The present invention relates to an aflatoxin production inhibitor and a method for
controlling aflatoxin contamination.
Description of the Related Art
While secondary metabolites of fungi contain useful compounds, they also contain
many toxic compounds, which are referred to as mycotoxin. At present, mycotoxin
contamination of crops has become a serious problem all over the world. In order to stably
obtain safe food, there is a need for ways to control mycotoxin contamination.
Among mycotoxin contamination of crops, the most serious one is aflatoxin
contamination of crops. It is known that aflatoxin has the strongest carcinogenicity and acute
toxicity among known natural substances. Since aflatoxin is a compound that is not degraded
by a normal food preparation method, its health hazard is serious. Death related to ingestion of
aflatoxin-contaminated crops has been frequently reported. In only China, hundreds of
thousands of people have died per year by liver cancer presumably caused by aflatoxin.
In order to prevent such health hazard, the value of aflatoxin allowed to be contaminated
in crops is regulated to as low as about 10 ppb. Thus, loss caused by disposal of crops
contaminated with aflatoxin comes to a large amount. For example, in United States, loss
caused by aflatoxin contamination comes to several ten billions yen per year. In Asia, there is
more loss caused by aflatoxin contamination, which is estimated to be over one hundred billion
yen per year.
Aflatoxin is mainly produced by Aspergillus flavus and Aspergillus parasiticus, which
are known to infect crops such as maize and peanut and produce aflatoxin in cultivation and
2
storage stages of the crops under tropical and subtropical environments (See, Council for
Agricultural Science and Technology., "Mycotoxins: Risks in Plant, Animal, and Human
Systems", CAST, Ames, Iowa, USA, 2003 and Shun-ichi Udagawa, Setsuko Tabata, and Mitsuo
Nakazato, "Mycotoxin", Chuohoki, 2002). Due to climate change by global warming in recent
years, it is concerned that contamination may extend to regions other than tropical and
subtropical regions.
Conventionally, in order to control aflatoxin contamination, there have been conducted
basic research such as genomic analysis of aflatoxin-producing fungi and identification of a gene
that is involved in aflatoxin production, and practical research such as acquisition of an
infection-resistant variety and reduction of contamination by competition with aflatoxin
non-producing fungi. However, an efficient and drastic method for controlling aflatoxin
contamination has not yet been established.
Possible methods for controlling aflatoxin contamination include, for example, a
method using an antifungal agent that inhibits growth of aflatoxin-producing fungi.
Specifically, it has been known that mycotoxin such as aflatoxin can be decreased in maize
before and after harvesting and during storage by using, in combination, one or two or more
selected from the group consisting of azoles such as cycloproconazole; strobilurins such as
azoxystrobin; and other microbicides such as boscalid and fludioxonil (see International
Publication No. W020091068 195). In W020091068 195, what conditions under which
mycotoxin such as aflatoxin is produced were specifically examined, and various
countermeasures against them were considered. Finally, it was concluded that the best way to
suppress production of mycotoxin itself is to suppress proliferation of mycotoxin-producing
fungi. However, use of microbicides that suppress proliferation of mycotoxin-producing fungi
may cause spread of fungi resistant to the microbicides.
Since aflatoxin is a secondary metabolite, it is considered that inhibition of aflatoxin
production does not affect growth of aflatoxin-producing fungi. Thus, an agent that specifically
inhibits only aflatoxin production can provide an effective method for controlling aflatoxin
contamination.
As a result of search for substances that inhibit aflatoxin production, it was found that
dichlorvos which is an organophosphorus insecticide, and tricyclazole which inhibits a melanin
biosynthetic enzyme, have aflatoxin production inhibitory activity (see L. L. Zaika and R. L.
Buchanan, J. Food. Prot., 1987,50,691). However, these compounds have not come into
practical use because the aflatoxin production inhibitory activity of these compounds is weak,
and there are problems in safety of the compounds themselves and in selectivity of the inhibitory
activity.
As mentioned above, there is a need for compounds that do not cause spread of resistant
fungi and have an activity to specifically inhibit only aflatoxin production. As a result of search,
some compounds have been found (see Japanese Patent Application Laid-Open (JP-A) Nos.
09-24 1 167 and 1 1-0799 1 1). However, in the present situation, a satisfactory aflatoxin
production inhibitor that is highly safe and highly practical has not been found yet.
SUMMARY OF THE INVENTION
The present invention aims to solve the above existing problems and to achieve the
following objects. That is, the present invention aims to provide an aflatoxin production
inhibitor that inhibits aflatoxin production specifically and efficiently, is highly safe, and is
practical; and a method for controlling aflatoxin contamination.
The present inventors conducted extensive studies to solve the above problems, and
have found that a respiratory inhibitor or a certain respiratory inhibitor has an excellent aflatoxin
production inhibitory activity, to thereby accomplish the present invention.
The present invention is based on the aforementioned finding by the present inventors,
and means for solving the aforementioned problems are as follows:
(1) An aflatoxin production inhibitor, including a respiratory inhibitor.
(2) The aflatoxin production inhibitor according to (I), wherein the respiratory inhibitor is a
respiratory inhibitor applied at a concentration at which an amount of fungus body of an
aflatoxin-producing fungus is 50% or more relative to that of an untreated group, or a respiratory
inhibitor by the action of which an amount of fungus body of an aflatoxin-producing fungus is
4
50% or more relative to that of an untreated group.
(3) The aflatoxin production inhibitor according to (1) or (2), wherein the respiratory
inhibitor is at least one selected from the group consisting of rotenone, siccanin, atpenin A5,
antimycin A, pyridaben, tolfenpyrad, fluacrypyrim, acequinocyl, bifenazate, mepronil,
metominostrobin, pyribencarb, and cyazofamid.
(4) The aflatoxin production inhibitor according to (1) or (2), wherein the respiratory
inhibitor is fluacrypyrim.
(5) The aflatoxin production inhibitor according to (1) or (2), wherein the respiratory
inhibitor is at least one selected from the group consisting of rotenone, siccanin, atpenin A5, and
antimycin A.
(6) A method for controlling aflatoxin contamination, including treating a plant to which an
aflatoxin-producing fungus may be attached, a fruit which is harvested from the plant, the h i t
during storage, or any combination thereof, with the aflatoxin production inhibitor according to
any one of (1) to (5).
The present invention can solve the above existing problems and achieve the above
objects. Specifically, the present invention can provide an aflatoxin production inhibitor that
inhibits aflatoxin production specifically and efficiently, is highly safe, and is practical; and a
method for controlling aflatoxin contamination.
DETAILED DESCRIPTION OF THE INVENTION
(Aflatoxin production inhibitor)
An aflatoxin production inhibitor of the present invention contains at least a respiratory
inhibitor; and, if necessary, may further contain other ingredients. The aflatoxin production
inhibitor refers to those inhibiting aflatoxin production. A degree of inhibition of the aflatoxin
production is not particularly limited as long as an amount of the aflatoxin production can be
suppressed compared to that of an untreated group.
The aflatoxin production inhibitor of the present invention is preferably a respiratory
inhibitor applied at a concentration at which an amount of fungus body of an aflatoxin-producing
5
fungus is 50% or more relative to that of an untreated group, or preferably a respiratory inhibitor
by the action of which an amount of fungus body of an aflatoxin-producing fungus is 50% or
more relative to that of an untreated group. The "respiratory inhibitor applied at a
concentration at which an amount of fungus body of an aflatoxin-producing fungus is 50% or
more relative to that of an untreated group" means that the respiratory inhibitor can suppress
survival or proliferation of the aflatoxin-producing fungus so that the amount of fungus body of
the aflatoxin-producing fungus is suppressed to be less than 50% relative to that of an untreated
group (i.e., control group) when it is applied at a high concentration, but can suppress aflatoxin
production itself even when it is applied at a low concentration. A degree of the amount of
fungus body is not particularly limited as long as an amount of fungus body of a treated group is
50% or more than relative to that of an untreated group. However, the amount of fungus body
of the treated group is preferably 70% or more, further preferably 95% or more. The
"respiratory inhibitor by the action of which an amount of fungus body of an aflatoxin-producing
fungus is 50% or more relative to that of an untreated group" means that an amount of fungus
body of an aflatoxin-producing fungus is always 50% or more relative to that of an untreated
group independent of concentration to be applied. A degree of the amount of fungus body is
not particularly limited as long as an amount of fungus body of a treated group is 50% or more
than relative to that of an untreated group. However, the amount of fungus body of the treated
group is preferably 70% or more, further preferably 95% or more. Additionally, a respiratory
inhibitor which normally has no activity against fungi is preferably used. In W020091068195,
azoxystrobin, fluoxastrobin, kresoxim-methyl, picoxystrobin, pyraclostrobin, trifloxystrobin, and
boscalid are described as respiratory inhibitors. However, as mentioned above, the invention of
W020091068 195 intends to suppress aflatoxin production by suppressing survival or
proliferation of an aflatoxin-producing fungus. Meanwhile, the present invention intends to
suppress aflatoxin production without suppressing survival or proliferation of an
aflatoxin-producing fungus. Accordingly, the present invention has obviously different
technical ideas from the invention of W020091068 195.

6
The respiratory inhibitor refers to those inhibiting respiration of a microorganism by
inhibiting the electron transport system of mitochondria or eliminating proton concentration
gradient.
The respiratory inhibitor is not particularly limited and may be appropriately selected
depending on the intended purpose.
Specific examples of the respiratory inhibitors on respective target regions will be
described.
(1) Complex I NADH oxidoreductase inhibitor:
diflumetorim, tolfenpyrad, fenazaquin, fenpyroximate, pyrimidifen, pyridaben, rotenone,
and tebufenpyrad;
(2) Complex I1 succinate dehydrogen&e inhibitor:
benodanil, benzovindiflupyr, bixafen, boscalid, carboxin, fenfuram, fluopyram,
fluxapyroxad, furametpyr, funnecyclox, flutolanil, isofetamid, isopyrazam, mepronil,
oxycarboxin, penflufen, penthiopyrad, sedaxane, thifluzamide, siccanin, atpenin A5, and
c y enop yrafen;
(3) Complex I11 ubiquinol oxidase Qo inhibitor:
methoxy acrylic acids such as azoxystrobin, coumoxystrobin, coumethoxystrobin,
enoxastrobin, flufenoxystrobin, picoxystrobin, pyraoxystrobin, and fluacrypyrim;
methoxycarbamates such as pyraclostrobin, pyrametostrobin, and triclopyricarb; oxyiminoacetic
acids such as kresoxim-methyl and trifloxystrobin; oxyiminoacetamidos such as dimoxystrobin,
fenaminstrobin, metominostrobin, and orysastrobin; oxazolidinediones such as famoxadone;
dihydrodioxazines such as fluoxastrobin; irnidazolinones such as fenamidone; benzylcarbamates
such as pyribencarb; and others such as acequinocyl;
(4) Complex I11 ubiquinol reductase Qi inhibitor:
amisulbrom, antimycin A, and cyazofamid;
(5) Oxidative phosphorylation uncoupling agent:
binapacryl, meptyldinocap, dinocap, chlorfenapyr, ferimzone, fluazinam, sulfluramid,
and DNOC;
0
(6) Oxidative phosphorylation inhibitor (ATP synthase inhibitor):
fentin acetate, fentin chloride, fentin hydroxide, azocyclotin, cyhexatin, diafenthiuron,
fenbutatin oxide, propargite, and tetradifon;
(7) ATP production inhibitor:
silthiofam;
(8) Complex I11 cytochrome bcl (ubiquinone reductase) Qx (unknown) inhibitor:
ametoctradin, and hydramethylnon;
(9) Other agents:
bifenazate.
These may be used alone or in combination.
Among the above respiratory inhibitors, rotenone, siccanin, atpenin A5, antimycin A,
pyridaben, tolfenpyrad, fluacrypyrim, acequinocyl, bifenazate, mepronil, metominostrobin,
pyribencarb, and cyazofamid are preferable because they do not suppress survival or
proliferation of the aflatoxin-producing fungus, and can suppress aflatoxin production.
Azoxystrobin, fluoxastrobin, picoxystrobin, pyraclostrobin, trifloxystrobin, and boscalid
described in W020091068 195 also can suppress aflatoxin production itself compared to that of
an untreated group, even when they are used at a concentration at which the number of
aflatoxin-producing fungi is only 50% or more compared to that of the untreated group. Thus,
the above-described compounds are also preferable when they are used under such conditions.
Among the above-described preferable respiratory inhibitors, more preferable are
fluacrypyrim and metominostrobin; and particularly preferable is fluacrypyrim from the
viewpoint of its excellent aflatoxin production inhibitory activity.
Also, rotenone, siccanin, atpenin A5, and antimycin A are derived from natural products,
and preferable from the viewpoint of safety. Among them, siccanin, atpenin A5, and antimycin
A are more preferable from the viewpoint of its excellent aflatoxin production inhibitory activity.
The respiratory inhibitor may be commercially available products, or those produced by
biosynthesis or chemosynthesis.
A method of the biosynthesis or chemosynthesis is not particularly limited and may be
8
appropriately selected from known methods.
An amount of the respiratory inhibitor contained in the aflatoxin production inhibitor is
not particularly limited and may be appropriately selected depending on the intended purpose.
The aflatoxin production inhibitor may be the respiratory inhibitor itself.

The other ingredients contained in the aflatoxin production inhibitor are not particularly
limited and may be appropriately selected depending on the intended purpose, unless they impair
effects of the present invention. Examples thereof include any agrochemical or medical
ingredients and agrochemical or medical adjuvants.
An amount of the other ingredients contained in the aflatoxin production inhibitor is not
particularly limited and may be appropriately selected depending on the intended purpose, unless
they impair effects of the present invention.
<>
The agrochemical ingredient is not particularly limited and may be appropriately
selected depending on the intended purpose. For example, active ingredients of the following
agrochemicals may be used.
Examples of the agrochemicals include a fungicide, a bactericide, an antivirus agent, a
plant resistance inducer, an insecticide, a miticide, a nematocide, an insect growth regulator, an
insect attractant, a herbicide, a plant growth regulator, a synergist, a safener, a bird repellent, a
fertilizer, and a soil conditioner. These may be used alone or in combination.
<>
The agrochemical adjuvant contains a carrier, a surfactant, and other adjuvants; and, if
necessary, further contains other ingredients.
-Carrier-
The carrier is not particularly limited and may be appropriately selected depending on
the intended purpose, as long as it can be used for agriculture or horticulture. Examples thereof
include a liquid carrier and a solid carrier. These may be used alone or in combination.
Examples of the liquid carrier include water; alcohols such as isopropyl alcohol, and
9
ethylene glycol; cyclohexanone; ketones such as methylethylketone; ethers such as propylene
glycol monomethylether, and diethylene glycol mono-n-butylether; aliphatic hydrocarbons such
as kerosene, and diesel oil; aromatic hydrocarbons such as xylene, trimethyl benzene,
tetramethyl benzene, methyl naphthalene, and solvent naphtha; amides such as
N-methyl-2-pyrrolidone; esters such as fatty acid glycerin ester; vegetable oils such as soybean
oil and canola oil.
Examples of the solid carrier include animal or plant powder such as starch, activated
charcoal, soybean flour, wheat flour, wood flour, fish flour, and powdered milk; mineral powder
such as talc, kaolin, bentonite, zeolite, diatomaceous earth, white carbon, clay, alumina, calcium
carbonate, potassium chloride, and ammonium sulfate.
-Surfactant-
The surfactant is not particularly limited and may be appropriately selected depending
on the intended purpose. Examples thereof include a nonionic surfactant, an anionic surfactant,
a cationic surfactant, and an amphoteric surfactant. These may be used alone or in
combination.
Examples of the nonionic surfactant include polyoxyethylene alkylether,
polyoxyethylene alkylarylether, polyoxyethylene styrylphenylether, polyoxyethylene alkylester,
polyoxyethylene sorbitan alkylate, polyoxyethylene phenylether polymer, polyoxyethylene
alkylenearylphenylether, polyoxyethylene alkyleneglycol, and polyoxyethylene
polyoxypropylene blockpolymer.
Examples of the anionic surfactant include lignin sulfonate, alkylaryl sulfonate, dialkyl
sulfosuccinate, polyoxyethylene alkylarylether sulfate, alkyl naphthalenesulfonate, and
polyoxyethylene styrylphenylether sulfate.
Examples of the cationic surfactant include alkylamine salts.
Examples of the amphoteric surfactant include quaternary ammonium salt alkylbetaine,
and mine oxide.
-Other adjuvants-
The other adjuvants are not particularly limited and may be appropriately selected
10
depending on the intended purpose. Examples thereof include a binder, a thickener, an
adhesive agent, an antiseptic fungicide, a solvent, a stabilizing agent of agrochemical active
ingredients, an antioxidant, an ultraviolet rays protective agent, a crystallization preventive agent,
an antifoaming agent, a physical property-improving agent, and a colorant.
The binder, the thickener, and the adhesive agent are not particularly limited and may be
appropriately selected depending on the intended purpose. Examples thereof include dextrin,
cellulose, methylcellulose, ethylcellulose, carboxymethylcellulose, hydroxyethylcellulose,
hydroxypropylcellulose, hydroxypropylmethylcellulose, carboxymethyl starch, pullulan, sodium
alginate, ammonium alginate, propylene glycol alginate, guar gum, locust bean gum, gum arabic,
xanthan gum, gelatin, casein, polyvinyl alcohol, polyethylene oxide, polyethylene glycol,
ethylenelpropylene block polymer, sodium polyacrylate, and polyvinyl pyrrolidone.

A dosage form of the aflatoxin production inhibitor is not particularly limited and may
be appropriately selected depending on the intended purpose. Examples thereof include an
emulsion, a suspension, a wettable ppwder, a water-soluble powder, a solution, a sol (a floable), a
wettable granule, a powder, a subtle granule, a granule, a tablet, an oil, a spray, a propellant, an
aerosol, and a paste. Among them, preferable is a solution.
A method for producing the above dosage form is not particularly limited and may be
produced by known methods.

The aflatoxin production inhibitor may be used in combination with or concomitant
with other microbicides (e.g., a fungicide, a bactericide, an antivirus agent, or a plant resistance
inducer), an insecticide, a miticide, a nematocide, an insect growth regulator, an insect attractant,
a herbicide, a plant growth regulator, a synergist, a safener, a bird repellent, a fertilizer, and a soil
conditioner.

The aflatoxin production inhibitor can be suitably used for a method for controlling
aflatoxin contamination because it has an excellent aflatoxin production inhibitory activity.
11
(Method for controlling aflatoxin contamination)
A method for controlling aflatoxin contamination of the present invention is a method
for controlling aflatoxin contamination, including treating a plant to which an
aflatoxin-producing fungus may be attached, a fruit which is harvested from the plant, the fruit
during storage, or any combination thereof, with any of the above described aflatoxin production
inhibitors.
The plant to which an aflatoxin-producing fungus may be attached refers to a plant
which serves as a host to the aflatoxin-producing fungus, to which the aflatoxin-producing
fungus may be attached, and which may be infected with the aflatoxin-producing fungus. The
plant may be a plant which has not been uninfected yet, or may be a plant which has been
already infected. In the case of the plant which has been already infected, the plant may be in a
state in which the plant has been already infected but not developed a disease yet, or may be in a
state in which the plant has already developed a disease. In addition, the plant may be in a state
I before or afler producing fruits. The aflatoxin production inhibitor may treat a fruit which has
I
I not been harvested yet, or a fruit which has already been harvested. In the case where the fiuit
is stored for a short or long period of time after harvesting, the aflatoxin production inhibitor
i may treat the fruit at beginning of storage, during storage, or before shipment. A treatment
1 method is not particularly limited and may be appropriately selected depending on the intended
purpose, as long as the production inhibitor can be brought into contact with a target to be
controlled. Specific examples thereof include a method in which the target is treated with the
aflatoxin production inhibitor in a neat form or in a diluted form (e.g., a water-diluted form) by
distribution (e.g., spraying, misting, atomizing, dusting, granule application, submerged
application or box application), a soil application (e.g., mixing or irrigation), a surface
application (e.g., coating, dressing or covering), or an immersion.
A target to be treated with the aflatoxin production inhibitor is not particularly limited
and may be appropriately selected depending on the intended purpose. Examples thereof
include a plant body and a crop.
Examples of the crop include cereals such as maize, rice, buckwheat, or adlay; nuts such
as peanut, pistachio nut, or Brazil nut; spices such as nutmeg, hot pepper, or paprika; beans such
as coffee bean; sesames; and cottonseed.
An application amount of the aflatoxin production inhibitor is not particularly limited
and may be appropriately selected depending on various conditions such as a formulation, a type
of disease or crop of interest, a degree of damage caused by a disease, an application place, an
application method, an application timing, a type or amount of an agent or fertilizer to be used in
combination or concomitantly, or weather.
An application concentration of the aflatoxin production inhibitor is not particularly
limited and may be appropriately selected depending on the intended purpose. However, the
aflatoxin production inhibitor is applied such that a concentration of the respiratory inhibitor
therein is preferably 0.001 mM or more, more preferably 0.01 mM or more. When the
application concentration is less than 0.01 mM, aflatoxin production may be inhibited
insufficiently. In particular, the aflatoxin production inhibitor is preferably applied at a
concentration at which an amount of fungus body of an aflatoxin-producing h g u s is 50% or
more, preferably 70% or more, further preferably 95% or more relative to that of an untreated
Examples
The present invention now will be specifically described with reference to Test Example,
but is note limited thereto.
(Test Example 1 : Evaluation of aflatoxin production inhibitory activity)
Aflatoxin production inhibitory activities of the following respiratory inhibitors were
evaluated as follows.

As~ernillusp arasiticus NRRL 2999 (USDA, obtained fiom Agricultural Research
Service, USA), as an aflatoxin B1- and GI-producing fungus, was incubated on a slant medium
of a potato dextrose agar medium (PDA medium, product of Difco Laboratories, Inc.) at 27S0C
for 7 days. After the incubation, spores were scrapped off fiom the flora thereof with a
13
platinum loop and suspended in a 0.1% by mass solution of Tween 20 in distillated water so as to
give a concentration of 6.5 x lo4 cells/p~t,o thereby prepare a spore suspension.

Each of respiratory inhibitors shown in Tables 1 and 5 was pipetted into wells of a
24-well microplate so as to give a final concentration of 0 mM (control) and 0.001 mM,
respectively. Each of respiratory inhibitors shown in Table 2 was pipetted into wells of a
24-well microplate so as to give a final concentration of 0 mM (control) and 0.01 mM,
respectively. Each of respiratory inhibitors shown in Table 3 was pipetted into wells of a
24-well microplate so as to give a final concentration of 0 mM (control) and 0.02 rnM,
respectively. Each of respiratory inhibitors shown in Table 4 was pipetted into wells of a
24-well microplate so as to give a final concentration of 0 mM (control) and 0.2 mM,
respectively. A potato dextrose liquid mediurn (product of Difco Laboratories, Inc.) was added
to each of the wells so as to give a total liquid volume of 1 mL.
To each of the wells, 10 pL of the spore suspension (A. parasiticus NRRL 2999) was
inoculated, followed by static incubation at 27S°C for 3 days.
After incubating for 3 days, a culture medium in each of the wells was subjected to
centrifugation to thereby be separated into fungus body and culture supernatant.
The fungus body was washed with 5 mL of distilled water, transferred to a 1 .S mL
microtube, and freeze-dried. After freeze-drying, a mass of the microtube containing the
fungus body was measured, from which a mass of an empty microtube was subtracted to thereby
determine a mass of the fungus body.
The culture supernatant (0.7 mL) was extracted with chloroform (0.2 mL), and the
chloroform was distilled off by air drying. Thereafter, the resultant was dissolved into a 90%
by volume solution of acetonitrile in water (0.25 mL). The resultant solution was subjected to
high performance liquid chromatography (HPLC) according to the following conditions, to
thereby quantifi a mass of aflatoxin.
[Measurement conditions of HPLC]
Apparatus: LC-2000 plus HPLC system (product of JASCO Corporation)
14
Column: Capcell-pak C18 column UG120 (length: 250 mm, diameter: 4.6 mm, product
of Shiseido Company, Limited)
Eluent: acetonitrile/methanol/water (1 0:30:60 (volume ratio))
Elution condition: isoractic elution for 20 min
Flow rate: 1 mL/min
Retention time: aflatoxin B1 (8.3 min)
aflatoxin G1 (1 1.1 min)
Detection: UV 365 nm
-Aflatoxin production inhibition rate (%)-
Aflatoxin production inhibition rate at each of the concentrations of the respiratory
inhibitors was calculated according to the following formula. Results are shown in Tables 1 to
5.
Aflatoxin production inhibition rate (%) = {(X - Y)/X) x 100
In the above formula, X denotes an aflatoxin production amount (pg/mL) in the case of
control (no respiratory inhibitor was added), and Y denotes an aflatoxin production amount
(pg/mL) in the case where a respiratory inhibitor was added.
The aflatoxin production amount was calculated using a total amount of aflatoxin B1
and aflatoxin GI, which were aflatoxins confirmed to be produced in a culture medium of A.
parasiticus NRRL 2999.
Influence of the respiratory inhibitors on a mass of fungus body was evaluated
according to the following criteria. Results are shown in Tables 1 to 5.
[Evaluation criteria]
- (no influence): a mass of fungus body is 95% or more relative to that of a control (no
respiratory inhibitor was added).
+ (almost no influence): a mass of fungus body is 70% or more but less than 95%
relative to that of a control (no respiratory inhibitor was added).
+ (some influence): a mass of fungus body is 50% or more but less than 70% relative to
that of a control (no respiratory inhibitor was added).
15
++ (strong influence): a mass of b g u s body is less than 50% relative to that of a
control (no respiratory inhibitor was added).
Table 1
Table 2
Influence on mass
offungus body
f
+
+
+
+
Respiratory
inhibitor
fluacrypyrim
boscalid
azoxystrobin
pyraclostrobin
pyribencarb
Aflatoxin production inhibition
rate (%)
-
0.001 mM
76
99
75
92
78
Respiratory
inhibitor
rotenone
siccanin
atpenin A5
antimycin A
pyridaben
tolfenpyrad
acequinocyl
Aflatoxin production inhibition
rate (%)
-
0.01 mM
25
54
75
57
90
82
46
Influence on mass
offungus body
-
f
-
-
+
+
-
Table 3
t
I I I
I metominostrobin I 93 I - I
Respiratory
inhibitor
I trifloxystrobin I 89 I + I
Aflatoxin production inhibition
Influence on mass
rate (%)
of fungus body
0.02 mM
Table 4
cyazofamid 74 -
Table 5
Respiratory
inhibitor
bifenazate
mepronil
picoxystrobin
Aflatoxin production inhibition
Influence on mass
rate (%)
of fungus body
0.2 mM
5 7 -
66 -
93 -
As shown in Tables 1 to 5, it was confirmed that each of the respiratory inhibitors had
Respiratory
inhibitor
kresoxim-methyl
an aflatoxin production inhibitory activity. It was also confirmed that respiratory inhibitors
Aflatoxin production inhibition
Influence on mass
rate (%)
of fungus body
0.001 mM
99 ++
other than kresoxim-methyl had almost no influence on growth of fungi, and selectively
inhibited aflatoxin production. In particular, fluacrypyrim had higher selectivity and stronger
aflatoxin production inhibitory activity than other inhibitors exhibiting the inhibitory activity at
0.001 rnM.
Dated: 6th Jan, 2014

CLAIMS
We Claim:
\ d
1. An aflatoxin production inhibitor, comprising:
a respiratory inhibitor.
' aRl~~~pL
2. The aflatoxin production inhibitor according to claim 1, wherein the respiratory
inhibitor is a respiratory inhibitor applied at a concentration at which an amount of fungus body
of an aflatoxin-producing fungus is 50% or more relative to that of an untreated group, or a
respiratory inhibitor by the action of which an amount of fungus body of an aflatoxin-producing
fungus is 50% or more relative to that of an untreated group.
3. The aflatoxin production inhibitor according to claim 1 or 2, wherein the respiratory
inhibitor is at least one selected from the group consisting of rotenone, siccanin, atpenin A5,
antimycin A, pyridaben, tolfenpyrad, fluacrypyrim, acequinocyl, bifenazate, mepronil,
metominostrobin, pyribencarb, and cyazofamid.
4. The aflatoxin production inhibitor according to claim 1 or 2, wherein the respiratory
inhibitor is fluacrypyrim.
5. The aflatoxin production inhibitor according to claim 1 or 2, wherein the respiratory
inhibitor is at least one selected from the group consisting of rotenone, siccanin, atpenin A5, and
antimycin A.
6. A method for controlling aflatoxin contamination, comprising:
treating a plant to which an aflatoxin-producing fungus may be attached, a fruit which is
harvested from the plant, the fruit during storage, or any combination thereof, with the aflatoxin
production inhibitor according to any one of claims 1 to 5.
Regd. Patent Agent [INPA-16091
Dated: 6" Jan, 2014