure to FB1 (P50 ppm) is carcinogenic to rodents:
hepatocarcinogenic in male BD IX rats (Gelderblom et al., 1991)
and female B6C3F1 mice (Howard et al., 2001a), and nephrocarcin-
ogenic in male F344 rats (Howard et al., 2001b). The intestine has
also been discussed as a possible target for FBs toxicity (Bouhet and
Oswald, 2007).
In horses, consumption of FB-contaminated feeds has been rec-
ognized as a cause of an illness known as equine leukoencephalo-
malacia (ELEM). ELEM syndrome is a fatal disease that apparently
occurs only in horses and related species, characterized by the
presence of liquefactive necrotic lesions in the white matter of
the cerebrum, although the grey matter may also be involved.
The first symptoms are lethargy, blindness, and decreased feed in-
take, followed by convulsions and death after several hours or days
(Morgavi and Riley, 2007). Similarly, FB1-contaminated feeds have
been shown to be cardiotoxic and cause pulmonary oedema in
pigs, a syndrome known as porcine pulmonary oedema or PPE.
Clinical signs usually include decreased feed consumption, dysp-
noea, weakness, cyanosis, and death. At necropsy, the animals ex-
hibit varying degrees of interstitial and interlobular oedema, with
pulmonary oedema and hydrothorax (Morgavi and Riley, 2007).
Cattle and poultry are considerably less sensitive to FBs than
horses, pigs, rabbits, or laboratory rodents.
Currently, there is no direct evidence that FBs cause adverse
health effects to humans. Available studies have only shown incon-
clusive associations between FBs and human cancer. Thus, human
exposure to FBs has been associated with oesophageal cancer in
South Africa (Sydenham et al., 1990) and liver cancer in China
(Yoshizawa et al., 1994).
FBs have also been associated with neural tube defects (NTDs)
in the Mexico–Texas border (Missmer et al., 2006). In this case,
FBs exposure increased NTDs risk, proportionate to dose, up to a
threshold level, at which point foetal death may be more likely
to occur. Interestingly, administration of folate may reverse the
FBs toxic effect. Thus, FBs-contaminated corn represents a poten-
tial risk for human NTD occurrence, especially in populations with
inadequate folate intake who rely on corn-based foods (Suarez
et al., 2012).
4.3.2. Zearalenone
ZEN, and some of its related metabolites, competitively binds to
oestrogen receptors. Thus, the toxicity is associated with reproduc-
tive problems in specific animal species and possibly in humans
(Gromadzka et al., 2009; Wood, 1992). Risk assessment of ZEN per-
formed by the SCF concluded on a temporary tolerable daily intake
(TDI) of 200 ng/kg bw, based on a short-term study in pigs (SCF,
2000a), whereas a PMTDI of 500 ng/kg bw was established by JEC-
FA (EFSA, 2004b). IARC carcinogenic evaluation of ZEN concluded
that it is not classifiable regarding its carcinogenicity to humans
(Group 3) (IARC, 1993).
However, it has been demonstrated that ZEN can potentially
stimulate growth of cells with estrogenic receptors in human
mammary glands (Minervini et al., 2005). These results together
with other epidemiologic studies support the hypothesis that
ZEN may have a role in the aetiology of human breast cancer (Yu
et al., 2005). Furthermore, ZEN was found in the endometrial tissue
of women suffering from adenocarcinoma and in some of endome-
trial hyperplasia cases (Tomaszewski et al., 1998). Finally, between
1978 and 1981, it was suspected that ZEN and zearalenol were the
causative agents of an epidemic episode of precocious puberty in
Puerto Rico (Sáenz de Rodríguez et al., 1985).
Few data are available on the acute toxicity of ZEN, although it
seems to be relatively low, with oral LD50 values ranging between
2000 and 20,000 mg/kg bw, depending on the tested species
(Flannigan, 1991).
In animals, toxic effects of prolonged dietary ZEN exposure in-
clude carcinogenicity, genotoxicity, reproductive toxicity, endo-
crine effects, and immunotoxicity, pig being one of the most
susceptible animals, with NOEL values of 40 mg/kg bw/day com-
pared to the 100 mg/kg bw/day in rats.
This susceptibility has severe effects on animal production.
Thus, during pregnancy, ZEN reduces embryo survival and foetal
weight. Additionally, ZEN produces vulvar dilatation and redness,
vulvovaginitis, retention or absence of milk, and rectal prolapse.
In males, ZEN can lower testosterone levels, testes weight, and
spermatogenesis, inducing feminization and reducing the libido
(Zinedine et al., 2007).
4.3.3. Trichothecenes (T-2/HT-2, DON)
T-2 toxin is a potent inhibitor of protein synthesis and mito-
chondrial function both in vivo and in vitro, and shows immunosup-
pressive and cytotoxic effects. Moreover, it has been reported that
this toxin has extremely toxic effects on skin and mucous mem-
branes (Eriksen and Pettersson, 2004; Sudakin, 2003; Visconti,
2001; Visconti et al., 1991). Toxicity of HT-2 has been less
investigated. However, because T-2 is rapidly metabolized in vivo
to HT-2, it is widely accepted that the in vivo toxicity of T-2
includes that of HT-2; thus the JECFA, after assessing the toxic effect
of T-2 and HT-2, concluded that the toxic effects of these mycotox-
ins cannot be differentiated.
The PMTDI for the combination of these two toxins, or each one
separately, was set at 0.06 lg/kg bw/day (JECFA, 2001); later the
TDI was set at 0.1 lg/kg bw/day by the expert panel of the CON-
TAM group of EFSA (EFSA, 2011c). The IARC evaluated the experi-
mental data on the carcinogenicity of T-2 toxin and classified it
in Group 3 (IARC, 1993).
Concerning DON, a temporary TDI of 1 lg/kg bw was estab-
lished by the EU Scientific Committee on Food (SCF, 2002), in line
with the temporary TDI established by the Nordic Group (Eriksen
and Alexander, 1998), and the PMTDI established by JECFA
(WHO, 2001). There is no experimental or epidemiological
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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
evidence for mutagenic and/or carcinogenic properties of DON and
it was classified by the IARC in Group 3 (IARC, 1993).
Prolonged exposure to trichothecenes by humans (mainly T-2
toxin) causes a disease known as alimentary toxic aleukia (ATA),
a disease characterized by the gradual appearance of the symp-
toms described below (Joffe, 1971):
– A few hours after the consumption of a contaminated food, a
burning sensation in the mouth and pharynx occurs, followed
later by the same symptoms in the oesophagus and the
stomach.
– After 1–3 days, the patient suffers gastric and intestinal swell-
ing, accompanied by diarrhoea, nausea, abdominal pain and
vomiting. In many cases, excessive salivation, headache, dizzi-
ness, weakness, fatigue, tachycardia, fever, and sweating occurs.
Gastroenteritis lasts for approximately nine days.
– After the gastroenteritis, an asymptomatic destruction of the
bone marrow begins, causing a progressive leukopenia, lym-
phocytosis, and granulopenia.
– Post-atrophy of the bone marrow, the immunosuppression
leads to sepsis and high fever. Local subcutaneous haemorrhag-
es appear mainly in the chest and other parts of the body, fol-
lowed by skin and muscle necrosis. At first, the petechiae are
localized in small areas, later spreading and becoming more
numerous. In severe cases, an intensive ulceration and gangrene
in the larynx develops, leading to aphonia and death by
asphyxia. Meanwhile, affected individuals have severe haemor-
rhages in the nasal, oral, gastric, and intestinal mucosa.
– Finally, bronchial pneumonia can develop that can be accompa-
nied by lung bleeding, and often the death of the patient.
Individuals affected by this disease require a prolonged conva-
lescence, needing at least 2 months to recover normal bone mar-
row function.
In pigs, T-2 toxin causes a decrease in feed intake and weight
gain, as well as the appearance of oral lesions. In dairy cows, rejec-
tion of food, low milk production, gastroenteritis, intestinal bleed-
ing are also observed and, in some cases, death. In both types of
animals, this toxin can cause immunosuppression (Gimeno and
Martins, 2011).
Although DON is not as toxic as other trichothecenes such as T-
2 or HT-2, it is one of the most common contaminants of cereals
worldwide. Acute effects of food poisoning in humans are abdom-
inal pains, dizziness, headache, throat irritation, nausea, vomiting,
diarrhoea, and bloody stools (Rotter et al., 1996).
Chronic administration of DON in animals, causes weight loss,
anorexia, and decreased nutritional efficiency. In pigs, the most
studied species, 1–2 ppm causes partial rejection of feed, while
12 ppm leads to total refusal. DON administration also results in
an increase of liver size, decline in serum proteins and albumin lev-
els, as well as a decrease in the hematocrit, and a reduction of ser-
um calcium and phosphorus. Other symptoms are the reduction of
thyroid size, increased levels of thyroxin, and immune disorders,
such as immunosuppression or immunostimulation (depending
of the dosage and exposure frequency), and increased susceptibil-
ity to facultative pathogens. In pigs, DON has also a neurotoxic
effect that produces an anorexic syndrome, by altering the concen-
tration of neurotransmitters in the hypothalamus, cerebellum, and
frontal cortex. It also increases serotonin, but has no effect on
norepinephrine and dopamine levels (Lori and Rizzo, 2007).
4.4. Patulin
Acute effects of PAT include nausea, vomiting and other gastro-
intestinal symptoms (gastric ulcers, intestinal haemorrhages, and
lesions in the duodenum), that are accompanied by kidney damage
229
(Speijers et al., 1988). It has been observed that at high concentra-
tions PAT has immunotoxic properties. In rodents, the oral LD50 of
PAT ranges from 29 to 55 mg/kg bw. Poultry seem less sensitive,
with an oral LD50 of 170 mg/kg bw. When administered by intrave-
nous, intraperitoneal, or subcutaneous routes, PAT is 3–6-times
more toxic (Puel et al., 2010).
In long-term studies with animals, PAT has been reported to be
mutagenic and to cause neurotoxic, immunotoxic, genotoxic, and
gastrointestinal effects in rodents (Hopkins, 1993). It has been also
demonstrated that PAT alters the intestinal barrier function (Mah-
foud et al., 2002).
There is some concern that similar effects may occur in humans
through chronic consumption of foods and beverages contami-
nated by this mycotoxin. As there is inadequate evidence for the
carcinogenicity of PAT in experimental animals, the IARC has clas-
sified PAT in Group 3 (IARC, 1993). Based on a non-observed effect
level (NOEL) of 43 lg/kg bw/day and a safety factor of 100, and due
to its toxicity, the JECFA established a PMTDI of 0.4 lg/kg bw/day
(WHO, 1995).
5. Exposure assessment to mycotoxins
Exposure assessment is one of the four steps included in the risk
assessment process. The process starts with two preliminary steps
known as hazard identification and hazard characterization, and
ends with risk characterisation. Although several scientific reports
have been published in order to propose the best methodologies
for the exposure assessment framework, to date harmonisation is
far from being achieved.
5.1. Methodological issues
A common approach to estimate exposure is the combination of
contamination data with consumption data. While contamination
data are often provided by researchers, consumption data is in
most of cases taken from national dietary surveys. These surveys
are designed mainly to assess the nutritional status of the popula-
tions concerning energy and/or nutrients. Thus, an important lim-
itation derived from the use of the national surveys is the low
specificity of several food categories susceptible to contamination
by mycotoxins. Concerning this limitation, large food consumption
information was collected by EFSA over the last 5 years at an
increasing level of detail. In 2008, following recommendations is-
sued by EFSA’s Scientific Committee (2005b), EFSA created the
‘‘Concise European Food Consumption Database’’ (EFSA, 2008),
subsequently improved by the Comprehensive European Food
Consumption Database to be used for detailed exposure calcula-
tions in 22 different Member States (EFSA, 2011d). Forty-three
per cent (43%) of the studies included in the database were carried
out by means of food records and 53% by 24-h recall (only one
study through a 48-h recall). Most of these studies (73%) were per-
formed at individual level and 27% at household level. The 24 h
duplicate diet has been proposed as the gold standard method,
increasing precision at the expense of loss of representativeness.
The 24 h duplicate diet approach was only used in two studies to
assess the exposure of children and adults to several mycotoxins
(Bakker et al., 2009; Sizoo and Van Egmond, 2005). Several studies
included a specific survey to assess the dietary habits concerning
foods susceptible to mycotoxin contamination, stressing the draw-
backs to obtain accurate consumption information of the target
population. For example, a food frequency questionnaire was spe-
cifically designed in Spain for the Catalan population, including
those foods consumed in the region and associated to mycotoxin
contamination. The main reason was that consumption data were
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230 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237

not available to run the exposure models for this population (Cano-
Sancho et al., 2011, 2012b).
Children and adults are the main population groups considered
in mycotoxin exposure studies, but also to assess the exposure to
other pollutants. However, there is not harmonisation concerning
the age ranges to classify the sub-populations such as infants, tod-
dlers, young children or older children, and adolescents. Exposure
assessment studies for all age groups have been reviewed in this
section, but only those studies including infants, children, and/or
adolescents have been highlighted because they represented the
main risk groups and the most vulnerable population. In some
cases, the exposure of adults was assessed with foods not con-
sumed by children, for example, beer or coffee intake can contrib-
ute significantly to global FB and OTA exposure. Celiac sufferers are
also suspected to be exposed to higher amounts of FBs because
most gluten-free cereal-based foods replace wheat flour by corn
flour. Different results have been found in Italy and Catalonia. In
the Italian study, celiac sufferers were exposed to higher levels of
FBs than the rest of population, in Catalonia the estimations for ce-
liac population were the lowest of the study (Cano-Sancho et al.,
2011, 2012c; Dall’Asta et al., 2012).
Regarding the collection of contamination data, EFSA supported
the total diet studies (TDS) to provide the most accurate estimates
of mean contamination by the chemicals in the food consumed by
the population or collective group of individuals. The aims of TDS
are: (1) to be representative of the whole diet, (2) to analyse com-
posite samples pooled from a variable number of individual sam-
ples, and (3) to analyse samples from foods commonly consumed
by the target population (EFSA, 2011e). Few TDS have been de-
signed to assess mycotoxin exposure in the population. One of
the pioneering studies was the First French Total Diet Study, which
included AFs (AFB1, B2, G1, and G2) and AFM1, OTA, PAT, trichothec-
enes (diacetoxyscirpenol, monoacetoxyscirpenol, neosolaniol,
HT-2, T-2, T2 triol, 3-acetyldeoxynivalenol (3-Ac-DON), 15-acety-
ldeoxynivalenol (15-Ac-DON), DON, fusarenon X, nivalenol (NIV),
ZEN, and FBs (FB1 and FB2) (Leblanc et al., 2005). Recently, the
study has been performed again to re-evaluate the exposure levels
of the French population, improving the sampling design based in
core foods and including several emergent mycotoxins and metab-
olites (Sirot et al., 2013; Sirot et al., 2009). Another study based on
TDS has been carried out in Catalonia to assess the exposure of the
most relevant groups to major mycotoxins (Cano-Sancho et al.,
2012a,b,c).
Management of left-censored contamination data is a main
source of uncertainty of exposure models (EFSA, 2006b). Despite
scientific reports methods other than substitution are recom-
mended, sophisticated methods such as Kaplan–Meier method or
Maximum Likelihood estimation method, have not been yet ex-
tended to mycotoxin risk assessments studies. Substitution meth-
ods replacing non-detects by limit of detection (LOD) divided by 2
or producing an upper and lower bound by substitution of non-de-
tects by LOD or 0, respectively, are the most common approaches
in mycotoxin studies (EFSA, 2010).
Contamination and consumption datasets have been often inte-
grated through deterministic methods; however, various authors
have applied information technology on stochastic or probabilistic
models to draw more sophisticated exposure scenarios. One of the
main drawbacks of the deterministic approach is that it does not
allow calculating complicated statistics such as high quantiles.
Defining high-level consumers is crucial for the outcome of risk
assessment. In practice, it determines the proportion of the popu-
lation that would exceed a health-based limit. The reliability of
high percentiles is related to the number of subjects included in
the calculation, not statistically robust with a limited number of
subjects. Therefore, when refinements are required, simulation
methods are proposed as the best approach, particularly for high
quantiles. Furthermore, simulation methods allow to know the
validity and accuracy of the high quantiles estimated for the simu-
lation method through the confidence interval. Gauchi and Leblanc
(2002) reported several stochastic approaches to quantitatively as-
sess the exposure to food chemicals, through the example of the
important mycotoxin OTA among the French population. The

Table 13
Estimates of dietary intake of aflatoxins in children and infants.

Country Consumption data Contamination data Population group Exposure estimates References
ng/kg bw/day
France National Survey N = 1018 TDS-1 Children 0.323–0.89 Leblanc
N = 456 et al. (2005)
(composite)
France National Survey N = 1444 TDS-2 ‘‘core Children 0.001–0.01 Sirot et al.
foods’’ (2013)
N = 457
(composite)
Greece Assumption Breakfast Children 0.07–10.75 Villa and
cereals Markaki
(2009)
Japan National Survey N = 334 Young 0.006–0.007# Kumagai et al.
children 0.005–0.006# (2008)
Older
children
Japan National N = 884 Young 0.013–0.014# Sugita-Konishi et al.
Survey 2005 children 0.011–0.012# (2010)
N = 17,827 Older
children
The Netherlands N = 123 Duplicate 24 h Children <0.02–0.44 Bakker et al.
diet samples (2009)
N = 123
Spain NA Infant formula Infants 0.08–37.47 Hernández-Martínez
and
Navarro-Blasco
(2010)
Catalonia (Spain) FFQ TDS N = 603 Adolescents 0.19–1.31# Cano-Sancho et al.
N = 1387 Children 0.03–0.34# (2013)
#
Stochastic approach to data. NA – Data no available.
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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237 231

comparison of the results from deterministic with probabilistic
models did not elucidate large differences or extreme estimations,
but in several cases provided confidence intervals, uncertainty and
variability estimations.
5.2. Exposure to mycotoxins
There are many publications on OTA, FBs, DON, AFs, and ZEN in
comparison to other mycotoxins. There are fewer studies on PAT or
AFM1, and limited data exists on the exposure to T-2 and HT-2,
NIV, or the emergent mycotoxins.
Despite the wide range of studies published during the last
three decades reporting the occurrence of AFs worldwide, not
much data are available about the exposure of populations to
this carcinogenic toxin. One of the common limitations is the
large percentage of left-censored data from occurrence studies,
which make the exposure estimation inaccurate and unreliable.
This fact is notorious when substitution of censoring is applied
and accordingly, large uncertainty intervals are derived. Another
common issue is the heterogeneity of AF contamination in the
samples. While most samples and sub-samples can present
undetectable levels of AFs, a reduced number can present very
high levels of contamination. Estimates for China, Korea,
Malaysia, and South Africa are high, with maximum levels of in-
take for total population of 2.69, 5.79, 8.89, and 133 ng/kg bw/
day (data not shown in tables) (Leong et al., 2011; Park et al.,
2004; Shephard, 2008; Sun et al., 2011). In European countries
and Japan, the mean estimates were in general below 1 ng/kg
bw/day, with the exception of Greece were large exposure was
derived from a worst case estimate (Table 13). As there is not
a tolerable daily intake for those substances that are both geno-
toxic and carcinogenic, the risk characterization approach recom-
mended by EFSA is based on the building of the margins of
exposure computed by division of a benchmark dose by the
exposure estimates. In the case of AFM1, only the studies from
France and Spain were found to draw reliable scenarios of expo-
sure. The dietary intake of AFM1 in Catalonia was similar to the
one found in the First French TDS, but higher than the latest
French TDS. The highest levels of exposure to AFM1 were found
in the 95th percentile for children, with maximum estimates of
0.48 and 0.55 ng/kg bw/day in Catalonia and France, respectively.
Other general estimations were produced by JECFA in the five re-
gional diets, showing similar results as in the aforementioned
studies with the European diet. Lowest estimates were derived
for the African region due to a lower milk intake among their
inhabitants (Henry et al., 2001).
Exposure to OTA has been mainly estimated through the con-
sumption of target foods such as coffee and wine or cereal-based
foods, alone or in combination, in case of the adult population,
and baby food or cereal-based food in the case of children and in-
fants (Table 14). The most exposed population groups to OTA are
expected to be infants and children, but in all cases, the estimates

Table 14
Estimates of dietary intake of ochratoxin A in children and infants.

Country Consumption data Contamination data Population group Exposure estimates (ng/kg bw/day) Reference
Canada National Secondary Children 2.60–9.48# Kuiper-Goodman et al.
Survey surveys* Infants 1.93–10.43# (2010)
(1994–1996, 1993–2006
1998) 12 food categories
N = 20,000
France National TDS N = 474 Children 0.23–5.25 Sirot et al. (2013)
Survey
N = 1444
France National TDS N = 343 Children 2.16–7.77 Leblanc et al. (2005)
Survey
N = 1018
Greece Assumptions Breakfast cereals Children 0.07–2.17 Villa and
(30–50 g/day) Markaki (2009)
a
Italy National N = 300 Cocoa, Adolescents 0.56–1.00 Brera et al. (2011)
Food chocolate
Consumption
Survey
N = 3323
Childrena 1.16–1.71
Infantsa 2.08–2.42
Japan National N = 2061 Young 2.21 1.51 Sugita-Konishi et al.
Health and (2004–2010) children (2012)
Nutrition Old children
Survey
(2007–2010)
N = 17,827
Morocco JECFA* N = 70 NA 235.7 Serrano et al.
(2012a)
The Netherlands N = 123 Duplicate 24 h Children 4.1 Bakker et al.
diet samples N = 123 (2009)
Catalonia (Spain) FFQ N = 1348 N = 784 Children Infants 0.09–0.98# Coronel et al.
(2012)
0.28–7.23#
Sweden FFQ N = 200 N = 158 Children 1.2.22 Thuvander et al.
National (2001)
Survey HULK
Turkey Assumption Baby food Infants 1.7 Kabak (2009)
RDI 40 g/day (N = 24)
UK N = 50 Duplicate NA 0.26–3.54 Gilbert et al. (2001)
N = 50  30 days
a
Weekly intake, Consumers.
#
Stochastic approach to data. NA – Data no available.
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232 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237

Table 15
Estimates of dietary intake of fumonisins in children and infants.

Country Consumption data Contamination Population group Exposure Reference

data estimates
(ng/kg bw/day)
Argentina Pacin et al. N = 38 Total FBs 15–25 1–5 3100–3500 11,300 Solovey et al. (1999)
(1998)
Brasil Household N = 208 Consumers 3180–7100 Caldas and Silva (2007)
Budget
Survey ND
France National TDS Children 15.4–106 Sirot et al. (2013)
Survey composite
N = 1444 N = 56
France National TDS Children 46–175 Leblanc et al. (2005)
Survey composite
N = 1018 N = 34
France NA NA All children 445.1 SCOOP (2003)
Italy SCOOP (2003) N = 82 Infants 5 D’Arco et al. (2009)
Japan National Survey N = 1505 Children 1.2–10.2 Sugita-Konishi et al. (2012)
N = 17,827
The Netherlands N = 123 Duplicate 24 h Children 28 Bakker et al. (2009)
diet samples
N = 123
Nordic Countries National N = 70 (1996) Children 400 Petersen and Thorup (2001)
Survey
N = 3098
Norway NA NA Infants 860 SCOOP (2003)
Catalonia (Spain) FFQ N = 928 TDS N = 370 Children Infants 85.30–293.66# Cano-Sancho et al. (2012)
composites 195.19–508.20#
Spain National survey N = 104 Baby 1.7–720 D’Arco et al. (2009)
Tanzania 24-h recall N = 273 (563) Kimanya et al. Infants 110–9090# Kimanya et al. (2007)
(2008)
#
Stochastic approach to data. NA – Data no available.

Table 16
Estimates of dietary intake of zearalenone in children and infants.

Country Consumption data Contamination Population group Exposure Estimates (ng/kg bw/ Reference
data day)
Canada NA NA NA 50–100 Kuiper-Goodman et al.
(1987)
France NA 7d record Children 42–84 SCOOP (2003)
France National Survey N = 245 Children 66–132 Leblanc et al. (2005)
N = 1018
France National Survey N = 339 Children 12–88 Sirot et al. (2013)
N = 1444
Germany Estimate NA Infants Children 17–62 SCOOP (2003)
The 2-day diet diary NA Children 46–50 SCOOP (2003)
Netherlands
Norway Q-FFQ NA Infants 12–1508 SCOOP (2003)
Portugal Food Balance Sheets NA Adults 14 SCOOP (2003)
Spain FFQ N = 485 composites Children Infants and toddler 2–17 12–52 Cano-Sancho et al. (2012a)
UK 7 day weighted record NA Infant Toddler Young 50 SCOOP (2003)
people
54
21–55
*
Assumption from the authors to normalise the information.
NA – Data no available.

were below the TDI of 17 ng/kg bw/day set by EFSA (2006a). Max-
imum estimates were found in infants from Catalonia and children
from France, with 95th percentile values of 7.2 and 7.8 ng/kg bw/
day, respectively, or 10.4 ng/kg bw/day for Canadian infants
(Coronel et al., 2012; Kuiper-Goodman et al., 2010; Leblanc et al.,
2005). In one study an extremely high estimation for the Moroccan
population was reported, reaching values of 236 ng/kg bw/day
(Serrano et al., 2012b).
Corn-based food is the primary category of election to
assess the exposure to fumonisins, including cornflakes, corn snacks,
or beer in the case of adults. Children and infants are the main risk
groups found in the studies, although immigrants living in Catalonia
were found to be a risk group (Cano-Sancho et al., 2012c). Celiac suf-
ferers were found to be more exposed than the rest of population in
Italy (Dall’Asta et al., 2012), but in Spain this group was expected to
be exposed to lower levels of these toxins than the other groups
(Cano-Sancho et al., 2012c). The highest estimations were deter-
mined in Brazil, Tanzania, Guatemala, South Africa, or Argentina,
exceeding the TDI of 2000 ng/kg bw/day (SCF, 2003). Lowest estima-
tions were found in Nordic European countries and Japan (Table 15).
Dietary intake of ZEN was also reported in the huge study of the
SCOOP (2003), along with other Fusarium mycotoxins, but few
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S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237 233

Table 17
Estimates of dietary intake of deoxynivalenol in children and infants.

Country Consumption data Contamination data Population Exposure estimates (lg/kg bw/ Reference
group day)
China NA NA Children 1.5–4.0 Wang et al. (2010)
China National Survey N = 74 NA 5.5 Fan et al. (2009)
France NA NA Children 0.73–1.51 SCOOP (2003)
France National Survey N = 1018 N = 238 composites TDS Children 0.45–0.93 Leblanc et al. (2005)

France National Survey N = 1444 N = 258 TDS core foods Children 0.54–1.03 Sirot et al. (2013)
Germany NA NA Young child 0.96–1.92 SCOOP (2003)
Germany NA NA Infants 0.51–1.02 SCOOP (2003)
India National Survey N = 100 (wheat, maize and barley NA 0.19–0.77 Mishra et al. (2013)
The Netherlands NA NA Infants 0.85 SCOOP (2003)
The Netherlands NA NA Infants 0.76 SCOOP (2003)
The Netherlands National Survey N = 6250 N = 2423 Children 0.46–1.0 Pieters et al. (2004)
The Netherlands N = 123 Duplicate 24 h diet samples N = 123 Children 0.29 Bakker et al. (2009)
Norway NA NA Infants 0.11–0.834 SCOOP (2003)
South Africa National survey N = 80 Children Infants 0.10–2.63 Shephard et al. (2010)
0.12–3.67
South Korea NA N = 689 0.07–0.14# Ok et al. (2009)
Catalonia FFQ N = 1390 N = 556 composite TDS Children Infants 0.68–2.49# Cano-Sancho et al. (2011)
(Spain) 0.90–3.57#
Spain Commercial recommendations N = 175 Children 0.18 Castillo et al. (2008)
UK NA NA Toddlers 0.48 SCOOP (2003)
UK NA NA Infants 0.37 SCOOP (2003)
#
Stochastic approach to data. NA – Data no available.

independent studies on ZEN exposure have been published to date
(Table 16). In line with the exposure profiles of the previous myco-
toxins, infants and children were the most exposed population
groups, but in most cases, including the highest percentiles, they
were considerably far from the TDI of 250 ng/kg bw/day (EFSA,
2011a). The maximum values were found in children from France
and Norway, 87.5 and 1508 ng/kg bw/day, respectively (Leblanc
et al., 2005).
DON is the most studied of the trichothecenes and little infor-
mation is available on NIV, 3-Ac-DON, 15-AC-DON, or T-2 and
HT-2 toxins. Exposure to DON is generalized around temperate
countries (Table 17). On one hand, trichothecenes-producing fungi
can grow and contaminate easily under these conditions, and on
the other hand, wheat (major substrate for these moulds) is widely
consumed by populations from Mediterranean countries. Overall,
infants and children were the most exposed population groups,
but high estimates were also obtained within the immigrant pop-
ulation (North Africans, South Americans, and Eastern Europeans)
in Spain. Although mean exposures have been estimated to be be-
low the 1 lg/kg bw/day TDI (SCF, 1999), the high percentiles can
reach and exceed this level in countries like Spain, France, or Ger-
many. The highest values were estimated for South African chil-
dren and young children (Shephard et al., 2010). The role of
global exposure to DON derivatives from food processing was
emphasized by JECFA (2011). However, Sirot et al. (2013) found
that the contribution of 3-Ac-DON and 15-Ac-DON represents only
0.1% of the mean lower bound exposure to DON and about 10% of
the upper bound. T2 and HT2 toxins have been recently evaluated
in France and Catalonia showing moderate exposure estimates for
their population. The mean exposure to f T-2 plus HT-2 in adults
and children from Catalonia ranged between 18–41 and 41–
91 ng/kg bw/day, respectively. The 95th percentile determined
for children ranged between 86 and 206 ng/kg bw/day depending
on the censoring management (Cano-Sancho et al., 2012b). In the
Second French TDS, the range of exposure was 8.94–51.8 and
14.5–91.1 ng/kg bw/day for the mean estimates in adults and chil-
dren, respectively. The 95th percentile for children was bounded
between 13.3 and 176.8 ng/kg bw/day, elucidating the large uncer-
tainty of the estimations with a wide range of censored data (Sirot
et al., 2013). Accordingly, children from France and Catalonia could
occasionally exceed the TDI of 100 ng/kg bw/day set by the EFSA
(2011c). Several estimations published in the JECFA monograph
(JECFA, 2011), reported lowest estimates for children in the UK
and Norway, where no risk was expected from their consumption.
EFSA estimated the total chronic dietary exposure to T-2 plus HT-2
toxins in toddlers across 14 European countries to be between 12
and 43 ng/kg bw/day for mean consumers and between 23 and
91 ng/kg bw/day for 95th percentile consumers (EFSA, 2011c).
The level of NIV intake was evaluated among the French popula-
tion through the First and Second TDS. The estimates for children
ranged between 163 and 300 ng/kg bw/day in the first TDS and be-
tween 31 and 119 in the second. Lower amounts were found for
adults with estimations between 88 and 157 ng/kg bw/day and
20–67 for the first and second TDS, respectively (Leblanc et al.,
2005; Sirot et al., 2013). In all cases, the estimations were far from
the 700 ng/kg bw/day TDI (SCF, 2002).
Estimated daily intakes of the emergent mycotoxins ENN, BEA,
and FUS were reported for infants in Spain through infant formu-
las. The mean levels for the sum of ENNs and FUS were 234.6
and 1.6 lg/kg bw/day, respectively. The authors could not assess
the risk for this group due to the lack of a safety level established
by JECFA or EFSA (Serrano et al., 2012b).
PAT is a mycotoxin mainly found in fruit-based foods. The cat-
egories commonly selected to assess the exposure are those con-
taining apple, such as apple juice or apple compote consumed by
children and infants (Table 18). Exposure assessment studies
showed high estimates for these vulnerable groups but in all cases
below the 400 ng/kg bw/day TDI (SCF, 2000b).
A challenge in the field of exposure assessment of mycotoxins is
to develop accurate and reliable biomarkers for human studies. The
biomarker approach may be a promising tool for directly measur-
ing toxin-mediated biological perturbations or to directly measure
the amounts of mycotoxins present in the organism. Biological
markers of aflatoxin, ochratoxin A, and fumonisin exposure have
attracted the attention for mycotoxin biomonitoring studies. How-
ever, while AFs and OTA biomarkers have been successfully applied
and validated over the last decade, large drawbacks remain to find
a suitable FB biomarker. The first studies in which biomarkers
where used to determine human exposure to food pollutants in-
volved AFB1. In these studies, correlation between AFB1 intake
and urinary AFM1 excretion was statistically achieved. Next, albu-
min bounded AFB1 and AFB1-DNA adducts in urine were explored
 Author's personal copy
234 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
Table 18
Estimates of dietary intake of patulin.
Country Consumption data Contamination data
Belgium FFQ 2095 children N = 177 apple juice
Italy NA N = 169
France NA N = 20 composites
France NA N = 83 composites
The Netherlands National survey N = 1802 NA
Catalonia (Spain) N = 206 (901) N = 384
Spain 200 ml/day N = 100 apple juice
130 ml/day
Assumption
South Africa National survey Apple juice N = 30
Sweden National survey HULK N = 100
a
Total intake if the consumption of juice consists of peach/pear juice.
b
Total intake if the consumption of juice consists of apple juice.
#
Stochastic approach to data. NA – Data no available.
for exposure assessment studies. The use of urinary AFB1-N7-guan-
ine adduct, primary DNA adduct of AFB1, provided a suitable mea-
sure of acute exposure to AFB1, and it has been validated for human
samples and laboratory models. Moreover, the long half-life of
AFB1-albumin adducts provides a measure of chronic exposure to
this toxin, also validated in human models (Crews et al., 2001). Di-
rect measure of OTA or the OTA metabolite OT-a in plasma or urine
has been shown to be a suitable indicator of exposure to the myco-
toxin (Coronel et al., 2010). Several factors such as age, gender, sea-
son, and geographic location have been shown to influence OTA
plasma levels. The main limitation to develop a FBs biomarker is
the short half-life of the mycotoxin in the organism. Due to the ra-
pid elimination and low bioavailability of FBs, an indirect indicator
of human exposure to these toxins is required. Based on the ability
of FB1 to inhibit ceramide synthase activity, the accumulation of
sphinganine (Sa) over sphingosine (So) (known as the Sa:So ratio)
has been proposed as an exposure biomarker. However, although it
has been reported to be a good indicator of FBs exposure in animals
and in vitro models, it has not yet been validated for humans
(Cano-Sancho et al., 2010). Several studies have evaluated DON
and DON-glucuronide content in urine to assess their applicability
as a biological indicator of DON exposure. Interesting correlations
among estimates of DON dietary intakes and DON and DON-glucu-
ronide levels in urine have been found, although some uncertain-
ties need to be solved to fully validate the method as a reliable
biomarker (Turner et al., 2012, 2011a, 2011b, 2008). DON-glucu-
ronides, such as DON-3-glucuronide and DON-15-glucuronide,
have been reported to be the primary urine metabolites in Austrian
people (Warth et al., 2012a). In the study, a high number of inhab-
itants were found to exceed the TDI even after consuming food
with DON contamination levels below EU limits. Recently, multi-
detection methods to assess the simultaneous level of a wide range
of mycotoxins and metabolites in biological matrices have been
proposed as promising multi-biomonitoring tools (Solfrizzo et al.,
2011; Warth et al., 2012b). Suitable performance characteristics
have been reached improving extraction and detection methods,
mainly based on LC–ESI-MS/MS chromatography. Despite the good
sensitivity, inter-individual variability needs to be characterized to
fully validate the method at individual level.
Conflict of Interest
The authors declare there are no conflicts of interest.
Acknowledgement
The authors are grateful to the Spanish government (projects
AGL2010-22182-C04-04 and AGL2011-24862), the Catalonian
Food Safety Agency of the ‘Generalitat de Catalunya’ Health Depart-
Population group Exposure estimates (ng/kg bw/day) Reference
#
Children 9–72 Baert et al. (2007)
Adolescents Children 1–4 3–14 Piemontese et al. (2005)
Children 29–106 Leblanc et al. (2005)
Children 1–97 Sirot et al. (2013)
Infants 17–307 Brandon et al. (2012)
Infants 8–22# Cano-Sancho et al. (2009)
Children Infants 155 Murillo-Arbizu et al. (2009)
252
Young children Infants 1–37 Shephard et al. (2010)
1–33
Children 5a–24b Thuvander et al. (2001)
ment, and the European Union (MYCORED KBBE-2007-2-5-05 pro-
ject) for funding.
References
Albassam, M.A., Yong, S.I., Bhatnagar, R., Sharma, A.K., Prior, M.G., 1987.
Histopathologic and electron microscopic studies on the acute toxicity of
ochratoxin A in rats. Vet. Pathol. 24, 427–435.
Baert, K., De Meulenaer, B., Verdonck, F., Huybrechts, I., De Henauw, S.,
Vanrolleghem, P.A., Debevere, J., Devlieghere, F., 2007. Variability and
uncertainty assessment of patulin exposure for preschool children in
Flanders. Food Chem. Toxicol. 45, 1745–1751.
Bakker, G., Sizoo, E., Jekel, A., Pereboom-De Fauw, D.P., Schothorst, R., Van Egmond,
H., 2009. Determination of mean daily intakes of aflatoxin B1, aflatoxin M1,
ochratoxin A, trichothecenes and fumonisins in 24-hour diets of children in the
Netherlands. World Mycotoxin J. 2, 451–459.
Barkai-Golan, R., 2008. Alternaria mycotoxins. In: Barkai- Golan, R., Nachman, P.
(Eds.), Mycotoxins in Fruits and Vegetables. Academic Press, San Diego, CA, USA,
pp. 185–203.
Bottalico, A., Logrieco, A., 1998. Toxigenic Alternaria species of economic
importance. In: Sinha, K.K., Bhatnager, D. (Eds.), Mycotoxins in Agriculture
and Food Safety. Marcel Dekker, New York, pp. 65–108.
Boudra, H., Le Bars, P., Le Bars, J., 1995. Thermostability of ochratoxin A in wheat
under two moisture conditions. Appl. Environ. Microbiol. 61, 1156–1158.
Bouhet, S., Oswald, I.P., 2007. The intestine as a possible target for fumonisin
toxicity. Mol. Nutr. Food Res. 51, 925–931.
Brandon, E.F.A., Baars, A.J., Biesebeek, J.D.T., Oomen, A.G., Bakker, M.I., De Heer, C.,
2012. Risk assessment of patulin intake from apple containing products by
young children. World Mycotoxin J. 5, 391–403.
Brera, C., Debegnach, F., De Santis, B., Iafrate, E., Pannunzi, E., Berdini, C., Prantera, E.,
Gregori, E., Miraglia, M., 2011. Ochratoxin A in cocoa and chocolate products
from the Italian market: occurrence and exposure assessment. Food Control 22,
1663–1667.
Bullerman, L.B., Bianchini, A., 2007. Stability of mycotoxins during food processing.
Int. J. Food Microbiol. 119, 140–146.
Caldas, E.D., Silva, A.C.S., 2007. Mycotoxins in corn-based food products consumed
in Brazil: an exposure assessment for fumonisins. J. Agric. Food Chem. 55, 7974–
7980.
Cano-Sancho, G., Gauchi, J.P., Sanchis, V., Marín, S., Ramos, A.J., 2011. Quantitative
dietary exposure assessment of the Catalonian population (Spain) to the
mycotoxin deoxynivalenol. Food Addit. Contam. A 28, 1098–1109.
Cano-Sancho, G., Marin, S., Ramos, A.J., Sanchis, V., 2009. Survey of patulin
occurrence in apple juice and apple products in Catalonia, Spain, and an
estimate of dietary intake. Food Addit. Contam.: B 2, 59–65.
Cano-Sancho, G., Marin, S., Ramos, A.J., Sanchis, V., 2010. Biomonitoring of Fusarium
spp. mycotoxins: perspectives for an individual exposure assessment tool. Food
Sci. Technol. Int. 16, 266–276.
Cano-Sancho, G., Marin, S., Ramos, A.J., Sanchis, V., 2012a. Occurrence of
zearalenone, an oestrogenic mycotoxin, in Catalonia (Spain) and exposure
assessment. Food Chem. Toxicol. 50, 835–839.
Cano-Sancho, G., Marín, S., Ramos, A.J., Sanchis, V., 2012b. Exposure assessment of
T2 and HT2 toxins in Catalonia (Spain). Food Chem. Toxicol. 50, 511–517.
Cano-Sancho, G., Ramos, A.J., Marín, S., Sanchis, V., 2012c. Occurrence of fumonisins
in Catalonia (Spain) and an exposure assessment of specific population groups.
Food Addit. Contam. A 29, 799–808.
Cano-Sancho, G., Sanchis, V., Marín, S., Ramos, A.J., 2013. Occurrence and exposure
assessment of aflatoxins in Catalonia (Spain). Food Chem. Toxicol. 51, 188–193.
Castillo, M.A., Montes, R., Navarro, A., Segarra, R., Cuesta, G., Hernández, E., 2008.
Occurrence of deoxynivalenol and nivalenol in Spanish corn-based food
products. J. Food Compos. Anal. 21, 423–427.
Cazzaniga, D., Basílico, J.C., González, R.J., Torres, R.L., De Greef, D.M., 2001.
Mycotoxins inactivation by extrusion cooking of corn flour. Lett. Appl.
Microbiol. 33, 144–147.
 Author's personal copy
S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
Chen, L.Y., Tian, X.L., Yang, B., 1990. A study on the inhibition of rat myocardium
glutathione-peroxidase and glutathione-reductase by moniliformin.
Mycopathologia 110, 119–124.
Coronel, M.B., Marín, S., Cano-Sancho, G., Ramos, A.J., Sanchis, V., 2012. Exposure
assessment to ochratoxin A in Catalonia (Spain) based on the consumption of
cereals, nuts, coffee, wine, and beer. Food Addit. Contam. A 29, 979–993.
Coronel, M.B., Sanchis, V., Ramos, A.J., Marin, S., 2010. Review. Ochratoxin A:
presence in human plasma and intake estimation. Food Sci. Technol. Int. 16, 5–
18.
Council for Agricultural Science and Technology (CAST), 2003. Mycotoxins, Risks in
Plant, Animal, and Human Systems, Ames, IA.
Crews, H., Alink, G., Andersen, R., Braesco, V., Holst, B., Maiani, G., Ovesen, L., Scotter,
M., Solfrizzo, M., van den Berg, R., Verhagen, H., Williamson, G., 2001. A critical
assessment of some biomarker approaches linked with dietary intake. Brit. J.
Nutr. 86, S5–S35.
Cundliff, E., Cannon, M., Davies, J., 1974. Mechanism of inhibition of eukaryotic
protein synthesis by trichothecene fungal toxins. Proc. Natl. Acad. Sci. USA 71,
30–34.
Cundliff, E., Davies, J., 1977. Inhibition of initiation, elongation, and termination of
eukaryotic protein synthesis by trichothecene fungal toxins. Antimicrob. Agents
CH. 11, 491–499.
D’Arco, G., Fernández-Franzón, M., Font, G., Damiani, P., Mañes, J., 2009. Survey of
fumonisins B1, B2 and B3 in conventional and organic retail corn products in
Spain and Italy and estimated dietary exposure. Food Addit. Contam.: B 2, 146–
153.
Dall’Asta, C., Scarlato, A.P., Galaverna, G., Brighenti, F., Pellegrini, N., 2012. Dietary
exposure to fumonisins and evaluation of nutrient intake in a group of adult
celiac patients on a gluten-free diet. Mol. Nutr. Food Res. 56, 632–640.
Dvorácková, I., Kusák, V., Veselý, D., Veselá, J., Nesnídal, P., 1977. Aflatoxin and
encephalopathy with fatty degeneration of viscera (Reye). Ann. Nutr. Aliment.
31, 977–989.
Dvorácková, I., Píchová, V., 1986. Pulmonary interstitial fibrosis with evidence of
aflatoxin B1 in lung tissue. J. Toxicol. Environ. Health 18, 153–157.
Edrington, T.S., Kamps-Holtzapple, C.A., Harvey, R.B., Kubena, L.F., Elissalde, M.H.,
Rottinghaus, G.E., 1995. Acute hepatic and renal toxicity in lambs dosed with
fumonisin-containing culture material. J. Anim. Sci. 73, 508–515.
Eriksen, G.S., Alexander, J., 1998. Fusarium Toxins in Cereals – A Risk Assessment.
Nordic Council of Ministers, TemaNord 502, Copenhagen, Denmark.
Eriksen, G.S., Pettersson, H., 2004. Toxicological evaluation of trichothecenes in
animal feed. Anim. Feed Sci. Technol. 114, 205–239.
European Food Safety Authority (EFSA), 2004a. Opinion of the scientific panel on
contaminants in the food chain on a request from the commission related to
aflatoxin B1 as undesirable substance in animal feed. EFSA J. 39, 1–27.
European Food Safety Authority (EFSA), 2004b. Opinion of the scientific panel on
contaminants in the food chain on a request from the commission related to
zearalenone as undesirable substance in animal feed. EFSA J. 43, 1–41.
European Food Safety Authority (EFSA), 2005a. Opinion of the Scientific Panel on
Contaminants in Food Chain on a request from the Commission related to
fumonisins as undesirable substances in animal feed. EFSA J. 235, 1–32.
European Food Safety Authority (EFSA), 2005b. Opinion of the Scientific Committee
on a request from EFSA related to Exposure Assessments (adopted on 22 June
2005). EFSA J. 249, 1–26.
European Food Safety Authority (EFSA), 2006a. Opinion of the Scientific Panel on
Contaminants in the Food Chain on a request from the Commission related to
ochratoxin A in food. EFSA J. 365, 1–56.
European Food Safety Authority (EFSA), 2006b. Guidance of the Scientific
Committee on a request from EFSA related to Uncertainties in Dietary
Exposure Assessment. EFSA J. 438, 1–54.
European Food Safety Authority (EFSA), 2007. Opinion of the Scientific Panel on
contaminants in the food chain related to the potential increase of consumer
health risk by a possible increase of the existing maximum levels for aflatoxins
in almonds, hazelnuts and pistachios and derived products. EFSA J. 446, 1–127.
European Food Safety Authority (EFSA), 2008. Guidance Document for the use of the
Concise European Food Consumption Database in Exposure Assessment, 54pp.
European Food Safety Authority (EFSA), 2010. Management of left-censored data in
dietary exposure assessment of chemical substances. EFSA J. 8, 1557.
European Food Safety Authority (EFSA), 2011a. Scientific opinion on the risks for
public health related to the presence of zearalenone in food. EFSA J. 9, 2197.
European Food Safety Authority (EFSA), 2011b. Scientific opinion on the risks for
animal and public health related to the presence of Alternaria toxins in food and
feed. EFSA J. 9, 2407.
European Food Safety Authority (EFSA), 2011c. Scientific opinion on the risks for
animal and public health related to the presence of T-2 and HT-2 toxin in food
and feed. EFSA J. 9, 2481.
European Food Safety Authority (EFSA), 2011d. GUIDANCE of EFSA Use of the EFSA
Comprehensive European Food Consumption Database in Exposure
Assessment. EFSA J. 9, 2097.
European Food Safety Authority (EFSA), 2011e. Towards a harmonised Total Diet
Study approach: a guidance document. EFSA J. 9, 2450.
European Food Safety Authority (EFSA), 2012. Scientific opinion on ergot alkaloids
in food and feed. EFSA J. 10, 2798.
Fan, P., Zhang, Y., Zhou, M., Chen, C., Wang, J., 2009. Incidence of trichothecenes in
wheat-based foods from China. Int. J. Environ. Anal. Chem. 89, 269–276.
Fitzpatrick, D.W., Picken, C.A., Murphy, L.C., Buhr, M.M., 1989. Measurement of the
relative binding-affinity of zearalenone, alpha-zearalenol and beta-zearalenol
235
for uterine and oviduct estrogen-receptors in swine, rats and chickens – an
indicator of estrogenic potencies. Comp. Biochem. Phys. C 94, 691–694.
Flannigan, B., 1991. Mycotoxins. In: D’Mello, J.P.F., Duffus, C.M., Duffus, J.H. (Eds.),
Toxic Substances in Crop Plants. The Royal Society of Chemistry, Cambridge, pp.
226–257.
Fuchs, R., Peraica, M., 2005. Ochratoxin A in human kidney diseases. Food Addit.
Contam. 22 (Suppl. 1), 53–57.
Gauchi, J.P., Leblanc, J.C., 2002. Quantitative assessment of exposure to the
mycotoxin Ochratoxin A in food. Risk Analysis 22, 219–234.
Gelderblom, W.C.A., Jaskiewicz, K., Marasas, W.F.O., Thiel, P.G., Hora, R.M., Vleggar, R.,
Kriek, N.P., 1988. Fumonisins. Novel mycotoxins with cancer-promoting activity
produced by Fusarium moniliforme. Appl. Environ. Microbiol. 54, 1806–1811.
Gelderblom, W.C.A., Kriek, N.P.J., Marasas, W.F.O., Thiel, P.G., 1991. Toxicity and
carcinogenicity of the Fusarium moniliforme metabolite, fumonisin B1 in rats.
Carcinogenesis 12, 1247–1251.
Gelderblom, W.C.A., Marasas, W.F.O., Vleggaar, R., Thiel, P.G., Cawood, M.E., 1992.
Fumonisins: isolation, chemical characterization and biological effects.
Mycopathologia 117, 11–16.
Gilbert, J., Brereton, P., MacDonald, S., 2001. Assessment of dietary exposure to
ochratoxin A in the UK using a duplicate diet approach and analysis of urine and
plasma samples. Food Addit. Contam. 18, 1088–1093.
Gimeno, A., Martins, M.L., 2011. Micotoxinas y Micotoxicosis en Animales y
Humanos. Special Nutrients, Florida, pp. 50–53.
Gromadzka, K., Waśkiewicz, A., Goliński, P., Świetlik, J., 2009. Occurrence of
estrogenic mycotoxin – zearalenone in aqueous environmental samples with
various NOM content. Water Res. 43, 1051–1059.
Grove, J.F., 1988. Non-macrocycle trichothecenes. Nat. Prod. Rep. 5, 187–209.
Grove, J.F., 1993. Macrocycle trichothecenes. Nat. Prod. Rep. 10, 429–448.
Gutleb, A.C., Morrison, E., Murk, A.J., 2002. Cytotoxicity assays for mycotoxins
produced by Fusarium strains: a review. Environ. Toxicol. Pharmacol. 11, 309–
320.
Hagler, W.M., Mirocha, C.J., Pathre, S.V., Behrens, J.C., 1979. Identification of the
naturally occurring isomer of zearalenol produced by Fusarium roseum
Gibbosum in rice culture. Appl. Environ. Microbiol. 37, 849–853.
Henry, S., Whitaker, T., Rabbini, I., Bowers, J., Park, D., Price, W.D., Bosch, F.X.,
Pennington, J., Verger, P., Yoshizawa, T., 2001. Aflatoxin M1. In: Safety
evaluation of certain mycotoxins in food. Prepared by the Fifty-sixth Meeting
of the Joint FAO/WHO Expert Committee on Food Additives (JECFA), FAO Food
and Nutrition Paper. Food and Agriculture Organization of the United Nations,
Rome, Italy.
Hernández-Martínez, R., Navarro-Blasco, I., 2010. Aflatoxin levels and exposure
assessment of spanish infant cereals. Food Addit. Contam.: B 3, 275–288.
Hope, J.H., Hope, B.E., 2012. A review of the diagnosis and treatment of ochratoxin A
inhalational exposure associated with human illness and kidney disease
including focal segmental glomerulosclerosis. J. Environ. Public Health 2012.
http://dx.doi.org/10.1155/2012/835059. Article ID 835059.
Hopkins, J., 1993. The toxicological hazards of patulin. Food Chem. Toxicol. 31, 455–
456.
Howard, P.C., Eppley, R.M., Stack, M.E., Warbritton, A., Voss, K.A., Lorentzen, R.J.,
Kovach, R.M., Bucci, T.J., 2001a. Fumonisin B1 carcinogenicity in a two-year
feeding study using F344 rats and B6C3F1 mice. Environ. Health Persp. 109
(Suppl. 2), 277–282.
Howard, P.C., Warbritton, A., Voss, K.A., Lorentzen, R.J., Thurman, J.D., Kovach, R.M.,
Bucci, T.J., 2001b. Compensatory regeneration as a mechanism for renal tubule
carcinogenesis of fumonisin B1 in the F344/N/Nctr BR rat. Environ. Health
Persp. 109 (Suppl. 2), 309–314.
IARC (International Agency for Research on Cancer), 1993. IARC Monographs on the
Evaluation of Carcinogenic Risks to Humans, Some Naturally Occurring
Substances, Food Items and Constituents, Heterocyclic Aromatic Amines and
Mycotoxins, vol. 56. Lyon, International Agency for Research on Cancer.
IARC (International Agency for Research on Cancer), 2002. IARC Monographs on the
Evaluation of Carcinogenic Risks to Humans, vol. 82. Lyon, International Agency
for Research on Cancer.
JECFA (Joint FAO/WHO Expert Committee on Food Additives), 2001. Safety
Evaluation of Certain Mycotoxins in Food. Rome, Italy, Food and Agriculture
Organization, pp. 281–320.
JECFA (Joint FAO/WHO Expert Committee on Food Additives), 2007. 68th Meeting.
Evaluation of Certain Food Additives and Contaminants. WHO, Geneva. 208.
JECFA (Joint FAO/WHO Expert Committee on Food Additives), 2011. Evaluation of
certain food additives and contaminants. 72nd Report of the Joint FAO/WHO
Expert Committee on Food Additive. WHO Technical Report Series, vol. 959.
Jestoi, M., Kokkonen, M., Uhlig, S., 2009. What about the ‘other’ Fusarium
mycotoxins? World Mycotoxin J. 2, 181–192.
Jiang, Y., Jolly, P.E., Preko, P., Wang, J.-S., Ellis, W.O., Phillips, T.D., Williams, J.H.,
2008. Aflatoxin-related immune dysfunction in health and in human
immunodeficiency virus disease. Clin. Dev. Immunol. vol. 2008, 12 pages
(Article ID 790309). doi:10.1155/2008/790309.
Joffe, A.Z., 1971. Alimentary toxic aleukia. In: Kadis, S., Ciegler, A., Ajl, S.J. (Eds.),
Microbiol Toxins, vol. 7. Academic Press, New York, pp. 139–189.
Juan, C., Mañes, J., Raiola, A., Ritieni, A., 2013. Evaluation of beauvericin and
enniatins in Italian cereal products and multicereal food by liquid
chromatography coupled to triple quadrupole mass spectrometry. Food Chem.
140, 755–762.
Kabak, B., 2009. Ochratoxin A in cereal-derived products in Turkey: occurrence and
exposure assessment. Food Chem. Toxicol. 47, 348–352.
 Author's personal copy
236 S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
Kamyar, M.R., Rawnduzi, P., Studenik, C.R., Kouri, K., Lemmens-gruber, R., 2004.
Investigation of the electrophysiological properties of enniatins. Arch. Biochem.
Biophys. 429, 215–223.
Kimanya, M., De Meulenaer, B., Tiisekwa, B., Ndomondo-Sigonda, M., Devlieghere,
F., Van Camp, J., Lachat, C., Baert, K., Kolsteren, P., 2007. Assessment of
fumonisin exposure to infants consuming maize based complementary foods in
Rombo District of Tanzania. Commun. Agric. Appl. Biol. Sci. 72, 13–17.
Kuiper-Goodman, T., Hilts, C., Billiard, S.M., Kiparissis, Y., Richard, I.D.K., Hayward,
S., 2010. Health risk assessment of ochratoxin a for all age-sex strata in a market
economy. Food Addit. Contam. A 27, 212–240.
Kuiper-Goodman, T., Scott, P.M., 1989. Risk assessment of the mycotoxin ochratoxin
A. Biomed. Environ. Sci. 2, 179–248.
Kuiper-Goodman, T., Scott, P.M., Watanabe, H., 1987. Risk assessment of the
mycotoxin zearalenone. Regulatory Toxicol. Pharmacol. 7, 253–306.
Kumagai, S., Nakajima, M., Tabata, S., Ishikuro, E., Tanaka, T., Norizuki, H., Itoh, Y.,
Aoyama, K., Fujita, K., Kai, S., Sato, T., Saito, S., Yoshiike, N., Sugita-Konishi, Y.,
2008. Aflatoxin and ochratoxin A contamination of retail foods and intake of
these mycotoxins in Japan. Food Addit. Contam. A 25, 1101–1106.
Leblanc, J.C., Tard, A., Volatier, J.L., Verger, P., 2005. Estimated dietary exposure to
principal food mycotoxins from The First French Total Diet Study. Food Addit.
Contam. 22, 652–672.
Leong, Y., Ahmad, R., Latiff, A.A., Nurul Izzah, A., 2011. Exposure assessment and risk
characterization of aflatoxin B1 in Malaysia. Mycotoxin Res. 27, 207–214.
Lewis, L., Onsongo, M., Njapau, H., Schurz-Rogers, H., Luber, G., Kieszak, S.,
Nyamongo, J., Backer, L., Dahiye, A., Misore, A., DeCock, K., Rubin, C., 2005.
Aflatoxin contamination of commercial maize products during an outbreak of
acute aflatoxicosis in Eastern and Central Kenya. Environ. Health Perspect. 113,
1763–1767.
Logrieco, A., Moretti, A., Castella, G., Kostecki, M., Golinski, P., Ritieni, A., Chelkowski,
J., 1998. Beauvericin production by Fusarium species. Appl. Environ. Microbiol.
64, 3084–3088.
Lori, G.A., Rizzo, I., 2007. Deoxinivalenol. In: Soriano, J.M. (Ed.), Micotoxinas en
Alimentos. Díaz de Santos, Madrid, pp. 269–292.
Mahfoud, R., Maresca, M., Garmy, N., Fantini, J., 2002. The mycotoxin patulin alters
the barrier function of the intestinal epithelium, mechanism of action of the
toxin and protective effects of glutathione. Toxicol. Appl. Pharmacol. 181, 209–
218.
Minervini, F., Giannoccaro, A., Cavallini, A., Visconti, A., 2005. Investigations on
cellular proliferation induced by zearalenone and its derivatives in relation to
the estrogenic parameters. Toxicol. Lett. 159, 272–283.
Mishra, S., Ansari, K.M., Dwivedi, P.D., Pandey, H.P., Das, M., 2013. Occurrence of
deoxynivalenol in cereals and exposure risk assessment in Indian population.
Food Control 30, 549–555.
Missmer, S.A., Suarez, L., Felkner, M., Wang, E., Merrill Jr, A.H., Rothman, K.J.,
Hendricks, K.A., 2006. Exposure to fumonisins and the occurrence of neural tube
defects along the Texas-Mexico border. Environ. Health Perspect. 114, 237–241.
Morales, H., Sanchis, V., Rovira, A., Ramos, A.J., Marin, S., 2007. Patulin accumulation
in apples during postharvest: effect of controlled atmosphere storage and
fungicide treatments. Food Control 18, 1443–1448.
Morgavi, D.P., Riley, R.T., 2007. An historical overview of field disease outbreaks
known or suspected to be caused by consumption of feeds contaminated with
Fusarium toxins. Anim. Feed Sci. Technol. 137, 201–212.
Murillo-Arbizu, M., Amézqueta, S., González-Peñas, E., de Cerain, A.L., 2009.
Occurrence of patulin and its dietary intake through apple juice consumption
by the Spanish population. Food Chem. 113, 420–423.
Nielsen, K.F., Grafenhan, T., Zafari, D., Thrane, U., 2005. Trichothecene production by
Trichoderma brevicompactum. J. Agric. Food Chem. 53, 8190–8196.
Norred, W.P., Plattner, R.D., Vesonder, R.F., Bacon, C.W., Voss, K.A., 1992. Effects of
selected secondary metabolites of Fusarium moniliforme on unscheduled
synthesis of DNA by rat primary hepatocytes. Food Chem. Toxicol. 30, 233–237.
Ok, H.E., Kim, H.J., Cho, T.Y., Oh, K.S., Chun, H.S., 2009. Determination of
deoxynivalenol in cereal-based foods and estimation of dietary exposure. J.
Toxicol. Environ. Health A 72, 1424–1430.
Ovchinnikov, Y.A., Ivanov, V.T., Evstratov, A.I., Mikhaleva, I.I., Bystrov, V.F., Portnova,
S.L., Balashova, T.A., Meshcheryakova, E.N., Tulchinsky, V.M., 1974. Enniatin
ionophores-conformation and ion binding properties. Int. J. Peptide Prot. Res. 6,
465–498.
Pacin, A., Martinez, E., Portela, M., Neira, S., 1998. Consumo de alimentos en la
población de la Universidad Nacional de Luján. Aporte energético y proteico. La
Alimentación Latinoamericana 221, pp. 28-36.
Palliyaguru, D.L., Wu, F., 2013. Global geographical overlap of aflatoxin and hepatitis
C, controlling risk factors for liver cancer worldwide. Food Add. Contam. – A. (in
press).
Park, D.L., 2002. Effect of processing on aflatoxin. Adv. Exp. Med. Biol. 504, 173–179.
Park, J.W., Kim, E.K., Kim, Y.B., 2004. Estimation of the daily exposure of Koreans to
aflatoxin B1 through food consumption. Food Addit. Contam. 21, 70–75.
Petersen, A., Thorup, I., 2001. Preliminary evaluation of fumonisins by the Nordic
countries and occurence of fumonisins (FB1 and FB2) in corn-based foods on the
Danish market. Food Addit. Contam. 18, 221–226.
Peterson, S.W., Ito, Y., Horn, B.W., Goto, T., 2001. Aspergillus bombycis, a new
aflatoxigenic species and genetic variation within its sibling species, Aspergillus
nomius. Mycologia 93, 689–703.
Pfohl-Leszkowicz, A., Petkova-Bocharova, T., Chernozemsky, I.N., Castegnaro, M.,
2002. Balkan endemic nephropathy and associated urinary tract tumours, a
review on aetiological causes and the potential role of mycotoxins. Food Add.
Contam. 19, 282–302.
Pfohl-Leszkowicz, A., Tozlovanu, M., Manderville, R., Peraica, M., Castegnaro, M.,
Stefanovic, V., 2007. New molecular and field evidences for the implication of
mycotoxins but not aristolochic acid in human nephropathy and urinary tract
tumor. Mol. Nutr. Food Res. 51, 1131–1146.
Piemontese, L., Solfrizzo, M., Visconti, A., 2005. Occurrence of patulin in
conventional and organic fruit products in Italy and subsequent exposure
assessment. Food Addit. Contam. 22, 437–442.
Pieters, M.N., Bakker, M., Slob, W., 2004. Reduced intake of deoxynivalenol in the
Netherlands: a risk assessment update. Toxicol. Lett. 153, 145–153.
Pirrung, M.C., Nauhaus, S.K., Singh, B., 1996. Cofactor-directed, time-dependent
inhibition of thiamine enzymes by the fungal toxin moniliformin. J. Org. Chem.
61, 2592–2593.
Placinta, C.M., D’Mello, J.P.F., MacDonald, A.M.C., 1999. A review of worldwide
contamination of cereal grains and animal feed with Fusarium mycotoxins. An.
Feed Technol. 78, 21–37.
Probst, C., Njapau, H., Cotty, P.J., 2007. Outbreak of an acute aflatoxicosis in Kenya in
2004, identification of the causal agent. Appl. Environ. Microbiol. 73, 2762–
2764.
Puel, O., Galtier, P., Oswald, I.P., 2010. Biosynthesis and toxicological effects of
patulin. Toxins 2, 613–631.
Rapid Alert System for Food and Feed (RASFF), 2012. Annual Reports 2012.
European Commission. <http://ec.europa.eu/food/food/rapidalert/rasff_publica-
tions_en.htm>.
Rice, L.G., Ross, F.B., 1994. Methods for detection and quantitation of fumonisins in
corn, cereal products and animal excreta. J. Food Protect. 57 (5), 36–40.
Richardson, K.E., Hagler, W.M., Mirocha, C.J., 1985. Production of zearalenone, a–
zearalenol and b–zearalenol by Fusarium spp. in rice culture. J. Agric. Food
Chem. 33, 862–866.
Rotter, B.A., Prelusky, D.B., Pestka, J.J., 1996. Toxicology of deoxinivalenol
(vomitoxin). J. Toxicol. Environ. Health – A 48, 1–34.
Ryu, D., Hanna, M.A., Bullerman, L.B., 1999. Stability of zearalenone during
extrusion of corn grits. J. Food Protect. 62, 1482–1484.
Sáenz de Rodríguez, C.A., Bongiovanni, A.M., Conde de Borrego, L., 1985. An
epidemic of precocious development in Puerto Rican children. J. Pediatr. 107,
393–396.
Santini, A., Meca, G., Uhlig, S., Ritieni, A., 2012. Fusaproliferin, beauvericin and
enniatins: occurrence in food – a review. World Mycotoxin J. 5, 71–81.
Scientific Committee on Food (SCF). 1999. Opinion on Fusarium Toxins-Part 1:
Deoxynivalenol (DON). <http://ec.europa.eu/food/fs/sc/scf/out44_en.pdf>
(accessed 07.02.13).
Scientific Committee on Food (SCF), 2000a. Opinion on Fusarium Toxins, Part 2,
Zearalenone. Scientific Committee on Food SCF/CS/CNTM/MYC/22 Rev. 3 Final.
<http://ec.europa.eu/food/fs/sc/scf/out65_en.pdf> (accessed 07.02.13.
Scientific Committee on Food (SCF), 2000b. Minute Statement on Patulin Expressed
by the Scientific Committee on Food During the Plenary Meeting. Brussels,
Belgium.
Scientific Committee on Food (SCF), 2002. Opinion of the Scientific Committee on
Food on Fusarium toxins. Part 6, Group Evaluation of T-2 Toxin, HT-2 Toxin,
Nivalenol and Deoxinivalenol. Scientific Committee on Food SCF/CS/CNTM/
MYC/27 Final. <http://ec.europa.eu/food/fs/sc/scf/out123_en.pdf> (accessed
07.02.13).
Scientific Committee on Food (SCF), 2003. Updated opinion of the Scientific
Committee on Food on Fumonisin B1, B2 and B3. Scientific Committee on Food
SCF/CS/CNTM/MYC/28 Final. <http://ec.europa.eu/food/fs/sc/scf/
out185_en.pdf> (accessed 07.02.13).
SCOOP, 2002a. Reports on Tasks for Scientific Cooperation – Report of Experts
Participating in Task 3.2.7 – Assessment of Dietary Intake of Ochratoxin A by the
Population of EU Member Status.
SCOOP, 2002b. Reports on tasks for scientific cooperation – Report of Experts
Participating in Task 3.2.8 – Assessment of Dietary Intake of Patulin by the
Population of EU Member States.
SCOOP, 2003. Reports on Tasks for Scientific Cooperation – Report of Experts
Participating in Task 3.2.10 – Collection of Occurrence Data of Fusarium Toxins
in Food and Assessment of Dietary Intake by the Population of EU Member
States.
Serrano, A.B., Font, G., Ruiz, M.J., Ferrer, E., 2012a. Co-occurrence and risk
assessment of mycotoxins in food and diet from Mediterranean area. Food
Chem. 135, 423–429.
Serrano, A.B., Meca, G., Font, G., Ferrer, E., 2012b. Risk assessment associated to the
intake of the emerging Fusarium mycotoxins BEA, ENs and FUS present in infant
formula of Spanish origin. Food Control 28, 178–183.
Shephard, G.S., 2008. Impact of mycotoxins on human health in developing
countries. Food Addit. Contam. 25, 146–151.
Shephard, G.S., Van Der Westhuizen, L., Katerere, D.R., Herbst, M., Pineiro, M., 2010.
Preliminary exposure assessment of deoxynivalenol and patulin in South Africa.
Mycotoxin Res. 26, 181–185.
Sirot, V., Fremy, J.M., Leblanc, J.C., 2013. Dietary exposure to mycotoxins and health
risk assessment in the second French total diet study. Food Chem. Toxicol. 52,
1–11.
Sirot, V., Volatier, J.L., Calamassi-Tran, G., Dubuisson, C., Ménard, C., Dufour, A.,
Leblanc, J.C., 2009. Core food of the French food supply: second total diet study.
Food Addit. Contam. A 26, 623–639.
Sizoo, E.A., Van Egmond, H.P., 2005. Analysis of duplicate 24-hour diet samples for
aflatoxin B1, aflatoxin M1 and ochratoxin A. Food Addit. Contam. 22, 163–172.
Solfrizzo, M., Chulze, S.N., Mallmann, C., Visconti, A., De Girolamo, A., Rojo, F., Torres,
A., 2004. Comparison of urinary sphingolipids in human populations with high
 Author's personal copy
S. Marin et al. / Food and Chemical Toxicology 60 (2013) 218–237
and low maize consumption as a possible biomarker of fumonisin dietary
exposure. Food Addit. Contam. 21, 1090–1095.
Solfrizzo, M., Gambacorta, L., Lattanzio, V.M.T., Powers, S., Visconti, A., 2011.
Simultaneous LC-MS/MS determination of aflatoxin M1, ochratoxin A,
deoxynivalenol, de-epoxydeoxynivalenol, alpha and beta-zearalenols and
fumonisin B1 in urine as a multi-biomarker method to assess exposure to
mycotoxins. Anal. Bioanal. Chem. 401, 2831–2841.
Solovey, M.M.S., Somoza, C., Cano, G., Pacin, A., Resnik, S., 1999. A survey of
fumonisins, deoxynivalenol, zearalenone and aflatoxins contamination in corn-
based food products in Argentina. Food Addit. Contam. 16, 325–329.
Speijers, G.J.A., Franken, M.A.M., van Leeuwen, F.X.R., 1988. Subacute toxicity study
of patulin in the rat, effects on the kidney and the gastro-intestinal tract. Food
Chem. Toxic. 26, 23–30.
Stefanovic, V., Polenakovic, M., Toncheva, D., 2011. Urothelial carcinoma associated
with Balkan endemic nephropathy: a worldwide disease. Pathol. Biol. 59, 286–
291.
Suarez, L., Felkner, M., Brender, J.D., Canfield, M., Zhu, H., Hendricks, K.A., 2012.
Neural tube defects on the Texas-Mexico Border, what we’ve learned in the 20
years since the Brownsville cluster. Birth Defects Res. A 94, 882–892.
Sudakin, D.L., 2003. Trichothecenes in the environment, relevance to human health.
Toxicol. Lett. 143, 97–107.
Sugita-Konishi, Y., Kamata, Y., Sato, T., Yoshinari, T., Saito, S., 2012. Exposure and
risk assessment for ochratoxin A and fumonisins in Japan. Food Addit. Contam.
– A. (in press).
Sugita-Konishi, Y., Sato, T., Saito, S., Nakajima, M., Tabata, S., Tanaka, T., Norizuki, H.,
Itoh, Y., Kai, S., Sugiyama, K., Kamata, Y., Yoshiike, N., Kumagai, S., 2010.
Exposure to aflatoxins in Japan: Risk assessment for aflatoxin B 1. Food Addit.
Contam. A 27, 365–372.
Sun, G., Wang, S., Hu, X., Su, J., Zhang, Y., Xie, Y., Zhang, H., Tang, L., Wang, J.S., 2011.
Co-contamination of aflatoxin B 1 and fumonisin B 1 in food and human dietary
exposure in three areas of China. Food Addit. Contam. A 28, 461–470.
Sydenham, E.W., Thiel, P.G., Marasas, W.F.O., Shephard, G.S., Schalkwyk, D.J., Koch,
K.R., 1990. Natural occurrence of some Fusarium mycotoxins in corn from low
and high esophageal cancer prevalence areas of Transkei, Southern Africa. J.
Agric. Food Chem. 38, 1900–1903.
Thrane, U., 2001. Developments in the taxonomy of Fusarium species based on
secondary metabolites. In: Summerbell, B.A., Leslie, J.F., Backhouse, D., Bryden,
W.L., Burgess, L.W. (Eds.), Fusarium. Paul E. Nelson Memorial Symposium. APS
Press, St. Paul, Minnesota, pp. 29–49.
Thuvander, A., Möller, T., Enghardt Barbieri, H., Jansson, A., Salomonsson, A.C., Olsen,
M., 2001. Dietary intake of some important mycotoxins by the Swedish
population. Food Addit. Contam. 18, 696–706.
Tomaszewski, J., Miturski, R., Semczuk, A., Kotarski, J., Jakowicki, J., 1998. Tissue
zearalenone concentration in normal, hyperplastic and neoplastic human
endometrium. Ginekologia Polska 69, 363–366.
Turner, P.C., Gong, Y.Y., Pourshams, A., Jafari, E., Routledge, M.N., Malekzadeh, R.,
Wild, C.P., Boffetta, P., Islami, F., 2012. A pilot survey for Fusarium mycotoxin
biomarkers in women from Golestan, Northern Iran. World Mycotoxin J. 5, 195–
199.
Turner, P.C., Hopton, R.P., White, K.L.M., Fisher, J., Cade, J.E., Wild, C.P., 2011a.
Assessment of deoxynivalenol metabolite profiles in UK adults. Food Chem.
Toxicol. 49, 132–135.
Turner, P.C., Ji, B.T., Shu, X.O., Zheng, W., Chow, W.H., Gao, Y.T., Hardie, L.J., 2011b. A
biomarker survey of urinary deoxynivalenol in China: the Shanghai Women’s
Health Study. Food Addit. Contam. A 28, 1220–1223.
237
Turner, P.C., Rothwell, J.A., White, K.L.M., Gong, Y., Cade, J.E., Wild, C.P., 2008.
Urinary deoxynivalenol is correlated with cereal intake in individuals from the
United Kingdom. Environ. Health Perspect. 116, 21–25.
Uhlig, S., Jestoi, M., Knutsen, A.K., Heier, B.T., 2006. Multiple regression analysis as a
tool for the identification of relations between semi-quantitative LC-MS data
and cytotoxicity of extracts of the fungus Fusarium avenaceum (syn.
F.arthrosporioides). Toxicon 46, 513–522.
Van der Merwe, K.J., Steyn, P.S., Fourie, L., 1965. Mycotoxins. Part II. The constitution
of ochratoxin A, B and C, metabolites of Aspergillus ochraceus Wilh. J. Chem. Soc.
5, 7083–7088.
Villa, P., Markaki, P., 2009. Aflatoxin B1 and ochratoxin A in breakfast cereals from
athens market: occurrence and risk assessment. Food Control 20, 455–
461.
Visconti, A., 2001. Problems associated with Fusarium mycotoxins in cereals. Bull.
Inst. Comprehens. Agric. Sci. Kinki Univ. 9, 39–55.
Visconti, A., Minervini, F., Lucivero, G., Gambatesa, V., 1991. Cytotoxic and
immunotoxic effects of Fusarium mycotoxins using a rapid colorimetric
bioassay. Mycopathologia 113, 181–186.
Voss, K.A., Smith, G.W., Haschek, W.M., 2007. Fumonisins, toxicokinetics,
mechanism of action and toxicity. Anim. Feed Sci. Technol. 137, 299–325.
Wang, W., Shao, B., Zhu, J., Yu, H., Li, F., 2010. [Dietary exposure assessment of some
important Fusarium toxins in cereal-based products in China]. Wei sheng yan
jiu. J. Hygiene Res. 39, 709–714.
Warth, B., Sulyok, M., Fruhmann, P., Berthiller, F., Schuhmacher, R., Hametner, C.,
Adam, G., Froehlich, J., Krska, R., 2012a. Assessment of human deoxynivalenol
exposure using an LC-MS/MS based biomarker method. Toxicol. Lett. 211, 85–
90.
Warth, B., Sulyok, M., Fruhmann, P., Mikula, H., Berthiller, F., Schuhmacher, R.,
Hametner, C., Abia, W.A., Adam, G., Froehlich, J., Krska, R., 2012b. Development
and validation of a rapid multi-biomarker liquid chromatography/tandem mass
spectrometry method to assess human exposure to mycotoxins. Rapid
Commun. Mass Spectrom. 26, 1533–1540.
WHO (World Health Organization), 1990. Selected Mycotoxins: Ochratoxins,
Trichothecenes, Ergot. Environ. Health Crit. 105. Geneva.
WHO (World Health Organization), 1995. 44th Report of the Joint FAO/WHO Expert
Committee on Food Additives. Technical Report Series 859, 36.
WHO (World Health Organization), 2001. Safety Evaluation of Certain Mycotoxins
in Food. WHO Food Additive Series 47, Geneva.
Wood, G.E., 1992. Mycotoxins in foods and feeds in the United States. J. Anim. Sci.
70, 3941–3949.
Wu, H.C., Santella, R., 2012. The role of aflatoxins in hepatocellular carcinoma.
Hepat. Mon. 12, 8–16.
Wolf-Hall, C.E., Hanna, M.A., Bullerman, L.B., 1999. Stability of deoxynivalenol in
heat-treated foods. J. Food Prot. 62, 962–964.
Yoshizawa, T., Yamashita, A., Luo, Y., 1994. Fumonisin occurrence in corn from high-
and low-risk areas for human esophageal cancer in China. Appl. Environ.
Microbiol. 60, 1626–1629.
Yu, Z., Zhang, L., Wu, D., Liu, F., 2005. Anti-apoptotic action of zearalenone in MCF-7
cells. Ecotoxicol. Environ. Safe. 62, 441–446.
Zinedine, A., Soriano, J.M., Moltó, J.C., Mañes, J., 2007. Review on the toxicity,
occurrence, metabolism, detoxification, regulations and intake of zearalenone,
an oestrogenic mycotoxin. Food Chem. Toxicol. 45, 1–18.