Document ID: EPA-HQ-OPP-2008-0270-0004
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2009-05-26T04:00Z

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460      

	OFFICE OF PREVENTION, PESTICIDE

	AND TOXIC SUBSTANCES

MEMORANDUM			

March 3, 2009									

SUBJECT:	Revised Acibenzolar-S-methyl Human Health Risk Assessment for
Proposed Use of Acibenzolar-S-methyl on Cucurbits and Bulb Onions

PC Code:  061402	DP Barcode: 361893

Decision No.:  393110	Registration No.: 100-922

Petition No.:  PP#8F7352, PP8E7337	Regulatory Action: Section 3
Registration 

Risk Assessment Type:  Single Chemical Aggregate	Case No.:  NA

TXR No.:  NA	CAS No.:  135158-54-2

MRID No.:  NA	40 CFR:  180.561

FROM:	Becky Daiss, Biologist 

James Scott Miller, Environmental Scientist

Bonnie Cropp-Kohlligian, Environmental Scientist

Gerome Burke, Toxicologist

Risk Assessment Branch 4

Health Effects Division (7509P)

THROUGH:	Susan V. Hummel, Branch Senior Scientist

Risk Assessment Branch 4, HED (7509P)

TO:		Tony Kish, Product Manager

		Fungicide Branch

		Susan Stanton, Product Manager

		RIMUER Branch

Registration Division (7505P)

	This document provides the revised Health Effects Division’s
(HED’s) risk assessment of use of acibenzolar-S-methyl on cucurbits
and bulb onion.  Supporting documents are listed below.

Occupational Exposure Assessment – J.S. Miller, D361894, 2/18/2009

Residue Chemistry Assessment – B. Cropp-Kohlligian, D352415, 1/26/09

Dietary and Drinking Water Exposure Assessment – B. Daiss, D362208
3/3/2009

Drinking Water Exposure Assessment – R. Miller, D351609, 8/12/08



TABLE OF CONTENTS				

 	 											      pg.

1.0 	EXECUTIVE SUMMARY	4

2.0	INGREDIENT PROFILE	7

2.1	Registered Products	7

2.2	Registered and Proposed New Uses	8

2.3  	Structure, Nomenclature and Physical/Chemical Properties	8

3.0	HAZARD CHARACTERIZATION/ASSESSMENT 	9

3.1	Hazard Characterization 	9

3.2	FQPA Hazard Considerations 	11

3.2.1	Developmental Toxicity  tc \l3 "4.2.3	Developmental Toxicity
Studies  	11

3.2.2	Reproductive Toxicity 	13

3.2.3  	Developmental Neurotoxicity	14

3.2.4	Pre-and/or Postnatal Toxicity tc \l3 "4.2.5  	Pre-and/or Postnatal
Toxicity 	15

	3.2.5    Immunotoxicity	15

	3.2.6    FQPA Factor	15

3.3	Toxicity Endpoint Selection	16

3.3.1   	Acute Reference Dose – Females Age 13-49 tc \l3 "4.4.1   
Acute Reference Dose - General Population 	16

3.3.2   	Acute Reference Dose – General Population tc \l3 "4.4.1   
Acute Reference Dose - General Population 	16

3.3.3	 tc \l3 "4.4.2	 Chronic Reference Dose – Females Age 13-49	16

3.3.4	 tc \l3 "4.4.3	 Chronic Reference Dose – All Populations	17

3.3.5	 tc \l3 "4.4.3	 Incidental Oral Exposure	19

3.3.6	 tc \l3 "4.4.4	 Dermal Absorption	19

3.3.7	 tc \l3 "4.4.5	 Dermal Exposure	19

3.3.8	 tc \l3 "4.4.6	 Inhalation Exposure	20

3.3.9	 tc \l3 "4.4.6	 Classification of Carcinogenic Potential	20

3.4	 tc \l3 "4.4.7	 Margins of Exposure	20

3.5	Recommendation for Aggregate Exposure	21

3.6	Summary of Endpoints Selected for Risk Assessment	21

3.7	Endocrine Disruption	22

4.0	DIETARY AND DRINKING WATER EXPOSURE ASSESSMENT	23

4.1	Residue Chemistry Profile	23

4.1.1 	Metabolism in Primary Crops	23

4.1.2 	Residue Analytical Method	23

4.1.3 	Residues in Crops	24

4.2 	Drinking Water Profile	25

4.2.1 	Environmental Fate and Transport	25

4.2.2 	Estimated Drinking Water Concentrations	25

4.3	Dietary and Drinking Water Exposure and Risk	26

4.3.1 	Acute Dietary and Drinking Water Analysis	26

4.3.2 	Chronic Dietary and Drinking Water Analysis	27

5.0 	AGGREGATE EXPOSURE AND RISK ASSESSMENT	27

6.0	OCCUPATIONAL EXPOSURE AND RISK	27

6.1 	Exposure Scenarios	27

6.1.1 	Handler Exposure Scenarios	27

6.1.2 	Post Application Exposure Scenarios	28

6.2	Occupational Exposure Data and Assumptions	28

6.2.1 	Exposure Data	28

	6.2.2    Exposure Assumptions	29

6.3	Occupational Exposure and Risk Estimates 	30

	6.3.1	Handler Exposure and Risk Estimates	30

	6.3.2	Post-Application Exposure and Risk Estimates	30

7.0	CUMULATIVE RISK	31

8.0	DATA NEEDS	31

8.1	Toxicity Data Requirements	31

8.2	Residue Chemistry Data Requirements	32

APPENDICES

1.0	GUIDELINE TOXICOLOGY DATA SUMMARY	33

	1.1  	Guideline Data Requirements	33

	1.2  	Toxicity Profiles	34

2.0	STUDIES REVIEWED FOR ETHICAL CONDUCT	39

3.0	ADDITIONAL TOXICITY STUDIES	40

4.0	REFERENCES FOR TOXICITY STUDIES	46

5.0	CHEMICAL NAMES AND STRUCTURES OF METABOLITES	49

	

1.0 	EXECUTIVE SUMMARY

This assessment provides information to support an amended Section 3
registration for the use of acibenzolar-S-methyl on cucurbit vegetables
and bulb onions.  This document addresses the exposures and risks
associated with exposures from currently registered uses and the
proposed new uses of acibenzolar-S-methyl.    

Use Profile

	  SEQ CHAPTER \h \r 1 Acibenzolar-S-methyl is a plant activator.  It is
a member of the benzothiadiazole class of compounds. 
Acibenzolar-S-methyl is registered to Syngenta Crop Protection, Inc for
use on leafy vegetables, Brassica leafy vegetables, fruiting vegetables,
and tobacco for the control or suppression of downy mildew, black rot,
white rust, bacterial leaf spot and speck, and blue mold. Permanent
tolerances ranging from 0.1-3.0 ppm are established under §180.561(a). 
Section 18 time-limited tolerances are established under §180.561(b)
for bulb and green onion (0.05 ppm).  Acibenzolar-S-methyl acts to
protect the plant against invading pathogen by inducing a natural
defense reaction in plants that enhances the plants ability to ward off
disease.  Applications may be made using ground or aerial equipment. 
There are no acibenzolar-S-methyl products registered for homeowner use
and no products registered for application to residential areas.  

   

Proposed New Uses

	The Interregional Research Project No. 4 (IR-4) and Syngenta Crop
Protection, Inc. are proposing new uses of Actigard® (EPA Reg. No.
100-922).  Actigard® is a water-dispersible granule (WG) formulation
containing 50% acibenzolar-S-methyl.  

	IR-4 proposes new uses of Actigard® on the bulb onion crop subgroup
3-07A.   The IR-4 petition proposes multiple foliar applications on the
onion subgroup using ground or aerial equipment at a maximum of 0.03 lb
ai/A per application, a maximum seasonal rate of 0.125 lb ai/A, and a
7-day preharvest interval (PHI).

	Syngenta proposes new uses of Actigard® on the cucurbit vegetable crop
group 9.  Actigard® is proposed for multiple foliar applications on
members of the cucurbit crop group using ground or aerial equipment at a
maximum of 0.06 lb ai/A per application, a maximum seasonal rate of 0.5
lb ai/A, and a 0-day PHI.  

Hazard Identification

The toxicology database for acibenzolar-S-methyl is adequate for
evaluating and characterizing toxicity and selecting endpoints for
purposes of this risk assessment.  

	Acibenzolar-S-methyl showed no significant toxicity in a battery of
acute toxicity tests (Toxicity Category III or IV in all tests) but
showed considerable skin-sensitivity.  The end-use product (ActigardTM
50Wg) displayed no significant acute toxicity and no dermal sensitizing
potential.  In subchronic and chronic oral studies in rats, dogs and
mice, signs of mild regenerative hemolytic anemia were consistently
observed in all three species.  Additional toxic effects observed in
these studies included decreases in body weight, body weight gain and/or
food consumption.  No other significant treatment-related effects of
toxicological concern were observed in these subchronic and chronic oral
studies.  No neurotoxic effects were seen at the highest dose tested in
a subchronic neurotoxicity study in rats.  In a 28 day dermal toxicity
study in rats no systemic or dermal effects were seen at the limit dose.
 

		In a developmental toxicity and developmental neurotoxicity studies in
rats, treatment related effects were observed in fetuses at levels that
were not toxic to the parent indicating increased sensitivity of rat
fetuses compared to adults.  Increased sensitivity was not observed in a
developmental toxicity study in rabbits, or in 1- and 2-generation
reproduction studies in rats.  In a 28 day dermal developmental toxicity
study in rats, no maternal or developmental toxicity was observed at
dose levels up to 500 mg/kg/day, the highest dose level tested.  

	Acibenzolar-S-methyl was classified by HED’s Hazard Identification
Assessment Review Committee (HIARC) as a “not likely” human
carcinogen based on the lack of evidence of carcinogenicity in male and
female rats and mice and lack of evidence of genotoxicity in an
acceptable battery of mutagenicity studies.  

An immunotoxicity study is required as part of new 40 CFR Part 158 the
data requirements for registration of a pesticide.  A special study is
required to generate specific data on the thyroid to protect the
developing nervous system from thyroid hormone disrupting chemicals.  A
battery of mutagenicity studies performed on technical-grade
acibenzolar-S-methyl prepared by the thiozole production process and an
Ames assay using the technical-grade acibenzolar-S-methyl prepared by
the thiozole production process are required.

Dose Response Assessment

Toxicological endpoints were selected for dietary/drinking water and
occupational exposure scenarios.  Acute and chronic reference doses
(RfDs) were selected for assessment of food and drinking water
exposures.  An acute RfD for the general population was not selected
because no effect attributable to a single day oral exposure was
observed in animal studies.  Acute and chronic RfD’s for females 13-49
were selected from a developmental neurotoxicity toxicity study in rats.
 A chronic RfD for the general population was selected from a chronic
feeding study in dogs with chronic studies in other species co-critical.
 Short and intermediate-term occupational exposures via the dermal and
inhalation routes may occur based on the use pattern and label
directions.  Toxicological endpoints for inhalation and dermal exposures
were selected from the developmental neurotoxicity study in rats.  An
uncertainty factor of 100X was applied to endpoints selected for all
exposure routes (10x for interspecies extrapolation, 10x for
intraspecies variation).   [Note: the toxicity endpoints and uncertainty
factors for acibenzolar-S-methyl have been revised from previous
assessments based on reevaluation of existing data and review of new
data.]

Exposure/Risk Assessment and Risk Characterization

	Risk assessments were conducted for dietary (food and water) and
occupational exposure pathways based on registered uses and requests for
new uses of acibenzolar-S-methyl on cucurbit vegetable and bulb onions. 
Worker exposures were assessed for handler and post-application
activities.  Refined acute and screening level chronic dietary and
drinking water risk assessments for acibenzolar-S-methyl conclude that
dietary and drinking water exposure estimates are below HED’s level of
concern the general population and all population subgroups.
Occupational exposure and risk estimates indicate that worker handler
and post-application exposures are not of concern at the maximum
allowable application rates for the proposed new uses.

Use of Human Studies

	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, listed in Appendix 2.0, have been determined
to require a review of their ethical conduct.  Some of these studies are
also subject to review by the Human Studies Review Board.  All of the
studies used have received the appropriate review.

Environmental Justice

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.  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.  

Tolerance Recommendation

	HED continues to recommend that the appropriate tolerance expression
for acibenzolar-S-methyl should be for “residues of
acibenzolar-S-methyl, benzo(1,2,3)thiadiazole-7-carbothioic
acid-S-methyl ester, convertible to benzo(1,2,3)thiadiazole-7-carboxylic
acid (CGA-210007), expressed as acibenzolar-S-methyl.”

Pending submission of confirmatory crop field trial data and/or a
revised Section B, the submission of a reference standard for the
metabolite CGA 210007, and a revised Section F, there are no residue
chemistry issues that would preclude granting a Section 3 registration
for the requested use of acibenzolar-S-methyl on crop subgroup 3-07A or
the establishment of tolerances for residues of acibenzolar-S-methyl as
follows:

Onion, bulb, subgroup 3-07A 	0.1 ppm

Pending submission of a revised Section B, the submission of a reference
standard for the metabolite CGA 210007, and a revised Section F, there
are no residue chemistry issues that would preclude granting a
time-limited acceptance of the proposed use of acibenzolar-S-methyl on
crop group 9, or the establishment of a tolerance for residues of
acibenzolar-S-methyl, as follows:

Vegetable, cucurbit, group 9 	2.0 ppm

	Pending receipt of required additional field trial/decline studies, the
conversion of the time-limited acceptance of the proposed use of
acibenzolar-S-methyl on cucurbit vegetables group 9 to an unconditional
registration may be considered.  An amended tolerance level may be
deemed appropriate once the required field trial/decline data are
submitted.

2.0	INGREDIENT PROFILE 

2.1	Registered Products	

	There are 7 active acibenzolar-S-methyl registrations, two Section 3
and five special local needs (SLNs) or 24(c) registrations. 

Table 1.  Summary Report of Supported Registered Acibenzolar-S-methyl 
Products

Reg #	Name	Company Name	%AI

100-921	Acibenzolar-S-Methyl Technical	Syngenta Crop Protection, Inc.	99

100-922	Actigard 50WG

50

FL030003	Actigard 50 WG Plant Activator

50

GA020005	Actigard 50 WG Plant Activator

50

NC020007	Actigard 50 WG Plant Activator

50

SC030004	Actigard 50 WG Plant Activator

50

VA030002	Actigard 50 WG Plant Activator

50

2.2		Registered and Proposed New Uses 

	Acibenzolar-S-methyl is a “plant activator” that protects the plant
against invading pathogens by stimulating the plant’s natural defense
mechanisms.  The single registered formulation, Actigard® (EPA Reg. No.
100-922), is a water-dispersible granule (WG) containing 50%
acibenzolar-S-methyl.  Actigard® is registered for use on a variety of
vegetable crops and tobacco for the control of downy mildew, black rot,
white rust, bacterial leaf spot and speck, and blue mold.  Two new uses
of Actigard® have been proposed.  IR-4 is proposing to amend the label
to add new uses on the bulb onion crop subgroup 3-07A.  Under the IR-4
petition, Actigard® is proposed for multiple foliar applications on the
onion subgroup using ground or aerial equipment at a maximum of 0.03 lb
ai/A per application, a maximum seasonal rate of 0.125 lb ai/A, and a
7-day preharvest interval (PHI).  Syngenta Crop Protection, Inc.
proposes a label amendment to add new uses on the cucurbit vegetable
crop group 9.  Under Syngenta’s petition, Actigard® is proposed for
multiple foliar applications on members of the cucurbit crop group using
ground or aerial equipment at a maximum of 0.06 lb ai/A per application,
a maximum seasonal rate of 0.5 lb ai/A, and a 0-day PHI.    Maximum
application rates for existing and proposed uses are provided in Table
2.  Maximum rates are based on a review of active and proposed labels.  

Table 2.  Maximum Application Rates for Acibenzolar-S-methyl Existing
and Proposed Uses

Application Site	Max % AI	Maximum AR	Reg No.

Leafy vegetables, Brassica leafy vegetables, Fruiting Vegetables,
Tobacco and Bulb Onion subgroup 3-07A	50	0.03 lb/ai A	100-922

Cucurbit vegetable crop group 9 

0.06 lb/ai A

	

2.3  	Structure, Nomenclature, and Physical/Chemical Properties   

	The nomenclature and physicochemical properties of acibenzolar-S-methyl
are provided in Tables 4 and 5. 

Common name	acibenzolar-S-methyl

Company experimental name	CGA-245704

IUPAC name	benzo[1,2,3]thiadiazole-7-carbothioic acid-S-methyl ester

CAS name	S-methyl, 1,2,3-bensothiadiazole-7-carbothioate

CAS registry number	135158-54-2

End-use product (EP)	Actigard® 50 WG (EPA Reg. No. 100-922)

Structure of metabolite 

CGA-210007  SEQ CHAPTER \h \r 1 	

1,2,3-benzothiadiazole-7-carboxylic acid

Table 5.	Physicochemical Properties of Acibenzolar-S-Methyl.

Parameter	Value	Reference

Melting point/range	133 ºC	DP# 250963, 2/10/99, H. Podall

pH	  SEQ CHAPTER \h \r 1 7.9 (1% solution in water) at 25 ºC

	Density	  SEQ CHAPTER \h \r 1 1.54 g/cc at 22 ºC (bulk density)

	Water solubility	0.0077 g/L

	Solvent solubility	Methanol	  4.2 g/L

Acetone		  28 g/L

Toluene		  36 g/L

n-Octanol	  5.4 g/L

n-Hexane	  1.3 g/L

Ethyl acetate	   25 g/L

Dichloromethane   160 g/L 	European Commission, 6506/VI/99-final,
5/21/02

Vapor pressure	3.6 x 10-6 Torr at 25 ºC	DP# 250963, 2/10/99, H. Podall

Dissociation constant, pKa	No dissociation constant in an

253 nm ε : 14.7 103 l mol-1 cm-1

288 nm ε : 5.4 103

324 nm ε : 7.2 103	European Commission, 6506/VI/99-final, 5/21/02

3.0	HAZARD CHARACTERIZATION/ASSESSMENT

3.1	Hazard Characterization

The toxicology database for acibenzolar-S-methyl is adequate for
evaluating and characterizing acibenzolar-S-methyl toxicity and
selecting endpoints for purposes of this risk assessment.  An
immunotoxicity study is required as part of new 40 CFR Part 158 data
requirements for registration of a pesticide.  An Ames assay and a
battery of mutagenicity studies performed on technical-grade
acibenzolar-S-methyl prepared by the thiozole production process are
required.

	Acibenzolar-S-methyl showed no significant toxicity in a battery of
acute toxicity tests (Toxicity Category III or IV in all tests). 
Considerable skin sensitizing (contact allergenic) potential was
demonstrated in a dermal sensitization study in guinea pigs.  The
formulated end-use product (Actigard™ 50 WG) also demonstrated no
significant acute toxicity; additionally, the end-use product did not
show dermal sensitization in guinea pigs.

	In subchronic and chronic oral studies in rats, dogs and mice, signs of
mild regenerative hemolytic anemia were consistently observed in all
three species.  These signs frequently included decreased erythrocyte
counts, decreased hemoglobin, decreased hematocrit, increased
reticulocyte counts, increased hemosiderosis in the spleen, liver and/or
bone marrow, extramedullary hematopoiesis in the spleen, and increased
spleen weights in both males and females.  A compensatory response
(increased erythrocyte production) regularly followed the initial
anemia.  Additional toxic effects observed in these same studies
included decreases in body weight, body weight gain and/or food
consumption.  No other significant treatment-related effects of
toxicological concern were observed in these subchronic and chronic oral
studies.  In a 28-day dermal study in rats, no systemic or dermal
effects were observed at dose levels up to 1000 mg/kg/day, the limit
dose.  No neurological effects were observed at any dose in a subchronic
neurotoxicity study in rats.

		Treatment-related developmental malformations, anomalies and
variations were observed in a developmental toxicity study in rats at or
below the no observable adverse effect level (NOAEL) for maternal
toxicity.  At the highest dose tested in this study (400 mg/kg/day),
both maternal toxicity (hemorrhagic perineal discharge) and considerable
developmental toxicity (including total litter resorptions, fetal
malformations, anomalies and variations) were observed.  The fetal
malformations noted at this dose included treatment-related effects on
nervous system tissues (hydrocephaly, craniorachisis and anophthalmia/
microphthalmia).  At the next lower dose tested (200 mg/kg/day),
treatment-related visceral malformations and skeletal variations were
demonstrated in the absence of significant maternal toxicity.  A similar
increased sensitivity of fetuses or pups (as compared to adults) was not
observed in a developmental toxicity study in rabbits or in 2-generation
and 1-generation (range-finding) studies in rats.  In a dermal
developmental toxicity study in rats, no maternal or developmental
toxicity was observed at dose levels up to 500 mg/kg/day, the highest
dose tested.  

		In a metabolism study in rats, acibenzolar-S-methyl was rapidly and
nearly completely absorbed and excreted in less than 48 hours.  The
major route of excretion was urine (88-95%).  The major metabolite in
urine was the carboxylic acid derivative of parent (79-92%).  

In a battery of mutagenicity studies, results were negative in all
studies except in an in vitro chromosome aberration study in Chinese
hamster ovary (CHO) cells, in which there was evidence of a clastogenic
response in the absence of S-9 activation.  This finding, however, can
only be considered to be suggestive evidence of a possible aneuploidy
effect since negative results were observed in an in vivo mouse
micronucleus assay.  In a S. typhimurium reverse gene mutation assay
performed on a structural isomer (CGA- 362020) of acibenzolar-S-methyl,
positive results were observed in strain TA1537 in the absence of S-9
activation.  This isomer is found at low levels (< 0.1%) in some batches
of technical-grade acibenzolar-S-methyl manufactured by a new
“thiazole process.”   Therefore, an Ames assay and a battery of
mutagenicity studies performed on technical-grade acibenzolar-S-methyl
prepared by that process are required.

		In a 2-year chronic toxicity/carcinogenicity study in rats and an
18-month carcinogenicity study in mice, acibenzolar-S-methyl was
negative for carcinogenicity when administered at dose levels adequate
for the testing of carcinogenic potential.   

The complete toxicity profile for acibenzolar-S-methyl is provided in
Appendix 1.2, Tables 2 and 3. 

3.2	FQPA Hazard Considerations tc \l2 "4.2	FQPA Hazard Considerations 

The toxicity database for acibenzolar-S-methyl is sufficient for a full
hazard evaluation and is considered adequate to evaluate risks to
infants and children.  Acceptable developmental toxicity studies in the
rat and rabbit, acceptable multi- and single generation reproduction
studies in the rat, and an acceptable developmental neurotoxicity study
in the rat are available.  An acceptable subchronic neurotoxicity study
in the rat is also available in the database.  An acute neurotoxicity
study is not available but not required at this time due to the
availability of the subchronic neurotoxicity study in which no
neurological effects were observed at any dose.  

3.2.1    Developmental Toxicity 

Rat Developmental Study  tc \l3 "4.2.3	Developmental Toxicity Studies 

Study 1 

In a developmental toxicity study (MRID 44014236), 24 pregnant Tif: RAI
f (Sprague-Dawley derived) rats per group were administered
acibenzolar-S-methyl by gavage in 0.5% (W/W) aqueous sodium
carboxymethyl-cellulose (vehicle) at daily doses of 0, 10, 50, 200, or
400 mg/kg/day on gestation days (GD) 6-15, inclusive. Controls were
treated only with 0.5% sodium carboxymethyl-cellulose. On GD 21, all
dams were sacrificed and subjected to gross necropsy and fetuses were
examined for external, visceral, and skeletal malformations and
variations.  All animals survived to scheduled sacrifice.  

Treatment-related hemorrhagic perineal discharge was observed in the 400
mg/kg/day group. No other treatment-related clinical signs of toxicity
were observed in any animal.  Interpretations of changes in body weight,
body weight gain and food consumption for the 400 mg/kg/day animals were
severely compromised because the data were based on only 6 animals (3 of
which had 50% resorptions compared to the number of implantation sites).
The maternal toxicity LOAEL is 400 mg/kg/day based on clinical signs of
hemorrhagic perineal discharge. The maternal toxicity NOAEL is 200
mg/kg/day.  

There were no differences between the control group and any treatment
group for number of animals pregnant, mean number of corpora lutea per
dam, or mean number of implantation sites per dam.  At 400 mg/kg/day
(highest dose tested), treatment-related complete litter resorptions,
increased postimplantation loss, decreased mean live fetuses/litter,
decreased mean fetal body weights, and several developmental
malformations, anomalies, and skeletal variations were observed. At 200
mg/kg/day, treatment-related developmental malformations and skeletal
variations were also observed.  The developmental toxicity LOAEL is 200
mg/kg/day based on developmental malformations, anomalies, and skeletal
variations. The developmental toxicity NOAEL is 50 mg/kg/day.  

[Note: the developmental LOAEL and NOAEL for this study have been
revised based on reevaluation of the existing data and receipt of new
data.  In previous assessments, the developmental LOAEL was 10 mg/kg/day
(the lowest dose tested) based on umbilical hernia observed in 3/327
fetuses from 2/23 litters at this dose and there was no NOAEL.  After
reevaluation of the developmental study, HED concluded that the
incidence of umbilical hernias at 10 mg/kg/day was not a
treatment-related adverse effect because the effect is not dose-related
(i.e., it was seen only at the low dose of 10 mg/kg/day), the effect was
not seen in dosed animals in other studies including developmental
toxicity studies and reproduction studies, umbilical hernia was observed
in the controls in the rat dermal developmental toxicity study (1/336
fetuses in 1 of 24 litters) and the effect is known to occur
spontaneously in the rat strain used in this study. The new studies,
including a developmental neurotoxicity study in rats (MRID 46046401), a
developmental toxicity study in rats (MRID 45089701) and two
non-standard investigative, phase specific studies (MRID 47430505),
support the finding that incidence of umbilical hernias are not
treatment related.] 

		Study 2

In a developmental toxicity study (MRID 45089701), acibenzolar-S-methyl
in aqueous 0.5% carboxymethylcellulose was administered via daily oral
gavage in a dose volume of 10 mL/kg to 22 presumed pregnant
Sprague-Dawley rats at doses of 0, 10, 75, 150, or 350 mg/kg/day from
gestation days (GD) 6-15. On GD 20, all dams were euthanized; each
dam’s uterus was removed via cesarean section and its contents
examined.  Fetuses were examined for external, visceral, and skeletal
malformations and variations.

All dams survived until scheduled termination, and there were no
clinical signs of toxicity during the study or gross lesions at
necropsy.  Furthermore, there were no effects of treatment on maternal
body weights, body weight gains, or food consumption.  A maternal LOAEL
was not observed.  The maternal NOAEL is 350 mg/kg/day.

There were no abortions, premature deliveries, or complete litter
resorptions.  There were no effects of treatment on the numbers of
litters, live fetuses, early resorptions, or late resorptions, resulting
in post-implantation loss values in the treated groups that were
comparable to controls.  There were no treatment-related effects on
placental weights, fetal body weights, or skeletal ossification. The
number of fetuses with skeletal variations was significantly increased
(p<0.05) at 350 mg/kg/day (60) compared to controls (18).  This increase
was accounted for by increased (p<0.05) incidences of fetuses with
rudimentary lumbar ribs (31 fetuses compared to 9 controls) and long
lumbar ribs (24 fetuses vs 0 controls).  There were no other
treatment-related external, visceral, or skeletal variations.  There
were no treatment-related external, visceral, or skeletal malformations.
 The developmental LOAEL is 350 mg/kg/day based on increased incidences
of rudimentary and long lumbar ribs.  The developmental NOAEL is 150
mg/kg/day.

	3.2.1.2 Rabbit Developmental Study 

In a developmental toxicity study (MRID 44014237), 17 pregnant Russian
Chbb:HM rabbits per group were administered acibenzolar-S-methyl by
gavage in an aqueous solution of carboxymethylcellulose (0.5% w/w) at
doses of 0 (vehicle), 10, 50, 300, and 600 mg/kg/day on gestation days
(GD) 7-19, inclusive.  On GD 29, all surviving does were sacrificed and
all fetuses were weighed and examined for external
malformations/variations and visceral abnormalities/ variations.  

Three 600 mg/kg/day does were found dead and three more were sacrificed
moribund during GD 13-28.  All exhibited moderate to extreme body weight
loss and reduced food consumption, two had perineal bleeding, two had
diarrhea, and one had reduced motor activity.  At 300 mg/kg/day, one doe
was found dead on GD 18.  This animal showed body weight loss and
reduced food consumption prior to death.  Gross necropsy findings in
animals that died intercurrently included stomach hemorrhage in three of
the 600 mg/kg/day does and in the 300 mg/kg/day doe.  No mortality or
treatment-related clinical signs of toxicity were observed for the 10
and 50 mg/kg/day treatment groups.  There were no significant
differences in maternal body weights, body weight changes, and food
consumption in the surviving animals from the treated and control groups
at any time during gestation.   Therefore, the maternal toxicity LOAEL
is 300 mg/kg/day based on mortality, clinical signs of toxicity, and
substantially decreased body weights and food consumption. The maternal
toxicity NOAEL is 50 mg/kg/day.

No treatment-related effects were observed for gravid uterine weights,
number of fetuses/litter, pre- and postimplantation loss, numbers of
corpora lutea/doe, number of implantations/doe, resorptions/doe, fetal
body weights, or fetal sex ratios.  At 600 mg/kg/day there was a slight
but significant increase in asymmetrically shaped caudal vertebral
centers (3 fetuses in 3 litters compared to 0 fetuses or affected
litters in the control group).  No statistically significant differences
in the incidence rates of any external or visceral
malformations/anomalies/ variations were observed in the treated litters
as compared to controls.  Therefore, the developmental toxicity LOAEL is
600 mg/kg/day based on a marginal increase in vertebral anomalies.  The
developmental NOAEL is 300 mg/kg/day.

	3.2.2	Reproductive Toxicity  tc \l3 "4.2.4	Reproductive Toxicity Study 

In a 2-generation reproduction toxicity study, acibenzolar-S-methyl was
administered to groups of 30 male and 30 female Tif: RAT f(SPF), hybrids
of RII/1 x RII/2 (Sprague-Dawley derived) rats at dietary concentrations
of 0, 20, 200, 2000, or 4000 ppm for two generations (MRID 44014241). 
Two litters were produced in the first generation and one litter was
produced in the second generation.  F1 adults were chosen from the F1a
pups and weaned onto the same diet as their parents.  Animals were given
test or control diet for 10 weeks before mating within the same dose
group. All animals were continuously exposed to test material either in
the diet or during gestation and lactation until sacrifice. 

No treatment-related clinical signs of toxicity were observed in males
or females of either generation at any time during the study.  Lower
body weights and food consumption for high dose dams was observed during
lactation.  No dose- or treatment-related effects on body weights, body
weight gains, or food consumption were observed for the F1 males or
females in the 20, 200, or 2000 ppm groups. Absolute body weights were
significantly reduced in high-dose F1 males throughout the study and in
high-dose F1 females through study day 57.  No significant differences
in body weights and food consumption for the F0 and F1 dams were
observed during gestation.  Absolute spleen weights were significantly
increased in both the 4000 and 2000 ppm F0 and F1 animals.  Relative
spleen weights were significantly increased in the 2000 and 4000 ppm F0
and F1 males and females.  High-dose F0 males also had significantly
increased absolute and relative kidney weights.  A dose-related increase
in the severity of splenic hemosiderosis was observed in F0 and F1 males
and females. This finding was associated with an increase in the
incidence of splenic congestion in the high-dose groups.  The LOAEL for
parental toxicity is 2000 ppm based on increased spleen weights and
hemosiderosis of the spleen in the F0 and F1 males and females. The
parental toxicity NOAEL is 200 ppm.  Mating, fertility, gestation, and
parturition indices, mean precoital interval, and mean gestation length
were similar between the treated and control groups of both generations.
 Mean litter size, live births, and pup survival were also similar
between the treated and control groups for both generations.  Therefore,
the NOAEL for reproductive toxicity is 4000 ppm (highest dose tested). 
A LOAEL for reproductive toxicity was not observed (>4000 ppm). 

No treatment-related clinical signs of toxicity were observed in the
F1a, F1b, or F2 pups during lactation.  Body weights of the 4000 ppm
group F1a, F1b, and F2 pups were significantly less than the controls on
lactation days 4,7, 14, and 21.  Body weights of the F1a, F1b, and F2
pups in the 2000 ppm group were significantly less than their respective
controls on lactation day 21.  Body weight gains by the high-dose F1a
and F2 pups throughout lactation and by the high-dose F1b pups beginning
on lactation day 4 were significantly less than the controls.  The LOAEL
for developmental toxicity is 2000 ppm based on reduced pup body weight
gains and lower pup body weights during lactation.  The developmental
toxicity NOAEL is 200 ppm.

 tc \l4 "4.2.6.2	Degree of Concern Analysis and Residual Uncertainties
for Pre and/or Post-natal Susceptibility 3.2.3	Developmental
Neurotoxicity Study 

		In a developmental neurotoxicity study (MRID 46046401)
acibenzolar-S-methyl was administered in the diet to pregnant Wistar
rats (30/dose) from gestation day (GD) 7 to lactation day (LD) 22 at
nominal doses of 0, 100, 1000, or 4000 ppm (equivalent to 0/0, 8.2/15.5,
82.0/153.6, and 326.2/607.8 mg/kg/day [gestation/lactation]).  Dams were
allowed to deliver naturally and were killed on LD 29.  On postnatal day
(PND) 5, litters were standardized to 8 pups/litter; the remaining
offspring and dams were sacrificed and discarded without further
examinations.  Subsequently, 1 pup/litter/group was allocated to subsets
for FOB, motor activity, acoustic startle response, learning and memory
evaluation, and neuropathological examination.  A maternal LOAEL was not
observed.  The maternal NOAEL is 4000 ppm (326.2 mg/kg/day).

	In the offspring, no treatment-related effects were seen on survival,
clinical signs, FOB, developmental land marks, brain weights or
neuropathology.  No conclusions can be drawn on the effects of the
compound on motor activity due the lack of consistency in the
observations. At the high dose, treatment-related effects were decreased
body weights and increased auditory startle response.  No differences
from controls were observed in any morphometric parameter in the treated
animals on PND 12, except for increased length from midline in the
hippocampus of the 4000 ppm females.  The following significant
differences in various morphometric measurements were noted on PND 63:
(i) decreased dorsal cortex 1 thickness at (100 ppm in males; (ii)
decreased dorsal cortex 2 thickness at (100 ppm in males; (iii)
decreased thickness of the molecular layer of the cerebellum at the
preculminate fissure in the 4000 ppm males; (iv) decreased thickness of
the molecular layer of the cerebellum at the prepyramidal fissure in the
(1000 ppm males; and (v) increased thickness of the corpus callosum in
the 4000 ppm females.  The biological significance of the reduction in
the dorsal cortex, which is not dose-dependent, is unclear.   The
offspring LOAEL is 100 ppm (82 mg/kg/day) based on changes in brain
morphometrics in the cerebellum.  The offspring NOAEL is (8.2
mg/kg/day).

3.2.4  	Pre-and/or Postnatal Toxicity tc \l3 "4.2.6  Pre-and/or
Postnatal Toxicity 

The toxicology database for acibenzolar-S-methyl is sufficient for FQPA
assessment.  In the rat developmental toxicity study treatment related
visceral malformations and skeletal variations were observed in fetuses
at 200 mg/kg/day, the NOAEL for maternal toxicity.  In the developmental
neurotoxicity study, offspring toxicity was observed at 82 mg/kg/day
while no maternal toxicity was observed at 326 mg/kg/day, the highest
dose tested.  Additional developmental toxicity studies in rats and
rabbits and reproduction studies in rats provided no indication of
increased susceptibility of rat or rabbit fetuses or neonates compared
to adult animals.  Based on the results of developmental toxicity and
developmental neurotoxicity studies in rats there is concern for
increased qualitative and/or quantitative susceptibility following in
utero exposure to acibenzolar-S-methyl.  There are no residual
uncertainties with regard to pre- and/or postnatal toxicity.  

Immunotoxicity

There are no indications in the available studies that organs associated
with immune function, such as the thymus and spleen, are affected by
acibenzolar-S-methyl.  While effects on the spleen were observed in
association with hematologic effects, these were considered to secondary
to the primary effects on blood hematology.  Effects on the thymus were
seen in only one study in one animal at a high dose (400 mg/kg/day).  An
immunotoxicity study is required.  This is a new data requirement under
40 CFR Part 158 as a part of the data requirements for registration of a
pesticide (food and non-food uses).

	3.2.6	FQPA Factor

	The HED Toxicology Science Advisory Committee (ToxSAC) recommended that
the 10X FQPA safety factor be reduced to 1X since endpoints protective
of sensitive populations are being used in the risk assessment.  

[Note: In previous risk assessments a 10X FQPA safety factor was added
for increased quantitative susceptibility (umbilical hernia) in the
developmental rat study and lack of a DNT study.  As previously noted,
based on reevaluation of existing data and review of newly submitted
data, the umbilical hernias are not considered to be treatment related
and a DNT study has been submitted.  Therefore, the 10X FQPA safety
factor may be reduced to 1X.]

3.3	Toxicity Endpoint Selection

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

	Selected Study: Developmental Neurotoxicity in Rats (MRID 46046401).  

	See Section 3.2.3

	Dose and Endpoint for Establishing an aRfD:  8.2 mg/kg/day based on an
offspring NOAEL of 8.2 mg/kg/day and an offspring LOAEL of 82 mg/kg/day
based on changes in brain morphometrics in the cerebellum.  

Uncertainty Factor (UF): 100 – 10x for interspecies extrapolation and
10x for intraspecies variation.

Comments about Study/Endpoint:  The developmental neurological effects
of concern are presumed to occur after a single exposure and are
relevant only to females aged 13-50 years old since they occur in utero.
 This endpoint has been revised from previous acibenzolar-S-methyl
assessments. The acute dietary endpoint selected for use in prior
acibenzolar-S-methyl assessments was based on a developmental rat study
(MRID 44014236).  LOAEL and NOAELs for this study have since been
revised.    

  

	3.3.2	Acute Reference Dose (aRfD) - General Population

No appropriate endpoint was identified for the general population which
might occur following a single dose or exposure.  

3.3.3 	Chronic Reference Dose (cRfD) - Females age 13-49

Selected Study: Developmental Neurotoxicity in Rats (MRID 46046401).  

See Section 3.2.3

	Dose and Endpoint for Establishing a cRfD:  8.2 mg/kg/day based on an
offspring NOAEL of 8.2 mg/kg/day and an offspring LOAEL of 82 mg/kg/day
based on changes in brain morphometrics in the cerebellum.  

Uncertainty Factor (UF): 100 – 10x for interspecies extrapolation and
10x for intraspecies variation.

Comments about Study/Endpoint:  The effects observed in this study can
occur after single or multiple doses.  Selection of a chronic endpoint
from this study ensures that the chronic exposure assessment is
protective of females of reproductive age and their offspring.   

3.3.4	Chronic Reference Dose (RfD) – General Population, including
infants and children

Selected Studies: Co-critical studies Chronic Feeding Study in Dogs
(MRID 44014234); Oncogenicity Study Mouse (MRID 44014235);
Chronic/Oncogenicity Study Rat (MRID 44014243); Reproductive Study Rat
(44014241)

Chronic Dog Study - In a chronic oral toxicity study
acibenzolar-S-methyl was administered to 4 beagle dogs per sex per dose
level in gelatin capsules at levels of 0, 5, 25, or 200 mg/kg/day for 52
weeks.  No biologically significant toxic effects were observed in
either males or females receiving 25 mg/kg/day or less of the test
material. At 200 mg/kg/day, there was an indication of hemolytic anemia
in dogs of both sexes i.e., effects consistent with prior or current
hemolytic anemia including hematological effects, hemosiderosis of the
liver and spleen, extraniedullary hematopoiesis in the spleen, and
increases in liver weight.  Significant treatment-related changes were
found in clinical chemistry in the dogs fed the test, substance at 200
mg/kg/day, (increase in triglycerides in both sexes, increase of
cholesterol in females at week 13, decrease in total protein in males at
weeks 26 and 52, and decrease in albumin in both sexes at week 13. These
clinical chemistry changes are suggestive of possible nutritional
deficiencies.  The LOAEL is 200 mg/kg/day in both sexes based on effects
consistent with prior or current hemolytic anemia.  The NOAEL is 25
mg/kg/day.  

Oncogenicity Mouse Study - In an oncogenicity study acibenzolar-S-methyl
was administered in the diet to groups of 60 male and 60 female Tif:MAGf
(SPF) mice at concentrations of 0, 10, 100, 2000, or 6000 ppm (1.14,
11.11, 237, and 698 mg/kg/day for males and 1.14, 10.8, 234, and 696
mg/kg/day for females) for 18 months.  Ten mice per group were reserved
for measurements of hematology parameters after 12 and 18 months of
treatment.  Treatment did not result in a significant decrease in
survival.  The overall food efficiency was decreased in females by 22%
at 6000 ppm.  The red blood cell count, hemoglobin, and hematocrit at 53
and 79 weeks were decreased in females at 2000 ppm and 6000 ppm.  Mean
corpuscular volume was decreased in high dose males at 79 weeks.  Mean
corpuscular hemoglobin concentration distribution increased at 6000 ppm.
 Reticulocyte counts in high-dose females was significantly increased. 
The hematology data are consistent with hemolytic anemia especially in
females at 2000 and 6000 ppm.  Macroscopic and microscopic findings
consistent with a mild hemolytic anemia in both sexes included increased
incidences of enlarged spleen, increased incidences of hemosiderosis in
liver, spleen, and bone marrow.  Incidence of extramedullary
hematopoiesis in the spleen and pancreatic exocrine hyperplasia was
increased in males at 2000 and 6000 ppm.  Incidences of alveolar foam
cells in the lungs were increased at 6000 ppm in both sexes.  The LOAEL
is 2000 ppm (237/234 mg/kg/day males/females), based on mild hemolytic
anemia and related microscopic changes (hemosiderosis) in the spleen,
liver, and bone marrow and extramedullary hematopoiesis in the spleen. 
The NOAEL was 100 ppm (11.1 mg/kg/day males; 10.8 mg/kg/day females).

Chronic/Oncogenicity Rat Study - In a chronic toxicity/oncogenicity
study, acibenzolar-S-methyl was administered in the diet to groups of 80
male and 80 female (Tif:RAIf (SPF)) Sprague-Dawley derived rats at
concentrations of 0, 20, 200, 2500, or 7500 ppm (0, 0.77, 7.77, 96.9,
and 312 mg/kg/day male and 0, 0.90, 9.08, 111, and 388 mg/kg/day
female).  Ten animals per sex per dose were sacrificed at 52 weeks for
interim evaluation; the remaining animals were used for hematologic and
clinical chemistry evaluation (20 per sex per group) and evaluation of
carcinogenic potential (50 animals per sex per group).  No
treatment-related effects were observed at 20, 200 or 2500 ppm for any
parameter.  Treatment related effects on body weight, weight gain, food
consumption, and food efficiency were observed in both sexes at 7500
ppm.  Other treatment-related effects at 7500 ppm included a slight
hemolytic anemia in both sexes as evidenced by decreases in erythrocyte
count, increased mean corpuscular volume, elevated reticulocyte counts,
increased serum bilirubin, increased severity of hemosiderin deposition
in the spleen, and increased incidence of hemosiderin deposition in
Kupifer cells of the liver.  Hemosiderin deposition in the spleen was
accompanied by a statistically significant increase in spleen weight. 
There was also a statistically significant increase in the incidence of
foam cells in the lungs of females fed the 7500-ppm diet.  The LOAEL was
7500 ppm (312/388 mg/kg/day males/females) based on decreased body
weights, body weight gain and food efficiency, mild hemolytic anemia,
and increased incidence of alveolar foam cells (females only). The NOAEL
was 2500 ppm (96.9 mg/kg/day males and 111 mg/kg/day females).  

Reproductive Study in Rats - See Section 3.2.2

Dose and Endpoint for Establishing cRfD:  25 mg/kg/day based on a NOAEL
of 25 mg/kg/day from the chronic dog study and a LOAEL of 105 mg/kg/day
from the rat reproduction study based on hemolytic anemia with
compensatory response. 

Uncertainty Factor(s): 100 – 10x for interspecies extrapolation and
10x for intraspecies variation.

	Comments about Study/Endpoint/Uncertainty Factor: The NOAEL of 25
mg/kg/day, taken from the chronic dog study, is based on the totality of
the database with a LOAEL of 105 mg/kg/day (the lowest LOAEL is from the
reproduction study) based on hemolytic anemia with compensatory
response.  The NOAEL and LOAEL recommended for use in the risk
assessment come from different studies conducted in different species. 
Typically, a combination of NOAELs/LOAELs from studies conducted in
different species is not recommended.  However, in the case of
acibenzolar-S-methyl the same endpoint (hemolytic anemia) is observed
across species at a relatively close range of LOAELs (105-312
mg/kg/day).  Therefore, the ToxSAC concluded that, in this instance, it
would be appropriate to base the point of departure for the risk
assessment on the NOAEL from the dog chronic toxicity study using the
LOAEL from the reproduction study in rats for the basis of the endpoint
(hemolytic anemia).

	

	3.3.5   	Incidental Oral Exposure (Short- and Intermediate-Term)

There are no residential uses for acibenzolar-S-methyl.  Therefore
incidental oral exposure endpoints are not required and not selected for
this assessment.   

3.3.6	Dermal Absorption

	There are no dermal absorption studies available with
acibenzolar-S-methyl.  However, the dermal absorption rate for
acibenzolar-S-methyl is estimated to be 10% by comparing  developmental
toxicity LOAEL (50 mg/kg/day) from an oral developmental toxicity study
in rats (MRID 44014236) to the developmental toxicity LOAEL (>500
mg/kg/day) from a dermal developmental toxicity study in rats (MRID
44014240).   

3.3.7	Dermal Exposure (Short and Intermediate-Term)

Selected Study: Developmental Neurotoxicity in Rats (MRID 44014236).  

See Section 3.2.3

	Dose and Endpoint:  8.2 mg/kg/day based on an offspring NOAEL of 8.2
mg/kg/day and an offspring LOAEL of 82 mg/kg/day based on changes in
brain morphometrics in the cerebellum.  

Uncertainty Factor (UF): 100 – 10x for interspecies extrapolation and
10x for intraspecies variation.

	Comments about Study/Endpoint:  Two acceptable dermal toxicity studies,
including a dermal developmental study were available.  However since
neurological effects observed in the oral developmental neurotoxicity
study are not measured in the dermal developmental study, the findings
from the dermal study may not be protective of these neurological
effects.  Since an oral NOAEL was selected for dermal risk assessment, a
dermal absorption factor of 10% should be used.  This factor is based on
the ratio of the dermal developmental NOAEL of 500 divided by the oral
developmental NOAEL of 50 which equals 10%.

3.3.8 	Inhalation Exposure (Short and Intermediate-Term)

Selected Study: Developmental Neurotoxicity in Rats (MRID 44014236).  

See Section 3.2.3

	Dose and Endpoint:  8.2 mg/kg/day based on an offspring NOAEL of 8.2
mg/kg/day and an offspring LOAEL of 82 mg/kg/day based on changes in
brain morphometrics in the cerebellum.  

Uncertainty Factor (UF): 100 – 10x for interspecies extrapolation and
10x for intraspecies variation.

	Comments about Study/Endpoint:  Since an oral NOAEL was selected for
inhalation risk assessment, an inhalation absorption factor of 100%
should be used

3.3.9	Classification of Carcinogenic Potential tc \l3 "4.4.8
Classification of Carcinogenic Potential 

The Hazard Identification Assessment Review Committee (HIARC) classified
acibenzolar-S-methyl as a “not likely” human carcinogen based on the
lack of evidence of carcinogenicity in rats and mice and on the lack of
unequivocal genotoxicity in an acceptable battery of mutagenicity
studies.  

[Note: The cancer classification has not been updated based on the
EPA’s 2005 Guidelines for Carcinogen Risk Assessment.]

3.4	Margins of Exposure

A summary of target Levels of Concern for risk assessment is provided in
Table 6.



Table 6.  Target Levels of Concern/Margin of Exposure for
Acibenzolar-S-Methyl

Route/Duration	Short-Term

(1-30 Days)	Intermediate-Term

(1 - 6 Months)	Long-Term

(> 6 Months)

Occupational (Worker) Exposure

Dermal	100	100	N/A

Inhalation	100	100	N/A

Residential (Non-Dietary) Exposure

Oral	N/A	N/A	N/A

Dermal	NA	N/A	N/A

Inhalation	N/A	N/A	N/A

3.5	Recommendation for Aggregate Exposure Risk Assessments

When there are potential residential exposures to the pesticide,
aggregate risk assessment must consider exposures from three major
sources: oral, dermal and inhalation exposures.  Since there are no
residential incidental oral exposures to acibenzolar-S-methyl, aggregate
exposure from food and non-food oral exposures is not required.  Dermal
and inhalation exposures to workers should be aggregated for
acibenzolar-S-methyl, however, because the same oral toxicity endpoint
was selected for both exposure routes.    

3.6	Summary of Endpoints Selected for Risk Assessment

Toxicological doses/endpoints selected for the acibenzolar-S-methyl risk
assessment are provided in Tables 7 and 8.

Table 7. Summary of Toxicological Doses and Endpoints for
Acibenzolar-S-methyl  for Use in Dietary Human Health Risk Assessments

Exposure/Scenario	Point of Departure	Uncertainty/FQPA Safety Factors
RfD, PAD, LOC for Risk Assessment	Study and Toxicological Effects

Acute Dietary (General Population)	An acute RfD for the general
population or any population subgroups was not selected  because no
effect attributable to a single (or few) day(s) oral exposure was
observed in animal studies.

Acute Dietary

(Females 13-49 years)	NOAEL = 8.2 mg/kg/day	UFA= 10x

UFH=10x

FQPA SF= 1x	Acute RfD = 0.082 mg/kg/day

aPAD = 0.082 mg/kg/day	Developmental Neurotoxicity Toxicity Study - Rat 

Developmental LOAEL = 82 mg/kg/day based on changes in brain
morphometrics in the cerebellum in offspring.  

Maternal LOAEL = was not observed NOAEL  = 4000 mg/kg/day HDT

Chronic Dietary 

(Females 13-49 years)	NOAEL = 8.2 mg/kg/day	UFA= 10x

UFH=10x

FQPA SF= 1x	Acute RfD = 0.082 mg/kg/day

aPAD = 0.082 mg/kg/day	Developmental Neurotoxicity Toxicity Study - Rat 

Developmental LOAEL = 82 mg/kg/day based on changes in brain
morphometrics in the cerebellum in offspring.  

Maternal LOAEL = was not observed 

NOAEL  = 4000 mg/kg/day HDT

Chronic Dietary 

(All Populations)	NOAEL = 25 mg/kg/day	UFA= 10x

UFH=10x

FQPA SF= 1x	Chronic RfD = 0.25

mg/kg/day

cPAD = 0.25 mg/kg/day	Chronic Toxicity - Dog; Co-critical;
Chronic/Cancer - Rat & Mouse, Reproduction Toxicity - Rat 

LOAEL = 105 mg/kg/day based on hemolytic anemia with compensatory
response.  

Incidental Oral 

Dermal

Inhalation	There are no residential uses for or exposures to
acibenzolar-S-methyl.  Therefore exposure endpoints are not required and
not selected for residential exposure assessment.   

Cancer (all routes)	A “not likely” human carcinogen

Point of Departure (POD) = A data point or an estimated point that is
derived from observed dose-response data and  used to mark the beginning
of extrapolation to determine risk associated with lower environmentally
relevant human exposures.  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).  UFL = use of a LOAEL to extrapolate a NOAEL.  UFS = use
of a short-term study for long-term risk assessment.  UFDB = to account
for the absence of key date (i.e., lack of a critical study).  FQPA SF =
FQPA Safety Factor.  PAD = population adjusted dose (a = acute, c =
chronic).  RfD = reference dose.  MOE = margin of exposure.  LOC = level
of concern.  N/A = not applicable.

 

Table 8.  Summary of Toxicological Doses and Endpoints for
Acibenzolar-S-methyl  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 (1-30 days) and Intermediate (1-6 months) Term

DAF = 10%	NOAEL= 8.2 mg/kg/day	UFA= 10x

UFH=10x

	Occupational LOC for MOE = 100	Developmental Neurotoxicity Toxicity
Study - Rat 

Developmental LOAEL = 82 mg/kg/day based on changes in brain
morphometrics in the cerebellum in offspring.  

Maternal LOAEL = was not observed 

NOAEL  = 4000 mg/kg/day HDT

Inhalation Short (1-30 days) and Intermediate (1-6 months) Term	NOAEL=
8.2 mg/kg/day	UFA= 10x

UFH=10x

	Occupational LOC for MOE = 100	Developmental Neurotoxicity Toxicity
Study - Rat 

Developmental LOAEL = 82 mg/kg/day based on changes in brain
morphometrics in the cerebellum in offspring.  

Maternal LOAEL = was not observed 

NOAEL  = 4000 mg/kg/day HDT

Cancer (all routes)	A “not likely” human carcinogen

DAF = Dermal Absorption Factor 

3.7	Endocrine Disruption

EPA is required under the FFDCA, as amended by FQPA, to develop a
screening program to determine whether certain substances (including all
pesticide active and other ingredients) “may have an effect in humans
that is similar to an effect produced by a naturally occurring estrogen,
or other such endocrine effects as the Administrator may designate.” 
Following recommendations of its Endocrine Disruptor and Testing
Advisory Committee (EDSTAC), EPA determined that there was a scientific
basis for including, as part of the program, the androgen and thyroid
hormone systems, in addition to the estrogen hormone system.  EPA also
adopted EDSTAC’s recommendation that the Program include evaluations
of potential effects in wildlife.  For pesticide chemicals, EPA will use
FIFRA and, to the extent that effects in wildlife may help determine
whether a substance may have an effect in humans, FFDCA authority to
require the wildlife evaluations.  As the science develops and resources
allow, screening of additional hormone systems may be added to the
Endocrine Disruptor Screening Program (EDSP).  

4.0	DIETARY AND DRINKING WATER EXPOSURE ASSESSMENT

4.1 	Residue Chemistry Profile

	4.1.1	Metabolism in Primary Crops 

	The nature of the residues in plants has been adequately delineated
based on metabolism studies with lettuce, tomato, rice, wheat, and
tobacco.  The HED MARC has determined that the residues of concern in
plants for tolerance expression are residues convertible to
benzo(1,2,3)thiadiazole-7-carboxylic acid (CGA-210007), expressed as
acibenzolar-S-methyl.  For purposes of risk assessment, residues of the
4- and 5-hydroxy metabolites (CGA-324041 and CGA-323060) of CGA-210007
should also be considered.  For this purpose, the toxicity of CGA-324041
and CGA-323060 should be considered to be equivalent to that of the
parent compound acibenzolar-S-methyl.  These residues occur in
significant quantities in lettuce and tomato.  At this time,
acibenzolar-S-methyl does not have common metabolites with other
agrochemicals.  The residues of concern for drinking water are the same
as those listed in the tolerance expression; that is, residues
convertible to CGA-210007.  Table 9 provides a summary of the MARC
decisions regarding residues of concern for acibenzolar-S-methyl.  

Table 9:  Summary of MARC Decisions for Acibenzolar-S-methyl 

Matrix	Residues of Concern 1

	For Risk Assessment 2	For Tolerance Expression

Plants	Residues of acibenzolar-S-methyl, benzo(1,2,3)
thiadiazole-7-carbothioic acid-S-methyl ester, convertible to
benzo(1,2,3)thiadiazole-7-carboxylic acid (CGA-210007), expressed as
acibenzolar-S-methyl &

4-hydroxy CGA-210007 (CGA-323060)

5-hydroxy CGA-210007 (CGA-324041)	Residues of acibenzolar-S-methyl,
benzo(1,2,3) thiadiazole-7-carbothioic acid-S-methyl ester, convertible
to benzo(1,2,3)thiadiazole-7-carboxylic acid (CGA-210007), expressed as
acibenzolar-S-methyl 

1 The residues of concern for drinking water are the same as those
listed in the tolerance expression

2 If a future petition is received for rice, residues of CGA-379019
should be included in the risk assessment

	4.1.2   Residue Analytical Methods

	4.1.2.1 Enforcement Methods	

	An adequate method, HPLC/UV Method AG-671A, is available for tolerance
enforcement.  This method has undergone a successful tolerance method
validation by the Analytical Chemistry Branch of BEAD.  The method was
forwarded to FDA for inclusion in PAM Volume II.  The method consists of
an initial hydrolysis with NaOH to convert acibenzolar-S-methyl to
CGA-210007 followed by methanol extraction.  Residues are then diluted
with HCl and purified by a series of solid-phase extraction steps. 
Prior to HPLC/UV analysis, residues are partitioned into ethyl acetate,
dried down, and redissolved in phosphoric acid.  This method has an LOQ
of 0.02 ppm.  The method includes optional detection via HPLC/MS, giving
a means of residue confirmation.  The HPLC/UV and LC/MS methods are
adequate for collecting data on residues of acibenzolar-S-methyl in/on
cucurbit vegetables and bulb onions, respectively.  

	4.1.2.2	Multiresidue Methods

	  SEQ CHAPTER \h \r 1  Data depicting the recovery of
acibenzolar-S-methyl and its metabolite, CGA-210007, using FDA
Multiresidue Methods, were previously submitted.  Acibenzolar-S-methyl
yielded adequate response to GLC analysis and was completely recovered
through Protocols D and E.  Partial recovery of CGA-210007 was achieved
via Protocol B after derivatization.

4.1.3	Residues in Crops

	The number and locations of field trials are in accordance with OPPTS
Guideline 860.1500 for bulb onion as the representative crop of the bulb
onion subgroup 3-07A, and for cantaloupe, cucumber, and summer squash as
the representative crops of the cucurbit vegetable group 9.  The trials
conducted reflect the proposed maximum seasonal rate and PHI, and there
are adequate storage stability data to validate sample storage
conditions and duration.  

	An adjuvant was not used in the onion field trials, however, and the
data do not support the use of a silicone adjuvant as proposed on the
Actigard® label.  If the petitioner desires to keep the use of the
surfactant on the label for bulb onions, then three confirmatory trials
using Actigard® with a surfactant should be tested side-by-side with
product applied without using a surfactant in the NAFTA Growing Zones 10
(2 sites) and 11 (1 site).  Alternatively, the petitioner may revise the
product label to disallow the use of silicone surfactant on the proposed
label.

	The results of the cucurbit vegetables residue decline studies
demonstrate that average residues of acibenzolar-S-methyl (i.e.,
residues convertible to CGA-210007) in/on cantaloupe remained generally
constant between the 0- and 3-day PHIs and increased in/on cucumber and
summer squash by approximately 10% and 100%, respectively.  Residues
generally decreased in cantaloupe, cucumber, and summer squash by the
7-10 day PHIs and were the lowest at the 10-day PHI.  These data suggest
that residues of acibenzolar-S-methyl in/on cucurbit vegetables
resulting from the proposed use rate and 0-day PHI may not represent the
worst case as residues may increase between 0- and 7-day PHIs and
generally decrease thereafter.  This phenomenon was noted in an earlier
acibenzolar-S-methyl review (D258680, M. Doherty, 05/02/2000) where
decline trials demonstrated that average residues of
acibenzolar-S-methyl in/on broccoli increased between the 0- and 5-day
PHIs by approximately 300% and than generally decreased thereafter.  An
explanation for this phenomenon is not readily apparent.  Given the
indication of variability within and between these cucurbit vegetable
data sets and the demonstrated potential for significant increases in
acibenzolar-S-methyl residues between 0- and 7-day PHIs, six additional
field trial/decline studies are required to support the proposed use of
acibenzolar-S-methyl on cucurbit vegetables.  Two side-by-side
cantaloupe and honeydew melon decline studies demonstrating the proposed
use rate and using an adjuvant must be conducted in Zones 6 and 10 (a
total of 4 trials), samples must be collected at 0, 1, 3, 5, 7, and 10
days after the last application (DALA).  Two cantaloupe decline studies,
using the same protocol, must be conducted in Zones 2 and 5 (a total of
2 trials). 

	Adequate confined rotational crop data have been submitted previously. 
Based on the available field rotational crop data for wheat, turnip, and
lettuce, it has been determined that tolerances are not needed for
rotational crops provided a 30-day plant-back interval is established
for all crops not listed as target crops.  The existing rotational crop
restrictions on the Actigard® label are adequate.

4.2	Drinking Water Profile

	4.2.1  	Environmental Fate and Transport

	Laboratory and field studies indicate that acibenzolar-S-methyl does
not have a high potential to leach to ground water nor move to surface
water based on its short resident times in soil and soil water.  The
major degradate observed was benzo[1,2,3]thiadiazole-7-carboxylic acid
(CGA 210007) in the hydrolysis and soil photolysis studies (up to 100
percent).  CGA-210007 has a longer soil residence time, lower Freundlich
Kd adsorption values, and is much more water soluble than
acibenzolar-S-methyl, thereby exhibiting greater potential than the
parent for lateral or vertical movement after application. 
Acibenzolar-S-methyl mainly degrades by photolysis and microbial action.
 Aqueous and soil photolysis irradiated half lives for parent are 0.6
hours and 1 day, respectively. The aerobic soil metabolism, anaerobic
aquatic metabolism and aerobic aquatic half lives are 5.3 hours, 4.0 and
< 1 day(s) respectively.  Field dissipation half-lives ranged from < 1
to 27 days.  Under aerobic soil, aerobic aquatic, and anaerobic aquatic
conditions acibenzolar quickly degrades to CGA 210007.  CGA 210007 is in
its anionic form at environmentally significant pH's (pH 6-9) and
appears to be more soluble than its parent, although measured at much
lower pH’s (approx. 225 ppm at 250C pH 3.6-3.8 v. 7.7 ppm at 250C pH
7.5-7.9).  CGA 210007 has longer resident field times than parent
(aerobic soil half-life at 16.5 days) and based on the batch equilibrium
and aged soil column leaching studies, is considerably more mobile. 
Acibenzolar is not considered a bioconcentrate nor is it highly
bioaccumulative.  

	4.2.2	Estimated Drinking Water Concentrations 

	Drinking water residues were incorporated directly into the acute and
chronic dietary analyses (“water, direct, all sources” and “water,
indirect, all sources”) and were provided by the Environmental Fate
and Effects Division (EFED; D351609, R. Miller, 8-Aug-2008). EFED
generated the surface water and ground water estimates using the   SEQ
CHAPTER \h \r 1 Pesticide Root Zone Model 3 (PRZM3; ver. 3.12.2;
2-May-2005)/Exposure Analysis Modeling System (EXAMS; ver. 2.98.04.06;
25-Apr-2005) and Screening Concentration in Ground Water (SCIGROW; Tier
1) models, respectively.  The residues of concern for drinking water
include acibenzolar-S-methyl and residues convertible to CGA-210007. 
EFED provided estimates for the proposed use of acibenzolar-S-methyl on
cucurbits because this use represents the highest use rate of the
proposed registered uses.  The SCI-GROW estimated ground water drinking
water concentrations for acibenzolar is not expected to exceed 4.08 x
10-5 µg/L and CGA 210007 is not expected to exceed 0.557 µg/L. 
Surface water EDWCs for acibenzolar-S-methyl, which were used in the
exposure assessment, are provided in Table 10.  CGA-210007 drinking
water residues were included in the dietary exposure assessment as
acibenzolar-S-methyl equivalents. 

 

 Table 10.  EDWCs for use in screening the Acibenzolar-S-Methyl Human
Health Risk Assessment 

Drinking Water Source	Chemical	Acute (μg/L)	NonCancer Chronic (μg/L)
Cancer Chronic(μg/L)

Surface Water	Acibenzolar1	0.74	0.10	0.05

Surface Water	CGA 2100072	14.21	9.48	5.47

1 Aerial application method

2 Degradation product formed after application 

 4.3 	Dietary and Drinking Water Exposure and Risk

cted using the Dietary Exposure Evaluation Model with the Food Commodity
Intake Database (DEEM-FCID™).  Dietary risk assessment incorporates
both exposure and toxicity of a given pesticide.  For acute and chronic
dietary assessments, the risk is expressed as a percentage of a maximum
acceptable dose (i.e., the dose which HED has concluded will result in
no unreasonable adverse health effects).  This dose is referred to as
the population adjusted dose (PAD).  The PAD is equivalent to the
reference dose (RfD) divided by the additional Safety Factor, if
applied. For acute and non-cancer chronic exposures, HED is concerned
when estimated dietary risk exceeds 100% of the PAD.  

	Available lettuce metabolism data for a 7-day PHI were used to estimate
the relative abundance of the hydroxy metabolites (CGA-323060 and
CGA-324041) to the residues of acibenzolar-S-methyl convertible to the
benzo(1,2,3) thiadiazole-7-carboxylic acid (CGA-210007).  Based on the
lettuce metabolism data, a factor of 1.5X was applied to estimates of
acibenzolar-S-methyl residues to account for all of the residues of
concern for dietary risk (including CGA-323060 and CGA-324041).  

	4.3.1	Acute Dietary and Drinking Water Analysis

A refined (probabilistic) acute dietary exposure analysis was performed
for the population subgroup females 13-49 only.  No acute endpoint was
identified for the remaining population subgroups.  The acute analysis
assumed a distribution of residues based on field trial data.  Empirical
and DEEM default processing factors were used to modify the field trial
data.  Maximum screening-level percent crop treated estimates were used
for commodities for which data were available.  If no percent crop
treated data were available, 100% crop treated was assumed.  The acute
analysis incorporated the 1 in 10 year peak surface drinking water
estimate from application of acibenzolar-S-methyl to cucurbit
vegetables.    SEQ CHAPTER \h \r 1 The resulting 99.9th percentile acute
exposure estimate for females 13-49 years old is not of concern to HED
(12% of the acute PAD).    

Table 11:  Summary of Acute Dietary Exposure and Risk for
Acibenzolar-S-Methyl

Population Subgroup	aPAD (mg/kg/day)	Acute (99th Percentile)

Exposure (mg/kg/day)	%aPAD

Females 13-49 years old	0.082	0.0010	12

	4.3.2	Chronic Dietary and Drinking Water Analysis

 A conservative chronic dietary exposure analysis was performed for the
general U.S. population and various population subgroups.  Tolerance
level residues and 100 crop treated assumptions were used.  DEEM default
and empirical processing factors were used to modify the tolerance
values.  The chronic analysis incorporated the 10 year average surface
drinking water estimate from application of acibenzolar-S-methyl to
cucurbit vegetables.  The population subgroup females 13-49 had a more
protective chronic population adjusted dose (cPAD, 0.082 mg/kg/day) than
the general U.S. population and all other population subgroups (0.25
mg/kg/day).  Chronic dietary risk estimates are not of concern for
general population or other population subgroups.  The subgroup with the
highest risk estimate was females 13-49 with a cPAD of 5%.  The cPAD for
the general U.S. population was 2%. 

Table 12:  Summary of Chronic Dietary Exposure and Risk for
Acibenzolar-S-Methyl 

Population Subgroup	cPAD (mg/kg/day)	Chronic

Exposure (mg/kg/day)	% cPAD

Females 13-49 years old	0.082	0.0038	5

Table 13:  Summary of Chronic Dietary Exposure and Risk for
Acibenzolar-S-Methyl

Population Subgroup	cPAD (mg/kg/day)	Chronic

Exposure (mg/kg/day)	% cPAD

General U.S. Population	0.25	0.0045	2

All Infants (< 1 year old)

0.0043	2

Children 1-2 years old

0.0093	4

Children 3-5 years old

0.0088	3.5

Children 6-12 years old

0.0059	2

Youth 13-19 years old

0.0043	2

Adults 20-49 years old

0.0038	1.5

Adults 50+ years old

0.0040	2

5.0 	AGGREGATE EXPOSURE AND RISK ASSESSMENT 

	In accordance with the FQPA, when there are potential residential
exposures to a pesticide, aggregate risk assessment must consider
exposures from three major routes: oral, dermal, and inhalation.  There
are three sources for these types of exposures:  food, drinking water,
and residential uses.  In an aggregate assessment, exposures from
relevant sources are added together and compared to quantitative
estimates of hazard (e.g., a NOAEL or PAD), or the risks themselves can
be aggregated.  When aggregating exposures and risks from various
sources, HED considers both the route and duration of exposure.  The
proposed/registered acibenzolar-S-methyl uses are not expected to result
in residential exposure.  Therefore, the acute and chronic exposure
estimates provided in the Dietary Exposure Section represent aggregate
exposure.  Dermal and inhalation exposures to workers should be
aggregated for acibenzolar-S-methyl because the same oral toxicity
endpoint was used to assess risk from both exposure routes.   

6.0	OCCUPATIONAL EXPOSURE AND RISK

6.1	 Exposure Scenarios

		Occupational handler and post-application exposure scenarios were
assessed for the risk assessment of the proposed new uses on bulb onion
and cucurbits.  

	6.1.1	Handler Exposure Scenarios

		The term “handler” applies to individuals who mix, load, and apply
the pesticide product.  The following handler exposure scenarios were
assessed for proposed new acibenzolar-S-methyl uses. 

	

	1)   	Mixing/Loading Water Soluble Granular for Aerial Application	

	2)   	Mixing/Loading Water Soluble Granular for Ground Boom Application

	3)   	Applying Sprays with Fixed-wing Aircraft (Closed cab)

	4)   	Applying Sprays with Ground boom (Open Cab)

	Flagging to support aerial applications was also considered.  However,
since flagging is typically done mechanically, exposures are minimal and
are covered by mixing, loading and application assessments.  

		

	6.1.2 	Post-Application Exposure Scenarios

Post-application exposures were assessed for the proposed new uses. 
There are no compound specific data with which to estimate
post-application exposures to agricultural workers.  Estimates of
post-application re-entry exposure to agricultural workers are based
upon the EXPOSAC Standard Operating Procedures (SOPs) (3.1, Reference
4).  This SOP lists a number of possible post-application agricultural
activities for the proposed crop uses that might result in
post-application. 

6.2 	Occupational Exposure Data and Assumptions

	6.2.1	Exposure Data

	6.2.1.1	Application Parameters 

	Maximum application rates for all of the exposure scenarios assessed
are based on information provided in the acibenzolar-S-methyl labels for
the proposed new uses. The maximum application rate for cucurbits is
0.06 lbs ai per acre and 0.03 lb ai per acre for onions.  

6.2.1.2	Occupational Exposure Data

	Data from the Pesticide Handlers Exposure Database (PHED) was used to
assess handler and post-application exposures in the absence of
chemical-specific data.  The transfer coefficients used in the
post-application exposure assessment are from an interim transfer
coefficient guidance document developed by HED’s Science Advisory
Council for Exposure using proprietary data from the Agricultural
Re-entry Task Force (ARTF) data base (SOP #3.1)

	6.2.2	Exposure Assumptions

		The following standard exposure assumptions were used in estimating
risks to workers from exposure to acibenzolar-S-methyl for the proposed
new uses on onion and cucurbits.

		6.2.2.1 Handler Exposures

	•	Average body weight of an adult handler is 60 kg.

	•	Exposure duration is short-term and intermediate-term for all
workers assessed.  

	•	Maximum application rates as determined by label review were used
for all types and methods of application.

	•	SOP daily volumes handled and/or area treated used for the
scenarios assessed are: 

	- 	350 acres treated per day for aerial application

	-  	80 acres treated per day for groundboom mixing, loading and
applying

				

		6.2.2.2 Post Application Exposures

	•	Average body weight is 60 kg.

	•	Transferrable residue is application rate times the fraction
initially available (20%). 

	•	Maximum transfer coefficient for post-application activities is
2500 cm2/hour (hand harvesting).

	•	Exposure duration is 8 hours day. 

	•	Exposure is assumed to occur on the day of application (day 0). 

6.3		Occupational Exposure and Risk Estimates

			

		6.3.1 	Handler Exposure and Risk Estimates

	A target LOC or MOE of 100 is considered adequate for inhalation and
dermal exposure. All worker exposures are assessed as short-and
intermediate-term based on label prescribed uses and expected exposure
durations.  Dermal and inhalation exposures to workers should be
aggregated for acibenzolar-S-methyl because the same oral toxicity
endpoint was selected for these exposure routes.  Exposure and risk
estimates indicate MOEs are not of concern (MOEs > 100) at the maximum
use rate for occupational handler activities for the proposed new uses. 
A summary of occupational handler exposure and risk calculations,
assumptions, and results is provided in Table 14.

Table 14.  Estimated Acibenzolar-S-methyl  Exposure & MOEs for
Occupational Handler Exposure - Dermal LOC/MOE = 100;  Inhalation
LOC/MOE = 100; 

Exposure 

Scenario1	Inhalation Unit Exposure   (ug/lb ai) 2	Dermal Unit Exp 

(mg/lb ai) 2	App Rate

(lbai/A) 3	Area Treated (A/day) 4	Inhalation Dose (m/k/d) 5	Inhal-

ation MOE6	Dermal

Dose

(m/k/d) 7	Dermal MOE8	 Aggreg MOE9

Mixing/Loading Water Soluble Granule (Dry Flowable) - Cucurbits

Aerial 	0.77	0.066	0.06	350	0.0003	29000	0.0024	3400	3000

Groundboom  

80	0.000065	130000	0.0006	150000	39000

Applying Sprays - Cucurbits

Fixed Wing Aircraft (Closed Cab) 	0.068	0.005	0.06	350	0.00003	330000
0.0002	450000	13000

Ground boom (Open Cab) 	0.014	0.74

80	0.00006	130000	0.0001	70000	46000

1  Use patterns are from the proposed labels Baseline PPE unit
exposures.  Values are reported in the PHED Surrogate Exposure Guide
dated August 1998 or are from data submitted by the Outdoor Residential
Exposure Task Force dated May 2000.

2 Baseline unit exposures.  Values are reported in the PHED Surrogate
Exposure Guide dated August 1998 or are from data submitted by the
Outdoor Residential Exposure Task Force dated May 2000.  

3 Application rates are based on maximum values based on proposed label.
 Most application rates upon which the analysis is based are presented
as lb ai/A.  In some cases, the application rate is based on applying a
solution at concentrations specified by the label (i.e., presented as lb
ai/gallon).  

4 Amount treated is based on the area or gallons that can be reasonably
applied in a single day for each exposure scenario of concern based on
the application method and formulation/packaging type. (Standard
EPA/OPP/HED values). 

5 Inhalation dose (mg/kg/day) = [unit exposure (ug/lb ai) * 0.001 mg/ug
unit conversion * Inhalation absorption (100%) * Application rate (lb
ai/acre or lb ai/gallon) * Daily area treated/amount handled (acres or
gallons)] / Body weight (60 kg).

6 Inhalation MOE = short-term endpoint for inhalation (NOAEL 8.2 mkd)/
Daily Inhalation Dose.

7 Dermal Dose (mg/kg/day) = [unit exposure (mg/lb ai) * Application rate
(lb ai/acre or lb ai/gallon) * Daily area treated/amount handled (acres
or gallons) * Dermal Absorption Factor (10%)] / Body weight (60 kg).

8 Dermal MOE = short-term and intermediate-term endpoint for dermal
(NOAEL 8.2 mkd)/Daily Dermal Dose.

9  Aggregate MOE = NOAEL (8.2 mkd)/Daily Inhalation Dose + Daily Dermal
Dose

	6.3.2 	Post-Application Exposure and Risk Estimates

		A target LOC or MOE of 100 is considered adequate for dermal exposure.
 Exposure and risk estimates indicate MOEs are not of concern (MOEs >
100) at the maximum use rate for occupational post-application exposure
activities for the proposed new uses.  A summary of post-application
exposure and risk calculations, assumptions, and results is provided in
Table 15.



Table 15.  Estimated Acibenzolar-S-methyl  Exposure & MOEs for
Occupational Post-application Exposure - Dermal LOC/MOE = 100 

Exposure Scenario	AR

(lb ai/A) 1	TTR/DFR (mg/cm2) 2	TC (cm2/hr) 3	Exposure Duration (hrs/day)
Dermal dose

(mg/kg/day) 4	Dermal MOE 5

Cucurbits, Hand weeding, scouting immature plants	0.06	0.000135	500	8
0.0008	9100

Cucurbits Scouting, irrigation mature plants

	1500

0.0027	3000

Cucurbits Hand harvesting

	2500

0.0045	1800

1 Application rates are based on maximum values based on proposed label.

2 DFR (mg/cm2) = Dislodgeable Foliar Residues corresponding to day 0.
Application Rate (lb ai/A) x CF (4.54E+5 mg/lb) x CF (2.47E-8 A/ cm2) x
20% (initial fraction of ai retained on foliage)

3 TC cm2/hr = Transfer coefficients and associated activities (ExpoSAC
Policy Memo #003.1)

4 Dermal Dose (mg/kg/day) = DFR (mg/cm2) x TC (cm2/hr) x 8 (hrs/day) *
10% DAF/ Body weight (60 kg).

5 Dermal MOE = short-term endpoint for dermal (NOAEL 8.2 mkd)/Dermal
Dose

7.0	CUMULATIVE RISK

Section 408(b)(2)(D)(v) of 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.”

	EPA does not have, at this time, available data to determine whether
acibenzolar-S-methyl has a common mechanism of toxicity with other
substances.  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
acibenzolar-S-methyl  and any other substances and, acibenzolar-S-methyl
 does not appear to produce a toxic metabolite produced by other
substances which have tolerances in the U. S.  For the purposes of this
tolerance reassessment action, therefore, EPA has not assumed that
acibenzolar-S-methyl  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 OPP concerning common mechanism determinations and procedures
for cumulating effects from substances found to have a common mechanism
on EPA’s website at   HYPERLINK
"http://www.epa.gov/fedrgstr/EPA_PEST/2002/January/Day_16/" 
http://www.epa.gov/fedrgstr/EPA_PEST/2002/January/Day_16/ .

8.0	DATA NEEDS

8.1		Toxicology Data Requirements  

		- 	Immunotoxicity (GLN 870.7800) 

		- 	Battery of mutagenicity studies performed on technical-grade
acibenzolar-S-methyl prepared by the thiazole production process.  

		- 	Ames assay using the technical-grade acibenzolar-S-methyl prepared
by the thiazole production process - including both plate incorporation
and pre-incubation. (GLN 870.5100, 870.5300, 870.375, 870.900) 

8.2  	Residue Chemistry Data Requirements

Directions for Use (GLN 860.

For bulb onions, if the petitioner desires to keep the use of the
surfactant on the label, then three confirmatory trials using Actigard®
with a surfactant should be tested side-by-side with product applied
without using a surfactant in the NAFTA Growing Zones 10 (2 sites) and
11 (1 site).  Alternatively, the petitioner may submit a revised Section
B to disallow the use of silicone surfactant on the proposed label. 

	      

	      For cucurbit vegetables, a revised Section B must be submitted to
remove the statement “allow 14 days between the last application and
harvest” since the proposed PHI is 0 days.  The label should also be
revised to specify a minimum retreatment interval of 7 days based on the
submitted data.

	

			Six additional field trial/decline studies are required to support
the proposed use of acibenzolar-S-methyl on cucurbit vegetables.  Two
side-by-side cantaloupe and honeydew melon decline studies demonstrating
the proposed use rate and using an adjuvant must be conducted in Zones 6
and 10 (a total of 4 trials), samples must be collected at 0, 1, 3, 5,
7, and 10 days after the last application (DALA).  Two cantaloupe
decline studies, using the same protocol, must be conducted in Zones 2
and 5 (a total of 2 trials).

	-    Proposed Tolerances (GLN 860.1550)

	

                     For the bulb onion petition, a revised Section F is
required to remove reference to “combined” residues in the tolerance
expression.  The tolerance level should be 0.1 ppm.  

	        

	       For the cucurbit vegetable petition, a revised Section F must be
submitted to reflect the correct commodity definition, Vegetable,
cucurbit, group 9.

	-    Submittal of Analytical Reference Standards (GLN 860.1650)

	       

	       The standard for the metabolite CGA 210007 (used for
quantitation) has expired (4/30/2008).  The registrant must either
recertify the lot in the repository and send in an updated certificate
of analysis (COA), or submit new standards (different lot #) if the
previous lots will not be recertified.

	

APPENDICES

TOXICOLOGY DATA SUMMARY

1.1 		Guideline Data Requirements

Table 1. Guideline Data Requirements

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-Day Dermal	

870.3250    90-Day Dermal	

870.3465    90-Day Inhalation		yes

yes

yes

no

yes	yes

yes

yes

-

yes

870.3700a  Developmental Toxicity (rodent)	

870.3700b  Developmental Toxicity (nonrodent)	

870.3800    Reproduction		yes

yes

yes	yes

yes

yes

870.4100a  Chronic Toxicity (rodent)	

870.4100b  Chronic Toxicity (nonrodent)	

870.4200a  Oncogenicity (rat)	

870.4200b  Oncogenicity (mouse)	

870.4300    Chronic/Oncogenicity		yes

yes

yes

yes

yes	yes

yes

yes

yes

yes

870.5100    Mutagenicity—Gene Mutation - bacterial	

870.5300    Mutagenicity—Gene Mutation - mammalian	

870.5375    Mutagenicity—Structural Chromosomal Aberrations	

870.5900    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  90-Day Neuro. Screening Battery (rat)	

870.6300    Develop. Neuro		no

no

yes

yes

no	-

-

yes

yes

-

870.7485    General Metabolism	

870.7600    Dermal Penetration	

870.7800    Immunotoxicity		yes

-

Yes	yes

-

no

1.2		Toxicity Profiles 

Table 2  Acute Toxicity Profile Acibenzolar-S-methyl  

Acibenzolar-S-methyl

Guideline No./

Study Type	MRID No.	Results	Toxicity Category

870.1100 Acute oral toxicity	44014214	LD50 >5000 mg/kg	IV

870.1200 Acute dermal toxicity	44014216	LD50 >2000 mg/kg	IV

870.1300 Acute inhalation toxicity	44014219	LC50 >5.022 mg/L	IV

870.2400 Acute eye irritation	44014220	minimal	III

870.2500 Acute dermal irritation	44014220	slight	IV

870.2600 Skin sensitization	44014225	positive	NA

Actigard 50WG

870.1100 Acute oral toxicity	44014215	LD50 >5000 mg/kg	IV

870.1200 Acute dermal toxicity	44014217	LD50 >2000 mg/kg	IV

870.1300 Acute inhalation toxicity	44014218	LC50 >2.79 mg/L	IV

870.2400 Acute eye irritation	44014221	minimal	III

870.2500 Acute dermal irritation	44014223	moderate	III

870.2600 Skin sensitization	44014224	negative	NA

Table 3  Subchronic, Chronic and Other Toxicity Profile for
Acibenzolar-S-methyl 

Guideline No.

Study Type	MRID No. (year)

Classification

Dose Levels	Results

870.3100

90-day oral toxicity rodents (rat)	44014230 (1993)

Acceptable/Guideline

M : 0, 2.42, 24.6, 126, 516 mg/kg/day;

F: 0, 2.64, 26.3, 131, 554 mg/kg/day	NOAEL:126 mg/kg/day Males or M);
131 mg/kg/day (Females or F)

LOAEL:516 mg/kg/day (M); 554 mg/kg/day females based decreased body
weight, reduced food intake and efficiency and increased liver and
spleen weights

870.3150

90-day oral toxicity non-rodents (dog)	44014232 (1994)

Acceptable/Guideline

M & F: 0, 10, 50, 200 mg/kg/day (capsules)

	NOAEL:50 mg/kg/day (M & F)

LOAEL:200 mg/kg/day (F) based on regenerative hemolytic anemia evidenced
by decreased erythroid parameters, pigmentation in the liver and spleen,
splenic congestion, and by activation of medullary (bone marrow) and
extramedullary (splenic) erythropoiesis.

870.3100

90-day oral toxicity rodents (mice)	44014228 (1993)

Acceptable/Guideline

0, 30.6, 152, and 624 mg/kg/day for males, and 0, 47.4, 220, and 803
mg/kg/day for females, respectively.	NOAEL: 30.6 mg/kg/day (M); 47.4
mg/kg/day (F)

LOAEL:152 mg/kg/day (M); 220 mg/kg/day females based decreased body
weight and body weight gain, increased spleen weight and microscopic
changes in the spleen

870.3200

21/28-day dermal toxicity (rat)	44014233 (1994)

Acceptable/guideline

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

LOAEL = not identified

870.3700a

Developmental Toxicity in rodents (rat)	44014236 (1994)

Acceptable/guideline

F: 0,10, 50, 200, 400 mg/kg/day

	Maternal NOAEL 200 mg/kg/day

Developmental NOAEL 50 mg/kg/day

Maternal LOAEL 400 mg/kg/day based on clinical signs of hemorrhagic
perineal discharge

Developmental LOAEL 200 mg/kg/day based on developmental malformations,
anomalies, and skeletal variations

870.3700a

Developmental Toxicity in rodents (rat)	45089701 (1998)

Acceptable/guideline

of 0, 10, 75, 150, or 350 mg/kg/day	Maternal NOAEL 350 mg/kg/day

Developmental NOAEL 200 mg/kg/day

Maternal LOAEL – not established (>350) 

Developmental LOAEL 350 mg/kg/day based on increased incidences of
rudimentary and long lumbar ribs

870.3700b

Developmental Toxicity in non- rodents (rabbit)	44014237 (1994)

Acceptable/guideline

F: 0, 10, 50, 300, 600 mg/kg/day

	Maternal NOAEL = 50 mg/kg/day

LOAEL = 300 mg/kg/day based on mortality, clinical signs of toxicity,
decreased maternal body weight and food consumption.

Developmental NOAEL = 300 mg/kg/day

LOAEL = 600 mg/kg/day based on a marginal increase in vertebral
anomalies.

870.6300

Developmental Neurotoxicity Toxicity in rodents (rat)	46046401 (2002)

Acceptable/non-guideline

0/0, 8.2/15.5, 82.0/153.6, and 326.2/607.8 mg/kg/day
[gestation/lactation].  	Maternal NOAEL 326.2 mg/kg/day

Developmental NOAEL 8.2 mg/kg/day

Maternal LOAEL Not Observed

Developmental LOAEL 82 based on changes in brain morphometrics in the
cerebellum

870.3800

Reproduction and fertility effects in rats	4014241 (1995)

Acceptable/guideline

M & F: 0, 1-3, 11-31, 105- 288, 223-604  mg/kg/day

	Parental/Systemic NOAEL = 11-31 mg/kg/day

LOAEL = 105-288 mg/kg/day based on increased weights and hemosiderosis
of the spleen.

Reproductive NOAEL = 223-604 mg/kg/day

LOAEL > 223-604 mg/kg/day based on no effects.

Offspring NOAEL = 11-31 mg/kg/day

LOAEL = 105-288 mg/kg/day based on reduced pup body weight gains and
lower pup body weights during lactation.

870.6200b 

Subchronic Neurotoxicity	45713601 (1997)

acceptable/nonguideline pending submission of positive control data for
neuropathological and FOB assessments

0/0, 24/26, 126/143, and 575/628 mg/kg/day [M/F], respectively

The study may be upgraded pending submission of acceptable positive
control data.	NOAEL 126/143 mg/kg/day, M/F

LOAEL 575/628 mg/kg/day, M/F based on decreased body weights, body
weight gains, and food consumption.  

870.4100

Chronic toxicity in dogs	44014234 (1996)

Acceptable/guideline

M & F: 0, 5, 25, 200 mg/kg/day (capsules)	NOAEL 25 mg/kg/day

LOAEL 200 mg/kg/day based on hematological effects, hemosiderosis of the
liver and spleen, extraniedullary hematopoiesis in the spleen, and
increases in liver weight.

870.4300

Chronic toxicity/ carcinogenicity in rodents (mice)	44014325 (1996)

Acceptable/guideline

M: 0, 1.14, 11.1, 237, 698 mg/kg/day

F: 0, 1.14, 10.8, 234, 696 mg/kg/day	NOAEL 11.1/10.8 mg/kg/day, M/F

LOAEL 237/234 mg/kg/day based on mild hemolytic anemia and related
microscopic changes (hemosiderosis) in the spleen, liver, and bone
marrow and extramedullary hematopoiesis in the spleen.

870.4200

Carcinogenicity study in rat	44014243 (1996)

Acceptable/guideline

M: 0, 0.77, 7.77, 96.9, 312 mg/kg/day

F: 0, 0.90, 9.08, 111, 388 mg/kg/day	NOAEL 96.9/111 (M/F)

LOAEL 312/388 / 191.0 mg/kg/d (M/F) based on decreased body weights,
body weight gain and food efficiency, mild hemolytic anemia, and
increased incidence of alveolar foam cells (females only).

870.5100

Bacterial reverse mutation assay (Ames test)	441014247 (1993)

Acceptable/guideline

312.5, 625, 1250, 2500, 5000 g/plate	Negative with and without S-9
activation at 5000 g/plate and less.

870.5100

Bacterial reverse mutation assay (Ames test)

Test Material: CGA- 362020 (isomer of acibenzolar-S-methyl)	44537025
(1998)

Acceptable/guideline

61.73 to 5000 g/plate (+S-9)

30-86 to 2500 g/plate (-S-9)	Positive in S. typhimurium strain TA1537
at 277.8 g/plate and higher in the absence of S-9.  Negative with S-9
activation at 5000 g/plate and less.

870.5100

Bacterial reverse mutation assay (Ames test)

Test Material: NOA- 419191 (by-product of acibenzolar-S-methyl)	44537024
(1998)

Acceptable/guideline

312.5 to 5000 g/plate (± S-9)	Negative with or without S-9
activation at 5000 g/plate and less

870.5100

Bacterial reverse mutation assay (Ames test)

Test Material: CGA- 323060 (plant metabolite of acibenzolar-S-methyl)
44537026 (1997)

Acceptable/guideline

312.5 to 5000 g/plate (± S-9)	Negative with or without S-9
activation at 5000 g/plate and less

870.5300

In vitro mammalian gene mutation assay 	44014246 (1993)

Acceptable/guideline

3.70 to 100 g/ml (-S-9), 37.04 to 1000 g/ml (+S-9)	Negative with
S-9 activation up to 1000g/ml.   Negative without S-9 activation up
to 100g/ml.

Compound tested to cytotoxic concentrations.

870.5375

In vitro mammalian chromosome aberration (CHO cells)	44014245 (1993)

Acceptable/guideline

7.5 to 60 g/ml (-S-9 and +S- 9)	Suggestive of clastogenicity in the
absence of S-9 activation at 30 and 60 g/mL at the 18-hour cell
harvest time; effect observed only in the presence of cytotoxicity. 
Increase in polyploid cells at 30 and 60 g/mL at the 42 hour harvest
time both with and without S-9.  Evidence of cell cycle arresting
activity at G2.

870.5395

Mammalian erythrocyte micronucleus test	44014244 (1993)

Acceptable/guideline

g/ml	Negative at 500 g/ml and less.

870.7485

Metabolism and pharmacokinetics

rats	44014250 (1995)

Acceptable/guideline

0.5, 100 mg/kg	Following oral treatment of rats, acibenzolar-S-methyl
was rapidly and nearly completely (>90% of administered dose) absorbed
from the gastrointestinal tract into the general circulation.   The
majority  (88- 95%) of the administered dose was excreted in the urine
within the first 48 hours.  The major metabolite (79- 92%) in the urine
was the carboxylic acid derivative of the parent.

870.7600

Dermal penetration	NA	NA

Special studies:

28-Day dietary 

rats

	

44014227 (1992)

Acceptable/nonguideline

M: 0, 45.9, 403, 1070 mg/kg/day; F: 0, 44.8, 376, 1000 mg/kg/day	

NOAEL =  M: 403 mg/kg/day; F: 376 mg/kg/day

LOAEL = M: 1070 mg/kg/day; F: 1000 mg/kg/day based on decreased mean
body weights, decreased liver weights, altered hematology parameters
accompanied by increased spleen weights.

28-Day oral gavage

rats	44014229 (1993) 

Acceptable/nonguideline

0, 10, 100, 800 mg/kg/day 	NOAEL = 100 mg/kg/day

LOAEL = 800 mg/kg/day based on decreased body weights, and decreased
hemoglobin-related parameters accompanied by hemosiderosis of the
spleen, increased liver and spleen weights, and decreased thymus
weights.

28-Day oral capsule dogs	44014231 (1994) 

Acceptable/nonguideline

0, 50, 250, 500 mg/kg/day	NOAEL = 50 mg/kg/day

LOAEL = 250 mg/kg/day based on decreased body weight, decreased
hemoglobin-related parameters, hepatic and splenic hemosiderosis.

90-Day Dietary 

mice	44014228 (1993)

Acceptable/nonguideline

M: 0, 30.6, 152, 624 mg/kg/day; F: 0, 47.4, 220, 803 mg/kg/day	NOAEL = 
M: 30.6 mg/kg/day; F: 47.4 mg/kg/day

LOAEL = M: 152 mg/kg/day; F: 220 mg/kg/day based on decreased mean body
weights and body weight gain in males, increased spleen weights and
splenic histopathology in both sexes. 

90-Day Dietary 

dog	44014232 (1994)

Acceptable/nonguideline

0, 10, 50, or 200 mg/kg/day	NOAEL =  50 mg/kg/day

LOAEL = 200 mg/kg/day in male and female dogs, based on regenerative
hemolytic anemia evidenced by decreased erythroid parameters,
pigmentation in the liver and spleen, splenic congestion, and by
activation of medullary (bone marrow) and extramedullary (splenic)
erythropoiesis.

90-Day Dietary 

rat	44014230 (1993)

Acceptable/nonguideline

males: 0,2.42, 24.6, 126, or 516 mg/kg/day; females: 0,2.64,26.3, 131,
or 554 mg/kg/day)	NOAEL = 126 mg/kg/day in males and 131 mg/kg/day

in females 

LOAEL = 516 mg/kg/day males and 554 mg/kg/day females based on decreased
mean body weights, reduced food intake, reduced food efficiency, and
increased liver and spleen weights with correlates for the liver and
spleen of glycogen deposition and hemosiderosis, respectively. 

Special

Developmental toxicity 

rats	44014238 (1994)

Acceptable/nonguideline

300 mg/kg/day, GD 6-15, 6-7, 8-9, 10-11, 12-13, or 14-15	Maternal and
developmental NOAELS and LOAELS could not be identified by this
protocol.  The most pronounced maternal and developmental toxicity
occurred when dams were treated on GD 6-15.

Special

Developmental toxicity

rats	44014239 (1994)

Acceptable/nonguideline

400 mg/kg/day, GD 6-7, 8-9, 10-11, 12-13, or 14-15	Maternal and
developmental NOAELS and LOAELS could not be identified by this
protocol.  The most pronounced maternal and developmental toxicity
occurred when dams were treated on GD 6-7 and 8-9.

Dermal developmental toxicity

rats	44014240 (1994)

Acceptable/nonguideline

0, 10, 100, 500 mg/kg/day, GD 6-15	Maternal NOAEL 500 mg/kg/day

LOAEL >500 mg/kg/day based on no effects.

Developmental NOAEL 500 mg/kg/day

LOAEL >500 mg/kg/day based on no effects.

Range-finding 

1-generation reproduction 

rats	44681301 (1993)

Acceptable/nonguideline

0, 199-209, 382-410, 700-728 mg/kg/day

	Parental/Systemic NOAEL = 209 mg/kg/day

LOAEL = 410 mg/kg/day based on decreased body weight gain and food
consumption in females.

Reproductive NOAEL = 410 mg/kg/day

LOAEL = 728 mg/kg/day based on total resorptions in all dams.

Offspring NOAEL = 209 mg/kg/day

LOAEL = 410 mg/kg/day based on reduced pup body weight gains and lower
pup body weights during lactation.

2.0	Studies Reviewed for Ethical Conduct

The PHED Task Force, 1995.  The Pesticide Handlers Exposure Database,
Version 1.1.  Task Force members Health Canada, U.S. Environmental
Protection Agency, and the National Agricultural Chemicals Association,
released February, 1995.

Agricultural Re-entry Task Force (ARTF) data base (SOP #3.1)

3.0	ADDITIONAL TOXICOLOGY STUDIES 

870.3700  Developmental Toxicity

In a non-guideline developmental toxicity study (MRID 44014238), twelve
presumed pregnant Tif:RAT f(SPF) (hybrids of RTT/1 x RII/2) rats per
group were administered CGA 245704 Technical (97.9%; Batch No. P.303011)
in a 0.5% aqueous solution of sodium carboxymethylcellulose by gavage at
a dose of 300 mg/kg/day. Groups 2-7 were administered test article on
gestation days (GD) 6-15,6- 7, 8-9, 10-11, 12-13, or 14-15,
respectively. Group 1 consisted of twelve animals which served as
controls and were treated with 0.5% sodium carboxymethylcellulose on GD
6-15. On GD 21, dams were sacrificed, subjected to gross necropsy, and
all fetuses examined externally. Visceral and skeletal examinations were
not performed.  All animals survived until scheduled sacrifice on GD 21.
Bloody, vaginal discharge was observed in 8/12 dams in Group 2, 3/12
dams in Group 3, 5/12 dams in Group 4, 4/12 dams in Group 5, and in 5/12
dams in Group 6. This finding was observed for one to four days starting
on GD 13 and was not observed in controls or in animals in Group 7. No
other treatment-related clinical signs of toxicity were observed. No
statistically significant differences in absolute body weights or body
weight changes occurred at any time between the treated groups and the
control group. Body weight gain by Group 2 was 80% of the control level
during the treatment interval (GD 6- 16), with the greatest reduction in
weight gain (77% of controls) during GD 11-16. Food consumption was not
affected. There were no statistically significant differences between
control and treatment groups for number of corpora lutea, number of
implantation sites, live fetuses/dam, pre- and post-implantation loss,
or fetal sex ratios. Fetal body weights from the Group 2 dams were
significantly (p <0.01) less than the controls. Although not
statistically significant, Group 2 dams also had fewer live fetuses per
litter (10.9 vs 14.8 for controls) and a greater post implantation loss
(30.1% vs 2.9% for controls) from early resorptions due to one dam with
whole liner resorption (17/17) and 2 additional dams with 13/16 and 8/9
resorptions/number of implantation sites. The overall incidence rates
for litters containing fetuses with major external malformations in
Groups 1-7 were 0/12, 1/10, 0/10, 1/11, 0/11, 0/10, and 0/12,
respectively. In one Group 2 litter, generalized edema was observed in
5/14 fetuses. In addition, two of the five also had a position anomaly
of the hindlimb and one of the two also had gastroschisis. These five
fetuses also had greatly reduced body weights. Generalized edema was
also seen in one fetus from Group 4.  Maternal and developmental
toxicity NOAELs and LOAELs could not be identified by this protocol,
which investigated the critical period of organogenesis. The most
pronounced maternal and developmental toxicity occurred when dams were
treated on GD 6-15, inclusive at a dose level of 300 mg/kg/day. This
study is classified as Acceptable/non-guideline as a developmental
toxicity study (83-3a) in rats.

In a non-guideline developmental toxicity study (MRID 44014239), eight
presumed pregnant Tif: RAT f (SPF) (hybrids of RII/1 x RII2) rats per
group were administered CGA 245704 Technical (97.9%; Batch No. P.303011)
by gavage at a dose of 400 mg/kg/day. Groups 2-6 were administered test
article on gestation days (GD) 6-7, 8-9, 10-11, 12-13, or 14-15,
respectively. Group 1 consisted of eight animals which served as
controls and were treated with 0.5% carboxymethylceliulose (vehicle) on
GD 6-15. On GD 21, dams were sacrificed, subjected to gross necropsy,
and all fetuses examined externally. Visceral and skeletal examinations
were not performed.  One Group 2 dam was found dead on GD 20. Prior to
death, this animal had bloody perineal discharge and necropsy findings
including mottled kidney and thymus. One Group 4 dam delivered on GD 20
and was sacrificed. All other animals survived until scheduled sacrifice
on GD 21. Bloody, perineal discharge was observed in 4/8 dams in Group
3. This finding was observed for three to four days starting on GD 16.
The same 4 dams were later observed to have total lifter resorptions. No
other treatment-related clinical signs of toxicity were observed. No
statistically significant differences in body weights, body weight
gains, or food consumption occurred at any time between the treated
groups and the control group.  There were no differences between the
control group and Groups 4, 5, and 6 for number of corpora lutea, number
of implantation sites, live fetuses/dam, pre- and post-implantation
loss, fetal body weights, or fetal sex ratios. Group 2 had a slightly
greater number of early resorptions/dam (2.7 vs 0.9 for controls) and an
increased postimplantation loss (18.7% vs 5.4% for controls). The four
Group 3 dams observed with bloody discharge had complete litter
resorption resulting in increased postimplantation loss (53.4%). Group 3
also had an increase in early rcsorptions (8.0/dam vs 0.9/dam for
controls) and decreased mean live fetuses/litter (8.5 vs 16.0 for
controls). Another Group 3 dam had fused placentae between two fetuses. 
The overall incidence rates for liners containing fetuses with major
external malformations in Groups 1-6 were 0/8, 0/6, 0/4, 1/7, 0/8, and
0/8, respectively. One Group 4 fetus had cleft palate and another fetus
from the same lifter had a kinked tail. No treatment-related external
malformations /variations were observed in any fetus from any group.
Maternal and developmental toxicity NOAELs and LOAELs could not be
identified by this protocol, which investigated the critical period of
organogenesis. Pronounced maternal and developmental toxicity occurred
when dams were treated on GD 6-7 and 8-9 at a dose level of 400
mg/kg/day.

870.7600 Dermal Penetration

In a dermal developmental toxicity study (MRID 44014240), 24 pregnant
Tif: RAIf (SPF) rats (Sprague-Dawley derived) per group were
administered acibenzolar-S-methyl (97.9% a.i.) in aqueous
carboxymethylcellulose (0.5% w/w) in 0.1% aqueous Polysorbate 80, by
dermal application at doses of 0 (vehicle), 10, 100, or 500 mg/kg/day on
gestation days (GD) 6-15, inclusive.  Exposure duration was 6 hours per
day. On GD 21, all dams were sacrificed and all fetuses were sexed,
weighed, and examined for external malformations/variations, then
sacrificed. Approximately one-half of each litter was placed in
Bouin’s fixative for subsequent visceral examination and the remainder
were prepared and stained for skeletal examination.  All animals
survived until scheduled sacrifice.  No clinical signs of toxicity were
observed at any treatment level.  Gestational body weights, body weight
changes, and food consumption were not statistically significantly
different for any treatment level throughout the gestational period.

Therefore, the maternal toxicity NOAEL is 500 mg/kg/day based on no
adverse maternal effects at the highest administered dose.  The maternal
toxicity LOAEL was >500 mg/kg/day.  No treatment-related effects were
observed for gravid uterine weights, number of fetuses/litter, pre- and
post-implantation losses, numbers of corpora lutea/dam, number of
implantations/dam, resorptions/dam, fetal body weights, or fetal sex
ratios.  No biologically relevant statistically significant differences
in the incidence rates of any external, visceral, or skeletal
malformations/ anomalies/variations were observed in the treated litters
as compared to the controls.  There were statistically significantly
decreased incidences of absent ossification of the proximal phalanx of
anterior digit-2 in all treatment groups and poor ossification of the
proximal phalanx of posterior digit-2 in the 500 mg/kg/day treatment
group as compared to the control group. However, these decreases were
not considered to be biologically significant.  Therefore, the
developmental toxicity NOAEL is 500 mgfkglday based on no adverse
developmental effects at the highest dose administered. The
developmental toxicity LOAEL was >500 mg/kg/day.

In a 28- day repeated dose dermal toxicity study (MRID 44014233), groups
of 5 females Tif: RAIf (SPF) albino rats were treated with
acibenzolar-S-methyl technical in 0.5% (w/v) carboxymethyl
cellulose/0.1% (w/v) aqueous polysobate 80 at does of 0, 1, 10, 100 or
1000 mg/kg/day.  Animals were treated by dermal occlusion for 6
hours/day, 5 days/week for 4 weeks.  No animals died during the study. 
There were no treatment- related clinical signs, dermal effects, effects
on body weight, food consumption, hematology, clinical chemistry, or
organ weight.  No gross or microscopic pathology were noted at necropsy.
 The systemic and dermal NOAEL is the limit dose of 1000 mg/kg/day.  A
dermal and systemic LOAEL were not identified.  

870.3800 Reproduction

In a 1-generation range-finding reproduction study in rats (Nonguideline
study, MRID 44014241)

CGA 245704 Technical (97.9% a.i.; Batch No. P.303011) was administered
to groups of 15 male and 15 female Tif: RAT f (SPF), hybrids of RTTI 1 x
P11/2 (Sprague-Dawley derived) rats at dietary concentrations of 0,
2000, 4000, or 8000 ppm for one generation (MRID 44681301). In this
range- finding study, F0 adults were administered test or control diet
for two weeks prior to mating. Premating doses were 199.4, 382.0, and
700.1 mg/kg/day, respectively, for F0 males and 208.7, 409.9, and 727.8
mg/kg/day, respectively, for F0 females. Males were terminated at the
end of mating and females were terminated at weaning of their pups. All
offspring, except five litters per dose group, were necropsied at
weaning. After weaning, five litters per dose group were retained on
treatment for an additional three weeks and then necropsied.  The only
clinical sign of toxicity in the parental animals was perineal
hemorrhage during late gestation in five high-dose females. Gross
necropsy was unremarkable. Absolute body weights, body weight gains, and
food consumption values for the low- and mid-dose males and low-dose
females were similar to the controls throughout the study. High-dose
males had significantly (p > 0.01; 88-91% of control) lower body weights
than the controls beginning on day 8 and continuing until study
termination. Overall weight gain by the high-dose males was 69% of the
control level. Body weights and body weight gains of the high-dose
females were significantly (p > 0.01; body weights 88-90% of controls)
Tess than the controls during the premating interval. Overall body
weight gain by the high- dose group was only 30% of the control group
level. Mid-dose females had significantly (p > 0.05) lower body weight
gains than the controls during days 1-8. Reduced body weight gains in
the high- dose males and females and mid-dose females correlated with
significantly (p > 0.05 or 0.01) reduced food consumption by these
treated groups during study days 1-8. Lower body weights, body weight
gains, and food consumption continued for the mid- and high-dose females
during gestation and lactation. Body weights of the high-dose group were
significantly (p > 0.05 or 0.01) lower than the controls throughout
gestation with overall weight gain by the high-dose females during
gestation 27% (p > 0.01) of the control group level. Mid-dose females
had significantly (p > 0.05 or 0.01) lower body weights than the
controls on GD 21 and on lactation days 7, 14, and 21. Weight gains by
the mid-dose group during gestation and lactation were 92% and 83%,
respectively of the control group levels. Food consumption by the
high-dose group was significantly (p > 0.05 or 0.01) less than the
controls throughout gestation. Food consumption by the mid-dose group
was significantly (p > 0.01) less than the controls throughout
lactation. Therefore, the LOAEL for parental toxicity is 4000 ppm based
on reduced food consumption and lower body weight gains in F0 females.
The parental toxicity NOAEL is 2000 ppm.  Mating and fertility indices,
mean precoital interval, mean gestation length, and mean numbers of
corpora lutea and implantation sites per dam were similar between the
treated and control groups. Mating and fertility indices for the
high-dose group were similar to the control group, but no live lifters
were produced by the high-dose group (i.e. all pregnancies were lost).
Live births and pup survival were similar between the low- and mid-dose
groups and the control group. Therefore, the LOAEL for reproductive
toxicity is 8000 ppm based on total resorptions in all dams. The
reproductive toxicity NOAEL is 4000 ppm.  No treatment-related clinical
signs of toxicity were observed in the pups during lactation. Absolute
body weights of pups in the low-dose group were similar to the controls
throughout lactation. Body weights of the mid-dose pups were
significantly (p > 0.05 or 0.01; 76-89% of controls) less than the
controls beginning on lactation day 7 and continuing throughout
lactation and postweaning. The lower body weights of the mid-dose pups
were a result of significantly (p > 0.05 or 0.01; 69-89% of controls)
lower body weight gains from postnatal day 4 through postnatal day 28.
Recovery of body weight gains occurred during the postweaning intervals
of days 28-35 and 35-42. Therefore, the LOAEL for developmental toxicity
is 4000 ppm based on reduced pup body weight gains and lower pup body
weights during lactation and early postweaning. The developmental
toxicity NOAEL is 2000 ppm.

870.6200b Subchronic Neurotoxicity

In a subchronic neurotoxicity study (MRID 45713601), CGA-245704
(Acibenzolar, 97.9% a.i., Batch No. P.303011) was administered in the
diet to 10 Sprague-Dawley derived rats/sex/group at dose levels of 0,
400, 2000, or 8000 ppm (equivalent to 0/0, 24/26, 126/143, and 575/628
mg/kg/day [M/F], respectively) for at least 90 days.  Neurobehavioral
assessment (functional observational battery [FOB] and motor activity
testing) was performed using all rats at pre-dosing and Weeks 4, 8, and
13.  At study termination, 5 rats/sex/group were anesthetized and
perfused in situ for neuropathological examination.  The tissues from
the perfused animals in the control and 8000 ppm groups were subjected
to histopathological evaluation of brain and peripheral nervous system
tissues.  Positive control data were not provided. No compound-related
effects were observed in mortality, clinical signs of toxicity, FOB,
motor activity, or gross or neuropathology. At 8000 ppm, body weights
were generally decreased throughout the study in the males (decr 11-14%,
not statistically significant) and females (decr 8-13%, attaining
statistical significance at Weeks 2-5, 7 and 8, and 11-13).  Also at
this dose, cumulative body weight gains were decreased throughout the
study in the males (decr 19-35%, attaining statistical significance at
Weeks 2-4 and 6-9), and in the females (decr 24-40%).  A decreasing
trend (p<0.01) in body weight and body weight gain was also generally
noted in both sexes at this dose.  These decreases in body weight gain
corresponded with decreases in food consumption (g/animal/day) which was
sporadically decreased in the 8000 ppm males (decr 9-21% at Weeks 1-3,
5-7, and 11, but only attaining statistical significance during Week 1),
and females (decr 14-18%, p<0.01 at Weeks 1 and 3).  A decreasing trend
(p<0.01) in food consumption was also generally noted in the males at
8000 ppm during the weeks reported above. No neurological effects were
observed at any dose in either sex.  The LOAEL was 8000 ppm (equivalent
to 575/628 mg/kg/day, M/F) based on decreased body weights, body weight
gains, and food consumption.  The NOAEL is 2000 ppm (equivalent to
126/143 mg/kg/day, M/F).  This study is classified as
acceptable/nonguideline pending submission of positive control data for
neuropathological and FOB assessments and does not satisfy the guideline
requirement (OPPTS 870.6200b) for a subchronic neurotoxicity study in
rats.  The study may be upgraded pending submission of acceptable
positive control data.

870.5100, 870-5300, 870.5375, 870.5900  Mutagenicity 

Five acceptable mutagenicity studies on technical grade
acibenzolar-S-methyl are available:

1. 	Reverse gene mutation, S. typhimuriumlE. Coli (MRID 44014247)

2. 	Forward gene mutation, Chinese hamster V79 lung cells in
culture/HGPRT locus (MRID

	44014246)

3. 	Chromosome aberration, Chinese hamster ovary (CHO) cells in culture
(MRID 44014245)

4. 	Unscheduled DNA synthesis assay, primary rat hepatocytes in culture
(MRID 44014248)

5. 	In vivo cytogenetics, micronucleus assay in mice (MRID 44014244)

	Results in all five studies were negative for genotoxicity. The five
studies on the technical grade product satisf’ the new revised
mutagenicity guideline requirements for a new chemical (published in
1991). 

Three additional acceptable mutagenicity studies on a) an isomer
(CGA-362020) of acibenzolar S-methyl, b) a by-product (NOA 419191) of
acibenzolar-S-methyl, and c) a plain metabolite (CGA-323060) of
acibenzolar-S-methyl are also available.

1. 	Reverse gene mutation, S. typhimuriumlE. Coli; Test Material:
CGA-362020 (MRID

	44537025). This study was POSITIVE for evidence of induced mutant
colonies over background in S. typhimurium strain TA1537 at
concentrations of 277.8 and higher in the absence of 59-mix.

2. 	Reverse gene mutation, S. typhimuriumlE. Coli; Test Material: NOA
419191 (MRID

	44537024). This study was negative.

3. 	Reverse gene mutation, S. typhimuriumfE. Coli; Test Material:
CGA-323060 (MRlD

	44537026). This study was negative

CGA-362020, an isomer of acibenzolar-S-methyl, is a low level impurity
arising from a new production process for acibenzolar-S-methyl, which
has not yet been submitted to the Agency. Trace levels of this isomer
(<0.1%) were found in some batches of technical grade
acibenzolar-S-niethyl produced via the new production process. See the
6(a)(2) letter from Novartis, dated 3/16/98, reporting the findings in
this mutagenicity study and the above information regarding the
formation of this isomer in the new production process. Presumably, the
same isomer does not occur in the old production process which produced
the test material (lot/batch # P.303011) that was used for virtually all
the toxicity testing on acibenzolar-S-methyl up to this point in
time.4.0	TOXICOLOGY STUDY REFERENCES 

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inding Rat Oral Teratogenicity: (Non-Standard Study): (Final Report):
Lab Project Number: 943004. Unpublished study prepared by Ciba-Geigy
Ltd. 118 p. 

44014240	Khalil, S. (1994) CGA-245704 Technical: Rat Dermal
Teratogenicity: (Final Report): Lab Project Number: 943001: 370440.
Unpublished study prepared by Ciba-Geigy Ltd. 381 p. 

45089701	Miyamoto, M. (1998) Teratology Study of Acibenzolar-S-methyl in
Rats: Final Report: Lab Project Number: AN98025: 1197-98. Unpublished
study prepared by Ina Research Inc. 131 p. {OPPTS 870.3700} 

44014241	Khalil, S. (1995) CGA-245704 Technical: Two-Generation
Reproduction Study in Rats: (Final Report): Lab Project Number: 923152.
Unpublished study prepared by Ciba-Geigy Ltd. 1628 p. 

44681301	Marty, J. (1993) CGA-245704 Technical: Rangefinding Rat Dietary
Reproduction Study: Lab Project Number: 923146: 634-93. Unpublished
study prepared by Novartis Crop Protection, Inc. 65 p. 

44014244	Hertner, T. (1993) CGA-245704 Technical: Micronucleus Test:
Mouse (OECD Conform) in vivo Study: Lab Project Number: 923143.
Unpublished study prepared by Ciba-Geigy Ltd. 40 p. 

44014245	Hertner, T. (1993) CGA-245704 Technical: Cytogenetic Test on
Chinese Hamster Cells in vitro (EC-Conform): Lab Project Number: 923142.
Unpublished study prepared by Ciba-Geigy Ltd. 45 p. 

44014246	Geleick, D. (1993) CGA-245704 Technical: Gene Mutation Test
with Chinese Hamster Cells V79 in vitro: Lab Project Number: 923141.
Unpublished study prepared by Ciba-Geigy Ltd. 53 p. 

44014247	Hertner, T. (1993) CGA-245704 Technical: Salmonella and
Escerichia/Liver Microsome Test: Gene Mutation Test: Lab Project Number:
923145. Unpublished study prepared by Ciba-Geigy Ltd. 67 p. 

44014248	Hertner, T. (1993) CGA-245704 Technical: Autoradiographic DNA
Repair Test on Rat Hepatocytes (OECD Conform) in vitro: Lab Project
Number: 923144. Unpublished study prepared by Ciba-Geigy Ltd. 86 p. 

44537024	Deparade, E. (1998) Salmonella and
Escherichia/Mammalian-Microsome Mutagenicity Test: NOA 419191 Technical:
Lab Project Number: 973097: 324-98: A98013. Unpublished study prepared
by Novartis Crop Protection, AG. 75 p. 

44537025	Deparade, E. (1998) Salmonella and
Escherichia/Mammalian-Microsome Mutagenicity Test: CGA-362020 Technical:
Lab Project Number: 973079: 325-98. Unpublished study prepared by
Novartis Crop Protection, AG. 74 p. 

44537026	Ogorek, B. (1997) Salmonella and
Escherichia/Mammalian-Microsome Mutagenicity Test: CGA-323060 Technical:
Lab Project Number: 963125: 761-97: A960043. Unpublished study prepared
by Novartis Crop Protection, AG. 74 p. 

45434101	Deparade, E. (1998) Salmonella and
Esherichia/Mammalian-Microsome Mutagenicity Test: CGA-245704: Final
Report: Lab Project Number: 983053: 1240-98. Unpublished study prepared
by Novartis CropProtection AG. 71 p. {OPPTS 870.5100} 

44014250	Molitor, E. (1995) The Metabolism of (U-(carbon 14))Phenyl
CGA-245704 in the Rat: Lab Project Number: 13/94: 016AM03. Unpublished
study prepared by Ciba-Geigy Ltd. 197 p. 

 	CHEMICAL NAMES AND STRUCTURES OF METABOLITES

Page  PAGE  1  of  NUMPAGES  49 

Acute RfD  (Females 13-49) =  (NOAEL) 8.2 mg/kg =  0.082 mg/kg

                                                          (UF) 100	

         Chronic RfD  =    25 mg/kg/day (NOAEL) = 0.25 mg/kg/day

 (General Population)	           100 (UF)

Chronic RfD  (Females 13-49) =  (NOAEL) 8.2 mg/kg =  0.082 mg/kg

                                                          (UF) 100