Document ID: EPA-HQ-OPP-2007-0234-0004
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2007-10-24T04:00Z

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

                         WASHINGTON, D.C.  20460

OFFICE OF	

PREVENTION, PESTICIDES

AND TOXIC SUBSTANCES

	

August 22, 2007

MEMORANDUM

SUBJECT:	Fluazinam: Human Health Risk Assessment for Proposed Use on
Edible-Podded Beans, Shelled Succulent and Dried Beans, Brassica Leafy
Vegetables, Bushberries, and Ginseng.  PC Code: 129098, Petition No:
6E7139, DP Barcode: D334949

		Regulatory Action:  Section 3 Registration Action

		Risk Assessment Type:  Single Chemical Aggregate

FROM:			Karlyn J. Bailey, Risk Assessor

			William Drew, Chemist, Residue Chemistry

			Michael Doherty, Chemist, Dietary Assessment 

			Zaida Figueroa, ORE Assessor

			Registration Action Branch 2

			Health Effects Division (7509P)

			John Doherty, Toxicologist

			Reregistration Branch 3 

			Health Effects Division (7509P)

THROUGH:	Richard Loranger, Branch Senior Scientist

		Christina Swartz, Branch Chief

		Registration Action Branch 2

Health Effects Division (7509P)

TO:		Daniel Rosenblatt/Shaja Brothers, PM 05

		Risk Integration Minor Use and Emergency Response Branch 

		Registration Division (7505P)

Table of Contents

  TOC \f  1.0	Executive Summary	4

2.0	Ingredient Profile	10

2.1	Summary of Registered/Proposed Uses	11

2.2	Structure and Nomenclature	12

2.3	Physical and Chemical Properties	12

3.0	Hazard Characterization/Assessment	13

3.1	Hazard and Dose-Response Characterization	13

3.1.1	Database Summary	13

3.1.1.1	Sufficiency of studies/data	13

3.1.1.2	Mode of action, metabolism, toxicokinetic data	13

3.1.2	Toxicological Effects	14

3.1.3	Dose-response	15

3.2	Absorption, Distribution, Metabolism, Excretion (ADME)	16

3.3	FQPA Considerations	16

3.3.1	Adequacy of the Toxicity Database	16

3.3.2	Evidence of Neurotoxicity	16

3.3.3	Developmental Toxicity Studies	18

3.3.4	Reproductive Toxicity Study	18

3.3.5	Additional Information from Literature Sources	18

3.3.6	Pre-and/or Postnatal Toxicity	19

3.3.6.1	Determination of Susceptibility	19

3.3.6.2	Degree of Concern Analysis and Residual Uncertainties	19

3.4	FQPA Safety Factor for Infants and Children	20

3.5	Hazard Identification and Toxicity Endpoint Selection	20

3.5.1	Acute Reference Dose (aRfD) - General Population	20

3.5.2	Acute Reference Dose (aRfD) - Females 13-49	21

3.5.3	Chronic Reference Dose (cRfD)	21

3.5.4	Dermal Absorption	22

3.5.5	Dermal Exposure (Short-, Intermediate- and Long-Term)	23

3.5.6	Inhalation Exposure (Short-, Intermediate- and Long-Term)	23

3.5.7	Level of Concern for Margin of Exposure	24

3.5.8	Recommendation for Aggregate Exposure Risk Assessments	24

3.5.9	Classification of Carcinogenic Potential	24

3.5.10	Summary of Toxicological Doses and Endpoints for fluazinam for
Use in Human Risk Assessments	25

3.6	Endocrine disruption	26

4.0	Public Health and Pesticide Epidemiology Data	27

5.0	Dietary Exposure/Risk Characterization	27

5.1  Pesticide Metabolism and Environmental Degradation	  PAGEREF
_Toc139001018 \h  27 

5.1.1	Metabolism in Primary Crops	  PAGEREF _Toc139001019 \h  27 

5.1.2	Metabolism in Rotational Crops	28

5.1.3	Metabolism in Livestock	28

5.1.4	Analytical Methodology	28

5.1.5	Environmental Degradation	29

5.1.6	Comparative Metabolic Profile	30

5.1.7	Toxicity Profile of Major Metabolites and Degradates	31

5.1.8	Pesticide Metabolites and Degradates of Concern	31

5.1.9	Drinking Water Residue Profile	32

5.1.10	Food Residue Profile	33

5.1.11	International Residue Limits	34

5.2  Dietary Exposure and Risk	35

5.2.1  Acute Dietary Exposure/Risk	35

5.2.2  Chronic Dietary Exposure/Risk	35

5.2.3  Cancer Dietary Risk	35

5.3 Anticipated Residue and Percent Crop Treated (%CT) Information	36

6.0	Residential (Non-Occupational) Exposure/Risk Characterization	36

7.0	Aggregate Risk Assessments and Risk Characterization	36

7.1	Acute Aggregate Risk	36

7.2	Short-Term Aggregate Risk	36

7.3	Intermediate-Term Aggregate Risk	37

7.4	Long-Term Aggregate Risk	37

7.5	Cancer Risk	37

8.0	Cumulative Risk Characterization/Assessment	37

9.0	Occupational Exposure/Risk Pathway	37

9.1	Agricultural Handler Risk	37

9.2	Postapplication Risk	41

10.0	Data Needs and Label Requirements	  PAGEREF _Toc139001049 \h  46 

10.1	Toxicology	46

10.2	Residue Chemistry	46

10.3	Occupational and Residential Exposure	46

References:	47

Appendix A:  Toxicology Assessment	49

A.1  Toxicology Data Requirements	49

A.2  Toxicity Profiles	50

A.3  Executive Summaries	55

A.4  Additional Toxicology Study	84

Appendix B:  Metabolism Assessment	85

B.1	Metabolism Guidance and Considerations	85

Appendix C:  Tolerance Reassessment Summary and Table	86

 

Executive Summary

The active ingredient (ai), fluazinam, is a preventive contact fungicide
with a multi-site mode of action.  It disrupts the production of energy
at several metabolic sites within the fungal cell.  Interregional
Research Project #4 (IR-4) has submitted petitions (PPs#6E7137, 6E7139)
proposing the use of a formulation on ginseng, dry beans and succulent
bean crops, edible-podded legume vegetables, bushberry and Brassica
leafy vegetables.  ISK Biosciences Corporation is the data submitter and
registrant for the active ingredient (ai), fluazinam, in the US.  The
following products have been assessed for occupational exposure: OMEGA®
500F and Allegro® 500F.  The products are formulated as a flowable
suspension/liquid containing 40.0% fluazinam and may be applied by
airblast or groundboom (aerial application of this EP is prohibited). 
Based on the anticipated application practices for the OMEGA® 500F and
Allegro® 500F Fungicide, product labels and information provided by the
registrant, handler exposures are expected to be short- and
intermediate-term in duration. 

HUMAN HEALTH RISK ASSESSMENT:

Toxicology/Hazard

In subchronic and chronic oral and dermal studies in rats, dogs and
mice, the liver appeared to be the primary target organ.  Signs of liver
toxicity included: changes in clinical chemistry (e.g. increased serum
alkaline phosphatase, increased aspartate aminotransferase), increased
absolute and/or relative liver weights, increased incidences of gross
lesions (e.g. pale, enlarged, pitted, mottled, accentuated markings) and
increased incidences of a variety of histopathological lesions.  

Treatment-related effects were also observed in other organs in
subchronic and chronic oral, dermal and inhalation studies in rats, dogs
and mice, but these effects were not regularly noted in all three
species or in all studies in a given species.  In rats, effects observed
were decreased body weight gain, decreased food consumption, mild
anemia, increased serum cholesterol, increased serum phospholipid,
increased serum aspartate aminotransferase, testicular atrophy,
increased testes weights (inhalation study), pancreatic exocrine
atrophy, increased lung weights, increased alveolar adenomatosis,
epithelialization and macrophages, thyroid gland follicular cell
hyperplasia, and an increased incidence of thyroid gland follicular cell
tumors in male rats, but not in female rats.  In dogs, these effects
included increased salivation, increased nasal dryness, grey mottling of
the retina, mild anemia, increased serum alkaline phosphatase and
gastric lymphoid hyperplasia.  In mice, these effects included increased
mortality (at high doses), decreased body weight gain, increased serum
glucose, increased kidney weights, cystic thyroid follicules, and an
increased incidence of both benign and malignant hepatocellular liver
tumors in male mice. 

In a developmental toxicity study in rats there was evidence of
increased qualitative susceptibility (skeletal
abnormalities/facial/palate clefts in fetuses vs. decreases in
bodyweight gain/food consumption in maternal animals) of fetuses to
fluazinam; there was no evidence of increased quantitative
susceptibility.  There was no evidence of increased quantitative or
qualitative susceptibility in a developmental toxicity study in rabbits
or a 2-generation reproduction study in rats.  Effects included:
increased incidences of total litter resorptions and fetal skeletal
abnormalities (eg. kinked tail tip, fused or incompletely ossified
sternebrae, and abnormalities of head bones) in rabbits and decreased
body weight gain in rats.  Reproductive effects were a decreased number
of implantation sites and decreased litter size.

In an acute neurotoxicity study in rats, there were decreases in motor
activity and soft stools at high doses (1000 mg/kg/day).  In two
subchronic neurotoxicity studies (evaluated together) there were no
signs of neurotoxicity observed up to 280 mg/kg/day.  A neurotoxic
lesion described as vacuolation of the white matter of the central
nervous system was observed in studies in mice and dogs; this lesion was
found to be reversible and is attributed to an impurity (impurity-5). 

A developmental neurotoxicity study in rats and a series of special
studies were submitted to address the issues of increased susceptibility
and the presence of neurotoxic lesions observed in the toxicological
database.  As a result, there are no residual uncertainties with regard
to pre- and/or postnatal toxicity and no additional factors are needed. 
Additionally, HED recommends the FQPA SF be reduced to 1X because the
toxicology database is complete in regard to pre-and postnatal toxicity
and neurotoxicity; the dietary food exposure assessment is based on
HED-recommended tolerance-level residues and assumes 100% crop treated
for all commodities, resulting in upper bound estimates of dietary
exposure; the drinking water assessment is based on values generated by
model and associated modeling parameters which are designed to provide
conservative, health protective upper bound estimates of water
concentrations; and there are no registered or proposed residential
uses.  

   

The Cancer Assessment Review Committee (CARC) classified fluazinam as
“Suggestive evidence of carcinogenicity, but not sufficient to assess
human carcinogenic potential,” and determined that quantification of
human cancer risk is not required.

For acute dietary exposure (females 13-49), the developmental toxicity
study in rabbits was used to calculate the acute reference dose (aRfD)
of 0.07 mg/kg/day.  The developmental NOAEL of 7 mg/kg/day and the LOAEL
of 12 mg/kg/day were based on increased incidences of total litter
resorptions and slight increased incidences of fetal skeletal
abnormalities.  The aRfD of 0.5 mg/kg/day calculated for general
population acute dietary exposure was based on a LOAEL of 1000 mg/kg/day
(NOAEL=50 mg/kg/day) from an acute neurotoxicity study in rats; adverse
effects seen were decreases in motor activity and soft stools on the day
of dosing.  For chronic dietary exposure (all populations), the
carcinogenicity study in mice was used to calculate the chronic
reference dose (cRfD) of 0.011 mg/kg/day.  The NOAEL of 1.1 mg/kg/day
and the LOAEL of 10.7 were based on adverse liver alterations (increased
liver weights and histopathology).  A 21-day dermal toxicity study in
rats was used to select the dose and endpoint for occupational short-
and intermediate-term dermal exposure.  The NOAEL of 10 mg/kg/day and
the LOAEL of 100 mg/kg/day were based on increased cholesterol and
aspartate aminotransferase.  For occupational short- and
intermediate-term inhalation exposure, a 7-day inhalation study in rats
was used.  The NOAEL of 1.38 and the LOAEL of 3.87 mg/kg/day were based
on increased liver weights and testes weights in males.  In the
inhalation study, a histopathological examination was not performed;
thus an additional factor of 10x was applied to the conventional
uncertainty factor of 100x.  This factor also addresses the use of a
short-term (7 days) study to evaluate intermediate-term inhalation
exposure.  There are no residential uses proposed for fluazinam;
therefore, incidental oral and residential dermal and inhalation risk
assessments were not conducted.  

Dietary Exposure (Food/Water)

HED performed both the acute and chronic analyses that are based on
tolerance-level residues, assume 100% crop treated, and incorporate
modeled estimated drinking water concentrations (EDWCs).  Therefore, the
resulting exposure and risk estimates should be considered high-end and
very conservative.  

The acute risk estimates are below HED’s level of concern for all
population subgroups, including those of infants and children. 
Generally, HED is concerned when risk estimates exceed 100% of the
population-adjusted dose (PAD).  The acute risk estimate for the U.S.
population, as a whole, is 1% of the acute PAD (aPAD).  For females
13-49 years of age, the risk estimate is 8% of their aPAD.  Risk
estimates for all other population subgroups are less than 8% aPAD. 
Likewise, chronic risk estimates are below HED’s level of concern for
all population subgroups.  The risk estimate for the U.S. population is
9% of the chronic PAD (cPAD).  The highest risk estimate is for All
Infants (< 1 year) population subgroup at 16% cPAD.

The nature of the residue in plants has been adequately delineated,
based on acceptable potato, peanut, and grape metabolism studies
reviewed previously (D257115; William Cutchin; 5/21/2001), along with an
acceptable apple metabolism study submitted recently (MRID# 46991301). 
At a meeting held on 11/28/2000, HED concluded that the residue of
concern (ROC) in potatoes and peanuts (for both tolerance expression and
dietary risk assessment purposes) was the parent compound only (D272624;
William Cutchin; 4/23/2001).  In wine grapes, both parent and AMGT were
included in the ROC for tolerance expression and risk assessment. 
Additionally, HED determined that data generated for potential new uses
on other crops (with the exception of root and tuber, and bulb
vegetables) should include analyses for both parent and AMGT; thus, the
ROC for the proposed primary crops are parent and AMGT.  

The nature of the residue in livestock is also understood, based on
adequate goat and hen metabolism studies (D257115; William Cutchin;
5/21/2001).  The fluazinam residues of regulatory interest in animals
were determined by HED to be parent plus the metabolites AMPA and DAPA,
and their sulfamate conjugates.  

The submitted gas chromatography with electron-capture detection
(GC/ECD) methods (modifications of the tolerance-enforcement method) are
adequate for collecting data and enforcing tolerances for residues of
fluazinam per se in the various crop commodities associated with these
petitions.  The tolerance-enforcement method, Fluazinam:  Method for the
Analysis in Peanut Nut Meat (MRID #43521016), was adequately
radiovalidated, and underwent a successful independent laboratory
validation (ILV) trial.  The method was forwarded to BEAD’s Analytical
Chemistry Branch (ACB) for a petition method validation (PMV) trial, and
was subsequently determined to be suitable as a tolerance-enforcement
method (D266802; Paul Golden; 6/22/2001).  

The submitted high performance liquid chromatography with ultraviolet
detection (HPLC/UV) method (a working method based on Method Evaluation
for the Analysis of AMGT in Grapes, MRID #45593101) is adequate for
collecting data on AMGT residues in blueberries.  The LLMV, limit of
detection (LOD), and LOQ were 0.020, 0.013, and 0.038 ppm, respectively,
for residues of AMGT in blueberries.  HED has previously determined that
residues of AMGT are to be regulated in wine grapes (D272624; William
Cutchin; 4/23/2001).  The Agency therefore requested that this method
undergo an ILV trial, and, potentially, a PMV trial by the ACB.  An ILV
study has not yet been submitted. 

 

The multiresidue method (MRM) testing data indicate that fluazinam is
partially recovered through Sections 302, 303, and 304 of PAM Volume I,
with its recovery being dependent on which Florisil elution system is
used.  The MRMs can serve as a confirmatory procedure for residues of
fluazinam.  Data should also be provided for the metabolite AMGT, since
it is included in the tolerance expression for grapes.  

ials were ≤ LOQ (≤ 0.010 ppm).  

The available crop field trial data are adequate, and support the
proposed uses.  However, it was noted that residue data for AMGT were
provided only for blueberries; AMGT data should also have been included
with the field trial studies for edible-podded beans, shelled succulent
and dried beans, and Brassica vegetables. 

 

There are no processed commodities for which residue data are required
associated with the proposed uses on the crops requested in the subject
petitions under review.  

There are no significant livestock feed items associated with the
proposed uses on the crops requested in the subject petitions under
review. 

 

Regulatory requirements pertaining to fluazinam residues in rotational
crops have been fulfilled, and the rotational crop restrictions on the
proposed label are adequate. 

 

  SEQ CHAPTER \h \r 1 There are no established or proposed Canadian or
Codex Maximum Residue Limits (MRLs) for residues of fluazinam in plant
or animal commodities.  There are Mexican MRLs established for residues
of fluazinam in potato at 0.05 ppm, and in beans at 0.1 ppm.

 

Occupational Exposure/Risk

Handlers

No chemical-specific data for assessing handler exposures were submitted
to the Agency in support of the proposed uses.  As a result, HED used
surrogate data from the Pesticide Handlers Exposure Data Base (PHED)
Version 1.1, and standard values established by the Health Effects
Division (HED) Science Advisory Council for Exposure, for acres treated
per day, body weight, and the level of personal protective equipment
(PPE) worn by handlers.  HED’s level of concern (LOC) for occupational
dermal exposures is 100 (i.e., MOE less than 100 is of concern).  The
level of concern for inhalation exposures is 1000 (i.e., MOE less than
1000 is of concern).  

All   SEQ CHAPTER \h \r 1 dermal risk estimates for short- and
intermediate-term handler exposure resulted in MOEs greater than 100
with the use of gloves.   All inhalation   SEQ CHAPTER \h \r 1 risk
estimates for short- and intermediate-term handler exposure resulted in
MOEs greater than 1000 with the use of a dust mist respirator. 
Summaries of the dermal and inhalation (MOEs) short- and
intermediate-term risks for handlers are provided in Table 9.1.2.

Postapplication 

Chemical-specific postapplication data were submitted in support of this
registration action. For purposes of comparison, a Tier 1 (HED standard
assumptions and defaults of 20% DFR) and Tier 2 (chemical-specific
foliar residue data) analysis were performed to ensure that potential
postapplication exposures are not of concern.  A comparison of Tier 1
and Tier 2 analyses resulted in similar postapplication exposure risks
of concern (MOE < 100).  

Since the Tier 1 and Tier 2 analyses resulted in similar exposures and
risks , HED based its postapplication assessment on the Tier 2 analysis.
 The only crop scenarios which resulted in MOEs greater than 100 on day
0 (immediately after application) were for low exposure activities
(i.e., scouting, hand weeding, thinning and irrigation) for beans and
ginseng.  All other crops (i.e. bushberries, Brassica and leafy
vegetables) did not reach a MOE greater than or equal to 100 for low
exposure activities until 3 to 13 days later.  All medium (i.e.,
scouting, hand weeding, and irrigation) and high (i.e., hand harvesting/
pruning/pinching/training) postapplication exposure activities for all
crops resulted in MOEs below 100 on day of application.  Crops did not
reach MOEs greater than or equal to 100 until 4 to 20 days later
depending on the specific crop.   

Since postapplication exposure resulting in MOEs below 100 may be an
indication of possible risk for re-entry of workers, HED provided a
comparison of the estimated number of  days required before a MOE of 100
is reached (i.e. restricted entry interval – REI) based on Tier 2
analysis to establish pre-harvest interval (PHI).  HED recommends that
the Registration Division ensure that the PHIs do not go below the
calculated REIs for harvesting.  

The Tier 2 postapplication estimates of exposure may be overestimating
residues on the proposed crops based on the methodology used to
determine dislodgeable residues.  However, HED cannot refine these
estimates without chemical specific data collected in accordance with
Agency guideline methods.  A possible option for the registrant would be
to repeat the DFR studies using guideline methods (i.e., leaf punch and
dislodgeable residues with surfactant as opposed to whole leaf
extraction). 

Restricted Entry Interval 

The technical material has an Acute Eye Irritation Toxicity Category I. 
Per the Worker Protection Standard (WPS), a 48-hr restricted entry
interval (REI) is required for chemicals classified under Toxicity
Category I.  The 48 hour REI appearing on the label is only appropriate
for postapplication activities for which the MOE reaches 100 on day 0. 
However, note that an interval of 3 to 20 days is necessary to reach a
MOE of 100 for medium and high postapplication exposure activities
(i.e., hand weeding/harvesting/pruning/pinching/training, and
irrigation).  HED recommends that the proposed label be revised to
ensure that the appropriate REI restrictions are clearly stated for all
crops and do not exceed pre-harvest intervals. 

Environmental Justice Considerations

Potential areas of environmental justice concerns, to the extent
possible, were considered in this human health risk assessment, in
accordance with U.S. Executive Order 12898, "Federal Actions to Address
Environmental Justice in Minority Populations and Low-Income
Populations"   HYPERLINK
"http://www.eh.doe.gov/oepa/guidance/justice/eo12898.pdf" 
http://www.eh.doe.gov/oepa/guidance/justice/eo12898.pdf ). 

As a part of every pesticide risk assessment, OPP considers a large
variety of consumer subgroups according to well-established procedures. 
In line with OPP policy, HED estimates risks to population subgroups
from pesticide exposures that are based on patterns of that subgroup’s
food and water consumption, and activities in and around the home that
involve pesticide use in a residential setting.  Extensive data on food
consumption patterns are compiled by the USDA under the Continuing
Survey of Food Intakes by Individuals (CSFII) and are used in pesticide
risk assessments for all registered food uses of a pesticide.  These
data are analyzed and categorized by subgroups based on age, season of
the year, ethnic group, and region of the country.  Whenever
appropriate, non-dietary exposures based on home use of pesticide
products and associated risks for adult applicators and for toddlers,
youths, and adults entering or playing on treated areas postapplication
are evaluated.  Further considerations are currently in development as
OPP has committed resources and expertise to the development of
specialized software and models that consider exposure to bystanders and
farm workers as well as lifestyle and traditional dietary patterns among
specific subgroups.

Review of Human Research

This risk assessment relies in part on data from studies in which adult
human subjects were intentionally exposed to a pesticide or other
chemical.  These studies, which comprise the Pesticide Handlers Exposure
Database (PHED), have been determined to require a review of their
ethical conduct, and have received that review.  The studies in PHED
were considered appropriate (ethically conducted) for use in risk
assessments.  

Additional Data Needs/Recommendations 

Toxicology

A 28-day subchronic inhalation study is recommended to support the
registration of fluazinam.  If an acceptable subchronic inhalation study
is submitted and determined to be more appropriate for endpoint
selection, the additional 10x safety factor can be reduced.   

Residue Chemistry

No major deficiencies were noted in the subject petition that would
preclude the establishment of permanent tolerances for fluazinam
residues in the requested crops.  Revised Sections F should be
submitted, so that the proposed tolerances reflect the recommended
tolerance levels, and correct commodity definitions, as specified in
Appendix C.  Issues pertaining to residue chemistry deficiencies should
be resolved (see below).  

Regulatory Recommendations

As a condition of registration, results of an ILV trial for the AMGT
analytical method (with wine grapes) should be submitted by the
registrant.  If the registrant agrees with the modifications made by
Ricerca to the original method (in MRID #45593101), these modifications
should be incorporated into a revised method for the ILV.  Sample sets
should include, at the minimum, 2 control (untreated) samples of wine
grapes, 2 samples fortified at the tolerance level (3.0 ppm), and 2
samples fortified at the LOQ (0.010 ppm). 

 

As a condition of registration, MRM recovery data should be provided for
the metabolite AMGT, since it is included in the tolerance expression
for wine grapes. 

The product label for Omega 500F should be amended to include a
restriction, stating that turnip roots from turnip plants treated with
this EP must not be used for human nor livestock consumption.  

The Agency has previously determined, and the registrant is hereby
advised again, that residue data for AMGT should be provided in the crop
field trial studies for all future requested plant commodities, except
root and tuber, and bulb vegetables.  

HED recommends in favor of establishing permanent tolerances for
fluazinam in the requested crops, at the levels specified in Appendix C
(Table C.1. Tolerance Summary of Fluazinam).

  TC \l2 "10.1	Toxicology Occupational and Residential Exposure

HED recommends that the Registration Division ensure that the PHIs do
not go below calculated REIs for harvesting.  Additionally, HED
recommends that the proposed label be revised to ensure that the
appropriate REI restrictions are clearly stated for all crops and
correspond to the postapplication activities and reentry intervals.

  TC \l1 "1.0	Ex

ecutive Summary 2.0	Ingredient Profile

Fluazinam (Omega 500F Agricultural Fungicide) is a non-systemic,
preventive, contact fungicide of the phenyl-pyridinamine class, with a
multi-site mode of action.  It disrupts the production of energy at
several metabolic sites within the fungal cell.  Fluazinam is a
protectant fungicide; when applied to plants, it remains primarily on
the plant surface, is not taken up to any extent by the plant, and is
not translocated within the plant.

2.1	Summary of Registered/Proposed Uses

Fluazinam is currently registered for use on peanuts and potatoes. 
There is also a tolerance established for fluazinam in imported wine
grapes (without US registration).  Permanent tolerances are established
for residues of fluazinam in peanuts and potatoes at 0.02 ppm (40CFR
§180.574[a][1]), and in imported wine grapes at 3.0 ppm (40CFR
§180.574[a][2]). 

Interregional Research Project #4 (IR-4) has submitted petitions
(PPs#6E7137, 6E7139) proposing the use of a formulation containing 4.17
pounds per gallon (lb/gal) of fluazinam (Omega 500F Agricultural
Fungicide; EPA Registration #71512-1) on various crops.  This end-use
product (EP) is formulated as a flowable-suspension (F) concentrate. 
ISK Biosciences Corporation is the data submitter and registrant for the
active ingredient (a.i), fluazinam, in the US.  Copies of the proposed
labels were provided, and the proposed uses on the requested crops are
summarized in Table 2.1 (below).  Applications of Omega 500F are to be
made using ground equipment or chemigation (application via irrigation
equipment) only; aerial application of this EP is prohibited.  

Table 2.1	Summary of Directions for Use of Fluazinam.

Application Timing; Type; and Equipment 1	Form-ulation 2

	Use Rate

(lb ai/A)	Maximum # of Uses per Season	Maximum Seasonal Use Rate (lb
ai/A)	PHI

(Days)	Use Directions and Limitations

Shelled Succulent and Dried Beans

At 10-30% bloom; foliar; spray.	Omega 500F

	0.26-0.45	2	0.90	30	RTI = 7-10 days. Volume adequate to cover foliage
and flowers.

Ginseng

At transplant (for root rot); broadcast; spray.	Omega 500F	0.52	6	3.1	30
RTI = 14 days.  Spray volume ≥ 100 gal/A.

At disease appearance or favorable conditions (for blight/white mold);
broadcast; spray.

0.52-0.78	4-6

	RTI = 7-14 days.  Spray volume ≥ 100 gal/A.

Edible-Podded Beans

At 10-30% bloom; foliar; spray.	Omega 500F	0.26-0.45	2	0.90	14	RTI =
7-10 days. Volume adequate to cover foliage and flowers.

Brassica (Cole) Vegetables

At transplant; soil drench; spray.	Omega 500F	0.055 lb ai/1000 plants	1
2.0	20/50 3	6.45 oz EP/100 gal water, 3.4 oz  (100 mL)/plant

Prior to transplant; soil incorporation; precision incorporator.

1.36

Band width ≥9”, soil depth 6-8”. Spray volume ≥ 50 gal/A.

Prior to forming bed; broadcast; spray.

2.0

Spray volume ≥ 50 gal/A.

Bushberries

1Green tip, 2pink tip, 3early bloom, 4full bloom, 5blossom drop, 6small
green fruit/some blue fruit; foliar; spray.	Omega 500F	0.65	6	3.9	30	RTI
= 7-10 days.  Volume adequate to cover foliage, flowers, and fruit.

1. Applications of Omega 500F are to be made using ground equipment or
chemigation (application via irrigation

    equipment) only.  Aerial application of this EP is prohibited.  

2. Omega 500F is a flowable suspension concentrate containing 4.17
lb/gal of fluazinam.  

3. PHI = 20 days for Brassica leafy greens, and 50 days for Brassica
heading vegetables.

2.2	Structure and Nomenclature

  SEQ CHAPTER \h \r 1 TABLE 2.2	Test Compound Nomenclature.

Compound	Chemical Structure

                              

Empirical Formula	

C13H4Cl2F6N4O4

Molecular Weight	465.1

Common Name	Fluazinam

Company Experimental Names	Fluazinam, IKF-1216

IUPAC Name
3-chloro-N-(3-chloro-5-trifluoromethyl-2-pyridyl)-α,α,α-trifluoro-2,6
-dinitro-p-toluidine

CAS Name
3-chloro-N-[3-chloro-2,6-dinitro-4-(trifluoromethyl)phenyl]-5-(trifluoro
methyl)-2-pyridinamine

CAS Number	79622-59-6

End-use Product/(EP)	Omega® 500F (USA); Allegro® 500F (Canada)

Physical and Chemical Properties

  SEQ CHAPTER \h \r 1 TABLE 2.3	Physicochemical Properties of Fluazinam

Parameter	Value	Reference

Melting Point/Range	115-117°C	The e-Pesticide Manual (13th Edition)
Version 3.1

pH	5.85	MRID #43521001

Density (25°C)	1.02 g/cm3*	LSS 2000_1973_2LS_rev

Water Solubility (25°C)	(pH buffered to 5) 0.131 mg/L

(pH buffered to 7) 0.157 mg/L

(pH buffered to 9) 3.384 mg/L	LSS 2000_1973_2LS_rev

Solvent Solubility (25°C)	Solvent

	Solubility (mg/mL)	LSS 2000_1973_2LS_rev

	acetone

dichloromethane

ethyl acetate

ethyl ether

hexane

methanol

octanol

toluene	853

675

722

231

8

192

41

451

	Vapor Pressure 	Temp (°C)	Vap. Press. (Pa)	LSS 2000_1973_2LS_rev

	25

35

45	2.3 x 10-5

1.3 x 10-4

6.7 x 10-5

	Dissociation Constant (pKa)	Average pKa = 7.22 in 50% ethanol/water
(v/v)	LSS 2000_1973_2LS_rev

Octanol/Water Partition Coefficient (Log [KOW])	1.08 x 104 (Log Kow =
4.03)	LSS 2000_1973_2LS_rev

UV/Visible Absorption Spectrum	pH	λmax (nm)	Regulatory Note REG2003-12

	5

7

>10	238

239, 342

260, 343, 482

	  SEQ CHAPTER \h \r 1 *REG2003-12 states the relative density as 1.76
g/cm3, temperature not stated.  

  TC \l2 "2.3	Physical and Chemical Properties 

3.0	Hazard Characterization/Assessment 

3.1	Hazard and Dose-Response Characterization

3.1.1	Database Summary  TC \l3 "3.1.1	Database Summary 

3.1.1.1	Sufficiency of studies/data

 

Based on the proposed use pattern, the toxicology database for fluazinam
is complete and adequate for risk assessment.  There are acceptable
studies available for endpoint selection that include: 1) subchronic
oral toxicity studies in rats, mice, and dogs; 2) a chronic oral
toxicity study in dogs and carcinogenicity studies in rats and mice; 3)
developmental and reproduction studies in rats and a developmental study
in rabbits; and  4) a subchronic dermal toxicity study in rats.  There
is also a complete mutagenicity battery, acute LD50 and neurotoxicity
studies (acute, subchronic, and developmental), as well as a metabolism
study in the rat.

Mode of action

Fluazinam is a preventive contact fungicide with a multi-site mode of
action.  It disrupts the production of energy at several metabolic sites
within the fungal cell.  Fluazinam is a protectant fungicide; when
applied to plants, it remains primarily on the plant surface.  It is not
taken up to any extent by the plant, and is not translocated within the
plant like systemic fungicides.

3.1.2	Toxicological Effects

In subchronic and chronic oral and dermal studies in rats, dogs and
mice, the liver appeared to be a primary target organ.  Signs of liver
toxicity included: changes in clinical chemistry (e.g. increased serum
alkaline phosphatase, increased aspartate aminotransferase), increased
absolute and/or relative liver weights, increased incidences of gross
lesions (e.g. pale, enlarged, pitted, mottled, accentuated markings),
and a variety of histopathological lesions.  Microscopic liver lesions
included: eosinophilic or basophilic hepatocytes, rarefied or vacuolated
hepatocytes, altered hepatocytic foci, hepatocytic single cell necrosis,
hepatocytic hypertrophy, hepatocellular fatty changes, increased brown
pigmented macrophages, sinusoidal chronic inflammation, pericholangitis,
and bile duct hyperplasia.    

Treatment-related effects were also observed in other organs in
subchronic and chronic oral, dermal and inhalation studies in rats, dogs
and mice, but these effects were not regularly noted in all three
species or in all studies in a given species.  In rats, effects observed
were decreased body weight gain, decreased food consumption, mild
anemia, increased serum cholesterol, increased serum phospholipid,
increased serum aspartate aminotransferase, testicular atrophy,
increased testes weights (inhalation study), pancreatic exocrine
atrophy, increased lung weights, increased alveolar adenomatosis,
epithelialization and macrophages, thyroid gland follicular cell
hyperplasia, and an increased incidence of thyroid gland follicular cell
tumors in male rats, but not in female rats.  In dogs, effects included
increased salivation, increased nasal dryness, grey mottling of the
retina, mild anemia, increased serum alkaline phosphatase and gastric
lymphoid hyperplasia.  In mice, increased mortality (at high doses),
decreased body weight gain, increased serum glucose, increased kidney
weights, cystic thyroid follicules, and an increased incidence of both
benign and malignant hepatocellular liver tumors (males) were seen. 

In a developmental toxicity study in rats there was evidence of
increased qualitative susceptibility of fetuses to fluazinam; there was
no evidence of increased quantitative susceptibility. Fetal exposure of
250 mg/kg/day resulted in decreases in body weights, decreased placental
weights, and increased incidences of facial/palate clefts, diaphragmatic
hernia and delayed ossification in several bone types.  There was also
greenish amniotic fluid and increases in late resorptions, as well as
postimplantation loss.   Maternal effects observed at the same dose
level were decreases in body weight gain and food consumption, increases
in water consumption, and increased urogenital staining.  

There was no evidence of increased quantitative or qualitative
susceptibility in a developmental toxicity study in rabbits or a
2-generation reproduction study in rats. 

In the developmental rabbit study, there were decreases in food
consumption and increased liver histopathology in maternal animals at 7
mg/kg/day.  At the higher dose of 12 mg/kg/day, fetal toxicity was
observed in the form of increased incidences of total litter resorptions
and a slight increased incidence of fetal skeletal abnormalities (eg.
kinked tail tip, fused or incompletely ossified sternebrae, and
abnormalities of head bones).  In the rat reproduction study, liver
pathology (hepatocytic fatty changes) was observed in parental F1 males
at 9.7 mg/kg/day.  Reproductive toxicity was manifested as a decreased
number of implantation sites and decreased litter sizes to day 4 post
partum for F1 females (F2 litters) at 53.6 mg/kg/day.  At 42.12
mg/kg/day, developmental effects observed were limited to decreased body
weight gain during lactation for both F1 and F2 pups.        

In an acute oral neurotoxicity study in rats, there were decreases in
motor activity and soft stools observed on the day of dosing at 1000
mg/kg/day.  These effects were considered due to systemic toxicity and
not a result of frank neurotoxicity.  In two subchronic neurotoxicity
studies (evaluated together) in rats there were no signs of
neurotoxicity observed up to 280 mg/kg/day.  A neurotoxic lesion
described as vacuolation of the white matter of the central nervous
system (CNS) was observed initially in long-term (1-2 year) chronic
studies on mice and dogs and later, upon careful re-examination of the
CNS, also in shorter-term (4-week to 90-day) subchronic studies on mice
and dogs.  This lesion was observed during the histopathology
examination of several tissues of the CNS and occurred most frequently
in the brain (sections of cerebrum and/or sections of cerebellum, pons,
medulla, and midbrain) and less frequently in the cervical spinal cord. 
This lesion is reversible and is attributed to an impurity-5 (see
section 3.3.2.).    

   

In a combined chronic/carcinogenicity study in rats, increased
incidences of thyroid gland follicular cell tumors were observed in male
rats; there were no treatment-related increases in female rats.  The
Cancer Assessment Review Committee (CARC) concluded that there was some
evidence that the thyroid tumors observed in the male rats were
treatment-related.  In two carcinogenicity studies in mice, increased
incidences of hepatocellular tumors were observed in males with no
treatment-related tumors observed in the females.  In one study, the
CARC concluded that there was clear evidence of treatment-related
increases in both benign and malignant liver tumors in the male mice. 
In the other study, it was concluded that there was equivocal/some
evidence for hepatocellular tumors in the male mice.  There is no
evidence of mutagenicity after exposure to fluazinam.  In accordance
with the EPA Draft Guidelines for Carcinogen Risk Assessment (July 2,
1999), the CARC classified fluazinam into the category “Suggestive
evidence of carcinogenicity, but not sufficient to assess human
carcinogenic potential.”  The CARC also determined that the
quantification of human cancer risk was not required.  

Dose-response

For acute dietary exposure (females 13-49), the developmental toxicity
study in rabbits was used to calculate the acute reference dose (aRfD)
of 0.07 mg/kg/day.  The developmental NOAEL of 7 mg/kg/day and the LOAEL
of 12 mg/kg/day were based on increased incidences of total litter
resorptions and slight increased incidences of fetal skeletal
abnormalities.  The aRfD of 0.5 mg/kg/day calculated for general
population acute dietary exposure was based on a LOAEL of 1000mg/kg/day
(NOAEL=50mg/kg/day) from an acute neurotoxicity study in rats; adverse
effects seen were decreases in motor activity and soft stools on the day
of dosing.  For chronic dietary exposure (all populations), the
carcinogenicity study in mice was used to calculate the chronic
reference dose (cRfD) of 0.011 mg/kg/day).  The NOAEL of 1.1 mg/kg/day
and the LOAEL of 10.7 were based on adverse liver alterations (increased
liver weights and histopathology).  A 21-day dermal toxicity study in
rats was used to select the dose and endpoint for occupational short-
and intermediate-term dermal exposure.  The NOAEL of 10 mg/kg/day and
the LOAEL of 100 mg/kg/day were based on increased cholesterol and
aspartate aminotransferase.  For occupational short- and
intermediate-term inhalation exposure, a 7-day inhalation study in rats
was used.  The NOAEL of 1.38 and the LOAEL of 3.87 mg/kg/day were based
on increased liver weights and testes weights in males.  In the
inhalation study, a histopathological examination was not performed;
thus an additional factor of 10x was applied to the conventional
uncertainty factor of 100x.  This factor also addresses the use of a
short-term (7 days) study to evaluate intermediate-term inhalation
exposure.  There are no residential uses proposed for fluazinam;
therefore, incidental oral and residential dermal and inhalation risk
assessments were not conducted.  

Absorption, Distribution, Metabolism, Excretion (ADME)

  TC \l2 "3.2	Absorption, Distribution, Metabolism, Excretion (ADME) 

Overall recovery of the administered radioactivity (reported in MRID
Nos. 43521006, 43521007, and 43521008) was acceptable (93.10-103.55%). 
Excretion via the urine was minor.  AMPA mercapturate and DAPA, the
major urinary metabolites, represented only 0.05-0.39% of the
administered dose.  Radioactivity in the feces represented most of the
administered dose (88.78-100.03%) as determined by review of MRID Nos.
43521006, 43521007, and 43521008.  Identified fecal metabolites,
however, represented from 11.20-68.59% of the administered dose. For all
dose groups, most of the fecal radioactivity appeared to reside with
unextractable components in the post-extraction solids (PES).  Further
analysis of the PES components using base hydrolysis indicated that most
of this radioactivity could be attributed to hydrolysis products of AMPA
and DAPA.   PES radioactivity was also greatest for the low-dose group
which was consistent with the lower overall accounting of identified
metabolites for this group.  Approximately 20-25% of the aqueous phase
of the fecal extraction was identified as a cysteine conjugate of DAPA
and represented <1% of the administered dose. With the exception of the
low-dose group, parent compound represented most of the identified
radioactivity in the feces.  AMPA and DAPA were identified in the feces
from all dose groups but these metabolites never represented more than
5% of the administered dose (except for high-dose female rats where AMPA
accounted for 10.22%).  

DAPA glucuronide  and AMPA mercapturate were the major biliary
metabolites but represented  < 4% of the administered dose.  Total
biliary radioactivity, however represented 25-34% of the administered
dose (MRID Nos. 43521006, 43521007, and 43521008).  Analysis of
chromatograms indicated that numerous other metabolites were present in
the bile but were individually of insufficient quantity to allow for
characterization.  

Metabolite profiles from administration of different label positions
(pyridyl and phenyl) indicated that there was no metabolic cleavage of
the ring structures.  Minor quantitative differences in metabolite
recovery were observed between genders but not of sufficient magnitude
to suggest biologically relevant differences in the metabolism of
IKF-1216

3.3	FQPA Considerations  TC \l2 "3.3	FQPA Considerations 

3.3.1	Adequacy of the Toxicity Database

	

3.3.2	Evidence of Neurotoxicity

 

There was no evidence of neurotoxicity observed in an acute
neurotoxicity study (MRID 44807210) in rats up to 1000 mg/kg/day. 
Systemic effects observed were decreases in motor activity and soft
stools observed on the day of dosing.  In two subchronic neurotoxicity
studies (MRIDs 44807217& 44807218), evaluated together, there were no
signs of neurotoxicity or systemic effects observed up to 280 mg/kg/day.
 A neurotoxic lesion described as vacuolation of the white matter of the
central nervous system (CNS) was observed initially in long-term (1-2
year) chronic studies in mice and dogs and later, upon careful
re-examination of the CNS, also in shorter-term (4-week to 90-day)
subchronic studies in mice and dogs.  This lesion was observed during
histopathological examination of several tissues of the CNS and occurred
most frequently in the brain (sections of cerebrum and/or sections of
cerebellum, pons, medulla, and midbrain) and less frequently in the
cervical spinal cord.  Although this lesion was also observed in control
animals, the increased incidence and/or severity of the lesion in
treated animals was clearly treatment-related and dose-related.  
Further investigation of this lesion in a series of special studies
demonstrated the same lesion could also be induced in rats.  In the
special studies, the following was also determined.

 

1.  Fluazinam, per se, was not responsible for the induction of this
lesion.  An analysis of the effects of impurities present in technical
grade fluazinam revealed that one single impurity, Impurity-5, was
solely responsible for the appearance of white matter vacuolation. 

 

2.  No significant differences in susceptibility or in incidence or
severity of vacuolation of the white matter of the CNS were observed
among species (mice, dogs, or rats).  Similarly, no significant
differences were attributed to sex.

3.  White matter vacuolation in the CNS was reversible.  Electron
microscopy of the white matter (cerebellum) of mice treated with
technical grade fluazinam indicated that treatment-related effects were
confined to the myelin sheaths.  Large vacuoles were observed in the
intramyelin sheaths due to the accumulation of fluid between the
sheaths.  The nucleus and mitochondria in oligodendroglia were observed
to remain intact, suggesting no damage to these cells. The myelin
sheaths appeared to recover completely during a recovery period of up to
56 days.

4.  There appears to be a non-linear dose-response with a clear
threshold below which no effect occurs.  It was concluded that a LOAEL
of 0.1 mg/kg/day and a NOAEL of 0.02 mg/kg/day for CNS effects could be
established for Impurity-5. 

A developmental neurotoxicity study (MRID 46534401) was submitted to
address the concerns regarding the white matter vacuolization observed
in the subchronic and chronic studies in mice and dogs, as well as the
increased qualitative susceptibility seen in the developmental rat
study.  In the study, there was no evidence of vacuolation of the brain
or any other treatment-related pathology seen in dams or pups up to 50
mg/kg/day.  Treatment-related effects observed in pups were decreases in
body weight and body weight gain (lactation), and delayed preputial
separation seen in the absence of maternal toxicity.    

As previously stated, the brain lesions (vacuolation of the brain and
nerve tissue) observed  are attributed to impurity #5. The level of
impurity 5 in the DNT study was 0.09%.  At the current maximum
concentration of Impurity-5 in technical grade fluazinam of 0.1% [see
memorandum from Indira Gairola, Technical Review Branch, RD (7505C) to
Cynthia Giles-Parker, Fungicide Branch, RD (7505C), dated May 18, 2001,
DP Barcode D272455], the NOAEL for CNS effects of 0.02 mg/kg/day for
Impurity-5 is equivalent to a NOAEL for CNS effects of 20 mg/kg/day for
technical grade fluazinam.  The aRfDs (0.07 and 0.5 mg/kg/day) and cRfD
(0.011 mg/kg/day), as well as risk assessments for dermal (10 mg/kg/day)
and inhalation (1.38 mg/kg/day+ additional 10x) exposures are set at
doses much lower than 20 mg/kg/day.  Therefore, they are protective of
any possible neurotoxic effects resulting from exposure to impurity #5. 

Based on the results of the developmental neurotoxicity study and the
overall weight of the evidence, additional neurotoxicity studies are not
needed.   

  TC \l3 "3.3.2	Evidence of Neurotoxicity 

3.3.3	Developmental Toxicity Studies

In a developmental toxicity study in rats, decreased body weight gain
and food consumption and increased water consumption and urogenital
staining were observed in maternal animals at the LOAEL of 250 mg/kg/day
(NOAEL= 50 mg/kg/day).  Additionally at 250 mg/kg/day, there were
treatment-related decreases in body weights and placental weights;
increases in facial/palate clefts, diaphragmatic hernia, and delayed
ossification; as well as slight increases in late resorptions and
postimplantation loss observed in fetuses.  In the developmental
toxicity study in rabbits, maternal effects included decreases in food
consumption and liver histopathology at the LOAEL of 7 mg/kg/day
(NOAEL=4 mg/kg/day).  In fetuses, skeletal anomalies and an increased
incidence of total litter resorptions were noted at 12 mg/kg/day
(NOAEL=7 mg/kg/day).  

3.3.4	Reproductive Toxicity Study

In a 2-generation reproduction study in rats, liver pathology
(periacinar hepatocytic fatty changes) was observed in parental F1 males
at the LOAEL of 9.7 mg/kg/day (NOAEL=1.9mg/kg/day).  Reproductive
toxicity was seen in F1 females (F2 litters) at the LOAEL of 53.6
mg/kg/day (NOAEL=10.6 mg/kg/day) as a decreased number of implantation
sites and decreased litter sizes up to day 4 post partum.  At the LOAEL
of 42.12 mg/kg/day (NOAEL=8.4 mg/kg/day), developmental effects observed
were limited to decreased body weight gain during lactation for both F1
and F2 pups.        

  TC \l3 "3.3.4	Reproductive Toxicity Study 

3.3.5	Additional Information from Literature Sources

	A PubMed literature search indicated a publication that suggests that
fluazinam 

	may have immunotoxic potential.  The reference is:

A. Draper et al (2003) Occupational asthma from fungicides fluazinam and
chlorothalonil. Occupational and Environmental Medicine 60:76-77.

Fluazinam is a skin sensitizer; thus, it is a potential inhalation
allergen with the potential to cause asthma.  Although the cited study
demonstrates that fluazinam causes asthmatic symptoms, the asthma effect
may be related to the sensitization potential for this chemical. 
Consequently, asthmatic symptoms may only be detected in a small
population of people who may develop hypersensitivity as a result of
chronic exposure to small amounts of the chemical in an industrial
setting.  Therefore, the potential development of asthma in the general
population is not anticipated and immunotoxicity tests are not required
at this time.  Additionally, current  immunotoxicity test guidelines
focus on immunosuppression and are not designed to test for asthma,
autoimmunity, or allergy (except dermal sensitization).  

  TC \l3 "3.3.5	Additional Information from Literature Sources 3.3.6
Pre-and/or Postnatal Toxicity  TC \l3 "3.3.6	Pre-and/or Postnatal
Toxicity 

Determination of Susceptibility

There was evidence of increased qualitative susceptibility of fetuses to
fluazinam observed in the developmental toxicity study in rats.  In this
study, increased incidences of facial/palate clefts and other rare
deformities in the fetuses were observed in the presence of minimal
maternal toxicity.  A developmental neurotoxicity study in rats was
submitted to address the increased susceptibility, as well as the
presence of neurotoxic lesions observed after fluazinam exposure.  In
the developmental neurotoxicity study, decreases in body weight and body
weight gain and a delay in completion of balano-preputial separation
were observed in pups.  These effects were seen in the absence of
maternal effects, suggesting increased quantitative susceptibility of
the offspring. 

3.3.6.2	Degree of Concern Analysis and Residual Uncertainties  TC \l4
"3.3.6.2	Degree of Concern Analysis and Residual Uncertainties  for Pre-
and/or Postnatal Susceptibility

The purposes of the Degree of Concern analysis are: (1) to determine the
level of concern for the effects observed when considered in the context
of all available toxicity data; and (2) to identify any residual
uncertainties after establishing toxicity endpoints and traditional
uncertainty factors to be used in the risk assessment.  If residual
uncertainties are identified, then HED determines whether these residual
uncertainties can be addressed by a FQPA safety factor and, if so, the
size of the factor needed.

Although there is qualitative evidence of increased susceptibility in
young in the developmental toxicity study in rats, there are no residual
uncertainties with regard to pre- and/or postnatal toxicity following in
utero exposure to rats or rabbits and pre and/or post-natal exposures to
rats. Considering the overall toxicity profile and the doses and
endpoints selected for risk assessment for fluazinam, the degree of
concern for the effects observed in the study is low.  There is a clear
NOAEL for the fetal effects seen, the effects occurred in the presence
of maternal toxicity, and they were only seen at the highest dose
tested.  Additionally, the NOAEL of 50 mg/kg/day identified in this
developmental toxicity study in rats is significantly higher than the
NOAEL used (7 mg/kg/day) to establish the acute Reference Dose (aRfD) of
0.07 mg/kg/day (females 13-49); thus, the aRfD is protective of any
potential developmental effects.  

Increased quantitative evidence of susceptibility was observed in a
developmental neurotoxicity study in rats (MRID 46534401).  In pups,
there were decreases in body weight and body weight gain during
lactation, and delayed preputial separation observed at 10 mg/kg/day
(NOAEL=2 mg/kg/day).  Although the NOAEL of 2 mg/kg/day is lower than
that used for the aRfD for females 13-49 (7 mg/kg/day), the effects
noted in the developmental neurotoxicity study are attributable to
multiple doses and are considered post-natal effects.  Therefore, the
study endpoint is not appropriate for acute dietary exposures.  The cRfD
of 0.011 mg/kg/day is based on a lower NOAEL of 1.1 mg/kg/day and is
considered protective of potential developmental effects. 

3.4	FQPA Safety Factor for Infants and Children

After evaluating the toxicological and exposure data, the fluazinam risk
assessment team recommends that the FQPA SF be reduced to 1x based on
the following: 

The toxicological database for fluazinam is complete in regard to
pre-and postnatal toxicity and neurotoxicity.  There are acceptable
developmental toxicity studies in rats and rabbits, and an acceptable
reproduction study in rats; there is an acceptable developmental
neurotoxicity study, as well as acute and subchronic neurotoxicity
studies.  In addition, there are a series of special studies
investigating the neurotoxic lesions observed after fluazinam exposure.

The toxicity data for fluazinam showed increased qualitative
susceptibility of fetuses to fluazinam in rats in a developmental
toxicity study and neurotoxic lesions in studies in rats, mice and dogs.
 However, the developmental neurotoxicity study in rats and the special
studies submitted adequately addressed the observed effects; thus, there
are no residual uncertainties with regard to pre- and/or postnatal
toxicity or neurotoxicity and no additional factors are needed (see
3.3.6.1 and 3.3.6.2). 

The dietary food exposure assessment is based on HED-recommended
tolerance- level residues and assumes 100% crop treated for all
commodities, which results in very high-end estimates of dietary
exposure.  Actual exposures and risks from fluazinam will likely be
lower.

The dietary drinking water assessment is based on values generated by
model and associated modeling parameters which are designed to provide
conservative, health protective, high-end estimates of water
concentrations.

No residential uses are registered or proposed at this time. 

  TC \l2 "3.4	Safety Factor for Infants and Children 

3.5	Hazard Identification and Toxicity Endpoint Selection  TC \l2 "3.5
Hazard Identification and Toxicity Endpoint Selection 

Acute Reference Dose (aRfD) - Females age 13-49

Study Selected:  Developmental toxicity study in rabbits	

MRID No:  46578987		

	Dose and Endpoint for Risk Assessment: NOAEL= 7 mg/kg/day  

	Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Comments about Study/Endpoint/Uncertainty Factors: A developmental
toxicity study in rabbits was used to select the dose and endpoint for
establishing the aRfD of 0.07 mg/kg/day.  The NOAEL of 7 mg/kg/day and
the LOAEL of 12 mg/kg/day were based on increased incidence of total
litter resorptions and a possibly increased incidence of fetal skeletal
abnormalities (including kinked tail tip, fused or incompletely ossified
sternebrae, and abnormalities of head bones).  The skeletal
abnormalities observed are considered effects that could occur after a
single dose of fluazinam; thus, the route and duration of exposure are
appropriate for this population . Uncertainty factors (100x) include:
10x interspecies extrapolation and 10x intraspecies variability.

  = 0.07 mg/kg/day

3.5.2	Acute Reference Dose (aRfD) - General Population

Study Selected:  Acute neurotoxicity study in rats	

MRID No:  44807210		

	Dose and Endpoint for Risk Assessment: NOAEL= 50 mg/kg/day  

	Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Study/Endpoint/Uncertainty Factors: An acute oral neurotoxicity study in
rats was used to select the dose and endpoint for establishing the aRfD
of 0.5 mg/kg/day.  The NOAEL of 50 mg/kg/day and the LOAEL of 1000
mg/kg/day were based on soft stools and decreased motor activity.  Due
to the large dose spread in this study between the NOAEL (50 mg/kg/day)
and the LOAEL (1000 mg/kg/day), the true NOAEL is probably much higher
than 50 mg/kg/day.  This study, however, provides the best data
available for determining an acute RfD for the general population
(including infants and children); the route and duration of exposure are
appropriate for this population.  Uncertainty factors (100x) include:
10x interspecies extrapolation and 10x intraspecies variability.

  = 0.5 mg/kg/day

3.5.3	Chronic Reference Dose (cRfD) 

Studies Selected: A 2-Year carcinogenicity study in mice and a 1-Year
chronic oral study in dogs (co-critical studies) were selected to
establish the cRfD.  The 2-Year  carcinogenicity study in mice, rather
than the 1-year chronic oral study in dogs, was used to establish the
RfD because the treatment-related effects at the LOAEL in the mouse
study were related to liver toxicity (the regularly observed target
organ for fluazinam in many studies); whereas, the effects at the LOAEL
in the dog study (increased incidence of nasal dryness in females and
increased incidence/severity of gastric lymphoid hyperplasia in males
and females) were unrelated to liver toxicity.  The NOAELs in the mouse
(1.12 mg/kg/day in males and 1.16 mg/kg/day in females) and the dog (1
mg/kg/day in males and females) studies and the LOAELs in the mouse
(10.72 mg/kg/day in males and 11.72 mg/kg/day in females) and dog (10
mg/kg/day in males and females) studies were similar. 

   

1st Study: 2-Year carcinogenicity study in mice 	

MRID No:  42208405, 44807220, 44807212 	

	Dose and Endpoint for Risk Assessment: NOAEL= 1.12 mg/kg/day  

	Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Study/Endpoint/Uncertainty Factors: A 2-Year carcinogenicity study in
mice was one of two co-critical studies used in selecting the dose and
endpoint for establishing the cRfD of 0.11 mg/kg/day.  The NOAEL of 1.12
mg/kg/day and the LOAEL of 10.72 mg/kg/day were based on increased
incidences of brown pigmented macrophages in the liver of both sexes,
increased incidences of eosinophilic vacuolated hepatocytes in males,
and increased liver weights in females.  The route and duration of
exposure are appropriate for this population.  Uncertainty factors
(100x) include: 10x interspecies extrapolation and 10x intraspecies
variability.

2nd Study: 1-Year chronic oral toxicity study in dogs	

MRID No:  42270603, 44807219	

	Dose and Endpoint for Risk Assessment: NOAEL= 1 mg/kg/day  

	Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Study/Endpoint/Uncertainty Factors: A 1-Year chronic oral toxicity study
in dogs was one of two co-critical studies used in selecting the dose
and endpoint for establishing the cRfD of 0.11 mg/kg/day. The NOAEL of 1
mg/kg/day and the LOAEL of 10 mg/kg/day were based on marginal increases
in the incidence of nasal dryness in females and the incidence/severity
of gastric lymphoid hyperplasia in both sexes.  The route and duration
of exposure are appropriate for this population.  Uncertainty factors
(100x) include: 10x interspecies extrapolation and 10x intraspecies
variability

  = 0.011 mg/kg/day

Comments:  A 2-year chronic feeding/carcinogenicity study in rats (MRID
42248620, 44807223) was also considered for establishing the cRfD based
on a lower NOAEL of 0.38 mg/kg/day.  The next highest dose level tested
in this study was 3.8 mg/kg/day in males and 4.9 mg/kg/day in females (a
10 fold higher dose).  A second 2-year chronic feeding study in rats
(MRID 44839901, 44807213) was subsequently performed with 2 intermediate
dose levels.  The doses used in the study were 0, 1.0, 1.9, and 3.9
mg/kg/day for males; 0, 1.2, 2.4, and 4.9 mg/kg/day for females.  The
NOAEL observed in the second study was 1.9 mg/kg/day for males and 4.9
mg/kg/day for females, which is higher than the NOAEL (1.1 mg/kg/day)
selected for establishing the chronic RfD. Therefore, this study was not
chosen for risk assessment.   

3.5.4	Dermal Absorption  TC \l3 "3.5.5	Dermal Absorption 

A dermal absorption study is not available for fluazinam.  A dermal
absorption factor of 25% was estimated by comparing the LOAEL from a
21-day dermal toxicity study in rats (42270602) to the LOAEL from a
4-week range-finding feeding study in rats (44807213) based on a common
endpoint (liver toxicity).  In the dermal toxicity study, liver effects
observed were increased aspartate aminotransferase (AST) and increased
cholesterol levels in males at the LOAEL of 100 mg/kg/day (NOAEL=10
mg/kg/day).  In the range-finding study, effects included increased
serum phospholipids in females, increased total cholesterol in males and
females, increased relative liver weights in females, liver
histopathology (periacinar hypertrophy) in males, as well as decreased
body weight gain and decreased food consumption in females at the LOAEL
of 26.4 mg/kg/day in males (25.9 mg/kg/day in females) and the NOAEL of
5.1 mg/kg/day in males (5.3 mg/kg/day in females).    

 

Estimated Dermal= Oral LOAEL x 100 = 25 mg/kg/day x 100 = 25% 

Absorption Factor	Dermal LOAEL	100 mg/kg/day

3.5.5	Dermal Exposure (Short-, Intermediate-Term) 

Study Selected:  21-Day dermal toxicity study in rats

MRID No:  42270602

		

		Dose and Endpoint for Risk Assessment: NOAEL= 100 mg/kg/day  

Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Study/Endpoint/Uncertainty Factors: A 21-day dermal toxicity study in
rats was used to select the dose and endpoint for short- and
intermediate-term dermal exposure. The NOAEL of 10 mg/kg/day and the
LOAEL of 100 mg/kg/day were based on increased AST and increased
cholesterol levels in males.  The route and duration of exposure are
appropriate for this population.  Uncertainty factors (100x) include:
10x interspecies extrapolation and 10x intraspecies variability.

 ≥ 12 mg/kg/day; however, since the endpoint chosen for dermal risk
assessments is based on a lower NOAEL (10 mg/kg/day) it is considered
protective of potential developmental effects.

3.5.6	Inhalation Exposure (Short-, Intermediate-Term) 

Study Selected:  7-Day range-finding inhalation study in rats (test
material: Frowncide(

 WP, containing 51.9% fluazinam) 

MRID No:  42248621

Dose and Endpoint for Risk Assessment: NOAEL= 1.38 mg/kg/day

Uncertainty Factor: 100x (10x interspecies extrapolation, 10x
intraspecies variability)

Study/Endpoint/Uncertainty Factors: A 7-day range-finding inhalation
study in rats was used to select the dose and endpoint for short- and
intermediate-term intermediate exposure. The NOAEL of 0.011 mg/L (2.76
mg/kg/day in males and 2.97 mg/kg/day in females) and the LOAEL of 0.032
mg/L (7.93 mg/kg/day in males and 8.50 mg/kg/day in females) were based
on slightly increased testes weights (males) and slightly increased
liver weights (females).  The test material used in the inhalation study
was not technical grade fluazinam, but a formulation (Frowncide( WP)
containing approximately 50% fluazinam.  Consequently, the NOAEL from
the study (0.011 mg/L or 2.76 mg/kg/day in males and 2.97 mg/kg/day in
females) was reduced by half to account for this (i.e. adjusted NOAEL =
1.38 mg/kg/day for males and 1.48 mg/kg/day for females); thus, for
inhalation risk assessment a NOAEL of 1.38 mg/kg/day was used.  A
histopathology examination was not performed in the study and it is
possible that the true NOAEL may be lower than that demonstrated in the
study.  An additional safety factor of 10x was applied to the
conventional uncertainty factor of 100x to account for the lack of
histopathology in the inhalation studies.  This factor also addresses
the use of a short-term (7 days) study to evaluate intermediate-term
inhalation exposure. Uncertainty factors (1000x) include: 10x
interspecies extrapolation, 10x intraspecies variability, and 10x for
lack of histopathological examination and use of a short-term study for
intermediate-term exposure.  The 7-day range-finding inhalation study is
the most appropriate route to use for evaluating inhalation risk.  The
target organ in the study is the liver, as seen in several other studies
in the toxicology database.  

Comments:  Developmental effects were noted in developmental toxicity
studies at ≥ 12 mg/kg/day; however, since the endpoint chosen for
inhalation risk assessments is based on a lower NOAEL (1.38 mg/kg/day)
it is considered protective of potential developmental effects.

Recommendations:  Since the last risk assessment, an inhalation waiver
request (subchronic inhalation study) has been submitted by the
registrant for fluazinam.  HED concludes that for the proposed new uses
of fluazinam, a subchronic inhalation study is not required.  However,
HED recommends that  a 28-day subchronic inhalation study be submitted
to further characterize and support the registration of fluazinam.  If
an acceptable subchronic inhalation study is submitted and determined to
be more appropriate for endpoint selection, the additional 10x safety
factor can be reduced.  HED reserves the right to require a subchronic
inhalation study in the future for any proposed new uses.

       

3.5.7	Level of Concern for Margin of Exposure  TC \l3 "3.5.8	Level of
Concern for Margin of Exposure 

Table 3.5.8.  Summary of Levels of Concern for Risk Assessment.

Route	Short-Term (1 - 30 Days)	Intermediate-Term (1 - 6 Months)
Long-Term (> 6 Months)

Occupational (Worker) Exposure

Dermal	100	100	N/A

Inhalation	1000	1000	N/A

Recommendation for Aggregate Exposure Risk Assessments

  TC \l3 "3.5.9	Recommendation for Aggregate Exposure Risk Assessments 

As per FQPA, 1996, when there are potential residential exposures to a
pesticide, aggregate risk assessment must consider exposures from three
major routes: oral, dermal, and inhalation exposures.  As there are no
registered or proposed residential uses an aggregate exposure (three
routes) risk assessment is not required.  Occupational dermal and
inhalation exposures were not added since they were based on different
endpoints.

 

Classification of Carcinogenic Potential

  SEQ CHAPTER \h \r 1 In accordance with EPA’s Draft Guidelines for
Carcinogen Risk Assessment (July, 1999), the Cancer Assessment Review
Committee (CARC) classified (HED Doc. No.: 014512, March 29, 2001)
fluazinam as “Suggestive evidence of carcinogenicity, but not
sufficient to assess human carcinogenic potential”, based on increases
in thyroid gland follicular cell tumors in male rats and increases in
hepatocellular tumors in male mice.  The Agency has determined that
quantification of human cancer risk is not required and the cRfD (0.011
mg/kg/day) is protective of potential cancer effects.

The cancer classification was based on the following weight-of-evidence
considerations:  

 at ≥ 100 ppm (3.8 mg/kg/day).  In one study in mice, there was clear
evidence that an increased incidence of hepatocellular tumors observed
in male mice was treatment related at 1000 ppm (107 mg/kg/day).  In
another study in mice, there was equivocal/some evidence that fluazinam
may have induced an increase in hepatocellular tumors in the male mice
at ≥ 3000 ppm (377mg/kg/day).  Increases in hepatocellular tumors
observed in the female mice in the latter study were not statistically
significant and some occurred at an excessively toxic dose level.  The
thyroid gland follicular cell tumors of concern were seen only in the
male rats and the hepatocellular tumors of concern were seen only in the
male mice. 

Fluazinam was negative in mutagenicity assays.

3.5.10	Summary of Toxicological Doses and Endpoints for Fluazinam for
Use in Human Risk Assessments  

Table 3.5a  Toxicological Doses and Endpoints for Fluazinam for Use in
Dietary and Non-Occupational Human Health Risk Assessments

Exposure/

Scenario	Point of Departure	Uncertainty/

FQPA Safety Factors	RfD, PAD, Level of Concern for Risk Assessment	Study
and Toxicological Effects

Acute Dietary (General population)	NOAEL= 50 mg/kg/day

	UFA= 10x

UFH=10x

FQPA SF=1x

Total UF=100x	Acute RfD =0.5 mg/kg/day

aPAD

= 0.5mg/kg/day	Acute Neurotoxicity-Rats.

LOAEL = 1000 mg/kg/day based on decreased motor activity and soft stools
on day of dosing. 

Acute Dietary

(Females 13-49 years of age)	NOAEL (developmental) = 7 mg/kg/day	UFA=
10x

UFH=10x

FQPA SF=1x

Total UF=100x	Acute RfD =0.07 mg/kg/day

aPAD

= 0.07mg/kg/day	Developmental Toxicity- Rabbits.

Developmental LOAEL = 12  mg/kg/day based on increased incidence of
total litter resorptions and possibly increased incidence of fetal
skeletal abnormalities.

Chronic Dietary (All Populations)	NOAEL= 1.1 mg/kg/day

	UFA= 10x

UFH=10x

FQPA SF=1x

Total UF=100x	Chronic RfD =0.011 mg/kg/day

cPAD

= 0.011mg/kg/day	Carcinogenicity-Mice.

LOAEL = 10.7 mg/kg/day based on liver histopathology and increased liver
weight.  

Cancer (oral, dermal, inhalation)	Classification: “Suggestive Evidence
of Carcinogenicity, but not sufficient to assess human carcinogenic
potential”

.

  TC \l3 "3.5.10	Classification of Carcinogenic Potential NOAEL = no
observed adverse effect level.  LOAEL = lowest observed adverse effect
level.  UF = uncertainty factor.  UFA = extrapolation from animal to
human (interspecies).  UFH = potential variation in sensitivity among
members of the human population (intraspecies). FQPA SF = FQPA Safety
Factor.  PAD = population adjusted dose (a = acute, c = chronic).  RfD =
reference dose. N/A = not applicable.  

Table 3.5b  Summary of Toxicological Doses and Endpoints for Fluazinam
for Use in Occupational Human Health Risk Assessments

Exposure/

Scenario	Point of Departure	Uncertainty Factors	Level of Concern for
Risk Assessment	Study and Toxicological Effects

Dermal Short-Term (1-30 days); Intermediate-Term (1-6 months)  	NOAEL
(systemic) 10 mg/kg/day	UFA=10x

UFH=10x

	Occupational LOC for MOE = 100 

	21-Day dermal, rats.

Systemic LOAEL = 100 mg/kg/day based on increased cholesterol, increased
aspartate aminotransferase.

Inhalation Short-Term (1-30 days)	Inhalation  study NOAEL= 1.38
mg/kg/day 

	UFA=10x

UFH=10x

UFDB=10x

IAF=100%	Occupational LOC for MOE = 1000

	7-Day inhalation, rats.

LOAEL = 3.97 mg/kg/day based on increased liver weights (females) and
increased testes weights.

Inhalation Intermediate-Term (1-6 months)  	Inhalation  study NOAEL=
1.38 mg/kg/day 

	UFA=10x

UFH=10x

UFDB/UFS

=10x

IAF=100%	Occupational LOC for MOE = 1000

	7-Day inhalation, rats.

LOAEL = 3.97 mg/kg/day based on increased liver weights (females) and
increased testes weights.

Cancer (oral, dermal, inhalation)	Classification: “Suggestive evidence
of carcinogenicity, but not sufficient to assess human carcinogenic
potential”

NOAEL = no observed adverse effect level.  LOAEL = lowest observed
adverse effect level.  UF = uncertainty factor.  UFA = extrapolation
from animal to human (interspecies).  UFH = potential variation in
sensitivity among members of the human population (intraspecies). UFS =
use of a short-term study for long-term risk assessment.  UFDB = to
account for the absence of key data (i.e., lack of a histopathological
examination).   MOE = margin of exposure.  LOC = level of concern.
IAF=inhalation absorption factor.

3.6	Endocrine disruption  TC \l2 "3.6	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 Screening 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 systems may be
added to the Endocrine Disruptor Screening Program (EDSP).

When additional appropriate screening and/or testing protocols being
considered under the Agency’s EDSP have been developed, fluazinam may
be subjected to further screening and/or testing to better characterize
effects related to endocrine disruption. 

   TC \l1 "3.0	Hazard Characterization/Assessment 

  TC \l2 "3.6	Endocrine disruption 4.0	Public Health and Pesticide
Epidemiology Data

No public health/epidemiology data were used in developing this risk
assessment.  

  TC \l2 "4.4	Other Pesticide Epidemiology Published Literature 5.0
Dietary Exposure/Risk Characterization

Acute and chronic aggregate dietary (food and drinking water) exposure
and risk assessments were conducted using the Dietary Exposure
Evaluation Model DEEM-FCID™, Version 2.03 which use food consumption
data from the U.S. Department of Agriculture’s Continuing Surveys of
Food Intakes by Individuals (CSFII) from 1994-1996 and 1998.  The
analyses were conducted as part of a human health aggregate risk
assessment for the requested uses of fluazinam on ginseng, Brassica
vegetables, legume vegetables, and bushberries.  An assessment of cancer
risk is not necessary for this chemical since it is classified as “not
likely to be carcinogenic to humans.”

Both the acute and chronic analyses are based on tolerance-level
residues, assume 100% crop treated, and incorporate modeled estimated
drinking water concentrations (EDWCs).  Therefore, the resulting
exposure and risk estimates should be considered high-end and very
conservative.  Actual exposures and risks from fluazinam will likely be
lower than the values presented in the analyses.

The acute risk estimates are below HED’s level of concern for all
population subgroups, including those of infants and children. 
Generally, HED is concerned when risk estimates exceed 100% of the
population-adjusted dose (PAD).  The acute risk estimate for the U.S.
population, as a whole, is 1% of the acute PAD (aPAD).  For females
13-49 years of age, the risk estimate is 8% of their aPAD.  Risk
estimates for all other population subgroups are less than 8% aPAD. 
Likewise, chronic risk estimates are below HED’s level of concern for
all population subgroups.  The risk estimate for the U.S. population is
9% of the chronic PAD (cPAD).  The highest risk estimate is for the All
Infants (<1year) population subgroup at 16% cPAD (Table 4).

The analyses indicate that dietary exposure considerations do not
preclude establishing the proposed tolerances for fluazinam.

  TC \l1 "5.0	Dietary Exposure/Risk Characterization 

5.1	Pesticide Metabolism and Environmental Degradation  TC \l2 "5.1 
Pesticide Metabolism and Environmental Degradation 

5.1.1	Metabolism in Primary Crops  TC \l3 "5.1.1	Metabolism in Primary
Crops 

The nature of the residue in plants has been adequately delineated,
based on acceptable potato, peanut, and grape metabolism studies
reviewed previously (D257115; William Cutchin; 5/21/2001), along with an
acceptable apple metabolism study submitted recently (MRID #46991301). 
The metabolism of fluazinam appears to be similar in potatoes, peanuts,
grapes, and apples.  Fluazinam undergoes reduction of one of the nitro
groups to an amine, forming AMPA.  AMPA may then be conjugated with
glutathione, with subsequent degradation of the glutathione moiety to
cysteine.  The AMPA-cysteine conjugate then undergoes transamination,
reduction, and conjugation with glucose to form AMGT.  In addition, both
rings of fluazinam appear to be labile to ring cleavage, and subsequent
degradation of the rings into small fragments that may then be
incorporated into a variety of natural plant components.  HED concluded
that the ROC in potatoes and peanuts (for both tolerance expression and
dietary risk assessment purposes) was the parent compound only (D272624;
William Cutchin; 4/23/2001).  In wine grapes, both parent and AMGT were
included in the ROC for tolerance expression and risk assessment. 
Additionally, HED determined that data generated for potential new uses
on other crops (with the exception of root and tuber, and bulb
vegetables) should include analyses for both parent and AMGT.  

5.1.2	Metabolism in Rotational Crops  TC \l3 "5.1.2	Metabolism in
Rotational Crops 

elated metabolites are likely to be detectable in rotational root crops,
and leafy vegetables planted ≥30 days (≥68 days for small grains,
and all other crops) following a 2.0 lb ai/A (1X) soil application of
fluazinam.  

Metabolism in Livestock

  TC \l3 "5.1.3	Metabolism in Livestock 

The nature of the residue in livestock is also understood, based on
adequate goat and hen metabolism studies (D257115; William Cutchin;
5/21/2001).  The metabolism of [14C]-fluazinam in ruminants and poultry
is similar, and involves reduction of one or both nitro groups on the
phenyl ring to form AMPA, MAPA, or DAPA.  Fluazinam also undergoes
dehalogenation and hydroxylation of the chlorine on the phenyl ring to
form HYPA.  These compounds may then undergo conjugation with
glutathione, and subsequent degradation of the glutathione component
yields a variety of polar compounds.  Although the ring structure of the
parent molecule remains intact, fluazinam per se was only a minor
component (≤2.7% TRR) of the [14C]-residues in poultry tissues and
eggs, and was not detected in ruminant tissues or milk.  The fluazinam
residues of regulatory interest in animals were determined by HED to be
parent plus the metabolites AMPA and DAPA, and their sulfamate
conjugates.  

5.1.4	Analytical Methodology

  TC \l3 "5.1.4	Analytical Methodology 

Fluazinam:  The tolerance-enforcement method, Fluazinam:  Method for the
Analysis in Peanut Nut Meat (MRID #43521016), was adequately
radiovalidated.  This GC/ECD method for determining residues of
fluazinam per se was originally reviewed in conjunction with the
time-limited tolerance petition for peanuts (D177127 and D177137; George
Herndon; 6/19/1992).  In brief, residues of fluazinam are extracted from
crop samples with MeOH/acetic acid (HOAc) (50:1, v/v), filtered,
acidified with 0.2N HCl, and partitioned into hexane.  Residues are then
partitioned into 0.5N NaOH, the aqueous phase is acidified, and residues
are partitioned back into hexane.  The resulting hexane fraction is
concentrated, and residues are purified using a Florisil column, then
analyzed by GC/ECD.  The petitioner achieved adequate recoveries of
fluazinam from peanut nutmeat samples fortified with fluazinam at
0.010-1.00 ppm.  This method has undergone a successful ILV trial
(D212612, D216941, and D217467; George Herndon; 9/5/1995) using peanut
nutmeats fortified with fluazinam at 0.010, 0.020, and 0.050 ppm. 
Recoveries at the 0.010 ppm level were low (56% and 68%) owing to an
interference peak; therefore, the validated LOQ would be 0.020 ppm. 
However, the independent laboratory noted that the method could possibly
be improved in the Florisil clean-up step.  The method was forwarded to
ACB for a PMV trial, and was subsequently determined to be suitable as a
tolerance-enforcement method (D266802; Paul Golden; 6/22/2001).  

	

The submitted GC/ECD methods (modifications of the tolerance-enforcement
method) are adequate for collecting data and tolerances enforcement on
residues of fluazinam per se in the various crop commodities associated
with this petition.  The LLMV and/or LOQ for residues of fluazinam per
se were 0.010 ppm in all plant matrices except snap beans and lima
beans, in which the LLMV and LOQ were 0.020 ppm.  

	

AMGT:  The submitted HPLC/UV method (a working method based on Method
Evaluation for the Analysis of AMGT in Grapes, MRID #45593101) is
adequate for collecting data on AMGT residues in blueberries. 
Blueberries were blended with acetonitrile (ACN)/water (4:1, v:v), and
filtered.  The filter paper with contents was extracted a second time. 
The combined solvent extract was then concentrated by evaporation.  The
sample was partitioned with 2% aqueous Na2SO4 and methylene chloride. 
The aqueous layer was acidified to a pH of <1 with 6N HCl, then
partitioned twice with EtOAc, and the organic phase was evaporated to
dryness.  The aqueous sample was applied to a C18 SPE column, and AMGT
was eluted with ACN/water (3:7; v:v).  After evaporation to dryness, the
sample was taken up in ACN/H2O/HOAc, and filtered through a 0.45 µm
PTFE disc prior to analysis by HPLC/UV at 256 nm.  The LLMV, LOD, and
LOQ were 0.020, 0.013, and 0.038 ppm, respectively, for residues of AMGT
in blueberries.  HED has previously determined that residues of AMGT are
to be regulated in wine grapes (D272624; William Cutchin; 4/23/2001). 
The Agency therefore requested that this method undergo an ILV trial,
and, potentially, a PMV trial by the ACB.  An ILV study has not yet been
submitted.

As there are currently no tolerances established in livestock
commodities, and none are needed as a result of the requested uses,
residue analytical methods for livestock commodities are not required.  

Environmental Degradation

Based on the properties of the chemical, applications of fluazinam are
likely to reach the target (the crop), but drift is also possible.  The
chemical has a low vapor pressure, and a moderate Henry’s Law
constant.  Due to the fact that it appears to show relatively short half
lives in aquatic media, and it binds to soils, EFED believes that the
chemical would not volatilize substantially.

EFED concludes that fluazinam appears to degrade at moderate to low
rates in aerobic soils, but it is more rapidly transformed into other
compounds of similar backbone structure in high pH solutions or in
aquatic media, both, aerobic or anaerobic.  Fluazinam may be photolyzed
relatively rapidly (2.5 days) to form a tricyclic compound (G-504).  The
total fluazinam residues,  fluazinam and its transformation products
(DCPA, CAPA, and DAPA) are persistent in most environments (aerobic
aquatic metabolism 51-71 days, relatively stable in anaerobic aquatic
environment) and are likely to reach aquatic media as a totality through
runoff.  Since fluazinam does not alter substantially its backbone
structure in the environment, but instead, goes through a slight
transformation of functional groups, EFED considered parent and
transformation products together when making assessments.

While the parent and two transformation products, HYPA and CAPA, have
relatively low mobility, indicating a relatively low potential for
ground water contamination, further information on the other
transformation products should be required in a new terrestrial field
dissipation study.

le fish; ≥67% of residues depurated in 21 days).

The fate and transport characterization also summarizes the various
degradation products formed by each process in the studies reviewed in
tabular form. (Table 5.1.5)

Table 5.1.5.  Summary of degradate formation from degradation of
fluazinam.

STUDY TYPE	DEGRADATE and MAXIMUM CONCENTRATION	SOURCE

	CAPA (% applied)	HYPA (% applied)	AMPA (% applied)

	  Hydrolysis	34% at 28 days pH 7; 84-85% at 20 days at pH 9	–	–	 
MRID: 42208412.

  Aqueous Photolysis	G-504 was 14.0-17.1% by 7-10 days	_	–	  MRID:
44807312, 43521009.

  Soil Photolysis	_	Detected at more than dark control	Detected at more
than dark control	  MRID: 44807313.

  Aerobic Soil Metabolism	_	Detected 	MAPA and DAPA also detected	 
MRID: 42208413.

  Aerobic Aquatic Metabolism	_	–	24.2% at 0.2 day; DAPA: at day 30;
SDS-67200 39.6%by day 14	MRID: 43521010.

 Anaerobic Aquatic Metabolism 

	12.6% at 72 hr	–	DAPA: 19.0% by 240 hr; DCPA: 11.3% at 24 hr	MRID:
44807314.

 Terrestrial Field Dissipation	MAPA, CAPA, and HYPA were monitored;
however, there were problems with the storage stability data	MRID:
various.

Comparative Metabolic Profile

Data depicting the metabolism of fluazinam in plants and animals, as
well as data on environmental degradates, have been submitted to the
Agency.  

In rats, fluazinam metabolism involves hydroxylation followed by
conjugation.  The major urinary metabolites identified were AMPA
mercapturate and DAPA at 0.05-0.39% of administered dose (AD). 
Radioactivity in the feces represented most of the AD (88.78-100.03%);
however, metabolites identified represented only 11.20-68.59% the AD. 
Fecal radioactivity (all dose groups) appeared to reside with
unextractable components in the post-extraction solids (PES), which upon
further analysis was attributed to hydrolysis products of AMPA and DAPA.
 Approximately 20-25% of the aqueous phase of the fecal extraction was
identified as a cysteine conjugate of DAPA and represented <1% of the
AD. With the exception of the low-dose group (greatest PES
radioactivity), parent compound represented most of the identified
radioactivity in the feces.  AMPA and DAPA were identified in the feces
from all dose groups but these metabolites never represented more than
5% of the AD (except for high-dose female rats where AMPA accounted for
10.22%).  DAPA glucuronide and AMPA mercapturate were the major biliary
metabolites (<4% of the AD). 

The metabolism of [14C]-fluazinam in ruminants and poultry is similar,
and involves reduction of one or both nitro groups on the phenyl ring to
form AMPA, MAPA, or DAPA.  Fluazinam also undergoes dehalogenation and
hydroxylation of the chlorine on the phenyl ring to form HYPA.  These
compounds may then undergo conjugation with glutathione, and subsequent
degradation of the glutathione component yields a variety of polar
compounds.  Although the ring structure of the parent molecule remains
intact, fluazinam per se was only a minor component (≤2.7% TRR) of the
[14C]-residues in poultry tissues and eggs, and was not detected in
ruminant tissues or milk 

  

In plants, fluazinam undergoes reduction of one of the nitro groups to
an amine, forming AMPA.  AMPA may then be conjugated with glutathione,
with subsequent degradation of the glutathione moiety to cysteine.  The
AMPA-cysteine conjugate then undergoes transamination, reduction, and
conjugation with glucose to form AMGT.  In addition, both rings of
fluazinam appear to be labile to ring cleavage, and subsequent
degradation of the rings into small fragments that may then be
incorporated into a variety of natural plant components.  

  

Toxicity Profile of Major Metabolites and Degradates of Concern

There is no toxicology information available on fluazinam metabolites
and degradates. 

Pesticide Metabolites and Degradates of Concern

Table 5.1.8	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

Plants

	Primary Crop: grapes	Parent fluazinam and AMGT	Parent fluazinam and
AMGT

	Primary Crop: peanuts, root/tuber vegetables 	Parent fluazinam	Parent
fluazinam

	Primary Crop: all others	Parent fluazinam and AMGT	Parent fluazinam

	Rotational Crop	N/A	Note: Tolerances not required based on the absence
of residues in rotational crops at the requested plant back interval.

Livestock

	Ruminant	Parent fluazinam, AMPA, and DAPA, and their sulfate conjugates
Parent fluazinam, AMPA, and DAPA, and their sulfate conjugates

	Poultry	Parent fluazinam, AMPA, and DAPA, and their sulfate conjugates
Parent fluazinam, AMPA, and DAPA, and their sulfate conjugates

Drinking Water

	Parent Fluazinam, CAPA, DAPA, DCPA 	Not Applicable

At a meeting held on 11/28/2000, HED concluded that the residue of
concern (ROC) in potatoes and peanuts (for both tolerance expression and
dietary risk assessment purposes) was the parent compound only (D272624;
William Cutchin; 4/23/2001).  In wine grapes, both parent and AMGT were
included in the ROC for tolerance expression and risk assessment. 
Additionally, HED determined that data generated for potential new uses
on other crops (with the exception of root and tuber, and bulb
vegetables) should include analyses for both parent and AMGT.  

The fluazinam residues of regulatory interest in animals were determined
by HED to be parent plus the metabolites AMPA and DAPA, and their
sulfamate conjugates. 

Estimated drinking water concentrations (EDWC’s) were calculated for
Total Fluazinam Residues along with EDWC’s for parent fluazinam since
the environmental fate studies indicated that the parent compound forms
transformation compounds (CAPA, HYPA, and AMPA) which are similar in
structure to the parent (under most conditions).  Given that HED is
unable to conclude all these degradates are significantly less toxic
than the parent, the drinking water assessment is based on total
residues of fluazinam and its major degradates.  

Drinking Water Residue Profile

The drinking water residue used in the dietary risk assessment was
provided by the Environmental Fate and Effects Division (EFED; J.
Meléndez, D334948, 7 Feb 2007) and incorporated directly into this
dietary assessment into the food categories “water, direct, all
sources” and “water, indirect, all sources.”  The estimated
drinking water concentration (EDWC) of 0.071 ppm is the estimated peak
concentration of fluazinam parent or total residues from the FIRST model
(for more information, see   HYPERLINK
"http://www.epa.gov/oppefed1/models/water/" 
http://www.epa.gov/oppefed1/models/water/ ) and was used for the acute
assessment.  The chronic assessment uses the EDWC of 0.0177 ppm based on
total residues of fluazinam in surface water from the FIRST model (Table
5.1.9).

Table 5.1.9.  Maximum Tier I Estimated Drinking Water Concentrations
(EDWCs) for drinking water assessment based on ground application of
fluazinam.

Drinking Water Source (Model Used) 	USE (Rate Modeled)	Maximum Estimated
Drinking Water Concentration  (EDWC; ppb) 

Groundwater

(SCI-GROW) Fluazinam and Total Residues of Fluazinam	Bushberries (3.90
lb a.i./A)	Acute and Chronic	0.187

Surface Water

(FIRST) Fluazinam	Bushberries (3.90 lb a.i./A)	Acute	71.0

	Bushberries (3.90 lb a.i./A)	Chronic	0.7

Surface Water

(FIRST) Total Residues of Fluazinam	Bushberries (3.90 lb a.i./A)	Acute
71.0

	Bushberries (3.90 lb a.i./A)	Chronic	17.7

Food Residue Profile

Residue chemistry issues relevant to the proposed new uses requested in
the current petitions were reviewed in the Summary of Analytical
Chemistry and Residue Data memorandum for fluazinam (D335640; W. Drew).

Adequate storage stability data were collected indicating that   SEQ
CHAPTER \h \r 1 fluazinam residues were stable under frozen storage in
blueberries, snap beans, and broccoli for the storage durations and
conditions of the samples from the respective crop field trials.  In
blueberries, AMGT residues were stable under frozen storage for the
storage durations and conditions of the samples from the blueberry field
trials.  However, storage stability studies indicated that there was
significant dissipation of fluazinam residues under frozen storage in
ginseng, lima beans, dried beans, cabbage, and mustard greens. 
Correction factors were therefore incorporated into the recommended
tolerances for fluazinam in ginseng, shelled succulent beans, and
shelled dried beans to account for dissipation during storage.  A
correction factor was not utilized when setting the recommended
tolerance in Brassica leafy vegetables, because fluazinam applications
made to cabbage and mustard greens were essentially identical to the
treatment of broccoli (which had acceptable storage stability), and all
residues in treated samples from the Brassica field trials were ≤LOQ
(≤0.010 ppm).  At the time of submission, the freezer storage
stability analyses were not completed for the AAFC cabbage field trial. 
A final report is expected shortly.  Pending submission of the final
report for AAFC Project AAFC03-066R, the storage stability data
generated for IR-4 Project 08796 are adequate to support the storage
conditions and durations of the cabbage samples from the AAFC field
trial.  

There are currently no tolerances for fluazinam established in livestock
commodities, and there are no significant livestock feed items
associated with the proposed uses.  

Regulatory requirements pertaining to fluazinam residues in rotational
crops have been fulfilled, and the rotational crop restrictions on the
proposed label are adequate.  

The crop field trial data are adequate, and support the proposed use
patterns.  Adequate numbers of trials were conducted in the appropriate
geographical regions, and samples were analyzed for the ROC using
adequate methods.  However, residue data for AMGT were provided only for
blueberries; AMGT data should also have been included with the field
trial studies for edible-podded beans, shelled succulent and dried
beans, and Brassica vegetables.  Residues of fluazinam in treated
blueberry samples ranged from 0.064 to 2.0 ppm, and residues of AMGT
ranged from 0.025 to 0.13 ppm (with combined residues of 0.166-2.094
ppm) at the target PHI of 30 days (23-32 days).  The maximum residue
observed in snap beans treated with a single application of fluazinam,
and harvested at PHIs of 14-28 days, was 0.029 ppm, detected in a single
sample.  All remaining samples had residues below the LOQ (<0.020 ppm). 
The maximum residue observed in snap beans treated with two applications
of fluazinam, and harvested at PHIs of 10-22 days, was 0.109 ppm.  No
quantifiable (<LOQ; <0.010 ppm) or detectable (<LOD; <0.003 ppm)
residues of fluazinam were reported in any broccoli sample harvested 50
to 113 days after a single root-drench application of fluazinam at the
time of transplant.  No quantifiable (<LOQ; <0.010 ppm) or detectable
(<LOD; <0.005 ppm) residues of fluazinam were reported in any cabbage
sample harvested 58 to 104 days after a single root-drench application
of fluazinam at the time of transplant.  No residues above the LLMV (the
maximum residue observed was 0.010 ppm) were reported in any mustard
greens sample harvested 22 to 78 days after a single root-drench
application of fluazinam at the time of transplant.  In the trials
performed at the 1X and 2X application rates, the residues of fluazinam
in ginseng ranged from 0.28 to 1.4 ppm, and 2.1 to 2.2 ppm,
respectively.  The storage stability study, however, raises the
possibility that actual residues in ginseng (at harvest) were up to 50%
greater than the quantitated results, based on in-storage dissipation of
fluazinam.  Fluazinam residues were less than the LLMV (<0.010 ppm) in
all dried bean samples from the field trials, except for one sample at
0.0114 ppm.  The storage stability study, however, raises the
possibility that actual residues in dried beans (at harvest) were up to
50% greater than the quantitated results, based on in-storage
dissipation of fluazinam.  Fluazinam residues were less than the LOQ
(<0.020 ppm) in all lima bean samples from the field trials.  The
storage stability study, however, raises the possibility that actual
residues in lima beans (at harvest) were up to 50% greater than the
quantitated results, based on in-storage dissipation of fluazinam.  

There are no processed commodities for which residue data are required
associated with the proposed uses on the crops requested in the subject
petitions under review. 

Table 5.1.10.  Summary of New-Use Food Crop Residue Levels Used in the
Acute and Chronic Dietary Exposure Analyses.

Crop/Crop Group	Recommended Tolerance Level, ppm	Residue Level for
Dietary Exposure Assessment, ppma

Ginseng	4.5	4.5b

Brassica Vegetables (Group 5)	0.01	0.0135

Edible Podded Legumes (except peas; Group 6A)	0.1	0.135

Succulent Shelled Pea and Bean (Group 6B)	0.04	0.054

Dried Shelled Pea and Bean (Group 6C)	0.02	0.027

Bushberries (Group 13B)	*Not needed	*Not needed

a Residue level = recommended tolerance × 1.35 (from grape metabolism
data)

b AMGT is not a significant residue in root and tuber crops and no
correction is necessary for risk assessment

c Residue level = recommended tolerance × 1.11 (from field trial data)

* Tolerance not needed, low bush blueberry is a member of the
bushberries  subgroup 13-B

5.1.11	International Residue Limits TC \l3 "5.1.11	International Residue
Limits 

There are no established or proposed Canadian or Codex MRLs for residues
of fluazinam in plant or animal commodities.  There are Mexican MRLs
established for residues of fluazinam in potato at 0.05 ppm, and in
beans at 0.1 ppm.  The recommended US tolerance of 0.10 ppm in
edible-podded beans will be in harmony with the existing 0.1 mg/kg MRL
for Mexico.  Tolerances on shelled beans are being established at lower
levels than the Mexican MRL based on residues reflecting the use
proposed in the U.S. 

Dietary Exposure and Risk

Fluazinam acute and chronic dietary exposure assessments were conducted
using the Dietary Exposure Evaluation Model software with the Food
Commodity Intake Database DEEM-FCID™,Version 2.03 which incorporates
consumption data from USDA’s Continuing Surveys of Food Intakes by
Individuals (CSFII), 1994-1996 and 1998. Both the acute and chronic
assessments are based on tolerance-level residues, with worst-case
assumptions regarding levels of the metabolite AMGT.  In addition, it
was assumed that all crops with registered or proposed uses of fluazinam
were treated (i.e., 100% crop treated).  These assumptions result in
highly conservative, health-protective estimates of exposure and risk.

5.2.1	Acute Dietary Exposure/Risk

  TC \l3 "5.2.2  Chronic Dietary Exposure/Risk 

The acute risk estimates are below HED’s level of concern for all
population subgroups, including those of infants and children. 
Generally, HED is concerned when risk estimates exceed 100% of the
population-adjusted dose (PAD).  The acute risk estimate for the U.S.
population, as a whole, is 1% of the acute PAD (aPAD).  For females
13-49 years of age, the risk estimate is 8% of the aPAD.  Risk estimates
for all other population subgroups are less than 8% of the aPAD.  

5.2.2	Chronic Dietary Exposure/Risk  TC \l3 "5.2.2  Chronic Dietary
Exposure/Risk 

The chronic risk estimates are also below HED’s level of concern for
all population subgroups.  The risk estimate for the U.S. population is
9% of the chronic PAD (cPAD).  The highest risk estimate is for the All
Infants (<1year) population subgroup at 16% cPAD.

5.2.3	Cancer Dietary Risk  TC \l3 "5.2.3  Cancer Dietary Risk 

Fluazinam is classified as demonstrating “suggestive evidence of
carcinogenicity, but not sufficient to assess human carcinogenic
potential.”  The chronic dietary exposure analysis is considered to be
protective of cancer effects.  As a result, a separate cancer risk
assessment was not performed.

Table 5.2.  Summary of Acute and Chronic Exposure and Risk Estimates for
Fluazinam.

Population Subgroup	Acute Assessment (95th Percentile)	Chronic
Assessment

	PAD, mg/day	Exposure Estimate, mg/day	% Pad	PAD, mg/day	Exposure
Estimate, mg/day	% Pad

U.S. Population	0.5	0.006015	1	0.011	0.000953	9

All infants	0.5	0.015211	3	0.011	0.001799	16

Children 1-2 yrs	0.5	0.007019	1	0.011	0.001133	10

Children 3-5 yrs	0.5	0.006323	1	0.011	0.000996	9

Children 6-12 yrs	0.5	0.004439	1	0.011	0.000650	6

Youth 13-19 yrs	0.5	0.003344	1	0.011	0.000438	4

Adults 20-49 yrs	0.5	0.005903	1	0.011	0.000996	9

Adults 50+ yrs	0.5	0.006933	1	0.011	0.001123	10

Females 13-49 yrs	0.07	0.005809	8	0.011	0.001016	9

5.3	Anticipated Residue and Percent Crop Treated (%CT) Information TC
\l2 "5.3 Anticipated Residue and Percent Crop Treated (%CT) Information 

The acute and chronic analyses assumed tolerance level residues, 100%
crop treated, and DEEM( (ver. 7.81) default processing factors for all
registered and proposed commodities.  For those processed commodities in
the DEEM-FCID( residue list which were not in DEEM( (ver 7.81) (e.g.,
flour, bran, etc.), a processing factor of 1 was assumed.

6.0	Residential (Non-Occupational) Exposure/Risk Characterization

Currently, there are no registered or proposed residential uses for
fluazinam; thus, there is no exposure via this pathway and an assessment
was not conducted.

Spray Drift

Based on the proposed label restrictions, DO NOT apply this product in a
way that will contact workers or other persons, either directly or
through drift. Aerial application of this product is prohibited.

Spray drift is a potential source of exposure for residents living in
close proximity to spraying operations.  This situation is particularly
the case with aerial application.  However, to a lesser extent, spray
drift resulting from the ground application of fluazinam could also be a
potential source of exposure.  The Agency has been working with the
Spray Drift Task Force (a membership of US pesticide registrants), EPA
Regional Offices, State Lead Agencies for pesticide regulation, and
other parties to develop the best spray drift management practices.  The
Agency is now requiring interim mitigation measures for aerial
applications that must be placed on product labels/labeling.  The Agency
has completed its evaluation of the new database submitted by the Spray
Drift Task Force, and is developing a policy on how to appropriately
apply the data and the AgDRIFT computer model to its risk assessments
for pesticides applied by air, orchard airblast, and ground hydraulic
methods.  After the policy is in place, the Agency may impose further
refinements in spray drift management practices to reduce off-target
drift, and risks associated with pesticide application.  

Aggregate Risk Assessments and Risk Characterization

Acute Aggregate Risk

 TC \l2 "7.1	Acute Aggregate Risk 

Acute aggregate risk consists of risks resulting from exposure to
residues in food and drinking water alone.  The acute dietary exposure
analysis included both food and drinking water.  As a result, the acute
aggregate risk assessment is equivalent to the acute dietary risk
assessment and risk estimates are below HED’s level of concern. 

Short-Term Aggregate Risk

As there are no residential uses for fluazinam, short-term aggregate
risk assessments were not conducted.

7.3	Intermediate-Term Aggregate Risk TC \l2 "7.3	Intermediate-Term
Aggregate Risk 

As there are no residential uses for fluazinam, intermediate-term
aggregate risk assessments were not conducted.

7.4	Long-Term Aggregate Risk TC \l2 "7.4	Long-Term Aggregate Risk 

The chronic aggregate risk assessment consists of risks resulting from
exposure to residues in food, drinking water, and residues resulting
from residential applications.  As there are no residential uses for
fluazinam, chronic aggregate risk consists of risks resulting from
exposure to residues in food and drinking water alone.  The chronic
dietary exposure analysis included both food and drinking water.  As a
result, the chronic aggregate risk assessment is equivalent to the
chronic dietary risk assessment and risk estimates are below HED’s
level of concern.   

7.5	Cancer Risk

Fluazinam was classified as demonstrating “suggestive evidence of
carcinogenicity, but not sufficient to assess carcinogenic potential.”
 The chronic aggregate assessment is considered to be protective of
cancer effects.  

8.0	Cumulative Risk Characterization/Assessment

Section 408(b)(2)(D)(v) of the FFDCA requires that, when considering
whether to establish, modify, or revoke a tolerance, the Agency consider
"available information concerning the cumulative effects” of a
particular pesticide's residues and "other substances that have a common
mechanism of toxicity.”

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 fluazinam and any other
substances, and fluazinam does not appear to produce a toxic metabolite
produced by other substances. For the purposes of this tolerance action,
EPA has not assumed that fluazinam has a common mechanism of toxicity
with other substances.  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

Agricultural Handler Risk

The following products have been assessed for occupational exposure:
OMEGA® 500F and Allegro® 500F.  The products are formulated as a
flowable suspension/liquid containing 40.0 % fluazinam.  It may be
applied by airblast or groundboom at application rates ranging from
0.45-1.36 lbs active ingredient (ai) per acre (A) for a single
application and rates from 0.91-3.9 lbs ai per acre per growing season. 
Based on the anticipated application practices for the OMEGA® 500F and
Allegro® 500F Fungicide, product labels and information provided by the
registrant, handler exposures are expected to be short- and
intermediate-term in duration.  The quantitative risk assessment
developed for handlers is based on the following exposure scenarios:

•	Mixing/Loading Liquid formulation for groundboom

•	Mixing/Loading Liquid formulation for airblast

•	Applying sprays using groundboom

•	Applying sprays using airblast

 

No chemical-specific data for assessing exposure during pesticide
handling activities were submitted to the Agency in support of this
Section 3 application.   Therefore, HED used data from the Pesticide
Handlers Exposure Database (PHED) Version 1.1 to assess handler
exposures for regulatory actions when chemical-specific data are not
available (HED Science Advisory Council for Exposure, SOP Number .007,
January 1999).

	

Handlers Exposure and Risk

HED’s level of concern (LOC) for occupational dermal exposures is 100
(i.e., MOE less than 100 is of concern).  The level of concern for
inhalation exposures is 1000 (i.e., MOE less than 1000 is of concern). 
All   SEQ CHAPTER \h \r 1 dermal risk estimates for short- and
intermediate-term handler exposure resulted in MOEs greater than 100
with the use of gloves.   All inhalation   SEQ CHAPTER \h \r 1 risk
estimates for short- and intermediate-term handler exposure resulted in
MOEs greater than 1000 with the use of a dust mist respirator (Table
9.1.2.).

As reflected in the calculations included in Table 9.1.2, PPE consisted
of the addition of chemical-resistant gloves and a dust/mist respirator
to the baseline attire in order for all scenarios to reach either dermal
MOEs of 100 or inhalation MOEs of 1000.  However, since fluazinam is
classified as a Toxicity Category I chemical for acute eye irritation,
HED recommends that the PPE requirements (i.e. chemical-resistant
gloves, chemical resistant footwear, coveralls, protective eyewear,
dust/mist respirator and chemical resistant apron when mixing and
loading) on the proposed label are followed by all handlers for
acute/local toxicity to reduce systemic exposure.

Table 9.1.2 Short- and Intermediate-term Handler Exposure and Risk for
Fluazinam

Exposure Scenario (Scenario #)	Mitigation

Level	Dermal Unit Exposure

(mg/lb ai)	Inhalation Unit Exposure  

 (µg/lb ai)	Crop	Single Application Rate	Amount Treated	Dermal Dose
(mg/kg/day)	Dermal MOE	Inhalation Dose (mg/kg/day)	Inhalation MOE

Mixing/Loading

Liquid for Groundboom Application (PHED)	Single Layer, No Gloves	2.9	1.2

(0.0012 mg/lb ai)

	Dry Bean and Succulent Bean Crop (Subgroup 6B except Peas);
Edible-podded Legume Vegetables (Subgroup 6A except Peas	0.45 lb ai/A	80
A	1.49	7	0.00062	2,200

Ginseng	0.78 lb ai/A

2.59	4	0.00107	1,300

Brassica (Cole) Leafy Vegetables (Group 5)	1.36 lb ai/A

4.51	2	0.00187	740

	Single Layer, Gloves	0.023

Dry Bean and Succulent Bean Crop (Subgroup 6B except Peas);
Edible-podded Legume Vegetables (Subgroup 6A except Peas	0.45 lb ai/A

0.012	830	0.00062	2,200

Ginseng	0.78 lb ai/A

0.021	480	0.00107	1,300

Brassica (Cole) Leafy Vegetables (Group 5)	1.36 lb ai/A

0.036	280	0.00186	740

	Single Layer, Gloves; Dust/Mist Respirator

0.00024 mg/lb ai	Brassica (Cole) Leafy Vegetables (Group 5)	1.36 lb ai/A

0.036	280	0.00037	3,700

Liquid for Airblast Application (PHED)	Single Layer, No Gloves	2.9	1.2

(0.0012 mg/lb ai)	Bushberry	0.65	40 A	1.07	10	0.00044	3,100

	Single Layer,  Gloves	0.023

	0.009	1,100	0.00044	3,100

Applicator

Sprays for Groundboom Application (PHED)	Single Layer, Gloves 	0.014
0.74

(0.00074 mg/lb ai)

	Dry Bean and Succulent Bean Crop (Subgroup 6B except Peas);
Edible-podded Legume Vegetables (Subgroup 6A except Peas)	0.45 lb ai/A
80 A

	0.0072	1,400	0.00038	3,600

Ginseng	0.78 lb ai/A

0.0124	810	0.00066	2,100

Brassica (Cole) Leafy Vegetables 

(Group 5)	1.36 lb ai/A

0.0218	460	0.00115	1,200

Sprays for Airblast Application (PHED)	Single Layer, No Gloves	0.36	4.5

(0.0045 mg/lb ai)	Bushberry	0.65 lb ai/A	40 A	0.134	70	0.00167	830

	Single Layer,  Gloves;

Dust/Mist Respirator	0.24	0.0009 mg/lb ai

0.089	110	0.00033	4,200

Dermal  and Inhalation Unit Exposures = PHED Version 1.1

Amount Treated = HED’s Exposure Science Advisory Committee SOP Number
9.1,

Short- and Intermediate-term Dermal (mg/kg/day) = [Rate (lb ai/A) x UE
(mg /lb ai ) x  DAF (100%) x Amount Treated (A/day)] / BW (70 kg)	

Short- and Intermediate-term Dermal MOE = [Dermal NOAEL (10 mg/kg/day)]
/ Dermal Dose (mg/kg/day) 

Short- and Intermediate-term Inhalation Dose (mg/kg/day)  = [ Rate (lb
ai/A) x UE (mg /lb ai ) x  Amount Treated (A/day)] / BW (70 kg)	

Short- and Intermediate-term Inhalation MOE = [Inhalation NOAEL (1.38
mg/kg/day)] / Inhalation Dose (mg/kg/day)9.2 	Postapplication Risk

9.2.1	Data and Assumptions for Postapplication Exposure Scenarios

Two chemical-specific postapplication studies were submitted in support
of this registration action: 

•	“Foliar Dissipation of Fluazinam from Potato Leaves Treated with
Omega® 500F – USA in 2004” (MRID# 469913-03); Report dated February
6, 2006. Author: Author: J.L. Wiedmann; Sponsor: Ishihara Sangyo Kaisha,
Ltd; Performing Laboratories: EN-CAS Analytical Laboratories and

•	“Foliar Dissipation of Fluazinam from Peanut Leaves Treated with
Omega® 500F – USA in 2004” (MRID# 469913-02); Report dated May 22,
2006. Author: J.L. Wiedmann; Sponsor: Ishihara Sangyo Kaisha, Ltd;
Performing Laboratories: EN-CAS Analytical Laboratories

These studies have been reviewed using the U.S. Environmental Protection
Agency’s (U.S. EPA) OPPT Series 875, Occupational and Residential
Exposure Test Guidelines, Group B: Dislodgeable Foliar Residue
Dissipation: Agricultural, Guideline 875.2100. These studies were
designed to determine the dissipation of foliar residues (FR) of
fluazinam and its metabolites in and on potato foliage and peanuts.  The
residue component of those studies has been extracted for
chemical-specific use in determining the dislodgeable foliar residue
(DFR) values for each sampling interval (Table 9.2.1a).

Table 9.2.1a:  Summary of Peanuts and Potatoes Dissipation Foliar
Residues Data

Location	Application Rate

(lb ai/A)	Application

Method	 R-Squared 	Slope

(Ln DFR vs. t)	T ½

(days)	Time after application

	Maximum value of Total Fluazinam Residue

(µg/cm2)	Coefficient of Variation (%)

Potatoes (MRID# 469913-03)

Fayette County, Ohio	0.45	Backpack Sprayer	0.958	-0.21342	3.25
(Application #4)	Immediately after sprays dried	3.01	6.98

0.33 day	2.91	5.08

1 day	2.71	19.1

5 days	0.58	16.9

8 days	0.351	18.6

13 days	0.125	17.4

22 days	0.034	32.2

Peanuts (MRID# 469913-02)

Sampson County, North Carolina	0.78	Tractor Mounted Sprayers	0.914
-0.07569	9.16 (Application #3)	Immediately after sprays dried	1.11	10.5

1 day	1.51	8.86

4 days	0.814	8.8

7 days	0.623	14.1

14 days	0.282	16.5

21 days	0.202	17.7

27 days	0.147	33

35 days	0.107	27.8

9.2.2	Postapplication Exposure and Risk

Chemical-specific postapplication data were submitted in support of this
registration action. For purposes of comparison, a Tier 1 (HED standard
assumptions and defaults of 20% DFR) and Tier 2 (dislodgeable foliar
residue data) analysis were performed to ensure that potential
postapplication exposures are not of concern.  A comparison of Tier 1
and Tier 2 analyses resulted in similar postapplication exposure risks
of concern (MOE< 100).  Tables 9.2.2a and b provide a summary of
postapplication exposure and risk resulting from Tier 1 and Tier 2
analysis.

Since the Tier 1 and Tier 2 analyses resulted in similar postapplication
exposure risks of concern, HED based its postapplication assessment on
the Tier 2 analysis.  The only crop scenarios which resulted in MOEs
greater than 100 on day 0 (immediately after application) were for low
exposure activities ((i.e., scouting, hand weeding, thinning and
irrigation) for beans and ginseng.  All other crops (i.e. bushberries,
brassica and leafy vegetables) did not reach a MOE greater than or equal
to 100 for low exposure activities until 3 to 13 days later.  All medium
(i.e., scouting, hand weeding and irrigation) and high (i.e., hand
harvesting/ pruning/pinching/training) postapplication exposure
activities for all crops resulted in MOEs below 100 on day of
application.  Crops did not reach MOEs greater than or equal to 100
until 4 to 20 days later depending on the specific crop.   

Since postapplication exposure resulting in MOEs below 100 may indicate
possible risk for re-entry of workers, HED provided a comparison of the
estimated number of  days required before a MOE of 100 is reached (i.e.
restricted entry interval – REI) based on Tier 2 analysis to establish
pre-harvest interval (PHI).  HED recommends that the Registration
Division ensure that the PHIs do not go below the calculated REIs for
harvesting.  Table 9.2.2c provides a summary of the REIs and PHIs for
each crop and activity.  

A comparison of Tier 1 and 2 analyses resulted in similar
postapplication exposure risks of concern (MOE< 100).  The tier 2
analyses resulted in slightly refined exposure risks by reducing the
number of days at which a MOE of 100 was achieved.  However, in a few
instances the Tier 2 approach resulted in an increase in the number of
days required before reaching that MOE.  The 2 postapplication estimates
of exposure may be overestimating residues on the proposed crops based
on the methodology used to determine dislodgeable residues.  However,
HED cannot refine these estimates without chemical specific data
collected in accordance with Agency guideline methods.  A possible
option for the registrant would be to repeat the DFR studies using
guideline methods (i.e., leaf punch and dislodgeable residues with
surfactant as opposed to whole leaf extraction). 

Fluazinam has a low volatility, with a vapor pressure of 1.7 x 10-7 mmHg
(2.3 x 10-5 Pa).  Short-term postapplication inhalation exposures are
expected to be minimal and less than the application exposures since
negligible fluazinam vapor is expected to volatilize from the field. 
Consequently, a quantitative postapplication inhalation exposure
assessment was not performed.

Restricted Entry Interval 

The technical material has an Acute Eye Irritation Toxicity Category I. 
Per the Worker Protection Standard (WPS), a 48-hr restricted entry
interval (REI) is required for chemicals classified under Toxicity
Category I.   The 48 hour REI appearing on the label is only appropriate
for postapplication activities for which the MOE reaches 100 on day 0. 
However, note that an interval of 3 to 20 days is necessary to reach a
MOE of 100 for medium and high postapplication exposure activities
(i.e., hand weeding/harvesting/pruning/pinching/training, and
irrigation).  HED recommends that the proposed label be revised to
ensure that the appropriate REI restrictions are clearly stated for all
crops and do not exceed the pre-harvest intervals.

Table 9.2.2a: Short- and Intermediate Term Postapplication Exposure and
Risk for Fluazinam Using HED Default of 20% Dislodgeable Residue

	

Crops	Application Rate

(lb ai/A)	Transfer Coefficients a	DATb	DFRc

(ug/cm2)	Daily Dosed (mg/kg/day)	MOEe

Dry Bean and Succulent Bean Crop 

Edible-podded Legume Vegetables 	0.45	Low (100)	0

	1.01

	0.012	870

Med (1,500)	5	0.595	0.102	100

Bushberry (Low and High bush)	0.65 	Low (400)

(Low bush)	0	1.45	0.067	150

High (1,500)

(Low bush)	9	0.564	0.097	100

Low (500)

(High bush)	0	1.456	0.083	120

High (1,100)

(High bush)	6	0.774	0.097	100

Ginseng	0.78 	Low (100)	0	1.75	0.02	500

Med (1,500)	10	0.609	0.104	96

Brassica (Cole) 	1.36 	Low (2,000)	18	0.457	0.105	96

High (5,000)	27	0.177	0.101	99

Leafy Vegetables

Low (500)	5	1.79	0.103	97

High (2500)	20	0.370	0.106	94

a.    Transfer Coefficient = estimated dermal transfer coefficients from
the Science Advisory Council For                                    
Exposure Policy Number 3.1: Agricultural Transfer Coefficients, August
2000

b.    DAT = Days After Treatment

c.    DFR = Dislodgeable Foliar Residue (For Tier I) = Application Rate
(lb ai/A) x (1- Daily Dissipation Rate) t                          	x
4.54E8 ug/lb x  2.47E-8 A/cm2  x 0.2

Daily Dose = [DFR (ug/cm2) x TC (cm2/hr) x 0.001 mg/ug x DAF (100%) x 8
hrs/day] ÷ Body Weight (70 kg),

MOE = NOAEL/Daily Dose   (NOAEL = 10 mg/kg/day) 

Table 9.2.2b: Short- and Intermediate Term Postapplication Exposure and
Risk for Fluazinam Using DFR Studies	

Crops	Study	Application Rate

(lb ai/A)	Transfer Coefficients	DATa	DFRb (ug/cm2)	Adjusted DFRc

(ug/cm2)	Daily Dosed (mg/kg/day)	MOEerounded

Dry Bean, Succulent Bean Crop , Edible-podded Legume Vegetables	Peanuts
data MRID# 46991302	0.45

	Low (100)	0	1.110	0.666	0.0076	1300

	Med (1,500)	4	0.814	0.4884	0.084	120

Bushberry (Low and High bush)	Potatoes data MIRD# 46991303

	0.65 	Low (400)

(Low bush)	3	1.645 *	2.3	0.105	95

	High (1,500)

(Low bush)	7	0.428 *	0.59	0.10	100

	Low (500)

(High bush)	4	1.1125 *	1.557	0.0889	110

	High (1,100)

(High bush)	5	0.58	0.812	0.105	100

Ginseng

0.78 	Low (100)	0	3.010	5.220	0.059	170

	Med (1,500)	8	0.351	0.607	0.104	100

Brassica (Cole) 

1.36 	Low (2,000)	13	0.125	0.380	0.0863	120

	High (5,000)	20	0.054 *	0.163	0.093	110

Leafy Vegetables

	Low (500)	5	0.58	1.75	0.10	100

	High (2500)	14	0.115*	0.348	0.099	100

a.  DAT = Days After Treatment;

b.  DFR = Dislodgeable Foliar Residue from the submitted studies;

b.* Back calculations for DFR values corresponding to days not listed on
Table 9.2.1a are provided in D340845 (Z. Figueroa).

c.  Adjusted DFR = DFRs were adjusted to compensate for difference in
application rates (AR) between study and actual label application rates 

= (AR label proposed rater/AR study rate ) x DFR study; 

d.  Daily Dose = [DFR (ug/cm2) x TC (cm2/hr) x 0.001 mg/ug x DAF (100%)
x 8 hrs/day] ÷ Body Weight (70 kg),

e.  MOE = NOAEL (10 mg/kg/day) /Daily Dose  (mg/kg/day)  

Table 9.2.2c: Comparison of Crop REIs to PHIs	

Crops	Transfer Coefficients	DAT 

REI	MOE	PHI

Dry Bean and Succulent Bean Crop 

Eddible podded legumes	Med (1,500)	4	120	30

14

Bushberry (Low bush)

	Low (400)	3	95	30

	High (1,500)	7	100

	Bushberry (High bush)	Low (500)	4	110

High (1,100)	5	100

	Ginseng	Med (1,500)	8	100	30

Brassica (Cole) 	Low (2,000)	13	120	50

	High (5,000)	20	110

	Leafy Vegetables	Low (500)	5	100	20

	High (2500)	14	100

	DAT = Days After Treatment required to reach MOE > 100 = REI

10.0	Data Needs and Label Recommendations  TC \l1 "10.0	Data Needs and
Label Requirements 

10.1	Toxicology

	

A 28-day subchronic inhalation study is recommended to support the
registration of fluazinam.  If an acceptable subchronic inhalation study
is submitted and determined to be more appropriate for endpoint
selection, the additional 10x safety factor can be reduced.   

	10.2	Residue Chemistry  TC \l2 "10.2	Residue Chemistry 

No major deficiencies were noted in the subject petition that would
preclude the establishment of permanent tolerances for fluazinam
residues in the requested crops.  Revised Sections F should be
submitted, so that the proposed tolerances reflect the recommended
tolerance levels, and correct commodity definitions, as specified in
Table C1 (Appendix C).  Issues pertaining to residue chemistry
deficiencies should be resolved (see below). 

 

As a condition of registration, results of an ILV trial for the AMGT
analytical method (with wine grapes) should be submitted by the
registrant.  If the registrant agrees with the modifications made by
Ricerca to the original method (in MRID #45593101), these modifications
should be incorporated into a revised method for the ILV.  Sample sets
should include, at the minimum, 2 control (untreated) samples of wine
grapes, 2 samples fortified at the tolerance level (3.0 ppm), and 2
samples fortified at the LOQ (0.010 ppm).  

As a condition of registration, MRM recovery data should be provided for
the metabolite AMGT, since it is included in the tolerance expression
for wine grapes.  

The product label for Omega 500F should be amended to include a
restriction, stating that turnip roots from turnip plants treated with
this EP must not be used for human nor livestock consumption.  

The Agency has previously determined, and the registrant is hereby
advised again, that residue data for AMGT should be provided in the crop
field trial studies for all future requested plant commodities, except
root and tuber, and bulb vegetables.  

  TC \l2 "10.1	Toxicology 10.3	Occupational and Residential Exposure

HED recommends that the Registration Division ensure that the PHIs do
not go below calculated REIs for harvesting.  Additionally, HED
recommends that the proposed label be revised to ensure that the
appropriate REI restrictions are clearly stated for all crops and
correspond to the postapplication activities and reentry intervals.

References:

Dietary and Residue Chemistry

Fluazinam.  Tolerance Petitions   SEQ CHAPTER \h \r 1 Requesting the
Establishment of Permanent Tolerances (Associated with Section 3
Registration) for Food Use of the Herbicide on Edible-Podded Beans
(Subgroup 6-A, Except Peas), Shelled Succulent Beans (Subgroup 6-B,
Except Peas), Shelled Dried Beans (Subgroup 6-C, Except Peas), Brassica
(Cole) Vegetables (Group 5), Bushberries (Subgroup 13-B), and Ginseng. 
Summary of Analytical Chemistry and Residue Data. W. Drew. D335640. W.
Drew. 2007.

Fluazinam: Acute and Chronic Aggregate Dietary (Food and Drinking Water)
Exposure and Risk Assessments for the Section 3 Registration Action. M
Doherty. D340854. 2007

Tier I Estimated Drinking Waters Concentrations of Fluazinam and Total
Residues for the Use in the Human Health Risk Assessment; IR4 Petition
for the Use of Fluazinam on Edible-Podded Legume Vegetables (except
peas), Bushberry (crop subgroup 13B), Brassica (Cole) Leafy Vegetables,
Ginseng, and Dry, and Succulent Bean Crop Subgroup 6B (except peas). J.
Meléndez. D334948, D334950. 2007

Human Health Risk Assessment for the Use of Fluazinam on Peanuts,
Potatoes and Wine Grapes. W. Cutchin. D275396. 2001 

PP#9F5079.  Fluazinam in/on Peanuts and Grapes.  Tolerance Method
Validation Report. D266802. Paul Golden. 6/22/2001.  

PP#9F5079.  Request for the Use of Fluazinam on Peanuts, Potatoes, and
Wine Grapes.  Evaluation of Analytical Chemistry and Residue Data.
D257115. W. Cutchin; 5/21/2001.  

Fluazinam.  Decision by Metabolism Assessment Review Committee (MARC).
D272624. William Cutchin. 4/23/2001.  

Temporary Tolerance Petition and Experimental Use Permit for Use of
Fluazinam on Peanuts; 050534-EUP-E. Submission Dated 1/23/95 in Response
to the Memo of G.J. Herndon Dated 6/19/92. D212612, D216941, and
D217467. George Herndon. 9/5/1995.  

Temporary Tolerance Petition and Experimental Use Permit for Use of
Fluazinam on Peanuts; 050534-EUP-E. D177127 and D177137. George Herndon.
6/19/1992.  

Occupational and Residential Exposure

  TC \l1 "References: 

	Fluazinam: Occupational Exposure/Risk Assessment for the Use on
Ginseng, Dry Beans, Edible-podded Legume Vegetables, Bushberry and
Brassica Leafy Vegetables. D340845. Z. Figueroa. 2007

Foliar Dissipation of Fluazinam from Potato Leaves Treated with Omega®
500F – USA in 2004” (MRID# 469913-03); Report dated February 6,
2006. Author: Author: J.L. Wiedmann; Sponsor: Ishihara Sangyo Kaisha,
Ltd; Performing Laboratories: EN-CAS Analytical Laboratories.

Foliar Dissipation of Fluazinam from Peanut Leaves Treated with Omega®
500F – USA in 2004” (MRID# 469913-02); Report dated May 22, 2006.
Author: J.L. Wiedmann; Sponsor: Ishihara Sangyo Kaisha, Ltd; Performing
Laboratories: EN-CAS Analytical Laboratories

Toxicology

Fluazinam-2nd Report of the Hazard Identification Committee. TXR
0051576. Data Package Submitted by E. Budd.  February, 19, 2003. 

Evaluation of the Carcinogenic Potential of Fluazinam. HED DOC. NO. 
014512. Data Package Submitted by E. Budd. March 29, 2001. 

	Fluazinam: PP # 9F05079.  EPA File Symbol 71512-R.  New Reduced Risk
Active Ingredient.  Toxicology Disciplinary Chapter for the Registration
Support Document and Data Evaluation Records (DERs) for All Recently
Submitted Toxicology Studies,  Toxicology Studies Not Previously
Reviewed, and Previously Reviewed Toxicology Studies for Which Amended
DERs or Updated Executive Summaries Have Been Prepared. D258235. E.
Budd. June 18, 2001.

	Memorandum from Indira Gairola, Technical Review Branch, RD (7505C) to
Cynthia Giles-Parker, Fungicide Branch, RD (7505C). DP Barcode D272455.
May 18, 2001.

  

Appendix A:  Toxicology Assessment  TC \l1 "Appendix A:  Toxicology
Assessment 

A.1	Toxicology Data Requirements TC \l2 "A.1  Toxicology Data
Requirements  

The requirements (40 CFR 158.340) for a food use for fluazinam are in
Table 1. Use of the new guideline numbers does not imply that the new
(1998) guideline protocols were used.

Test 

	

Technical

	

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

yes

yes

yes

yes

yes	

yes

yes

yes

yes

yes

yes

870.3100	Oral Subchronic (rodent)	

870.3150	Oral Subchronic (nonrodent)	

870.3200	21/28-Day Dermal	

870.3250	90-Day Dermal	

870.3465	90-Day Inhalation		

yes

yes

yes

no

no	

yes

yes

no

---

---

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	

870.5375	Mutagenicity—Structural Chromosomal Aberrations	

870.5395	Mutagenicity—Other Genotoxic Effects		

yes

yes

yes

yes	

yes

yes

yes

yes

870.6100a	Acute Delayed Neurotox. (hen)	

870.6100b	90-Day Neurotoxicity (hen)	

870.6200a	Acute Neurotox. Screening Battery (rat)	

870.6200b	Chronic Neurotox. Screening Battery (rat)	

870.6300	Develop. Neuro		

no

no

yes

yes

yes	

---

---

yes

yes 

yes

870.7485	General Metabolism	

870.7600	Dermal Penetration		

yes

no	

yes

---

A.2  Toxicity Profiles

 

Table 1. Acute Toxicity Data on Fluazinam Technical

Guideline No./ Study Type	Test Substance	MRID No.	Results	Toxicity
Category

870.1100 

Acute oral toxicity  

rats	Technical grade fluazinam (lot #109; 95.3%)	42248603	

M: LD50 = 4500 mg/kg

F: LD50 = 4100 mg/kg	III

	Technical grade fluazinam (lot #8412-20; 95.3%)	42248602	M: LD50 >5000
mg/kg

F: LD50 >5000 mg/kg	

IV

	Technical grade fluazinam (lot #1/87; 97.9%)	42248604	M: LD50 >5000
mg/kg

F: LD50 >5000 mg/kg	

IV

870.1200 

Acute dermal toxicity

rats	

Technical grade fluazinam (lot #8303-2; 98.5%)	

42248605	

M: LD50 >2000 mg/kg

F: LD50 >2000 mg/kg	

III

870.1300

Acute inhalation toxicity

rats	

Technical grade fluazinam (lot #109; 95.3%)	

42270601	

M: LC50 = 0.463 mg/L

F: LD50 = 0.476 mg/L	

II

870.2400

Acute eye irritation

rabbits	

Technical grade fluazinam (lot # SNPE B-1216, No. 1006; 97.9%)	

42248606	

Extremely irritating.  Corneal opacity did NOT reverse in 21 days.	

I

870.2500 Acute dermal irritation

rabbits	

Technical grade fluazinam (lot # SNPE B-1216, No. 1006; 97.9%)	

42248607	

Slightly irritating	

IV

870.2600 Dermal sensitization

guinea pigs	

Technical grade fluazinam (lot # 1030/91; 96.7%)	

42274401	

Positive	

NA

	

Ultra-purified fluazinam (lot #Y910401; 100%)	

42248608	

Negative	

NA

Table 2. Acute Toxicity Data on Fluazinam (formulation)

Guideline No./ Study Type	Test Substance	MRID No.	Results	Toxicity
Category

870.1100 

Acute oral toxicity  

rats	

Omega 500F 

(40% fluazinam)	

42974907	

M: LD50 >5000 mg/kg

F: LD50 >5000 mg/kg	

IV

870.1200 

Acute dermal toxicity

rabbits	

Omega 500F

(40% fluazinam)	

42974908	

M: LD50 >2000 mg/kg

F: LD50 >2000 mg/kg	

III

870.1300

Acute inhalation toxicity

rats	

Fluazinam 50% WP (1)

(51.3% fluazinam)	

42311001	

M: LC50 = 3.0 mg/L

F: LD50 = 3.4 mg/L	

IV

870.2400

Acute eye irritation

rabbits	

Omega 500F

(40% fluazinam)	

42974910	

Slightly irritating	

III

870.2500 Acute dermal irritation

rabbits	

Omega 500F

(40% fluazinam)	

42974911	

Moderately irritating	

II

870.2600 Dermal sensitization

guinea pigs	

Omega 500F

(40% fluazinam)	

42974912	

Positive	

NA

(1)   Study satisfies requirement for testing on Omega 500F.

Table 3. Subchronic, Chronic and Other Toxicity Table	

Guideline No.

 Study Type	

MRID No./ (year) Classification /Doses	

Results

870.3100

90-Day oral toxicity rats	

42248610 (1984); 44807214 (1998)

Acceptable/guideline

M : 0, 0.15, 0.77, 3.8, 38 mg/kg/day;

F: 0, 0.17, 0.86, 4.3, 44 mg/kg/day	

NOAEL: Males: 3.8 mg/kg/day; Females: 4.3 mg/kg/day

LOAEL Males = 38 mg/kg/day; Females = 44 mg/kg/day based on increased
liver weights and liver histopathology in males, and increased lung and
uterus weights in females.

870.3150

90-Day oral toxicity 

dogs	

42248611 (1991); 44807215 (1998)

Acceptable/guideline

M & F: 0, 1, 10, 100 mg/kg/day	

NOAEL = 10 mg/kg/day

LOAEL = 100 mg/kg/day based on retinal effects, increased relative liver
weight, liver histopathology and possible increased serum alkaline
phosphatase in females and possible marginal vacuolation of the cerebral
white matter (equivocal)

870.3200

21-Day dermal toxicity

rats	

42270602 (1985)

Acceptable/guideline

M & F: 0, 10, 100, 1000 mg/kg/day	

Systemic NOAEL = 10 mg/kg/day

 LOAEL = 100 mg/kg/day based on increased AST and cholesterol levels in
clinical chemistry determinations (males)

Dermal NOAEL = not identified

LOAEL = 10 mg/kg/day based on erythema, acanthosis, and dermatitis

870.3250

90-Day dermal toxicity	

NA	

NA

870.3465

90-Day inhalation toxicity	

NA	

NA

870.3700a

Prenatal developmental toxicity 

rats	

42248613 (1985)

Acceptable/guideline

F: 0,10, 50, 250 mg/kg/day	

Maternal NOAEL = 50 mg/kg/day

LOAEL = 250 mg/kg/day based on decreased body weight gain and food
consumption and increased water consumption and urogenital staining

Developmental NOAEL = 50 mg/kg/day

LOAEL = 250 mg/kg/day based on decreased fetal body weights and
placental weights, increased facial/cleft palates, diaphragmatic hernia,
and delayed ossification in several bone types, greenish amniotic fluid
and possible increased late resorptions and postimplantation loss

870.3700b

Prenatal developmental toxicity

rabbits	

42248616 (1988)

Acceptable/guideline

F: 0, 2, 4, 7, 12 mg/kg/day	

Maternal NOAEL = 4 mg/kg/day

LOAEL = 7 mg/kg/day based on decreased food consumption and increased
liver histopathology.

Developmental NOAEL = 7 mg/kg/day

LOAEL = 12 mg/kg/day based on an increase in total litter resorptions
and possible fetal skeletal abnormalities

870.3700b

Prenatal developmental toxicity 

rabbits	

42248615 (1985); 42248614 (1984); 42248617 (1984)

Unacceptable/guideline

F: 0, 0.3, 1, 3 mg/kg/day	

Maternal NOAEL = 3 mg/kg/day

LOAEL = not identified (>3 mg/kg/day)

Developmental NOAEL = 3 mg/kg/day

LOAEL = not identified (>3 mg/kg/day)

870.3800

Reproduction and fertility effects

rats	

42248619 (1987); 42208406 (1985); 42248618 (1986)

Acceptable/guideline

F0 males: 0, 1.5, 7.3, 36.6 mg/kg/day

F0 females: 0, 1.7, 8.4, 42.1 mg/kg/day

F1 males: 0, 1.9, 9.7, 47.3 mg/kg/day

F1 females: 0, 2.2, 10.6, 53.6 mg/kg/day	

Parental/Systemic NOAEL = 1.9 mg/kg/day

LOAEL = 9.7 mg/kg/day based on liver pathology in F1 males

Reproductive NOAEL = 10.6 mg/kg/day

LOAEL = 53.6 mg/kg/day based on decreased number of implantation sites
and decreased litter sizes to day 4 post-partum for F1 females (F2
litters).

Offspring NOAEL = 8.4 mg/kg/day

LOAEL = 42.1 mg/kg/day based on reduced F1 and F2 pup body weight gains
during lactation.

870.4100a

Chronic toxicity 

rats	

44839901 (1993)

Acceptable/guideline

M: 0, 1.0, 1.9, 3.9 mg/kg/day

F: 0, 1.2, 2.4, 4.9 mg/kg/day	

NOAEL = Males: 1.9 mg/kg/day; Females: 4.9 mg/kg/day

LOAEL = Males: 3.9 mg/kg/day; Females: not identified (>4.9 mg/kg/day) 
based on increased testicular atrophy in males and no effects in females

870.4100b

Chronic toxicity 

dogs	42270603 (1987); 44807219 (1998)

Acceptable/guideline

M & F: 0, 1, 10, 50 mg/kg/day	

NOAEL = 1 mg/kg/day

LOAEL = 10 mg/kg/day based on gastric lymphoid hyperplasia in both sexes
and nasal dryness in females

870.4300

Combined chronic toxicity/carcino-genicity

rats	

42248620 (1988); 44807223 (1999); 45150201 (2000)

Acceptable/guideline

M: 0, 0.04, 0.38, 3.8, 40 mg/kg/day

F: 0, 0.05, 0.47, 4.9, 53 mg/kg/day	

NOAEL = Males: 0.38 mg/kg/day; Females: 0.47 mg/kg/day

LOAEL = Males: 3.8 mg/kg/day; Females: 4.9 mg/kg/day based on liver
toxicity in both sexes, pancreatic exocrine atrophy in females and
testicular atrophy in males.

Some evidence of carcinogenicity (thyroid gland follicular cell tumors)
in male rats, but not in females.

870.4200b

Carcinogenicity 

mice	

42208405 (1988); 4807220 (1996)

Acceptable/guideline

M: 0, 0.12, 1.1, 10.7, 107 mg/kg/day

F: 0, 0.11, 1.2, 11.7, 117 mg/kg/day	

NOAEL =  Males:1.1 mg/kg/day; Females: 1.2 mg/kg/day

LOAEL = Males: 10.7 mg/kg/day; Females: 11.7 mg/kg/day based on
increased incidences of brown macrophages in the liver of both sexes,
eosinophilic vacuolated hepatocytes in males, and increased liver weight
in females

Clear evidence of carcinogenicity (hepatocellular tumors) in male mice,
but not in females

870.4200b

Carcinogenicity 

mice	

44807222 (1996); 44807221 (1998); 45201301 (2000)

Acceptable/guideline

M: 0, 126, 377, 964 mg/kg/day

F: 0, 162, 453, 1185 mg/kg/day	

NOAEL =  Males:<126 mg/kg/day, Females: <162 mg/kg/day

LOAEL = Males: 126 mg/kg/day; Females: 162 mg/kg/day based on increased
liver weights and liver and brain histopathology in both sexes

Equivocal/some evidence of carcinogenicity (hepatocellular tumors) in
male mice, but not in females

870.5100

Bacterial reverse mutation assay (Ames test)	

42270605 (1988)

Acceptable/Guideline

Up to 2 μg/plate for S. typhimurium strains and up to 250 μg/plate for
E. coli (-S9).  Up to 100 μg/plate for S. typhimurium strains and up to
500 μg/plate for E. coli (+ S9)	

Negative with and without S9 up to cytotoxic concentrations. 

870.5100

Bacterial reverse mutation assay (Ames test)	

42270604 (1989)

Acceptable/Guideline

Up to 1μg/plate for S. typhimurium strains and up to 250 μg/plate for
E. coli (-S9) and  up to 100 μg/plate for S. typhimurium strains and up
to 500 μg/plate for E. coli (+S9)	

Negative with and without S9 up to cytotoxic concentrations. 

870.5300

In vitro mammalian gene mutation assay 	

45261801 (2000)

Acceptable/guideline

Up to 9 μg/ml (+S9), up to 0.3  μg/ml (-S9) 	

Negative with S9 activation up to 9 μg/ml.  Negative without S9
activation up to 0.3 μg/ml.

Compound tested to cytotoxic concentrations.

870.5300

In vitro mammalian gene mutation assay 	

45156902 (1986)

Acceptable/guideline

Up to 5 μg/ml (+/-S9)	

Negative with and without S9 activation up to 5 μg/ml.   

Compound tested to cytotoxic concentrations.

870.5375

In vitro mammalian chromosome aberration (CHL cells)	

42270606 (1988)

Acceptable/guideline

Up to 9.5 μg/ml (+S9) up to 4 μg/ml (-S9)	

Negative with and without S9 up to cytotoxic concentrations.  Cells
harvested at 24 and 48 hours in nonactivated studies and at 24 hours in
activated studies.

870.5395

Mammalian erythrocyte micronucleus test	

44807224 (1999)

Acceptable/guideline

500, 1000, 2000 mg/kg (oral gavage)	

Negative at 24 hour sacrifice (500, 1000, 2000 mg/kg).

Negative at 24, 48, and 72 hour sacrifices (2000 mg/kg).

870.5550

UDS in primary rat hepatocytes	

45156901 (1984)

Unacceptable/guideline

0.05 to 6.25 μg/ml	

Negative; however there were several serious study deficiencies:
treatment time shorter than recommended, no data supporting the claim of
cytotoxicity, data variability for major endpoints.

870.5550

Differential killing/growth inhibition in B. subtilis	

42270607 (1988)

Unacceptable/guideline

0.003 to 0.3 μg/disk (-S9)

0.3 to 30 μg/disk (+S9)	

Negative, however only one replicate plate/dose was used. 

870.6200a

Acute neurotoxicity screening battery 

rats	

44807210 (1995)

Acceptable/guideline

M & F: 0, 50, 1000, 2000 mg/kg	

Systemic NOAEL = 50 mg/kg

LOAEL = 1000 mg/kg based on soft stools and decreased motor activity on
day of dosing.

Neurotoxicity NOAEL = 2000 mg/kg

LOAEL = not identified (>2000 mg/kg)

870.6200b

Subchronic neurotoxicity screening battery 

rats	

44807217 (1998); 44807218 (1998)

Acceptable/guideline

M :0, 21, 69, 74, 149, 233 mg/kg/day;

F: 0, 23, 81, 89, 175, 280 mg/kg/day	

Neurotoxicity NOAEL = Males: 233 mg/kg/day; Females: 280 mg/kg/day

LOAEL = not identified (Males: >233 mg/kg/day; Females: > 280 mg/kg/day)
 

870.6300

Developmental neurotoxicity

	

46534401, 47018301 and 47037001. 

Acceptable/Non-Guideline

0, 2, 10 or 50 mg/kg/day	

Maternal:

NOAEL not established.  No toxicity at highest dose tested. 

Developmental:

NOAEL = 2 mg/kg/day.

LOAEL = 10 mg/kg/day based on decreased pup weight and gain and delayed
balano-preputial separation. 

870.7485

Metabolism and pharmacokinetics

rats

	

44807233 (1995); 43521004 thru 43521008, 43553001 (1993-1995)

Acceptable/guideline

0.5, 50 mg/kg	

Only 33-40% of the administered dose was absorbed.  Most of the
administered dose was recovered in the feces (>89%).   Excretion via the
urine was minor (<4%).  Total biliary radioactivity, however,
represented 25-34% of the administered dose, indicating considerable
enterohepatic circulation. 

A.3 Executive Summaries TC \l2 "A.3  Executive Summaries 

A.3.1	Subchronic Toxicity

870.3100		90-Day Oral Toxicity – Rat

Executive Summary: In a subchronic oral toxicity study (MRID 42248610,
44807214), technical grade fluazinam (98.5% a.i.) was administered in
the diet to 10 CD (remote Sprague-Dawley strain) rats/sex/dose level at
dose levels of 0, 2, 10, 50, or 500 ppm for 13 weeks (0, 0.15, 0.77,
3.8, or 38 mg/kg/day for males; 0, 0.17, 0.86, 4.3, or 44 mg/kg/day for
females).  Slides of brain and cervical spinal cord from all control and
500 ppm rats were later re-examined to assess for vacuolation of the
white matter in the central nervous system (MRID 44807214). 

No treatment-related mortalities, clinical signs of toxicity, changes in
body weights or body weight gains, differences in food or water
consumption, or ophthalmological findings were observed.  No
treatment-related effects in hematology, clinical chemistry, or
urinalyses parameters were noted.  Gross necropsies were negative.  At
termination, statistically significant treatment-related increases were
observed in the liver of 500 ppm males (absolute weights increased 8 %
(not significant) and relative liver/body weight ratios increased 11% in
comparison to controls), in the lungs of 500 ppm females (absolute
weights increased 18 % and relative lung/body weight ratios increased
25% in comparison to controls), and in the uterus of 500 ppm females
(absolute weights increased 36 % and relative uterus/body weight ratios
increased 43% in comparison to controls).  Statistically significant
compound-related histopathological lesions were observed in the livers
of 500 ppm males (increased incidences of periacinar hepatocyte
hypertrophy and sinusoidal chronic inflammation).  There was no effect
of treatment on the incidence or severity of vacuolation of the white
matter of the brain or cervical spinal cord in the 500 ppm rats as
compared with the controls.     

The LOAEL is 500 ppm (38 mg/kg/day in males and 44 mg/kg/day in
females), based on increases in absolute and relative liver weights in
males, increases in absolute and relative lung and uterus weights in
females, and increases in histopathology in the liver of males
(increased incidences of periacinar hepatocyte hypertrophy and
sinusoidal chronic inflammation).  The NOAEL in this study is 50 ppm
(3.8 mg/kg/day in males and 4.3 mg/kg/day in females).   

This subchronic oral toxicity study in rats is classified
Acceptable/Guideline and satisfies the Subdivision F guideline
requirement for a subchronic oral toxicity study [OPPTS 870.3100
((82-1a) in rats. 

870.3100		90-Day Oral Toxicity – Mouse 

Refer to carcinogenicity study. 

870.3150		90-Day Oral Toxicity – Dog

Executive Summary: In a subchronic oral toxicity study (MRID 42248611,
44807215), fluazinam (98.5% a.i.) was administered to 4 beagle
dogs/sex/dose daily via gelatin capsule at doses of 0, 1, 10 or 100
mg/kg/day for 90 days.   Slides of brain tissue were later reexamined
(Addendum to the original report, MRID 44807215) to assess vacuolation
of white matter according to criteria for severity used in mouse
carcinogenicity studies.

At 100 mg/kg/day, retinal effects of slight hyper-reflection and
slight-to-moderate grey mottling of the tapetal fundus in all males and
females during at least two of the three ophthalmologic examinations (7,
10 and/or 13 weeks), increased serum alkaline phosphatase levels
((2-fold, largely due to 1 female), increased SGPT in 1 female ((2 to
3-fold), increased absolute/relative liver weights (males 31%/34% and
females 33%/36% above controls), hepatic coagulative necrosis (1 male,
focal; 2 females, multifocal, all slight, vs. 0, controls) and slight to
moderate bile duct hyperplasia with/without cholangiofibrosis (2 males
and 2 females) were observed.  A possible marginal increase in cerebral
white matter vacuolation was observed in the reevaluation reported in
the Addendum (1 male and 2 females, severity graded as trace vs. 0,
controls).   There were no treatment-related effects observed for
clinical signs of toxicity, body weight/weight gain, food consumption,
hematology or urinalysis parameters or gross pathology.  No
treatment-related findings were observed at 1 or 10 mg/kg/day.  The
LOAEL is 100 mg/kg/day, based on retinal effects, possible increased
serum alkaline phosphatase in females, increased relative liver weight
and liver histopathology and possible marginal increase in vacuolation
of the cerebral white matter.  The NOAEL is 10 mg/kg/day.

This study is classified Acceptable/Guideline and satisfies the
guideline requirement for a subchronic oral toxicity study [OPPTS
870.3150((82-1b) in the dog. 

870.3200		21/28-Day Dermal Toxicity – Rat

Executive Summary: In a 21-day repeated dose dermal toxicity study (MRID
42270602), groups of 10 male and 10 female CD (Sprague-Dawley) rats were
treated with Fluazinam technical (98.0% a.i.; lot no. 8303-2) in 0.5%
methylcellulose in distilled water at doses of 0, 10, 100 or 1000
mg/kg/day.  Animals were treated by dermal occlusion for 6 hours per
day, 7 days per week, for 3 weeks.  

No treatment-related mortalities occurred.  At 1000 mg/kg/day, decreased
body weight gain in males (19% compared to controls, p <0.05) was
observed.  Liver damage in both males and females was also evident at
1000 mg/kg/day as demonstrated by increased absolute liver weights
(17-26%), increased relative liver/body weight ratios (27-30%),
statistically significant increases in aspartate aminotransferase (AST)
and cholesterol levels, and highly increased incidences of periacinar
hepatocellular hypertrophy in males and females.  At 100 mg/kg/day,
statistically significant increases in AST and cholesterol levels were
observed in males.  The LOAEL for systemic toxicity is 100 mg/kg/day
based on increased AST and increased cholesterol levels in males.  The
NOAEL for systemic toxicity is 10 mg/kg/day.     

At 1000 mg/kg/day, slight to severe erythema and atonia were observed
after 11-13 days and encrustation and/or staining at 21 days in males
and females.  At 100 mg/kg/day, slight erythema was observed after 14
days in males and females and encrustation and/or staining at 21 days in
females.  At 10 mg/kg/day, slight erythema was noted after 13 days in
one male.  Histologically, at 1000 mg/kg/day and 100 mg/kg/day,
acanthosis, dermatitis, scabs and ulceration were noted in males and
females.  At 10 mg/kg/day, acanthosis and dermatitis were observed in
males and females.  At 10 mg/kg/day, the test material was considered to
be a very mild irritant.  The LOAEL for dermal toxicity is <10 mg/kg/day
based on erythema, acanthosis and dermatitis in males and/or females. 
No NOAEL for dermal toxicity was determined in this study (< 10
mg/kg/day).  

This 21-day dermal toxicity study in rats is classified
Acceptable/Guideline and satisfies the Subdivision F guideline
requirement for a 21/28-day dermal toxicity study [OPPTS 870.3200 (OPP
82-2)]. 

870.3465	28- 	90-Day Inhalation – Rat

No subchronic inhalation study is available.  A 28-day study has been
requested. 

A.3.2		Prenatal Developmental Toxicity

870.3700a 	Prenatal Developmental Toxicity Study – Rat

Executive Summary: In a developmental toxicity study (MRID 42248613), 20
presumed pregnant Sprague-Dawley CD rats per group were administered
Fluazinam (98.5% a.i, Lot No.: 8303-2) by gavage in corn oil at doses of
0, 10, 50 and 250 mg/kg/day on gestation days (GD) 6-15, inclusive. 
Controls were treated with corn oil (vehicle).  On GD 20, all dams were
sacrificed and examined.  All fetuses were weighed, sexed and examined
for external malformations and variations.  Approximately half of the
fetuses from each litter were examined for soft tissue effects and half
were stained with Alizarin red S and examined for skeletal effects.  

Maternal Toxicity.  At 250 mg/kg/day, statistically significant
reductions in body weight gain during treatment (30 gm vs. 51 gm for
controls on GD 6-15; p < 0.01; most pronounced during GD 6-8),
statistically significant reductions in food consumption during
treatment (13 mg/kg/day vs. 17 mg/kg/day for controls on GD 6-8; p
<0.01), increased water consumption (during GD 6-11) and an increased
incidence of urogenital staining (most pronounced during GD 6-8) were
considered to be treatment-related.  The maternal toxicity LOAEL is 250
mg/kg/day based on decreased body weight gain , decreased food
consumption, increased water consumption, and increased urogenital
staining during treatment.  The maternal toxicity NOAEL is 50 mg/kg/day.
   

Developmental Toxicity. At 250 mg/kg/day, statistically significant
decreased mean fetal body weights (2.81 gm vs. 3.19 gm for controls, p
<0.001, below historical control range), statistically significant
decreased placental weights (0.47 gm vs. 0.54 gm for controls, p <0.05,
within historical control range), increased fetal incidence of
facial/palate clefts (10 fetuses in 3 litters vs. none in controls), 
increased fetal incidence of diaphragmatic hernia (7 fetuses in 2
litters vs. none in controls), delayed ossification in a number of bone
types, greenish amniotic fluid (10.5% fetal incidence vs. 0.0% in
controls) and possible increased late resorption/postimplantation loss
(0.55 late resorptions/dam vs. 0.05 late resorptions /dam for controls,
within historical control range;  and 11.0% postimplantation loss vs.
4.2% postimplantation loss for controls, within historical control
range) were considered to be treatment-related.  The developmental
toxicity LOAEL is 250 mg/kg/day based on decreased fetal body weights;
decreased placental weights; increased fetal incidences of facial/palate
clefts, diaphragmatic hernia and delayed ossification in several bone
types; greenish amniotic fluid and possible increased late resorptions
and postimplantation loss.  The developmental toxicity NOAEL is 50
mg/kg/day.  

This developmental toxicity study in rats is classified
Acceptable/Guideline and satisfies the Subdivision F guideline
requirement for a developmental toxicity study in rats [OPPTS 870.3700
(OPP 83-3a)].  No major deficiencies were noted in this study.   

Non-guideline  	Developmental Range-Finding Toxicity - Rat	

Executive Summary: In a developmental range-finding toxicity study (MRID
42248612), 7 pregnant CD (Sprague-Dawley origin) rats per group were
administered B-1216 (98.5%; Lot No. 8303-2) by gavage in corn oil at
doses of 0, 1, 10, 100, and 1000 mg/kg/day on gestation days (GD) 6-15,
inclusive.  On GD 20, dams were sacrificed and necropsied, and the
number and location of viable and nonviable fetuses, early and late
resorptions, and the number of total implantations and corpora lutea
were recorded, as well as the weights of the ovaries, empty uteruses,
and adrenal and pituitary glands.  All fetuses were weighed, sexed and
examined externally, and approximately half of each litter was processed
for visceral examination, and the remaining half of each litter was
examined by fresh dissection then processed for skeletal examination.

Maternal toxicity was evident at 1000 mg/kg/day.  Two animals were found
dead on GD 13 and the remaining 5 were sacrificed in extremis between GD
12 and 14.  Prior to death the high-dose animals exhibited clinical
signs of stained and ungroomed coats, lethargy, hunched posture, ataxia,
flaccid muscles, and salivation. Post mortem findings included decreased
thymus size and gastrointestinal tract disturbances.  Marked weight loss
was observed at 1000 mg/kg/day after GD 7, and mean absolute body
weights were 74-86% of those of controls during GD 10-13.  Body weight
and survival were not affected in the 1, 10, and 100 mg/kg/day groups.

There were no differences between the control group and the 1, 10, or
100 mg/kg/day groups for number of corpora lutea, number of implantation
sites, live fetuses/dam, pre- and post-implantation losses, resorptions,
or fetal sex ratios.  At 100 mg/kg/day, mean fetal weight was marginally
decreased as compared with concurrent controls but fell within the range
of historical control data.  The incidence of incomplete ossification of
sternebrae was increased in the 100 mg/kg/day group as compared to
concurrent and historical controls (38.9% of fetuses and 7/7 litters vs.
11.8% and 3/7 litters for concurrent controls and a historical control
range of 1.1-28.3%); however, there was no evidence of delayed
ossification in any other bone types.  The incidence rate for litters
containing fetuses with additional (14th) rib(s) was 1/7, 2/7, 2/7, and
3/7 for the 0, 1, 10, and 100 mg/kg/day groups, respectively, with the
percentage of affected fetuses slightly increased in all treated groups
as compared with concurrent and historical controls.  Treatment with
B-1216 did not result in an increased incidence of fetal malformations.

Therefore, it was concluded that an appropriate high dose for the main
developmental toxicity study (MRID 42248613) would be greater than 100
mg/kg/day but less than 1000 mg/kg/day.  The dose levels chosen were 0,
10, 50, and 250 mg/kg/day.

This study is classified as Acceptable/Nonguideline and fulfills its
intent as a range finding study for a developmental toxicity study
[870.3700 ((83-3a)] in rats.  Despite numerous deficiencies in the
conduct of this study, an assessment of appropriate doses of the test
article for the main developmental toxicity study was possible.

870.3700b 	Prenatal Developmental Toxicity Study - Rabbit

Executive Summary: In a developmental toxicity study (MRID 42248616),
16-18 presumed pregnant New Zealand White rabbits per group were
administered Fluazinam (95.3% a.i, Lot No.: 8412-20) by gavage in 1% w/v
aqueous methyl cellulose at doses of 0, 2, 4, 7 and 12 mg/kg/day on
gestation days (GD) 6-19, inclusive.  Controls were treated with 1% w/v
aqueous methyl cellulose  (vehicle).  On GD 29, all surviving does were
sacrificed and necropsied and all fetuses were weighed, and examined for
external malformation/variations.  Each fetus was examined viscerally by
fresh dissection and the sex determined.  All carcasses were eviscerated
and processed for skeletal examination.

Maternal Toxicity.  At 12 mg/kg/day, statistically significant
reductions in body weight gain during treatment ( 0.0 kg vs. +0.25 kg
for controls on GD 10-20), decreased food consumption (268 g/animal/day
vs. 368 g/animal/day for controls on GD 6-19) and an increased incidence
of liver histopathology (cellular hypertrophy, single cell necrosis,
binucleate hepatocytes, increased brown pigment deposition, apoptosis)
were considered to be treatment-related.  At 7 mg/kg/day, slightly
depressed food consumption (139 g/animal/day vs. 186 g/animal/day for
controls on GD 13-19) and an increased incidence of liver histopathology
(cellular hypertrophy, single cell necrosis, binucleate hepatocytes,
increased brown pigment deposition, apoptosis) were also considered to
be treatment-related.  The maternal toxicity LOAEL is 7 mg/kg/day based
on decreased food consumption and increased liver histopathology.  The
maternal toxicity NOAEL is 4 mg/kg/day.

Developmental Toxicity. At 12 mg/kg/day, an increased incidence of total
litter resorptions (0, 0, 0, 1 and 5 for the 0, 2, 4, 7 and 12 mg/kg/day
groups respectively) was considered to be treatment-related.  Several
abortions were also observed (0, 0, 2, 2 and 1 for the 0, 2, 4, 7 and 12
mg/kg/day groups respectively).  The total number of litters lost was
such that the numbers of litters born were 15, 13, 10, 10 and 7 for the
0, 2, 4, 7 and 12 mg/kg/day groups respectively.  Also at 12 mg/kg/day,
there was an increased incidence of placental anomalies (0.7, 3.2, 0.0,
0.0 and 18.2 percent fetal incidence for the 0, 2, 4, 7 and 12 mg/kg/day
groups respectively) and a slight increase in some skeletal
abnormalities including kinked tail tip, fused or incompletely ossified
sternebrae, and abnormalities of head bones.  The developmental toxicity
LOAEL is 12 mg/kg/day based on an increased incidence of total litter
resorptions and a possibly increased incidence of fetal skeletal
abnormalities.  The developmental toxicity NOAEL is 7 mg/kg/day.  

This developmental toxicity study in rabbits is classified
Acceptable/Guideline and satisfies the Subdivision F guideline
requirement for a developmental toxicity study in rabbits [OPPTS
870.3700 (OPP 83-3b)].  No major deficiencies were noted in this study. 
 

870.3700b 	Prenatal Developmental Toxicity Study - Rabbit

Executive Summary: In another developmental toxicity study (MRID
42248615, 42248614, 42248617), 20-24 presumed pregnant New Zealand White
rabbits per group were administered Fluazinam (98.5% a.i, Lot No.:
8303-2) by gavage in 1% w/v aqueous methyl cellulose at doses of 0, 0.3,
1.0, and 3.0 mg/kg/day on gestation days (GD) 6-19, inclusive.  Controls
were treated with 1% w/v aqueous methyl cellulose (vehicle).  On GD 29,
all surviving does were sacrificed and all fetuses were weighed, and
examined for external malformation/variations.  Each fetus was examined
viscerally by fresh dissection and the sex determined.  All carcasses
were eviscerated and processed for skeletal examination.

Four control group animals and one each in the 1.0 and 3.0 mg/kg/day
groups died or were killed in extremis following observations of reduced
food intake, reduced fecal output, weight loss, nasal discharge,
tremors, inactivity, abdominal bloat, abnormal respiration, and/or
soiled, stained, wet, or matted fur.  Deaths were not considered to be
treatment-related and may be related to disease or intubation error. One
female each in the 0.3 and 3.0 mg/kg/day groups aborted and were
subsequently sacrificed on GD 21 and 28, respectively, following weight
loss.  An additional control group female that exhibited weight loss and
scouring was killed on GD 6 and categorized as (removed from study.  No
statistically or biologically significant differences in body weight or
food consumption were noted.  Therefore, the maternal toxicity LOAEL is
not identified, and the maternal toxicity NOAEL is greater than or equal
to 3.0 mg/kg/day.

There were no differences between the control group and the 0, 0.3, 1.0
or 3.0 mg/kg/day groups for number of corpora lutea, number of
implantation sites, live fetuses/dam, pre- and post-implantation losses,
or mean fetal weight.  The percentage of small fetuses (weighing less
than 32.0 g.) were 19.6% (8/16), 13.0% (4/15), and 20.8% (6/14 litters),
respectively, for the 0.3, 1.0, and 3.0 mg/kg/day groups as compared to
4.1% (5/18 litters) for concurrent controls.  However, these fetal
incidence rates did fall within the historical control range of 0.0 to
30.4% (mean 13.36%).  No other statistically or biologically significant
differences between treated and control groups were noted.

The overall incidence rates for litters containing grossly abnormal
fetuses in the 0, 0.3, 1.0 and 3.0 mg/kg/day groups were 2/18, 1/16,
0/15, and 1/14, respectively.  No single malformation apparently
occurred at a statistically significant incidence in the treated groups,
and the incidence of total major malformations was also not
significantly increased for any of the treated groups as compared to
controls.   However, this assessment may be flawed since data from two
separate studies were combined and disease and intubation errors were
clearly apparent in the data.   It is unclear how disease differed in
the two studies and how this may have impacted on study interpretation. 
Therefore, the developmental toxicity LOAEL was not determined, and the
developmental toxicity NOAEL is greater than or equal to 3.0 mg/kg/day.

This study is classified as Unacceptable/Guideline and does not satisfy
the requirements for a developmental toxicity study 870.3700 ((83-3) in
rabbits.  At the high dose, significant maternal effects were not
reported, and developmental toxicity did not occur.   In addition, due
to the small number of animals available for examination due to disease
and intubation errors, an additional study was started at a later date
and these data were later combined and presented in this report. The use
of sick animals and the methods of combining different populations of
animals into a larger study as noted in this study are not acceptable. 

A.3.3	Reproductive Toxicity 

870.3800 Reproduction and Fertility Effects - Rat

Executive Summary: Technical grade fluazinam (95.3 % a.i.) was
administered to groups of 24 male and 24 female Sprague-Dawley rats at
dietary concentrations of 0, 20, 100, or 500 ppm for two generations
(MRID 42248619, 42208406, 42248618).  One litter was produced in each
generation.  Mean premating doses were 1.5, 7.3, and 36.6 mg/kg/day,
respectively for F0 males and 1.7, 8.4, and 42.1 mg/kg/day, respectively
for F0 females.  Mean premating doses were 1.9, 9.7, and 47.3 mg/kg/day
respectively, for F1 males and 2.2, 10.6, and 53.6 mg/kg/day,
respectively, for F1 females.  F1 adults were chosen from the F1 pups
and weaned onto the same diet as their parents.  Animals were given test
or control diet for 11 weeks before mating within the same dose group. 

There were no deaths or clinical signs of toxicity that were
attributable to the presence of fluazinam in the diet.  Mean body
weight, body weight gain, food consumption and food efficiency among all
groups of F0 males and F0 females treated with 20 or 100 ppm and F0
males treated with 500 ppm were similar to the control group means.  The
F0 females treated with 500 ppm of the test diet had significantly
decreased (82% of control value, p< 0.001) overall body weight gain and
food consumption (96% of control value, p< 0.05) for the premating
period.  The F1 males and females treated with 20 or 100 ppm had mean
body weights, body weight gains, food consumption, and food efficiencies
that were similar to their respective control group means.  The F1
animals treated with 500 ppm had significantly decreased mean body
weight gain and food consumption values that were 88% and 92% (p< 0.001
and p< 0.01) and 85% and 93% (p< 0.001 and p< 0.01) of the control
values for males and females, respectively for the premating period. 
The decreased body weights continued into gestation for females treated
with 500 ppm of both generations; some recovery was made during
lactation.  The relative liver weights of F0 and F1 males and F0 females
treated with 500 ppm were significantly increased compared to the
control group.  Histopathological findings included an increased
incidence of periacinar hepatocytic fatty changes and a decreased
incidence of hepatic glycogen pallor among F0 males treated with
500 ppm compared to the control group.  Males in the F1 generation
treated with 100 or 500 ppm also had significantly increased incidences
of periacinar hepatocytic fatty changes compared to the control groups. 
The LOAEL for parental toxicity is 100 ppm (9.7 mg/kg/day), based on
liver pathology (increased incidences of periacinar hepatocytic fatty
changes) in F1 males.  The NOAEL is 20 ppm (1.9 mg/kg/day).  

The fertility index for males and females treated with 500 ppm of the
test substance was slightly decreased (n.s.) for F1 parents compared to
the control group.  The number of implantation sites observed in F1 dams
was decreased significantly (p< 0.05) at 500 ppm (12.2 vs. 15.3 in
controls) and marginally (n.s.) at 100 ppm (13.1 vs. 15.3 in controls). 
Mean litter size on day 1 was slightly decreased (n.s.) in the 500 ppm
groups compared to the control groups in both generations.  Mean litter
size on day 4 was slightly decreased (n.s.) in the 500 ppm group for F1 
litters, but was significantly decreased (p<0.05) in the 500 ppm group
for F2 litters (9.8 + 3.7 for 500 ppm vs. 12.4 + 3.0 for controls).  Pup
survival was similar between the treated and control groups for both
generations.  The LOAEL for reproductive toxicity is 500 ppm (53.6
mg/kg/day), based on a decreased number of implantation sites and
decreased litter sizes to day 4 post partum for F1 females (F2 litters).
 The NOAEL is 100 ppm (10.6 mg/kg/day). 

Mean overall body weight gain during lactation was significantly
decreased (10-13%), among pups in the 500 ppm groups in both
generations.  The most pronounced effect on pup weight gains occurred
between lactation days 7-21.  Absolute body weights, however, were not
significantly decreased compared to the control groups at any time point
during lactation.  A slightly decreased developmental time for pinna
unfolding, hair growth and eye opening, particularly in the F2 pups, was
observed.  The LOAEL for developmental toxicity is 500 ppm (42.1
mg/kg/day), based on decreased body weight gain during lactation for
both F1 and F2 pups.  The NOAEL is 100 ppm (8.4 mg/kg/day).   

This study is classified as Acceptable/Guideline and satisfies the
requirements for a 2-generation reproduction study [OPPTS 870.3800
((83-4)] in rats.  No major deficiencies were noted in this study.

A.3.4	Chronic Toxicity

870.4100a (870.4300) Chronic Toxicity – Rat

Executive Summary: In a chronic oral toxicity study (MRID 44839901,
44807213), technical grade fluazinam (95.3%  a.i., Batch # 8412-20) was
administered to 25 Crl:CD((SD)BR rats/sex/dose in the diet at dose
levels of 0, 25, 50, or 100 ppm for 104 weeks (0, 1.0, 1.9, or 3.9
mg/kg/day for males; 0, 1.2, 2.4, or 4.9 mg/kg/day for females).  A
four-week range-finding study in rats was also conducted using 0, 10,
50, 250, or 3000 ppm (MRID 44807213).

No clinical signs of toxicity were observed, and survival rates were
unaffected by treatment.  At study termination, survival rates for the
0, 25, 50, or 100 ppm groups were  32, 52, 28, and 36% for the males,
respectively, and 72, 56, 72, and 52% for the females, respectively.  No
treatment-related effects on mean absolute body weights, body weight
gain, food consumption, or water consumption were noted.  

No treatment-related differences were observed in hematology analysis or
urinalysis, and treatment-related clinical chemistry changes were
limited to a transient increase in total serum cholesterol in high-dose
females at Week 52 (154% of controls, p<0.01).  Relative liver weights
were increased in high-dose females at study termination (124% of
controls; p<0.01), but were not accompanied by any histopathological
correlates.  The increased relative liver weights and transient increase
in cholesterol were therefore not considered adverse.  Treatment with
fluazinam appeared to affect the testes.  High-dose decedent males
(those not surviving to study termination) had an increased incidence of
small and/or flaccid testes (11/16 or 69%, 8/16 or 50%, respectively;)
as compared with decedent controls (4/17 or 24% for both) during
macroscopic examination.   Microscopic examination revealed a
corresponding increased incidence of marked tubular atrophy in the
testes of these 100 ppm decedent males (8/16; 50%) as compared with
decedent controls (3/17; 18%); however, this increase was not
statistically significant and did not show a dose-response when
considering the average severity ranking for the decedent males (2.5,
3.4, 2.6, and 3.4 for the 0, 25, 50, and 100 ppm decedent males,
respectively).  When considering all 100 ppm males (decedent +
terminal), macroscopic examination did not reveal an increased incidence
of small and/or flaccid testes (16/25 vs. 16/25 controls), but the
overall severity of testicular tubular atrophy was increased in the
high-dose males (3.1) as compared with controls (2.6).  Because
testicular atrophy was also an effect noted in male rats following
dietary treatment with 100 or 1000 ppm in another chronic
toxicity/carcinogenicity study (MRID 42248620), it is considered an
effect of treatment in this study.

At the doses tested, there was not a treatment-related increase in tumor
incidences when compared to controls.

A LOAEL of 100 ppm (3.9 mg/kg/day) was identified for male rats based on
increased testicular atrophy, with a corresponding NOAEL of 50 ppm (1.9
mg/kg/day).  A LOAEL could not be identified for females. The NOAEL for
females was therefore 100 ppm (4.9 mg/kg/day).

This chronic oral toxicity study in the rat is classified as
Acceptable/Guideline and satisfies the Subdivision F guideline
requirement for a chronic oral toxicity study [OPPTS 870. 4100 ((83-1a)]
in rats.

Executive Summary: In a 4-week oral range-finding study (MRID 44807213),
technical grade fluazinam (96.3% a.i., Batch # 8203) was administered to
10 CD (Sprague-Dawley) rats/sex/dose in the diet at dose levels of 0,
10, 50, 250, or 3000 ppm for 4 weeks (0, 1.0, 5.1, 26.4, or 302
mg/kg/day for males; 0, 1.1, 5.3, 25.9, or 309 mg/kg/day for females).  

The following treatment-related effects were observed at 3000 ppm in
both males and females: decreased body weight gain, decreased food
consumption, increased serum phospholipid, increased total cholesterol,
increased absolute liver weights, and increased relative liver/body
weight ratios.  In addition, increased incidences of  histopathological
findings were observed in the liver of males (increased periacinar
hypertrophy) and in the liver of females (increased single cell
necrosis).  The following treatment-related effects were also observed
at 250 ppm: decreased body weight gain (females), decreased food
consumption (females), increased serum phospholipid (females), increased
total cholesterol (males and females), increased relative liver/body
weight ratios (females), and increased histopathology in the liver of
males (increased periacinar hypertrophy).  In an addendum to this study,
there was no effect of treatment on the incidence or degree of white
matter vacuolation in the brain of male or female rats of the high dose
group (3000 ppm), compared with controls.  

The LOAEL was 250 ppm (26.4 mg/kg/day for males; 25.9 mg/kg/day for
females), based on decreased body weight gain (females), decreased food
consumption (females), increased serum phospholipid (females), increased
total cholesterol (males and females), increased relative liver/body
weight ratios (females), and increased histopathology in the liver of
males (increased periacinar hypertrophy).  The NOAEL was 50 ppm (5.1
mg/kg/day for males; 5.3 mg/kg/day for females).

This range-finding study is classified as Acceptable/Nonguideline.  It
does not satisfy the Subdivision F guideline requirement for a
subchronic oral toxicity study [OPPTS 870.3100 (82-1)] in rats.  

     

Executive Summary: In a combined chronic toxicity/carcinogenicity study
(MRID 42248620, 44807223, 45150201), fluazinam technical (95.3% a.i.,
lot number 8412-20) was administered to groups of 60 male and 60 female
Sprague-Dawley rats at dietary concentrations of 0, 1, 10, 100, or 1000
ppm (0, 0.04, 0.38, 3.8, or 40 mg/kg/day for males and 0, 0.05, 0.47,
4.9, or 53 mg/kg/day for females) for up to 104 weeks.  Groups of 10
rats of each sex per dose group were sacrificed at 52 weeks for interim
evaluations.

No treatment-related effects were observed in rats receiving the 1 ppm
or 10 ppm diets.  No treatment-related effect on mortality was observed
in rats receiving any dose of the test material.  The only clinical
signs observed were straw-discoloration of the fur in all rats receiving
the 1000 ppm diet and an increased incidence of alopecia in females
receiving the 1000 ppm diet.

Males receiving the 1000 ppm diet weighed 16% (p<0.01) less than
controls from week 8 to study termination, gained 15% less weight
overall, and consumed (8% less food than controls at each weekly
interval.  Females receiving the 1000 ppm diet weighed 24% (p<0.01) less
than controls from week 2 to termination, gained 35% less weight
overall, and consumed 18% less food than controls at each weekly
interval.  The food utilization factor or the food efficiency suggested
that reduced body weight gain was due in part to toxicity of the test
material.  No treatment-related effects were observed on body weights,
body weight gain, food consumption, or food utilization/efficiency in
male or female rats receiving the 1-, 10- or 100-ppm diets.  No
treatment-related effects were observed on the eyes at any dose at any
time during the study.  Clinical pathology evaluations showed only mild
anemia and elevated cholesterol in both sexes receiving the 1000 ppm
dose.  Treatment-related microscopic findings showed that the test
material was toxic to the following organs: liver, pancreas, lungs, and
possibly thyroid gland in both sexes, kidneys and testes in males, and
lymph nodes in females, but the primary target appeared to be the liver.
 Liver toxicity was manifested by increased organ weight and increased
incidences of gross and microscopic lesions.  Absolute liver weights
were increased by at least 7-27% and relative weights by 17-63% in both
sexes at 1000 ppm.  Gross lesions in the liver consisted of pale,
swollen or accentuated markings on the livers at 1000 ppm in males and
enlarged, pitted, or mottled livers at 1000 ppm in females. 
Treatment-related microscopic liver lesions at 100 ppm consisted of
eosinophilic hepatocytes in 22% of females (8% in controls),
centrilobular hepatocyte rarefaction and vacuolation in 8% of each sex
(0% for controls), centrilobular sinusoidal dilatation in 10% of males
and 18% of females (0% for male and 2% for female control), and
pericholangitis in 18% of males and 14% of females (4% for male and 2%
for female controls).  Additional treatment-related liver lesions at
1000 ppm in main study group consisted of centrilobular hepatocyte
vacuolation in males, centrilobular hepatocyte necrosis in females, and
centrilobular fat and bile duct hyperplasia in both sexes. 
Centrilobular hepatocyte vacuolation and centrilobular fat was also seen
in 1000 ppm group male and female rats at interim sacrifice.

The incidences of exocrine atrophy of the pancreas in both sexes and
acinal epithelial vacuolation or fat accumulation in females were
increased at 1000; the incidence of exocrine atrophy was also increased
at 100 ppm in females compared with that of control rats.  The incidence
of exocrine degranulation was increased in 1000 ppm group female rats at
interim sacrifice but not in the main study.  An increased incidence of
thyroid follicular hyperplasia was observed in males at 1000 ppm (8% vs.
2% for controls) and in females at 1000 ppm (10% vs. 2% in controls). 
This  finding may possibly be related to treatment with the test
material.  In male rats, the incidence of cortical tubular basophilia in
the kidney was increased at 1000 ppm compared with that of the controls.
 Other treatment-related lesions included  pneumonitis, alveolar
adenomatosis, and alveolar epithelialization in 1000 ppm group males,
alveolar epithelialization and alveolar macrophage aggregates in 1000
ppm group females, testicular atrophy in 100 ppm and 1000 ppm group
males, and spermatocele granuloma also in 1000 ppm males.  The incidence
of sinus histiocytosis in the lymph nodes was increased in1000 ppm group
females.  Histopathologic assessment of the brain and spinal cord of
rats in the control and 1000 ppm dose groups showed no treatment-related
effect on vacuolation of white matter.

The lowest-observed-adverse-effect level (LOAEL) for fluazinam was 100
ppm (3.8 mg/kg/day for males and 4.9 mg/kg/day for females) based on
liver toxicity in both sexes, testicular atrophy in males and pancreatic
exocrine atrophy in females.  The corresponding
no-observed-adverse-effect level (NOAEL) was 10 ppm (0.38 mg/kg/day for
males and 0.47 mg/kg/day for females). 

In this study, there were statistically significant positive trends for
thyroid gland follicular cell adenocarcinomas and combined follicular
cell adenomas/adenocarcinomas for the male rats.   There was also a
statistically significant increase by pair-wise comparison of the male
high dose group (1000 ppm) with the controls for combined follicular
cell adenomas/adenocarcinomas (23% vs. 8% in controls).  In addition to
an Exact Trend Test and a Fisher's Exact Test, a Peto's Prevalence Test
was also conducted (which excluded animals that died or were sacrificed
before observation of the first tumor at week 68).  For follicular cell
adenocarcinomas in males, results of the Peto's Prevalence Test showed a
statistically significant positive trend and a borderline statistically
significant (p= 0.056) increase by pair-wise comparison of the 1000 ppm
male group with the controls (7% vs. 0% in controls), indicating the
increased incidence of thyroid tumors had a malignant component to it. 
For combined follicular cell adenomas/adenocarcinomas, Peto's Prevalence
Test also showed a statistically significant increase by pair-wise
comparison of the high dose male group with the controls (26% vs. 9% in
controls).  The incidences of thyroid gland  adenomas at 100 ppm and
1000 ppm (15% and 17%, respectively) and adenocarcinomas at 1000 ppm
(6%) were slightly outside  their respective ranges in the historical
control data (range: adenomas, 0%-13%; adenocarcinomas, 0%-5%).  Animals
in the lower dose groups were not microscopically examined for thyroid
lesions unless abnormalities were observed in that organ at gross
necropsy.  Therefore, percentage incidences of thyroid tumors in these
lower dose groups may have been somewhat misleading (too high).  The
highest dose level tested in this study was considered to be adequate
and not excessive because there were decreased  body weight gains (up to
15% and 35% in males and females, respectively), decreased food
consumption, decreased food efficiency, increased thyroid weights at 52
weeks, enlarged thyroids and a slightly increased incidence of thyroid
gland follicular cell hyperplasia  at 104 weeks in males. The survival
of the animals was not decreased by treatment with the test material. 
There was no treatment-related increase in the thyroid tumor incidence
in the female rats in this study.  In addition, slightly increased
incidences of pancreatic islet cell adenomas were also observed in the
treated male rats, but these incidences were not dose-related and did
not exceed the upper range of historical control data for the same tumor
type (20 comparable studies at the same testing laboratory).  Further,
this type of tumor in this strain of rats is a common spontaneously
occurring neoplasm.  Therefore, the increased incidence of pancreatic
islet cell adenomas observed in the treated male rats in this study was
considered not likely to be related to treatment with the test material.
 High incidences of pituitary gland adenomas in males and females and of
mammary gland fibroadenomas in females were also observed in all groups,
including controls.  These neoplasms are not considered to be
treatment-related.  A toxicologically significant increase in tumors was
not observed in any other tissues in the treated male or female rats in
this study.

This combined chronic toxicity/carcinogenicity study in the rat is
Acceptable/Guideline and satisfies guideline requirements for a chronic
toxicity/carcinogenicity study [OPPTS 870.4300 ((83-5)] in rats.

870.4100b 	Chronic Toxicity – Dog

Executive Summary: In a chronic oral toxicity study (MRIDs 42270603,
main study and 44807219, addendum), Fluazinam (Lot No. 8412-20, 95.3%
purity) was administered to groups of six male and six female beagle
dogs/dose for 52 weeks at doses of 0, 1, 10, or 50 mg/kg/day in gelatin
capsules. 

No animals died as a result of treatment. The most notable clinical
signs were increased incidence of salivation and nasal dryness, mainly
in the high-dose dogs but nasal dryness was also slightly increased in
females at 10 mg/kg/day. Body weight was mildly decreased at high dose
(-4%, males and -9%, females; not analyzed statistically), and total
body weight gain was significantly reduced (29%, p<0.05; -13% when
calculated as a percentage of initial body weight) only in females but
was also lower in males (-19%; -9% as a percentage of initial body
weight). Hematocrit, hemoglobin, and RBC counts of high-dose dogs were
consistently lower (8-17%; p<0.05, 0.01, or 0.001) than controls
throughout the treatment period, and WBC counts were elevated (32-64%,
p<0.05 or 0.001) at study end (these findings considered
treatment-related but not biologically significant). Alkaline
phosphatase was significantly increased (52-183%; p<0.05, 0.01, or
0.001) in high-dose dogs throughout the treatment period. 

Absolute liver weight (males, 37%; females, 16%; p<0.05) and the
liver/body weight ratio (males, 45%; females, 47%; p<0.01) were
increased in high-dose dogs. In the reexamination of brain and spinal
cord tissues, incidence  of vacuolation of white matter in the brain was
increased in both sexes at the high dose (6/6 animals/sex affected vs.
2-4/6, controls) , along with increased severity (1.5-2.17 vs. 1.0,
controls).  In addition, vacuolation of the white matter of the spinal
cord was seen in high-dose females (4/6 affected vs. 0, controls). An
increase in liquefied GI tract contents and incidence/severity of
stomach mucosal lymphoid hyperplasia was seen in mid- and high-dose dogs
of both sexes, although in females, neither incidence nor mean severity
of the hyperplasia at these dose levels showed a dose-related increase.

The LOAEL is 10 mg/kg/day for both male and female dogs, based on
marginal increases in the incidence of nasal dryness in females and the
incidence/severity of gastric lymphoid hyperplasia in both sexes.  The
NOAEL is 1 mg/kg/day. 

This chronic toxicity study is classified as Acceptable/guideline and
satisfies the guideline requirement for a chronic oral study [OPPTS
870.4100 ((83-1b)] in dogs. No major deficiencies were noted in this
study.

A.3.5		Carcinogenicity

870.4200b 	Carcinogenicity (feeding) – Mouse

Executive Summary: In a carcinogenicity study (MRID 42208405, 44807220,
44807212), Fluazinam (95.3% a.i., lot no. 8412-20) was administered to
groups of 52 male and 52 female CD(-1 mice in the diet at concentrations
of 0, 0, 1, 10, 100, or 1000 ppm.  There were 2 control groups.  The
test diets were given for 104 weeks.  These concentrations resulted in
mean daily compound intakes of 0.12, 1.1, 10.7, and 107 mg/kg/day for 1
ppm, 10 ppm, 100 ppm, and 1000 ppm, respectively, for males and 0.11,
1.2, 11.7, and 117 mg/kg/day, respectively, for females.  Additional
microscopic review of brain and spinal cord was presented in MRID
44807220.  A four-week-range finding study (MRID 44807212) using 0, 10,
50, 250, or 3000 ppm in the diet was also conducted.

Treatment with Fluazinam did not result in treatment-related changes in
survival, clinical signs, body weights, body weight gains, food
consumption or hematology parameters.  The group mean liver weights
adjusted for body weight were increased in males and females by 45% and
30%, respectively, at 1000 ppm compared to the controls, and by 15% in
females at 100 ppm after 104 weeks of treatment (p<0.01).  Microscopic
examination showed increased incidences of liver areas containing
basophilic hepatocytes (controls, 12%; 1000 ppm, 38%, p<0.01) and/or
eosinophilic vacuolated hepatocytes (controls, 1%; 100 ppm, 8%, p<0.05;
1000 ppm 19%, p<0.01) in treated males compared to the controls. 
Increased incidences of granulomatous hepatitis of minimal severity were
seen in high-dose males (controls, 11%; 1000 ppm, 37%, p<0.01) and
females (controls, 11%; 1000 ppm, 21%, p<0.01).  Higher incidences of
aggregates of brown pigmented macrophages were seen in the livers of
treated males (controls, 13%; 100 ppm, 27%, p<0.05; 1000 ppm, 19%,
p<0.01) and females (controls, 15%; 1 ppm, 40%, p<0.01; 10 ppm, 21%, NS;
100 ppm, 38%; 1000 ppm, 50%, p<0.01).  Granulomatous hepatitis and brown
pigmented macrophage aggregates were most commonly seen in mice that
survived until the end of the study.  The only effects that were not
associated with the liver were an increased incidence of thymic
hyperplasia in high-dose females (controls, 5%; 1000 ppm, 21%, p <0.01),
and increased incidences of cystic thyroid follicles in high-dose males
(controls, 23%; 1000 ppm, 52%, p<0.01) and high-dose females (controls,
16%; 1000 ppm, 33%, p<0.01). 

The central nervous systems of the animals were re-examined and the
results reported in an addendum to the main study (MRID 44807220). 
Treatment-related increases in the incidences and severity of
vacuolation of white matter occurred in the brains of males at 1000 ppm
and increased severity of white matter vacuolation was seen in the
brains of females at 1000 ppm compared to the control groups.  No clear
effect of treatment on the incidence or severity of white matter
vacuolation in the spinal cord was seen in either sex, and no
treatment-related effects were seen at 1, 10, or 100 ppm.

 

The LOAEL is 100 ppm in the diet (10.7 mg/kg/day for males; 11.7
mg/kg/day for females), based on increased incidences of brown pigmented
macrophages in the liver of both sexes, increased incidences of
eosinophilic vacuolated hepatocytes in males, and increased liver
weights in females.  The NOAEL was 10 ppm (1.1 mg/kg/day for males; 1.2
mg/kg/day for females).

In this study, there were statistically significant positive trends for
hepatocellular adenomas, carcinomas and combined adenomas/carcinomas for
the male mice.  There were also statistically significant  increases by
pair-wise comparison of the male high dose group (1000 ppm) with the
controls for hepatocellular adenomas (34% vs. 16% in controls), for
hepatocellular carcinomas (34% vs. 19% in controls) and for combined
hepatocellular adenomas/carcinomas (62% vs. 33% in controls).  The
incidence of hepatocellular adenomas (34%) at the highest dose level for
males exceeded the highest incidence in the historical control data for
1981-1983 (4-27%) and for 1986-1988 (8-23%), and the incidence of
hepatocellular carcinomas (34%) for the highest dose level for males
exceeded the highest incidence in the historical control data for
1986-1988 (5-13%), but not for 1981-1983 (12-38%).  There were no
treatment-related tumors observed in the female mice in this study.  
The highest dose level tested was considered to be adequate but not
excessive because liver and brain toxicity were observed in the male and
female mice at 1000 ppm.  Although there were no significant changes in
survival or body weight gain, mean liver weight gains were increased in
males and females and histopathological changes were observed in the
livers and brain (vacuolation of the white matter) of males and females.

This carcinogenicity study in the mouse is Acceptable/Guideline and does
satisfy the guideline requirement for an carcinogenicity study [OPPTS
870.4200 (83-2b)] in mice. An additional study has been done following
this study with higher concentrations of fluazinam (see MRID 44807222). 

In a range-finding study (MRID 44807212), groups of 10 male and 10
female CD(1 mice were given concentrations of 0, 10, 50, 250, or 3000
ppm fluazinam (B-1216) in the diet for 4 weeks.  There were no
significant changes in body weights between treated and control animals;
however, group mean body weight gain was slightly less in both sexes at
250 and 3000 ppm.  The body weight change did not show a clear dose
dependency especially in females.  Platelet counts were marginally
elevated in males at 3000 ppm and total blood cholesterol was slightly
higher in both sexes at 3000 ppm compared to the controls.  Phospholipid
concentration was slightly increased in females and marginally increased
in males at 3000 ppm.  Blood glucose concentrations were increased in
females at 250 and 3000 ppm compared to the control group.  Absolute and
relative (to body weight) liver weights were increased at 3000 ppm in
both sexes and the absolute and relative kidney weights were increased
in females at 3000 ppm.  Hepatocyte periacinar hypertrophy incidences
and severity were increased in both sexes at 3000 ppm compared to the
control groups.

870.4200b 	Carcinogenicity (feeding) - Mouse

Executive Summary: In another carcinogenicity study (MRID 44807222,
44807221, 45201301, 44807211), technical grade Fluazinam (97.0% a.i.,
lot no. 1030/91) was administered to groups of 50 male and 50 female
Crl:CD(-1 mice in the diet at concentrations of 0, 1000, 3000, or 7000
ppm.  The test diets were given for 97 weeks to females and for 104
weeks to males.  These concentrations resulted in mean daily compound
intakes of 126, 377, and 964 mg/kg/day for males and 162, 453, and 1185
mg/kg/day for females for 1000 ppm, 3000 ppm, and 7000 ppm,
respectively.  Twenty mice were added to the control and 7000 ppm groups
for a satellite study in which the mice were killed and necropsied after
treatment for 78 weeks.  A Pathology Working Group (PWG) report
presenting revised incidences for hepatocellular tumors in the male mice
in this study was later submitted (MRID 45201301).  A four-week
range-finding study in mice was also conducted using 0, 3000, 5000, and
7000 ppm in the diet (MRID 44807211).

Treatment with fluazinam resulted in a significant decrease in survival
in females at 7000 ppm (control, 58%; 7000 ppm, 26%, p <0.01).  All
females were terminated after 97 weeks of treatment because of low
survival at the high-dose.  At 7000 ppm, body weight gain was decreased
in males during weeks 4-36 by 32% (p< 0.01) and food conversion ratios
over weeks 9-13 were increased by 86% compared to the controls
indicating decreased efficiency of food utilization.  At termination,
relative liver/body weight ratios were increased in males by 54%, 113%
and 182% and in females by 21%, 45%, and 109% at 1000, 3000, and 7000
ppm, respectively, compared to the controls (p<0.01).  Microscopic
examination showed increased incidences of altered hepatocyte foci at
all concentration levels in males and in high-dose females (males:
control, 12/50; 1000 ppm 24/50, p<0.05; 3000 ppm, 36/50; 7000 ppm,
33/50, p<0.01; females: control, 3/50; 7000 ppm 15/50, p<0.01). 
Incidences of hepatocyte enlargement, pale or vacuolated hepatocyte
cytoplasm, and brown pigmented macrophage aggregates were increased in
all treated males and females compared to the control groups (p<0.01). 
The pigmented macrophage aggregates also increased in severity from
0-22% of lesions in the controls to 41-58% of lesions at 7000 ppm graded
moderate or marked.  Incidences of brown pigmented centrilobular
hepatocytes and parenchymal inflammatory cells increased in males at
3000 ppm (both 6/50, p<0.05) and 7000 ppm (11-16/50, p<0.01) compared to
the controls (0-1/50).  Males were more sensitive to the hepatotoxic
effects of fluazinam than females.  Incidences of vacuolation of white
matter in the brains of all treated animals were increased compared to
the controls (p<0.01).  Vacuolation of white matter was also increased
in the cervical spinal cord of males at 3000 and 7000 ppm (control,
18/50; 3000 ppm, 37/50, p<0.05; 7000 ppm, 46/50, p<0.01) and marginally
in females (control, 37/50; 3000 and 7000 ppm, 45/50, NS).  The severity
of the vacuolation of white matter in the brain and spinal cord
increased with increasing dose from 0% graded moderate or marked in the
controls to 33-60% of lesions at 7000 ppm.  Incidences of left atrial
thrombus in the hearts of high-dose males and females were increased
compared to the controls and contributed to the unscheduled deaths of
about 46% of high-dose males and 30% of high-dose females during the
study.  

The LOAEL is 1000 ppm in the diet (126 mg/kg/day for males; 162
mg/kg/day for females), based on increased liver weights in males and
females and on histopathological changes in the liver and brain in males
and females.  A NOAEL was not determined (<1000ppm).

In this study, there were no statistically significant positive trends
for hepatocellular adenomas, carcinomas or combined adenomas/carcinomas
for the male mice.  For the mid-dose male mice (3000 ppm), however,
there was a statistically significant increase by pair-wise comparison
with the controls for hepatocellular adenomas (47% vs. 17% in controls)
and for combined hepatocellular adenomas/carcinomas (49% vs. 18% in
controls).  For the high-dose male mice (7000 ppm), there was no
statistically significant increase by pair-wise comparison with controls
for hepatocellular adenomas or carcinomas when considered separately,
but there was a statistically significant increase for combined
adenomas/carcinomas (33% vs. 18% in controls).   The incidence of
hepatocellular adenomas at the mid-dose level for males (47%) exceeded
the highest incidence in the historical control data for 1991-1993
(8-34%) and for 1987-1993 (0-31%), but the incidence at the high-dose
level for males (30%) did not exceed the highest incidences in the
comparable historical control data.  Similarly, the incidence of
combined hepatocellular adenomas/carcinomas at the mid-dose level for
males (49%) exceeded the highest incidence in the historical control
data for 1987-1993 (4-42%), but the incidence at the high-dose level for
males (33%) did not exceed the highest incidences in the comparable
historical control data.  For the male mice in this study, the
tumorigenic response did not appear to be dose-related because the
response at 7000 ppm was less than that observed at 3000 ppm.  The
highest dose level tested for the male mice in this study was considered
to be adequate but not excessive.  In this study, hepatocellular tumors
were also observed in the female mice at the high-dose (7000 ppm). 
These tumors, however, occurred at an excessively toxic dose which may
have resulted in indirect effects that may not have been present at
lower doses.  There was a treatment-related increased mortality for the
high-dose females in this study.  Although a statistically significant 
positive trend was observed for combined hepatocellular
adenomas/carcinomas for the female mice in this study, this calculation
included the response at 7000 ppm.  At the next lower dose level (3000
ppm), there was no statistically significant pair-wise increase in
hepatocellular adenomas, carcinomas or combined adenomas/carcinomas for
the female mice in this study.   

           

This carcinogenicity study in the mouse is Acceptable/Guideline and does
satisfy the guideline requirement for an carcinogenicity study [OPPTS
870.4200 (83-2b)] in mice.

Executive Summary: Treatment levels of 0, 3000, 5000, or 7000 ppm in the
diet were given to mice in a 4-week study (MRID 44807211). 
Treatment-related changes were seen in the liver of mice at all
treatment levels and kidney effects were seen in males at 5000 and 7000
ppm.  Treatment-related increased incidences and severity of vacuolation
of white matter of the brain were seen at four weeks in males at 3000,
5000, and 7000 ppm and in females at 7000 ppm.  Treatment-related
increased incidences and severity of vacuolation of white matter of the
spinal cord were also seen in males at 5000 and 7000 ppm.  

The LOAEL for vacuolation of white matter in the brain of male mice in
this four-week study was 3000 ppm (555 mg/kg/day; lowest dose level
tested).  A NOAEL was not demonstrated for this effect (i.e. NOAEL <555
mg/kg/day).

870.4300 	Combined Chronic Carcinogenicity Study – Rat

EXECUTIVE SUMMARY:  In a combined chronic toxicity/carcinogenicity study
(MRID 42248620, 44807223, 451450201), fluazinam technical (95.3% a.i.,
lot number 8412-20) was administered to groups of 60 male and 60 female
Sprague-Dawley rats at dietary concentrations of 0, 1, 10, 100, or 1000
ppm (0, 0.04, 0.38, 3.8, or 40 mg/kg/day for males and 0, 0.05, 0.47,
4.9, or 53 mg/kg/day for females) for up to 104 weeks.  Groups of 10
rats of each sex per dose group were sacrificed at 52 weeks for interim
evaluations.

No treatment-related effects were observed in rats receiving the 1 ppm
or 10 ppm diets.  No treatment-related effect on mortality was observed
in rats receiving any dose of the test material.  The only clinical
signs observed were straw-discoloration of the fur in all rats receiving
the 1000 ppm diet and an increased incidence of alopecia in females
receiving the 1000 ppm diet.

8% less food than controls at each weekly interval.  Females
receiving the 1000 ppm diet weighed 7–24% (p<0.01) less than controls
from week 2 to termination, gained 35% less weight overall, and consumed
18% less food than controls at each weekly interval.  The food
utilization factor or the food efficiency suggested that reduced body
weight gain was due in part to toxicity of the test material.  No
treatment-related effects were observed on body weights, body weight
gain, food consumption, or food utilization/efficiency in male or female
rats receiving the 1-, 10- or 100-ppm diets.  No treatment-related
effects were observed on the eyes at any dose at any time during the
study.  Clinical pathology evaluations showed only mild anemia and
elevated cholesterol in both sexes receiving the 1000 ppm dose.  

Treatment-related microscopic findings showed that the test material was
toxic to the following organs: liver, pancreas, lungs, and possibly
thyroid gland in both sexes, kidneys and testes in males, and lymph
nodes in females, but the primary target appeared to be the liver. 
Liver toxicity was manifested by increased organ weight and increased
incidences of gross and microscopic lesions.  Absolute liver weights
were increased by at least 7-27% and relative weights by 17-63% in both
sexes at 1000 ppm.  Gross lesions in the liver consisted of pale,
swollen or accentuated markings on the livers at 1000 ppm in males and
enlarged, pitted, or mottled livers at 1000 ppm in females. 
Treatment-related microscopic liver lesions at 100 ppm consisted of
eosinophilic hepatocytes in 22% of females (8% in controls),
centrilobular hepatocyte rarefaction and vacuolation in 8% of each sex
(0% for controls), centrilobular sinusoidal dilatation in 10% of males
and 18% of females (0% for male and 2% for female control), and
pericholangitis in 18% of males and 14% of females (4% for male and 2%
for female controls).  Additional treatment- related liver lesions at
1000 ppm in main study group consisted of centrilobular hepatocyte
vacuolation in males, centrilobular hepatocyte necrosis in females, and
centrilobular fat and bile duct hyperplasia in both sexes. 
Centrilobular hepatocyte vacuolation and centrilobular fat was also seen
in 1000 ppm group male and female rats at interim sacrifice.

The incidences of exocrine atrophy of the pancreas in both sexes and
acinal epithelial vacuolation or fat accumulation in females were
increased at 1000; the incidence of exocrine atrophy was also increased
at 100 ppm in females compared with that of control rats.  The incidence
of exocrine degranulation was increased in 1000 ppm group females rats
at interim sacrifice but not in the main study.  An increased incidence
of thyroid follicular hyperplasia was observed in males at 1000 ppm (8%
vs 2% for controls) and in females at 1000 ppm (10% vs 2% in controls). 
This  finding may possibly be related to treatment with the test
material.  In male rats, the incidence of cortical tubular basophilia in
the kidney was increased at 1000 ppm compared with that of the controls.
 Other treatment-related lesions included  pneumonitis, alveolar
adenomatosis, and alveolar epithelialization in 1000 ppm group males,
alveolar epithelialization and alveolar macrophage aggregates in 1000
ppm group females, testicular atrophy in 100 ppm and 1000 ppm group
males, and spermatocele granuloma also in 1000 ppm males.  The incidence
of sinus histiocytosis in the lymph nodes was increased in1000ppm group
females.  Histopathologic assessment of the brain and spinal cord of
rats in the control and 1000 ppm dose groups showed no treatment-related
effect on vacuolation of white matter.

The lowest-observed-adverse-effect level (LOAEL) for fluazinam was 100
ppm (3.8 mg/kg/day for males and 4.9 mg/kg/day for females) based on
liver toxicity in both sexes, testicular atrophy in males and pancreatic
exocrine atrophy in females.  The corresponding
no-observed-adverse-effect level (NOAEL) was 10 ppm (0.38 mg/kg/day for
males and 0.47 mg/kg/day for females).

CARC Comments: In this study, there were statistically significant
positive trends for thyroid gland follicular cell adenocarcinomas and
combined follicular cell adenomas/adenocarcinomas for the male rats.  
There was also a statistically significant increase by pair-wise
comparison of the male high dose group (1000 ppm) with the controls for
combined follicular cell adenomas/adenocarcinomas (23% vs 8% in
controls).  In addition to an Exact Trend Test and a Fisher’s Exact
Test, a Peto’s Prevalence Test was also conducted (which excluded
animals that died or were sacrificed before observation of the first
tumor at week 68).  For follicular cell adenocarcinomas in males,
results of the Peto’s Prevalence Test showed a statistically
significant positive trend and a borderline statistically significant
(p= 0.056) increase by pair-wise comparison of the 1000 ppm male group
with the controls (7% vs 0% in controls), indicating the increased
incidence of thyroid tumors had a malignant component to it.  For
combined follicular cell adenomas/adenocarcinomas, Peto’s Prevalence
Test also showed a statistically significant increase by pair-wise
comparison of the high dose male group with the controls (26% vs 9% in
controls).  The incidences of thyroid gland  adenomas at 100 ppm and
1000 ppm (15% and 17%, respectively) and adenocarcinomas at 1000 ppm
(6%) were slightly outside  their respective ranges in the historical
control data (range: adenomas, 0%-13%; adenocarcinomas, 0%-5%).  Animals
in the lower dose groups were not microscopically examined for thyroid
lesions unless abnormalities were observed in that organ at gross
necropsy.  Therefore, percentage incidences of thyroid tumors in these
lower dose groups may have been somewhat misleading (too high).  The
highest dose level tested in this study was considered to be adequate
and not excessive because there were decreased body weight gains (up to
15% and 35% in males and females, respectively), decreased food
consumption, decreased food efficiency, increased thyroid weights at 52
weeks, enlarged thyroids and a slightly increased incidence of thyroid
gland follicular cell hyperplasia at 104 weeks in males. The survival of
the animals was not decreased by treatment with the test material. 
There was no treatment-related increase in the thyroid tumor incidence
in the female rats in this study.  In addition, slightly increased
incidences of pancreatic islet cell adenomas were also observed in the
treated male rats, but these incidences were not dose-related and did
not exceed the upper range of historical control data for the same tumor
type (20 comparable studies at the same testing laboratory).  Further,
this type of tumor in this strain of rats is a common spontaneously
occurring neoplasm.  Therefore, the increased incidence of pancreatic
islet cell adenomas observed in the treated male rats in this study was
considered not likely to be related to treatment with the test material.
 High incidences of pituitary gland adenomas in males and females and of
mammary gland fibroadenomas in females were also observed in all groups,
including controls.  These neoplasms are not considered to be
treatment-related.  A toxicologically significant increase in tumors was
not observed in any other tissues in the treated male or female rats in
this study.

This combined chronic toxicity/carcinogenicity study in the rat is
Acceptable/Guideline and satisfies guideline requirements for a chronic
toxicity/carcinogenicity study [OPPTS 870.4300 (§83-5)] in rats.

A.3.6	Mutagenicity

Gene Mutation

Guideline 870.5100 

Bacterial reverse mutation assay (Ames Test) with S. typhimurium and E.
coli

MRID 42270605

Acceptable	

ut S9 up to cytotoxic concentrations.  Compound was tested up 2μg/plate
for S. typhimurium strains and up to 250 μg/plate for E. coli in the
absence of S9, and up to 100 μg/plate for S. typhimurium strains and up
to 500 μg/plate for E. coli in the presence of S9.

Guideline 870.5100 

Bacterial reverse mutation assay (Ames Test) with S. typhimurium and E.
coli

MRID 42270604

Acceptable	

Negative with and without S9 up to cytotoxic concentrations.  Compound
was tested up 1μg/plate for S. typhimurium strains and up to 250
μg/plate for E. coli in the absence of S9, and up to 100 μg/plate for
S. typhimurium strains and up to 500 μg/plate for E. coli in the
presence of S9.

Guideline 870.5300 

Mammalian cells in culture forward gene mutation assay with mouse
lymphoma L5178Y/TK +/- cells

MRID 45156902

Acceptable	

Negative with and without S9 up to 5 μg/mL.  Compound was tested up to
cytotoxic concentrations.

Guideline 870.5300 

Mammalian cells in culture forward gene mutation assay with mouse
lymphoma L5178Y/TK +/- cells

MRID 45261801

Acceptable	

Negative with S9 up to 9 μg/mL and without S9 up to 0.3 μg/mL. 
Compound was tested up to cytotoxic concentrations.

Cytogenetics

Guideline 870.5375

In vitro mammalian chromosome aberration (CHL cells)

MRID 42270606

Acceptable	

Negative with and without S9 up to cytotoxic concentrations.  Compound
was tested up 4 μg/mL in the absence of S9, and up to 9.5 μg/mL in the
presence of S9.  Cells harvested at 24 and 48 hours in nonactivated
studies and at 24 hours in activated studies.

Guideline 870.5395

Mammalian erythrocyte micronucleus test (mouse)

MRID 44807224

Acceptable	

Negative.  Test compound at 500, 1000, or 2000 mg/kg (oral gavage) with
a 24 hour sacrifice or at 2000 mg/kg with 24, 48, and 72 hour sacrifices
did not induce the formation of micronuclei in polychromatic
erythrocytes from bone marrow.  Clinical signs included piloerection,
decreased motor activity, loose stools, and/or soiled fur.

Other Genotoxicity

Guideline 870.5550

Unscheduled DNA synthesis in primary rat hepatocytes

MRID 45156901

Unacceptable	Negative; however, there were several serious study
deficiencies: treatment time shorter than recommended, no data
supporting the claim of cytotoxicity, and data variability for major
endpoints.

Guideline 870.5500 

Differential Killing/Growth Inhibition in B. subtilis

MRID 42270607

Negative without S9 up to 0.3 μg/disk and with S9 up to 30 μg/disk. 
Deficiency: only one replicate per plate dose was used.

A.3.7	Neurotoxicity

870.6200 	Acute Neurotoxicity Screening Battery

Executive Summary: In an acute oral neurotoxicity study (MRID 44807210),
single gavage doses of 0, 50, 1000 or 2000 mg/kg fluazinam (96.8%, Lot
No.: 1030/91) in 1.5% (w/v) aqueous methylcellulose were administered to
groups of fasted Sprague-Dawley rats (10/sex/dose).   Functional
Observational Battery (FOB) and Motor Activity (MA) assessments were
performed before test substance administration, between 5 and 7 hours
post-administration (time of peak effect) and on days 7 and 14.  Body
weights were measured weekly and clinical signs were recorded daily.  
The animals were sacrificed and grossly examined 14 days after
administration of the test material.  Five rats/dose/sex were perfused
in situ for neuropathological evaluation.

There were no treatment-related deaths, clinical signs, or body weight
effects during the study.  Treatment-related soft stools were observed
in mid- and high-dose males and females, but only on the day of
treatment.  Treatment-related mean motor activity values were
significantly decreased (23-65%) in mid- and high-dose females compared
to controls (not dose-related), but only on the day of treatment.   No
other treatment-related effects of any kind were observed at any time
during the study.  No treatment-related gross effects or histopathology
were observed. Since the decreased mean motor activity values were
observed in one sex only (females), on one occasion only (on the day of
dosing), at high doses only (>1000 mg/kg), and were not dose-related,
and were observed in one study only (not observed in the subchronic
neurotoxicity study in rats, MRID 44807217, 44807218), it is likely that
this treatment-related effect (decreased mean motor activity values in
females only on the day of dosing only) is a manifestation of acute
general systemic toxicity and not a direct neurotoxic response to the
administration of the test material. 

Under the conditions of this study, the acute general systemic toxicity
LOAEL is 1000 mg/kg for male and  female rats based on soft stools and
decreased motor activity.  The acute general systemic toxicity NOAEL is
50 mg/kg for male and female rats.  The LOAEL for neurotoxic effects is
not identified (> 2000 mg/kg).  The NOAEL for neurotoxic effects is 2000
mg/kg. 

 

This acute oral neurotoxicity study is classified Acceptable/ Guideline.
This study does satisfy the guideline requirement for an acute oral
neurotoxicity study [OPPTS 870.6200 (81-8ss)] in rats.

870.6200 	Subchronic Neurotoxicity Screening Battery

Executive Summary: In two subchronic oral neurotoxicity studies (MRID
44807217 & MRID 44807218), groups of 10 male and 10 female Crl:CD BR
rats were fed diets containing 0, 300, or 1000 ppm fluazinam (MRID
44807217, 96.9%, Lot No. 6109) or 0, 1000, 2000, or 3000 ppm fluazinam
(MRID 44807218, 98.4%, Lot No. 9601-2) for 13 weeks.   Achieved doses
were 20.7, 69-74, 149, and 233 mg/kg/day for males in the 300, 1000,
2000, and 3000 ppm groups, respectively; and 23.4, 81-89, 175, and 280
mg/kg/day for females in the 300, 1000, 2000, and 3000 ppm groups,
respectively.  Functional Observational Battery (FOB) and Motor Activity
(MA) assessments were performed prior to treatment and during weeks 4,
8, and 13 of treatment.  Body weights, food consumption, and clinical
signs were monitored throughout the study.   At the end of the treatment
period, all rats were perfused in situ.  The brain from all rats was
removed, weighed, and measured and 5 males and 5 females from the
control and high-dose groups of each study were subjected to
neuropathological evaluation.

There were no treatment-related deaths or clinical signs.  At the end of
the study, group mean body weight gains were significantly (p<0.01)
decreased in females in and above the 1000 ppm groups and in males in
the 2000 and 3000 ppm groups.  Similarly, cumulative food consumption
was decreased in males (p<0.01) and females (p<0.05) fed 2000 and 3000
ppm fluazinam.  Food efficiency was decreased in males at 3000 ppm and a
dose-related decrease in food efficiency was observed in females in all
treatment groups.

No treatment-related FOB or MA effects were observed.  Brain weights of
females in the 3000 ppm group were 8% lower (p<0.01) than controls;
however, no supporting pathology was observed.  No treatment-related
gross effects or histopathology were observed. 

Under the conditions of these studies, the neurotoxicity NOAEL is 3000
ppm (233 mg/kg/day) for male rats and 3000 ppm (280 mg/kg/day) for
female rats.  A neurotoxicity LOAEL was not identified. 

These subchronic neurotoxicity studies are classified as
Acceptable/Guideline. When considered together, these studies satisfy
the guideline requirement for a subchronic neurotoxicity study [OPPTS
870.6200 (82-7ss)] in rats.

870.6300 	Developmental Neurotoxicity Study

EXECUTIVE SUMMARY: In a developmental neurotoxicity study (2005, MRID
46534401), Fluazinam (97.8% a.i.; lot # A629/1995, impurity #5 0.09%)
was administered by gavage to 24 Crl:CD® (SD) IGS BR rats/sex/dose at
0, 2, 10 or 50 mg/kg/day from gestation day (GD) 6 through lactation day
(LD) 20.  The pups were administered the same doses by gavage from
postnatal days (PND) 7 to 20 or 21. Maternal evaluation consisted of a
Functional Operational Battery (FOB) was performed on GDs 12 and 18, and
on post partum days (PPDs) 35, 45 and 60. Additional behavioral
assessments included: motor activity on GD 15 and PPD 60; auditory
startle habituation on GD 19 and PPD 58; and learning and memory on LD
16 and PPD 61.  From each maternal group, 12 were sacrificed on both LD
21 and PPD 66; of these, 10/group were selected for neuropathology
procedures. The additional assessment of the dams is related to trying
to further characterizing neurotoxicity that is attributed to impurity
#5. On postnatal day (PND) 4, litters were culled to yield four males
and four females (as closely as possible).  Offspring were allocated for
detailed clinical observations (FOB) and assessment of motor activity,
auditory startle reflex habituation, learning and memory (watermaze
testing), and neuropathology at days 23/24 and on PND day 66.   On PND
21, the whole brain was collected from 10 pups/sex/dose group for
micropathologic examination and morphometric analysis.  Pup physical
development was evaluated by body weight.  The age of sexual maturation
(vaginal opening in females and preputial separation in males) was
assessed.

Maternal parameters.  No effects on absolute body weight were noted.
Mean body weight gain for GDs 6-14 was significantly decreased (14%) at
10 and 50 mg/kg/day and during GDs 6-20 at 50 mg/kg/day (10%).  Mean
body weight gain during lactation was not affected.  Food consumption
during gestation was comparable to controls but during lactation was
significantly decreased at 10 (10 to 14%) and 50 mg/kg/day (7 to 11%).
Mean body weight and body weight gain post weaning (days 28-63) were not
affected. The weight gain and food consumption data that occurs in the
absence of absolute weight differences for the dams are not considered
to be of sufficient magnitude to be included as a true toxic response.
Reproductive parameters and behavioral assessments, including FOB, motor
activity, auditory startle reflex habituation and learning and memory,
were not affected by treatment.  No treatment-related changes were
observed at either the necropsy on LD 21 or PPD 66.  The characteristic
grey matter vacuolation attributed to fluazinam in previous studies was
not seen at LD 21 or PPD 66.  The maternal LOAEL for Fluazinam was not
established.  The maternal NOAEL is > 50 mg/kg/day. The weight gain and
food consumption data that occurs in the absence of absolute weight
differences for the dams are not considered to be sufficient of
sufficient magnitude to be included as a true toxic response.

Offspring parameters. No treatment-related effects were observed on
litter size at birth or survival to weaning.  Birth weight was lower in
females at 10 and 50 mg/kg/day (both 6%, p < 0.01) but not in males.
Mean offspring body weight was significantly decreased in males and
females at 10 (6-11%) and 50 mg/kg/day (6-16%) during lactation.  Mean
body weight gain was significantly decreased in male and female pups at
10 (4-24%) and 50 mg/kg/day (16-35%) during lactation. During the
post-weaning period (PNDs 28-63), mean body weight was significantly
decreased in males and females at 10 (3-7%) and 50 mg/kg/day (7-15%). 
However, post weaning body weight gain was essentially comparable to the
control group.  The mean age of completion of balano-preputial
separation was significantly delayed at 10 and 50 mg/kg/day.   Rearing
counts in female pups were decreased on day 21 in the 10 (to a mean of
3.5 vs. 8.1 in the controls) and 50 (mean 3.7) mg/kg/day dose groups. 
Dark and/or distended abdomens observed in a total of 12 offspring from
4 litters at 50 mg/kg/day were considered treatment-related since
similar signs were observed in the preliminary study. The peak amplitude
in the auditory startle response was affected in males in the high dose
group at day 23/24.  Absolute brain weight for high dose males was 6.1%
decreased on PND 21 and slight changes in brain width were noted.  No
treatment-related effects were observed on other behavioral assessments,
including FOB, motor activity or learning and memory.  Grey matter
vacuolation was not seen in the LD21 neuropathology assessment.  A
single isolated incident of grey matter vacuolation in the high dose
group at day 66 was not considered related to treatment.  The offspring
LOAEL is 10 mg/kg/day based on decreased body weight and body weight
gain and delay in completion of balano-preputial separation.  The
offspring NOAEL is 2 mg/kg/day.

This developmental neurotoxicity is classified as
Acceptable/Non-Guideline and may be used for regulatory purposes.  It
does not, however, satisfy the guideline requirement for a developmental
neurotoxicity study in rats (OPPTS 870.6300, §83-6): OECD 426 (draft)
due to the pending review of the positive control data. 

A.3.8		Metabolism

870.7485		Metabolism – Rat

Executive Summary: In a metabolism characterization study (MRID No.
44807233), Fluazinam (IKF-1216) was administered by gavage at single
doses of 0.5 mg/kg or 50 mg/kg, or 14-day repeated doses of 0.5
mg/kg/day.  In addition to nonlabeled IKF-1216 (lot no. T9002, 99.6%
purity), [14C]-IKF-1216 labeled on the phenyl moiety (lot. No. 93-5,
purity 98%, sp. act. 57.3 mCi/mmol) or pyridyl moiety (lot. No. 93-90,
98% purity; sp. act. 66.2 mCi/mmol)  were also administered in some
studies to assess metabolic cleavage of the phenyl or pyridyl ring of
the test material.  Experimental groups were established for overall
distribution/excretion assessment and for analysis of biliary secretion.
The metabolite profiles of urine, feces, and bile were examined and
major metabolites were identified.

There were no treatment-related deaths in the rats.  Overall recovery of
the administered radioactivity (reported in MRID Nos. 43521006,
43521007, and 43521008 and evaluated in a separate DER) was acceptable
(93.10-103.55%).  Excretion via the urine was minor.  AMPA mercapturate
and DAPA, the major urinary metabolites, represented only 0.05-0.39% of
the administered dose.  Radioactivity in the feces represented most of
the administered dose (88.78-100.03%) as determined by review of MRID
Nos. 43521006, 43521007, and 43521008 and evaluated in a separate DER. 
Identified fecal metabolites, however, represented only 11.20-68.59% of
the administered dose. For all dose groups, most of the fecal
radioactivity appeared to reside with unextractable components in the
post-extraction solids (PES).  Further analysis of the PES components
using base hydrolysis indicated that most of this radioactivity could be
attributed to hydrolysis products of AMPA and DAPA.   PES radioactivity
was also greatest for the low-dose group which was consistent with the
lower overall accounting of identified metabolites for this group. 
Approximately 20-25% of the aqueous phase of the fecal extraction was
identified as a cysteine conjugate of DAPA and represented <1% of the
administered dose. With the exception of the low-dose group, parent
compound represented most of the identified radioactivity in the feces. 
AMPA and DAPA were identified in the feces from all dose groups but
these metabolites never represented more than 5% of the administered
dose (except for high-dose female rats where AMPA accounted for 10.22%).
 

DAPA glucuronide  and AMPA mercapturate were the major biliary
metabolites but represented  <4% of the administered dose.  Total
biliary radioactivity, however represented 25-34% of the administered
dose (MRID Nos. 43521006, 43521007, and 43521008 evaluated in a separate
DER).  Analysis of chromatograms indicated that numerous other
metabolites were present in the bile but were individually of
insufficient quantity to allow for characterization.  

Metabolite profiles from administration of different label positions
(pyridyl and phenyl) indicated that there was no metabolic cleavage of
the ring structures.  Minor quantitative differences in metabolite
recovery were observed between genders but not of sufficient magnitude
to suggest biologically relevant differences in the metabolism of
IKF-1216.

This metabolism study is Acceptable/Guideline.  When considered together
with the previously submitted general metabolism studies on IKF-1216
(MRIDs 43521004 through 43521008 and MRID 43553001), the requirement for
a general metabolism study in rats [OPPTS 870.7485 ((85-1)] is
satisfied. 

870.7600	Dermal Absorption 

There is no dermal absorption study with fluazinam and no study has been
requested. 

A.3.9			Special/Other Studies 

Nonguideline  	90-Day Oral Toxicity – Rat (Special Liver Toxicity
Study)

Executive Summary: In a 90-day oral toxicity study (special liver
toxicity study) (MRID 42248609), groups of 10 male and 10 female CD
rats were given Fluazinam (a.i. 98.5%, Lot/Batch # 8303-2) administered
at 0 or 500 ppm (equivalent to average mg/kg/day levels of 0 and 37.63
in males and 0 and 44.71 in females) in the diet.  An additional group
of 10 male and 10 female rats were given Fluazinam at 0 or 500 ppm in
the diet for 90 days, then given no Fluazinam for 4 weeks in a
reversibility study.

≤ 0.01) and females (+15%, p≤ 0.01), and periacinar hepatocytic
hypertrophy in 10/10 males (0/10 controls).  These effects  resolved
after the 4-week reversibility phase.

This special liver toxicity study in rats is classified as Acceptable
/Nonguideline.  The study did not fully meet its objective of assessing
the hepatotoxic effects of the test material or determine their
reversibility because only one dose (500 ppm) of Fluazinam was utilized,
and the modest liver changes observed were of questionable toxicological
significance.

Nonguideline  	11-Week Oral Toxicity – Dog (Special Retinal Toxicity
Study)	

Executive Summary: In an 11-week oral toxicity study (special retinal
toxicity study) (MRID 44807216), Fluazinam (2 batches from Lot no.
8303-2: 98.0% and 98.1% a.i.) was administered in gelatin capsules for
11 weeks to 6 male beagle dogs/group at 0 or 200 mg/kg/day (dose 
reduced to 150 mg/kg/day in two animals at week 3, and in the remaining
four animals at week 5, due to excessive toxicity).  Three
animals/dose group were terminated on study day 77 (Group II;
corresponding controls Group I), and the remaining 3 animals/group
remained on study without treatment for 5 weeks before termination on
study day 112 (Group IIW; corresponding controls Group IW).  This study
was designed to evaluate retinal changes seen at 100-150 mg/kg/day in
previous 28- and 90-day studies, follow the time course of any changes,
assess retinal function and assess reversibility.  In addition to
ophthalmologic evaluation, fundic photography and electroretinography
were performed pretest, weekly during treatment and at weeks 2 and 4 of
compound withdrawal.  Eyes were examined microscopically and by electron
microscopy.

At 150 mg/kg/day (maximum tolerance level), dogs experienced frequent
loose/liquid feces, vomiting, inappetence, excess salivation,
vasodilation (as noted by pink or reddened ears and/or abdomens); these
findings were observed sporadically or not at all in controls.  Unusual
behavioral (2 males) and motor changes (1 male)were observed on the
first day of dosing only.  Decreased mean body weight/weight gains
during treatment (at termination of treatment, -15%/-89% less than
corresponding control group, Group II; -6%/weight loss of -0.61 kg,
Group IIW), were also observed.  Food consumption was also decreased
during treatment (mean estimated at >10%, Group II and >6%, Group IIW;
see Results for details) but not withdrawal (measurements were not
possible for several weeks due to mixing with unspecified amounts of
supplemental diet).  Mean ALP, ALT, AST and blood urea levels were
elevated due to increases above the provided reference range in 1-2
animals/dose group.  Instead of grey mottling of the retinal tapetal
fundus noted in previous studies, increased brown granularity of the
tapetal fundus was observed in the tapetal fundus of one main study
animal and in one withdrawal group treated animal, although this effect
was too subtle to be demonstrated photographically.  Electroretinography
(ERG) demonstrated that dosing was associated with decreased a- and
b-wave amplitudes of (50%, but waveform was not altered. Other
measurements of scotopic (dark or dim light) vision were not
significantly altered and there was no evidence of neural damage. The
ERG amplitudes in 2/3 treated animals recovered to some degree during
withdrawal.

This eleven-week oral toxicity in the dog is Unacceptable/Nonguideline
(not upgradeable).   This study did not repeat the ophthalmologic
findings observed in the previous 28- and 90-day studies (grey mottling
of the retina), although this did not necessarily render this special
study results invalid and brown granularity in the retina was observed
in 2 treated dogs. However, several factors compromised the integrity of
the study:  unexplained incongruity in the ERG data between control
groups and between the control and treated animals and other technical
problems in the ERG evaluation, double-penning of the animals and the 
possibility of illness due to an infectious agent, unspecified technical
problems, errors and uncertainties in food consumption data, a striking
parallelism of mean body weights for control and treated animals and
unexplained decreased gain in the withdrawal control animals.  Despite
these problems, the study does appear to support the conclusion that the
retina is a target tissue for fluazinam and that functional as well as
morphological alterations may result from exposure.  

Nonguideline  	7-Day Inhalation Toxicity (Range-Finding)- Rat	

                              (Test Material:  Frowncide( WP (51.9%
fluazinam, a.i.)

Executive Summary: In a 7-day inhalation toxicity (range-finding) study
(MRID 42248621), groups of five male and five female young adult CD rats
 were exposed nose-only to Frowncide( WP (51.9% Fluazinam, a.i., Batch
No. 004) for two 3-hour periods per day for 7 days at concentrations of
0, 0.003, 0.011, 0.032, or 0.110 mg/L. The estimated achieved dosages of
Frowncide( WP over the 7 days of treatment were calculated to be 0.72,
2.76, 7.93 and 27.43 mg/kg/day for males and 0.75, 2.97, 8.50 and 29.23
mg/kg/day for females for the concentrations of 0, 0.003, 0.011, 0.032,
and 0.110 mg/L, respectively.  The mass median aerodynamic diameter
(MMAD) was estimated to be 3.22-3.98 (m and the geometric standard
deviation was 2.04-2.69 (m.  Approximately 60-70% of particles had an
aerodynamic diameter < 6.0 (m.  The animals were observed daily. 
Hematology, clinical chemistries and urinalyses were performed.  All
animals were necropsied after completion of exposure, but no
histopathology was performed.

No rats died during the study.  No clinical signs of toxicity were noted
from any rat.  The body weight changes of all groups were similar to
that of the control group.  No toxicologically significant effects of
the test material were noted on food consumption, water consumption,
food efficiency, hematology, clinical chemistries, or urinalyses.  At
0.110 mg/L, slightly increased lung weights (males and females),
slightly increased testes weights (males), and slightly increased liver
weights (females) were observed.  At 0.032 mg/L, slightly increased
testes weights (males), and slightly increased liver weights (females)
were also observed.  No macroscopic changes attributed to treatment with
test material were noted at necropsy.  Histopathological examination of
tissues was not performed.  

The LOAEL is 0.032 mg/L (7.93 mg/kg/day in males and 8.50 mg/kg/day in
females), based on slightly increased testes weights (males) and
slightly increased liver weights (females).  The NOAEL is 0.011 mg/L
(2.76 mg/kg/day in males and 2.97 mg/kg/day in females).  

This inhalation study is classified as Acceptable/Nonguideline.  It does
not satisfy the subdivision F guideline requirements for a repeated dose
inhalation study in the rat because histopathological examination of
tissues was not performed.  The study was conducted as a range-finding
study (for a four-week inhalation study with Frowncide( WP in rats) and
is acceptable for that purpose.

Nonguideline:  A Review of 8 Special Mechanistic Studies Conducted to
Assess                                                  the CNS White
Matter Vacuolation Produced by Impurity-5 Present In                    
               Fluazinam Technical

Background:  A neurotoxic lesion described as vacuolation of the white
matter of the central nervous system (CNS) was observed initially in
long-term (1-2 year) guideline chronic studies on mice and dogs and
later, upon careful re-examination of the CNS, also in shorter-term
(4-week to 90-day) subchronic studies on mice and dogs.  This lesion was
observed during the (light) microscopic examination of several tissues
of the CNS, occurring most frequently in brain (sections of cerebrum
and/or sections of cerebellum, pons, medulla, and midbrain) and less
frequently in cervical spinal cord.  Although this lesion was also
observed in control animals, the increased incidence and/or severity of
the lesion in test animals was clearly treatment-related and
dose-related.  It is noteworthy that the lesion was not observed in any
guideline studies on rats, even though a careful re-evaluation of the
CNS was performed for all critical studies, including the major chronic
(MRID 42248620) and subchronic (MRID 42248610) studies.  Further
investigation of this lesion in a series of 8 additional special
mechanistic studies, however, demonstrated the lesion could also be
induced in rats at higher dose levels than used in the guideline
studies.  These 8 additional special mechanistic studies were designed
to determine, if possible, the etiology of the vacuolation of the white
matter of the CNS observed in the guideline studies on fluazinam and to
further evaluate several additional characteristics of the lesion.  

Summary:  Eight special studies (MRIDs 44807225, 44807226, 44807227,
44807228, 44807229, 44807230, 44807231, and 44807232)  conducted on the
etiology of an impurity in Fluazinam Technical which produced
vacuolation of the white matter in the CNS of mice, rats, and dogs have
revealed several important toxicological features.  In the mouse, rat,
and dog, vacuolation of the white matter in the brains (and optic nerves
of mice) was observed only when high doses of fluazinam technical were
administered.  Fluazinam technical itself was not responsible for the
induction of this aberration.  An analysis of the effects of nine
impurities present in Fluazinam Technical revealed that one impurity,
Impurity-5, is responsible for the appearance of white matter
vacuolation.  With respect to the ability of Impurity-5 to produce white
matter vacuolation, there seems to be a non-linear dose-response with a
clear threshold below which no effect occurs.  No significant
differences between species/sex susceptibility were observed.  White
matter vacuolation in the CNS was reversible, and no progression of this
abnormality was observed with time.  An age-related increased
sensitivity was identified in mice and rats at 10 weeks compared to 3
weeks of age.  Electron microscopy of the white matter (cerebellum) of
mice treated with fluazinam technical indicated that treatment-related
effects were confined to the myelin sheaths.  Large vacuoles were
observed in the intramyelin sheaths due to the accumulation of fluid
between the sheaths.  The nucleus and mitochondria in oligodendroglia
were observed to remain intact, suggesting no damage to these cells. The
myelin sheaths appeared to recover completely during a recovery period
of up to 56 days.  Macroscopic changes in the liver of animals treated
with fluazinam technical and the analytical standard of fluazinam
revealed that liver effects are due to fluazinam itself and not to the
presence of Impurity-5.      

Review: A brief synopsis of the results in the special mechanistic
studies is presented below.

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e for inducing the lesion in mice, dogs

             and rats

Fluazinam, per se, was not responsible for the induction of this lesion.
 An analysis of the effects of nine impurities present in fluazinam
technical revealed that one single impurity, Impurity-5, is solely
responsible for the appearance of white matter vacuolation.  Impurity-5
was present in the various lots of fluazinam technical used for toxicity
testing at concentrations ranging from <0.005% to 0.2% w/w.  With
respect to the ability of Impurity-5 to produce white matter
vacuolation, there seems to be a non-linear dose-response with a clear
threshold below which no effect occurs.	 

Single oral (gavage) doses of 5 mg/kg of Impurity-5 (99.5% purity) given
to fasted mice  caused course fur, staggering gait, sedation for 20
hours and decreased body weight.  All mice were sacrificed in extremis
at 24 hours.  Increased brain weights, edema of the brain and
vacuolation of the brain were observed in the treated animals. 

In rats, single oral (gavage) doses of 5000 mg/kg of the analytical
standard of fluazinam (containing <0.0005% Impurity-5)  caused no
vacuolation of the white matter of the brain whereas similar doses of
fluazinam technical did cause vacuolation of the white matter of the
brain.      

2.   Determination of dose-response relationships for fluazinam
technical

In the mouse and rat, vacuolation of the white matter in the brains (and
optic nerves of mice) was observed only when high doses of fluazinam
technical were administered.    

In mice, single oral (gavage) doses of 3000 mg/kg of fluazinam technical
(95.3% purity) caused decreased locomotor activity, prone position,
paralysis of hind legs, tremors,  staggering gait and moribundity. 
Edema of the brain and vacuolation of the white matter of the brain were
observed in the treated animals.  Assuming a concentration of 0.1 %
Impurity-5 in fluazinam technical, this dose (3000 mg/kg) is
approximately equal to a dose of 3 mg/kg of Impurity-5.    

In mice, administration of fluazinam technical (containing 0.12%
Impurity-5) in the diet for 4 days at 20,000 ppm resulted in abnormal
behavior on day 4 and trace edema of the brain.   In mice,
administration of fluazinam technical (containing 0.12% Impurity-5) in
the diet for 4 days at 7,000 ppm resulted in trace edema of the brain. 
In mice, administration of fluazinam technical (containing 0.12%
Impurity-5) in the diet for 28 days at 7,000 ppm resulted in vacuolation
of the white matter of the brain.  Vacuolation of white matter was
observed in the brains of all treated animals sacrificed at the end of
treatment.

In rats, single oral (gavage) doses of 5000 mg/kg of fluazinam technical
(containing 0.12% to 0.20% Impurity-5) caused decreased locomotor
activity, prone position, paralysis of hind legs, tremors, staggering
gait and moribundity.  Edema of the brain and vacuolation of the white
matter of the brain were observed in the treated animals.  Assuming a
concentration of 0.12 % Impurity-5 in fluazinam technical, this dose
(5000 mg/kg) is approximately equal to a dose of 6 mg/kg of Impurity-5. 
In rats, administration of fluazinam technical (containing 0.12%
Impurity-5) in the diet for 14 days at 30,000 ppm (1742 mg/kg/day) and
at 10,000 ppm (714 mg/kg/day) resulted in edema of the brain and minimal
to moderate vacuolation of the white matter of the brain at 30,000 ppm,
and trace vacuolation of the white matter of the brain at 10,000 ppm.

3.   Reversibility of the CNS lesion

White matter vacuolation in the CNS was  reversible, and no progression
of this abnormality was observed with time.  In the 14-day dietary study
in rats described above, recovery from the CNS lesion was also studied. 
Some rats were allowed to recover for an additional 25 days (no
fluazinam in the diet) and then examined.  For 30,000 ppm animals, only
trace vacuolation of the white matter of the brain was observed.  For
10,000 ppm animals, no vacuolation of the white matter of the brain was
observed.

In the mouse study described above in which fluazinam technical was
administered in the diet for 4 days at 20,000 ppm, or for 4 days at
7,000 ppm, or for 28 days at 7,000 ppm, vacuolation of white matter was
observed in the brains of all treated animals sacrificed at the end of
treatment.  This abnormality was not observed after a 24-day recovery
period among animals treated at 7,000 ppm for 4 days or after 56 days
among those treated at 7.000 ppm for 28 days or at 20,000 ppm for 4
days.

4.  Differences in species and sex susceptibility

In a series of studies, no significant differences in susceptibility or
in incidence or severity of vacuolation of the white matter of the CNS
were observed between species (mice, dogs, or rats).  Similarly, no
significant differences were attributed to sex.

5.   Differences in age-related susceptibility

An age-related increased sensitivity was identified in mice and rats at
10 weeks compared to 3 weeks of age.

Impurity-5 was administered to groups of male mice aged 3, 10, or 24
weeks by a single oral gavage at 2.5 mg/kg.  The severity of white
matter vacuolation increased with age until about ten weeks, then
plateaued at 24 weeks as observed under the limitations of this study. 

The difference in age sensitivity was comparable between rats and mice
as displayed in another oral gavage study using 3 and 10 week-old mice
and rats dosed with 0 or 0.5 mg/kg of Impurity-5.  Microscopic
observation of the brains of treated animals revealed white matter
vacuolation with slightly different severity between the respective age
groups.  The severity of these lesions was similar for both species of
the same age, but greater in 10 week old animals as compared to 3 week
old animals.  

	6.  Electron Microscopy of the CNS lesion

Electron microscopy of the white matter (cerebellum) of mice treated
with fluazinam technical indicated that treatment-related effects were
confined to the myelin sheaths.  Large vacuoles were observed in the
intramyelin sheaths due to the accumulation of fluid between the
sheaths.  The nucleus and mitochondria in oligodendroglia were observed
to remain intact, suggesting no damage to these cells. The myelin
sheaths appeared to recover completely during a recovery period of up to
56 days.

Comments on Threshold Dose for Vacuolation of the White Matter of the
CNS:  There appears to be a non-linear dose-response with a clear
threshold below which no effect occurs.  When the guideline data were
analyzed and presented graphically [see Figure 1 (page 100) in the
Overview Document (MRID 44807207) prepared by the applicant], it was
apparent that no white matter vacuolation occurred when the dose of
Impurity-5 was below about 0.1 mg/kg/day.  The lowest effect level for
white matter vacuolation was observed in the dog chronic study at 0.1
mg/kg/day of Impurity-5 (equivalent in that study to 50 mg/kg/day of
fluazinam technical containing 0.2% of Impurity-5).  

Based on a consideration of all the available data and information
relating to this treatment-related neurotoxic lesion, the HIARC
concluded that a LOAEL of 0.1 mg/kg/day and a NOAEL of 0.02 mg/kg/day
for CNS effects could be established for Impurity-5.  

NOAEL (for CNS effects)  =  0.02 mg/kg/day of Impurity-5

At the current maximum concentration of Impurity-5 in technical grade
fluazinam of 0.1% w/w [see memorandum from Indira Gairola, Technical
Review Branch, RD (7505C) to Cynthia Giles-Parker, Fungicide Branch, RD
(7505C), dated May 18, 2001, DP Barcode D272455], this is equivalent to:

     NOAEL (for CNS effects) = 20.0 mg/kg/day of technical grade
fluazinam

Calculation:   0.02 mg/kg/day   =   x  mg/kg/day

                   	0.1%		      	100%		          x  = 20.0

  

This NOAEL (for CNS effects) of 20.0 mg/kg/day for technical grade
fluazinam is to be compared to:

NOAEL (for chronic effects for chronic RfD)  =  1.1 mg/kg/day of
technical grade fluazinam

The chronic RfD of 0.011 mg/kg/day for (all populations(, including
infants and children, is therefore protective of the CNS effects caused
by Impurity-5 present in technical grade fluazinam at levels up to 0.1%
w/w.  

A.4	Additional Toxicology Study

Fulcher, S. (2005) Technical Fluazinam: Developmental Neurotoxicity
Study in the Rat by Oral (Gavage) Administration. Project Number:
ISK/272/042019. Unpublished study prepared by Huntingdon Life Sciences,
Ltd. 1471 p.

Appendix B:  Metabolism Assessment

B.1. Metabolism Guidance and Considerations

 TC \l1 "Appendix C:  Tolerance Reassessment Summary and Table 

Chemical Name (other names in parenthesis)	Matrix	

Structure

Parent

Fluazinam	Primary crop, Ruminant, Rat, and Drinking water	

*AMGT	Primary crop

(AMGT data only available for blueberries)	Not available

AMPA 	Ruminant, Rat (+ AMPA hydrolysis products)	Not available

DAPA	Ruminant, Rat (+ DAPA hydrolysis,conjugation products), Drinking
water	

CAPA

	Drinking water	

DCPA	Drinking water	

Appendix C:  Tolerance Reassessment Summary and Table

Table C.1	Tolerance Summary for Fluazinam.

Commodity	Proposed Tolerance (ppm)	Recommended Tolerance (ppm)	Comments

[Correct Commodity Definition]

Ginseng	3.00	4.5

	Bean, dry	0.01	0.02	[Pea and bean, dried shelled, except soybean,
subgroup 6-C (except peas)]

Succulent-shelled legume vegetables subgroup 6B, except pea	0.02	0.04
[Pea and bean, succulent shelled, subgroup 6-B (except peas)]

Edible-podded legume vegetables subgroup 6A, except peas	0.15	0.10
[Vegetable, legume, edible-podded, subgroup 6-A (except peas)]

Leafy Brassica greens subgroup	0.02	0.01	Crop group tolerance is
appropriate. [Vegetable, Brassica leafy, group 5]

Head and stem Brassica subgroup	0.01

Turnip, leaves	0.02	0.01	[Turnip, tops]

Bushberry subgroup 13B	4.5	7.0	[Bushberry subgroup 13-B]

Aronia berry	4.5	7.0

	Blueberry, lowbush	4.5	Not needed	Low bush blueberry is a member of the
bushberries  subgroup 13-B

Buffalo currant	4.5	7.0

	Chilean guava	4.5	7.0

	European barberry	4.5	7.0

	Highbush cranberry	4.5	7.0

	Honeysuckle	4.5	7.0

	Jostaberry	4.5	7.0

	Juneberry	4.5	7.0

	Lingonberry	4.5	7.0

	Native currant	4.5	7.0

	Salal	4.5	7.0

	Sea Buckthorn	4.5	7.0

	

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