Page
1
of
87
UNITED
STATES
ENVIRONMENTAL
PROTECTION
AGENCY
WASHINGTON,
D.
C.
20460
OFFICE
OF
PREVENTION,
PESTICIDES
AND
TOXIC
SUBSTANCES
Date:
6/
29/
2006
MEMORANDUM:

SUBJECT:
Benthiavalicarb­
Isopropyl:
Human
Health
Risk
Assessment
for
Proposed
Uses
on
Imported
Grapes
and
Tomatoes.
PC
Code:
098379,
Petition
No:
3E6545,
DP
Barcode:
D330555.

Regulatory
Action:
Section
3
Registration
Action
Risk
Assessment
Type:
Single
Chemical
Food
Only
(
Import
Tolerances)

FROM:
Douglas
Dotson,
Ph.
D.,
Chemist
Kim
Harper,
Toxicologist
Registration
Action
Branch
2
Health
Effects
Division
(
7509C)

AND
Dennis
McNeilly,
Chemist
Karlyn
Bailey,
Toxicologist
Michael
Doherty,
Ph.
D.,
Chemist
Registration
Action
Branch
2
Health
Effects
Division
(
7509C)

THROUGH:
Richard
Loranger,
Ph.
D.,
Branch
Senior
Scientist
Christina
Swartz,
Branch
Chief
Registration
Action
Branch
2
Health
Effects
Division
(
7509C)

TO:
Tamue
Gibson/
Mary
Waller,
RM
21
Fungicide
Branch
Registration
Division
(
7505C)

This
document
is
a
revised
version
of
the
3/
2/
2006
risk
assessment
(
D322937).
It
has
been
prepared
in
response
to
comments
from
the
Office
of
General
Council
regarding
cumulative
risk
as
well
as
the
use
of
average
field
trial
residues
and
percent
crop
imported
data.
There
have
been
no
changes
in
recommended
tolerances
or
estimates
of
dietary
risk.
Page
2
of
87
Table
of
Contents
1.0
Executive
Summary.......................................................................................................
4
2.0
Ingredient
Profile...........................................................................................................
9
2.1
Summary
of
Registered/
Proposed
Uses
................................................................
9
2.2
Structure
and
Nomenclature.................................................................................
9
2.3
Physical
and
Chemical
Properties
.......................................................................
10
3.0
Metabolism
Assessment...............................................................................................
11
3.1
Comparative
Metabolic
Profile
...........................................................................
11
3.2
Nature
of
the
Residue
in
Foods
..........................................................................
11
3.2.1.
Description
of
Primary
Crop
Metabolism................................................
11
3.2.2
Description
of
Livestock
Metabolism......................................................
12
3.2.3
Description
of
Rotational
Crop
Metabolism,
including
identification
of
major
metabolites
and
specific
routes
of
biotransformation......................
12
3.3
Environmental
Degradation................................................................................
12
3.4
Toxicity
Profile
of
Major
Metabolites
and
Degradates
........................................
12
3.5
Summary
of
Residues
for
Tolerance
Expression
and
Risk
Assessment
...............
13
3.5.1
Tabular
Summary
...................................................................................
13
3.5.2
Rationale
for
Inclusion
of
Metabolites
and
Degradates............................
13
4.0
Hazard
Characterization/
Assessment..............................................................................
14
4.1
Hazard
Characterization.....................................................................................
14
4.2
FQPA
Hazard
Considerations.............................................................................
25
4.2.1
Adequacy
of
the
Toxicity
Data
Base
.......................................................
25
4.2.2
Evidence
of
Neurotoxicity
......................................................................
25
4.2.3
Developmental
Toxicity
Studies..............................................................
25
4.2.4
Reproductive
Toxicity
Study
..................................................................
26
4.2.5
Additional
Information
from
Literature
Sources......................................
26
4.2.6
Pre­
and/
or
Postnatal
Toxicity..................................................................
26
4.2.6.1
Determination
of
Susceptibility....................................................
26
4.2.6.2
Degree
of
Concern
Analysis
and
Residual
Uncertainties
for
Pre
and/
or
Post­
natal
Susceptibility
...................................................
26
4.3
Recommendation
for
a
Developmental
Neurotoxicity
Study
...............................
26
4.4
Hazard
Identification
and
Toxicity
Endpoint
Selection........................................
27
4.4.1
Acute
Reference
Dose
(
aRfD)
­
All
Populations......................................
27
4.4.2
Chronic
Reference
Dose
(
cRfD)
.............................................................
27
4.4.3
Non­
Dietary
Exposure
for
Handlers........................................................
27
4.4.4
Margins
of
Exposure
..............................................................................
27
4.4.5
Recommendation
for
Aggregate
Exposure
Risk
Assessments..................
27
4.4.6
Classification
of
Carcinogenic
Potential
..................................................
27
4.5
Special
FQPA
Safety
Factor
...............................................................................
27
4.6
Endocrine
disruption
..........................................................................................
28
Page
3
of
87
5.0
Public
Health
Data
......................................................................................................
29
6.0
Exposure
Characterization/
Assessment......................................................................
29
6.1
Dietary
Exposure/
Risk
Pathway
.........................................................................
29
6.1.1
Residue
Profile
.......................................................................................
29
6.1.2
Acute
and
Chronic
Dietary
Exposure
and
Risk........................................
31
6.2
Water
Exposure/
Risk
Pathway
...........................................................................
34
6.3
Residential
(
Non­
Occupational)
Exposure/
Risk
Pathway....................................
34
7.0
Aggregate
Risk
Assessments
and
Risk
Characterization
...........................................
34
7.1
Acute
Aggregate
Risk
........................................................................................
34
7.2
Short­,
Intermediate­,
and
Long­
Term
Aggregate
Risk.......................................
34
7.3
Cancer
Risk........................................................................................................
34
8.0
Cumulative
Risk
Characterization/
Assessment..........................................................
34
9.0
Occupational
Exposure/
Risk
Pathway........................................................................
35
10.0
Data
Needs
and
Label
Requirements..........................................................................
35
10.1
Toxicology.........................................................................................................
35
10.2
Residue
Chemistry
.............................................................................................
35
10.3
Occupational
and
Residential
Exposure
..............................................................
36
11.0
Summary
of
Proposed
and
Recommended
Tolerances
..............................................
36
References:
..............................................................................................................................
37
Appendices
..............................................................................................................................
38
Page
4
of
87
1.0
Executive
Summary
Benthiavalicarb­
isopropyl
(
hereafter
in
this
document
referred
to
solely
as
benthiavalicarb)
is
a
new
valinamide
carbamate
type
of
fungicide
being
proposed
for
use
on
grapes
and
tomatoes
(
both
field
and
greenhouse)
in
the
EU
for
the
control
of
downy
mildew.
Its
mode
of
action
is
through
the
inhibition
of
phospholipid
biosynthesis.
The
R­
L
stereoisomer
of
benthiavalicarb
(
KIF­
230RL
is
the
main
and
pesticidally
active
component;
however,
the
S­
L
stereoisomer
(
KIF­
230S­
L),
which
is
not
pesticidally
active,
is
also
present
as
a
minor
impurity.
The
benthiavalicarb
products
being
proposed
for
use
in
the
EU
on
grapes
and
tomatoes
include
a
15%
water­
dispersible
granule
(
WDG),
for
use
only
on
tomatoes,
and
a
multiple
active
ingredient
WDG
formulation
containing
both
benthiavalicarb
(
1.75%)
and
mancozeb
(
70%)
for
use
on
both
grapes
and
tomatoes.
These
formulations
are
labeled
for
multiple
foliar
applications.
There
are
currently
no
food/
feed
uses
or
tolerances
for
benthiavalicarb
in
the
U.
S.

The
toxicology
database
for
benthiavalicarb
is
complete
and
adequate
for
the
purposes
of
risk
assessment.
None
of
the
toxicity
study
endpoints
were
the
result
of
a
single
oral
exposure
to
benthiavalicarb.
As
a
result,
no
acute
dietary
endpoint
was
identified
and
an
acute
dietary
exposure
assessment
is
not
needed.
The
main
target
organ
of
toxicity
is
the
liver
in
all
four
species
tested
(
rat,
mouse,
rabbit,
and
dog).
Chronic
exposure
to
benthiavalicarb
also
resulted
in
kidney
toxicity
in
rats
and
mice.
Prolonged
exposure
to
benthiavalicarb
resulted
in
the
formation
of
several
tumor
types:
liver
tumors
in
male
and
female
mice,
thyroid
follicular
cell
adenomas
in
male
mice,
and
uterine
tumors
in
rats.
This
compound
is
classified
as
"
likely
to
be
carcinogenic
to
humans."
HED's
Cancer
Assessment
Review
Committee
(
CARC)
concluded
that
the
mechanistic
studies
submitted
for
this
chemical
were
insufficient
to
support
definitive
modes
of
action
for
the
individual
tumor
types.
Therefore,
the
Committee
recommended
using
a
low­
dose
extrapolation
approach
(
Q1*)
for
estimating
the
human
cancer
risk
based
on
the
most
potent
tumor
type
(
liver
tumors
in
mice).

Benthiavalicarb
is
not
mutagenic,
nor
does
this
chemical
produce
developmental,
reproductive,
or
neurotoxic
effects.

Benthiavalicarb
was
absorbed
and
excreted
rapidly
in
rats,
with
nearly
full
recovery
of
radioactive
label
within
168
hours.
Analysis
of
the
metabolites
in
bile,
urine,
and
feces
of
cannulated
rats
identified
a
total
of
13
compounds,
including
the
parent,
which
accounted
for
43­
60%
of
the
radiolabeled
dose.
Based
on
the
components
identified
in
excreta,
the
metabolism
of
benthiavalicarb
in
rats
primarily
involves
either
(
1)
hydroxylation
of
the
phenyl
ring
followed
by
glucuronic
acid
conjugation
or
glutathione
conjugation,
or
(
2)
hydroxylation
of
the
phenyl
ring
followed
by
glutathione
conjugation
with
subsequent
degradation
of
the
glutathione
moiety,
resulting
in
a
variety
of
metabolites
present
in
small
quantities.
Cleavage
and
hydroxylation
of
the
valyl
side
chain
also
occurs
to
a
limited
extent.
Parent
(
excreted
in
feces
only)
and
metabolites
M15
(
urine
and
feces)
and
M18
(
urine
only)
were
also
the
principal
components
detected
in
plasma,
liver,
and
kidneys.

The
risk
assessment
team
concluded
that
none
of
the
observed
effects
were
the
result
of
a
single
exposure
to
benthiavalicarb.
As
a
result,
an
appropriate
endpoint
for
acute
dietary
risk
assessment
is
not
available
and
an
acute
dietary
exposure
assessment
is
not
needed.
For
chronic
Page
5
of
87
dietary
risk
assessment,
the
endpoint
selected
is
based
on
liver
effects
seen
in
the
chronic
rat
study.
The
liver
effects
are
the
primary
effects
of
concern.
The
NOAEL
of
9.9
mg/
kg/
day
is
protective
of
tumorigenic
effects
observed
at
doses
at,
or
above,
300
mg/
kg/
day
(
uterine
tumors
in
rats
at,
or
above,
318
mg/
kg/
day
and
liver
and
thyroid
tumors
in
mice
at,
or
above,
358
mg/
kg/
day).
The
standard
100x
uncertainty
factor
for
inter­
and
intra­
species
variabilities
is
being
used,
and
the
special
FQPA
safety
factor
was
reduced
to
1x
because
of
the
absence
of
neurotoxic
and
developmental
effects.
Therefore,
the
chronic
RfD
and
cPAD
are
0.099
mg/
kg/
day.
The
target
MOE
is
100.
For
the
purpose
of
cancer
risk
assessment
the
Q1*
is
0.063
(
mg/
kg/
day)­
1.
This
value
was
based
on
increases
in
male
mouse
liver
combined
adenomas,
carcinomas,
and/
or
blastomas.

For
purposes
of
this
risk
assessment,
the
nature
of
the
residue
in
plants
has
been
adequately
delineated
based
on
grape,
tomato,
and
potato
metabolism
studies.
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
residue
in
fruit
and
foliage.
Parent
was
also
the
principal
14C­
residue
in
potato
foliage,
but
other
minor
metabolites
were
identified.
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
Secondary
metabolism
occurs
via
conjugation
of
sugars
with
hydroxy
groups.
Although
two
major
(>
10%)
residues
were
observed
in
potato
tubers,
the
absolute
levels
of
these
metabolites
(
suggested
to
be
sugar
conjugates)
were
<
0.006
ppm.
Chiral
analysis
of
potato
foliage
extracts
also
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).
Based
on
the
results
of
the
metabolism
studies,
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.
The
nature
of
the
residue
in
livestock
was
not
determined,
and
is
not
required
at
this
time,
as
there
are
no
significant
livestock
feed
items
associated
with
grape
and
tomato
commodities.

An
adequate
GC/
NPD
Method
(
RCC
No.
665943)
is
available
for
collecting
data
on
residues
of
both
the
R­
L
and
S­
L
isomers
of
benthiavalicarb
in/
on
grape
and
tomato
commodities.
For
this
method,
residues
are
extracted
with
acetone,
partitioned
into
hexane,
purified
using
an
amino
SPE
cartridge,
and
determined
by
GC/
NPD.
The
lower
limit
of
method
validation
(
LLMV)
is
0.01
ppm
for
each
analyte
in
grape
and
tomato
matrices.
The
limit
of
detection
(
LOD)
in
grapes
is
0.001
ppm.
The
LOD
in
grape
processed
commodities,
tomatoes,
and
tomato
processed
commodities
is
0.002
ppm.
The
method
was
adequately
validated
by
the
petitioner,
and
is
being
proposed
as
the
tolerance
enforcement
method.
The
method
was
forwarded
to
the
Analytical
Chemistry
Branch
(
ACB)
of
the
Biological
and
Economic
Assessment
Division
(
BEAD)
for
a
petition
method
validation
(
PMV)
trial.
ACB
concluded
that
the
method
does
not
need
to
be
laboratory­
validated
by
EPA,
and
appears
to
be
suitable
for
tolerance
enforcement.
As
the
method
is
not
highly
specific,
the
petitioner
should
either
develop
a
confirmatory
method
for
the
analytes
of
interest
or
conduct
an
interference
study
for
any
future
tolerances.

Adequate
storage
stability
data
are
available
indicating
that
benthiavalicarb
is
stable
under
frozen
( ­
18
°
C)
conditions
for
at
least
13
months
in
grapes
and
8
months
in
tomatoes.
These
data
support
the
maximum
storage
intervals
(
7­
8
months)
from
the
grape
and
tomato
field
trials.

As
there
are
no
significant
animal
feed
items
associated
with
the
proposed
use
on
grapes
and
tomatoes,
livestock
feeding
studies
are
not
required
for
this
petition.
Tolerances
do
not
need
to
Page
6
of
87
be
established
on
animal
commodities.

The
available
field
trial
data
are
adequate
and
will
support
the
proposed
use
patterns
on
grapes
and
tomatoes.
For
grapes,
34
field
tests
were
conducted
throughout
the
EU
in
which
grapes
were
treated
at
1x
the
maximum
proposed
rate
and
harvested
at
the
minimum
PHI
of
28
days.
For
tomatoes,
12
tests
were
conducted
at
the
1x
application
rate
throughout
the
EU
on
both
greenhouse­
and
outdoor­
field­
grown
tomatoes.
The
recommended
tolerances
for
grapes
and
tomatoes
were
generated
using
HED's
statistical
tolerance
generator
for
NAFTA­
harmonized
tolerances.
The
recommended
tolerance
for
grapes
is
0.25
ppm,
even
though
the
highest
field
trial
value
is
0.27
ppm.
The
recommended
tolerance
for
tomatoes
is
0.45
ppm.

The
available
processing
data
are
adequate.
A
separate
tolerance
is
required
for
raisins.
HED
recommends
in
favor
of
the
proposed
tolerance
of
1.0
ppm
for
grape,
raisin.
As
residues
do
not
concentrate
in
wine,
the
0.25
ppm
grape
tolerance
is
adequate
to
cover
residues
in
wine.
A
separate
tolerance
for
wine
is
not
needed.
The
tomato
processing
studies
demonstrate
that
separate
tolerances
are
not
required
for
processed
tomato
fractions.
Storage
stability
data
were
not
submitted
for
processed
grape
and
tomato
commodities.
These
data
are
not
required
for
the
current
tolerance
requests
because
acceptable
storage
stability
data
were
submitted
for
grapes
and
tomatoes.
Residues
were
stable
in
grapes
for
13
months
and
in
tomatoes
for
8
months.
In
the
tomato
storage
stability
studies,
tomatoes
were
homogenized
prior
to
storage.
Homogenized
tomatoes
are
comparable
in
both
composition
and
texture
to
processed
tomato
commodities.
The
matrix
is
an
acidic
one
which
would
very
likely
maximize
residue
decay.
As
a
result,
the
tomato
data
are
considered
adequate
to
demonstrate
storage
stability
in
processed
grape
commodities
as
well.

As
the
proposed
uses
are
for
grapes
and
tomatoes
grown
in
the
EU,
confined
and
field
rotational
crop
studies
are
not
required
for
the
current
petition.

A
chronic
dietary
risk
assessment
was
conducted
using
the
Dietary
Exposure
Evaluation
Model
(
DEEM­
FCID,
Version
2.03),
which
uses
food
consumption
data
from
the
USDA's
Continuing
Surveys
of
Food
Intakes
by
Individuals
(
CSFII)
from
1994­
1996
and
1998.
The
result
of
the
chronic
dietary
exposure
assessment
for
the
general
U.
S.
population
is
being
used
along
with
the
cancer
Q1*
value
to
estimate
the
cancer
risk
to
the
U.
S.
population.
The
chronic
dietary
exposure
analysis
is
based
on
average
field
trial
values
and
estimates
of
percent
crop
imported.
Empirical
processing
factors
were
used
for
most
processed
commodities.
When
empirical
factors
were
not
available,
DEEM
(
Version
7.81)
default
processing
factors
were
used.
The
general
U.
S.
population
and
all
population
subgroups
have
risk
estimates
that
are
below
HED's
level
of
concern
(
i.
e.,
100%
of
the
chronic
population
adjusted
dose
(
cPAD)).
The
risk
estimates
for
all
population
subgroups
are
<
1%
cPAD.

In
an
unrefined
cancer
dietary
exposure
analysis,
the
cancer
risk
to
the
general
U.
S.
population
exceeds
HED's
level
of
concern
(
i.
e.,
1.0
x
10­
6
(
or
one
in
one
million)).
As
a
result,
refinements
were
made
to
the
analysis.
The
refinements
include
the
use
of
average
field
trial
values
and
estimates
of
percent
crop
imported.
With
the
use
of
these
refinements,
the
lifetime
cancer
risk
to
the
U.
S.
population
is
1.6
x
10­
6.
Although
refinements
were
made
to
the
analysis,
it
is
considered
to
be
conservative.
First,
average
field
trial
values
from
field
trials
almost
always
exceed
the
expected
residue
levels
found
on
crops
at
the
time
of
consumption.
When
field
trials
are
Page
7
of
87
performed,
the
maximum
allowable
application
rate
is
used
and
crops
are
harvested
at
the
minimum
PHI.
Samples
are
stored
frozen
until
analysis
to
ensure
minimal
degradation
of
residues.
In
actual
practice,
however,
growers
will
not
usually
use
the
maximum
application
rates
for
economic
reasons.
In
addition,
most
crops
are
not
harvested
and
immediately
stored
frozen.
From
previous
dietary
exposure
analyses,
HED
has
noted
that
monitoring
data
are
often
one
to
two
orders
of
magnitude
lower
than
field
trial
data.
The
second
reason
this
analysis
is
considered
to
be
conservative
is
that
the
registrant
is
planning
on
marketing
benthiavalicarb
in
Europe
only.
In
the
dietary
analysis,
percent
crop
imported
estimates
were
used
for
all
imports,
not
just
those
from
Europe.
The
third
reason
this
analysis
is
considered
to
be
conservative
is
that
HED
assumed
that
100%
of
the
imported
crop
will
be
treated.
The
actual
percent
crop
treated
will
very
likely
be
considerably
lower
than
that.
For
these
reasons,
HED
is
confident
that
this
analysis
does
not
underestimate
risk
to
the
general
U.
S.
population.
HED
is
also
confident
that
average
field
trial
values
and
percent
crop
imported
values
used
in
the
assessment
will
not
be
exceeded
in
the
future.
For
the
reasons
stated
above,
the
field
trial
values
considerably
overestimate
residues
found
on
food
at
the
time
of
consumption.
It
is
unlikely
that
average
residue
levels
in
imported
grape
and
tomato
commodities
at
the
time
of
consumption
would
ever
exceed
the
average
field
trial
values.
As
far
as
percent
crop
imported
values
being
exceeded
in
the
future,
this
situation
is
unlikely
as
well.
As
stated
above,
the
conservative
assumptions
were
made
that
100%
of
the
grape
and
tomato
commodities
imported
from
all
over
the
world
would
be
treated.
It's
very
unlikely
that
the
quantities
of
imports
actually
treated
will
ever
increase
to
those
levels.
Data
for
grape
commodities
were
provided
by
BEAD.
The
economist
who
provided
the
data
stated
that
it
was
unlikely
the
estimates
of
import
shares
would
be
exceeded
in
the
near
future.
The
data
for
grape
commodities
provided
to
HED
were
five
year
averages,
and
were
compared
to
the
annual
data
points
of
each
data
source,
when
available.
Import
share
data
do
vary
from
year
to
year,
but
crop
production
patterns
and
consumption
and
preferences
are
more
stable
and
change
more
slowly.
For
tomatoes,
the
worldwide
imports
changed
as
follows
from
1998
to
2004:
37%,
31%,
30%,
33%,
32%,
37%,
and
36%.
Clearly,
no
increase
occurred
over
that
time
period.
For
processed
tomato
commodities,
the
worldwide
imports
changed
as
follows
from
2000
to
2004:
2.9%,
5.6%,
7.1%,
5.4%,
and
5.8%.
Between
2001
and
2004
there
was
no
significant
increase
in
processed
tomato
commodity
imports.

The
tolerances
being
granted
in
conjunction
with
these
requests
are
for
imported
commodities
only.
As
a
result,
residues
in
water
are
not
being
considered
in
this
assessment.
In
addition,
there
are
no
residential
uses
for
benthiavalicarb
(
including
home
uses,
recreational
uses,
spray
drift,
etc.);
therefore,
short­
term
and
intermediate­
term
aggregate
risk
assessments
are
not
needed.
Because
of
the
proposed
use
pattern,
the
chronic
aggregate
risk
assessment
is
equivalent
to
the
chronic
dietary
exposure
analysis.
The
general
U.
S.
population
and
all
population
subgroups
had
risk
estimates
that
were
well
below
HED's
level
of
concern.

The
aggregate
cancer
risk
assessment
is
equivalent
to
the
cancer
dietary
exposure
analysis.
The
lifetime
cancer
risk
to
the
U.
S.
population
is
1.6
x
10­
6.
As
stated
above,
the
analysis
is
considered
to
be
a
conservative
one.
HED
believes
that
the
cancer
dietary
exposure
analysis
almost
certainly
overestimates
exposure.
HED's
rationale
for
this
belief
is
given
in
the
discussion
of
cancer
dietary
exposure.
The
cancer
risk
to
the
general
U.
S.
population
does
not
exceed
HED's
level
of
concern.
Page
8
of
87
No
major
deficiencies
were
noted
in
the
subject
petition
that
would
preclude
establishing
tolerances
for
benthiavalicarb
on
grape
and
tomato
commodities.
However,
before
any
further
tolerances
are
granted,
the
petitioner
should
either
develop
a
confirmatory
analytical
method
for
the
analytes
of
interest
or
conduct
an
interference
study.
The
recommended
tolerances
are
as
follows:
0.25
ppm
for
grape,
0.45
ppm
for
tomato,
and
1.0
ppm
for
grape,
raisin.
The
recommended
tolerances
were
generated
using
HED's
statistical
tolerance
generator
for
NAFTAharmonized
tolerances.
The
residues
to
be
included
in
the
tolerance
expression
are
the
combined
R­
L
and
S­
L
isomers
of
benthiavalicarb.
The
petitioner
has
recommended
that
only
the
R­
L
isomer
be
included
in
the
tolerance
expression
because
the
S­
L
isomer
is
a
minor
impurity
in
the
commercial
formulations
and
the
potato
metabolism
study
indicates
that
the
active
R­
L
isomer
is
not
converted
to
the
S­
L
isomer.
In
addition,
the
residue
definition
proposed
for
the
EU
MRLs
includes
only
the
R­
L
isomer.
The
registrant
has
proposed
EU
MRLs
of
0.2
ppm
for
grapes
and
0.3
ppm
for
tomatoes.
It's
possible
that
if
EU
MRLs
are
established,
the
residue
definition
might
be
benthiavalicarb­
isopropyl
only,
and
no
distinction
will
be
made
as
to
the
isomer.
HED
concludes
that
the
S­
L
isomer
should
be
included
in
the
tolerance
expression,
however.
The
results
of
the
field
trials
indicate
that
the
residue
levels
of
the
S­
L
isomer
are
often
non­
negligible
compared
to
the
levels
of
the
R­
L
isomer.
In
the
grape
field
trials
several
samples
had
S­
L
isomer
levels
that
were
10­
20%
of
the
R­
L
isomer
levels.
A
small
number
of
samples
had
S­
L:
R­
L
ratios
that
exceeded
20%.
In
the
tomato
field
trials,
the
S­
L
residue
levels
were
generally
nondetectable
In
a
few
cases,
however,
the
S­
L
isomer
was
present
at
6­
7%
of
the
level
of
the
R­
L
isomer.
The
technical
material
contains
93%
R­
L
isomer
and
3.4%
S­
L
isomer.
The
analytical
method
determines
both
isomers
separately.

HED
concludes
that
there
are
no
residue
chemistry
or
risk
issues
that
would
preclude
establishing
tolerances
for
the
combined
residues
of
isopropyl[(
S)­
1­[[[(
1R)­
1­(
6­
fluoro­
2­
benzothiazolyl)
ethyl]
amino]
carbonyl]­
2­
methylpropyl]
carbamate
and
isopropyl[(
S)­
1­
[[[(
1S)­
1­(
6­
fluoro­
2­
benzothiazolyl)
ethyl]
amino]
carbonyl]­
2­
methylpropyl]
carbamate
at
0.25
ppm
in/
on
grape,
0.45
ppm
in/
on
tomato,
and
1.0
ppm
in/
on
grape,
raisin.
As
residues
do
not
concentrate
in
wine,
a
separate
tolerance
for
wine
is
not
needed.

EPA
has
not
made
a
common
mechanism
of
toxicity
finding
as
to
benthiavalicarb
and
any
other
substances,
and
benthiavalicarb
does
not
appear
to
produce
a
toxic
metabolite
produced
by
other
substances.
As
a
result,
for
the
purposes
of
this
tolerance
action,
EPA
has
not
assumed
that
benthiavalicarb
has
a
common
mechanism
of
toxicity
with
other
substances.
Although
benthiavalicarb
is
a
carbamate
compound,
it
is
not
a
member
of
the
class
of
insecticides
known
as
the
N­
methyl
carbamates
for
which
the
Agency
is
presently
conducting
a
cumulative
risk
assessment.
The
substituents
on
the
benthiavalicarb
nitrogen
atom
are
much
larger
than
the
methyl
group
in
the
insecticides.
While
the
N­
methyl
carbamates
are
neurotoxicants
based
on
their
ability
to
inhibit
the
enzyme
cholinesterase,
there
is
no
evidence
of
neurotoxicity
or
neuropathology
in
the
hazard
database
for
benthiavalicarb.
Benthiavalicarb
is
also
not
a
member
of
the
thiocarbamate
class
of
herbicides
or
the
dithiocarbamate
class
of
fungicides.

As
benthiavalicarb
will
be
used
outside
the
U.
S.
to
treat
grapes
and
tomatoes,
there
will
be
no
occupational
risk
to
U.
S.
workers.
As
a
result,
an
occupational
assessment
is
not
relevant
to
this
tolerance
petition.
Page
9
of
87
2.0
Ingredient
Profile
Benthiavalicarb
is
a
new
valinamide
carbamate
type
of
fungicide
being
proposed
for
use
on
grapes
and
tomatoes
(
both
field
and
greenhouse)
in
the
EU
for
the
control
of
downy
mildew.
Its
mode
of
action
is
through
the
inhibition
of
phospholipid
biosynthesis.
The
R­
L
stereoisomer
of
benthiavalicarb
(
KIF­
230R­
L)
is
the
main
and
pesticidally
active
component;
however,
the
S­
L
stereoisomer
(
KIF­
230S­
L),
which
is
not
pesticidally
active,
is
also
present
as
a
minor
impurity.

The
benthiavalicarb
products
being
proposed
for
use
in
the
EU
on
grapes
and
tomatoes
include
a
15%
water­
dispersible
granule
(
WDG),
for
use
only
on
tomatoes,
and
a
multiple
active
ingredient
WDG
formulation
containing
both
benthiavalicarb
(
1.75%)
and
mancozeb
(
70%)
for
use
on
both
grapes
and
tomatoes.
These
formulations
are
labeled
for
multiple
foliar
applications.

2.1
Summary
of
Registered/
Proposed
Uses
Table
2.1.
Summary
of
Directions
for
Use
of
Benthiavalicarb
Applic.
Timing,
Type,
and
Equip.
Formulation
Applic.
Rate
(
lb
ai/
A)
Max.
No.
Applic.
per
Season
Max.
Seasonal
Applic.
Rate
(
lb
ai/
A)
PHI
(
days)
Use
Directions
and
Limitations
Grapes
Broadcast
foliar
application
Ground
or
aerial
equipment
1.75%
WDG
1
0.031
6
0.187
28
Minimum
application
volumes
and
RTIs
were
not
specified.

Tomatoes
Broadcast
foliar
application
Ground
or
aerial
equipment
1.75%
WDG
1
15%
WDG
0.067
(
0.031)
2
6
0.40
(
0.19)
2
greenhouse
­
3
field
­
7
Minimum
application
volumes
and
RTIs
were
not
specified.

1
The
1.75%
WDG
is
a
MAI
that
also
contains
70%
mancozeb.
2
When
used
in
conjunction
with
mancozeb,
the
maximum
single
and
seasonal
application
rate
is
lower.

2.2
Structure
and
Nomenclature
Table
2.2.
Nomenclature
of
Benthiavalicarb
Compound
S
N
N
H
H
CH
3
NH
O
C
H
3
CH
3
O
H
CH
3
C
H
3
F
O
Page
10
of
87
Table
2.2.
Nomenclature
of
Benthiavalicarb
Common
name
Benthiavalicarb
(
R­
L
isomer)

Company
experimental
names
KIF­
230R­
L
IUPAC
name
Isopropyl[(
S)­
1­{[(
1R)­
1­(
6­
fluoro­
1,3­
benzothiazol­
2­
yl)
ethyl]
carbamoyl}­
2­
methylpropyl]
carbamate
CAS
name
Isopropyl[(
S)­
1­[[[(
1R)­
1­(
6­
fluoro­
2­
benzothiazolyl)
ethyl]
amino]
carbonyl]­
2­
methylpropyl]
carbamate
CAS
#
177406­
68­
7
End­
use
products/
EP
15%
WDG,
1.75
WDG
(
MAI
with
70%
Mancozeb)

Compound
S
N
N
H
CH
3
H
NH
O
C
H
3
CH
3
O
H
CH
3
C
H
3
F
O
Common
name
Benthiavalicarb
(
S­
L
isomer)

Company
experimental
names
KIF­
230S­
L
IUPAC
name
Isopropyl[(
S)­
1­{[(
1S)­
1­(
6­
fluoro­
1,3­
benzothiazol­
2­
yl)
ethyl]
carbamoyl}­
2­
methylpropyl]
carbamate
CAS
name
Isopropyl[(
S)­
1­[[[(
1S)­
1­(
6­
fluoro­
2­
benzothiazolyl)
ethyl]
amino]
carbonyl]­
2­
methylpropyl]
carbamate
CAS
#
Not
available
2.3
Physical
and
Chemical
Properties
Table
2.3.
Physicochemical
Properties
of
Benthiavalicarb
Parameter
Value
Reference
Melting
point
167
±
0.5
°
C
pH
4.35
at
25
°
C
(
1%
w/
v
aqueous
suspension)

Density
1.25
g/
cm3
at
20.5
±
0.5
°
C
Water
solubility
(
20
°
C)
10.96
mg/
L
at
pH
5
13.14
mg/
L
at
unadjusted
pH
12.76
mg/
L
at
pH
9
Solvent
solubility
(
g/
L
at
20
°
C)
Not
reported
Vapor
pressure
at
25
°
C
<
3.0
x
10­
4
Pa
Dissociation
constant
(
pKa)
No
dissociation
from
pH
1.12­
12.81
at
20
°
C
Octanol/
water
partition
coefficient
Log(
KOW)
2.52­
2.59
at
pH
5­
9
UV/
visible
absorption
spectrum
(
 max,
nm)
Not
reported
Section
A
of
PP#
3E6545
Page
11
of
87
3.0
Metabolism
Assessment
3.1
Comparative
Metabolic
Profile
The
metabolism
of
benthiavalicarb
in
rats
primarily
involves
either
(
1)
hydroxylation
of
the
phenyl
ring
followed
by
glucuronic
acid
conjugation
or
glutathione
conjugation,
or
(
2)
hydroxylation
of
the
phenyl
ring
followed
by
glutathione
conjugation
with
subsequent
degradation
of
the
glutathione
moiety
resulting
in
a
variety
of
metabolites
present
in
small
proportions.
Cleavage
and
hydroxylation
of
the
valyl
side
chain
also
occurs
to
a
limited
extent.
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
residue
in
fruit
and
foliage
(
55­
95%
TRR).
Parent
was
also
the
principal
14C­
residue
in
potato
foliage
(
88­
90%
TRR),
but
other
minor
metabolites
were
identified.
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
Secondary
metabolism
occurs
via
conjugation
of
sugars.
Chiral
analysis
of
potato
foliage
extracts
also
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).
Based
on
the
results
of
the
metabolism
studies,
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.
The
nature
of
the
residue
in
livestock
was
not
determined,
and
is
not
required,
as
there
are
no
significant
livestock
feed
items
associated
with
grapes
and
tomatoes.

The
metabolism
of
benthiavalicarb
is
more
extensive
in
rats
than
it
is
in
grapes,
tomatoes,
and
potatoes.
In
all
plant
species
tested,
metabolism
of
parent
benthiavalicarb
occurred
to
a
very
limited
extent.
In
both
plants
and
animals,
the
primary
metabolic
pathways
are
hydroxylation
of
the
phenyl
ring
with
subsequent
conjugation
with
sugars
or
glutathione.
The
only
metabolic
pathway
occurring
in
plants
that
did
not
occur
in
rats
was
the
defluorination
that
occurred
in
potatoes.

3.2
Nature
of
the
Residue
in
Foods
3.2.1.
Description
of
Primary
Crop
Metabolism
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
residue
in
fruit
and
foliage
(
55­
95%
TRR).
Parent
was
also
the
principal
14C­
residue
in
potato
foliage
(
88­
90%
TRR),
but
other
minor
metabolites
were
identified.
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
Secondary
metabolism
occurs
via
conjugation
of
sugars.
Chiral
analysis
of
potato
foliage
extracts
also
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).
Based
on
the
results
of
the
metabolism
studies,
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.

In
grapes,
the
total
radioactive
residues
(
TRR)
were
0.241
ppm
in/
on
grapes
and
21.66
ppm
in/
on
grapevine
foliage
from
the
[
Val­
14C]
test
and
0.327
ppm
in
grapes
and
14.01
ppm
in
foliage
from
the
[
Bz­
14C]
test.
HPLC
analyses
identified
parent
(
94.0­
96.5%
TRR)
as
the
main
component
in
both
fruit
and
foliage.
No
other
metabolites
were
identified,
and
isolated
unknowns
accounted
for
Page
12
of
87
 
2.0%
of
the
TRR
in
each
matrix.
Enzymatic
hydrolysis
of
foliage
extracts
with
 ­
glucosidase
did
not
show
evidence
of
glucoside
conjugation.
In
tomatoes,
HPLC
analyses
identified
parent
as
comprising
54.8­
95.1%
of
the
TRR
in
tomatoes
and
tomato
foliage.
Parent
was
the
only
compound
detected
in
either
fruit
or
foliage.
In
the
56­
DAT
fruit,
unknowns
accounted
for
a
total
of
40.9%
of
the
TRR,
but
each
unknown
accounted
for
 
9.4%
of
the
TRR
(<
0.001
ppm).

In
potatoes,
after
a
single
foliar
application
to
immature
plants,
parent
(
65­
75%
TRR)
was
the
only
component
detected
in
foliage
14
DAT.
Following
a
single
postemergence
soil
application,
14C­
residues
in
tubers
were
too
low
for
analysis,
and
the
major
component
detected
in
foliage
(
90
DAT)
was
the
sugar
conjugate
Metabolite
1
(
28­
30%
TRR).
Parent
also
accounted
for
10­
11%
of
the
TRR
in
foliage
and
Metabolites
2,
3,
and
6
each
accounted
for
3­
17%
of
the
TRR
(
0.001­
0.011
ppm).
Following
the
last
of
six
broadcast
foliar
applications,
the
primary
component
identified
in
foliage
at
14
DAT
was
parent
(
88­
90%
TRR),
along
with
minor
amounts
of
Metabolites
1,
2,
and
6
(
0.8­
3.2%
TRR).
Treatment
with
 ­
glucosidase
and
LC/
MS
analysis
identified
Metabolites
1,
2,
and
6,
and
sugar
conjugates
of
ring­
hydroxylated
metabolites.
In
tubers,
the
primary
metabolites
detected
were
Unknown
Metabolites
4
(
40%
TRR)
and
5
(
23%
TRR),
along
with
trace
amounts
of
parent
(
4.7%
TRR).
Treatment
with
 ­
glucosidase
suggests
that
Metabolites
4
and
5
are
also
sugar
conjugates;
however,
concentrations
of
these
metabolites
were
too
low
(<
0.006
ppm)
for
additional
analysis.

3.2.2
Description
of
Livestock
Metabolism
The
raw
agricultural
commodities
(
RAC)
and
processed
commodities
associated
with
the
proposed
uses
on
imported
grapes
and
tomatoes
will
not
be
used
as
significant
animal
feed
items.
Therefore,
a
determination
of
the
nature
of
the
residue
in
livestock
is
not
required
for
this
petition.

3.2.3
Description
of
Rotational
Crop
Metabolism,
including
identification
of
major
metabolites
and
specific
routes
of
biotransformation
The
proposed
uses
are
for
crops
grown
in
Europe
for
importation
to
the
U.
S.;
therefore,
rotational
crop
studies
are
not
required
for
this
petition.

3.3
Environmental
Degradation
The
tolerances
being
granted
in
conjunction
with
these
requests
are
for
imported
commodities
only.
As
a
result,
residues
in
water
are
not
being
considered
in
this
risk
assessment.

3.4
Toxicity
Profile
of
Major
Metabolites
and
Degradates
In
grapes,
tomatoes,
and
potato
foliage,
parent
benthiavalicarb
was
the
principal
14C
residue.
In
potato
foliage,
minor
metabolites
were
identified.
In
rats,
there
were
no
major
metabolites.
In
plants
and
rats,
the
minor
metabolic
products
that
formed
include
hydroxylated
compounds
in
which
the
OH
group
appears
on
the
phenyl
ring
and
to
a
small
extent
on
the
valyl
side
chain.
Other
metabolites
include
sugar
and
glutathione
conjugates
(
of
the
phenyl
ring).
It
is
unlikely
that
these
hydroxy
metabolites,
sugar
conjugates,
and
glutathione
conjugates
will
be
more
toxic
than
the
parent
compound
itself.
Page
13
of
87
3.5
Summary
of
Residues
for
Tolerance
Expression
and
Risk
Assessment
3.5.1
Tabular
Summary
Table
3.5.
Summary
of
Metabolites
and
Degradates
to
be
Included
in
the
Risk
Assessment
and
Tolerance
Expression
Matrix
Residues
included
in
Risk
Assessment
Residues
included
in
Tolerance
Expression
Primary
Crop
Parent
Benthiavalicarb­
Isopropyl,
both
R­
L
and
SL
isomers
Parent
Benthiavalicarb­
Isopropyl,
both
R­
L
and
SL
isomers
Plants
Rotational
Crop
N/
A
N/
A
Ruminant
N/
A
N/
A
Livestock
Poultry
N/
A
N/
A
Drinking
Water
N/
A
N/
A
3.5.2.
Rationale
for
Inclusion
of
Metabolites
and
Degradates
Based
on
the
results
of
the
metabolism
studies,
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
residue
in
fruit
and
foliage
(
55­
95%
TRR).
Parent
was
also
the
principal
14C­
residue
in
potato
foliage
(
88­
90%
TRR),
but
other
minor
metabolites
were
identified.
Chiral
analysis
of
potato
foliage
extracts
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).
Although
two
metabolites
in
potato
tubers
comprised
23%
and
40%
of
the
TRR,
these
residues
were
present
at
very
low
levels
(<
0.006
ppm),
and
appear
to
be
sugar
conjugates.
In
plants
and
rats,
the
metabolic
products
include
hydroxylated
compounds
in
which
the
OH
group
appears
on
the
phenyl
ring
and
to
a
small
extent
on
the
valyl
side
chain.
Other
minor
metabolites
include
sugar
and
glutathione
conjugates
(
of
the
phenyl
ring).
It
is
unlikely
that
these
hydroxy
metabolites,
sugar
conjugates,
and
glutathione
conjugates
will
be
more
toxic
than
the
parent
compound
itself.
As
a
result,
taking
into
account
the
low
%
TRR
or
low
ppm
levels
of
the
metabolites,
a
risk
assessment
of
the
parent
compound
will
be
protective
of
the
metabolites
as
well.

Both
the
R­
L
and
S­
L
isomers
should
be
included
in
the
tolerance
expression
as
well.
The
results
of
the
field
trials
indicate
that
the
residue
levels
of
the
S­
L
isomer
are
often
non­
negligible
compared
to
the
levels
of
the
R­
L
isomer.
In
the
grape
field
trials
several
samples
had
S­
L
isomer
levels
that
were
10­
20%
of
the
R­
L
isomer
levels.
A
small
number
of
samples
had
S­
L:
R­
L
ratios
that
exceeded
20%.
In
the
tomato
field
trials,
the
S­
L
residue
levels
were
generally
non­
detectable.
In
a
few
cases,
however,
the
S­
L
isomer
was
present
at
6­
7%
of
the
Page
14
of
87
level
of
the
R­
L
isomer.
The
technical
material
contains
93%
R­
L
isomer
and
3.4%
S­
L
isomer.
The
analytical
method
determines
both
isomers
separately.

4.0
Hazard
Characterization/
Assessment
4.1
Hazard
Characterization
The
toxicology
database
for
benthiavalicarb
is
complete,
including
the
studies
needed
for
assessment
of
the
FQPA
Safety
Factor.
The
liver
is
the
target
organ
of
toxicity
for
benthiavalicarb.
Increased
absolute
and
relative
liver
weights
were
seen
in
four
species
(
rat,
mouse,
rabbit,
dog),
accompanied
by
hepatocyte
hypertrophy
and
lesions.
Chronic
exposure
resulted
in
kidney
toxicity
in
rats
and
mice.
During
the
carcinogenicity
studies
in
mice,
prolonged
exposure
to
benthiavalicarb
was
linked
to
increased
incidences
of
hepatocellular
adenomas,
carcinomas,
and
blastomas.
Benthiavalicarb
was
also
linked
to
thyroid
tumors
in
male
mice
and
uterine
tumors
in
female
rats.
Benthiavalicarb
is
not
mutagenic,
nor
does
it
produce
adverse
developmental,
reproductive,
or
neurotoxic
effects.

In
acute
oral
toxicity
studies,
no
mortalities
were
observed
up
to
the
limit
dose
of
2000
mg/
kg.
This
chemical
is
classified
as
Toxicity
Category
III.
No
other
acute
studies
were
available
for
this
chemical.

Benthiavalicarb
was
administered
to
mice
(
MRID
45835007),
rats
(
MRID
45835004),
and
dogs
(
MRID
45835006)
for
90
days
via
their
diet.
All
three
species
had
increased
absolute
and
relative
liver
weights
and
hepatocyte
hypertrophy
at
their
respective
LOAELs.
The
adverse
effects
observed
included
decreases
in
hematocrit,
hemoglobin,
serum
albumin,
and
albumin/
globulin
ratios
in
females
and
hemosiderin
pigment
deposits
in
the
spleen
of
males
at
200
mg/
kg/
day.
At
the
LOAEL
of
352
mg/
kg/
day
in
rats,
increases
in
total
cholesterol
and
GGT
were
evident
in
both
sexes;
free
cholesterol
and
phospholipids
were
increased
in
the
females;
and
total
protein
was
increased
in
males.
Body
weight
gains
and
food
consumption
were
decreased
in
male
and
female
mice
dosed
above
1293
mg/
kg/
day.
In
mice,
liver
lesions
that
occurred
at
or
above
a
dose
of
1293
mg/
kg/
day
included
black
colored
livers
in
females
and
necrosis,
hypertrophy,
and
anisonucleosis
in
males.

In
the
chronic
rat
(
MRID
45835017)
and
mouse
(
MRID
45835016)
studies,
the
effects
were
found
primarily
in
the
liver.
Both
species
had
increased
absolute
and
relative
liver
weights,
hepatocyte
hypertrophy,
and
gross
liver
lesions.
The
chronic
study
in
dogs
(
MRID
45835015)
demonstrated
minimal
increases
in
absolute
and
relative
liver
weights.
In
mice,
liver
lesions
included
brown
patches
and
nodules
in
both
sexes,
enlarged
livers
in
females,
intermediate
fatty
change,
and
hepatic
foci
of
cellular
alteration
in
both
sexes.
In
male
mice,
anisonucleosis,
necrosis,
single
cell
necrosis,
multinucleated
cells,
accumulation
of
macrophages,
cellular
infiltration,
lymphocyte
infiltration,
bile
duct
proliferation,
and
extramedullary
hematopoiesis
were
also
present
at
the
LOAEL
of
358
mg/
kg/
day.
Similar
liver
lesions
were
present
in
rats
at
doses
greater
than
250
mg/
kg/
day.
In
both
sexes
of
mice,
thyroid
cell
hyperplasia
was
present
at
the
LOAEL.
In
males,
dilatated
thyroid
follicles
were
present
at
the
LOAEL.
In
the
chronic
rat
study,
the
kidney
was
also
a
target
organ
of
toxicity.
The
following
kidney
lesions
were
observed
at
the
LOAEL
of
250
mg/
kg/
day:
chronic
nephropathy
and
tubular
dilatation
in
the
males,
Page
15
of
87
glomerulosclerosis;
calculus
and
hyaline
droplets
in
both
sexes;
lymphocytic
cellular
infiltration,
basophilic
tubules,
hyaline
casts,
and
brown
pigment
deposits
in
females.

Several
tumor
types
were
seen
in
the
carcinogenicity
studies
in
both
rats
(
MRID
45835017)
and
mice
(
MRID
45835016).
In
the
rat
carcinogenicity
study,
animals
were
exposed
to
0,
50,
200,
5,000,
and
10,000
ppm
of
benthiavalicarb
via
dietary
admix.
Male
rats
in
the
high­
dose
group
had
a
statistically
higher
incidence
of
hepatocellular
adenomas
than
did
the
control
group.
The
incidence
rate
was
within
the
range
of
historical
control
data;
therefore,
the
CARC
concluded
that
this
tumor
type
in
rats
may
not
be
directly
related
to
exposure
to
benthiavalicarb.
However,
a
similar
increase
in
hepatocellular
tumors
occurred
in
mice,
and
the
CARC
concluded
that
the
liver
tumor
data
in
rats
may
provide
some
supporting
evidence
of
carcinogenic
potential
for
liver
tumors.
In
the
mouse
carcinogenicity
study,
animals
were
administered
0,
20,
100,
2500,
or
5,000
ppm
of
benthiavalicarb
in
their
diets.
Male
mice
in
the
2,500
and
5,000
ppm
dose
groups
had
statistically
higher
incidences
of
hepatocellular
adenomas,
carcinomas,
blastomas,
and
combined
tumors
than
controls.
Female
mice
in
the
top
two
dose
groups
had
increased
incidences
of
hepatocellular
adenomas.
The
increases
in
hepatocellular
tumors
in
mice
were
considered
to
be
treatment­
related,
and
this
effect
was
used
to
calculate
the
Q1*
for
risk
assessment.
All
doses
in
the
rat
and
mouse
studies
were
considered
adequate,
but
not
excessive,
for
determination
of
carcinogenic
potential,
except
for
the
5,000
ppm
dose
in
mice,
which
was
considered
to
be
excessive
because
it
resulted
in
severe
systemic
toxicity.

Female
rats
in
the
top
two
dose
groups
had
a
significantly
increased
incidence
of
uterine
adenocarcinomas
compared
to
concurrent
controls.
The
incidence
rate
also
exceeded
the
rates
in
historical
control
data.
Therefore,
uterine
tumors
in
rats
were
considered
to
be
treatment­
related.
Similarly
the
increased
incidence
of
follicular
cell
adenomas
in
male
mice
in
the
2,500
and
5,000
ppm
dose
groups
were
considered
to
be
treatment
related.
Follicular
cell
hyperplasia
was
also
present
in
these
dose
groups.

Benthiavalicarb
is
classified
as
"
likely
to
be
carcinogenic
to
humans."

Eleven
mechanistic
studies
were
submitted
in
order
to
determine
a
mode
of
action
for
benthiavalicarb.
After
thorough
consideration
by
the
CARC,
it
was
concluded
that
the
data
were
insufficient
to
support
definitive
modes
of
action
for
each
tumor
type.
Therefore,
the
CARC
recommended
using
a
linear
low­
dose
extrapolation
approach
(
Q1*)
for
estimating
human
cancer
risk
based
upon
the
most
potent
tumor
type.
After
quantitative
analysis
of
the
tumor
types,
the
Q1*
was
based
on
combined
hepatocellular
tumors
in
male
mice.
The
Q1*
is
6.2795x10­
2.

More
than
20
mutagenicity
studies
were
submitted
for
benthiavalicarb,
testing
the
mutagenic
potential
of
various
lots
of
the
parent
and
a
number
of
congeners.
All
in
vitro
and
in
vivo
studies,
except
two,
were
negative
for
the
induction
of
mutagenicity
up
to
the
limit
dose
or
precipitation
of
the
compound.
The
two
exceptions
of
positive
results
were
with
two
lots
of
the
parent
compound
in
the
TA98
strain
of
S.
typhimurium
in
the
presence
of
S9­
activation.
However,
later
studies
demonstrated
that
these
positive
results
were
not
due
to
the
parent,
but
to
a
mutagenic
impurity
ordinarily
at
low,
ineffective
levels
in
all
but
these
two
batches.

During
a
metabolism
study
in
rats
(
MRID
45835108),
both
the
low
and
high
dose
radiolabeled
forms
were
absorbed
rapidly,
with
nearly
full
recovery
of
radioactive
label
(
89­
100%)
within
168
Page
16
of
87
hours.
Radioactivity
was
recovered
primarily
in
the
feces
(
62­
83%)
and
urine
(
7­
24%),
with
up
to
85%
recovery
in
48
hours.
Urinary
excretion
was
slightly
higher
in
females
than
males.
Absorption
was
greater
in
the
low
dose
group
than
in
the
high
dose
group,
indicating
incomplete
absorption
in
the
high
dose
group.
Radiolabeled
residues
preferentially
partitioned
into
plasma.
In
both
dose
groups
(
at
all
time
points),
the
concentration
of
radioactivity
was
higher
in
the
kidneys
and
liver
than
it
was
in
plasma.

Analysis
of
the
metabolites
in
bile,
urine,
and
feces
of
cannulated
rats
identified
a
total
of
13
compounds,
including
the
parent,
which
accounted
for
43­
60%
of
the
radiolabeled
dose.
The
metabolite
profile
was
similar
between
the
sexes.
A
total
of
11
metabolites
were
identified
in
the
urine,
most
present
in
concentrations
<
2%
of
the
administered
dose.
A
total
of
8
metabolites
were
found
in
the
bile,
with
the
principal
metabolites
being
isomers
of
hydroxylated
glutathione
conjugates
and
a
glucuronide
conjugate.
Parent
compound
was
not
present
in
the
urine
or
bile.

Based
upon
the
components
identified
in
excreta,
the
metabolism
of
benthiavalicarb
in
rats
primarily
involves
either
hydroxylation
of
the
phenyl
ring
followed
by
glucuronic
acid
conjugation
or
glutathione
conjugation
and
hydroxylation
of
the
phenyl
ring
with
subsequent
degradation
of
the
glutathione
moiety,
resulting
in
a
variety
of
metabolites
present
in
small
quantities.
Cleavage
and
hydroxylation
of
the
valyl
side
chain
also
occurs
to
a
limited
extent.
Parent
(
excreted
in
feces
only)
and
metabolites,
M15
(
urine
and
feces)
and
M18
(
urine
only),
were
also
the
principal
components
detected
in
plasma,
liver,
and
kidneys.

Table
4.1.
Acute
Toxicity
Profile
­
KIF­
230
Guideline
No.
Study
Type
MRID(
s)
Results
Toxicity
Category
870.1100
Acute
oral
rat
KIF­
230
Tech:
45834909
KIF­
230
S­
L:
45834910
KIF­
230­
1­
KR:
45835011
KIF­
230­
1­
1(
S):
45835002
KIF­
230­
1­
13:
45835003
LD50
>
2000
mg/
kg
III
870.1200
Acute
dermal
[
species]
NA
870.1300
Acute
inhalation
[
species]
NA
870.2400
Acute
eye
irritation
[
species]
NA
870.2500
Acute
dermal
irritation
[
species]
NA
870.2600
Skin
sensitization
[
species]
NA
NA
=
not
available
Page
17
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
Subchronic
Studies
870.3100
90­
Day
oral
toxicity
rodents
(
Fischer
rats)
45835004
(
1998)
Acceptable/
Guideline
0,
50,
200,
7000,
or
20,000
ppm
M:
0,
3.5,
14.1,
353,
or
1444
mg/
kg/
day
F:
0,
3.9,
15.3,
379,
or
1552
mg/
kg/
day
NOAEL
=
14.1/
15.3
mg/
kg/
day
LOAEL
=
353/
379
mg/
kg/
day
based
on
hepatocyte
hypertrophy
in
both
sexes
and
increases
in:
absolute
and
relative­
to­
body
liver
weights,
total
cholesterol,
and
GGT
in
both
sexes;
free
cholesterol
and
phospholipids
in
the
females;
and
total
protein
in
the
males.

870.3100
90­
Day
oral
toxicity
rodents
(
B6C3F1
mice)
45835007
(
1998)
Acceptable/
guideline
0,
50,
200,
7000,
or
20,000
ppm
M:
0,
8.4,
33.0,
1293,
or
4031
mg/
kg/
day
F:
0,
11.3,
45.2,
1620,
or
4946
mg/
kg/
day
NOAEL
=
33.0/
45.2
mg/
kg/
day
LOAEL
=
1293/
1620
mg/
kg/
day
based
on
decreased
body
weight,
body
weight
gain,
and
food
efficiency
in
males,
increased
absolute
and
relative
liver
weight,
enlarged
liver,
black­
colored
liver
(
females
only),
and
histopathological
liver
effects
(
necrosis,
hypertrophy,
and
anisonucleosis
[
males
only]).

870.3150
90­
Day
oral
toxicity
in
dogs
(
beagle)
45835005
(
1999)
Acceptable/
Guideline
0,
40,
200,
or
1000
mg/
kg/
day
NOAEL
=
40
mg/
kg/
day
LOAEL
=
200
mg/
kg/
day
based
on
decreases
in
hematocrit,
hemoglobin,
serum
albumin
and
albumin/
globulin
ratio
in
females
and
increases
in
liver­
to­
body
weight
ratios,
hepatocyte
hypertrophy
(
both
sexes)
and
hemosiderin
pigment
deposits
in
the
spleen
of
males.

Developmental
and
Reproduction
Studies
870.3700a
Prenatal
developmental
in
rodents
(
Sprague­
Dawley)
45835012
(
2000)
Acceptable/
Guideline
0,
10,
100,
or
1000
mg/
kg/
day
Maternal
NOAEL
=
100
mg/
kg/
day
LOAEL
=
1000
mg/
kg/
day
based
on
increased
absolute
and
relative
liver
weights
and
on
increased
incidence
of
enlarged
liver.
Developmental
NOAEL
=
1000
mg/
kg/
day
LOAEL
=
not
observed.

870.3700b
Prenatal
developmental
in
rabbits
(
New
Zealand
White)
45835010
(
2000)
Acceptable/
Guideline
0,
10,
20,
or
40
mg/
kg/
day
Maternal
NOAEL
=
20
mg/
kg/
day
LOAEL
=
40
mg/
kg/
day
based
on
increased
absolute
and
relative
liver
weights.
Developmental
NOAEL
=
40
mg/
kg/
day
LOAEL
=
not
observed.

870.3800
Reproduction
and
fertility
effects
(
Sprague­
Dawley)
45835014
(
1999)
Acceptable/
Guideline
0,
100,
1000,
or
10,000
ppm
M:
0,
8.4,
84.1,
or
880
mg/
kg/
day
F:
0,
8.8,
91.0,
or
943
mg/
kg/
day
Parental/
Systemic
NOAEL
=
880/
943
mg/
kg/
day
LOAEL
=
not
observed.
Reproductive
NOAEL
=
880/
943
mg/
kg/
day
LOAEL
=
not
observed.
Offspring
NOAEL
=
880/
943
mg/
kg/
day
LOAEL
=
not
observed.

Chronic
and
Carcinogenicity
Studies
Page
18
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
870.4100a
Chronic
toxicity
rats
(
Fischer)
45835017
(
2001)
Acceptable/
Guideline
0,
50,
200,
5000,
or
10,000
ppm
M:
0,
2.5,
9.9,
249.6,
or
518.3
mg/
kg/
day
F:
0,
3.2,
12.5,
318.2,
or
649.4
mg/
kg/
day
NOAEL
=
9.9/
12.5
mg/
kg/
day
LOAEL
=
249.6/
318.2
mg/
kg/
day
based
on
nephrotoxicity
and
hepatotoxicity
in
both
sexes.
In
the
kidney,
absolute
and
relative
weights
were
increased
in
both
sexes.
Kidney
lesions
included
chronic
nephropathy
and
tubular
dilatation
in
males;
glomerulosclerosis,
calculus,
and
hyaline
droplets
in
both
sexes;
lymphocytic
cellular
infiltration,
basophilic
tubules,
hyaline
casts,
and
brown
pigment
deposits
in
females.
Liver
findings
included
transient
increased
_­
glutamyltranspep­
tidase
in
both
sexes;
increased
levels
of
total
and
free
cholesterol
and
phospholipids
in
females;
increased
absolute
and
relative
liver
weights
in
both
sexes;
increased
incidences
of
brown
and/
or
white
patches
on
the
liver
in
males,
and
red
patches
on
the
liver
in
females;
increased
incidences
of
hepatocellular
hypertrophy;
and
increased
incidences
of
fatty
liver
in
both
sexes.
At
week
104,
increased
incidences
of
foci/
area
of
cellular
alteration
in
the
liver
and
spongiosis
hepatis
were
observed
in
the
males.

Increased
incidences
of
hepatocellular
tumors
in
males
and
uterine
tumors
in
females
were
observed.

870.4100b
Chronic
toxicity
dogs
(
beagle)
45835015
(
2001)
Acceptable/
Guideline
0,
4,
40,
or
400
mg/
kg/
day
NOAEL
=
40
mg/
kg/
day
LOAEL
=
400
mg/
kg/
day
based
on
minimal
increases
in
absolute
and
relative
liver
weights
(
not
significant)
at
the
highest
dose
level.

870.4300
Carcinogenicity
mice
(
B6C3F1)
45835016
(
2001)
Acceptable/
Guideline
0,
20,
100,
2500,
or
5000
ppm
M:
2.7,
13.7,
358.4,
or
731.3
mg/
kg/
day
F:
0,
3.7,
18.6,
459.3,
or
927.8
mg/
kg/
day
NOAEL
=
13.7/
18.6
mg/
kg/
day
LOAEL
=
358.4/
459.3
mg/
kg/
day
based
on
clinical
signs
(
abdominal
masses
and
distention,
and
pallor)
in
males,
decreased
body
weight
gain
and
food
efficiency
in
males;
gross
liver
lesions
(
brown
patches
and
nodules
in
both
sexes,
and
enlarged
livers
in
females);
increased
absolute
and
relative
to
body
liver
weight
in
both
sexes;
non­
neoplastic
liver
lesions
(
hepatocyte
hypertrophy,
hepatic
intermediate
fatty
change,
hepatic
foci
of
cellular
alteration
in
both
sexes,
anisonucleosis,
necrosis,
single
cell
necrosis,
multinucleated
cells,
accumulation
of
macrophages,
cellular
infiltration,
lymphocyte
infiltration,
bile
duct
proliferation
and
extramedullary
hematopoiesis
in
males);
thyroid
follicular
cell
hyperplasia
in
both
sexes;
and
dilated
thyroid
follicles
in
males.
no
evidence
of
carcinogenicity
Neurotoxicity
Studies
870.6200a
Acute
neurotoxicity
screening
battery
in
rats
(
Sprague­
Dawley)
45869735
(
2002)
Acceptable/
Guideline
0
or
2000
mg/
kg
NOAEL
=
2000
mg/
kg
LOAEL
=
not
observed
Subchronic
neurotoxicity
study
in
rats
(
CD
rats)
45869736
(
2002)
Acceptable/
Guideline
0,
200,
2000,
or
20,000
NOAEL
=
174.1/
185.7
mg/
kg/
day
LOAEL
=
1853.7/
1845.8
mg/
kg/
day
based
on
decreased
body
weight
and
body
weight
gains
in
the
males.
Page
19
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
ppm
M:
0,
17.7,
174.1,
or
1853.7
mg/
kg/
day
F:
0,
19.3,
185.7,
or
1845.8
mg/
kg/
day
Gene
Mutation
Studies
Gene
Mutation
870.5300
Mutation
at
the
Thymidine
Kinase
Locus
of
Mouse
Lymphoma
Cells
45869728
(
1999)
Acceptable/
Guideline
KIF­
230
technical
Negative
for
increases
in
mutant
frequency
up
to
the
highest
soluble
concentration
in
the
presence
or
absence
of
activation.

Gene
Mutation
Nonguideline
Gene
Mutation
in
Transgenic
Mice
45869730
(
2000)
Acceptable/
Non­
guideline
KIF­
230
technical
Negative
for
the
induction
of
in
vivo
gene
mutation
in
mice
treated
up
to
the
limit
dose.

Reverse
Gene
Mutation
870.5100
45869722
(
1999)
Acceptable/
Guideline
KIF­
230
technical
Positive
for
gene
mutation
in
the
presence
of
rat
liver
activation
when
assayed
up
to
5000
_
g/
plate
(
because
of
impurity).

Reverse
Gene
Mutation
870.5100
45835106
(
2001)
Acceptable/
Guideline
KIF­
230S­
L
Negative
for
reverse
gene
mutation
up
to
the
limit
of
solubility.

Reverse
Gene
Mutation
870.5100
45869717
(
2002)
Acceptable/
Guideline
KIF­
230
Lot
G51­
47­
190
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869718
(
2002)
Acceptable/
Guideline
KIF­
230
Lot
G51­
48­
190
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869719
(
2002)
Acceptable/
Guideline
KIF­
230
Lot
G51­
49­
190
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869720
(
2002)
Acceptable/
Guideline
KIF­
230
Lot
G51­
50­
190
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869721
(
2002)
Acceptable/
Guideline
KIF­
230
Lot
G51­
51­
190
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869723
(
2001)
Acceptable/
Guideline
KIF­
230
Lot
G51­
35­
184
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
45869724
(
2001)
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
Page
20
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
Mutation
870.5100
Acceptable/
Guideline
KIF­
230
Lot
G51­
37­
184
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869725
(
2001)
Acceptable/
Guideline
KIF­
230
Lot
G51­
36­
184
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869726
(
2001)
Acceptable/
Guideline
KIF­
230
Lot
G51­
08­
158
This
lot
had
innate
mutagenic
activity
in
one
bacterial
strain
in
the
presence
of
S9
activation
when
exposed
to
concentrations
<
320
mg/
plate.
In
a
repeat
study,
the
positive
response
was
demonstrated
to
be
due
to
a
mutagenic
impurity
(
45835107)

Reverse
Gene
Mutation
870.5100
45869732
(
2001)
Acceptable/
Guideline
KIF­
230­
I­
1
(
R)
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45839733
(
2001)
Acceptable/
Guideline
KIF­
230­
I­
1
(
S)
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Reverse
Gene
Mutation
870.5100
45869734
(
2001)
Acceptable/
Guideline
KIF­
230­
I­
13
Negative
for
inducing
gene
mutations
in
the
presence
and/
or
absence
of
activation
up
to
the
limit
dose
of
5000
_
g/
plate.

Structural
Chromosomal
Aberration
In
vitro
Chromosomal
Aberration
Test
870­
5375
45869727
(
1998)
KIF­
230
technical
Unacceptable/
Not
upgradeable
There
were
no
increased
incidences
in
chromosomal
aberrations
at
any
dose
compared
to
concurrent
or
historical
controls.
However
this
study
is
classified
as
unacceptable/
not
upgradeable
due
to
the
presence
of
a
"
white
powdery
test
substance"
at
all
concentrations
in
this
assay.

Other
Genotoxic
Effects
In
vivo/
In
vitro
Unscheduled
DNA
Synthesis
870.5550
45869731
(
2001)
Acceptable/
Guideline
Negative
for
UDS
induction
in
hepatocytes
isolated
from
rats
treated
up
to
the
limit
dose,
2000
mg/
kg.

Measurement
of
Unscheduled
DNA
Synthesis
in
vitro
870.5550
45869729
(
1999)
Acceptable/
Guideline
KIF­
230
Negative
for
UDS
induction
in
hepatocytes
isolated
from
rats
treated
up
to
the
limit
dose,
2000
mg/
kg.

Induction
of
Micronuclei
in
the
Bone
Marrow
of
Treated
mice
870.5385
45835105
(
2000)
Acceptable/
Guideline
KIF­
230
Negative
for
the
induction
of
micronucleated
polychromatic
erythrocytes
in
bone
marrow
of
mice
treated
up
to
the
limit
dose,
2000
mg/
kg.

Identification
of
a
Mutagenic
Substance
Isolated
from
KIF­
45835107
(
2002)
Summary
The
positive
results
with
Lot
Nos.
G­
51­
15­
162
and
G51­
08­
158
in
TA98
in
the
presence
of
S9
activation
previously
reported
appears
to
be
due
to
a
mutagenic
component
identified
Page
21
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
230
Nonguideline
as
6,6'­
difluoro­
2,2'­
bibenzothiazole.

Non­
Guideline
Mechanistic
Studies
Hepatocarcinogenicity
Study
in
Rats
45835021
(
2000)
Acceptable/
Non­
guideline
Five
groups
of
12
male
Fischer
rats
were
used
for
this
study.
Groups
1,
3,
and
5
were
initiated
by
i.
p.
injection
of
diethylnitrosamine
(
DEN)
in
0.9%
saline,
while
groups
2
and
4
were
dosed
with
0.9%
saline
only.
Group
1
rats
were
fed
a
normal
basal
diet,
while
groups
2
and
3
received
a
diet
with
10,000
ppm
(
586.5­
634.1
mg/
kg/
day)
KIF­
230,
and
groups
4
and
5
received
diets
including
0.05%
phenobarbital
sodium.
At
week
3
a
partial
hepatectomy
was
performed
to
stimulate
hepatic
cell
division
No
treatment­
related
effects
were
observed
on
mortality
or
clinical
signs.
Minor
decreases
in
body
weights,
cumulative
body
weight
gains,
and
food
efficiency
were
observed,
but
were
not
considered
treatment
related.

Rats
in
groups
2
and
3,
which
were
dosed
with
KIF­
230,
had
increased
absolute
and
relative
liver
weights
compared
to
controls.
Enlarged
liver,
hepatocyte
hypertrophy,
and
increased
number
of
GST­
positive
foci
were
seen
in
all
treatment
groups
(
saline/
KIF­
230,
DEN/
KIF­
230,
saline/
PB,
and
DEN/
PB).
Groups
4
and
5,
which
received
PB,
had
increased
incidence
of
fatty
change
in
the
liver.
Increased
mitosis
was
observed
in
10/
12
animals
dosed
with
DEN/
KIF­
230
and
12/
12
animals
dosed
with
DEN/
PB
compared
to
4/
12
animals
treated
with
DEN
alone.
Increased
incidence
of
acidophilic
cell
focus
was
noted
in
10/
12
animals
dosed
with
DEN/
KIF­
230
and
9/
12
animals
dosed
with
DEN/
PB,
compared
to
4/
12
animals
treated
with
DEN
alone.

Data
indicate
that
administration
of
KIF­
230
in
the
diet
at
10,000
ppm
to
male
rats
caused
a
promoter
effect
in
the
liver
similar
to
that
caused
by
phenobarbital,
a
known
tumor
promoter
in
rat
liver
in
a
two­
stage
experimental
model
of
hepatocarcinogenesis.

Hepatocarcinogenicity
Study
Examining
the
Initiator
Effect
of
KIF­
230
in
Male
Rats
45869715
(
2000)
Acceptable/
Non­
guideline
Male
Fischer
rats
were
placed
into
groups
of
12
each.
A
partial
hepatectomy
was
performed
on
all
groups
to
stimulate
cell
division.
Twelve
hours
later,
the
animals
were
initiated
with
a
single
injection
of
20
mg/
kg
DEN,
a
single
gavage
dose
of
2000
mg/
kg
KIF­
230,
or
a
single
gavage
dose
of
carboxymethyl
cellulose
sodium
salt
(
CMC­
Na).
All
rats
were
fed
a
basal
diet
containing
phenobarbital
sodium
salt
(
PB)
from
weeks
2­
10
of
the
study.
At
week
3,
all
rats
were
given
a
single
injection
of
300
No
treatment
related
signs
were
observed
on
mortality,
clinical
signs
of
toxicity,
body
weights,
body
weight
gains,
or
gross
pathology.

Absolute
and
relative
liver
weights
were
increased
in
the
KIF­
230
group
compared
to
the
DEN
group.
Fatty
change
was
observed
in
all
animals
in
all
groups.
Hepatocyte
hypertrophy
was
seen
in
the
DEN
and
KIF­
230
groups.
Mitosis
was
observed
in
6/
12
animals
in
the
DEN
group,
compared
to
0/
12
and
1/
12
animals
in
the
KIF­
230
and
CMC­
Na
groups,
respectively.
Clear
cell
foci
(
3/
12)
and
eosinophilic
cell
foci
(
8/
12)
were
observed
in
the
DEN
group
compared
to
0/
12
animals
in
the
KIF­
230
or
CMC­
Na
groups.
The
number
of
GST­
P
positive
foci
and
the
area
of
the
GST­
P
positive
foci
were
greater
in
the
DEN
groups
compared
to
the
KIF­
230
and
CMC­
Na
groups.

The
data
indicated
that
administration
of
KIF­
230
as
a
single
dose
of
2000
mg/
kg
to
male
rats
did
not
demonstrate
an
initiator
effect
in
the
liver
when
compared
to
DEN,
a
known
initiating
agent
in
the
two­
stage
hepatocarcinogenic
model.
However,
an
additional
group
of
saline/
PB
treated
rats
should
have
been
included
in
this
study
to
allow
for
full
comparison
of
treated
Page
22
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
mg/
kg
D­
galactosamine.
animals
with
untreated
controls.

Two­
stage
Transformation
Assay
45835103
(
2000)
Acceptable/
Non­
guideline
Initiation:
BALB/
c
3T3
A31­
1­
1
cells
were
exposed
to
KIF­
230
at
concentrations
of
0,
10.4,
17.3,
28.8,
48.0,
or
80.0
g/
mL
for
three
days.
Promoter:
cells
were
exposed
to
KIF­
230
at
concentrations
of
0,
3.0,
6.0,
9.0,
12.0,
or
15.0
g/
mL
for
three
days.
The
test
compound
did
not
act
as
an
initiating
or
promoting
agent
under
conditions
of
this
study.

Mechanism
Study
of
Thyroid
Tumors
in
Rats
45835018
(
2002)
Acceptable/
Non­
guideline
12
male
Fisher
rats/
sex/
dose
were
fed
KIF­
230
in
their
diets
at
concentrations
of
0,
13.3,
or
661.4
mg/
kg/
day
for
14
days.
The
data
indicated
that
the
administration
of
KIF­
230
in
the
diet
at
a
dose
of
661.4
mg/
kg/
day
resulted
in
decreased
serum
T4
levels
(
11­
18%)
in
male
rats.
Absolute
and
relative­
to­
body
liver
weights
were
increased
(
22­
24%)
compared
to
controls,
and
macroscopic
examination
revealed
enlarged
liver
in
all
rats
in
this
dose
group,
which
is
consistent
with
enzyme
induction.
TSH
levels
were
non­
significantly
increased
compared
to
controls,
and
T3
levels
remained
similar
to
controls.
The
observed
increase
in
UDP­
GT
activity
(
16%)
on
T4
could
cause
a
decrease
in
circulating
T4
levels,
resulting
in
increased
serum
TSH
levels
via
a
feedback
mechanism.
This
mechanism
could
ultimately
cause
hyperthyroidism
following
a
sufficient
duration
of
exposure
in
rats.

Mechanism
Study
of
Thyroid
Tumors
in
Mice
45835019
(
2002)
Acceptable/
Non­
guideline
12
Scl:
B6C3F1
male
mice/
dose
were
exposed
to
KIF­
230
in
their
diet
at
concentrations
of
0,
17.0,
or
855.0
mg/
kg/
day
for
14
days.
No
effects
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
food
consumption.

Increased
absolute
and
relative
liver
weights
were
observed
at
855
mg/
kg/
day,
which
is
consistent
with
enzyme
induction
and
the
increased
UDP­
GT
activity
on
T4
seen
in
this
study.
While
serum
T4
levels
were
decreased
25­
29%,
T3
and
TSH
remained
similar
to
controls
throughout
the
study.

Typical
UDP­
GT
inducers
are
known
to
cause
hyperthyroidism
by
inducing
TSH
through
a
decrease
in
serum
thyroid
hormone
levels;
and
thyroid
hyperplasia
is
a
potential
mechanism
for
the
induction
of
thyroid
tumors.

The
lack
of
an
appropriate
antibody
(
anti­
mouse
TSH)
could
cause
inaccurate
measurements
of
TSH
in
the
radioimmunoassay
used
to
determine
serum
levels
of
TSH.

While
the
mechanism
is
plausible,
no
definitive
conclusions
can
Page
23
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
be
reached
concerning
this
mechanistic
study.

Drug
­
Metabolizing
Enzymes
and
Hepatocyte
Proliferation
in
Mice
45835102
(
2001)
Acceptable/
Non­
guideline
Eight
B6C3F1
mice
/
sex/
dose
were
exposed
to
KIF­
230
by
gavage
at
concentrations
of
0,
10,
or
1000
mg/
kg/
day
for
seven
days.
Five
mice/
sex/
dose
were
then
checked
for
hepatocyte
P­
450
and
specific
P­
450
isoenzyme
levels.
Three
mice/
sex/
dose
were
evaluated
for
hepatocyte
proliferation.
No
effects
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
on
the
BrdU
labeling
index.

At
1000
mg/
kg/
day,
absolute
and
relative
liver
weights
were
increased
13­
31%.
Microscopic
examination
revealed
hypertrophy
of
the
hepatocytes
in
the
liver
of
both
males
and
females
compared
to
controls.
Total
microsomal
P­
450
levels
were
increased
(
71­
92%)
in
both
sexes,
CYP1A2
was
increased
63­
149%,
CYP2B1
(
2B2)
increased
180­
553%,
and
CYP3A2
increased
104­
172%.
CYP1A1,
CYP2E1,
and
CYP4A1
levels
were
similar
to
control.

The
increased
liver
weights
and
increased
liver
enzymes
are
consistent
with
enzyme
induction
and
hepatocyte
hypertrophy.
The
level
of
enzyme
induction
observed
was
low
compared
to
the
induction
seen
with
classical
induction
agents
like
phenobarbital.

Drug
­
Metabolizing
Enzymes
and
Hepatocyte
Proliferation
in
Rats
45835101
(
2001)
Acceptable/
Non­
guideline
Eight
Fischer
rats/
sex/
dose
were
exposed
to
KIF­
230
by
gavage
at
concentrations
of
0,
10,
or
1000
mg/
kg/
day
for
seven
days.
Five
rats/
sex/
dose
were
then
checked
for
hepatocyte
P­
450
and
specific
P­
450
isoenzyme
levels.
Three
mice/
sex/
dose
were
evaluated
for
hepatocyte
proliferation.
No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
gains,
gross
or
microscopic
pathological
examinations,
or
on
the
hepatic
BrdU
labeling
index.

At
1000
mg/
kg/
day,
absolute
and
relative
liver
weights
were
increased
8­
22%.
In
males,
total
hepatic
microsomal
P­
450
levels
were
increased
18%,
and
several
specific
CYP
enzymes
were
increased101­
160%.
In
females,
specific
CYP
enzymes
were
increased
76­
130%.
Levels
of
CYP2E1
and
CYP4A1
were
similar
to
controls
in
both
sexes.

KIF­
230
at
1000
mg/
kg/
day
for
seven
days
resulted
in
increased
liver
weights
and
enzyme
induction.
Unchanged
levels
of
BrdU
indicate
that
there
was
no
treatment­
related
effect
on
hepatocyte
proliferation.
The
level
of
P­
450
induction
in
males
was
greater
than
in
females,
but
the
magnitude
of
difference
does
not
clearly
indicate
a
sex
difference.

Oxidative
DNA
in
Mice
45835022
(
2001)
Acceptable/
Non­
guideline
Five
Scl:
B6C3F1
mice/
sex/
dose
were
exposed
to
KIF­
230
in
their
diets
at
concentrations
of
0,
100,
or
5000
ppm
(
0,
19.4/
26.1,
or
1031.2/
1203.7
mg/
kg/
day
male/
female)
for
up
to
2
weeks.
No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
food
consumption.

At
5000
ppm,
absolute
and
relative
body
weights
were
increased
(
75­
78%).
No
effects
were
observed
on
8­
OHdG
levels
measured
in
the
liver
tissue
DNA.

Liver
enlargement
was
consistent
with
enzyme
induction.
There
was
no
indication
of
hepatic
oxidative
DNA
damage,
as
measured
by
8­
OHdG
production.
Page
24
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
Oxidative
DNA
Damage
in
Rat
Liver
45869716
(
2001)
Acceptable/
Non­
guideline
Five
Fischer
rats/
sex/
dose
were
exposed
to
KIF­
230
in
their
diets
at
concentrations
of
0,
200,
or
10,000
ppm
(
0,
17.4/
17.1,
or
797.5/
914.5
mg/
kg/
day
male/
female)
for
up
to
2
weeks.
No
treatment­
related
effects
were
observed
on
mortality
or
clinical
signs.

At
10,000
ppm,
absolute
and
relative
liver
weights
were
increased
(
73­
113%)
in
both
sexes.
At
200
ppm,
relative
liver
weights
were
increased
slightly
in
females
(
9%).
8­
OHdG
levels
were
decreased
in
the
200
and
10,000
ppm
males
(
20%
each)

Body
weights
and
cumulative
body
weight
gains
were
decreased
(
12­
50%)
in
the
high
dose
males.
Decreased
food
consumption
was
also
observed
in
these
animals.

Liver
enlargement
was
consistent
with
enzyme
induction.
There
was
no
indication
of
hepatic
oxidative
DNA
damage,
as
measured
by
8­
OHdG
production.

Mechanism
Study
of
Uterine
Cancer
in
Rats
45835020
(
2002)
Acceptable/
Non­
guideline
12
Fischer
female
rats/
dose
were
exposed
to
KIF­
230
in
the
diet
at
concentrations
of
0,
200,
or
10,000
ppm
(
0,
11.6,
or
576.4
mg/
kg/
day)
for
up
to
8
weeks.
Microsomal
aromatase
activity
was
measured
at
week
8,
while
serum
estradiol
(
E2),
progesterone
(
P4),
and
luteinizing
hormone
(
LH)
levels
were
measured
at
weeks
2,
4,
6,
and
8.
No
treatment
related
effects
were
observed
on
mortality,
clinical
signs
of
toxicity,
body
weight,
cumulative
body
weight
gains,
or
food
consumption.

At
576.4
mg/
kg/
day,
microsomal
aromatase
activity
was
increased
in
the
liver
36%
and
slightly
increased
in
the
uterus
(
15%),
but
was
decreased
in
the
ovary
( 
10%).
No
treatmentrelated
effects
were
observed
on
serum
sex
hormone
levels.
Ovary
and
uterine
weights
were
unaffected
by
treatment,
and
no
effects
were
noted
during
necropsy.
Absolute
and
relative
liver
weights
were
increased
(
28­
31%),
and
macroscopic
examination
revealed
dark
liver
in
10/
11
rats
and
enlarged
liver
in
11/
11
rats
compared
to
0/
10
controls.

The
data
indicate
that
KIF­
230
in
the
diets
of
female
rats
at
10,000
ppm
does
cause
increased
absolute
and
relative
liver
weights
and
increased
aromatase
activity
in
the
liver,
but
it
does
not
appreciably
alter
aromatase
levels
in
organs
other
than
the
liver.
Results
were
consistent
with
enzyme
induction,
which
could
result
in
upregulation
of
hepatic
aromatase,
but
this
increase
was
insufficient
to
increase
circulating
E2
levels
in
this
study.
The
lack
of
increased
aromatase
activity
in
the
ovary
and
uterus
suggested
little
direct
participation
of
E2
in
the
development
of
uterine
cancer
by
KIF­
230.

Uterotrophic
Effects
in
Ovariectomized
Rats
45869714
(
2001)
Unacceptable/
Nonguideline
Six
ovariectomized
rats
per
dose
were
exposed
to
KIF­
230
daily
by
gavage
for
14
days
at
doses
of
0,
10,
100,
or
1000
mg/
kg/
day.
A
positive
control
group
was
dosed
with
0.001
No
treatment­
related
effects
were
observed
on
mortality,
clinical
signs,
body
weight,
overall
body
weight
gains,
absolute
or
relative
uterine
weights,
microscopic
uterine
pathology,
or
uterine
cellular
proliferation
as
determined
by
BrdU
incorporation.

The
lack
of
a
uterotropic
response
in
the
positive
controls
makes
it
impossible
to
determine
if
the
lack
of
response
in
the
test­
compound­
treated
animals
is
a
true
response
or
is
due
to
an
error
in
experimental
design.
Page
25
of
87
Guideline
No./
Study
Type
MRID
No.
(
year)/
Classification
/
Doses
Results
mg/
kg/
day
E2.

4.2
FQPA
Hazard
Considerations
4.2.1
Adequacy
of
the
Toxicity
Database
The
toxicology
database
for
benthiavalicarb
is
complete
and
adequate
for
FQPA
assessment.
Acceptable/
guideline
studies
for
developmental
toxicity
in
rats
and
rabbits
and
a
2­
generation
reproduction
study
in
rats
were
available
for
consideration
during
endpoint
selection.

4.2.2
Evidence
of
Neurotoxicity
Neurotoxicity
was
not
observed
in
either
the
acute
or
subchronic
neurotoxicity
studies
up
to
the
limit
dose
of
2000
mg/
kg/
day.
Furthermore,
no
evidence
of
neurotoxicity
was
seen
throughout
the
entire
toxicology
database.

4.2.3
Developmental
Toxicity
Studies
Developmental
effects
were
not
seen
in
either
the
developmental
rat
or
rabbit
studies.
There
were
no
effects
on
survival,
body
weights,
incidences
of
abortions,
premature
deliveries,
or
litter
resorptions.
There
were
no
effects
on
external,
visceral,
or
skeletal
malformations,
variations,
or
retardations.

The
maternal
NOAEL
(
100
mg/
kg/
day)
in
rats
was
based
upon
liver
effects
(
increased
absolute
and
relative
liver
weights
and
increased
incidence
of
enlarged
liver)
observed
at
the
LOAEL
(
1000
mg/
kg/
day).
Microscopic
examination
of
the
livers
was
not
performed
in
the
developmental
study;
however,
when
compared
to
the
liver
lesions
observed
in
the
subchronic
rat
study,
the
reviewer
noted
that
the
effects
on
the
liver
were
similar
to
the
liver
changes
in
other
studies
and
were
considered
adverse.

The
maternal
NOAEL
(
20
mg/
kg/
day)
in
the
rabbit
developmental
study
was
based
upon
increased
absolute
(
10%)
and
relative
(
11%)
liver
weights
at
the
LOAEL
of
40
mg/
kg/
day.

4.2.4
Reproductive
Toxicity
Study
No
adverse
effects
(
maternal,
reproductive,
or
developmental)
were
seen
in
the
2­
generation
reproduction
study
in
rats.
Increased
liver
weights,
slight
hepatocyte
hypertrophy,
and
two
incidences
of
enlarged
liver
were
observed
in
the
high
dose
group.
However,
in
the
absence
of
other
corroborating
pathological
indications
of
toxicity,
the
effect
on
liver
size
was
regarded
as
an
adaptive
response.

4.2.5
Additional
Information
from
Literature
Sources
Page
26
of
87
An
open
literature
review
for
benthiavalicarb
has
not
been
conducted.
However,
benthiavalicarb
is
registered
for
use
in
Europe,
and
the
European
monograph
is
available.
The
monograph
was
consulted
and
used
as
supplementary
information.

4.2.6
Pre­
and/
or
Post­
natal
Toxicity
Developmental
studies
in
two
species
(
both
rat
and
rabbit)
and
a
2­
generation
reproduction
study
in
rats
did
not
show
any
evidence
of
developmental
or
reproductive
toxicity.

4.2.6.1
Determination
of
Susceptibility
The
toxicology
database
is
complete
and
adequate
to
assess
questions
of
increased
qualitative
and/
or
quantitative
susceptibility
in
infants
and
children.
Evidence
suggests
that
there
is
no
concern
for
fetuses
exposed
to
benthiavalicarb
in
utero
or
post­
natally.
Maternal
toxicity
is
observed
in
both
the
rat
and
rabbit
developmental
studies
in
the
absence
of
developmental
effects
to
the
fetuses.
No
adverse
effects
were
observed
in
the
2­
generation
reproduction
study.

4.2.6.2
Degree
of
Concern
Analysis
and
Residual
Uncertainties
for
Pre
and/
or
Post­
natal
Susceptibility
There
is
no
concern
for
increased
risk
to
developing
fetuses
resulting
from
exposure
in
utero
and/
or
post­
natally.
Therefore,
the
FQPA
factor
has
been
reduced
to
1x.

4.3
Recommendation
for
a
Developmental
Neurotoxicity
Study
The
registrant
submitted
acceptable
acute
and
subchronic
neurotoxicity
studies.
There
was
no
evidence
of
neurotoxicity
up
to
the
limit
dose
of
2000
mg/
kg/
day.
Because
of
the
absence
of
neurotoxicity
throughout
the
entire
toxicology
database
and
the
absence
of
adverse
developmental
and
reproductive
effects,
a
developmental
neurotoxicity
study
is
not
necessary
at
this
time.
Page
27
of
87
4.4
Hazard
Identification
and
Toxicity
Endpoint
Selection
4.4.1
Acute
Reference
Dose
(
aRfD)
­
All
Populations
Because
of
the
absence
of
toxic
effects
attributable
to
a
single
dose,
no
acute
endpoint
has
been
selected
for
risk
assessment.

4.4.2
Chronic
Reference
Dose
(
cRfD)

The
chronic
reference
dose
was
selected
from
the
chronic
rat
study
based
upon
nephrotoxicity
and
hepatotoxicity.
In
particular,
hepatotoxicity
is
prevalent
throughout
the
entire
toxicology
database
and
is
the
main
effect
of
concern.
Furthermore,
the
NOAEL
of
9.9
mg/
kg/
day
is
protective
of
tumor
effects
seen
in
both
rats
and
mice
at
doses
greater
than
250
mg/
kg/
day.
The
standard
10x
intraspecies
variability
and
10x
interspecies
extrapolation
safety
factors
apply.
Therefore,
the
cRfD
=
0.099
mg/
kg/
day.

4.4.3
Non­
Dietary
Exposure
for
Handlers
The
registrant
is
only
seeking
import
tolerances
for
benthiavalicarb.
As
a
result,
the
only
relevant
route
of
exposure
is
through
food.
Occupational
and
residential
exposures
do
not
need
to
be
assessed
at
this
time.

4.4.4
Margins
of
Exposure
The
level
of
concern
for
risk
assessment
is
margins
of
exposure
that
are
less
than
100,
based
on
the
standard
10x
interspecies
and
10x
intraspecies
extrapolation
factors.

4.4.5
Recommendation
for
Aggregate
Exposure
Risk
Assessments
The
registrant
is
seeking
import
tolerances
on
grapes
and
tomatoes.
Therefore,
the
risk
assessment
only
includes
dietary
exposure
to
benthiavalicarb
via
imported
grapes
and
tomatoes.

4.4.6
Classification
of
Carcinogenic
Potential
In
accordance
with
EPA's
Final
Guidelines
for
Carcinogen
Risk
Assessment
(
March,
2005),
the
CARC
classified
benthiavalicarb
as
"
likely
to
be
carcinogenic
to
humans,"
based
on
the
presence
of
malignant
uterine
tumors
in
rats,
liver
tumors
in
both
sexes
of
mice
(
with
some
supporting
evidence
found
in
liver
tumors
in
male
rats),
and
thyroid
follicular
cell
tumors
in
male
mice,
at
doses
adequate
to
assess
carcinogenicity.
Benthiavalicarb
is
not
mutagenic.

4.5
Special
FQPA
Safety
Factor
The
toxicology
database
is
complete
and
adequate
for
FQPA
analysis.
There
is
no
concern
for
developmental
and/
or
reproductive
effects,
increased
susceptibility,
or
neurotoxicity.
Therefore,
HED
has
reduced
the
FQPA
safety
factor
to
1x.
The
endpoints
selected
for
risk
assessment
are
based
on
the
effects
of
concern
(
hepatotoxicity),
and
are
protective
of
tumor
types
observed
in
the
chronic/
carcinogenicity
studies.
There
is
little
residual
uncertainty
with
regard
to
the
toxicology
Page
28
of
87
database.

Table
4.5.
Summary
of
Toxicological
Doses
and
Endpoints
for
Use
in
Human
Health
Risk
Assessments
Exposure
Scenario
Dose
Used
in
Risk
Assessment,
UF
Special
FQPA
SF*
and
Level
of
Concern
for
Risk
Assessment
Study
and
Toxicological
Effects
Acute
Dietary
ALL
populations
including
infants
and
children
when
applicable
NOAEL=
N/
S
UF
=
N/
A
Acute
RfD
=
N/
A
FQPA
SF
=
N/
A
None
of
the
study
endpoints
were
the
result
of
one
or
two
exposures
to
benthiavalicarb;
therefore
an
acute
endpoint
was
not
selected
for
dietary
exposure.

Chronic
Dietary
all
populations
NOAEL=
9.9
mg/
kg/
day
UF
=
100
Chronic
RfD
=
0.099
mg/
kg/
day
FQPA
SF
=
1X
cPAD
=
0.099
mg/
kg/
day
Chronic
Oral
Toxicity
in
Rats
LOAEL
=
249.6
mg/
kg/
day
based
on
nephrotoxicity
and
hepatotoxicity.

Cancer
(
oral,
dermal,
inhalation)
Likely
to
be
carcinogenic
to
humans
Q1*
=
6.2795x10­
2
Based
on
increases
in
male
mouse
liver
combined
adenomas
and/
or
carcinomas
and/
or
blastomas.

UF
=
uncertainty
factor,
FQPA
SF
=
Special
FQPA
safety
factor,
NOAEL
=
no
observed
adverse
effect
level,
LOAEL
=
lowest
observed
adverse
effect
level,
PAD
=
population
adjusted
dose
(
a
=
acute,
c
=
chronic)
RfD
=
reference
dose,
MOE
=
margin
of
exposure,
LOC
=
level
of
concern,
NA
=
Not
Applicable
*
Refer
to
Section
4.5
4.6
Endocrine
disruption
EPA
is
required
under
the
FFDCA,
as
amended
by
FQPA,
to
develop
a
screening
program
to
determine
whether
certain
substances
(
including
all
pesticide
active
and
other
ingredients)
"
may
have
an
effect
in
humans
that
is
similar
to
an
effect
produced
by
a
naturally
occurring
estrogen,
or
other
such
endocrine
effects
as
the
Administrator
may
designate."
Following
recommendations
of
its
Endocrine
Disruptor
and
Testing
Advisory
Committee
(
EDSTAC),
EPA
determined
that
there
was
a
scientific
basis
for
including,
as
part
of
the
program,
the
androgen
and
thyroid
hormone
systems,
in
addition
to
the
estrogen
hormone
system.
EPA
also
adopted
EDSTAC's
recommendation
that
the
Program
include
evaluations
of
potential
effects
in
wildlife.
For
pesticide
chemicals,
EPA
will
use
FIFRA
and,
to
the
extent
that
effects
in
wildlife
may
help
determine
whether
a
substance
may
have
an
effect
in
humans,
FFDCA
authority,
to
require
the
wildlife
evaluations.
As
the
science
develops
and
resources
allow,
screening
of
additional
hormone
Page
29
of
87
systems
may
be
added
to
the
Endocrine
Disruptor
Screening
Program
(
EDSP).
In
the
available
toxicity
studies
on
benthiavalicarb,
there
was
no
estrogen,
androgen,
and/
or
thyroid
mediated
toxicity.
When
additional
appropriate
screening
and/
or
testing
protocols
being
considered
under
the
Agency's
EDSP
have
been
developed,
benthiavalicarb
may
be
subjected
to
further
screening
and/
or
testing
to
better
characterize
effects
related
to
endocrine
disruption.

5.0
Public
Health
Data
As
benthiavalicarb
will
not
be
used
in
the
U.
S.,
public
health
data
are
not
relevant
to
this
tolerance
petition.

6.0
Exposure
Characterization/
Assessment
6.1
Dietary
Exposure/
Risk
Pathway
6.1.1
Residue
Profile
A
discussion
of
benthiavalicarb
residue
chemistry
is
given
in
a
memorandum
prepared
for
these
tolerance
requests
(
D322934,
D.
Dotson,
3/
2/
2006).
The
nature
of
the
residue
has
been
adequately
delineated
for
the
proposed
uses
based
on
the
acceptable
grape
and
tomato
metabolism
studies.
The
available
potato
metabolism
study
is
also
acceptable
pending
submission
of
supporting
storage
stability
data
(
21
months).
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
residue
in
fruit
and
foliage
(
55­
95%
TRR).
Parent
was
also
the
principal
14C­
residue
in
potato
foliage
(
88­
90%
TRR),
but
other
minor
metabolites
were
identified.
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
Secondary
metabolism
occurs
via
conjugation
of
sugars
with
hydroxy
groups.
Chiral
analysis
of
potato
foliage
extracts
also
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).
Based
on
the
results
of
the
metabolism
studies,
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.
The
nature
of
the
residue
in
livestock
was
not
determined,
and
is
not
required,
as
there
are
no
livestock
feed
items
associated
with
grapes
and
tomatoes.

An
adequate
GC/
NPD
Method
(
RCC
No.
665943)
is
available
for
collecting
data
on
residues
of
both
the
R­
L
and
S­
L
isomers
of
benthiavalicarb
in/
on
grape
and
tomato
matrices.
For
this
method,
residues
are
extracted
with
acetone,
partitioned
into
hexane,
purified
using
an
amino
SPE
cartridge,
and
determined
by
GC/
NPD.
The
lower
limit
of
method
validation
(
LLMV)
is
0.01
ppm
for
each
analyte
in
grape
and
tomato
matrices.
The
reported
limit
of
detection
(
LOD)
in
grapes
is
0.001
ppm.
The
reported
LOD
in
grape
processed
commodities,
tomatoes,
and
tomato
processed
commodities
is
0.002
ppm.
Although
the
submitted
reports
state
that
the
LOQ
was
0.01
ppm,
an
inspection
of
the
chromatograms
indicates
that
the
LOQ
would
probably
be
in
the
range
of
0.002­
0.003
ppm
for
the
various
grape
and
tomato
commodities.
In
many
cases
residue
values
were
reported
as
being
between
0.001
and
0.01
ppm
(
i.
e.,
actual
values
were
reported
as
0.002
ppm,
0.003
ppm,
etc.).
The
reporting
of
such
values
is
further
evidence
that
the
LOQ
is
Page
30
of
87
indeed
below
0.01
ppm.
The
method
was
adequately
validated
by
the
petitioner,
and
is
being
proposed
as
the
tolerance
enforcement
method.
The
method
was
successfully
validated
in
an
independent
laboratory
method
validation
(
ILV)
trial,
and
radiovalidation
data
are
not
required,
given
the
similarity
between
the
extraction
procedures
in
the
method
and
the
metabolism
studies.
However,
because
the
method
is
not
highly
specific,
the
petitioner
should
either
develop
a
confirmatory
method
for
the
analytes
of
interest
or
conduct
an
interference
study.
The
method
was
forwarded
to
BEAD/
ACB
for
a
petition
method
validation
(
PMV)
trial.
The
method
was
forwarded
to
the
Analytical
Chemistry
Branch
(
ACB)
of
the
Biological
and
Economic
Assessment
Division
(
BEAD)
for
a
petition
method
validation
(
PMV)
trial.
ACB
concluded
that
the
method
does
not
need
to
be
laboratory­
validated
by
EPA,
and
appears
to
be
suitable
for
tolerance
enforcement.

Adequate
storage
stability
data
are
available
indicating
that
benthiavalicarb
is
stable
under
frozen
( ­
18
°
C)
conditions
for
at
least
13
months
in
grapes
and
8
months
in
tomatoes.
These
data
support
the
maximum
storage
intervals
(
7­
8
months)
from
the
grape
and
tomato
field
trials.

As
there
are
no
significant
animal
feed
items
associated
with
the
proposed
use
on
grapes
and
tomatoes,
livestock
feeding
studies
are
not
required
for
this
petition.
Tolerances
do
not
need
to
be
established
on
animal
commodities.

The
available
field
trial
data
are
adequate
and
will
support
the
proposed
use
patterns
on
grapes
and
tomatoes.
The
number
and
geographic
distribution
of
the
field
trials
are
adequate.
For
grapes,
a
total
of
34
field
tests
were
conducted
throughout
the
EU,
in
which
grapes
were
treated
at
1x
the
maximum
proposed
rate
and
harvested
at
the
minimum
PHI
of
approximately
28
days.
Maximum
combined
residues
were
<
0.27
ppm
in/
on
grapes,
and
the
HAFT
residue
value
was
0.22
ppm.
For
tomatoes,
12
tests
were
conducted
at
1x
the
maximum
rate
throughout
the
EU
on
both
greenhouse­
and
outdoor­
field­
grown
tomatoes.
Nine
of
these
12
field
trials
were
performed
on
greenhouse­
grown
tomatoes
and
three
were
performed
on
outdoor­
grown­
tomatoes.
Maximum
combined
residues
were
0.27
ppm
in/
on
greenhouse
tomatoes
3
days
after
treatment
(
DAT)
and
<
0.015
ppm
in/
on
field­
grown
tomatoes
(
7
DAT).
The
recommended
tolerances
for
grapes
and
tomatoes
were
generated
using
HED's
statistical
tolerance
generator
for
NAFTAharmonized
tolerances.
The
recommended
tolerance
for
grapes
is
0.25
ppm,
even
though
the
highest
field
trial
value
is
0.27
ppm.
The
recommended
tolerance
for
tomatoes
is
0.45
ppm.

Overall,
22
grape
field
trials
were
performed
in
which
samples
were
taken
at
1­
week
intervals.
The
results
of
these
decline
studies
can
be
found
in
Table
A.
6.
The
samples
were
taken
at
0,
7,
14,
21,
and
28
DAT.
In
four
of
those
trials,
samples
were
not
taken
at
28
days.
Average
residues
declined
as
follows:
0.088
ppm,
0.070
ppm,
0.055
ppm,
0.049
ppm,
and
0.046
ppm
at
0,
7,
14,
21,
and
28
days,
respectively.
Residues
declined
by
0.018
ppm,
0.015
ppm,
0.06
ppm,
and
0.03
ppm
over
the
four
intervals.
In
effect,
residues
declined
with
the
greatest
decreases
occurring
in
the
early
stages
of
the
field
trials.

The
available
processing
data
are
adequate,
pending
submission
of
supporting
storage
stability
data
for
processed
matrices.
The
average
processing
factors
for
the
combined
residues
were
3.7x
in
raisins,
0.47x
in
juice,
and
0.97x
in
wine.
Based
on
the
3.7x
processing
factor
for
raisins
and
the
HAFT
residue
value
of
0.22
ppm
for
grapes,
the
maximum
expected
residues
in
raisins
would
be
0.81
ppm,
which
is
above
the
recommended
grape
tolerance.
Therefore,
a
separate
tolerance
is
Page
31
of
87
required
for
raisins.
HED
recommends
in
favor
of
the
proposed
tolerance
of
1.0
ppm
for
grape,
raisin.
Storage
stability
data
were
not
submitted
for
processed
grape
and
tomato
commodities.
These
data
are
not
required
for
the
current
tolerance
requests
because
acceptable
storage
stability
data
were
submitted
for
grapes
and
tomatoes.
Residues
were
stable
in
grapes
for
13
months
and
in
tomatoes
for
8
months.
In
the
tomato
storage
stability
studies,
tomatoes
were
homogenized
prior
to
storage.
Homogenized
tomatoes
are
comparable
in
both
composition
and
texture
to
processed
tomato
commodities.
The
matrix
is
an
acidic
one
which
would
very
likely
maximize
residue
decay.
As
a
result,
the
tomato
data
are
considered
adequate
to
demonstrate
storage
stability
in
processed
grape
commodities
as
well.

In
the
tomato
processing
studies,
the
average
processing
factors
from
tests
having
quantifiable
residues
were
0.49x
in
juice,
1.2x
in
puree,
and
1.4x
in
ketchup.
As
residue
data
were
not
available
for
paste,
the
5.5x
theoretical
concentration
factor
based
on
loss
of
water
was
used
for
paste.
The
HAFT
residue
value
of
0.015
ppm
from
the
tests
on
field­
grown
tomatoes
was
used
to
assess
the
need
for
tolerances
on
processed
tomato
fractions
as
greenhouse
tomatoes
are
not
utilized
for
processing.
Based
on
the
above
processing
factors
and
the
highest
field
trial
value
of
0.015
ppm,
the
maximum
expected
residues
in
puree,
ketchup,
juice,
and
paste
would
be
0.018,
0.021,
0.0074
ppm,
and
0.083
ppm,
respectively,
which
are
all
below
the
recommended
tomato
tolerance
of
0.45
ppm.
Therefore,
separate
tolerances
are
not
required
for
processed
tomato
fractions.

As
the
proposed
uses
are
for
grape
and
tomatoes
grown
in
the
EU,
confined
and
field
rotational
crops
studies
are
not
required
for
the
current
petition.

6.1.2
Acute
and
Chronic
Dietary
Exposure
and
Risk
A
chronic
dietary
risk
assessment
was
conducted
using
the
Dietary
Exposure
Evaluation
Model
(
DEEM­
FCID,
Version
2.03).
The
dietary
exposure
analysis
and
its
results
are
discussed
in
Memo,
D322935,
D.
Dotson,
3/
2/
2006.
The
DEEM­
FCID
Model
uses
food
consumption
data
from
the
USDA's
Continuing
Surveys
of
Food
Intakes
by
Individuals
(
CSFII)
from
1994­
1996
and
a
supplemental
children's
survey
conducted
in
1998.
The
result
of
the
chronic
dietary
exposure
assessment
for
the
general
U.
S.
population
was
used
along
with
the
cancer
Q1*
value
to
determine
the
cancer
risk
to
the
U.
S.
population.

In
order
to
determine
the
percent
crop
imported
for
individual
grape
and
tomato
commodities,
data
are
needed
for
the
following
three
quantities:
(
1)
domestic
production,
(
2)
the
quantity
exported,
and
(
3)
the
quantity
imported.
These
data
were
not
available
for
all
commodities
in
the
analysis.
For
grapes,
these
data
were
available
for
grape
juice
and
raisins
only.
Percent
imported
data
for
grapes
and
wine
are
directly
available
from
the
USDA
Economic
Research
Service
(
ERS),
however.
For
unprocessed
tomatoes,
the
three
quantities
listed
above
are
available.
Data
were
not
used
for
the
individual
processed
tomato
commodities
because
a
significant
amount
of
processing
occurs
after
importation.
Data
are
available
for
domestic
tomato
production
utilized
for
processing
and
processed
tomato
imports.
The
import
data
include
canned
tomatoes,
a
significant
fraction
of
which
is
utilized
for
processing
in
the
U.
S.
Dividing
the
imported
by
the
total
yields
a
value
of
5.84%
imported.
The
USDA
ERS,
Vegetable
and
Melons
Outlook
(
VGS­
308)
of
April
21,
2005
states
the
following:
"
According
to
ERS
estimates,
nearly
6
percent
of
the
Page
32
of
87
tomato
products
consumed
by
Americans
in
2004
were
imported."
The
value
of
5.84%
was
used
for
paste,
puree,
and
juice.
A
value
of
100%
was
used
for
dried
tomatoes.

The
data
for
grape
commodities
were
provided
by
BEAD
(
electronic
communication,
D.
Donaldson,
2/
1/
2006).
USDA
ERS
data
for
2002
and
2003
were
used.
USDA
ERS
data
from
2004
were
used
for
tomatoes.
The
following
percent
crop
imported
values
were
used
in
the
analysis:
tomatoes
(
36.3%),
processed
tomato
commodities
(
5.84%),
grapes
(
45%),
grape
juice
(
43.9%),
wine
(
23%),
and
raisins
(
10.7%).

Chronic
Dietary
Exposure
Results
and
Characterization
The
chronic
dietary
exposure
analysis
is
based
on
average
field
trial
values
and
estimates
of
percent
crop
imported.
Empirical
processing
factors
were
used
for
most
processed
commodities.
When
empirical
factors
were
not
available,
DEEM
(
Version
7.81)
default
processing
factors
were
used.
The
general
U.
S.
population
and
all
population
subgroups
had
risk
estimates
that
were
below
HED's
level
of
concern
(
i.
e.,
100%
of
the
cPAD).
In
fact,
the
U.
S.
population
and
all
population
subgroups
had
risk
estimates
that
were
less
than
1%
of
the
cPAD.
For
the
chronic
assessment,
all
population
subgroups
would
have
had
risk
estimates
that
were
below
HED's
level
of
concern
if
no
refinements
had
been
made
to
the
analysis,
i.
e.,
if
tolerance
level
residues
and
100%
crop
imported
assumptions
had
been
used.
However,
it
was
necessary
to
refine
the
inputs
for
the
cancer
assessment.
For
simplicity,
the
same
refinements
were
used
for
the
chronic
analysis.

Cancer
Dietary
Exposure
Results
and
Characterization
As
stated
above,
the
CARC
determined
that
benthiavalicarb
is
likely
to
be
carcinogenic
to
humans
and
assigned
it
a
Q1*
value
of
0.063
(
mg/
kg/
day)­
1.
In
an
unrefined
cancer
dietary
exposure
analysis,
the
estimated
cancer
risk
to
the
general
U.
S.
population
exceeds
HED's
level
of
concern
(
i.
e.,
1.0
x
10­
6
or
one
in
one
million).
As
a
result,
refinements
were
made
to
the
cancer
dietary
exposure
analysis.
The
refinements
made
are
the
same
as
those
made
for
the
chronic
analysis
discussed
above:
the
use
of
average
field
trial
values
and
estimates
of
percent
crop
imported.
With
the
use
of
these
refinements,
the
estimated
lifetime
cancer
risk
to
the
U.
S.
population
is
1.6
x
10­
6.
Refinements
were
made
to
the
analysis;
however,
it
is
still
considered
to
be
conservative.
First,
average
field
trial
values
from
field
trials
almost
always
exceed
the
expected
residue
levels
found
on
crops
at
the
time
of
consumption.
When
field
trials
are
performed,
the
maximum
allowable
application
rate
is
used
and
crops
are
harvested
at
the
minimum
PHI.
Samples
are
stored
frozen
until
analysis
to
ensure
minimal
degradation
of
residues.
In
actual
practice,
however,
growers
will
not
usually
use
the
maximum
application
rates
for
economic
reasons.
In
addition,
most
crops
are
not
harvested
and
immediately
stored
frozen.
From
previous
dietary
exposure
analyses,
HED
has
noted
that
monitoring
data
are
often
one
to
two
orders
of
magnitude
lower
than
field
trial
data.
The
second
reason
this
analysis
is
considered
to
be
conservative
is
that
the
registrant
is
planning
on
marketing
benthiavalicarb
in
Europe
only.
In
the
dietary
analysis,
percent
crop
imported
estimates
were
used
for
all
imports,
not
just
those
from
Europe.
The
third
reason
this
analysis
is
considered
to
be
conservative
is
that
HED
assumed
that
100%
of
the
imported
crop
will
be
treated.
The
actual
percent
crop
treated
will
very
likely
be
considerably
lower
than
that.
For
these
reasons,
HED
is
confident
that
this
analysis
overestimates
risk
to
the
general
U.
S.
population.
The
cancer
risk
to
the
general
U.
S.
population
does
not
exceed
HED's
Page
33
of
87
level
of
concern.

Table
6.1.
Summary
of
Dietary
Exposure
and
Risk
for
Benthiavalicarb
DEEM:
Chronic
Analysis
Population
Subgroup
Acute
Analysis
Dietary
Exposure
(
mg/
kg/
day)
%
cPAD
Cancer
Analysis
General
U.
S.
Population
0.000026
<
0.1
1.6
x
10­
6
All
Infants
(<
1
year
old)
Not
Applicable:
No
Acute
Dietary
Endpoint
0.000017
<
0.1
Children
1­
2
years
old
0.000071
0.1
Children
3­
5
years
old
0.000053
0.1
N/
A:
Cancer
risk
is
determined
for
the
general
U.
S.
population
only
Children
6­
12
years
old
0.000030
<
0.1
Youth
13­
19
years
old
0.000019
<
0.1
Adults
20­
49
years
old
0.000022
<
0.1
Adults
50+
years
old
0.000023
<
0.1
Females
13­
49
years
old
0.000021
<
0.1
The
commodities
that
made
the
greatest
contribution
to
the
risk
estimates
are
tomatoes,
grape
juice,
grapes,
wine,
and
raisins.
For
the
general
U.
S.
population,
these
commodities
contribute
a
total
of
98%
of
the
entire
cancer
risk.
The
individual
contributions
are:
tomatoes
(
61%),
grape
juice
(
14%),
grapes
(
12%),
wine
(
8%),
and
raisins
(
3%).

HED
is
confident
that
this
analysis
does
not
underestimate
risk
to
the
general
U.
S.
population.
HED
is
also
confident
that
average
field
trial
values
and
percent
crop
imported
values
used
in
the
assessment
will
not
be
exceeded
in
the
future.
For
the
reasons
stated
above,
the
field
trial
values
considerably
overestimate
residues
found
on
food
at
the
time
of
consumption.
It
is
unlikely
that
residue
levels
in
imported
grape
and
tomato
commodities
at
the
time
of
consumption
would
ever
exceed
the
average
field
trial
values.
As
far
as
percent
crop
imported
values
being
exceeded
in
the
future,
this
situation
is
unlikely
as
well.
As
stated
above,
the
conservative
assumptions
were
made
that
100%
of
the
grape
and
tomato
commodities
imported
from
all
over
the
world
would
be
treated.
It's
very
unlikely
that
the
quantities
of
imports
actually
treated
will
ever
increase
to
those
levels.
Data
for
grape
commodities
were
provided
by
BEAD.
The
economist
who
provided
the
data
stated
that
it
was
unlikely
the
estimates
of
import
shares
would
be
exceeded
in
the
near
future.
The
data
for
grape
commodities
provided
to
HED
were
five
year
averages,
and
were
compared
to
the
annual
data
points
of
each
data
source,
when
available.
Import
share
data
do
vary
from
year
to
year,
but
crop
production
patterns
and
consumption
and
preferences
are
more
stable
and
change
more
slowly.
For
tomatoes,
the
worldwide
imports
changed
as
follows
from
1998
to
2004:
37%,
31%,
30%,
33%,
32%,
37%,
and
36%.
Clearly,
no
increase
occurred
over
that
time
period.
For
processed
tomato
commodities,
the
worldwide
imports
changed
as
follows
from
2000
to
2004:
2.9%,
5.6%,
7.1%,
5.4%,
and
5.8%.
Between
2001
and
2004
there
was
no
significant
increase
in
processed
tomato
commodity
imports.
Page
34
of
87
6.2
Water
Exposure/
Risk
Pathway
The
tolerances
being
granted
in
conjunction
with
these
requests
are
for
imported
commodities
only.
As
a
result,
residues
in
water
are
not
being
considered
in
this
assessment.

6.3
Residential
(
Non­
Occupational)
Exposure/
Risk
Pathway
There
are
no
residential
uses
for
benthiavalicarb
and
the
food
uses
are
for
imported
commodities.
As
a
result,
residential
uses
(
including
home
uses,
recreational
uses,
spray
drift,
etc.)
are
not
relevant
to
this
assessment.

7.0
Aggregate
Risk
Assessments
and
Risk
Characterization
7.1
Acute
Aggregate
Risk
None
of
the
toxicity
study
endpoints
were
the
result
of
a
single
oral
exposure
to
benthiavalicarb.
As
a
result,
no
acute
dietary
endpoint
was
identified,
and
an
acute
risk
assessment
is
not
needed.

7.2
Short­,
Intermediate­,
and
Long­
Term
Aggregate
Risk
As
the
current
petition
is
for
import
tolerances
on
grapes
and
tomatoes
only,
there
are
no
residential
uses,
and
residues
in
drinking
water
are
not
of
concern.
As
a
result,
short­
and
intermediate­
term
aggregate
risk
assessments
are
not
needed.
The
chronic
aggregate
risk
assessment
is
equivalent
to
the
chronic
dietary
exposure
analysis.
See
Section
6.1.2.
for
a
discussion
of
the
dietary
exposure
analysis.
The
general
U.
S.
population
and
all
population
subgroups
had
risk
estimates
that
were
well
below
HED's
level
of
concern.

7.3
Cancer
Risk
The
cancer
aggregate
risk
assessment
is
equivalent
to
the
cancer
dietary
exposure
analysis.
See
Section
6.1.2.,
above
for
a
discussion
of
this
analysis.
With
the
use
of
some
refinements,
the
lifetime
cancer
risk
to
the
U.
S.
population
is
1.6
x
10­
6.
HED
believes
this
risk
estimate
is
still
an
overestimate,
however,
and
that
the
cancer
risk
is
not
of
concern.

8.0
Cumulative
Risk
Characterization/
Assessment
Unlike
other
pesticides
for
which
EPA
has
followed
a
cumulative
risk
approach
based
on
a
common
mechanism
of
toxicity,
EPA
has
not
made
a
common
mechanism
of
toxicity
finding
as
to
benthiavalicarb
and
any
other
substances,
and
benthiavalicarb
does
not
appear
to
produce
a
toxic
metabolite
produced
by
other
substances.
For
the
purposes
of
this
tolerance
action,
therefore,
EPA
has
not
assumed
that
benthiavalicarb
has
a
common
mechanism
of
toxicity
with
other
substances.
Although
benthiavalicarb
is
a
carbamate
compound,
it
is
not
a
member
of
the
class
of
insecticides
known
as
the
N­
methyl
carbamates
for
which
the
Agency
is
presently
conducting
a
cumulative
risk
assessment.
The
substituents
on
the
benthiavalicarb
nitrogen
atom
are
much
larger
than
the
methyl
group
in
the
insecticides.
While
the
N­
methyl
carbamates
are
Page
35
of
87
neurotoxicants
based
on
their
ability
to
inhibit
the
enzyme
cholinesterase,
there
is
no
evidence
of
neurotoxicity
or
neuropathology
in
the
hazard
database
for
benthiavalicarb.
Benthiavalicarb
is
also
not
a
member
of
the
thiocarbamate
herbicides
or
dithiocarbamate
fungicides.

For
information
regarding
EPA's
efforts
to
determine
which
chemicals
have
a
common
mechanism
of
toxicity
and
to
evaluate
the
cumulative
effects
of
such
chemicals,
see
the
policy
statements
released
by
EPA's
Office
of
Pesticide
Programs
concerning
common
mechanism
determinations
and
procedures
for
cumulating
effects
from
substances
found
to
have
a
common
mechanism
on
EPA's
website
at
http://
www.
epa.
gov/
pesticides/
cumulative/.

9.0
Occupational
Exposure/
Risk
Pathway
As
benthiavalicarb
will
be
used
outside
the
U.
S.
to
treat
grapes
and
tomatoes,
there
will
be
no
occupational
risk
to
U.
S.
workers.
As
a
result,
an
occupational
assessment
is
not
relevant
to
this
tolerance
petition.

10.0
Data
Needs
and
Label
Requirements
10.1
Toxicology
The
toxicology
database
is
complete
and
adequate
for
risk
assessment.
There
are
no
further
toxicology
requirements
at
this
time.

10.2
Residue
Chemistry
No
major
deficiencies
were
noted
in
the
subject
petition
that
would
preclude
establishing
tolerances
for
benthiavalicarb
on
grape
and
tomato
commodities.
However,
before
any
further
tolerances
are
granted,
the
petitioner
should
either
develop
a
confirmatory
analytical
method
for
the
analytes
of
interest
or
conduct
an
interference
study.

There
are
no
residue
chemistry
issues
that
would
preclude
establishing
tolerances
of
0.25
ppm
for
grape,
0.45
ppm
for
tomato,
and
1.0
ppm
for
grape,
raisin.
The
recommended
tolerances
for
grape
and
tomato
were
generated
using
HED's
statistical
tolerance
generator
for
NAFTAharmonized
tolerances.
The
residues
to
be
included
in
the
tolerance
expression
are
the
combined
R­
L
and
S­
L
isomers
of
benthiavalicarb.
The
petitioner
has
recommended
that
only
the
R­
L
isomer
be
included
in
the
tolerance
expression
because
the
S­
L
isomer
is
a
minor
impurity
in
the
commercial
formulations
and
the
potato
metabolism
study
indicates
that
the
active
R­
L
isomer
is
not
converted
to
the
S­
L
isomer.
In
addition,
the
residue
definition
proposed
for
the
EU
MRLs
includes
only
the
R­
L
isomer.
The
registrant
has
proposed
EU
MRLs
of
0.2
ppm
for
grapes
and
0.3
ppm
for
tomatoes.
It's
possible
that
if
EU
MRLs
are
established,
the
residue
definition
might
be
benthiavalicarb­
isopropyl
only,
and
no
distinction
will
be
made
as
to
the
isomer.
HED
concludes
that
the
S­
L
isomer
should
be
included
in
the
tolerance
expression,
however.
The
results
of
the
field
trials
indicate
that
the
residue
levels
of
the
S­
L
isomer
are
often
non­
negligible
compared
to
the
levels
of
the
R­
L
isomer.
In
the
grape
field
trials
several
samples
had
S­
L
isomer
levels
that
were
10­
20%
of
the
R­
L
isomer
levels.
A
small
number
of
samples
had
S­
L:
R­
L
ratios
that
exceeded
20%.
In
the
tomato
field
trials,
the
S­
L
residue
levels
were
generally
non
Page
36
of
87
detectable.
In
a
few
cases,
however,
the
S­
L
isomer
was
present
at
6­
7%
of
the
level
of
the
R­
L
isomer.
The
technical
material
contains
93%
R­
L
isomer
and
3.4%
S­
L
isomer.
The
analytical
method
determines
both
isomers
separately.

10.3
Occupational
and
Residential
Exposure
There
are
no
data
needs
or
label
requirements.

11.0
Summary
of
Proposed
and
Recommended
Tolerances
Table
11.1
gives
the
proposed
tolerances,
the
recommended
tolerances,
and
the
correct
commodity
definitions
for
the
commodities
associated
with
this
risk
assessment.
The
registrant
did
not
propose
a
tolerance
for
wine
and
none
is
needed.
Residues
do
not
concentrate
in
wine;
therefore,
the
0.25
ppm
tolerance
recommended
for
grape
is
adequate
to
cover
wine.

Table
11.1
Tolerance
Summary
for
Benthiavalicarb­
Isopropyl
(
PP#
3E6545)

Commodity
Proposed
Tolerance
(
ppm)
Recommended
Tolerance
(
ppm)
Correct
commodity
definition
and
comments
Grapes
0.5
0.25
Grape
The
available
data
are
adequate.
Maximum
residues
in/
on
grapes
were
0.27
ppm.

Raisins
1.0
1.0
Grape,
Raisin
The
available
data
are
adequate.
Based
on
an
average
processing
factor
of
3.7x
for
raisins
and
the
HAFT
residue
value
in
grapes
of
0.22
ppm,
maximum
residues
in
raisins
would
be
0.81
ppm.

Tomatoes
0.5
0.45
Tomato
The
available
data
are
adequate.
Maximum
residues
were
0.26
ppm
in/
on
greenhouse
grown
tomatoes
and
<
0.015
ppm
in/
on
field
grown
tomatoes.

Tomato
Paste
1.5
None
Tomato,
Paste
The
available
data
are
adequate.
Average
processing
factors
were
1.2x
and
1.4x
for
tomato
puree
and
ketchup,
respectively,
and
were
estimated
to
be
5.5x
for
paste
(
the
maximum
theoretical
concentration
factor).
Based
on
the
highest
field
trial
residue
value
of
0.015
ppm
for
field­
grown
tomatoes,
the
maximum
expected
residues
in
tomato
paste
would
be
0.083
ppm.
Page
37
of
87
References:

Benthiavalicarb­
Isopropyl:
Report
of
the
Cancer
Assessment
Review
Committee,
TXR
No.
0053803,
J.
Kidwell,
10/
18/
2005
Benthiavalicarb:
Quantitative
Risk
Assessment
(
Q1*)
Based
on
Fischer
F­
344/
DuCrj
Rat
and
SLC:
B6C3F1(
C57BL/
6xC3H)
SPF
Mouse
Dietary
Studies
With
¾
'
s
Interspecies
Scaling
Factor,
TXR
No.
0053755,
Lori
Brunsman,
9/
26/
2005
Benthiavalicarb­
Isopropyl.
PP#
3E6545.
Request
for
Tolerances
on
Imported
Grapes
and
Tomatoes.
Summary
of
Analytical
Chemistry
and
Residue
Data,
D322934,
D.
Dotson,
3/
2/
2006.

Benthiavalicarb­
Isopropyl:
Chronic
and
Cancer
Dietary
Exposure
Assessments
for
the
Section
3
Registration
Action
on
Imported
Grapes
and
Tomatoes,
D322935,
D.
Dotson,
3/
2/
2006.
Page
38
of
87
Appendices
1.0
TOXICOLOGY
DATA
REQUIREMENTS
The
requirements
(
40
CFR
158.340)
for
import
tolerance
on
food
crops
for
benthiavalicarb
are
in
Table
1.
Use
of
the
new
guideline
numbers
does
not
imply
that
the
new
(
1998)
guideline
protocols
were
used.

Table
A.
1.
Toxicology
Data
Requirements
Technical
Test
Required
Satisfied
870.1100
Acute
Oral
Toxicity
870.1200
Acute
Dermal
Toxicity
870.1300
Acute
Inhalation
Toxicity
870.2400
Primary
Eye
Irritation
870.2500
Primary
Dermal
Irritation
870.2600
Dermal
Sensitization
yes
no
no
no
no
no
yes
­
­
­
­
­

870.3100
Oral
Subchronic
(
rodent)
870.3150
Oral
Subchronic
(
nonrodent)
870.3200
21­
Day
Dermal
870.3250
90­
Day
Dermal
870.3465
90­
Day
Inhalation
yes
yes
no
no
no
yes
yes
­
­
­

870.3700a
Developmental
Toxicity
(
rodent)
870.3700b
Developmental
Toxicity
(
nonrodent)
870.3800
Reproduction
yes
yes
yes
yes
yes
yes
870.4100a
Chronic
Toxicity
(
rodent)
870.4100b
Chronic
Toxicity
(
nonrodent)
870.4200a
Oncogenicity
(
rat)
870.4200b
Oncogenicity
(
mouse)
870.4300
Chronic/
Oncogenicity
yes
yes
yes
yes
yes
yes
yes
yes
yes
yes
870.5100
Mutagenicity_
Gene
Mutation
­
bacterial
870.5300
Mutagenicity_
Gene
Mutation
­
mammalian
870.5xxx
Mutagenicity_
Structural
Chromosomal
Aberrations
870.5xxx
Mutagenicity_
Other
Genotoxic
Effects
yes
yes
yes
yes
yes
yes
yes
yes
870.6100a
Acute
Delayed
Neurotoxicity.
(
hen)
870.6100b
90­
Day
Neurotoxicity
(
hen)
870.6200a
Acute
Neurotoxicity.
Screening
Battery
(
rat)
870.6200b
90
Day
Neurotoxicity.
Screening
Battery
(
rat)
870.6300
Develop.
Neurotoxicity
no
no
no
no
no
­
­
yes
yes
­

870.7485
General
Metabolism
870.7600
Dermal
Penetration
yes
no
yes
­

Special
Studies
for
Ocular
Effects
Acute
Oral
(
rat)
Subchronic
Oral
(
rat)
Six­
month
Oral
(
dog)
no
no
no
no
­
­
­
­
Page
39
of
87
2.0
TOXICOLOGY
STUDIES
2.1.
Subchronic
Toxicity
Studies
2.1.1.
Inoue,
H.
(
1998)
KIF­
230
Technical:
subchronic
toxicity
study
in
rats
by
dietary
administration
for
13
weeks
followed
by
a
4­
week
recovery
study.
Biosafety
Research
Center,
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center)
Shizuoka,
Japan.
Laboratory
Study
No.:
3386,
December
18,
1998.
MRID
45835004.
Unpublished.

EXECUTIVE
SUMMARY
­
In
a
subchronic
oral
toxicity
study
(
MRID
45835004),
Benthiavalicarb
(
KIF­
230;
87.7%
a.
i.,
Lot
#:
G51­
02­
152)
was
administered
to
10
Fischer
rats/
sex/
group
in
the
diet
at
dose
levels
of
0,
50,
200,
5000,
or
20,000
ppm
(
equivalent
to
0/
0,
3.5/
3.9,
14.1/
15.3,
353/
379,
and
1444/
1552
mg/
kg/
day
[
M/
F])
for
13
weeks.
An
additional
10
rats/
sex/
group
were
concurrently
administered
the
0,
5000,
and
20,000
ppm
diets
for
13
weeks
and
were
retained
for
a
recovery
period
of
4
weeks.

There
were
no
adverse
effects
of
treatment
on
survival,
body
weights,
body
weight
gains,
food
consumption,
food
efficiency,
ophthalmoscopy,
or
hematology.

Absolute
and
relative
(
to
body)
liver
weights
were
increased
(
p 
0.01)
in
both
sexes
at
 
5000
ppm
at
Week
13
and
remained
increased
(
p 
0.01)
in
the
20,000
ppm
females
at
Week
17.
Hepatocyte
hypertrophy
was
observed
in
both
sexes
at
5000
ppm
(
3/
10
treated
vs
0/
10
controls)
and
20,000
ppm
(
10/
10
treated).
Additionally
in
both
sexes
at
20,000
ppm,
black
and
enlarged
liver
was
observed
macroscopically.
These
findings
were
not
observed
at
the
end
of
the
recovery
period.

At
Week
13,
the
following
clinical
chemistry
parameters
were
dose­
dependently
increased
(
p 
0.05)
over
controls
and
corroborated
the
toxicity
to
the
liver:
(
i)
total
cholesterol
and
GGT
at
 
5000
ppm
in
both
sexes;
(
ii)
free
cholesterol
and
phospholipids
in
the
20,000
ppm
males
and
 
5000
ppm
females;
(
iii)
total
protein
in
the
 
5000
ppm
males
and
20,000
ppm
females;
and
(
iv)
increased
albumin
in
both
sexes
at
20,000
ppm.
At
the
end
of
the
recovery
period,
increased
(
p 
0.05)
GGT
and
albumin
were
still
observed
in
both
sexes
at
 
5000
ppm,
and
a
minor
increase
in
total
protein
(
p 
0.05)
was
noted
in
the
20,000
ppm
females,
although
to
a
lesser
extent
than
in
the
main
study
animals
at
Week
13.
In
general,
serum
protein
electrophoresis
demonstrated
increases
in
globulin
that
exceeded
increases
in
albumin,
resulting
in
a
decreased
A/
G
ratio
that
may
support
an
effect
on
the
liver
at
 
5000
ppm.

The
LOAEL
for
this
study
is
5000
ppm
(
equivalent
to
352.6/
378.5
mg/
kg/
day
[
M/
F])
based
on
hepatocyte
hypertrophy
in
both
sexes
and
increases
in:
absolute
and
relative
to
body
liver
weights,
total
cholesterol,
and
GGT
in
both
sexes;
free
cholesterol
and
phospholipids
in
the
females;
and
total
protein
in
the
males.
The
NOAEL
is
200
ppm
(
equivalent
to
14.1/
15.3
mg/
kg/
day
[
M/
F]).

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.3100a;
OECD
408)
for
a
subchronic
oral
toxicity
study
in
the
rat.

2.1.2.
Inoue,
H.
(
1998)
KIF­
230
Technical:
Preliminary
oncogenicity
study
in
mice
by
dietary
administration
for
13
weeks.
Biosafety
Research
Center,
Shizuoka,
Japan.
Laboratory
Report
Page
40
of
87
No.:
3385,
November
27,
1997.
MRID
45835007.
Unpublished.

EXECUTIVE
SUMMARY
­
In
a
subchronic
oral
toxicity
study
(
MRID
45835007),
KIF­
230
(
benthiavalicarb;
87.7%
a.
i.,
Lot
#:
G51­
02­
152)
was
administered
to
10
B6C3F1
mice/
sex/
dose
in
the
diet
at
doses
of
0,
50,
200,
7000,
or
20,000
ppm
(
equivalent
to
0/
0,
8.4/
11.3,
33.0/
45.2,
1293/
1620,
or
4031/
4946
mg/
kg/
day
[
M/
F])
for
up
to
90
days.

Urinalysis,
clinical
chemistry,
neurological
evaluations,
and
ophthalmoscopic
examinations
were
not
performed.
No
compound­
related
effects
on
mortality,
clinical
signs,
or
hematology
were
observed
at
any
dose
in
either
sex.

Body
weights
were
decreased
(
p 
0.05)
in
the
7000
ppm
males
at
Weeks
6­
13,
and
at
20,000
ppm
in
the
males
at
Weeks
3­
13
and
in
the
females
at
Weeks
4­
10.
Overall
(
Weeks
0­
13)
body
weight
gains
were
decreased
(
p 
0.01)
in
the
 
7000
ppm
males
and
in
the
20,000
ppm
females
(
not
statistically
significant).
Food
efficiency
was
decreased
(
p 
0.05)
in
the
7000
ppm
males
at
Weeks
4,
7,
9­
10,
and
13.
At
20,000
ppm,
food
consumption
was
sporadically
increased
(
p 
0.05)
in
the
males
and
females,
and
overall
food
consumption
was
slightly
increased
(
p 
0.01)
in
the
males.
Food
efficiency
was
decreased
throughout
the
study
in
the
males,
with
statistical
significance
(
p 
0.05)
achieved
at
Weeks
3­
5,
9,
and
13,
and
during
Weeks
1­
5
in
the
females,
with
statistical
significance
(
p 
0.01)
being
achieved
at
Week
5.
Overall
food
efficiency
was
decreased
(
p 
0.05)
in
the
 
7000
ppm
males
and
in
the
20,000
ppm
females.

The
target
organ
was
the
liver.
At
 
7000
ppm,
absolute
liver
weights
were
increased
(
p 
0.01)
in
both
sexes,
and
relative
(
to
body)
liver
weights
were
also
increased
(
p 
0.01)
in
these
animals.
Gross
lesions
observed
at
 
7000
ppm
included
enlarged
liver
in
all
animals
and
black­
colored
liver
in
7­
10
females.
Additionally
at
20,000
ppm,
black­
colored
liver
was
noted
in
all
males
and
a
white
patch/
zone
in
the
liver
was
noted
in
6/
10
females.
Histopathological
effects
noted
in
the
liver
included:
(
i)
necrosis
in
the
 
7000
ppm
males
and
females;
(
ii)
slight
bile
duct
hyperplasia
in
the
20,000
ppm
males
and
females;
(
iii)
fatty
change
in
the
20,000
ppm
males;
(
iv)
slight
to
moderate
anisonucleosis
in
the
 
7000
ppm
males;
(
v)
multinucleated
giant
cell
formation
in
the
20,000
ppm
males;
and
(
vi)
slight
to
marked
hepatocellular
hypertrophy
in
the
 
7000
ppm
males
and
females.
Electron
microscopy
of
the
liver
samples
from
the
7000
and
20,000
ppm
groups
revealed
increases
in
rough
endoplasmic
reticulum
associated
with
the
hepatocellular
hypertrophy.

Additional
findings
included
brown­
colored
thyroid
in
all
20,000
ppm
males.

No
treatment­
related
effects
were
observed
at
 
200
ppm
in
either
sex.

The
LOAEL
is
7000
ppm
(
equivalent
to
1293/
1620
mg/
kg/
day
[
M/
F];
greater
than
the
limit
dose)
based
on
decreased
body
weight,
body
weight
gain,
and
food
efficiency
in
males,
increased
absolute
and
relative
liver
weight,
enlarged
liver,
black­
colored
liver
(
females
only),
and
histopathological
liver
effects
(
necrosis,
hypertrophy,
and
anisonucleosis
[
males
only]).
The
NOAEL
is
200
ppm
(
equivalent
to
33.0/
45.2
mg/
kg/
day
[
M/
F]).
This
study
is
classified
acceptable/
guideline
and
satisfies
the
guideline
requirement
(
OPPTS
870.3100;
OECD
408)
for
a
90­
day
oral
toxicity
study
in
the
mouse.
Page
41
of
87
2.1.3.
Subchronic
Toxicity
Study
in
Dogs
EXECUTIVE
SUMMARY:
In
a
subchronic
oral
toxicity
study
(
MRID
45835006),
Benthiavalicarb
(
KIF­
230);
(
88.8%
a.
i.,
Lot/
Batch
#:
G51­
08­
158)
was
administered
daily
to
4
beagle
dogs/
sex/
group
in
gelatin
capsules
at
dose
levels
of
0,
40,
200,
or
1000
mg/
kg/
day
for
13
weeks.

At
1000
mg/
kg/
day,
liquid
stool
was
observed
in
three
males
and
one
female
at
1000
mg/
kg/
day.
Hematocrit,
hemoglobin,
and
red
blood
cell
counts
were
decreased
(
p 
0.05),
while
the
percentage
of
reticulocytes
and
the
concentration
of
serum
total
bilirubin
were
increased
in
both
sexes.
These
changes
were
indicative
of
a
hemolytic
anemia.
Moderate
deposits
of
hemosiderin
pigment
were
also
observed
in
the
spleen
and
Kupffer
cells
of
the
liver
in
all
animals
at
this
dose,
indicative
of
secondary
effects
of
a
hemolytic
anemia.
Serum
gamma­
glutamyl
transferase,
alkaline
phosphatase,
and
platelet
counts
were
increased
(
p 
0.05)
in
both
sexes.
Serum
albumin
in
both
sexes
and
creatine
kinase
and
albumin/
globulin
ratio
in
the
females
were
decreased
(
p 
0.05).
In
females,
absolute
and
relative
(
to
body)
thymus
weights
were
decreased
(
p 
0.01).
Absolute
and
relative
(
to
body)
liver
weights
were
increased
(
p 
0.01)
and
enlarged
liver
was
grossly
observed
in
4/
4
males
and
3/
4
females
at
this
dose
compared
to
0/
4
in
controls.
Slight
diffuse
lobular
hepatocyte
hypertrophy
was
also
observed
in
all
animals
at
this
dose.

At
200
mg/
kg/
day,
hematocrit,
hemoglobin,
serum
albumin,
and
albumin/
globulin
ratio
were
decreased
(
p<
0.05)
in
females.
Additionally,
relative
(
to
body)
liver
weights
were
increased
and
slight
diffuse
lobular
hepatocyte
hypertrophy
was
observed
in
two
males
and
two
females
at
this
dose
compared
to
0/
0
in
the
controls.
Absolute
and
relative
(
to
body)
thymus
weights
were
decreased.
A
greater
incidence
of
hemosiderin
pigment
deposits
were
observed
in
the
spleen
of
males
and
the
lesion
was
less
severe
than
at
1000
mg/
kg/
day.

The
only
effects
observed
at
40
mg/
kg
were
decreased
(
p 
0.05)
hematocrit,
hemoglobin,
serum
albumin,
and
relative
(
to
body)
thymus
weight
in
females
(
p 
0.05).

The
LOAEL
is
200
mg/
kg/
day
based
on
decreases
in
hematocrit,
hemoglobin,
serum
albumin
and
albumin/
globulin
ratio
in
females
and
increases
in
liver­
to­
body­
weight
ratios,
hepatocyte
hypertrophy
(
both
sexes)
and
hemosiderin
pigment
deposits
in
the
spleen
of
males.
The
NOAEL
is
40
mg/
kg/
day.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.3150;
OECD
409)
for
a
subchronic
oral
toxicity
study
in
the
dog.

2.2.
Chronic/
Carcinogenicity
Studies
2.2.1.
Murata,
K.
(
2001)
KIF­
230
Technical:
chronic
toxicity
study
and
oncogenicity
study
in
rats.
Biosafety
Research
Center,
Shizuoka,
Japan.
Laboratory
Experimental
No.:
3822
(
001­
208),
September
14,
2001.
MRID
45835017.
Unpublished.
EXECUTIVE
SUMMARY
­
In
this
combined
chronic
toxicity/
carcinogenicity
study
(
MRID
45835017),
KIF­
230
technical
(
benthiavalicarb;
88.8­
89.1%
a.
i.;
dose
levels
adjusted
for
purity;
Lot
No.:
G51­
08­
158)
was
administered
in
the
diet
for
up
to
2
years
to
Fischer
F344/
DuCrj
rats
Page
42
of
87
(
80/
sex/
dose)
at
nominal
concentrations
of
0,
50,
200,
5000,
or
10,000
ppm
(
equivalent
to
0/
0,
2.5/
3.2,
9.9/
12.5,
249.6/
318.2,
and
518.3/
649.4
mg/
kg/
day
in
males/
females).
Ten
rats/
sex/
dose
were
sacrificed
at
Weeks
26,
52,
and
78,
and
the
remaining
survivors
were
sacrificed
at
Week
104.

No
treatment­
related
effects
were
observed
on
mortality,
clinical
signs,
food
consumption,
food
efficiency,
ophthalmoscopic
examination,
or
on
hematology
parameters.

At
 
5000
ppm,
nephrotoxicity
and
hepatotoxicity
were
observed.
In
the
kidney,
absolute
and
relative
(
to
body)
weights
were
increased
throughout
the
study
in
both
sexes.
At
Week
52,
increased
incidences
of
basophilic
tubules
were
noted
in
the
males.
At
Week
104,
increased
incidences
of
the
following
lesions
were
observed
in
the
kidney:
(
i)
chronic
nephropathy
and
tubular
dilatation
in
the
males;
(
ii)
glomerulosclerosis,
calculus,
and
hyaline
droplets
in
both
sexes;
and
(
iii)
lymphocytic
cellular
infiltration,
basophilic
tubules,
hyaline
casts,
and
brown
pigment
deposits
in
the
females.
Urinary
protein
levels
were
increased
at
Week
52;
however,
this
transient
effect
may
have
been
incidental.
Liver
findings
included
increased
gamma­
glutamyltranspeptidase
in
the
males
at
Weeks
26
and
104
and
in
the
females
throughout
the
study.
Increased
levels
of
total
and
free
cholesterol
and
phospholipids
were
observed
in
the
females
throughout
the
study.
Absolute
and
relative
(
to
body)
liver
weights
were
increased
throughout
the
study
in
both
sexes.
Grossly,
hepatic
enlargement
was
noted
at
Week
52
(
in
males
only)
and
at
Week
78
(
in
both
sexes),
and
was
also
observed
in
the
10,000
ppm
group
at
Weeks
26
and
52.
Additionally,
at
Week
104,
increased
incidences
were
observed
of
brown
and/
or
white
patches
on
the
liver
in
the
males,
and
red
patches
on
the
liver
in
the
females.
An
increased
incidence
in
hepatocellular
hypertrophy
was
observed
in
both
sexes
at
Weeks
26,
52,
and
78;
and
in
the
females
at
Week
104
(
and
in
the
10,000
ppm
males
at
Week
104).
A
fatty
change
was
noted
in
the
males
at
Week
52,
in
both
sexes
at
Week
78,
and
in
females
at
Week
104.
Additionally
at
Week
104,
increased
incidences
of
foci/
area
of
cellular
alteration
in
the
liver
and
spongiosis
hepatis
were
observed
in
the
males.

At
10,000
ppm,
the
following
additional
effects
were
observed
in
the
kidney
and
liver.
In
the
kidney,
an
increased
incidence
of
granular
kidney
was
noted
grossly
in
males
at
Week
104.
Glomerusclerosis,
hyaline
casts,
and
hyaline
droplets
were
observed
in
the
males
at
Week
52.
Additionally,
at
Week
104,
increased
incidences
of
transitional
cell
hyperplasia,
fibrosis,
and
lymphocytic
cellular
infiltration
were
noted
in
males.
In
both
sexes,
an
increase
in
the
amount
of
smooth
endoplasmic
reticulum
was
noted
in
the
liver
at
Weeks
26
and
52.
Only
minor
decreases
in
body
weight
and
body
weight
gain
were
observed
in
this
group.

The
LOAEL
is
5000
ppm
(
equivalent
to
249.6/
318.2
mg/
kg/
day
in
males/
females),
based
on
nephrotoxicity
and
hepatotoxicity
in
both
sexes.
The
NOAEL
is
200
ppm
(
equivalent
to
9.9/
12.5
mg/
kg/
day
in
males/
females).

There
was
a
treatment­
related
increase
in
tumor
incidence
when
compared
to
controls.
Increased
incidences
of
hepatocellular
adenoma
were
observed
(%
treated
vs
%
controls)
in
the
10,000
ppm
males
at
Weeks
78
(
10
vs
0;
NS)
and
104
(
14
vs
2;
p 
0.05),
but
fell
within
the
range
of
historical
controls
(
0­
18%).
Mutagenicity
tests
of
this
compound
and
concurrently
submitted
mechanistic
studies
(
MRID
45835021
and
45869715)
suggest
that
KIF­
230
is
acting
as
a
promoter,
but
not
an
initiator,
in
the
development
of
rat
liver
neoplasia.
Additionally,
uterine
adenocarcinoma
was
Page
43
of
87
increased
at
 
5000
ppm
at
Weeks
78
(
20%
vs
0%;
NS)
and
104
(
20­
22%
vs
6%;
p 
0.05)
and
exceeded
the
historical
control
range
(
0­
8%).
Increased
(
p 
0.05)
estradiol
levels
were
observed
in
the
 
5000
ppm
females
at
Weeks
26
and
78
( 
27­
71%).
However,
the
Sponsor
indicated
that
a
more
excessive
increase
in
estradiol
levels
with
corroborating
histological
effects
would
have
been
expected
if
elevated
hormone
levels
were
the
causative
factor
in
the
uterine
neoplasia.
The
Sponsor
stated
that
the
uterine
adenocarcinoma
was
most
likely
due
to
a
direct
effect
on
the
organ.
Dosing
was
considered
to
be
adequate
based
on
increased
lesions
in
the
kidneys
and
liver.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.4300;
OECD
453)
for
a
combined
chronic
toxicity/
carcinogenicity
study
in
rats.

2.2.2.
Murata,
K
(
2001)
KIF­
230
Technical:
Oncogenicity
study
in
mice.
Biosafety
Research
Center,
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center)
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
ID.:
3823
(
001­
209),
September
7,
2001.
MRID
45835016.
Unpublished.

EXECUTIVE
SUMMARY
­
In
a
carcinogenicity
study
(
MRID
45835016),
benthiavalicarb
(
KIF­
230);
88.8­
89.1%
(
w/
w),
dose
levels
adjusted
for
purity,
Lot/
Batch
#
G51­
08­
158)
was
administered
to
70
SLC:
B6C3F1
(
C57BL/
6xC3H)[
SPF]
mice/
sex/
group
in
the
diet
at
dose
levels
of
0,
20,
100,
2500,
or
5000
ppm
(
equivalent
to
0/
0,
2.7/
3.7,
13.7/
18.6,
358.4/
459.3,
731.3/
927.8
mg/
kg/
day
[
M/
F],
respectively)
for
104
weeks,
with
interim
sacrifices
of
10
mice/
sex/
dose
at
52
and
78
weeks.
No
adverse
treatment­
related
effects
were
noted
on
food
consumption.

At
 
2500
ppm,
greater
incidences
of
clinical
signs
of
toxicity
(
abdominal
masses
and
distention,
and
pallor)
were
observed
in
males
during
Weeks
79­
104.
Body
weight,
overall
(
Week
0­
104)
body
weight
gain,
and
overall
(
Weeks
0­
52)
food
efficiency
were
also
decreased
(
p 
0.05)
in
males.
At
5000
ppm,
survival
at
Week
104
was
significantly
(
p 
0.01)
decreased
in
males.
At
5000
ppm,
wasting,
piloerection,
and
tachypnea
were
clinically
observed
in
males
during
Weeks
79­
104.
At
 
2500
ppm,
hepatotoxicity
and
thyroid
gland
toxicity
were
observed.
Absolute
and
relative
to
body
liver
weights
were
increased
(
p 
0.05)
in
both
sexes.
The
incidences
of
brown
patches
and
nodules
of
the
liver
in
both
sexes,
and
enlarged
liver
in
females
were
increased
(
p 
0.05)
at
 
2500
ppm.
At
5000
ppm,
red
patches
in
males
and
white
patches
in
females
were
grossly
observed
in
the
liver.
Slight
intermediate
fatty
change,
slight
hepatocyte
hypertrophy,
slight
to
marked
foci
of
cellular
alteration
were
observed
in
both
sexes.
Necrosis,
single
cell
necrosis,
anisonucleosis,
cellular
infiltration,
lymphocyte
infiltration,
multinucleated
cells,
bile
duct
proliferation,
extramedullary
hematopoiesis,
and
microgranuloma
were
observed
in
males
at
 
2500
ppm
and
were
slight
in
severity.
Additionally,
slight
to
moderate
accumulation
of
macrophages
were
observed
in
males
at
 
2500
ppm.
At
5000
ppm,
slight
necrosis,
single
cell
necrosis,
anisonucleosis,
and
cellular
infiltration
were
observed
in
females
and
slight
angiectasis
was
observed
in
males.

In
the
thyroid
gland,
increased
(
p 
0.01)
incidences
of
follicular
cell
hyperplasia
in
both
sexes
and
dilated
thyroid
follicles
in
males
at
 
2500
ppm
and
in
females
at
5000
ppm
were
observed.
In
the
ovary,
absolute
and
relative
to
body
weight
were
decreased
(
p 
0.05)
and
ovarian
atrophy
was
observed
in
females.
Page
44
of
87
Platelet
count
was
increased
(
p 
0.05)
at
 
2500
ppm
in
both
sexes.
Clotting
time
parameters
were
not
measured;
therefore,
the
functional
impact
of
increased
platelet
count
was
not
demonstrated.
However,
the
platelet
count
difference
was
corroborated
by
an
increase
in
the
number
of
megakaryocytes
in
the
5000
ppm
males.
The
toxicological
relevance
of
this
finding
was
considered
equivocal.

The
only
effects
observed
at
100
ppm
were
abdominal
distention
during
Weeks
79­
104
and
slight
microgranuloma
in
the
liver
in
males.

The
LOAEL
is
2500
ppm
(
equivalent
to
358.4/
459.3
mg/
kg/
day
[
M/
F])
based
on
clinical
signs
(
abdominal
masses
and
distention,
and
pallor)
in
males,
decreased
body
weight
gain
and
food
efficiency
in
males,
gross
liver
lesions
(
brown
patches
and
nodules
in
both
sexes,
and
enlarged
liver
in
females),
increased
absolute
and
relative
(
to
body)
liver
weight
in
both
sexes,
nonneoplastic
liver
lesions
(
hepatocyte
hypertrophy,
hepatic
intermediate
fatty
change,
and
hepatic
foci
of
cellular
alteration
in
both
sexes,
and
anisonucleosis,
necrosis,
single
cell
necrosis,
multinucleated
cells,
accumulation
of
macrophages,
cellular
infiltration,
lymphocyte
infiltration,
bile
duct
proliferation,
and
extramedullary
hematopoiesis
in
males),
thyroid
follicular
cell
hyperplasia
in
both
sexes,
and
dilated
thyroid
follicles
in
males.
The
NOAEL
is
100
ppm
(
equivalent
to
13.7/
18.6
mg/
kg/
day
in
M/
F).

At
 
2500
ppm,
there
was
a
treatment­
related
increase
(
p 
0.01)
in
tumor
incidence
of
the
liver
and
thyroid
gland
compared
to
controls.
In
the
liver,
hepatocellular
adenomas
(
20­
47/
50
treated
vs
4­
15/
50
control)
in
both
sexes
and
hepatoblastoma
(
9­
12/
50
treated
vs
0/
50
control)
and
hepatocellular
carcinomas
(
35­
43/
50
treated
vs
12/
50
control)
in
males
were
increased.
Increased
incidences
of
hepatocellular
adenomas
were
also
observed
at
Week
52
in
the
5000
ppm
males
(
7/
10
treated
vs
1/
10
control)
and
at
Week
78
in
the
5000
ppm
males
(
10/
10
treated
vs
5/
10
control)
and
2500
ppm
females
(
6/
10
treated
vs
1/
10
controls).
In
the
thyroid
gland,
follicular
cell
adenomas
were
increased
in
males
at
5000
ppm
(
9/
50
treated
vs
0/
50
control).
Additionally,
the
incidence
of
malignant
lymphoma
was
higher
in
all
treated
females
compared
to
control;
increases
were
statistically
significant
(
p 
0.05)
in
the
100
and
5000
ppm
groups
(
8/
50
treated
vs
2/
50
control).
A
clear
dose­
response
was
not
observed
for
this
increase;
however,
these
lesions
may
have
been
related
to
treatment.
Dosing
was
considered
adequate
based
on
clinical
signs,
increased
liver
weight,
gross
and
microscopic
lesions
of
the
liver,
and
follicular
cell
hyperplasia
of
the
thyroid.

This
carcinogenicity
study
in
mice
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.4200;
OECD
451)
for
a
chronic
oral
toxicity
study
in
the
mouse.

2.2.3.
Hasegawa,
K.
(
2001)
KIF­
230
Technical:
Chronic
toxicity
study
by
oral
administration
to
beagle
dogs
for
52
weeks.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center)
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
ID.:
4551(
001­
242).
March
12,
2001.
MRID
45835015.
Unpublished.

EXECUTIVE
SUMMARY
­
In
a
chronic
oral
toxicity
study
(
MRID
45835015),
benthiavalicarb
(
KIF­
230)
technical;
(
87.5­
87.9%
a.
i.;
Lot/
Batch
#
G51­
15­
162)
was
administered
to
4
beagle
dogs/
sex/
group
in
gelatin
capsules
at
dose
levels
of
0,
4,
40,
or
400
mg/
kg/
day
for
52
weeks.
Page
45
of
87
At
400
mg/
kg/
day,
lower
(
p 
0.05)
fibrinogen
values
in
the
400
mg/
kg
males
at
weeks
26
(
179
mg/
dL)
and
52
(
158
mg/
dL)
were
noted
as
compared
to
controls
(
300
and
267
mg/
dL
at
weeks
26
and
52,
respectively).
Serum
free
fatty
acids
in
males
were
higher
than
controls
at
Weeks
13
(
p 
0.01),
26
(
p 
0.05),
and
52
(
NS).
Absolute
liver
weights
were
increased
(
p 
0.05)
and
relative
(
to
body)
liver
weights
were
higher
(
not
statistically
significant)
in
both
sexes
at
400
mg/
kg
as
compared
to
controls.
Histopathology
findings
associated
with
administration
of
the
test
material
were
not
observed
in
this
study.

The
LOAEL
was
determined
to
be
 
400
mg/
kg/
day
based
upon
minimal
increases
in
absolute
(
p 
0.05)
and
relative
liver
weights
(
N.
S.)
at
the
highest
dose
level,
400
mg/
kg/
day.
In
addition,
lower
(
p 
0.05)
fibrinogen
values
in
the
400
mg/
kg
males
at
weeks
26
(
179
mg/
dL)
and
52
(
158
mg/
dL)
were
noted
as
compared
to
controls
(
300
and
267
mg/
dL
at
weeks
26
and
52,
respectively.
Serum
free
fatty
acids
were
increased
( 
73­
198%;
p 
0.05)
in
the
400
mg/
kg
males
at
weeks
13
and
26.
At
week
52,
free
fatty
acids
were
also
high
( 
64%,
NS).
The
toxicological
significance
of
this
finding
is
unclear.

The
NOAEL
is
40
mg/
kg/
day.
Although
minimal
toxicity
(
absolute
and
relative
liver
weight
increase)
was
observed
at
the
400
mg/
kg/
day
dose
level,
findings
were
not
supported
by
histopathological
findings
and
the
toxicological
significance
of
the
reported
clinical
chemistry
findings
remains
unclear.

The
dose
levels
in
this
study
were
selected
based
on
findings
in
the
concurrently
submitted
90­
day
subchronic
study
(
MRID
45835006).
The
LOAEL
in
this
study
was
determined
to
be
200
mg/
kg/
day.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.4100b;
OECD
452)
for
a
non­
rodent
chronic
oral
toxicity
study.
However,
it
is
recognized
that
dose
levels
in
this
study
could
have
been
higher
eliciting
a
more
definitive
toxicological
response
at
the
highest
dose.

2.3.
Developmental
and
Reproductive
Toxicity
2.3.1.
Itoh,
K.
(
2000)
KIF­
230
Technical:
teratogenicity
study
in
rats.
Biosafety
Research
Center,
Shizuoka,
Japan.
Laboratory
Experimental
No.:
4541
(
001­
240),
April
12,
2000.
MRID
45835012.
Unpublished.

Tanaka,
R.
(
1998)
KIF­
230
Technical:
preliminary
teratogenicity
study
in
rats.
Biosafety
Research
Center,
Sizuoka,
Japan.
Laboratory
Experimental
No.:
3387,
December
11,
1998.
MRID
45835011.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
developmental
toxicity
study
(
MRID
45835012),
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.,
dose
levels
adjusted
for
purity,
Lot/
Batch
#
G51­
08­
158)
was
administered
daily
via
oral
gavage
to
25
presumed
pregnant
Sprague­
Dawley
rats/
group
at
dose
levels
of
0,
10,
100,
or
1000
mg/
kg/
day
from
gestation
day
(
GD)
7
through
19.
All
dams
were
killed
on
GD
20;
their
fetuses
were
removed
by
cesarean
section
and
examined.
There
were
no
effects
of
treatment
on
survival,
clinical
signs,
body
weights,
body
weight
gains,
or
food
Page
46
of
87
consumption.

Enlarged
liver
was
noted
in
2/
23
dams
at
100
mg/
kg/
day
and
6/
24
dams
at
1000
mg/
kg/
day.
Additionally
at
1000
mg/
kg/
day,
absolute
and
relative
liver
weights
were
increased
(
p 
0.01)
by
11­
12%.
The
liver
was
not
examined
microscopically
in
this
study.
In
order
to
determine
if
the
effects
on
these
organs
were
an
adaptive
response
to
the
presence
of
the
test
substance
or
if
these
organs
were
adversely
affected,
the
reviewers
compared
these
findings
to
those
of
the
subchronic
rat
study
(
MRID
45835004).
The
reviewers
concluded
that
the
effects
on
the
liver
at
1000
mg/
kg/
day
in
the
developmental
study
were
adverse
based
on
similar
increases
in
liver
weights
with
hypertrophy
and
corroborating
changes
in
clinical
chemistry
at
comparable
doses
in
the
subchronic
study.

The
maternal
LOAEL
is
1000
mg/
kg/
day
(
limit
dose)
based
on
increased
absolute
and
relative
liver
weights
and
on
incidences
of
enlarged
liver.
The
maternal
NOAEL
is
100
mg/
kg/
day.

There
were
no
abortions,
premature
deliveries,
or
complete
litter
resorptions.
There
were
no
effects
of
treatment
on
the
numbers
of
litters,
fetuses
(
live
or
dead),
or
resorptions
(
early
or
late)
or
on
sex
ratio,
fetal
weights,
placental
weights,
or
post­
implantation
loss.
There
were
no
treatment­
related
external,
visceral,
or
skeletal
malformations,
variations,
or
retardations.

The
developmental
LOAEL
was
not
observed.
The
developmental
NOAEL
is
1000
mg/
kg/
day
(
limit
dose).

This
study
is
classified
acceptable/
guideline
(
OPPTS
870.3700a)
and
satisfies
the
guideline
requirements
for
a
developmental
study
in
the
rat.

2.3.2.
Itoh,
K.
(
2000)
KIF­
230
Technical:
teratogenicity
study
in
rabbits.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4762
(
001­
255),
November
30,
2000.
MRID
45835010.
Unpublished.

Itoh,
K.
(
1999)
KIF­
230
Technical:
preliminary
teratogenicity
study
in
rabbits.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4542
(
001­
241),
December
28,
1999.
MRID
45835008.
Unpublished.

Tanaka,
R.
(
1998)
KIF­
230
Technical:
preliminary
teratogenicity
study
in
rabbits.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
3388,
December
11,
1998.
MRID
45835009.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
developmental
toxicity
study
(
MRID
45835010),
KIF­
230
(
benthiavalicarb;
87.5­
87.9%
a.
i.,
dose
levels
adjusted
for
purity,
Lot/
Batch
#
G51­
15­
162)
in
0.5%
CMC­
Na
containing
1%
Tween
80
was
administered
daily
via
oral
gavage
at
a
dose
volume
of
10
mL/
kg
to
22
presumed
pregnant
New
Zealand
White
rabbits/
group
at
dose
levels
of
0,
10,
20,
or
40
mg/
kg/
day
from
gestation
day
(
GD)
6
through
28.
All
does
were
sacrificed
on
GD
29;
Page
47
of
87
their
fetuses
were
removed
by
cesarean
section
and
examined.

There
were
no
effects
of
treatment
on
maternal
survival,
clinical
signs,
body
weights,
body
weight
gains,
food
consumption,
or
gross
pathology.

At
40
mg/
kg/
day,
two
does
aborted
on
GD
25
(#
2315)
and
GD
28
(#
2317)
following
dramatically
decreased
food
consumption
and
body
weights.
These
abortions
were
considered
to
be
secondary
to
the
decreased
food
consumption
and
subsequent
body
weight
losses
in
these
animals.
Because
there
were
no
treatment­
related
effects
on
group
mean
food
consumption,
body
weights,
or
body
weight
gains
in
the
treated
groups
compared
to
controls,
the
decreases
in
these
two
animals
were
not
considered
treatment­
related.
Furthermore,
in
the
concurrently
submitted
preliminary
studies
(
MRIDs
45835008
and
45835009),
there
were
no
abortions
at
40
or
60
mg/
kg/
day,
and
abortion
was
noted
in
one
doe
at
30
mg/
kg/
day.
Thus,
when
the
three
studies
are
examined
in
conjunction,
the
incidences
of
abortion
are
unrelated
to
dose.

Additionally
at
40
mg/
kg/
day,
relative
(
incr.
11%;
p 
0.05)
and
absolute
(
incr.
10%;
not
significant)
liver
weights
were
increased.
The
liver
was
not
examined
microscopically
in
this
study.
In
other
concurrently
reviewed
toxicity
studies
on
this
chemical,
the
doses
at
which
liver
toxicity
was
observed
in
rats
(
353­
1444
mg/
kg/
day),
mice
(
358­
1293),
and
dogs
(
1000
mg/
kg/
day)
are
considerably
higher
than
the
highest
dose
tested
in
the
rabbit
study
(
40
mg/
kg/
day).
However,
given
the
overall
weight
of
evidence
and
the
known
mode
of
action
for
affecting
the
liver,
the
reviewers
consider
the
increased
liver
weights
in
the
rabbits
to
be
adverse.

The
maternal
LOAEL
is
40
mg/
kg/
day
based
on
increased
absolute
and
relative
liver
weights.
The
maternal
NOAEL
is
20
mg/
kg/
day.

There
were
no
premature
deliveries
or
complete
litter
resorptions;
and
there
were
no
effects
of
treatment
on
the
numbers
of
abortions,
litters,
fetuses
(
live
or
dead),
or
resorptions
(
early
or
late)
or
on
fetal
weights,
placental
weights,
sex
ratio,
or
post­
implantation
loss.
There
were
no
treatment­
related
external,
visceral,
or
skeletal
malformations,
variations,
or
retardations.

The
developmental
LOAEL
was
not
observed.
The
developmental
NOAEL
is
40
mg/
kg/
day.

This
study
is
classified
acceptable/
guideline
and
satisfies
the
guideline
requirements
for
a
developmental
toxicity
study
in
the
rabbit
(
OPPTS
870.3700b).

2.3.3.
Tanaka,
R.
(
1999)
KIF­
230
technical:
two­
generation
reproduction
toxicity
study
in
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Experimental
No.:
3820
(
001­
206),
December
21,
1999.
MRID
45835014.
Unpublished.

Tanaka,
R.
(
1998)
KIF­
230
technical:
preliminary
two­
generation
reproduction
toxicity
study
in
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Experimental
No.:
3389,
December
11,
1998.
MRID
45835013.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
two­
generation
reproduction
toxicity
study
(
MRIDs
45835014
Page
48
of
87
and
45835013),
KIF­
230
(
benthiavalicarb;
88.8­
89.1%
a.
i.,
dose
levels
adjusted
for
purity);
Lot
No.
G51­
08­
158)
was
administered
continuously
in
the
diet
to
Sprague­
Dawley
rats
(
25
rats/
sex/
dose)
at
dose
levels
of
0,
100,
1000,
or
10,000
ppm
(
equivalent
to
0/
0,
8.4/
8.8,
84.1/
91.0,
and
880/
943
mg/
kg
bw/
day
[
M/
F]).
The
P
and
F1
parents
were
dosed
for
at
least
10
weeks
before
they
were
mated
to
produce
the
F1
and
F2
litters,
respectively.
The
F1
pups
were
weaned
on
postnatal
day
(
PND)
22,
and
21­
22
pups/
sex/
group
(
1
pup/
sex/
litter
as
nearly
as
possible)
were
randomly
selected
as
parents
of
the
F2
generation.

No
treatment­
related
adverse
effect
was
observed
in
the
parents
on
mortality,
clinical
signs,
body
weight,
body
weight
gain,
food
consumption,
food
efficiency,
gestation
index,
or
on
gross
or
histological
pathology.

Only
adaptive
changes
were
observed
in
the
liver
(
generally
at
10,000
ppm),
including
gross
enlargement
in
one
P
and
one
F1
female,
slight
hepatocyte
hypertrophy,
and
increased
absolute
and
relative
to
body
organ
weights.

The
LOAEL
for
parental
toxicity
was
not
observed.
The
NOAEL
is
10,000
ppm
(
equivalent
to
880/
943
mg/
kg/
day
in
males/
females).

No
treatment­
related
effect
was
observed
on
the
estrous
cycle;
sperm
enumeration,
activity,
motility,
or
morphology;
male
or
female
fertility
or
fecundity
indices;
the
duration
of
gestation;
number
of
stillborn
pups;
the
delivery
indices;
or
the
number
of
implantation
sites.

The
LOAEL
for
reproductive
performance
was
not
observed.
The
NOAEL
is
10,000
ppm
(
equivalent
to
880/
943
mg/
kg/
day
in
males/
females).

No
treatment­
related
adverse
effects
were
observed
on
the
following:
number
of
pups
born
alive
or
dead,
and
deaths
during
Days
0­
4
and
5­
21;
sex
ratio;
litter
size
at
Days
0,
4,
7,
14,
and
21;
the
birth,
viability,
or
weaning
indices
in
the
F1
generation;
time
to
detachment
of
the
auricle,
eruption
of
the
lower
incisor,
or
separation
of
eyelids;
body
weight;
time
to
vaginal
opening
or
preputial
separation;
organ
weights;
or
gross
and
histological
pathology.
It
was
stated
that
only
sporadic
clinical
signs
were
observed
among
the
offspring.

Increases
in
absolute
and
relative
to
body
liver
weights
were
observed
at
10,000
ppm,
and
this
was
considered
an
adaptive
response.

The
LOAEL
for
offspring
toxicity
was
not
observed.
The
NOAEL
is
10,000
ppm
(
equivalent
to
880/
943
mg/
kg/
day
in
males/
females).
This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.3800;
OECD
416)
for
a
two­
generation
reproduction
study
in
the
rat.

2.4.
Neurotoxicity
2.4.1.
Rush,
R.
E.
(
2002)
An
acute
oral
neurotoxicity
study
of
KIF­
230
(
TGAI)
in
rats.
Springborn
Laboratories,
Inc.,
Spencerville,
OH.
Laboratory
Project
ID:
3404.12,
June
10,
2002.
MRID
45869735.
Unpublished.
Page
49
of
87
EXECUTIVE
SUMMARY:
In
an
acute
oral
neurotoxicity
study
(
MRID
45869735),
KIF­
230
(
benthiavalicarb;
92.3%
a.
i.,
Lot
#
G51­
24­
176)
in
0.5%
(
w/
v)
aqueous
carboxymethylcellulose
was
administered
in
a
single
dose
by
gavage
(
10
mL/
kg)
to
non­
fasted
Sprague­
Dawley
rats
(
5/
sex/
dose)
at
doses
of
0
or
2000
mg/
kg
(
limit
dose).
All
animals
were
observed
for
up
to
14
days
post­
dosing.
Functional
observational
battery
(
FOB)
and
motor
activity
were
evaluated
pretreatment
and
on
Days
1
(
at
the
time
of
peak
effect,
1
hour
post­
dosing),
8,
and
15.
At
termination,
5
rats/
sex/
group
were
perfused
in
situ
for
neurohistological
examination.
Positive
control
data
were
provided.

No
compound­
related
effects
on
clinical
signs,
body
weight,
body
weight
gain,
FOB,
motor
activity,
or
gross
and
histopathology
parameters
were
observed
in
either
sex
at
2000
mg/
kg
(
limit
dose).

The
LOAEL
was
not
observed.
The
NOAEL
is
2000
mg/
kg
(
limit
dose).

No
evidence
of
neurotoxicity
was
observed.

The
submitted
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirements
(
OPPTS
870.6200a;
OECD
424)
for
an
acute
neurotoxicity
screening
battery
in
rats.

2.4.2.
Cooper,
S.
(
2002)
KIF­
230
(
TGAI):
Neurotoxicity
study
by
dietary
administration
to
CD
rats
for
4
weeks.
Huntingdon
Life
Sciences
Ltd.,
Huntingdon,
UK.
Laboratory
Project
ID:
KCI
207/
022387,
May
27,
2002.
MRID
45869736.
Unpublished.

EXECUTIVE
SUMMARY
­
In
a
subchronic
neurotoxicity
study
(
MRID
45869736),
KIF­
230
(
benthiavalicarb;
92.6%
a.
i.,
Batch
#
G51­
36­
184)
was
administered
in
the
diet
to
10
CD
rats/
sex/
group
at
doses
of
0,
200,
2000,
or
20,000
ppm
(
equivalent
to
0/
0,17.7/
19.3,
174.1/
185.7,
and
1853.7/
1845.8
mg/
kg/
day
[
M/
F],
respectively)
for
4
weeks.
All
animals
were
subjected
to
a
complete
FOB
and
motor
activity
assessment
during
pretreatment
and
at
Week
4.
An
abbreviated
FOB
(
excluding
sensory,
neuromuscular,
and
physiological
observations)
was
performed
weekly
throughout
treatment.
At
termination,
5
rats/
sex/
group
were
perfused
in
situ,
and
tissues
from
the
control
and
20,000
ppm
groups
were
examined
microscopically.
Positive
control
data
were
provided.

No
compound­
related
effects
on
mortality,
clinical
signs,
food
consumption,
FOB,
motor
activity,
brain
weight
and
measurements,
gross
necropsy,
or
neuropathology
parameters
were
observed.
In
the
20,000
ppm
males,
body
weights
were
decreased
(
not
statistically
significant;
NS)
by
5­
7%
compared
to
controls
throughout
the
study,
and
body
weight
gains
were
decreased
(
p<
0.05)
by
33%
during
Weeks
0­
1
and
by
18%
overall
(
Weeks
0­
4).
Additionally,
food
efficiency
was
minimally
decreased
(
NS)
throughout
treatment
at
Weeks
1­
4
(
14.3­
23.6%
treated
vs
17.1­
28.5%
controls)
and
overall
(
Weeks
1­
4;
18.5%
treated
vs
23.1%
controls).

No
treatment­
related
effects
were
observed
in
the
 
2000
ppm
males
or
in
the
females
at
any
dose.

The
LOAEL
is
20,000
ppm
(
equivalent
to
1853.7/
1845.8
mg/
kg/
day
[
M/
F];
>
limit
dose)
based
on
decreased
body
weight
and
body
weight
gain
in
the
males.
The
NOAEL
is
2000
ppm
(
equivalent
to
174.1/
185.7
mg/
kg/
day
[
M/
F]).
Page
50
of
87
There
was
no
evidence
of
neurotoxicity
at
any
dose
tested.

The
study
is
classified
as
acceptable/
guideline
and
satisfies
the
guideline
requirement
(
870.6200b)
for
a
subchronic
neurotoxicity
study
in
the
rat.
Although
the
study
was
only
4
weeks
in
duration
(
not
the
typical
13
weeks),
it
was
acceptable
because
there
were
no
signs
of
neurotoxicity
in
the
concurrently
reviewed
acute
neurotoxicity
study
(
MRID
45869735)
at
2000
mg/
kg
(
limit
dose),
or
in
the
chronic
oncogenicity
rat
study
(
MRID
45835017)
at
518/
649
mg/
kg/
day
[
M/
F],
and
the
animals
were
dosed
at
nearly
2x
the
limit
dose
for
the
subchronic
neurotoxicity
study.

2.5.
Metabolism
Study
2.5.1.
Dow,
P.,
and
G.
Lappin.
(
2001)
(
14C)­
KIF­
230R­
L:
absorption,
distribution,
metabolism
and
excretion
following
oral
administration
to
the
rat.
Covance
Laboratories,
Inc.,
Harrogate,
North
Yorkshire,
England.
Laboratory
Project
Identification:
Covance
Report
No.
535/
55­
D1141,
September
19,
2001.
MRID
45835108.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
series
of
rat
metabolism
studies
(
MRID
45835108),
[
14C]
KIF­
230R­
L
in
0.5%
aqueous
carboxymethylcellulose
plus
Tween­
80
was
administered
to
Fischer
(
CDF
Crl:
BR
[
F344])
rats
by
gavage.
In
the
main
excretion/
mass
balance
study,
five
rats/
sex/
dose
were
given
either
[
Bz­
14C]
KIF­
230R­
L
(
Batch
#
CP­
2176;
radiochemical
purity
96.5­
96.7%)
or
[
Val­
14C]
KIF­
230R­
L
(
Batch
#
CP­
2179;
radiochemical
purity
96.8­
100.7%)
as
a
single
oral
dose
of
5
or
400
mg/
kg.
Rats
were
dosed
similarly
in
the
pharmacokinetic
study
(
five
rats/
sex/
dose),
the
tissue
distribution
study
(
four
rats/
sex/
dose/
sampling
interval),
and
the
biliary
excretion
study
(
two
to
three
rats/
sex/
dose).
The
distribution
of
radioactivity
between
tissues
and
excreta
was
determined.
Metabolites
were
identified
and
quantified
in
the
urine,
feces,
and
bile
from
the
excretion/
mass
balance
and
biliary
excretion
studies.
Metabolites
in
plasma,
liver,
and
kidney
were
identified
from
the
tissue
distribution
study.

The
low
and
high
doses
of
both
radiolabeled
forms
were
absorbed
rapidly.
As
indicated
by
the
biliary
excretion
data
discussed
below,
absorption
of
the
low
dose
was
essentially
complete,
but
absorption
of
the
high
dose
was
limited.
Blood
and
plasma
parameters
were
similar
within
radiolabeled
forms
and
doses.
Animals
dosed
with
the
[
Val­
14C]
form
demonstrated
greater
maximum
concentrations,
terminal
elimination
half­
lives,
and
AUCs
than
animals
dosed
with
the
[
Bz­
14C]
form,
particularly
at
the
high
dose.

In
the
excretion/
mass
balance
groups,
total
recoveries
ranged
from
89.4­
100.4%
of
the
dose
by
168
h
post­
dose.
No
differences
were
observed
in
the
distribution
of
radioactivity
between
the
[
Bz­
14C]
and
[
Val­
14C]
labels.
Radioactivity
was
recovered
primarily
in
the
feces
(
62.7­
83.1%)
and
urine
(
7.1­
24.9%),
with
the
majority
of
the
dose
being
excreted
within
48
h
(
72.7­
85.7%
dose).
Although
a
minor
route
of
elimination,
urinary
excretion
was
higher
in
the
females
(
11.5­
24.9%
dose)
than
males
(
7.1­
14.3%
dose),
and
was
also
higher
in
the
low
dose
females
(
22.3­
24.9%
dose)
than
the
high
dose
females
(
11.5­
13.1%
dose).
Cage
washes,
cage
debris,
carcass,
and
tissues
each
accounted
for
low
amounts
( 
1.5%
dose)
of
the
administered
doses,
and
data
from
a
preliminary
study
indicated
that
<
0.1%
of
the
dose
would
be
expected
in
exhaled
air.
At
168
h
post­
dose,
the
concentration
of
radioactivity
was
notably
higher
in
the
kidneys
and
liver
in
all
groups.
Residual
radioactivity
was
also
consistently
detected
in
the
trachea,
whole
blood,
Page
51
of
87
adrenals,
aorta,
plasma,
bile
duct,
heart,
Harderian
gland,
lung,
and
gastrointestinal
tract.
Low
concentrations
of
radioactivity
were
sporadically
detected
in
other
tissues
of
individual
animals.

The
time
course
study
on
the
distribution
of
radioactivity
in
tissues
showed
that
radioactivity
was
initially
detected
in
a
number
of
tissues,
but
concentrations
decreased
steadily
over
time.
Concentrations
in
plasma
were
higher
than
in
whole
blood,
indicating
that
[
14C]
residues
preferentially
partitioned
into
the
plasma.
With
the
exception
of
the
gastrointestinal
tract,
bile
duct,
bladder,
and
residual
carcass,
the
concentration
of
radioactivity
was
highest
in
the
kidneys
and
liver
in
both
sexes
at
all
time
points,
regardless
of
dose
group
and
[
14C]
label
position.
In
other
tissues
(
such
as
adrenals,
epididymis,
lymph
nodes,
pancreas,
prostate
gland,
trachea,
and
thyroid),
increases
over
levels
in
plasma
were
observed
in
both
sexes
at
the
first
time
point
examined,
but
concentrations
decreased
to
approximately
plasma
levels
by
24/
48
h.
All
other
tissues
had
concentrations
of
radioactivity
that
were
comparable
to
or
below
the
concentrations
in
plasma
throughout
the
time
course.

In
the
biliary
excretion
study,
the
excretion
and
distribution
of
radioactivity
were
similar
between
the
sexes
and
was
generally
similar
between
the
two
[
14C]
labels,
although
biliary
excretion
was
slightly
higher
and
urinary
excretion
was
slightly
lower
for
the
[
Bz­
14C]
label
compared
to
the
[
Val­
14C]
label.
For
all
groups,
the
total
recovery
of
the
administered
dose
was
85.8­
107.3%.
At
the
low
dose,
biliary
excretion
accounted
for
63.6­
90.4
%
of
the
dose,
urinary
excretion
accounted
for
4.2­
19.1%
dose,
and
feces
accounted
for
1.1­
3.8%
dose.
At
the
high
dose,
levels
of
urinary
excretion
were
similar
(
2.1­
13.1%
dose),
but
biliary
excretion
(
27.8­
40.3%
dose)
decreased
with
a
concomitant
increase
in
fecal
excretion
(
32.2­
60.9%
dose).
Based
on
the
recovery
of
radioactivity
in
the
bile
and
urine,
82.7­
94.7%
of
the
low­
dose
was
absorbed
within
48
hours
of
dosing
compared
to
34.6­
47.7%
of
the
high
dose.

Thirteen
compounds
were
identified
through
extensive
analysis
and
quantitation
of
metabolites
in
bile,
urine,
and
feces
of
bile
duct­
cannulated
rats.
The
metabolites
included
parent,
which
accounted
for
43.5­
60.2%
of
the
dose.
The
metabolite
profile
was
qualitatively
similar
between
the
sexes,
dose
levels,
and
[
14C]
labels,
and
the
quantitative
levels
of
metabolites
were
generally
similar
between
the
sexes,
particularly
for
the
major
metabolites.

In
urine,
a
total
of
11
metabolites
were
identified,
including
U6,
B7,
B8,
B9,
B11,
B13,
M3,
M19,
M15,
M18
and
U37.
Parent
was
not
detected
in
urine,
and
most
of
the
metabolites
were
present
at
low
levels
(<
2%
dose).
The
primary
urinary
metabolites
in
each
dose
group
included:
B8,
B11,
B13,
and
M15.
In
feces,
parent
compound
was
the
major
[
14C]
residue,
accounting
for
0.19­
1.18%
of
the
low
dose
and
27.74­
42.44%
of
the
high
dose,
further
indicating
that
absorption
was
incomplete
at
the
high
dose.
Other
minor
metabolites
(<
1%
dose)
detected
in
feces
included
B11,
M3,
M15,
and
M18.
For
both
urine
and
feces,
individual
unknown
components
each
accounted
for
<
2%
of
the
dose.

A
total
of
eight
metabolites
were
identified
in
bile,
with
the
principal
metabolites
being
isomers
of
hydroxylated
glutathione
conjugates
(
B7,
B8,
B9)
and
a
glucuronide
conjugate
(
B11);
parent
was
not
detected
in
bile.
Metabolite
B11
was
the
major
biliary
metabolite
in
both
low­
dose
(
13.16­
20.56%
dose)
and
high­
dose
rats
(
5.71­
9.41%
dose),
and
the
isomeric
metabolites
B7,
B8,
and
B9
together
accounted
for
16.57­
23.68%
of
the
dose
in
low­
dose
rats
and
5.82­
10.88%
of
the
dose
in
high­
dose
rats.
All
other
identified
metabolites
were
generally
present
in
minor
amounts
Page
52
of
87
(<
5%
dose).

Based
upon
the
components
identified
in
excreta,
the
metabolism
of
KIF­
230R­
L
in
rats
primarily
involves
either
hydroxylation
of
the
phenyl
ring
followed
by
glucuronic
acid
conjugation
or
glutathione
conjugation
and
hydroxylation
of
the
phenyl
ring
with
subsequent
degradation
of
the
glutathione
moiety
resulting
in
a
variety
of
metabolites
present
in
small
proportions.
Cleavage
and
hydroxylation
of
the
valyl
side
chain
also
occurs
to
a
limited
extent.

The
proposed
pathway
was
also
supported
by
results
from
the
analysis
of
urine
and
fecal
extracts
from
the
main
excretion/
mass
balance
groups
and
analysis
of
plasma,
liver,
and
kidney
samples
from
the
tissue
distribution
groups.
The
major
components
detected
in
urine
were
metabolites
M15,
M19,
M18,
B11,
and
U6
([
Bz­
14C]
label
only),
and
the
major
components
detected
in
feces
were
parent
and
metabolite
M15.
Parent
and
metabolites
M18
and
M15
were
also
the
principal
components
detected
in
plasma,
liver,
and
kidneys.

This
metabolism
study
in
the
rat
is
classified
acceptable/
guideline
and
satisfies
the
guideline
requirement
for
a
Tier
1
metabolism
study
[
OPPTS
870.7485,
OPP
85­
1]
in
rats.

2.6.
Mechanistic
Studies
2.6.1.
Hirouchi,
Y.
(
2000)
KIF­
230
technical:
two
stage
hepatocarcinogenicity
study
using
rats
for
examining
the
initiator
effect.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4906
(
001­
261),
November
8,
2000.
MRID
45869715.
Unpublished.

EXECUTIVE
SUMMARY:
In
this
non­
guideline
hepatocarcinogenicity
study
(
MRID
45869715),
male
Fischer
(
F344/
DuCrj
[
SPF])
rats
were
placed
in
three
groups
containing
12
animals
each.
A
b
partial
hepatectomy
was
performed
on
all
groups
to
stimulate
hepatic
cell
division.
Twelve
hours
later,
animals
were
initiated
with
a
single
i.
p.
injection
of
20
mg/
kg
NNitrosodiethylnitrosamine
(
DEN)
in
0.9%
saline
(
Group
1),
a
single
gavage
dose
of
2000
mg/
kg
KIF­
230
(
benthiavalicarb;
88.8­
89.1%
a.
i.;
Lot
#:
G51­
08­
158)
in
0.5%
carboxymethylcellulose
sodium,
1.0%
Tween­
80
(
CMC­
Na)
(
Group
2)
at
a
dose
volume
of
5
mL/
kg,
or
a
single
gavage
dose
of
CMC­
Na
only
(
Group
3).
All
rats
were
fed
basal
diet
for
the
first
two
weeks,
then
diet
containing
0.05%
(
w/
w)
phenobarbital
sodium
(
PB)
from
Study
Weeks
2­
10.
Additionally,
on
Week
3,
all
rats
were
given
a
single
i.
p.
injection
of
300
mg/
kg
D­
galactosamine,
a
hepatic
toxicant.
Eight
weeks
after
PB
dosing
was
started
(
Week
10),
the
rats
were
killed,
and
immunohistochemically­
stained
liver
sections
were
examined
for
the
presence
of
the
placental
form
of
glutathione­
S­
transferase
(
GST­
P)
positive
cell
foci.
The
purpose
of
this
study
was
to
examine
the
potential
for
KIF­
230
to
act
as
a
hepatic
tumor
initiator
by
using
an
established
twostage
model
of
hepatocarcinogenesis.

No
effects
of
treatment
were
observed
on
mortality,
clinical
signs
of
toxicity,
body
weights,
body
weight
gains,
or
gross
pathology.

Compared
to
the
DEN
animals,
food
consumption
was
greater
in
the
KIF­
230
and
CMC­
Na
groups
during
Study
Weeks
1,
2,
and
5;
total
(
Weeks
0­
10)
food
consumption
was
also
greater
in
the
KIF­
230
and
CMC­
Na
animals
compared
to
the
DEN
group.
Food
efficiency
was
greater
in
Page
53
of
87
the
CMC­
Na
group
compared
to
the
DEN
animals
during
Study
Week
1,
but
generally,
food
efficiency
was
lower
in
the
KIF­
230
and
CMC­
Na
groups
compared
to
the
DEN
animals.
Additionally,
overall
(
Weeks
0­
10)
mean
food
efficiency
was
lower
in
the
KIF­
230
and
CMC­
Na
animals
compared
to
the
DEN
group.
Absolute
and
relative
(
to
body)
liver
weights
were
increased
in
the
KIF­
230
group
compared
to
the
DEN
group.
Fatty
change
was
observed
in
12/
12
animals
in
all
groups.
Hepatocyte
hypertrophy
was
noted
in
12/
12
DEN
and
KIF­
230
animals,
but
not
in
the
CMC­
Na
group
(
0/
12;
p 
0.05).
Mitosis
was
observed
in
6/
12
animals
in
the
DEN
group
compared
to
0/
12
(
p 
0.01)
KIF­
230
animals
and
1/
12
(
p 
0.05)
CMC­
Na
animals.
Clear
cell
foci
(
3/
12)
and
eosinophilic
cell
foci
(
8/
12)
were
observed
in
the
DEN
group
compared
to
0/
12
animals
in
the
KIF­
230
or
CMC­
Na
groups.
The
number
of
GST­
P
positive
foci/
cm2
was
greater
in
the
DEN
animals
(
23.695)
compared
to
the
KIF­
230
(
0.304;
p 
0.01)
or
CMC­
Na
(
0.473;
p 
0.01)
groups.
The
area
(
mm2/
cm2)
of
the
GST­
P
positive
foci
was
also
greater
in
the
DEN
animals
(
0.647)
compared
to
the
KIF­
230
(
0.002;
p 
0.01)
or
CMC­
Na
(
0.004;
p 
0.01)
groups.

The
data
indicated
that
administration
of
KIF­
230
as
a
single
gavage
dose
of
2000
mg/
kg
to
male
rats
did
not
demonstrate
an
initiator
effect
in
the
liver
when
compared
to
DEN,
a
known
initiating
agent
in
a
two­
stage
experimental
model
of
hepatocarcinogenesis.
However,
an
additional
group
of
saline/
PB­
treated
rats
should
have
been
included
in
this
study
to
allow
full
comparison
of
treated
animals
with
untreated
controls.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.2.
Hirouchi,
Y.
(
2000)
KIF­
230
technical:
two
stage
hepatocarcinogenicity
study
using
rats
for
examining
the
promotor
effect.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4905
(
001­
260),
November
8,
2000.
MRID
45835021.
Unpublished.

EXECUTIVE
SUMMARY:
At
the
start
of
this
non­
guideline
study
(
MRID
45835021),
male
Fischer
(
F344/
DuCrj
[
SPF])
rats
were
placed
in
5
groups
containing
12
animals
each.
Groups
1,
3,
and
5
were
initiated
by
i.
p.
injection
with
200
mg/
kg
N­
Nitrosodiethylnitrosamine
(
DEN)
in
0.9%
saline,
and
rats
in
Groups
2
and
4
were
dosed
with
0.9%
saline
vehicle
only.
From
Study
Week
2
through
Week
8,
rats
were
fed
basal
diet
(
Group
1),
or
diets
with
either
10,000
ppm
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158;
equivalent
to
586.5­
634.1
mg/
kg/
day)
(
Groups
2
and
3)
or
0.05%
(
w/
w)
phenobarbital
sodium
(
PB)
(
Groups
4
and
5).
A
b
partial
hepatectomy
was
performed
in
all
groups
on
Week
3
to
stimulate
hepatic
cell
division.
Eight
weeks
after
initiation,
the
rats
were
killed,
and
the
livers
were
examined
immunohistochemically
for
the
presence
of
placental
glutathione­
S­
transferase
(
GST­
P)
positive
cell
foci.
The
purpose
of
this
study
was
to
examine
the
potential
for
benthiavalicarb
to
act
as
a
hepatic
tumor
promotor
by
using
an
established
two­
stage
model
of
hepatocarcinogenesis.

No
treatment­
related
effects
were
observed
on
mortality
or
clinical
signs
of
toxicity.
Minor
decreases
in
body
weights,
cumulative
(
Weeks
0­
8)
body
weight
gains,
food
consumption,
and
food
efficiency
were
observed,
but
these
were
considered
to
have
resulted
from
the
administration
of
DEN
or
the
partial
hepatectomy
that
was
performed
on
all
animals,
and
not
as
a
consequence
of
exposure
to
the
test
compound.
Page
54
of
87
Absolute
and
relative
(
to
body)
liver
weights
were
increased
in
rats
dosed
with
the
test
compound
(
Groups
2
and
3)
and
in
rats
dosed
with
PB
(
Groups
4
and
5),
compared
to
rats
only
initiated
with
DEN
(
Group
1).
Enlarged
liver
was
observed
in
11/
12
rats
dosed
with
saline/
PB
(
Group
4)
and
12/
12
rats
dosed
with
saline/
KIF­
230,
DEN/
KIF­
230,
and
DEN/
PB
(
Groups
2,
3,
and
5),
but
was
not
noted
in
animals
only
initiated
with
DEN
(
Group
1).
Increased
(
p 
0.01)
incidence
of
fatty
change
was
observed
in
12/
12
animals
dosed
with
PB
(
Groups
4
and
5)
compared
to
0/
12
animals
only
initiated
with
DEN
(
Group
1).
Hepatocyte
hypertrophy
was
noted
in
all
(
12/
12)
animals
dosed
with
either
KIF­
230
(
Groups
2
and
3)
or
PB
(
Groups
4
and
5)
compared
to
0/
12
animals
only
initiated
with
DEN
(
Group
1).
Increased
(
p 
0.05)
mitosis
was
observed
in
10/
12
animals
dosed
with
DEN/
KIF­
230
(
Group
3)
and
in
12/
12
animals
dosed
with
DEN/
PB
(
Group
5),
compared
to
4/
12
animals
only
initiated
with
DEN
(
Group
1).
Increased
(
p 
0.05)
incidence
of
acidophilic
cell
focus
was
noted
in
10/
12
animals
dosed
with
DEN/
KIF­
230
(
Group
3)
and
in
9/
12
animals
dosed
with
DEN/
PB
(
Group
5),
compared
to
4/
12
animals
only
initiated
with
DEN
(
Group
1).
The
number
of
GST­
P
positive
foci
was
increased
(
p 
0.05)
in
animals
treated
with
DEN/
KIF­
230
and
DEN/
PB
(
Groups
3
and
5)
compared
to
animals
only
initiated
with
DEN
(
Group
1).
The
area
of
the
GST­
P
positive
foci
was
also
increased
in
the
DEN/
KIF­
230­
(
Group
3;
not
significant)
and
DEN/
PB­
treated
(
Group
5;
p 
0.05)
animals
compared
to
animals
only
initiated
with
DEN
(
Group
5).
Animals
that
did
not
receive
DEN
(
Groups
2
and
4)
demonstrated
small
numbers
of
GST­
P
positive
foci
that
had
small
areas.

The
data
indicate
that
administration
of
benthiavalicarb
in
the
diet
at
a
dose
of
10,000
ppm
to
male
rats
caused
a
promoter
effect
in
the
liver
similar
to
that
caused
by
phenobarbital,
a
known
tumor
promoter
in
rat
liver
in
a
two­
stage
experimental
model
of
hepatocarcinogenesis.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.3.
Nakajima,
M.
(
2000)
KIF­
230
technical:
two­
stage
transformation
assay
on
BALB/
c
3T3
cells.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4909
(
001­
262),
August
1,
2000.
MRID
45835103.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835103),
BALB/
c
3T3
A31­
1­
1
cells
(
mouse)
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
DMSO
in
a
two­
stage
transformation
assay.
To
assess
initiation
potential,
cells
were
exposed
to
the
test
compound
at
concentrations
of
0,
10.4,
17.3,
28.8,
48.0,
and
80.0
µ
g/
mL
for
three
days.
To
assess
promoter
activity,
cells
were
exposed
to
the
test
compound
at
concentrations
of
0,
3.0,
6.0,
9.0,
12.0,
and
15.0
µ
g/
mL
for
three
days.
The
purpose
of
this
study
was
to
assess
the
potential
of
the
test
compound
to
act
as
an
initiator
or
promoter
in
vitro.

For
the
initiator
studies,
BALB/
c
3T3
cells
were
seeded
in
culture
dishes
and
incubated
for
24
h.
Solvent,
test
compound
solutions,
or
3­
methylcholanthrene
(
MCA,
the
initiator
positive
control)
were
added,
and
the
dishes
were
then
incubated
for
three
days.
The
culture
medium
was
removed
and
replaced,
and
the
cells
were
incubated
for
three
more
days.
The
medium
was
again
removed
and
replaced
with
medium
containing
either
solvent
or
12­
O­
tetradecanoyl
13
acetate
(
TPA,
the
promoter
positive
control),
and
the
cells
were
incubated
for
3­
4
days.
This
incubation
was
repeated
three
times
for
a
total
incubation
time
of
11
days.
The
cells
were
then
cultured
twice
for
3­
4
days
each,
then
fixed
in
methanol,
stained
with
Giemsa
solution,
rinsed
with
water,
and
Page
55
of
87
allowed
to
dry.

For
the
promoter
studies,
BALB/
c
3T3
cells
were
seeded
in
culture
dishes
and
incubated
for
24
hours.
Solvent
or
MCA
was
added,
and
the
dishes
were
then
incubated
for
three
days.
The
culture
medium
was
removed
and
replaced,
and
the
cells
were
incubated
for
three
more
days.
The
medium
was
again
removed
and
replaced
with
medium
containing
either
solvent,
test
compound
solutions,
or
TPA,
and
the
cells
were
incubated
for
3­
4
days.
This
incubation
was
repeated
three
times
for
a
total
incubation
time
of
11
days.
The
cells
were
then
cultured
twice
for
3­
4
days
each,
then
fixed
in
methanol,
stained
with
Giemsa
solution,
rinsed
with
water,
and
allowed
to
dry.

The
test
compound
did
not
initiate
formation
of
foci
or
promote
formation
of
foci
in
cells
initiated
with
0.2
µ
g/
mL
MCA
at
any
dose
level.
In
the
initiation
studies,
one
of
10
plates
exposed
to
28.8
µ
g/
mL
of
the
test
compound
was
observed
to
have
one
focus
of
cellular
growth,
as
did
one
solvent
control
plate.
In
contrast,
eight
of
10
plates
exposed
to
0.2
µ
g/
mL
MCA
exhibited
foci
(
p 
0.01)
with
a
total
of
14
foci
noted.
The
cell
survival
rates
were
 
40%
for
the
highest
dose
and
 
80%
for
the
lowest
dose.
In
the
promoter
studies,
four
of
10
plates
exposed
to
3.0
µ
g/
mL
of
the
test
compound
were
observed
to
have
eight
foci;
four
of
10
plates
exposed
to
6.0
µ
g/
mL
of
the
test
compound
were
observed
to
have
five
foci;
and
two
of
10
plates
exposed
to
9.0
µ
g/
mL
of
the
test
compound
were
observed
to
have
three
foci.
In
contrast,
eight
of
10
plates
initiated
with
0.2
µ
g/
mL
MCA
and
promoted
with
0.1
µ
g/
mL
TPA
exhibited
foci
(
p 
0.01)
with
a
total
of
14
foci
noted.
The
cell
survival
rates
were
 
90%
for
the
three
lowest
doses.

In
summary,
the
data
indicate
that
benthiavalicarb
did
not
act
as
an
initiating
or
promoting
agent
under
the
conditions
of
this
in
vitro
study.
The
methodologies
and
controls
used
were
appropriate.

This
study
is
classified
as
acceptable/
non­
guideline.

2.6.4.
Inagaki,
S.
(
2001)
Oxidative
DNA
damage
study
of
KIF­
230
in
liver
of
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5433
(
001­
284),
May
11,
2001.
MRID
45869716.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45869716),
5
F344/
DuCrj
rats/
sex/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
the
diet
at
concentrations
of
0,
200,
or
10,000
ppm
(
equivalent
to
0/
0,
17.4/
17.1,
and
797.5/
914.5
mg/
kg/
day
[
M/
F])
for
two
weeks.
The
animals
were
killed
at
the
end
of
Week
2;
the
liver
was
removed
and
weighed;
and
the
levels
of
8­
hydroxy­
2'­
deoxyguanosine
(
8­
OHdG)
was
measured
in
hepatic
DNA.
The
purpose
of
this
study
was
to
assess
the
potential
for
the
test
compound
to
induce
oxidative
DNA
damage
in
the
liver
by
measuring
the
levels
of
8­
OHdG.

At
10,000
ppm,
absolute
and
relative
(
to
body)
liver
weights
were
increased
( 
73­
113%;
p 
0.01)
in
both
sexes
compared
to
controls.
At
200
ppm,
a
minor
increase
in
relative
liver
weight
was
noted
in
the
females
( 
9%;
p 
0.01).
Page
56
of
87
8­
OHdG
levels
were
decreased
(
p 
0.05)
in
the
200
and
10,000
ppm
males
( 
20%
each).

No
effects
of
treatment
were
observed
on
mortality
or
clinical
signs
of
toxicity.

Body
weights
and
cumulative
(
Weeks
0­
2)
body
weight
gains
were
decreased
( 
12­
50%;
p 
0.01)
in
the
10,000
ppm
males.
Additionally,
decreases
(
p 
0.01)
in
food
consumption
during
Week
1
( 
31%)
and
total
(
Week
0­
2)
food
consumption
( 
20%)
were
also
observed.

The
data
indicate
that
administration
of
benthiavalicarb
in
the
diet
at
doses
of
up
to
10,000
ppm
(
equivalent
to
797.5/
914.5
mg/
kg/
day
in
males/
females)
did
not
induce
oxidative
hepatic
DNA
damage,
as
measured
by
8­
OHdG
production.
The
increased
absolute
and
relative
(
to
body)
liver
weights
observed
in
the
current
study
are
consistent
with
hepatic
enlargement
associated
with
enzyme
induction
as
described
in
a
concurrently
submitted
special
mechanistic
study
(
MRID
45835101).

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.5.
Inagaki,
S.
(
2001)
Oxidative
DNA
damage
study
of
KIF­
230
in
liver
of
mice.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5434
(
001­
285),
May
11,
2001.
MRID
45835022.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835022),
5
Scl:
B6C3F1
(
C57Bl/
6
x
C3H)
[
SPF]
mice/
sex/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
the
diet
at
concentrations
of
0,
100,
or
5000
ppm
(
equivalent
to
0/
0,
19.4/
26.1,
and
1031.2/
1203.7
mg/
kg/
day
male/
female)
for
up
to
two
weeks.
The
animals
were
killed
at
the
end
of
Week
2;
the
liver
was
weighed;
and
the
presence
of
8­
hydroxy­
2'­
deoxyguanosine
(
8­
OHdG)
was
measured
in
hepatic
DNA.
The
purpose
of
this
study
was
to
assess
the
potential
for
the
test
compound
to
cause
oxidative
DNA
damage
in
the
liver
after
administration
in
the
diet
for
up
to
two
weeks.

No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
food
consumption.

At
5000
ppm,
absolute
and
relative
(
to
body)
liver
weights
were
increased
(
incr.
75­
78%;
p 
0.05).
No
effects
of
treatment
were
observed
on
8­
OHdG
levels
measured
in
liver
tissue
DNA.

No
effects
of
treatment
were
noted
at
100
ppm.

In
summary,
the
data
indicated
that
administration
of
benthiavalicarb
in
the
diet
at
a
dose
of
5000
ppm
resulted
in
increased
absolute
and
relative
(
to
body)
liver
weights.
These
increases
were
consistent
with
hepatic
enlargement
associated
with
enzyme
induction
as
described
in
a
concurrently
submitted
special
mechanistic
study
(
MRID
45835102).
There
was
no
indication
of
oxidative
hepatic
DNA
damage,
as
measured
by
8­
OHdG
production.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.
Page
57
of
87
2.6.6.
Cellular
Proliferation
in
Rats
Murata,
K.
(
2001)
KIF­
230
technical:
a
study
on
the
induction
of
drug­
metabolic
enzyme
and
proliferation
of
hepatocytes
in
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4900
(
001­
259),
March
1,
2001.
MRID
45835101.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835101),
8
Fischer
(
F344/
DuCrj)
[
SPF]
rats/
sex/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G51­
08­
158)
daily
by
oral
gavage
at
concentrations
of
0,
10,
or
1000
mg/
kg/
day
in
a
dose
volume
of
10
mL/
kg
for
up
to
seven
days.
After
dosing
on
Day
7,
all
animals
were
fasted
for
16
hours.
Five
rats/
sex/
dose
were
killed,
the
liver
removed
and
weighed,
and
hepatic
microsomes
were
prepared.
The
total
hepatic
P­
450
levels
and
specific
P­
450
isoenzyme
levels
were
then
determined.
The
remaining
three
rats/
sex/
dose
were
given
a
single
intraperitoneal
injection
of
5­
bromo­
2'­
deoxyuridine
(
BrdU)
one
hour
before
termination,
and
then
were
killed,
necropsied,
and
evaluated
for
hepatocyte
proliferation.
The
purpose
of
this
study
was
to
examine
whether
repeated
administration
of
KIF­
230
Technical
caused
induction
of
microsomal
cytochrome
P­
450
drug­
metabolizing
enzymes
in
the
liver
and/
or
affected
hepatocyte
proliferation
as
measured
by
BrdU
incorporation.

No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
gross
or
microscopic
pathological
examinations,
or
on
the
hepatic
BrdU
labeling
index.

At
1000
mg/
kg,
absolute
and
relative
(
to
body)
liver
weights
were
increased
in
all
animals
(
incr.
8­
22%;
p 
0.05).
In
the
males,
total
hepatic
microsomal
P­
450
levels
were
increased
(
incr.
18%;
p 
0.01),
and
increases
(
p 
0.05)
were
observed
in
CYP1A1
(
1A2)
(
incr.
160%),
CYP2B1
(
2B2)
(
incr.
158%),
and
CYP3A2
(
incr.
101%).
In
the
females,
increases
were
noted
in
CYP1A1
(
1A2)
(
incr.
97%;
not
significant
[
NS]),
CYP2B1
(
2B2)
(
incr.
76%;
NS)
and
CYP3A2
(
incr.
130%;
p 
0.01).
Levels
of
CYP2E1
and
CYP4A1
were
similar
to
controls
in
both
sexes.

No
treatment­
related
effects
were
noted
in
the
10
mg/
kg
animals.

In
summary,
the
data
indicated
that
administration
of
benthiavalicarb
to
rats
by
daily
oral
gavage
at
a
dose
of
1000
mg/
kg
resulted
in
increased
absolute
and
relative
(
to
body)
liver
weights,
increased
total
hepatic
P­
450,
and
induction
of
several
specific
CYP
isoenzymes.
BrdU
labeling
index
indicated
there
was
no
treatment­
related
effect
on
hepatocyte
proliferation.
The
level
of
P­
450
induction
in
males
was
higher
than
in
females;
however,
the
reviewers
do
not
consider
the
magnitude
of
difference
great
enough
to
indicate
a
clear
sex
difference.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.7.
Murata,
K.
(
2001)
KIF­
230
technical:
a
study
on
the
induction
of
drug­
metabolic
enzyme
and
proliferation
of
hepatocytes
in
mice.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
4899
(
001­
258),
March
1,
2001.
MRID
45835102.
Unpublished.
Page
58
of
87
EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835102),
8
B6C3F1
(
C57BL/
6xC3H)
[
SPF]
mice/
sex/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G51­
08­
158)
daily
by
oral
gavage
at
concentrations
of
0,
10,
or
1000
mg/
kg/
day
in
a
dose
volume
of
10
mL/
kg
for
up
to
seven
days.
After
dosing
on
Day
7,
all
animals
were
fasted
for
16
hours.
Five
mice/
sex/
dose
were
killed,
the
liver
removed
and
weighed,
and
hepatic
microsomes
were
prepared.
The
total
hepatic
P­
450
levels
and
specific
P­
450
isoenzyme
levels
were
then
determined.
The
remaining
three
mice/
sex/
dose
were
given
a
single
intraperitoneal
injection
of
5­
bromo­
2'­
deoxyuridine
(
BrdU)
one
hour
before
termination,
and
then
were
killed,
necropsied,
and
evaluated
for
hepatocyte
proliferation.
The
purpose
of
this
study
was
to
examine
whether
repeated
administration
of
KIF­
230
technical
caused
induction
of
microsomal
cytochrome
P­
450
drug­
metabolizing
enzymes
in
the
liver
and/
or
affected
hepatocyte
proliferation
as
measured
by
BrdU
incorporation.

No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
on
the
BrdU
labeling
index.

At
1000
mg/
kg,
absolute
and
relative
(
to
body)
liver
weights
were
increased
in
all
animals
(
incr.
13­
31%;
p 
0.05;
not
significant
[
NS]
in
all
females).
At
necropsy,
enlarged
liver
was
observed
in
1/
3
males
compared
to
0/
3
controls.
Microscopic
examination
revealed
hypertrophy
of
the
hepatocytes
in
the
liver
of
the
males
(
3/
3;
p 
0.05)
and
females
(
2/
3;
NS),
both
compared
to
0
controls.
Total
microsomal
P­
450
levels
were
increased
in
both
sexes
(
incr.
71­
92%;
p 
0.01),
and
increases
(
p 
0.05)
were
observed
in
CYP1A2
(
incr.
63­
149%),
CYP2B1
(
2B2)
(
incr.
180­
553%),
and
CYP3A2
(
incr.
104­
172%).
Levels
of
CYP1A1,
CYP2E1,
and
CYP4A1
were
similar
to
controls
in
both
sexes.

In
summary,
the
data
indicate
that
administration
of
benthiavalicarb
to
mice
by
daily
oral
gavage
at
a
dose
of
1000
mg/
kg
results
in
increased
absolute
and
relative
(
to
body)
liver
weights
with
hypertrophy
of
the
hepatocytes.
However,
the
BrdU
labeling
index
indicated
there
was
no
proliferation
of
hepatocytes
resulting
from
treatment
with
the
test
compound.
There
was
induction
of
several
specific
CYP
isoenzymes
along
with
increased
total
P­
450
levels.
The
level
of
induction
of
the
CYP
isoenzymes
observed
was
low
compared
to
the
induction
seen
with
classical
inducing
agents
like
phenobarbital.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.8.
Inagaki,
S.
(
2002)
KIF­
230
technical:
mechanism
study
of
potential
effects
on
the
thyroid
gland
in
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5903
(
001­
323),
March
1,
2002.
MRID
45835018.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835018),
12
F344/
DuCrj[
SPF]
male
rats/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.6­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
the
diet
at
concentrations
of
0,
200,
or
10,000
ppm
(
equivalent
to
0,
13.3,
and
661.4
mg/
kg/
day)
for
up
to
14
days.
Total
and
free
triiodothyronine
(
T3)
and
thyroxine
(
T4),
and
thyroid
stimulating
hormone
(
TSH)
levels
were
measured
in
serum
on
Days
7
and
14.
The
animals
were
sacrificed
and
necropsied
on
Day
14,
the
thyroid
gland
and
liver
were
weighed,
and
hepatic
microsomes
were
prepared.
Uridine
diphosphate­
glucuronosyltransferase
(
UDP­
GT)
Page
59
of
87
activity
on
T4
in
hepatic
microsomes
was
then
determined.
The
purpose
of
this
study
was
to
examine
the
mechanism
of
hyperthyroidism
observed
in
treated
male
rats
by
measuring
TSH,
T3,
and
T4
levels,
and
microsomal
UDP­
GT
activity
after
administration
of
the
test
compound
in
the
diet
for
up
to
14
days.

No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
cumulative
body
weight
gains,
or
food
consumption.

At
10,000
ppm,
on
Days
7
and
14
decreases
(
p 
0.01)
in
both
total
and
free
T4
( 
11­
18%)
were
observed,
while
increases
(
not
significant)
in
TSH
were
noted
( 
19­
29%).
Additionally
at
this
dose,
an
increase
(
p 
0.01)
in
hepatic
microsomal
UDP­
GT
activity
on
T4
was
observed
( 
16%).
Absolute
and
relative
(
to
body)
liver
weights
were
increased
( 
22­
24%;
p 
0.01),
and
macroscopic
examination
revealed
enlarged
liver
in
all
rats.

No
effects
of
treatment
were
observed
at
200
ppm.

In
summary,
the
data
indicated
that
administration
of
benthiavalicarb
in
the
diet
at
a
dose
of
10,000
ppm
resulted
in
decreased
serum
T4
levels
in
male
rats.
The
increased
absolute
and
relative
(
to
body)
liver
weights
and
increased
size
noted
during
macroscopic
examination
are
consistent
with
hepatic
enlargement
associated
with
enzyme
induction.
The
observed
increased
UDP­
GT
activity
on
T4
could
cause
a
decrease
in
circulating
T4
levels,
resulting
in
increased
serum
TSH
levels
via
a
feedback
mechanism.
This
mechanism
could
ultimately
cause
hyperthyroidism
following
a
sufficient
duration
of
exposure.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.9.
Inagaki,
S.
(
2002)
KIF­
230
technical:
mechanism
study
of
thyroid
gland
tumors
in
mice.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5904
(
001­
324),
March
1,
2002.
MRID
45835019.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835019),
12
Slc:
B6C3F1
(
C57Bl/
6
x
C3H)
[
SPF]
male
mice/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.6­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
the
diet
at
concentrations
of
0,
100,
or
5000
ppm
(
equivalent
to
0,
17.0,
and
855.0
mg/
kg/
day)
for
up
to
14
days.
An
additional
12
mice/
dose
were
treated
with
the
test
compound
as
above
for
measurement
of
thyroid
stimulating
hormone
(
TSH),
and
it
was
stated
that
anti­
rat
TSH
antibody
was
used
instead
of
anti­
mouse
TSH
antibody,
as
the
antimouse
antibody
was
not
available.
Total
and
free
triiodothyronine
(
T3)
and
thyroxine
(
T4),
and
TSH
levels
were
measured
in
serum
on
Days
7
and
14.
The
animals
were
sacrificed
and
necropsied
on
Days
7
and
14,
and
the
thyroid
gland
and
liver
were
weighed.
Hepatic
microsomes
were
prepared
only
on
Day
14.
Uridine
diphosphate­
glucuronosyltransferase
(
UDP­
GT)
activity
on
T4
in
hepatic
microsomes
was
then
determined.
The
purpose
of
this
study
was
to
examine
the
mechanism
of
oncogenicity
for
thyroid
gland
tumors
observed
in
male
mice
by
measuring
TSH,
T3,
and
T4
levels
and
UDP­
GT
activity
after
administration
of
the
test
compound
in
the
diet
for
up
to
14
days.

No
effects
of
treatment
were
observed
on
mortality,
clinical
observations,
body
weights,
Page
60
of
87
cumulative
body
weight
gains,
or
food
consumption.

At
5000
ppm,
hepatic
microsomal
UDP­
GT
activity
on
T4
was
increased
(
incr.
65%;
p 
0.01)
on
Day
14.
Additionally
at
this
dose,
decreases
(
p 
0.05)
in
both
total
and
free
T4
(
decr.
25­
29%)
were
observed
on
Days
7
and
14.
Absolute
and
relative
(
to
body)
liver
weights
were
increased
(
incr.
53­
71%;
p 
0.01)
on
Days
7
and
14,
and
macroscopic
examination
on
Days
7
and
14
revealed
enlarged,
dark
liver
in
5­
6/
6
treated
mice
compared
to
0/
6
controls.

No
effects
of
treatment
were
noted
at
100
ppm.

In
summary,
the
data
indicated
that
administration
of
benthiavalicarb
in
the
diet
at
a
dose
of
5000
ppm
resulted
in
decreased
serum
T4
levels
in
male
mice.
The
increased
absolute
and
relative
(
to
body)
liver
weights
and
increased
size
noted
during
macroscopic
examination
are
consistent
with
hepatic
enlargement
associated
with
enzyme
induction.
The
observed
increased
UDP­
GT
activity
on
T4
could
cause
a
decrease
in
circulating
T4
levels.
Typical
UDP­
GT
inducers
are
known
to
cause
hyperthyroidism
by
inducing
TSH
through
a
decrease
in
serum
thyroid
hormone
levels
in
mice,
and
thyroid
hyperplasia
is
a
potential
mechanism
for
the
induction
of
thyroid
tumors.
However,
thyroid
hyperplasia
in
rodents
is
driven
by
an
increase
in
circulating
TSH
levels;
in
this
study,
TSH
levels
were
unaffected
by
treatment.
The
lack
of
an
appropriate
antibody
(
anti­
mouse
TSH)
could
cause
inaccurate
measurements
of
TSH
in
the
radioimmunoassay
used
to
determine
serum
TSH
levels,
and
no
information
was
provided
concerning
the
cross­
reactivity
of
the
anti­
rat
TSH
antibody
used.
While
the
mechanism
suggested
is
plausible,
it
is
the
opinion
of
the
reviewers
that
no
definitive
conclusions
can
be
reached
concerning
this
mechanistic
study.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.10.
Inagaki,
S.
(
2002)
KIF­
230
technical:
mechanism
study
of
uterine
cancer
in
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5914
(
001­
325),
March
1,
2002.
MRID
45835020.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45835020),
12
F344/
DuCrj
[
SPF]
female
rats/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.6­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
the
diet
at
concentrations
of
0,
200,
or
10,000
ppm
(
equivalent
to
0,
11.6,
and
576.4
mg/
kg/
day)
for
up
to
8
weeks.
Microsomal
aromatase
activity
was
measured
at
Week
8
using
1 ­
[
3H]­
androstenedione
as
a
substrate,
and
serum
estradiol
(
E2),
progesterone
(
P4),
and
luteinizing
hormone
(
LH)
levels
were
measured
at
Weeks
2,
4,
6,
and
8.
The
purpose
of
this
study
was
to
examine
the
mechanism
of
oncogenicity
for
uterine
cancer
by
measuring
aromatase
activity
in
liver,
ovaries,
and
uterus,
and
serum
levels
of
E2,
P4,
and
LH
after
administration
of
the
test
compound
in
the
diet
for
up
to
8
weeks.
Increased
aromatase
activity
could
result
in
increased
E2
levels,
which
by
acting
in
either
an
autocrine
or
paracrine
manner,
could
drive
the
growth
of
female
reproductive
tissues
such
as
the
ovary
and
uterus
that
express
estrogen
receptors.

No
treatment­
related
effects
were
observed
on
mortality,
clinical
signs
of
toxicity,
body
weight,
cumulative
(
Weeks
­
1
to
8)
body
weight
gains,
or
food
consumption.

No
effects
of
treatment
were
observed
at
200
ppm.
Page
61
of
87
At
10,000
ppm,
microsomal
aromatase
activity
was
increased
in
the
liver
( 
36%;
p 
0.01),
slightly
increased
in
the
uterus
(
incr.
15%;
not
significant
[
NS]),
but
was
decreased
in
the
ovary
( 
10%;
NS).
No
treatment­
related
effects
were
observed
on
serum
sex
hormone
levels.
Ovary
and
uterus
weights
were
unaffected
by
treatment,
and
no
effects
were
noted
during
necropsy
on
the
ovary
or
uterus.
Additionally
at
this
dose,
absolute
and
relative
(
to
body)
liver
weights
were
increased
( 
28­
31%;
p 
0.01),
and
macroscopic
examination
revealed
dark
liver
in
10/
11
rats
and
enlarged
liver
in
11/
11
rats
compared
to
0/
10
controls.

In
summary,
the
data
indicate
that
administration
of
benthiavalicarb
in
the
diet
at
a
dose
of
10,000
ppm
does
cause
increased
absolute
and
relative
liver
weights,
but
does
not
appreciably
alter
aromatase
activity
in
organs
other
than
liver.
The
increased
liver
weights
and
increased
size
noted
during
macroscopic
examination
are
consistent
with
hepatic
enlargement
associated
with
enzyme
induction.
Hepatic
enzyme
induction
could
also
result
in
an
upregulation
of
hepatic
aromatase,
but
this
increase
was
insufficient
to
increase
circulating
E2
levels
in
this
study.
The
investigator
stated
that
the
lack
of
increased
aromatase
activity
in
the
ovary
and
uterus
suggested
little
direct
participation
of
E2
in
the
development
of
uterine
cancer
by
KIF­
230.
However,
E2
levels
and
estrogen
receptor
levels
were
not
determined
in
the
specific
tissues.

The
submitted
study
is
classified
as
acceptable/
non­
guideline.

2.6.11.
Itoh,
K.
(
2001)
The
uterotrophic
study
of
KIF­
230
in
ovariectomized
rats.
Biosafety
Research
Center,
Foods,
Drugs,
and
Pesticides
(
An­
Pyo
Center),
Arahama
Shioshinden,
Fukude­
Cho
Iwata­
Gun,
Shizuoka,
Japan.
Laboratory
Project
No.:
5642
(
001­
299),
September
7,
2001.
MRID
45869714.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
non­
guideline
study
(
MRID
45869714),
6
ovariectomized
female
Fischer
(
F344/
DuCrj)
rats/
dose
were
exposed
to
benthiavalicarb
(
KIF­
230;
88.8­
89.1%
a.
i.;
Lot
#:
G­
51­
08­
158)
in
0.5%
(
w/
v)
carboxymethylcellulose
sodium,
1.0%
(
v/
v)
Tween­
80
vehicle
(
CMC­
Na)
by
daily
oral
gavage
at
0,
10,
100,
or
1000
mg/
kg
in
a
dose
volume
of
10
mL/
kg
for
14
days.
An
additional
group
of
6
ovariectomized
females
was
dosed
with
0.001
mg/
kg
 ­
estradiol
(
E2)
in
CMC­
Na
by
oral
gavage
to
serve
as
a
positive
control.
One
hour
before
termination
on
Study
Day
14,
rats
were
given
a
single
intraperitoneal
injection
of
5­
bromo­
2'­
deoxyuridine
(
BrdU).
The
uterus
was
then
removed
and
weighed,
and
examined
microscopically.
Uterine
cellular
proliferation
was
determined
by
BrdU
incorporation.
The
purpose
of
this
study
was
to
examine
potential
estrogenic
effects
of
the
test
compound
on
the
uterus
of
ovariectomized
rats.

No
treatment­
related
effects
were
observed
on
mortality,
clinical
signs
of
toxicity,
body
weight,
overall
(
Study
Days
0­
14)
body
weight
gains,
absolute
or
relative
uterus
weights,
microscopic
uterine
pathology,
or
uterine
BrdU
labeling
index.

Positive
control
animals
dosed
with
E2
demonstrated
increased
(
p 
0.05)
body
weight
on
Days
13­
14
( 
5­
6%);
increased
overall
body
weight
gains
( 
36%;
calculated
by
reviewers)
was
also
observed.
Relative
(
to
body)
blotted
uterus
weight
was
decreased
(
p 
0.05)
in
the
E2­
treated
animals
(
34
mg
%)
compared
to
vehicle
controls
(
39
mg
%).
The
reviewers
consider
this
reduced
Page
62
of
87
organ
weight
to
be
a
reflection
of
the
increased
body
weight.

The
submitted
study
is
classified
as
unacceptable/
non­
guideline.
Although
the
animals
were
dosed
to
the
limit
dose
(
1000
mg/
kg/
day),
the
lack
of
a
uterotrophic
response
in
the
positive
control
animals
makes
it
impossible
to
determine
if
the
lack
of
response
in
test
compound­
treated
animals
is
a
true
response
or
due
to
an
error
in
experimental
design.

2.7.
Mutagenicity
Studies
2.7.1.
Riley,
S.
(
2000).
KIF­
230
(
TGAI):
Induction
of
Micronuclei
in
the
Bone
Marrow
of
Treated
Mice,
performed
at
Covance
Laboratories,
Ltd.,
North
Yorkshire,
England.
Report
No.:
535/
47­
D5140,
dated
January
11,
2000.
MRID
45835105.
Unpublished.

EXECUTIVE
SUMMARY:
In
an
in
vivo
mammalian
cytogenetic
(
micronucleus)
assay
(
MRID
45835105),
groups
of
eight
male
mice
were
administered
KIF­
230
technical
(
Batch
No.
G51­
15­
162,
87.9%
a.
i.
suspended
in
0.5%
(
w/
v)
carboxymethylcellulose,
CMC),
at
2000
mg/
kg
by
oral
gavage
once
daily
on
two
consecutive
days,
or
CMC
(
negative
control)
on
two
consecutive
days.
Both
groups
were
sacrificed
24
hours
after
the
second
administration
of
the
test
article,
or
the
vehicle.
A
known
mutagen,
cyclophosphamide
(
CPA),
dissolved
in
physiological
saline,
was
administered
to
eight
mice
at
a
single
dose
of
40
mg/
kg,
orally,
and
sacrificed
24
hours
later
(
positive
control).
Following
sacrifice,
bone
marrow
was
withdrawn
from
each
animal,
and
processed
to
demonstrate
micronuclei
in
polychromatic
erythrocytes
(
mPCE).
The
incidence
of
mPCEs
in
KIF­
230
(
TGAI)­
treated
mice
was
compared
to
that
in
vehicle
(
negative)
controls.

The
sponsor
notified
the
performing
laboratory
that
the
test
compound
showed
no
evidence
of
clinical
toxicity
up
to
an
LD50
 
5000
mg/
kg.
Therefore,
the
internationally
accepted
limit
dose
of
2000
mg/
kg/
day
for
in
vivo
studies
was
selected
for
the
main
micronucleus
assay.
Additionally,
the
sponsor
reported
that
there
was
no
substantial
difference
in
clinical
toxicity
between
the
sexes;
accordingly,
the
assay
was
performed
only
with
male
mice.

Bone
marrow
from
KIF­
230­
treated
animals
showed
both:
a
group
mean
ratio
of
PCE
to
normochromatic
erythrocytes
(
NCE)
comparable
to
vehicle
controls
(
indicating
no
erythropoetic
toxicity);
and
an
incidence
of
mPCE,
that
was
not
significantly
different
from
concurrent
vehicle
controls,
or
the
laboratory's
historical
negative
control
range.
Positive
controls
exhibited
markedly
increased
mPCEs,
significantly
greater
than
concurrent
controls.

Therefore,
KIF­
230
(
TGAI)
was
cytogenetically
negative
for
the
induction
of
micronucleated
polychromatic
erythrocytes
in
bone
marrow
of
mice
treated
at
the
limit
dose,
2000
mg/
kg.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vivo
cytogenetic
mutagenicity
data.

2.7.2.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
Test
of
KIF­
230S­
L
with
Bacteria,
performed
in
the
Biosafety
Research
Center,
Food,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experiment
No.
5502
(
001­
292),
June
28,
2001.
MRID
45835106.
Unpublished.
Page
63
of
87
EXECUTIVE
SUMMARY:
In
a
reverse
mutation
assay
in
bacteria
(
MRID
45835106),
four
histidine­
deficient
(
his­)
strains
of
Salmonella
typhimurium
(
TA100,
TA98,
TA1535,
and
TA1537)
and
one
tryptophan­
deficient
(
try­
­)
strain
of
Escherichia
coli
(
WP2
uvrA)
were
exposed
for
48
hours
to
KIF­
230S­
L
(
Lot
No.
9,
100%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO)
at
six
concentrations
ranging
from
156
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
derived
from
induced
hepatic
microsomes
drawn
from
male
rats
treated
intraperitoneally
(
i.
p.)
with
phenobarbital
and
benzoflavone,
plus
generating
co­
factor
(
±
S9­
mix).
In
addition
to
cultures
exposed
to
the
solvent
(
negative
control),
other
cultures
were
treated
with
strain­
specific
mutagens
(
positive
controls).
At
termination
of
incubation,
the
incidence
of
revertant
colonies
(
his+,
try+)
was
compared
to
that
of
the
negative
control.

In
a
"
dose­
finding
study"
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9­
mix,
no
cytotoxicity
was
encountered
in
cultures
treated
up
to
2000
µ
g/
plate,
but
growth
inhibition
was
evident
in
all
strains
at
the
highest
dose,
5000
µ
g/
plate.
Precipitation
of
the
test
compound
was
observed
at
51.2
µ
g/
plate
without
S9­
mix,
and
at
128
µ
g/
plate
under
S9­
activation,
but
at
no
concentration
up
to
the
"
limit
dose"
(
5000
µ
g/
plate)
were
increased
revertant
colonies
observed
over
control
values.

In
the
main
study,
growth
inhibition
occurred
in
all
cultures
treated
with
5000
µ
g/
plate
of
the
test
compound,
and
precipitation
of
the
test
article
was
observed
at
all
concentrations.
However,
at
no
concentration
up
to
5000
µ
g/
plate
were
significant
increases
in
revertants
(
his+,
try+)
found
in
treated
cultures
compared
to
those
in
concurrent
solvent
controls,
or
the
laboratory's
historical
control
data.

Therefore,
the
data
from
both
the
"
dose­
finding
study"
and
the
main
assay
indicated
that
KIF­
230S­
L
did
not
induce
reverse
gene
mutation
in
four
strains
of
S.
typhimurium
and
one
strain
of
E.
coli.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.3.
Saito,
Y.
(
2002).
Identification
of
a
mutagenic
substance
isolated
from
KIF­
230,
performed
at
the
Life
Science
Research
Institute,
Kumiai
Chemical
Industry
Company,
Ltd.,
Shizuoka,
Japan.
Report
No.
202L
S097,
dated
29
March
2002.
MRID
45835107.
Unpublished.

EXECUTIVE
SUMMARY:
As
stated
by
the
investigator,
this
report
was
submitted
"
to
clarify
the
positive
response
induced
by
KIF­
230,
Lot
No.
G51­
15­
162
in
a
reverse
mutation
assay
with
Salmonella
typhimurium
TA98
in
the
presence
of
metabolic
activation,
S9"
(
initially
reported
in
December,
1999
­
see
MRID
45869722).

Ten
lots
of
KIF­
230
technical
[
Lots
G51­
08,
G51­
15,
G51­
35,
G51­
36,
G51­
37,
G51­
47,
G51­
48,
G51­
49,
G51­
50,
G51­
51]
were
fractionated
using
high
performance
liquid
chromatography
(
HPLC)­
external
standard
method
(
Znt
PTS,
or
modified
PTS)
by
the
Research
Laboratory
of
the
Ihari
Chemical
Industry
Co.,
Ltd.,
Shizuoka,
Japan.
Of
the
10
lots
of
technical
KIF­
230
analyzed,
a
"
fraction
1a"
was
detected
in
Lots
G51­
08
and
G51­
15,
containing
respectively
76.6
ppm
and
204.3
ppm
of
the
impurity
(
described
as
a
"
greenish
white
powder"),
which
induced
Page
64
of
87
positive
mutagenicity
in
TA98
+
S9.
The
amount
of
impurity
in
the
other
eight
lots
each
contained
<
5
ppm,
the
limit
of
quantification
(
and
appeared
as
"
white
powders")
negative
for
mutagenicity
in
TA98
+
S9
at
a
concentration
equivalent
to
1000
µ
g/
plate
of
the
technical
grade
of
KIF­
230.
The
mutagenic
impurity,
fraction
Ia,
was
identified
as
6,6'­
difluoro­
2,2'­
dibenzothiazole
by
gas
chromatography­
mass
spectrometry
(
GE­
MS).
The
specific
activity
as
an
index
of
the
mutagenicity
of
1a
was
also
tested
in
this
assay
using
the
TA­
98
strain
of
S.
typhimurium
+
S9,
and
discovered
to
be
equal
to
independently
synthesized
6,6'­
difluoro­
2,2'­
bibenzothiazole
(
I­
6),
as
well
as
to
the
reference
mutagen,
benzo(
a)
pyrene.

The
threshold
of
the
impurity
that
produces
positive
results
was
estimated
as
~
50
ppm.

In
reconstruction
experiments,
when
two
lots
of
the
TGAI
(
Nos.
G15­
35,
G15­
47),
initially
negative
for
reverse
mutation
(
no
increase
in
revertants)
were
mixed
with
200
ppm
I­
6,
positive
mutagenicity
was
obtained
with
both
lots.

Therefore,
the
positive
results
with
Lots
No.
G51­
15­
162
and
G51­
08­
158
in
TA98
in
the
presence
of
S9­
metabolic
activation
previously
reported
appears
to
be
due
to
a
mutagenic
component,
identified
as
6,
6'­
difluoro­
2'­
dibenzothiazole.
This
study
is
classified
as
acceptable/
non­
guideline.

2.7.4.
Mizuhashi,
F.
(
2002).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
G5I­
47­
190)
With
Bacteria,
performed
at
the
Biosafety
Research
Center,
Food,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No:
6239(
001­
351),
dated
March
26,
2002.
MRID
45869717.
Unpublished.

In
a
reverse
mutation
assay
(
MRID
45869717),
cultures
of
the
histidine­
deficient
(
auxotrophic
his
 
)
Salmonella
typhimurium
strain
TA98
were
exposed
by
the
pre­
incubation
procedure
for
48
hours
to
technical
KIF­
230
(
Lot
No.
G5I­
47­
190,
90.5%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
at
5
concentrations
ranging
from
313
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
provided
by
rat
hepatic
microsomes
(
±
S9­
mix).
In
addition
to
cultures
exposed
to
the
solvent,
acting
as
"
negative"
control,
other
cultures
were
treated
with
the
mutagens,
2­(
2­
furyl)­
3­(
5­
nitro­
2­
furyl)
acrylamide
(
AF­
2,
0.1
µ
g/
plate)
and
2­
aminoanthracene
(
2­
AA,
0.5
µ
g/
plate),
to
serve
as
positive
controls,
respectively,
for
the
nonactivated
and
activated
test
series.
At
harvest,
bacterial
growth
(
background
lawn)
was
examined
for
toxicity
using
a
stereo
microscope
at
a
magnification
of
60X,
and
revertant
colonies
(
his
+
)
were
counted
as
a
measure
of
mutation.

Precipitation
of
the
test
article
occurred
at
all
test
concentrations,
increasingly
denser
at
 
625
µ
g/
plate,
obscuring
visibly
accurate
colony
counts,
necessitating
manual
counting.
Scoring
of
revertants
could
be
accomplished
at
the
lowest
test
concentration
employing
an
automatic
colony
counter.

In
a
preliminary
preincubation
dose­
finding
study
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9,
precipitation
of
the
test
article
was
seen
at
 
800
µ
g/
plate
±
S9,
necessitating
manual
counting
at
these
higher
concentrations.
Revertants
at
lower
concentrations
were
scored
by
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect
at
any
solublle
concentration
(
8.19
to
128
µ
g/
plate).
There
was
no
indication
of
a
cytotoxic
or
Page
65
of
87
mutagenic
effect
on
any
soluble
concentration
(
8.19
to
128
µ
g/
plate
±
S9).

The
precipitation
at
all
levels
in
the
main
assay
renders
this
phase
of
testing
invalid,
but
the
dosefinding
study,
which
included
soluble
levels
provided
acceptable
evidence
of
a
negative
response
in
treated
TA98
cultures.

However,
at
no
utilizable
concentration
of
S.
typhimurium
TA98
with
or
without
S9­
activation
in
either
the
dose
finding
study
or
main
assay
did
the
incidence
of
revertants
(
to
prototrophic
his
+
)
differ
significantly
from
negative
control.
By
contrast,
a
marked
increase
of
revertants
was
found
in
both
positive
controls.

Therefore,
the
G5I­
47­
190
lot
of
KIF­
230
(
TGAI)
was
negative
for
inducing
gene
mutations
in
S.
typhimurium
strain
TA98
in
the
presence
or
absence
of
metabolic
activation.

This
study
is
classified
as
acceptable/
non­
guideline
as
only
one
strain
(
TA98)
of
the
customary
battery
of
Salmonella
strains
was
assayed
with
the
test
article.
The
findings
support
the
negative
data
from
an
earlier
bacterial
reverse
gene
mutation
assay
with
100%
of
the
test
material
(
see
MRID
45835106).
It
is
not
upgradable,
and
does
not
fully
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.5.
Mizuhashi,
f.
(
2002).
Reverse
Mutation
Test
of
Kif­
230
TGAI
(
Lot
G51­
48­
190)
With
Bacteria,
performed
at
the
biosafety
Research
Center,
Food,
drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.:
6240(
001­
352).
Dated
March
26,
2002.
MRID
45869718.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
mutation
assay
(
MRID
45869718),
cultures
of
the
histidine­
deficient
(
auxotrophic
his
­)
Salmonella
typhimurium
strain
TA98
were
exposed
by
the
preincubation
procedure
for
48
hours
to
technical
KIF­
230
(
Lot
No.
G51­
48­
190,
92.4%
a.
i.,
dissolved
in
DMSO),
at
five
concentrations
ranging
from
313
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
provided
by
rat
hepatic
microsomes.
In
addition
to
cultures
exposed
to
the
solvent,
acting
as
"
negative"
control,
other
cultures
were
treated
with
the
mutagens,
2­(
2­
furyl)­
3­(
5­
nitro­
2­
furyl)
acrylamide
(
AF­
2,
0.1
µ
g/
plate)
and
2­
aminoanthracene
(
2­
AA,
0.5
µ
g/
plate),
to
serve
as
positive
controls,
respectively,
for
the
nonactivated
and
activated
test
series.
At
harvest,
bacterial
growth
(
background
lawn)
was
examined
for
toxicity
using
a
stereo
microscope
at
a
magnification
of
60X,
and
revertant
colonies
(
his
+)
were
counted
as
a
measure
of
mutation.

Precipitation
of
the
test
article
occurred
at
all
test
concentrations,
increasingly
denser
at
 
625
µ
g/
plate,
obscuring
visibly
accurate
colony
counts,
and
necessitating
manual
counting.
Scoring
of
revertants
could
be
accomplished
at
the
lowest
test
concentration
employing
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.

In
a
preliminary
preincubation
dose­
finding
study
at
8
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9
and
including
positive
and
negative
controls,
precipitation
of
the
test
article
was
seen
at
±
320
µ
g/
plate
±
S9,
necessitating
manual
counting
at
higher
concentrations.
Revertants
at
lower
concentrations
were
scored
by
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.
Page
66
of
87
Precipitation
at
all
concentrations
renders
the
main
assay
invalid,
but
the
data
at
soluble
levels
in
the
"
dose­
finding
study"
are
acceptable.

Therefore,
the
G51­
48­
190
lot
of
KIF­
230
(
TAGAI)
was
negative
for
inducing
gene
mutations
in
S.
typhimurium
strain
TA98
in
the
presence
or
absence
of
metabolic
activation.
This
study
is
classified
as
acceptable/
non­
guideline,
but
only
for
the
one
Salmonella
strain
(
TA98)
of
the
customary
battery
of
Salmonella
strains.
It
is
not
upgradable,
and
does
not
fully
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
Mutagenicity
(
bacterial
reverse
gene
mutation)
data.
However,
the
data
support
the
findings
from
an
acceptable
and
negative
assay
from
a
reverse
gene
mutation
assay
in
bacteria
with
100%
of
the
active
ingredient
(
MRID
45869706).

2.7.6.
Mizuhashi,
F.
(
2002).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
G5I­
49­
190)
With
Bacteria,
performed
at
the
Biosafety
Research
Center,
Food,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No:
6241(
001­
353),
dated
March
26,
2002.
MRID
45869719.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
mutation
assay
(
MRID
45869719),
cultures
of
the
histidine­
deficient
(
auxotrophic
his
­)
Salmonella
typhimurium
strain
TA98
were
exposed
by
the
preincubation
procedure
for
48
hours
to
technical
KIF­
230
(
Lot
No.
G5I­
49­
190,
92.7%
a.
i.),
dissolved
in
dimethyl
sulfoxide
(
DMSO),
at
five
concentrations
ranging
from
3.13
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
provided
by
rat
hepatic
microsomes.
In
addition
to
cultures
exposed
to
the
solvent,
acting
as
"
negative"
control,
other
cultures
were
treated
with
the
mutagens,
2­(
2­
furyl)­
3­(
5­
nitro­
2­
furyl)
acryla­
mide
(
AF­
2,
0.1
g/
plate)
and
2­
aminoanthracene
(
2­
AA,
0.5
µ
g/
plate),
to
serve
as
positive
controls,
respectively,
for
the
non­
activated
and
activated
test
series.
At
harvest,
bacterial
growth
(
background
lawn)
was
examined
for
toxicity
using
a
stereomicroscope
at
a
magnification
of
60X,
and
revertant
colonies
(
his
+
)
were
counted
as
a
measure
of
mutation.

Precipitation
of
the
test
article
occurred
at
all
test
concentrations
( 
3.13
µ
g/
plate
±
S9),
increasingly
denser
at
 
625
µ
g/
plate,
obscuring
visibly
accurate
colony
counts,
necessitating
manual
counting.
Scoring
of
revertants
could
be
accomplished
at
the
lowest
test
concentration
employing
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.

In
a
preliminary
preincubation
dose­
finding
study
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9
and
including
negative
and
positive
controls,
precipitation
of
the
test
article
was
seen
at
 
320
µ
g/
plate
±
S9,
necessitating
manual
counting
at
these
concentrations.
Revertants
at
lower
concentrations
were
scored
by
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.

No
utilizable
concentrations
of
KIF­
230
TGAI
with
or
without
S9­
activation
were
achieved
in
the
main
assay.
However,
the
data
from
the
dose­
finding
study
indicate
that
the
test
material
was
not
mutagenic.
By
contrast,
marked
increases
of
revertants
were
found
in
both
positive
controls.

Therefore,
the
G5I­
49­
190
lot
of
KIF­
230
(
TGAI)
was
negative
for
inducing
gene
mutations
in
Page
67
of
87
S.
typhimurium
strain
TA98
in
the
presence
or
absence
of
metabolic
activation.

This
study
is
classified
as
acceptable/
non­
guideline
as
only
one
Salmonella
strain
(
TA98)
of
the
customary
battery
of
Salmonella
strains
was
assayed
with
the
test
article.
It
is
acceptable
for
TA98
but
does
not
fully
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.
However,
the
data
support
the
findings
of
the
acceptable
and
negative
reverse
gene
mutation
assay
with
100%
of
the
test
material
(
see
MRID
45835106).

2.7.7.
Mizuhashi,
F.
(
2002).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
G5I­
50­
190)
With
Bacteria,
performed
at
the
Biosafety
Research
Center,
Food,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No:
6242(
001­
354),
dated
March
26,
2002.
MRID
45869720.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
mutation
assay
(
MRID
45869720),
cultures
of
the
histidine­
deficient
(
auxotrophic
his
 
)
Salmonella
typhimurium
strain
TA98
were
exposed
by
the
preincubation
procedure
for
48
hours
to
technical
KIF­
230
(
Lot
No.
G5I­
50­
190,
91.7%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
at
five
concentrations
ranging
from
313
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
provided
by
rat
hepatic
microsomes
(
±
S9­
mix)
from
male
Sprague­
Dawley
rats
induced
with
phenobarbital
and
5,6­
benzoflavone.
In
addition
to
cultures
exposed
to
the
solvent,
acting
as
"
negative"
control,
other
cultures
were
treated
with
the
mutagens,
2­(
2­
furyl)­
3­(
5­
nitro­
2­
furyl)
acrylamide
(
AF­
2,
0.1
µ
g/
plate)
and
2­
aminoanthracene
(
2­
AA,
0.5
µ
g/
plate),
to
serve
as
positive
controls,
respectively,
for
the
non­
activated
and
activated
test
series.
At
harvest,
bacterial
growth
(
background
lawn)
was
examined
for
toxicity
using
a
stereomicroscope
at
a
magnification
of
60X,
and
revertant
colonies
(
his
+
)
were
counted
as
a
measure
of
mutation.

Precipitation
of
the
test
article
occurred
at
all
test
concentrations,
increasingly
denser
at
 
625
µ
g/
plate,
obscuring
visibly
accurate
colony
counts,
necessitating
manual
counting.
Scoring
of
revertants
could
be
accomplished
at
the
lowest
test
concentration
employing
an
automatic
colony
counter.
There
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.

In
a
preliminary
preincubation
dose­
finding
study
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9
and
including
negative
and
positive
controls,
precipitation
of
the
test
article
was
seen
at
 
320
µ
g/
plate
±
S9,
necessitating
manual
counting
at
these
concentrations.
Revertants
at
lower
concentrations
were
scored
by
an
automatic
colony
counter.
Again,
there
was
no
indication
of
a
cytotoxic
or
mutagenic
effect.

Precipitation
at
all
concentrations
renders
the
main
assay
invalid,
but
the
dose­
finding
test
at
soluble
concentrations
validates
the
test
material
as
being
non­
mutagenic.

Therefore,
the
G5I­
50­
190
lot
of
KIF­
230
(
TGAI)
was
negative
for
inducing
gene
mutations
in
S.
typhimurium
strain
TA98
in
the
presence
or
absence
of
metabolic
activation.

This
study
is
classified
as
acceptable/
non­
guideline
as
only
one
Salmonella
strain
(
TA98)
of
the
customary
battery
of
Salmonella
strains
was
assayed
with
the
test
article.
It
is
not
upgradable,
and
does
not
fully
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
Page
68
of
87
(
bacterial
reverse
gene
mutation)
data.
However,
the
data
support
the
findings
of
the
acceptable
and
negative
reverse
gene
mutation
assay
with
100%
of
the
active
ingredient
(
MRID
45869706).

2.7.8.
Mizuhashi,
F.
(
2002).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
G5I­
51­
190)
with
Bacteria,
performed
at
the
Biosafety
Research
Center,
Food,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No:
6243(
001­
355),
dated
March
26,
2002.
MRID
45869721.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
mutation
assay
(
MRID
45869721),
cultures
of
the
histidine­
deficient
(
auxotrophic
his
 
)
Salmonella
typhimurium
strain
TA98
were
exposed
by
the
preincubation
procedure
for
48
hours
to
technical
KIF­
230
(
Lot
No.
G5I­
51­
190,
93.3%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
at
five
concentrations
ranging
from
313
to
5000
µ
g/
plate,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
provided
by
hepatic
microsomes
(
±
S9­
mix)
from
male
Sprague­
Dawley
rats
induced
with
phenobarbitol
and
5,6­
benzoflavone.
In
addition
to
cultures
exposed
to
the
solvent,
acting
as
"
negative"
control,
other
cultures
were
treated
with
the
mutagens,
2­(
2­
furyl)­
3­(
5­
nitro­
2­
furyl)
acrylamide
(
AF­
2,
0.1
µ
g/
plate)
and
2­
aminoanthracene
(
2­
AA,
0.5
µ
g/
plate),
to
serve
as
positive
controls,
respectively,
for
the
non­
activated
and
activated
test
series.
At
harvest,
bacterial
growth
(
background
lawn)
was
examined
for
toxicity
using
a
stereo
microscope
at
a
magnification
of
60X,
and
revertant
colonies
(
his
+
)
were
counted
as
a
measure
of
mutation.

Precipitation
of
the
test
article
occurred
at
all
test
concentrations
in
the
main
assay
( 
313
µ
g/
plate
±
S9),
increasingly
denser
at
 
1250
µ
g/
plate,
obscuring
visibly
accurate
colony
counts,
necessitating
manual
counting.
Scoring
of
revertants
could
be
accomplished
at
the
lower
test
concentrations
employing
an
automatic
colony
counter.

In
a
preliminary
preincubation
dose­
finding
study
in
TA100
only
at
five
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9
and
including
positive
and
negative
controls,
toxicity
(
reduction
in
revertants)
was
seen
at
the
maximum
concentration.
There
was
no
indication
of
a
mutagenic
effect.

No
utilizable
concentrations
of
S.
typhimurium
TA98
with
or
without
S9­
activation
were
achieved
in
the
main
assay,
and
as
with
the
dose­
finding
study,
there
were
no
increased
incidences
of
revertants
(
to
prototrophic
his+)
colonies.
By
contrast,
marked
increases
of
revertants
were
found
in
both
positive
controls.

Therefore,
the
G5I­
51­
190
lot
of
KIF­
230
(
TGAI)
was
negative
for
inducing
gene
mutations
in
S.
typhimurium
strain
TA98
in
the
presence
or
absence
of
metabolic
activation.

This
study
is
classified
as
acceptable
but
non­
guideline.
Whereas
only
one
Salmonella
strain
(
TA98)
of
the
customary
battery
of
Salmonella
strains
was
assayed,
the
data
do
support
the
findings
of
the
acceptable
and
negative
gene
mutation
assay
with
100%
of
the
active
ingredient
(
MRID
45869706).
It
does
not
fully
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.9.
Dawkes,
N.
(
1999).
KIF­
230
(
TGAI):
Reverse
mutation
in
Four
Histidine­
requiring
Strains
of
Salmonella
typhimurium
and
One
Tryptophan­
requiring
Strain
of
Escherichia
coli,
Page
69
of
87
performed
at
Covance
Laboratories,
Limited,
North
Yorkshire,
England.
Report
No.
535/
44­
D5140,
dated
December,
1999.
MRID
No.
45869722.
Unpublished.

EXECUTIVE
SUMMARY:
In
independent
reverse
mutation
assays
in
bacteria
(
MRID
45869722),
cultures
of
four
histidine­
deficient
(
auxotrophic
his
­
)
strains
of
S.
typhimurium
(
TA1535,
TA1537,
TA98
and
TA100)
and
the
tryptophan­
deficient
(
auxotrophic
try
­
)
WP2
uvrA
strain
of
Escherichia
coli
were
exposed
by
the
preincubation
procedure
for
72
hours
to
KIF­
230
TGAI
(
Batch
No.
G51­
15­
162,
87.9%
a.
i.,
dissolved
in
dimethyl
formamide,
DMF)
at
five
concentrations
ranging
from
8
to
5000
µ
g/
plate
in
the
first
experiment;
at
seven
concentrations
ranging
from
32
to
5000
µ
g/
plate
in
the
second
experiment,
both
trials
in
the
presence
(+
S9)
and
absence
(­
S9)
of
hepatic
microsomes
isolated
from
male
Sprague­
Dawley
rats
injected
intraperitoneally
with
Aroclor
1254,
plus
generating
co­
factors
(
S9­
mix).
In
addition
to
bacterial
cultures
exposed
to
the
solvent
(=
negative
control),
others
were
treated
with
strain­
specific
mutagens
(
and
served
as
positive
controls).
At
harvest,
the
numbers
of
revertant
colonies
(
prototrophic
his
+
and
try
+)
in
test
cultures
were
compared
to
those
in
solvent
controls.

In
a
preliminary
dose­
finding
study,
TA­
100
was
exposed
to
five
concentrations
of
the
test
chemical
ranging
from
8
to
5000
µ
g/
plate
±
S9,
together
with
solvent
and
positive
controls
±
S9.
Cytotoxicity
(
marked
reduction
in
revertants)
was
seen
at
the
maximum
test
concentration
(
HCT),
5000
µ
g/
plate,
both
in
the
presence
and
absence
of
S9.

In
the
first
main
trial
employing
all
strains
of
bacteria
and
the
same
concentration
as
in
the
rangefinding
study,
cytotoxicity
was
again
observed
at
the
HCT
in
all
strains
with/
without
activation.
While
retaining
the
HCT
for
Trial
2,
a
narrower
range
of
seven
lower
concentrations
(
32
to
5000
µ
g/
plate)
was
employed.
Evidence
of
cytotoxicity
was
evident
at
the
HCT
in
all
treated
strains
of
Trial
2
±
S9.
In
addition,
precipitation
of
the
test
article
was
observed
at
concentrations
 
500
µ
g/
plate
­
S9
and
 
1000
µ
g/
plate
+
S9.

Evidence
of
mutagenicity
was
found
in
treated
TA98
in
the
presence
of
S9­
metabolic
activation
as
shown
by
statistically
significant
(
p
<
0.01,
by
Dunnett's
Test),
concentration­
related,
and
reproducible
increases
in
revertants.
The
response
in
Experiment
I
was
~
5­
fold
at
1000
µ
g/
plate
+
S9
(
120
revertants/
plate
vs.
25
revertants/
plate
for
DMF).
In
Experiment
II,
the
response
was
~
3.2­
fold
at
2000
µ
g/
plate,
4­
fold
at
1000
µ
g/
plate
and
3­
fold
at
500
µ
g/
plate
which
corresponds
to
83,
106
and
80
revertants
vs.
26
for
DMF,
respectively.
Neither
in
the
treatment
of
TA98
in
the
absence
of
activation,
nor
in
the
treatment
of
any
other
exposed
bacterial
strain
±
S9
was
such
definitive
positive
mutagenic
activity
shown.

Therefore,
it
is
concluded
that
the
G51­
15­
162
batch
of
KIF­
230
TGAI
produced
mutation
in
S.
typhimurium
TA98
in
the
presence
of
a
rat
liver
activation
(+
S9)
when
assayed
at
concentrations
up
to
5000
µ
g/
plate.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.10.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
No.
G51­
35­
184,
91.9%
a.
i.),
performed
at
the
Biosafety
Research
Center,
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoko,
Japan.
Laboratory
Experiment
No.
5839
(
001­
320),
dated
September
11,
Page
70
of
87
2001.
MRID
45869723.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
gene
mutation
assay
in
bacteria
(
MRID
45869723),
cultures
of
four
histidine­
requiring
(
auxotrophic
his
­)
strains
of
Salmonella
typhimurium
(
TA100,
TA98,
TA1535,
TA1537
)
and
the
tryptophan­
requiring
WP2
uvrA
strain
of
Escherichia
coli
(
auxotrophic
try
­)
were
exposed
for
48
hours
to
KIF
230
TGAI
(
Lot
No.
651­
35­
184,
91.9%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
at
six
concentrations
ranging
from
156
to
5000
µ
g/
plate
in
the
presence
of
a
rat
metabolic
activation
system
(+
S9)
with
TA100,
TA1535,
and
WP2
uvrA,
and
at
six
concentrations
ranging
from
39.1
to
1250
µ
g/
plate
with
TA98
and
TA1537
in
its
absence.

In
addition
to
cultures
exposed
to
the
solvent
("
negative"
control),
other
cultures
were
treated
with
strain­
specific
reference
mutagens
(
positive
controls).
At
harvest,
the
numbers
of
revertant
colonies
(
prototrophic
his+
and
try+
)
in
test
article­
treated
cultures
and
positive
control
cultures
were
compared
to
those
in
the
negative
control
cultures.

The
preliminary
dose­
finding
study
included
negative
and
positive
controls
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9
and
revealed
cytotoxicity
(
growth
inhibition)
and
increasingly
denser
precipitation
at
 
800
µ
g/
plate.

In
the
main
(
mutagenicity)
assay,
growth
inhibition
and
increasingly
denser
precipitation
of
the
test
article
were
evident
at
 
625
µ
g/
plate
±
S9.
All
positive
controls
induced
the
expected
response
in
the
corresponding
strain.
However,
in
neither
the
dose­
finding
study
nor
the
main
assay
did
the
test
article
increase
the
incidence
of
revertants
over
negative
control
at
any
concentration
in
any
bacterial
strain,
in
the
presence
or
absence
of
S9­
activation.

Therefore,
Lot
No.
G51­
35­
184
technical
KIF­
230
did
not
induce
gene
mutation
in
the
conventional
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.11.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
No.
G51­
37­
184,
95.8%
a.
i.),
performed
at
the
Biosafety
Research
Center,
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experiment
No.
5840
(
001­
321),
dated
September
11,
2001.
MRID
45869724.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
gene
mutation
assay
in
bacteria
(
MRID
45869724),
cultures
of
four
histidine­
requiring
(
auxotrophic
his
­
)
strains
of
Salmonella
typhimurium
(
TA100,
TA98,
TA1535,
and
TA1537)
and
the
tryptophan­
requiring
(
auxotrophic
try
­)
WP2
uvrA
strain
of
Escherichia
coli
were
exposed
for
48
hours
to
KIF­
230
TGAI
(
Lot
No.
G51­
37­
184,
95.8%
a.
i.),
dissolved
in
dimethyl
sulfoxide,
DMSO,
in
the
presence
(+
S9)
and
absence
(­
S9)
of
a
rat
metabolic
activation
system
(
S9­
mix),
at
seven
concentrations
ranging
from
78.1
to
5000
µ
g/
plate.
In
addition
to
cultures
exposed
to
the
solvent
alone
("
negative"
controls),
other
cultures
were
treated
with
strain­
specific
reference
mutagens
(
positive
controls).
At
harvest,
the
numbers
of
revertant
colonies
(
prototrophic
his
+
and
try+)
in
test
article­
treated
and
positive
control
cultures
were
compared
to
those
in
the
negative
controls,
to
ascertain
the
mutagenicity
of
KIF
Page
71
of
87
230
Lot.
No.
G51­
37­
184,
and
the
adequacy
of
the
strain­
specific
mutagens.

A
preliminary
dose­
finding
study
at
eight
concentrations,
ranging
from
8.19
to
5000
µ
g/
plate
±
S9,
reduced
revertant
colony
counts
for
all
strains
up
to
the
highest
concentration
tested
(
HCT)
­
S9,
and
produced
increasingly
denser
precipitation
at
 
800
µ
g/
plate.
In
the
main
(
mutagenicity)
assay,
reduced
revertant
colonies
were
observed
at
2500
and
5000
µ
g/
plate
in
the
absence
of
S9­
activation,
and
increasingly
denser
precipitation
of
the
test
article
was
evident
at
 
313
µ
g/
plate
­
S9
and
at
 
625
µ
g/
plate
+
S9.
The
positive
controls
induced
the
expected
response
in
the
corresponding
strains.

However,
in
neither
the
dose­
finding
study
nor
the
main
assay
did
the
test
article
increase
the
incidence
of
revertants
over
negative
control
at
any
concentration
in
any
bacterial
strain,
in
the
presence
and
absence
of
S9­
activation.

Therefore,
Lot
No.
G51­
37­
184
of
technical
KIF­
230
did
not
induce
gene
mutation
in
the
conventional
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline,
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation
data.

2.7.12.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
No.
G51­
36­
184,
performed
at
the
Biosafety
Research
Center:
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experiment
No.
5918
(
001­
326),
dated
October
29,
2001.
MRID
45869725.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
gene
mutation
assay
in
bacteria
(
MRID
45869725),
cultures
of
four
histidine­
requiring
(
auxotrophic
his
­
)
strains
of
Salmonella
typhimurium
(
TA100,
TA98,
TA1535,
and
TA1537),
and
the
tryptophan­
requiring
(
auxotrophic
try
­)
WP2
uvrA
strain
of
Escherichia
coli,
were
exposed
for
48
hours
employing
the
preincubation
procedure,
to
KIF­
230
TGAI
(
Lot
No.
G51­
36­
184,
92.6%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
in
the
presence
(+
S9)
and
absence
(­
S9)
of
a
rat
metabolic
activation
system
(
S9­
mix),
to
eight
concentrations
ranging
from
39.1
to
5000
µ
g/
plate
­
S9,
and
to
six
concentrations
ranging
from
156
to
5000
µ
g/
plate
+
S9.
In
addition
to
cultures
exposed
to
the
solvent
alone
(
designated
as
"
negative"
controls),
other
cultures
were
treated
with
strain­
specific
reference
mutagens
(
positive
controls).
At
the
end
of
treatment,
the
numbers
of
revertant
colonies
(
prototrophic
his
+

and
try+)
in
test
article­
treated
and
positive
control
cultures
were
compared
to
those
in
the
negative
controls,
to
ascertain
the
mutagenicity
of
KIF­
230
Lot.
No.
G51­
36­
184,
and
the
adequacy
of
the
results
from
the
positive
controls.

In
a
preliminary
dose­
finding
study
which
included
negative
and
positive
controls,
at
eight
concentrations
ranging
from
8.19
to
5000
µ
g/
plate
±
S9,
cytotoxicity
(
growth
inhibition)
was
observed
at
 
800
µ
g/
plate,
and
precipitation
was
encountered
at
 
320
µ
g/
plate
­
S9.
In
treated
S9­
activated
cultures,
cytotoxicity
was
found
only
at
the
highest
concentration,
5000
µ
g/
mL,
and
precipitation
was
visible
at
 
800
µ
g/
plate
+
S9.
In
the
main
(
mutagenicity)
assay,
growth
inhibition
was
observed
at
625
µ
g/
plate
in
TA98,
and
1250
µ
g/
mL
in
TA1537
in
the
absence
of
S9­
activation,
accompanied
by
increasingly
denser
precipitation
of
the
test
article
at
 
313
Page
72
of
87
µ
g/
plate;
whereas
increasing
precipitation
was
seen
at
 
625
µ
g/
plate
+
S9,
but
no
growth
inhibition
up
to
the
limit
dose,
5000
µ
g/
plate.

However,
in
neither
the
dose­
finding
study
nor
the
main
assay
did
the
test
article
increase
the
incidence
of
revertants
over
negative
control
at
any
concentration
in
any
bacterial
strain,
in
the
presence
or
absence
of
S9­
activation.
All
positive
controls
induced
the
expected
marked
increases
in
revertant
counts.

Therefore,
KIF­
230
TGAI
(
Lot
No.
G51­
36­
184)
did
not
induce
gene
mutation
in
the
conventional
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline,
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.13.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
Test
of
KIF­
230
TGAI
(
Lot
No.
G51­
08­
158,
88.6%
a.
i.),
performed
at
the
Biosafety
Research
Center
of
the
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.
5919
(
001­
327),
dated
October
29,
2001.
MRID
45869726.
Unpublished.

EXECUTIVE
SUMMARY:
In
a
reverse
gene
mutation
assay
in
bacteria
(
MRID
45869726),
four
histidine­
requiring
(
auxotrophic
his
­)
strains
of
Salmonella
typhimurium
(
TA100,
TA98,
TA1535
and
TA1537),
and
the
tryptophan­
requiring
(
auxotrophic
try
­)
strain
of
Escherichia
coli
WP2
uvrA,
were
exposed
for
48
hours,
employing
the
preincubation
procedure,
to
KIF­
230
TGAI
(
Lot
No.
51­
08­
158,
88.6%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
in
the
presence
(+
S9)
and
absence
(­
S9)
of
a
metabolic
activation
system,
at
12
concentrations
ranging
from
28.4
to
5000
µ
g/
plate
­
S9
and
eight
concentrations
ranging
from
186
to
5000
µ
g/
plate
+
S9.
Cultures
of
all
strains
were
also
exposed
to
the
solvent
(
serving
as
"
negative"
controls),
and
other
cultures
were
treated
with
strain­
specific
mutagens
(
serving
as
positive
controls).
At
harvest,
the
numbers
of
revertant
colonies
(
by
reverse
mutation,
i.
e.,
to
prototrophic
his
+
and
try
+)
in
the
test
article
and
positive
controls
were
compared
to
the
negative
controls
to
ascertain
mutagenic
activity
and
validity
of
the
results
from
the
positive
controls.

In
a
preliminary
dose­
finding
(
pilot)
study,
which
included
negative
and
positive
controls,
at
eight
concentrations
of
the
test
article
ranging
from
8.19
to
5000
µ
g/
plate
in
the
presence
(+
S9)
and
absence
(­
S9)
of
metabolic
activation
(
±
S9­
mix),
increasingly
denser
precipitation
of
the
test
compound
was
present
at
 
320
µ
g/
plate
±
S9,
whereas
reduced
revertant
colonies
were
observed
in
all
treated
S.
typhimurium
strains
at
and
above
320
µ
g/
plate.

In
the
main
(
mutagenicity)
assay,
precipitation
was
visible
at
 
298
µ
g/
plate
±
S9,
and
growth
inhibition
in
all
treated
strains.

However,
in
both
the
pilot
study
and
the
main
assay,
the
numbers
of
revertant
colonies
in
S9­
activated
TA98
cultures
treated
at
test­
article
concentrations
approximating
 
300
µ
g/
plate
were
substantially
increased
compared
to
solvent
control
values.
Such
increases
in
revertants
were
not
found
in
any
other
strain
at
any
other
treatment,
and
were
not
observed
in
non­
activated
TA98
test
cultures.
All
positive
controls
induced
the
expected
marked
increases
over
solvent
controls
in
revertants.
Fold­
increases
in
the
pilot
study
ranged
from
3.7X
at
5000
µ
g/
plate
+
S9
to
3X
at
320
Page
73
of
87
µ
g/
plate
+
S9,
and
reached
a
maximum
of
6.7X
at
800
µ
g/
plate
(
177
revertants
vs.
26
in
the
solvent
control.
In
agreement
with
these
findings,
fold
increases
in
the
main
assay
were
3.5X
at
477
and
1953
µ
g/
plate
+
S9,
with
a
maximum
increase
(
4.81X)
at
763
µ
g/
plate
+
S9
(
160
revertants
vs.
33
in
the
solvent
control).

Therefore,
it
is
concluded
that
technical
KIF­
230
TGAI
(
Lot
No.
G51­
08­
158)
has
innate
mutagenic
activity
in
one
bacterial
strain,
in
the
presence
of
S9­
activation.
However,
in
a
recent
repeat
study,
dated
29
March
2002,
the
positive
response
in
S9­
activated
TA98
cultures
was
demonstrated
to
be
due
to
a
mutagenic
impurity
(
see
MRID
45835107).

The
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
for
Mutagenicity
(
bacterial
reverse
mutation)
data.

2.7.14.
Nakajima,
M.
(
1998).
In
Vitro
Chromosomal
Aberration
Test
on
CHL
Cells
Treated
With
KIF­
230
Technical,
performed
at
the
Biosafety
Research
Center
of
the
Foods,
Drugs
and
Pesticides
(
An
Pyo
Center),
Shizuoko,
Japan.
Laboratory
Experimental
No.
3391
(
001­
200),
dated
September
17,
1998.
MRID
45869727.
Unpublished.

EXECUTIVE
SUMMARY:
In
independent
in
vitro
chromosome
aberration
assays
(
MRID
45869727),
cultures
of
Chinese
hamster
lung
(
CHL)
fibroblasts
were
exposed
to
KIF­
230
TGAI
(
Lot
No.
G51­
02­
152,
87.7%
a.
i.
and
9%
inactive
optimal
isomer,
suspended
in
1%
carboxymethyl
cellulose
sodium
salt,
1%
CMC­
Na).
Chromosomal
preparations
were
made
following
two
culturing
procedures:

1.
The
"
direct
method,"
in
which
cells
were
first
incubated
continuously
for
3
days
at
37
°
C
under
5%
CO2,
either
for
24
hours
at
concentrations
of
955,
1910
and
3820
µ
g/
mL,
or
for
48
hours
at
the
same
concentrations
plus
a
lesser
concentration
of
478
µ
g/
mL.
This
pre­
incubation
was
followed
by
the
addition
of
the
vehicle
(
1%
CMC.
Na),
or
a
positive
control
plus
the
test
substance,
mixed
thoroughly,
and
the
mixture
incubated
for
an
additional
24
or
48
hours,
at
the
end
of
which
chromosomal
spreads
were
prepared
2.
The
"
activation
method,"
in
which
non­
activated
cells
were
exposed
for
6
hours
to
955,
1910
and
3820
µ
g/
mL
test
material,
or
to
the
vehicle,
or
the
positive
control.
In
the
presence
of
S9­
activation,
cells
were
exposed
to
identical
concentrations
of
the
test
material,
or
the
vehicle,
or
positive
control.
Then,
the
culture
medium
was
replaced
with
fresh
medium,
and
the
cells
washed
with
phosphate­
buffered
saline
(
PBS),
incubated
for
an
additional
18
hours,
at
the
end
of
which
chromosome
specimens
were
prepared
In
addition
to
cultures
exposed
to
the
vehicle,
CMC.
Na
in
both
of
these
assay
methods,
other
cultures
were
treated
with
the
mutagenic
clastogens,
mitomycin
C
(
MMC),
0.05
and
0.025
µ
g/
mL
for
the
two
phases
of
the
direct
method,
and
with
cyclophosphamide
(
CP)
12.5
µ
g/
mL
for
the
activation
method.
Two
hours
before
harvest,
cell
division
was
halted
in
"
c­
metaphase"
by
the
addition
of
the
alkaloid,
colcemid,
following
which
the
conventional
array
of
structural
chromosome
aberration
plus
polyploidy
were
scored.

In
a
preliminary
cytotoxicity
"
pilot"
test
to
ascertain
cell
growth
inhibition,
CHL
cells
were
exposed
to
10
concentrations
of
the
test
material
ranging
from
38.5
to
3820
µ
g/
mL
in
the
"
direct"
procedure
(
for
both
the
24­
hour
and
48­
hour
treatments),
as
well
as
in
the
non­
activated
(­
S9)
Page
74
of
87
assay,
and
treated
for
6
hours.
Eight
concentrations
ranging
from
107
to
3820
µ
g/
mL
(
the
latter,
equivalent
to
a
limit
of
10
mM)
were
tested
under
S9­
activation.
The
remaining
details
of
both
procedures
were
essentially
the
same
as
stated
above
for
the
main
(
cytotoxicity)
assay.

The
50%
cell
growth
inhibition
concentrations
(
IC50)
were
2147
and
1088
µ
g/
mL
for
the
24­
hr
and
48­
hr
"
direct"
exposures,
respectively,
but
no
cytotoxicity
was
seen
in
the
"
activation"
procedure.
A
"
white,
powdery
test
substance"
(
as
described
by
the
investigator)
was
observed
at
all
assayed
levels
in
all
tests/
trials
of
the
main
study,
down
to
the
lowest
concentration
tested,
38.5
µ
g/
mL.

At
no
concentration
in
either
procedure
(
or
in
the
pilot"
test)
up
to
the
highest
concentration
tested,
3820
µ
g/
mL,
were
chromosome
aberrations
(
structural
or
numerical)
in
test
cultures
found
to
differ
significantly
from
concurrent
controls,
or
from
the
range
of
the
laboratory's
historical
controls
(
as
stated
by
the
investigator,
but
no
data
were
presented).
Both
positive
controls
induced
a
markedly
increased
incidence
of
structural
aberrations
in
both
methods
(
except,
obviously,
for
non­
activated
cultures
treated
with
CP,
as
there
was
no
metabolic
inhibition
in
this
part
of
the
study).

This
study
is
classified
as
unacceptable
because
of
the
presence
of
a
"
white
powdery
test
substance"
at
all
concentrations
in
all
assays.
Therefore,
the
study
cannot
be
upgraded,
and
does
not
satisfy
the
requirement
for
FIFRA
Test
Guideline
for
in
vitro
cytogenetic
data.
(
see
DER,
Section
III
for
further
discussion).

2.7.15.
Fellows,
M.
(
1999).
KIF­
230
(
TGAI):
Mutation
at
the
Thymidine
Kinase
(
tk)
Locus
of
Mouse
Lymphoma
L5178Y
(
MLA)
Cells
using
the
Microtitre
®
Fluctuation
Technique,
performed
at
Covance
Laboratories,
Ltd.,
North
Yorkshore,
England.
Report
No.:
535/
45­
D5140,
dated
10
December
1999.
MRID
45869728.
Unpublished.

EXECUTIVE
SUMMARY:
In
independent
repeat
mammalian
cell
gene
mutation
assays,
(
MRID
45869728),
cultures
of
mouse
lymphoma
cells,
heterozygous
for
the
thymidine
kinase
locus
(
tk+/
­)
were
exposed
for
3
hours
by
the
fluctuation
procedure,
to
KIF­
230
TGAI
(
Batch
No.
G51­
15­
162,
87.9%
a.
i.,
dissolved
in
dimethyl
sulfoxide,
DMSO),
in
the
presence
and
absence
of
a
metabolic
activation
system
(
±
S9),
at
six
concentrations
ranging
from
3.75
to
120
µ
g/
mL
(
Experiment
1),
and
at
five
concentrations
ranging
from
7.5
to
120
µ
g/
mL
(
Experiment
2).
In
addition
to
cultures
exposed
to
DMSO
(
solvent,
"
negative"
control),
other
cultures
were
treated
with
the
mutagens,
4­
nitroquinoline­
1­
oxide
(
4­
NQO,
0.05
and
0.1
µ
g/
mL)
and
benzo(
a)
pyrene
(
BP,
2.0
and
3.0
µ
g/
mL).
At
harvest,
the
treatment
medium
was
replaced
with
fresh
medium
and
the
cells
maintained
for
2
days
to
express
the
mutant
genotype,
TK­/­
determined
by
tolerance
to
the
nucleoside
analogue,
5­
trifluorothymidine
(
TFT),
whereas
TK+/­
and
TK+/+
cells
cannot
exist
in
its
presence.

In
a
preliminary
cytotoxicity
range­
finding
test,
cultures
were
treated
with
six
concentrations
of
the
test
material
ranging
from
7.81
to
250
µ
g/
mL
±
S9.
The
limit
of
its
solubility
was
established
at
250
µ
g/
mL.
Cells
survived
all
concentrations;
the
highest
concentration
yielded
57.39%
survival
relative
to
the
negative
control
(
RS)
­
S9,
and
29.45%
+
S9.

In
Experiment
1,
the
highest
test
article
concentration
(
120
µ
g/
mL
)
produced
a
RS
of
23.66%
­
Page
75
of
87
S9,
and
37.82%
+
S9.
In
Experiment
2,
with
a
slightly
altered
concentration
range,
RS
at
120
µ
g/
mL
KIF­
230
TGAI
was
57.40%
­
S9
and
59.58%
+
S9.
Both
positive
controls
induced
the
expected
marked
increases
in
MF.

However,
in
neither
experiment
at
any
concentration
up
to
the
highest
soluble
concentration
in
the
presence
or
absence
of
activation
were
significant
increases
in
mutant
frequency
(
MF)
found.

This
study
is
classified
as
acceptable/
guideline,
and
satisfies
the
requirement
for
the
FIFRA
Test
Guidelines
for
mutagenicity
(
bacterial
reverse
mutation
data).

2.7.16.
Fellows,
M.
(
1999).
KIF­
230
(
TGAI):
Measurement
of
Unscheduled
DNA
Synthesis
in
Isolated
Rat
Hepatocytes
In
Vitro,
performed
at
Covance
Laboratories,
Ltd.,
North
Yorkshire,
England.
Report
No.:
535/
46­
D5140,
dated
10
December
1999.
MRID
45869729.
Unpublished.

EXECUTIVE
SUMMARY:
In
independent
in
vitro
unscheduled
DNA
synthesis
(
UDS)
assays
(
MRID
45869729),
primary
rat
hepatocyte
cultures
were
exposed
to
KIF­
230
TGAI
[
Batch
No.
G51­
15­
162,
87.9%
a.
i.,
dissolved
in
dimethyl
sulfoxide
(
DMSO)]
at
five
concentrations
ranging
from
5
to
500
µ
g/
mL
(
EXPERIMENT
1),
and
at
six
concentrations
ranging
from
15.625
to
500
µ
g/
mL
(
EXPERIMENT
2).
In
addition
to
cultures
exposed
to
the
solvent
("
negative"
control),
other
cultures
were
treated
with
the
mutagen,
2­
acetoaminofluorene
(
2­
AAF,
2.5
and
5.0
µ
g/
mL),
as
positive
control.
All
cultures
with
a
mean
viability
of
>
70%
were
treated
with
the
solvent,
positive
control,
or
the
test
article
plus
radioactive
[
3H]­
thymidine,
and
incubated
overnight
at
37
°
C
in
a
5%
CO2
in
air
(
v/
v)
atmosphere.
Following
incubation,
autoradiograms
of
the
hepatocytes
were
prepared,
and
UDS
determined
(
from
net
nuclear
grain
counts)
in
each
concentration
group.

In
preliminary
cytotoxicity
testing,
viability
(
percent
of
control
<
50%)
was
limited
to
concentrations
 
250
µ
g/
mL.
Because
of
concentration­
reduced
relative
cell
counts
and
precipitation
of
the
test
article,
the
UDS
analyses
(
2
assays)
were
conducted
using
a
maximm
concentration
of
500
µ
g/
mL.

In
both
UDS
experiments,
no
concentration
up
to
that
limited
by
solubility
caused
a
reproducible
increase
in
the
number
of
net
nuclear
grain
counts
(
NNGC)
or
the
percent
of
cells
in
repair.
In
contrast,
the
positive
control
induced
marked
increases
of
both
NNGC
and
the
percent
of
cells
in
repair.
Therefore,
there
was
no
evidence
(
or
a
dose­
related
positive
response)
that
UDS
as
determined
by
radioactive
tracer
procedures
(
nuclear
silver
grain
counts)
was
induced
by
KIF­
230
TGAI
Batch
No.
G51­
15­
162.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
other
genotoxic
mutagenicity
data.

2.7.17.
Nakajima,
M.
(
2000).
KIF­
230
Technical:
Gene
Mutation
in
Transgenic
Mice,
performed
at
the
Biosafety
Research
Center
[
of
the]
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.:
4911
(
001­
264),
dated
August
25,
2000.
MRID
45869730.
Unpublished.
Page
76
of
87
EXECUTIVE
SUMMARY:
In
an
in
vivo
gene
mutation
assay
with
transgenic
mice
(
MRID
45869730),
groups
of
five
MUTA
 
mice
were
administered
single
doses
for
5
days
of
1000
or
2000
mg/
kg/
day
KIF­
230
Technical
(
Batch
No.
G51­
08­
158,
88.8%/
89.1%
a.
i.,
suspended
in
0.5
w/
v%
carboxymethylcellulose,
sodium
salt
(
CMC.
Na),
or
a
dose
of
0.1/
0.2
mL
of
the
vehicle
(
to
serve
as
"
negative"
control)
by
gavage,
or
with
5
mg/
kg/
day
dimethylnitrosamine
(
DMN)
injected
intraperitoneally
(
to
serve
as
positive
control).
The
incidence
of
gene
mutation
in
hepatic
genomic
DNA
was
determined
in
all
mice
28
days
after
final
dosing.

Following
isolation
of
genomic
DNA
from
the
liver,
the
target
gene,
LacZ
was
"
packaged"
into
lambda
bacteriophage
(
 
phage),
mixed
with
Escherichia
coli
C
(
gal
E)
and
plated.
Up
to
4
x
105
pfu's
(
plaque­
forming­
units)
per
animal
were
counted.
In
the
absence
of
any
toxicity
(
as
determined
by
changes
in
body
weight,
and
clinical
observation),
the
incidences
of
mutation
frequency
(
MF)
at
either
dose
level
in
the
two
KIF­
230
treatment
groups
were
comparable
to
the
negative
control
value.
The
incidence
in
the
positive
control
group
was
markedly
increased
over
the
CMC.
Na
control
group.

Therefore,
the
potential
of
KIF­
230
to
induce
gene
mutation
in
this
study
is
considered
negative.

This
study
is
classified
as
acceptable/
non­
guideline.
No
FIFRA
Test
Guideline
is
presently
available
for
this
type
of
study.

2.7.18.
Mizuhashi,
F.
(
2001).
KIF­
230:
In
Vivo/
In
Vitro
Unscheduled
DNA
Synthesis
(
UDS)
Assay
in
Rat
Hepatocytes,
performed
at
the
Biosafety
Research
Center
[
of
the]
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.:
5336
(
001­
282),
dated
April
03,
2001.
MRID
45869731.
Unpublished.

EXECUTIVE
SUMMARY:

In
an
in
vivo/
in
vitro
unscheduled
DNA
synthesis
(
UDS)
assay
(
MRID
45869731),
groups
of
four
male
rats
were
administered
single
oral
doses
of
1000
or
2000
mg/
kg
KIF­
230
TGAI
[
Lot
No.
G51­
24­
176,
92.3%
a.
i.,
suspended
in
0.5
w/
v%
carboxymethyl
cellulose­
sodium
salt
(
CMC.
Na)],
or
the
vehicle,
CMC.
Na
("
negative"
control),
and
hepatocytes
isolated
by
liver
perfusion
two
and
16
hours
later.
In
addition,
four
males
each
were
injected
with
single
5
mg/
kg
doses
of
the
mutagens,
dimethylnitrosamine
(
DMN)
in
water
or
single
100
mg/
kg
doses
of
2­
acetoaminofluorene
(
2­
AAF)
in
corn
oil
(
serving
as
positive
controls),
and
hepatocytes
were
isolated,
respectively,
2
and
16
hours
later.

At
harvest,
the
isolated
hepatocytes
were
cultured,
and
UDS
measured
by
determining
nuclear
grain
counts
after
radioactive
labeling.
Since
no
toxicity
was
observed,
results
from
only
three
animals
per
group
were
analyzed.

The
mean
numbers
of
net
nuclear
grain
counts,
NNGC,
i.
e.,
the
incidences
of
gross
nuclear
grains
minus
the
averages
of
three
similar­
sized
cytoplasmic
areas
surrounding
each
nucleus
in
hepatocytes
from
groups
treated
with
both
doses
of
KIF­
230
were
comparable
to
that
in
the
concurrent
negative
control
group,
and
also
did
not
differ
from
the
laboratory's
historical
control
values.
The
incidence
of
cells­
in­
repair
in
the
two­
hour
treatment
group
was
slightly
higher
at
the
Page
77
of
87
2000
µ
g/
kg
dose,
but
no
increase
was
observed
in
any
other
group.
By
contrast,
the
incidences
of
NNGC
and
cells­
in­
repair
in
both
positive
control
groups
were
markedly
higher.

Therefore,
KIF­
230
TGAI
Lot
No.
G51­
24­
176
was
negative
for
UDS
in
rat
hepatocytes
drawn
from
males
treated
up
to
the
in
vivo
limit
dose.

There
was
no
evidence
(
or
a
dose­
related
positive
response)
that
unscheduled
DNA
synthesis,
as
determined
by
radioactive
tracer
procedures
(
nuclear
silver
grain
counts),
was
induced.

This
study
is
classified
as
acceptable/
guideline.
It
does
satisfy
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
other
genotoxic
mutagenicity
data.

2.7.19.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
of
KIF­
230­
I­
1(
R)
with
Bacteria,
performed
at
the
Biosafety
Research
Center
[
of
the]
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.:
5504(
001­
274),
dated
May
24,
2001.
MRID
45869732.
Unpublished.

EXECUTIVE
SUMMMARY:
In
a
bacterial
reverse
mutation
assay
(
MRID
45869732),
histidine­
deficient
(
auxotrophic,
his
­)
cells
of
four
Salmonella
typhimurium
strains
(
TA100,
TA98,
TA1535,
and
TA1537)
and
the
tryptophan­
deficient
(
auxotrophic,
try­
­)
cells
of
Escherichia
coli
WP2
uvrA
were
exposed
by
the
preincubation
procedure
to
KIF­
230­
I­
1(
R),
Lot
No.
G51­
25­
178,
99.6%,
dissolved
in
dimethyl
sulfoxide
(
DMSO),
at
eight
concentrations
ranging
from
156
to
5000
µ
g/
plate,
in
the
presence
and
absence
of
a
metabolic
activation
system
(
±
S9).
In
addition
to
cells
exposed
to
the
solvent
(
to
serve
as
"
negative"
controls),
other
cells
were
treated
with
strain­
specific
mutagens
(
to
serve
as
positive
controls).
At
harvest,
the
incidence
of
revertant
colonies
of
cells
(
his
+,
try­
+)
in
test
cultures
were
compared
to
the
negative
control.

In
a
preliminary
dose­
finding
("
pilot")
study,
bacteria
were
exposed
to
eight
concentrations
of
test
article
ranging
from
8.19
to
5000
µ
g/
plate
with/
without
S9
activation.
Precipitation
of
test
article
and
cytotoxicity
were
encountered
at
the
higher
concentrations.

In
the
main
(
mutation)
assay,
increasingly
heavier
precipitation
at
concentrations
preventing
clear
colony
counts
were
also
observed
following
treatment
in
the
range
 
625
to
800
µ
g/
plate.
As
well,
cytotoxicity
(
cells
growth
inhibition)
was
observed.
However,
at
no
concentration
of
test
article
in
the
presence
or
absence
of
S9
in
the
main
assay
(
or
in
the
pilot
study)
were
any
substantial
increases
in
revertants
(
his+
,
try+
colonies)
over
negative
control
values
found.
In
contrast,
marked
increases
in
revertants
were
clearly
produced
in
all
positive
controls
±
S9.

Therefore,
techincal
KIF­
230­
I­
1
(
Lot
No.
G51­
25­
178)
did
not
induce
reverse
gene
mutation
in
the
standard
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.20.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
of
KIF­
230­
I­
1(
S)
with
Bacteria,
performed
at
the
Biosafety
Research
Center
[
of
the]
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Page
78
of
87
Laboratory
Experimental
No.:
5592(
001­
298),
dated
June
28,
2001.
MRID
45869733.
Unpublished.

EXECUTIVE
SUMMMARY:
In
a
bacterial
reverse
mutation
assay
(
MRID
45869733),
histidine­
deficient
(
auxotrophic,
his
­)
cells
of
four
Salmonella
typhimurium
strains
(
TA100,
TA98,
TA1535,
and
TA1537)
and
the
tryptophan­
deficient
(
auxotrophic,
try­
­)
cells
of
Escherichia
coli
WP2
uvrA
were
exposed
by
the
preincubation
procedure
to
KIF­
230­
I­
1(
S),
Lot
No.
G51­
30­
182,
99.5%,
dissolved
in
dimethyl
sulfoxide
(
DMSO),
at
seven
concentrations
ranging
from
78.1
to
5000
µ
g/
plate,
in
the
presence
and
absence
of
a
metabolic
activation
system
(
±
S9).
In
addition
to
cells
exposed
to
the
solvent
(
to
serve
as
"
negative"
controls),
other
cells
were
treated
with
strain­
specific
mutagens
(
to
serve
as
positive
controls).
At
harvest,
the
incidence
of
revertant
colonies
of
cells
(
his
+,
try­
+)
in
test
cultures
were
compared
to
the
negative
control.

In
a
preliminary
dose­
finding
("
pilot")
study,
bacteria
were
exposed
to
eight
concentrations
of
test
article
ranging
from
8.19
to
5000
µ
g/
plate
with/
without
S9
activation.
Precipitation
of
test
article
and
cytotoxicity
were
encountered
at
the
higher
concentrations.

In
the
main
(
mutation)
assay,
increasingly
heavier
precipitation
at
concentrations
preventing
clear
colony
counts
were
also
observed
following
treatment
in
the
range
 
625
µ
g/
plate.
However,
at
no
concentration
of
test
article
in
the
presence
or
absence
of
S9
in
the
main
assay
(
or
in
the
pilot
study)
were
any
substantial
increases
in
revertants
(
his+
,
try+
colonies)
over
negative
control
values
found.
In
contrast,
marked
increases
in
revertants
were
clearly
produced
in
all
positive
controls
±
S9.

Therefore,
techincal
KIF­
230­
I­
1(
S)
(
Lot
No.
G51­
30­
182)
did
not
induce
reverse
gene
mutation
in
the
standard
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

2.7.21.
Mizuhashi,
F.
(
2001).
Reverse
Mutation
of
KIF­
230­
I­
13
with
Bacteria,
performed
at
the
Biosafety
Research
Center
[
of
the]
Foods,
Drugs
and
Pesticides
(
An­
Pyo
Center),
Shizuoka,
Japan.
Laboratory
Experimental
No.:
5504(
001­
274),
dated
May
24,
2001.
MRID
45869734.
Unpublished.

EXECUTIVE
SUMMMARY:
In
a
bacterial
reverse
mutation
assay
(
MRID
45869734),
four
histidine­
deficient
(
auxotrophic,
his
­)
Salmonella
typhimurium
strains
(
TA100,
TA98,
TA1535,
and
TA1537)
and
the
tryptophan­
deficient
(
auxotrophic,
try­)
cells
of
Escherichia
coli
WP2
uvrA
were
exposed
by
the
preincubation
procedure
to
KIF­
230­
I­
13,
Lot
No.
G51­
31­
183,
98.7%
a.
i.,
dissolved
in
dimethyl
sulfoxide
(
DMSO),
in
the
presence
(+
S9)
and
absence
(­
S9)
of
a
metabolic
activation
system
(
±
S9),
at
eight
concentrations
ranging
from
15.6
to
2000
µ
g/
plate
­
S9,
and
to
seven
concentrations
ranging
from
31.3
to
2000
µ
g/
plate
+
S9.
In
addition
to
cells
exposed
to
the
solvent
(
to
serve
as
"
negative"
controls),
other
cells
were
treated
with
strain­
specific
mutagens
(
to
serve
as
positive
controls).
At
harvest,
the
incidence
of
revertant
colonies
of
cells
(
his
+,
try­
+)
in
test
cultures
were
compared
to
the
negative
control.

In
a
preliminary
dose­
finding
("
pilot")
study,
bacteria
were
exposed
to
eight
concentrations
of
test
Page
79
of
87
article
ranging
from
8.19
to
5000
µ
g/
plate
with/
without
S9
activation.
Precipitation
of
test
article
and
cytotoxicity
were
encountered
at
the
higher
concentrations.

In
the
main
(
mutation)
assay,
increasingly
heavier
precipitation
at
concentrations
preventing
clear
colony
counts
were
also
observed
following
treatment
at
 
125
µ
g/
plate
­
S9,
and
 
250
µ
g/
plate
+
S9,
as
well
as
cytotoxicity
(
cells
growth
inhibition)
in
the
majority
of
strains
 
250
µ
g/
plate
­
S9
and
at
 
1000
µ
g/
plate
+
S9.
However,
at
no
concentration
of
test
article
in
the
presence
or
absence
of
S9
in
the
main
assay
(
or
in
the
pilot
study)
were
any
substantial
increases
in
revertants
(
his+
,
try+
colonies)
over
negative
control
values
found.
In
contrast,
marked
increases
in
revertants
were
clearly
produced
in
all
positive
controls
±
S9.

Therefore,
techincal
KIF­
230­
I­
13
(
Lot
No.
G51­
31­
183)
did
not
induce
reverse
gene
mutation
in
the
standard
battery
of
bacterial
strains.

This
study
is
classified
as
acceptable/
guideline
and
satisfies
the
requirement
for
FIFRA
Test
Guideline
84­
2
for
in
vitro
mutagenicity
(
bacterial
reverse
gene
mutation)
data.

3.0
METABOLISM
CONSIDERATIONS
1.0
Introduction
1.1
Description
of
Issues
As
this
tolerance
petition
is
for
imported
grapes
and
tomatoes,
and
there
are
no
significant
livestock
feed
items
associated
with
these
commodities,
the
metabolites
of
concern
in
plants
are
the
only
metabolites
that
need
to
be
addressed.
In
the
grape
and
potato
metabolism
studies,
which
used
both
[
Bz­
14C]
and
[
Val­
14C]­
labeled
benthiavalicarb,
metabolism
of
the
two
14C­
labels
was
qualitatively
and
quantitatively
similar,
indicating
that
the
parent
molecule
remains
relatively
intact.
In
grapes
and
tomatoes,
the
metabolism
of
benthiavalicarb
was
minimal
with
parent
being
the
only
major
14C­
residue
in
fruit
and
foliage.
Parent
was
also
the
principal
14C­
residue
in
potato
foliage,
but
other
metabolites
were
identified.
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
Secondary
metabolism
occurs
via
conjugation
with
sugars.
Chiral
analysis
of
potato
foliage
extracts
also
indicates
that
there
was
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
possible
isomers
(
S­
L,
R­
D,
or
S­
D).

1.2
Team
Proposal
The
risk
assessment
team
proposes
that
the
residues
of
concern
in
grapes
and
tomatoes
for
both
tolerance
expression
and
risk
assessment
are
the
combined
residues
of
the
R­
L
and
S­
L
isomers
of
benthiavalicarb.

2.0
Nature
of
the
Residue
Studies
in
Plants
2.1.1
Executive
Summary
of
Plant
Study
No.
1
Page
80
of
87
In
a
grape
metabolism
study,
the
R­
L
isomer
of
[
14C]
benthiavalicarb,
radiolabeled
at
the
2­
valyl
position
[
Val­
14C]
or
uniformly
in
the
benzene
ring
[
Bz­
14C]
was
applied
as
six
broadcast
foliar
applications
to
grape
vines
during
fruit
development,
at
rates
equivalent
to
0.089
lb
ai/
A/
application
at
retreatment
intervals
(
RTIs)
of
7­
14
days,
for
a
total
of
0.534
lb
ai/
A/
season.
Grapevine
foliage
and
grapes
were
harvested
17
days
after
the
last
application
(
17
DAT).

The
total
radioactive
residues
(
TRR)
were
0.241
ppm
in
grapes
and
21.66
ppm
in
foliage
from
the
[
Val­
14C]
test.
TRR
were
0.327
ppm
in
grapes
and
14.01
ppm
in
foliage
from
the
[
Bz­
14C]
test.
Samples
were
extracted
sequentially
with
acetonitrile
(
ACN),
water,
and
acetone.
The
ACN
and
water
extracts
were
combined
and
partitioned
with
ethyl
acetate
(
EtOAc),
and
the
resulting
fractions
were
analyzed
by
HPLC
and/
or
TLC.
Aqueous
phase
extracts
from
vine
foliage
samples
were
further
subjected
to
 ­
glucosidase
enzymatic
hydrolysis
and
analyzed
by
HPLC.

Initial
solvent
extractions
solubilized
90­
98%
TRR
from
grape
matrices,
and
another
5%
of
the
TRR
was
extracted
from
foliage
with
water.
Subsequent
acetone
extraction
solubilized
an
additional
1­
2%
TRR.
Nonextractable
residues
accounted
for
1.1­
2.4%
TRR
in
grape
matrices.
HPLC
analyses
identified
parent
(
94.0­
96.5%
TRR)
as
the
main
component
in
both
fruit
and
foliage.
No
other
metabolites
were
identified,
and
isolated
unknowns
accounted
for
 
2.0%
of
the
TRR
in
each
matrix.
Enzymatic
hydrolysis
of
foliage
extracts
with
 ­
glucosidase
did
not
show
any
evidence
of
glucoside
conjugation.
In
addition,
no
differences
were
noted
in
the
level
and
distribution
of
14C­
residues
between
the
two
14C­
labels.
Based
on
this
residue
profile,
the
metabolism
of
benthiavalicarb
in
grapes
was
minimal.

2.1.2
Tabular
Summary
of
Plant
Study
No.
1
TABLE
A.
2.
Summary
of
Characterization
and
Identification
of
Solvent
Extracted
14C­
Residues
in
Grape
Matrices
Following
Six
Foliar
Applications
of
[
14C]
Benthiavalicarb
Totaling
0.534
lb
ai/
A.

[
Val­
14C]­
label
[
Bz­
14C]­
label
Grapes
Foliage
Grapes
Foliage
TRR
=
0.241
ppm
TRR
=
21.663
ppm
TRR
=
0.327
ppm
TRR
=
14.010
ppm
Compound
1
%
TRR
ppm
%
TRR
ppm
%
TRR
ppm
%
TRR
ppm
KIF­
230R­
L
(
parent)
96.5
0.2325
94.0
20.359
95.7
0.3131
94.6
13.2492
Total
identified
96.5
0.233
94.0
20.359
95.7
0.313
94.6
13.249
Unknowns
(
each
 
1%
TRR)
1.5
0.0038
0.9
0.1948
2.0
0.0066
1.0
0.1408
Total
characterized
98
0.2368
94.9
20.5538
97.7
0.3196
95.6
13.3898
Unallocated
background
<
0.2
0.0003
<
0.4
0.0614
<
0.7
0.0020
<
0.5
0.0538
Total
extractable
98.2
0.2371
95.3
20.6152
98.4
0.3216
96.1
13.4436
Unextractable
(
PES)
2
1.4
0.0034
2.4
0.5223
1.1
0.0035
2.3
0.3195
Accountability
3
99.7
97.6
99.4
98.2
1
Metabolites
were
identified
by
HPLC
and
identities
were
confirmed
by
LC/
MS.
2
Post
extraction
solids
(
PES);
residues
remaining
after
exhaustive
extractions.
3
Accountability
=
(
Total
extractable
+
Total
unextractable)/(
TRR
from
combustion
analysis;
see
TABLE
C.
2.1)
*
100.
Page
81
of
87
2.1.3.
Executive
Summary
of
Plant
Study
No.
2
In
a
tomato
metabolism
study,
the
R­
L
isomer
of
[
U­
phenyl­
14C]
benthiavalicarb
[
Bz­
14C]
was
applied
as
six
broadcast
foliar
applications
to
tomato
plants
during
fruit
development
at
a
rate
equivalent
to
0.089
lb
ai/
A/
application
at
retreatment
intervals
(
RTIs)
of
7­
14
days,
for
a
total
of
0.534
lb
ai/
A/
season.
Tomato
fruit
was
harvested
at
14,
28,
35,
49,
and
56
days
after
the
last
application
(
DAT),
and
foliage
was
harvested
at
56
DAT.

Following
the
final
application,
total
radioactive
residues
(
TRR)
in/
on
fruit
declined
slowly
from
0.018­
0.021
ppm
at
14
DAT
to
0.007
ppm
at
56
DAT.
TRR
was
2.33
ppm
in/
on
tomato
foliage
harvested
56
DAT.
Samples
were
extracted
sequentially
with
acetonitrile
(
ACN)
and
water.
The
ACN
extract
from
fruit
samples
was
diluted
with
water,
further
cleaned
up
using
solid
phase
extraction
(
SPE),
then
analyzed
by
HPLC
and
TLC.
The
aqueous
SPE
phase
was
concentrated,
reconstituted
in
water:
MeOH
(
4:
1,
v/
v),
and
analyzed
by
HPLC
and
TLC.
The
ACN
and
water
extracts
from
the
foliage
samples
were
combined,
concentrated,
and
analyzed
by
HPLC
and
TLC.

Initial
ACN
extractions
solubilized
63­
98%
TRR
from
tomato
matrices,
and
another
0­
34%
of
the
TRR
was
extracted
with
water.
Nonextractable
residues
accounted
for
2.4­
4.0%
TRR
in
tomato
matrices.
HPLC
analyses
identified
parent
as
comprising
54.8­
95.1%
of
the
TRR
in
tomatoes
and
tomato
foliage.
Parent
was
the
only
compound
detected
in
either
fruit
or
foliage.
In
the
56­
DAT
fruit,
unknowns
accounted
for
a
total
of
40.9%
of
the
TRR,
but
each
unknown
accounted
for
 
9.4%
of
the
TRR
(<
0.001
ppm).
The
metabolism
of
benthiavalicarb
in
tomatoes
was
minimal.
Page
82
of
87
2.1.4
Tabular
Summary
of
Plant
Study
No.
2
TABLE
A.
3.
Summary
of
Characterization
and
Identification
of
14C­
Residues
in
Tomato
Fruit
and
Foliage
Following
Six
Foliar
Applications
of
[
Bz­
14C]
Benthiavalicarb
Totaling
0.534
lb
ai/
A.

Tomatoes
(
14
DAT)
Tomatoes
(
56
DAT)
Foliage
(
56
DAT)

TRR
=
0.0212
ppm
TRR
=
0.0067
ppm
TRR
=
2.3306
ppm
Compound
1
%
TRR
ppm
%
TRR
ppm
%
TRR
ppm
KIF­
230R­
L
(
parent)
88.8
0.0188
54.8
0.0034
95.1
2.2163
Total
identified
88.8
0.0188
54.8
0.0034
95.1
2.2163
Unknowns
(
each
 
9.4%
TRR)
8.0
0.0017
40.9
0.0027
ND
­­

Total
characterized
96.8
0.0205
95.7
0.006
95.1
2.2163
Unallocated
background
0.7
<
0.0002
1.4
<
0.0003
0.9
0.0206
Total
extractable
97.5
0.0207
97.1
0.006
96
2.2369
Unextractable
(
PES)
2
2.4
0.0005
3.0
0.0002
4.0
0.0942
Accountability
3
100
92.5
100
1
Metabolites
were
identified
by
HPLC
and
identities
were
confirmed
by
LC/
MS.
Structures
and
codes
are
found
in
Table
B.
2.1.
2
Post
extraction
solids
(
PES);
residues
remaining
after
exhaustive
extractions.
3
Accountability
=
(
Total
extractable
+
Total
unextractable)/(
TRR
from
combustion
analysis;
see
TABLE
C.
2.1)
*
100.

2.1.5.
Executive
Summary
of
Plant
Study
No.
3
The
metabolism
of
benthiavalicarb
in
potatoes
was
examined
in
three
different
tests
using
both
[
U­
phenyl­
14C]­
and
[
Valyl­
2­
14C]­
labeled
benthiavalicarb
in
each
test.
In
a
preliminary
study
(
Test
I),
each
14C­
labeled
compound
was
applied
as
a
single
broadcast
foliar
application
to
immature
potato
plants
at
a
rate
equivalent
to
~
0.089
lb
ai/
A,
and
foliage
samples
were
collected
12
days
after
treatment
(
DAT).
In
the
second
study
(
Test
II),
each
[
14C]­
label
was
applied
as
a
single
post­
emergence
soil
application
at
a
rate
equivalent
to
~
0.089
lb
ai/
A,
and
samples
of
tubers
and
foliage
were
collected
90
DAT.
In
the
third
study
(
Test
III),
each
[
14C]­
label
was
applied
as
six
broadcast
foliar
applications,
at
retreatment
intervals
of
7
days,
to
mature
potato
plants
at
rates
equivalent
to
0.088­
0.091
lb
ai/
A,
for
a
total
of
approximately
0.54
lb
ai/
A/
season.
Mature
tubers
and
foliage
were
then
harvested
14
days
after
the
last
application.

The
levels
of
radioactivity
were
similar
for
the
[
Bz­
14C]
and
[
Val­
14C]­
labeled
benthiavalicarb
in
samples
from
each
test.
In
Test
I,
TRR
in
immature
potato
foliage
was
0.198­
0.330
ppm
12
days
after
a
single
foliar
application.
In
Test
II,
TRR
was
 
0.001
ppm
in
tubers
and
0.041­
0.078
ppm
in
foliage
harvested
90
days
after
a
single
soil
application.
In
Test
III,
TRR
was
0.003­
0.014
ppm
in
tubers
and
4.57­
5.86
ppm
in
foliage
harvested
14
days
after
the
last
of
six
foliar
applications.
The
low
levels
of
radioactivity
in
tubers
from
Tests
II
and
III
indicate
that
transport
of
the
14C­
residues
from
foliage
is
minimal.

For
analysis,
foliage
and
tuber
samples
were
extracted
sequentially
with
acetonitrile
(
ACN),
water,
and
acetone.
The
initial
ACN
and
water
extractions
solubilized
82­
98%
of
the
TRR
from
tubers
and
foliage,
and
acetone
solubilized
another
0.5­
1.4%
of
the
TRR.
Unextractable
Page
83
of
87
14C­
residues
accounted
for
1.2­
9.8%
of
the
TRR
from
foliage
and
17.5%
of
the
TRR
(<
0.003
ppm)
from
tubers.
14C­
Residues
in
the
ACN
and
water
extracts
were
cleaned
up
and
analyzed
by
HPLC
and/
or
TLC,
and
identities
of
parent
and
metabolites
were
confirmed
by
LC/
MS.

Following
a
single
foliar
application
to
immature
plants,
parent
(
65­
75%
TRR)
was
the
only
component
detected
in
foliage
at
14
DAT.
Following
a
single
postemergence
soil
application,
14C­
residues
in
tubers
were
too
low
for
analysis,
and
the
major
component
detected
in
foliage
(
90
DAT)
was
the
sugar
conjugate
Metabolite
1
(
28­
30%
TRR).
Parent
also
accounted
for
10­
11%
of
the
TRR
in
foliage
and
Metabolites
2,
3,
and
6
each
accounted
for
3­
17%
of
the
TRR
(
0.001­
0.011
ppm).
Following
the
last
of
six
broadcast
foliar
applications,
the
primary
component
identified
in
foliage
at
14
DAT
was
parent
(
88­
90%
TRR),
along
with
minor
amounts
of
Metabolites
1,
2,
and
6
(
0.8­
3.2%
TRR).
Treatment
with
 ­
glucosidase
and
LC/
MS
analysis
identified
Metabolites
1,
2,
and
6
as
sugar
conjugates
of
ring­
hydroxylated
metabolites.
In
tubers,
the
primary
metabolites
detected
were
Unknown
Metabolites
4
(
40%
TRR)
and
5
(
23%
TRR),
along
with
trace
amounts
of
parent
(
4.7%
TRR).
Treatment
with
 ­
glucosidase
suggests
that
Metabolites
4
and
5
are
also
sugar
conjugates;
however,
concentrations
of
these
metabolites
were
too
low
(<
0.006
ppm)
for
additional
analysis.

In
each
test,
the
metabolism
of
the
two
14C­
labels
was
qualitatively
and
quantitatively
similar,
indicating
that
the
parent
molecule
remains
relatively
intact
on
foliage.
In
tubers,
two
major
metabolites
(>
10%)
were
observed,
but
at
very
low
absolute
levels
(<
0.006
ppm).
In
potatoes,
the
metabolism
of
benthiavalicarb
occurs
primarily
via
direct
hydroxylation
of
the
phenyl
ring
or
defluorination
and
hydroxylation
of
the
phenyl
ring.
The
hydroxylated
metabolites
are
subsequently
conjugated
with
sugars.
In
addition,
chiral
analysis
of
foliage
extracts
indicates
that
there
is
no
isomeric
conversion
of
the
active
R­
L
isomer
to
other
stereoisomers
(
SL,
RD,
or
SD).

2.1.6.
Tabular
Summary
of
Plant
Study
No.
3
TABLE
A.
4.
Distribution
of
the
14C­
Residues
in
Potato
Foliage
Harvested
90
Days
After
a
Single
Soil
Application
of
[
14C]
Benthiavalicarb
at
~
0.089
lb
ai/
A.

[
Val­
14C]­
label
[
Bz­
14C]­
label
TRR
=
0.0619
ppm
TRR
=
0.0488
ppm
Metabolite
Fraction
1
%
TRR
ppm
%
TRR
ppm
ACN
extract
84.8
0.0525
84.4
0.0412
Water
extract
(
combined
with
ACN
extract)
5.1
0.0032
4.5
0.0022
EtOAc
(
HPLC
analysis)
61.4
0.0380
60.9
0.0297
Parent
10.9
0.0067
8.9
0.0043
Met
1
20.8
0.0128
20.5
0.0100
Met
2
10.7
0.0067
7.4
0.0036
Met
3
5.2
0.0032
2.6
0.0013
Met
6
8.8
0.0055
8.1
0.0040
Other
Unknowns
(
each
 
7.2%
TRR)
4.6
0.0029
13.2
0.0064
Page
84
of
87
TABLE
A.
4.
Distribution
of
the
14C­
Residues
in
Potato
Foliage
Harvested
90
Days
After
a
Single
Soil
Application
of
[
14C]
Benthiavalicarb
at
~
0.089
lb
ai/
A.

[
Val­
14C]­
label
[
Bz­
14C]­
label
TRR
=
0.0619
ppm
TRR
=
0.0488
ppm
Metabolite
Fraction
1
%
TRR
ppm
%
TRR
ppm
Unallocated
background
0.4
0.0003
0.2
0.0001
Aqueous
(
HPLC
analysis)
28.4
0.0176
28.0
0.0137
Parent
ND
­­
1.3
0.0006
Met
1
8.7
0.0054
7.4
0.0036
Met
2
3.9
0.0024
3.2
0.0015
Met
3
0.5
0.0003
ND
­­

Met
6
8.4
0.0052
7.8
0.0038
Other
Unknowns
(
each
 
1.7%
TRR)
6.7
0.0041
8.1
0.0040
Unallocated
background
0.1
0.0001
0.2
0.0001
Acetone
1.3
0.0008
1.4
0.0007
Residual
Solids
8.8
0.0055
9.8
0.0048
1
Metabolites
were
quantified
by
HPLC
and
identified
by
LC/
MS.
Structures
for
the
parent
and
metabolites
are
presented
in
Table
C.
3.1
ND
=
not
detected.

3.0
Nature
of
the
Residue
in
Livestock
No
significant
animal
feed
items
are
associated
with
grape
and
tomato
commodities,
as
a
result,
the
nature
of
the
residue
in
livestock
is
not
relevant
to
this
assessment.

4.0
Confined
Rotational
Crop
Studies
As
all
proposed
uses
in
this
tolerance
petition
are
on
imported
commodities,
confined
rotational
crop
studies
are
not
relevant
to
this
assessment.
Page
85
of
87
5.0
Analytical
Methodology
Table
A.
5.
Analytical
Methodology
Method
Name
Applicable
Commodities
Analytes
Extraction
Solvent(
s)
Clean­
up
Step(
s)
Determinative
Step
LLMV*
ppm
LOD,
ppm
RCC
665943
Grapes
R­
L
and
S­
L
isomers
Acetone,
Hexane
Amino
SPE
Cartridge
GC/
NPD
0.01
0.001
RCC
665943
Grape
juice,
raisins,
wine
R­
L
and
S­
L
isomers
Acetone,
Hexane
Amino
SPE
Cartridge
GC/
NPD
0.01
0.002
RCC
665943
Tomatoes
and
all
processed
tomato
commodities
R­
L
and
S­
L
isomers
Acetone,
Hexane
Amino
SPE
Cartridge
GC/
NPD
0.01
0.002
*
Although
the
submitted
reports
state
that
the
LOQ
was
0.01
ppm,
an
inspection
of
the
chromatograms
indicates
that
the
LOQ
would
probably
be
in
the
range
of
0.002­
0.003
ppm
for
the
various
grape
and
tomato
commodities.
In
many
cases
residue
values
were
reported
as
being
between
0.001
and
0.01
ppm
(
i.
e.,
actual
values
were
reported
as
0.002
ppm,
0.003
ppm,
etc.).
The
reporting
of
such
values
is
further
evidence
that
the
LOQ
is
indeed
below
0.01
ppm.

6.0
Summary
of
Magnitude
of
Residue
Studies
6.1
Plants
Table
A.
6.
Summary
of
Magnitude
of
Residue
Studies
for
Benthiavalicarb
R­
L
Isomer
S­
L
Isomer
Combined
Residues
Commodity
App
Rate
x
#
of
App
(
total)

(
lb
ai/
A)
PHI
days
No
of
trials
Range
of
Residues
(
ppm)
Avg.
(
ppm)
Range
of
Residues
(
ppm)
Avg.
(
ppm)
Range
of
Residues
(
ppm)
Avg.
(
ppm)

Grapes
0.031x6
(
0.188)
0
22
0.020­
0.226
0.081
<
0.001­
0.023
0.0068
0.031­
0.243
0.088
Grapes
0.031x6
(
0.188)
3
4
0.044­
0.073
0.060
0.006­
0.014
0.0090
0.050­
0.081
0.069
Grapes
0.031x6
(
0.188)
7
22
0.023­
0.158
0.0654
<
0.001­
0.021
0.0043
0.024­
0.168
0.070
Grapes
0.031x6
(
0.188)
14
22
<
0.001­
0.170
0.0515
<
0.001­
0.025
0.0034
<
0.002­
0.195
0.055
Grapes
0.031x6
(
0.188)
21
22
0.007­
0.138
0.0459
<
0.001­
0.019
0.0029
0.008­
0.157
0.049
Page
86
of
87
Table
A.
6.
Summary
of
Magnitude
of
Residue
Studies
for
Benthiavalicarb
R­
L
Isomer
S­
L
Isomer
Combined
Residues
Commodity
App
Rate
x
#
of
App
(
total)

(
lb
ai/
A)
PHI
days
No
of
trials
Range
of
Residues
(
ppm)
Avg.
(
ppm)
Range
of
Residues
(
ppm)
Avg.
(
ppm)
Range
of
Residues
(
ppm)
Avg.
(
ppm)

Grapes*
0.031x6
(
0.188)
28
18
0.011­
0.140
0.0424
<
0.001­
0.008
0.0036
0.012­
0.161
0.046
Grapes
(
Total)**
0.031x6
(
0.188)
28
34
0.011­
0.208
0.0621
<
0.001­
0.061
0.0077
0.012­
0.269
0.070
Greenhouse
Tomatoes
0.067x6
(
0.402)
0
4
0.050­
0.156
0.110
<
0.002­
<
0.002
0.002
0.052­
0.158
0.11
Greenhouse
Tomatoes
0.067x6
(
0.402)
1
4
0.023­
0.161
0.0893
<
0.002­
<
0.002
0.002
0.025­
0.163
0.091
Greenhouse
Tomatoes
0.067x6
(
0.402)
3
9
0.039­
0.254
0.108
<
0.002­
0.014
0.0049
0.041­
0.268
0.11
Greenhouse
Tomatoes
0.067x6
(
0.402)
7
4
0.041­
0.135
0.0898
<
0.002­
<
0.002
0.002
0.043­
0.137
0.092
Outdoor
Tomatoes
0.067x6
(
0.402)
0
3
0.017­
0.058
0.0367
<
0.002­
<
0.002
0.002
0.019­
0.060
0.039
Outdoor
Tomatoes
0.067x6
(
0.402)
1
3
0.019­
0.049
0.0293
<
0.002­
<
0.002
0.002
0.021­
0.051
0.031
Outdoor
Tomatoes
0.067x6
(
0.402)
3
3
<
0.002­
0.022
0.0127
<
0.002­
<
0.002
0.002
0.004­
0.024
0.015
Outdoor
Tomatoes
7
3
<
0.002­
0.013
0.0057
<
0.002­
<
0.002
0.002
0.004­
0.015
0.0077
Grapes*:
Grape
samples
from
the
residue
decline
studies
only.
Grapes
(
Total)**:
Grape
samples
from
all
studies
including
the
residue
decline
studies.
Trials
in
bold
print
are
those
performed
at
a
1x
application
rate
and
the
proposed
PHI.

6.2
Livestock
As
no
significant
animal
feed
items
are
associated
with
grape
and
tomato
commodities,
livestock
feeding
studies
are
not
relevant
to
this
assessment.

7.0
International
Considerations
There
are
currently
no
Canadian,
Mexican,
Codex,
or
EU
MRLs
for
benthiavalicarb.
The
registrant
has
proposed
EU
MRLs
of
0.2
ppm
for
grapes,
0.3
ppm
for
tomatoes,
and
0.01
ppm
for
potatoes.
The
MRL
status
sheet
is
attached.
Page
87
of
87
8.0
Environmental
Degradation
As
all
proposed
uses
in
this
tolerance
petition
are
on
imported
commodities,
environmental
degradation
is
not
relevant
to
this
assessment.

INTERNATIONAL
RESIDUE
LIMIT
STATUS
Chemical
Name:
Isopropyl[(
S)­
1­
{[(
1R)­
1­(
6­
fluoro­
1,3­
benzothiazol­
2­
yl)
ethyl]
carbamoyl}­
2­
methylpropyl]
carbamate
Common
Name:
Benthiavalicarb
Isopropyl
 
Proposed
tolerance

Reevaluated
tolerance

Other
Date:
10/
18/
2005
Codex
Status
(
Maximum
Residue
Limits)
U.
S.
Tolerances
XNo
Codex
proposal
step
6
or
above

No
Codex
proposal
step
6
or
above
for
the
crops
requested
Petition
Number:
3E6545
DP
Barcode:
D
Other
Identifier:

Reviewer/
Branch:
Doug
Dotson/
RAB2
Residue
definition
(
step
8/
CXL):
Not
applicable
Residue
definition:
parent
benthiavalicarb
isopropyl
Crop
(
s)
MRL
(
mg/
kg)
Crop(
s)
Tolerance
(
ppm)

Grape
0.20
Grape,
Raisin
1.0
Tomato
0.45
Tomato
Paste
0.45
Limits
for
Canada
Limits
for
Mexico
X
No
Limits

No
Limits
for
the
crops
requested
XNo
Limits

No
Limits
for
the
crops
requested
Residue
definition:
N/
A
Residue
definition:
N/
A
Crop(
s)
MRL
(
mg/
kg)
Crop(
s)
MRL
(
mg/
kg)

Notes/
Special
Instructions:
S.
Funk,
10/
19/
2005.
