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Mycotoxins
Introduction
1
Mycotoxins are secondary metabolites (secondary metabolite: A
compound that is not necessary for growth or maintenance of cellular functions but is synthesized,
generally, for the protection of a cell or micro-organism, during the stationary phase of the growth
cycle. Many are used in foods, pharmaceuticals, and other industrial applications
.) of fungi
that are recognized as toxic to other life forms.
1.Fungal growth
• a. Field fungi : fungi that attack plants that grow in the field (occurring prior to harvest)
grow under special conditions. (Fusarium)
• b. Storage fungi : Storage fungi usually invade grain or seed during storage and are
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generally not present in large quantities before harvest in the field. The most common storage fungi
are species of Aspergillus and Penicillium. Contamination occurs through spores
contaminating the grain as it is going into storage from the harvest. The development of fungi is
influenced by the:
Moisture content of the stored grain
Temperature
Condition of the grain going into storage
Length of time the is grain stored and
Amount of insect and mite activity in the grain
2. Characteristics of mycotoxin induced
disease
a. not transmitted among animals
b. Pharmaceutical treatment does not alter the
course of disease
c. Mycotoxicosis most often presents as a
uncertain, sub-acute or chronic condition
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3.Treatment of mycotoxininduced disease
a. For most mycotoxins, there is no
specific treatment or antidote
b. Supplement with vitamins & selenium
may be helpful, and provision of adequate
high-quality protein
4
4.Prevention of mycotoxininduced disease
a. Avoiding
b. Diluting
c. Cleaning
d. Testing
e. Drying
f. Adding (organic acids will prevent mold
growth)
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A- Aflatoxin
• 1. Sources :
Aspergillus flavus & A.paraciticus :
Corn, peanuts
• 2. Factor favoring production of aflatoxins
a. Temperature : 25-30 ๐c
b. Grain moisture
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3. Chemical characteristics
• Exhibit intense blue or green
fluorescence under UV.
: aflatoxins B1, B2, G1 and G2
also
: aflatoxin M1 is a metabolites of
AFB1 found in animal urine, milk or
tissues.
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5.Mechanism of toxicologic
damage
Also called steatosis, fatty liver can be a temporary or long-term condition, which is
not harmful itself, but may indicate some other type of problem. Left untreated, it can
contribute to other illnesses. It is usually reversible once the cause of the problem is
diagnosed and corrected. The liver is the organ responsible for changing fats eaten in
the diet to types of fat that can be stored and used by the body. Triglycerides are one
of the forms of fat stored by the body and used for energy and new cell formation. The
break down of fats in the liver can be disrupted by alcoholism, malnutrition, pregnancy,
or poisoning. In fatty liver, large droplets of fat, containing mostly triglycerides, collect
within cells of the liver. The condition is generally not painful and may go unnoticed for
a long period of time. In severe cases, the liver can increase to over three times its normal size and
may be painful and tender
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• a. Loss of enzyme
• b. Lack of formation of lipid acceptor
protein in liver
• c. Decreased cellulose digestion,
volatile fatty acid formation &
proteolysis
• d. Necrosis
(breakdown of proteins )
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6.Toxicity
• a. Young animals are more susceptible than
adult.
• b. Nutrition deficiency increase
susceptibility
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7. Diagnosis
• Clinical sign : decreased growth rate,
reduced feed efficiency,,, mild anemia,
and increased susceptibility to infectious
disease.
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8.Treatment & Prevention
• a. Detoxification : Hydrated sodium
calcium aluminosilicate (HSCAS) can
absorb aflatoxins
• b. Supportive : Vitamin .E & selenium
• c. Prevention
- Mold inhibitor
- Treatment of grain with anhydrous
ammonia for 10-14 days.
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B- Zearalenone
• 1. Sources : Fusarium roseum
( F.graminearum ):
corn, wheat, barley, oats
• 2. Factor favoring production
a. High moisture 22% - 25%
b. Alternating high and low temp. (7-21 ๐c)
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3. Mechanism of toxicological damage
a. initiating specific RNA synthesis
b. Function as a weak estrogen.
4.Toxicity
a. Swine are most susceptible
b. low for all effects except reproductive
function.
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C-Ergot
1.Source :
Claviceps purpurea :
barley, wheat & oats
2. Factor favoring :
Warm & humid
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3.Mechanism of toxic
• a. potent initiators of contraction in smooth muscle
• b. mimic the action of dopamine.
4.Clinical sign
a. necrosis of the feet, ears and tail
b. increased temperature., pulse & respiration rate
c. lactation does not occur
d. hyper-excitability & tremors
e. heat intolerance in cattle
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E.Treatment
a. animals should be provided with a
warm, clean, stress-free environment
b. Control secondary bacterial
infection
c. milk supplement
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D- Ochratoxin & Citrinin
• 1.Sources : Aspergillus orchraceus &
• Penicillium viridicatum
• 2. Mechanism of toxic :
target the renal proximal tubule
• - Disrupt protein synthesis
• -Bind strongly to protein (albumin)
• -Interfere with synthesis of tRNA & mRNA
• -Disrupt carbohydrate metabolism
• -Increase the generation of free radical
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4.Clinical sign
• a. Acute : vomiting, diarrhea, dehydration &
depression
• b. Subacute to chronic : weight loss, feed efficiency,
& dehydration. Immunosupression, teratogenicity,
carcinogenesis & hemorrhage
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Mycotoxins
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Mycotoxin
Mycotoxin is a convenient generic term describing the toxic
secondary metabolites produced by fungi. “Myco” means
fungal (mold) and “toxin” represents poison.
They encompass a considerable variety of low molecular
weight compounds with diverse chemical structures and
biological activities.
Some mycotoxins could also be toxic to plants or other
microorganisms; but these compounds are not classified as
antibiotics of fungal origin.
Like most microbial secondary metabolites, the benefit of
mycotoxins for the fungi themselves is still not clearly defined.
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• In considering the effects of mycotoxins on
animals, it is important to distinguish
between “mycotoxicosis” and “mycosis.”:
• Mycotoxicosis is used to describe the
action of mycotoxin(s) and is frequently
mediated through a number of organs,
notably the liver, kidney, lungs, and the
nervous, endocrine, and immune systems.
• Mycosis” refers to a generalized invasion
of living tissue(s) by growing fungi.
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• Due to their diverse chemical structures,
mycotoxins may exhibit a number of
biological effects, including both acute and
chronic toxic effects as well as carcinogenic,
mutagenic, genotoxic, and immunotoxic
effects.
• The interaction of mycotoxins with cellular
macromolecules plays a dominant role in
their toxic actions. Recent studies on the
effect of mycotoxins on apoptosis have
further revealed their mode of action at the
cellular level.
23
Historical
• Modern mycotoxicology was not developed until the discovery
of aflatoxins in the early 1960s as the causative agent in the
• peanut meal causing the “Turkey X” disease that killed more
than 10,000 turkeys fed with the contaminated meal.
• Because aflatoxins are a series of highly potent carcinogens
produced by commonly occurring Aspergillus flavus and A.
parasiticus, research has focused new attention on mycotoxins.
• In the last 40 years, many new mycotoxins have been identified
and characterized, and their biosynthetic origin in various fungi
elucidated. It has been estimated that at least 25% of the
world’s agricultural product is contaminated with mycotoxins
and certain diseases have been linked to ingestion of food and
feed
contaminated with mycotoxins.
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Economic Impact of Mycotoxin Contamination
• The most obvious negative economic impact of
mycotoxins is an outright loss of crops and affected
animals.
• Also, humans may encounter severe health hazard or
high mortality rates in countries with less regulation or
monitoring programs.
• Thus, the negative economic impact resulting from
mycotoxin contamination is certainly very significant
and estimated to be $932 million annually.
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PRODUCTION OF MYCOTOXINS BY TOXICOGENIC
FUNGI
Invasion by fungi and production of mycotoxins in commodities can occur
under favorable conditions in the field, at harvest, and during processing,
transportation and storage
Fungi that are frequently found in the field include: A. flavus, Alternaria
longipes, A. alternata, Claviceps purpura, Fusarium verticillioides
(previously called moniliforme), F. graminearum, and a number of other
Fusarium spp.
Species most likely introduced at harvest include:
F. sporotrichioides, Stachybotrys atra, Cladosporium sp., Myrothecium
verrucaria, Trichothecium roseum, as well as A. alternata.
Most penicillia are storage fungi. These include: Penicillium citrinum, P.
cyclopium, P. citreoviride, P. islandicum, P. rubrum, P. viridicatum, P. urticae,
P. verruculosum, P. palitans, P. puberulum, P. expansum, and P. roqueforti.
All of which are capable of producing mycotoxins in grains and foods.
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• Other toxicogenic storage fungi are: Aspergillus.
parasiticus, A. flavus, A. versicolor, A. ochraceus, A.
clavatus, A. fumigatus, A. rubrum, A. chevallieri,
Fusarium verticillioides, F. tricinctum, F. nivale, and
several other Fusarium spp.
• It is apparent, most of the mycotoxin producing fungi
belong to three genera: Aspergillus, Fusarium, and
Penicillium. However, not all species in these
genera are toxicogenic
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Factors Affecting Mycotoxin Production
• Genetics and environmental and nutritional factors
greatly affect the formation of mycotoxins.
• Depending on the susceptibility of the crop, geographic
and seasonal factors, as well as cultivation, harvesting,
storage, and transportation practices, mycotoxins are
found worldwide.
• In the field, weather conditions, plant stress,
invertebrate vectors, species and spore load of
infective fungi, variations within plant and fungal
species, and microbial competition all significantly
affect mycotoxin production.
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Continue Factors Affecting…….
• Physical factors such as time of exposure,
temperature during exposure, humidity, and
extent of insect or other damage to the
commodity prior to exposure determine
mycotoxin contamination in the field or during
storage.
• Chemical factors including the nutritional
status of the crops or chemicals (such as
fungicides) used in crop management could
affect fungal populations, and consequently
toxin production
29
Continue Factors Affecting…….
• In general, mycotoxins are optimally produced
at 24–28C, but some toxins such as T-2 toxin
is maximally produced at 15C.
• Contamination during crop storage may be
affected by changes in temperature and water
activity, that allow ecological succession of
different fungi as water activity and temperature
of stored grain changes.
30
Water activity = It is defined as the vapor pressure of water above a sample divided by that of pure water at the
same temperature;
Continue Factors Affecting…….
• During storage and transportation, water
activity (aw), temperature, crop damage, and a
number of physical and chemical factors, such
as aeration (O2, CO2 levels), types of grains,
pH, and presence or absence of specific
nutrients and inhibitors are important.
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Aflatoxins
A
Chemical structure of flatoxins
(A) The B-type aflatoxins are
characterized by a cyclopentane Ering. These compounds have a
blue fluorescence under longwavelength ultraviolet light.
(B) The G-type aflatoxins, with a green
fluorescence, have a xanthone ring
in place of the cyclopentane.
(C) Aflatoxins of the B2 and G2 type
have a saturated bis-furanyl ring.
Only the bis-furan is shown.
(D) Aflatoxin of the B1a and G1a type
have a hydrated bis-furanyl
structure.
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R
3
R
1
C
R
2
R
1
B
R
4
D
R
1
R
1
• At least 16 structurally related toxins in this group
are produced by Asparagillus flavus and A.
parasiticus and infrequently by A. pseudotamarii and
A. nominus
• A. ochraceoroseus has also been found to
produce aflatoxins
• The optimal temperatures and water activity (aw) for
the growth of A. flavus and A. parasiticus are around
35–37C (range from 6–54C) and 0.95 (range from
0.78–1.0), respectively; whereas for aflatoxin
production, they are 28–33C and 0.90–0.95 (range
from 0.83–0.97), respectively.
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• Aflatoxin B1 is most toxic in this group and is
one of the most potent naturally occurring
carcinogens).
• Other significant members of the aflatoxin
family, such as M1 and M2, are metabolites
of AFB1 and AFB2, respectively, and
originally isolated from bovine milk.
34
Natural Occurrence
• Aflatoxins have been found in corn, peanuts ‫ فول سوداني‬and
peanut products, cotton seeds, peppers, rice, pistachios, ‫فستق‬
tree nuts, pumpkin ‫ قرع‬seeds, sunflower seeds and other oil
seeds, copra, ‫ جوز هند‬spices, and dried fruits (figs, raisins).
• Among these products, frequent contamination with high
levels of AF in peanuts, corn, and cottonseed, mostly due to
infestation with fungi in the field, are of most concern.
• Soybeans ‫الصويا‬, beans ‫الفاصولياء‬, pulses (Pea)‫البازالء‬, cassava
‫منيهوت‬, sorghum ‫ الذرة‬, millet ‫الدخن‬, wheat ‫القمح‬, oats ‫القطن‬, barley
‫الشعير‬, and rice‫ رز‬are resistant or only moderately susceptible
to AF contamination in the field.
•
It should be reiterated that resistance to AF contamination in
the field does not guarantee that the commodities are free of
AF contamination during storage. Inadequate storage
conditions, such as high moisture and warm temperatures
(25–308C), can create conditions favorable for the growth of
fungus and production of AF.
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Toxic Effects
• Aflatoxins are mutagenic, teratogenic, and hepatocarcinogenic.
• Aflatoxin B1 is one of the most potent naturally occurring
carcinogen, extensive research was primarily done on this
toxin. The main target organ of AF is the liver.
• AFB1 also affects other organs and tissues including the lungs
and the entire respiratory system.
•
For the carcinogenic effects, rats, rainbow trout, monkeys, and
ducks are most susceptible and mice are relatively resistant.
• Consumption of AFB1-contaminated feed by dairy cows results
in the excretion of AFM1 in milk. AFM1, a hydroxylated
metabolite of AFB1, is about 10 times less toxic than AFB1; but
its presence in milk is of concern for human health.
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Impact on Human Health
• Whereas AFB1 has been found to be a potent carcinogen in
many animal species, the role of AF in carcinogenesis in
humans is complicated by hepatitis B virus (HBV) infections in
humans).
• Epidemiological studies have shown a strong positive
correlation between AF levels in the diet and primary
hepatocellular carcinoma.
•
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Since multiple factors are important in carcinogenesis and
environmental contaminants such as AFs and other
mycotoxins may, either in combination with HBV or
independently.
Ochratoxins
• Ochratoxins, are produced by a number of fungi in
the genera Aspergillus and Penicillium. The largest
amounts ochratoxins are made by A. ochraceus and
P. cyclopium.
• A. ochraceus and P. viridicatum (reclassified as P.
verrucosum), two species that were first reported as
ochratoxin A (OA) producers, occur most frequently
in nature.
• Other fungi, such as Petromyces alliceus, A.
citricus, and A. fonsecaeus (both in A. niger group),
have also been found to produce OA. Most of the
OA producers are storage fungi and preharvest
fungal infection.
38
• Although most OA producers can grow in a range from 48C to
37C and at aw as low as 0.78, optimal conditions for toxin
production are narrower with temperature at 24–25C and aw
values .0.97.
• Ochratoxins are produced primarily in cereal grains (barley,
oats, corn, wheat) and mixed feed during storage in temperate
climatic conditions, with levels higher than 1 ppm being
reported.
• OA has been found in other commodities, including beans,
coffee, nuts, olives, raisin, cheese, fish, pork, milk powder, fruit
juices wine beer, peppers.
•
OA can be carried through the food chain because of the
presence of OA residues in animal products as result of its
binding with serum albumin. Natural occurrence of OA in
kidneys, blood serum, blood sausage.
39
Structure of the ochratoxins. These metabolites form different classes depending on the nature of the
amide group (a–c), and the presence or absence of a chlorine moiety at R2 in the phenyl group
40
.
Ochratoxin A, the most toxic member of this
group of mycotoxins, has been found to be a
potent nephrotoxin causing kidney damage
as well as liver necrosis and enteritis in
many animal species
The OA inhibits carboxypeptidase A, renal
phosphoenolpyruvate carboxykinase,
phenylalaninetRNA synthetase, and phenylalanine
hydroxylase activity.
Formation of free radicals has been considered as
one of the mechanisms for the carcinogenic/toxic
effects of OA.
41
Fumonisins
Fumonisins (Fm) are a group of toxic metabolites produced primarily by F.
verticillioides, F. proliferatum and other related species readily colonize corn
all over the world. Although F. anthophilum, F. nupiforme, and F. nygamai
are capable of producing Fms.
More than 11 structurally related Fms (B1, B2, B3, B4, C1, C4, A1, A2, etc.),
have been found since the discovery of FmB1.
Fumonisins are most frequently found in corn, corn-based foods, and other
grains (such as sorghum and rice). The level of contamination varies
considerably with different regions and year, ranging from negligible to more
than 100 ppm; but is generally reported to be between 1 and 2 ppm.
FmB1 is the most common Fm in naturally contaminated samples; FmB2
generally accounts for 1/3 or less of the total. Although production of the
toxin generally occurs in the field, continued production of toxin during
postharvest storage also contributes to the overall levels.
42
Toxicologic Effects
Fumonisin B1 is primarily a hepatotoxin and carcinogen in rats. Feeding
culture material from F. verticillioides or pure FmB1 to rats resulted in
‫تليف كبدي‬
cirrhosis and hepatic nodules, carcinoma. Kidney is also a target organ.
Mechanistically, Fms are inhibitors of ceramide synthase
(sphinganine/sphingosine N-acyltransferase), a key enzyme involved in the
biosynthesis of sphingolipids, which are heavily involved in cellular
regulation, including cell differentiation, mitogenesis and apoptosis
The ability of FmB1 to alter gene expression and signal transduction
pathways are considered necessary for its carcinogenic and toxic effects.
FmB1 is a good example of an apparently non-genotoxic (non-DNA
reactive) agent producing tumors through the regulation of apoptosis
43
Trichothecenes (TCTCs)
Several species of Fusaria infect corn, wheat, barley, and rice.
Under favorable conditions, they elaborate a number of different types of
mycotoxins (look figure).
(TCTCs) are generally classified as macrocyclic (Type C) or nonmacrocyclic
(Types A and B). Although more than 100 TCTCs have been identified, only a
few frequently found in foods and feeds are potentially hazardous to human
and animal health.
Trichothecenes
Other fungal genera elaborate TCTCs are: Myrothecium, Trichoderma,
Trichothecium, Cephalosporium, Verticimonosporium, and Stachybotrys. In
addition to fungi, extracts from a Brazilian shrub, Baccharis megapotamica,
also contain macrocyclic TCTCs. The term TCTCs is derived from
trichothecin, the first compound isolated in this group.
44
The TCTC mycotoxicoses affect many organs, including the gastrointestinal
tract, hematopoietic, nervous, immune, hepatobiliary, and cardiovascular
systems.
Mechanistically, inhibition of protein synthesis is one of the earlier events in
manifestation of TCTC toxic effects and they act at different steps in the
translation process. Inhibitory effects of these mycotoxins vary considerably
with the chemical structure of the side chain.
a) T-2 toxin,
T-2 toxin, a highly toxic type A TCTC, is produced by F.
tricinctum, F. sporotrichioides (major), F. poae,
F. sulphureum, F. acuminatum, and F. sambucinum.
Unlike most mycotoxins, which are usually synthesized
near 25C, the optimal temperature for T-2 toxin
production
is around 15C.
45
Almost all the major TCTCs, including T-2 toxin,
are cytotoxic and cause hemorrhage, edema,
and necrosis of skin tissues.
b) Deoxynivalenol (DON)
The DON is a major type B TCTC mycotoxin produced
by F. graminearum (major) and other related fungi
such as F. culmorum and F. crookwellense. Because
DON causes feed refusal and emesis in swine, the
name “vomitoxin” is also used for this mycotoxin.
46
• Worldwide frequent natural occurrence of DON in
cereal grains has been reported. Contamination of this
toxin in corn and wheat is generally high.
• Also, contamination of barley, oats, sorghum, rye,
safflower seeds, and mixed feeds has also been
reported.
• Although inadequate storage may lead to the
production of some TCTC mycotoxins, infestation of
fusaria in wheat and corn in the field is of most
concern for the DON problem.
• With wet and cold weather during maturation,grains
are especially susceptible to F. graminearum infection,
which causes so-called “scabby wheat” and
simultaneously produces the toxin. The optimal
temperature for DON production is about 248C.
47
• Toxicologically, DON induces anorexia and emesis both in
humans and animals. Swine are most sensitive to feed
contaminated with DON. Whereas most TCTCs are
immunosuppressors, DON is a hyperinducer of cytokines.
48
• Other Selected Mycotoxins
• In addition to the mycotoxins discussed above, a number of other
mycotoxins occur naturally.
• Other Mycotoxins Produced by Aspergillus:
• Sterigmatocystin (ST) is a naturally occurring hepatotoxic and
carcinogenic mycotoxin produced by fungi in the genera Aspergillus,
Bipolaris, and Chaetomium as well as P. luteum.
• Structurally related to AFB1 ST is known to be a precursor of AFB1.
• ST is a mutagen and genotoxin and has been found in cereal grains
(barley, rice, and corn), coffee beans, and cheese.
Structure of sterigmatocystin. The bis-furanyl structure is similar to
that of the aflatoxins except that the E-ring is a substituted phenol.
49
• A. terreus and several other fungi (e.g., A. flavus and A.
fumigatus) have been found to produce the tremorgenic
toxins, territrems, aflatrem, and fumitremorgin.
•
A. terreus, A. fumigatus, and Trichoderma viride also
produce gliotoxin, In addition, A. flavus, A. wentii, and
A. oryzae, are capable of producing nitropropionic acid
(NPA), a mycotoxin causing apnea, convulsions,
congestion in lungs and liver.
• Production of NPA in sugarcanes by Arthrinium sacchari,
Arth. saccharicola, and Arth. Phaeospermum has been
found to be involved in fatal food poisoning in humans
50
Other Mycotoxins Produced by Penicillium
Penicillia produce many mycotoxins with diverse toxic effects.
Cyclochlorotine, luteoskyrin (LS), and rugulosin (RS) have long been
considered to be possibly involved in the yellow rice disease during the
Second World War. They are hepatotoxins. Several other mycotoxins,
including patulin (PT) penicillic acid (PA) citrinin (CT), cyclopiazonic acid
(CPA, citreoviridin, and xanthomegnin, which are produced primarily by
several species of Penicillia .PT and PA are produced by many species in
the genera Aspergillus and Penicillium. Byssochlamys nivea also produces
PT
Patulin
51
penicillic acid
Chemical structure of cyclopiazonic acid
Stucture of zearalenone
52
Structure of alternariol
• Other Mycotoxins Produced by Fusarium
• Some fusaria are capable of producing mycotoxins other than
TCTCs and Fm. Zearalenone (ZE) a mycotoxin produced by
the scabby wheat fungus, F. graminearum (roseum), is of most
concern. Also called F-2, ZE is a phytoestrogen causing
hyperestrogenic effects and reproductive problems such as
premature onset of puberty in female animals,especially swine.
• ZE has been shown to bind with the estrogen and steroid
receptors, and stimulates protein synthesis by mimicking
hormonal action.
•
Zearalenone can be toxic to plants; it can inhibit seed
germination and embryo growth at low concentrations.
• Natural contamination with ZE primarily occurs in cereal grains
such as corn and wheat
53
• Fusarium verticillioides and related species, also produce
several other mycotoxins, including fusarins A-F, moniliformin,
fusarioic, and fusaric acid, fusaproliferin and beauvericin.
• Although the impact of these mycotoxins on human health is
still not known, fusarin C (FC) has been identified as a potent
mutagen and is also produced by F. subglutinans, F.
graminearium and several other Fusaria.
• Moniliformin, which causes cardiomyopathy in test animals,
may be involved in the Keshan disease in humans in regions
where dietary selenium deficiency is also a problem.
• Among many fungi, F. verticilioides is also most capable of
reducing nitrates to form potent carcinogenic nitrosamines.
These observations further suggest that the contamination of
foods with this fungus could be one of the etiological factors
involved in human carcinogenesis in certain regions of the
world.
54
Zearalenone
Alternariol
Mycotoxins Produced by Alternaria Species
Alternaria has been known for centuries to cause various plant diseases.
Species of this fungus are widely distributed in soil and on aerial plant parts.
More than 20 species of Alternaria are known to produce about 70 secondary
metabolites belonging to a diverse chemical group. However, only alternariol
tenuazonic acid, altertoxin-I, alternariol monomethyl ether (AME), altenuene
are common contaminants in consumable items like fruits (apples), vegetables
(tomato), cereals (sorghum, barely, oat), and other plant parts (such as
leaves)
The most common species of Alternaria, A. alternata (formerly known as A.
kikuchiana) produces all important Alternaria toxins including the five
mentioned above and tentoxin, alteniusol, alternaric acid, altenusin,
dehydroaltenusin
55
• Mycotoxins Produced by Other Fungi
• Sporidesmines, a group of hepatotoxins discovered
in the 1960s. These mycotoxins, causing facial
eczema in animals, are produced by Pithomyces
chartarum and Sporidesmium chartarum and are
very important economically to the sheep industry.
• Slaframine, a significant mycotoxin produced by
Rhizoctonia leguminicola (in infested legume forage
crops).
56
PREVENTIVE MEASURES
Management of Mycotoxin Contamination
• The economic implications of the mycotoxin problem and its potential
health threat to humans have clearly created a need to eliminate or at
least minimize mycotoxin contamination of food and feed.
• While an association between mycotoxin contamination and inadequate
storage conditions has long been recognized, studies have revealed that
seeds are contaminated with mycotoxins prior to harvest . Therefore,
management of mycotoxin contamination in commodities must include
both pre- and postharvest control measures
57
• Preharvest Control
• Mycotoxin contamination can be reduced somewhat by using
of resistant varieties (most effective, but not all are
successful) and earlier harvest varieties:
– crop rotation,
– adequate irrigation,
– control of insect pests.
•
Significant control of toxin contamination is expected to be
dependent on a detailed understanding of the:
– physiological and environmental factors that affect the
biosynthesis of the toxin,
– the biology and ecology of the fungus,
– the parameters of the host plant–fungal interactions.
• Efforts are underway to study these parameters primarily for
the most agriculturally significant toxins, namely AFs, Fms,
and TCTCs
58
• Use of atoxigenic biocompetitive, native A. flavus strains to
out-compete the toxigenic isolates has been effective in
significantly reducing preharvest contamination with
aflatoxin in cotton and peanuts.
• However, the aflatoxin contamination process is so complex
that a combination of approaches will be required to
eliminate or even control the preharvest toxin contamination
problem.
59
Mycotoxins and food chain
Fungal contamination
Vegetables
Many accidents
Animal
Production
Elimination
60
rare accidents (cancer)
Man
Products
(Animal origin)
Postharvest Control
After harvest, crop should not be allowed to over-winter
in the field as well as subjected to birds and insects
damage or mechanical damage. Grains should be
cleaned and dried quickly to less than 10–13%
moisture and stored in a clean area to avoid insect and
rodent infestation.
Postharvest mycotoxin contamination is prevalent in
most tropical countries due to:
•a hot, wet climate coupled with
•subadequate methods of harvesting,
(handling, and storage practices),
which often lead to severe fungal growth and mycotoxin
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contamination of food and feed.
• Sometimes contaminated food has been diverted to
animal feed to prevent economic losses and health
concerns. However, this is not a solution to the
contamination problem.
• Irradiation has been suggested as a possible means
of controlling insect and microbial populations in
stored food, and consequently, reducing the hazard
of mycotoxin production under these conditions .
• Significant emphasis has been placed on detoxification
methods to eliminate the toxins from the contaminated lots or
at least reduce the toxin hazards by bringing down the
mycotoxin levels under the acceptable limits.
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• I. Removal or Elimination of Mycotoxins.
– Since most of the mycotoxin burden in contaminated commodities
is localized to a relatively small number or seeds or kernels
removal of these contaminated seeds/kernels is effective in
detoxifying the commodity.
• Methods currently used include:
– (a) physical separation by:
–
–
–
–
identification and removal of damaged seed;
mechanical or electronic sorting;
flotation and density separation of damaged or contaminated seed;
physical screening and subsequent removal of damaged kernels by air
blowing;
– washing with water
– use of specific gravity methods
All these methods have shown some effect for some mycotoxins,
including DON, FmB, and AFB1
– (b) removal by filtration and adsorption onto filter pads, clays,
activated charcoal, etc.,
– (c) removal of the mycotoxin by solvent extraction
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II. Inactivation of Mycotoxins.
When removal or elimination of mycotoxins is not
possible, mycotoxins can be inactivated by:
(a) physical methods such as thermal inactivation, photochemical or gamma
irradiation,
(b) chemical methods such a treatment of commodities with acids, alkalies,
aldehydes, oxidizing agents, and gases like chlorine, sulfur dioxide, NaNO2, ozone
and ammonia,
(c) biological methods such as fermentations and enzymatic digestion that cause
the breakdown of mycotoxins. The commercial application of some of these
detoxifying mechanisms is not feasible because, in a number of cases, the methods
will be limited by factors such as the toxicity of the detoxifying agent, nutritional or
aesthetic losses of commodities during treatment, and the cost of the sophisticated
treatment.
Although several detoxification methods have been established for aflatoxins, only
the ammoniation process is an effective and practical method. Other chemicals such
as ozone, chlorine, and bisulfite have been tested and some effect for some
mycotoxins was shown in it. Solvent extractions have been shown to be effective but
are not economically feasible.
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III. Removal of Mycotoxins During Food
Processing.
• While cooking generally does not destroy
mycotoxins, some mycotoxins can be detoxified
or removed by certain kinds of food processing.
– For example, extrusion cooking appears to be
effective for detoxifying DON but not AFB.
FmB1 can form Schiff’s bases with reducing
sugars such as fructose under certain
conditions and lose its hepato-carcinogenicity;
but the hydrolyzed FmB1 was found to be still
65 toxic.
• Avoiding Human Exposure
Role of Rigorous Monitoring Programs
While it is impossible to remove mycotoxins completely from foods and
feeds, effective measures to decrease the risk of exposure depend on a
rigorous program of monitoring mycotoxins in foods and feeds.
Consequently, governments in many countries have set limits for
permissible levels or tolerance levels for a number of mycotoxins in foods
and feeds.
Over 50 countries of the world have developed such guidelines. For
example, levels varying from zero tolerance to 50 ppb have been set for
total AFs.
A tolerance level of 1 ppm for DON in grains for human consumption has
been set by a number of countries, including the United States. The FmB1
levels established by FDA in 2000 are limited to 5, 20, 60 100, 30, and 10
ppm, in corn and corn by-products to be used for horse and rabbit, catfish
and swine, and mink, poultry,
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• Among 77 countries which have regulations for
different mycotoxins, eight have specific regulations
for OA, with limits ranging from 1 to 20 mg/kg in
different foods.
• Regulatory guidelines to limit the presence of PT to
50mg/kg in various foods and juices have been
established by at least ten countries worldwide.
Details on worldwide regulatory issues and
permissive levels of mycotoxins in foods and feeds
have appeared in a number of recent reviews.
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• Detection and Screening of Mycotoxins
•
•
Because of the diverse chemical structures of mycotoxins, the presence of trace
amounts of toxins in very complicated matrices that interfere with analysis, and
the uneven distribution of the toxins in the sample, analysis of mycotoxins is a
difficult task.
Because many steps are involved in the analysis, it is not uncommon that the
analytical error can amount to 20–30%
•
To obtain reliable analytical data, an adequate sampling program and an accurate
analytical method are both important.
•
To minimize the errors, studies have led to many improved and innovative
analytical methods for mycotoxin analysis over the years.
•
New, more sensitive TLC, HPLC, and GC techniques are now available.
•
The MS methods have also been incorporated into HPLC systems.
•
New chemical methods, including capillary electrophoresis and biosensors are
emerging and have gained application for mycotoxin analysis.
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• After a number of years of research, immunoassays have
gained wide acceptance as analytical tools for mycotoxins in
the last decade. Antibodies against almost all the mycotoxins
are now available. Some quantitative and qualitative
immunoassays have been approved. Many
immunoscreening kits, which require less than 15 min. per
test, are commercially available.
•
Rather than analysis of toxin, PCR methods, based on the
primers of key enzymes involved in the biosynthesis of
mycotoxins, have been introduced for the determination of
toxicogenic fungi present in foods.
• Detailed protocols for mycotoxin analysis can be seen in
several of the most recent reviews and books and the most
recent edition of AOAC .
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• Dietary Modifications
• Dietary modification greatly affects the absorption, distribution, and
metabolism of mycotoxin and subsequently affect its toxicity. For
example, the carcinogenic effect of AFB1 is affected by nutritional factors,
dietary additives, and anticarcinogenic substances. Diet containing
chemoprotective agents and antioxidants such as ascorbic acid, and
even green tea, have also been found to inhibit carcinogenesis caused
by AFB1 in test animals.
•
The toxic effect of OA and FmB to test animals was minimized when antioxidants
such as vitamins C and E are added to the diet. Ascorbic acid also provided
protective effect against AFs.
• Most mycotoxins have a high affinity for hydrated sodium calcium
aluminasilicate (HSCAS) and other related products.
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CONCLUSIONS
• Mycotoxins are low molecular weight secondary
metabolites of fungi that are contaminants of agricultural
commodities, foods, and feeds.
• Fungi that produce these toxins do so both prior to
harvest and during storage. Although contamination of
commodities by toxigenic fungi occurs frequently in
areas with a hot and humid climate, they can also be
found in temperate conditions.
• Production of mycotoxins is dependent upon the type of
producing fungus and environmental conditions such as
the substrate, water activity (moisture and relative
humidity), duration of exposure to stress conditions, and
microbial, insect, or other animal interactions.
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• Although outbreaks of mycotoxicoses in humans have been
documented, several of these have not been well
characterized, neither has a direct correlation between the
mycotoxin and resulting toxic effect been well established in
vivo.
• Even though the specific modes of action of most of the
toxins are not well established, acute and chronic effects in
prokaryotic and eukaryotic systems, including humans have
been reported.
• The toxicity of the mycotoxins varies considerably with the
toxin, the animal species exposed to it, and the extent of
exposure, age, and nutritional status.
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• Most of the toxic effects of mycotoxins are limited to specific
organs, but several mycotoxins affect many organs. Induction
of cancer by some mycotoxins is a major concern as a chronic
effect of these toxins.
• It is nearly impossible to eliminate mycotoxins from food and
feed in spite of the regulatory efforts at the national and
international levels to remove the contaminated commodities.
This is because mycotoxins are highly stable compounds, the
producing fungi are ubiquitous, and food contamination can
occur both before and after harvest. Nevertheless, good farm
management practices and adequate storage facilities
minimize the toxin contamination problems.
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• A combination of natural biocontrol competition fungi and
enhancement of host-resistance against fungal growth or toxin
production could prevent toxin formation to a very significant
extent.
• Rigorous programs for reducing the risk of human and animal
exposure to contaminated food and feed also include:
• economically feasible
• safe detoxification processes
• dietary modifications.
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• Additional, systematic epidemiological data for
human exposure is needed for establishing
toxicological parameters for mycotoxins and the
safe dose for humans.
• It is unreasonable to expect complete elimination of
the mycotoxin problem. But multiple approaches will
be needed to minimize the negative economic
impact of the toxins on the entire agriculture industry
as well as their harmful effects on human and
animal health.
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