Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 3 of 176 5.1.2 Summary of Environmental Degradation

................................ ............................... 32

5.1.3 Comparison of Metabolic Pathways

................................ ................................ ....... 33

5.1.4 Residues of Concern Summary and Rationale

................................ ........................ 33

5.2 Food Residue Profile

................................ ................................ ................................ ..... 36

5.3 Water Residue Profile

................................ ................................ ................................ ... 36

5.4 Dietary Risk Assessment

................................ ................................ .............................. 41

5.4. 1 Description of Residue Estimates Used in Dietary Assessment

............................. 41

5.4.2 Percent Crop Treated Used in Dietary Assessment

................................ ................ 41

5.4.3 Acute Dietary Risk Assessment

................................ ................................ .............. 41

5.4.4 Chronic Dietary Risk Assessment

................................ ................................ .......... 42

5.4.5 Cancer Dietary Risk Assessment

................................ ................................ ............ 42

5.4.6 Summary Table

................................ ................................ ................................ ....... 42

6.0 Residential (Non

- Occupational) Exposure/Risk Characterization

................................ ... 43

6.1 Residential Bystander Post

- Application Inhalation Exposure

................................ ...... 43

6.2 Spray Drift

................................ ................................ ................................ .................... 43

7.0 Aggregate Exposure/Risk Characterization

................................ ................................ ...... 44

8.0 Cumulative Exposure/Risk Characterization

................................ ................................ .... 44

9.0 Occupational Exposure/Risk Characterization

................................ ................................ . 44

9.1 Short

- and Intermediate

- Term Handler Risk Estimates

................................ ................ 45

9.2 Short

- and Intermediate

- Term Post

- Application Risk Estimates

................................ . 50

9.2.1 Dermal Post

- Application Risk Estimates

................................ ................................ 50

9.2.2 Inhalation Post

- Application Risk Estimates

................................ ........................... 53

10.0 References

................................ ................................ ................................ ......................... 54

Appendix A. Toxicology Profile and Executive Summaries

................................ ....................... 56

Appendix B. Sulfoxaflor Mode

- of Action for Reproductive and Developmental Effects

........ 132

Appendix C. Metabolism Summary Table

................................ ................................ ................. 156

Appendix D. Physical/Chemical Propert ies and Summaries of Field Trial and Processing Study

Results

................................ ................................ ................................ ................................ ......... 158

Appendix E. Review of Human Research

................................ ................................ ................. 166

Appendix F. Occupational Exposure/Risk Summary Tables

................................ .................... 167

Appendix G. Anticipated Post

- Application Activities and Dermal Transfer Coefficients

........ 174Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 4 of 176 1.0 Executive Summary

Dow AgroSciences has petitioned the Agency to grant registrations and establish tolerances for

the new insecticide sulfoxaflor { CAS name: cyanamide, N

- [methyloxido[1

- [6

- (trifluoromethyl)

- 3

- pyridinyl]ethyl ] λ 4

- sulfanylidene] }

. Sulfoxaflor is the first member of a new class of insecticides, the sulfoximines , which act through a unique interaction with the nicotinic acetylcholine receptor (nAChR) in insects. Sulfoxaflor is a highly efficacious agonist of that
receptor

. The registration request is for one technical

- grade product (File Symbol 62719

- AGR )

and two end

- use products, TransformTM WG ( 62719

- AEL ) and GF

- 2032 SC ( 62719

- AEG ). The requested use sites are identical for the two end

- use products and incl ude numerous agricultural crops (39 crops/crop groups in all), turfgrass (sod farms only), and ornamental plants. At this time, no uses have been requested that would result in residential exposure.

T he toxicology database for sulfoxaflor is complete for risk assessment purposes, including the assessment of potential increased susceptibility of infants and children. Toxicity and mechanistic studies in rats, rabbits, dogs and mice indicate that the liver and the nervous systems

are target organ systems. Liver effects in subchronic and chronic studies include organ weight and enzyme changes, hypertrophy, proliferation, and tumors ; long

- term studies resulted in hepatic effects at doses lower than in short

- term studies

. Based on the weight of evidence

, incl uding mode

- of

- action data , the C ancer A ssessment R eview C ommittee (CARC) determined that there is a “Suggestive Evidence of Carcinogenic Potential” for sulfoxaflor and that quantification of ri sk using a non

- linear approach [ i.e., reference dose (RfD) ] wil l adequately account for all chronic toxicity, including carcinogenicity, that could result from exposure to

sulfoxaflor. The nervous system effects were observed as frank neurotoxicity as well as secondary developmental effects due to prolonged contractu re of fetal muscle resulting in limb

flexure and bent clavicle in fetuses

, and decreased survival of neonatal rats. As these developmental effects were observed only in rats, the petitioner conducted numerous non

guideline studies to investigate the cause of these effects and to determine the ir human relevance. Those studies are adequate to delineate the mode of action of the developmental effects and provide information regarding interspecies uncertainty. The database indicates that the developmental ef

fects are unlikely to be relevant to humans; however, an additional study is required in order to make a final conclusion. Therefore, p ending that study , the developmental

effects are considered to be relevant to humans . Data are sufficient to support re ducing the interspecies uncertainty factor to 3X for the developmental effects

. The developmental, neuromuscular, or liver effects are the basis for this risk assessment, depending on the duration of exposure and the population group of interest.

HED has recommended that the required 10X FQPA Safety Factor be reduced to 1X based on the completeness of the toxicological database, the adequacy of the submitted neurotoxicity and developmental neurotoxicity studies, the selection of endpoints and doses select ed for risk assessment that are protective of observed developmental effects, special studies to address HED’s concern regarding human relevance for the developmental effects, and exposure

assessments that are unlikely to underestimate exposure

.

No direct residential uses or commercial/professional uses at residential sites are being requested

for sulfoxaflor at this time. Although t here is a proposed use on ornamental plants

, the proposedSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 5 of 176 use is only for nurseries and commercial greenhouses. Furthermore , exposure from homeowners buying turfgrass (sod) and planting it as a residential turf would be minimal. At this time, there are no uses of sulfoxaflor that will result in residential exposures. Therefore, i f the requested registrations are granted, exp osure to sulfoxaflor may occur via residues in food and/or drinking

water. Data depicting the fate of sulfoxaflor in plants

, animals , and the environment indicates

that in systems that include an aerobic soil component (e.g., soil

- directed applications, rotational crops, groundwater systems), breakdown of sulfoxaflor is quite extensive and rapid, with very little parent compound remaining. In those scenarios, the metabol ite designated X11719474 is the major component. Contrary to that, the parent compound undergoes very little

transformation in animals or via hydrolytic or photolytic processes . Some metabolism of the parent compound occurs in plants following foliar app lication, resulting in two major metabolites (X11719474 and X11721061). Plant metabolism studies also demonstrate that residues are distributed throughout the plant and are not restricted to treated surfaces. In residue field studies using foliar treatme nt (which is the requested use pattern), residues of sulfoxaflor were generally well above the limit of quantitation (LOQ) of 0.01 ppm in most commodities. The exception to that is in root crops and rotational crops where residues were approximately at or below the LOQ ; p resumably due to the involvement of the soil matrix in those situations . T ypical food processing activities, such as baking, boiling and canning

, are unlikely to significantly affect the nature and magnitude of sulfoxaflor residues. In p rocessing studies, sulfoxaflor generally concentrated during production of dried foods and tended to be reduced in juices, though exceptions to those trends are evident in the database. In the case of drinking water, degradates are expected to be signific antly greater in groundwater than in surface water. In addition, the residue profile for these two drinking water sources are expected to be quite different, with residue in surface water being comprised of sulfoxaflor almost exclusively and residues in

g roundwater being made up primarily of the metabolites X11719474 and, to a lesser extent, X11519540; parent sulfoxaflor is expected to be absent from groundwater. The different residue profiles, combined with toxicological information and risk assessment e ndpoints for sulfoxaflor

versus these two metabolites, results in different strategies for risk assessment. In short, this assessment focuses on the following three combinations for evaluating aggregate exposure: Neuromuscular effects (acute dietary expos ure, general population): Assess sulfoxaflor in food and the metabolites designated X11719474 and X11519540 in groundwater; Developmental effects (acute dietary exposure, women of child

- bearing age; short

- and

intermediate

- term dermal and inhalation expos ure): Assess sulfoxaflor in food and surface water (note that at this time in the absence of residential uses, the dermal and inhalation exposure routes are not relevant to an aggregate assessment); Liver effects (chronic dietary exposure, all populations ): Assess sulfoxaflor in food and the metabolites designated X11719474 and X11519540 in groundwater, factoring in

information on the relative potency of the metabolites to elicit the liver effects. Relative to sulfoxaflor, these factors are 0.3X for X117 19474 and 10X for X11519540.

HED has used moderately refined, health

- protective estimates of residue levels in foods and

upper

- bound residue estimates for drinking water, taking into account the toxicological endpoint/residue profile considerations described above, in combination with an extensive database of food and water consumption for the U.S. population

, to estimate dietary exposure and risk that may result from the use of sulfoxaflor. Across all representative population subgroups, including those of infants and children, the maximum d ietary risk estimates are 16 %Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 6 of 176 and 18 % of the acute and chronic population

- adjusted dose (aPAD, cPAD) , respectively. These estimates are well below 100% and are, therefore, below HED’s level of concern. In the absence of residential use sites, aggregate risk e s timates are equivalent to dietary risk estimates and are, by extension, below HED’s level of concern. There are no aggregate risk issues that would preclude granting the requested registrations and establ ishing tolerances for residues of

sulfoxaflor at the levels recommended in Table 2.2.3.

HED has also estimated risks for persons exposed to sulfoxaflor during occupational activities.

These activities include application

- related tasks (mixing, loading, a pplying, etc.) as well as post

application tasks (scouting, weeding, irrigating, etc.). The current risk metric for assessing short

and intermediate

- term occupational exposure to sulfoxaflor is a margin of exposure (MOE) that

is less than 30

. At a basel ine level of personal protective equipment (PPE), e stimated occupational MOEs range from 80 to 4,700,000, and the vast majority of the estimates are greater than 300. All estimated occupational MOEs indicate that risks are below HED’s level of

concern.

Although the proposed labels specify a 24

- hour Restricted Entry Interval (REI) , the acute toxicity categories for technical

- grade sulfoxaflor indicate that a 12

- hour REI is appropriate to protect agricultural workers from post

- application exposures.

T his risk assessment relies

, in part

, on data from studies in which adult human subjects were intentionally exposed to a pesticide or other chemical. These data, which include the Pesticide Handlers Exposure Database Version 1.1 (PHED 1.1); the Agricultural H andler Exposure Task Force (AHETF) database; the Outdoor Residential Exposure Task Force (ORETF) database; the Agricultural Re

- entry Task Force (ARTF) database; other registrant

- submitted exposure studies ( MRID Nos. 47832415, 47832416 ); are subject to ethi cs review pursuant to 40 CFR 26, have received that review, and are compliant with applicable ethics requirements. For certain studies that review may have included review by the Human Studies Review Board. Descriptions of data sources as well as guidanc e on their use can be found at : http://www.epa.gov/pesticides/science/handler

- exposure

- data.html and http://www.epa.gov/pesticides/science/post

- app

- exposure

- data.html

.

2.0 HED Recommendations

HED is recommending for granting the registrations for use of sulfoxaflor on the requested crops and for the establishment of permanent tolerances for residues of sulfoxaflor as specified in

Section 2.2.3. HED further recommends that the labels be revised as specified in Section 2.3.

2.1 Data Deficiencies/Conditions of Registration

None.

2.2 Tole rance Considerations

2.2.1 Enforcement Analytical MethodSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 7 of 176 High performance liquid chromatographic ( HPLC ) methods with positive

- ion electro spray (ESI) tandem mass spectrometry (LC/MS/MS) were developed for data collection and enforcement of

sulfoxaflor re sidues and the two metabolites X11719474 and X11721061. Method 091116 was developed for plant commodities, and Method 091188 was developed for livestock commodities.

Both methods use solvent extraction (acetonitrile+water, 80+20, v/v), solid

- phase extrac tion clean up, and reverse

- phase LC

- MS/MS for separation and analysis. In addition, the plant method incorporates base

- and enzymatic

- hydrolysis steps to liberate conjugated residues. Successful validation of both methods was reported by the registrant a nd by an independent laboratory. Additionally, successful radiovalidation was reported for both methods. The lowest level of method validation (LLMV) was 0.01 mg/kg for each of sulfoxaflor, X11719474, and

X11721061 in all matrices. The limit of detectio n (LOD) was reported as 0.003 mg/kg for each analyte in all matrices.

FDA multiresidue methods are not suitable for analysis of sulfoxaflor; however, data were provided which indicate that the DFG S

- 19 multiresidue method may provide satisfactory

results.

2.2.2 International Harmonization

At this time, there are no MRLs for sulfoxaflor. As a result of the global joint review process, HED expects that sulfoxaflor MRLs covering the majority of the commodities addressed in this

assessment will be established by Australia, Canada and Codex.

T he U.S., Canada, and Australia have agreed on the residue definition for sulfoxaflor and the global partners are committed to harmonizing regulatory levels to the maximum extent allowed by each regulatory authority’s policies. In addition, we note that s ufloxaflor was presented at the Joint Meeting on Pesticide Residues (JMPR, 2011)

. As a pilot process to maximize harmonization, r ecommendations from that meeting were forwarded to the Codex Committee on Pesticide Residues (CCPR) prior to registration by any regulatory authority in an a ttempt to set
benchmark MRLs. To date, final CCPR recommendations have not been made and negotiations

are ongoing between the JMPR and CCPR . All tolerances recommended by HED are harmonized with the MRLs being recommended by Canada.

2.2.3 Recommended To lerances

HED recommends that 40 CFR be amended by establishing tolerances for residues of sulfoxaflor

in the plant and livestock commodities listed in the T able 2.2.3 . As the residue definitions for plant and animal commodities are the same , HED recommen ds that the same entr y be used for

both types of commodities. T he tolerances should be defined as follows:

Tolerances are established for residues of the insecticide sulfoxaflor , including its metabolites and degradates, in or on the commodities in the table below. Compliance with the tolerance levels specified below is to be determined by measuring only

sulfoxaflor ( N

- [methyloxido[1

- [6

- (trifluoromethyl)

- 3

- pyridinyl]ethyl]

- γ 4

- sulfanylidene]cyanamide )

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 8 of 176 Table 2.2.3. Tolerance Summary for Sulfoxaflor. Commodity Tolerance, ppm Comments (correct commodity definition) Proposed Recommended
Almond, hulls 4.0 6.0 Barley, grain 0.15 0.40 Barley, hay 0.8 1.0 Barley, straw 1.5 2.0 Tolerance is higher than the HAFT, but

lower than the maximum residue. Since

the CODEX Global MRL recommended by JMPR is 2.0 ppm, this value would be

harmonized. Bean, dry seed 0.25 0.20
Bean, succulent 0.7 4.0 Beet, sugar, dried pulp

-- 0.07 Beet, sugar, molasses 0.03 0.25 Berry, low growing, subgroup 13

- 07G 0.06 0.70

Cauliflower 0.08 0.08 Individual tolerance is required because it

exceeds a 5x factor. Citrus, dried pulp 0.9 3.60
Cotton, hulls 0.4 0.35 Cotton, gin byproducts 8.0 6.0 Cottonseed subgroup 20C 0.2 0.20 Fruit, citrus, group 10

- 10 0.6 0.70 Fruit, pome, group 11

- 10 0.4 0.50 Fruit, small, vine climbing, subgroup 13

- 07F, except fuzzy kiwi fruit 1.3 2.0

Fruit, stone, group 12 0.6 3.0 Grain, aspirated fractions

-- 20.0 Grape, raisin 5.0 6.0 The calculated tolerance was 5.4 ppm; however, HED recommends for a 6.0 ppm

tolerance in order to harmonize with pending CODEX Global MRL. Leafy greens, subgroup 4A 5.0 6.0 Leafy petiole, subgroup 4B 1.0 2.0 Nuts, tree, group 14 0.02 0.015 Onion, bulb, subgroup 3

- 07A 0.01 0.01 Onion, green, subgroup 3

- 07B 0.6 0.70

Pistachio 0.02 0.015 Pistachio requires a separate tolerance.

Data from tree nuts are translated to pistachio. Plum, prune, dried

-- 1.90 Processing factor for dried cherry used to

calculate concentration of residues. Rapeseed, meal 0.5 0.50 Rapeseed subgroup 20A 0.3 0.30 Soybean, seed 0.2 0.20 Soybean, hay 1.8 3.0 Soybean, forage 1.9 2.0 Tomato, paste 1.6 2.60 Tomato, puree 0.7 1.20 Vegetable , brassica , leafy, group 5, except cauliflower 1.0 (5A) 1.6 (5B) 2.0

Vegetable, cucurbit, group 9 0.3 (except 0.40Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 9 of 176 Table 2.2.3. Tolerance Summary for Sulfoxaflor. Commodity Tolerance, ppm Comments (correct commodity definition) Proposed Recommended
squash) Vegetable, fruiting, group 8

- 10 1.2 0.70 The calculated tolerance was 0.9 ppm; however, HED recommends for a 0.7 ppm

tolerance in order to harmonize with

CODEX. Vegetable, leaves of root and tuber, group 2 4.0 3.0 Based on sugar beet and radish top data

Vegetable, legume, foliage, group 7

-- 3.0 Vegetable, root and tuber, group 1 0.01 0.05

Watercress

-- 6.0 Translated from subgroup 4A data, not a member of subgroup 4A. Wheat, grain 0.07 0.08 Wheat, forage 0.8 1.0 Tolerance is higher than the HAFT, but

lower than the maximum residue. Wheat hay 1.1 1.50
Wheat, straw 2.0 2.0

Livestock Commodities *

Cattle, meat 0.1 0.15
Cattle, fat 0.04 0.10 Cattle, kidney 0.2

-- See Cattle, meat byproducts Cattle, meat byproducts 0.25 0.40

Milk 0.08 0.15

Milk fat NA

-- Goat, meat 0.1 0.15
Goat, fat 0.04 0.10 Goat, kidney 0.2

-- See Goat, meat byproducts Goat, meat byproducts (except kidney) 0.3

-- See Goat, meat byproducts
Goat, meat byproducts

-- 0.40 Hog, meat 0.01 0.01 Hog, fat 0.01 0.01 Hog, meat byproducts 0.04 0.01
Horse, meat 0.1 0.15
Horse, fat 0.04 0.10 Horse, kidney 0.2

-- See Horse, meat byproducts Horse, meat byproducts (except kidney) 0.3

-- See Horse, meat byproducts Horse, meat byproducts

-- 0.40 Poultry, meat 0.01 0.01 Poultry, fat 0.01 0.01 Poultry, meat byproducts 0.03 0.01 Poultry, eggs 0.01 0.01 Sheep, meat 0.1 0.15 Sheep, fat 0.04 0.10 Sheep, kidney 0.2

-- See Sheep, meat byproducts Sheep, meat byproducts (except kidney) 0.3

-- See Sheep, meat byproducts Sheep, meat byproducts

-- 0.40

* Quantifiable residues of sulfoxaflor are not expected in poultry or swine commodities; for harmonization purposes, however, H ED is recommending tolerances at the LOQ level (0.01 ppm) for sulfoxaflor in those commodities

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 10 of 176 2.2.4 Revisions to Petitioned

- For Tolerances

M any of the proposed tolerances are different from th ose recommended by HED . The reason for the differences is that the registrant determined the proposed tolerances using the NAFTA (North American Free

- Trade Agreement) tolerance calculat ion procedures rather than by using

the OECD calculat ion procedures

. In addition, the regi strant proposed tolerances for some crops as both an individual crop and as members of a crop group. The registrant should submit a revised Section F in which the proposed tolerances are the same as those recommended by HED.

2.3 Label Recommendations

2.3.1 Recommendations from Residue Reviews

The sulfoxaflor labels should be revised to specify that no more than two applications may be

made to soybean forage, as supported by the available residue data.

2.3.2 Recommendations from Occupational Assessmen t

Sulfoxaflor is classified as Toxicity Category IV via the dermal route and for skin irritation potential. It is not a skin sensitizer. Short

- and intermediate

- term post

- application risk estimates were not a concern on day 0 (12 hours following applica tion) for all post

- application activities. Under 40 CFR 156.208 (c) (2) (iii), ai’s classified as Acute III or IV for acute dermal, eye irritation and primary skin irrigation are assigned a 12

- hour REI. Therefore, the [156 subpart K]

Worker Protection St atement interim REI of 12 hours is adequate to protect agricultural workers from post

- application exposures to sulfoxaflor. It should be noted that the petitioner has proposed a REI of 24 hours for the proposed products based on their interpretation of th e acute ocular toxicity study of the TransformTM WG formulated product resulting in a Toxicity Category II for eye irritation. HED recommends that RD ensure that the proper REI is listed on both proposed labels according to the acute toxicity profile of th e active ingredient.

2.3.3 Recommendations f rom Residential Assessment

At this time, there are no residential uses for sulfoxaflor.

3.0 In troduction

3.1 Chemical Identity

Table 3 .1 . Sulfoxaflor Nomenclature

. Chemical StructureSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 11 of 176 Common name Sulfoxaflor

Company experimental names XDE

- 208

IUPAC name 1

- (6

- trifluoromethylpyridin

- 3

- yl)ethyl](methyl)

- oxido

- λ 6

- sulfanylidenecyanamide

CAS name cyanamide, N

- [methyloxido[1

- [6

- (trifluoromethyl)

- 3

- pyridinyl]ethyl] λ 4

- sulfanylidene]

Molecular formula C 10 H 10 F 3 N 3 OS

CAS registry number 946578

- 00

- 3

End

- use products (EP) GF

- 2032 SC, Transform WG

Chemical Class Sulfoximine

Known Impurities of Concern None

3.2 Physical/Chemical Characteristics

The physical/chemical properties of sulfoxaflor (Appendix D ) do not indicate that there are any special considerations that need to be made in conducting this risk assessment. The compound is not particularly volatile (vapor pressure ≤ 1.4 x 10 - 6 Pa) and based on the K OW (~6.35) it is not expected to partition into fatty tissue or bioaccumulate. In the soil, sulfoxaflor and its major degradate, X11719474, are expected to be highly mobile.

3.3 Pesticide Use Pattern

Sulfoxaflor products are intended to be used for insect control i n a wide variety of crops as well as turf grass and ornamentals. Sulfoxaflor is generally applied 2

- 4 times with retreatment intervals ranging from 5 to 14 day s , depending on crop, pest, and country/region. The end

- use products may be applied through a variety of application methods including aerial, chemigation, ground

- boom sprayer

, and air

- blast sprayer

. The recommended application rate s for agricultural

crops range from 0.016

- 0.133 lb ai/A. A summary of the use directions is provided in Table 3.3

.

Table 3.3. Summary of Proposed Directions for Use of Sulfoxaflor.

Applic. Timing, Type, and Equip. Formulation Applic.

Rate

, lb ai/A Max. No.

Applic.

per Season Max. Seasonal

Applic. Rate

, lb ai/A [g ai/ha] PHI

(days) Use Directions and

Limitations

Barley/Triticale/Wheat

Ground, air blast,

and aerial SC/WDG 0.012

- 0.043 2 0.09 [101] 7 (forage/ fodder/hay) 14 (grain/ straw) 14

- day retreatment

interval.

Brassica (Cole) Leafy Vegetables
Ground, air blast,

and aerial SC/WDG 0.023

– 0.09 0 4 0.266 [298] 3 7

- day retreatment

interval. Bulb Vegetables (Crop Group 3

- 07)
Ground, air blast,

and aerial SC/WDG 0.09 0 3 0.266 [298] 7 7

- day retreatment

interval. Canola (Rapeseed) Subgroup 20A
Ground, air blast, SC/WDG 0.012

- 2 0.09 [101] 14 14

- day retreatmentSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 12 of 176 Table 3.3. Summary of Proposed Directions for Use of Sulfoxaflor.

Applic. Timing, Type, and Equip. Formulation Applic.

Rate

, lb ai/A Max. No.

Applic.

per Season Max. Seasonal

Applic. Rate

, lb ai/A [g ai/ha] PHI

(days) Use Directions and

Limitations

and aerial 0.043 interval. Citrus (Crop Group 10)
Ground, air blast,

and aerial SC/WDG 0.023

- 0.133 4 0.266 [298] 1 7

- day retreatment

interval.

Cotton

Ground, air blast,

and aerial SC/WDG 0.012

- 0.09 0 4 0.266 [298] 14 5

- day retreatment

interval.

Cucurbit Vegetables (Crop Group 9)
Ground, air blast,

and aerial SC/WDG 0.023

- 0.09 0 4 0.266 [298] 1 7

- day retreatment

interval. Fruiting Vegetables (Crop Group 8)
Ground, air blast,

and aerial SC/WDG 0.022

- 0.09 0 4 0.266 [298] 1 7

- day retreatment

interval. Leafy Vegetables (except Brassica) Crop Group 4
Ground, air blast,

and aerial SC/WDG 0.023

- 0.09 0 4 0.266 [298] 3 7

- day retreatment

interval. Leaves of Root and Tuber Vegetables (Crop Group 2)
Ground, air blast,

and aerial SC/WDG 0.023

- 0.09 0 4 0.266 [298] 7 7

- day retreatment

interval. Pome Fruits (Crop Group 11)
Ground, air blast,

and aerial SC/WDG 0.023

- 0.09 0 4 0.266 (298) 7 7

- day retreatment

interval. Root and Tuber Vegetables (Crop Group 1)
Ground, air blast, aerial, and chemigation. SC/WDG 0.023

- 0.09 0 4 0.266 [298] 7 7

- day retreatment

interval.

Small Fruit Vine Climbing, except Fuzzy Kiwifruit (Subgroup 13

- 07F)
Ground, air blast,

and aerial SC/WDG 0.043

- 0.09 0 4 0.266 [298] 7 7

- day retreatment
interval. Low Growing Berry (Subgroup 13

- 07G)
Ground, air blast,

and aerial SC/WDG 0.043

- 0.09 0 4 0.266 [298] 1 7

- day retreatment

interval. Soybean
Ground, air blast,

and aerial SC/WDG 0. 012

- 0.09 0 4* 0.266 [298] 7 7

- day retreatment

interval. Stone Fruit (Crop Group 12)
Ground, air blast,

and aerial SC/WDG 0.023

- 0.133 4 0.266 [298] 7 7

- day retreatment

interval. Succulent, Edible Podded, and Dry Beans
Ground, air blast,

and aerial SC/WDG 0.023

- 0.09 0 4 0.266 [298] 7 7

- day retreatment
interval.

Tree Nuts
Ground, air blast,

and aerial SC/WDG 0.022

- 0.133 4 0.266 [298] 7 7

- day retreatment

interval. Turf grass/ Ornamentals

Ground and a erial ( ornamentals only ) SC/WDG 0.043

- 0.133 Not

specified 0.266 [298] NA 7

- day retreatment

interval. GreenhouseSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 13 of 176 Table 3.3. Summary of Proposed Directions for Use of Sulfoxaflor.

Applic. Timing, Type, and Equip. Formulation Applic.

Rate

, lb ai/A Max. No.

Applic.

per Season Max. Seasonal

Applic. Rate

, lb ai/A [g ai/ha] PHI

(days) Use Directions and

Limitations

use. *The residue data support two applications for soybean forage.

3.4 Anticipated Exposure Pathways

The requested uses may result in human exposure to sulfoxaflor and/or its metabolites in food and drinking water. At this time, there are no existing, pending, or requested registrations that would result in exposure from use in a residential setting.

Sulfoxaflor presents a somewhat unique situation regarding the food and drinking water exposure pathway. Environmental fate data indicate that use of sulfoxaflor is likely to result in

different residue profiles in surface water and groundwater. In surface water, the principal residue is expected to be sulfoxaflor whereas in groundwater, the expected residues are the metabolites X11719474 and, to a lesser extent, X11519540.

3.5 Consideration of 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 Population s," ( 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

- establ ished procedures. In line with OPP policy, HED estimates risks to population subgroups from pesticide exposures that are based on patterns of that subgroup’s food and water consumption, and activities in and around the home that involve pesticide use in a residential setting. Extensive data on food consumption patterns are compiled by the USDA under the Continuing Survey of Food Intake by Individuals (CSFII) and are used in pesticide risk assessments for all registered food uses of a pesticide. These dat a are analyzed and categorized by subgroups based on age, season of the year, ethnic group, and region of the country. Additionally, OPP is able to assess dietary exposure to smaller, specialized subgroups and exposure assessments are performed when condi tions or circumstances warrant. 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 post

- application are

e valuated. 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 die tary patterns among specific subgroups.

4.0 Hazard Characterization and Dose

- Response Assessment

Sulfoxaflor is a member of the sulfoximine class of insecticides. It controls insects via a unique interaction with the insect nicotinic acetylcholine recept or. The proposed insecticidal mode

- of

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 14 of 176 action (MOA) is similar to neonicotinoid insecticides

, as both are nicotinic acetylcholine receptor (nAChR) agonists. However, sulfoxaflor differs from the neonicotinoids because it is not susceptible to in vitro met abolism and is proposed to have a different adverse outcome pathway.

4.1 Toxicology Studies Available for Analysis

The hazard database for sulfoxaflor is extensive and provides sufficient information to complete

a human health risk assessment. Sulfoxafl or was evaluated for acute, subchronic and chronic oral toxicity in rats, mice and dogs, dermal toxicity in rats, teratogenicity in rats and rabbits, reproductive and postnatal toxicity in rats, carcinogenicity in mice and rats, neurotoxicity in rats,

and immunotoxicity in rats. Additional guideline stu dies included bacterial reverse

- mutation test, micronucleus test, and an in vitro chromosome aberration test. The database does not contain a subchronic inhalation study. HED has determined that th is study is not required for

sulfoxaflor ( TXR# 0056310 ).

During the conduct of the guideline studies, limb flexure and /or increased deaths in neonatal rats were observed in the developmental study, the reproduction and fertility study, and the developmental neuro toxicity stud y . In an attempt to further elucidate the biological processes underlying these effects and their potential relevance to humans, the registrant conducted

mechanistic and pharmacokinetic studies beyond the guideline requirements. These studies included development and reproduction studies in the rat and rabbit, as well as in vitro studies in rat and human tissues. Mechanistic studies were also provided to support MOA proposals for Leydig cell tumors, hepatocellular tumors, and pre putial gland tumors that were observed in the reproduction and chronic guideline studies. A detailed discussion of these studies can be found in the Cancer Assessment Review Committee (CARC) memorandum (TXR # 0056301)

. The guideline and mechanistic studi es have been designed in such a manner as to be mutually supportive. For example: identical animal strains, vehicles, and routes of administration are used throughout the studies; plasma sulfoxaflor concentrations were measured in all studies for

estimate s of internal doses; and the same dose levels were used in multiple studies ( e.g.,. the 2 generation reproduction study and the DNT were both conducted at 0, 25, 150, or 400 ppm). This allows for a more integrated evaluation of studies across the sulfoxaf lor toxicity profile

.

As described in the following section, sulfoxaflor is resistant to in v iv o metabolism. However, X11719474 is a major plant metabolite and environmental degradate of sulfoxaflor with a potential for human exposure through drinking w ater. A battery of studies have been conducted including an oral acute study, absorption and metabolism studies in rats; 28

- day and 90

- day oral studies in rats, rabbits, and dogs; a reproduction study in rats; a complete battery of mutagenicity studies; a nd in vitro rat and human receptor binding studies. Limited studies are also available

for several other metabolites

.

4.2 Absorption, Distribution, Metabolism, & Elimination (ADME)

Several studies are available to evaluate the ADME of sulfoxafl or in the rat, including single

dose studies, repeated

- dose studies, and an intravenous

- dose study to determine systemic availability. In addition, sulfoxaflor plasma concentrations were determined in many of theSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 15 of 176 studies, including guideline and non

- guideline stud ies in the rat, rabbit, and mouse. Offspring and fetal sulfoxaflor plasma concentrations were determined in the reproduction and development studies in the rat and rabbit. This information allows the comparison of toxicokinetics across species, route of administration, duration

, and life

- stages

. Furthermore, data are available reflecting ADME of the plant/environmental metabolites X11719474 and

X11519540. Note that these metabolites were not found in the rat metabolism study.

ADME of Sulfoxaflor in the Rat

Sulfoxaflor is extensively and quickly absorbed following an oral dose. In single

- and repeated

dose studies using radiolabeled sulfoxaflor, absorption estimates ranged between 92 and 96%. Maximum plasma concentrations are reached within 2 hours. When compared to an intravenous dose, 92% of an oral dose is available systemically, confirming the hi gh level of absorption. There were no significant differences between the low (5 mg/kg), high (100 mg/kg), or repeated (14 days @ 5 mg/kg) dose groups and plasma concentrations were dose

- related indicating absorption was not saturated up to 100 mg/kg when given by oral gavage.

Following absorption, sulfoxaflor is widely distributed throughout the body. When tissue

distribution was determined at C

max (i.e., maximum concentration of sulfoxaflor in the plasma), the tissues containing the largest proportion s of radioactivity were the gastrointestinal tract, liver, kidney, and urinary bladder: a ll organs consistent with point

- of

- entry and excretion. By 168 hours, an average of <1.5% of the administered dose remained in the body and that was

distributed throu ghout the body in minor quantities.

The primary route of elimination for sulfoxaflor is through urine. Within 24 hours of an oral dose, 89

- 94% of the administered dose is eliminated through urine, compared to 5

- 8% through

feces. The removal of sulfoxafl or from plasma occurs in 2 distinct phases; a rapid phase with a

half

- life of 4

- 6 hours followed by a much slower phase with a half

- life of 39

- 45 hours. In the 90

- day oral study, the plasma half

- lives were 8 and 9 hours in the males and females, longer th an the

28

- day study of 4

- 5 and 7

- 8 hours for the males and females, respectively. Despite the longer second phase and the observation that elimination of sulfoxaflor is marginally slower in longer

term studies , little sulfoxaflor remains in vivo after the initial 24 hours and therefore there is no

concern for bioaccumulation.

In vivo metabolism of sulfoxaflor is minimal. As previously indicated, greater than 90% of an oral dose is eliminated within 24 hours; at least 93% of which is eliminated as the parent compound . A glucuronide conjugate of the urea metabolite was identified at 2

- 4%. Five additional metabolites were observed; however, they were not identified and each accounted for <1% of the administered dose. Similarly, no major metabolites we re identified in either the rat

or mouse 28

- day and 90

- day oral studies.

Fetal Exposure / Maternal Transfer in the Rat

To support the proposed MOA for developmental effects, the pharmacokinetics of sulfoxaflor were determined in a number of developmental and reproductions studies. Pharmacokinetics is a summation of the processes that control how sulfoxaflor is distributed in vivo

, i.e

., how it isSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 16 of 176 metabolized or eliminated, and

, of critical importance to the developmental MOA, how it is presented to developing fetuses or newborn offspring. The distribution and clearance parameters

of sulfoxaflor were described in the metab olism studies discussed above . The parameters relevant to fetal or offspring exposure were captured in the following reproduction and developmental studies.

In the 2

- generation reproduction study in the rat, plasma concentrations were determined in

off spring and dams 4 days after parturition. Offspring levels were ≈ 30% of maternal levels across all dose levels indicating that the offspring internal dose was ≈ 1/3 of the maternal internal

dose. In a cross

- fostering and a developmental toxicity study i n rats, the maternal and fetal

s ulfoxaflor blood concentrations were similar on GD 21

. Similar findings regarding the comparison of maternal vs. offspring s ulfoxaflor blood concentrations were reported i n the rabbit developmental study. This suggests that sulfoxaflor moves readily across the placenta.

Lactational transfer was also determined in the cross

- fostering study in the rat. O n Lactation Day ( LD ) 0

, t he measured concentrations of sulfoxaflor in m ilk from sulfoxaflor

- exposed dams were approximately half the corresponding concentrations in plasma . Although milk

concentrations were not determined in the 2

- generation study, the offspring plasma concentrations of sulfoxaflor were approximately 30% of the maternal animals, supporting the observed reduction in lactational transfer. In a goat metabolism study, less than 2% of the dietary sulfoxaflor was found in the milk. Thus

, lactational transfer ranges between 2 and 50% in goats and rats, respectivel y.

These studies clearly indicate that fetal plasma levels of sulfoxaflor are similar to the dams during gestation. However, after parturition when offspring exposure to sulfoxaflor is limited to lactational transfer through the milk, exposure to the rat pups is 2 to 3

- fold lower than maternal

doses.

Sulfoxaflor Metabolite s X11719474 and X11519540

Despite the lack of in vivo metabolism, there is a potential for exposure to environmental degradates of sulfoxaflor. In particular, X11719474 is a major degradate found in environmental fate studies, and is expected to be the major degradate in drinking water from groundwater

sources . The toxicokinetics of X11719474 are similar to the parent sulfoxaflor; following an oral dose, X11719474 is highly absorbe d (>98%), reaches a maximum plasma concentration quickly (≈1 hour), and is quickly eliminated through the urine >90% within 12 hours. Limited ADME data are also available for a second metabolite, X11519540, which has been observed in

limited amounts in fi eld and animal feeding studies. The elimination half

- life of X11 5 19540 is approximately 27 hours, longer than sulfoxaflor and X11719474. Both metabolites are largely

absorbed and excreted unmetabolized in in vivo studies.

4.2.1 Dermal Absorption

The Registrant submitted an in vivo study in rats and an in vitro study utilizing split

- thickness rat and human skin in which sulfoxaflor was applied at doses of 2360 μg/cm 2 , 4.8 μg/cm 2 , and 0.25 μg/cm 2

. These doses represent sulfoxaflor concentrate, the highest in

- use concentration, and theSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 17 of 176 lowest in

- use concentration, respectively. The in vivo data showed that skin

- bound residues of sulfoxaflor were absorbed for up to 144 hours after skin washing. The dermal absorption factor (DAF) for sulfoxaflor afte r 10 hours of dermal exposure (that included absorbable skin

- bound residues) was 11% for both the highest and lowest in

- use concentrations. These values were larger in magnitude than the concentrate. The in vitro data for rat skin after 24 hours of derma l exposure showed the largest magnitude potentially absorbable dose for the lowest in

- use concentration. The Agency considered comparison of data at the 24

- hour time point in the in

vitro study to the 10

- hour time point in the in vivo study on the basis t hat the estimates at the 24

- hour time point would be conservative surrogates for a 10

- hour dermal exposure (because dermal absorption generally increases with time). The DAF for rat skin in vitro was 8%, a value that

does not differ from the 11% DAF for r at skin from the in vivo study after standard errors in

measurement are taken into account. This result indicated that the in vitro study design and execution was predictive of the in vivo results. The Agency ’s policy is that if an in vitro study is

show n to be predictive of an in vivo result, the Agency will use human skin from the in vitro study to refine the DAF for human exposures. The DAF for human skin in the in vitro study at

the lowest in

- use concentration was 2.4% (Table 4.2.1)

. The Agency is u sing this human value of 2.4% as the DAF for determining dermal

- equivalent doses in the sulfoxaflor risk assessment.

Table 4.2.1. Derivation of the Dermal Equivalent Dose for Human Health Risk Asssessment.

Parameters In Vivo Rat In Vitro Rat In Vitro Human Study ID MRID No. 47832413

TXR No. 0055507 MRID No.47832414

TXR No.0055507 MRID No.47832414

TXR No.0055507

Test Material SC Formulation (24%) SC Formulation (24%) SC Formulation (24%) Exposure 10 hours 24 hours 24 hours

Doses tested (μg/cm 2 ) Low Mid Hi Low Mid High Low Mid High

0.25 4.8 2360 0.25 4.8 2360 0.25 4.8 2360

% Absorbed 10.77 a 11.35 1.22 8.02 b 8.72 1.67 2.38 b 2.44 0.35 Point of Departure Oral POD = Oral NOAEL = 1.8 mg/kg/day based on decreased neonatal survival at 7.1

mg/kg/day observed in the DNT Dermal Equivalent Dose (DED) For short

- and intermediate

- term dermal exposure, the POD is 1.8 mg/kg/day

.

POD = oral NOAEL = 1.8 mg/kg/day DAF ( In Vivo rat ) ≈ DAF ( In Vitro rat) ≈ 11%

In Vitro Human dermal absorption = 2.4%

Dermal Equivalent Dose = Oral POD/human DAF=1.8 mg/kg/day/ 0.024= 75 mg/kg/day

Comments: Same formulation products were tested in all three studies.

Doses are identical in the in vitro rat and human and the in vivo rat study.

In vitro absorption predicts in vivo absorption in the rat. DED is calculated using the human dermal absorption factor.

a

– Absorbed dose after 144 hours

b

- Potential absorbable dose after 24 hours

4.3 Toxicological Effects

Sulfoxaflor is a member of the sulfoximine class of insecticides. It controls insects via agonism of the insect nicotinic acetylcholine receptor (nAChR) leading to sustained muscle contracture, paralysis, and death. In mammals, the pattern of toxicity at tributed to sulfoxaflor exposure includes developmental effects (manifested as neuromuscular abnormalities and neonatal death),Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 18 of 176 hepatotoxicity, and generalized toxicity ( e.g., decreases in body weight, body weight gain, and/or food consumption).

4.3.1 De velopmental/Reproductive Effects, Mode of Action, and Human Relevance

Developmental/offspring toxicity

, manifested as skeletal abn ormalities and neonatal death s
, was observed in rats only

. The skeletal abnormalities, forelimb flexure, bent clavicles, and hindlimb

rotation likely result from skeletal muscle contraction due to agonism of the muscle nAChR in

utero . Similarly, contraction of the diaphragm muscle prevents normal breathing in neonates resulting in increased mortality. The skeletal abnormaliti es were observed at high doses in the developmental and reproduction studies and decreased neonatal survival was consistently observed in the reproduction and developmental neurotoxicity studies. These developmental

effects were not observed in the rabbit

.

4.3.1.1 Mode of Action Framework

Given the known insecticidal MOA for sulfoxaflor, it was hypothesized that these effects could be related to an interaction between sulfoxaflor and the nAChR. In an attempt to test this hypothesis and further elucidate the biological processes underlying these effects and their potential relevance to humans, the registrant conducted numerous mechanistic studies beyond the guideline requirements. These studies included but were not limited to development and reproduction studies in the rat and rabbit, as w ell as in vitro studies in rat and human tissues. These studies were designed to assess if sulfoxaflor binds and activates the nAChR in rats, rabbits, and humans. Ultimately, the objective was to determine if the differences observed in
sulfoxaflor’s dev elopmental toxicity profile could be ascribed to a differential interaction with the nAChRs in these three species.

HED has used existing, peer reviewed frameworks in the analysis of the special data. In keeping with EPA’s Framework for Determining a Mut agenic Mode of Action and the International Programme on Chemical Safety (IPCS) Framework f or Analyzing the Relevance of a Non

Cancer Mode of Action for Humans, the Agency conducted a mode of action framework analysis considering the relevance of the mode of action (MOA) to humans. This is consistent with the approach es taken by the Joint F AO /W HO Meetings on Pesticide Residues (JMPR) and the Organization for Economic Cooperation and Development (OECD). The MOA analysis framework relies on a weight

- of

- evid ence approach , which is based on modified Bradford Hill

criteria for causality (Hill, 1965) . In line with the 2007 NRC report on Toxicity testing in the

21 st Century, some have co nsidered the concept of adverse

- outcome pathway s (AOP) in addition

to MOA. These concepts are fundamentally similar in that each considers key events (including an initiating event) leading from exposure to an adverse outcome. This document uses the MOA terminology instead of the AOP terminology. In brief, the MOA framework beg ins with a proposed set of key events and then describes the evidence for such events with particular focus on temporal and dose

- response concordance. Consistent with the Bradford Hill criteria, the MOA analysis also takes into consideration the consisten cy, specificity, and biological plausibility of the findings. The analysis also involves explicit evaluation of alternative hypotheses, alternative MOAs, and remaining uncertainties.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 19 of 176 Proposed Key Events

In light of observations in the guideline toxicity studies and the basic biological understanding of

muscle nAChR ontogeny and function in mammalian systems, the registrant proposed a MOA by

which the muscle effects (forelimb flexure, bent clavicles, and r otated hindlimbs) and deaths

observed in rat neonates are caused by sulfoxaflor’s sustained agonism at the fetal

- type

muscle nAChR and subsequent sustained muscle contracture of the limb, shoulder girdle and diaphragm, respectively. This proposed MOA can be expressed in terms of four key events depicted in the Figure 4.3.1. 1:

Figure 4.3. 1 .1

. Key events of the sulfoxaflor MOA. Each box represents a key event , beginning with the availability of sulfoxaflor at the neuromuscular nAChR and concluding with the reproductive effects observed in the reproduction and development studies (KE =Key Event ).

Summary of the Evidence for the Key Events:

Key Event #1: Internal dose at the target tissue: As described in the ADME section of

t his document, sulfoxaflor is extensively and quickly absorbed following oral exposure and distributed systemically. It is, therefore

, available to interact with the target tissue.

Key Event #2: Muscle receptor binding: Two types of nAChRs are present in mammals, neuronal and muscle. The muscle nAChRs are found in the intramuscular junctions of skeletal muscles and are involved in muscle contraction; it is the muscle nAChRs which are the targets for sulfoxaflor. Disregulation of these receptors can resu lt in improper muscle contraction, difficulties in breathing, and ultimately death (due to sustained diaphragm contracture). Two isoforms of the muscle nAChRs have been identified in mammals, a fetal isoform and an adult isoform. The transition from feta l to adult isoform

occu rs shortly after birth in rats and late during the 3 rd trimester in humans. Furthermore, sulfoxaflor has been shown to bind the fetal isoforms of the nicotinic receptors of rats, rabbits, and humans in a series of mechanistic studie s. These studies demonstrated a

dose

- dependent increase in sulfoxaflor binding to the fetal muscle nAChR in all three species.

Key Event #3: Receptor agonism: Sulfoxaflor binds the nAChR in the rabbit, rat, and human. A study was undertaken to determin e if sulfoxaflor causes activation of the
receptors

. Rat and human muscle nAChRs were expressed in Xenopus oocytes

– a model system used to evaluate activation of receptors. Rabbit receptors were not evaluated because the encoding proteins are not currently available for this species. The ability of

sulfoxaflor to activate both fetal and adult muscle nAChRs w as examined. This study

indica ted that the rat fetal

- type nAChR was activated by sulfoxaflor. However, there was no activation in the adult rat, fetal human, or the adult human receptors. The implications Target Tissue

KE 1 nAChR

Binding

KE 2 nAChR

Agonism

KE 3 Increased

Muscle

Contraction

KE 4 In Vivo

Clinical Signs

KE 4Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 20 of 176 for the lack of response in humans are discussed below in the human relevance section.

Key Event #4: Muscle contraction and clinical signs: Sustained contraction of fetal rat diaphragm muscles in the presence of sulfoxaflor was demonstrated in vitro . Contraction of the diaphragm during late gestation leads to asphyxiation and the observed increase in

neonatal deaths.

The muscle nAChRs transition from the fetal to the adult isoforms and the timing of this shift provides insight into the potential role of the fetal nAChR in the developmental

effects observed in rats after sulfoxaflor exposure. Both the skeletal effects and the offspring deaths in rats are limited to the very early postnatal period when the fetal isoform is the predominant isoform present. However, beyond post

- natal day 4 there is

no increase in pup death and the skeletal effects noted shortly after birth (forelimb flexure, bent clavicles, and rotated hindlimbs) begin to recover. Thus, it appears that

there is a time

- concordance between the appearance of the adult isoform and recover y from the developmental effects strongly suggesting that the fetal isoform of the muscle nAChR is critical to the manifestation of these types of effects. For a more detailed description of the agonism experiments refer to Appendix B.

Alternative MOAs: As discussed in Appendix B, a variety of alternative hypotheses were considered (i.e., neuronal nicotinic receptor activation, muscarinic receptor activation, fetal nicotinic receptor inactivation, and ACE inhibition). However, none of these were

consist ent with sulfoxaflor’s toxicological profile, particularly the adverse outcomes

observed.

Weigh t of the Evidence: In conclusion, there is compelling evidence that the proposed MOA is operative in the fetal rat and leads to the neonatal death and the s keletal effects seen in the guideline toxicity studies involving that species. As such, HED believes the developmental and reproduction MOA based on agonism of the fetal nAChR has been

established.

4.3.1.2. Human Relevan ce

As summarized above and dis cussed in detail in Appendix B, the MOA for the neonatal death and limb flexures based on agonism of muscular nicotinic receptor has been established. Given this finding, HED has used the Human Relevance Framework (HRF) to consider the degree to

which the development al and reproduction MOA is relevant to humans and thus relevant for risk

assessment. The HRF entails both a qualitative and quantitative evaluation and comparison of the key events in animals and humans. Like the MOA analysis, the HRF analysis includes an

evaluation of uncertainties and consideration of alternatives. An HRF analysis may be

undertaken fo r a variety of reasons including:

S ituations where the effects in animals would have potentially serious consequences if they occurred in humans

, and

S ituations where there are interspecies differences in either the type of effect or the dose

levels at wh ich an effect occurs.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 21 of 176 In addition to the guideline toxicity studies conducted with sulfoxaflor, an extensive mechanistic data set has been generated to further evaluate the interspecies sensitivity to sulfoxaflor’s developmental effects. These studies inc lude both binding and agonism in vitro/ex vivo studies to ascertain if: (a) sulfoxaflor binds to the muscle nAChR receptor in rats, rabbits, and humans and (b) if the bound sulfoxaflor can activate the receptor. Data from these experiments indicate

that a lthough sulfoxaflor does bind to the fetal nAChR in rats, rabbits, and humans, it does not activate the human receptor.

Table 4.3.1.1. Key Events of the Developmental and Reproduction Sulfoxaflor MOA based on agonism of the rat fetal nAChR and the supporting events observed in rats and human tissues. Key Event Evidence in Rodents Evidence in Humans #1 Binding to the fetal

- type muscle nAChR Yes Yes

#2 Agonism at the fetal

- type muscle nAChR Yes No; Plausible, but no agonism occurs

with sulfoxaflor either at the fetal

- or adult

- type muscle nAChR.

#3 Sustained fetal

- type muscle nAChR agonism/sustained muscle contraction Yes No data in humans. Presumably not possible via this MoA as key event #2

does not occur.

Apical Endpoints Forelimb Flexure, Hindlimb Rotation, Bent Clavicle, Neonatal Death Yes No data in humans. Presumably not possible via this MoA as key event #2

does not occur.

Close examination of the key events indicates significant d ifferences in the key events between

the rat and human. Key Event #1: Qualitatively, sulfoxaflor has demonstrated the ability to bind both rat and human nAChRs. As previously described, sulfoxaflor has been shown to bind the fetal

isoforms of the nico tinic receptors of rats, rabbits, and humans.

Key Event #2: Despite binding, s ulfoxaflor does not activate the human fetal muscle

nAChR. An in vitro study using fetal isoform for rat and human of the muscle nAChR expressed in Xenopus oocytes, examined the ability of sulfoxaflor to perturb the functionality of rat and human muscle nAChR isoforms. Treatment with sulfoxaflor resulted in increased response in the rat fetal isoform. In contrast, there was no indication

of agonism by sulf

oxaflor in the human isoform. Figure 4.3.1. 2 is a graphical representation of the agonism of the rat and human fetal NAChRs by sulfoxaflor. Fetal rat and human nAChRs were expressed in Xenopus oocytes followed by exposure to sulfoxaflor (blue line). The endogenous agonist acetylcholine (ACh; red line) was also tested to ensure the integrity of the expressed receptors and for a quantitative comparison

to sulfoxaflor

- induced activity. ACh induced a clear and potent response at 10 - 6 M in the rat and human receptors. However, considerably higher concentrations of sulfoxaflor were required to initiate activity in the rat receptor (10 - 4 M), and the response was relatively low. It is especially important to note that the human receptor was refractory to

sulfoxaflor

- induced activity, even at concentrations exceeding 10 - 3 M, or approximately

500

- fold greater than the concentrations that resulted in activation in the rat receptor.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 22 of 176 Figure 4.3.1. 2. Agonist activation of muscle nAChRs expre ssed in Xenopus oocytes. Data are shown for the (A) rat fetal , (B) human fetal muscle nAChR. AChRs were expressed by microinjection of cDNA or cRNA in Xenopus oocytes. A C h = acetylcholinesterase; IMI = imidacloprid ; Sulfoxaflor metabolite X11719474.

Al though the technology for cloning the rabbit muscle nAChR is not yet available, the lack of developmental effects in the rabbit is an indirect indicator of the unique sensitivity of the rat. In the rabbit developmental study, (MRID 47832066) and non

- guide line rabbit survival study (MRID 47832138) there is no indication of the fetal anomalies observed in the rat developmental toxicity study, in particular the forelimb flexure, bent clavicle, and hindlimb rotation seen in the rat developmental toxicity. Fur thermore, the sulfoxaflor plasma concentrations on gestation day (GD) 27 were ≈ 4

- fold higher than the plasma

concentration associated with neonatal death and neuromuscular/skeletal effects in the rat pups, indicating that the absence of developmental eff ects in the rabbit studies is due to a pharmacodynamic rather than a pharmacokinetic difference between rabbits and rats.

Key Events #3 and #4: Muscle contraction and in vivo clinical signs are not available for evaluation in humans. However, in the abs ence of Key Event #2, these final key events are not expected to occur in humans.

Weight of Evidence: Based on the lack of receptor activity, t here is compelling evidence that the developmental and reproduction MOA based on agonism of the fetal nAChR is not likely to be relevant to humans. Furthermore, a comparison of the amino acid sequence of the rat and human γ subunit (specific to the fetus) revealed that, although the two subunits are similar (approximately 90% identical), they contain 53 amino acid differences ( Millar, 2010 ). There are several precedents for species

- selective agonist activity of nAChR ligands. Given the evidence that as few as one or two amino acid differences can confer species

- selective agonist activ ity upon nicotinic ligands, it seems entirely plausible that the differences in agonist activity of sulfoxaflor can be explained by differences in the amino acid sequence of the rat and human muscle nAChR γ subunits. The γ and ε subunits (fetal and adult isoforms, respectively) show even greater sequence differences tha n the human and rat γ subunit (even from the same species), where these subunits share only about 50% identity in amino acid sequence in the rat.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 23 of 176 Uncertainties: The use of Xenopus oocytes in the agonism study represents a residual uncertainty, albeit extremely low, that precludes the Agency from discounting the possibility of human relevance on a qualitative basis. The heterologous expression of mammalian receptors in Xenopus oocytes is a common technique that has been used for determining the structure and function of nicotinic receptors (Mi llar 2009). Expression in Xenopus oocy t es offers several advantages over native mammalian cells including the following: limited expression of endogen ous ion channels and receptors so that exogenous proteins can be studied with minimal contamination; some channels can only be expressed in oocytes and not in mammalian cells; and oocytes are well suited for electrophysiological recording . Past studies ha ve indicate d that the ac tivity of muscle nAChRs expressed in either oocytes and HEK were similar for muscle (Stern et. al. 1994) and neuronal (Luetje and Patrick 1991) type receptors. However, the unique matrixes provided by the oocytes and mam malian cell s can lead to differential expression of receptors or channels. For example, the expression of the human neuronal nAChR α3β4 results in different ratios of the α and β subunits for oocytes and HEK cells (Krashia e.t al. 2010; Boorman et. al. 2000). Alter ed subunit ratios have been shown to result in different sensitivities to the acetylcholine and other agonists (Krashia et al. 2010; Moroni et. al. 2006; Moroni et. al. 2008). Due to the severity of the endpoint ( i.e , neonatal deaths), HED has a requested a confirmatory study; that is a study confirming the refractory response of a fetal human nicotinic receptor expressed in a mammalian cell. Pending the results of the confirmatory study in human receptors expressed in mammalian cells, HED considers the M OA relevant to humans.

4. 3

. 1.3

. Interspecies Uncertainty Factor

The available information comparing rat and human muscle nAChR response to sulfoxaflor has bearing on the interspecies uncertainty factor (UF A ) when assessing nAChR

- mediated risks

associated with sulfoxaflor exposure. HED recommends that the 10X interspecies factor be reduced to 3X for acute dietary risk assessment for women of reproductive age and for other scenarios assessed using the neonatal death endpoint ( i.e., dermal a nd inhalation risk

assessments ) . This recommendation is based on the evidence gathered from the mechanistic data which indicate that rats may be uniquely sensitive to the developmental effects of sulfoxaflor.

HED has used the 2005 IPCS document on Chemic al Specific Adjustment Factors and the Agency’s 2011 external peer review draft of the Guidance on Data Derived Extrapolation Factors. In short, both approaches endorse the use of mechanistic and pharmacokinetic data to inform data derived extrapolation f actors for inter

- and intra

- species extrapolation. In both approaches, the typical 10X factor is split between a PD and PK component. USEPA typically uses an even split of 3X and 3X. As discussed in 4.1.1, the developmental and reproduction

MOA based on agonism of the fetal nAChR has been established for skeletal anomalies (e.g., forelimb flexure, bent clavicle, and rotated hindlimbs) and neonatal deaths in the rat and HED cannot currently rule out this MOA as being not relevant to humans. As such, the neonatal mortality is relevant for POD selection (see above). Although, HED believes that additional data are necessary to fully demonstrate the lack of human relevance, the existing mechanistic are high quality and provide a robust evaluation of the resp onse to rat and human tissues for the agonism of the nAChR. Moreover, the nature and quality of these data are consistent with the kinds of data recommended by the International Programme on Chemical Safety (IPCS) Chemical

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 24 of 176 Specific Adjustment Factor ( CSA F ) and the EPA D ata D erived E xtrapolation Factors guidance

documents.

Quantitatively

, the sulfoxaflor concentration required to initiate receptor activation in human

fetal nAChR is at least 500

- fold greater than that required to activate rat fetal nAC hR. The absence of nAChR activation precludes the manifestation of skeletal abnormalities and neonatal deaths. Therefore, the data indicate that rats are uniquely sensitive to the developmental and reproduction MOA leading to neonatal death and neuromusc ular effects. The Agency is proposing that the PD portion of the interspecies UF be reduced to 1X. Pharmacodynamics can

be defined as what the chemical does to the bod y

. In the case of sulfoxaflor, PD is expressed as the ability of the sulfoxaflor to in teract with the nAChR. Since agonism of the nAChR is the critical key event, interspecies extrapolation can be assessed using the agonism data ( i.e., data demonstrating sulfoxaflor

- induced activity in the oocyte expressed rat and human nAChRs). A s descri bed in 4.3.1.2 , sulfoxaflor does not elicit a response in the human nAChR at concentrations

500

- fold higher than the concentrations eliciting a response in the uniquely susceptible fetal rat
receptor. In other words, based on this assay, human tissue is a t minimum 500

- fold less sensitive

than is the rat tissue. Since humans are clearly not more sensitive pharmacodynamically to sulfoxaflor compared to the rat, reduction of the UF for PD to 1X, which assumes an equal response between humans and rats, is con servative.

The PK portion of the inter

- species factor is retained at 3X since there are PK studies in human tissues or physiologically

- based pharmacokinetic models which allow for refinement of the PK component of the extrapolation factor.

4.3.2. Oth er Adverse Outcomes Observed for Sulfoxaflor

In terms of adults, sulfoxaflor systemic toxicity is manifested primarily in the form of hepatotoxicity in rat, mouse, and dog studies. Moreover, effects were observed following exposures as short as 8 days as well as in studies continuing for up to 2 years . Increased liver weight, hepatic hypertrophy, fatty liver, single cell necrosis, and changes in clinical chemistry (i.e., cholesterol, AST, or ALT) were consistently observed throughout the toxicology database. The longer term carcinogenicity studies re sulted in hepatic effects at lower doses compared to

the short

- term studies. In addition, males appeared to be more sensitive to the hepatic toxicity. As indicated in section 4.2, male rats had a slower clearance rate compared to females. The

slower cle arance rate in males in essence increases exposure and might account for the increased male sensitivity to hepatotoxicity in the longer

- term studies.

Aside from the developmental and hepatic effects, other effects seen in the sulfoxaflor database

includ e decreases in food consumption and body weight as well as changes in the male reproductive system. Decreased body weight, body weight changes, and food consumption were observed during the first few days of several oral studies at the mid

- and high

- dose levels. As a result of decreased feeding early in the studies, body weights were typically lower in the mid

and high

- dose groups compared to the controls, although the differences were not generally statistically significant. Decreased palatability is a likely contributor to this effect as body weight decreases were often observed at study initiation but were comparable to control animals

within several weeks.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 25 of 176 Effects in the male reproductive organs were observed in the chronic/carcinogenicity study in rats that included increased testicular and epididymal weights, atrophy of seminiferous tubules, and decreased secretory material in the coagulating glands, prost ate, and seminal vesicles. Additionally, there was an increased incidence of interstitial cell (Leydig cell) tumors. The Leydig cell tumors observed after exposure to sulfoxaflor are not considered treatment related due to the lack of dose response , the lack of statistical significance for the combined tumors (unilateral and bilateral)

, and t he high background rates for this tumor type in F344 rats. T he primary effects on male reproductive organs are considered secondary to the loss of normal

testicular function due to the size of the interstitial cell (Leydig Cell) adenomas. Consequently, the secondary effects to the male reproductive organs are also not considered treatment related.

Clinical indications of neurotoxicity were only observed at high dose s in the acute neurotoxicity study in rats. At the highest dose tested, clinical signs included increased muscle tremors and twitches, convulsions, hindlimb splaying, increased lacrimation and salivation, decreased pupil size and response to touch, gait a bnormalities and decreased rectal temperature. Decreased

motor activi ty was also observed in the mid

- and high

- dose group s. While the muscle effects are consistent with perturbation of the neuromuscular nAChR, salivation, lacrimation, and pupil

response are typically associated with muscarinic receptor activity. Since the neurotoxicity was observed only at a very high dose and is not entirely consistent with nicotinic receptor toxicity, it is unlikely that these effects are due to agonism of the nAChR.

Finally, tumors were observed in chronic rats and mouse studies. In the rat, a significant

increase in the incidence of hepatocellular adenomas and combined adenomas and/or carcinomas in the high

- dose males was observed when compared to controls. As previously indicated, Leydig cell tumors were also observed in the high

- dose group, but it was determined that these were not related to treatment with sulfoxaflor. There was also a significant increase in the incidence of preputial gland tumors in males in the high

- dose group. Marginal increases were

also observed in the low

- and mid

- dose groups; however, the incident values for these groups were within the range of historical control values. In male mice , treatment

- related changes in the high

- dose gro ups consisted of increased incidences of hepatocellular adenomas and/or

carcinomas.

4. 3

. 3 . Metabolites

As indicated previously, the petitioner has provided several additional studies on sulfoxaflor metabolites and degradates. Sulfoxaflor has six known p rimary metabolites; all are formed by metabolism of the parent’s ethyl

- sulfanylidene

- cyanamide side chain and/or the side chain of its

metabolites.

X11719474 is a major metabolite in terms of magnitude of residue and/or presence across

matrices

- soil, water and edible crops. The estimated half

- life of X11719474 exceeds 1000 days

in US soils and the compound has the potential to leach into ground water. Of the metabolites,

X11719474 has the most complete toxicology database , which includ e s oral acute, subchronic, developmental, reproduction, and metabolism studies. Unlike sulfoxaflor, X11719474 does not

cause agonism of the fetal rat muscle nAChR and prenatal exposure does not result in increasedSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 26 of 176 neonatal deaths or skeletal abnormalities. The major ta rget organ of X11719474 is liver and the LOAELs are based on hepatotoxicity.

X11519540 is a minor soil and animal metabolite . It is also found as a low

- level impurity of the manufacturing process. Repeat

- dose studies in the rat show that the primary t arget organ is the liver. In addition, X11519540 resulted in a dose

- dependent increase in the adrenal gland weight.

When the LOAELs for the 28

- day oral studies for sulfoxaflor and X11519 540 are compared, the

LOAEL for X11519 540 is 10

- fold lower than the LOAEL for sulfoxaflor. The oral LD 50 for this

metabolite is approximately half of the LD 50 for sulfoxaflor. Therefore, it is apparent that

X11519 540 is more potent than sulfoxaflor. The 28

- day rat study also indicates that X11519540 has a longer half

- li fe; 24

- 35 hours compared to 4

- 8 hours for sulfoxaflor, which could contribute to its increased potency.

Other metabolites include X11721061, X11596066, X11579457, and X11718922. None of these metabolites are expected to contribute significantly to the p otential toxicity of sulfoxaflor.

As indicated above, the target organ for X11719474 and X11519540 is the liver. To determine if

X11719474

- induced hepatotoxicity is also mediated by nuclear receptor

- mediated MOA, rats were given either 0 or 8000 ppm (583 mg/kg/day) X11719474 for 7 days in a non

- guideline study. After 7 days, the key events identified in the sulfoxaflor MOA were determined to also occur after X11719474 exposure. Based on these observations, CARC determined that the

metabolite X11719474 induced hepatocellular gene expression, enzyme activity, and proliferation in a manner very similar to sulfoxaflor. A similar study providing mechanistic data on the pathway leading to hepatotoxicity is not av ailable for X11519540 . However, due to structural similarity between X11519540 and X11719474, and the similar MOAs for X11719474 and sulfoxaflor, HED assumes that the events leading to hepatotoxicity in X11519540 are similar

to those for sulfoxaflor and X 11719474.

As previously indicated, there is a complete database of toxicological studies for sulfoxaflor and there are numerous studies describing the toxicity of X11719474. However, the database for

X11519540 is limited to two studies, a 28

- day oral tox icity study in the rat and an acute LD 50 study, also in the rat. Since the rat 28

- day oral toxicity studies are common to all three chemical
databases and the studies’ LOAEL s are all based on the common toxic effect of hepatotoxicity,

HED has estimated th e relative potency of X11719474 and X11519540 based on results from the

28

- day oral toxicity study in the rat. The study LOAELs are 79.4, 244, and 7.7 mg/kg/day for sulfoxaflor, X11719474, and X11519540 , respectively. Therefore the relative potency facto rs with respect to liver toxicity are 0.3 for X11719474 and 10 for X11519540

.

4.4 Safety factor for Infants and Children (FQPA Safety Factor)

HED has recommended that the required 10X FQPA Safety Factor be reduced to 1X based on the following considerati ons: 1) the toxicology database for sulfoxaflor is complete with regard to FQPA consideration, including the required developmental and reproductive toxicity studies; 2) the required neurotoxicity studies, including the developmental neurotoxicity study (D NT) have been submitted, are considered adequate; 3) although there is evidence of quantitative susceptibility in the DNT study, based on decreased survival of offspring up to postnatal day 4,Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 27 of 176 the endpoints and doses selected for risk assessment are protec tive for these effects; further, HED’s degree of concern for human susceptibility is reduced based on the special studies submitted in support of the mode of action; and 4) although some refinements are used in the exposure assessment, the dietary and drin king water assessments still result in upper

- bound estimates of exposure.

4.4 .1 Completeness o f the Toxicology Database

The toxicology database for sulfoxaflor is adequate for FQPA consideration including a developmental toxicity study in rats and rabb its, a 2

- generation reproduction study in rats, and acute, subchronic, and developmental neurotoxicity studies in rats

.

An acceptable guideline inhalation study is required for all food

- use registrations ; however in the case of sulfoxaflor, t he use of an oral study is considered protective for inhalation exposure for the following reasons: 1) The acute inhalation toxicity (Category IV) is lower than the acute oral LD 50 (Category III); 2) T he vapor pressure is low (≤1.4×10

- 6 Pa = 1.05×10 - 8 mm Hg at 20°C); 3) T here is adequate characterization of toxicity; 4) O ral absorption is >90%

– substantially higher ab s orption via the inhalation route is not expected; 5) O ral absorption is rapid; 6) S ulfoxaflor is not subject to enterohepatic circulat ion, making the absorbed oral dose available systemically; and

7 ) S ulfoxaflor is not subject to in vivo metabolism and therefore potential first

- pass effects are not relevant.

Therefore, the subchronic inhalation study is waived for sulfoxaflor. The lack of a subchronic inhalation study is not considered a data gap and a database uncertainty factor is not required ( TXR# 0056310)

.

4.4.2 Evidence of N eurotoxicity

The level of concern for neurotoxicity is low because the effects are well characterized , the dose

response curve for these effects is well characterized, and clear NOAELs have been identified

.

4.4.3 Evidence o f Sensitivity/Su sceptibility in the Developing o r Young Animal

Although there was quantitative susceptibility observe d in the DNT, there is no residual uncertainty because the effects are well characterized, a clear NOAEL was identified, and the endpoints chosen for risk assessment are protective of potential in utero and developmental

effects. Quantitative susceptibili ty in the DNT was based on an increased rate of neonatal deaths at a dose where no maternal toxicity was observed. However, the apparent enhanced sensitivity

may be due to the limited number of evaluations conducted in dams in the study rather than a

true sensitivity of the young. Qualitative susceptibility was observed in the 2

- generation reproduction study since neonatal deaths were observed at the same dose that resulted inSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 28 of 176 hepatotoxicity in parental animals. However, these effects occurred at a highe r dose compared to the offspring effects observed in the DNT. Finally, there was no evidence of quantitative or qualitative susceptibility in the developmental studies in the rat or rabbit.

4.4.4 Residual Uncertainty in the Exposure Datab ase

There are n o residual uncertainties with regard to dietary and occupational exposure. T he dietary exposure assessments are based on high

- end health protective residue levels (that account for parent and metabolites of concern), processing factors, and percent crop t reated assumptions (100%). Actual exposure to sulfoxaflor will likely be lower than the estimated exposure

. Furthermore, conservative, upper

- bound assumptions were used to determine exposure through drinking water sources, such that these exposures have not been underestimated.

4.5 Toxicity Endpoint and Point of D eparture Selections

4.5.1 Dose

- Response Assessment

Based on the use pattern and the toxicological profile of sulfoxaflor, HED selected endpoints and doses for acute and chronic dietary risk assessment. Endpoints and doses were not selected for

non

- dietary non

- occupational exposures (i.e., incidental oral, dermal, or inhalation), since there are no proposed uses which would result in these exposure pathways. For occupational handlers

and wor kers re

- entering treated areas, HED selected endpoints and doses for dermal and inhalation exposure and risk assessment.

Endpoint Selection

Acute Dietary Endpoint for the General Population: The endpoint used for establishing the aPAD for the general po pulation was selected from the acute neurotoxicity study in rats. An aPAD of 0.25 mg/kg/day was derived from a NOAEL of 25 mg/kg/day and a 100

- fold factor

that included 10x for inter

- species extrapolations, 10x for intra

- species variations, and a 1x FQPA SF. The LOAEL of 75 mg/kg was based on decreased motor activity on D ay 1. More severe effects were observed at the high dose (750 mg/kg)

, including increased muscle tremors and twitches, convulsions, hindlimb splaying, increased lacrimation and salivati on, decreased pupil size and response to touch, gait abnormalities and decreased rectal temperature. None of these effects were observed either 8 or 15 days after dosing. This endpoint is appropriate for the acute dietary exposure assessment because it i s the result of a single dose, is an appropriate route of exposure (oral) and is protective of the general population.

Acute Dietary Endpoint for Females 13

- 49 years of age: The endpoint used for establishing the

aPAD for females 13

- 49 years of age was s elected from the DNT study in rats. An aPAD of 0.06 mg/kg/day was derived from a developmental NOAEL of 1.8 mg/kg/day and a 30

- fold

factor that included 3x for inter

- species extrapolation, 10x for intra

- species variation, and 1x FQPA SF. The endpoint of concern is decreased neonatal survival in PND 0

- 4 offspring at the LOAEL of 7.1 mg/kg/day. Through a series of non

- guideline developmental studies in the rat, the critical window of susceptibility for skeletal effects and neonatal deaths was determined to be gestation days (GD) 19

- 20. Therefore, t his endpoint is appropriate because a single d ose d uringSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 29 of 176 GD 19

- 20 can be adverse. Furthermore, it is of a relevant route of exposure (oral), and is protective for effects seen in the most sensitive subgroup ( fetuses )

.

Chronic Dietary: The endpoint used for establishing the cPAD was selected from the chronic/carcinogenicity combined study in the rat. A cPAD of 0. 0 5 mg/kg/day was derived from

a NOAEL of 5.13 mg/kg/day and a 100

- fold factor that included 10x for inter

- species extrapolation, 10x for intra

- species variability, and a 1x FQPA SF. The LOAEL of 21.3 mg/kg was based on liver effects including increased blood cholesterol, liver weight, hy pertrophy, fatty change, single

- cell necrosis and macrophages i n males and females.

Dermal Absorption Factor: HED is using 2.4% as the factor for determining dermal

- equivalent doses in the sulfoxaflor risk assessment based on a comparison of a rat in vivo and rat and human

in vitro studies (see Section 4.2)

.

Inhala tion Absorption Factor: HED is assuming an inhalation absorption factor of 100%.

Dermal and Inhalation (short

- and intermediate

- term): The endpoint used for short

- and

intermediate

- term dermal exposure was selected from the DNT study in rats. The endpo int of

concern is decreased neonatal survival in PND 0

- 4 offspring with a NOAEL of 1.8 mg/kg/day and a LOAEL of 7.1 mg/kg/day. A 30

- fold uncertainty factor that includes a 3X factor for inter

species extrapolation and a 10x factor for intra

- species variab ility will be retained for risk assessment. Although a route

- specific dermal study was available for endpoint selection, the developmental endpoint was chosen from the DNT because it is protective of reproductive and developmental parameters not measured in the dermal study. As previously noted, in the case of sulfoxaflor, the use of an oral study is considered protective for assessing inhalation exposures.

4.5.2 Recommendation for Combining Routes of Exposures for Risk Assessment

T he developmental neur otoxicity study was selected for evaluating oral exposure as well as

short

- and intermediate

- term dermal and inhalation exposure s . T herefore, exposure estimates

from the se routes should be combined when assessing short

- and intermediate

- term risks based on the decreased neonatal survival endpoint

.

4.5.3 Cancer Classification and Risk Assessment Recommendation

The CARC in accordance with the EPA's Final Guidelines for Carcinogen Risk Assessment (March, 2005), determined that there is a “Suggestive Evidence of Carcinogenic Potential” for sulfoxaflor, based on the preputial gland tumor response seen in rats. When there is suggestive evidence, the Agency does not attempt a dose

- response assessment as the nature of the data generally would not support one. Therefore, the Agency has determined that quantification of risk using a non

- linear approach (i.e., reference dose (RfD) will adequately account for all chronic toxicity, including carcinogenicity, that could result from exposure to sulfoxaflor. The CARC considered the following factors in their weight

- of

- evidence deliberation on assessing the carcinogenic potential of sulfoxaflor.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 30 of 176 (i) The liver tumors observed in male rats and male and female mice were considered to be

treatment

- related;

(ii) The hypothesized mode of action (CAR mediated, mitogenic) for liver tumors was adequately supported by studies that clearly identified the sequence of key events, dose

response concordance and temporal relationship to the tumor type. There is convincing

ev idence that the hepatocarcinogenic effects are not likely to occur below a defined dose range. The mode of action data met the criteria established by the Agency;

(iii) The Leydig cell tumors were not considered to be treatment

- related in male rats beca use there is a high background rate for this tumor type in the particular strain of rats (F344) tested;

(iv) There was evidence of a response for carcinomas of the preputial glands in male rats. Preputial gland tumors are not commonly diagnosed in bioass ay studies. The available data were inadequate to draw confident conclusions regarding this response due to the small sample size and lack of histopathology data on all animals. Thus, it was not possible to determine whether the preputial gland tumors are due to treatment. If the response is positive, this would be an unusual finding. Because the tumors could not be discounted, they were used in the suggestive classification; and

(v) There is no mutagenicity concern from the in vivo or in vitro genetic t oxicity assays.

4. 5

. 4 Summary of Points of Departure and Toxicity Endpoints Used in Human Risk

Assessment

Table 4.5.4.1 Summary of Toxicological Doses and Endpoints for Sulfoxaflor for Use in Dietary and Non

Occupational Human Health Risk Assessments

. Exposure/

Scenario Point of

Departure Uncertainty/

FQPA Safety

Factors RfD & PAD for Risk

Assessment Study and Toxicological Effects

Acute Dietary (General Population, including Infants and Children) NOAEL=

25 mg/kg/day UF A = 10x

UF H =10x FQPA SF= 1x RfD = 0.25 m g/kg/day

aPAD =0.25 mg/kg/day Acute Neurotoxicity Study

LOAEL = 75 mg/kg/day based on decreased motor activity.

Acute Dietary (Females 13

- 49 years of age) NOAEL =

1.8 mg/kg/day UF A = 3x

UF H =10x FQPA SF= 1x RfD = 0.06 mg/kg/day

aPAD =0.06 mg/kg/day Developmental Neurotoxicity Study

LOAEL = 7.1 mg/kg/day based on decreased neonatal survival (PND 0

- 4)

Chronic Dietary (All Populations) NOAEL=

5.13 mg/kg/day UF A = 10x

UF H =10x FQPA SF= 1x RfD = 0.05

mg/kg/day

cPAD = 0.05 mg/kg/day Chronic/Carcinogenicity Study

- Rat

LOAEL = 21.3 mg/kg/day based on liver effects including increased blood cholesterol, liver weight, hypertrophy, fatty change, single cell necrosis and macrophages. Cancer (oral, dermal, inhalation) Sulfoxaflor is classi fied as “Suggestive Evidence of Carcinogenic Potential.” Quantification of

risk using a non

- linear approach (i.e., reference dose (RfD) will adequately account for all chronic toxicity, including carcinogenicity, that could result from exposure to sulfoxa flor.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 31 of 176 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. UF A = extrapolation from animal to human (interspecies). UF H = potential variation in sensitivity among members of the human population (intraspe cies). 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 4.5.4.2 Summary of Toxicological Doses and Endpoints f or Sulfoxaflor 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

- and

Intermediate Term NOAEL= 1.8

mg/kg/day

DAF: 2.4% UF A =3x

UF H =10x Occupational LOC

for MOE = 30 Developmental Neurotoxicity Study

LOAEL = 7.1 mg/kg/day based on decreased neonatal survival (PND 0

- 4)

. Inhalation Short

and Intermediate

Term NOAEL= 1.8

mg/kg/day * UF A =3x

UF H =10x Occupational LOC

for MOE = 30 Developmental Neurotoxicity Study

LOAEL = 7.1 mg/kg/day based on decreased neonatal survival (PND 0

- 4)

. Cancer (oral, dermal, inhalation) Sulfoxaflor is classified as “Suggestive Evidence of Carcinogenic Potential.” Quantification of

risk us ing a non

- linear approach (i.e., reference dose (RfD) will adequately account for all chronic toxicity, including carcinogenicity, that could result from exposure to sulfoxaflor.

* Inhalation absorption assumed equivalent to oral absorption.

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 ad verse effect level. LOAEL = lowest observed adverse effect level. UF = uncertainty factor. UF A = extrapolation from animal to human (interspecies). UF H = potential variation in sensitivity among members of the human population (intraspecies). MOE = ma rgin of exposure. LOC = level of concern. N/A = not applicable.

5.0 Dietary Exposure and Risk Assessment

5.1 Metabolite/Degradate Residue Profile

5.1.1 Summary of Plant and Animal Metabolism Studies

HED has reviewed studies depicting the metabolis m of sulfoxaflor in plants, livestock, and rats. In plants, metabolism was investigated with lettuce, pea, rice, and tomato (with foliar and soil application for each). In addition, metabolism was studied in radish, lettuce, and wheat grown as

rotational crops. Livestock metabolism studies were conducted with lactating goat and laying

hen with both sulfoxaflor and the plant metabolite X11719474 . All of the submitted studies meet guideline requirements and are considered to be acceptable.

In plants, m et abolism of sulfoxaflor was similar in all four target crops . O xidation of the cyano

carbon bond results in the form ation of X11719474, followed by loss of the sulfur side chain to form X11721061. X117211061 is then conjugated with glucose, which may then be conjugated with malonic acid. Metabolism continues through incorporation of the radiolabeled carbon into natural plant constituents, such as lignin and starch. A very minor pathway (<1%) included degradation of the X11719474 urea side chain to form X 1159540 and X11479457, which are further reduced to give X11721061. The primary difference noted between the foliar and soil metabolism studies was that parent sulfoxaflor was the predominant residue found from foliar applications, while X11719474 was the major resi due found after application of sulfoxaflor toSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 32 of 176 the soil

. Similarly, X11719474 was the predominant residue observed in the rotational crop

studies.

Metabolism studies with goats, hens, and rats consistent ly showed that sulfoxaflor is rapidly

abs orbed and eliminated (chiefly via urine in mammals), and undergoes very little in vivo

transformation. The results of studies in which X11719474 was administered to goats and hens

mirrored the studies conducted with sulfoxaflor in that the administered do se was rapidly absorbed and eliminated, primarily as unmetabolized X11719474.

5.1.2 Summary of Environmental Degradation

The fate of sulfoxaflor in the environmental is highly dependent on whether or not it is in a soil system, groundwater system, or surface water system. Environmental fate studies show sulfoxaflor to be stable to photolysis (soil and aqueous) as well as hydrolysis across a broad pH

range (estimated t 1⁄2 = 141 days in aerobic water, 637 days in anaerobic water) . In soil systems,

howeve r, sulfoxaflor is rapidly degraded to X11719474 (estimated t 1⁄2 = 0.4 days). Figure 5.1.2 depicts the fate of sulfoxaflor in the environment. In terms of drinking water, the fate pathways and their kinetic properties result in sulfoxaflor being the expecte d predominant residue in

surface water. Contrariwise , and despite its high mobility , sulfoxaflor is not expected in groundwater due to its rapid degradation to X11719474 and, to a much lesser extent,

X11519540.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 33 of 176 Figure 5.1.2. Expected environmental degradation pathways and transformation profiles for

sulfoxaflor . Diagram taken from D 382617 , M. Ruhman, 1 August 2012

.

5.1.3 Comparison of Metabolic Pathways

Although some metabolism was noted in studies with target crops (foliar application), livestock, and rats, sulfoxaflor does not generally show significant metabolism in those systems. Crops treated foliarly with sulfoxaflor tend to show more metabolism th an the other two systems, but

even then , there is little breakdown beyond oxidative loss of the cyano group and formation of X11719474. Environmental fate and rotational crop studies indicate that once sulfoxaflor is in

contact with the soil

, the parent c ompound is short lived and X11719474 predominates.

X11719474 was not detected in livestock or rats.

5.1.4 Residues of Concern Summary and Rationale
E. Holman, 6 April 2012 , D398294

HED’s Residues of Concern Knowledgebase Subcommittee (ROCKS) has recomme nded the

residues of concern summarized in Table 5.1.4 for sulfoxaflor. Although t he global review partners concur with the recommended residue definition in terms of regulatory enforcement

,

r isk assessment strategies differ in terms of metabolites of con cern. X 11519 540 Rapid Bio

- degradation

(90 th percentile t1⁄2= 0.4 days) (Aerobic Soil System)

Slow Aqueous Photolysis (t1/2= 261 days) Very Slow Aqueous Photolysis ( Practically Stable) Sulfoxaflor

X117189

- 22 X 11719 474

X115794

- 57 Very Slow Biodegradation* (90 th percentile t1⁄2= 825 days) (Aerobic Soil System) X117210

- 61Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 34 of 176 Table 5.1.4. Summary of Sulfoxaflor Residue Definitions for Risk Assessment and Tolerance Enforcement.

Matrix Risk Assessment Tolerance Expression

Plants Cotton Sulfoxaflor Sulfoxaflor

Rotational Crop Sulfoxaflor 1 Sulfoxaflor

Livestock Ruminant Sulfoxaflor Sulfoxaflor

Poultry Sulfoxaflor Sulfoxaflor Drinking Water Sulfoxaflor + X11719474 + X11519540 NA

1 Based on the proposed later season foliar application use pattern; decision may need to be revisited for future early season soil application uses.

Hazard Considerations

HED has determined that X11719474 does not share the reproductive MOA that was presented

for sulfoxaflor ( CARC Memo, TXR 0056301 )

. However, exposure to X11719474 does results in hepatotoxicity comparable to th at observed for sulfoxaflor. Based on several toxicology studies, the hepatotoxicity of X11719474 is 3

- to 7

- times less potent than sulfoxaflor. X11579457, a minor soil metabolite, exhibits no genotoxicity

, and no acute oral toxicity. Structurally, it is quite similar to X11719474, and is therefore considered to be substantially less

toxic than parent sulfoxaflor

. The other primary plant metabolite, X11721061, is also considered to be significantly less toxic than the parent based on subchronic toxicity s tudies and is not mutagenic. Little toxicity data are available for X11596066 , however, it has no structural alerts for genotoxicity, is negative in the Ames test , is structurally similar to X11721061 , and therefore,

is of low concern for human health. C ontrary to the other metabolites, X11519540 appears to have a relative hepatotoxic potency that is greater than sulfoxaflor, as demonstrated in LD50 and

28

- day studies in the rat.

Exposure Considerations

Water. Due to rapid degradation of sulfoxaflor to X11719474, f or the purposes of drinking water modeling, initial X11719474 levels are expected to be essentially equivalent to the amount of parent that reaches the soil directly following application (not intercepted by foliage) and

amounts washed

- off fol iage later. In the long term, there is potential for X11719474 to degrade

to metabolites X11519540 and X115794 57 . Although not detected in U.S. aerobic soil metabolism studies, these metabolites were observed at higher levels in European studies. X11579457, a minor soil metabolite, is structurally quite similar to X11719474, which exhibits significantly less toxicity than parent sulfoxaflor

. Therefore, X11579457 is excluded from the residue definition due to its decreased toxicity and low overall exposure levels. As previously noted, X11519540 appears to be more toxic than sulfoxaflor in terms of liver toxicity; t herefore, despite the generally very low overall exposure to X11519540, this metabolite should be included, along with parent sulfoxaflo r and X11719474 , for the purposes of the drinking water risk assessment and modeling.

Plants (Primary Crops). The parent compound together with the two main plant metabolites, X11719474 and X11721061, are present in most of the primary crops tested in crop field trials. It is noted that all field trials were conducted at higher rates than those proposed for actual use, making the anticipated exposure levels to these metabolites even lower than recorded in the field studies. Due to the lower toxicity o f X11719474 and X11721061 , as well as generally lower overall exposure compared to parent sulfoxaflor, these metabolites are excluded from the residueSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 35 of 176 definition. The metabolite X11519540 was only observed at low concentrations (<1% of the TRR) in the let tuce metabolism study and was not observed at all in the other plant metabolism studies. Consequently, X11519540 is excluded from the residue definition due to low potential for exposure through plant matrices. Therefore, the inclusion of only parent sul foxaflor should be adequate for dietary risk assessment. For tolerance enforcement, parent sulfoxaflor is also considered an adequate indicator of misuse

. Although the parent may not be a good marker for misuse in crops like potatoes, the metabolites wou ld not be much better indicators based on the

field trial results showing that residues in root crops are likely to be very low (<0.04 ppm)

.

Rotational Crops

. Although X11719474 is the predominant residue observed in rotational crops

, assessing dietary r isk based on exposure to parent only is appropriate. F ield rotational crop studies demonstrate that the maximum amount of X11719474 taken up by succeeding crops is similar to the maximum amount present in primary crops after foliar treatments. As these

s ummed levels of X11719474 are still low relative to the amount of parent sulfoxaflor in most primary crops, risk assessments based on parent sulfoxaflor will be protective for any X11719474 residues taken up by rotational crops. An exception to the preced ing generalization regarding relative metabolite levels was noted for the root and tuber vegetables. However, for these crops, the maximum residue levels are low (<0.04 ppm). Thus, a separate residue of

concern definition for rotational root and tuber ve getable crops was not considered necessary. Also, because there are few potential rotational crops that are not proposed for primary crop tolerances, concerns regarding rotational crop issues are low. Based on the above considerations

combined with its s ubstantially lower toxicity relative to sulfoxaflor, X11719474 is excluded from the residue definition. For tolerance enforcement, parent sulfoxaflor is considered an adequate indicator of misuse for the proposed application methods

. Should there ever be soil directed applications of sulfoxaflor to primary crops, there is a potential for significant residues

of X11719474 in rotated crops , requiring this decision to be revisited.

Livestock

. Based on the available information, the inclusion of parent sulfoxaflor only should be sufficient for dietary risk assessment. With the exception of liver, t he majority of the residue in milk, eggs, and edible tissues consisted of the parent compound , generally >80% of the residue. X11596066 was a major metabolite in ruminant and hen liver (14

- 18% TRR), but has been determined to have low overall toxicity; therefore, it is excluded from the residue definition.

X11 5 19540 was detected at trace levels in most of the edible samples (<2% of the TRR in the

ruminant and ≤ 8% of the TRR in the hens). The actual ppm levels of X11 5 19540 were <0.01 ppm in all matrices. However, due to the potential for increased toxicity associated with

X11 5 19540 , the sulfoxaflor team consult ed with the HED Toxicology Science A dvisory Council (ToxSAC) in order to adequately account for the relative potency of this metabolite. Following consultation with ToxSAC, the sulfoxaflor team has determine d that it is appropriate to exclude

residues of X11 5 19540 in livestock commodities i n the dietary risk assessment. It is noted that

the draft 2011 JMPR decision excludes this metabolite from their residue definition due to the low overall observed concentrations. For tolerance enforcement, parent sulfoxaflor is considered an adequate in dicator of misuse

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 36 of 176 5 .2 Food Residue Profile
D. McNeilly, 12 September 2012 , D398756

The petitioner is requesting establishment of tolerances for residues of sulfoxaflor in/on 39 crops/crop groups. In support of that request, the petitioner has provided data from 583 field trials as well as supporting data related to metabolism, storage stability, analytical methods, processed commodities, residue transfer to livestock, and uptake in rotational crops. All of the studies are considered to be acceptable an d adequate for evaluating regulatory levels and assessing risk.

Field trials were generally conducted in accordance with the proposed use directions; although application rates in the trials were somewhat exaggerated (1.4X) relative to the proposed labels

. Generally, residues of sulfoxaflor were well above the limit of quantitation (LOQ) of 0.01 ppm in most commodities. The exception to that is in root crops where maximum residues were only slightly greater than the LOQ and rotational crops where residue s were generally non

quantifiable. Studies indicate that residues are distributed throughout the crops and are not restricted to treated surfaces. Sulfoxaflor was stable in studies designed to mimic boiling, cooking, and pasteurization . T ypical food pro cessing activities, such as baking, boiling and canning

, are unlikely to significantly affect the nature and magnitude of sulfoxaflor residues. In commodity

- specific processing studies, sulfoxaflor generally concentrated during production of

dried foods a nd tended to be reduced in juices, though exceptions to those trends are evident in

the database. A summary of the field trial and processing study results are provided in Appendix

D

.

The request for registration of sulfoxaflor includes uses on a large n umber of commodities that are considered to be significant feed items. To account for transfer of sulfoxaflor residues from livestock feeds into livestock commodities, HED has derived maximum, reasonably balanced dietary burden s for beef cattle, dairy cat tle, poultry, and swine and combined those estimates with data from dairy cattle and laying hen feeding studies in order to calculate anticipated

residues in livestock commodities. For poultry and swine, quantifiable residues are not expected in any commo dity. Nevertheless, HED is recommending for tolerances at the LOQ (0.01 ppm) for harmonization purposes. Quantifiable residues are more likely in commodities, including milk, derived from cattle, goat s, horses, and sheep, and HED is recommending for tole rances ranging from 0.10 to 0.40 ppm in those commodities.

5.3 Water Residue Profile
M. Ruhman, 1 August 2012 , D 382617

Based on abiotic laboratory studies, sulfoxaflor is stable to photolysis on soil and to hydrolysis in acidic, neutral and alkaline syst ems. Aqueous photolysis is expected to be slow and may not be considered an important transformation route in the natural environment (t1⁄2 >1,000 days in de

ionized water and 637 days in natural water). In the aerobic soil system, sulfoxaflor degrades rather quickly to its major degradate X 11719 474 (90 th percentile t1⁄2= 0.4 days). Other residues, forming as a result of aerobic soil biotransformation, include limited amounts of X

- 1157957 and

X 11519 540. Anaerobic soil data suggest that anaerobic conditi o ns produces the main degradate

, X 11719 474. In contrast to the rapid degradation of sulfoxaflor in aerobic soil systems, theSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 37 of 176 chemical degrades rather slowly to X 11719 474 in both aerobic and anaerobic aquatic conditions (90 th percentile t1⁄2= 141 days).

Sul foxaflor is considered to be non

- persist ent in soil and exhibits low affinity to soil/sediment particles. The chemical is not expected to partition significantly into the air from wet or dry

soils

. Sources of surface water contamination include drift for parent and run

- off for X11719474 and X11519540 following the rapid degradation of parent in the soil. Parent compound that drifts into surface water is expected to degrade rather slowly to X11719474. In contrast, the source of ground water contamination is expected to be leaching of parent and/or its degradation products X11719474 and X11519540, all of which are considered to be highly mobile.

Monitoring data depicting residues of sulfoxaflor in drinking water are not available. The

Environmental Fate and Effects Division has used modeling to estimate upper

- bound concentrations of sulfoxaflor in surface and ground water. Two scenarios were modeled: (1)

Use of sulfoxaflor on non

- aquatic row and orchard crops and (2) use of sulfoxaflor on

watercress. F or the non

- aquatic crops, screening

- level estimated drinking water concentrations (EDWCs) were calculated using Tier II PRZM (Pesticide Root Zone Model) and EXAMS (Exposure Analysis Modeling System) for surface water sources and the Tier I SCI

- GROW ( S creen ing C oncentration in Gro und W ater) model was used for calculating the EDWCs in ground water sources. Of the requested non

- aquatic

- crop uses, those on citrus resulted in the highest EDWCs and are presented in Table 5.3.1 a

.

Table 5.3.1 a . Estimated Drinking Water Concentrations (EDWCs) for Total Toxic Residues of Sulfoxaflor in

Surface and Ground Waters from Use on Citrus

.

Water Source * Total Concentration, ppb [Sulfoxaflor + X11719474 + X11519540]

Acute Peak 1 Non

- cancer/Chronic 2 Cancer 3

Surface (PRZM/EXAMS) 26.4 [ 9.4 + 15.0 + 2.0 ] 13.5 [ 1.9 + 10.2 + 1.4 ] 9.3 [ 1.5 + 6.9 + 0.9 ]

Ground (SCI

- GROW) 69.2 [ 0.0 + 60.9 + 8.3 ]

* Surface water EDWCs include sulfoxaflor + X11719474 + X11519540. Due to rapid degradation of sulfoxaflor in aerobic soil systems (half

- life = 0.4 days), the EDWC for groundwater includes X11719474 + X11519540.

1 Peak value , from Florida citrus ; 2 1

- in

- 10

- year annual mean values , Ca lifornia citrus ; 3 1

- in

- 10

- year overall annual

mean values , California citrus

.

The Tier I rice model was used t o obtain surface water EDWC from the use of sulfoxaflor on

watercress . The results of those analyses are presented in Table 5.3.1b and include the accumulation of residues that is expected from multiple applications.

Table 5.3.1 b . Estimated Drinking Water Concentrations (EDWCs) for Total Toxic Residues of Sulfoxaflor in

Surface and Ground Waters from Use on Watercress

.

Water Source * Total Concentration, ppb [ Sulfoxaflor + X11719474] One Application Two Applications Three Applications

Surface ( Tier 1 Rice Model ) 91.3 [26.5 + 64.8] 182.5 [52.9 + 129.6] 273.8 [79.4 + 194.4]

* Aquatic

- use EDWCs include sulfoxaflor + X11719474.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 38 of 176 The values in Table s 5.3. 1 a and 5.3. 1 b represent upper

- bound estimates of residue concentrations that might be found in drinking water due to the use of sulfoxaflor on non

- aquatic crops and watercress, respectively

.

Risk Assessment Considerations

Although the EDWCs associated with the use of sulfoxaflor on watercress are higher than those

for the non

- aquatic crops, HED’s risk assessment is focusing on the non

- aquatic

- crop EDWCs. The 2007 census of agriculture estimates that approximately 680 acres of the U.S. are used for watercress production ; thus, this use represents a very small fraction of the potential crop acreage that may be treated with sulfoxaflor. Moreover, the inputs to the Tier 1 rice model are quite

conservative , especially with rega rd to application efficiency (the model assumes that there is no interception of the applied material by the watercress plants) and the 10

- cm water column at the time of application (information from watercress growers indicates that watercress fields are drained prior to pesticide applications). Finally, the rice model predicts pesticide concentrations in water in the field and not drinking water per se where concentrations are expected to be lower

due to dissipation processes such as degredation, stream flow, and dilution

. While the use on
watercress may impact drinking water for a few individuals , HED does not believe that the EDWCs and residue profiles associated with the watercress use give a representative depiction of

the potential exposure profile for the vast majority of the U.S. population. HED has quantitatively estimated risks based on the watercress EDCWs and found the risk estimates to be below our level of concern ( D. McNeilly, D401670, 12 September 2012 ). We note that if a

future use for s ulfoxaflor is requested on rice or cranberry, the EDWCs of primary interest would likely be those coming from the rice model.

Based on the modeled estimates associated with non

- aquatic crops, the potential exposure through drinking water is much higher from groundwater than from surface water. Due to the different relative toxicities of the X11719474 and X11519540 metabolites, HED requested that EFED provide estimates for each compound separately. Environmental fate data indicate that the residue in groundwater will be comprised, predominantly, of the X11719474 metabolite. Quantitatively, the 69.2 ppb EDWC for groundwater can be divided into 60.9 ppb from X11719474 and 8.3 ppb from X1 1519450. HED has used these estimates and the relative toxicities of the metabolites to adjust the drinking water component of the dietary risk assessment, as outlined below. As noted earlier, sulfoxaflor presents a somewhat unique situation regarding th e food and drinking water exposure pathway. Environmental fate data indicate that use of sulfoxaflor is likely to result in different residue profiles in surface water and groundwater , namely that sulfoxaflor is likely to be the predominant residue in sur face water and X11719747/X11519540 will predominate in groundwater . When the residue profiles are coupled with the toxicological database, it becomes apparent that the EDWCs for assessing acute dietary exposure for the general population, acute dietary ex posure for women of child

- bearing age, and chronic dietary exposure for all populations need to be addressed differently. An explanation of the three scenarios and the rationale for the approaches taken by HED is provided

below . T he EDWCs used in the var ious dietary assessments are summarized in Table s 5.3.2 a and 5.3.2b

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 39 of 176 Acute Exposure

Separate acute endpoints were selected for the general population and females 13

- 49 years of

age . For the general population, the point of departure is based on decreased motor activity observed in the acute neurotoxicity study. As there are no data available to examine the potency

of X11719474 and X11519540 with respect to this endpoint, HED has a ssumed that the two

metabolites possess similar toxicity relative to sulfoxaflor in order to assess acute dietary risk for the general population. The EDWC for groundwater is significantly greater than the acute

estimate for surface water and, per HED pol icy, is being used in the acute dietary assessment for the general population. As it is a groundwater EDWC, it represents residues of the metabolites.

For females 13

- 49 years of age , the developmental endpoint of increased neonatal deaths was chosen because a single exposure during late gestation can adversely affect the developing fetus and the age group represents women of child

- bearing age

. As discussed in Section 4.3.4, the

meta bolites X11719474 and X1159540 are not likely to result in agonism of the muscle nicotinic receptor and , therefore, should be excluded from assessment scenarios using the developmental endpoint

. Since the groundwater EDWC represents residues of these meta bolites only, the acute surface water EDWC becomes the appropriate estimate for assessing dietary exposure for

women of child

- bearing age

.

Chronic Exposure

The endpoint for assessing chronic dietary exposure is hepatotoxicity. As discussed in Section

4. 3.4, HED has determined that it is appropriate to combine residues of sulfoxaflor, X11719474, and X11519540 when assessing chronic exposure and, furthermore, there is sufficient evidence to adjust the assessment to account for the different potencies of th e metabolites. Based on

NOAELs in the 28

- day oral toxicity studies in rats, the potencies of the metabolites, relative to sulfoxaflor, are 0.3X for X11719474 and 10X for X11519540. To account for the relative toxicity, the EDWCs for each metabolite are m ultiplied by their respective potency factor.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 40 of 176 Table 5.3.2 a . Adjusted Estimated Drinking Water Concentrations (EDWCs) from Use on C itrus for Estimating Dietary Risk. Drinking Water

Scenario Water Source Toxic Residue, ppm Adjustment

Factor Adjusted

Residue

, ppm Rationale

Acute

– General Population Groundwater Sulfoxaflor 0 1 0 EDWC: Groundwater >

Surface Water

Endpoint is decreased motor activity. No information is

available to rule out applicability or estimate relative toxicity. X11719474 = 0.0 609 1 0.0 609

X11519540 = 0.00 83 1 0.00 83

Total 0.0 692

Acute

– Females

13

- 49 years old Surface water Sulfoxaflor = 0.00 94 1 0.00 94 Residues in groundwater are not appropriate for assessing the endpoint.

Chronic

– All Populations Groundwater Sulfoxaflor 0 1 0 EDWC: Groundwater >

Surface Water

Endpoint is liver effects. 28

- day studies are available for sulfoxaflor, X11719474, and

X11519540 to derive relative

toxicities. X11719474 = 0.0 609 0.3 0.01 83

X11519540 = 0.00 83 10 0.0 83

Total 0. 1013

Table 5.3.2b. Adjusted Estimated Drinking Water Concentrations (EDWCs) from Use on Watercress for Estimating Dietary Risk. Drinking Water

Scenario Water Source Toxic Residue, ppm Adjustment

Factor Adjusted

Residue,

ppm Rationale

Acute

– General Population Groundwater Sulfoxaflor = 0.0794 1 0.0794 EDWC: Groundwater >

Surface Water

Endpoint is decreased motor activity. No information is

available to rule out applicability or estimate relative toxicity. X11719474 = 0. 1944 1 0. 1944

X11519540 = 0.0 1 0.0

Total 0. 2738

Acute

– Females

13

- 49 years old Surface water Sulfoxaflor = 0.0 794 1 0.0 794 Residues in groundwater are not appropriate for assessing the endpoint.

Chronic

– All Populations Groundwater Sulfoxaflor = 0.0794 1 0.0794 EDWC: Groundwater >

Surface Water

Endpoint is liver effects. 28

- day studies are available for sulfoxaflor, X11719474, and

X11519540 to derive relative

toxicities. X11719474 = 0. 1944 0.3 0.0 583

X11519540 = 0.0 10 0.0

Total 0. 1377Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 41 of 176 5.4 Dietary Risk Assessment
D. McNeilly, 12 September 2012 , D 401670

Acute and chronic aggregate dietary (food and drinking water) exposure and risk assessments were conducted using the Dietary Exposure Evaluation Model DEEM

- FCIDTM (v. 2.03). This

model uses food consumption data from the U.S. Department of Agriculture’s (USDA’s) Continuing Surveys of Food Intakes by Individuals (CSFII) from 1994

- 1996 and 1998. Based on analysis of the 1994

- 96, 98 CSFII consumption data, which took into account dietary patter ns and survey respondents, HED concluded that it is most appropriate to report risk for the following population subgroups: the general U.S. population, all infants (<1 year old), children

1

- 2, children 3

- 5, children 6

- 12, youth 13

- 19, adults 20

- 49, female s 13

- 49, and adults 50+ years

old.

5.4.1 Description of Residue Estimates Used in Dietary Assessment

The data used in the acute assessment reflect several refinements relative to a screening

- level

assessment. Maximum residue values from field trials were used rather than tolerance

- level residue estimates. For crop groups, th e res idue values were translated from representative crops to the other crops in the group using HED SOP 2000.1. F or processed commodities , empirical processing factors were used for all commodities unless an empirical factor was not available, in

which case the DEEM default estimate was used. Residue estimates for livestock were derived using maximum observed residues in the cattle and hen feeding studies.

For the chronic asses sment , the same refinements were made as those described for the acute assessment, with two exceptions: (1) average residue levels from crop field trials were used rather than maximum values and (2) average residues from feeding studies, rather than maxim um values, were used to derive residue estimates for livestock commodities.

Estimates of residues in drinking water were incorporated directly into the assessments

. As previously discussed (Section 5.3), HED is focusing on the dietary risk estimates that incorporate

the non

- aquatic

- crop EDWCs rather than the watercress EDWCs. Nevertheless, HED has estimated dietary risk s using the watercress EDWCs and all estimates are below HED’s level of

concern.

5.4.2 Percent Crop Treated Used in Dietary Assessment

HED has assumed 100% of crops covered by the registration request are treated with sulfoxaflor.

5.4.3 Acute Dietary Risk Assessment

Acute dietary exposure was estimated for all of the standard, representative population groups

described above. As previo usly noted, the assessment for women of child

- bearing age (females

13

- 49 years old) i s based on a different endpoint / point of departure and includes a different set of assumptions regarding residues in drinking water than were used for the other groups. G iven the assumptions used in conducting the acute assessment, exposure and risk estimates (Table 5.4.6) should be considered moderately refined. The use of the highest residue values from field trials give some refinement in comparison to use of tolerance

- level residues . H owever, crop fieldSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 42 of 176 trials are designed to result in worst

- case residue levels and the assumption that all crops with proposed uses of sulfoxaflor are treated (100% crop treated) and that they bear residues at

maximum level s is likely to overestimate actual acute exposure. Significant refinement to the dietary exposure estimates could likely be obtained by using the full distribution of field trial data in a probabilistic assessment.

Acute dietary risk estimates range fro m 4 % to 16 % of the acute population

- adjusted dose (aPAD), with the highest risk estimate s being for children 1

- 2 years old and females 13

- 49 years

old

. Generally, HED is concerned when exposure estimates exceed 100% of the population

adjusted dose (PAD). Even with the conservatisms in the assessment, acute dietary risk

estimates are below HED’s level of concern.

5.4.4 Chronic Dietary Risk Assessment

Chronic dietary exposure was estimated for all of the standard, representative population groups

descri bed in Section 5.4 above. The chronic dietary exposure estimates reflect significant refinement with respect to residue levels, although overall, the use of field trial data and the assumption of 100% crop treated results in estimates that are moderately refined. Monitoring data, if available, and incorporation of use statistics (i.e., % crop treated) have the potential to provide significant refinements to the assessment.

Chronic dietary risk estimates range from 5 % to 1 8 % of the chronic population

- adju sted dose (cPAD). T he highest risk estimate is for infants and all of the risk estimates are below HED’s

level of concern.

5.4.5 Cancer Dietary Risk Assessment

Sulfoxaflor is classified as having "suggestive evidence of carcinogenicity" and HED has

conc luded that assessments using a non

- linear approach (e.g., a chronic RfD

- based assessment) will adequately account for all chronic toxicity, including carcinogenicity that could result from exposure to sulfoxaflor. Chronic dietary risk estimates are below HED’s level of concern; therefore, cancer risk is also below HED’s level of concern.

5.4.6 Summary Table

Table 5.4.6. Summary of Acute and Chronic Dietary Exposure and Risk Estimates for Sulfoxaflor.

Population Subgroup Acute (95 th Percentile) Chronic Cancer

Estimated Exposure (mg/kg/day) Estimated

Risk, %

aPAD Estimated

Exposure (mg/kg/day) Estimated

Risk, %

cPAD Exposure

(mg/kg/day) Estimated

Risk
U.S. Population 0.0 15832 6 0.00 3343 7

See Chronic All infants 0.0 31158 12 0.009110 18

Children 1

- 2 yrs 0.04 4887 1 6 0.007867 16

Children 3

- 5 yrs 0.0 32523 1 3 0.00 6210 12

Children 6

- 12 yrs 0.0 18338 7 0.00 3892 8

Youth 13

- 19 yrs 0.0 10829 4 0.00 2456 5

Adults 20

- 49 yrs 0.0 11137 4 0.00 2812 6 Adults 50+ yrs 0.01 1446 5 0.00 2992 6Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 43 of 176 Table 5.4.6. Summary of Acute and Chronic Dietary Exposure and Risk Estimates for Sulfoxaflor.

Population Subgroup Acute (95 th Percentile) Chronic Cancer

Estimated Exposure (mg/kg/day) Estimated

Risk, %

aPAD Estimated

Exposure (mg/kg/day) Estimated

Risk, %

cPAD Exposure

(mg/kg/day) Estimated

Risk

Females 13

- 49 yrs 0.009495 16 0.00 2809 6 Entries with the highest acute and chronic risk estimates are bolded

.

6.0 Residential (Non

- Occupational) Exposure/Risk Characterization

N o residential uses, and no commercial/professional uses at residential sites are being requested

for sulfoxaflor at t his time; therefore, no residential risk assessment has been conducted. There

are proposed use s on ornamental plants and turfgrass , but those uses are for

nurseries / commercial greenhouses and for sod farms , respectively

. For sulfoxaflor

, HED

believe s that the post

- application exposure for homeowners buying and planting ornamental plants or turf

grass (sod) would be minimal based on the exposure data available , the time

between application and purchase, and the watering

- in of turf

. At this time, there are no uses of sulfoxaflor that will result in residential exposures.

6. 1 Residential Bystander Post

- A pplication Inhalation Exposure

Based on the Agency's current practices, a quantitative post

- application inhalation exposure assessment was not performed for sulfoxaflor at this time primarily because of the acute inhalation toxicity (Toxicity Category III and IV), low vapor pressure ( ≤1.4×10 - 6 Pa = 1.05×10 - 8
mm Hg at 20°C ) , and the maximum seasonal proposed use rate ( 0. 266 lb ai/A /season ). However, volatilization of pesticides may be a source of post

- application inhalation exposure to individuals nearby pesticide applications. The Agency sought expert advice and input on issues related to volatilization of pesticides from its Federal Insect icide, Fungicide, and Rodenticide Act Scientific Advisory Panel (SAP) in December 2009, and received the SAP’s final report on March 2, 2010 (http://www.epa.gov/scipoly/sap/meetings/2009/120109meeting.html). The Agency is in the process of evaluating the SAP report and may, as appropriate, develop policies and procedures to identify the need for and, subsequently, the way to incorporate post

application inhalation exposure into the Agency's risk assessments. If new policies or procedures are developed, th e Agency may revisit the need for a quantitative post

- application inhalation exposure assessment for sulfoxaflor

.

6. 2 Spray Drift

Spray drift is always a potential source of exposure to residents nearby to spraying operations. This is particularly the c ase with aerial application, but, to a lesser extent, could also be a potential source of exposure from the ground application method employed for sulfoxaflor . The Agency has been working with the Spray Drift Task Force, EPA Regional Offices and State

Lea d Agencies for pesticide regulation and other parties to develop the best spray drift management practices (see the Agency’s Spray Drift website for more information at http://www.epa.gov/opp00001/factsheets/spraydrift.htm). On a chemical by chemical basi s, the Agency is now requiring interim mitigation measures for aerial applications that must be placed on product labels/labeling. The Agency has completed its evaluation of the new databaseSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 44 of 176 submitted by the Spray Drift Task Force, a membership of U.S. pe sticide registrants, and is developing a policy on how to appropriately apply the data and the AgDRIFT computer model to its risk assessments for pesticides applied by air, orchard airblast and ground hydraulic methods. After the policy is in place, the A gency may impose further refinements in spray drift management practices to reduce off

- target drift with specific products with significant risks

associated with drift.

Although a quantitative residential post

- application inhalation exposure assessment wa s not performed as a result of pesticide drift from neighboring treated agricultural fields, an inhalation exposure assessment was performed for flaggers. This exposure scenario is representative of a worse case inhalation (drift) exposure and may be cons idered protective of most outdoor agricultural and commercial post

- application inhalation exposure scenarios. The risk estimate for flaggers (MOE ≥ 4,200, Table 9.1 ) is not of concern

.

7.0 Aggregate Exposure/Risk Characterization

As there are no regist ered or requested residential uses of sulfoxaflor, aggregate exposure is equivalent to the dietary exposure discussed in Section 5. Therefore aggregate risk estimates are
below HED’s level of concern.

8.0 Cumulative Exposure/Risk Characterization

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 sulfoxaflor and any other substances and sulfoxaflor does not appear to produce a toxic metabolite produced by other substances. For the purposes of this tolerance action, therefore,

EPA has not assumed that sulfoxaflor has a common mechanism of toxicity with other

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

9.0 Occupational Exposure/Risk Characterization
Z. Figueroa, 18 September 2012, D 382606

T wo agricultural products are proposed for use on the various crops , turf grass and ornamentals : GF

- 2032 SC and Transform TM WG.

GF

- 2032 SC is formulated as a suspension concentrate (SC) which contains 21.8 % of the active ingredient, sulfoxaflor

. The Personal Protective Equipment (PPE) statement on the proposed

label requires applicators and other handlers to wear : long

- sleeved shirt and long pants, and shoes plus socks

.

Transform TM WG is formulated as a w ater dispersible granule (WG) which contai ns 50% of the active ingredient, sulfoxaflor

. The Personal Protective Equipment (PPE) statement on the proposed label adds protective eyewear to the PPE found on the GF

- 2032 SC label (i.e., long

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 45 of 176 sleeved shirt and long pants, shoes plus socks, and protecti ve eyewear ) due to the Category II finding in the acute ocular toxicity testing with Transform TM WG

.

These end use products may be applied through a variety of application methods including: aerial, chemigation (for potatoes only), ground boom, airblast, b ackpack, and hand held equipment. There is a potential for occupational exposure associated with handler activities (i.e., mixing, loading, and applying) as well as with post

- application activities (i.e., re

- entering treated areas). No data on the number of exposure days per year was provided. Based on the proposed labels’ use pattern and directions (e.g., 2

- 4 applications per season with a minimum 5

- to 14

- day spray interval), only short

- (1 to 30 days) and intermediate

- term (1 to 6 months) exposures are expected for handlers.

In the past HED has considered ornamentals and greenhouse uses to have the potential for long

term dermal exposure. However, the pests that are targets for sulfoxaflor have biological and

life

- stage properties that provide for short windows of opportunity for control. T o avoid development of sulfoxaflor

- resistant pests, the sulfoxaflor labels require switching periodically to another insecticide. Therefore, based on the proposed use pattern and resistance managemen t strategies (i.e., avoid consecutive applications of insecticides with the same mode of action), long

- term exposure is not expected to occur.

9.1 Short

- and Intermediate

- Term Handler Risk Estimates

HED uses the term handlers to describe those individu als who are involved in the pesticide application process. HED believes that there are distinct job functions or tasks related to applications

, and that exposures can vary depending on the specifics of each task. Job requirements ( e.g., amount of chemica l used in each application), types of equipment used, the

area treated or amount handled , and the level of protection used by a handler can cause exposure levels to differ in a manner specific to each application event. Detailed descriptions of the

handle r assessment are available in D 382606 (Z. Figueroa, 18 September 2012 ) and a summary table of the handler exposure and risk estimates is provided in Appendix F

. T he occupational

hander exposure and risk estimates rely on generic data from the Pesticide Handlers Exposure Database Version 1.1 (PHED 1.1), the Outdoor Residential Exposure Task Force (ORETF)

database and the Agricultural Handler Exposure Task Force (AHETF) database.

All combined (dermal + inhalation) occupational handler scenarios for the proposed uses resulted in estimated MOEs that are greater than 30 (i.e., not of concern) at the baseline level of PPE (i.e.

, baseline clothing, no gloves

, and no respirator) and engineering controls (enclosed cockpit) for aerial applications (Table 9.1) . The highest risk estimate ( MOE = 80 ) is associated with handlers conducting mixer/loader activities with dry flowable formulation (TransformTM WG) for aerial applications.

HED has no data to assess exposures to pilots using open cockpits. The on ly available data are for exposure to pilots in enclosed cockpits. Therefore, risks to pilots are assessed using the engineering control (enclosed cockpits) and baseline attire (long

- sleeve shirt, long pants, shoes, and socks); pilots are not required to wear protective gloves. With this level of protection, there

are no ri sks of concern for applicators.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 46 of 176Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 47 of 176 Table 9.1. Summary of Occupational Handler Exposure and Risk Estimates for Sulfoxaflor. *
Activity Scenario Use Site Formulation Dermal Inhalation

Dose (mg/kg/day) MOE

(LOC=30) Dose

(mg/kg/day) MOE

(LOC=30) Combined

MOE (LOC=30)

Mixer/Loader Groundboom

Application (AHETF) Brassica (Cole) Leafy Veg., Bulb Veg.

(CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG

13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry

Beans Transform TM WG 0.00057 3,200 0.00089 2,000 1,200

GF

- 2032 SC 0.00057 3,100 0.000023 79,000 3,000

Ornamentals, Turf grass (Sod Farm) Transform TM WG 0.00088 2,100 0.00140 1,300 800

GF

- 2032 SC 0.00085 2,100 0.000034 53,000 2,000 Barley, Tricale, Wheat, Canola (Sub CG 20A) Transform TM WG 0.00077 2,300 0.00120 1,500 900

GF

- 2032 SC 0.00069 2,600 0.000027 66,000 2,500 Cotton, Soybean Transform TM WG 0.0014 1,300 0.00220 810 490

GF

- 2032 SC 0.00140 1,300 0.000057 32,000 1,200

Aerial Application (AHETF) Brassica (Cole) Leafy Veg., Bulb Veg.

(CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG

13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry

Beans Transform TM WG 0.00250 730 0.00390 460 280

GF

- 2032 SC 0.00250 720 0.00010 18,000 690

Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) Transform TM WG 0.00380 470 0.00600 300 180

GF

- 2032 SC 0.00370 490 0.00015 12,000 470

Barley, Tricale, Wheat, Canola (Sub CG 20A) Transform TM WG 0.00460 390 0.00730 250 150

GF

- 2032 SC 0.00410 440 0.00016 11,000 420 Cotton, Soybean Transform TM WG 0.00850 210 0.01340 130 80

GF

- 2032 SC 0.00860 210 0.00034 5,300 200 Chemigation (AHETF) Potatoes Transform TM WG 0.00250 730 0.00390 460 280

GF

- 2032 SC 0.00250 720 0.00100 18,000 690Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 48 of 176 Table 9.1. Summary of Occupational Handler Exposure and Risk Estimates for Sulfoxaflor. *
Activity Scenario Use Site Formulation Dermal Inhalation

Dose (mg/kg/day) MOE

(LOC=30) Dose

(mg/kg/day) MOE

(LOC=30) Combined

MOE (LOC=30)

Airblast Application (AHETF) Citrus (CG 10), Pome Fruits (CG 11),

Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) Transform TM WG 0.00044 4,100 0.00069 2,600 1,600

GF

- 2032 SC 0.00042 4,200 0.000017 110,000 4,100

Applicator Groundboom

Application (AHETF) Brassica (Cole) Leafy Veg., Bulb Veg.

(CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG

13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Dry Podded,

Beans Transform TM WG 0.00020 9,200 0.000034 53,000 7,800

GF

- 2032 SC 0.00021 8,800 0.000035 51,000 7,500

Ornamentals, Turf grass (Sod Farm) Transform TM WG 0.00030 5,900 0.000052 34,000 5,100

GF

- 2032 SC 0.00030 5,900 0.000052 34,000 5,100 Barley, Tricale, Wheat, Canola (Sub CG

20A) Transform TM WG 0.00027 6,700 0.000046 39,000 5,700

GF

- 2032 SC 0.00024 7,300 0.000042 42,000 6,300 Cotton, Soybean Transform TM WG 0.00049 3,700 0.000085 21,000 3,100

GF

- 2032 SC 0.00051 3,500 0.000089 20,000 3,000

Aerial Application (AHETF) Brassica (Cole) Leafy Veg., Bulb Veg.

(CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG

13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Dry Podded,

Beans Transform TM WG 0.000055 33,000 0.000030 61,000 21,000

GF

- 2032 SC 0.000057 32,000 0.000031 58,000 25,000

Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) Transform TM WG 0.000084 21,000 0.000046 39,000 14,000

GF

- 2032 SC 0.000084 21,000 0.000046 39,000 19,000

Barley, Tricale, Wheat, Canola (Sub CG

20A) Transform TM WG 0.00010 18,000 0.000056 32,000 11,000

GF

- 2032 SC 0.000093 19,000 0.000051 35,000 12,000 Cotton, Soybean Transform TM WG 0.00019 9,600 0.0001 18,000 6,200

GF

- 2032 SC 0.00020 9,200 0.00011 17,000 6,000Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 49 of 176 Table 9.1. Summary of Occupational Handler Exposure and Risk Estimates for Sulfoxaflor. *
Activity Scenario Use Site Formulation Dermal Inhalation

Dose (mg/kg/day) MOE

(LOC=30) Dose

(mg/kg/day) MOE

(LOC=30) Combined

MOE (LOC=30)

Airblast Application (AHETF) Citrus (CG 10), Pome Fruits (CG 11),

Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) Transform TM WG 0.00340 530 0.00036 5,000 480

GF

- 2032 SC 0.00340 530 0.00036 5,000 480

Flagger Flagging for

Aerial Application (PHED) All Low Acreage Crops Transform TM WG 0.00019 9,700 0.00024 7,600 4,200

GF

- 2032 SC 0.00019 9,700 0.00024 7,600 4,200 All High Acreage Crops Transform TM WG 0.00012 15,000 0.00015 12,000 6,700

GF

- 2032 SC 0.00013 14,000 0.00016 11,000 6,300

Mixer/ Loader/ Applicator Mechanically

pressurized Handgun Sprayer Ornamentals, Greenhouses Transform TM WG 0.00169 1,100 0.00231 780 450

GF

- 2032 SC 0.00169 1,100 0.00231 780 450

Turfgrass (Sod farms) Transform TM WG 0.00047 3,800 0.00040 4,400 2,100

GF

- 2032 SC 0.00047 3,800 0.00040 4,400 2,100

Backpack Sprayer Ornamentals Transform TM WG 0.00025 7,200 0.00011 16,000 5,000

GF

- 2032 SC 0.00025 7,200 0.00011 16,000 5,000

Turfgrass (Sod farms) Transform TM WG 0.00016 11,000 0.000002 900,000 11,000

GF

- 2032 SC 0.00016 11,000 0.000002 900,000 11,000 Manually

pressurized

Handwand (PHED) Ornamentals

, Turfgrass (Sod farms) Transform TM WG 0.00193 930 0.000023 78,000 920

GF

- 2032 SC

0.00193 930 0.000023 78,000 920

* From Z. Figueroa, 18 September 2012, D 382606Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 50 of 176 9.2 Short

- and Intermediate

- Term Post

- Application Risk Estimates

9.2.1 Dermal Post

- A pplication Risk Estimates

Occupational workers who enter treated fields to perform post

- application activities such as hand weeding and scouting may be exposed to sulfoxaflor residues

. Based on the use pattern, workers may be exposed to short

- and intermediate

- term exposure duration s.

Dislodgeable Foliar Residue Data: I t is HED ’s policy to use the best available data to assess
post

- application exposure. The registrant submit ted dislodgeable foliar r esidue (DFR) studies for

wheat and broccoli in support of th e proposed agricultural uses. Both studies demonstrated that dislodgeable foliar residue values declined over time for all sites and both formulations

. The available DFR data were matched to the proposed agricultural crop s (excep t cotton; see below) as surrogate data based on available information about foliage type, region of use, use pattern,

etc. Dermal post

- application exposure resulting from application of sulfoxaflor was estimated using the best available surrogate DFR

- pred icted Day

- 0 residue value ; Day

- 0 DFR values were adjusted by application rate. Maximum single application rates for each crop/crop groups were used in this assessment.

Dislodgeable residue data were not submitted for cotton. Calculation of dermal post

- application exposure resulting from application of sulfoxaflor to cotton during harvesting activities (e.g., module builder, picker operator, raker, tramper) requires an estimate of the "dislodgeable"

residue on the cotton bolls. Since that information w as unavailable, exposure was estimated using cotton field trial residues (see Appendix D). Additionally, since the proposed PHI for cotton is 14 days, the Day

- 14 residue value from the cotton field trials is considered adequate to estimate dermal post

- app lication exposures and risks during harvesting activities. Adjusted to the proposed maximum seasonal application rate, the Day

- 14 residue is 0.000129 mg ai/g cotton.

Note also that the cotton field trial residue data reflect solvent

- extractable residues, which are likely to be significantly greater than dislodgeable cotton boll residue during harvesting activities. Therefore, use of the residue data from the cotton field trials, even after being reduced to account for the difference in application rate, results in a conservative estimate for the daily

dose.

Turf transferable residue (TTR) data were not submitted for turfgrass (sod farms); therefore,

default TTR values were used to assess dermal post

- application exposure and risk.

Transfer Coefficients: Sources of generic post

- application exposure data, used as surrogate data

in the absence of chemical

- specific data, are derived from Agricultural Reentry Task Force (ARTF) exposure monitoring studies, and, as proprietary data, are subject to the data protection provisions of FIFRA. The standard values recommended for use in predicting post

- application exposure that are used in this assessment, known as “transfer coefficients, Tc ” are presented in the “Science Advisory Council for Exposure (ExpoSAC) Po licy 3” (http://www.epa.gov/pesticides/science/exposac_policy3.pdf), which, along with additional information about the ARTF data, can be found at: http://www.epa.gov/pesticides/science/post

app

- exposure

- data.html.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 51 of 176 Dermal exposures during post

- application activities were estimated using standard default and surrogate contact activity values (see Appendix G ) , chemical

- specific DFR for all agricultural crops except cotton, default TTR for sod farms, and field trial data for cotton

.

A summary of the post

- ap plication exposure and risk estimates for all proposed crops except

cotton is provided in Table 9.2.1

. Only risk estimates for the highest contact activity are shown

below. Post

- application dermal exposure and risk estimates resulted in MOEs greater than 30

and are not of concern on Day

- 0 (i.e., 12 hours after application)

.

A summary of the post

- application exposure and risk estimates from use of sulfoxaflor on cotton during harvesting activities is provided in Table 9.2.2 . Post

- application dermal exp osure and risk estimates resulted in MOEs greater than 30 and are not of concern.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 52 of 176 Table 9.2.1 . Post

- Application Exposure and Risk Estimates for use of Sulfoxaflor. *

Crop Group Proposed Uses/Groups Proposed

Maximum Application

Rate (lb ai/A) Activity Daily Dose (mg/kg/d) MOE

DFR Data on Treated Wheat (MRID No. 47832415)

Field / Row Crop,

Low/Medium Barley, Wheat

Canola 0.047 Scouting 0.0000007 2,600,000

Cotton 0.090 Scouting 0.0000003 7,000,000 Soybean 0.090 Scouting 0.0000014 1,300,000

Succulent Edible Podded and Dry Beans 0.090 Irrigation (Hand Set) 0.0000023 770,000

DFR Data on Treated Broccoli (MRID No. 47832416) Vegetable, head

and stem

Brassica Brassica (Cole) Leafy

Vegetables 0.090 Hand Weeding 0.00001 130,000

Vegetable,

"Root" Bulb Vegetables (Crop

Group 3

- 07) 0.090 Hand Weeding 0.00001 130,000

Leaves of Root and Tuber Vegetables (Crop Group 2); Root and Tuber Vegetables (Crop Group 1) 0.090 Irrigation (Hand Set) 0.000006 280,000

Tree Fruit, Evergreen Citrus (Crop Group 10) 0.133 Thinning Fruit 0.00002 100,000

Vegetable,

Cucurbit Cucurbits Vegetables

(Crop Group 9) 0.090 Irrigation (Hand Set) 0.00001 280,000

Vegetable, Fruting Fruiting Vegetables

(Crop Group 8) 0.090 Irrigation (Hand Set) 0.00001 280,000

Vegetable, Leafy Leafy Vegetables

(except Brassica

– Crop Group 4 & Watercress) 0.090 Irrigation (Hand Set) 0.000006 280,000

Nursery/

Greenhouse (Unassigned) Ornamentals 0.133 Irrigation (Hand Set) 0.00001 190,000

Tree, "Fruit",

Deciduous Pome Fruits (Crop

Group 11); Stone Fruits (Crop Group 12) 0.133 Thinning Fruit 0.00002 100,000

Vine/Trellis Small Fruit Vine

Climbing (except fuzzy

kiwifruit

– Subgroup

13

- 07F) 0.090 Turning, Girdling 0.0001 28,000

Berry, Low Low Growing Berry

(Subgroup 13

- 07G) 0.090 Irrigation (Hand Set) 0.000006 280,000

Tree, “Nut” Tree Nuts (Crop Group

14 & Pistachio) 0.133 Hand Harvesting 0.000007 260,000

Default TTR Approach
Turf, Sod Turfgrass, Sod farms 0.133 Maintenance, Harvesting, Slab Transplanting/ Planting 0.0000004 4,700,000

* Taken from Z. Figueroa, 18 September 2012, D 382606Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 53 of 176 Table 9.2.2 . Post

- Application Exposure and Risk Estimates for use of Sulfoxaflor on Cotton during Harvesting

Activities. *

Crop Proposed Maximum

Seasonal Application Rate (lb ai/A) Activity Days

After

Treatment Residue

from Cotton

Field Trial 2
(mg ai/g cotton) Daily Dose

(mg/kg/d) 3 MOE 4

Cotton 0.266 Harvesting Mechanical (Module Builder)

14 0.000129 0.00034 5,300

Harvesting Mechanical (Picker Operator; Raker) 0.00089 2,000

Harvesting Mechanical (Tramper) 0.00189 950

* Taken from Z. Figueroa, 18 September 2012 , D 382606

. 1. Transfer Coefficients from HED Policy No. 3.0., Non

- Foliar Transfer Coefficient Table, March 2012 (p 122).

2. Residue Value from Cotton Field Trial (Table 9) adjusted to Max. Seasonal Application Rate (mg ai/g cotton) = (0.182 mg ai/kg cotton) (0.001 mg/g) ((0.266 lb ai/A)/(0.375 lb ai/A)) = 0.000129 mg ai/g

cotton.
3. Dermal Dose (mg/kg/day) = [Cotton Field Trial Residue Value adjusted (mg ai/g cotton) x TC (g cotton/hr) x hr/day (8 hrs/day) x DAF (2.5%)]/ BW (69 kg).

4. Dermal MOE = NOAEL (1.8 mg/kg/day)/Dermal Dose (mg/kg/day); LOC = 30.

Restricted Entry Interval (REI)

Although acute toxicity testing of technical

- grade sulfoxaflor and the GF

- 2032 SC end

- use product support an REI of 12 hours, the petitioner has proposed REIs of 24 hours for both end

use products based on their interpretation of the TransformTM WG acute ocular toxicity study resulting in a Toxicity Category II. The petitioner believes that the higher toxicity of Transform TM WG is due to an inert ingredient in that product. It is the petitioner’s understanding that the REI for all products containing a particular active ingredient must be the maximum dictated by testing of the individu al products.

HED recommends that RD ensure that the proper REI is listed on both labels according to the acute toxicity profile and the formulation for each product.

9.2.2 Inhalation Post

- A pplication Risk Estimates

Based on the Agency's current practices, a quantitative post

- application inhalation exposure assessment was not performed for sulfoxaflor at this time primarily because it has a low vapor pressure ( 1.05 × 10 - 8 mm Hg at 25°C ) and it is applied at low application rates ( maximum single

ap plication rate of 0.133 lb ai/A). However, volatilization of pesticides may be a potential source of post

- application inhalation exposure to individuals nearby to pesticide applications. The Agency sought expert advice and input on issues related to vola tilization of pesticides from its Federal Insecticide, Fungicide, and Rodenticide Act Scientific Advisory Panel (SAP) in December 2009. The Agency received the SAP’s final report on March 2, 2010 (http://www.epa.gov/scipoly/SAP/meetings/2009/120109meeting .html).Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 54 of 176 Although a quantitative occupational post

- application inhalation exposure assessment was not performed, an inhalation exposure assessment was performed for occupational handlers. This

assessment resulted in risk estimates that did not exceed HE D’s level of concern at baseline inhalation PPE. Handler exposure resulting from application of pesticides outdoors is likely to result in higher exposure than post

- application exposure. Therefore, it is expected that these handler inhalation exposure es timates would be protective of most occupational post

- application inhalation exposure scenarios.

Furthermore, the Worker Protection Standard for Agricultural Pesticides contains requirements for protecting workers from inhalation exposures during and afte r greenhouse applications through the use of ventilation requirements [40 CFR 170.110, (3) (Restrictions associated with pesticide applications)].

10.0 References

Memoranda

Holman, E. 6 April 2012. D398294. “ Sulfoxaflor. Report of the Residues of Co ncern Knowle d gebase Subcommittee (ROCKS). ” McNeilly, D. 12 September 2012 . D398756. “ Sulfoxaflor. Petition for the Section 3 Registration and Establishment of T olerances for the New Insecticide Sulfoxaflor on:

B arley, Brassica Leafy Vegetables, Bulb Vegetables, Canola (Rapeseed), Citrus, Cotton, Cucurbit Vegetables, Fruiting Vegetables, Leafy Vegetables (Except Brassica ), Leaves of Root and Tuber Vegetables, Low Growing Berry, Okra, Ornamentals (Herbaceous and Woody), Pistachio , Pome Fruits, Root and Tuber Vegetables, Small Fruit Vine Climbing (Except Fuzzy Kiwifruit), Soybean, Stone Fruits, Beans Succulent, Edible Podded, and Dry Beans, Tree Nuts, Triticale, Turf Grass, Watercress, Wheat. Summary of Analytical Chemistry and Re sidue Chemistry Data. ” Figueroa, Z. 18 September 2012

. D382606 . “ Sulfoxaflor

. Occupational and Residential Exposure and Risk Assessment to Support the Registration of the New Active Ingredient on a Variety of Food Crops , Turfgrass (Sod Farms) and Ornam entals

. ” Mcneilly, D. 12 September 2012. D401670. “ Sulfoxaflor: Acute and Chronic Aggregate Dietary Exposure and Risk Assessment for Section 3 Uses on Barley, Brassica Leafy Vegetables, Bulb Vegetables, Canola (Rapeseed), Citrus, Cotton, Cucurbit Vegetables, Fruiting Vegetables, Leafy Vegetables (Except Brassica), Leaves of Root and Tuber Vegetables, Low

- Growing Berries, Pistachio, Pome Fruits, Root and Tuber Vegetables, Small Fruit Vine Climbing (Except Fuzzy Kiwifruit), Soybean, Stone Fruits, Suc culent Beans, Edible Podded, and Dry Beans, Tree Nuts, Triticale, Watercress, and Wheat. Ruhman, M. 1 August 2012. D382617. “ Estimated Drinking Water Concentrations of the New Chemical Sulfoxaflor from Proposed use on Beans, Berries, Canola, Citrus, Frui ts (Pome & Stone), Ornamentals, Soybeans, Tree nuts, Turf grass, Vegetables (Brassica, Bulb, Cucurbits, Fruiting, Okra, Leafy, Root and Tuber) and Watercress for the Use in

Human Health Risk Assessment. ” Kidwell, J. 26 April 2012. TXR No. 0056301. “SULF OXAFLOR : Report of the Cancer

Assessment Review Committee .”Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 55 of 176 Van Alstine J. 14 September 2012 . TXR# 0056310. Sulfoxaflor: Summary of Hazard and Science Policy Council (HASPOC) Meeting of March 15, 2012: Recommendation on

the need for a 28

- day inhalatio n study.

Literature Studies

Boorman, J. P. B., P. J. GrootKormelink and L. G. Sivilotti (2000). "Stoichiometry of human recombinant neuronal nicotinic receptors containing the β3 subunit expressed in Xenopus oocytes." The Journal of Physiology 529 (3): 565

- 577. Hill, A. B. (1965). "The Environment and Disease: Association or Causation?" Proceedings of the Royal Society of Medicine 58 : 295

- 300. Krashia, P., M. Moroni, S. Broadbent, G. Hofmann, S. Kracun, M. Beato, P. J. Groot

Kormelink and L. G. Sivilott i (2010). "Human α3β4 Neuronal Nicotinic Receptors Show Different Stoichiometry if They Are Expressed in <italic>Xenopus</italic> Oocytes or

Mammalian HEK293 Cells." PLoS ONE 5 (10): e13611. Luetje, C. W. and J. Patrick (1991). "Both alpha

- and beta

- subunit s contribute to the agonist sensitivity of neuronal nicotinic acetylcholinesterase receptors." The Journal of

Neuroscience 11 (3): 837

- 845. Millar, N. (2010). "Characterization of the Agonist Effects of XDE

- 208 on Mammalian Muscle Nicotinic Acetylcholine Re ceptors." Report of the University College London, London, UK. Millar, N. S. (2009). "A review of experimental techniques used for the heterologous expression of nicotinic acetylcholine receptors." Biochemical Pharmacology 78 (7): 766

- 776. Moroni, M. and I. Bermudez (2006). "Stoichiometry and pharmacology of two human α4β2 nicotinic receptor types." Journal of Molecular Neuroscience 30 (1): 95

- 96. Moroni, M., R. Vijayan, A. Carbone, R. Zwart, P. Biggin and I. Bermudez (2008). "Non

Agonist

- Binding Subunit Inte rfaces Confer Distinct Functional Signatures to the Alternate Stoichiometries of the alpha4beta2 Nicotinic Receptor: An alpha4

– alpha4 Interface Is Required for Zn2+ Potentiation." The Journal of Neuroscience 28 (27): 6884

- 6894. Stern, P., M. Cik, D. Colquho un and F. A. Stephenson (1994). "Single channel properties of cloned NMDA receptors in a human cell line: comparison with results from Xenopus oocytes." The Journal of Physiology 476 (3): 391

- 397.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 56 of 176 A ppendix A . Toxicology Profile and Executive Summaries

A.1 Toxicology Data Requirements

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

Study 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

- waived 870.3700a Developmental Toxicity (rodent)

................. 870.3700b Developmental Toxicity (nonrodent)

........... 870.3800 Reproduction

................................ ................ yes

yes

yes yes

yes

yes 870.4100a Chronic Toxicity (rodent)

............................. 870.4100b Chronic Toxicity (nonrodent)

...................... 870.4200a Oncogenicity (rat)

................................ ........ 870.4200b Oncogenicity (mouse)

................................ .. 870.4300 Chronic/Oncogenicity

................................ .. yes

yes

yes

yes

yes yes

yes

yes

yes

yes 870.5100 Mutagenicity

— Gene Mutation

- bacterial

... 870.5300 Mutagenicity

— Gene Mutation

- mammalian

................................ ................................ ...... 870.5xxx Mutagenicity

— Structural Chromosomal

Aberrations

................................ ................................ ...... 870.5xxx Mutagenicity

— Other Genotoxic Effects

..... yes

yes

yes

yes yes

yes

yes

yes

870.6100a Acute Delayed Neurotoxicity (hen)

............. 870.6100b 90

- Day Neurotoxicity (hen)

......................... 870.6200a Acute Neurotoxicity Screening Battery (rat)

870.6200b 90

- Day Neurotoxicity Screening Battery (rat)

................................ ................................ .................. 870.6300 Develop. Neurotoxicity

................................ no

no

yes

yes

yes -

- yes

yes

yes

870.7485 General Metabolism

................................ ..... 870.7600 Dermal Penetration

................................ ...... 870.7800 Immunotoxicity

................................ ............ yes

yes

yes yes

yes

yesSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 57 of 176 Study Technical

Required Satisfied
Special Studies for Ocular Effects

Acute Oral (rat)

................................ ............. Subchronic Oral (rat)

................................ .... Six

- month Oral (dog)

................................ .... no

no

no -

-

-

A.2 Toxicity Profiles

Sulfoxaflor Acute Toxicity Profile

Guideline

No. MRID

(Annex II Point) Route/

method Species/

strain (sex) Result Classification

Sulfoxaflor

870.1100 47832144

(5.2.1) Oral/

gavage Rat/

Fischer 344 (both) LD 50 =1000

mg/kg Category III

47832040 (5.2.1) Oral/

gavage Mouse/ CD1

(male) LD 50 =750

mg/kg Category III

870.1200 47832051

(5.2.2) Dermal/

topical Rat/

Fischer 344 (both) LD 50 >5000

mg/kg Category IV

870.1300 47832052

(5.2.3) Inhalation/

nose only Rat/

Fischer 344 (both) LC 50 >2.09 mg/L

(4hr) Category IV

870.2400 47832055

(5.2.5) Eye/

instillation Rabbit/

NZW (female) Slight Irritation Category III

870.2500 47832053

(5.2.4) Dermal/

topical Rabbit/

NZW (female) Minimal

Irritation Category IV

870.2600 47832041

(5.2.6) Dermal/

LLNA Mouse/ CBA/J

(female) No Sensitization None

Metabolites

Metabolite X11719474

Non

guideline 47832308

( 5.8.1.2/1 ) Oral/gavage

300 mg/kg/day Rat/F344

(female) No Observed

Effects Not Applicable

870.1100 47832131

(5 .8.1.2/2 ) Oral/gavage

2000, 5000 mg/kg/day Rat/F344

(female) No death LD50

> 5000. Signs for 48

hours Category IV

Non

guideline 47832309

( 5

. 8.1.2/3 ) Dermal

1000 mg/kg/day Rat/F344

(female) No death.

Irritation for 48

hours Not Applicable

Non

guideline 47832309

(5.8.1.2/3) Eye

0.1 ml (0.069 g) Rabbit/NZW

(female) Conjunctivitis

for 24 hours Not ApplicableSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 58 of 176 Sulfoxaflor Toxicity Profile Table

Guideline

No. Study Dosages Effects at lowest observed effect level

870.3050 Oral 28

- day F344 Rat

MRID 47832045

Annex 5.3.1/2

Acceptable/Guideline 0, 300, 1000, 2000, 3000 ppm

M/F: 0/0, 24.8/26.5, 79.4/88.3, 155/170, 205/192 mg/kg/day LOAEL

– 88.3 mg/kg/day based on liver effects including serum cholesterol, liver weight and hepatocellular hypertrophy with altered tinctorial properties.

NOAEL

– 26.5 mg/kg/day

870.3050 Oral 28

- day CD1Mouse

MRID 47832042

Annex 5.3.1/4

Acceptable/Guideline M: 0, 300/350, 1500, 3500

ppm F: 350, 1500, 3500 ppm

0/0, 43.9/53, 230/273,

524638 mg/kg/day LOAEL

– 230/273 M/F based on liver effects including ALT, AST, liver weight, hepatocellular hypertrophy in males and females. Fatty change, multifocal single cell necrosis and mitotic figures in mal es only

NOAEL

- 43.9/53 mg/kg/day M/F

Non

Guideline Oral 28

- day in Beagle

Dogs

MRID 47832043

Annex 5.3.1/6

Acceptable Group 1 (dietary)

- F: 500 ppm The only tolerable dose and route was 15 mg/kg/day technical grade XDE

- 208 via oral gavage in Methocel vehicle. Exposure at 100 ppm via diet was well tolerated as determined by adequate food consumption, but test material intake was insufficient to justif

y its use as a high dose level for subsequent studies. Group 2 (gelatin capsule)

- F: 15 mg/kg/day Group 3 (oral gavage

analytical grade XDE 208 in Methocel)

- F: 15 mg/kg/day Group 4 (dietary)

- F: 100 ppm 870.2600 47832130

( 5.8.1.2/4 ) Dermal

50% / rLLNA CBA/J Mice

(female) Negative Category IV

Metabolite X11721061

870.1100 47832086

( 5.8.2.2/1 ) Oral/Gavage

1000, 2000, 5000 mg/kg/day Rat/F344

(female) LD50 > 2000 Category III

Metabolite X11596066

870.1100 47832067

( 5.8.3.2/1 ) Oral/gavage

2000 mg/kg/day Rat/F344

(female) No deaths.;

LD50 > 2000

Piloerection for

24 hours Category III

Metabolite X11579457

870.1100 47832145

( 5.8.4.2/1 ) Oral/gavage

2000 mg/kg/day Rat/F344

(female) No death; LD50

> 2000

Piloerection for

24 hours Category III

Metabolite X11519540

870.1100 47832069

( 5.8.5.2/1 ) Oral/gavage

320,1000,2000 mg/kg/day Rat/F344

(female) LD50 565; No

death at 320.

Death at 1000

and 2000 Category IIISulfoxaflor Human Health Risk Assessment DP No. D382604

Page 59 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

Group 5 (oral gavage

technical grade XDE 208 in Methocel)

- F: 15 mg/kg/day
870.3100; 870.6200;

870.7800 90 day oral toxicity

Neurotoxicity Screening Immunotoxicity

Oral 90

- day F344 Rat

with 28

- day Recovery

MRID 47832046

Annex 5.3.2/1

Acceptable/Guideline 0, 100, 750, 1500 ppm

M/F: 0/0, 6.36/6.96, 47.6/51.6, 94.9/101 mg/kg/day LOAEL

– 47.6/51.6 mg/kg/day (M/F)

based on 11

- 13% body weight gain.

Liver

- serum cholesterol, liver weight, hepatocellular hypertrophy with altered tinctorial properties, and multifocal single

cel l necrosis;Fatty change in males.

NOAEL = 6.36/6.96 mg/kg/day (M/F)

There were no indications of neurotoxicity (NOAEL ≥ 94.9 mg/kg/day)

There were no indications of immunotoxicity (NOAEL ≥ 94.9 mg/kg/day)

870.3100 Oral 90

- day CD1Mouse

MRID 47832057

Annex 5.3.2/2

Acceptable/Guideline M: 0, 100, 750, 1250 ppm (0, 12.8, 98, 166 mg/kg/day)

F: 0, 100, 1500, 3000 ppm (0, 16.2, 247, 489 mg/kg/day) LOAEL

– 98/247 mg/kg/day (M/F) based on liver effects including liver weight, hepatocellular hypertrophy with altered tinctorial properties, multifocal single cell

necrosis.

Adrenal

- very slight hypertrophy/vacuolization of zona

fasciculata.

Females also had ALT, AST and triglycerides

NOAEL

– 12.8/16.2 mg/kg/day (M/F)

870.3150 Oral 90

- day Dog/Beagle

(gavage)

MRID 47832058

Annex 5.3.3/1

Acceptable/Guideline 0, 1, 3, 6 mg/kg/day (10 mg/kg/day days 1

- 5) LOAEL

– 10 mg/kg/day based on food consumption

NOAEL = 6 mg/kg/day

870.3200 28

- day Dermal F344 Rat

MRID 47832059

Annex 5.3.7/1

Acceptable/Guideline 0, 100, 500, 1000 mg/kg/day LOAEL

- > 1000 mg/kg/day ;

Females: No effects

Males : 17% serum cholesterol; 6.5%

absolute and 4.4% relative liver weights; very slight centrilobul ar to midzonal hepatocellular hypertrophy with altered tinctorial properties . Although

treatment

- related, these effects were not

considered adverse.

NOAEL = 1000 mg/kg/daySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 60 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

870.3700 Developmental Toxicity

in CD Rats (Definitive)

MRID 47832140

Annex 5.6.10/2

Acceptable/Guideline 0, 25, 150, 1000 ppm

(0, 2, 11.5, 70.2 mg/kg/day) Dam : LOAEL

– 70.2 mg/kg/day based on reduced feed consumption and body weight gain; increased liver weight.

NOAEL = 11.5 mg/kg/day

Litter : LOAEL

– 70.2 mg/kg/day based on decreased fetal body weight; fetal abnormalities (forelimb flexure, bent clavicle, hindlimb rotation, convoluted/hydro

- ureter).

NOAEL = 11.5 mg/kg/day

870.3700 Developmental Toxicity

Dietary in NZW Rabbits (Definitive)

MRID 47832066

Annex 5.6.11/3

Acceptable 0, 30, 150, 750 ppm

(0, 1.3, 6.6, 31.9 mg/kg/day ) Dam : LOAEL

– 31.9 mg/kg/day based on decreased feed consumption (8

- 21%), body weight gain, (12%) and fecal output.

NOAEL = 6.6 mg/kg/day

Litter : LOAEL > 31.9 mg/kg/ day

NOAEL = 31.9 mg/kg/day

Toxicokinetics: AUC α Dose, dose adjusted AUC similar

to rat 870.3800;

OECD 421 Reproduction/

Developmental Probe

CD Rat

MRID 47832143

Annex 5.6.1/1

Acceptable/Guideline 0, 100, 500, 1000 ppm Reproduction : LOAEL = 39.5 mg/kg/day based on

decreased neonatal survival. Postnatal day (PND) 4 survival 81.2% (95.4% in controls).

NOAEL = 8.1 mg/kg/day (PND 4 pup survival 7.3%.)

Parental toxicity : NOAEL > 78.2 mg/kg/day

– D ecreased body weight gain was observed at 500 and 1000 ppm in males and females. Increased liver weight in males at 500 and 1000 ppm. Hepatocellular hypertrophy in males at 500 and 1000 ppm and females at 1000 ppm. These effects were

considered minimal and not adverse.

Offspring : LOAEL

– 39.5 mg/k g/day based on decreased neonatal survival (see above).

NOAEL = 8.1 mg/kg/daySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 61 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

870.3800 2

- Generation

Reproduction in CD Rats

MRID 47832142

Annex 5.6.1/2

Acceptable/Guideline 0, 25, 100, 400 ppm

(1.52/1.91, 6.07/7.82, 24.6/30.5 mg/kg/day [m/f]) Reproduction : LOAEL

– 24.6 mg/kg/day based on decreased neonatal survival (ca. 2

- 5%)

NOAEL = 6.07 mg/kg/day

Parental toxicity : LOAEL

– 24.6 mg/kg/day based on increased liver weight in with correlating histopathologic changes

NOAEL = 6.07 mg/kg/day

Off spring : LOAEL

– 24.6 mg/kg/day based on decreased neonatal survival and a slight delay in preputial separation (puberty onset) in F1 males

NOAEL = 6.07 mg/kg/day

870.4100 Oral 1

- year Beagle Dog

(gavage)

MRID 48288906

Annex 5.3.4/1

Acceptable/Non

guideline 0, 1, 3, 6 mg/kg/day LOAEL > 6 mg/kg/day NOAEL = 6 mg/kg/day

870.4200 18

- month carcinogenicity

CD1 Mouse

MRID 47832054

Annex 5.5.3

Acceptable/Guideline ♂: 0, 25, 100, 750 ppm

0, 2.54, 10.4, 79.6 mg/kg/day LOAEL

– 79.6 mg/kg/day based on liver effects including adenomas and carcinomas; Increased weight, hypertrophy with eosinophilia, fatty change, single cell necrosis, eosinophilic/ vacuolated foci,

mitosis.

NOAEL = 10.4 mg/kg/day ♀: 0, 25, 250, 1250 ppm

0, 3.43, 33.9, 176 mg/kg/day LOAEL

– 176 mg/kg/day based on liver effects including carcinomas; increased weight, hypertrophy with eosinophilia, fatty change, single cell necrosis and eosinophilic/ vacuolated foci.

NOAEL = 33.9 mg/kg/daySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 62 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

870.4300 2

- year combined toxicity

and carcinogenicity dietary

F344 Rat

MRID 47832060

Annex 5.5.1 & 5.5.2

Acceptable/Guideline 0, 25, 100, 500 ppm

M/F: 0/0, 1.04/1.28, 4.24/5.13, 21.3/39 mg/kg/day Males:

1

- year: LOAEL

– 21.3 mg/kg/day based on liver effects including increased blood cholesterol, liver weight, hypertrophy, fatty change, single cell necrosis and macrophages.

2

- year: LOAEL

– 21.3 mg/kg/day based on

increased serum cholesterol concentrations, and histopa thological liver

effects in males ; increased incidence and size of Leydig cell adenomas with secondary effects including preputial gland tumours; liver adenomas.

NOAEL = 4.24 mg/kg/day

Females: 1

- and 2

- year LOAEL

– 39 mg/kg/day based on liver

effects in cluding increased blood cholesterol, liver weight, hypertrophy, fatty change, single cell necrosis and macrophages.

NOAEL = 5.13 mg/kg/day

870. 5100 Bacterial Reverse

Mutation Test

MRID 47832047

Annex 5.4.1

Acceptable/Guideline S.typhimurium TA 98, TA 100, TA 1535 & TA 1537

E. coli , WP2 uvr

A

100, 333, 1000, 2500, and

5000 μg per plate +/

- S9 Negative

870. 5300 In vitro Mammalian Cell

Gene Mutation Test

MRID 47832049

Annex 5.4.3/1

Acceptable/Guideline Chinese hamster ovary cells

CHO/HGPRT

0, 173.3, 346.6, 693.3, 1386.5,

and 2773 g/ml +/

- S9 Negative

870. 5375 In Vitro Mammalian

Chromosome Aberration

Test

MRID 47832050

Annex 5.4.2/1

Acceptable/Guideline Rat lymphocytes

4 hr treatment 0, 693.3,
1386.5, and 2773 g/ml +/

- S9; 24 hr treatment 0, 173.3,
346.6, and 693.3 g/ml NegativeSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 63 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

870.5395 Mammalian Erythrocyte

Micronucleus Test

MRID 47832071

Annex 5.4.4 Mouse bone marrow polychromatic erythrocytes

Male and female CD

- 1 mice, 6/sex/dose, single oral gavage

on two consecutive days at 0, 100, 200, and 400 mg/kg/day Negative

870.6200 Acute neurotoxicity in

F344 Rats

MRID 47832134

Annex 5.7.1/1

Acceptable/Guideline 0, 7.5, 75, 750 mg/kg;

Additional motor activity work at 0, 2.5, 7.5, 25 mg/kg LOAEL

– 75 mg/kg/day. On Test Day 1, decreased motor activity in 750 and 75 mg/kg groups. Effect was considered equivocal in 7.5 mg/kg group. Clinical findings on test days 2,3, or 4

– decreased feces, red perioral soiling, perineal urine soiling (females) 750 mg/kg

– muscle tremors and twitches, convulsions, splayed hindlimbs and perineal urine soiling, increased lacrimation and salivation, decreased pupil size and response to touch, decreased level of open

- filed activity and gait abnormalities, decreased r ectal temperature. One female death on day 1

– cause unknown

No treatment

- related histopathological findings.

NOAEL = 25 mg/kg

870.6300 Developmental

neurotoxicity in CD Rats

MRID 47832133

Annex 5.7.5/1

Acceptable/Guideline 0, 25, 100, 400 ppm;

0, 1.8/1.9, 7.1/7.6, 27.7/29.8 mg/kg/day
(gestation/lactation) Maternal:

LOAEL

– > 27.7 mg/kg/day. No

treatment

- related systemic or reproduction

related effects observed in the dams NOAEL = 27.7 mg/kg/day

Offspring:

LOAEL

– 7.1mkd based on reduction in

pos tnatal survival (PND 0

- 4) NOAEL = 1.8 mg/kg/day

Non

Guideline Toxicokinetic

– Rat and

Mouse

Probe looking at ADME: single oral dose

Rat/F344 & Mouse/CD1

MRID 47832038

Annex 5.1.1/1

Acceptable Oral gavage/
100 mg/kg bw Rapid and almost complete absorption

and elimination without detectable

metabolism. Most (rat: 87

- 98%; mouse: 80

- 85%) of

the elimination was via urine.

Plasma t1⁄2 was 9h in male rats and 7h in

female rats.

report also includes integrated plasma

and urine TK data from 28

- and 90

- day rat, and 28

- day mouse dietary toxicity

studiesSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 64 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

870.7485 Toxicokinetic

– F344 Rat

ADME: single oral dose

with examinations at

C

max and 1⁄2C

max

MRID 47832039

Annex 5.1.2/1

Acceptable/Guideline Oral gavage/
5, 100 mg/kg bw Rapid absorption, wide tissue distribution. Radioactivity detected mainly in portal of entry (GI tract, liver) and excretion (kidney, urinary bladder) tissues. No metabolite in tissues or plasma. Kinetics in tissues were parallel to plasma showing no pote ntial for bioaccumulation.

870.7485 Toxicokinetic

– F344 Rat

ADME: oral

– single and repeat dose; IV

– single

dose with examinations at 168 h (7 days )

MRID 47832034

Annex 5.1.3/1

Acceptable/Guideline Oral gavage/ 5 mg/kg bw for 1 and 15 days

100 mg/kg bw for 1 day
Intravenous 5 mg/kg bw for 1 day Rapid and almost complete (at least 92

- 96%) absorption of the oral dose. Low metabolism (<7%) and almost complete (89

- 94%) and rapid elimination of the oral

dose in urine in the 1 st 24 hours.

No tissue a ccumulation, ≤1.2% remained in tissues 7 days after dosing.

Non

Guideline Ex vivo gene expression

and cell proliferation analyses

CD1Mouse (♀) & F344

Rat

MRID 47832033

Annex5.5.4/1

Acceptable Mice: 0, 3000, 4500. Rats: 0, 2000 XDE

- 208

- induced gene expression profile in mice and liver (hepatocellular) proliferation in both mice and rats characteristic of phenobarbital

- like CAR agonism.

Non

Guideline Targeted gene

expression, cell proliferation and cytochrome P450 enzymatic activity in

F344

MRID 47832061

Annex 5.5.4/2

Acceptable 0, 100, 750, 1500 for 3 or 7 days XDE

- 208

- induced liver effects were PB

like. Males were affected more than

females. Neither AhR nor PPARα were

involved.

Non

Guideline Liver Weight Effects in

crl:cd1(icr) CD1 Mice

MRID 47832062

Annex 5.5.4/3

Acceptable Males: 0, 500, 750. Females: 0, 1000, 1500 for 7 days XDE

- 208

- induced liver effects were consistent with CAR activation resulting

in a PB

- like MoA; males were more

sensitive than females. Neither AhR nor

PPARα were involved.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 65 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

Non

Guideline A Study to Characterize

the Induction Profile of

XDE

- 208 in the Livers

Of C57BL/6J Mice

MRID 47832276

Annex 5.5.4/4

Acceptable 0, 750, 1500 ppm XDE

- 208

- induced liver effects in C57Bl/6J mice were similar to previously

observed effects in CD1 mice

Non

Guideline Liver Effects Observed

In Regulatory Toxicology Studies By

Use Of Dual Car

- PXR

Knockout And

Humanised Mice (Mouse/C57Bl/6J WT,

Humanised and KO PXR/CAR)

MRID 47832255

Annex 5.5.4/5

Acceptable 0, 750 ppm for 7 days In WT C57BL/6J XDE

- 208 caused the

same liver effects seen in CD1 mice. In PXR/CAR KO mice, XDE

- 208 did not induce any liver changes, demonstrating

that activation of one or both of these receptors is required to elicit the liver

effects seen in WT mice.

In PXR/ CAR humanised mice slight liver hypertrophic effects occurred but not hepatocellular proliferation. This study demonstrated that XDE

- 208, like PB, acts via a CAR

- mediated MoA and that mice carrying the human PXR and CAR receptors did not develop hepatocell ular proliferation responsible

for liver tumor induction.

Non

Guideline A Dietary Reproductive

Toxicity Cross

- Fostering Study in Crl:CD(SD)

Rats

MRID 47832063

Annex 5.6.12/1

Acceptable Group 1: 0/0 ppm

Group 2: 0/1000 ppm

Group 3: 1000/0 ppm

Group 4: 1000/1000 ppm Groups 1 and 2 (negative control and lactation exposure only) had no effects Groups 3 and 4 (gestation exposure only and positive control) had decreased

neonatal survival

Dam : Reduced feed consumption and body weight gain when dams given 1000 ppm

Litter : All offspring exposed prior to birth died by PND 4. No effect on neonatal survival

with lactation

- only exposure. Therefore, pup survival effect due to in utero

exposure.
Non

Guideline Neonatal Survival in

New Zealand White

Rabbits

MRID 47832138

Annex 5.6.12/2

Acceptable 0, 750 ppm

(0, 29 mg/kg/day) Dam : Decreased feed consumption and body weight gain.

Litter : No EffectsSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 66 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

Non

Guideline Characterization of the

agonist effects of XDE

208 on mammalian

muscle nicotinic acetylcholine receptors

MRID 47832035

Annex 5.6.12/3

Acceptable 0

- 3mM XDE

- 208 was an agonist to the rat fetal

muscle nAChR

XDE

- 208 was not an agonist to human fetal, human adult, or rat adult muscle

nAChR

Non

Guideline Phase 1; Critical window

of exposure for fetal

abnormalities and

neonatal survival effects in Crl:CD(SD) rats

MRID 47832137

Annex 5.6.12/4

Acceptable Group 1: 0 ppm

Group 2: 1000 ppm GD 6

- 16

Group 3: 1000 ppm GD 16

- birth Litter : 1000 ppm from GD 16

- birth caused

neonatal death and limb abnormalities in offspring. Also, all fetal abnormalities observed in developmental toxicity study reversed by PND 4.

1000 ppm from GD 6

- 16 produced no

effects.

Non

Guideline Phase 2; Critical window

of exposure for fetal

abnormalities and

neonatal survival effects in Crl:CD(SD) rats

MRID 47832136

Annex 5.6.12/5

Acceptable Group 1: 0 ppm

Group 2: 1000 ppm GD 16

- 18

Group 3: 1000 ppm GD 18

- 20

Group 4:

1000 ppm GD 20

- 22/LD0 Litter : 1000 ppm exposure for 48 hours starting

on GD 20 caused neonatal death and limb abnormalities in offspring. All fetal abnormalities observed in developmental toxicity study reversed by PND 4.

1000 ppm from GD 16

- 18 or 18

- 20 produ ced no effects.

Non

Guideline Observations on the

effects of XDE

- 208 on the phrenic nerve

hemidiaphragm preparation from new

born rat.

MRID 47832064

Annex 5.6.12/6

Acceptable 0, 0.1, 1.0mM XDE

- 208 induced nAChR

- dependent

sustained contracture of the neonatal rat diaphragm.

Non

Guideline Histopathological

Evaluation Of Fetal Lung Samples From The Developmental Toxicity Study In Crl:Cd(Sd) Rats

MRID 47832135

Annex 5.6.12/7

Acceptable 0, 1000 ppm Fetal lungs from the rat developmental toxicity study were normal.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 67 of 176 Guideline

No. Study Dosages Effects at lowest observed effect level

Non

Guideline

(GF

- 3032 Formulation) Comparative Dermal

Absorption, in vitro rat
and human skin

MRID 47832414

Annex IIIA 7.6.2

Acceptable 0.024, 0.48, 2.4 g/L for 24

hours % AD absorbed for 0.024, 0.48, 2.4 g/L, respectively; Human; 1.15, 1.54, 0.26% Rat: 4.34, 8.72, 1.3%

Non

Guideline

(GF

- 3032 Formulation) Dermal Absorption in the

rat, in vivo

MRID 47832413

Annex IIIA 7.6.3

Acceptable 0.024, 0.48, 2.4 g/L for

10hours then skin washed with mild soap. Tissue collected for up to 192 hours. The amount of applied dose removed by washing was 96%, 71%, and 67% of the applied dose. After 10 hours, 2%, 11%,

and 13% of the AD was absorbed across the dose groups, respectively.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 68 of 176 Study type Route/method

Dose levels* Effects at Lowest Observed Adverse Effects

Level

1. Metabolite X11719474 ADME probe in F344 Rats

MRID 47832132

Annex 5.8.1.1/1 Acceptable/Non

- guideline Oral/gavage 100 mg/kg/day n/a

870.3050

28

- day Oral in F344 Rats

MRID 47832099

Annex 5.8.1.3/2 Acceptable/Guideline Oral/diet 0, 1000, 2000, 3000, 8000 ppm 0/0, 83.4/90.1, 167/184, 244/278, 662/734 mg/kg/day (M/F) LOAEL= 3000 ppm (241mg/kg /day) based on decreased triglycerides in the males

NOAEL= 2000 ppm (167 mg/ kg/day).

870.3100 ; 870.3100

90

- day Oral in F344 Rats

MRID 47832147

Annex 5.8.1.3/3 Acceptable/Guideline Oral/diet 0, 500, 1000, 5000 ppm At 5000 ppm ( 71.8 mg/kg/day) ↑Cholesterol, ↑liver wt. with hypertrophy, eosinophilia, fatty change & SCN

870.3150

90

- day Oral in Beagle Dogs (males)

MRID 47832037

Annex 5.8.1.3/5 Acceptable/Guideline Oral/gavage 0, 10, 25, 50 mg/kg/day No Observed Effects

870. 5100

Ames S. typhimurium E. coli

MRID 47832097

Annex 5.8.1.4/1 Acceptable/Guideline In vitro S9

0

- 5000 μg per plate Negative

870. 5375 Lymphocyte chromosome aberration in CD Rats (males)

MRID 47832096

Annex 5.8.1.4/2 Acceptable/Guideline In vitro S9

0

- 2953 g/ml Negative

870. 5300

HGPRT Chinese hamster

ovary MRID 47832095

Annex 5.8.1.4/3 Acceptable/Guideline In vitro S9

0

- 2953 g/ml Negative

Targeted Gene Expression, Cell Proliferation, and Cytochrome p 450 Enzymatic

Activities in Rats.

MRID 47832151

Annex 5.8.1.5/1 Acceptable/Non

- Guideline 0 or 8000 ppm Increased expression of several CAR

- related transcripts, a long with the pregnane X receptor (PXR)

- related transcript Cyp3a, and PROD enzyme activity, suggests that X11719474 may

be agonist ligand for CAR,

870.3550; OECD 421 Dietary reproduction & Developmental Toxicity Screening Test in CRL:CD(SD) Rats

MRID 47832094

Annex 5.8.1.6/1 Acceptable/Guideline 0, 1000, 2000, 5000 ppm (0, 80.8/81.7, 162/167, 396/451 mg/kg/day (M/F) No Observed EffectSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 69 of 176 Study type Route/method

Dose levels* Effects at Lowest Observed Adverse Effects

Level

870.3700 Dietary Developmental Toxicity Study in CRL:CD(SD) Rats

MRID 47832087

Annex 5.8.1.6/2 Acceptable/Guideline 0, 1000, 2000, 5000 ppm No Observed Effect

2. Metabolite X11721061

870.3050

28

- day Dietary in F344 Rats

MRID 47832036

Annex 5.8.2.3/2 Acceptable/Guideline Oral/diet 0, 1000, 3000, 8000 ppm LOAEL= 8000 ppm (662/734 mg/kg/day (M/F)) based on increased liver weights, hepatic hypertrophy, and increased cholesterol levels

NOAEL = 3000 ppm (244/278 mg/kg /day (M/F)).

870. 5100

Ames S. typhimurium E. coli

MRID 47832085

Annex 5.8.2.4/1 Acceptable/Guideline In vitro S9

0

- 5000 μg per plate Negative

870. 5375 Lymphocyte chromosome aberration in CD Rats (males)

MRID 47832078

Annex 5.8.2.4/2 Acceptable/Guideline In vitro S9

0

- 1920 g/ml Negative

870. 5300

HGPRT Chinese hamster

ovary MRID 47832077

Annex 5.8.2.4/3 Acceptable/Guideline In vitro S9

0

- 1920 g/ml Negative

3. Metabolite X11596066

870. 5100

Ames S. typhimurium E. coli

MRID 47832076

Annex 5.8.3.4/1 Acceptable/Guideline In vitro S9

0

- 5000 μg per plate Negative

4. Metabolite X11579457

870. 5100

Ames S. typhimurium E. coli

MRID 47832070

Annex 5.8.4.4/1 Acceptable/Guideline In vitro S9

0

- 5000 μg per plate Negative

870. 5375 Lymphocyte chromosome aberration in CD Rats (males)

MRID 48288932

Annex 5.4.2/2 Acceptable/Guideline In vitro S9

0

– 2525 g/ml NegativeSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 70 of 176 Study type Route/method

Dose levels* Effects at Lowest Observed Adverse Effects

Level

870. 5300

HGPRT CHO

MRID 48288933

Annex 5.8.4.4/3 Acceptable/Guideline In vitro S9

0

– 2525 g/ml Negative

5. Metabolite X11519540

870.3050

28

- day dietary in F344 Rats

Annex 5.8.5.3/2

MRID 48288941 Acceptable/Non

- guideline 0, 100, 300, 1000, 2000 ppm (0/0, 7.7/8.5, 23.1/24.9, 74/77, 140/152 mg/kg/day (M/F)) LOAEL = 300 ppm (23.1 mg/kg/day) based on changes increased liver weight and histopathology along with degeneration of kidney tubules (in males).

NOAEL= 100 ppm (7.7 mg/kg/day).

870. 5100

Ames S. typhimurium E. coli

MRID 47832068

Annex 5.8.5.4/1 Acceptable/Guideline In vitro S9

0

- 5000 μg per plate Negative

870. 5375 Lymphocyte chromosome

aberration in CD Rats (males)

MRID 48288935

Annex 5. 8.5.4 / 2 Acceptable/Guideline In vitro S9

0

– 2540 g/ml Negative

870. 5300

HGPRT CHO

Annex 5.8 .5.4 / 3

MRID 48288934 Acceptable/Guideline In vitro S9

0

– 2540 g/ml NegativeSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 71 of 176 A.3 Hazard Identification and Endpoint Selection

A.3.1 Acute Reference Dose (aRfD)

- Females age 13

- 49

Study Selected: Developmental Neurotoxicity Study in the Rat

MRID No.: 47832133 Executive Summary : See Appendix A, Guideline § 870.6 3 00 Dose and Endpoint for Risk Assessment: NOAEL = 1.8 mg/kg/day based on decreased

neonatal survival PND 0

- 4 seen at LOAEL = 7.1 mg/kg/day. Uncertainty Factors : 30 (3 X for interspecies (UF A ) and 10 X for intraspecies (UF H ) variability). Comments about Study/Endpoint/Uncertainty Factors: The DNT was sele cted because the effect is a single dose effect, and can be provided the exposure occurs between gestation days 20 to birth. This endpoint is protective of decreased neonatal survival observed in the 2

- generation reproduction study with a NOAEL of 6.63 mg /kg/day and offspring effects observed in the rat developmental study with a NOAEL of 11.5 mg/kg/day. This endpoint is appropriate for the females age 13

- 49 since it is protective of potential in utero and reproductive effects.

A.3.2 Acute Reference Dose (aRfD)

- General Population, excluding females 13+

Study Selected: Acute Neurotoxicity Study in the Rat

MRID No.: 47832134 Executive Summary : See Appendix A, Guideline § 870.6200 Dose and Endpoint for Risk Assessment: NOAEL = 25 mg/kg/day based on decreased motor activity on day 1 seen at LOAEL = 75 mg/kg/day. Uncertainty Factors : 100 (10 X for interspecies (UF A ) and 10 X for intraspecies (UF H ) variability). Comments about Study/Endpoint/Uncertainty Factors: The ACN was selected because of

the f ollowing: the effect was observed on the day of exposure, the effect, neurotoxicity, is consistent with the proposed MOA for sulfoxaflor; and the oral route of exposure is appropriate for risk assessment. This study is appropriate for the general populati on because the effect is not specific to a single sex or lifestage. Decreased food consumption was also observed within the first 4 days of exposure in several studies. However, decreased food consumption was attributed to the palatability of sulfoxaflor and was transient.

A.3.3 Chronic Reference Dose (cRfD ) `

Study Selected: Combine Chronic/Carcinogenicity Study in the Rat

MRID No.: 47832060 Executive Summary : See Appendix A, Guideline § 870. 43 00Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 72 of 176 Dose and Endpoint for Risk Assessment: NOAEL = 5.13 mg/kg/day (females) based on liver effects including increased serum cholesterol, hypertrophy, necrosis of centrilobular hepatocytes, vacuolization, and increased severity of aggregates of macrophages/histiocytes observed in males (21.3 mg/kg/day) and fema les (39 mg/kg/day). Although the NOAEL for males was slightly lower at 4.24 mg/kg/day, HED is using the female NOAEL of 5.13 mg/kg/day as a PoD for risk assessment. HED considers 4.24 mg/kg/day to be artificially low as a result of dose

spread and the NO AEL of 5.13 is supported by similar rat NOAELs in the 2

- generation reproduction study (6.07 mg/kg/day), the 90

- day oral study (6.36 mg/kg/day), and the development study (11.5 mg/kg/day).

Uncertainty Factors : 100 (10 X for interspecies (UF A ) and 10 X for intraspecies (UF H ) variability). Comments about Study/Endpoint/Uncertainty Factors: The combined chronic/carcinogenicity study was selected because the effects were observed of 2 years of exposure and the route of exposure of is appropriate. Hepatot oxicity was also observed in several other oral studies including the 28

- day, 90

- day, and chronic studies in the rat and mouse. However, the selected endpoint occurs at a lower dose and is therefore protective of the observed hepatotoxicity.

A.3.4 Dermal Exposure (Short

- and Intermediate

- Term )

Study Selected: Developmental Neurotoxicity Study in the Rat

MRID No.: 47832133 Executive Summary : See Appendix A, Guideline § 870. 6300 Dose and Endpoint for Risk Assessment: NOAEL = 1.8 mg/kg/day based on decreased

neonatal survival PND 0

- 4 seen at LOAEL = 7.1 mg/kg/day. Uncertainty Factors : 30 (3 X for interspecies (UF A ) and 10 X for intraspecies (UF H ) variability). Comments about Study/Endpoint/Uncertainty Factors: The endpoint used for short

- and

int ermediate

- term dermal exposure was selected DNT study in ra t s. The endpoint of concern is

decreased neonatal survival in PND 0

- 4 offspring with a NOAEL of 1.8 mg/kg/day and a LOAEL of 7.1 mg/kg/day. This study is appropriate for the durations of exposure as well as it is protective for the populations of concern (fetuses in utero who may be exposed when mothers are exposed in occupational settings).

The dermal equivalent dose (DED) using a dermal absorption value of 2.5% is 25 mg/kg/day (1000 mg/kg/day x 0.025 = 25 mg/kg/day). The NOAEL for the route

- specific 28

- day dermal study is 1000 mg/kg/day (no effects in females up to the limit dose). The DED using the 28

- dermal study is greater than the dose where offspring effects were observed in the DNT and is therefore not protective of the developmental effects observed in the DNT. As such, use of the

28

- day dermal study is not appropriate for dermal exposure assessment.

A.3.5 Inhalation Exposure (Short

- and Intermediate

- Term)

Study Selected: Developme ntal Neurotoxicity Study in the Rat

MRID No.: 47832133Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 73 of 176 Executive Summary : See Appendix A, Guideline § 870. 6300 Dose and Endpoint for Risk Assessment: NOAEL = 1.8 mg/kg/day based on decreased

neonatal survival PND 0

- 4 seen at LOAEL = 7.1 mg/kg/day. Uncertainty Factors : 30 (3 X for interspecies (UF A ) and 10 X for intraspecies (UF H ) variability). Comments about Study/Endpoint/Uncertainty Factors: The endpoint used for short

- and

intermediate

- term inhalation exposure was selected DNT study in ra t s. The endpoint of concern

is decreased neonatal survival in PND 0

- 4 offspring with a NOAEL of 1.8 mg/kg/day and a LOAEL of 7.1 mg/kg/day. This study is an appropriate duration of exposure and it is protective for the populations of concern (fetuses in uter o who may be exposed when mothers are exposed in occupational settings). A default inhalation absorption factor of 100% is assumed in the absence of an inhalation study.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 74 of 176 A.4 Executive Summaries

A.4.1 Subchronic Toxicity

870.350 28

- Day Oral Toxicity

IIA 5.3.1/2 28

- Day Oral Toxicity (feeding) Fischer 344 Rats;

In a 28

- day dietary toxicity study (MRID 47832045), groups of five male and five female Fischer 344 rats were given test diets formulated to supply 0, 300, 1000, 2000, or 3000 ppm

sulfoxaflor f

or 28 days. These dose levels corresponded to 0, 24.8, 79.4, 155, or 205 mg/kg/day for males and 0, 26.5, 88.3, 170 or 192 mg/kg/day for females, respectively. Toxicokinetic analysis of blood plasma was completed.

Administration of sulfoxaflor to male and female rats at 3000 ppm resulted in excessive reductions in feed consumption (31% and 36%) and body weight loss (21% and 19%) compared to controls after 9 days of administration, respectively. The 3000 ppm dose groups were euthanized (day 9). The low er feed consumption was attributed to decreased palatability of rodent feed containing sulfoxaflor and was responsible for the decreased body weights. Animals in all other dose groups exhibited a dose

- related decrease in feed consumption at the

start of ex posure, which was due to decreased palatability of treated diets containing sulfoxaflor. Males given 300, 1000 or 2000 ppm consumed 5%, 29% or 54% less feed than control animals after one day. Females given 300, 1000, or 2000 ppm consumed 7%, 26% or 48% l ess than control animals. Feed consumption increased in animals given 1000 or 2000 ppm for the remainder of the study. Feed consumption for males given 300 or 1000 ppm was comparable to controls by the end of the study and was 6% decreased at 2000 ppm. Females given 300, 1000 or 2000 ppm consumed 8%, 6% or 11% less than controls at the end of the study. Animals given 1000 or 2000 ppm had decreased body weight after one day of exposure, and was considered a secondary effect of the reduced feed consumption. By day 28, males and females given 2000 ppm weighed 8.5% and 10% less, respectively, than controls. Males and females given 300 or 1000 ppm had body weights comparable to controls on day 28. Males and females (1000 or 2000 ppm) had dose related increases in serum total cholesterol levels, and both sexes given 1000 or 2000 ppm, also had total serum pr otein levels higher than controls. Albumin and globulin levels were higher than concurrent and historical controls and were considered to be treatment related in males given 1000 or 2000 ppm and females given 2000 ppm.

Males and females given 2000 ppm had treatment

- related decreases in final body weights. Males and females given 1000 or 2000 ppm had increased absolute and relative liver weights

that were dose and treatment related.

There were a number of organ weights of males and females given 2000 p pm that were altered and included: relative brain (males and females), relative kidney (males), relative testes (males), relative thyroid (males and females), absolute heart (males), and absolute spleen (males and females). These differences in organ weig hts were considered to be secondary to the lower body weights of this dose group. This conclusion is supported by the absence of histopathological changes in these organs.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 75 of 176 Treatment

- related histological effects were observed in the livers of males and fem ales given 1000 or 2000 ppm and consisted of a dose related increase in the severity (very slight to moderate) of hepatocellular hypertrophy, with altered tinctorial properties, involving the centrilobular to midzonal regions of the hepatic lobule. Effect s were more prominent in males, compared to females, increasing to moderate severity in 2000 ppm males. Vacuolization, consistent with fatty change, involving hepatocytes primarily in the right lateral lobe was also occasionally seen in a multifocal distr ibution in males given 1000 or 2000 ppm and in one female given 2000 ppm. The restriction of this alteration to only one liver lobe, the minor nature of the effect (very slight or slight) and the lack of a clear dose response relationship in regards to se verity suggests that this may not be a significant effect.

Toxicokinetic analysis of the plasma showed that levels of sulfoxaflor (AUC24h) were effectively proportional to dose; 3.3 to 3.6 fold increase between 300 and 1000 ppm groups and

a 2.0

- fold inc rease between 1000 and 2000 ppm dose groups. Females were more efficient in eliminating the test material than males. The 24 hour systemic dose as measured by the AUC24h was 21, 15 and 14% lower in females than males at 300, 1000 and 2000 ppm dose

groups , respectively (corresponding to 210, 693, 1371 μg h/ml versus 167, 591 and 1183 μg h/ml at the low, middle and high doses, respectively). Plasma elimination half

- life of sulfoxaflor in male rats was between 7 and 8 hours; whereas it was 32

- 43% lower in f emales (between 4

- 5 hours).

The no

- observed

- adverse

- effect

- level (NOAEL) for both sexes is 300 ppm (26.5 mg/kg/day)

and the lowest

- observed

- adverse

- effect

- level (LOAEL) is 1000 ppm (88.3 mg/kg/day), based on enlarged livers (size and weight) with hepato cellular hypertrophy, slight vacuolization (males), and increased cholesterol levels. This study is Fully reliable ( acceptable/n on

Guideline): GLP compliant and acceptable for use as a range

- finding study

IIA 5.3.1/4 28

- Day Oral Toxicity [feeding] CD1( ICR) mice

I n a 28 day toxicity study in mice (MRID 47832042, PMRA 1941272), groups of five male and

five female CD

- 1 mice were fed diets supplying 0, 300, 1500, or 3500 ppm sulfoxaflor (purity, 98.1%; Lot # C2120

- 16, TSN105885), equivalent to 0, 44, 230, 524 mg/kg/day in males and 0, 53, 273, and 638 mg/kg/day in females, for at least 28 days. Parameters evaluated were daily

cage

- side observations, weekly detailed clinical observations, ophthalmic examinations, body weights, feed consumption, hematology, clinical chemistry, selected organ weights, and gross and histopathologic examinations.

Feed consumption values for males and females given 1500 or 3500 ppm were lower than the controls during days 1

- 2, whereas feed consumption values were comparable to the controls after day 4. Slight treatment

- related decreases in body weights and body weight gains were observed in males and females given 1500 or 3500 ppm during the first week of the study, however, they were comparable to controls through the rest of the study.

Toxicokinetic analysis of the plasma showed that dose

- proportional intake of sulfoxaflor was

translated into a dose

- proportional increase in plasma concentrations of sulfoxaflor. The systemic exposure of sulfoxaflor was ~40% higher in males than in female s. The 24

- hour urineSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 76 of 176 urinary elimination of sulfoxaflor in males was between 33 and 44% and in females was between 23 and 65% of what they consumed during a 24

- hour period, indicating that the majority of the dietary administered sulfoxaflor was excreted essentially unchanged in the urine.

At termination on day 30, males and females given 1500 or 3500 ppm had treatment

- related elevations in mean serum ALP, ALT, AST activities and triglycerides. There were treatment

related increases in the mean liver wei ghts and liver histopathology of males and females given 1500 or 3500 ppm compared to controls. Males given 300 ppm had marginal treatment

- related increases in liver weights over the respective control values, but were not associated with detectable hepat ocyte hypertrophy or clinical chemistry changes and therefore, considered a

non

- adverse effect. Males given 3500 ppm had treatment

- related elevations in absolute and relative adrenal gland weights that corresponded to hypertrophy of the zona fasciculata o f the adrenal cortex. Males given ≥ 1500 ppm had treatment

- related lower absolute and relative kidney weights. In summary, due to higher systemic exposure in male mice, higher toxicity of

sulfoxaflor was observed.

The no

- observed

- adverse

- effect

- level (NOAEL) in male and female mice is 300 ppm (44 and 53 and mg/kg/day, respectively). The lowest

- observed

- adverse

- effect

- level (LOAEL) in male and female mice is 1500 ppm (230 and 273 mg/kg/day, respectively), based on increased liver weights,

increased liv er enzymes (ALT, AST), and hepatocellular hypertrophy in both sexes. Liver histopathology (necrosis) in male mice was also seen at this dose. At 3500 ppm (524 and 638 mg/kg/day, respectively), the same effects were seen along with increased triglycerides and ALP in both sexes, liver histopathology in males (mitotic cells, vacuolization/fatty change) and females (necrosis), and adrenal effects (increased weights and hypertrophy) in males.

This study is fully reliable (acceptable/guideline).

IIA 5.3.1/6 28

- day Palatability/Tolerability Probe Study in Beagle dogs ; In a 28

- day oral palatability probe study (MRID 47832043), sulfoxaflor (XDE

- 208 technical grade, 95.6% a.i.; lot no. E2162

- 34 or analytical grade, 99.7% a.i., lot no. 20062

- 464

- 9) was

administe red to Beagle dogs via dietary, capsule or oral gavage administration for up to 28 days. Using the same animals (with 1 or 2

- week breaks of basal diet only between dosing regimens), routes of test material administration and dosing regimens were altered f or a total of five dosing groups, during the course of the study as follows. Three female dogs received technical grade test material via the diet, ad libitum, for 6 consecutive days at a dose level of 500 ppm (Group 1). The dosing route for these animal s was changed to analytical grade test material via oral gavage, once daily for 28 consecutive days, at a dose level of 15 mg/kg/day (Group 3). Another group of three female dogs received the technical test material via capsule, twice daily for 6 consecut ive days, at a dose level of 15 mg/kg/day (Group 2). The dosing route for these animals was changed to dietary, ad libitum (technical grade) for 5 consecutive days, at a dose level of 100 ppm (Group 4). The dosing route for these animals was once again c hanged to oral gavage (technical grade), once daily for 28 consecutive days, at a dose level of 15 mg/kg/day (Group 5). For both oral gavage groups, the vehicle was 0.5% methylcellulose in deionized water and the dose volume was 10 mL/kg. A previous, pre liminary palatability probe study (MRID 47832056) did not identify adequate oral acceptance by diet and capsule administration.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 77 of 176 Observations for morbidity, mortality, injury and the availability of food and water were conducted twice daily for all animals. Clinical observations were conducted daily. Body weights and food consumption were measured and recorded daily. Blood samples for clinical pathology evaluations were collected from all animals pretest and on Days 6 and 29 (prior to necropsy), a nd urine samples were collected at necropsy from all animals on Day 29. Blood samples for determination of the plasma concentrations of the test article were collected from animals in Group 4 at designated time points on Day 5 (the last day of dietary dos ing), and from animals in Groups 3 and 5 at designated time points on each respective Day 28 (prior to necropsy). At study termination, necropsy examinations were performed and organ weights were recorded. A complete set of tissues from Group 5 animals w as sent to the Sponsor for microscopic evaluation. Findings were compared with historical range data due to the lack of

concurrent controls.

All animals survived until the scheduled termination intervals. However, there were clinical findings and body w eight findings that were associated with reduced food consumption (50% or more compared to pretreatment). In some instances, the reduced food consumption was accompanied by a lack of sufficient fluid intake which resulted in a loss of skin elasticity. In addition, intermittent emesis and fecal alterations (discolored, mucoidal, and soft/watery feces) was likely test material

- related.

Oral gavage administration at 15 mg/kg/day was a tolerable route and concentration for the technical grade test material exposure as determined by adequate food consumption and tolerable

in

- life clinical signs. Exposure at 100 ppm via dietary route was well tolerated as determined by adequate food consumption, but the amount of test material consumed was not sufficient to justify it as a potential highest dose for subsequent studies. Exposure at 500 ppm via dietary route or 15 mg/kg/day via capsule route was not well tolerated as determined by insufficient food consumption. Therefore, the dietary route could not be consid ered a viable method of test material exposure over a sustained duration. Based on this information, oral gavage exposure was determined to be the most appropriate route for a sustained duration of test material exposure in Beagle dogs.

This non

- guidelin e study is reliable with restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to GLP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific

in tegrity of the data. It was conducted to determine palatability and the appropriate method of oral administration for subchronic and chronic oral toxicity studies and not to satisfy guideline requirements.

IIA 5.8.1.3/2 X11719474: 28

- D ay Dietary Toxici ty Study in F344/DuCrl Rats;

In a 28

- day dietary toxicity study (MRID 47832099) , five male and five female Fischer 344 rats per group were given test diets formulated to supply 0, 1000, 2000, 3000 or 8000 ppm X11719474 (Purity 99.5%; Lot # E

- 2695

- 1, TSN030626

- 0003) for 29 days. These concentrations corresponded to time

- weighted aver age doses of 0, 83.4, 167, 244 or 662 mg/kg bw/day for males and 0, 90.1, 184, 278 or 734 mg/kg bw/day for females, respectively. Standard parameters were evaluated as well as toxicokinetic analyses of plasma and urine.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 78 of 176 There were no treatment

- related e ffects on clinical signs, ophthalmic observations, hematologic or coagulation parameters.

In the 8000 ppm dose group, males and females exhibited reduced body weight gain compared to controls during the first five days of treatment, and this effect was c onsidered secondary to reduced feed consumption. After five days of treatment, body weight gains for males and females were 27% or 20% lower than controls, respectively. By Day 29, body weights and body weight gains were similar to control values. Decre ased in feed consumption during the first five days of the study was considered to reflect decreased palatability of the treated diet. Males and females consumed ~11% or ~7% less feed than controls, respectively, during this period. However, feed consump tion was comparable to controls by the end of the study. Feed consumption values for males and females given 0, 1000, 2000 or 3000 ppm were either slightly higher or comparable to controls throughout the study.

Also in the 8000 ppm dose group, blood cho lesterol levels (25% in males and 22% in females), and liver weights (33% in males and 14% in females) were higher compared to controls. Triglyceride levels decreased 2

- fold in males given 3000 and 8000 ppm X11719474. Males and females given 8000 ppm and males given 3000 ppm had slight treatment

- related hypertrophy of hepatocytes. All other histopathologic observations were considered to be spontaneous alterations unassociated with exposure to X11719474.

Absorption of the X11719474 from the diet was do se

- proportional for both genders. The ingested X11719474 was rapidly eliminated from plasma upon the removal of the fortified diet (fasting) and dropped below the limit of quantification at the time of sacrifice in most of the

rats. The elimination half

- life of X11719474 from plasma was between 2.7 and 3.6 hours. Male rats eliminated an average of 54 ± 21% of what they consumed during a 24

- hour period (range: 40 to 85%), while elimination by females averaged 81 ± 10% (range: 66 to 89%) of the

test materi al consumed in 24 hours. Overall, these data show that X11719474 was significantly absorbed following dietary administration and quickly eliminated, primarily via urine as parent compound. The ratio of diastereomers was not substantially different in the plasma and urine samples when compared with the matrix samples added with the test material.

The lowest

- adverse

- effect

- level (LOAEL) was 3000 ppm (241mg/kg bw/day) based on decreased triglycerides in the males. The no

- observed

- effect

- level (NOAEL) was 2000 ppm (167 mg/kg bw/day).

This subc hronic toxicity study in the rats is Fully Reliable (acceptabl e/guideline) and satisfies the guideline requirement (OECD 407) for a repeat

- dose oral study in rats.

IIA 5.8.2.3/2 X11721061: A 28

- Day Oral Dietary Toxicity Study i n Fischer 344/DuCrl Rats; This study (MRID 47832036) was conducted to evaluate the potential toxicity of X11721061, a metabolite of the experimental insecticide sulfoxaflor , in rats when administered orally via the

diet for 28 days. Three treatment groups of five male and five female Fischer 344 CDF ®

[F

- 344/DuCRL] rats were administered the test article at dose levels of 1000, 2000, 3000, andSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 79 of 176 8000 ppm. One additional group of five animals/sex served as the co ntrol and received untreated

diet.

Standard parameters Observations for morbidity, mortality, injury, and the availability of food and water were conducted twice daily for all animals. Cageside observations were conducted daily. Detailed clinical observ ations were conducted weekly. Body weights were measured and recorded prior to randomization (Day

– 2 and

- 3) and on Days 1, 5, 8, 15, 22, and 29. Food consumption was measured and recorded pretest (Days

- 4 to

- 1) twice during the first week, and

once weekl y for the remainder of the study. Compound consumption was calculated during each interval food consumption was measured, beginning on Day 1. Ophthalmoscopic examinations were conducted pretest and prior to the terminal necropsy. Blood and urine samples fo r clinical pathology evaluations were collected from all animals prior to the terminal necropsy. Blood samples for plasma analysis were collected from all animals at designated time points on Days 28 and 29. At study termination, necropsy examinations were performed and organ weights were recorded. Tissues were microscopically examined for animals at 0 (control) and 8000 ppm.

No treatment

- related deaths occurred. There were no treatment

- related effects on clinical signs, ophthalmic observations, hematologi c or coagulation parameters. Slight reductions in food consumption noted in males and females at the highest dose (8000 ppm) during the first 5 days of treatment only were considered to reflect transient reduced palatability of the diet. No

treatment

- rela ted clinical signs of toxicity or affects on body weight were observed.

Blood cholesterol levels for males and females given 8000 ppm were higher than the controls, statistically significant, and treatment related. Males and females given 8000 ppm had

sta tistically

- identified higher absolute (33% and 14%, respectively) and relative (37% and 18%, respectively) liver weights than the controls. These differences were interpreted to be treatment

related. There were no other treatment

- related alterations or sta tistical differences in the organ weights for males or females.

Males and females given 8000 ppm and males given 3000 ppm had a slight treatment

- related hypertrophy of hepatocytes in the centrilobular/midzonal region of the hepatic lobules. This

effect w as not considered adverse at 3000 ppm since liver weight and clinical chemistry were not significantly impacted at this dose. All other histopathologic observations were considered to be spontaneous alterations unassociated with exposure to X11721061.

Un der the conditions of this study, the lowest

- observed

- adverse

- effect

- level (LOAEL) for males and females was 8000 ppm (662/734 mg/kg/day (M/F)) based on increased liver weights, hepatic hypertrophy, and increased cholesterol levels. The no

- observed

- adver se

- effect level (NOAEL) for males and females was 3000 ppm (244/278 mg/kg /day (M/F)).

IIA 5. 8.5.3/2 X11519540: 28

- day Dietary Toxicity Study in F344/DuCrl Rats;

In a 28

- day repeated oral dose study (MRID 48288941), groups of five male and five female F344/DuCrl rats were given test diets formulated to supply 0, 100, 300, 1000 or 2000 ppm X11519540 (targeted doses of 0, 10, 30, 100, or 200 mg/kg/day, respectively) for at least 28 days. These dose levels corresponded to time

- we ighted average doses of 0, 7.7, 23.1, 74.0, and 140 mg/kg/day for males and 0, 8.5, 24.9, 77.2 and 152 mg/kg/day for females, respectively.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 80 of 176 Standard parameters were evaluated, as well as toxicokinetic analysis of plasma and urine, and gene expression anal ysis of liver tissue.

Rats given 1000 or 2000 ppm had treatment

- related lower body weights and body weight gains throughout the study, relative to controls. At study termination, the body weight gains of males given 1000 or 2000 ppm were 17.4 and 29.6% l ower than controls, and the body weight gains of females given 1000 or 2000 ppm were 17.0 and 26.4% lower than controls, respectively. Males and females given 1000 or 2000 ppm also had treatment

- related decrements in feed consumption. The lower feed cons umption was interpreted to be caused by reduced palatability of the diets, resulting in lower body weight and body weight gains. There were no treatment

related effects on body weights, body weight gains, or feed consumption in male or female rats given 1 00 or 300 ppm.

Males and females given 1000 or 2000 ppm had treatment

- related lower red blood cell counts, hemoglobin concentrations and hematocrit levels. The decrements in erythrocytic parameters

were dose

- related, and were interpreted to be secondary to lower feed consumption at these dose

levels.

Males given 1000 or 2000 ppm had treatment

- related higher serum alanine aminotransferase (ALT) activity, aspartate aminotransferase (AST) activity, total protein, albumin, and cholesterol concentrations, rel ative to controls. In addition, males given 2000 ppm had a

treatment

- related higher gamma

- glutamyl transpeptidase (GGT) activity. Males given 300 ppm

had treatment

- related higher total protein, albumin and cholesterol concentrations. Females given 1000 or 2000 ppm had treatment

- related higher GGT activity, total protein, albumin, globulin and cholesterol concentrations. Females given 300 ppm had treatment

- related higher total protein, albumin and cholesterol concentrations.

The urine pH of males given 300, 1000 or 2000 ppm, and of females given 1000 or 2000 ppm, was slightly lower than controls. Males given 2000 ppm had slightly higher protein concentration in the urine, relative to controls. The alterations in urine pH and protein were interpreted to be treatment related.

Treatment

- related decreases in final body weights occurred in both males and females given 1000 (6.5 and 6.6%, respectively) and 2000 ppm (11.1% and 6.8%, respectively) when compared to controls. Treatment

- related increases in the mean absolute and relative liver weights were noted in males and females given 100 (33.1 and 30.6%, and 19.9 and 25.7%, respectively), 300 (69.8 and 67.4%, and 53.3 and 59.8%, respectively), 1000 (101 and 116%, and 117.5 and 132.7%, respectively) or 2000 p pm (106.8 and 133.5%, and 143.4 and 161.4%, respectively), relative to controls. The higher liver weights corresponded with hypertrophy of centrilobular and midzonal hepatocytes at all dose levels.

Males and females from all dose levels had treatment

- related higher absolute and relative adrenal gland weights. The higher adrenal weights corresponded with histopathologic findings of very slight or slight hypertrophy of the zona fasciculata in males and females given 300, 1000 or 2000 ppm, and very slight or slight increased vacuolization of cortical cells in males from all dose groups and females given 300, 1000 or 2000 ppm.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 81 of 176 One female given 2000 ppm died, and one male given 1000 ppm and one female given 2000 ppm were euthanized in a moribund condition, following blood collection for toxicokinetic plasma samples on test day 28. All three animals had large axillary hematomas secondary to jugular vein bleeding. Excessive hemorrhage was interpreted to be the cause of moribundity or

death in thes e animals, and may have been reflective of a treatment

- related impairment of coagulation. However, the prothrombin times and activated partial thromboplastin times were unaffected by treatment in males and females at all dose levels.

The liver was the pr imary target organ. Male and female rats from all dose levels had

treatment

- related hepatocellular hypertrophy with altered tinctorial properties (increased cytoplasmic eosinophila) of centrilobular and midzonal hepatocytes. The severity of this

alterati on (very slight to moderate) increased in a dose

- related manner. Other treatment

- related liver effects consisted of: very slight increased numbers of mitotic figures (males at all dose levels, females at 300, 1000 or 2000 ppm), very slight multifocal necr osis of individual hepatocytes (males and females at 300, 1000 and 2000 ppm), and very slight multifocal hepatocellular necrosis with accompanying inflammation (males at 1000 and 2000 ppm and females at 1000 ppm). The other primary treatment

- related effec t was an increase in the incidence of very slight multifocal degeneration of tubules in the kidneys, which occurred in males given 300, 1000 or 2000 ppm, and in females given 2000 ppm.

Males from all dose levels had treatment

- related very slight or slight diffuse follicular cell hypertrophy of the thyroid gland, and females given 1000 or 2000 ppm had very slight diffuse hypertrophy of the thyroid gland. The thyroid hypertrophy may have been caused by a

treatment

- related induction of liver microsomal enzym es responsible for biliary excretion of thyroid hormones, with resultant chronic stimulation of the thyroid by thyroid stimulating hormone through disruption of the hypothalamic

– pituitary

– thyroid axis.

Males and females given 300, 1000 or 2000 ppm had treatment

- related very slight or slight hypertrophy of the zona fasciculata of the adrenal glands. In addition, males from all dose levels, and females given 300, 1000 or 2000 ppm had treatment

- related very slight or slight

increased vacuolization of the adrenal cortex (zona glomerulosa, zona fasciculata and zona reticularis). Males given 1000 or 2000 ppm and females given 2000 ppm had very slight diffuse acinar cell hypertrophy of the submandibular salivary gland. The adrenal and salivary effects

were interpreted to be secondary, stress

- related effects.

Males and females given 1000 or 2000 ppm had treatment

- related very slight or slight erythroid cell hyperplasia of the bone marrow. Males given 2000 ppm had a treatment

- related increase in

the incidenc e of very slight extramedullary hematopoiesis (erythroid cell) of the spleen. The bone marrow and spleen effects were interpreted to be reflective of a regenerative response to the lower red blood cell counts, hemoglobin concentrations and hematocrits in males and females given 1000 or 2000 ppm.

Males and females given 1000 or 2000 ppm had treatment

- related very slight or slight atrophy of the mesenteric adipose tissue. Females given 1000 or 2000 ppm had treatment related very slight or slight decreased size of the uterus. Females given 2000 ppm also had treatment

- relatedSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 82 of 176 slight decreased size of the cervix and vagina. The atrophy of mesenteric adipose tissue and decreased size of the female reproductive tract were interpreted to be secondary to lower f eed consumption and lower body weight gains of animals given 1000 or 2000 ppm.

Toxicokinetic analysis of plasma after 28 days of dietary administration to X11519540 showed that the systemic exposure of X11519540 (as plasma AUC 24h ) deviated from linearity (in both sexes) between the low (100 ppm) and mid

- low (300 ppm) dose groups. The mean plasma

elimination half

- lives of X11519540 were calculated to be 27 and 28 hours in male rats of the 1000 and 2000 ppm dose groups, respectively; plasma elimination half

- lives in female rats, of the 1000 and 2000 ppm dose groups, were 35 and 24 hours, respectively. Plasma elimination

half

- lives could not be reliably calculated for animals from the 100 and 300 ppm dose groups as the X11519540 concentrations in plasma at t erminal sacrifice were in most cases roughly equivalent to plasma levels measured at the time at which the test material formulated feed was

withdrawn from the animals.

The amount of X11519540 eliminated in urine, over a 24 hour collection interval, dev iated from linearity (in both sexes) between the low (100 ppm) and mid

- low (300 ppm) dose groups. In male rats 47%, 31%, 18%, and 13% of the calculated daily TMI was eliminated over 24 hours in urine of low, mid

- low, mid

- high, and high dose groups. A sim ilar pattern was observed in female rats with 51%, 28%, 15%, and 11% of the calculated daily TMI eliminated in urine of low, mid

- low, mid

- high, and high dose groups.

Targeted gene expression analysis of Cyp2b1

, Cyp2b2 , and Cyp3a3 in the liver tissue sho wed significantly elevated transcripts at 100, 300 and 1000 ppm (2000 ppm was not tested for these endpoints). These data suggest that X11519540 may stimulate gene expression consistent with constitutive androstane nuclear receptor (CAR) activation, and w ere similar to those seen in previous studies with sulfoxaflor (the parent compound of X11519540) and X11719474 (another metabolite of sulfoxaflor).

The Lowest

- observed

- adverse

- effect

- level (LOAEL) is 300 ppm (23.1 mg/kg/day) based on changes increased li ver weight and histopathology along with degeneration of kidney tubules (in males). The no

- observed

- adverse

- effect

- level (NOAEL) is 100 ppm (7.7 mg/kg/day).

This subchronic toxicity study in the rats is acceptable and satisfies the guideline requirement (OECD 40 7) for a repeat

- dose oral study in rats.

II 5.8.1.3/2 X11719474: 28

- D ay Dietary Toxicity Study in F344/DuCrl Rats ; In a 28

- day dietary toxicity study (MRID 47832099) , five male and five female Fischer 344 rats per group were given test diets formulated to supply 0, 1000, 2000, 3000 or 8000 ppm X11719474 (Purity 99.5%; Lot # E

- 2695

- 1, TSN030626

- 0003) for 29 days. These concentrations corresponded to time

- weighted aver age doses of 0, 83.4, 167, 244 or 662 mg/kg/day for males and 0, 90.1, 184, 278 or 734 mg/kg/day for females, respectively. Standard parameters were evaluated as well as toxicokinetic analyses of plasma and urine.

There were no treatment

- related effect s on clinical signs, ophthalmic observations, hematologic or coagulation parameters.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 83 of 176 In the 8000 ppm dose group, males and females exhibited reduced body weight gain compared to controls during the first five days of treatment, and this effect was consid ered secondary to reduced feed consumption. After five days of treatment, body weight gains for males and females were 27% or 20% lower than controls, respectively. By Day 29, body weights and body weight gains were similar to control values. Decreased in feed consumption during the first five days of the study was considered to reflect decreased palatability of the treated diet. Males and females consumed ~11% or ~7% less feed than controls, respectively, during this period. However, feed consumption was comparable to controls by the end of the study. Feed consumption values for males and females given 0, 1000, 2000 or 3000 ppm were either slightly higher or comparable to controls throughout the study.

Also in the 8000 ppm dose group, blood choleste rol levels (25% in males and 22% in females), and liver weights (33% in males and 14% in females) were higher compared to controls. Triglyceride levels decreased 2

- fold in males given 3000 and 8000 ppm X11719474. Males and females given 8000 ppm and males given 3000 ppm had slight treatment

- related hypertrophy of hepatocytes. All other histopathologic observations were considered to be spontaneous alterations unassociated with exposure to X11719474.

Absorption of the X11719474 from the diet was dose

- pr oportional for both genders. The ingested X11719474 was rapidly eliminated from plasma upon the removal of the fortified diet (fasting) and dropped below the limit of quantification at the time of sacrifice in most of the

rats. The elimination half

- life of X11719474 from plasma was between 2.7 and 3.6 hours. Male rats eliminated an average of 54 ± 21% of what they consumed during a 24

- hour period (range: 40 to 85%), while elimination by females averaged 81 ± 10% (range: 66 to 89%) of the

test material co nsumed in 24 hours. Overall, these data show that X11719474 was significantly absorbed following dietary administration and quickly eliminated, primarily via urine as parent compound. The ratio of diastereomers was not substantially different in the plas ma and urine samples when compared with the matrix samples added with the test material.

The no

- observed

- adverse effect level (NOAEL) for males and females was 2000 ppm (167/184 mg/kg/day (M/F)) based on increased triglyceride at 3000 ppm. Increased chol esterol levels along liver changes (increased weight and histopathology) were observed at 8000 ppm. .

This subc hronic toxicity study in the rats is Fully Reliable (acceptabl e/guideline) and satisfies the guideline requirement (OECD 407) for a repeat

- dose oral study in rats.

870.3100 90

- Day Oral Toxicity

– Rat

IIA 5.3.2 /1 90

- Day Subchronic Oral Toxicity in rats ; In a 90

- day oral toxicity study (MRID 47832046), ten male and ten female Fischer 344 rats per group were given test diets formulated to supply 0, 100, 750 or 1500 ppm sulfoxaflor (purity 96.6%; Lot # E2198

- 17, TSN106108) corresponding to time

- weighted average concentrations of 0, 6.36, 47.6, or 94.9 mg/k g/day for males and 0, 6.96, 51.6, or 101 mg/kg/day for females, respectively. Parameters evaluated were daily cage

- side observations, weekly detailed clinical observations, ophthalmic examinations, functional observational battery (FOB, pre

- exposure and prior to necropsy, comprising cage

- side, hand

- held, and open field observations, rectalSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 84 of 176 temperature, fore

- and hindlimb grip performance, landing foot splay, and motor activity), body weights, feed consumption, prothrombin time, hematology, urinalysis, cli nical chemistry, selected organ weights and gross and histopathologic examinations, which included a specifically detailed review of the nervous system. The study also included integrated toxicokinetics and an assessment of immunotoxicity.

Males (all tre ated doses) and females (750 ppm and 1500 ppm) exhibited dose

- related lower feed consumption, which was due to decreased palatability of diets containing sulfoxaflor. During the first four days males given 100, 750 or 1500 ppm consumed 5%, 12% or 24% less feed than controls, and females given 750 or 1500 ppm consumed 8% or 21% less feed than controls. However, feed consumption for males at all dose levels was comparable to controls by the end of the study. Female feed consumption in the 750 and 1500 ppm groups was 5% or 8% lower than controls at 90 days, respectively, and statistically identified.

Male and females given 750 or 1500 ppm gained less weight than controls during the first four days of treatment and were dose related. After four days of treatment, body weight gains of males given 750 or 1500 ppm were 24% or 45% lower than controls, resp ectively, and body weight gains of females given 750 or 1500 ppm were 13% or 60% lower than controls, respectively. These animals gained weight for the remainder of the study. By day 90, males and females given 750 or 1500 ppm weighed 8% or 9% and 3% or 8% less than controls, respectively. By day 90, the body weight gain of males and females given 750 or 1500 ppm was 11% or 13% and 9% or 20% less than controls, respectively. All body weight effects were considered secondary to the lower feed consumption due to decreased palatability of the test material in the feed. Males and females given 100 ppm had body weights comparable to the controls at the end of the 90

- day study. Serum cholesterol levels in males (127%) and females (83%) given 1500 ppm were si gnificantly greater than controls and considered adverse. All other clinical pathology values were comparable to control values.

Rats given 750 or 1500 ppm had statistically identified higher absolute and relative liver weights that were dose related, and interpreted to be treatment related. The absolute liver weights of males and females given 750 or 1500 ppm was 6% or 5% and 25% or 17% higher than controls, respectively and the relative liver weights of males and females given 750 or 1500 ppm was 14% or 8% and 41% or 27% higher than controls, respectively. There were a number of additional differences in organ weights of males and females given 750 or 1500 ppm that were statistically identified. These differences in organ weights were secondary to t he lower body weights of these dose groups and did not reflect a primary target organ effect of sulfoxaflor. This conclusion was supported by the absence of histopathological changes in these organs.

Treatment

- related histological liver effects occurre d in male and females given 750 or 1500 ppm

and consisted of a dose

- related increase in the severity (slight to moderate) of hepatocellular hypertrophy (with altered tinctorial properties) involving the centrilobular to midzonal regions of the hepatic lobu le. Individual hepatocyte necrosis was also observed in the centrilobular region with a multifocal distribution to a very slight or slight degree. All effects were seen in both sexes but were more prominent in males compared to females. Vacuolization of hepatocytes, consistent with fatty change, was also observed in all males in the 750 and 1500 ppm groups at very slight, slight or moderate degrees. In addition, in the rats with the greatest degree ofSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 85 of 1 76 hepatocellular hypertrophy, necrosis and vacuolizati on, there was an increase in the incidence of rats with multifocal aggregates of macrophages

- histocytes. The microscopic changes were present in all three lobes of the liver examined in male and female rats; however, they were more readily apparent in th e right lateral lobe. The microscopic changes in the liver were consistent with the increased liver weights and cholesterol levels noted for these rats.

Assessment of immunotoxicity as measured by immune responsiveness in the sheep red blood

cell antib ody

- forming cell assay indicated there was no effect on immune responsiveness in female rats up to and including the high dose level of 1500 ppm. There was no effect on immune responsiveness for male rats in the 100 and 750 ppm groups, while the 1500 ppm group displayed a lower, non

- statistically significant, response (26% lower) when compared to the vehicle control group. The lower AFC response in the high dose male group coincided with considerable general toxicity, including decreased body weights and increases in liver weight (absolute and relative), hepatocellular hypertrophy, necrosis, vacuolization consistent with fatty change, multi

- focal aggregates of macrophages, and elevated serum cholesterol, for which the overall NOEL was 100 ppm. Therefore, the lower AFC response in the high dose males was considered secondary to systemic toxicity and thus does not reflect primary immunotoxic potential for sulfoxaflor.

The potential to recover from the effects induced by sulfoxaflor was demonstrated in male and female rats given 0 or 1500 ppm for 90 days, followed by control feed for 28 days. Nearly complete recovery was seen in body weights (only ~5% lower for both sexes). Feed consumption during the 28

- day recovery period was comparable to controls in bot h sexes. Also, the serum cholesterol levels that were elevated during the 90

- day study in both males and females were normal following the 28

- day recovery period and a complete recovery was seen in the absolute and relative liver weights of males and fema les given 1500 ppm. There was partial recovery of the microscopic hepatic effects. Two male rats in the 1500 ppm group still had recognizable hepatocellular hypertrophy of a very slight degree in the centrilobular and midzonal regions. One of these two rats also had multifo cal, very slight individual hepatocellular necrosis. Multifocal, very slight or slight, hepatocellular vacuolization consistent with fatty change was present in most of the recovery males; however, the degree of involvement was substantially less severe i n the recovery group. There were no microscopic

treatment

- related changes present in the liver in females given 1500 ppm.

Toxicokinetic analysis of the plasma showed that the systemic exposure of sulfoxaflor was dose proportional. An ~8

- fold increase i n AUC 24h was found between 100 and 750 ppm groups and a ~2

- fold increase between 750 and 1500 ppm doses. Females were more efficient at eliminating the test material from their system than males. The 24

- hour systemic dose as measured by the plasma

AUC 24h was 15, 16 and 14 percent lower in females than males at 100, 750 and 1500 ppm nominal dose groups, respectively. Plasma elimination half

- life of sulfoxaflor in male rats was ~9 hours; whereas, it was ~8 hours in females (12% lower). The chromatograms o f the plasma samples taken from sulfoxaflor dosed rats (via diet) contained up to 5 minor peaks in addition to the parent compound. These peaks may represent metabolites of the test material or metabolites of test material impurities. Absolute quantitati on of the minor metabolites could not be made, due to lack of reference standards. Elimination of sulfoxaflor in urine over 24 hours ranged between 51 and 61% of the ingested dose, with the exception of high dose in males which was 37% of the ingestedSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 86 of 176 dos e, 26 days after the initiation of the study. Elimination of test material in 0

- 24 hr urine on days 84 and 85 ranged between 52

- 69% for the lower two dose levels, but was somewhat lower at the high dose for both sexes (32

- 36% of ingested dose). In additi on to parent sulfoxaflor, four urinary metabolites were detected. One peak was a known impurity in this lot of the test material. No definitive quantitation of the other three metabolites was obtained.

In conclusion, during the first few days of the st udy decreased feed consumption and body weight gain was observed at all dose levels. Subsequently feed consumption and body weight gain returned to normal in the 100 ppm group. There were no other observations noted in the parameters evaluated at this do se level. At 750 ppm and 1500 ppm the primary toxicologically significant findings were increased serum cholesterol levels, increased liver weight and histopathological findings in the liver. All of the effects due to ingestion of the test material had

r ecovered during the 28

- day recovery phase, except minor histopathologic liver effects noted only in males given 1500 ppm.

The lowest

- observed

- adverse

- effect level (LOAEL) for systemic toxicity is based on the observation of decreased body weight, elevated cholesterol levels, hepatoxicity (increased weight, hypertrophy, necrosis, and vacuolization) at 750 ppm (47.6 mg/kg/day). The no

- observed

- adverse

effect

- level (NOAEL) is 100 ppm (6.36 mg/kg/day).

There were no indications of neurotoxicity at any dose level, therefore, a LOAEL was not determined. The NOAEL is ≥ 1500 ppm (94.9 mg/kg/day) for neurotoxic effects. There were no indication of immunotoxicity at any dose level, therefore, the LOAEL was not determined. The NOAEL is ≥ 1500 ppm (94.9 mg/kg/day ) for immunotoxic effects.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement for a subchronic oral study (OPPTS 870.3100; OECD 408), subchronic neurotoxicity (OPPTS 870.6200; OECD 424), and immunotox icity (OPPTS 870.7800) study in

rats.

870.3100 90

- Day Oral Toxicity

- Mouse

IIA 5.3.2/2 90

- Day Dietary Toxicity Study in Crl:CD1(ICR)

– Mice; In a 90 day oral toxicity study (MRID 47832057, PMRA 1941277), groups of ten male and ten

female CD

- 1 mice w ere fed diets supplying 0, 100, 750 or 1250 ppm and 0, 100, 1500, 3000 ppm sulfoxaflor (purity 96.6%; Lot # E2198

- 17, TSN106108), respectively for at least 90 days. These concentrations supplied average doses of 0, 12.8, 98.0 or 166 mg/kg/day for males a nd 0, 16.2, 247 or 489 mg/kg/day for females. Parameters evaluated were: daily cage

- side observations, weekly detailed clinical observations, ophthalmic examinations, body weights, feed consumption, hematology, clinical chemistry, selected organ weights, and gross and histopathologic examinations. In addition, toxicokinetic analyses were conducted on urine (day 80) and terminal blood plasma.

There were no treatment

- related effects on clinical signs, ophthalmic parameters, body weights or feed consumption.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 87 of 176 Toxicokinetic analysis of the plasma showed that dose

- proportional intake of sulfoxaflor

translated into a dose

- proportional increase in plasma concentrations of sulfoxaflor only up to the mid dose for both male (750 ppm, 92 mg/kg/day) and female (1500 ppm, 227 mg/kg/day) mice. In males, the systemic exposure, as measured by the plasma concentration of sulfoxaflor, became supra

- linear betw een the mid (92 mg/kg/day) and high (152 mg/kg/ day ) doses (3.9

- fold

increase instead of 1.6

- fold expected from the difference in the test material intake between the mid and high doses). Plasma concentrations of sulfoxaflor in females reached a plateau,

r emaining almost unchanged between the mid (227 mg/kg/day) and the high (467 mg/kg/day)

doses. Total elimination of sulfoxaflor in 24

- hour urine remained dose

- proportional only up to

the mid dose and showed less than dose

- proportional increase at the highe st dose, both in male

and female mice. These data are consistent with a saturation of elimination of sulfoxaflor in male mice at the highest dose and a saturation of absorption of sulfoxaflor from the gastrointestinal tract in female mice at the highest d ose. On the basis of these results, the kinetically

- derived maximum dose (KMD) i.e ., the dose above which kinetics become non

linear was considered to be 92 (750 ppm) and 227 (1500 ppm) mg/kg/day for male and female mice, respectively.

Males given 750 or 1250 ppm and females given 1500 or 3000 ppm had treatment

- related increases in absolute (20% and 74%; 36% and 54%, respectively) and relative liver weights (26% and 85%; 40% and 50%, respectively) compared to controls. Treatment

- related organ weight chan ges consisted of increased absolute and relative adrenal weights in males given 1250 ppm, and lower absolute kidney weights in males given 750 or 1250 ppm. The lower kidney weights were however, considered non

- adverse. Males given 1250 ppm had a treatmen t

- related 200% increase in serum alanine aminotransferase (ALT), 43% increase in aspartate aminotransferase (AST) and 142% increase in alkaline phosphatase (ALP) activities. Serum

total cholesterol and total bilirubin concentrations were decreased in males given 750 and 1250 ppm, and were attributed to treatment. Females given 1500 or 3000 ppm had treatment

- related elevations in serum ALT (125% and 171% increase, respectively) and AST activities (44% and 31% increase, respectively), and decreased ALP acti vity (3000 ppm only) compared to controls. Serum triglycerides were elevated in females given 1500 or 3000 ppm and serum cholesterol concentration was elevated in females given 3000 ppm, and attributed to treatment. There was a minor, treatment

- related r eduction in hematocrit and hemoglobin concentration in females given 1500 or 3000 ppm.

Males given 750 or 1250 ppm had slight or moderate, and females given 1500 or 3000 ppm had very slight, slight or moderate, treatment

- related, centrilobular to midzonal hepatocyte hypertrophy with altered tinctorial properties. Other treatment

- related his tologic liver effects consisted of an overall, very slight or slight increase in the numbers of mitotic figures (hepatocytes in mitosis) in the liver of males given 1250 ppm, and very slight or slight fatty change in hepatocytes of males given 750 or 1250 ppm. Males given 750 or 1250 ppm had

treatment

- related, very slight or slight necrosis of scattered, individual hepatocytes, whereas this change in females given 1500 or 3000 ppm was infrequent or minimal. Males given 750 or 1250 ppm and females given 1 500 or 3000 ppm had treatment

- related, very slight hypertrophy of the zona fasciculata of the adrenal cortex. A very slight, treatment

- related fatty change was also present in the zona fasciculata of the adrenal cortex in some females given 1500 or 3000 p pm.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 88 of 176 Four out of ten females given 3000 ppm had very slight, treatment

- related increase in extramedullary erythrocytic hematopoiesis in the spleen.

The No

- Observed

- Adverse

- Effect Level (NOAEL) for males and females is 100 ppm (13 and 16 mg/kg/day, respect ively). The Lowest

- Observed

- Adverse

- Effect

- Level (LOAEL) for males is 750 ppm (98 mg/kg/day), based on increased liver weights and hypertrophy, liver histopathology (necrosis/fatty change), increased cholesterol and bilirubin, and adrenal hypertrophy. At 1250 mg/kg/day (169 mg/kg/day), the same effects were observed, along with increased liver enzymes (ALT, AST, ALP), mitotic figures (hepatocytes), and increased adrenal weights.

The LOAEL in females is 1500 ppm (247 mg/kg/day), based on increased liver weights, liver hypertrophy, increased liver enzymes (ALT, AST), increased triglycerides, decreased hemoglobin and hematocrit, and adrenal changes (hypertrophy and fatty change/vacuolization). At 3000 mg/kg/day (489 mg/kg/day), the same effects were obser ved, along with decreased ALP, increased cholesterol, and hematopoiesis in the spleen. The toxicokinetics data indicates that there is a saturation of absorption from the intestinal tract at 1500 ppm in females and a maximal excretory level at 750 ppm in males.

This study is fully reliable ( US EPA Acceptable/Guideline). The subchronic toxicity study in the mice satisfies the guideline requirement for a subchronic oral study (OPPTS 870.3100; OECD 408) in mice.

870.3150 90

- Day Oral Toxicity

– Dog

IIA 5.3.3/1 90

- Day Oral Gavage Toxicity Study in Beagle Dogs ; In a 90 day oral dog study (MRID 47832058), sulfoxaflor (XDE

- 208 technical, 95.6% a.i., lot

no. E2162

- 34) was administered by gavage to 4 beagle dogs/sex/dose at dose le vels of 0 (vehicle only), 1, 3 or 10 mg/kg/day. The 10 mg/kg/day dose level was reduced to 6 mg/kg/day on Day 5 of the study due to intolerance (lack of food consumption). Controls received the vehicle, 0.5% methylcellulose (Methocel A4C) in water. The test material or vehicle was given once a day for 90 consecutive days, at a dose volume of 10 mL/kg.

Observations for morbidity, mortality, injury, availability of food and water were conducted twice daily. Clinical observations were recorded weekly. Food consumption was measured daily from Days 1 to 7 and 9 to 35, twice weekly from Week 6 to 12, and once during Week 13. Blood and urine samples were collected from all animals pretest and during Weeks 6 and 13. Blood and urine samples were collected from all animals at intervals d uring Week 13 for plasma

and urine concentrations of the test material

. At termination, necropsies were performed, organ weights recorded, and selected tissues were microscopically examined.

All animals survived the study and there were no treatment

- rela ted clinical signs. Treatment

related decreased mean body weights were observed at the high dose level of 10 mg/kg/day in both sexes, primarily during the first week of exposure. However, mean body weights at this exposure level did not have any signific ant decreases beyond Week 2, after the dose was reduced to 6 mg/kg/day on Day 5. By approximately Week 9, the mean body weights at this exposure level had returned to pre

- exposure values. The transient nature of the decreased mean bodySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 89 of 176 weights was a clear indication that the effect was related to exposure at 10 mg/kg/day and not related to exposure at 6 mg/kg/day.

Sulfoxaflor at 10 mg/kg/day was not tolerated, as determined by significant and unacceptable decreases in food consumption . However, reduction to 6 mg/kg/day after 5 days allowed for food consumption in the affected group comparable to controls for the remainder of the study.

Treatment

- related, decreased food consumption values at the high dose were observed in males

and fe males during the first 2 weeks of the study. By Week 3, the mean food consumption values had stabilized and were similar to control values with males appearing to recover slightly sooner than females. Based on the data, the effect on food consumption was considered to be related to exposure at 10 mg/kg/day, with residual effects prolonging the instability in several animals into Week 3 of the study. There were no other notable food consumption findings related to intake of

the test material for the remain der of the study. There were no other parameters under evaluation

that showed treatment

- related effects.

In Week 13, the steady

- state systemic dose (AUC 24 h ) of sulfoxaflor to dogs after doses of 1, 3, 6 mg/kg/day was 32 ± 6, 84 ± 23 and 147 ± 13 μ g*h/ml in males and 22 ± 3, 71 ± 26 and 119 ± 19

μ g*h/ml in females, respectively. The increase in systemic dose was dose

- proportional in female dogs and was approximately dose proportional in male dogs across all three dose levels. The

dose

- corrected AUC 24 aft er oral gavage to dogs was 3

- to 4

- fold higher than that observed in dogs after 28 days of oral gavage and 3 to 5

- fold higher than that observed in rats after 90 days of dietary exposure. Urinary elimination of sulfoxaflor was 70 ± 6, 76 ± 12, and 59 ± 33 percent of administered dose in male and 69 ± 4, 80 ± 6, and 74 ± 13 percent in female dogs at 1, 3, and 6 mg/kg/day doses, respectively. The mean plasma elimination half

- life of XDE

- 208 in Week 13 of dosing was between 17 and 28 hours.

The LOAEL is 10 mg/kg/day, based on excessively reduced food consumption and body weight loss prior to reduction of the dose on Day 5. The NOAEL is 6 mg/kg/day. This study is classified Fully Reliable (Acceptable/Guideline) and satisfies the guideline requirement for a subchronic oral toxicity (oral gavage)

- Beagle dogs ( OPPTS 870.3150 (nonrodent); OECD 409; EEC Part B.27; JMAFF Subchronic Oral Toxicity Study ).

IIA 5.8.1.3/5 X11719474: A 90

- Day Oral Gavage Toxicity Study In Male Beagle Dogs; This study (MRID 478320 37) was conducted for The Dow Chemical Company on behalf of Dow ArgoSciences, LLC., to act as a bridging study from the parent compound, sulfoxaflor, for which a full guideline compliant 90

- day oral toxicity study in dogs was conducted. This study

was req uired to enable a valid comparison of toxicity to be made between the test material, X11719474, and the parent compound using the minimum number of animals. In this respect, the study was limited to a single sex, males. Three treatment groups of four mal e beagle dogs were administered the test material at respective dose levels of 10, 25, and 50 mg/kg/day. One additional group of four animals served as the control and received the vehicle,

0.5% Methylcellulose (Methocel A4C) in reverse osmosis water. Th e vehicle and test material were administered to all groups via oral gavage, once a day for 90 consecutive days, at a dose volume of 10 mL/kg/dose.

Observations for morbidity, mortality, injury, and the availability of food and water were conducted twice daily for all animals. Clinical observations were conducted and body weightsSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 90 of 176 were measured and recorded weekly. Food consumption was measured and r ecorded weekly beginning with Week

- 1. Physical examinations were conducted by a veterinarian pretest to confirm good

- health status of each animal placed on study. Blood and urine samples for clinical pathology evaluations were collected from all animals pretest and during Weeks 6 and 13. Blood samples for determination of the plasma concentrations of the test article were collected from all animals at 0.5, 2, 4, 24 hours postdose and once during Week 13. At study termination, necropsy

examinations were performed, organ weights were recorded, and tissues were collected for microscopic examination.

No treatment

- related effects were noted during clinical examinations and no early deaths occurred. Body weight, food consumption, and clinical pathology param eters were unaffected by treatment with X11719474. There were no treatment

- related histopathologic effects at any dose concentration. Toxicokinetic analysis of plasma after 13 weeks of oral gavage

administration of sulfoxaflor metabolite X11719474 to mal e dogs showed that the systemic exposure of sulfoxaflor metabolite X11719474 (AUC 24hr ) was proportional across all dose levels. The mean plasma elimination half

- lives of sulfoxaflor metabolite X11719474 were 8.4 ± 2.4 hours, 7.8 ± 1.0 hours, and 7.7 ± 1.1 hours from dogs of the 10, 25, and 50 mg/kg/day dose groups, respectively.

In summary, X11719474 administered orally at doses up to 50 mg/kg/day for 90 days was well tolerated by male beagle dogs. There were no treatment

- related histopathologic effects at any dose concentration. The no

- observed

- adverse

- effect level (NOAEL) is 50 mg/kg/day. A lowest

observed

- adverse

- effect

- level (LOAEL) was not identified.

This study was conducted using the following guidance, but it does not meet the specific criteria

of u sing both sexes of Beagle dogs ( OPPTS 870.3150 (nonrodent); OECD 409; EEC Part B.27; JMAFF Subchronic Oral Toxicity Study; Acceptable/Non

- Guideline ).

870.3200 21/28

- Day Dermal Toxicity

– Rat

IIA 5.3.7/1 28

- Day Dermal Toxicity Study in Rats;

In a repeated

- dose dermal toxicity study (MRID 47832059), sulfoxaflor (XDE

- 208 technical grade, 9 5.6 % a.i., lot no. E2162

- 34, TSN003725

- 0001 ) in 0.5% methylcellulose was applied dermally to 10 Fischer DuCrl 344 rats/sex/dose, exposed at a semi

- occluded, shaved skin test site to 0, 100, 500 or 1000 mg/kg body weight/day for six hours per day for 28 consecutive days. Parameters evaluated were daily cage

- side and weekly detailed clinical observations, dermal observations, ophthalmic examinations, body weight, feed consumption, hematology, clinical chemistry, urinalysis, toxicokinetics of blood plasma, selected organ weights, and gross and histopathologic examinations.

Systemic toxicity: At 1000 mg/kg/day, males showed marginal increases in absolute and relative

l iver weights (6.5% and 4.4%, respectively above controls, p<0.05). A treatment

- related histopathologic change was observed in the livers of 6 of 10 males, consisting of very slight hepatocyte hypertrophy, with altered tinctorial properties (increased cyto plasmic eosinophilia), involving the centrilobular/midzonal regions of the hepatic lobule. These were minor changes, not associated with increases in liver enzymes in the blood and hence, considered adaptive and

non

- adverse. Recent mechanistic studies ha ve demonstrated a phenobarbital

- like mode of action

for sulfoxaflor

- induced liver effects. Mean serum cholesterol was 17% higher compared toSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 91 of 176 controls. This effect was considered treatment related but non

- adverse. Toxicokinetic analysis

showed that the a verage plasma concentration of test material at high dose (1000 mg/kg/day) was greater than dose proportional. Similar plasma concentrations were found prior to and 16 hr after test material removal, indicating some retention of test material at the appli cation site. There

were no treatment

- related systemic effects observed in females at any dose. The NOAEL for systemic toxicity is 1000 mg/kg/day (mild liver effects observed in males indicated some dermal absorption, but were not considered adverse). A LOAEL for systemic toxicity was not determined (>1000 mg/kg/day).

Local dermal toxicity: There were no treatment

- related gross or microscopic dermal effects at the application site. The NOAEL for local dermal toxicity is 1000 mg/kg/day. A LOAEL for

loca l dermal toxicity was not determined (>1000 mg/kg/day).

This dermal toxicity study in the rat is classified as Fully Reliable (acceptable/guideline) and satisfies the guideline requirement for a repeat

- dose dermal toxicity study (OPPTS 870.3200; OECD 410) in rats

.

870.3465 90

- Day Inhalation

– Rat

A.4.2 Prenatal Developmental Toxicity

870.3700a Prenatal Developmental Toxicity Study

– Rat

IIA 5.6.10/2 Prenatal Developmental Study in Rats ; In a prenatal developmental study (MRID 47832140), g roups of 26 time

- mated female Crl:CD(SD) rats were administered diets containing 0, 25, 150, or 1000 ppm sulfoxaflor (Purity 95.6%; Lot # E2162

- 34) on gestation day (GD) 6 through 21 corresponding to time

- weighted average doses of 0, 1.95, 11.5, or 70.2 mg/kg/day, respectively, in order to evaluate the potential maternal and developmental toxicity of this compound. In

- life maternal study parameters included clinical observations, body weight, body weight gain, and feed consumption. On GD 21, all rats were euthaniz ed and each dam and fetus was examined for gross pathologic alterations. In addition, blood was collected from dams and fetuses to determine blood levels of the test material. Liver, kidneys and gravid uterine weights were recorded, along with the

number of corpora lutea, uterine implantations, resorptions and live/dead fetuses. All fetuses were weighed, sexed and examined for external alterations. Approximately one half of the fetuses were examined for visceral and craniofacial alterations, while skele tal examinations were conducted on the remaining fetuses.

Administration of 1000 ppm sulfoxaflor in rodent feed resulted in maternal and developmental toxicity. Maternal toxicity was evidenced by decreases in body weight and body weight gains,

relative t o controls, with concomitant decreased feed consumption, throughout the treatment period, and increased relative liver weights. Developmental toxicity was evidenced by decreases in fetal body weight and gravid uterine weight. In addition, clear increases in several fetal abnormalities (forelimb flexure, bent clavicle, hindlimb rotation, convoluted ureter, and hydroureter) occurred, which have subsequently been shown to reverse by postnatal day four. The terminal plasma concentrations of sulfoxaflor in bo th dam and fetal blood were dose

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 92 of 176 proportional throughout the entire range of dietary exposure concentrations with similar levels

between the maternal and fetal blood compartments. Administration of 150 or 25 ppm

sulfoxaflor in rodent feed produced no trea tment

- related maternal toxicity and no indications of embryo/fetal toxicity or teratogenicity.

The LOAEL is 1000 ppm (70.2 mg/kg bw/day) for developmental toxicity based on increased resorption, increased postimplantation loss, decreased number of viabl e fetuses/litter, decreased fetal weights, convoluted ureter and hydroureter, forelimb flexure and hindlimb rotation, and skeletal alterations including bent clavicles. The NOAEL is 150 ppm (11.5 mg/kg bw/day).

The developmental toxicity study in the rat is classified as Fully Reliable (acceptable/guideline) and satisfies the guideline requirement for a developmental toxicity study (OPPTS 870.3700; OECD 414) in rats.

I IA 5.8.1.6/2 X11719474: Dietary Developmental Toxicity Study in CRL:CD(SD) Rats;

In a di etary developmental toxicity study (MRID 47832087), groups of 26 time

- mated female CD rats were given the s ulfoxaflor metabolite X11719474 (purity 95.5% a.i. wt/wt, Lot # E2695

- 1, TSN030626

- 0003) in feed at concentrations of 0, 1000, 2000, or 5000 ppm on g estation days (GD) 6 through 21, which corresponded to time

- weighted average doses of 0, 74, 152, or 368 mg/kg/day, in order to evaluate the maternal and developmental toxicity potential of this compound.

Treatment

- related effects were limited to a tran sient decrease in body weight gain in the 5000 ppm group at the initiation of treatment, with a concomitant decrease in feed consumption. This finding was deemed of no toxicological significance as it was minor in nature, isolated to the first three days of treatment, and likely due to decreased palatability of the diet. There was no

treatment

- related maternal toxicity at 1000 or 2000 ppm. There was no treatment

- related organ weight or gross pathologic findings at any tested dose level. Toxicokinetic an alyses of dam and fetal plasma X11719474 concentrations revealed dose proportionality across all groups and concentrations that were similar in dams and fetuses from the same groups. There was no evidence of developmental toxicity at any dose level tested in this study.

There was no indication of maternal or developmental toxicity at the highest dose tested and

therefore a no

- observed

- adverse

- effect

- level (LOAEL) was not identified. The no

- observed

adverse

- effect level (NOAEL) for maternal and developm ental toxicity was 5000 ppm (368 mg/kg/day).

The developmental toxicity study in the rat is classified as fully reliable ( acceptable /guideline) and satisfies the guideline requirement for a Prenatal Developmental Toxicity Study (OPPTS 870.3700; OECD 414) in rats.

870.3700b Prenatal Developmental Toxicity Study

– Rabbit

IIA 5.6.11/3 Prenatal Developmental Study

– Rabbit; In a dietary developmental toxicity study (MRID 47832066), groups of 26 time

- mated female

rabbits were administered sulfoxaflor (purity 95.6% a.i. wt/wt, Lot # E2162

- 34, TSN003725

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 93 of 176 001) at dietary concentrations of 0, 30, 150, or 750 ppm, which corresponded to time

- weighted average doses of 0, 1.3, 6.6, or 31.9 mg/kg/day, on gestation days (GD) 7

- 28. In

- life parameters

evaluated fo r all rabbits included: clinical observations, body weight, body weight gain, and feed consumption. Maternal blood was collected for sulfoxaflor analysis from four rabbits/group

over a 24

- hour period starting on the morning of GD 27 , and also at terminat ion on GD 28. Fetal

umbilical cord blood was also taken at termination. All rabbits surviving to GD 28 were euthanized and examined for gross pathologic alterations and changes in liver, kidney, and gravid uterine weight. The number of corpora lutea, ut erine implantations, resorptions, and live/dead fetuses were determined. All fetuses were weighed, sexed, and examined for external and visceral alterations. Also, the heads were examined for craniofacial alterations by serial sectioning in approximately one half of the fetuses in each litter, while skeletal examinations were performed on all fetuses.

Animals in the 750 ppm dose group exhibited treatment

- related maternal toxicity in the form of decreased feces in 7 of 26 animals, decreased mean body weig ht gain (55%) from GD 7

- 13, decreased mean body weight gain (12%) throughout treatment (GD 7

- 28), and decreased mean feed consumption (8

- 21%) from GD 7

- 17. There was no treatment

- related maternal toxicity for animals in the 30 or 150 ppm dose groups. The re was no treatment

- related developmental toxicity in any dose group.

The daily systemic dose of sulfoxaflor on GD 27

- 28 was dose

- proportional as indicated by the

near identical mean dose

- corrected AUC 24 h values of 18, 19, and 19 μg sulfoxaflor /h/kg - 1 f or animals given 30, 150, and 750 ppm, respectively. Levels of sulfoxaflor in maternal and fetal blood were similar. The daily systemic dose in this dietary study was similar to that measured in prior gavage studies with sulfoxaflor.

The maternal lowest

- adverse

- effect

- level observed in this study was 750 ppm (31.9 mg/kg/day) based on decreased body weight and body weight gain. The no

- observed

- adverse

- effect

- level was 150 ppm (6.6 mg/kg/day). There was no treatment

- related developmental toxicity; therefore the developmental NOAEL was > 750 ppm (31.9 mg/kg/day).

This study is fully reliable (Acceptable/Guideline) and fulfills the guideline requirements for a prenatal developmental study in rabbits (OPPTS 870.3700, OECD 414 Test Guideline).

IIA 5.6.12/1 A Dietary Reproductive Toxicity Cross

- Fostering Study in Crl:CD(SD) Rats;

In a non

- guideline cross

- fostering dietary developmental toxicity study (MRID 47832063), groups of female Crl:CD(SD) rats (n=16) were administered s ulfoxaflor (purity 95.6% wt/wt,

Lot # E2162

- 34, TSN003725

- 001) at dietary concentrations of 0 or 1000 ppm. The purpose of this study was to determine whether the previously observed decreased survival of pups born to

s ulfoxaflor

- treated dams resulted from in utero and/or lactation al exposure. As part of this study, effects on general toxicity, toxicokinetic analysis of blood and milk, reproductive function and prenatal/early neonatal growth and survival were assessed.

Groups of female Crl:CD(SD) rats were fed diets supplying 0 (control) or 1000 ppm s ulfoxaflor for two weeks prior to mating through weaning on lactation day (LD) 21. As the control and treated females mated, they were subdivided into Foster dams and Donor dams. Cesarean

-Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 94 of 176 section was performed on gestation day (GD) 21 Donor dams, at which time, one or more batches of two of their offspring/sex were immediately cross

- fostered to a Foster dam(s) that had their own litter removed that day (i.e., on LD 0). After cross

- fostering was complete, each

control and s ulfoxaflo r

- treated Foster dam had mixed litters comprised of two pups/sex that originated from control Donor dams (five litters) and two pups/sex that originated from

s ulfoxaflor

- treated Donor dams (eight litters). This design controlled for litter of origin effec ts, and enabled comparison of the survival of pups exposed to s ulfoxaflor during gestation alone or during lactation alone with unexposed control pups and pups exposed during both gestation and

lactation.

Dams given 1000 ppm s ulfoxaflor had treatment

- re lated effects on body weight, body weight gain, and feed consumption consistent with effects seen at this dose level in the previous reproduction/developmental toxicity screening study (MRID 47832143) . Time weighted average

doses for treated animals were 81.2, 74.5, and 59.5 mg/kg/day in the pre

- mating, gestation, and lactation periods, respectively. These corresponded to maternal s ulfoxaflor blood concentrations

of 23.0

- 29.3 μg/g plasma on GD 21 and 19.6

- 25.0 μg/g plasma on LD 0. The average measured pl asma concentration of s ulfoxaflor of male/female pups on GD 21 and LD 0 from these dams was 24.8/24.8 and 25.3/25.9 μg s ulfoxaflor /g plasma, respectively. Thus, fetal and pup plasma

levels of s ulfoxaflor were very similar to one another, and very similar to dam plasma levels. The measured milk concentrations from the same dams on LD 0 were approximately half the corresponding plasma levels and ranged from 12.3

- 14.0 μg s ulfoxaflor /g milk (mean = 13.3 μg/g)

.

All offspring from dams exposed to 1000 ppm su lfoxaflor prior to birth died by postnatal day (PND) 4, irrespective of whether they were cross

- fostered to control

- or treated

- foster dams (see results Table 1 below). Consistent with reduced viability, some offspring were cold to the touch,

had bluish s kin, autolyzed and cannibalized, and stomach void of milk. Conversely, there was no effect on neonatal survival for pups exposed to s ulfoxaflor only after birth. Furthermore, PND 1 pup body weights were significantly decreased in prenatally exposed offsp ring.

In conclusion, these data demonstrate that the effect of sulfoxaflor on pup survival was due to in

utero , not lactational, exposure.

This cross

- fostering study in the rat is classified as Fully Reliable (acceptable/non

- guideline).

IIA 5.6.12/2 A Study of the Effect of Sulfoxaflor on Neonatal Survival in New Zealand

White Rabbits ; In a non

- guideline study (MRID 47832138), sulfoxaflor (purity 95.6%; Lot # E2162

- 34) , was offered on a continuous basis in the diet (with 0.5% apple flavoring) to a gr oup of 12 litter

experienced, time

- mated female New Zealand White [Hra:(NZW)SPF] rabbits from gestation day (GD) 7 through the initiation of parturition (25

- 26 consecutive days). Actual test material intake in the 750 ppm group was 29 mg/kg/day during GD 7

- 28. A concurrent control group of

12 time

- mated females received the apple

- flavored control diet on a comparable regimen.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 95 of 176 With the exception of one F 0 female in the control and 750 ppm groups euthanized on LD 3 due to total litter loss, all females s urvived to the scheduled necrops ies

. No test substance

- related maternal macroscopic findings were noted.

Lower mean body weight gains (24.2%) and food consumption (7.3%) were noted in the

750 ppm group during the gestation exposure period compared to the control group. Corresponding incidences of decreased defecation were noted for three females in this group. Although mean body weights remained within 2.9% of control group values throughout gestation , the reductions in mean body weight gains and food c onsumption were attributed to test substance exposure

. Mean body weights, body weight gains, and food consumption in the 750 ppm group were similar to the control group during LD 1

- 4.

No test substance

- related effects were observed on the mean number of offspring born, offspring survival, or the general physical condition of the offspring.

When sulfoxaflor was continuously in the diet from GD 7 through the initiation of parturition to pregnant New Zealand White rabbits, t here was no indication of materna l or developmental

treatment

- related toxicity; therefore the lowest

- adverse

- effect

- level (LOAEL) was not

determined. The no

- adverse

- effect

- level (NOAEL) is 750 ppm (29 mg/kg/day).

This study is Fully Reliable (acceptable/non

- guideline)

A.4.3 Reproductive Toxicity

870.3800 Reproduction and Fertility Effects

– Rat

IIA 5.6.1/1 Dietary Reproduction/Developmental Toxicity Screening Test in Crl:CD(SD) Rats; In a reproduction/developmental toxicity screening study (MRID 47832143) sulfoxaflor ( 95.6%

a.i. wt/wt, Lot # E2162

- 34, TSN003725

- 0001) was administered to Crl:CD (SD) rats (12/sex/dose group) at concentrations of 0, 100, 500 or 1000 ppm in the diet

. These dose levels corresponded to time

- weighted average doses for males of 0, 8.26, 40.7 or 79.1 mg/kg /day. The time weighted average doses for females during the various study phases ranged from 0, 8.12

- 8.30, 39.5

- 44.1, and 78.2

- 81.6 mg/kg/day, respectively. Males were fed the test diets for two weeks prior to breeding and continuing throughout breeding until termination. The females were fed the test diets for two weeks prior to breeding, continuing through breeding (up to two weeks), gestation, lactation and weaning; pups were weaned on postnatal day 21. Effects on gonadal function, mating behavior, con ception, development of the conceptus, parturition and postnatal growth and survival were evaluated. In addition, a gross necropsy and histopathological examination of the adults were conducted with an emphasis on organs of the reproductive system. In the offspring, litter size, pup survival, sex, body weight and the presence of gross external morphological alterations were assessed.

Males in the 1000 and 500 ppm dose groups had a treatment

- related decrease in food consumption and body weight (1000 ppm only) during the first week of treatment. Females in the 1000 and 500 ppm dose groups had treatment

- related decreases in body weight gain duringSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 96 of 176 the first week of gestation, and females in the 1000 ppm dose group had slightly lower food consumption during the pre

- breeding and gestation phases. Body weight changes were minimal

and not considered adverse.

Males of the 1000 and 500 ppm dose groups had increased absolute and relative liver weights that were dose and treatment related (liver weights for female s at 1000 ppm were not recorded due to an effect on litter survival; there was no effect at 500 ppm). Treatment

- related histological effects were observed in the livers of males given 1000 and 500 ppm and females given 1000 ppm and

consisted of a dose

- rela ted increase in the severity of hepatocellular hypertrophy, with altered tinctorial properties, involving the centrilobular to midzonal regions of the hepatic lobule. Histological effects were very slight or slight severity in males and very slight in fem ales. In addition, there was treatment

- related multifocal hepatocyte vacuolization (slight severity) in 1000 ppm females. These hepatic effects were minimal and not considered adverse.

There were no reproductive or developmental toxicity effects observed in any group up to PND 0 (birth). Shortly after birth, there was a significant decrease in pup survival in the 1000 ppm dose group such that PND 1 survival was 46.3%, compared to 98.3% i n controls. In addition, PND 1 pup body weights were significantly decreased (22

- 25%) relative to controls. By PND 4, eleven of twelve dams had total litter loss resulting in 7.3% pup survival, compared to 95.4% in controls. Because only one of twelve lit ters remained, this dose group was terminated on PND 6. Pup survival was also affected in the 500 ppm group with 4 of 12 dams losing approximately half of their litters by PND 4, which resulted in a pup survival rate of 81.2% compared to 95.4%

in controls. There were no effects on pup body weight in the 500 ppm group. There were no

treatment

- related effects on any other reproductive parameters at 1000 or 500 ppm, and no reproductive effects whatsoever at 100 ppm.

The concentration of XDE

- 208 in pup and d am plasma was a dose proportionate from the 100, 500, and 1000 ppm groups. Sulfoxaflor plasma levels in PND 4 pups were ≈35% of adult plasma concentrations. Individual (n = 3) concentrations of sulfoxaflor were equivalent in dam plasma and milk from dams of the 1000 ppm dose group.

The systemic LOAEL is 1000 (78.2 mg/kg/day) based on decreased pup body weight. The NOAEL is 500 ppm (39.5 mg/kg/day).

The developmental LOAEL is 500 ppm (39.5 mg/kg/day) based on decreased pup survival. The NOAEL is 100 p pm (12.0 mg/kg/day).

This study is fully reliable (acceptable/guideline) and fully satisfies the guideline requirements for a reproduction/development screening study (OPPTS 870.3550; OECD 421) in rats.

IIA 5.8.1.6/1 X11719474: Dietary Reproduction/Deve lopmental Toxicity Screening Test in CRL:CD(SD) Rats; Groups of 12 male and 12 female Crl:CD(SD) rats were administered the sulfoxaflor metabolite X11719474 via the diet at concentrations supplying 0, 1000, 2000, or 5000 ppm, which corresponded to time

- wei ghted average doses of 0, 80.8, 162, or 396 mg/kg/day for males

, and ranged from 0, 81.7

- 114, 167

- 212, and 451

- 507 mg/kg/day during the female pre

- breeding, gestationSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 97 of 176 and lactation phases (MRID 47832094) . Males were fed the test diets for two weeks prior to breeding and continuing through breeding (up to two weeks) up until necropsy (test day 39). Females were fed the test diets for two weeks prior to breeding, through breeding (up to two weeks), gestation and lactation up until necropsy on post

- partum da y 22

- 24 . Effects on gonadal function, mating behavior, conception, development of the conceptus, parturition, and postnatal growth and survival were evaluated. In addition, a gross necropsy and histopathologic examination of the adults

were conducted wit h an emphasis on organs of the reproductive system. O ffspring were evaluated through postnatal day (PND) 21 for litter size, survival, sex, body weight and the presence of gross external morphological alterations. Plasma concentrations of X11719474 were measured in PND 4 culled pups to demonstrate systemic exposure.

There were no treatment

- related clinical findings or adverse effects on feed consumption at any dose level. Body weight and/or body weight gain in the 2000 and 5000 ppm groups were slightly l ower than controls during isolated gestation and lactation intervals. However, these findings were considered of no toxicological significance based on the lack of a dose

- response relationship, the small magnitude of change, and lack of consistency across intervals.

There was a treatment

- related increase in the liver weights of males and females (15 and 7% increase in relative weight, respectively) in the 5000 ppm group. These findings corresponded with an increased incidence of very slight centrilobular and midzonal hepatocellular hypertrophy in all males and the majority of females (10/12) in the 5000 ppm group. There were no treatment

- related organ weight or pathologic effects in the 1000 and 2000 ppm groups of either sex. Toxicokinetic data

demonstr ated dose

- proportional systemic exposure in PND 4 pups.

There were no treatment

- related effects on reproductive endpoints or development of the offspring at any dose level.

There was no indication of maternal toxicity at the highest dose tested of 5000 ppm (396 mg/kg/day)

and therefore a lowest

- observed

- adverse

- effect

- level (LOAEL) was not identified. The no

observed

- adverse

- effect

- level (NOAEL) for maternal toxicity was 500 0 ppm (396 mg/kg/day). There was no indication of reproductive or developmental toxicity at the highest dose tested of 5000 ppm (396 mg/kg/day) and therefore a LOAEL was not identified. The reproductive and developmental NOAEL was 5000 ppm (396 mg/kg/day ).

This study is fully reliable (acceptable/guideline) and satisfies the guideline requirements for a dietary screening reproduction/developmental study in rats (OPPTS 870.3550; OECD 421).

IIA 5.6.1/2 Reproduction and Fertility Effects

– Rats ; In a two

- generation dietary reproduction toxicity study (MRID 47832142) sulfoxaflor (95.6% a.i. wt/wt, Lot # E2162

- 34, TSN003725

- 0001) was administered to Crl:CD (SD) rats (27/sex/dose group) at concentrations of 0, 25, 100 or 400 ppm in the diet for approximately ten weeks prior to breeding, and continuing through breeding, gestation and lactation for two generations. In

- life parameters included clinical observations, feed consumption, body weights, estrous cyclicity, reproductive performance, pup survival, pup b ody weights, puberty onset and anogenital distance. In addition, post

- mortem evaluations included gross pathology and organ weights in weanlings, toxicokinetic analyses, gross pathology, organ weights, oocyte quantitation and sperm count, motility and morp hology, and histopathology, in adults.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 98 of 176 Parental toxicity at 400 ppm consisted of increased absolute and relative liver weights in the P1 (12.8 and 10.9%, respectively) and P2 (6.5 and 7.8%, respectively) males. This effect on liver weight correlated with histopathological findings of very slight to slight centrilobular hepatocyte hypertrophy, often with a very slight increase in individual cell necrosis of centrilobular hepatocytes. No other systemic effects were noted at 400 ppm, and there were no treatm ent

related effects on P1 or P2 parameters in male or female rats at 25 or 100 ppm.

Offspring effects were limited to 400 ppm and comprised slightly decreased neonatal survival in both generations; this in turn led to a lower percentage of live pups up to culling on PND 4. In addition, there was an apparent treatment

- related delay in preputial separation (PPS) for 400 ppm F1 males. This external marker of male puberty onset is androgen dependent, but the underlying

reason for how sulfoxaflor induced this finding is not known; however, there were no other indications of androgenic or anti

- androgenic effects. This included no treatment

- related effects on anogenital distance, no effects on testis or accessory sex gland (i.e., prostate, seminal vesicle,

and e pididymis) weight or histopathology, no evidence of malformations like hypospadias or ectopic testes, no effects on mating, fertility, time to mating, or gestation length, and no

treatment

- related effects on preputial separation at the same dose level in a developmental neurotoxicity study with sulfoxaflor. Taken together, the weight of evidence across androgen

sensitive endpoints led to the conclusion that the data do not support any other sulfoxaflor

- mediated anti

- androgenic effects. There were no effe cts on puberty onset or any other parameter of reproductive performance or offspring growth and survival at 25 or 100 ppm.

Toxicokinetic data from LD 4 dams and culled PND 4 pups in the second generation show dose

proportional systemic exposure to sulfox aflor in dams and their offspring. Plasma concentrations of sulfoxaflor in rat pups were, on average, 32% of the levels measured in the dams.

The lowest

- observed

- adverse

- effect

- level (LOAEL) for systemic toxicity was 400 ppm (24.6 mg/kg/day) based on hepa tic toxicity (increased weight, hypertrophy, and necrosis) in the P1 and

P2 males. The no

- observed

- adverse

- effect

- level (NOAEL) was 100 ppm (6.07 mg/kg/day).

The reproductive LOAEL is established at 400 ppm (24.6 mg/kg/day) based on decreased

neonatal su rvival (ca. 2

- 5%)

.

The offspring toxicity LOAEL is 400 ppm (24.6 mg/kg/day) based on decreased neonatal

survival and delay in preputial separation in F1 females . The NOAEL is established at 100 ppm (6.07 mg/kg/day).

This study is classified as fully reli able (acceptable/guideline) and satisfies the guideline requirement (OPPTS 8700.3800; OECD 416) for a reproduction and fertility study.

A.4.4 Chronic Toxicity

870.4100a (870.4300) Chronic Toxicity

– RatSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 99 of 176 IIA 5.5.1/1 Combined Chronic Toxicity/Oncogenicity Study in F344/DuCrl Rats test by the oral route in the rat;

In a combined chronic carcinogenicity study in rats (MRID 47832060), g roups of 60 mal e and 60

female F344/DuCrl rats were treated orally by dietary administration with sulfox aflor (96.5%,

Lot # E2162

- 34, TSN003725

- 0001) at dose levels of 0, 25, 100, 500 (males only) or 750 (females only) ppm for up to two years. The time

- weighted average dosages ingested, based upon mean feed consumption and body weight data for 24

- months wer e 0, 1.04, 4.24 or 21.3 mg/kg/day for males and 0, 1.28, 5.13 or 39.0 mg/kg/day for females. Ten rats/sex/dose level were necropsied after one year (chronic toxicity group) and the remaining 50 rats/sex/dose level were fed the respective diets for up to t wo years.

Animals were evaluated by daily cage

- side examinations, bi

- weekly clinical observations, periodic detailed clinical observations, body weights, feed consumption and toxicokinetics (plasma and urine). Clinical pathology examinations (hematology , clinical chemistry, and urinalysis) were conducted at regular intervals throughout the study. Ophthalmic examinations were conducted on all rats prestudy, and all surviving rats prior to the scheduled necropsies. All rats had a complete necropsy, with weights of multiple organs collected from all rats at the scheduled necropsies. Histopathological examination of an extensive set of organs was performed on all control and high

- dose level rats and all rats that died spontaneously or were

euthanized due t o their moribund condition. Histopathological examination of rats from the low

and intermediate

- dose levels of the 12

- month chronic toxicity group was limited to the liver, adrenal glands, testes and relevant gross lesions. Histopathological examination of survivors

from the low

- and intermediate

- dose levels of the 24

- month oncogenicity group was limited to the liver, testes, epididymides, coagulating glands, prostate, seminal vesicles and relevant gross

lesions.

No treatment related mortality or clinica l signs were observed after sulfoxaflor exposure. Males given 500 ppm had decreases in body weights at most measurement intervals starting on Day 512 and continuing through Day 729. On Day 729, the mean body weight and body weight gain for males given 50 0 ppm were 5.0% and 5.7% lower than controls, respectively. The decreases were statistically significant and lower than historical control for most time points; therefore, they were considered treatment related. Females given 750 ppm had treatment

- related

, statistically significant decreases in body weights, first noted on Day 57, and continuing through Day 729. On Day 729, the mean body weight and body weight gain for females given 750 ppm were 6.3% and 9.1% lower than controls, respectively. The decreas es were outside of the historical control range. The body weights of males and females given 100 or 25 ppm were unaffected by treatment with sulfoxaflor

.

High

- dose level males and females had treatment

- related , statistically significant increases in

chol esterol concentrations at 3, 6 and 12 months, and 3, 6, 12 and 18 months, respectively, with increases ranging from 17.5 to 32.9%

.

The liver was the primary target organ for histopathologic effects for males given 500 ppm and females given 750 ppm at 12 and 24 months. The absolute and relative liver weights for high

dose males (500 ppm) and females (750 ppm) were only increased at 12 months, in the range of

3.2 to 17%. Non

- neoplastic liver effects at 12 and 24 months consisted of hypertrophy ofSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 100 of 176 centrilobular and midzonal hepatocytes, necrosis of individual centrilobular hepatocytes, vacuolization ( females only ) consistent with fatty change of hepatocytes, and an increase in the severity of aggregates of macrophages/histiocytes in both sexes at 500 and 750 ppm . An

additional treatment

- related liver effect in females given 750 ppm at 24 months consisted of a lower number of basophilic foci of altered hepatocytes. A treatment

- related neoplastic liver

effect at 24 months consisted of a statistically significant increase in the incidence of benign hepatocellular adenomas in males given 500 ppm. Females given 750 ppm did not have a

treatm ent

- related increase in the incidence of liver tumors. There were no treatment

- related liver effects in males or females given 25 or 100 ppm. A phenobarbital (PB) like mode of action ( MOA ) has been postulated for sulfox a flor induced rodent liver tumors and studies were conducted (MRID 47832062) with sulfoxa flor investigat ing the proposed MOA , including a study with dual CAR/PXR knockout mice . A formal framework analysis investigating the proposed MOA is available (MRID 48288939) and the data adequately supports a threshold based, mitogenic MOA for rodent liver tumors

.

At 24 months, males given 100 or 500 ppm had statistically significant increases in absolute and relative testes weights, and statistically significant decreases in absolute and relative epididymal weights. Absolute testes weights of males given 100 or 500 ppm were approximately 46% and 62% higher than controls, respectively. The higher testes weights were reflective of larger interstitial (Leydig) cell adenomas in the testes of males at these dose levels.

The incidence of interstitial cell adenomas in at least one testis of males from all dose groups was comparable to controls at 24 months. However, males given 500 ppm had a statistically significant increase in the incidence of bila teral interstitial cell adenomas of the testes, and a corresponding decrease in the incidence of unilateral interstitial cell tumors, relative to controls.

An additional testicular effect consisted of a statistically significant, increase in the incidence of severe bilateral atrophy of seminiferous tubules in males given 100 or 500 ppm. The seminiferous tubular atrophy was reflective of the generally larger interstitial cell adenomas in these dose groups.

At 24 months, males given 100 or 500 ppm had sta tistically significant decreases in absolute and relative epididymal weights. The lower epididymal weights were associated with a higher incidence of decreased spermatic elements (bilateral, severe) in the lumen of the epididymides of

these males.

Males given 500 ppm had statistically significant increases in the incidence of decreased secretory material in the coagulating glands (severe), prostate (moderate), and seminal vesicles (severe) at 24 months. Males given 500 ppm also had an increased incidence of carcinoma of the preputial gland. T he primary effect was interpreted to be the interstitial cell adenomas, and the effects on seminiferous tubules, epididymides, accessory sex glands were considered secondary

to loss of normal testicular function due to the size of the interstitial cell adenomas, and possible

alterations in the endocrine balance of these rats. Although the mode of action (MOA) for the

effects on reproductive organ was not investigated as part of this study, MOA data for the interstitial cells were submitted separately (MRID 48288942). A similar (shared) MOA with interstitial cell adenomas was postulated for the increased incidence of preputial glandSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 101 of 176 carcinomas; however, the MOA remains unclear since the avail able data are not fully supportive of the proposed MOA.

Toxicokinetic analyses of plasma samples at 3 and 12 months indicated dose proportionality in systemic dose levels that were comparable between the two time points. There were no gender

differences in plasma concentrations of sulfoxaflor across the dose levels and times analyzed. Urinary elimination of sulfoxaflor was also dose proportional for both male and female rats at 3, 6 and 12 months, representing 58

- 127% of the average test material consum ed in a 24

- hour period.

The no observed adverse effect level (NOAEL) is 100 ppm (4.24 mg/kg/day) and the lowest observed adverse effect level (LOAEL) is 500 ppm ( 21.3 mg/kg/day), based on increased serum cholesterol concentrations, and histopathological l iver effects in males.

Note: Although male reproductive effects were seen at doses lower than the determined NOAEL, the effects were considered secondary to the increased incidence of interstitial cell (Leydig Cell)

tumors. The CARC determined that the increased incidence of interstitial cell tumors were not treatment related; therefore, the male reproductive effects (seen in the presence of the tumors)

are also considered to be not treatment

- related. In contrast, PMRA determined that the LOAEL

is 100 ppm, based on the male reproductive effects.

This study is fully reliable (US EPA Acceptable/Guideline): GLP compliant and fully compliant with the combined chronic/carcinogenicity study in rats (OPPTS 870.4300); OECD 453:

870.4100b Chronic Toxicity

– D og

IIA 5.3.4 Oral 1

- year Chronic Toxicity (Dog); In a chronic oral toxicity study (MRID 48288906) , four Beagle dogs/sex/dose were administered sulfoxaflor (XDE

- 208 technical, 99.7% a.i., lot no. E2162

- 34) at doses of 0, 1, 3 or 6 mg/kg/day

for 52 weeks. Test material was administered by a daily gavage dose in 0.5% aqueous methylcellulose vehicle (Methocel A4C; dosing volume 10 mL/kg). Dietary or capsule administration was not performed due to unsatisfactory food consumption using these methods.

Observ ations for morbidity, mortality, injury and the availability of food and water were conducted twice daily for all animals. Toxicity was assessed by weekly detailed clinical observations, food consumption and body weight measurements, ophthalmoscopic exami nations, and clinical pathology evaluations. Blood and urine samples for determination of plasma concentrations of the test material were collected from all animals at Weeks 13, 26 and 52. Toxicokinetic (TK) parameters were determined for the test articl e from concentration

- time data. At study termination, necropsy examinations were performed, select organ weights were recorded, and tissues were preserved for subsequent microscopic examination.

No treatment

- related deaths occurred. The increased frequency of soft and/or watery feces in two males at 6 mg/kg/day was considered treatment

- related due to the high incidence in these animals relative to other groups, but was not considered adverse beca use these findings had no effect on food consumption or body weight/weight gain. A transient, treatment

- related decrease in food consumption and body weight was observed in females at 6 mg/kg/day during the first two weeksSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 102 of 176 of dosing. The decreases in foo d consumption and body weight were considered non

- adverse findings due to their transient nature. No definitive effects on hematological or clinical pathology parameters were noted. The systemic exposure of sulfoxaflor (AUC 24h ) in plasma was proportional across all dose levels in both sexes. With the exception of females at 52 weeks of exposure (urine values of high dose females fell below the regression line fitted to the mean values), toxicokinetic analysis of parent in urine showed that the systemic e xposure of sulfoxaflor (AUC 24h ) was proportional across all dose levels and time points. There were no treatment

- related organ weight changes or gross/histopathologic effects at any dose.

The NOAEL is 6 mg/kg/day (soft/watery feces in males and a trans ient decrease in food consumption and body weight in females during the initial two weeks were considered treatment

related, but not adverse). A LOAEL was not identified (> 6 mg/kg/day).

This study is classified fully reliable (US EPA acceptable/non

- gu ideline) and satisfies the guideline requirement for a chronic oral toxicity study in the dog ( OPPTS 870.4100; OECD 452)

.

A.4.5 Carcinogenicity

870.4200a Carcinogenicity Study

– R at (See 870.4100a (p. 106))

870.4200b Carcinogenicity (feeding)

– Mous e

IIA 5.5.3/1 Carcinogenicity

– Mice [feeding] ; In a carcinogenicity study in mice (MRID 47832054, PMRA 1941285), groups of 50 male Crl:CD1(ICR) mice were treated orally, by dietary administration, with sulfoxaflor (purity 95.6%, Lot # E2162

- 34, TSN003725

- 0001), at dose levels of 0, 25, 100, or 750 ppm for 18 months ; groups of 50 female Crl:CD1(ICR) mice were similarly treated at dose levels of 0, 25, 250, or 1250 ppm. These concentrations corresponded to time

- weighted average doses of 0, 2.54, 10.4 or 79.6 mg/kg/day for males, and 0, 3.43, 33.9, or 176 mg/kg/day for females, respectively.

Animals were evaluated by daily cage side, bi

- weekly observations, periodic handheld detailed clinical examinations, body weights, and feed consumption. Opht halmic examinations were conducted pre

- exposure and prior to necropsy. All mice had a complete necropsy examination with total white blood cell (WBC) and differential WBC counts, and organ weight recording at the scheduled necropsy. An extensive set of o rgans was examined histopathologically from all control and high

- dose group mice, as well as those that died or were euthanized in moribund condition. The kidneys, liver, lungs, adrenal glands (females only) and all relevant gross lesions

from the low

- an d intermediate

- dose groups from the terminal necropsy were also examined histopathologically.

There were no treatment

- related changes in clinical observations, body weights and body weight gains, feed consumption, ophthalmologic observations, or total a nd differential WBC counts in any of the sulfoxaflor treated groups.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 103 of 176 Toxicokinetic analysis indicated that the daily systemic dose (steady

- state plasma concentration) of sulfoxaflor in male and female mice remained essentially proportional to dose during the course of the study (3 and 12 months after study start) and concentrations of sulfoxaflor in urine also increased proportionally with dose for both male and female mice, and at both collection times (3 and 12 months).

The liver was the primary target of sulfoxaflor. The absolute and relative liver weights of males given 750 ppm were increased 87 and 79% respectively; in females given 1250 ppm they were increased 51 and 47%, respectively as compared to controls. At necropsy, there was a treatment

rela ted increase in the incidence of mass nodules and multifocal pale foci in the liver of males given 750 ppm. Females given 1250 ppm had a treatment

- related increased incidence of mass nodules in the liver, albeit at lower numbers compared to the high

- dose males (750ppm). Histopathologic treatment

- related changes consisted of hepatocellular adenomas and/or carcinomas in 60% of male mice given 750 ppm and in 10% of female mice given 1250 ppm. Although there were no statistically significant differences in t he overall mortality rates between the controls and any of the sulfoxaflor treated groups, hepatocellular carcinomas or adenomas were attributed as the cause of death or moribundity in a small proportion of males (6 of 50) given 750 ppm. Treatment

- related non

- neoplastic liver effects consisted of increases in the incidences of eosinophilic and vacuolated foci of cellular alteration in males given 750 ppm; slight to moderate centrilobular/midzonal or panlobular hepatocellular hypertrophy with altered

tincto rial properties (increased cytoplasmic eosinophilia) consistent with liver enzyme induction in males and females given 750 or 1250 ppm; very slight or slight multifocal individual cell necrosis of hepatocytes in males given 750 ppm and females given 1250 p pm (very slight); very slight fatty change in centrilobular/midzonal hepatocytes in males and females given 750 or 1250 ppm and increased incidence of hepatocytes in mitosis in males given 750 ppm.

The only treatment

- related change in females given 250 p pm was an increased incidence of slight centrilobular/midzonal hepatocyte hypertrophy with altered tinctorial properties (increased cytoplasmic eosinophilia) consistent with liver enzyme induction. This was considered to be an adaptive response and non

- ad verse per se due to a complete lack of any associated changes including increase in liver weight or any other treatment

- related histopathologic findings.

Males given 750 ppm had an exacerbation in the cumulative incidence of spontaneous dermatitis

which is common in CD

- 1 mice. Histologically, these were characterized by subacute to chronic inflammation, variable epidermal ulceration and acanthosis. Associated with the ulcerative dermatitis was an increased incidence of reactive plasmacytosis of the loca l submandibular lymph nodes of males given 750 ppm. The exacerbation of spontaneous dermatitis in males given 750 ppm was interpreted to be secondary to the general unthriftiness of these mice due to the excessive stress induced by the malignant and/or be nign hepatocellular neoplasms. In females, a non

- statistically significant increase in lymphosarcoma was seen at the HDT of 1250

ppm.

A phenobarbital (PB) like mode of action (MoA) has been postulated for sulfoxaflor induced

rodent liver tumors and sev eral studies were conducted (MRID 47832062) with sulfoxaflor that investigate the proposed MoA, including a study with dual CAR/PXR knockout mice. Additionally, a formal framework analysis has been submitted (MRID 48288939) . TheSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 104 of 176 available toxicity data and the framework analysis will be used to evaluate the carcinogenic potential of sulfox a flor.

The no

- observed

- adverse

- effect levels (NOAELs) for males and females are 100 (10.4 mg/kg/day) and 250 ppm (33.9 mg/kg/day), respectively. The lowest

- observed

- adverse

- effect level (LOAEL) for males is 750 ppm (79.6 mg/kg/day) and for females is 1250 ppm (176 mg/kg/day), based on increased liver weights, increased incidence of liver nodules, liver hypertrophy, and liver histopathology (necrosis, fatty change). In males, liver foci (eosinophilic and vacuolated) and gross (dermatitis) and histopathological changes (chronic inflammation, epidermal ulceration, and acanthosis) in the skin were also seen. This was accompanied by reactive plasmacytosis of the local sub mandibular lymph nodes.

This study is fully reliable (US EPA Acceptable/Guideline): GLP compliant and fully compliant with the carcinogenicity study in mice (OPPTS870.4200); OECD 451 (US EPA Acceptable/Guideline).

IIA 5.5.4/1 Ex vivo gene expression a nd cell proliferation analyses The purpose of this study (MRID 47832033) was to obtain preliminary information on the potential mode of action responsible for the liver weight increases observed in mice and rats administered Sulfoxaflor [N

- (Methyloxido(1

- (6

- (trifluoromethyl)

- 3 pyridinyl )ethyl)sulfanylidene)

- cyanamide] via the diet. To accomplish this, specific gene expression was assessed by real

- time PCR in liver samples from female CD1 mice exposed to 0 and 4500 ppm sulfoxaflor in diet for four days in a previously conducted palatabil ity probe study (MRID 47832048). In total, eight genes were selected for this study. Five genes, primarily Cyp2b10 but with four additional genes, were chosen to address whether sulfoxaflor induces a phenobarbital (PB)

- like gene expression response. Two genes which are also induced by PB were selected specifically to investigate the effect on blood cholesterol seen in sulfoxaflor

treated rodents. One gene, Cyp4a10 , was included as a marker of peroxisome proliferator

activated receptor alpha (PPAR alpha) to examine this potential mode of action. In addition, cell proliferation (seen as an early response to PB treatment) was assessed by Ki

- 67 immunohistochemical staining in liver tissue from male and female F344 rats from a 28

- day rat study (MRID 47832045; 0, and 2000 ppm dose) and CD1 mice from the mouse palatability study (0, 3000, and 4500 ppm doses). Sulfoxaflor

- treated mice showed gene expression similar to that reported in the literature following PB exposure. The primary PB

- like marker gene, Cyp2b 10

, was induced 148.5

- fold. In addition, three of the four remaining PB

- like response genes ( Cyp3a11

, Alas1 , and NADPH

- Cyp

- reductase ) were induced. The remaining PB

- like gene,

Slco1b2 , was not induced in sulfoxaflor

- treated mice. Sulfoxaflor stimulated the cholesterol synthesis

- related genes, Dhcr7 and Sqle1 , at levels similar to those reported in the literature following PB exposure. The lack of induction for both Cyp4a10 and an internal homeostasis control gene, Scd1 , suggests that sulfoxaflor is not acting as a peroxisome proliferator. Mice exposed to 3000 ppm sulfoxaflor for seven days showed increased proliferation in both the centrilobular and midzonal regions, whereas both sexes of rat exposed to 2000 ppm sulfox aflor for 28 days showed increased proliferation only in the centrilobular region.

In conclusion, treatment with sulfoxaflor resulted in a gene expression profile similar to that reported for phenobarbital. Additionally, increased hepatocellular prolifer ation in both the centrilobulor and midzonal regions was seen in female mice following 3000 ppm treatment,Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 105 of 176 whereas both sexes of rats exposed to 2000 ppm sulfoxaflor showed increased proliferation only in the centrilobulor region. The carcinogenic potenti al of sulfoxaflor, along with the proposed MOA for liver tumors will be evaluated by the Cancer Assessment Review committee (CARC).

Although this study does not satisfy EPA guideline requirements, it provides supplementary information regarding the propos ed mechanism of action for carcinogenicity of sulfoxaflor in

rats and mice. This study fully reliable (acceptable/non

- guideline).

IIA 5.5.4/2 Targeted gene expression, cell proliferation and cytochrome P450 enzymatic activity in rats; In previous studie s with sulfoxaflor, increased liver weight was observed. Targeted gene expression data in mice and hepatocellular proliferation data in both mice and rats suggest a possible phenobarbital (PB)

- like mode of action (MoA) could be responsible for the observe d liver weight increases, and subsequent liver tumors. The molecular response stimulated by PB is mediated by the constitutive androstane nuclear receptor (CAR), with peripheral involvement of the pregnane X receptor (PXR), and typically results in increa sed hepatocellular proliferation and hypertrophy, with associated increased liver weight, inhibition of apoptosis, hypertrophy, and eventual development of altered hepatic foci. In this study (MRID 47832061), five male and

five female Fischer 344/DuCrl ra ts per group were given sulfoxaflor in the diet at 0, 100, 750, or

1500 ppm for 3 or 7 days. The primary endpoints examined were liver weight, targeted gene expression, liver enzyme analysis, and hepatocellular proliferation.

Decreased body weights an d body weight gains were seen in males and females at 1500 ppm after 3 and 7 days. After 3 days, body weight was decreased 7% in males and 9% in females and body weight gain was decreased 60% and 100%, respectively. After 7 days, there was a 7%

decrease in male body weight and 43% and 47% decrease in body weight gain in males and female, respectively. Food consumption in males at 750 and 1500 ppm for 3 days and 1500 ppm for 7 days was significantly lower (15 and 30%

- day 3; 23%

- day 7, respectively). A similar pattern of reduced food consumption was seen in females exposed to 750 and 1500 ppm for 3 days and 1500 ppm for 7 days (18 and 35%, day 3; 20% day 7, respectively).

Elevated cholesterol levels were seen in males on day 3 at 750 or 1500 ppm (19 and 29%, respectively) and day 7 at 750 or 1500 ppm (33% and 87%, respectively). In females, elevated cholesterol levels were seen after 7 days at 1500 ppm (41%). Relative liver weights were increased for males and females in the 1500 ppm group at 3 day s (14 and 3%, respectively) and at 750 and 1500 ppm at 7 days (11 and 23% for males and 6 and 18% for females, respectively). Absolute liver weights were increased in male and females at 1500 ppm after 7 days (14 and 16%, respectively).

Cy p2b1 gene expres sion, the prototypical gene response following PB exposure, was induced

over 800

- fold in both male and female rats exposed to 1500 ppm sulfoxaflor for 3 and 7 days.

To further examine similarities in PB

- like response, Cyp2b2 and Cyp3a3 (CAR

- and PXR

relat ed genes, respectively) expression levels, together with PROD and BROD enzyme activity were evaluated at 3 and 7 days. These molecular markers were elevated in males and females exposed to 750 and 1500 ppm at both time periods in a PB

- like manner. At 7 d ays, male rats exposed to 750 and male and female rats at 1500 ppm sulfoxaflor showed significantSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 106 of 176 hepatocellular proliferation; for males it occurred in the centrilobular and midzonal regions, whereas for females it was evident in the centrilobular region.

An additional aim of this study was to determine if other nuclear receptors might have played a

role in sulfoxaflor

- induced liver effects. Four nuclear receptors are primarily responsible for

xenobiotic

- induced liver weight increase; the aryl hydroc arbon receptor (AhR), CAR, PXR, and the peroxisome proliferator

- activated receptor alpha (PPARα). Cyp1a1 gene expression and EROD enzyme activity were slightly but significantly elevated at day 3; however, EROD enzyme activity returned to control levels b y day 7. Furthermore, gene expression of Cyp4a22 was not elevated in this study. From these data it is implied that sulfoxaflor is not likely an AhR or PPARα agonist.

In conclusion, the data suggests that sulfoxaflor may be an agonist ligand for CA R, and this activation results in the observed phenobarbital

- like response in the liver. However, the carcinogenic potential of sulfoxaflor, along with the proposed MOA for liver tumors will be evaluated by the Cancer Assessment Review Committee (CARC).

T his non

- guideline study is reliable with restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to G LP standards. Additionally, the study was well conducted and the Agency has confidence in t he scientific integrity of the data.

IIA 5.8.1.5/1 X11719474: Targeted Gene Expression, Cell Proliferation, and Cytochrome p 450 Enzymatic Activities in Rats;

T he purpose of this study (MRID 47832151) was to generate information to aid in understanding the mode of action for liver effects in rats induced by X11719474 a metabolite of sulfoxaflor and to compare the results to those observed following exposure to su lfoxaflor . In this study, male F344/DuCrl rats were given test diets formulated to supply 0 or 8000 ppm X11719474 in the diet for 7 days which corresponded to time

- weighted average doses of 0 or 583 mg/kg/day, respectively. The treatment level of 8000 ppm was the high dose level used in the rat 28

- day toxicity study. The parameters evaluated were daily cageside observations, body weights, body weight gains, feed consumption, liver weights, histopathology of the liver, focused gene expression, liver cytochro me P450 enzyme activity, and hepatocellular proliferation using BrdU immunohistochemical methods. Results from this study were compared to an earlier work in which male F344/DuCrl rats were exposed to 1500 ppm sulfoxaflor for 7 days, resulting in a

time

- we ighted average dose of 102 mg/kg/day.

At the end of the 7

- day treatment period, males exposed to 8000 ppm X11719474 had body weight and body weight gains decrements of approximately 2% and 22% relative to controls, respectively. In addition, males given 8000 ppm X11719474 had statistically identified,

treatment

- related increases in liver weights (absolute: 16%; relative: 19%). X11719474 induced liver weight increases that correlated with treatment

- related microscopic observations of very slight centrilob ular/midzonal hypertrophy of hepatocytes that were present in all treated animals.

At the end of the 7

- day treatment period, males exposed to 8000 ppm X11719474 had the same types of treatment

- related effects as parent sulfoxaflor , but were consistently le ss marked despite

the almost 6

- fold higher dose level.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 107 of 176 The constitutive androstane receptor (CAR)

- related transcripts, Cyp2b1 and Cyp2b2, along with the pregnane X receptor (PXR)

- related transcript Cyp3a3, were increased 806, 32, and 4

- fold, respectively. Sulfoxaflor

- induced gene expression showed a similar profile of 848, 21, and 9

- fold, respectively. The level of hepatocellular PROD enzyme activity, a CAR

- associated biomarker, was increased 15

- fold following X11719474 exposure. A similar level of induct ion (10

- fold) was seen for sulfoxaflor

- induced PROD enzyme activity. Hepatocellular proliferation, as measured by BrdU incorporation, was significantly elevated 4.5, 2.5, and 2.1

- fold in the centrilobular, midzonal, and periportal regions, respectively. A n earlier hepatocellular proliferation study with sulfoxaflor , as measured by Ki

- 67, showed increased proliferation in the centrilobular and midzonal regions of 3.4 and 2.9

- fold, respectively.

In conclusion, the data suggests that sulfoxaflor and its metabolite X11719474 may be agonist ligand for CAR, and this activation results in the observed phenobarbital

- like response in the liver. However, the carcinogenic potential of sulfoxaflor, along with the proposed MOA for liver tumors will be evaluated by the Cancer Assessment Review Committee (CARC).

This non

- guideline study is reliable with restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to GLP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific integrity of the data.

IIA 5.5.4/3 Mode of Action Study Investigating Liver Weight Effects in crl:cd1(icr) Mice;

In a non

- guideline study, (MRID 47832062) 5 male and 5 female Crl:CD

- 1 (ICR ) mice were exposed to different dose levels of sulfoxaflor in the diet for 7 days. In previous studies with sulfoxaflor, increased liver weight was observed. Targeted gene expression data in mice and hepatocellular proliferation data in both mice and rats suggested a phenobarbital (PB)

- like mode of action (MoA) could be responsible for the observed liver weight increase. The molecular response stimulated by PB is mediated by the constitutive androstane nuclear receptor (CAR), with peripheral involvement of the pregnane X receptor (PXR), and typically results in increased hepatocellular proliferation and hypertr ophy, with associated increased liver weight, inhibition of apoptosis, hypertrophy, and eventual development of altered hepatic foci. In this study, male

CD

- 1® mice were exposed to 0, 500, or 750 ppm, while female mice were exposed to 0 , 1000, or 1500 ppm sulfoxaflor in the diet for 7 days. Endpoints evaluated were daily cage

- side observations, body and liver weights, feed consumption, serum clinical chemistries, focused gene expression, liver cytochrome P450 enzyme activity, and hepatocellular proliferat ion using

BrdU and Ki

- 67 methods. Archived liver samples from previously conducted 28 and 90

- day sulfoxaflor mouse studies were analyzed for targeted gene expression, liver enzyme activity, and hepatocellular proliferation (Ki

- 67).

In the 7

- day study, males exposed to 750 ppm had a statistically identified increase (17%) in relative liver weights compared to controls. Females exposed to 1000 and 1500 ppm had a statistically identified increase in relative (38 and 43%, respectively) and absolute (43 and 47%, respectively) liver weights compared to controls. These liver weight increases correlated with

treatment

- related observations of centrilobular and midzonal hepatocyte hypertrophy with very slightly increased cytoplasmic eosinophilia. Analysis by Oi l Red O stain demonstrated slightlySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 108 of 176 increased amounts of lipid in the hepatocytes in males exposed to 750 ppm sulfoxaflor, however no changes were observed in females.

Cyp2b10 expression, considered to be the prototypical gene response following PB exposu re through activation of CAR, was induced 42.1 and 54.8

- fold in males exposed to 500 and 750 ppm, respectively, and 20.0 and 30.8

- fold, in females exposed to 1000 and 1500 ppm, respectively, relative to controls. Cyp2b10 gene expression in males from the 28 and 90

- day studies was increased 61.7 (300 ppm) and 56.5

- fold (750 ppm), respectively. Females exposed to 1500 ppm showed elevated Cyp2b10 expression at 28 and 90 days of 93.9

- and 53.9

- fold, respectively. Cyp3a11 , a PB

- and PXR

- related gene, was sign ificantly elevated in the 750 ppm males at 7 and 90 days, while in female mice it was significantly elevated at all doses and time periods (range of 3.4 to 6.6

- fold). PROD and BROD liver enzyme activities, which give a measure of Cyp2b enzyme induction, w ere elevated in both male and female mice at all time

points (range of 2.6 to 9.5

- fold).

Both BrdU and Ki

- 67 immunohistochemical techniques were used to measure hepatocellular proliferation in the 7

- day study. For the BrdU analysis, males exposed to 50 0 ppm sulfoxaflor had elevated proliferation in the centrilobular region and at 750 ppm in both the centrilobular and midzonal regions. Females exposed to 1000 and 1500 ppm showed significant induction in all three regions. For Ki

- 67 analysis, males exp osed to 500 and 750 ppm showed significant induction in the centrilobular region, however unlike the BrdU analysis, increased proliferation was not observed at 750 ppm in the midzonal region. In females, increased proliferation was not evident at any dose or zone by Ki

- 67 analysis. Ki

- 67 analysis of hepatocellular proliferation in

the 28 and 90

- day studies showed no induction at either time point in male or female mice. An additional aim of this study was to determine if other nuclear receptors might ha ve played a

role in sulfoxaflor

- induced liver effects. Four nuclear receptors are primarily responsible for

xenobiotic

- induced liver weight increase; the aryl hydrocarbon receptor (AhR; Cyp1a), CAR (Cyp2b), PXR (Cyp3a), and the peroxisome proliferator

- ac tivated receptor alpha (PPARα; Cyp4a). AhR

- related EROD liver enzyme activity was slightly elevated in this study at all time

points in both male and female mice; however, the degree of induction was mild (none greater

than 2.3

- fold) and was most likely a ssociated with the large induction of Cyp2b enzyme. Cyp4a10 , a PPARα related gene, was not significantly altered in this study.

These data suggests that sulfoxaflor may be an agonist ligand for CAR, and this activation results

in the observed PB

- like response and increased liver weight. However, the carcinogenic potential of sulfoxaflor, along with the proposed MOA for liver tumors will be evaluated by the Cancer Assessment Review committee (CARC).

This non

- guideline study is reliable with restricti ons (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to G LP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific integrity of the data.

IIA 5.5.4/ 4 Induction profile and gene expression in mice livers;Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 109 of 176 In an induction profile study (MRID 47832 276 ), sulfoxaflor (95.6% pure; batch number E2162

- 34) was administered in the diet to 15 male mice at dose levels of 0, 750, or 1500 ppm (equivalent to 0, 160, and 310 mg/kg/day respectively) for 7 days.

Administration of dietary sulfoxaflor to mice for 7 days resulted in decreased body weight at the high dose of 1500 ppm compared to control. There was a treatment

- related increase in absolute

and relative li ver weights and ALT following seven days of exposure to sulfoxaflor. However,

no significant increases in plasma AST, cholesterol and triglycerides were seen . Administration

of sulfoxaflor at 750 ppm and 1500 ppm elicited a 3

- and 5

- fold increase in tot al hepatic P450, respectively, a 33

- fold increase in PROD activity at both concentrations, a 47

- and 82

- fo lding increase in BROD activity , respectively, and a 4

- and 7

- fold increase in BQ activity, respectively.

The study examined the expression of two ge nes, Cyp2b10 and Cyp3a11 . Gene expression indicated the presence of Cyp2b10 mRNA in only the treatment groups as it is constitutively expressed. The change in expression of Cyp2b10 from sulfoxaflor 750 ppm treatment to

sulfoxaflor 1500 ppm treatment was 9.2

- fold. Cyp3a11 is constitutively expressed in the mouse, and results can be expressed as a relative fold change over control values. Administration of 750 and 1500 ppm sulfoxaflor resulted in a 2.4

- and 5.6

- fold increase in Cyp3a 11 relative to controls.

Sulfoxaflor induced gene expression data was confirmed by sodium dodecyl sulphate

polyacrylamide gel electrophoresis (SDS

- PAGE) and Western blotting.

In conclusion, the data suggest that sulfoxaflor exerts its enzyme induction p roperties via CAR and possibly PXR. However, the carcinogenic potential of sulfoxaflor, along with the proposed MOA for liver tumors will be evaluated by the Cancer Assessment Review Committee (CARC).

This study is Reliable with Restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to GLP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific integrity of the data.

IIA 5.5.4/5 A St udy To Investigate The Mode Of Action For Liver Effects Observed In Regulatory Toxicology Studies By Use Of Dual Car

- PXR Knockout And Humanised Mice ; The aim of this study (MRID 47832255) was to investigate: (1) if the constitutive androstane receptor (C AR) or pregnane X receptor (PXR) mediate sulfoxaflor

- induced hypertrophy and hyperplasia in mice; and (2) if the human receptors (CAR and PXR) support these processes to a similar extent as the murine receptors. The mouse models used were wild type C57BL/6 J (WT) mice, C57BL/6J mice null for PXR and CAR (PXRKO/CARKO) and C57BL/6J mice “humanised” for PXR and CAR (hPXR/hCAR).

In this study, groups of ten male mice of each strain were offered diets containing 0 or nominally 750 ppm sulfoxaflor for seven days. The measured dietary concentration of sulfoxaflor was 740 ppm (approximately 98.6% of target) with good homogeneity. Endpoints evaluated were daily clinical observations, body weights, body weight gain, feed consumption, plasma clinical chemistries, focu sed gene expression, protein quantification, liver cytochrome P450 enzyme activity, hepatocellular proliferation using nuclear incorporation of BrdU, and liver histopathology.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 110 of 176 There were no treatment

- related clinical observations or effects on body weig ht or body weight gain in any strain of mouse. There were treatment

- related increases in absolute and relative liver weights in WT and hPXR/hCAR mice but not in the PXRKO/CARKO animals. Absolute liver weights for WT and hPXR/hCAR mice in the 750 ppm sulf oxaflor groups were 124 and 109% of controls respectively, and relative liver weights were 125 and 112% of controls respectively.

In WT mice, sulfoxaflor treatment increased hepatocellular proliferation (as measured by approximately 4

- fold but no such c hanges in proliferation were seen in either the hPXR/hCAR or

PXRKO/CARKO mice. Treatment

- related hepatocyte hypertrophy was observed in WT and hPXR/hCAR mice while increased mitotic figures were observed only in WT mice. Neither hepatocyte hypertrophy nor increased mitotic figures were seen in PXRKO/CARKO mice.

Sulfoxaflor behaved as a phenobarbital

- like inducer in WT mice. A marked induction of total cytochrome P450, markedly increased PROD and BROD (Cyp2b selective substrates), increased expression of C

yp2b10 mRNA (demonstrated by RT

- PCR), increases in Cyp2b10 protein (immunoblotting data) were observed after exposure to sulfoxaflor. However, in the hPXR/hCAR under the same conditions, induction of Cyp2b10 activity, protein and mRNA was markedly less th an observed in the WT animals following treatment with sulfoxaflor. In PXRKO/CARKO animals, sulfoxaflor had no significant effect on Cyp2b10 expression or catalytic activity.

Sulfoxaflor

- mediated Cyp3a11 induction, as determined by BQ activity (Cyp3a sel ective

r e action), RT

- PCR and immunoblotting was observed in the “humanised” and WT mice, to similar extents, but was not seen in the PXRKO/CARKO mice.

In conclusion, sulfoxaflor exhibited markedly more activity towards the mouse CAR than the human CAR and relatively weak activity towards the mouse and human PXR. This suggests that the difference in hepatic response between wild type and humanised mice in this study may be mediated via CAR. Additionally, the data show that the human CAR/PXR support sulfoxaflor

induced hypertrophy but not hyperplasia.

In conclusion, the data suggests that sulfoxaflor acts via a CAR

- mediated mode of action in mice

and it i s possible that exposure would not result in human carcinogenicity. However, these findings, along with those in other mechanistic studies, will be used by the Cancer Assessment Review Committee (CARC) to evaluate the carcinogenic potential of sulfoxaflor and the proposed MOA (and human relevance) for liver tumors.

This non

- guideline study is reliable with restrictions ( acceptable/non

- guideline ). The study is not GLP compliant. However, all experiments were done according to GLP standards. Additionally , the study was well conducted and the Agency has confidence in the scientific integrity of the data.

A.4.6 Mutagenicity

Gene MutationSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 111 of 176 IIA 5.4.1 In vitro bacterial ( Salmonella typhimurium and E. coli ) / mammalian activation gene mutation assay ; In independent trials of a reverse gene mutation assay (MRID 47832047), sulfoxaflor (Purity 96.6%; Lot/Batch No. E2198

- 17), prepared in dimethyl sulfoxide (DMSO), was tested in 4

strains of S. typhimurium (TA100, TA98, TA1535 and TA1537) and in E. coli WP2 uvrA in a

pre

- incubation reverse mutation assay at concentrations ranging from 100 to 5000 μg/plate (both trials) with and without S9 activation. The S9 fraction was derived from the livers of male Sprague

- Dawley rats induced with Aroclor 1254.

In both a ssays, the test material was not cytotoxic or mutagenic for any strain at concentrations up to the limit dose for this test system either in the absence or presence of S9 activation. The

numbers of revertant colonies in all strain

- specific positive control groups were clearly

increased.

Under the conditions of this study, sulfoxaflor did not induce gene mutation in any of the strains employed, either with or without metabolic activation, at concentrations up to the limit dose for this test system.

The st udy is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirements (OPPTS 870.5100; OECD 471) for in vitro mutagenicity (bacterial reverse gene mutation) data.

IIA 5.8.1.4/1 In vitro bacterial ( Salmonella typhimurium ) / mamm alian activation gene mutation assay; In independent trials of a reverse gene mutation assay (MRID 47832097), X11719474 (Purity

99.6% ; Lot/Batch No. XS9

- 36891

- 82, TSN106429 and Lot /Batch No. XS9

- 37307

- 78,

TSN030626

- 0001 ) , prepared in dimethyl sulfoxide (DM SO), was tested in 4 strains of S. typhimurium (TA100, TA98, TA1535 and TA1537) and in E. coli WP2 uvrA in a pre

- incubation reverse mutation assay at concentrations ranging from 100 to 5000 μg/plate (both trials) with and

without S9 activation. The S9 frac tion was derived from the livers of male Sprague

- Dawley rats

induced with Aroclor 1254.

In both assays, the test material was not cytotoxic or mutagenic for any strain at concentrations up to the limit dose for this test system either in the absence or pr esence of S9 activation. The

numbers of revertant colonies in all strain

- specific positive control groups were clearly

increased.

Under the conditions of this study, X11719474 did not induce gene mutation in any of the strains employed, either with or without metabolic activation, at concentrations up to the limit dose for this test system.

The study is classified as fully reliable (acceptable/guideline) and satisfies the guide line requirements (OPPTS 870.5100; OECD 471) for in vitro mutag enicity (bacterial reverse gene mutation) data.

IIA 5.8.2.4/1 Salmonella

– Escherichia coli /mammalian

- microsome reverse mutation assay preincubation method with a confirmatory assay with X11721061;Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 112 of 176 In independent trials of a reverse gene mutation assay (MRID 47832085), [X11721061 Purity 99%; Lot/Batch No. GF908627 (TSN030701

- 0003)], prepared in dimethyl sulfoxide (DMSO)

was tested in 4 strains of S. typhimurium (TA100, TA98, TA1535 and TA1 537) and in E. coli

WP2 uvr A in a pre

- incubation reverse mutation assay at concentrations ranging from 1.5 to 5000 μg/plate (Initial cytotoxicity

- mutation assay) and 50 to 5000 μg/plate (Confirmatory assay) both

with and without S9 activation. The S9 fract ion was derived from the livers of male Sprague

Dawley rats induced with Aroclor 1254.

In the initial assay, there was some indication of a slight cytotoxic effect (reduced revertant colony counts) for several strains at the highest concentration; however , this was not reproduced in the confirmatory trial. Nevertheless, the data are in good agreement and indicate that the test material was not mutagenic in any strain at concentrations up to the limit dose for this test system either in the absence or pres ence of S9 activation. The numbers of revertant colonies in

all strain

- specific positive control groups were clearly increased.

Under the conditions of this study, X11721061 did not induce gene mutation in any of the strains employed, either with or witho ut metabolic activation, at concentrations up to the limit dose for this test system

.

The study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirements (OPPTS 870.5100; OECD 471) for in vitro mutagenicity (bacterial reverse gene mutation) data.

IIA 5. 8.3.4 / 1 Salmonella

– Escherichia coli /mammalian

- microsome reverse mutation assay preincubation method with a confirmatory assay with X11596066; In independent trials of a reverse gene mutation assay (MRID 47832076), the su lfoxaflor

metabolite X11596066 [Purity 98%; Lot/Batch No. SYN

- 6268

- 05

- 06 (TSN031296

- 0002)], prepared in dimethyl sulfoxide (DMSO), was tested in 4 strains of S. typhimurium (TA100, TA98, TA1535 and TA1537) and in E. coli WP2 uvr A in a pre

- incubation reverse mutation assay at concentrations ranging from 1.5 to 5000 μg/plate (Initial cytotoxicity

- mutation assay) and 15 to 5000 μg/plate (Confirmatory assay) both with and without S9 activation. The S9 fraction was derived from the livers of male Sprague

- Dawley rats induced with Aroclor 1254.

In both assays, concentrations ≥1500 μg/plate +/

- S9 were cytotoxic for any strains. However, there were no increases in the number of revertant colonies at any noncytotoxic level, in any strains, both with and without S9. The numbers of revertant colonies in all strain

- specific positive control groups were clearly increased.

Under the conditions of this study, X11596066 did not induce gene mutation in any of the strains employed, either with or without metabolic activation , at concentrations up to those producing evidence of cytotoxicity.

The study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirements (O PPTS 870.5100; OECD 471) for in vitro mutagenicity (bacterial reverse gene mutation) data.

IIA 5.8.4.4/1 Salmonella

– Escherichia coli /mammalian

- microsome reverse mutation assay preincubation method with a confirmatory assay with X11579457;Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 113 of 176 In independent trials of a reverse gene mutation assa y (MRID 47832070), the sulfoxaflor

metabolite X11579457 ( Purity 99%; Lot/Batch No. SYN

- 5866

- 65 (TSN030941

- 0002), was prepared in distilled H 2 O, and tested in 4 strains of S. typhimurium (TA100, TA98, TA1535 and TA1537) and in E. coli WP2 uvrA in a pre

- incub ation reverse mutation assay at concentrations ranging from 1.5 to 5000 μg/plate (Initial cytotoxicity

- mutation assay) and 50 to 5000 μg/plate (Confirmatory assay) both with and without S9 activation. The S9 fraction was derived from the livers of male Sp rague

- Dawley rats induced with Aroclor 1254.

In both assays, the test material was not cytotoxic or mutagenic for any strain at concentrations up to the limit dose for this test system either in the absence or presence of S9 activation. The

numbers of rev ertant colonies in all strain

- specific positive control groups were clearly

increased.

Under the conditions of this study, X11579457 did not induce gene mutation in any of the strains employed, either with or without metabolic activation, at concentratio ns up to the limit dose for this test system.

The study is classified as fully reliable (acceptable/guideline) an d satisfies the guideline requirements (OPP TS 870.5100; OECD 471) for in vitro mutagenicity (bacterial reverse gene mutation) data.

IIA 5.8.5 .4/1 Salmonella

– Escherichiacoli /mammalian

- microsome reverse mutation assay preincubation method with a confirmatory assay with X11519540; In independent trials of a reverse gene m utation assay (MRID 47832068), the sulfoxaflor

metabolite X11519540 ( Purity 98%; Lot/Batch No. 37307

- 15 (TSN106498), prepared in dimethyl sulfoxide (DMSO), was tested in 4 strains of S. typhimurium (TA100, TA98, TA1535 and TA1537) and in E. coli WP2 uvr A in a pre

- incubation reverse mutation assay at concentrations ranging fr om 1.5 to 5000 μg/plate (Initial cytotoxicity

- mutation assay) and 50 to 5000 μg/plate (Confirmatory assay) both with and without S9 activation. The S9 fraction was derived from the livers of male Sprague

- Dawley rats induced with Aroclor 1254.

In both as says, the test material was not cytotoxic or mutagenic for any strain at concentrations up to the limit dose for this test system either in the absence or presence of S9 activation. The

numbers of revertant colonies in all strain

- specific positive control groups were clearly

increased.

Under the conditions of this study, X11519540 did not induce gene mutation in any of the strains employed, either with or without metabolic activation, at concentrations up to the limit dose for this test system.

The study is classified as fully reliable ( acceptable/guideline) and satisfies the guideline requirements (O PPTS 870.5100; OECD 471) for in vitro mutagenicity (bacterial reverse gene mutation) data.

Cytogenetics

IIA 5.4.3/1 In Vitro Gene Mutation assay in Chinese hamster ovary cells;Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 114 of 176 In independent trials of a mammalian cell forward gene mutation assay (MRID 47832049), Chinese hamster ovary (CHO/HGPRT) cells were exposed to sulfoxaflor (Purity 96.6% Lot #

E2198

- 17, TSN106108), prepared in dimethyl sulfoxide (DMSO) , at concentrations ranging from 173.3 to 2773 μg/ml in the absence and presence of S9. The S9 liver homogenate was derived from the livers of Sprague

- Dawley rats induced with Aroclor 1254. The highest concentration was the 10 mM limit for the assay system . Positive control chemicals used in this assay were ethyl methanesulfonate (EMS) in the absence of S9 and 20

- methylcholanthrene (20

- MCA) in the presence of S9.

Sulfoxaflor was tested up to the limit dose for this test system and failed to induce either a cytotoxic or mutagenic effect in either the absence or the presence of S9 activation. The expected responses were obtained with the negative and positive controls either with or without S9

activation . It was, therefore, concluded that sulfoxaflor was not active in this test system.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement (OPPTS 870.5300, OECD 476) for in vitro mutagenicity (mammalian forward gene mutation) data.

IIA 5.8.1.4/3 Evaluation of X11719474 in the Chinese hamster ovary cell/ hypoxanthine

guanine

- phosphoribosyl transferase (CHO/HGPRT) forward mutation assay; In independent trials of a mammalian cell forward gene mutation assay (MRID 47832095), Chinese hamster ovary (CHO /HGPRT) cells were exposed to X11719474 (Purity 100%; Lot #

XS9

- 36891

- 82, TSN106429), prepared in dimethyl sulfoxide (DMSO) at concentrations ranging from 184.6 to 2953 μg/ml in the absence and the presence of S9. The highest concentration was

based on th e 10 mM limit of the assay system. The S9 liver homogenate was derived from the livers of Sprague

- Dawley rats Aroclor 1254

- induced. The highest concentration was the 10 mM limit for the assay system. Positive control chemicals used in this assay were eth yl methanesulfonate (EMS) in the absence of S9 and 20

- methylcholanthrene (20

- MCA) in the presence of S9.

X11719474 was tested up to the limit dose for this test system and failed to induce either a cytotoxic or mutagenic effect in either the absence or the presence of S9 activation. The expected responses were obtained with the negative and positive controls either with or without S9 activation. It was, therefore, concluded that X11719474 was not active in this test system.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement (OPPTS 870.5300, OECD 476) for in vitro muta genicity (mammalian forward gene mutation) data.

IIA 5.8.2.4/3 Evaluation of X11721061 in the Chinese hams ter ovary cell/ hypoxanthine

guanine

- phosphoribosyl transferase (CHO/HGPRT) forward mutation assay; In independent trials of a mammalian cell forward gene mutation assay (MRID 47832077), Chinese hamster ovary (CHO/HGPRT) cells were exposed to the sulfoxafl or metabolite

X11721061 (Purity 99%; Lot # GF908627, TSN030701

- 0003), prepared in distilled H 2 O, at concentrations ranging from 120 to 1920 μg/ml (both trials) in the absence and the presence of

S9. The S9 liver homogenate was derived from the livers of S prague

- Dawley rats induced with Aroclor 1254. The highest concentration was the 10 mM limit for the assay system. PositiveSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 115 of 176 control chemicals used in this assay were ethyl methanesulfonate (EMS) in the absence of S9

and 20

- methylcholanthrene (20

- MCA) in t he presence of S9.

X11721061 was tested up to the limit dose for this test system and failed to induce either a cytotoxic or mutagenic effect in either the absence or the presence of S9 activation. The expected responses were obtained with the negative and positive controls either with or without S9

activation. It was, therefore, concluded that X11721061 was not active in this test system.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement (OCSPP 870.5300, OECD 476) for in vitro mutagenicity (mammalian forward gene mutation) data.

I IA 5.8.4.4/3 Evaluation of X11579457 in the Chinese hamster ovary cell/ hypoxanthine

guanine

- phosphoribosyl transferase (CHO/HGPRT) forward mutation assay; In independent trials of a mammalian cell forward gene mutation assay (MRID No. 48288933), Chinese hamster ovary (CHO/HGPRT) cells were exposed to X11579457 (Purity 99%; Lot#

SYN

- 5866

- 65, TSN030941

- 0002), prepared in distilled H 2 O, at concentrations rangin g from 157.8 to 2525 μg/mL (both trials) in the absence and the presence of S9. The S9 liver homogenate was derived from the livers of Sprague

- Dawley rats induced with Aroclor 1254. The highest concentration was the 10 mM limit for the assay system. Posi tive control chemicals used in this assay were ethyl methanesulfonate (EMS) in the absence of S9 and 20

- methylcholanthrene (20

- MCA) in the presence of S9.

X11579457 was tested up to the limit dose for this test system and failed to induce either a
cytot oxic or mutagenic effect in either the absence or the presence of S9 activation. The expected responses were obtained with the negative and positive controls either with or without S9 activation. It was, therefore, concluded that X11579457 was not active i n this test system.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement (OPPTS 870.5300, OECD 476) for in vitro mutagenicity (mammalian forward gene mutation) data.

IIA 5.8.5.4/3 Evaluation of X1151 9540 in the Chinese hamster ovary cell/ hypoxanthine

guanine

- phosphoribosyl transferase (CHO/HGPRT) forward mutation assay; In independent trials of a mammalian cell forward gene mutation assay (MRID No. 48288934), Chinese hamster ovary (CHO/HGPRT) cells w ere exposed to the sulfoxaflor metabolite X11519540 (Purity 98%; Lot # 37307

- 15, TSN106498), prepared in dimethyl sulfoxide (DMSO) concentrations ranging 158.8 to 2540 μg/ml in the absence and the presence of S9. The highest

concentration was based on the 10 mM limit of the assay system. The S9 liver homogenate was derived from the livers of Sprague

- Dawley rats Aroclor 1254

- induced. The highest concentration was the 10 mM limit for the assay system. Positive control chemicals used in this assay were ethy l methanesulfonate (EMS) in the absence of S9 and 20

- ethylcholanthrene (20

- MCA) in the presence of S9.

X1151940 was tested up to the limit dose for this test system and failed to induce either a cytotoxic or mutagenic effect in either the absence or the presence of S9 activation. The expectedSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 116 of 176 responses were obtained with the negative and positive controls either with or withou t S9 activation. It was, therefore, concluded that X11519540 was not active in this test system.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requireme nt ( OPPTS 870.5300, OECD 476) for in vitro mutagenici ty (mammalian forward gene mutation) data

Other Genotoxicity

IIA 5.4.2/1 Evaluation of sulfoxaflor in an in vitro chromosomal aberration assay utilizing rat lymphocytes ; In an in vitro chromosome aberration test (MRID 47832050), primary rat lymphocytes, derived

from 10

- 11 week old Sprague

- Dawley male rats, were exposed to sulfoxaflor (Purity 96.6%

Batch/Lot No. E2198

- 17, TSN106108) prepared in dimethyl sulfoxide (DMSO) at 0, 86.7, 173.3,
346.6, 693.3, 1389.5, and 2773 g/mL without and with S9 activation for 4 hours and continuously for 24 hours at 0, 21.7, 43.3, 86.7, 173.3, 346.6, 693.3, 1386.5 and 2773 g/mL without S9. The highest concentration tested approximates the limit dose of 10 mM and the S9 liver homogenate was p repared from Aroclor 1254

- induced male Sprague

- Dawley rats. Base on the analysis of mitotic indices (MIs), cultures treated for 4 hours with 0, 693.3, 1386.5, and 2773 g/mL +/

- S9 and cultures treated continuously for 24 hours with 173.3, 346.6 and 693.3 g/mL

– S9 were scored for structural and numerical chromosome aberrations.

Relative MIs (RMIs) were ≥ 50% of control at 2773 g/mL

– S9 (4

- hour treatment); 2200 g/mL +S9 (4

- hour treatment); and 346.6 g/mL

– S9 (24

- hour treatment). There were no significan t increases in the frequencies of cells with aberrations in either the presence or absence of S9 activation. Cultures treated with the positive control chemicals (mitomycin C without S9 and cyclophosphamide with S9) had significantly higher incidences of abnormal cells in all assays.

Based upon these results, sulfoxaflor was not considered to be clastogenic in this in vitro chromosomal aberration assay utilizing rat lymphocytes.

This study is classified as totally reliable (acceptable/guideline) and sa tisfies the guideline requirement (OPPTS 870.5375, OECD 473) for in vitro mammalian cytogenetics (chromosome aberrations) data.

IIA 5.8.1.4/2 Evaluation of X11719474 in an in vitro chromosomal aberration assay utilizing rat lymphocytes;

In an in vitro chr omosome aberration test (MRID 47832096), lymphocytes, derived from male Sprague

- Dawley rats w ere exposed to X11719474 (Purity 100%; Batch/Lot No. XS9

- 36891

- 82, TSN106429), prepared in dimethyl sulfoxide (DMSO) at co ncentrations ranging from 184.6 to

2953 g/mL for 4 hours in the presence and absence of S9 activation and for 24 hours without S9. The highest concentration was based on the limit dose of 10 mM in this assay system. The

S9 was derived from the livers of male Sprague

- Dawley rats induced with A roclor 1254. Metaphases, harvested from cultures treated with 738.25, 1476.5 and 2953 g/mL

- S9 (4

- and 24

- hour treatments) and 369.1, 738.25 and 2953 g/mL +S9 (4

- hour treatment), w ere examined for

t he incidences of cells with structural aberrations and n umerical aberrations.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 117 of 176 X11719474 was not severely cytotoxic nor clastogenic at any concentration up to the limit dose for this test system following a 4

- hour exposure. For the prolonged, 24

- exposure, the MI was

reduced to 32.3% of control at 2953 g/mL

– S9; however, the test material was not clastogenic
at any non

- cytotoxic concentration. The expected responses were induced by the non

- activated

and S9

- activated positive controls, mitomycin C and cyclophosphamide, respectively. Based upon these results, X11719474 was considered not to be clastogenic in this in vitro chromosomal aberration assay utilizing rat lymphocytes.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement (OPPTS 870.5375, OECD 473) for in vitro mammalian cytogenetics (chromosome aberrations) data.

IIA 5.8.2.4/2 Evaluation of X11721061 in an in vitro ch romosomal aberration assay utilizing rat lymphocytes;

In an in vitro chromosome aberration test (MRID 47832078), lymphocytes, derived from male Sprague

- Dawley rats w ere exposed to X11721061 (Purity 99%; Batch/Lot No. GF908627,TSN030701

- 0003), prepared in d istilled water

, at concentrations ranging from 30 to

1920 g/mL for 4 hours in the presence and absence of S9 activation and at concentrations ranging from 15 to 1920 g/mL for 24 hours without S9. The highest concentration was based

on the limit dose of 10 mM in this assay system. The S9 was derived from the livers of male Sprague

- Dawley rats induced with Aroclor 1254. Metaphases, harvested from cultures treated
with 480, 960 and 1920 g/mL +/

- S9 (4

- hour treatment) and 120, 240 and 480 g/mL

– S9 (24

- hour treatment), w ere examined for t he incidences of cells with structural aberrations and

numerical aberrations.

X11721061 was not severely cytotoxic nor clastogenic at any concentration up to the limit dose for this test system following a 4

- hour e xposure. For the prolonged, 24

- exposure, MIs were
≥50% of control at concentrations ≤480 g/mL

– S9; however, the test material was not clastogenic at any non

- cytotoxic concentration. The expected responses were induced by the

non

- activated and S9

- activat ed positive controls, mitomycin C and cyclophosphamide, respectively. Based upon these results, X11721061 was considered not to be clastogenic in this in vitro chromosomal aberration assay utilizing rat lymphocytes.

This study is classified as totally re liable (acceptable/guideline) and satisfies the guideline requirement (OPPTS 870.5375, OECD 473) for in vitro mammalian cytogenetics (chromosome aberrations) data.

IIA 5.8.5.4/2 Evaluation of X11519540 in an in vitro chromosomal aberration assay utilizing rat lymphocytes;

In an in vitro chromosome aberration test (MRID 48288935), lymphocytes, derived from male Sprague

- Dawley rats w ere exposed to X11519540 (Purity 98%; Batch/Lot No. 37307

- 15,TSN106498), prepared in dimethyl sulfoxide (DMSO), at concentratio ns ranging from 39. 7 to

25 40 g/mL for 4 hours in the presence and absence of S9 activation and at concentrations ranging from 19.8 to 25 40 g/mL for 24 hours without S9. The highest concentration was based on the limit dose of 10 mM in this assay system. The S9 was derived from the livers of maleSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 118 of 176 Sprague

- Dawley rats induced with Aroclor 1254. Metaphases, harvested from cultures treated
with 635, 1270 and 2540 g/mL +/

- S9 (4

- hour treatment) and 158.8, 317.5 and 635 g/mL

– S9 (24

- hour treatment), w ere exa mined for t he incidences of cells with structural aberrations and

numerical aberrations.

X11579457 was not severely cytotoxic nor clastogenic at any concentration up to the limit dose for this test system following a 4

- hour exposure. For the prolonged, 24

- hour exposure, MIs were
≥50% of control at concentrations ≤635 g/mL

– S9; however, the test material was not clastogenic at any non

- cytotoxic concentration. The expected responses were induced by the

non

- activated and S9

- activated positive controls, mi tomycin C and cyclophosphamide, respectively. Based upon these results, X11519540 was not clastogenic in this in vitro chromosomal aberration assay utilizing rat lymphocytes.

This study is classified as totally reliable (acceptable/guideline) and satisfies the guideline

req uirement (OPPTS 870.5375, OECD 473) for in vitro mammalian cytogenetics (chromosome aberrations) data.

IIA 5.4.2/2 Evaluation of X11579457 in an in vitro chromosomal aberration assay utilizing rat lymphocytes;

In an in vitro chr omosome aberration test (MRID 48288932), lymphocytes, derived from male Sprague

- Dawley rats w ere exposed to X11579457 (Purity 99%; Batch/Lot No. SYN

- 5866

- 65,TSN030941

- 0002), prepared in distilled H 2 O, at concentrations ranging from 39.5 to 2525 g/mL for 4 hours in the presence and absence of S9 activation and at concentrations ranging

from 19.7 to 2525 g/mL for 24 hours without S9. The highest concentration was based on the limit dose of 10 mM in this assay system. The S9 was derived from the livers of male Sprague

Dawley rats induced with Aroclor 1254. Metaphases, harvested from cultures treated with
631.3, 1262.5 and 2525 g/mL +/

- S9 (all treatment times), w ere examined for t he incidences of

cells with structural aberrations and numerical aberrations.

X11579457 was neither cytotoxic nor clastogenic at any concentration up to the limit dose for this test system. The expected responses were induced by the nonactivated and S9

- activated positive controls, mitomycin C and cyclophosp hamide, respectively. Based upon these results, X11579457 was considered not to be clastogenic in this in vitro chromosomal aberration assay utilizing rat lymphocytes.

This study is classified as fully reliable (acceptable/guideline) and satisfies the gu ideline requirement ( O PPTS 870.5375, OECD 473) for in vitro mammalian cytogenetics (chromosome aberrations) data.

IIA 5.4 .4 /1 In vivo mammalian cytogenetics

- erythrocyte micronucleus assay in mice ; In a bone marrow micronucleus assay (MRID 47832071), gro ups of 6 male and 6 female CD

- 1 mice were treated orally, by gavage, with 0, 100, 200 or 400 mg/kg/day of sulfoxaflor (Purity 95.6%; Lot/Batch No. E2162

- 34, TSN003725

- 0001) prepared in METHOCELTM on 2 consecutive days. The highest dose level of 400 mg/kg bw was selected for the main assay based on the results of a range

- finding test in which doses ≥1000 mg/kg bw/day caused more than 50% mortality in both sexes and 500 mg/kg bw/day caused unacceptable body temperature decreasesSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 119 of 176 in males only. Therefore, both sexes were evaluated in the main study. Groups of animals were

sacrificed at 24 hours after the second treatment for the collection of femoral bone marrow and evaluation of polychromatic erythrocytes (PCE, 2000 PCE/animal) with micronuclei (MN

- PCE)

from the first five animals in each group. The proportion of PCE was determined based upon 200 erythrocytes per animal and the results expressed as a percentage. Mice treated with 120

- mg/kg bw cyclophosphamide monohydrate by a single oral gavage dose and sacr ificed at 24 hours served as the positive control.

All animals survived to the end of the observation period. Treatment related clinical signs (decreased activity) occurred in 3/6 male mice at 400 mg/kg /day and two of these mice also had body temperat ure decreases of up to 5.8°C five hours post

- dosing. There were no statistically significant increases in the frequencies of MN

- PCE in groups treated with the test material as compared to the negative controls. There were no statistically significant dif ferences in the percent PCE in groups treated with the test material compared to negative controls. By contrast, a significant increase in the frequency of MN

- PCE and a significant decrease in the relative proportion of PCE: NCE (p<0.05) was seen in the p ositive control group as compared to the negative control group.

Under the experimental conditions used, sulfoxaflor was not genotoxic in the mouse bone

marrow micronucleus test.

This study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirement ( USEPA OPPTS 870.5395; OECD 474) for an i n vivo mammalian cytogenetics

– micronucleus assay in mice

.

A.4.7 Neurotoxicity

870.6100 Delayed Neurotoxicity Study

- Hen

870.6200 Acute Neurotoxicity Screening Battery

IIA 5.7.1/1 Acute Neurotoxicity

– Rats; In an acute neurotoxicity study (MRID 47832134), t en male and ten female F344/DuCrl rats per group were given a single, oral gavage dose of 0, 7.5, 75, or 750 mg sulfoxaflor/kg body weight to evaluate the potential for acute neurotoxicity. Body weights were recorded and a functional observational battery (FOB) and test for motor activity were conducted pre

- exposure (baseline), the day of dosing (day 1, time

- of

- peak effect), day 8, and day 15. The FOB included hand

- held and open

- field observations as well as measurements of grip performance, landing foot splay and rectal temperature. Clinical observations were conducted on days 2, 3, and 4. At the end of the study all rats from the control and high

- dose group and f ive rats/sex/low

- and mid

- dose group were perfused for histopathologic evaluation of the central and peripheral nervous systems, which was conducted on all of the control and high

- dose group rats. A second motor activity study at dose levels of 0, 2.5, 7. 5 and 25 mg/kg was conducted to investigate whether an apparent decrease in motor activity at 7.5 mg/kg was reproducible or treatment

- related, and toSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 120 of 176 establish a clear no

- observed

- adverse

- effect level (NOAEL). Motor activity was the only endpoint examined in this study phase.

One female rat given 750 mg/kg died following dosing on day 1, but the cause of death could

not be determined. Treatment

- related categorical observations on day 1 in males and females given 750 mg/kg included increased incidences of muscle tremors and twitches, convulsions, splayed hindlimbs and perineal urine soiling. Treatment

- related ranked FOB observations on day 1 in males and females given 750 mg/kg were as follows: increased lacrimation and salivation, decreased pupil size an d response to touch, increased level of urination (females only), decreased level of open

- field activity and gait abnormalities. There were no treatment

related ranked or categorical FOB observations present on day 8 or day 15 in males or females given 75 0 mg/kg. There were no treatment

- related ranked or categorical observations in males or females given 7.5 or 75 mg/kg during any FOB.

There was a treatment

- related decrease in body weight of the 750 mg/kg group when compared to controls on days 8 and 1 5, which was more prominent in males than in females. There was a

treatment

- related decrease in rectal temperature of the 750 mg/kg group when compared to controls on day 1, which was not present in the subsequent examinations on days 8 and 15.

There wer e no treatment

- related effects in grip performance or landing foot splay.

There was a treatment

- related decrease in the day 1 total motor activity and an effect on the distribution of motor activity counts of males and females given 75 or 750 mg/kg. The e ffect on total motor activity of animals given 7.5 mg/kg was considered equivocal on day 1. There were no effects on motor activity on days 8 or 15 in rats of any dose group. In the follow

- up motor activity study, there were no treatment

- related effects on total motor activity or on the distribution of motor activity counts for males and females given 2.5, 7.5 or 25 mg/kg when compared to controls.

Treatment

- related clinical findings on days 2, 3 or 4 were limited to males and females given 750 mg/kg, and included decreased or absent feces, red perioral soiling, and perineal urine soiling (females only). There were no treatment

- related gross or histopathologic findings in the central or peripheral nervous system.

The lowest

- observed

- adverse

- effect

- level (LOAEL) for neurotoxicity was 75 mg/kg based on decreased motor activity observed on day 1. The no

- observed

- adverse

- effect

- level (NOAEL) was 25 mg/kg. No treatment

- related effects were observed for neuropathology; therefore the NOAEL for neuropathology was ≥ 750 mg/kg, the highest dose level tested.

The study is classified as fully reliable (acceptable/guideline) and satisfies the guideline requirements (OPPTS 870.6200; OECD 424) for an acute neurotoxicity study in rats

.

870.6200 Subchronic Neurotoxicity Screening Battery (See 870.3100 (p. 85))

870.6300 Developmental Neurotoxicity Study

IIA 5.7.5 /1 Developmental Neurotoxicity StudySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 121 of 176 In a developmental neurotoxicity study in rats ( MRID 478321333), sulfoxaflor (purity 95.6%;

Lot # E2162

- 34) was offered on a continuous basis in the diet to 3 groups of 25 bred female Crl:CD(SD) rats daily from gestation day 6 through lactation day 21. Target test substance concentrations were 25, 100, and 400 parts per million (ppm), which corresponded to predicted dosage levels of 2, 8, and 32 mg/kg/day, respectively. Actual overall mean test substance consumption in the 25, 100, and 400 ppm groups was 1.8, 7.1, and 27.7 mg/kg/day through gestation a nd 1.9, 7.6, and 29.8 mg/kg/day through lactation, respectively. A concurrent control group composed of 25 bred females received the basal diet on a comparable regimen. Dams were approximately 13 weeks of age at the beginning of test diet exposure.

There were no test substance

- related mortalities in the dams during the study. There were no test

substance

- related clinical findings noted during the daily examinations. Detailed clinical observation parameters, as well as maternal body weights and food cons umption during gestation and lactation were unaffected by test substance exposure.

There were no test substance

- related differences noted between groups when comparing the mean length of gestation, the process of parturition, and internal macroscopic path ologic findings. The mean numbers of former implantation sites and unaccounted for sites, as well as maternal kidney and liver weights were similar across groups.

There were no test substance

- related effects on F 0 maternal parameters in this study. Ther efore,

the no

- observed

- adverse

- effect level (NOAEL) for F 0 systemic toxicity and F 0 maternal reproductive toxicity (process of parturition and duration of gestation) of sulfoxaflor when administered orally in the diet was considered to be 400 ppm (equivale nt to 2 7.7 mg/kg/day).

There were no test substance

- related effects on the mean number of pups born, live litter size, or the percentage of males at birth at any maternal exposure level. However, there was a reduction in postnatal survival from birth t o PND 4 compared to the control group in the 100 and 400 ppm dose groups. Furthermore, mean pup body weights in the 400 ppm group were 11.8% and 6.5% lower than the control group at birth (PND 1) and on PND 4, respectively. The reduced pup body weights re sulted in a statistically significant delay in surface righting response for pups in the 400 ppm group. Pup body weights in the 400 ppm group did not differ from the control group values on PND 7 or later time points. The decrease in postnatal survival a t 100 and 400 ppm is consistent with results from a previous probe reproduction study, in which dietary exposures of 500 and 1000 ppm resulted in decreased pup survival. The high dose level of 400 ppm in this study was based on the treatment

- related decre ase in survival observed in the probe study. Postnatal survival and pup body weights and body weight gains in the 25 and 100 ppm groups were unaffected by maternal test substance exposure. The age of attainment of surface righting response in the 25 and 100 ppm groups and eye opening in the 25, 100, and 400 ppm groups were similar to the control group. The attainment of sexual developmental landmarks (balanopreputial separation and vaginal patency) were unaffected by maternal test substance

exposure.

No remarkable clinical observations or macroscopic findings were noted for offspring at any exposure level. No test substance

- related effects were observed with respect to detailed clinical observations, locomotor activity, auditory startle response, and learning and memory. Furthermore, there were no test substance

- related effects on morphometric parameters or histopathology of the brain and/or central and peripheral nervous systems for offspring on PNDSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 122 of 176 21 and 72. On PND 72, slight, statistically signifi cant changes in brain weight (a decrease of 5% in males relative to controls; an increase of 4% in females relative to controls)) and brain length (a decrease of 4% in males relative to controls) were noted at 400 ppm.

The LOAEL for offspring is 100 ppm ( 7. 1 mg/kg bw/day) based on the reduction in pup survival. The NOAEL is 25 ppm (1.8 mg/kg bw/day).

The study is classified as fully reliable (acceptable/guideline) and may be used for regulatory purposes. It does not, however, satisfy the guideline requ irement for a developmental neurotoxicity study in rats (OPPTS 870.6300; OECD 426 (draft)) due to the pending review of the positive control data.

A.4.8 Metabolism

870.7485 Metabolism

– Rat

IIA 5.1.1/1 Single and repeat dose oral study in rats (and m ice);

In a metabolism probe study (MRID 47832038), the absorption, distribution, metabolism and

elimination of 14 C

- sulfoxaflor (>97% radiochemical purity, specific activity 42.0 mCi/mmol; lot no. INV 2058) in Fischer 344 DuCrl rats and Crl:CD1 (ICR) mice w as evaluated following a single oral gavage dose in 0.5% aqueous methylcellulose at a dosing volume of 5 mL/kg. One rat/sex and two mice/sex were administered 100 mg/kg 14 C

- sulfoxaflor. Time

- course blood and excreta were collected from rats, whereas only excreta was collected from mice up to 72 hours
post

- dosing. The only tissues collected at the 72 hr sacrifice were plasma and RBC. Urine samples collected during the first 12 hours post

- dosing were analyzed for parent compound and metabolite(s). Additi onally, two rats (1/sex) and four mice (2/sex) were orally dosed with 100 mg/kg 14 C

- sulfoxaflor and plasma collected at C

max (2 hour post

- dosing for rats and an estimated 11⁄2 hour post

- dosing for mice) and analyzed for parent and metabolite(s). This repor t also included results of the analysis of plasma and urine samples collected from 28

- day rat, 90

- day rat, and 28

- day mouse dietary subchronic toxicity studies.

Orally administered 14 C

- sulfoxaflor was rapidly absorbed from the GI tract both by rats and

mic e. Total urinary elimination accounted for 87

- 98% of the dose in rats and 80

- 85% of the dose in mice, most of which occurred within the first 24 hours post

- dosing. Fecal elimination accounted for only 5% in rats and 13% in mice, mostly apparently represe nting unabsorbed dose due to its recovery in feces within the GI transit time of 24 hours. Negligible or no radioactivity was eliminated via expiration. Elimination of radioactivity from blood was mono

- exponential

with t 1⁄2 of 9 and 11 hours from the plasma and RBC of male rats, respectively; the t 1⁄2 of

elimination of 14 C

- sulfoxaflor from the plasma and RBC of female rats was 7 and 8 hours, respectively. Female rats eliminated 14 C

- sulfoxaflor from the body at a 1.2

- fold faster rate than males, resulting in t he same difference in the AUC. No metabolism of the test material was observed following a single oral dose of 14 C

- sulfoxaflor to rats or mice. Sulfoxaflor was excreted mostly as unmetabolized parent, with only low levels of metabolites identified. Both parent sulfoxaflor and the X11602627 (XR

- 208) urea metabolite were observed in rat and mouse urine samples, but only parent sulfoxaflor was observed in rat plasma (mouse plasma not evaluated due to low radioactivity). The concentration of the urea metabo lite was similar to thatSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 123 of 176 detected in the test material itself as an impurity. In addition to the urea metabolite, 5 additional minor metabolites in the rat plasma and 3 possible minor metabolites rat urine were observed,

whereas one metabolite was observe d in mouse urine. Metabolite profiles were similar in urine and plasma samples from the 28

- or 90

- day dietary studies.

This study is classified as reliable with restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experim ents were done according to GLP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific integrity of the

data. It was conducted as a metabolism probe study and not to satisfy guideline requirements for

sulfox aflor.

IIA 5.1. 2/1 Toxicokinetic studies

– Second si ngle dose, oral route, in rats;

In a supplemental rat metabolism study (MRID 47832039), the tissue distribution of sulfoxaflor ( 14 C

- sulfoxaflor, radiochemical purity 99.3% a.i.; lot no. INV 027474

- 0001) was evaluated following a single oral gavage administration of 5 or 100 mg/kg body weight to groups of four

F344/DuCrl rats/sex/dose/sacrifice time. The 14 C

- sulfoxaflor test material was administered in 0.5% aqueous methylcellulose vehic le (dosing volume 5 mL/kg body weight). Animals were sacrificed at predetermined times of either C

max and 1⁄2C

max for plasma radioactivity levels (i.e., ranges 0.5

- 2 and 6

- 8 hours post

- dosing, respectively). Radioactivity was also determined in urine, fece s and cage wash for the 1⁄2 C

max groups. Additionally, the profile of sulfoxaflor

derived metabolites in three representative tissues (liver, kidney and plasma) was characterized by HPLC/ESI/MS for groups dosed at 100 mg/kg. A probe study was performed on 3 rats/sex at 100 mg/kg single gavage dose (sacrificed at C

max ) to develop extraction procedures for liver, kidney and plasma for identification of metabolites in the main study.

Orally administered 14 C

- sulfoxaflor was rapidly absorbed without any apparent lag time based

on C
max times of 0.5

- 1 hr (5 mg/kg) and 2 hr (100 mg/kg). Total recovery of radioactivity from all the animals averaged 98% in both groups.

Test material

- derived radioactivity (as μg 14 C

- sulfoxaflor eq/g) was highest in the GI tract, liver, kidney and urinary bladder, consistent with portal of entry and primary excretion tissues. Plasma levels were approximately 50

- 60% of those found in the kidney and liver in both groups. Comparison of the C

max and C 1/2 max tissue radioact ivity levels, together with comparisons of tissue to blood ratios, did not suggest the potential for bioaccumulation.

Parent sulfoxaflor, identified as its two diastereomers, was the only component detected in the kidney, liver and plasma samples above th e limits of detection of 0.286, 0.563 and 0.146 μg equivalent sulfoxaflor /g tissue, respectively, indicating no metabolism of sulfoxaflor.

In summary, administered sulfoxaflor was rapidly absorbed, widely distributed without metabolism, with the highest levels in portal of entry and excretory tissues. Test material

derived radioactivity in tissues (other than portal of entry and excretory) tracked that of blood and did not indicate potential for bioaccumulation.

This study is classified Fully Reliable ( a cceptable/ n on

- guideline). The study was conducted to provide supplemental information on tissue distribution and metabolite identification ofSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 124 of 176 sulfoxaflor at C
max and 1⁄2C

max and on its own does not by itself satisfy guideline requirements for a general met abolism study in the rodent.

IIA 5.1.3/1 Toxicokinetic studies

– Repeated dose, oral route, in rats ; In a rodent general metabolism study, (MRID 47832034), 14 C

- sulfoxaflor (radiochemical purity 99.7% a.i.; lot no. INV 2066) was administered to four g roups of male and female F344/DuCrl rats (4/ group/sex) at the following doses: low dose of 5 mg/kg via gavage as a single dose or as multiple doses (14 daily doses of unlabeled sulfoxaflor followed by a single radiolabeled dose); single gavage high dose of 100 mg/kg, or a single intravenous (IV) low dose of 5 mg/kg) . All oral doses were administered in 0.5% aqueous methylcellulose vehicle (5 mL/kg body weight).

Animals were monitored for 168 hours (7 days) post

- dosing to determine absorption,

distributi on, metabolism and excretion of 14 C

- sulfoxaflor.

Orally administered sulfoxaflor was rapidly absorbed without any apparent lag time. The percent absorption of the administered dose in all three oral groups (single 5 mg/kg, multiple 5 mg/kg or single 1 00 mg/kg) was at least 92

- 96%, based on recovery in urine, non

- GI tissues and expired air. Total recovery of radioactivity from all the animals averaged 102 ± 4 and 108 ± 4% in the oral and IV dose groups, respectively.

The oral doses were rapidly excreted in urine (92

- 97% of the administered dose) without any gender difference. The majority of the urinary elimination (89

- 94%) occurred within the first 24 hours post

- dosing. Only a small percent (5

- 8%) of the oral dose w as eliminated in feces, also mostly within the first 24 hrs post

- dosing.

The IV

- administered test material was also rapidly excreted in urine (97

- 101% of administered dose). The majority of the urinary elimination (91

- 95%) occurred in the first 24 hour s post

dosing. Only a small percent (6

- 9%) of the IV dose was eliminated in feces, mostly in the first 24 hours post

- dosing. Systemic bioavailability, calculated from the dose

- corrected plasma AUC data for the low oral and IV dose groups, was 94% for bot h male and female rats.

Only 0.2

- 1.2% of the administered 14 C

- sulfoxaflor (oral administration: low single, high single or multiple dose) remained in the tissues after 168 hours (7 days) post

- dosing. An average of

0.6

- 1.3% of the IV dosed 14 C

- sulfoxa flor remained in the tissues of the animals sacrificed 168 hours post

- dosing.

In the rats dosed orally with 14 C

- sulfoxaflor, elimination of the radioactivity from plasma was rapid during the elimination phase (t 1⁄2α = 4

- 6 hours) followed by relatively sl ow elimination during the terminal phase (t 1⁄2 = 39

- 45 hours). The area under curve (AUC) and C

max of radioactivity in plasma were essentially dose proportional between the low and high dose groups, suggesting unsaturated kinetic behavior of sulfoxaflor up to 100 mg/kg body weight. The fate of

the test

- material derived radioactivity in RBC was similar to plasma. The time

- course of radioactivity in RBC was essentially parallel to the plasma time

- course concentration profiles

with similar concentrations. Kinetics (absorption, elimination t 1⁄2 , AUC, clearance) of radioactivity in RBC were similar to plasma, except that detectable levels of radioactivity were found in RBC for an extended period of time compared to plasma.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 125 of 176 A total of seven radiolabeled compo nents were identified in urine and/or fecal samples. Parent sulfoxaflor was the primary component in both urine and fecal samples, accounting for >93% of the administered dose. A glucuronide conjugate of sulfoxaflor metabolite X11721061 was tentatively i dentified in urine samples only, comprising 2

- 4% of the administered dose. Three other unidentified minor components, all <1% of administered dose, were found only in pooled urine samples. Two different minor components, both <1% of the administered dose , were present only in extracts of fecal samples.

In summary, sulfoxaflor was rapidly and highly absorbed, poorly metabolized and readily eliminated primarily in the urine from the rat, with low tissue residues. ADME of sulfoxaflor did not show significa nt gender differences.

This study is classified as fully reliable (acceptable/g uideline) and satisfies the guideline requirements for a general metabolism study in the rodent.

IIA 5.8.1 .1/1 X11719474 : Probe Study to Determine Absorption, Metabolism and Elimination in F344/DuCrl Rats;

In a non

- guideline metabolism probe study (MRID 47832132), absorption, metabolism and

excretion of the sulfoxaflor metabolite 14 C

- X11719474 (radiochemical purity 98.8% a.i., lot #

XS9

- 100040

- 6, INV030930

- 0002 ) following a si ngle oral dose of 100 mg test material/kg body weight was evaluated in 1 F344/DuCrl rat /sex . Pharmacokinetics, mass balance and urine metabolite profiles were determined. The profile of circulating metabolites in plasma was

determined in 1 additional rat /sex.

Orally administered 14 C

- X11719474 was rapidly absorbed without any apparent lag time. T
max occurred at 1 hour. Total absorption was 95% of the administered dose in the male rat and 98%

in the female. A total of 95

- 97% of the administered dose was excreted in the urine after 168 hours post

- dosing , with t he majority (91

- 95%) rapidly eliminated by 12 hours post

- dosing. A much lower percentage (2

- 3%) of the dose was eliminated in feces by 168 hours post

- dosing. Total recovery of radioactivity from b oth animals was 98 and 99%.

Elimination of radioactivity from plasma was biphasic

, with most occurring during the rapid ( ) elimination phase (t 1⁄2α ≤2 hours), with a slower ( ) elimination phase (t 1⁄2 = 36

- 41 hours). The AUC of radioactivity in plasma was 505 and 351 μg h g - 1 for male and female, respectively.

Test

- material derived radioactivity was also present in RBC, with C

max concentrations ~27

- 31% lower levels than seen in plasma. Elimination of the radioactivity from RBC was monophasic with all of the elimination occurring during the rapid ( ) elimination phase (t 1⁄2α <3 hours). The AUC of radioactivity in RBC was 377 and 240 μg h g - 1 , in the male and female, respectively.

The absorbed test material was essentially un

- metabolized in the male and female rat. The primary component found in urine was parent compound, with two metabolites, or test material impurities, present at less than 1% of the administered dose. Only parent compound was fo und

in the C

max plasma samples.

In summary, administered 14 C

- X11719474 was rapidly absorbed, mostly un

- metabolized and

eliminated very quickly from the rat.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 126 of 176 This study is classified Fu lly Reliable (acceptable/n on

- guideline). I t was conducted as a prob e metabolism study on a metabolite of sulfoxaflor and was not submitted to satisfy guideline requirements for a rodent general metabolism study.

870.7600 Dermal Absorption

– Rat

IIIA 7.6.2 Comparative dermal absorption, in vitro using rat and human skin ; GF

- 2032 is a suspension concentrate (SC) formulation containing the active substance sulfoxaflor, an experimental insecticide. The nominal concentration of the active substance in

GF

- 2032 is 240 g/L. The highest in

- use spray dilution concentration is 0.48 g/L and the lowest

in

- use spray dilution concentration is 0.024 g/L. In an in vitro dermal penetration study (MRID 47832414), sulfoxaflor was applied as GF

- 2032 to the dermal surface of groups of 5 human skin samples/group, or 8 rat skin samples/grou p at concentrations of 0.024, 0.48, or 2.4 g/L for 24

hours at an application volume of 10 μL/cm 2

.

The overall recovery of the radiolabeled sulfoxaflor was good (95

- 97%) for all the groups. Application of 0.024 and 0.48 g/L resulted in similar rates of absorbed dose, 1.15, and 1.54 % of the administered dose in the human and 4.34 and 8.72% in the rat skin, respectively. The absorbed dose for the 240 g/L group was lower, 0.26 and 1.3%, in the human and rat skin, respectively, indicating saturation of th e skin at this level. In all dose groups, a greater percentage of sulfoxaflor was absorbed through the rat skin compared to the human skin. The rates were most similar in the 0.48 g/L group, with a rat absorbing 2.6

- fold greater percentage of the applied dose compared to human skin. The differences in absorption rates were greater in the 0.024 g/L (3.8

- fold) and 240 g/L (5

- fold) groups.

Although this study does not satisfy EPA guideline requirements, this dermal penetration study was done according to O ECD 428 guidelines. This study fully reliable (acceptable/non

guideline).

IIIA 7.6.3 Dermal Absorption, In Vivo in the Rat; 5 groups of 4 male rats were dosed with radiolabeled sulfoxaflor as GF

- 2032 in a dermal penetration study (MRID 48732413). GF

- 2032 is a suspension concentrate (SC) formulation containing the active substance sulfoxaflor. The nominal concentration of the active substance in

GF

- 2032 is 240 g/L. The highest in

- use spray dilution concentration is 0.48 g sulfoxaflor/L.

The low est in

- use spray dilution concentration is 0.024 g sulfoxaflor/L. Rats were dosed with 0.024, 0.48, or 240 g/L sulfoxaflor as GF

- 2032. [ 14 C]

- Sulfoxaflor was applied to 10 cm 2 patch of shaved skin for 10 hours, and then washed off with a mild soap

solution . Groups of rats were sacrificed at 24, 48, 96, 144, and 192 hours after dermal application and the extent of dermal penetration was determined. The amount of applied dose removed by washing the skin was 96%, 71% and 67% of the applied dose across the do se groups, respectively.

Following a dermal exposure period of 10 h to sulfoxaflor formulation concentrate (240 g/L), 93

- 99% of the applied radioactivity was readily removed from the skin surface by a mild detergent wash. Approximately 1

- 2% of the applied dose was absorbed over the exposure interval andSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 127 of 176 subsequent post exposure collection periods up to 192 h after exposu re. The highest absorption was observed within the 48 hour group with 1.7%.

After a 10 h dermal exposure to the 0.48g/L spray dilution, the dosed material was not as readily

removed from the skin surface with 62

- 79% of the applied radioactivity removed b y a mild detergent wash. Approximately 2

- 3% of the applied radioactivity was absorbed after 24 h and 48 h, increasing to 8% at 96 h. By 144 h and 192 h the absorption levels were similar at 11%, indicating that absorption of the dose was essentially comp lete.

After a 10 hour dermal exposure to the 0.024g/L spray, the amount of dislodgeable dose was similar to the 0.48g/L spray dilution with 3

- 71% of the applied radioactivity removed from the skin surface by the detergent wash. Approximately 1

- 3% of the applied radioactivity was absorbed after 24 h and 48 h increasing to 6% at 96 h. By 144 h and 192 h the absorption levels were similar at 13% and 11%, respectively, indicating that absorption of the dose was essentially complete.

This study is considered Fully Reliable (acceptable/guideline) and fulfills the requirements (OPPTS 870.7600; OECD 427) for a dermal penetration study in the rat.

A.4.9 Immunotoxicity

870.7800 Immunotoxicity (See 870.3100 (p.85))

A.4.9 Special/Other Studies

IIA 5. 6.12/3 Characterization of the agonist effects of XDE

- 208 on mammalian muscle nicotinic acetylcholine receptors ; Sulfoxaflor is a compound with insecticidal activity that acts as an agonist of insect nicotinic acetylcholine receptors (nAChRs). The aim of the wor k described in this report was to examine the influence of sulfoxaflor on mammalian muscle nAChRs. Competition radioligand binding was used to examine the ability of sulfoxaflor to bind to nAChRs from three mammalian species (human, rabbit and rat) (MRID 47832035). In addition, two

- electrode voltage

- clamp recording was used (with human and rat nAChRs) to examine whether binding of sulfoxaflor resulted in functional activation of muscle nAChRs. Radioligand binding experiments demonstrated that

sulfoxaflor binds to human, rabbit and rat fetal muscle nAChRs. Electrophysiological studies revealed that sulfoxaflor is a partial agonist of the rat fetal muscle nAChR. In contrast, sulfoxaflor has no detectable agonist activity on the human fetal muscle nAChR or on the adult muscle nAChR (from either human or rat). In contrast to the clear agonist activity of sulfoxaflor on the rat fetal muscle nAChR, no agonist activity was observed with X11719474, a soil

metabolite of sulfoxaflor.

This study demonstrates that sulfoxaflor is an agonist of the rat fetal muscle nAChR. In contrast, sulfoxaflor has no agonist activity on the equivalent human fetal nAChR or on the rat or human adult muscle nAChR. From these findings, it seems reasonable to conclude that the select ive agonist activity of sulfoxaflor is due to species and age

- dependent differences in the amino acidSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 128 of 176 sequences of the nAChR subunits. Furthermore, the soil metabolite, X11719474, had no agonistic activity on either the rat or human nAChR subtypes.

This study is reliable with restrictions (acceptable/non

- guideline).

IIA 5.6.12/4 Investigation of the critical window of exposure for fetal abnormalities and neonatal survival effects in Crl:CD(SD) rats;

In a non

- guideline reproductive/developmental t oxicity study (MRID 47832137) sulfoxaflor (95.6% a.i. wt/wt; Lot # E2162

- 34, TSN003725

- 0001 ) was administered to pregnant female Crl:CD(SD) rats (12/group) at concentrations of 0 (Group 1, control) or 1000 ppm (Groups 2 and 3) in the diet (corresponding to doses of 0, 38.6, and 76.5 mg/kg/day for Groups 1, 2, and 3, respectively). Group 2 was fed the test substance from gestation day (GD) 6

- 16 to cover all of embryogenesis up to, but not including, the start of early fetal movements; Group 3 was fed the

te st substance from GD 16

- birth to cover development of the muscle nAChR and its role in development of synchronized fetal limb movements up to onset of parturition.

Treatment

- related decreases in body weight were observed in dams in Group 1, resulting in decreased body weight gain from GD 6

- 16; similarly Group 2 dams had treatment

- related decreases in mean body weight and body weight gain during the test substance treatment interval. The body weight effects were accompanied by lower feed consumption (23.3 % and 22.6% lower than controls during the treatment interval for Groups 2 and 3, respectively). No gross anatomical abnormalities were observed in any dams in any treatment group.

There were no treatment

- related differences in the number of pups born al ive or dead in Groups 2 or 3 when compared to the control. There was a treatment

- related 18.8

- 20.8% decrease in Group 3 mean pup body weight in both sexes (alpha=0.05)

Offspring from animals in Group 2 did not display previously described fetal abnorma lities or reduced neonatal survival. Offspring in Group 3 had the same gross effects of limb contractures and reduced neonatal survival (alpha=0.05) seen in previous studies that had treatment with 1000 ppm sulfoxaflor throughout gestation.

Daily examin ation of Group 3 offspring born with limb abnormalities indicated that these were fully reversible shortly after withdrawal of maternal dietary exposure to sulfoxaflor

. In some cases, full reversal of the limb abnormalities was evident the day after birth and occurred for all affected animals that survived to postnatal day (PND) 4; reversal also occurred in some animals that subsequently died before PND 4. The visceral and skeletal findings of abnormal ureter and bent clavicle, the latter of which had a h igh incidence (30.1% of fetuses) in a previous definitive developmental toxicity study, were not present in this study at necropsy on PND 4 despite similar maternal sulfoxaflor blood concentrations and pup limb abnormality indices between these two studies

.

This study demonstrates that the critical period of developmental susceptibility to

sulfoxaflor

- induced fetal abnormalities ( forelimb flexure and hindlimb rotation) and

reduced neonatal survival is between GD 16

- birth, and that all of the fetal abnormalities are rapidly reversible after birth.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 129 of 176 This study is Fully Reliable (acceptable/non

- guideline).

IIA 5.6.12/5 Investigation of the critical window of exposure for fetal abnormalities and neonatal survival effects in Crl:CD(SD) rats (Phase 2);

In a non

- guideline reproductive/developmental toxicity study (MRID 47832136) sulfoxaflor (95.6% a.i. wt/wt; Lot # E2162

- 34, TSN003725

- 0001 ) was administered in the diet to pregnant female Crl:CD(SD) rats (10/group) at concentrations of 0 ppm (Group 1, cont rol) or 1000 ppm on gestation days (GD) 16

- 18 (Group 2), GD 18

- 20 (Group 3), or GD 20

- 22/LD 0 (Group 4), corresponding to mean intakes of 0, 63.9 , 42.5, and 35.7 mg/kg/day for Groups 1, 2, 3, and 4 respectively.

Dams in all sulfoxaflor treatment groups (Groups 2

- 4) had treatment

- related decreases in body weight gain during their respective treatment intervals. These body weight effects were attributed to lower feed consumption, consistent with decreased palatability of test material

fortified diet obser ved in previous studies at 1000 ppm. No treatment

- related clinical signs were observed in any dams in any test substance treatment group.

There were no sulfoxaflor treatment

- related effects on gestation survival or sex ratios of the pups. There were no apparent treatment

- related differences in the number of pups born alive or dead, pup body weight, or litter size.

Offspring from animals given 1000 ppm sulfoxaflor for 48 hours starting on the morning of GD 16 or 18 (Groups 2 and 3) were similar to contr ols and did not display previously described fetal abnormalities or reduced neonatal survival. Offspring given 1000 ppm sulfoxaflor for 48 hours starting on the morning of GD 20 (Group 4) had fetal limb abnormalities (forelimb flexure and

hindlimb rotatio n) as well as reduced neonatal survival (89.6% survival), demonstrating that exposure shortly before birth (GD 21 or 22) is sufficient to induce developmental toxicity.

In addition, daily examination of surviving Group 4 offspring born with limb abnormali ties indicated that these were fully reversible in surviving offspring shortly after withdrawal of maternal dietary
exposure to sulfoxaflor . In some cases, full reversal of the limb abnormalities was evident the day

after birth and occurred for all affect ed animals that survived to postnatal day (PND) 4; reversal also occurred in some animals that subsequently died before PND 4. Likewise, the visceral and skeletal findings of abnormal ureter and bent clavicle, the latter of which had a high incidence (30. 1% of fetuses), in the definitive developmental toxicity study were not present in this study at necropsy on

PND 4.

This study demonstrated that sulfoxaflor exposure between GD20

- GD22/LD0 is sufficient to cause fetal abnormalities (forelimb flexure and hindlimb rotation) and reduced neonatal survival. Furthermore, these fetal abnormalities are reversible by PND 4.

This study is F ully R eliable (acceptable/non

- guideline)

.

IIA 5. 6.12/6 Observations on the effects of XDE

- 208 on the phrenic nerve

- hemidiaphragm preparation from new

- born rat;Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 130 of 1 76 Sulfoxaflor, a compound targeted to the insect nicotinic acetylcholine receptor (nAChR) has been shown to cause foetal limb contractions and reduced neonatal survival in rats following dietary exposure during gestation. The aim of the work described in this report was to make a qualitative investigation of the action of sulf oxaflor on isolated phrenic nerve

- hemidiaphragm preparations from new

- born rats (MRID 47832064)

.

Sulfoxaflor consistently (n=5) produced a reversible, concentration

- dependent contracture of the diaphragm that was blocked by the selective muscle

- type nAC hR antagonist, tubocurarine (10 μM), showing that the contracture induced by sulfoxaflor is mediated via nAChR activation, rather than via a post

- receptor mechanism. Furthermore, prolonged application of sulfoxaflor

caused a sustained muscle contracture. Muscle twitches in response to phrenic nerve stimulation were not affected at low sulfoxaflor concentration (100 μM) but were reduced at high concentration (1 mM), demonstrating that sulfoxaflor can cause inhibition of nerve

- evoked contraction of the diap hragm during sustained contracture, consistent with the observed impairment of respiration in the neonatal rat.

The results of these experiments demonstrate that sulfoxaflor caused a contracture of the new

born rat diaphragm by acting on the nAChR. Pro longed application caused a sustained muscle

contracture and a contracture

- associated inhibition of the phrenic nerve

- evoked muscle twitch, which is considered analogous to the situation in vivo which resulted in foetal limb contractions (sustained muscle contractions) and compromised respiration at birth (contracture

- associated inhibition of the muscle twitch).

Therefore, the results described in this report are entirely consistent with, and add additional support to, the hypothesis that sulfoxaflor cau ses neonatal death (and foetal abnormalities) via

activation of the foetal muscle

- type nAChR.

This study is Reliable with Restrictions (acceptable/non

- guideline).

IIA 5. 6.12/7 Histopathological Evaluation Of Fetal Lung Samples From The Developmental To xicity Study In Crl:Cd(Sd) Rats; Dietary administration of 1000 ppm sulfoxaflor to Crl:CD(SD) rats during gestation has been previously shown to cause neonatal pup death. In order to determine if morphological alterations (e.g., increased collagen deposit ion) in any region of the lungs were responsible for pup death, one fetus/sex from five control and four 1000 ppm litters (18 samples total) from the definitive developmental toxicity study were collected and preserved in neutral, phosphate buffered 10%

fo rmalin (MRID 47832135). Sections from these preserved tissues were processed such that each slide contained sections of the trachea, bronchi, bronchioles, and alveoli. Slides were stained with hematoxylin and eosin and evaluated for histopathological cha nges. Tissues were archived with the developmental toxicity study.

There were no sulfoxaflor induced lesions in the trachea, bronchi, bronchioles and alveoli in any

of the treated fetuses examined. There were no treatment

- related increases in collagen deposition around the airways or alveolar walls or any other changes. All observations were

considered within normal limits.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 131 of 176 This non

- guideline study is reliable with restrictions (acceptable/non

- guideline). The study is not GLP compliant. However, all experiments were done according to GLP standards. Additionally, the study was well conducted and the Agency has confidence in the scientific integrity of the data.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 132 of 176 Appendix B . Sulfoxaflor Mode

- of Action for Reproductive and Developmental Effects

The following Mode

- of

- Action was presented to an HED committee for comment on its acceptability and plausibility on Dec. 7, 2011. The general consensus of the committee was that the data supported the overall MOA in rats but

t he MOA was applicable to huma ns

.

Sulfoxaflor is a member of a new class of insecticides, the sufoximines, with proposed uses on cotton, soybeans, cereals, citrus, leafy and fruiting vegetables, cole crops, grapes, apples, and a variety of other crops. It provides control for a broad spectrum of piercing/sucking insects. Sulfoxaflor is a nicotinic receptor agonist, similar to neonicotinoid insecticides such as acetamiprid, imidacloprid, and thiacloprid. However, unlike neonicotinoids, sulfoxaflor is not susceptible to in vivo metabo lism by monooxygenase enzymes, which causes virtually all known

cases of resistance in the field to this class of insecticides.

Toxicity and mechanistic studies in rats, rabbits, dogs and mice highlight the liver and the
nervous systems as target organ sy stems. Liver effects in subchronic and chronic studies include organ weight and enzyme changes, hypertrophy, proliferation, and tumors. Neurological effects are manifested as limb flexure, bent clavicle and convoluted/hydroureter in rat fetuses and

decre ased neonatal survival. These effects were not observed at similar maternal and fetal systemic exposure levels in NZW rabbits. Finally, long term studies produced treatment related tumors in the rat and mouse including liver, Leydig, and preputial gland tumors.

In rats, sulfoxaflor exposure during gestation caused effects on neonatal survival and fetal abnormalities, but neither effect occurred in rabbits. Similar maternal and fetal blood data showed that the interspecies difference between rats and ra bbits was not due to toxicokinetics; therefore, the species difference is likely due to toxicodynamics.

Since the insecticidal M O A is based on agonism of the nicotinic acetylcholine receptor (nAChR), it is likely that the developmental effects were also r elated to nAChR binding. There are two types of mammalian nAChRs, neuronal and muscular. The neuronal nAChRs are principally located in the central and peripheral nervous system and disregulation is often manifested as the modification of biological setp oints such as tachycardia, blood pressure, inflammation, cognitive learning, arousal motor control and analgeisa. The muscle nAChRs are found in the intramuscular junctions of skeletal muscles. Disregulation of these receptors can

result in muscle contra ction, difficulties in breathing, and ultimately death. Furthermore, there are two subtypes of muscle nAChRs, a fetal form and an adult form. The switch from fetal to adult occurs shortly before birth in humans and shortly after birth in rats. Based upo n the types of skeletal effects observed and their appearance during early development, the registrant has proposed a MoA involving the interaction of sulfoxaflor with the fetal musc le nAChRs.

Postulated MOA for sulfoxaflor

- induced muscle contracture an d death in neonatal offspring

Given that sulfoxaflor targets the insect nAChR, and that functional expression of the fetal

- type neuromuscular junction nAChR occurs in late gestation and is involved with limb muscle function, it was hypothesized that the neonatal offspring abnormalities and deaths occur via a single MoA: sulfoxaflor’s sustained agonism at the fetal

- type muscle nAChR andSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 133 of 176 subsequent sustained muscle contracture of the limb, shoulder girdle and diaphragm, respectively (MRID 48288940).

A sin gle M O A is considered to be responsible for the neonatal offspring findings since the

sulfoxaflor

- induced limb contracture, clavicle abnormalities, and neonatal deaths occurred at the same dose with similar incidences (Figure 1). In order for a single mod e of action to be plausible, the findings should have a similar incidence across doses from all studies. Based upon

similar dose

- response curves for both effects (i.e., limb abnormalities and neonatal deaths), the data from multiple studies have a high de gree of correlation (R 2 values of 0.93 and 0.91), supporting a single M O A with different apical end points rather than a different M O A for each of the two major effects in rats.

Figure 1. Dose

- response curves with non

- linear regression analyses for the percent of neonatal offspring affected by death and/or limb contracture in rats. Study type abbreviations: 1 st gen and 2 nd gen, 1 st and 2 nd generations of the two

- generation reproductive toxicity study; 421

- like, reproductive toxicity screening study; X

- foster, cross

foster study; CW 1, critical window study phase 1; CW 2, critical window study phase 2; DNT, developmental neurotoxicity study; DT, prenatal developmental toxicity study.

This M O A is proposed to progress through the following key events: (1) sulfoxaflor binding and (2) agonism at the fetal

- type muscle nAChR, resulting in (3) sustained agonism/sustained muscle

contracture in neona tal offspring (fetus and pup). This sustained muscle contracture results in limb contracture, bent clavicles, and abnormal neonatal respiration after birth with ensuing

neonatal deaths.

The sequence of events in sulfoxaflor’s proposed MOA for muscle co ntracture and neonatal pup

death:

Specific binding to the fetal

- type muscle nAChR (Key Event #1)Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 134 of 176 Agonism at the fetal

- type muscle nAChR ( Key Event #2)

Sustained agonism/sustained muscle contracture (Key Event #3)

Key Event #1: Specific binding to the fet al

- type muscle nAChR

Competition radioligand binding was used to examine the ability of sulfoxaflor to bind nAChRs from three mammalian species (human, rabbit and rat). Initial radioligand binding studies were performed with [ 3 H]

- sulfoxaflor, the aim bein g to examine whether sulfoxaflor is able to bind to mammalian muscle nAChRs. However, technical difficulties were encountered in performing binding experiments with [ 3 H]

- sulfoxaflor that were associated with high levels of non

- specific binding. Therefore , a series of experiments was performed to examine whether unlabelled sulfoxaflor was able to displace binding of the high

- affinity nAChR radioligand [ 3 H]

- epibatidine from tissue homogenates prepared from rabbit fetal forelimb muscle, rat fetal forelimb mu scle and human fetal recombinant nAChRs expressed in HEK (human embryonic kidney) cells (MRID 47832035).

Dose

- dependent displacement of [ 3 H]

- epibatidine binding was observed with increasing concentrations of sulfoxaflor for all three preparations examine d (human, rabbit and rat). At higher concentrations of sulfoxaflor, almost complete displacement of bound [ 3 H]

- epibatidine was observed (Fig. 2). Although data obtained with human nAChRs and rabbit muscle are well fitted by the single

- site model (solid l ines in Fig. 2 ), the binding data from rat muscle were fitted

with a two

- site model (dotted line in Fig. 2 ) that revealed two distinct binding sites of different affinities (0.01 mM and 8.9 mM). The ability of sulfoxaflor to bind either a single (human an d rabbit) or multiple (rat) sites on the nAChR is ambiguous. However, demonstration of the capacity of sulfoxaflor to bind mammalian nAChRs and potentially affect the functionality of the receptors is of great significance.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 135 of 176 Figure 2. Sulfoxaflor displacement of [ 3 H]

- epibatidine in (A) rabbit fetal muscle tissue, (B) rat fetal muscle tissue demonstrateing the 1

- site and 2

- site binding models, and (C) and in HEK

- 293 cells expressing recombinant human fetal nAChRs

Table 1. XDE

- 208: Dose Response for MoA Key Event #1: Fetal

- type Muscle nAChR Binding

Media Conc. (μM) Key Event 1 Fetal

type Muscle

#

0.3 μM 5 +

3 μM 0

10 μM 0

30 μM 21

100 μM 31

300 μM 31

1000 μM 37

3000 μM 51 DoseSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 136 of 176 10000 μM 100

# Percent inhibition of maximum epibatidine binding

+ Within background of system and not considered biologically

Key Event #2: Agonism at the fetal

- type muscle nAChR

The ability of sulfoxaflor to act as an agonist of fetal and adult muscle nAChRs was examined by expression of rat and human nAChRs in Xenopus oocytes (MRID 47832035)

. In mammalian muscle cells, nAChRs are expressed at the neuromuscular junction (NMJ) and are composed of five nAChR subunits (α1, β1, γ, δ and ε). Transcription of the γ and ε subunits is differentially regulated during development, with the γ subunit expressed in "fetal" muscle and the ε subunit expressed in "adult" muscle ( Mishina et al., 1986 ) (Figure 3).

Figure 3. Subunit composition of the fetal

- type and adult

- type muscle nAChR.

Expression of the fetal

- and adult

- type nAChRs was achieved by microinjection of cDNA or cRNA encoding the appropriate rat or human muscle subunits in Xenopus oocytes. To generate the fetal form of the muscle nAChRs, α1, β1, γ and δ cDNAs or cRNAs were injected, whereas α1, β1, δ and ε were injected to generate th e adult form. In each case, functional responses (i.e, membrane currents) were detected in response to application of the endogenous agonist acetylcholine (ACh). There have been no reports of the molecular cloning of nAChR subunits

from rabbit and so rab bit muscle nAChRs were not examined in this manner. Two

- electrode voltage

- clamp recording from Xenopus oocytes was performed 1

- 5 days after micro

- injection

with cDNA or cRNA.

Clear agonist

- evoked responses were observed with sulfoxaflor (XDE

- 208) at the rat fetal

- type nAChR (Fig. 4). Dose

- dependent agonism at the fetal

- type muscle nAChR was demonstrated; the lowest tested concentration having no agonism while incubation of the oocytes with higher concentrations of sulfoxaflor showed increasing agonism. There was no apparent agonism in either the adult rat or human (fetal and adult) type receptors (Table 2 ).

Fetal

- Type nAChR Adult

- Type nAChRSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 137 of 176 Figure 4. Representative whole

- cell current responses in a Xenopus oocyte cell expressing

fetal

- type rat (α1) 2 β1γδ nAChRs. Inward currents are show from the same oocyte in response to application of acetylcholine (100 μM) and XDE

- 208 (3 mM). The length of agonist application (5 secs.) is indicated by the horizontal bar . Agonist activation is associated with downward deflection in the trace. Note, there was recovery of response after ACh or sulfoxaflor were washed off ( Millar, 2010 ).

Table 2 . Agonist

- activation of nAChRs expressed in Xenopus oocytes by sulfoxaflor (3mM). Maximum response (% + SEM; normalized to max. ACh response)

Rat fetal muscle 39 ± 2.4%

Rat adult muscle No response

Human fetal muscle No response

Human adult muscle No responseSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 138 of 176 Figure 5. Agonist activation of nAChRs expressed in Xenopus oocytes. Data are shown for the rat fetal (α1) 2 β1γδ nAChR (A) and rat adult (α1) 2 β1δε nAChR (B). AChRs were expressed by microinjection of cRNA in Xenopus oocytes. Dose

- response curves are shown in which ag onist

- evoked responses are normalized to the maximal response detected with the endogenous agonist, acetylcholine (ACh). Note that no receptor desensitization occurs, which would have been seen as a diminished agonist response at higher doses. Data point s are means of 3

- 7 responses. Adapted from Millar, 2010

.

Table 3 . Sulfoxaflor

- induced agonism of rat Fetal

- type muscle nAChRs expressed in Xenopus Oocytes

Media Conc. (μM sulfoxaflor) % Response ( normalized to

maximum. ACh response) 10 μM 0 30 μM 1.0 100 μM 5.1 300 μM 28 1000 μM 71 3000 μM 100 10000 μM Not Determined Sulfoxaflor Agonism at the Adult

- type Muscle nAChR 100 to 3000 μM 0

Key Event #3: Sustained agonism at the fetal

- type muscle nAChR and sustained muscle

contracture

Sulfoxaflor readily perfuses into muscle from the blood (MRID 47832039) and therefore muscle sulfoxaflor concentrations would also be maintained at effective levels as long as dietarySulfoxaflor Human Health Risk Assessment DP No. D382604

Page 139 of 176 treatment continued. At the neuromuscular junction, the neurotransmitter ACh binds to Ach receptors on muscle fibers causing contraction. ACh undergoes tightly regulated release followed by rapid hydrolysis by acetylcholinesterase (AChE), unlike sulfoxaflor which would remain at the nAChR synaptic cleft d ue to its lack of hydrolysis by AChE. Thus, upon fetal

- type muscle nAChR activation, a sulfoxaflor

- induced muscle contracture would be sustained for as long as sufficient sulfoxaflor molecules remain available for receptor binding.

To directly assess mus cle contracture at the neonatal diaphragm, sulfoxaflor was tested for agonist action on the isolated phrenic nerve

- hemidiaphragm preparation (Bulbring, 1946) from

new

- born rats (MRID 47832064). A number of issues were addressed in this study including 1)

Concentration

- dependence of the contracture produced by sulfoxaflor

, 2) Block of the response to sulfoxaflor by tubocurarine , and 3) Response to prolonged sulfoxaflor application . To accomplish this, diaphragm muscle preparations were mounted in a recording chamber and bathed in a physiological salt solution. A myograph wire attached to the muscle tendon was connected to a Harvard isometric strain gauge transducer to record musc le tension. The phrenic nerve was stimulated using a bipolar stimulating electrode placed at the nerve entry point to the muscle with supra

- maximal rectangular voltage pulses of 0.2 ms duration at a frequency of 0.5 Hz. Strain gauge transducer and stimul ating electrode were mounted on micromanipulators in order to allow accurate positioning relative to the muscle. Acetylcholine, tubocurarine, and sulfoxaflor were applied by manually switching taps controlling solutions flowing to the inflow manifold of th e recording chamber. Recorded measures include 1) changes in muscle twitch, and 2) muscle contracture following test material application to the bath perfusion system.

1) Concentration

- dependence of the contracture produced by sulfoxaflor

1mM sulfoxafl or consistently produced a contracture of the neonatal diaphragm muscle and a decrease in muscle twitch response. At 100 μM sulfoxaflor, there was little effect on

twitch tension.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 140 of 176 Figure 6. Rat fetal diaphragm tissue demonstrates contraction in the p resence of

sulfoxaflor.

Expanded time scale display of the sulfoxaflor response illustrates the muscle contracture and the accompanying decrease in twitch tension induced by sulfoxaflor. In five preparations, the muscle twitch response decreased to 34 ± 3.2% of control during responses to 1 mM sulfoxaflor.

Figure 7. Expanded time scale demonstrating decreased twitch tension in the presence of

sulfoxaflor.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 141 of 176 2) Block of the response to sulfoxaflor by tubocurarine

,

The response to sulfoxaflor was sho wn to be antagonized by the nicotinic antagonist,

tubocurarine. Pre

- incubation of the muscle tissue with tubocurarine prevents sulfoxaflor

- induced muscle contractions, indicating that sulfoxaflor is acting via the nAChR and not via a post

receptor mechani sm.

Figure 8 . Pre

- application of 10μM tubocurarine (Tubo) effectively blocks the muscle twitches and antagonises responses to 100μM or 1mM XDE

- 208 ( Gibb, 2010 ).Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 142 of 176 3) Response to prolonged sulfoxaflor application During application of a high concentration of sulfoxaflor (1mM), the muscle contracture is sustained, suggesting little desensitization of the muscle nAChRs in response to this agonist. Although the muscle contracture is sustained in the presence of sulfo xaflor, the muscle recovers normal function on removal of sulfoxaflor from the solution bathing the muscle.

Figure 9. Prolonged application (7 minutes) of 1 mM XDE

- 208 shows a sustained contracture by the diaphragm muscle ( Gibb, 2010 ).

Dose Response of sustained muscle contracture (i.e., forelimb flexure and hindlimb rotation)

Increased receptor response with increasing sulfoxaflor exposure has been demonstrated for several key events including receptor binding, receptor agonism, diaphragm muscle contraction,

and fetal effects in vivo.

Table 4. Dose response observations from in vitro mechanistic studies

Effect Sulfoxaflor conc.(μM) Key Event #1 Increased displacement of nAChR

radiolignad [H3]

- epibatidine from 0

to near 100% 0.3

- 1000

Key Event#2 Increased agonism in fetal

- type

nAChR from 0 to 100% 10

- 3000

Key Event #3 Greater relative contraction of fetal

rat diaphragm muscle at 1mM compared to 100 μM sulfoxaflor 100 μM and 1 mMSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 143 of 176 Temporal concordance of sulfoxaflor levels, nAChR fetal

- type muscle receptors, muscle flexures,

and neonatal deaths

Ontogeny of fetal

- type nAChRs In rats, the muscle nAChR develops functional subunit expression at the neuromuscular junction between GD 15 and 17 (Kues et al., 1995) resulting in synchronized fetal limb movements (Robinson and Smotherman, 1988) and diaphragmatic responsiveness between GD 16 and 17 (Bennett and Pettigrew, 1974), the latter being critical for the transition to extrauterine respiration. Replacement of the γ subunit by the ε subunit initiates late during the first postnatal week in rats and is largely complete by the end o f the second postnatal week in limb and respiratory muscles (Missias et al., 1996).

Cross fostering

– Gestation

A cross

- fostering study demonstrated that the effect of sulfoxaflor on pup survival was due to in

utero , not lactational exposure (MRID 4783206 3). Groups of female Crl:CD(SD) rats were fed diets supplying 0 (control) or 1000 ppm sulfoxaflor for two weeks prior to mating through weaning on lactation day (LD) 21. As the control and treated females mated, they were

subdivided into Foster dams and Donor dams. Cesarean

- section was performed on gestation day (GD) 21 Donor dams, at which time, one or more batches of two of their offspring/sex were immediately cross

- fostered to a Foster dam(s) that had their own litter removed that day (i.e., on LD 0). After cross

- fostering was complete, each control and sulfoxaflor

- treated Foster dam had mixed litters comprised of two pups/sex that originated from control Donor dams (five litters) and two pups/sex that originated from sulfoxaflor

- treated Donor dams (e ight litters).

All offspring from dams exposed to 1000 ppm sulfoxaflor prior to birth died by postnatal day (PND) 4, irrespective of whether they were cross

- fostered to control

- or treated

- foster dams (see results Table 5 below). Consistent with reduce d viability, some offspring were cold to the touch, had bluish skin, autolyzed and cannibalized, and stomach void of milk. Conversely, there was no effect on neonatal survival for pups exposed to sulfoxaflor only after birth. Furthermore, PND 1 pup body weights were significantly decreased in prenatally exposed offspring.

Table 5 . Cross Foster or Treated Foster Dams Results

Foster

Dams Donor

Pups Hypotheses for Pup Survival Outcome

Control Control in utero No effect expected (negative control) No effect

Treated in utero If pups die, effect comes from treated pups (i.e. in utero effect ) All pups died By PND4

sulfoxaflor 1000 ppm Control in utero If pups die, effect comes from treated

dams (i.e. lactational effect ) No effect

Treated in utero Pups death expected (positive control) All pups died By PND4

Critical window studies

– late gestation effectSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 144 of 176 The cross fostering study clearly indicated that sulfoxaflor

– induced deaths were dependent upon

in utero exposure. In order to appropriately frame the incidence data presented in this mode

- of

action experiment, the findings presented here were compared to the guideline definitive developmental toxicity study in Crl:CD(SD) rats and the first critical window study. Treatment

related findings in the offspring were found at 1000 ppm in the developmental toxicity study (sulfoxaflor exposure GD 6

- birth; MRID 47832140), Group 3 in critical window 1 (sulfoxaflor exposure GD 16

- birth; MRID 47832137) and in Group 4 i n the critical window 2 study (sulfoxaflor exposure GD 20

- 22/birth; MRID 47832136). See Table 6. When exposure was

limited to between GD 6

- 20, regardless of dose, they were no treatment

- related offspring effects. However, when exposure spanned the GD 20

- birth, and were sufficiently high (i.e., 1000 ppm), offspring treatment related effects were observed.

Table 6. Comparison to Developmental Toxicity Study

Developmental Toxicity Study(MRID 47832140) Critical Window 1 (MRID 47832137) Critical Window 2 (MRID 47832136)

Group: Group 2 Group

3 Group 4 Group 2 Group 3 Group 2 Group 3 Group 4

Dose (ppm): 25 150 1000 1000 1000 1000 1000 1000

Treatment period: GD 6

- 21 GD 6

- 21 GD 6

- 21 GD 6

- 16 GD 16

- Birth GD 16

-

18 GD 18

- 20 GD 20

-

22/LD 0 Avg.TMI (mg/kg/day) 1.95 11.5 70.2 76.5 38.6 62.9 42.5 35.7

TK plasma

conc. (μg/g) 0.843

± 0.09 4.938

± 0.87 35.245

± 5.43 35.4

–

40.9 32.1

- 43.2 16.4

-

33.3 23.0

-

30.2 5.41

- 16.1 a

Offspring

Effects NO NO YES NO YES NO NO YES

GD gestation day

a Three of the four sampled rats had undergone parturition prior to blood collection

Reversibility when sulfoxaflor removed from diet

The incidence of limb abnormalities decreased over the first two

- to

- four days of postnatal life. While a portion of the decreased incidence was due to pup deaths, reversibility of the limb abnormalities was directly demonstrated in the two critical window studies as evidenced by a 0% incidence of forelimb flexure and hindlimb rotation by PND 2 or 4 (Table 7). Reversal of the bent clavicles was indirectly demonstrated by an absence of bent clavicles in PND 4 pups (of 86 pups evaluated in the affected groups), which is significant as these findings were observed at a relatively high incidence (30 .1%) in GD 21 fetuses from dams exposed to 1000 ppm sulfoxaflor in the developmental toxicity study. These reversibility data suggest that the limb abnormalities were transient, consistent with a pharmacologic MoA for sulfoxaflor.

Table 7. Reversibilit y of limb abnormalities in aging pups.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 145 of 176 Critical Window 1 (MRID 47832137) Critical Window 2 (MRID 47832136) Treatment period GD 16

- birth GD 20

- 22/LD0

PND 0 to 1 11 5

PND 1 to 2 6 2

PND 2 to 3 3 0

PND 3 to 4 1 0

Total reversals 21 7

Furthermore, the timing of the neonatal deaths indicates a distinct change in the pharmacodynamic activity of sulfoxaflor after PND 4. In the 2

- generation reproduction study (MRID 47832142), pup deaths were slightly, but significantly, greater in the high dose group (400 ppm) compared to controls; 95.4% v. 97.2% for the F1 and 93.0% v. 98.5% in the F2 generation prior to PND 4. However, survivability exceeded 99% for both generations after PND 4, despite the continued exposure of the pups to sulfoxaflor t hrough LD 21.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 146 of 176 Table 8. Concordance and dose response of offspring effects observed following treatment with sulfoxaflor

Internal

Dose (μM) Applied

Dose (ppm) Exposure

Days KE #1

Fetal

- type
Muscle

nAChR Binding # KE #2

Fetal

- type
Muscle

nAChR Agonism $ KE #3

Sustained Agonism & Sustained Muscle

Contracture Neonatal Apical Endpoints Forelimb Flexure, Hindlimb Rotation, Bent Clavicle, Death & Neonatal Diaphragm $

(Fetal

- type Muscle nAChR) Reversible? Neonatal Offspring

Limb

Contracture ^ Reversible?

2.3 25 1 GD 6

- 21

- -

4.0 25 2 Premating

LD 21 - -

10 μM in vitro 4

- - - 1 5 150 1 GD 6

- 21

- -

16 100 2 Premating

LD 21 - -

ND 100 3 X

- - 30 μM in vitro 4 + +

57 400 2 Premating

LD 21 - +

ND 500 3 ND + +

60 1000 6 + Y +

89 1000 7 ND + +++

100 μM in vitro 4 ++ +
100 μM ex vivo 8 + Y

101 1000 3 X ND + +++
108 1000 1 GD 6

- 21 +++ +++
136 1000 5 ++ Y +++
300 μM in vitro 4 ++ ++
1000 μM in vitro 4 +++ +++
1000 μM ex vivo 8 +++ Y

3000 μM in vitro 4 +++ +++

1 Rat Developmental Toxicity, 2 Two

- Generation Reproduction, 3 One

- Generation Reproduction, 4 Xenopus Oocytes

- Recombinant Fetal

- type Muscle nAChR, 5 Critical Phase 1 Mode of Action, 6 Critical Phase 2 Mode of Action, 7 Cross Foster, 8 Neonatal Diaphragm Ex Vivo Electrophysiology

4$ Dose

- response curve evaluated for muscle nAChR agonism (100

- 3000 μM). 

### DoseSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 147 of 176 4* Full dose

- response data for fetal

- type muscle nAChR binding and agonism showed in Table 3.

9 Highest dose level from the One

- Generation Reproduction and Rat Developmental Toxicity studies.

# Percent inhibition of epibatidine binding: +, 0

- 25%; ++, 25

- 50%; +++, 50

- 100%

$ Percent of maximum XDE

- 208 response: +, 0

- 25%; ++, 25

- 50%; +++, 50

- 100%

^ Incidence (fetal/pup basis): +, 0

- 25%; ++, 25

- 50%; +++, 50

- 100%

& Maximum incidence (fetal/pup basis): +, 0

- 25%; ++, 25

- 50%; +++, 50

- 100 % ND, No Data

ND + , No data as early studies did not examine this parameter
Y, YesSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 148 of 176 Biological plausibility and coherence

The postulated MoA for the neonatal deaths in rats is considered biologically plausible and

coherent. The observed sulfoxaflor

- induced limb and shoulder girdle contracture abnormalities and neonatal death are biologically consistent with the functional ontogeny of the fetal

- type
muscle nAChR in the rat.

Alternative Modes of Action

The registrant has proposed a number of alternative MoAs, including consideration of

sulfoxaflor

- induced limb abnormalities and neonatal offspring deaths due to: agonism at other AChR types (adult

- type muscle nAChR, neuronal nAChR, muscarinic AChR), action

downstream of the fetal

- type muscle nAChR, antagonism at the fetal

- type muscle nAChR, maternally

- mediated fetal immobilization, inhibition of acetylcholinesterase and inhibition of angiotensin

- converting enzyme. A more detailed examination of each of the alternative MoAs

can be found in Mo de of Action Evaluation and Human Relevance Framework Analysis for

XDE

- 208

- Induced Fetal Abnormalities and Neonatal Death in Rats (MRID 48288940).

a) Agonism at other Acetylcholine Receptor Types (Adult

- type muscle nAChR, Neuronal nAChRs, Muscarinic AChR)

Adult

- type nAChRs

In an in vitro study (MRID 47832035) in which recombinant adult

- type nAChR were expressed in Xenopus oocytes, sulfoxaflor was found to bind to the receptor, however no functional agonism was demonstrated up to sulfoxaflor’s limit of sol ubility. Furthermore, in in vivo studies in adult rats or rats older than PND 4 in the two generation reproduction study, there was no evidence of nACHr related toxicity such as muscle fasciculation or limb contractions. Rather, continued dietary exposur e to sulfoxaflor in older animals resulted in hepatic toxicity along decreased body weight and body weight gains.

Neuronal nAChRs Although the interaction of sulfoxaflor with neuronal nAChRs, as opposed to the proposed skeletal muscle nAChRs is plausible, the absence of historic neuronal nicotinic manifestations such as muscle weakness or flaccidity makes this MoA unlikely. In contrast, sulfoxaflor

- induced limb contractures result in rigid, contracted limbs. Breathing movements are also known to be parti ally regulated by neuronal nAChRs in the brainstem. Although, prenatal nicotine exposure has been demonstrated to alter fetal lung development in rats ( Maritz and van Wyk, 1997 ), it does not cause death in normoxic conditions ( Geller, 1959; Sobrian et al., 1995 ). Increased deaths in neonatal rats following exposed to sulfoxaflor and normal histopathology of GD 21 fetal lung tissue (MRID 47832135) decrease the plausibility of this MoA.

Muscarinic ACh Rs Agonism at muscarinic AChRs has also been explored as an alternative MoA. The most plausible target by which a compound could cause death in neonatal offspring would be via muscarinic activity would be the heart or lung. However, activity at these rece ptors would be accompanied by systemic clinical signs of mAChR activity such as salivation, lacrimation, andSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 149 of 176 convulsions, none of which were observed in the Guideline or MoA studies. Hearts were grossly normal on GD 21 visceral examination (including inte rnal structures) in the prenatal developmental study in rats, and there were no adverse histological findings in the lungs.

b) Agonsim downstream of the fetal

- type muscle nAChR Downstream agonism of diaphragm muscle has been demonstrated with the herbici de cartap, causing respiratory failure and irreversible contracture of adult

- type diaphragm preparations. Rather than acting at the muscle nAChR directly, the diaphragm contracture is via post

- receptor induction of extracellular Ca2+ influx, release of in ternal Ca2+, and an inhibition of [3H]ryanodine binding to the Ca2+ release channel of sarcoplasmic reticulum (Liao et al. , 2000). In the phrenic

- nerve hemidiaphragm experiments conducted with sulfoxaflor where sustained

muscle contracture was demonstrate d, a post

- nAChR MoA would have been manifested by continued contraction in the presence of co

- application with the nAChR antagonist tubocurarine.

The sulfoxaflor

- induced diaphragm contracture was completely eliminated with co

- incubation with tubocurarine, thereby discounting a post

- receptor MoA in the induction of the developmental effects of sulfoxaflor.

c) Fetal

- type muscle nAChR inactivation Nongenetic, pharmacologic antagonism of the fetal

- type muscle nAChR also results in neonatal offspring death and limb abnormalities. A historical example of this includes the use of tubocurarine. Exposure to tubocurarine, a non

- depolarizing neuromuscular blocking agent, during the last week of gestation in rats causes direct fetal paralysis capable of causing fore limb contracture, hindlimb rotation and death (Shoro, 1977). While this may appear similar to the effects seen with sulfoxaflor there are several important distinctions. First of all, neuromuscular blocking agents do not readily cross the placenta (Evans and Waud, 1973). Secondly, the limb contracture effects result in skeletal alterations in the limb cartilage, something that is not observed with sulfoxaflor. Finally, fetal paralysis additionally results in pulmonary hypoplasia, which has previously bee n mentioned to not occur following sulfoxaflor exposure.

d) Acetylcholinesterase Inhibition Following sublethal exposures in rats, cholinergic signs of AChE toxicity are generally observed as clinical signs and in the absence of external, visceral, or ske letal abnormalities in fetuses. Therefore AchE inhibition does not fit the profile observed following sulfoxaflor exposure.

e) Angiotensin

- converting enzyme (ACE) inhibition Angiotensin

- converting enzyme (ACE) inhibition presents another MoA that has been demonstrated to result in fetal limb contractures when exposure occurs during fetal development in humans (Buttar, 1997). The limb contactures which occur are secondary to renal failure associated oligohydramniosis. Other secondary fetal anomalies i nclude potentially fatal hypotension, anuria, craniofacial deformities and hypoplastic lung development. Furthermore, rodents are relatively resistant to the teratogenic effects of ACE inhibitors and high doses of ACE inhibitors typically result only in f etal growth retardation and occasionally increased pup

death.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 150 of 176 Table 9 . Summary Evaluation for Other Possible M O As in the Rat

Alternative

MoA Example MoA Characteristics Plaubility/

Coherence Evidence

Adult

- type
NMJ

nAChR agonism Acetyl

choline Muscle fasciculations,

tonic or clonic limb contractions; Fetal

immobilization due to

uterine contraction - Plausibility/ Strong

coherence

across studies XDE

- 208 causes no

agonism at the adult

- type
NMJ in vitro ; no clinical signs in dev. tox. and
repro. to x. studies

Neuronal

nAChR subtype agonism Nicotine,

Epibatidine Clinical signs of

cholinergic stimulation followed by

desensitization (inhibition); Lung hypoplasia - Plausibility/

Strong

coherence

across studies No indication of

representative clinical signs in adults or offspring; normal lungs (including histopathology) in GD 21

fetuses

Muscarinic

nAChR agonism Muscarine,

Carbamyl

choline Clinical signs of

cholinergic stimulation and/or inhibition; death

caused by action at heart or lungs - Plausibility/ Strong

coherence

across studies No indication of

representative clinical signs in adults or offspring; normal hearts (including internal structures) and lungs (incl. histopathology) in

GD 21 fetuses

Agonism

downstrea

m of the

fetal

- type
NMJ

nAChR Cartap Muscle contracture

associated with

intracellular calcium or ATP regulation; not blocked by NMJ AChR antagonists - Plausibility/

Limited coherence, only evaluated

in ex vivo study Diaphragm contracture

complete ly blocked by

co

- exposure to the NMJ nAChR antagonist, α

tubocurarine

Fetal

- type
NMJ

nAChR

inactivation α

Tubocurari

ne Decreased or

eliminated fetal activity; Limb

contracture characterized by hypotonia; Altered limb skeletal structures; Lung hypoplasia - Plausibility/

Good

coherence across studies; Agents have

limited placental transfer, requiring direct fetal injection XDE

- 208

- induced

decreased activity likely due to respiratory difficulties; Limb tone

and skeletal structures normal; Lungs normal

AChE

inhibition Neostigmin

e Clinical signs of

cholinergic toxicity resulting in maternal - Plausibility/ Strong

coherence No cholinergic clinical

signs in adults or neonatal offspring in dev.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 151 of 176 death; Sublethal exposures generally

without across studies tox . and repro. tox. studies; no maternal

death

ACE

inhibition Captopril,

Enalapril In humans, limb

contractures secondary

to renal failure

associated oligohydramniosis; In rats, fetal growth

retardation and occasionally increased pup death without limb

co ntractures - Plausibility/

Strong

coherence

across studies Lack of similarity in

neonatal abnormalities

and resistance of rodents

to ACE inhibition

induced limb

contractures

f) Conclusion of consideration of alternative M O As There is sufficient information to preclude many of the alternative M O As . However, much of the evidence is indirect and does directly assess the mechanisms. For example, it is not likely that neuronal nicotinic receptors are responsible for the increased pup deaths; since the same isoforms of neuronal nicotinic receptors are present in both juveniles and adults, toxicity would be evident in adults as well as juveniles, which is not the case. Empirically determining the inability of sulfoxaflor to bind an d adulterate the functionality of these neuronal receptors would provide a great deal more confidence in the decision to exclude this M O A

.

Human Relevance DAS has postulated that the proposed MoA is not relevant to humans based upon data demonstrating f

undamental qualitative differences in the agonism of sulfoxaflor at the rat or

human fetal

- type muscle nAChR. Specifically, binding but no agonism was evident with

sulfoxaflor at the human fetal

- type or human adult

- type muscle nAChR (MRID 47832035) (Figure 10.) . Both muscle receptor types were examined as the transition from the fetal

- type to
adult

- type human muscle nAChR occurs prenatally.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 152 of 176 Figure 10

. Agonist activation of nAChRs expressed in Xenopus oocytes. Data are shown for the human fetal (α1 )2β1γδ nAChR (A) and human adult (α1)2β1δε nAChR (B). AChRs were expressed by microinjection of cDNA or cRNA in Xenopus oocytes. Dose

- response curves are shown in which agonist

- evoked responses are normalized to the maximal response detected with the endog enous agonist, acetylcholine (ACh). Data points are means of 3

- 7 responses. Adapted from Millar, 2010.

Table 10

. Concordance of Key Events for Induction of Muscle Contracture and Associated

Death in Neonatal Offspring in Rodents and Humans.

Key Event Evidence in Rodents Evidence in Humans

#1 B inding to the fetal

type muscle nAChR Yes Yes

#2 Agonism at the

fetal

- type muscle

nAChR Yes No

P lausible

, but no agonism

occurs with XDE

- 208 either

at the fetal

- or adult

- type
muscle nAChR

. #3 Sustained fetal

- type
muscle nAChR agonism/sustained

muscle contraction Yes No data in humans. Presumably n ot possible via

this MoA as key event #2

does not occur. Apical Endpoints Forelimb Flexure, Hindlimb Rotation, Bent Clavicle,

Neonatal Death Yes No data in hu mans. Presumably n ot possible via

this MoA as key event #2

does not occur.

The species

- specificity of the effects in the rat is supported by the finding that although sulfoxaflor binds to the fetal rabbit muscle nAChR, it does not induce any developmental effects in this species despite similar systemic exposure. Furthermore, th e amino acid sequence of theSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 153 of 176 rat and human γ subunit indicate they are 90% identical, and therefore sufficiently unique that pharmacodynamic differences can be expected.

Conclusion The Agency agrees that the proposed MoA is reasonable and likely. There is sufficient evidence with dose and time concordance to support the postulated MoA for skeletal abnormalities and increased neonatal pup deaths in rats. However, for the purposes of risk assessment, the Agency

considers this MoA relevant to humans based on uncertainty regarding alternative MoAs.

Area of concern and potential study designs DAS should consider to reduce the uncertainty and increase confidence in the proposed MoA include the following:

Demonstrate the inability of sulfoxaflor to cause sus tained contracture in rabbit fetal diaphragm muscle. Alternatively, DAS can explore the functional response of rabbit nicotinic receptors (fetal and adult isoforms) expressed in HEK cells. Conclusively demonstrate that sulfoxaflor does not perturb the fun ctionality of neuronal nicotinic receptors

III. Sulfoxaflor Metabolite

- 474 Considerations

X11719474 (

- 474) is the main soil metabolite of sulfoxaflor; it is found in treated crops and is predicted to have the potential to occur in ground and surface wa ter (t 1⁄2 > 1000 days). Due to the high potential for exposure, numerous toxicity studies were conducted with

- 474 including acute studies, 28

- and 90

- day studies in the rat and dog, a mutagenicity battery, a reproduction and development study, liver toxici ty mechanistic studies, and an nAChR binding study.

- 474 was consistently less toxic than sulfoxaflor, approximately 7

- fold, and does not have acute or developmental toxicity. Unlike sulfoxaflor,

- 474 does not bind the rat fetal muscle nAChR (MRID 478320 35) and therefore does not share the developmental MoA described for sulfoxaflor. Otherwise,

- 474 and sulfoxaflor have similar toxicity profiles; the most reliable indicators of the relative toxicity are effects to the liver (e.g., liver weight increase, hepatocellular hypertrophy, fatty change (vacuolation), single cell necrosis and aggregates of macrophages).

IV. Global Partner Comments

PMRA has reviewed the MoA characterizing the pup deaths as not relevant to humans. The overall evidence in suppo rt the proposed MOA for the fetal abnormalities and neonatal death was considered to be weak by the PMRA reviewer due to the inconsistencies noted in the findings as they relate to the proposed key events, and the fact that the possibility of alternate MoA s has not been fully discounted.

Key Event 1: Specific binding to the fetal

- type muscle nAChR (Key Event #1)

Sulfoxaflor binding to other AChR types was discussed in the MoA rationale; however, these alternate MoAs were not fully explored and could not be discounted. It was postulated in the Leydig cell MoA that there was binding of sulfoxaflor to neuronal nAChRs which increases dopamine release; this effect was demonstrated in the recently submitted microdialysis assay.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 154 of 176 Furthermore, although there was a lack of muscarinic nAChR

- mediated clinical signs, heart arrhythmia may be asymptomatic or the clinical signs could have been easily missed, especially when neonatal deaths occur very shortly after birth.

Agonism at the fetal

- type muscle nAChR ( Key Event #2)

Expression of the recombinant rat and human (fetal and adult) muscle nAChRs in a mammalian cell culture would have provided a more native cellular environment compared to Xenopus ooctyes. Furthermore, the temperature at which the receptors are express ed would be significantly different (18 o C in ooctyes and 37 o C in mammalian cells) and may have a potential impact on the function of the receptor. Also, investigation of sulfoxaflor

- induced agonist activity on fetal and adult rabbit muscle nAChRs could pro vide a useful comparison between species and additional weight of evidence to the proposed MoA.

Sustained agonism/sustained muscle contracture (Key Event #3)

In the rat developmental toxicity and mechanistic studies, there was a lack of consistency in o bservations for fetal abnormalities;

o Forelimb flexure and hindlimb rotation were not seen in the one

- generation reproductive toxicity range

- finding study

o There were no treatment

- related developmental findings reported in the reproductive toxicity range

- fin ding study up to 1000 ppm

o Necropsy of (PND 4) neonates in the critical window mechanistic studies did not reveal any observations of bent clavicle, convoluted ureter, or hydroureter. However, there are literature studies of chemicals demonstrating postnat al remodeling (based on delay ossification) which was found to be reversible by PND 6 (caffeine toxicity) and PND 63 (ethylene glycol).Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 155 of 176 References:

Bennett, M. R. and A. G. Pettigrew (1974). "The formation of synapses in striated muscle during developmen t." J Physiol 241 : 515

- 545. Bulbringe, E. (1946). "Observations on the isolated phrenic nerve diaphragm preparation of the

rat." Br. J. Pharmacol. Chemother. 1 : 38

- 61. Buttar, H. S. (1997). "An overview of the influence of ACE inhibitors on fetal

- placental circulation and perinatal development." Molecular and Cellular Biochemistry 176 (1): 61

- 71. Evans, C. A. and D. R. Waud (1973). "Do Maternally Administered Neuromuscula r Blocking Agents Interfere With Fetal Neuromuscular Transmission?" Anesthesia & Analgesia

52 (4): 548

- 552. Geller, L. M. (1959). "Failure of nicotine to affect development of offspring when administered to pregnant rats." Science 129 : 212

- 214. Gibb, A. (2010). "Observations on the effects of XDE

- 208 on the phrenic nerve

- hemidiaphragm preparation from new

- born rat." Report of the University College London, London, UK. Kues, W. A., B. Sakmann and V. Witzemann (1995). "Differential expression patte rns of five acetylcholine receptor subunit genes in rat muscle during development." Eur J Neurosci

7 : 1376

- 1385. Liao, J.

- W., J.

- J. Kang, S.

- H. Liu, C.

- R. Jeng, Y.

- W. Cheng, C.

- M. Hu, S.

- F. Tsai, S.

- C. Wang and V. F. Pang (2000). "Effects of Cartap on Isol ated Mouse Phrenic Nerve Diaphragm

and Its Related Mechanism." Toxicological Sciences 55 (2): 453

- 459. Maritz, G. S. and G. v. Wyk (1997). "Influence of maternal nicotine exposure on neonatal rat lung structure: protective effect of ascorbic acid." Comp Bio chem Physiol C Pharmacol

Toxicol Endocrinol 117 : 159

- 165. Millar, N. (2010). "Characterization of the Agonist Effects of XDE

- 208 on Mammalian Muscle Nicotinic Acetylcholine Receptors." Report of the University College London, London,

UK. Mishina, M., T. Ta kai, K. Imoto, M. Noda, T. Takahashi, S. Numa, C. Methfessel and B. Sakman (1986). "Molecular distinction between fetal and adult forms of muscle acetylcholine receptor." Nature 313 : 364

- 369. Missias, A. C., G. C. Chu, B. J. Klocke, J. R. Sanes and J. P. M erlie (1996). "Maturation of the Acetylcholine Receptor in Skeletal Muscle: Regulation of the AChR γ

- to

- ε Switch." Developmental Biology 179 (1): 223

- 238. Robinson, S. R. and W. P. Smotherman (1988). Behavior of the Fetus . Caldwell, New Jersey, The

Telford Press. Shoro, A. A. (1977). "Intra

- uterine growth retardation and limb deformities produced by neuromuscular blocking agents in the rat fetus." J. Anat. 123 : 341

- 350. Sobrian, S. K., S. F. Ali, W. S. Jr. and R. R. Holson (1995). "Interactive effects of pre natal cocaine and nicotine exposure on maternal toxicity, postnatal development and behavior

in the rat." Mol Neurobiol 11 : 121

- 143.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 156 of 176 A ppendix C . Metabolism Summary Table

Table C .1. Tabular Summary of Metabolites and Degradates

Chemical Name (other names in parenthesis) Matrix Approximate Percent TRR

Structure Matrices

- Major Residue (>10%TRR) Matrices

-

Minor Residue (<10%TRR)

Parent Tomato 33

--

Lettuce 17

-- Succulent Pea (pods) 60

-- Rice (grain) 35

-- Rotational Crops

-- < 3

Ruminant > 80

-- Poultry > 80

-- Rat (urine and feces) > 93

-- Surface Water ~100

-- Groundwater

-- -- X11719474 Tomato 20

--

Lettuce 31

-- Succulent Pea (pods) 13

-- Rice (grain)

-- 9 Rotational Crops ~40

- 90

-- Ruminant

-- -- Poultry

-- -- Rat (urine and feces)

-- -- Surface Water

-- -- Groundwater ~88

-- X11721061 (including glucose and glucose/malonyl conjugates) Tomato 16

--

Lettuce

-- 9

Succulent Pea (pods) 14

-- Rice (grain) 13

-- Rotational Crops

-- Generally < 9

Ruminant

-- < 3 Poultry

-- -- Rat (urine and feces)

-- < 4

Surface Water

-- -- Groundwater

-- -- X11519540 Tomato

-- --

Lettuce

-- -- Succulent Pea (pods)

-- -- Rice (grain)

-- -- Rotational Crops

-- < 5

Ruminant

-- < 2 Poultry

-- < 8

Rat (urine and feces)

-- -- Surface Water

-- -- Groundwater 12

-- X11596066 Tomato

-- --

Lettuce

-- -- Succulent Pea (pods)

-- -- Rice (grain)

-- -- Rotational Crops

-- -- Ruminant < 18

--Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 157 of 176 Table C .1. Tabular Summary of Metabolites and Degradates

Chemical Name (other names in parenthesis) Matrix Approximate Percent TRR

Structure Matrices

- Major Residue (>10%TRR) Matrices

-

Minor Residue (<10%TRR) Poultry < 14

-- Rat (urine and feces)

-- -- Surface Water

-- -- Groundwater

-- -- All crop metabolism studies were conducted at approximately a 2

- fold exaggerated rate relative to the draft labels. Lettuce was harvested 7 days after the last treatment; tomatoes, peas, and rice were harvested 14 days after the last treatment. The proposed label lists PHIs ranging from 1 to 14 days, depending on th e crop.

Livestock metabolism studies were conducted at levels of 12

- 14 mg/kg in the feed. The estimated dietary burdens are approximately 2 ppm for dairy cattle, 0.45 ppm for beef cattle, and 0.054 ppm for poultry.

Rats received a gavage dose at either 5 mg/kg (single and multiple doses) or 100 mg/kg, or a single intravenous dose at 5 mg/kg.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 158 of 176 Appendix D. Physical/Chemical Properties and Summaries of Field Trial and Processing Study Results

Parameter Value Melting point/range (°C) 112.9 pH (24 oC

, 1.00% w/w aqueous solution) 5.82 Density (g/cm 3

, Technical grade @ 19.7 oC) 1.5378 Water solubility @ 20° C (g/L, 99.7% pure active ingredient ) pH 5 1

. 380 pH 7 0. 570 pH 9 0. 550

Unbuffered 0. 670 Solvent solubility @ 20 ° C (g/L, technical

- grade material )

Methanol 93.1

Acetone 217

p

- Xylene 0.743
1,2

- dichloroethane 39.6 ethyl acetate 95.2

n

- heptane 0.000242

n

- octanol 1.66 Vapor Pressure (Pa) (99.5% pure @ 25 oC) (technical grade @ 20 oC0) 2.7 x 10 - 7

1.1 x 10 - 7

Dissociation constant (pK

a ) NA Octanol/water partition coefficient Log(K OW )

Unbuffered NA pH 5 0.806 pH 7 0.802 pH 9 0.799 UV/visible absorption spectrum wavelength max absorption: 192, 211, and 260 nm

N/A = not available

Summary of Residues from the Crop Field Trials with Sulfoxaflor (SC formulation unless otherwise specified). Crop Matrix Applic. Rate

lb ai/A (g a.i./ha) PHI

(days) n Residues (ppm)

Min Max HAFT 1 Median Mean Std. Dev

Crop Group 1 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI)

Carrot roots 0.357

- 0.376

(401

- 423) 7 24 <0.01 0.032 0.031 0.010 0.015 0.008

Potato roots 0.355

- 0.374

(399

- 420) 6

- 8 36 <0.01 <0.01 0.01 0.01 0.01 NA

Radish roots

(WDG formulation) 0.360

- 0.363

(404

- 407) 7 12 <0.01 0.013 0.012 0.010 0.011 0.001

0.360

- 0.364 (404

- 407) 7 6 <0.01 0.016 0.014 0.010 0.011 0.002

Sugar beet roots 0.351

- 0.374

(394

- 420) 7 26 <0.01 0.025 0.023 0.010 0.011 0.003

Crop Group 2 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Carrot tops 0.360

- 0.366

(405

- 411) 7 8 0.311 2.283 2.069 0.515 0.857 0.764

Radish tops 0.360

- 0.364 7 12 0.183 0.506 0.478 0.258 0.297 0.108Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 159 of 176 Summary of Residues from the Crop Field Trials with Sulfoxaflor (SC formulation unless otherwise specified). Crop Matrix Applic. Rate

lb ai/A (g a.i./ha) PHI

(days) n Residues (ppm)

Min Max HAFT 1 Median Mean Std. Dev

(WDG formulation) (404

- 407)

0.360

- 0.364 (404

- 407) 7 6 0.242 0.465 0.441 0.398 0.370 0.099

Sugar beet tops 0.351

- 0.374

(394

- 420) 7 25 0.141 1.685 1.615 0.716 0.744 0.452

Crop Group 3 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Dry Bulb Onion 0.357

- 0.365

(400

- 409) 7 12 <0.01 <0.01 0.01 0.01 0.01 NA

Green Onion 0.360

- 0.366

(404

- 410) 7

- 8 12 <0.01 0.440 0.387 0.105 0.132 0.130

Crop Group 4 (proposed use = 0.266 lb ai/A total application rate, 3

- day PHI) Celery 0.360

- 0.361

(404

- 405) 2

- 3 24 0.058 0.804 0.771 0.143 0.229 0.235

Head Lettuce 0.328

- 0.388

(369

- 436) 2

- 3 27 <0.01 0.528 0.494 0.040 0.148 0.177

Leaf Lettuce

(WDG formulation) 0.342

- 0.377

(384

- 424) 2

- 3 36 0.05 3.07 2.74 0.495 0.744 0.659

0.360

- 0.363 (404

- 407) 3 6 0.379 0.885 0.806 0.577 0.595 0.189

Spinach 0.343

- 0.372

(385

- 418) 3 16 0.039 3.256 2.863 0.820 1.076 0.927

Crop Group 5 (proposed use = 0.266 lb ai/A total application rate, 3

- day PHI)

Broccoli 0.347

- 0.379

(390

- 426) 3 29 <0.01 1.600 1.584 0.070 0.201 0.395

Cauliflower 0.320

- 0.373

(360

- 419) 3

- 4 20 <0.01 0.07 0.055 0.014 0.020 0.014

Cabbage 0.341

- 0.383

(383

- 430) 2

- 3 28 <0.01 0.400 0.377 0.058 0.092 0.103

Mustard Greens 0.354

- 0.367

(397

- 411) 2

- 4 16 0.278 1.167 0.899 0.666 0.680 0.234

Crop Group 6 Beans (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI)

Succulent edible podded bean 0.352

- 0.372

(393

- 411) 7 12 0.024 2.019 1.938 0.104 0.422 0.716

Dry shelled bean 0.350

- 0.365

(393

- 411) 7 12 0.020 0.112 0.104 0.078 0.068 0.029

Crop Group 7 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Soybean, Forage 0.178

- 0.183

(200

- 205) 7

- 8 38 0.013 1.734 1.690 0.219 0.346 0.386

Soybean, Hay 0.358

- 0.368

(401

- 414) 6

- 8 38 0.057 1.378 1.238 0.725 0.670 0.383

Crop Group 8 (proposed use = 0.266 lb ai/A total application rate, 1

- day PHI)

Tomato 0.317

- 0.383

(356

- 430) 1 88 <0.01 0.762 0.602 0.051 0.091 0.123

Pepper, bell 0.321

- 0.364

(361

- 409) 1 24 <0.01 0.284 0.256 0.092 0.106 0.095

Pepper, non

- bell 0.343

- 0.428

(385

- 481) 1 15 0.017 0.46 0.44 0.090 0.156 0.145

Crop Group 9 (proposed use = 0.266 lb ai/A total application rate, 1

- day PHI) Cucumber (revised 0.355

- 0.374 1 36 <0.01 0.172 0.152 0.047 0.055 0.049Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 160 of 176 Summary of Residues from the Crop Field Trials with Sulfoxaflor (SC formulation unless otherwise specified). Crop Matrix Applic. Rate

lb ai/A (g a.i./ha) PHI

(days) n Residues (ppm)

Min Max HAFT 1 Median Mean Std. Dev

from Tier II removing Zucchini samples) (399

- 420)

Squash, summer /Zucchini (revised from Tier II adding Zucchini samples)

Squash, winter 0.357

- 0.367

(400

- 412) 1 10 <0.10 0.10 0.10 0.010 0.031 0.039

0.357

- 0.364 (400

- 408) 1 6 <0.01 0.021 0.018 0.011 0.013 0.004

Melon 0.356

- 0.375

(400

- 421) 0

- 2 31 <0.01 0.304 0.266 0.031 0.054 0.067

Crop Group 10 (proposed use = 0.266 lb ai/A total application rate, 1

- day PHI) Grapefruit 0.360

(404) 1 16 <0.01 0.186 0.130 0.014 0.041 0.056

Lemons 0.360

(404) 1 12 0.025 0.317 0.136 0.050 0.098 0.100

Oranges 0.263

- 0.368

(296

- 413) 1 61 0.037 0.46 0.44 0.114 0.167 0.120

C rop Group 11 Pome Fruit (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Apples 0.289

- 0.374

(325

- 420) 6

- 8 56 <0.01 0.297 0.266 0.070 0.086 0.057

Pears 0.279

- 0.379

(314

- 426) 7

- 8 32 0.044 0.267 0.261 0.142 0.146 0.063

Crop Group 12 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Apricot 0.337

- 0.345

(379

- 388) 7 8 0.13 0.45 0.39 0.16 0.21 0.12

Cherry 0.337

- 0.365

(379

- 410) 7 32 0.26 1.60 1.49 0.820 0.824 0.379

Nectarine 0.341

- 0.351

(383

- 394) 7 18 0.074 0.247 0.232 0.150 0.154 0.045

Peach 0.338

- 0.373

(380

- 419) 6

- 8 48 <0.01 0.636 0.541 0.121 0.164 0.118

Plum

(WDG formulation) 0.342

- 0.370

(384

- 415) 7 14 0.021 0.221 0.212 0.049 0.074 0.064

0.358

- 0.366 (401

- 410) 7 12 0.014 0.362 0.358 0.076 0.115 0.117

Crop Group 15 (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI) Barley Grain 0.083

- 0.095

(94

- 108) 12

- 17 50 <0.010 0.37 0.32 0.048 0.060 0.060

Barley Hay 0.086

- 0.096

(97

- 108) 6

- 8 40 <0.010 0.643 0.642 0.213 0.249 0.174

Barley Straw 0.083

- 0.095

(94

- 108) 12

- 17 52 <0.010 2.2 1.46 0.189 0.322 0.413

Wheat Grain

(WDG formulation) 0.084

- 0.098

(94

- 110) 12

- 17 67 <0.01 0.067 0.059 0.013 0.019 0.013

0.089

- 0.093 (100

- 104) 14 6 <0.01 0.063 0.063 0.010 0.028 0.027

Wheat Forage 0.084

- 0.098

(94

- 110) 6

- 8 60 <0.01 1.400 1.400 0.108 0.187 0.225Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 161 of 176 Summary of Residues from the Crop Field Trials with Sulfoxaflor (SC formulation unless otherwise specified). Crop Matrix Applic. Rate

lb ai/A (g a.i./ha) PHI

(days) n Residues (ppm)

Min Max HAFT 1 Median Mean Std. Dev

(WDG formulation) 0.084

- 0.098

(94

- 110) 7 6 0.041 0.221 0.197 0.095 0.111 0.073

Wheat Hay

(WDG formulation) 0.084

- 0.096

(94

- 108) 6

- 8 60 0.024 1.024 0.848 0.246 0.292 0.202

0.089

- 0.093 (100

- 104) 7 6 0.101 0.167 0.159 0.150 0.139 0.030

Wheat Straw

(WDG formulation) 0.084

- 0.097

(94

- 109) 12

- 17 69 0.030 1.562 1.489 0.250 0.400 0.393

0.088

- 0.093 (99

- 104) 14 6 0.060 1.641 1.600 0.105 0.588 0.784

Crop Groups 13

- 07F & 13

- 07G (proposed use = 0.266 lb ai/A total application rate, 1

- day or 7

- day PHI)

13

- 07G

Strawberries 0.339

- 0.369

(381

- 414) 1 26 0.02 0.50 0.49 0.184 0.182 0.109

13

- 07F Grape (all) 0.276

- 0.390

(310

- 438) 7

- 8 66 0.01 1.9 1.6 0.133 0.268 0.336

Crop Group 14 Tree Nuts (proposed use = 0.266 lb ai/A total application rate, 7

- day PHI)

Almond nutmeat 0.355

- 0.367 7 12 <0.01 0.013 0.123 <0.01 0.011 0.001

Almons hulls 0.355

- 0.367 7 12 1.04 3.12 3.05 1.68 1.86 0.681

Pecan nutmeat 0.360

- 0.363

(404

- 407) 7 12 <0.01 <0.01 <0.01 <0.01 <0.01 NA

C rop Group 20 (proposed use = 0.266 lb ai/A total application rate, 14

- day PHI) Cotton gin byproducts (14

- Day PHI) 0.346

- 0.375

(389

- 422) 14

- 15 42 0.037 4.20 4.03 0.476 0.869 1.12

Cotton, seed (14

- Day PHI) 0.346

- 0.375

(389

- 422) 14

- 15 44 <0.01 0.182 0.176 0.019 0.037 0.038

Canola, seed (14

- Day PHI) 0.078

- 0.099

(88.1

- 110.8) 13

- 15 36 <0.01 0.224 0.215 0.042 0.054 0.046

Canola, forage (14

- Day PHI) 0.078

- 0.099

(88.1

- 110.8) 13

- 14 8 0.03 1.5 1.3 0.11 0.39 0.57

Soybean, Seed 0.357

- 0.371

(400

- 416) 6

- 8 36 <0.01 0.214 0.199 0.010 0.026 0.044

1 HAFT = Highest Average Field Trial

Identification of Radioactivity in Hydrolyzed Buffered Water*. Analyte 14C

- sulfoxaflor 14C

- X11719474 14C

- X11721061

pH 4 pH 5 pH 6 pH 4 pH 5 pH 6 pH 4 pH 5 pH 6

Sulfoxaflor 99.6 100.0 100.4

- - - - - - X11719474

- - - 99.0 96.9 89.1

- - - X11721061

- - - - - - 100.0 99.9 99.7

X11579457

- - - 0.4 3.8 11.6**

- - - Accountability 99.6 100.0 100.4 99.4 100.7 100.7 100.0 99.9 99.7 *Only the pH is reported in this table. Durations and Temperatures were pH 4 = 20 minutes at 90°C, pH 5 = 60 minutes at 100°C, pH 6 = 20 minutes at 120°C. **Confirmed by LC/MS/MS.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 162 of 176 Residue data from processing studies with Sulfoxaflor (XDE

- 208)

RAC Processed Commodity Total Rate

lb a.i./A (g a.i./ha) PHI a

(day) Residues (ppm) b,c,d Processing Factor e

Apple Apple Fruit RAC *

0.716 (802.2) 7 0.343

-- Washed apples 0.246 0.7x Apple sauce 0.202 0.6x

Juice 0.128 0.4x Wet pomace 0.365 1.1x Dry pomace 1.431 4.2x Canned apples (0.0098) 0.03x Dried apples 0.118 0.3x * Apple fruit RAC samples analyzed after collection had similar residues (0.362 ppm, 0.028 ppm, and 0.009 ppm) to RAC samples analyzed prior to processing.

Barley Grain (RAC) *

0.277 (311.7) 16 0.174

-- Pearl barley 0.130 0.7x Pot barley 0.152 0.9x

Bran 0.167 1.0x

Flour 0.132 0.8x Cleaned barley 0.165 0.9x Brewing malt 0.163 0.9x Malt sprouts 0.224 1.3x

Beer 0.029 0.2x Spent grains 0.029 0.2x Brewer’s yeast 0.026 0.1x

* Barley RAC samples analyzed after collection had similar residues (0.16 ppm, 0.014 ppm, and 0.041 ppm) to RAC samples analyzed prior to processing.

Cabbage Heads (upon collection of field

samples)

0.734 (824) 3 0.429

--
Heads prior to processing (inc. wrapper leaves) 0.180

--

Inner leaves 0.014 0.1x Outer (wrapper) leaves 0.322 1.8x

Cooked head (0.009) 0.1x Cooking liquid (0.007) 0.1x

Sauerkraut 0.017 0.1x Sauerkraut juice 0.013 0.1x These values are not used in calculation of the processing factor. The cabbage RAC was stored under “cool” conditions prior to processing, and was not kept in frozen storage.

Oil seed, rape/

Taurus Seeds prior to processing

0.264 (296.4) 14 0.033

-- Cleaned seeds 0.037 1.1x

Meal 0.063 1.9x

Crude oil ND 0.3x

Refined oil ND 0.3x

Solvent

- extracted meal 0.072 2.2x Canola RAC samples analyzed after collection had similar residues (0.038 ppm, ND, and <LOQ) to RAC samples analyzed prior to processing.

Carrot Carrot root (RAC) 1.092 (1,224) 7 0.0101

-- Washed and peeled root (0.0082)

, ND <1.0x

Cooked carrot ND

, ND <1.0x Cooking liquid 0.0130

, (0.0085) Avg. 0.012 1.1x

Carrot juice 0.0234

, 0.0246 Avg. 0.0240 2.4xSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 163 of 176 Residue data from processing studies with Sulfoxaflor (XDE

- 208)

RAC Processed Commodity Total Rate

lb a.i./A (g a.i./ha) PHI a

(day) Residues (ppm) b,c,d Processing Factor e

Canned carrot ND

, ND <1.0x

Cherry Whole cherries* (RAC)

0.715 (803) 7 0.765

-- Washed cherries 0.648 0.8x

Canned cherries 0.737 1.0x

Juice 0.653 0.9x

Jam 0.838 1.1x

Dried cherries 3.950 5.2x *Cherry fruit RAC samples analyzed after collection had similar residues (0.616 ppm, 0.035 ppm, and 0.039 ppm) to RAC samples analyzed prior to processing.

Cotton Seed (RAC)

1.08 (1,213) 15 0.1001

-- Aspirated seed fractions 2.3197 23x

Delinted seed 0.0983 1.0x

Hulls 0.1846 1.8x

Meal 0.0822 0.8x Meal press cake 0.0781 0.8x

Crude oil (0.0059) <0.1x

Refined oil ND <0.1x

Grape/

Pinot Noir RAC

0.739 (830) 7 0.243

-- Raisins 0.856 3.5x

Juice 0.164 0.7x

Wine bottled 0.171 0.7x

Pomace 0.237 1.0x

Head Lettuce Heads with wrapper leaves (RAC)

0.748 (837.9) 3 0.592

-- Wrapper leaves 0.593 1.0X Unwashed heads without wrapper

leaves 0.354 0.6X

Washed heads 0.146 0.2X Washings 0.045 0.1X Head lettuce RAC samples analyzed after collection had similar residues (0.507 ppm, 0.081 ppm, and 0.07 ppm) to RAC samples analyzed prior to processing.

Leaf Lettuce Unwashed lettuce (RAC) 0.728 (818) 3 0.654

-- Washed lettuce 0.488 0.7x Washings 0.159 0.2x

Onion Whole onion (RAC) 1.09 (1,225) 7 ND

-- Peeled onion ND NC

Dried onion ND NC

Orange (Valencia) Pulp RAC*

1.79 (2.00) 1 (0.0039)

-- Peel RAC* 0.3661

-- Whole orange RAC* (peel +pulp) 0.0625

--

Juice (0.0042) <0.2X Wet pulp 0.1567 2.5X Dried pulp 0.5159 8.3X

Peel 0.5714 9.1X

Oil ND <0.2X

Marmalade ND <0.2X

Canned slices ND <0.2X *Oranges were separated into peel and pulp prior to analysis. Processing factors were calculated using the whole orange RAC (peel + pulp) residues, which were calculated by the study reviewer as the sum of the peel and pulp residues, adjusted for the indi vidual weights of the fractions.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 164 of 176 Residue data from processing studies with Sulfoxaflor (XDE

- 208)

RAC Processed Commodity Total Rate

lb a.i./A (g a.i./ha) PHI a

(day) Residues (ppm) b,c,d Processing Factor e

Potato (Kuras) Potato tubers* (RAC)

1.05 (1,176) 7 0.012

-- Washed potatoes 0.014 1.2X Peeled potatoes 0.019 1.6X

Peel 0.022 1.8X

Potato flakes 0.030 2.5X

Micro

- waved potatoes 0.013 1.1X

Boiled potatoes 0.012 1.0X Cooking water (0.006) <0.8X Potato chips 0.025 2.1X Dried potatoes 0.043 3.6X

French fries 0.019 1.6X *Unpeeled and unwashed; sampled just prior to processing.

Soybean Seed (RAC)

1.10 (1,236) 7 0.0727

-- AGF 6.8871* 95X

Meal 0.0964 1.3X

Hulls 0.1079 1.5X

Pressed cake 0.0793 1.1X Expeller crude oil 0.0220 0.3X

Solvent

- extracted crude oil 0.0203 0.3X

Refined oil ND <0.1X

*Results are the mean of re

- analysis.

Strawberry Unwashed strawberry

0.724 (812) 1 0.2003

--

Washed strawberry 0.1670

0.1859 [0.1765] 0.9X

Juice 0.0498

0.0564 [0.0531] 0.3X

Canned strawberry 0.1215

0.1180 [0.1198] 0.6X

Jam 0.0830

0.0702 [0.0766] 0.4X

Samples were analyzed in duplicate; average residues are bolded and reported in brackets. Residues <LOQ were assigned a value of 0.01 ppm when averaged with a detectable residue.

Sugar beet Sugar beet root (RAC)

1.06 (1,191) 7 0.013

-- Pulp (0.005) <0.8x

Press water (0.006) <0.8x Raw juice 0.018 1.4x Thin juice 0.014 1.1x Lime sludge (0.010) <0.8x Thick juice 0.061 4.7x Raw sugar 0.023 1.8x White sugar (0.006) <0.8x

Molasses 0.135 10x Dried pulp 0.039 3.0x

Tomato Tomato RAC 0.719 (807.8) 1 0.119

-- Fruit, washed and peeled 0.141 1.2X

Juice 0.117 1.0XSulfoxaflor Human Health Risk Assessment DP No. D382604

Page 165 of 176 Residue data from processing studies with Sulfoxaflor (XDE

- 208)

RAC Processed Commodity Total Rate

lb a.i./A (g a.i./ha) PHI a

(day) Residues (ppm) b,c,d Processing Factor e

Canned 0.093 0.8X Ketchup 0.250 2.1X

Puree 0.235 2.0X

Paste 0.518 4.4X

Wheat Wheat Grain (RAC)

0.444 (499) 15 0.0650

-- Aspirated Grain Fraction 1.3817 21X

Total bran 0.0230 0.4X

Germ 0.0307 0.5X

Bran 0.0279 0.4X Middlings 0.0109 0.2X

Shorts 0.0132 0.2X

Whole meal flour 0.0136 0.2X

Refined white flour (0.0079) <0.2X Whole grain bread (0.0091) <0.2X

White bread (0.0046) <0.2X

Gluten ND <0.2X

Gluten feed meal (0.0073) <0.2X

Starch ND <0.2X

a PHI = Preharvest Interval. ND = Non Detectable where residues were below the limit of detection (<LOD). LOD = 0.003 ppm.

b Residue values in parenthesis are <LOQ but >LOD. The LOQ was 0.010 ppm and the LOD was 0.003 ppm.

c Control specimens were collected and analyzed. No residues were detected for the unfortified control samples nor were any analytical interferences present in any untreated control sample.

d Processing factors were ca lculated using the equation: Processing Factor = [Measured residue for analyte in the processed fraction] / [Measured residue for analyte in the RAC prior to processing]. Residues <LOQ and ND were assigned a value of 0.01 ppm for calculation purposes.

e NC = Not calculated because residues were not detected above the LOQ in the RAC and processed fraction.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 166 of 176 Appendix E. Review of Human Research

Aside from the Pesticide Handlers Exposure Database Version 1.1 (PHED 1.1)

, the Agricultural Handler Exposure Task Force (AHETF) database

, the Outdoor Residential Exposure Task Force (ORETF) database

, and the Agricultural Re

- entry Task Force (ARTF) database , which were described at the end of the Executive Summary, no data from s tudies in which adult human subjects were intentionally exposed to a pesticide or other chemical were used in this risk

assessment.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 167 of 176 Appendix F. Occupational Exposure/Risk Summary Tables

Short

- and Intermediate

- term Occupational Handler Exposu re and Risk Estimates for Proposed U se s of Sulfoxaflor ( Transform TM WG ). Exposure

Scenario Crop or Target (Crop Group #) Dermal

Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated or

Amount Handled Daily

3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

Mixer/Loa der Dry Flowable

for

Groundboom Application (AHETF) Brassica (Cole) Leafy Veg., Bulb

Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small F ruit Vine (Sub CG 13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.227 (Baseline) 0.00896

(No respirator) 0.086 80 A 0.00057 3,200 0.00089 2,000 1,200

Ornamentals, Turf grass (Sod Farm) 0.227 (Baseline) 0.00896

(No respirator) 0.133 80 A 0.00088 2,100 0.00140 1,300 800

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.227 (Baseline) 0.00896

(No respirator) 0.047 200 A 0.00077 2,300 0.00120 1,500 900

Cotton, Soybean 0.227 (Baseline) 0.00896

(No respirator) 0.086 200 A 0.0014 1,300 0.0022 0 810 49 0

Dry Flowable

for Aerial Application (AHETF) Brassica (Cole) Leafy Veg., Bulb

Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.227 (Baseline) 0.00896

(No respirator) 0.086 350 A 0.00250 730 0.00390 460 280

Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 0.227 (Baseline) 0.00896

(No respirator) 0.133 350 A 0.00380 470 0.00600 300 180

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.277 (Baseline) 0.00896

(No respirator) 0.047 1200 A 0.00460 390 0.00730 250 150

Cotton, Soybean 0.277 (Baseline) 0.00896

(No respirator) 0.086 1200 A 0.0085 0 210 0.0134 0 130 80

Dry Flowable

for Chemigation (AHETF) Potatoes 0.227 (Baseline) 0.00896

(No respirator) 0.086 350 A 0.00250 730 0.00390 460 280

Dry Flowable

for A irblast Citrus (CG 10), Pome Fruits (CG

11), Ornamentals, Stone Fruits (CG 0.227 (Baseline) 0.00896

(No respirator) 0.133 40 A 0.00044 4,100 0.00069 2,600 1,600Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 168 of 176 Short

- and Intermediate

- term Occupational Handler Exposu re and Risk Estimates for Proposed U se s of Sulfoxaflor ( Transform TM WG ). Exposure

Scenario Crop or Target (Crop Group #) Dermal

Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated or

Amount Handled Daily

3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

Application (AHETF) 12), Tree Nuts (CG 14 & Pistachio)

Applicator Sprays for

Groundboom Application (AHETF) Brassica (Cole) Leafy Veg., Bulb

Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 0 7F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.0786 (Baseline) 0.00034

(No respirator) 0.086 80 A 0.00020 9,200 0.000034 53,000 7,800

Ornamentals, Turf grass (Sod Farm) 0.0786 (Baseline) 0.00034

(No respirator) 0.133 80 A 0.00030 5,900 0.000052 34,000 5,100

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.0786 (Baseline) 0.00034

(No respirator) 0.047 200 A 0.00027 6,700 0.000046 39,000 5,700

Cotton, Soybean 0.0786 (Baseline) 0.00034

(No respirator) 0.086 200 A 0 .00049 3,700 0.000085 21,000 3,1 00

Sprays for

Aerial Application (PHED ) Brassica (Cole) Leafy Veg., Bulb

Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.005 (Engineerin g Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.086 350 A 0.000055 33,000 0.000030 61,000 21,000

Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 0.005 (Engineerin g Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.133 350 A 0.000084 21,000 0.000046 39,000 14,000

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.005 (Engineerin g Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.047 1200 A 0.00010 18,000 0.000056 32,000 11,000

Cotton, Soybean 0.005 (Engineerin g Controls

- Enclosed 0.000068

(Engineering

Controls

- Enclosed 0.086 1200 A 0.00019 9,600 0.0001 18,000 6,200Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 169 of 176 Short

- and Intermediate

- term Occupational Handler Exposu re and Risk Estimates for Proposed U se s of Sulfoxaflor ( Transform TM WG ). Exposure

Scenario Crop or Target (Crop Group #) Dermal

Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated or

Amount Handled Daily

3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

cockpit) cockpit) Sprays for

A irblast Application (AHETF) Citrus (CG 10), Pome Fruits (CG

11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 1.77 (Baseline) 0.00471

(No respirator) 0.133 40 A 0.00340 530 0.00036 5,000 480

Flagger Flagging for

Aerial Application (PHED) All Low Acreage Crops 0.011 (Baseline) 0.00035

(No respirator) 0.133 350 A 0.00019 9,700 0.00024 7,600 4,200

Flagging for

Aerial Application (PHED) All High Acreage Crops 0.011 (Baseline) 0.00035

(No respirator) 0.086 350 A 0.00012 15,000 0.00015 12,000 6,700

Mixer/Loader/Applicator

Mechanically

pressurized Handgun Sprayer Ornamentals , Greenhouses 3.5 (Baseline) (PHED) 0.120

(No respirator) (PHED) 0.00 133 1,000 gal/day 0.00169 1,100 0.00231 780 450

Turf grass (Sod farms) 1.96 (Baseline) (ORETF) 0.042

(No respirator) (ORETF) 0.133 5 A 0.00047 3,800 0.00040 4,400 2,100

Backpack Sprayer Ornamentals 13.2 (Baseline) (AHETF) 0.140

(No respirator) (AHETF) 0.00 133 40 gal/day 0.00025 7,200 0.00011 16,000 5,000

Turf grass (Sod farms) 8.26 (Baseline) (MRID 44339801) 0.00258

(No respirator) (MRID 44339801) 0.00 133 40 gal/day 0.00016 11,000 0.000002 900,000 11,000

Manually

pressurized

Handwand (PHED) Ornamentals

, Turf grass (Sod farms) 100

(Baseline) 0.030

(No respirator) 0.00 133 40 gal/day 0.00193 930 0.000023 78,000 920

1. Occupational Pesticide Handler Unit Exposure Surrogate Reference Table (OPHED; March, 2012).

2. Application Rate is based on the proposed label.

3. Acres Treated per Day is based on Exposure SAC SOP #9: Standard Values for Daily Acres Treated.

4. Dermal Dose = [Dermal Unit Exposure (mg/lb ai) x Application Rate (lb ai/A) x Acres Treated (A/day) x % DAF (2.5%)]/%)/BW (69 kg).

5. Dermal MOE = Dermal NOAEL (1.8 mg/kg/day)/Dermal Dose (mg/kg/day) ; LOC = 30

. 6. Inhalation Dose = [Inhalation Unit Exposure (mg/lb ai) x Application Rate (lb ai/A) x Acres Treated (A/day)]/)/BW (69 kg).

7. Inhalat ion MOE = NOAEL (1.8 mg/kg/day)/Inhalation Dose (mg/kg/day) ; LOC = 30

. 8. Combined MOE = NOAEL (1.8 mg/kg/day)/Combined Dose (mg/kg/day) ; LOC = 30

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 170 of 176 Short

- and Intermediate

- term Occupational Handler Exposure and Risk Estimates for Proposed U se s of Sulfoxaflor ( GF

- 2032 SC ). Exposure Scenario Crop or Target (CG #) Dermal Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated

or Amount

Handled Daily 3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

Mixer/Loader Liquids for

Groundboom Application (AHETF) Brassica (Cole) Leafy

Veg., Bulb Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1 ), Small Fruit Vine (Sub CG 13

- 07F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.220 0.000219 0.090 80 A 0.00057 3,100 0.000023 79,000 3,000

Ornamentals, Turf grass (Sod Farm) 0.220 0.000219 0.133 80 A 0.00085 2,100 0.000034 53,000 2,000

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.220 0.000219 0.043 200 A 0.00069 2,600 0.000027 66,000 2,500

Cotton, Soybean 0.220 0.000219 0.090 200 A 0.0014 0 1,300 0.00 0057 32,000 1,2 00

Liquids for Aerial Application (AHETF) Brassica (Cole) Leafy

Veg., Bulb Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 07F), Low Growing Berry (Sub 13

- 07G),

Suc culent, Edible Podded, Dry Beans 0.220 0.000219 0.090 350 A 0.00250 720 0.00010 18,000 690

Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 0.220 0.000219 0.133 350 A 0.00370 490 0.00015 12,000 470

Barley, Tricale, Wheat, 0.220 0.000219 0.043 1200 A 0.00410 440 0.00016 11,000 420Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 171 of 176 Short

- and Intermediate

- term Occupational Handler Exposure and Risk Estimates for Proposed U se s of Sulfoxaflor ( GF

- 2032 SC ). Exposure Scenario Crop or Target (CG #) Dermal Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated

or Amount

Handled Daily 3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

Canola (Sub CG 20A) Cotton, Soybean 0.220 0.000219 0.090 1200 A 0.00860 210 0.00034 5,300 200 Liquids for Chemigation (AHETF) Potatoes 0.220 0.000219 0.0 90 350 A 0.00250 720 0.00100 18,000 690

Liquids for

A irblast Application (AHETF) Citrus (CG 10), Pome

Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 0.220 0.000219 0.133 40 A 0.00042 4,200 0.000017 110,000 4,100

Applicator Sprays for

Groundboom Application (AHETF) Brassica (Cole) Leafy

Veg., Bulb Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 0 7F), Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans 0.0786 (Baseline) 0.00034

(No respirator) 0.090 80 A 0.00021 8,800 0.000035 51,000 7,500

Ornamentals, Turf grass (Sod Farm) 0.0786 (Baseline) 0.00034

(No respirator) 0.133 80 A 0.00030 5,900 0.000052 34,000 5,100

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.0786 (Baseline) 0.00034

(No respirator) 0.043 200 A 0.00024 7,300 0.000042 42,000 6,300

Cotton, Soybean 0.0786 (Baseline) 0.00034

(No respirator) 0.090 200 A 0.00051 3,500 0.000089 20,000 3,000

Sprays for Aerial Application (PHED ) Brassica (Cole) Leafy

Veg., Bulb Veg. (CG 3

- 07), Cucurbit Veg. (CG 9), Fruiting Veg. (CG 8), Leafy Veg. (Except

Brassica CG 4 & Watercress), Leaves Root/Tuber Veg. (CG 2), Root/Tuber Veg. (CG 1), Small Fruit Vine (Sub CG 13

- 07F), 0.005 (Engineering

Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.090 350 A 0.000057 32,000 0.000031 58,000 25,000Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 172 of 176 Short

- and Intermediate

- term Occupational Handler Exposure and Risk Estimates for Proposed U se s of Sulfoxaflor ( GF

- 2032 SC ). Exposure Scenario Crop or Target (CG #) Dermal Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated

or Amount

Handled Daily 3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

Low Growing Berry (Sub 13

- 07G), Succulent, Edible Podded, Dry Beans Citrus (CG 10), Pome Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 0.005 (Engineering

Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.133 350 A 0.000084 21,000 0.000046 39,000 19,000

Barley, Tricale, Wheat, Canola (Sub CG 20A) 0.005 (Engineering

Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.043 1200 A 0.000093 19,000 0.000051 35,000 12,000

Cotton, Soybean 0.005 (Engineering

Controls

- Enclosed cockpit) 0.000068

(Engineering

Controls

- Enclosed cockpit) 0.090 1200 A 0.00020 9,200 0.00011 17,000 6,000

Sprays for A irblast Application (AHETF) Citrus (CG 10), Pome

Fruits (CG 11), Ornamentals, Stone Fruits (CG 12), Tree Nuts (CG 14 & Pistachio) 1.77 (Baseline) 0.00471

(No respirator) 0.133 40 A 0.00340 530 0.00036 5,000 480

Flagger Flagging for Aerial Application (PHED) All Low Acreage Crops 0.011 (Baseline) 0.00035

(No respirator) 0.133 350 A 0.00019 9,700 0.00024 7,600 4,200

Flagging for Aerial Application (PHED) All High Acreage

Crops 0.011 (Baseline) 0.00035

(No respirator) 0.090 350 A 0.00013 14,000 0.00016 11,000 6,300

Mixer/Loader/Applicator

Mechanically

pressurized Handgun Sprayer Ornamentals

,

Greenhouses 3.5 (Baseline) (PHED) 0.120

(No respirator) (PHED) 0.00 133 1,000 gal/day 0.00169 1,100 0.00231 780 450

Turf grass (Sod farms) 1.14 (Baseline) 0.0019

(Baseline) (ORETF) 0.133 5 A 0.00047 3,800 0.00040 4,400 2,100Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 17 3 of 176 Short

- and Intermediate

- term Occupational Handler Exposure and Risk Estimates for Proposed U se s of Sulfoxaflor ( GF

- 2032 SC ). Exposure Scenario Crop or Target (CG #) Dermal Unit Exposure 1
(mg/lb ai) (Mitigation Level) Inhalation Unit

Exposure 1
(mg/lb ai) (Mitigation Level) Max Single

App Rate (lb ai/A or lb ai/gal ) 2 Area Treated

or Amount

Handled Daily 3 Dermal Inhalation Combined

MOE 8
(LOC=30) Dose 4
(mg/kg/day) MOE 5 Dose 6 (mg/kg/day) MOE 7

(ORETF)

Backpack Sprayer Ornamentals 13.2 (Baseline) (AHETF) 0.140

(No respirator) (AHETF) 0.00 133 40 gal/day 0.00025 7,200 0.00011 16,000 5,000

Turfgrass (Sod farms) 8.26 (Baseline) (MRID 44339801) 0.00258

(No respirator) (MRID 44339801) 0.00 133 40 gal/day 0.00016 11,000 0.000002 900,000 11,000

Manually

pressurized

Handwand (PHED) Ornamentals

, Turfgrass

(Sod farms) 100 (Baseline) 0.030

(No respirator) 0.00 133 40 gal/day 0.00193 930 0.000023 78,000 920

1. Occupational Pesticide Handler Unit Exposure Surrogate Reference Table (OPHED; March, 2012).

2. Application Rate is based on the proposed label.

3. Acres Treated per Day is based on Exposure SAC SOP #9: Standard Values for Daily Acres Treated.

4. Dermal Do se = [Dermal Unit Exposure (mg/lb ai) x Application Rate (lb ai/A) x Acres Treated (A/day) x % DAF (2.5%)]/%)/BW (69 kg).

5. Dermal MOE = Dermal NOAEL (1.8 mg/kg/day)/Dermal Dose (mg/kg/day) ; LOC = 30

. 6. Inhalation Dose = [Inhalation Unit Exposure (mg/lb ai) x Application Rate (lb ai/A) x Acres Treated (A/day)]/)/BW (69 kg).

7. Inhalation MOE = NOAEL (1.8 mg/kg/day)/Inhalation Dose (mg/kg/day) ; LOC = 30

. 8. Combined MOE = NOAEL (1.8 mg/kg/day)/Combined Dose (mg/kg/day) ; LOC = 30

.Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 174 of 176 Appendix G. Anticipat ed Post

- Application Activities and Dermal Transfer Coefficients

Anticipated Post

- A pplication Activities and Dermal Transfer Coefficients. † Proposed Crop Crop Group Transfer Coefficients

,

cm 2 /hr Activities EPA MRID No.

Field / row crop,

low / medium Barley, Wheat 70 Hand weeding 42851302

, 42689103

, 42689104
1,100 Scouting 45005904

, 45005908 Canola (Subgroup 20A) 1,100 Scouting 45005904

, 45005908

Cotton 70 Hand weeding 42851302, 42689103, 42689104

210 Scouting 42851302

900 g cotton/hr Harvesting Mechanical (Module Builder) 42701601

2400 g cotton/hr Harvesting Mechanical (Picker Operator; Raker) 42701601

5050 g cotton/hr Harvesting Mechanical (Tramper) 42701601 Soybean 70 Hand weeding 42851302

, 42689103

, 42689104
1,100 Scouting 45005904

, 45005908

Succulent Edible Podded and Dry Beans 70 Hand weeding 42851302

, 42689103

, 42689104

210 Scouting 42851302

, 42974501

1100 Scouting 45005904

, 45005908

1900 Irrigation (hand set) 45224801 Vegetable, head and stem

Brassica Brassica (Cole) Leafy Vegetables 230 Transplanting 45344501

, 45469501

, 45469502

330 Scouting 42974501

1400 Hand Weeding 45530102

1900 Irrigation (hand set) 45224801

4200 Weeding, Hand 45005906

, 45005907

, 45191701 Vegetable, " R oot" Bulb Vegetables (Crop Group 3

- 07) 330 Scouting 42974501

1400 Hand Weeding 45530102

1900 Irrigation (hand set) 45224801

4200 Weeding, Hand 45005906

, 45005907

, 45191701

Leaves of Root and Tuber Vegetables (Crop Group 2) 70 Weeding, Hand 42851302

, 42689103

, 42689104

210 Scouting 42851302

, 42974501

230 Transplanting 45344501

, 45469501

, 45469502

1100 Harvesting, Hand 43013003

, 40966503

, 40966502

, 45530103

1900 Irrigation (hand set) 45224801

Root and Tuber Vegetables (Crop Group 1) 70 Weeding, Hand 42851302

, 42689103

, 42689104

210 Scouting 42851302

, 42974501

1100 Harvesting, Hand 43013003

, 40966503

, 40966502

, 45530103

1900 Irrigation (hand set) 45224801 Tree Fruit, Evergreen Citrus (Crop Group 10) 10 0 Hand weeding 43297602

230 Transplanting 45344501

, 45469501

, 45469502

580 Hand Pruning 45175102

, 45480302Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 175 of 176 Anticipated Post

- A pplication Activities and Dermal Transfer Coefficients. † Proposed Crop Crop Group Transfer Coefficients

,

cm 2 /hr Activities EPA MRID No.

1400 Hand Harvesting 42830002

, 40975308

, 40966501

, 45138202

, 45175101

, 45432301

, 45432302

3600 Thinning Fruit 42428101 Vegetable, C ucurbit Cucurbits Vegetables (Crop Group 9) 9 0 Scouting 45005909

230 Transplanting 45344501

, 45469501

, 45469502

550 Turning 45480301

, 45491902

1900 Irrigation (hand set) 45224801 Vegetable, Fruting Fruiting Vegetables (Crop Group 8) 70 Weeding, Hand 42851302

, 42689103

, 42689104

9 0 Scouting 45005909

210 Scouting 42851302

, 42974501

230 Transplanting 45344501

, 45469501

, 45469502

550 Tying/Training , Hand Harvesting 45480301

, 45491902

1100 Harvesting, Hand 43013003

, 40966503

, 40966502

, 45530103

1900 Irrigation (hand set) 45224801 Vegetable, Leafy Leafy Vegetables except

Brassica

– (Crop Group 4)& Watercress 70 Weeding, Hand 42851302

, 42689103

, 42689104

210 Scouting 42851302

, 42974501

230 Transplanting 45344501

, 45469501

, 45469502

1100 Harvesting, Hand 43013003

, 40966503

, 40966502

, 45530103

1900 Irrigation (hand set) 45224801 Nursery/Greenhouse (Unassigned) Ornamentals 230 Harvesting, Hand 45344501

, 45469501

, 45469502

1900 Irrigation (hand set) 45224801
Tree, "F ruit", D eciduous Pome Fruits (Crop Group 11) 100 Orchard M aintenance , Propping, Hand Weeding 43297602

230 Transplanting 45344501

, 45469501

, 45469502

580 Scouting, Hand Pruning, Training 45175102

, 45480302

1400 Harvesting, Hand 42830002

, 40975308

, 40966501

, 45138202

, 45175101

, 45432301

, 45432302

3600 Thinning Fruit 42428101 Stone Fruits (Crop Group 12) 100 Orchard M aintenance , Propping, Hand Weeding, Thinning Fruit, Bird Control 43297602

230 Transplanting 45344501

, 45469501

, 45469502

580 Scouting, Hand Pruning, Training 45175102

, 45480302

1400 Harvesting, Hand 42830002

, 40975308

, 40966501

, 45138202

, 45175101

, 45432301

, 45432302

3600 Thinning Fruit 42428101

Vine /T rellis Small Fruit Vine Climbing (except fuzzy

kiwifruit

– Subgroup 13

- 07F) 230 Transplanting 45344501

, 45469501

, 45469502

640 Scouting, Hand Pruning, Hand Weeding,

Propagating, Bird Control, Trellis Repair, Tying/Training 45005910

1900 Irrigation (hand set) 45224801Sulfoxaflor Human Health Risk Assessment DP No. D382604

Page 176 of 176 Anticipated Post

- A pplication Activities and Dermal Transfer Coefficients. † Proposed Crop Crop Group Transfer Coefficients

,

cm 2 /hr Activities EPA MRID No.

5500 Leaf Pulling , T ying/Training , Hand Harvesting 43223907

, 40985601

10100 Leaf Pulling

, Tying/Training , Hand Harvesting 45491901

19300 Turning, Girdling 40975304 Berry, L ow Low Growing Berry (Subgroup 13

- 07G) 70 Hand Weeding, Hand Pruning (shears)

, Canopy Management 42851302

, 42689103

, 42689104

210 Scouting 42851302

, 42974501

230 Transplanting 45344501

, 45469501

, 45469502

1100 Hand Harvesting (ranking)

, Scouting 43013003

, 40966503

, 40966502

45530103

, 45005904

, 45005908

1900 Irrigation (hand set) 45224801 Tree, “Nut” Tree Nuts (Crop Group 14 & Pistachio) 100 Orchard M aintenance , Poling, Hand Weeding 43297602

190 Harvesting, Mechanical (shaking) 41848604

230 Transplanting 45344501

, 45469501

, 45469502

580 Scouting 45175102

, 45480302

1400 Harvesting, Hand (net) 42830002

, 40975308

, 40966501

45138202

, 45175101

, 45432301

45432302

Turf/Sod Turfgrass, Sodfarms 6700 Maintenance

, Harvesting, Slab Transplanting/Planting 45432303

† The information in the table is based on proprietary and non

- proprietary data.

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