Post on 23-Jan-2022
rf FAOIUNEPIUSSR
nternationai TraIning Course 'TRAlNINO ACTIVJTIES ON FOOD CONTAMINATION CONTROL -
AND MONITORING WITH SPECIAL REFERENCE TO MYCO1OXINS
L. V. KRAVCHENKO
AFLATOXINS AND THEIR BIOLOGICAL
ACTIVITY
Centre of International Projects, GKNT
Moscow, 1984
AFLATOXINS AND T1lIR BIOLOGICAL AC1IVITY
L. V • CRAVCHEM(0
The group of aflatoxine (secondary metaboittea of micro-
soopie fungi of the Aepergillus genus)includea more than 10
compounds of similar chemical structure and biological, action.
It in mostly aflatoxins 81, B, 1' and 02 that are found in
natural conditions as oontaminante of food products and feed..
Out of four main representatives of eflatoxine, B 1 is most to-
xic and, as a rule, U is ayntheeized in the largest amounts.
The results of studying aflatoxine over 20 years mince
their discovery have shown that most mammals (including pri-
uiatee), birds, some species of fish, insects, and aicroorga-
nines are susceptible, in verioua degrees, to the toxic action
of efistoxins.
g.tsboliea of aflatoxin.
The alimentary pathway is the main form of entry of nfl.-
toxins in the organism, the principal and, in most of cases,
the only damaged organ being the liver.
Aflatoxin B 1 is found in the liver of rats as early as
30 minutes after being administered, and its concentration in
liver reaches the maximum level within two hours. After single
intraperitonsal administration, approximately 20% of labelled
toxin is retain.d in the organism of rats within 24 hourej the
highest concentration of the toxin is found in the liver. Si-
ailer re ults were obtained in experiments on mice, hamsters,
sheep, pigs, and poultry.
gxperiiente with labelled nflatoxin B 1 have shown that
I-I
-2- the main pathway of withdrawal of the toxin (in an unaltered
form or as a etabolite) Is its excretion with bile and urine,
and only a small anount is isolated with the exhaled air in
the form of CO2 . Most of investigators, studying the rate of
5etBp,olizRtiOfl of aflatoxine in different species of animals,
have found that the half-life of aflatoxin B 1 in the organism
is 12-15 hours.
Fxperiaier,te conducted in recent years, both in vitro, and
in vivo, have shown that afletoxins are metabolized by the ma-me enzymatic systems as other xenobiotice. Aflatoxin B may
be subject to hydroxylation by zircosomal oxidaaee with a mi-
xed function o give lees toxic metabolites -- aflatoxins
Q 1 1 and P 1 . Af].atoxin M 1 , in many species of animals, is one
of the main metabolitee which are found in milk and urine.Tkiva,
it has been detected in milk of cows, sheep, and goats which
had consumed feed contaminated with aflatoxin B 1 ,
The second possible pathway of detoxication of aflatoxin
B 1 in the organism is the reduction of cyclopentenone to afla-
toxicol with the participation of soluble cytosol dehydrogena-
sea. This reaction is revereable and therefore many authors
consider aflatoxicol as a "reserve" toni of aflatoxin B the cell.
Finally, atlatoxin B 1 , with the engagement of the eaae
enzymatic ayotea of the liver aicrosoisea, can be "activated"
i.e. it can be turned into compounds with a more pronounced
toxicity. It is supposed that one of such active forms of afla-
toxin B Is its hesiacetal-aflatoxth B2.,the other Corm being
ito 2,3-epoxide. It is also believed that epoxidation affecte '-4
-3-
the double bond of the terminal furane ring of the molecules
of the most toxic representatives of the 8flatoxin family--
aflatoxins B 1 , G1. and M 1 , whereas aflatoxinu B 2 and 02 whose
molecules do not have that double bond, possess a ich lower
biological activity. It should be emphasized that acute toxic
effect (aflatoxin B 25 ) and carcinogenic activity (2,3-epoxide
of aflatoxin Bi) of aflatoxins are primarily aesociatad with
these ctiveCmataboljtes of aflatoxin B 1 .
The 2 9 3-dihydrodiol of aflatoxin B 1 which is formed from
the epoxide, just as other derivativea of aflatoxin B 1 , may
produce in liver cello conjugates with glutathione, cysteine,
gluouronic and sulphuric acids and in this form be excreted
from the organism with bile or urine.
Biological activity of aflatoxino
Acute aflatoxicosis
Acute alimentary toxicoses aesociated with the ingestion
of contaminated feeds have been described in 1960 almost con-
currently for turkey-poulto, ducklinga, pigs, and calves. The
most susceptibie among farm animals are 3-12 weeks old piglets,
pregnant saws, and 1-6 months old calves. As for puultry, high
susceptibility to aflatoxine is found in turkey-poults and
ducklings; lees susceptible are young pheasants while chicken
are characterized by a relative resiatanca.
The leading clinical symptoma of acute intoxication with
aflatoxins is the absence of appetite, loss of body mase, and
a reduction in weight gain. It is necessary to emphasiz, a
1-2
-4- rapid development of ayiaptouia of intoxication and a high death
rate anong animal.. Aflatoxiooees in bird, are di.tingut.hed
by eymptome of the damage to the flerVOUe eyetem, in calves by
the dieruption of the funotion of the ga.tro-int..tinal tract,
in pigs and dogs by the development of Jaundice. Characteristic
symptoms of aoute intoxication are multiple haesorrhagio and
old ema8.
Aflatoxine are hepatotropto toxin., the target organ in
all species of animals being the liver. Aflatoxina oauee dif-
ferently expressed and differently localized neoroses of the
liver parenchyma and also adipoge and albuminoue degeneration
of hepatocytes. A oharacteriatio feature of the cotton of afla-
toxins is rapid proliferation of the epitheliva of biliary
duct..
Table 1 sums up eome data on the action of aflatoxin-
contaminated f..d on farm sni.male and poultry.
Changes in the liver, .iailar to those observed in tarn
animal., are found in experimental condition, in most of test
animal,. The LD50 values for some species are shown in Table 2.
Depending on the susceptibility to aflatoxin B 1 , the animali
may be aorted into three groupsa 1) very eu.ceptible for which
LD50 4 1 wg/kg; 2) suaceptible forwhtoh LD in 1-10 mg/kg;
and 3) re.istant for which LI)50> 10 mg/kg.
It should be noted that aflotoxin B 1 in most active among
aflatoxine. The relation between the toxtcitu of individual
repreaentUt ives of this group may be demonstrated on an example
of LD50 values for one-day old dicklirigs, which are 0.36, 1.70,
0.18, and 2.83 mg/kg, for aflatoxin. B, B, G I and 02' zeap.o-
-5- lively. LD of aflatxtn B1 for rots of the Fischer line in
1.16 .g/kg,a*d that of aflatoxin 01 is 1.5-2.0 !*g/kg; aflato-
zinc B2 and 0 2 are weakly toxic at doses in excess of 200 mg/kr
Interaction on the toxic action of aflatoxina upon prian-
te• is of definite interest. It has been danonetrated in •xp-
rla.nts on Theses aonk.y* and czab.atsre that aflatoxin Si
in doses ranging from 62 1gAg of the body mans to 5 rr4/kg
induses changes in the liver which are cbaiaotertstic of afla-
toxicosis and rapid (at high doe..) death of animals. A ope-
del syndrome ha* bien d..cribsd in young 9acnca faac1culari
f.aale, otter the int.rnal ad.tni.trs*ion of aflatoxin
in a dose of 0.5 or 1.5 mg/kg. The characteristic clinical
symptoms were aaughtaig, vomiting, diarrhea, and coma. The
changes in liver included osetrilobular neorosin, moderate
proliferation of btliarj, ducts, and nasal?, adipose degenera-
tion which was also observed in the heart and kidneys. Osdema
of the brain and the degen.ret ive changes of nary, cells were
also observed. Soma of these change, were sintlar to symptoms
noted in chtldsn who suffered from Beja's syndrome which will
be described later on. Thug, the esnaitivity of monkeys to
acute action of aflatoxins in indubitable.
As seen from rable 1 and 2. there are con.id.rable inter-
specific differ.ncea in eusc.ptibtltty to e.flatxina. The rea-
son, according to anny tnvcctigatore, in due to the differences
in the rates of afletoxin metabolization between the epectea;
other authors believe it to be associated with different path-
ways of metabolism.
It is noteworthy that there are differences in euscepti-
1-3
Cattle
calves
bulls
adult oow•
: 08//day day
0.5 aflatoxin B 1] kg/day
0.7 mg/kg feed 1.0 mg/kg feed
2.0 ag/kg feed 16.0-46.0 rnpg/kg
0.6-0.9 mg/day/oow
iforses 0.015 aflatoxin B 1 kg/day
0.15 mg/kg/day 0.3 mg/kg/day
Piga mase 6.5 kg 0.62 mg/kg
mae. 20 kg 0.26 mg/kg feed 0.065 mg/kg/day
mass 22 kg
Poultry chicken
broilers
2 - 4 mg/kg feed
0.25 mg/kg feed i-i mg/kg f.ed
0.25 mg/kg feed
0.6 mg/kg feed 1.5 mg/kg feed 2.5 mg/kg feed 5 - 10 mg/kg feed
laying hens 2 - 8 mg/kg feed
20 mg/kg feed
-6-
Table I Toxic action of feeds contaminated with aflatozins on
farm animals and poultry
Animal
Doe. 1ff.ot
Drop in weight gain Drop in weight gain, ooagulopathy Coagulopathy. n.oroeee of liver, death
Drop in wght Death (59
eiday.)
gain
Drop in milk yield Deteotion of aftatoxin in milk Detection of aflatoxin in milk
Disruption of liv.r funo-tion, jaundice, death on the 37th day Ditto, death on 26th day Ditto, death on 12-15th day
Corresponds to LD 0 (internally) Growth retardation Suppression of i,mnuno-gene aim Acute toxicosis, death
Drop in weight gain Necrosis of the liver, death
3uppression of iiwnuno-genesis Drop in resistance Drop in weight gain Coagulopathy Necrosis of the liver, death Reduction in egg laying capacity Reduction in egg-laying capacity, detection of aflatoxin in eggs
'-I
-7- Table 2
Value. of LD50 of aflatoxin B 1 for some epecise of farm
and laboratory animal. (aingi. adasini.tration)
Animal LD50 , ag/kg of body mace
Duckling. 0.34-0.56
Rabbita 0.3 -0.5
Rainbow trout 0.5
Oat. 0.55
tnk 0.5 -0.6
Pig. 0.62
Dog. 1.0
Guinea pies 1.4 -2.0
3h..p 2.0
Monk.y. 2.2
Rates
newborn 0.56
w.enitnge 5.5
adult male. 7.2
adult female. 17.9
Chicken 6.5 -16.5
tc. 9.0
flasneter. 10.2
Chicken embryo. 0.025pgA,ggj
1-4
-8- bility to aflatoxina even among breede of one and the ammo
epectee. For inetance, the otudy of 18 etraine of chicken,
turkey-poulte and quiii indicated that only one breed -- New
Hampshire-- is diotinguishad by high euaceptibility to aflato-
un B 1 . A comparteon of the auoaptjbtiiti.e of the enbryoa
of different breede of hen to aflatoxin B revealed that most
remietnat to this aflatoxin are Rhode Inland enbryoe and the
leaet euoceptible are the enbryoe of the White I'iymouthrock
breed.
In vitro etudiem of different oyeteae have greatly faci-
litated the elucidation of the btoloical activity of efiato-
xina. Thum, toxic propertlee of aflatoxina have been proved in
relation to culturee of chick embryo lIver, baby rut liver,
lunje, and kidneya, calf and monkey kidneya,human liver, eta.
In theae eyatemo, the activity of aflatoxin 8 I was damonatrat-
ad at a done ranging from 0.01 to 10 jig/mi. Ito toxicity was
exprenoed in the changes in the morphology of cultivated cells
and in the dieruption of their functional activitys auppreasion
of the myntheota of nucleic ooida and protein.
The information about high eumoeptibUtty of tienue oalle
of man --liver, lunge, blood cello, and akin fibrobleete --to
aflatoxing is of intereet. Along with eflatoxin B 1 , toxic cotton
on the celia of hunan embryo liver is exerted by aflatoxia.
20 a 2 .and Gi. Numeroua obeervatiOne and experimento have indicated
that afletoxina are atrong immunodepreasante, which woetly
affect cellular immunity. The P ayetem of cellular immunity
is diotinguiehed by extremely hig eunceptibility to aflatoxine '
- 9- 9 1 and m i . Afletoxine also influence the meohanieme df non-
.p.oifio r..ietance of the orantme - the nyntheete of some
freotione of the ooepl.ment. the production of interferon.
etc.
Chronic afluitoxtoocie
Chronic intoxication with aflatoxins entails the develop-
meat of malignant tuui3ure of the liver. At prenent, aflatoxinu,
primarily eflatoxin B, are damned with the ntron.et chemical
ceneirog.na.
When adsinietered internally, aflatoxine induce hepatomna
in all thus far studied epeciee of animals, mp.cificeuly in
rat, of the Pincher, Wistar, and Porton eli-nine, ducklinpm,
chicken, rainbow trout, malson, guppi.e, pole cat., doge and
monkeys.
A linsar depndence of the frequency of h.patocellular
cercinoase upon the do.s of afla toxin BI in the ret ion was ob-
ssrv.d in rate of the Pincher strain: at aflatoxin concentra-
tion of 1 pg/kg the fr.qu.ncy of tumoure was 10%, at a conoen-
tretion of 100 pg/kg it was 100% (?abl. 3). The frequency mdi-
cen of cancer in the lifetim, of rate, theoretically calculat-
.d by the results of various experimental studien, were 240/10
at a concentration of aflatozin B 1 in the ration 0.1 pg/kg and
1100i105 rate at a concentration of 0.3pjkg of feed.
1-5
Tabi. 3
Dependence of carcinogenic activity of aflatoxin
in male rate of the Fischer strain upon its cont.nt
in the ration
Aflatoxin Duration of Frequency garliest time
concentration, feeding, weeks of liver of the develop-
g/kg carcinoma meat of the tumour, weeka
0 74-109 0/18 -
1 78 - 105 2/22 104
5 65-93 1/22 93
15 69-96 4/21 96
50 71 - 97 20/25 82
100 54 - 88 28/28 54
single intraperitoneal administration of aflatxin
in fewale rate at a does correepondSng to LD50 aloe led to
the development of h.pato.as in coven out of 13 rat. within
60-128 week..
Carcinogenic action of afletoxine may manifest itself
H1SO in the progenys the development of cholangiooarotaomae
has been observed in bady rate subjected to prenatal (intrau-
term.) or poet-natal (through mother's milk) exposure to afia-
toxin Da
There are reports about the development of tumour, out-
cide the liver as a reault of the attni.tratiOn of aflatoxin
carcinoama of the stomach, ad.nocaroinomas of the large
-11- intestine, kidnwys and lungs, tumoure or the salivary glands,
tDngue, and oeaophagus, The cases of the development of ear-
comes in the place of subcutaneous edainiatration of eflato-
un are also described.
0arcinogentc properties of other aflatoxine --B 20 0 1,
020 M i are lees pronounced. When afletoxin 01 was administered
to rete with dringkng water it induced not only tunoure of the
liver but also tumoure of the kidneys. Aflatoxin B2 in a total
do.s of 150 ag per animal induced hepatocellular carcinomas in
three out of nine rate within of 57-59 weeka. In the name as
rice of experiments, aflatoxin Bi administered in a done of
1.3 ag per animal induced hepatomas in 9 out of 9 rate within
46 weeks. These results indicate that the effective done of
afletoxin B2 in 115 times higher than the dose of qflatoxtn B 1
giving ris, to hapatoinae in ret..
Unlike rats, nice manifest wall-pronounced resistance to
the carcinogenic action of aflatoxine administered internally.
Prolonged consumption of aflatoxin Bi by aloe at a concentra-
tion up to 1000 pg/kg of the feed failed to induce any tum3urs.
At the seas time when aflatoxtn Bi was intraperitoneally admi-
nistered to baby mice in a doe, of 1,25 pg/kg during the first
7 days of life or in a dose of 6 pg/kg during three day., h.pa-
tome, were found within 80 weeks (Table 4).
Rainbow trout I. known for high susceptibility to aflato-
xinn. Th. inclusion of aflatoxin B 1 into its feed at a rate of
0.1 pg/kg induc.i the development of hepatoaae withIn 20 month..
Aflatoxi'i K 1 manifests Itself an a weaker hapatocaroinogen in
experiments i4th trout (Table 5).
1-6
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-13..
Table 5
H.pstocsroinog.n.city of aflatoxin 9 in rainbow
trout as ooapor.d to aflatoxin
Aflatoxin Liver tumzr frequence
1v.l in males females the ration,
4.0 4/28 (14%) 13/27 (48%)
16.0 22/21 (81%) 11/14 (79%)
i 32.0 24/25 (96%) 13/14 (93%)
K I 64.0 21/24 (88%) 9/10 (90%)
5 14.0 15/22 (68%) 18/23 (78%)
For a long time (up to 1971) primate, were believed to
be r..iutant to carcinogenic action of aflatozina. It was
only in 1972 that Gopsian and collaborators publiuhed a report
about the development of hepatooellular carcinoma in a male
rh.aus monkey which had been fed with partially purified afla-
toxin. B, and 01 for 5.5 y.ar.j the carcinoma was detected 8
years aft.r the comeenoement of the experiment. Purtheron,
the.ei re.ults were repeatedly confirmed by other authore. The
h.pathocarcinog.nio •ffect of aflatozine 8I and of a mixture
of aflatoxin. B 1 • 82 + 01 + 02 ha-s been deiaonatrated for
marmoaete and tupatae of both aexea. It ahould be nted that
attempts at determining the doac dependence of the effect have
not been undertaken for prinatem.
I-?
-14- Teratogenic and nutagento action of afintoxins
Teratogenic.properti.s of aflatoxin Bi have been deter-
mined in experimental conditions for hamsters, rate, mice,
chicken, and Jepaname smooth snake (Ory2iue latipee).
Atlatoxin administered to hamsters on the 8th day of pre-
gnanoy induced malformations in 29.4% of tootua.e. The add-
nietration of aflatoxin B 1 at a concentration of only 1.0
g/kg to female wiatar rate every other day for the firet 14
days of pregnancy entailed the death of 1% of embryos, resorp-
tion of 6.8% of embryoe, while 3.5% of fetuses had various mal-
formations of developments iicrooephalia, hernias, bradideoty-
lies.
In 11.5% of mice embryos exposed to aflatoxin B 1 , at the
8th day of intrauterine development various malformations were
observeds brain herniae, anomalies of the gastrointestinal
tract.
The administration of aflatoxin B 1 to the yolk sack of hen
eggs at the 6th day of incubation entailed the development of
malformations in 65-90% (depending on the doss) of embryo..
Aflatoxin 81 induces ohro.osoaal aberration, and breakups
of the DNA in plant and animal cell.. It has been likewise de-
monstrated that it produces mutationa of genes in bacterial
test-systems (the Ames test) after mstaboliaatton ("motivation")
by mioromomal preparations from the rat or buman liver.
Studies of recent years have demonstrated that autagenio
and txigenio propertise of the precursors of aflatoxin B 1
(during its biosynthesis) increase as their structure becomes
more complex and reach their peak in aflatoxin Hi. The dipsat-
-15-
.no. of mutagento prop.rtise of aflatoxin B 1 upon the presence of the laotons ring in its structure has been demonstrated.
Factors. influencing the biological activity
of aflatoxin.
A lerge amount of data nocumulated till now demonstrates
the possibility of modification of toxic, oaroinogenio, and
other manifeetationi of the biological activity of aflatoxine
within broad limit, by using different factors.
The toxic action of aflatoxine denda considerably upon
the age and sex of animale. The common feature for all species
is a decrease in their susceptibility be aflatozin.e with age.
A. was seen in Table 2, LD50 for newborn rate is almost 1/10
of that for weaned rate and is 1/13 of that for the adult male
rat..
Numerous studies have demonstrated that females are more
resistant both to the acute toxic and to the carcinogenia ac-
tion of atlatoxinu compared to males. Table 2 also shows that
adult mal, rats are approximately 2.5 times more susceptible
to aflatoxin B 1 than female. (LD50 7.2 and 17.9 mg/kg of body
mass, reepectively). It is not without interest that when afla-
toxin B 1 I. included in the ration, of rate of both sexes, the
frequency of pre-cancer changes in the liver is practically
the same for male, and females, but the period between the
appearance of these change, and the development of heatic
carcinoma in females in much longer than in males. Table 6
gives the estimated figures of the total amounts of aflatoxin
later I the organism of animals (male and female rats) in
the period preceding the appearance of the liver tumoure.
1-8
-16- Table 6
The dependence of carcinogenic activity of aflatoxin
upon eax of rate
Concentration Total dome of afla- Time of detection of the of eflatoxin Bi toxin B 1 , mg/kg liver tumoure, daym mg/kg of the anital
ration malem femalea malea fernalem
1.0 2.9 5.9 245 448
0.015 0.095 0.115 476 560
There are all grounde to believe that the revealed sex
distinctions in the eunceptibility of animals to aflatoxine
are determined by differencee in the hormonal background. The
adininiatration of diethyletilbeetrol concurrently with afla-
toxin 81 to male rate entatle coneiderable lowering in the
frequency of liver tumoura (8 out of 40 as against 25 out of
35 in the control). The development of tunoura in the liver
in mal, rate under the influence of aYlatoxin Bi was prev.nted
by a preliminary hypophyeotomy and also by oaatration of the
animala. The adminiatrntton of teetoeteron along with afletoxin
to omatrated rata entailed death of all experimental animale.
There is special interest in the information about the
influence of the factore of nutrition upon the biological ac-
tivity of tiflmtoxine. It has been demonstrated that under the
conditions of protein inaufficienoy the frequenoy of tumou.ra
of the liver under the action of aflatoxin 81 is much higher
than against the background of full-value nutrition and the •
tine of the development of tumours is conniderably ieee.
-17- Thu., in rate which were fed on rations containing 9% of pro-
tein, tumoure of the liver developed in 11 out of 15 animals
after the passage of 8 months whereas in rate which consumed
ratione with 22% of protein tumours developed in 7 out of 14
animal. within 10 months . In the case of protein insufficien-
cy, the acute toxic effect of aflatoxina is also more pronoun-
ced. At the same time there is a different information mdi-
onting a decrease in the carcinogenic effect of aflatoxine un-
der the conditions of protein deficiency in the rations. It
was shown that not only protein insufficiency but also the de-
ficiency of certain amino acids (tryptophan, specifically) may
aggravate aflatoxtoosie.
Speaking of other alimentary factors capable of modifying
the biological activity of aflatoxins, we should single out
lipotropic agents, some vitamins, and mioroelenants. The in-
sufficiency of .ethionine and choline in the rations has a
protective action in relation to the acute toxic effect of
aflatoxin B 1 , but reliably intensifies its carcinogenic acti-
vity in experiments on rate. At the e&ne ttae, a more pronounc-
ed deficiency of these lipotropto substances in the rations
rather decreases, than increases, the frequency of hapatocarci-
no.ae in rate which were given aflatoxin Bi. It should be lIk,e-
wise etr.seed that the effeot of lipotropto substance, which
modify carcinogenic activity depends largely upon the amount
and quality of fats in the ration of experimental animal..
The final biological effect of aflatoxine is strongly
influenced by the availability of vitamins. For instance, the
defl.cency of vitamin. A and C produces inhibition of the mete-
-18-
boltam of afletoxin B 1 . Som authors etrese that whenever the-
re is a deficiency of vitamin A, there is a greater frequency
of carcinomas of the large intestin, in rate.
Pronounced protective action in relation to aflatoxina
has been d.aonetrat.d by selenium and copper added at defini-
te oonc.ntratione to the rations of rate.
inally, attention should be drawn to the activation of
inicrosoinal oxidases with a mixed function under the influence
of ethanol which may intensify the formation of the waotiv.
form of aflatoxin B 1 its 2,3-spoxide-- and increase thereby
the hepatocarcJnogenio activity of atlatoxin B 1 .
It seems that most of factors modifying the aflatoxin
toxicity act by changing the activity of enzymatic systems
which metabolize aflatoxins. This supposition is confirmed by
the results of study of the influenc, of inductor. and Inhibi-
tors of rnicroeoaal oxidaaee with a mixed function upon the
biological activity of aflatoxins. Thus, when test animals
are given phenobarbital which inducee the activity of oxidas,.,
the toxic effect of aflatoxin B 1 decreases which manifests it-
self in the reduction of the inhibiting action of the toxin on
the iyntheaie of the protein, the prevention of the liver nec-
roses, the weakening of aflatoxin's carcinogenic properties.
At the same time, the administration of an inhibitor of microso-
mel oxidasee (5KP525A) was accompanied by an intensification
of the damaging action of afletoxin B1 upon the liver.
Thus, the results of eudes of aflatoxicosee in farm ani
male and the experimental findings enable us to class aflato-
xina with most potent hepatotoxic and hepatocarciaogenic to-
-19- xins. It should be stressed that the extent of biological acti-
vity of these microtoxins depede creatly upon the species
of animal., their age, sex, and the nature or nutrition.
Action of aflatoxina on man
Clinical observations
Since eflatoxine were found to possess strong hepatotdxio.
and hepatooaroinogenio propert tee, the relatively high frequen-
cy and level of the contamination of food products with atm-
toxins and the abundance of aflatoxin producer in natural con-
dit tons make us to olaee these aiorotoxine with biological pol-
lutants of the environment which are potentially dangerous for
man's health.
cut, aflatoxicoees in humane are rare and are associated
with high concentrations of aflatoxine in food (ranging from
0.2 to several mg/kg). All cases of poisonings occurred in
countries distinguished by a high level of contaia.nation of
food products with aflatoxina (Table 7). The sicknee of child-
ren in Senegal was caused by peanut flour containing aflatoxin
at a concentration of up to 1.0 mg/kg. In India, children in
the ae group from 1.5 to 5.0 years when treated for the e'n-
droms of protein insufficiency - kwashiorkor—were given pea-
mid flour which contained aflatoxin B 1 at a concentration of
0.3 mg/kg. The average daily dose of the toxin in this case
was 1.1 /kg of body mess. In on. of described case., the
liver of a 15 year-old boy which died of acute hepatitis de-
monstrated changes which are obaraotertotiâ of aflatoxiconia.
20- I .
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-21-
The diaeaee was caused by maniac containing aflatoxin B,at a
concentration of 1.7 mg/kg.
A convincing instance of the association of aflatoxine
with acute hepatitis in people was the outbreak of toxic hepa-
titis in the North-West districts of India in 1974. The disea-
se was characterized by a subacute commencement with a fever an
a subsequent rapid development of jaundice (98% of cases) and
aecttis. The patients demonstrated hepatoeplenomengalia. Liver
section obtained by biopsy and autopsy demonstrated a charao-
t.riatio proliferation of biliary ducts. The death rate was
very high. The analysis of food products indicated that the
cause of the dioeaee aen maize which contained aflatoxin 8 1
at a concentration of up to 15.6 pg/kg. with this level of
contamination, the daily ingestion of aflatoxin ranged from
2 to 6 mg/per person which corresponds to daily doses of up
to 120g/kg of body mass.
Special mention should be made of information about the
influence of aflatoxin upon man in industry. In one of the
instance, 7 people out of 55 who were angaged in the process-
ing of peanuts and other oil-bearing crops, demonstrated the
development of cancer of varying localization (the period of
observation was 11 y.ars, the time of exposure was 2-3 years).
The concentration of aflatoxins in air in this case could have
been in a range of 0.87 to 72 ig/a3. Two oases of pulnonary
adenoizatosie with a lethal outoome have been described in pe-
ople handling Brazilian peanuts. Changes characteriutio of the
action of aflatoxin B 1 were found in their lungs. Finally, a
report is known about finding carcinoma of the large inteutine
-22- in two reeearch workers who for a nuaber of years were engaged
in the isolation and purification of afletoxine.
The study of patients suffering of cirrhosis of the liver,
residing in the regions of Iran where this disease is believed
to be frequent, revealed the presence of ntis toxin 9 in the
urine of six out of 26 patients. Aflatoxin was never found in
the urine of patients with other diagnoses. The patients came
to the clinic from villages which were noted for a high con-
centration of aflatoxin 9 in milk.
The poesible relation of Rays's syndrome with the oontamt-
nhtion of food by eflatoxtha is still debatable. Out of four
countries where this relationship was studied (Thailand, New
Zealand, USA, and Czechoslovakia) and where aflatoxin B 1 had
been found in the liver of patients, only Thailand is situated
in an area with a high frequency of contamination of food with
aflatoxime. The three following reporte are noteworthy.
In Thailand, the autopsy of 23 children deceased as a
result of Rays's syndrome revealed considerable amounts of
aflatoxin B 1 (up to 93/kg) in the liver, in the content of
the stomach and the intestines (up to 127)og/kg), and in bill.
Traces of aflatoxin B 1 have been also detected in other tis-
sues-brain, kidneys- and in the urine.
Reports from Czechoslovakia present results of clinical
observation of 27 children in tte age group from 3 days to 8
years who also died of Reye'F syndrome. The disease was cha-
racterized by an acute oneet. In some cases, brain ey.ptoas
prevailed and the disease lasted from 2 to 3 months. In suba-
cute canes, periportal fibrosis and the oroitferation of the
bile ducts were observed in the liver: when the disease con-
-23- ttnu.d for 4 month., there were syaptoma of cirrhosis. In all
cases, aflatoxin B 1 was found in liv.r tissues at a concentra-
tion of from 20 to 2760 /kg; in three paces aflatoxin U 1
was also found.
In 1979 Rajan and collaborators published results of an
anaijais of aflatoxine in the liver of 8 children with Raye'a
syndrome. In six cases, the concentration of aflatoxin 21
ranged from 2.23 to 17.33g/kg of the tissue. In two children 0
during the acute stage of the disease, the aflatoxin was found
also in blood at a concentration of 11.93 and 31.3 mg/aL There
are reports of other investigators about cases when aflatoxin
B 1 was found in blood serum of patients suffering from Reye's
syndrome.
Thus, available inforinatin makes it possible to conclude
that aflatoxins may play a definite part in the development of
Rays's syndrome in humans in some areas. At the same time we
cannot exclude the possibility that the pathological changes
peculiar to Rays's syndrome, may land to the disruption of
metabolism and elimination of aflatoxis, thereby retaining
them in Ih. organism.
Bpideniological studies
The results of studying a possible correlation between
the level of contamination of food products with aflatoxthe
and the frequency of primary cancer of the liver inhumans
are of considerable interest.
Primary cancer of the liver in guropsan countries compri-
ses 1.2 per cant of all cancer cases, in USIA 2.5-2.8 per cent,
in African countries 14%. The highest frequency of this disease
-24- in obnerved in Bantu male, in South Africa (Moznmbtque) up to
68-77% of all eancr case..
Epide,iologtcal studien indicate a correlation between the
level of daily ingestin of aflatoxine and the frequency of pri-
mai'y ennoer of the liver in some pert, of Ic.nya, Mosambiqus,
Swaziland, and Thailand (Table 8).
Most interesting are the date about the age distribution
of the frequency of prihery cancer of the liver among the p0-
pulatian of Nozaabique. It is noteworthy thr.t in areas with
a high frequency of this dinease the peak In found in the early
age period - 20-29 year., whereas in areas with a low frequency
of primary cancer of the liver, the morbidity increased with
age gradually. This pronounced "juvenation" of prinary cancer
of the liver may be explained by a greater susceptibility of a
growing organism to the carcinogenic action of aflatoxine.
Some authors believe that the viru, of hepatitie B which
in widely spread in countries with a high incidence of primary
cancer of the liver say also act as a cofactor in the etiology
of this disease.
In studies undertaken in Indoneeta on 71 patients euffer-
ing of primary cancer of the liver, aflatoxine were found in
biopsy samples of the liver in 57.7 case.. Anatnnesttc data in-
dicatod the consumption of contaminated food products, includ-
ing a prolonged daily consumption of peanuts. Aflatoxin B 1 was
found in eeapiee of food products at concentrations ranging
from 17 to 1190 pg/kg while afla toxin G, was found at concen-
trations ranging from 5 to 630 , g/kg.
In the United States, we know of a case when aflatoxin B1
00 C U 5-'
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U 1p
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4.
1-4 0
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0 0 U v-s 14 0
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U U U ' U ' aS C U N • .0 U P U U . U 0 b4 El Cl) I')w 0 1-4 Ct) .
-26- at a ooneentratton of 520g/kg of wet weight was found in
the lifer ttseu• of a patient suffering of carcinoma of the
rectum and liver.
Published epideiologioal studies are limited in volume
end beidee, they evaluate but one of the possible etiol3gic-
.1 factorej the influence of aflatoxine. There are no doubts
that other factors • such as malnutrition, viruses, other my-
cotoxine, plant alkaloids, tielminthiasee, may also play an
etiological or a modifying part in the development of cancer
of the liver.
Conclusion
Thus, the observations of aliientary aflatoxicosee among
farn animals in many countries and the results of numerous
experimental studies demonstrated that alfetoxins are highly
toxic compounds which affect mainly the liver. Acute aflatoxi-
cosia to characterized by the development of necroees of h.pato-
cytea and the proliferation of the biliary ducts; chronic into-
xication may entail cirrhosis of the liver and the dsvelpment
of hops tomas.
Afletoxin B 1 induces chromosomal aberrations and ruptu-
res of the DNA in plant and animal cells, and in some baot.-
riml teat systems - gene mutations after activation with 'Uc-
rosoniel enzymatic systems. High concentrations of afletozias
have a teratoganio action upon some species.
The biological activity o' aflatoxins is derendent on the
age and sex of aniiale. The nature of nutrition, and priaarily
the amounts of protein, lipotropic substances, and some vita-
-27-
mina, may materially modify the course of aflatoxicoses.
A correlation has been found between the level of afla-
toxin B 1 arriving with food and the incidence of cancer of
the liver in man, in areas with high frequency and high level
of contamination of food produots with atlatoxine and high
frequency of the primary onnoer of the liver.
At the macne time, the number of trends in the study of
biologial aotivity of aflatoxine call for further develop-
ment. It is necessary to have a more detailed information about
the abaorpt ion of nflatoxina in the gastrointestinal tract and
about the rate of tJeir withdrawal from tissues. We find it
important to study the modify!ng role of alimentary factore
and of other biologically active substances in the manifesta-
tion of the toxicity of aflatoxina. This aspect should be ta-
ken into ooneideration also in epiderniological studies related
to the connection between primary oanoer of the liver in man
and intoxication with low concentrations of aflatoxins.
The suppooition about the casual relation between the
consumption of aflatoxina and primary Cancer of the liver in
man and also about the role of eflatoxins in the developaent
if Reye'e eundroae requires a further substantiation.
28
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3. 1230 flHK BHHHTFI