UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460

MEMORANDUM

Date: Se pt e mb er 15

, 2015 OFFICE OF

CHEMICAL SAFETY AND

POLLUTION PREVENTION

SU BJ EC T : Tribufos: Human Health Draft Risk Assessment for R eg istration Re v iew.

PC Co d e: 074801

D ecis ion No.:**40L~o** 

### t;

Petition No.: NA DP Barcode: D 357537 Registration No.: NA

Re g ulator y Action: Reg

Review

Risk Assess ment Type: Single Chemical Aggregate

T XRNo.: NA Case No.: 2145

CAS No

. : 78-48-8

40 CF R: §180.299 M RID No

. : NA

FROM : Evisabel Craig, Ph.D.

, DABT

, Toxicologi st/ Risk Assessor, RAB YI Bridgett Bobowiec

, Biologist

, ORE**Asse~or,** J>AB VJ**~,..-4** Sheila Piper

, Chemist, RAB**Vl ~~pA--**  Health Effects Div i sion (7509P)

T HRO UGH: Donna Da vis, Acting Branch Chief 

## C'\1/1(/ /'

- /\ ,/

Risk Assessment Branch YI

### ~VCA-V**~** Health Effects Division (7509P)

# A~~

::stina Swa rtz

, RARC

# evicwe~k,**11--. ~**  William Irwin

, RARC Reviewer

### (,,(}~ 

## d\-**~** "YL

TO: Ke ll y Sherman, Branch Ch i ef

Marianne Mannix

, Chemical Re v iew Manager

Risk Management and Impl ementation Branch Ill

Pesticide R e -Evaluation Divi s ion ( PRO) (750 8P )

Page I of8-I1.0 Executive Sum1nary

..... ... .. . .. ... .... ...... .. ....... . ..... .. ........ .. ............... .. ..... .. ..... ... .... . .. .. ..... .. ........ 4

2

. 0 1-I ED Re co mm e ndati ons

.. .. ................................................................................................... 7

2.1 Data Defic i encies

............................................................................................................. 7

2.2 Tolerance Co n s id era ti o n s

. .. . . . ..... .. . .. ........ . . ... .. ...... . ... ....... .. . ... . . . . . .. .. .. ...... . .. . ..... .. . .. . ... . . .. ... . 7

2.2. l E n forcement Ana l ytical Method

.................................................................................. 7

2.2.2 Int ernat i ona l Harrn o nj za tion

................ . .. ... . . ....... .. .. ..... . . .. . .. . . .... .. .. ...... . . .. ..... .. . .. . .... . ... .. . 7

2

. 2.3 RecomJTiended Tolerances

. . . .. ... .. .. .... ... ....... .. ....... . .... ..... ... .. .. . ...... .. ..... .... ... .... ..... .. ..... ... 7

2.3 Labe l Recommendations

... .. .... .. .. . . .. ................................................................................. 8

2

. 3. l Recommendations from Residue R eviews

... ...... . . . .. .. .. ... .. . .. ... . .. .. . . .. . .. . . ........................ 8

3.0 In troduction

.... .. ..... .. ..... ... ..... ... .... ...... ... ............ ... ............... . ................................... . ..... .. . .. .. . 8

3. 1 C hemi ca l I dent i t y

.................................................................... . . ...... . ....... .... .... ..... .... ..... .. . 8

3.2 Ph ysical/Che mi ca l Characteristics

........... ............. ... ..... .... ..... ... .... .... ...... .. ..... .... ..... . ........ 9

3

. 3 P es ti c id e Use P atte rn

........................................................................................................ 9

3.4 Anticipated Exposure P athways

. ........ .. ... ....... ... ... ...... .. ...... ... ...... .. ..... ..... .. .... ..... .... ..... ... 12

3

. 5 Conside rati o n of E n viro nment al Ju st i ce

............ .. .. . ..... .. ...... .. ...... ..... ... . . . . ...................... 1 2

4. 0 H azard C ha racte ri za tion an d Dose-Response Assessment

.......... .. ...... ... .... .. ........ .. .... ..... .. 12

4 .1 Toxicology Studies Avai l able for Ana l ys i s

.. .. .... . .. ... ... ..... ... ..... ... ... . .............................. 1 3

4. 2 Absorption

, Distribution, Metabolism

, & E limination (ADME)

..... . ...... ...... .... .. .... .... ... 13

4.2. l D erma l Absorption

.... ..... .. ...... ... ... ... ....... .. .... ...... ... .. . .. ... .. . ... .. ..... ... .... ... .... ..... ..... .. .... . . 14

4 .3 Toxicological E ffec t s

.. . .. . .. .. .. . .. ... . . .. .... . ........ .. ........ . .. . ... . ... .. .... . ... .. . ... ..... .. ..... . ................ 14

4.3.1 T oxicology Profile

.. ...... .. .......................................................................... .. ................ . .... 14

4.3 .2 Cr iti ca l Durati ons of Expos ur e

........... .. ..... . ..... . .. . ....... .. ...... .. ...... .. ... .. .. .. . . . ... .. ..... . ... .. .. 1 5

4.4 Literature Review on Ne urode velopme nt Effec t s

.. ... ..... ... ..... ... . . ... . . .............................. 15

4.5 Safety Factor fo r I nfants a nd Children (FQ PA SF)

........ . ....... . ................. . .... . . . ....... .. ..... 20

4

. 5 .1 Co mpl eteness of th e To x i cology Databa se

......................................................... . ...... 21

4.5.2 Evide n ce of Neurotoxic it y

.......................................................................................... 2 1

4.5.3 Evide nc e of Sensi ti v it y / Susceptib ilit y in the Developing or Youn g Animal.

...... .. .... 21

4.5.4 Residual U nc er tainty in the Exposu r e Databa se

...... .. .. .. .. .. ...... .. ..... .. . .. . . .. . .. ... ... .... ...... 21

4

. 6 Toxicity Endpoint and Point of Dep art ur e Se le ct ion s

... .. ...... .. ...... . ................ . ... .. ... . . . .. . . 22

4.6. I Dose-Re spo n se Assessment

....................................................................................... 22

4.6.2 Recommendation for Co mbinjn g Route s of Expos ure s for Ri sk Assess m e nt

. .... . ..... 24

4

. 6

. 3 Cance r Class i fication and Risk Assessment Recommendat i on

.................................. 25

4.6.4 Summary of P o int s of Departure and Toxicity E ndpoint s Used in Hwnan Ri sk

Assessment.

.. ... . ...... . ..... . . . ..... .. .. .. . ..... . . ... ....... ... . . . ... .. .. . . .... .. ....... ... ..... .. ...... .. ............... . ........ . ...... 25

4.7 Endocr i n e Disruption

..................................................................................................... 26

5.0 Di etary Ex po s ure and Ri sk Assessme n t

........... .. ..... .. . .. ....... . . ... ... . . ..... .. ............................. 27

5

. 1 Metabolite / Degradate Residue Profile

. .. ....... .. ....... .. ... ..... ....... .. ..... ... .... ... . . ..... ... .... .... .... 27

5.2 Food R esidue Pr ofi l e

............... .. .............. .. ..... . .. ... .. .. ....... . . .. .. . . . ....... . ... ..... . . ...... . .. ..... . .. ... 28

5.3 Water Residue Profil e

................................................................................................ .. .. 28

5.4 Dietary Ri sk As sess ment..

.. ....... ... .. .... .. . ... ... ... ........ . ....... .. ... ..... ..... . . .... .. . .... . .... . ...... . ... .. .. 29

5.4. 1 Description of R es idu e Data Used in Di eta r y Assessment..

............... . ............... .. ..... . 29

5.4.2 Per ce n t Crop Treated Used in Di etary Assessment..

.................................................. 30

5.4.3 Ac ut e Di e t a r y Risk Assessment

. ........ .. ............. .. ....... .. ...... .. .. .. . . .. . .. .. ..... ..... . . . .. . ..... .... 30

5.4.4 Steady State Dietar y Ri s k As sessme nt

................................................ . . ..... .. . .. .. .. . . .... . 31

5.4.5 Ca n ce r Di etary Ri sk Assessme nt..

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

6

. 0 R esidential (Non- Occupational ) Exposure/R i sk Characterization

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

Pa ge 2 of84Page 3 of 84

6.1 Non

- Occupational Bystander Post

- Application Inhalation Exposure and Risk Estimates

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

6.2 Non

- Occupational Spray Drift Exposure and Risk

................................ ........................ 34 6.2.1 Combined Risk Estimates from Lawn Deposition Adjacent to Applications

.................. 35

7.0 Aggregate Exposure/Risk Characterization

................................ ................................ ....... 39

7.1 Acute Aggregate Risk

................................ ................................ ................................ .... 39

7.2 Steady State Aggregate Risk

................................ ................................ .......................... 39

7.3 Cancer Aggregate Risk

................................ ................................ ................................ ... 39

8.0 Cumulative Exposure/Risk Characterization

................................ ................................ ..... 39

9.0 Occupational Exposure/Risk Characterization

................................ ................................ .. 40

9.1 Steady

- State Handler Risk

................................ ................................ ............................. 40

9.2 Steady State Post

- Application Risk

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

9.2.1 Occupational Post

- application Inhalation Exposure/Risk Estimates

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

9.2.2 Occupational Post

- application Dermal Exposure/Risk Estimates

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

10.0 Review of the Incident Report

................................ ................................ ........................... 49

11.0 References

................................ ................................ ................................ .......................... 49 Appendix 1. Summary of OPP’s ChE Policy & Use of BMD Modeling

................................ .... 52 Appendix 2

– Results for BMD/BMDL modeling for Tribufos

................................ ................... 53 Appendix 3. Toxicology Profile and Executive Summaries

................................ ........................ 55

A.3.1 Toxicology Data Requirements

................................ ................................ .................. 55

A.3.2 Toxicity Profiles

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

A.3.2 Hazard Identification and Endpoint Selection............................................. 61 Appendix 4. IRL Sheet

..................................................................................... 8 2 Appendix 5. Summary of Steady State Non Cancer Occupational Handler Dermal Risk

Estimates for Tribufos .......................................................................................8 3Page 4 of 84

1.0 Executive Summary

Tribufos [S,S,S

- tributyl phosphorotrithioate] is an organophosphate (OP) herbicide used as a defoliant on cotton plants prior to harvest and as a bottom defoliant to reduce or prevent losses from boll rot organisms. There are no registered non

- occupation al (residential uses) for tribufos.

Humans may be exposed to tribufos in food and drinking water, since tribufos may be applied directly to growing crops and application may result in tribufos reaching surface and ground water sources of drinking water. There are no registered uses for tribufos that will result in direct

non

- dietary (i.e., residential) exposures. However, there is the potential for indirect non

- dietary exposure (incidental oral and dermal) to tribufos related to spray drift. In an occu pational setting, applicators may be exposed while handling the pesticide prior to application, as well as during application. There is a potential for post

- application exposure for workers re

- entering

treated fields. Tribufos is formulated as an emulsif iable concentrate (EC). The maximum application rates for the use on cotton are 1.875 pounds (lbs) active ingredient (ai) per acre (A) for the states of California (CA) and Arizona (AZ)

, and 1.125 lb ai/A for all other states. Application can be made by ground or aerial equipment

. Cotton h arvesting must be mechanical. Tribufos labels have a restricted entry interval (REI) of 7 days and require mixer/loaders to wear coveralls over l ong

- sleeved shirt, lon g pants, chemical resistant gloves, chemical

- resistant footwear plus socks, chemical

- resistant apron, and protective eyewear.

Tribufos is a member of the OP class of pesticides. Like other OPs, the initiating event in the

mammalian adverse outcome pathwa y (AOP)/ mode of action (MOA), for tribufos involves inhibition of the enzyme acetylcholinesterase (AChE) via phosphorylation of the serine residue at the active site of the enzyme. This inhibition leads to accumulation of acetylcholine and ultimately to neurotoxicity in the central and/or peripheral nervous system . For tribufos , AChE inhibition is the most sensitive endpoint in the toxicology database in multiple species, durations, lifestages, and routes. OPs also exhibit a phenomenon known as steady s tate AChE inhibition. After repeated dosing at the same dose level, the degree of inhibition comes into equilibrium with the production of new, u ninhibited enzyme. The “steady state” endpoint selection for tribufos overlaps HED’s traditional short

- term e xposure duration endpoint selection as well as being appropriately protective for workers that are exposed over longer periods of time (i.e.,

intermediate

- term exposures) as well as chronic exposure

. Therefore, steady state exposure

asses sments were condu cted instea d of the traditional chronic, long

- term, intermediate

- term

, and/ or short

- term assessments

.

The tox icology database for tribufos is complete for risk assessment . There are acceptable data

available f

or toxicity endpoint selection from studies across multiple lifestages, durations, and

routes for both red blood cell ( RBC ) and brain AChE inhibition . Clinical signs of neurotoxicity can be found throughout the database of toxicity studies at doses much higher than those causing

inhibition of AChE. Q uantitative susceptibility was observed in pups compared to adults in the acute comparative ChE (CCA) studies but no q uantitative or qualitative susceptibility was observed in repeated

- dosing studies. However, the Food Quality Protection Act (FQPA) Saf ety Factor (SF) has been retained for infants, children, youths, and women of childbearing age for all exposure scenarios due to uncertainty in the human dose

- response relationship for neurodevelopmental effects (See Section 4.4). As a result, a total unce rtainty factor of 1000XPage 5 of 84

was applied for all exposure scenarios, except dietary exposures for the adult population subgroup 50

- 99 years old where the FQPA SF does not apply (total uncertainty factor = 100X) and inhalation exposures where the interspecies un certainty factor has been reduced to 3X (total uncertainty factor = 300X).

Tribufos is classified as “likely” at high doses but “unlikely” to be a human carcinogen at low doses since all tumor increases in mice occurred only at the highest dose tested (48 mg/kg/day in males and 63 mg/kg/day in females) and were accompanied by severe ChE inhibition and increased mortality. In addition, there was no concern for mutagenicity, and no structural analogs of concern were identified. Tribufos is categorized as h aving high acute toxicity via the

oral and dermal route s (Category II ) and is moderately toxic via the inhalation route (Category III)

. Tribufos induce s mild eye and skin irritation (Category IV) and is not a dermal sensitizer.

The residue chemistry database for tribufos is complete. Acceptable data were submitted depicting residues in the target crop , cotton

. The qu alitative nature of the residues of tribufos in plants (cottonseed) and livestock has been adequately identified/ characterized and is understood. The HED Metabolism Assessment Review Committee (MARC) reviewed the tribufos toxicology

and metabolism data and recommend ed that the residue of concern for both tolerance expression and risk assessment for plants and livest ock should consist of tribufos only. In addition, HED previously determined that tolerances for tribufos residues in eggs and poultry tissue s were not required based on the results of the respective poultry metabolism and feeding studies (CFR §180.6(a)(3) ). These conclusions have not changed for the purpose of registration review. Additionally, based on the livestock feeding study and on an updated livestock diet, there are no reasonable expectation s of finite residues of tribufos in/ on any livestock comm odity. Therefore,

t he established tolerances for residues of tribufos in/on milk, eggs, cattle, goat, horse, hog, poultry, and sheep commodities (40 CFR §180.272) should all be revoked.

Tribufos is moderately mobile and persistent in the soil suggesting the potential for runoff and/or leaching . Based on the environmental fate parameters, there is the potential for tribufos to reach drinking water resources. The Environmental Fate and Effects Division (EFED) performed surface water and groundwater modeling using the highest application rates. The highest estimated drinking water concentrations were from surface water (Texas cotton [TX] ) and to

further refine t he dietary assessment, average rates were considered.

The acute aggregate (food and drinking water) dietary exposure estimates are below HED’s level

of concern ( i.e., <100% of the a cute population adjusted dose ( aPAD ) ) for the U.S. population and all population subgroups. Combined dietary exposure from cottonseed oil and drinking

water concentrations based on maximum rates in TX at the 95 th percentile of exposure is 15 % of the aPAD for the U.S. population and 48 % of th e aPAD for all infants (<1 yr old), the most highly exposed population subgroup. Also, the combined dietary exposure from cottonseed oil and drinking water concentrations based on average rates in TX at the 95 th percentile of exposure

is 7% of the aPAD for the U.S. population and 22% of the aPAD for all infants (<1 yr old), the most highly exposed population subgroup. The food only exposure contributes <1 for and drinking water only contributes 48% of the aPAD for all infants.Page 6 of 84

For the steady state assess ment, 2

- day average food residues, representing average food exposure, and 21

- day forward

- rolling water averages were compared to the steady state (ss) point of departure

. Combined dietary exposure from cottonseed oil and drinking water concentrations

based on maximum rates in TX at the 95 th percentile of exposure is 72 % of the ssPAD for the U.S. population and 226 % of the ssPAD for all infants (<1 yr old) , the most hig hly exposed population subgroup

. T he combined dietary exposure from cottonseed oil and TX cotton

average rate for drinking water at the 95 th percentile of exposure is 34 % of the ssPAD for the U.S. population and 105 % of the ssPAD for all infants (<1 yr old) , the most hig hly exposed populat ion subgroup

. The food only exposure contributes <2% of the ssPAD and drinking water only contributes 226 % of the ssPAD for all infants.

T here are no registered residential uses ; therefore, the acute and steady state aggregate

assessments are the same a s the acute and steady state dietary assessments discussed above

.

A spray drift assessment was completed for the currently registered occupational use of tribufos on cotton, to determine the potential exposure and risk for those exposed via drift from b oth aerial and ground applications . The spray drift assessment was completed using both registered maximum application rates for cotton (1.875 lbs ai/acre and 1.125 lb ai/A) for aerial and groundboom application methods. The resulting risk estimates were still a concern at 300 feet from the field edge (MOE < 1,000) for adult dermal exposures as well as children (1<2 years of age) combined dermal and incidental oral exposure s (MOE < 1,000).

For occupational handler exposure, a total aggregate risk index (ARI) approach was used as a risk metric since the toxicological effects for dermal and inhalation exposure routes were similar but the levels of concern (LOCs) are different (LOC = 1000 for dermal and 300 for inhalation exposure) . The occupational handler assessment identified risk estimates of concern (ARIs < 1) for all mixer/loader and applicator scenarios despite the engineering control and application rate mitigations made in the previous RED.

For the occupational post

- application assessment, H ED relied on chemical

- specific data depicting the residues of tribufos available for transfer to workers re

- entering treated areas or conducting specific post

- application activities. HED has determined that there are steady

- state
duration post

- applicatio n dermal risk estimates of concern (i.e., MOEs < 1

, 00 0 ) on the day of treatment ( DAT 0)

. Current tribufos labels require a Restricted Entry Interval (REI) of 7 days after application

, however

, activities such as module builder operator, rakers, and trampe rs result in risks of concern up to as many as 30 days following application

.

This risk assessment relies in part on data from studies in which adult human subjects were intentionally exposed to a pesticide or other chemical. These data, which include studies from Pesticide Handler Exposure Database (PHED) 1.1; the Agricultural Handlers Exposure Task Force (AHETF) database; the Agricultural Reentry Task Force (ARTF) database ; and other registrant

- submitted studies (MRID 427016 01 and MRID 42685901) submitted to the Agency by the registrant are (1) subject to ethics review pursuant to 40 CFR 26, (2) have received that review, and (3) are compliant with applicable ethics requirements. For certain studies, the ethicsPage 7 of 84

review may hav e included review by the Human Studies Review Board. Descriptions of data sources, as well as guidance on their use, can be found at the Agency website 1

.

2.0 HED Recommendations

2.1 Data Deficiencies

There are no residue chemistry or toxicology data deficiencies. A biomonitoring study was requested in the 2000 IRED to confirm risk management decisions. In 2001, the request for the biomonitoring study was placed on hold, pending submission of the reques ted neurotoxicity

studies. These neurotoxicity studies have since been submitted and reviewed

, and found not to change the dermal toxicity endpoint. Accordingly, a biomonitoring study is no longer required.

2.2 Tolerance Considerations

2.2.1 Enforcemen t Analytical Method

Adequate methods are available for enforcement and data collection purposes for residues of

tribufos in cottonseed commodities and milk. The Pesticide Analytical Method Volume II (PAM II) lists four gas

- liquid chromatography (GLC) /fla me ionization detector methods (Methods I, II, A and C). Method B is available for analysis of cottonseed commodities and milk, and uses an electron capture detector. Method II is used to determine tribufos residues in milk. The limit of quantitation (LOQ) is 0.05 ppm for cottonseed and 0.002 ppm for milk. There is no single

residue method included in PAM Vol. II for tribufos residues in livestock commodities.

2.2.2 International Harmonization

There are no CODEX, Canadian, or Mexican maximum residue levels (MRL s ) established for

tribuf os

. Therefore, international harmonization is not an issue

.

2.2.3 Recommended Tolerances

Permanent tolerances are currently established under 40 CFR§180. 272 for residue s of tribuphos (i.e. tribufos) in/on cott on and livestock commodities

. However, based on the recalculated diets for ruminants and swine, there is no reasonable expectation of finite residues of tribufos on any livestock commodity (Category 3 of 40 CFR §180.6(a)). The established tolerances for residues of tribufos in/on cattle, goat, horse, hog, and sheep commodities (40 CFR §180.272) should thus

be revoked. HED recommends that the residue definition for the tribufos tolerance expression should read as following :

“Tolerances are established fo r the residues of the defoliant tribuphos, including its metabolites and degradates, in or on the commodities in the listed below. Compliance

1 http://www.epa.gov/pesticides/science/handler

- exposure

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

- app

- exposure

- data.htmlPage 8 of 84

with the tolerance levels specified below is to be determined by measuring only tribuphos (S,S,S

- tributyl phosph orotrithioate), in or on the commodity.”

Table 2.2.2 . Tolerance Summary for Tribuphos , §180. 272 Commodity Proposed Tolerance (ppm) HED

- Recommended

Tolerance (ppm) Comments

( correct commodity definition )
Cattle, fat 0.15 revoke
Cattle, meat 0.02 revoke Cattle, meat byproducts 0.02 revoke Cotton, gin byproducts 40.0 40.0
Cotton, undelinted seed 4.0 4.0
Goat, fat 0.15 revoke
Goat, meat 0.02 revoke Goat, meat byproducts 0.02 revoke Hog, fat 0.15 revoke
Hog, meat 0.02 revoke
Hog, meat byproducts 0.02 revoke
Horse, fat 0.15 revoke

Horse, meat 0.02 revoke Horse, meat byproducts 0.02 revoke

Milk 0.01 revoke Sheep, fat 0.15 revoke Sheep, meat 0.02 revoke Sheep, meat byproducts 0.02 revoke

2.3 Label Recommendations

2.3.1 Recommendations from Residue Reviews

No label changes are needed for the residue chemistry.

3.0 In troduction

Tribufos is an OP herbicide registered for use on cotton as a defoliant

. It was first registered in

the United States in 1961 to defoliat e cotton in preparation for machine harvesting

. The technical registrant for tribufos is Bayer CropScience

. The most recent human health risk

assessment for tribufos was conducted in 200 0 (D266540, R. Travaglini, 06/26/2000).

3.1 Chemical Identity

Tab le 3.1. Tribufos Nomenclature

Chemical StructurePage 9 of 84

Tab le 3.1. Tribufos Nomenclature

Empirical Formula C 12 H 27 OPS 3
Common Name Tribuphos; Tribufos

IUPAC name S,S,S

- tributyl phosphorotrithioate

CAS Name S,S,S

- tributyl phosphorotrithioate
CAS Registry

Number 78

- 48

- 8

Chemical Class Organophosphate Defoliant

3.2 Physical/Chemical Characteristics

Tribufos is a colorless to yellow liquid with a mercaptan

- like odor. It is practically insoluble in

water, but is completely miscible in dichloromethane, n

- hexane, 2

- propanol, and toluene. Tribufos is relatively stable to heat and under acidic conditions, but slowly hydrolyzes under

alkaline conditions. Tribufos ’ moderately low vapor pressure (1.7 x 10

- 6 torr) and Henry’s Law constant (3.1 x10

- 7 atm

- m 3 /mol ) indicate that the compou nd is semi

- volatile. The water solubility, soil adsorption, hydrolysis and aerobic soil metabolism data suggest that tribufos is not likely to be a ground water contaminant.

Table 3.2. Tribufos Physical and Chemical Properties

Parameter

Value

Molecular Weight 314.5 g/mole

Water solubility (20°C) 2.3mg/L

Solvents

- Completely miscible with n

- hexane, dichloromethane, toluene, and 2

- propanol Vapor pressure (20°C) 1.7 x 10

- 6 Torr Melting and Boiling Point <

- 25 0 C; 150 ° C at 0.3 mm Hg Density 1.02 g/cm 3 @ 20 o C Henry’s Law Constant (25°C) 3.1 x 10

- 7 atm

- m 3 /mol Octanol/water partition coefficient (25°C) 3.31 x 10 5 ; l og Kow = 5.7

3.3 Pesticide Use Pattern

Tribufos is registered for use as a cotton defoliant

. Tribufos accelerates the defoliation process by stimulating the formation of the abscission layer where the stem joins the stalk, causing the leaves and stems to drop cleanly to allow mechanical harvesting of the crop without staining the

lint. Leaves may begin to drop in green c ondition 3 day s after application with satisfactory leaf drop in 5

- 7 days under favorable conditions. Tribufos is formulated as a liquid in emulsifiable concentrate (70 % active ingredient , ai ), and as a liquid technical grade (97 % ai ). ProductsPage 10 of 84

contain 6 lbs of tribufos per United States (US) gallon of product. Tribufos containing products currently registered include; 1) Folex 6 EC Cotton (EPA Reg. No. 5481

- 504), 2) Def 6 Emulsifiable Defoliant (5481

- 903 or EPA Reg. No. 264

- 730) [AMVAC Chemical Corpora tion], 3) DFT 6 EC Cotton Defoliant (EPA Reg. No. 34704

- 867) [Loveland Products, Inc.], and 4) Tribufos 6 (EPA Reg. No. 85678

- 15) [Red Eagle International Corporation]. Tribufos is applied by ground and aerial equipment. Application by chemigation is pro hibited. California and Arizona have a higher single maximum application rate of 1.875 lb ai/A than the rest of the U.S. states with a maximum application rate of 1.125 lb ai/A.

Labels indicate mixer/loaders must wear personal protective equipment ( PPE ) including coveralls over long

- sleeved shirt, long pants, chemical resistant gloves, chemical

- resistant footwear plus socks, chemical

- resistant apron, and protective eyewear. One product, FOLEX 6

EC (EPA reg # 5481

- 504) also has a respirator requirement. Mixers and loaders for aerial applications must use a closed mechanical transfer system that meets requirements listed in the worker protection standard (WPS) for agricultural pesticides [40 CFR 170.240 (d)(4)] for providing dermal protection. A erial app lications must be conducted in an enclosed cockpit. Cotton must be mechanically harvested. Hand harvesting is prohibited. Module building equipment does not appear to be prohibited on tribufos labels. See Table 3.3 for use profile.Tribufos Human Health Risk Assessment DP No. D357537

Page 11 of 84 Table 3.3 . Summary of Directions for Use of Tribufos.

Registrant Company Product

and EPA (Reg. No) Percent

active ingredient (ai) Application.

Timing, Type, and Equip. Application.

Rate (lb ai/A) Max. No.

Application. per Season REI PPE Limits and Directions

Cotton

AMVAC

CHEMICAL

CORPORAT

ION FOLEX 6

EC (5481

- 504)

70.5 % ai.

Contain 6 lbs/gallon Ground and

Aerial California

and Arizona

1.875 lb

ai/A

Other U.S.

states 1.125

lb ai/A. 2 7 days Mixers/loaders for aerial application must use mechanical transfer system. Applicators and

other handlers must wear coveralls over baseline dermal wear plus gloves, protective eyewear, chemical resistant headgear for overhead exposure. Chemical

- resistant a pron when cleaning, mixing or loading. In addition a PF10 respirator is required for enclosed areas and PF5 for outdoor applications

. Can be used for bottom or complete

defoliation. Cotton must be mechanically harvested . Hand harvesting prohibited. Aerial application must be in enclosed cockpits. Do not apply through any type of irrigation. Do not graze treated fields. Diesel oil may be

used instead of water in adverse

conditions. DEF 6 EC

Defoliant

5481

- 9023 (264

- 730) Mixers/loaders for aerial application must use mechanical transfer system. Mixer/loaders, applicators must wear coveralls over baseline dermal wear plus gloves, protective eyewear, chemical resistant headgear of overhead exposure and chemical

- resistant apron when cleaning, mixing o r loading

DEF Technical Defoliant (5481

- 9022) 97.9 % ai.; For use in the manufacture of cotton defoliants.

LOVELAND PRODUCTS,

INC DFT 6 EC

Cotton

Defoliant (34704

- 867) 70.5 % ai. Ground and

Aerial California

and Arizona

1.875 lb

ai/A

Other U.S.

states 1.125

lb ai/A. 2 7 days Mixers/loaders for aerial application must use mechanical transfer system. Mixer/loaders, applicators must wear coveralls over baseline dermal wear plus gloves, protective eyewear, chemical resistant headgear of overhead

expo sure and chemical

- resistant apron when cleaning, mixing or loading. Can be used for bottom or complete defoliation. Aerial application must be in enclosed cockpits. Cotton must be mechanically harvested. Hand harvesting prohibited. Do not apply through an y type of irrigation. Do not graze treated fields. Diesel oil may be used instead of water in

adverse conditions. REDEAGLE

INTERNATI

ONAL LLC Tribufos 6

(85678

- 15)

Tribufos Technical (85678

- 10) 99.5 % ai. For use in the manufacture of cotton defoliants

AXION AG PRODUCTS,

LLC AX

Tribufos 6 (89167

- 43) 70.5 % ai.

Contain 6 lbs/gallon Ground and

Aerial California

and Arizona

1.875 lb

ai/A

Other U.S.

states 1.125

lb ai/A. 2 7 days Applicators and other handlers must wear

coveralls over baseline dermal wear plus gloves, protective eyewear, chemical resistant headgear of overhead exposure. Chemical

resistant apron when cleaning, mixing or loading. Can be used for bottom or complete defoliation. Aerial application must be in enclosed cockp its. Cotton must be mechanically harvested

. Hand harvesting prohibited. Do not apply through any type of irrigation. Do not graze treated fields. Diesel oil may be used instead of water in

adverse conditions.Tribufos Human Health Risk Assessment DP No. D357537

Page 12 of 84 3.4 Anticipated Exposure Pathways

The Pesticide Re

- Evaluation Division has requested a registration review assessment of human health risk to support the registered use s of tribuf os . Humans may be exposed to tribufos in food and drinking water, since tribufos may be applied directly to growing crops and application may

result in tribufos reaching surface and ground water sources of drinking water. There are no registered uses for t ribufos that will result in direct non

- dietary (i.e., residential)

exposures. However, there is the potential for indirect non

- dietary exposure (incidental oral and dermal) to tribufos related to spray drift. In an occupational setting, applicators may b e exposed while handling the pesticide prior to application, as well as during application. There is a potential for post

- application exposure for workers re

- entering treated fields.

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://epa.gov/compliance/ej/resources/policy/exec_order_12898.pdf )

. As a part of every pesticide risk assessment, OPP considers a large variety of consumer subgroups a ccording to

well

- established procedures. In line with OPP policy, HED estimates risks to population subgroups from pesticide exposures that are based on patterns of that subgroup’s food and water consumption, and activities in and around the home that invo lve 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 data are analyzed and categorized by subgroups based on age and ethnic group. Additionally, OPP is able to assess dietary exposure to smaller, specialized subgroups and exposure assessments are performed when

conditions or circumstances wa rrant. 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 evaluated. Further

consider ations are currently in development as OPP has committed resources and expertise to the development of specialized software and models that consider exposure to bystanders and farm workers as well as lifestyle and traditional dietary patterns among specifi c subgroups.

4.0 Hazard Characterization and Dose

- Response Assessment

Tribufos is a member of the organophosphate (OP) class of pesticides which functions by plant esterase inhibition causing elevated ethylene levels and leaf abscission

. Like other OPs, the initiating event in the mammalian adverse outcome pathway (AOP)/mode of action (MOA), for tribufos involves inhibition of the enzyme cholinesterase (ChE) via phosphorylation of the serine residue at the active site of the enzyme. This inhibition leads to accumulation of acetylcholine and ultimately to neurotoxicity in the central and/or peripheral nervous system (see Figure 1).

For tribufos , ChE inhibition is the most sensitive endpoint in the toxicology database in multiple species, durations, lifestages, and routes. ChE inhibition is the focus of this hazardTribufos Human Health Risk Assessment DP No. D357537

Page 13 of 84 characterization; the availability of reliable ChE inhibition dose response data is one of the key determinants in evaluating this toxicology database.

Figure 1. Adverse outcome pathway for OPs

4.1 Toxicology Studies Available for Analysis

The toxicology database for tribufos is complete for risk assessment. There are acceptable studies available for toxicity endpoint selection; they include:

 chronic oral toxicity studies in rats and dogs

 carcinogenicity studies in rats and mice

 developmental studies in rats and rabbits

 r eproduction study in rats

 acute and subchronic neurotoxicity studies in rats

 developmental neurotoxicity (DNT) study in rats

 acute and repeated comparative cholinesterase (ChE) studies in juvenile and adult

rats

 repeated, gestational ChE study in pregnant rats and fetuses

 delayed neurotoxicity study in hens

 subchronic dermal toxicity in rabbits

 subchronic inhalation study in rats

 immunotoxicity study in rats

 gene mutation, chromosome aberrations and unscheduled DNA synthesis studies

 metabolis m studies in rats

4.2 Absorption, Distribu tion, Metabolism, & Elimination (ADME)

Some OPs require metabolic activation to the oxon metabolite. However, tribufos is not one of these OPs (see chemical structure in Table 3.1). Instead, tribufos as the pa rent active ingredient,

is a ChE

- inhibiting chemical. Generally, absorption and distribution are rapid with extensive

metabolism and no accumulation in the tissues.

In a rat metabolism study (MRID 42034501)

, [1

- C 14 ] tribufos (97 % ai) was administered to male

and female Sprague

- Dawley rats (5 animals/sex/dose) by gavage as a single dose at levels of 5 or

100 mg/kg, or as a single dose at 5 mg/kg following a 14

- day pretreatment with unlabeled

tribufos at 5 mg/kg/day. Five additional rats/sex received a single 5 mg/kg int ravenous dose

. Target

Tissue Dose Phospho rylation

of the active site
of ChE Neurotoxicity

Accumulation of acetylcholineTribufos Human Health Risk Assessment DP No. D357537

Page 14 of 84 Fifty

- five to 80% of tribufos was absorbed into the bloodstream

. The majority of the radioactive dose was excreted within 72 hours through urine (55

- 80%), followed by feces (15

- 42%), with

< 1% in the expired air. Females showed more radioactive compound in the urine than males for each dose regimen. There was no significant tissue residue . Only two excreted compounds could be identified, unmetabolized tribufos , the major compound found in t he feces and butyl

gamma

- glutamylcysteinylglycine disulfide , a minor metabolite, in the urine. The data indicate complete metabolic breakdown of absorbed tribufos with extensive breakdown of the S

- butyl moieties. No evidence of sulfonation or glucuronidati on was observed based on treatment with

sulfatase or b

- glucuronidase.

4.2.1 Dermal Absorption

No guideline dermal penetration study was available; however, the dermal absorption factor was

not needed for tribufos because a 21

- day dermal toxicity study, which showed RBC ChE inhibition in rabbits at 2 mg/kg/day, was available (MR I D # 42007201 ). The similarity of the dermal and oral PODs following BMD modeling indicates that tribufos is highly absorbed via the dermal route.

4.3 Toxicological Effects

4.3. 1 Toxicology Profile

The toxicology database for tribufos is complete with respect to the regulatory requirements and provides evidence that like other organophosphates (OPs), tribufos inhibits cholinesterase in dogs, rabbits, rats, and mice. Furthermore, ChE inhibition is readily seen via all exposure routes

and at all durations. T he toxicology profile for tribufos has acceptable dose

- response data across multiple lifestages, durations, and routes for both RBC and brain ChE inhibition . Many of these studies have been evaluated using benchmark dose ( BMD ) modeling techniques. The data

indicate that RBC ChE inhibition is generally more sensitive than brain ChE inhibition. Also, RBC data provided the best fit for BMD modeling. There is no evidence to suggest sex

- specific sensitivity or that pregnant females are more sensitive than non

- pregnant females. Juveniles

appear to be more sensitive than adults following acute, but not repeated dosing. Also, fetuses do not appear to be more sensitive than pregnant females. Dermal and inhalation studies with a ChE component are available and allow route

- specific evaluations

.

In the subchronic neurotoxicity study, brain and RBC ChE activity were decreased starting at the

mid

- dose (2.89/3.54 mg/kg/day [M/F]) but no cholinergic signs were observed up to the highest dose tested (36.8/42.6 mg/kg/day [M/F]). In the developmental ne urotoxicity study, rat brain and RBC ChE activity were decreased starting at 3.4 mg/kg/day while cholinergic signs (tremors) were observed at the highest dose tested (16.4 mg/kg/day) in pregnant rats. Decreased body weights, clinical signs (delayed surfac e righting reflex, decreased motor activity, decreased startle response) as well as decreased plasma (but not RBC or brain) ChE were also observed in the offspring at 16.4 mg/kg/day. Tribufos is not a developmental or a reproductive toxicant. In the developmental neurotoxicity study, neurotoxic effects were observed in the offspring at the

same dose as maternal effects. In chronic studies conducted wit h mice, rats and dogs, tox icity

was manifested as inhibition of plasma, RBC and/or b rain ChE activity a nd ocular damage (rats only)

. Tribufos i s no t immunotoxic

.Tribufos Human Health Risk Assessment DP No. D357537

Page 15 of 84 The HED Cancer Peer Review Committee (CPRC) [now designated as Cancer Assessment Review Committee (CARC)] classified tribufos as “likely” at high doses but “unlikely” to be a human carcinogen at low doses

. A ll tumor increases in mice occurred only at the highest dose tested (48 mg/kg/day in males and 63 mg/kg/day in females) and were accompanied by severe ChE inhibition and increased mortality in the later weeks of the study (TXR# 0012242, E. Rinde

and R. Zen dzian, 22

- MAY

- 1997). No tumors were found in rats. The CPRC recommended a

non

- quantitative approach for the purpose of risk characterization utilizing the most sensitive toxic endpoint. In addition, there was no apparent concern for mutagenicity, and no s tructural analogs of concern were identified.

With regard to acute toxicity, technical tribufos is placed in Toxicity Category II by the oral and dermal routes and Category III by the inhalation route. Dermal and eye irritation response is assigned to Toxicity Category IV. Tribufos is not a dermal sensitizer. In an acute neurotoxicity study, a single oral administration to rat s resulted in signs of neurotoxicity (decreased motor activity) and inhibition of plasma and RBC ChE activ ity in both sexes at the same dose

.

4.3.2 Critical Durations of Exposure

One of the key elements in risk assessment is the appropriate integration of temporality between the exposure and hazard assessments. One advantage of an AOP understanding is that human health risk assessments can be refined, focused on the most relevant durations of exposure. OPs exhibit a ph enomenon known as steady state ChE inhibition. A fter repeated dosing at the same dose level, the degree of inhibition comes into equilibrium with the production of new, uninhibited enzyme. At this po int, the amount of ChE inhibition at a given dose remains consistent across

duration. In general, OPs reach steady state within 2

- 3 weeks and as such, this is considered a

relevant duration of ex posure.

4.4 Literature Review on Neurodevelopment Effects

For the OPs, historically the Agency has used inhibition of AChE as the POD for human health risk assessment; at present time, this policy continues. This science policy is based on decades

of work which shows that AChE inhibition is the initial event in the pathway to acute cholinergic neurotoxicity. The use of AChE inhibition data for deriving PODs was supported by the FIFRA SAP (2008, 2012) for chlorpyrifos as the most robust source of dose

- response data for extrapolating risk and is the source of data for PODs for tribufos. A detailed review of the epidemiological studies used in this review can be found either in the 2014 chlorpyrifos risk assessment (D424485, D. Drew et al, 12/29/2014) or in the 2015 literature review for other organophosphates (OPP /USEPA; D331251; 9/15/15).

Newer lines of research on OPs in the areas of potential AOPs, in vivo animal studies, and notably epidemiological studies in mothers and children, have raised some uncertainty about the agency’s risk assessment approach with regard to the potential for neurodevelopmental effects in fetuses and children. Many of these studies have been the subject of review by the agency over the last several years as part of efforts to develop a risk assessment for chlorpyrifos (D424485,

D. D rew et al, 12/29/2014). Initially, t he agency focused on studies from three US cohorts: 1)Tribufos Human Health Risk Assessment DP No. D357537

Page 16 of 84 The Mothers and Newborn Study of North Manhattan and South Bronx performed by the Columbia Children’s Center for Environmental Health (CCCEH) at Columbia Universit y; 2) the

Mt. Sinai Inner

- City Toxicants, Child Growth and Development Study or the “Mt. Sinai Child Growth and Development Study;” and 3) the Center for Health Assessment of Mothers and Children of Salinas Valley (CHAMACOS) conducted by researchers at Uni versity of California Berkeley. The agency has evaluated these studies and sought external peer review (FIFRA SAP reviews in 2008 and 2012; federal panel, 2013 2 ) and concludes they are of high quality. In the 3 US cohort epidemiology studies, mother

- infan t pairs were recruited for the purpose of studying the potential health effects of environmental exposures during pregnancy on subsequent child development. Each of these cohorts evaluated the association between prenatal chlorpyrifos and/or OP exposure (w ith adverse neurodevelopmental outcomes in children through age 7 years. For the 2014 chlorpyrifos revised human health risk assessment (D424485, D. Drew et al, 12/29/ 2014) , EPA included epidemiologic research results from these three US prospective

bir th cohort studies but primarily focused on the results of CCCEH since this cohort has published studies on the association between cord blood levels of chlorpyrifos and neurodevelopmental outcomes. The agency retained the FQPA 10X Safety Factor (SF) in the 2014 chlorpyrifos revised risk assessment, in large part, based on the findings of these studies.

In the 2015 updated literature review (OPP/USEPA; D331251; 9/15/15), the agency conducted a systematic review expanding the scope of the 2012/2014 review fo cused on US cohort studies with particular emphasis on chlorpyrifos. The expanded 2015 review includes consideration of the epidemiological data on any OP pesticide, studies designs besides prospective cohort studies,

and non

- U.S. based studies. The updat ed literature review identified eight studies which were

relevant (Bouchard et al., 2010; Fortenberry et al., 2014; Fortenberry et al., 2014a; Furlong et al., 2014; Guodong et al., 2012; Oulhote et al., 2013; Zhang et al., 2014; Shelton et al., 2014).

Th ese eight studies have been evaluated in context with studies from the 2012/2014 review ( D424485, D. Drew et al, 12/29/2014 ). Only a brief summary is provided below.

The OP exposure being assessed in many of these studies used concentrations of urinary d ialkyl phosphate metabolites (DAPs) as the urinary biomarker. Total DAPs is a non

- specific measure of OP exposure and is the sum of six separate molecules

- three dimethyl alkylphosphate (DMAP) molecules of DMP, DMTP, DMDTP, and three diethyl alkylphospha te (DEAP) molecules of DEP, DETP, and DEDTP. Each metabolite is a breakdown product from multiple OPs (Table 4.4.1; CDC, 2008) 3 . Specifically, DMP, DMTP, and DMDTP are associated with 18, 13, and 5 OPs, whereas DEP, DETP, and DEDTP are associated with 10 , 10, and 4 OPs, respectively. Thus, using urinary DAPs alone as an exposure measure, it is not possible to separate the exposure and associated effects for single, specific OPs.

Table 4.4.1.CDC Table of organophosphate pesticides and their dialkyl pho sphate metabolites (2008).

Pesticide DMP DMTP DMDTP DEP DETP DEDTP

Azinphos methyl X X X
Chlorethoxyphos X X Chlorpyrifos X X

2 http://www.regulations.gov/#!documentDetail;D=EPA

- HQ

- OPP

- 2008

- 0850

- 0170

3 http://www.cdc.gov/nchs/data/nhanes/nhanes_03_04/l26opd_c_met_organophosphorus_pesticides.pdfTribufos Human Health Risk Assessment DP No. D357537

Page 17 of 84 Chlorpyrifos methyl X X Coumaphos X X
Dichlorvos (DDVP) X

Diazinon X X Dicrotophos X

Dimethoate X X X

Disulfoton X X X

Ethion X X X

Fenitrothion X X

Fenthion X X Isazaphos

- methyl X X

Malathion X X X

Methidathion X X X Methyl parathion X X

Naled X Oxydemeton

- methyl X X

Parathion X X

Phorate X X X

Phosmet X X X Pirimiphos

- methyl X X Sulfotepp X X Temephos X X

Terbufos X X X Tetrachlorviphos X

Trichlorfon X
DMP = dimethylphosphate; DEP = diethylphosphate; DMTP = dimethylthiophosphate; DMDTP = dimethyldithiophosphate; DETP = diethylthiophosphate; DEDTP = diethyldithiophosphate.

For studies which measured urinary 3,5,6

- trichloro

- 2

- pyridinol ( TCPy) (e.g., Fortenberry et al, 2014; Eskenazi et al, 2007; Whyatt et al, 2009), this metabolite can be d erived from chlorpyrifos, chlorpyrifos

- methyl, and the herbicide triclopyr. TCPy is also the primary environmental degradate of chlorpyrifos, chlorpyrifos

- methyl, and triclopyr; thus exposure can be found directly on food treated with these pesticides. C CCEH studies have largely used chlorpyrifos measured in cord blood as the specific biomarker (e.g., Lovasi et al, 2010; Whyatt et al, 2004; Rauh et al, 2011). The CHARGE study (Shelton et al, 2015) did not measure biomarkers but instead used geospatial an alysis to focus on the residential proximity to OP exposure using data from the California Department of Pesticide Regulation, with five OPs accounting for a total of 73% of the pesticide applied near residential settings (chlorpyrifos, acephate, diazinon, bensulide, and dimethoate).

Similarly, DAPs can be found directly on food following OP applications (Zhang et al, 2008; Chen et al, 2012). Specifically, studies have shown that DAPs may form as environmental degradates abiotic hydrolysis, photolysis, and plant metabolism (Zhang et al, 2008; Chen et al, 2012; Racke et al, 1994). Furthermore, since these DAPs are excreted more rapidly and extensively than the parent OPs (Zhang et al, 2008; Forsberg et al, 2008), direct exposure to DAPs may lead to an ov erestimate of OP exposure when using urinary DAPs as a biomarker ofTribufos Human Health Risk Assessment DP No. D357537

Page 18 of 84 OP exposure. The agency recognizes that this is a source of uncertainty when using DAPs for assessing OP exposure and will continue to monitor this issue in future assessments.

With re spect to neurological effects near birth, the CHAMACOS and Mt. Sinai cohorts measured neurological effects at birth (the Brazelton index), and observed a putative association with total DEAP, total DMAP, and total DAP exposure (Engel et al., 2007; Young et al., 2005) . Similarly, a Chinese study (Zhang et al., 20 14) reported statistically significant associations between for total DEAPs, total DMAPs, and total DAPs prenatal OP pesticide exposure and neonatal neurodevelopment assessed 3 days after birth. However, another cross

- sectional Chinese study, Guodong et a l (2012), observed no association with these exposures and a developmental quotient score for 23

- 25 month old children.

The 3 US cohorts (CCCEH, Mt. Sinai, CHAMACOS) each reported evidence of impaired mental and psychomotor development, albeit not consist ent by age at time of testing (ranging from 6 month to 36 months across the three cohorts). Attentional problems and ADHD were reported by three prospective cohorts [Rauh et al, 2006; Eskenazi et al., 2007; Marks et al, 2010 ; and Fortenberry et al (2014, 2014a)] investigators with additional support from a case control study, Bouchard et al. (2010). The exposure metric varied among these studies. Specifically, Fortenberry et al (2014) found suggestive evidence of an association with TCPy and ADHD in boys whereas statistically significant associations were observed by Rauh et al (2006) with chlorpyrifos exposure and ADHD

. Eskenazi et al (2007) reported associated with total DMAPs and total DAPs and ADHD; Marks et al (2010) reported associations with total DEAP, DMAP, and total DAP exposure and ADHD. In a national cross

- sectional study of Canadian children

2007

- 2009 data for age 6

- 11 years (Oulhote et al., 2013), there were no overall statistically significant associations observed between child urinary DE AP, DMAP, or total DAP metabolite levels and parentally reported behavioral problems. In contrast, Bouchard et al. (2010), looking at U.S. children age 8

- 15 years in the 2000

- 2004 National Health and Nutrition Examination Survey (NHANES), observed a posit ive association between attention and behavior problems and total DAPs and DMAPs, but not DEAPs. As part of their analysis, Oulhote et al (2013) noted that their outcome assessment for behavioral problems may not have been as sensitive as Bouchard et al ( 2010), which may in part account for the difference in the observed results from

these studies.

In addition, the 3 US cohorts and the CHARGE study have reported suggestive or positive associations between OP exposure and autism spectrum disorders (Rauh et al., 2006; Shelton et al., 2014; Eskenazi et al, 2007; Furlong et al., 2014). Specifically, Fur long et al (2014) documented suggestive evidence of an association between total DEAP exposure and reciprocal social responsiveness among blacks and boys. Eskenazi et al (2007) reported a statistically significant association between pervasive development al disorder (PDD) and total DAP exposure, whereas Eskenazi et al (2010) reported non

- significant, but suggestive, increased odds of PDD of 2.0 (0.8 to 5.1; p=0.14). Rauh et al (2006) documented a significant association between PDD and specifically chlorp yrifos exposure. Both PDD and reciprocal social responsiveness are related to the autism spectrum disorder. Using a different exposure assessment method (geospatial analysis and residential proximity to total OP exposure), Shelton et al (2014) also showe d statistically significant associations between total OP exposure and ASD. While these studies vary in the magnitude of the overall strength of association, they haveTribufos Human Health Risk Assessment DP No. D357537

Page 19 of 84 consistently observed a positive association between OP exposure and ASD. Finally, CCC EH, Mt. Sinai, CHAMACOS have reported an inverse relation between the respective prenatal measures of chlorpyrifos and intelligence measures at age 7 years (Rauh et al. ,2011; Engel et al.,
2011; Bouchard et al., 2011 )

.

Across the epidemiology database of studies, the maternal urine, cord blood, and other (meco nium) measures provide evidence that exposure did occur to the fetus during gestation but the actual level of such exposure during the critical window(s) of susceptibility is not known. While significant uncertainties remain about the actual exposure leve ls experienced by mothers and infant participants in the children’s health cohorts, it is unlikely that these exposures resulted in AChE inhibition. As part of the CHAMACOS study, Eskenazi et al. (2004) measured AChE activity and showed that no difference s in AChE activity were observed. The biomarker data (chlorpyrifos) from the Columbia University studies are supported by the agency’s dose reconstruction analysis using the PBPK

- PD model. Following the recommendation of the FIFRA SAP (2012), the agency conducted a dose reconstruction analysis of residential uses available prior to 2000 for pregnant women and young children inside the home. The PBPK

- PD model results indicate for the highest exposure considered (i.e., indoor broadcast use of a 1% chlorpyr ifos formulation) <1% RBC AChE inhibition was produced in pregnant women. While uncertainty exists as to actual OP exposure at (unknown) critical windows of exposure, EPA believes it is unlikely individuals in the epidemiology studies experienced RBC AChE inhibition.

A review of the scientific literature on potential modes of action/adverse outcome pathways (MOA/AOP) 4 leading to effects on the developing brain was conducted for the 2012 FIFRA SAP meeting (USEPA, 2012) and updated for the December 2014 chl orpyrifos revised risk assessment (D424485, D. Drew et al, 12/29/2014). In short, multiple biologically plausible hypotheses and pathways are being pursued by researchers that include targets other than AChE inhibition, including cholinergic and non

- choli nergic systems, signaling pathways, proteins, and others. However, no one pathway has sufficient data to be considered more credible than the

others. The fact that there are, however, sparse AOP data to support the in vitro to in vivo extrapolation, or t he extrapolation from biological perturbation to adverse consequence significantly limits their quantitative use in risk assessment. The SAP concurred with the agency in 2008 and 2012 about the lack of definable key events in a MOA/AOP leading to developm ental neurobehavioral effects. However, since the 2014 literature review, there are no substantive changes in the ability to define and quantitate steps in an MOA/AOP leading from exposure to effects on the developing brain. Published and submitted guide line DNT laboratory animal studies have been reviewed for OPs as part of the 2012/2014 review (D424485, D. Drew et al, 12/29/2014) and the updated 2015 review (OPP/USEPA; D331251; 9/15/15). Neurobehavioral alterations in laboratory animals were often repo rted, albeit at AChE inhibiting doses, but there was generally a lack of consistency in terms of pattern, timing, or dose

- response for these effects, and a number of studies were of lower quality. However, this information does provide evidence of long

- la sting neurodevelopmental disorders in rats and mice following gestational exposure.

4 Mode of action (MOA) and adverse outcome pathways (AOPs) describe a set of measureable key events that make up the biological processes leading to an adverse outcome and the causal linkages between such events.Tribufos Human Health Risk Assessment DP No. D357537

Page 20 of 84 At this time, a MOA(s)/AOP(s) has/have not been established for neurodevelopmental outcomes. This growing body of literature does demonstrate, however, that OPs are biologically active on a number of processes that affect the developing brain. Moreover, there is a large body of in vivo laboratory studies which show long

- term behavioral effects from early life exposure, albeit at doses which cause AChE inhibition. EPA considers the results of the toxicological studies relevant to the human population, as qualitatively supported by the results of epidemiology studies. The agency acknowledges the lack of established MOA/AOP pathway and uncertainties associated lack of ability to make strong causal linkages and unknown window(s) of susceptibility. These unce rtainties do not undermine or reduce the confidence in the findings of the epidemiology studies. The epidemiology studies reviewed in the 2012/2014 and 2015 literature reviews represent different investigators, locations, points in time, exposure assessme nt procedures, and outcome measurements. Despite all these differences in study design, with the exception of two negative studies in the 2015 literature review (Guodong et al, 2012; Oulhote et al, 2013), authors have identified associations with neurodev elopmental outcomes associated with OP exposure across four cohorts and twelve study citations. Specifically, there is evidence of delays in mental development in infants (24

- 36 months), attention problems and autism spectrum disorder in early childhood, a nd intelligence decrements in school age children who were exposed to OPs during gestation. Investigators reported strong measures of statistical association across several of these evaluations (odds ratios 2

- 4 fold increased in some instances),

and observ ed evidence of exposures

- response trends in some instances, e.g. , intelligence

measures.

As section 408(b)(2)(C) of the FFDCA instructs EPA, in making its “reasonable certainty of no harm” finding, that in “the case of threshold effects, an additional ten fold margin of safety for the pesticide chemical residue and other sources of exposure shall be applied for infants and children

to take into account potential pre

- and postnatal toxicity and completeness of data with respect to exposure and toxicity to in fants and children .” Section 408 (b)(2)(C) further states that “the Administrator may use a different margin of safety for the pesticide chemical residue only if, on the basis of reliable data, such margin will be safe for infants and children.” Given the totality of the evidence, there is sufficient uncertainty in the human dose

- response relationship for neurodevelopmental effects which prevents the agency from reducing or removing the statutory 10X FQPA Safety Factor. For the tribufos DRA, a value of 10 X has been applied. Similarly, a database uncertainty factor of 10X will be retained for occupational risk assessments. The agency will continue to evaluate the epidemiology studies and pursue approaches for quantitative

or semi

- quantitative comparisons between doses which elicit AChE inhibition and those which are associated with neurodevelopmental outcomes prior to a revised human health risk

assessment.

4.5 Safety Factor for Infants and Children (FQPA SF)

As noted above, the lack of an established MOA/AOP makes quantitative use of the epidemiology studies in risk assessment challenging, particularly with respect to determining

dose

- response, critical duration of exposure, and window(s) of susceptibility. However, exposure levels in the range measu red in the epidemiology studies are likely low enough that they are unlikely to result in AChE inhibition. Epidemiology studies consistently identified associations with neurodevelopmental outcomes associated with OP exposure such as delays inTribufos Human Health Risk Assessment DP No. D357537

Page 21 of 84 mental dev elopment in infants (24

- 36 months), attention problems and autism spectrum disorder in early childhood, and intelligence decrements in school age children. Therefore, there is a need to protect children from exposures that may cause these effects; this ne ed prevents the agency from reducing or removing the statutory FQPA Safety Factor. Thus, the FQPA 10X Safety Factor will be retained for tribufos for the population subgroups that include infants, children, youths, and women of childbearing age for all ex posure scenarios.

4.5 .1 Completeness of the Toxicology Database

The database of toxicology studies for tribufos is complete and adequate to assess potential risk to

infants and children. It consists of developmental studies in rat and rabbit, a developmental neurotoxicity study, a reproductive toxicity study, dermal and inhalation subchronic studies, and

oral comparative ChE stud ies with three components (acute, repeated, gestational).

As discussed in Section 4.4, there is uncertainty in the human dose

- response relationship for neurodevelopmental effects and this warrants retention of the FQPA Safety Factor for the population subgroups that include infants, children, youths, and women of childbearing age for all exposure scenarios.

4.5 .2 Evi dence of Neurotoxicity

Tribufos is an OP with a neurotoxic AOP ; neurotoxicity is the most sensitive effect in all species, routes, and life stages . Consequently, ChE inhibition, as an indicator of neurotoxicity, is being used

to derive the PoDs

. CNS neuro pathy was observed in hens and retinal damage was observed in the chronic rat studies but these effects occurred at doses much higher than the PoDs.

4. 5 .3 Evidence of Sensitivity/Susceptibility in the Developing or Young Animal

A reproductive toxicity st udy and a developmental neurotoxicity (DNT) study in rats as well as

r at and rabbit developmental studies are available and do not show any quantitative susceptibility . The CCA studies are available and provide ChE dose

- response data from potentially susc eptible lifestages (i.e., fetuses, pregnant dams, post

- natal pups). Q uantitative susceptibility was observed in pups compared to adults in the acute comparative ChE (CCA)

st udies but no quantitative or qualitative susceptibility was observed in repeated

- dosing studies

.

As discussed in Section 4.4, there is uncertainty in the human dose

- response relationship for neurodevelopmental effects and this warrants retention of the FQPA Safety Factor for the population subgroups that include infants, ch ildren, youths, and women of childbearing age for all exposure scenarios.

4. 5 .4 Residual Uncertainty in the Exposure Database

There are no residual uncertainties in the exposure database. Since the dietary and non

- dietary exposure estimates were based on several conservative assumptions, HED does not believe the exposure assessment underestimates risk. These assumptions include tolerance

- level residues and 100% crop treated estimates for all commodities. Furthermore, conservative, upper

- boundTribufos Human Health Risk Assessment DP No. D357537

Page 22 of 84 assumptions were used to determine exposure through drinking water and residential sources, such that these exposures have not been under estimated.

4. 6 Toxicity Endpoint and Point of Departure Selections

4. 6 .1 Dose

- Response Assessment

Although multiple datasets provided a poor fit for BMD modeling (see appendix 2), sufficient information was available for endpoint selection. Table 4.5. 4.1 summarizes the tribufos toxicity endpoints and PoDs selected from an evaluation of the database. This endpoint selection was based on a weight of the evidence evaluation using the following considerations:

 Relative sensitivity of the brain and RBC com partments : For tribu fos, across most studies, durations, life stages, and routes, the RBC compartment tend s to be more sensitive than the brain. For the OP cumulative risk assessment , OPP used brain data in

P o D derivations as these data represent ed t he target tissue and tend ed to be less variable

than RBC data. In the case of tribufos; however, the brain data in general did not model

well because often there was no dose

- response . In cases where brain data did model, RBC data provided more protective endpoints.

 Potentially susceptible populations (fetuses, juveniles, pregnancy ): The available ChE data across multiple lifestages (adults, pregnant adults, fetuses, juveniles) show evidence of quantitative life stage sensitivity following acute exposur e . This finding has driven the selection of the PND 11 pup RBC BMD L s to derive the PoDs for the acute dietary endpoint

.

 Route of exposure: It is preferred to match, to the degree possible, the route of exposure in the toxicity study with that of the expo sure scenario(s) of interest . In the case of

tribufos , there are oral , dermal, and inhalation studies which contain adequate dose

response ChE data.

 Duration of exposure: It is preferred to match, to the degree possible, the duration of toxicity study w ith that of the exposure duration of interest. In the case of tribufos , there are single day and steady

- state oral studies as well as steady

- state dermal and inhalation

studies

.

 Consistency across studies: In cases where multiple datasets are available for a single duration, it is important to evaluate the extent to which data are consistent (or not) across

studies. Although there were limited data sets that modeled, t he tribufos database has generally consisten t data across studies which allows the P o D s to be derived from several

critical studies.

De scription s of the primary toxicity studies used for selecting toxicity endpoints and PoDs for various exposure scenarios are presented in A ppendix 3 of this document. Summary tables of BMD analyses can be found in Appendix 2 and the technical details of the analysis can be found

in the BMD memo ( R. Bever and E. Holman

, D423990, 6/17/ 201 5 ).Tribufos Human Health Risk Assessment DP No. D357537

Page 23 of 84 Consistent with risk assessments for other ChE

- inhibitin g compounds, OPP has used a benchmark response (BMR) level of 10% and has thus calculated BMD 10 s and BMDL 10 s (See Appendix 1 for summary of OPP’s ChE policy). The BMD 10 is the estimated dose where ChE is inhibited by 10% compared to background. The BMDL 1 0 is the lower confidence bound on the

BMD 10 value

. As a matter of science policy, the agency uses the BMDL, not the BMD, for

selection of the P o D s (USEPA, 2012). All BMD/BMDL modeling for all individual datasets was completed using USEPA BMD S oftware, ve rsion 2.4 ; an exponential model was used to fit the data, with the assumption of constant variance across each dataset . BMD results from the OP Cumulative Risk Assessment ( CRA 2002, 2006) were included in the endpoint selection weight

of evidence evaluati on.

Acute Dietary ( all populations, including females 13

- 49 )

As shown in Table 4.5.4.1, results of the acute CCA study (MRID # 48707704 ) with tribufos provide the best dataset for this exposure scenario (RBC ChE data in the PND 11 male pups).

The BMDL 10 of 1.05 mg/kg /day associated with RBC ChE inhibition was selected as a suitable PoD for the acute dietary (all populations) exposure scenario. The corresponding BMD 10 was 1.41 mg/kg /day

.

RBC cholinesterase inhibition data were selected for the Po D because it provided the best fit for BMD modeling and RBC cholinesterase appear s to be more sensitive than the brain ChE

. Findings from the PND 11 pups represent the highly exposed sub

- populations (infants and young children) and are, thus, appropriate f or the PoD derivation.

An uncertainty factor of 100X (10X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10 to obtain an aRfD of 0.01 mg/kg /day . Based on the FQPA safety factor of 1 0 X due to uncerta inty in the human dose

- response relationship for neurodevelopmental effects (See Section 4.4)

, the acute population adjusted dose (aPAD) is

0. 0 01 mg/kg /day , except for the population subgroup of adults 50

- 99 where the FQPA SF does not apply (aPAD for this subgroup = 0.01 mg/kg/day)

.

Steady

- State Dietary (all populations)

Table 4.3.2.1 also shows that the PoD for the steady

- state dietary (all populations) scenario was derived from the subchronic oral neurotoxicity study (MRID 45369101 ), based on RBC ChE

inhibition in adult female rat s. The BMDL 10 of 0.19 mg/kg/day associated with RBC ChE was selected as a suitable PoD for the steady

- state dietary exposure assessment (all populations). No difference in sensitivity to RBC ChE inhibition was observed between pups and adult animals following repeated exposure, thus the selected endpoint is protective of all populations. The corresponding BMD 10 was 0.27 mg/kg/day; this value represents the lowest BMD in the

database.

An uncertainty facto r of 100X (10X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10 to obtain an ss RfD of 0.002 mg/kg/day. Based on the FQPA safety factor of 1 0 X due to uncertainty in the human dose

- response relationship forTribufos Human Health Risk Assessment DP No. D357537

Page 24 of 84 neurodevelopmental effects (See Section 4.4) , the steady

- state PAD (ssPAD) is 0. 0 002

m g/kg/day, except for the population subgroup of adults 50

- 99 where the FQPA SF does not apply (aPAD for this subgroup = 0.01 mg/kg/day)

.

Incidental Oral, Steady Sta te

For the purpose of assessing potential risk associated with incidental oral exposure from steady state durations resulting from spray drift, HED selected the subchronic oral neurotoxicity study (MRID # 45369101), based on RBC ChE inhibition in adult fem ale rat s . Quantitation of incidental oral risks was performed using the BMDL 10 value of 0.19 mg/kg /day and the BMD 10 of 0.27 mg/kg /day

. Because no difference in sensitivity was observed between pups and adult rats in repeated

- dosing studies (see table A.5), these values are considered protective of the sub

populations of interest (infants, toddlers, young children) for incidental oral exposure. An uncertainty factor of 1 0 00X (10X to account for interspecies extrapolation

, 10X for intraspecies

variation , and a 10X FQPA SF ) is applied for incidental oral exposure.

Dermal, Steady

- State

A 21

- day repeated dose dermal study in rabbits (MRID # 42007201) that measured parameters in the functional observational battery (FOB) as well as ChE inhibition was th e only study available for the dermal risk assessment. Consequently, the PoD for the steady

- state exposure scenario was selected from this study and was based on RBC ChE inhibition (BMDL 10 = 0.16 mg/kg/day)

in adult female rats. An uncertainty factor of 1 00 0 X (10X to account for interspecies extrapolation

, 10X for intraspecies variation , and a 1 0 X FQPA SF for residential assessments or a database uncertainty factor in occupational assessments due to uncertainty in the human dose

response relationship for n eurodevelopmental effects (see Section 4.4) ) is applied for dermal

exposure.

Inhalation , Single Day and Steady

- State

The PoD for the inhalation steady

- state exposure scenario was selected from the 13

- week inhalation study in rats (MRID # 4 2399801) based on RBC ChE inhibition in adult male rats (BMDL 10 = 0. 79 mg/m 3 /day; BMD 10 = 1.00 mg/m 3 /day)

. Inhalation dosimetry was applied to the derived PoDs in accordance with EPA guidelines (EPA, 1994) to obtain a human equivalent concentration (HEC) and human equivalent dose (HED). HECs were generated from th is study because exposure was via an aerosol, which resulted in a regional deposited dose ratio ( RDDR )

of 2.926 . HECs were then converted to HEDs, which are 0.1 mg/kg/day for the BMDL 10 and 0.12 mg/kg/day f

or the BMD, respectively. For steady state inhalation exposures, a total

uncertaint y factor of 30 0 X was applied [3 X for interspecies extrapolation, 10X for intraspecies

variation and a 1 0 X FQPA SF for residential assessments or a database uncertainty facto r in occupational assessments due to uncertainty in the human dose

- response relationship for neurodevelopmental effects (see Section 4.4) ].

4. 6 .2 Recommendation for Combining Routes of Exposures for Risk Assessment

P o Ds for the oral, dermal, and inhalation routes are all derived from RBC ChE inhibition. Therefore, there is a common endpoint and exposures can be combined

.Tribufos Human Health Risk Assessment DP No. D357537

Page 25 of 84 4. 6 .3 Cancer Classification and Risk Assessment Recommendation

The HED Cancer Peer Review Committee (CPRC) [now designated as Cancer Assessment Review Committee (CARC)] classified tribufos as “likely” at high doses but “unlikely” to be a human carcinogen at low doses since all tumor increases in mice occurred only at the highest dose tested (48 mg/kg/day in males and 63 mg/kg/day in females) and were accompanied by severe ChE inhibition and increased mortality in the later weeks of the study (TXR# 0012242, E. Rinde and R. Zendzian, 22

- MAY

- 1997). The CPRC concluded th at a low

- dose linear extrapolation risk assessment based on the tumors is not recommended because of the severe accompanying toxicity ( ChE inhibition) typical of OPs, which occurred at all doses tested in mice. Therefore, a non

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

4. 6 .4 Summary of Points of Departure and Toxicity Endpoints Used in Human Risk

Assessment

Table 4.5.4.1 Summary of Toxicological Dos es and Endpoints and Points of Departure

for Tribuf

os for Use in Dietary, Non

- Dietary and Occupational Human Health Risk

Assessments

Exposure

Scenario Point of

Departure (mg/kg/day) Uncertainty/FQPA

Safety Factors a RfD, PAD, Level of

Concern for Risk

Assessment Study and Toxicological

Effects

Acute Dietary (all populations, including

females 13

- 49) BMDL 10 = 1.05

mg/kg/day UF A = 10x

UF H = 10x FQPA SF = 1 0 x Acute RfD = 0.01

mg/kg/day

Acute PAD = 0.001 mg/kg/day Acute CCA Study (MRID 48707704) in

the rat

– PND 11

BMD 10 = 1.41 mg/kg/day

Inhibition of RBC ChE

in male rat pups.

Steady

- State Dietary (all populations) BMDL 10 = 0.19

mg/kg/day UF A = 10x

UF H = 10x FQPA SF = 1 0 x Steady State RfD =

0.002 mg/kg/day

Steady State PAD =

0.0002 mg/kg/day Subchronic oral neurotoxicity study (MIRD 45369101)

BMD 10 = 0.27 mg/kg/day

Inhibition of RBC ChE

in adult female rat s

Incidental Oral Steady State BMDL 10 = 0.19

mg/kg/day UF A = 10x

UF H = 10x FQPA SF = 1 0 x LOC for MOE = 1000 Subchronic oral

neu rotoxicity study (M R I D 45369101)

BMD 10 = 0.27 mg/kg/dayTribufos Human Health Risk Assessment DP No. D357537

Page 26 of 84 BMD = Benchmark dose. BMDL = Benchmark dose lower confidence limit. HED = Human equivalent dose. 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. LOC = level of concern. MOE = margin of exposure. UF = uncertainty factor. UF A = extrapolation from animal to human (interspecies). UF H = potential variation in sensitivity among members of the human population (intraspecies). UF DB = database uncertainty factor. FQPA SF = FQPA Safety Factor. PAD = population adjusted

dose.

a FQPA SF retained for infants, children, youths, and women of childbearing age for al l exposure scenarios due to uncertainty in the human dose

- response relationship for neurodevelopmental effects (see Section 4.4). This includes all exposure scenarios, except the dietary exposure scenarios for the population subgroup adults 50

- 99 for whic h the FQPA SF has been reduced to 1X (aPAD = 0.01 mg/kg/day; ssPAD = 0.002 mg/kg/day). In occupational dermal and inhalation assessments, a 10X database uncertainty factor is applied due to this uncertainty.

4. 7 Endocrine Disruption

As required by FIFRA and FFDCA, EPA reviews numerous studies to assess potential adverse outcomes from exposure to chemicals. Collectively, these studies include acute, subchronic and chronic toxicity, including assessments of carcinogenicity, neurotoxicity, development al, reproductive, and general or systemic toxicity. These studies include endpoints which may be susceptible to endocrine influence, including effects on endocrine target organ histopathology, organ weights, estrus cyclicity, sexual maturation, fertility, pregnancy rates, reproductive loss, and sex ratios in offspring. For ecological hazard assessments, EPA evaluates acute tests and chronic studies that assess growth, developmental and reproductive effects in different taxonomic groups. As part of its mos t recent registration decision for tribufos, EPA reviewed these data and selected the most sensitive endpoints for relevant risk assessment scenarios from the existing hazard database. However, as required by FFDCA section 408(p ), tribufos is subject

to t he endocrine screening part of the Endocrine Disruptor Screening Program (EDSP). Inhibition of RBC ChE

in adult female rat s

Dermal

S teady State BMDL 10 = 0.16

mg/kg/day UF A = 10x

UF H = 10x FQPA SF /UF DB = 1 0 x LOC for MOE = 1000 21

- day derm al toxicity study (MRID 42007201) in rabbits

BMD 10 = 0.22 mg/kg/day

Inhibition of RBC ChE

in adult female rats

.

Inhalation

S teady State BMDL 10 = 0.79

mg/m 3
(HED = 0.10 mg/kg/day) UF A = 3x

UF H = 10x FQPA SF /UF DB = 1 0 x LOC for MOE = 300 13

- Week inhalati on toxicity study (MRID

42399801 )

BMD 10 = 1.00 mg /m 3
( HED = 0.12 mg/kg/day)

I nhibition of RBC ChE

in adult male rats. Cancer (oral, dermal, inhalation) Classification: "Likely at high doses but unlikely to be carcinogenic to humans at low doses ”Tribufos Human Health Risk Assessment DP No. D357537

Page 27 of 84 EPA has developed the EDSP to determine whether certain substances (including pesticide active and other ingredients) may have an effect in humans or wildlife similar to an effect produced by a “naturally occurring estrogen, or other such endocrine effects as the Administrator may designate.” The EDSP employs a two

- tiered approach to making the statutorily required determinations. Tier 1 consists of a battery of 11 screening assays to identify the potential of a chemical substance to interact with the estrogen, androgen, or thyroid (E, A, or T) hormonal

systems. chemicals that go through Tier 1 screening and are found to have the potential to interact with E, A, or T hormona l systems will proceed to the next stage of the EDSP where EPA will determine which, if any, of the Tier 2 tests are necessary based on the available data. Tier 2 testing is designed to identify any adverse endocrine

- related effects caused by the substance , and

establish a dose

- response relationship between the dose and the E, A, or T effect.

Under FFDCA section 408(p), the Agency must screen all pesticide chemicals. Between October 2009 and February 2010, EPA issued test orders/data call

- ins for the fir st group of 67 chemicals, which contains 58 pesticide active ingredients and 9 inert ingredients. A second list of

che micals identified for EDSP screening was published on June 14, 2013 5 and includes some pesticides s che duled for Registration Review and ch e micals found in water

. Neither of these lists should be construed as a list of known or likely endocrine disruptors. For fur ther information on the status of the EDSP, the policies and procedures, the list s of che micals, future lists, the test guidelines and the Tier 1 screening battery, please visit our website. 6

5.0 Dietary Exposure and Risk Assessment

5.1 Metabolite/Degradate Residue Profile

The qu alitative nature of the residues of tribufos in plants (cottonseed) and livestock has been adequately identified/characterized and is understood. The HED Metabolism Assessment Review Committee (MARC) reviewed the tribufos toxicology and metabolism data and

recommend ed that the residue of concern for both tolerance expression and risk assessment for plants and livestock should consist of tribufos only. In addition, HED previously determined that

tolerances for tribufos residues in eggs and poultry tissue s were not required based on the results of the respective poultry metabolism and f eeding studies (CFR §180.6(a)(3)). These conclusions have not changed for the purpose of registration review.

Table 5.1 . Summary of Metabolites and Degradates of Tribufos

. Matrix Residues included in Risk

Assessment Residues included in Tolerance Expression

Plants Primary Crop

Tribufos Tribufos Rotational Crop

5 See http://www.regulations.gov/#!documentDetail;D=EPA

- HQ

- OPPT

- 2009

- 0477

- 0074 for the final second list of

chemicals.

6 http://www.epa.gov/endo/Tribufos Human Health Risk Assessment DP No. D357537

Page 28 of 84 Table 5.1 . Summary of Metabolites and Degradates of Tribufos

. Matrix Residues included in Risk

Assessment Residues included in Tolerance Expression Poultry 1 Not Required Not Required

Ruminant 1 Tribufos Tribufos

Drinking Water Tribufos Not Applicable

1 Tolerances are no longer needed for livestock commodities.

5 .2 Food Residue Profile

Tribufos is only registered for use on cotton, and dietary exposure to tribufos can occur via residues present in cottonseed oil or as a result of transfer of residues from livestock feed items (cotton gin

- byproducts, cottonseed hulls and cottonseed meal) to meat and milk. Residues are most likely to be surface residues from ground or foliar application on cotton and quantifiable residues in harvested crops. Environmental fate data suggest that residues are persistent in soil.

Based on the animal metabol ism data, residues of tribufos may end up in ruminant tissues and milk at low levels as a result of feeding livestock treated feed items. However, since ruminant s are fed only 5% of cottonseed and cotton gin byproducts, the expected secondary residues are

minimal. Additionally, based on the livestock feeding study and on an updated livestock diet

, there are no reasonable expectation s of finite residues of tribufos in/ on any livestock commodity. Therefore, t he established tolerances for residues of tribufos in/on milk, eggs, cattle, goat, horse, hog, poultry, and sheep commodities (40 CFR §180.272) should all be revoked.

5.3 Water Residue Profile

The estimated drinking water concentration s (EDWCs) for trib ufos residues in surface water were generated using the Surface Water Concentration Calculator (SWCC) computer model, while the Pesticide Root Zone Model for Groundwater (PRZM

- GW) and Screening Concentration in Ground Water (SCI

- GROW) models were used to g enerate groundwater EDWCs. The residue of concern (ROC) for tribufos is limited to the parent compound. Some OPs require metabolic activation to the oxon metabolite. However, tribufos is not one of these OPs and is already in the oxon form.

The labeled maximum annual and single application rate in the U.S. ( except CA and AZ ) is 1.125 lbs a.i./A, allowing only one application per season. To further refine the drinking water assessment in response to a request fr om the HED , EFED utilized average applicat ion rates rather than the maximum application rates that were used in the initial assessment. Additionally, a lower spray drift fraction (13.5%, decreased from 16%) was utilized, per current EFED guidance (USEPA, 2013). Only the scenario (Texas cotton OP ; aerial spray application) which resulted in the maximum surface water EDWC in the prior assessment was modeled for this refinement. In this refinement, as in the initial assessment, EFED assume d that only a single growing season for cotton may occur wit hin a single year and a single application is made (See Table 5. 3 )

.Tribufos Human Health Risk Assessment DP No. D357537

Page 29 of 84 For the acute and steady state assessments, the entire 30

- year distribution of estimated daily concentrations was incorporated into the Pesticide Root Zone Model and Exposure Analysis

Mo deling System (PRZM/EXAMs) and used in the probabilistic analyses. For steady state the

TX cotton maximum and average rate daily time series were recalculated using the 21

- day forward rolling averages. In the 21

- day rolling average distributions, the first data point is the average of days 1

- 21, the second data point is the average of days 2

- 22, the third data point is the average of days 3

- 23, etc.

Table 5.3 . EDWCs for the Maximum and Average Use of Tribufos on Cotton

Drinking Water

Source (Model Used) Use (Rate

Modeled) Maximum Estimated Drinking Water Concentration (EDWC)

Acute ( μg/L ) Chronic ( μg/L ) Cancer ( μg/L )
Maximum EDWC for the Use of Tribufos on Cotton at Maximum Application Rate

Surface water (SWCC; TX

Cotton OP scenario) Cotton

1.125 lbs a.i./A x 1 application/yr;

aerial 7.26 3.34 2.46

Groundwater (PRZM

- GW) Cotton

1. 87 5 lbs a.i./A x 1 application/yr 1.83E

- 09 1.8E

- 10  ̄

Groundwater (SCI

- GROW) Cotton

1. 87 5 lbs a.i./A x 1 application/yr 1.12E

- 02 1.12E

- 02  ̄

Surface Water EDWCs for the Aerial Use of Tribufos on Cotton at the Average Application Rate

Surface water (SWCC; TX

Cotton OP scenario) Cotton

0.54 lbs a.i./A x 1 application/yr;

aerial 3.36 1.56 1.15

Surface water (SWCC; CA Cotton w/irrig scenario) Cotton

0.64 lbs a.i./A x 1 application/yr;

aerial 1.36 2 0.90 0.67

1 While the higher application rate for CA and AZ was also modeled, the lower ra t e modeled using the TX scenario resulted in higher surface water EDWCs.

2 Modeled as a single application at the average rate, although the average number of applications for CA is reported as 1.2.

5.4 Dietary Risk Assessment

5.4.1 Description of Residue Data Used in Dietary Assessment

A cute and steady state dietary (food and drinking water) exposure and risk assessments were

conduct ed using the Dietary Exposure Evaluation Model software with the Food Commodity Intake Database (DEEM

- FCID) Version 3.16. This software uses 2003

- 2008 food consumption

data from the USDA’s NHANES/W WEIA. The drinking water estimates provided by EFED are modeled estimates, and were generated using the most conservative model available for the existing uses. The labeled maximum annual and average application rate s for cotton wereTribufos Human Health Risk Assessment DP No. D357537

Page 30 of 84 considered as a refin ement to the initial assessment

. The partially refined assessments assumed anticipated residues (average residue in cottonseed oil )

, an empirical processing factor for

cottonseed oil , and 100% crop treated (PCT) for the acute assessment and steady state

a ssessments.

5.4.2 Percent Crop Treated Used in Dietary Assessment

Percent crop treated information were not used in these assessments.

5.4.3 Acute Dietary Risk Assessment

The acute aggregate (food and drinking water) dietary exposure estimates are below HED’s level

of concern ( i.e., <100% of the aPAD) for the U.S. population and all population subgroups. Combined dietary exposure from cottonseed oil and drinking water concentrations based on

maximum rates in TX at the 95 th percentile of exposure is 15 % of the aPAD for the U.S. population and 48 % of the aPAD for all infants (<1 yr old), the most highly exposed population subgroup. Also, the combined dietary exposure from cottonseed oil and drinking water concentrations based on “average” rates in TX at the 95 th percentile of exposure is 7% of the aPAD for the U.S. population and 22% of the aPAD for all infants (<1 yr old), the most highly exposed population subgroup. The food only exposure contributes <1 for and drinking water only contributes 48% of the aPAD for all infants. The summary results for the general population and all population subgroups are shown b elow in Table s 5. 4.3 and 5.4.4.

Table 5 .4.3 . Results of Acute Dietary Exposure Analysis for Tribufos Food only and Drinking Water only, and Food plus Drinking Water using Maximum Rate EDWCs. Population Subgroup aPAD

(mg/kg/day) 1 Food only Drinking Water only (Maximum Rate) Food plus Drinking Water (Maximum Rate)

95 th Percentile 95 th Percentile 95 th Percentile Exposure (mg/kg/day) %

aPAD Exposure (mg/kg/day) %

aPAD Exposure

(mg/kg/day) %

aPAD

General U.S. Population 0.001 0.000002 <1 0.000153 15 0.000154 15

All Infants (<1 year old) 0.001 0.000001 <1 0.000476 48 0.000476 48

Children 1

- 2 years old 0.001 0.000005 <1 0.000232 23 0.000234 23

Children 3

- 5 years old 0.001 0.000005 <1 0.000189 19 0.000192 19

Children 6

- 12 years old 0.001 0.000004 <1 0.000141 14 0.000143 14

Youth 13

- 19 years old 0.001 0.000002 <1 0.000123 12 0.000124 12

Adults 20

- 49 years old 0.001 0.000002 <1 0.000151 15 0.000151 2

Adults 50

- 99 years old 0.01 0.000001 <1 0.000138 1 0.000139 14Tribufos Human Health Risk Assessment DP No. D357537

Page 31 of 84 Females 13

- 49 years old 0.001 0.000002 <1 0.000153 15 0.000154 15

1 Highest exposure at the 95th percentile is in bold.

2 mkd = mg/kg/day. aPAD = acute population

- adjusted dose.

Table 5.4.4 . Results of Acute Dietary Exposure Analysis for Tribufos Food only and Drinking Water only, and Food plus Drinking Water using Average Rate EDWCs. Population Subgroup aPAD

(mg/kg/day) 1 Food only Drinking Water only ( Average Rate) Food plus Drinking

Water ( Average Rate)

95 th Percentile 95 th Percentile 95 th Percentile Exposure (mg/kg/day) %

aPAD Exposure (mg/kg/day) %

aPAD Exposure

(mg/kg/day) %

aPAD

General U.S. Population 0.001 0.000002 <1 0.000071 7 0.000072 7

All Infants (<1 year old) 0.001 0.000001 <1 0.000222 22 0.000222 22

Children 1

- 2 years old 0.001 0.000005 <1 0.000108 11 0.000111 11

Children 3

- 5 years old 0.001 0.000005 <1 0.000088 9 0.000091 9

Children 6

- 12 years old 0.001 0.000004 <1 0.000066 7 0.000068 7

Youth 13

- 19 years old 0.001 0.000002 <1 0.000057 6 0.000058 6

Adults 20

- 49 years old 0.001 0.000002 <1 0.000070 7 0.000071 7

Adults 50

- 99 years old 0.01 0.000001 <1 0.000064 <1 0.000065 <1

Females 13

- 49 years old 0.001 0.000002 <1 0.000071 7 0.000072 7

1 Highest exposure at the 95th percentile is in bold.

2 mkd = mg/kg/day. aPAD = acute population

- adjusted dose.

5.4.4 Steady State Dietary Risk Assessment

The DEEM acute module was used to conduct the steady state assessment. 2

- day average food residues, representing average food exposure, and 21

- day forward

- rolling water averages were compared to the steady state (ss) point of departure

. The 2

- day average dietary exposure assessment incorporated anticipated residues for cotton seed oil , empirical processing factors for

cottonseed oil , and 100 percent crop treated (CT).

Combined dietary exposure from cottonseed oil and drinking water concentrations based on

maximum rates in TX at the 95 th percentil e of exposure is 72 % of the ssPAD for the U.S. population and 226 % of the ssPAD for all infants (<1 yr old) , the most hig hly exposed population subgroup

. The combined dietary exposure from cottonseed oil and TX cotton “average” rate for drinking water at the 95 th percentile of exposure is 34 % of the ssPAD for theTribufos Human Health Risk Assessment DP No. D357537

Page 32 of 84 U.S. population and 105 % of the ssPAD for all infants (<1 yr old) , the most hig hly exposed population subgroup

. The food only exposure contributes <2% of the ssPAD and drinking water only contribu tes 226 % of the ssPAD for all infants. The summary results for the general population and all population subgroups are shown below in Table 5.4.5 and 5.4.6.

Table 5.4.5 . Results of Steady

- State Dietary Exposure Analysis for Terbufos Food only, Drinking

Water only, and Food plus Drinking Water

- using Maximum Rate EDWCs. Population Subgroup ssPAD

(mg/kg/day) 1 Food only Drinking Water only (Maximum Rate) Food plus Drinking Water (Maximum Rate)

95 th Percentile 95 th Percentile 95 th Percentile Exposure (mg/kg/day) %

ssPAD Exposure (mg/kg/day) %

ssPAD Exposure

(mg/kg/day) %

ssPAD

General

U.S. Population 0.0002 0.000002 <1 0.000143 72 0.000144 72

All Infants (<1 year old) 0.0002 0.000001 <1 0.000451 226 0.000452 226

Children 1

- 2 years old 0.0002 0.000005 2 0.000216 108 0.000218 109

Children 3

- 5 years old 0.0002 0.000005 2 0.000172 86 0.000175 87

Children 6

- 12 years old 0.0002 0.000003 2 0.000129 64 0.000131 65

Youth 13

- 19 years old 0.0002 0.000002 <1 0.000114 57 0.000115 57

Adults 20

- 49 years old 0.0002 0.000002 <1 0.000141 70 0.000142 71

Adults 50

- 99 years old 0.002 0.000001 <1 0.000132 7 0.000132 7

Females 13

- 49 years old 0.0002 0.000002 <1 0.000144 72 0.000144 72

1 Highest exposure at the 95th percentile is in bold.

2 mkd = mg/kg/day. aPAD = acute population

- adjusted dose.

Table 5.4.6 . Results of Steady

- State Dietary Exposure Analysis for Terbufos Food only, Drinking

Water only, and Food plus Drinking Water

- using Average Rate EDWCs. Population Subgroup ssPAD

(mg/kg/day) 1 Food only Drinking Water only ( Average Rate) Food plus Drinking

Water ( Average Rate)

95 th Percentile 95 th Percentile 95 th Percentile Exposure (mg/kg/day) %

ssPAD Exposure

(mg/kg/day) %

ssPAD Exposure

(mg/kg/day) %

ssPAD

General U.S. Population

0.00001 0.000002 <1 0.000067 33 0.000068 34Tribufos Human Health Risk Assessment DP No. D357537

Page 33 of 84 All Infants (<1 year old)

0.00001 0.000001 <1 0.000211 105 0.000211 105

Children 1

- 2 years old 0.00001 0.000005 2 0.000101 50 0.000103 52

Children 3

- 5 years old 0.00001 0.000005 2 0.000080 40 0.000083 41

Children 6

- 12 years old 0.00001 0.000003 2 0.000060 30 0.000062 31

Youth 13

- 19 years old 0.00001 0.000002 <1 0.000053 27 0.000054 27

Adults 20

- 49 years old 0.00001 0.000002 <1 0.000066 33 0.000066 33

Adults 50

- 99 years old 0.0001 0.000001 <1 0.000061 3 0.000062 3

Females 13

- 49 years old 0.00001 0.000002 <1 0.000067 34 0.000068 34

1 Highest exposure at the 95th percentile is in bold.

2 mkd = mg/kg/day. aPAD = acute population

- adjusted dose.

5.4.5 Cancer Dietary Risk Assessment

Tribufos is classified as “ Likely at high doses, but unlikely to be carcinogenic to humans at low

doses ” (TXR# 0012242, E. Rinde and R. Zendzian, 22

- MAY

- 1997). The steady state dietary exposure and risk estimates are protective of potential carcinogenicity.

6.0 Residential (Non

- Occupational) Exposure/Risk Characterization

There are currently no registered residential uses for tribufos; therefore a residential handler and

post

- application exposure assessment was not performed. However, an assessment was

conducted in consider ation of potential bystander exposures to spray drift following tribufos applications to cotton.

6 .1 Non

- Occupational Bystander Post

- Application Inhalation Exposure and Risk

Est imates

Volatilization of pesticides may be a source of post

- application in halation exposure to individuals nearby pesticide applications. The agency sought expert advice and input on issues related to volatilization of pesticides from its Federal Insecticide, Fungicide, and Rodenticide Act Scientific Advisory Panel (SAP) in Dec ember 2009, and received the SAP’s final report on
March 2, 2010 (http://www.epa.gov/scipoly/SAP/meetings/2009/120109meeting.html). The agency has evaluated the SAP report and has developed a Volatilization Screening Tool and a subsequent Volatilization S creening Analysis (http://www.regulations.gov/#!docketDetail;D=EPA

- HQ

- OPP

- 2014

- 0219). During Registration Review, the agency will utilize this analysis to determine if data (i.e., flux studies, route

- specific inhalation toxicological studies) or further analysis is required for tribufos.Tribufos Human Health Risk Assessment DP No. D357537

Page 34 of 84 6. 2 Non

- Occupational Spray Drift Exposures and Risk Estimates

Off

- target movement of pesticides can occur via many types of pathways and it is governed by a variety of factors. Sprays that are released and do not deposit in the application area end up off

target and can lead to exposures to those it may directly contact. They can also deposit on surfaces where contact with residues can eventually lead to indirect exposures ( e.g. , children playing on lawns where res idues have deposited next to treated fields). The potential risk estimates from these residues can be calculated using drift modeling onto 50 feet wide lawns coupled with methods employed for residential risk assessments for turf products.

The approach to be used for quantitatively incorporating spray drift into risk assessment is based on a premise of compliant applications which, by definition, should not result in direct exposures to individuals because of existing label language and other regulatory re quirements intended to prevent them. 7 Direct exposures would include inhalation of the spray plume or being sprayed directly. Rather, the exposures addressed here are thought to occur indirectly through contact with impacted areas, such as residential la wns, when compliant applications are conducted. Given this premise, exposures for children (1 to 2 years old) and adults who have contact with turf where residues are assumed to have deposited via spray drift thus resulting in an indirect exposure are the focus of this analysis analogous to how exposures to turf products are

considered in risk assessment.

In order to evaluate the drift potential and associated risks, an approach based on drift modeling coupled with techniques used to evaluate residentia l uses of pesticides was utilized. Essentially, a residential turf assessment based on exposure to deposited residues has been completed to address drift from the agricultural applications of tribufos. In the spray drift scenario, the deposited residue va lue was determined based on the amount of spray drift that may occur at varying distances from the edge of the treated field using the AgDrift ( v2.1.1) model and the Residential Exposure Assessment Standard Operating Procedures Addenda 1: Consideration of Spray Drift Policy

. Once the deposited residue values were determined, the remainder of the spray drift assessment was based on the algorithms and input values specified in the recently revised (2012) Standard Operating Procedures For Residential Risk Asse ssment (SOPs)

.

A screening approach was developed based on the use of the AgDrift model in situations where specific label guidance that defines application parameters is not available. 8 AgDrift is appropriate for use only when applications are made by a ircraft, airblast orchard sprayers, and ground boom sprayers. When AgDrift was developed, a series of screening values (i.e., the Tier 1 option) were incorporated into the model and represent each equipment type and use under

varied conditions. The scree ning options specifically recommended in this methodology were selected because they are plausible and represent a reasonable upper bound level of drift for common application methods in agriculture. These screening options are consistent with how spray d rift is considered in a number of ecological risk assessments and in the process used to develop drinking water concentrations used for risk assessment. In all cases, each scenario is to be evaluated unless it is not plausible based on the anticipated use pattern (e.g., herbicides are

7 This approach is consistent with the requirements of th e EPA’s Worker Protection Standard.

8 http://www.agdrift.com/Tribufos Human Health Risk Assessment DP No. D357537

Page 35 of 84 not typically applied to tree canopies) or specific label prohibitions (e.g., aerial applications are not allowed).

In many cases, risks are of concern when the screening level estimates for spray drift are used as

the bas is for the analysis. In order to account for this issue and to provide additional risk management options additional spray drift deposition fractions were also considered. These drift estimates represent plausible options for pesticide labels.

6. 2 .1 Com bined Risk Estimates from Lawn Deposition Adjacent to Applications

The spray drift risk estimates are based on an estimated deposited residue concentration as a result of the screening level agricultural application scenarios. Tribufos is used on cotton and can be applied via ground boom and aerial equipment at a maximum application rate of 1.875 lbs

ai/acre for the states CA and AZ and 1.125 lb ai/acre for other US states. The recommended drift scenario screening level options are listed below:

 Groundboom applications are based on the AgDrift option for high boom height and using very fine to fine spray type using the 90 th percentile results.

 Aerial applications are based on the use of AgDrift Tier 1 aerial option for a fine to medium spray typ e and a series of other parameters which will be described in more detail below (e.g., wind vector assumed to be 10 mph in a downwind direction for entire application/drift event). 9

In addition to the screening level spray drift scenarios described above, additional results are provided using viable drift reduction technologies (DRTs)

. In particular, different spray qualities have been considered as well as the impact of other application conditions (e.g., boom height, use of a helicopter instead of fixed wing aircraft, crop canopy conditions).

For adults, when an endpoint is not sex

- specific (i.e., the endpoints are based on developmental or fetal effects) a body weight of 80 kg is typically used in risk assessment; however, in this case,

a female

- specif

ic body weight of 69 kg was used. While the endpoint of concern, RBC AChE inhibition, is not sex

- specific, the female

- specific body weight was used to protect for pregnant women due to uncertainty in the human dose

- response relationship for neurodevelopme ntal effects (See Section 4.4).

Adult (dermal only) and children 1 to <2 years of age (combined dermal and incidental oral) risk estimates indicate that there are concerns for the indirect tribufos exposure on turf near aerial and ground applications to cotton. Dermal and incidental oral risk estimates for children 1 to <2 years old were combined because the toxicity endpoint for each route of exposure is the inhibition of brain cholinesterase (ChE). The total applicable LOC is 1,000

, so MOEs < 1,000

r epresent risk estimates of concern. Adult dermal risk estimates from indirect exposure to tribufos related to spray drift result in buffers of over 300 feet for groundboom and aerial applications for the application rate s of 1.875 lb ai/A and 1.125 lb ai/ A. Children’s (1 to <2

9 AgDrift allows for consideration of even finer spray patterns characterized as very fine to fine. However, this spray pattern was not selected as the common screening basis since it is used less commonly for most agriculture.Tribufos Human Health Risk Assessment DP No. D357537

Page 36 of 84 year s old) combined dermal and incidental oral risk estimates from indirect exposure to tribufos are of concern at 300 feet for groundboom and aerial applications based on the screening level scenarios for both application rates. A ppropriate DRTs such as changing the spray type/nozzle configuration to coarser spray applications may result in less drift and reduced risk concerns (i.e., higher MOEs) from aerial applications. Similarly, using coarser sprays and lowering boom height fo r ground boom sprayers reduces risk concerns. Spray drift analysis r esults are presented

in Table s 6. 2 .1.1 and 6. 2 .1.2.Tribufos Human Health Risk Assessment DP No. D357537

Page 37 of 84 Tables: 6. 2 .1 .1 : Adult Dermal Spray Drift Analysis for Tribufos

Crop/Rate Group Spray Type/ Nozzle

Configuration Application Rate (lb ai/A) Estimated or

Adjusted

TTR (ug/cm2) At Edge 10 Feet 25 Feet 50 Feet 75 Feet 100 Feet 125 Feet 150 Feet 200 Feet 250 Feet 300 Feet

Dermal MOE Dermal

MOE Dermal

MOE Dermal MOE Dermal

MOE Dermal

MOE Dermal

MOE Dermal

MOE Dermal

MOE Dermal

MOE Dermal

MOE

Aerial Fine to Medium

1.875 0.2084063 1 1 1 2 2 3 3 4 5 6 7

Medium to Coarse 1 1 2 2 3 5 6 7 9 12 15

Coarse to Very Coarse 1 2 2 4 5 7 9 11 15 20 25 Very Fine to Fine 1 1 1 1 1 1 1 1 2 2 2
AT401, M, 10 mph, 37% SD 1 1 1 2 3 3 4 5 6 8 9
WASP, M, 10 mph, 37% SD 1 1 2 2 3 4 5 6 8 9 11

AT401, C, 10 mph, 25% SD 1 1 2 3 4 5 7 8 12 15 18

WASP, C, 10 mph, 25% SD 1 2 2 4 5 7 9 11 15 20 22

AT401, VC, 10 mph, 20% SD 1 2 3 5 6 9 11 14 20 25 33
WASP, VC, 10 mph, 20% SD 1 2 3 5 8 10 14 16 25 28 33

Groundboom High Boom Very fine to Fine 1 2 4 6 8 10 12 14 18 25 28

Low Boom Very fine to Fine 2 6 10 15 20 25 28 33 39 49 65 High Boom Fine to

Medium/Coarse 4 10 15 22 28 33 39 39 49 65 65

Low Boom Fine to

Medium/Coarse 6 16 25 33 39 49 65 65 98 98 98

Aerial Fine to Medium

1.125 0.1250438 1 2 2 3 3 4 5 6 8 10 12

Medium to Coarse 2 2 3 4 6 7 9 11 16 20 25
Coarse to Very Coarse 2 3 4 6 9 12 15 18 25 33 41 Very Fine to Fine 1 1 1 1 1 2 2 2 3 3 3
AT401, M, 10 mph, 37% SD 1 2 2 3 4 5 7 8 10 13 16
WASP, M, 10 mph, 37% SD 2 2 3 4 5 7 8 10 13 16 18 AT401, C, 10 mph, 25% SD 2 2 3 5 7 9 11 14 19 25 30

WASP, C, 10 mph, 25% SD 2 3 4 6 9 12 14 18 25 33 36

AT401, VC, 10 mph, 20% SD 2 3 5 7 11 14 18 23 33 41 54 WASP, VC, 10 mph, 20% SD 2 4 6 9 13 17 23 27 41 47 54

Groundboom High Boom Very fine to Fine 2 4 6 9 13 16 19 23 30 41 47

Low Boom Very fine to Fine 4 10 16 25 33 41 47 54 65 82 110

High Boom Fine to

Medium/Coarse 7 17 25 36 47 54 65 65 82 110 110

Low Boom Fine to

Medium/Coarse 10 27 41 54 65 82 110 110 160 160 160

Bolded number are risk estimates of conc ern (MOE <1,000)Tribufos Human Health Risk Assessment DP No. D357537

Page 38 of 84 Tables: 6. 2 .1.2 : Tribufos Combined Incidental Oral and Dermal Spray Drift Analysis for Children (Ages 1<2 Years Old)

Crop/Rate Group Spray Type/ Nozzle

Configuration Application

Rate (lb ai/A) Estimated

or Adjusted

TTR t (ug/cm2) At Edge 10 Feet 25 Feet 50 Feet 75 Feet 100 Feet 125 Feet 150 Feet 200 Feet 250 Feet 300 Feet

Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE Combined

MOE

Aerial Fine to Medium

1.875 0.208406 0 1 1 1 1 2 2 2 3 3 4

Medium to Coarse 1 1 1 1 2 3 3 4 5 7 9 Coarse to Very

Coarse 1 1 1 2 3 4 5 6 9 11 14

Very Fine to Fine 0 0 0 0 1 1 1 1 1 1 1
AT401, M, 10 mph,

37% SD 0 1 1 1 1 2 2 3 4 4 5

WASP, M, 10 mph,

37% SD 1 1 1 1 2 2 3 3 4 5 6

AT401, C, 10 mph,

25% SD 1 1 1 2 2 3 4 5 7 9 10

WASP, C, 10 mph,

25% SD 1 1 1 2 3 4 5 6 9 11 13

AT401, VC, 10 mph,

20% SD 1 1 2 3 4 5 6 8 11 14 19

WASP, VC, 10 mph,

20% SD 1 1 2 3 5 6 8 9 14 16 19

Groundboom High Boom Very

fine to Fine 1 1 2 3 5 6 7 8 10 14 16

Low Boom Very fine

to Fine 1 4 6 9 11 14 16 19 23 28 38

High Boom Fine to

Medium/Coarse 2 6 9 13 16 19 23 23 28 38 38

Low Boom Fine to

Medium/Coarse 3 9 14 19 23 28 38 38 56 56 56

Aerial Fine to Medium

1.125 0.125043 1 1 1 2 2 3 3 4 5 6 7

Medium to Coarse 1 1 2 2 3 4 5 7 9 12 14 Coarse to Very

Coarse 1 2 2 4 5 7 9 10 14 19 24

Very Fine to Fine 1 1 1 1 1 1 1 1 2 2 2

AT401, M, 10 mph,

37% SD 1 1 1 2 2 3 4 5 6 7 9

WASP, M, 10 mph,

37% SD 1 1 2 2 3 4 5 6 7 9 10

AT401, C, 10 mph,

25% SD 1 1 2 3 4 5 7 8 11 14 17

WASP, C, 10 mph,

25% SD 1 2 2 4 5 7 8 10 14 19 21

AT401, VC, 10 mph,

20% SD 1 2 3 4 6 8 10 13 19 24 31

WASP, VC, 10 mph,

20% SD 1 2 3 5 8 10 13 16 24 27 31

Groundboom High Boom Very

fine to Fine 1 2 3 5 8 9 11 13 17 24 27

Low Boom Very fine

to Fine 2 6 9 14 19 24 27 31 38 47 63

High Boom Fine to

Medium/Coarse 4 10 14 21 27 31 38 38 47 63 63

Low Boom Fine to

Medium/Coarse 6 16 24 31 38 47 63 63 94 94 94

Bolded number are risk estimates of concer n. (MOE <1,00 0)Tribufos Human Health Risk Assessment DP No. D357537

Page 39 of 84 7.0 Aggregate Exposure/Risk Characterization

In accordance with the FQPA, when there are potential residential exposures to a pesticide, aggregate risk assessment must consider exposures from three major routes: oral, dermal, and inhalation. There are three sources for these types of exposures: food , drinking water, and residential uses. In an aggregate assessment, exposures from relevant sources are added together and compared to quantitative estimates of hazard (e.g., a NOAEL or PAD), or the risks themselves can be aggregated. When aggregating ex posures and risks from various sources, HED considers both the route and duration of exposure. The registered tribufos uses are not anticipated to result in residential exposure.

7.1 Acute Aggregate Risk

The acute aggregate risk assessment is equivale nt to the acute dietary risk assessment discussed

in Section 5.4.3 above.

7.2 Steady State Aggregate Risk

The steady state aggregate risk assessment is equivalent to the steady state dietary risk

assessment discussed in Section 5.4.4 above.

7. 3 Cancer Aggregate Risk

Tribufos is classified as “likely” at high doses but “unlikely” to be a human carcinogen at low

doses

. T he Agency has determined that the quantification of risk using a non

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

8.0 Cumulative Exposure/Risk Characterization

OPs, like tribufos, share the ability to inhibit AChE through phosphorylation of the serine residue on the enzyme leading t o accumulation of acetylcholine and ultimately cholinergic neurotoxicity. This shared MOA/AOP is the basis for the OP common mechanism grouping per OPP’s Guidance For Identifying Pesticide Chemicals and Other Substances that have a Common Mechanism of Tox icity (USEPA, 1999). The 2002 and 2006 CRAs used brain AChE inhibition in female rats as the source of dose response data for the relative potency factors and PODs for each OP, including tribufos. Prior to the completion of Registration Review, OPP will update the OP CRA on AChE inhibition to incorporate new toxicity and exposure information available

since 2006.

As described in Section 4.4, OPP has retained the FQPA Safety Factor for OPs, including tribufos, due to uncertainties associated with neurode velopmental effects in children and exposure to OPs. There is a lack of an established MOA/AOP for the neurodevelopment

outcomes which precludes the agency from formally establishing a common mechanism group per the Guidance For Identifying Pesticide Chemicals and Other Substances that have a Common Mechanism of Toxicity (USEPA, 1999) based on that outcome. Moreover, the lack of aTribufos Human Health Risk Assessment DP No. D357537

Page 40 of 84 recognized MOA/AOP and other uncertainties with exposure assessment in the epidemiology studies prevent the agency from es tablishing a causal relationship between OP exposure and neurodevelopmental outcomes. The agency will continue to evaluate the epidemiology studies associated with neurodevelopmental outcomes and OP exposure prior to the release of the

revised PRA. Durin g this period, the agency will determine whether or not it is appropriate to apply the draft guidance document entitled, Pesticide Cumulative Risk Assessment: Framework for Screening Analysis for the neurodevelopment outcomes.

9.0 Occupational Exposure/R isk Characterization

9.1 Steady

- State Handler Risk

HED uses the term handlers to describe those individuals who are involved in the pesticide application process. HED believes that there are distinct job functions or tasks related to applications and exposures can vary depending on the specifics of each t ask. Job requirements (amount of chemical used in each application), the kinds of equipment used, the target being treated, and the level of protection used by a handler can cause exposure levels to differ in a manner specific to each application event.

Based on the anticipated use patterns and current labeling, types of equipment and techniques that can potentially be used, occupational handler exposure is expected from the registered uses of tribufos. The quantitative exposure/risk assessment develop ed for occupational handlers is based on the following scenarios:

 Mixer/Loader for aerial and groundboom equipment;

 Applicators for aerial and groundboom equipment; and

 Flagger for aerial spray applications.

The above exposure scenarios best represent t he registered use pattern (i.e., cotton defoliant application) for tribufos. Chemigation applications are prohibited on the product labels.

Occupational Handler Exposure Data and Assumptions

A series of assumptions and exposure factors served as the ba sis for completing the occupational handler risk assessments. Each assumption and factor is detailed below on an individual basis.

Application Rate : Product labels indicate that tribufos can be used at a maximum single application rate of 1.875 lbs ai/A for the states CA and AZ and 1.125 lb a i/A for other states

. The occupational handler assessment was completed assuming both single maximum application

rates.

Unit Exposures: It is the policy of HED to use the best available data to assess handler exposure. Sources of generic handler data, used as surrogate data in the absence of chemical

- specific data, include PHED 1.1, the AHETF database, or other registrant

- submitted occupa tional exposure studies. Some of these data are proprietary (e.g., AHETF data), and subject to the data protection provisions of FIFRA. The standard values recommended for use in predicting handlerTribufos Human Health Risk Assessment DP No. D357537

Page 41 of 84 exposure that are used in this assessment, known as “uni t exposures”, are outlined in the “Occupational Pesticide Handler Unit Exposure Surrogate Reference Table 10 ”, which, along with additional information on HED policy on use of surrogate data, including descriptions of the various sources, can be found at the Agency website 11

.

A chemical

- specific handler exposure study (MRID 42685901) was reviewed and used in the 2000 risk assessment. Data from the study were combined as per HED’s policy with other surrogate data from the Pesticide Handlers Exposure Database (PHED) to increase the sample size and numbers of studies (T. Leighton, D252221, January 26, 1999). The study was designed to determine the dermal and inhalation exposures to the workers. The study was conducted in California and Mississippi wit h application rates ranging from 1.127 lb ai/A to 1.875 lb ai/A. The worker exposures in this study, and subsequent MOEs, were determined from dosimetry

data

. The exposure assessment component of the study on which OPP is relying does not
measure or iden tify a toxic effect; therefore, review of the exposure assessment research by the Human Studies Review Board (HSRB) is not required (M. Lydon, Ethics Review of Exposure Research from 1993 Study of Worker Exposure to Tribufos during Aerial and Ground Applic ation of DEF 6 to Cotton, MRID 42685901, 07/02/ 2015)

.

Area Treated or Amount Handled: Based on HED EXPOSAC Policy 9.1, the amount treated or handled per day was assumed to be:

 200 acres for mixing/loading ground boom applications and applying via grou nd equipment on cotton;

 1,200 acres for mixing/loading for aerial applications and applying via aircraft on cotton;

and

 350 acres for flagging for aerial application on cotton crops.

Body Weight: For adults, when an endpoint is not sex

- specific (i.e., the endpoints are based on developmental or fetal effects) a body weight of 80 kg is typically used in risk assessment; however, in this case, a female

- specific body weight of 69 kg was used. While the endpoint of concern, RBC AChE inhibition, is not sex

- specific, the female

- specific body weight was used to protect for pregnant women due to uncertainty in the human dose

- response relationship for neurodevelopmental effects (See Section 4.4).

Since the steady

- state dermal point of departure P o D was based on a route

- specific toxicity study, no dermal absorption factor was necessary to estimate exposure. A route

- specific inhalation study is available and was used in the assessment.

Exposure Duration: HED typically classifies exposures from 1 to 30 days as short

- term and exposures 30 days to six months as intermediate

- term. Exposure duration is determined by many things, including the exposed population, the use site, the pest pressure triggering the use of the pesticide, and the cultural practi ces surrounding that use site. For the organophosphate pesticides, the match between the toxicological endpoint selection and the exposure duration is different than HED’s approach for conventional chemicals. The endpoint selection for organophosphate pe sticides involves a “single day” and “steady state” approach based on an

10 Available: http://www.epa.gov/opp00001/science/handler

- exposure

- table.pdf

11 Available: http://www.epa.gov/pesticides/science/handler

- exposure

- data.htmlTribufos Human Health Risk Assessment DP No. D357537

Page 42 of 84 individual chemical’s cholinesterase inhibition. T he steady state approach is appropriate for

tribufos given the toxicological and exposure profile. The “steady state” endpoint sele ction for

tribufos overlaps HED’s traditional short

- term exposure duration endpoint selection and is

considered protective for occupational handlers that apply commercially over longer periods of time (i.e., intermediate

- term exposures).

Existing Mitigati on/Personal Protective Equipment: Estimates of dermal and inhalation exposure were calculated based on the mitigation of agricultural uses from the July 2006 tribufos

RED. Current tribufos labels require closed systems for mixers/loaders for aerial applic ations

and all applicators and handlers must wear coveralls over baseline dermal wear in addition to gloves, protective eyewear, chemical resistant headgear of overhead exposure and chemical

resistant apron when cleaning, mixing or loading

.

Combining Exp osures/Risk Estimates:

A total aggregate risk index (ARI) was used since the LOC values for dermal exposure (1

, 00 0 ) and inhalation exposure ( 300 ) are different. The target ARI is 1; therefore, ARIs of less than 1 are of concern. The aggregate risk index (ARI) was calculated as follows.

Aggregate Risk Index (ARI) = 1÷ [(Dermal LOC ÷ Dermal MOE) + (Inhalation LOC ÷ Inhalation MOE)]

Summary of Occupational Handler Non

- Cancer Exposure and Risk Estimates

HED has no data to assess exposure to pilots using open cockpits. The only data available is for exposure to pilots in enclosed cockpits. Therefore, risks to pilots are assessed assuming the use

of engineering control s (enclosed cockpits) and baseline at tire (long

- sleeve shirt, long pants, shoes, and socks); pilots are not required to wear protective gloves.

HED does not have unit exposure data for assessing dermal and inhalation exposure for tribufos flaggers with engineering controls. HED has presen ted flagger risk estimates at double layer

baseline PPE (coveralls over long

- sleeve shirt, long pants, shoes, socks, gloves) and PF10 respirator.

The Agency matches quantitative occupational exposure assessment with appropriate characterization of exposur e potential. While HED presents quantitative risk estimates for human flaggers where appropriate, agricultural aviation has changed dramatically over the past

two decades. According the 2012 National Agricultural Aviation Association (NAAA) survey of

the ir membership, the use of GPS for swath guidance in agricultural aviation has grown steadily

from the mid 1990’s. Over the same time period, the use of human flaggers for aerial pesticide applications has decreased steadily from ~15% in the late 1990’s to only 1% in the most recent (2012) NAAA survey. The Agency will continue to monitor all available information sources to best assess and characterize the exposure potential for human flaggers in agricultural aerial applications.

In the Appendix 5 , dermal and inhalation risk estimates are presented individually with varying

PPE levels ranging from baseline (i.e., no gloves and no respirator) through PF10 respirator and engineering controls. Dermal MOEs appear to be the driver and range from 0.1 7 to 3.9 withTribufos Human Health Risk Assessment DP No. D357537

Page 43 of 84 labeled PPE 0.57 to 9.6 with ECs. Inhalation MOE s range d from 54 to 1,000 with labeled PPE (See Appendix Table A.5.1 and Table A.5.2 highlighted MOEs).

A ll of the occupational handler exposure scenarios resulted in combined dermal and inhalation

r isk estimates of concern (i.e., ARIs are < 1) at the label

- defined PPE for occupational handlers,

and all additional levels of PPE and ECs assessed

. Occupational handler r isk estimates are presented in the following table 9. 1.1

.Tribufos Human Health Risk Assessment DP No. D357537

Page 44 of 84 Table 9 .1.1. Steady

- State Occupational Exposure and Risk Estimates for Tribufos Total ARI

Exposure Scenario Application

Rate 1 Amount

Handled /

Area Treated 2 ARI 3

Worker Activity Formula

tion Applic

ation Equip

ment Value Units Value Units SL/No G

+ No

- R SL/G +

No

- R DL/G +

No

- R SL/No

G +

PF5 R SL/G

+ PF5

R DL/G +

PF5 R SL/No

G +

PF10 R SL/G

+ PF10

R DL/G

+ PF10

R EC

M/L L/SC/EC Aerial 1.875

lb ai/A 1200

A 0.00002 0.00013 0.00017 0.00002 0.00013 0.00017 0.00002 0.00013 0.00017 0.00057

1.125 0.00004 0.00022 0.00028 0.00004 0.00022 0.00028 0.00004 0.00022 0.00028 0.00095

Ground

boom 1.875

200 0.00013 0.00078 0.00100 0.00013 0.00078 0.00100 0.00013 0.00078 0.00100 0.00340

1.125 0.00022 0.00130 0.00170 0.00022 0.00130 0.00170 0.00022 0.00130 0.00170 0.00570

Applicator

Spray Aerial 1.875

lb ai/A 1200

A No Data No Data No Data No

Data No

Data No

Data No

Data No

Data No

Data 0.00240

Aerial 1.125 1200 No Data No Data No Data No

Data No

Data No

Data No

Data No

Data No

Data 0.00390

Ground

boom 1.875 200 0.00037 0.00180 0.00230 0.00037 0.00180 0.00230 0.00037 0.00180 0.00230 0.00580

Ground

boom 1.125 200 0.00062 0.00300 0.00380 0.00062 0.00300 0.00390 0.00062 0.00300 0.00390 0.00960

Flagger Aerial 1.875

lb ai/A 350 A 0.00150 0.00140 0.00160 0.00150 0.00140 0.00160 0.00150 0.00140 0.00160 No Data

1.125 0.00250 0.00230 0.00260 0.00250 0.00230 0.00260 0.00250 0.00230 0.00260 No Data

1. Based on registered label (e.g. Reg. No. 5481

- 504, 5481

- 903 or EPA Reg. No. 264

- 730, 34704

- 867, and 85678

- 15).

2. Exposure Scienc e Advisory Council Policy #9.1

3. Total Aggregate Risk Index (ARI) = 1÷ [(Dermal LOC ÷ Dermal MOE) + (I nhalation LO C ÷ Inhalation MOE). Bolded numbers are risk estimates of concern (ARI<1 ). Highlighted risk

estimates are label defined PPE

. SL = single layer, DL = double layer, G = gloves, EC = engineering controls, R = respiratorTribufos Human Health Risk Assessment DP No. D357537

Page 45 of 84 9.2 Steady State Post

- Application Risk

HED uses the term post

- application to describe exposures that occur when individuals are present in an environment that has been previously treated with a pesticide (also referred to as re

entry exposure). Such exposures may occur w hen workers enter previously treated areas to perform job functions, including activities related to crop production, such as scouting for pests or harvesting. Post

- application exposure levels vary over time and depend on such things as the type of activi ty, the nature of the crop or target that was treated, the type of pesticide application, and the chemical’s degradation properties. In addition, the timing of pesticide applications, relative to harvest activities, can greatly reduce the potential for po st

- application exposure.

9.2.1 Occupational Post

- application Inhalation Exposure/Risk Estimates

There are multiple potential sources of post

- application inhalation exposure to individuals performing post

- application activities in previously treated field s. These potential sources include volatilization of pesticides and resuspension of dusts and/or particulates that contain pesticides. The Agency sought expert advice and input on issues related to volatilization of pesticides from its Federal Insecticid e, Fungicide, and Rodenticide Act Scientific Advisory Panel (SAP) in December 2009, and received the SAP’s final report on March 2, 2010 12 . The Agency has evaluated the SAP report and has developed a Volatilization Screening Tool and a subsequent Volatiliz ation Screening Analysis 13 . During Registration Review, the agency will utilize this analysis to determine if data (i.e., flux studies, route

- specific inhalation toxicological studies) or further analysis is required for tribufos.

In addition, the Agency i s continuing to evaluate the available post

- application inhalation exposure data generated by the Agricultural Reentry Task Force. Given these two efforts, the Agency will continue to identify the need for and, subsequently, the way to incorporate

occupat ional post

- application inhalation exposure into the agency's risk assessments.

9.2.2 Occupational Post

- application Dermal Exposure/Risk Estimates

Occupational Post

- application Dermal Exposure Data and Assumptions

A series of assumptions and exposure factors served as the basis for completing the occupational

post

- application risk assessments. Each assumption and factor is detailed below on an individual

basis.

Exposure Duration: The endpoint selection for organophosphate pesticides involves a “singl e day” and “steady state” approach based on an individual chemical’s cholinesterase inhibition. The steady state approach is appropriate for tribufos given the toxicological and exposure profile. The “steady state” endpoint selection for tribufos overlap s HED’s traditional short

- term exposure

12 http://www.epa.gov/scipoly/SAP/meetings/2009/120109meeting.html

13 http://www.regulations.gov/#!docketDetail;D= EPA

- HQ

- OPP

- 2014

- 0219.Tribufos Human Health Risk Assessment DP No. D357537

Page 46 of 84 duration endpoint selection as well as being appropriately health protective for workers that are exposed over longer periods of time (i.e., intermediate

- term exposures).

Transfer Coefficients: It is the policy of t he HED to use the best available data to assess post

application exposure. Sources of generic post

- application data, used as surrogate data in the

absence of chemical

- specific data, are derived from ARTF exposure monitoring studies, and, as proprietary da ta, are subject to the data protection provisions of FIFRA. HED standard values for “transfer coefficients”, are presented in the EXPOSAC Policy 3 14 ” which, along with additional information about the ARTF data, can be found at the Agency website 15 . Policy 3 includes the standard transfer coefficient (TC) values for mechanically harvesting cotton based

on a tribufos chemical specific study (MRID No. 42701601). The study was reviewed by EPA (B. Tarplee, D227007, 02/12/19 9 7) and then re

- reviewed in 1999 (T. Leighton, D252221

01/26/199 9 ) and was a two part worker monitoring study that determined dermal and inhalation exposures of workers conducting mechanical harvesting activities for cotton, and, in addition, collected cotton boll samples for residues. The transfer coefficient values are as follows; picker operator 2,400 g/hr, module builder operator 900 g/hr, rakers 2,400g/hr, and trampers 5,050 g /hr. The exposure assessment component of the study on which OPP is relying does not measure or identify a toxic effect; therefore, review of the exposure assessment research by the Human

Studies Review Board (HSRB) is not required. If OPP needs to rely on this study’s other type of research, namely its research on cholinesterase levels, then HSRB review of the cholinesterase research would be required (M. Lydon, Ethics Review of Exposure Research from 1993 Study of Worker Exposure to Tribufos during Harv esting of Cotton Treated with DEF 6, MRID 42701601, 07/2/2015)

. Because tribufos is a defoliant, other post

- application activities such as weeding and scouting were not considered in this assessment

.

Table 9 .2.2.1: Anticipated Post

- Application Activi ties and Dermal Transfer Coefficients.

Crops Policy Crop

Group Category Transfer

Coefficients Activities
g/hr

Cotton Field/ Row

Crop,

Low/Medium 2,400 Harvesting, Mechanical, Picker Operator Harvesting,

900 Mechanical, Module Builder Operator
2,400 Harvesting, Mechanical, Raker
5,050 Harvesting, Mechanical, Tramper

Application Rate: The occupational post

- application assessment was conducted at the maximum single application rates of 1.125 lb ai/A and 1.875 lb ai/A.

Body Weight: For adults, when an endpoint is not sex

- specific (i.e., the endpoints are based on developmental or fetal effects) a body weight of 80 kg is typically used in risk assessment; however, in this case, a female

- specific body weight of 69 kg was used. While t he endpoint of concern, RBC AChE inhibition, is not sex

- specific, the female

- specific body weight was used to

14 Available: http://www.epa.gov/pesticides/science/ ExpoSAC _policy3.pdf

15 Available: http://www.epa.gov/pesticides/science/post

- app

- exposure

- data.htmlTribufos Human Health Risk Assessment DP No. D357537

Page 47 of 84 protect for pregnant women due to uncertainty in the human dose

- response relationship for neurodevelopmental effects (See Section 4.4).

Absorpt ion Factor: Since the steady

- state dermal PoD was based on a route

- specific toxicity study, no absorption factor was necessary to estimate exposure.

Exposure Time: The average occupational workday is assumed to be 8 hours.

Dislodgeable Cotton Boll Res idues: As stated above in the transfer coefficient section, a

chemical

- specific study was submitted (MRID 42701601, Eberhart, D.; Ellisor, G. (1993) Evaluation of Worker Exposure to Tribufos During Harvesting of Cotton Treated with DEF 6: Lab Project Numb er: DE202302: 105103. Unpublished study prepared by Miles Inc. 307 p. ). Triplicate cotton samples (50 grams each) from four fields at two different geographical areas (California and Mississippi ) were collected at various days after treatment (DAT) of DEF 6 at the maximum application rate of 1.875 lb ai/acre. Cotton boll samples were collected prior to treatment DAT 0, and up to DAT 20. This assessment uses the ground application to the California site for DFR analysis because it was the most complete dat a set from the four sites. This analysis generated an initial concentration value of 1.803 μg/g bolls on day one after

treatment. Table 9 .2.2.2 below provides a summary of the data for use in the post

- application exposure calculations.

Table 9 .2.2.2. DFR Values and Regression Analysis Results for Cotton Treated with DEF 6 (EPA REG #

3125

- 282) Tribufos (MRID# 42701601).

Cotton

Input/Parameter Site: California Ground Application Application Rate (lb ai/A) a 2.5 pints/acre or 1.875 lb ai/acre Spray Volume (GPA) 20 GPA Measured Average Day 0 Residue (μg/cm 2 ) 1.803 μg/g bolls a. The applications were conducted using ground equipment.

Occupational dermal post

- application risk assessments were conducted at application rates of

1.875 lb ai/A for the states CA and AZ and for the lower rate of 1.125 lb ai/A using the standardized dermal transfer coefficients for mechanically harvesting cotton and the dislodgeable foliar residue values collected from the ground CA application. Usin g this information there were risks estimates of concern (MOE < 1,000) for the mechanical module picker, mechanical picker, raker and tramper from DAT 0 through DAT 30 for the higher application rate of 1.875

lb ai/A and for the lowe r application rate. Se e Table 9.2.2.3 and 9 .2.2.4 for post application risk

estimates.Tribufos Human Health Risk Assessment DP No. D357537

Page 48 of 84 Table 9 .2.2.3: Summary of Occupational/Commercial Mechanically Harvesting Post

- application Risk Estimates for Tribufos on Cotton 1.125 lb ai/A with Steady State PoD

Activity Transfer

Coefficient (cm 2 /hr) 1 MOE 2
DAT (Day After Treatment)

0 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30

Module

Builder Operator 900 1.4 5.5 6.7 8.1 9.8 12 15 18 21 26 32 38 46 56 68 83 100 120 150 180 22

0 270 320 390 480

Picker Operator 2400 0.53 2.1 2.5 3 3.7 4.5 5.4 6.6 8 9.7 12 14 17 21 26 31 38 46 56 68 82 100 120 150 180

Raker 2400 0.53 2.1 2.5 3 3.7 4.5 5.4 6.6 8 9.7 12 14 17 21 26 31 38 46 56 68 82 100 120 150 180

Tramper 5050 0.25 0.98 1.2 1.4 1.8 2.1 2.6 3.1 3.8 4.6 5.6 6.8 8.3 10 12 15 18 22 26 32 39 47 58 70 85

1. DFR and TC values for each activity taken from taken from chemical

- specific study data (MRID 42685901 and MRID 42701601).

2. MOE = POD (mg/kg/day) / Daily Dermal Dose. Daily Dermal Dose = [DFR/TTR (μg/g boll \ ) x TC x 0.001 mg/μg x 8 hrs. /day]  69 (kg).

Table 9 .2.2.4: Summary of Occupational/Commercial Mechanically Harvesting Post

- application Risk Estimates for Tribufos on Cotton 1.875 lb ai/A with Steady State PoD

Activity Transfer

Coefficient (cm 2 /hr) 1 MOE 2
DAT (Day After Treatment)

0 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30

Module

Builder Operator 900 0.85 3.3 4 4.9 5.9 7.2 8.7 11 13 16 19 23 28 34 41 50 61 73 89 110 130 160 190 240 290

Picker Operator 2400 0.32 1.2 1.5 1.8 2.2 2.7 3.3 4 4.8 5.8 7.1 8.6 10 13 15 19 23 28 33 41 49 60 73 88 110

Raker 2400 0.32 1.2 1.5 1.8 2.2 2.7 3.3 4 4.8 5.8 7.1 8.6 10 13 15 19 23 28 33 41 49 60 73 88 110

Tramper 5050 0.15 0.59 0.71 0.87 1.1 1.3 1.6 1.9 2.3 2.8 3.4 4.1 5 6 7.3 8.9 11 13 16 19 23 28 35 42 51

1. DFR and TC values for each activity taken from taken from chemical

- specific study data (MRID 42685901 and MRID 42701601).

2. MOE = POD (mg/kg/day) / Daily Dermal Dose. Daily Dermal Dose = [DFR/TTR (μg/g boll \ ) x TC x 0.001 mg/μ g x 8 hrs. /day]  69 (kg).Tribufos Human Health Risk Assessment DP No. D357537

Page 49 of 84 Restricted Entry Interval

Tribufos is classified as Toxicity Category II via the dermal route and for eye irritation and Toxicity Category IV for skin irritation potential. It is not a skin sensitizer. Under 40 CFR 156.208 (c) (2), i f the active ingredient is an organophosphorus ester that inhibits cholinesterase and that may be applied outdoors in an area where the average rainfall is less than 25 inches per year, the restricted interval should be listed as 72 hours. The current tribufos restricted entry interval (REI) was changed from 24 hours to 7 days per the 2000 IRED mitigation, however

, there remains risk estimates of concern for all mechanical post

- application activities using the updated PoDs and SOPs.

10.0 Review of the Incident Report

Tribufos was previously reviewed in 2008 in the Updated Review of Tribuphos Incident Reports (M. Hawkins and J. Co rdova, 10/30/2008). In this earlier review, Main Incident Data Sy s tem ( IDS ) was queried from 2002 to October 2008. Two incidents were reported, one was classified as major and one was classified as moderate. This current Tier I update looks at IDS incident reports from 2007 through 2014 and found two additional single active ingredient (ai) cases which were both minor in severity (incident report reference) . SENSOR

- Pesticides identified 19 single ai cases from 1998 to 2010. B ased on the low frequency and relatively low severity of incident cases reported for Tribufos in both IDS and National Institute of Occu pa tional Safety and Health Sentinel Event Notification System for Occupational Risk ( NIOSH SENSOR )

Pesticides in the current analysis , there does not appear to be a concern at this time with respect

to incident cases that would warrant further investigation. The Agency will continue to monitor the incident information and if a concern is triggered, additional analysis will be included in the

risk assessment.

11.0 References

Bouchard MF, Chevrier J, Harley KG, Kogut K, Vedar M, Calderon N, et al. Prenatal Exposure to Organophosphate Pesticides and IQ in 7

- Year

- Old Children. Environ Health Perspectives.
2011;119:1189

- 1195.

Engel SM, Berkowitz GS, Barr DB, Teitelbaum SL, Siskind J, Meisel SJ, Wetmur JG, Wolff MS. Prenatal Organophosphate Metabolite and Organo chlorine Levels and Performance on the Brazelton Neonatal Behavioral Assessment Scale in a Multiethnic Pregnancy Cohort. American Journal of Epidemiology. 2007;165:1397

- 1404.

Engel SM, Wetmur J, Chen J, Zhu C, Barr DB, Canfield RL, Wolff MS. Prenatal Expos ure to Organophosphates, Paraoxonase 1, and Cognitive Development in Childhood. Environmental Health Perspectives. 2011;119:1182

- 1188.

Eskenazi B, Harley K, Bradman A, Weltzien E, Jewell NP, Barr DB, Furlong CE, Holland NT. Association of in Utero Organop hosphate Pesticide Exposure and Fetal Growth and Length of Gestation in an Agricultural Population. Environmental Health Perspectives. 2004;115:792

– 798.Tribufos Human Health Risk Assessment DP No. D357537

Page 50 of 84 Eskenazi B, Marks AR, Bradman A, Harley K, Barr DB, Johnson C, Morga N, Jewell NP. Organophosphate Pest icide Exposure and Neurodevelopment in Young Mexican

- American Children. Environmental Health Perspectives. 2007;115:792

– 798.

FIFRA Scientific Advisory Panel. (2012). Transmittal of the Meeting Minutes from the FIFRA Scientific Advisory Panel Meeting of Apr il 10

- 12, 2012. FIFRA Scientific Advisory Panel, Office of Science Coordination and Policy, Office of Prevention, Pesticides and Toxic Substances, U.S. Environmental Protection Agency. Washington, DC. Available: http://www.epa.gov/scipoly/sap/meetings/2012/041012meeting.htmll

Fortenberry, GZ, Meeker, JD, Sánchez, BN, Barr, DB, Panuwet, P, Bellinger, D, Schnaas, L,

Solano

- González, M, Ettingera, AS, Hernandez

- Avila, M, Hu, H, Tellez

- Rojo j, MM. Urinary 3,5,6

- trichloro

- 2

- pyridinol (TCPY) in pregnant women from Mexico City: Distribution, temporal variability, and relationship with child attention and hyperactivity. International Journal of Hygiene and Environmental Health 217 (2014): 405

– 412

. Fortenberry, GZ, Meeker, JD, Sánchez, BN, Barr, DB, Peterson, K, Bellinger, D, Schnaas, L,

Solano

- González, M, Ettinger, AS, Hernandez

- Avila, M, Hu, H, Tellez

- Rojoj, MM. Paraoxonase I polymorphisms and attention/hyperactivity in school

- age children from Mexico City, Mexico. (2014a.) Environmental Research 132:342

– 349.

Furlong, Melissa A., Engel, Stephanie M., Boyd Barr, Dana, Wolff, Mary S. Prenatal exposure to organophosphate pesticides and reciprocal social behavior in childhood. 2014. Environment

Inter national 70:125

- 131.

Guodong, Ding, Pei, Wang, Ying, Tian, Jun, Zhang, Yu, Gao, Xiaojin, Wang, Rong, Shi, Guoquan, Wang, and Xiaoming, Shen. Organophosphate Pesticide Exposure and Neurodevelopment in Young Shanghai Children. Environmental Science and Techn ology

46:2911

- 2917, 2012.

Oulhote, Youssef and Bouchard, Maryse F. Urinary Metabolites of Organophosphate and Pyrethroid Pesticides and Behavioral Problems in Canadian Children. Environmental Health Perspectives. 121:1378

- 1384 (2013)

Rauh VA, Garfinkel R, Perera FP, Andrews HF, Hoepner L, Barr DB, Whitehead R, Tang D, Whyatt RW. Impact of Prenatal Chlorpyrifos Exposure on Neurodevelopment in the First 3 Years of Life Among Inner

- City Children. Pediatrics. 2006;18(6): 1835

- 1859.

Rauh V, Arunajadai S, Horton M, Perera F, Hoepner L, Barr DB, et al. 2011. Seven

- Year Neurodevelopmental Scores and Prenatal Exposure to Chlorpyrifos, a Common Agricultural Pesticide. Environmental Health Perspectives. 2011;119:1196

- 1201.

Shelton, Janie F., Geraghty, Estella M., Tanc redi, Daniel J., Delwiche, Lora D., Schmidt, Rebecca J., Ritz, Beate, Hansen, Robin L., and Hertz

- Picciotto, Irva. Neurodevelopmental disorders and prenatal residential proximity to agricultural pesticides: The CHARGE Study. Environmental Health Perspectiv es 122:1103

– 1109, 2014.

U.S. Environmental Protection Agency. (2008). Science Issue Paper: Chlorpyrifos Hazard and Dose Response Characterization

. http://www.epa.gov/scipoly/sap/meetings/2008/091608_mtg.htmTribufos Human Health Risk Assessment DP No. D357537

Page 51 of 84 U.S. Environmental Protection Agency. (2012). Draft Issue Paper: Scientific Issues Concerning Health Effects of Chlorpyrifos. http://www.regulations.gov/#!documentDetail;D=EPA

- HQ

- OPP

2012

- 0040

- 0002

Young JG, Eskenazi B, Gladstone EA, Bradman A, Pedersen L, Johnson C, Barr DB, Furlong CE, Holland NT. Association Between In Utero Organophosphate Pesticide Exposure and Abnormal Reflexes in Neonates. NeuroToxicology. 2005;26:199

– 209.

Zhang, Ying, Han, Song, Liang, Duohong, Shi, Xinzhu, Wang, Fengzhi, Wei Liu, Zhang, Li, Chen, Lixin, Gu, Yingzi, Tian, Yin g. Prenatal exposure to organophosphate pesticides and neurobehavioral development of neonates: A birth cohort study in Shenyang, China. PlosOne

47:80

– 85, 2014.

Tribufos: Human Health Risk Assessment. D266540, June 26, 2000, R. Travaglini.

Tribufos: Occ upational and Residential Exposure Assessment for the Registration Review Risk

Assessment. D424281

, 9/15/2015

, B. Bobowiec.

Tribufo s: Updated Dietary Burden for Livestock Commodities, D427650, June 4, 2015, S. Piper.

Tribufos: Acute and Steady State Aggregate Dietary (Food and Drinking Water) Exposure and Risk Assessments to Support Re gistration Review. D42796, September 8 , 2015, S. Piper.

Tribufos: Tier 1 Update Review of Human Incidents fo r Preliminary Risk Assessment.
D428263, November 25, 2014, S. Recore.

Tribufos: Drinking Water Exposure Assessment in Support of the Registration Review for the Organophosphate Tribufos. D424579, January 20, 2015, C. Sutton.

Tribufos: Amendment to Refine the Drinking Water Exposure Assessment in Support of the Registration Review for the Organophosphate Tribufos . July 14 , 2015, D424 903 , C, Sutton.

Tribufos: Benchmark Dose Analysis of Acute, Subchronic, and Chronic Studies to Support Derivation of Points of Departure. D423990 , 06/17/ 1 5

. R. Bever & E. Holman

.

Ethics Review of Exposure Research from 1993 Study of Worker Exposure to Tribufos during Aerial and Ground Application of DEF 6 to Cotton. 07/01/2015. M. Lydon.

Ethics Review of Exposure Research from 1993 Study of Worker Exposure to Tribufos during

Harve sting of Cotton Treated with DEF 6. 07/02/2015. M. Lydon.Tribufos Human Health Risk Assessment DP No. D357537

Page 52 of 84 Appendix 1. Summary of OPP’s ChE Policy & Use of BMD Modeling

OPP’s cholinesterase ( ChE ) policy (USEPA, 2000 16 ) describes the manner in which ChE data are

used in human health risk assessment. The following text provides a brief summary of that document to provide context to points of departure selected.

Acetylcholinesterase (AChE) can be inhibited in the central or peripheral nervous tissue.

Measurem ents of AChE or ChE inhibition in periph eral tissues (e.g., liver, diaphragm, heart, lung etc) are rare. As such, experimental laboratory studies generally measure brain (central) and blood (plasma and red blood cell, RBC) ChE. Blood measures do not represent the target tissue, per se, but are instead used as surrogate measures for peripheral toxicity in studies with laboratory animals or for peripheral and/or central toxicity in humans

. In addition, RBC measures represent AChE

, whereas plasma measures are predominately butyryl

- ChE (BuChE)

. T hus, RBC AChE data may provide a better representation of the inhibition in target tissues. As part of the dose response assessment, evaluations of neurobehavior and clinical signs are performed to consider the dose response linkage between AChE inhibitio n and apical outcomes.

Refinements to OPP’s use of ChE data have come in the implementation of BMD approaches in dose response assessment. Beginning with the OP CRA, OPP has increased its use of BMD modeling to derive PoDs for AChE inhibiting compounds. Most often the decreasing exponential empirical model has been used.

OPP does have not a defined benchmark response (BMR) for OPs. However, the 10% level has been used in the majority of dose response analyses conducted to date. This 10% level

repre sents a 10% reduction in AChE activity (i.e., inhibition) compared to background (i.e., controls). Specifically, t he BMD 10 is the estimated dose where ChE is inhibited by 10% compared to background. The BMDL 10 is the lower confidence bound on the BMD 10

.

The use of the 10% BMR is derived from a combination of statistical and biological considerations. A power analysis was conducted by the Office of R esearch and Development (ORD) on over 100 brain AChE datasets across more than 25 OPs as part of the OP CRA (USEPA, 2002). This analysis demonstrated that 10% is a level that can be reliably measured in the majority of rat toxicity studies . In addition, the 10% level is generally at or near the limit of sensitivity for discerning a statistically significant decrease in ChE activity in the brain compartment and is a response level close to the background brain ChE level. With respect to biological conside rations, a change in 10% brain AChE inhibition is protective for downstream clinical signs and apical neurotoxic outcomes. With respect to RBC AChE inhibition, these data tend to be more variable than brain AChE data. OPP begins its BMD analyses using th e 10% BMR for RBC AChE inhibition but BMRs up to 20% could be considered on a case by case basis as long as such PoDs are protective for brain AChE inhibition, potential peripheral inhibition, and clinical signs of neurotoxicity.

16 USEPA (2000) Office of Pesticide Programs, US Environmental Protection Agency, Washington DC 20460. August 18, 2000 Office of Pesticide Programs Science Policy of The Use of Data on Cholinesterase Inhibition for Risk Assessments of Organophosphorous and Carbamate Pesticides.Tribufos Human Health Risk Assessment DP No. D357537

Page 53 of 84 Appendix 2

– Results for BMD/BMDL modeling for Tribufos

Please note that confidence in the accuracy of reported BMD 10 and BMDL 10 values were not the same for all submitted tests or
specific data sets within each test. The most reliable values are shown i n bold

. Less trusted values are indicated in gray

.

For all the tables: NDR = No Dose Response;

--- = a trustworthy value could not be calculated, usually due to too much variance;
CCA = Comparative Cholinesterase Assay; M# = Model number chosen; C = constant variance model; NC = modeled without

constant va riance; FT# = Failed statistical test number x ; RD = Repeated Dose (study); RF = Range Finder

Appendix Table 2.1

. BMD results of single dose studies in rats.

Test Sex & age Compartment BMD Results

Comments BMD 10 (mg/kg) BMDL 10 (mg/kg)

MRID 48709901

Acute CCA RF, 24h Male PND11 Brain

--- --- Too much variance

Female PND11 Brain 5.56 4.31 M2, C, Too much variance

MRID 48709901

Acute CCA RF, 24h Male PND11 RBC

--- --- Too much variance

Female PND11 RBC 1.22 0.98 M4 a , NC

MRID 48709901

Acute CCA RF, 48h Male PND11 Brain 6.47 5.51 M2, C, Too much variance

Female PND11 Brain 6.90 6.02 M2, NC

MRID 48709901

Acute CCA RF, 48h Male PND11 RBC 2.95 1.70 M4, NC

Female PND11 RBC 3.25 2.43 FT3, M2, NC, Too much variance

MRID 48709904

Acute CCA a , 24h Female Brain 77.97 56.03 M2, C, Not Accurate

Female RBC 5.98 4.68 FT3, M2, NC

MRID 48709904

Acute CCA RF, 6h Male PND11 Brain 9.21 4.74 FT3, M5 a , NC

Female PND11 Brain 8.62 5.68 FT3, M5, NC, Accuracy?

MRID 48709904

Acute CCA RF, 6h Male PND11 RBC 1.41 1.05 M4, NC

Female PND11 RBC

--- --- NF

a The Chi

- Square test for fit is not valid, because degrees of freedom ≤0.Tribufos Human Health Risk Assessment DP No. D357537

Page 54 of 84 Appendix Table 2.2

. BMD results of repeated dos e oral studies in rats

.

Test Sex & age Compartment BMD Results

Comments BMD 10 (mg/kg) BMDL 10 (mg/kg)

MRID 48709901

RD CCA RF, 6 h Male PND11 Brain 2.40 2.19 M2, C, Too much variance

Female PND11 Brain 2.59 2.07 M4, C

MRID 48709901

RD CCA RF, 6 h Male PND11 RBC

--- --- Too much variance

Female PND11 RBC

--- --- Too much variance

MRID 48709905

RD CCA a , 24h Female Brain

--- --- NDR

Female RBC 0.54 0.31 M4, C, Not accurate

MRID 48709905

RD CCA , 6h Male PND11 Brain 3.49 2.69 M3, C

Female PND11 Brain 2.20 1.79 M4, C

MRID 48709905

RD CCA , 6h Male PND11 RBC 4.08 0.66 M3, NC, Too much variance

Female PND11 RBC 0.47 0.29 M4, NC, Not accurate

MRID 48709906

Gestational CCA , 24h Dam Brain 3.70 3.05 M3, C

Dam RBC 5.32 1.06 M5 c , NC

MRID 48709906

Gestational CCA b , 24h Fetus Brain

--- --- FT1

Fetus RBC 22.30 13.92 FT3, M2, NC, Too much variance

MRID 45369101

Subchr

. N T 4W Male RBC 0.28 0.21 FT3, M4, NC

Female RBC 0.27 0.19 M4, NC

MRID 42007201 21D

Dermal Tox Male Brain 6.67 4.97 M4

Female Brain 7.73 6. 50 FT3, M2, NC

MRID 42007201 21D

Dermal Tox Male RBC 0.87 0.48 M4, C, Too much variance

Female RBC 0.22 0.16 M4, NC

MRID 42399801

Subc hr. Inhal

. 13W Male RBC 1.00 mg/m 3 0.79 mg/m 3 FT3, M4, NC

Female RBC 1.01 mg/m 3 0.85 mg/m 3 FT3, M4, NC

a Adult male not tested

b Fetus not more sensitive than dam

c The Chi

- Square test for fit is not valid, because degrees of freedom ≤0Tribufos Human Health Risk Assessment DP No. D357537

Page 55 of 84 Appendix 3. Toxicology Profile and Executive Summaries

A. 3. 1 Toxicology Data Requirements
The requirements (40 CFR 158.500) for the non

- food use for Tribufos are in Tabl e A.1. Use of the new guideline numbers does not imply that the new (1998) guideline protocols were used.

Table A. 3. 1 Toxicology Data Requirements for Tribufos

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 21/28

- Day Dermal

................................ ......................... 870.3250 90

- Day Dermal

................................ .............................. 870.3465 90

- Day Inhalation

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

yes

yes

no CR yes

yes

yes

yes

yes

870.3700a Developmental Toxicity (rodent)

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

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

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

yes

yes yes

yes

yes

870.4100a Chronic Toxicity (rodent)

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

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

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

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

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

no

yes

yes

yes yes

yes

yes

yes

yes

870.5100 Mutagenicity

— Gene Mutation

- bacterial

..................... 870.5300 Mutagenicity

— Gene Mutation

- mammalian

................ 870.5375 Mutagenicity

— Structural Chromosomal Aberrations

... 870.5395 Mutagenicity

— Mammalian erythrocyte micronucleus. 870.5550 Mutagenicity

— Unscheduled DNA synthesis............ yes

yes

yes

yes

no yes

no

yes

no

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 Developmental Neurotoxicity

................................ ....... CR

no

yes

yes CR yes

yes

yes

yes

yes

870.7485 General Metabolism

................................ ...................... 870.7600 Dermal Penetration

................................ ........................ 870.7800 Immunotoxicity

................................ ............................. yes CR

yes yes

yes

yes

Special Studies

Comparative Cholinesterase (rat)

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

CR = conditionally requiredTribufos Human Health Risk Assessment DP No. D357537

Page 56 of 84 A. 3. 2 Toxicity Profiles

Table A. 3. 2.1 Acute Toxicity of Tribufos

Guideline

Number Study Type MRID Results Toxicity Category

870.1100 Acute Oral

- Rat 41954903 LD 50 =192

- 235 mg/kg II

870.1200 Acute Dermal

- Rabbit 41954902 LD 50 =>1000 mg/kg (m) <2000 mg/kg (f) II

870.1300 Acute Inhalation

- Rat 41782301 LC 50 =4650 mg/m 3 (m) 2460 mg/m 3 (f) III

870.2400 Primary Eye Irritation

- Rabbit 42441601 Signs of irritation present 1 hour after exposure but resolved by day 7 IV

870.2500 Primary Skin Irritation

- Rat 41896203 Mild to moderate erythema, dry cracked skin, edema IV

870.2600 Dermal Sensitization

- Guinea Pig 41618812 negative NA

The toxicology data for repeated

- dose toxicity studies with tribufos are listed in Table A. 2.2.

Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

870.3100 90

- Day oral toxicity

rodents See Subchronic

Neurotoxicity study
870.6200b

870.3200 21

- Day dermal

toxicity

(New Zealand White) rabbits 42007201 ( 1991 )

Acceptable/guideline

0, 2, 11, or 29 mg/kg/day Dermal and systemic NOAEL: 2 mg/kg/day Dermal and systemic LOARL: 11 mg/kg/day based on dermal irritation, muscle fasciculation and anal staining in males and females.

NOAEL = Not established LOAEL = 2 mg/kg/day based on decreased plasma

ChE in male rats and decreased RBC ChE in female

rats.

BMD 10 = 0.22 mg/kg/day

BMDL 10 = 0.16 mg/kg/day based on RBC ChE

inhibition in adult female rats.

870.3465 90

- Day inhalation

toxicity

(Wist a r) rats 42399801 (1992)

Acceptable/guideline

0, 0.93, 2.43, 12.2, or 59.5 mg/m 3
( 0, 0.3, 0.9, 4.5 or 22 mg/kg/day ) NOAEL = 2.43 mg/m 3 (0.9 mg/kg/day) LOAEL = 12.2 mg/m 3 (4.5 mg/kg/day) based on

decreased RBC ChE in male and female rats.

BMD 10 = 1.00 mg/m 3 (HED = 0.12 mg/kg/day)Tribufos Human Health Risk Assessment DP No. D357537

Page 57 of 84 Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

BMDL 10 = 0.79 mg/m 3 (HED = 0.10 mg/kg/day)

based on RBC ChE inhibition in adult male rats

870.3700a Developmental

toxicity

(SD) rats 40190601 (1986 )

Acceptable/guideline

0, 1, 7, 28 m g/kg/day M aternal toxicity NOAEL = 1 mg/kg/day.

M aternal toxicity LOAEL = 7 mg/kg/day based on decreased plasma and RBC ChE.

D evelopmental toxicity NOAEL = 28 m g/kg/day

D evelopmental toxicity LOAEL = Not established

870.3700b D evelopmental

toxicity in (American Dutch )

rabbits 40190602 ( 1987 )

Acceptable/Guideline

0 , 1, 3, or 9 mg/kg/day Maternal toxicity NOAEL = Not established

M aternal toxicity LOAEL = 1 mg/kg/day based on decreased plasma and RBC ChE.

Developmental toxicity NOAEL = 9 mg/kg/day

D evelopmental toxicity LOAEL = Not established

870.3800 Re productive

toxicity ( SD) rats 42040101 ( 1991 )

Acceptable/Guideline

0, 4

, 32 or 250 ppm (0, 0.2, 1.7, or 15 mg/kg/day) for two generations Parental/Systemic NOAEL = Not established Parental/Systemic LOAEL = 4ppm ( 0.2 mg/kg/day )

based on decreased plasma (F0 females) and RBC ChE (F0 females, adult F1 males/females).

Reproductive NOAEL = 32 ppm (1.7 mg/kg/day) Reproductive LOAEL = 250 ppm (15 mg/kg/day)

based on increased number of litters with still born pups and pup death (F1 and F2 generation); decrease in F1 and F2 pup body weight; and increased F1 gestation period.

870.4100b Chronic toxicity

dogs 42007203 (1991 )

Acceptable/Guideline

0, 4, 16, or 64 ppm (0, 0.1, 0.4, 1.7 mg/kg/day in males and 0, 0.1, 0.4, or 2.0 mg/kg/day in females) for 52

weeks Systemic NOAEL = 64 ppm (1.7 mg/kg/day in males and 2.0 mg/kg/day in females) Systemic LOAEL = Not established

Cholinesterase Inhibition NOAEL = 16 ppm (0.4 mg/kg/day) Cholinesterase Inhibition LOAEL = 64 ppm (1.7 mg/kg/day) based on statistically significantly

decreased RBC ChE in males and fem ales.

870.4200a Chronic toxicity /

Carcinogenicity (Fisher) rat 42335101 ( 1992 )

Acceptable/Guideline

0, 4, 40, or 320 ppm (0, 0.2, 1.8, or 16.8 mg/kg/day for male rats and 0, 0.2, 2.3, or 21.1 mg/kg/day for female rats) for 24 months. Systemic NOAEL = 40 ppm (1.8 mg/kg/day for males and 2.3 mg/kg/day for females ). Systemic LOAEL = 320 ppm (16.8 mg/kg/day for

males and 21.1 mg/kg/day for females) based on ocular damage

.

Cholinesterase Inhibition NOAEL = 4 ppm (0.2 mg/kg/day for males and females) Cholinesterase Inhibition LOAEL = 40 ppm (1.8 mg/kg/day for males and 2.3 mg/kg/day for females ) based on statistically significantly decreased RBC

ChE.Tribufos Human Health Risk Assessment DP No. D357537

Page 58 of 84 Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

No evidence of carcinogenicity was observed in

male or female ra ts

.

870.4200b Carcinogenicity

mice 41171001 (1 989 )

Acceptable/Guideline

0, 10, 50, or 250 ppm (0, 1.64, 8.28, or 48.02 mg/kg/day for males and 0,
2.08, 11.14, or 63.4 mg/kg/day for females ) for

90 weeks. Systemic NOAEL = Not established Systemic LOAEL = 10 ppm ( 1.64 mg/kg/day in males and 2.08 mg/kg/day in females) based on

decreased MCV/MCH in males and decreased

hematocrit in females

Cholinesterase Inhibition NOAEL = Not established Cholinesterase Inhibition LOAEL = 10 ppm (1.64 mg/kg/day in males and 2.08 m g/kg/day in females)

based on statistically significantly decreased plasma,

RBC and brain ChE.

Evidence of carcinogenicity was observed at the highest dose only (250 ppm = 48.02 mg/kg/day in males and 63.4 mg/kg/day in females), with increased hemangiosar comas and adenocarcinomas

of the small intestine in males and increased

alveolar/bronchiolar adenomas in females.

870.6100 a Acute delayed

neurotoxicity

- hen Not available. Literature references and an acceptable subchronic study in the hen

show that tribufos produces organophosphate

- induced delayed neuropathy. Therefore, an acute study in the hen is not required.

870.6100b Delayed Subchronic

Neurotoxicity Study

Hen 42007202 (1991)

Acceptable/Guideline

0, 2.6, 11, or 42 mg/kg/day for 90 days Neuropathy NOAEL = 11 mg/kg/day

Neuropathy LOAEL = 42 mg/kg/day based on delayed neuropathy observed in the brain and spinal

cord of hens.

Cholinesterase Inhibition NOAEL = Not established Cholinesterase Inhibition LOAEL = 2.6 mg/kg/day

based on whole bloo d ChE inhibition

870.6200a Acute Neurotoxicity

(Wistar) Rat

(Oral gavage) 45194401 (2000)

Acceptable/Guideline

0, 2, 20, or 100 mg/kg (single dose). Systemic NOAEL = 2 mg/kg Systemic LOAEL = 20 mg/kg based on decreased motor activity.

Cholinesterase Inhibition NOAEL = 2 mg/kg Cholinesterase Inhibition LOAEL = 20 mg/kg based on plasma and RBC ChE inhibition.

870.6200b Subchronic

Neurotoxicity

(Wistar) Rat

(Oral dietary) 45369101 (2001)

Acceptable/Guideline

0, 2, 40 or 500 ppm (00.14, 2.89, or 36.8 mg/kg/day in males and 0, 0.17, 3.54. or 42.6 mg/kg/day in females)

for 13 weeks. Systemic NOAEL = 2 ppm (0.17 mg/kg/day in females)

Systemic LOAEL = 40 ppm (3.54 mg/kg/day in females), based on decreased body weight and food consumption in females.

Chol inesterase Inhibition NOAEL = 2 ppm (0.14 mg/kg/day in males and 0.17 mg/kg/day in females)

Cholinesterase Inhibition LOAEL = 40 ppm (2.89 in males and 3.54 mg/kg/day in females) based on ChETribufos Human Health Risk Assessment DP No. D357537

Page 59 of 84 Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

inhibition in plasma, and RBCs (both sexes) and brain (in female s only).

BMD 10 = 0.27 mg/kg/day

BMDL 10 = 0.19 mg/kg/day based on RBC ChE

inhibition in adult female rats

870.6300 Developmental

Neurotoxicity

(Wistar) Rat

(Oral dietary) 45499501 (2001)

Acceptable/Non

- guideline

0, 4, 40 or 200 ppm ( 0, 0.4,

3.4 , or 16.4 mg/kg/day ) from gestation day 0 through postnatal day 21. Maternal systemic NOAEL = 40 ppm (3.4 mg/kg/day).

Maternal systemic LOAEL = 200 ppm (16.4 mg/kg/day) based on clinical signs (tremor) and decreased body weight.

Maternal c holinesterase Inhibit ion NOAEL = 4 ppm (0.4 mg/kg/day)

Maternal c holinesterase Inhibition LOAEL = 40 ppm (3.4 mg/kg/day) based on decreases in plasma, RBC

and brain ChE activity.

Offspring systemic NOAEL = 40 ppm (3.4 mg/kg/day)

Offspring systemic LOAEL = 200 ppm (16.4 mg/kg/day), based on decreased body weight, body weight gain, delay in preputial separation, delayed surface righting reflex, decreased motor activity, decreased startle response amplitude, decreased absolute brain weights, and changes in the brain morphom etrics (cerebrum and cerebellum).

Offspring c holinesterase Inhibition NOAEL = 200 ppm (16.4 mg/kg/day)

Offspring c holinesterase Inhibition LOAEL = Not

established

870.7485 Metabolism and

pharmacokinetics

(SD) rat 42034501 (199 1 )

Acceptable/Guideline

Rats received single oral dose, 5mg/kg or 100 mg/kg or 5 mg/kg/day X 14 days cold tribufos followed by 5 mg/kg [1

- C 14 ] tribufos. Fifty

- five to 80% was absorbed of which 90+% was excreted in 72 hours. There was no significant tissue

residue. Absorbed m aterial was extensively and completely metabolized.

Tribufos is already in the oxon form

870.7800 Immunotoxicity

Female (Wistar) rat

(oral dietary) 48709904 (2011)

Acceptable/Non

- guideline 0, 5, 50, or 500 ppm (0, 0.43,
4.32, or 44.62 mg/kg/day) for

4 weeks Immunotoxicity NOAEL = 500 ppm (equivalent to 44.62 mg/kg/day)

Immunotoxicity LOAEL = Not established

Cholinesterase Inhibition NOAEL= 5 ppm (0.43 mg/kg/day)

Cholinesterase Inhibition LOAEL = 50 ppm ( 4.32 mg/kg/day) based on significant decreased in RBC

ChE activity.

Special Studies

- Comparative Cholinesterase StudiesTribufos Human Health Risk Assessment DP No. D357537

Page 60 of 84 Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

Comparative

Cholinesterase Studies (PND 11 rats single & repeated dosing

-- dose

- range finding)

(oral gavage) 48709901 (2011)

Acceptable/Non

- guideline

Single

- dosing study: 0, 20, 40 or 50 mg/kg.

Cholinesterase assessments

made at 24 and 48 hours

post

- dosing.

Repeated

- dosing study:

0, 5, 10, 15, or 20 mg/kg/day

for 11 consecutive doses.

Cholinesterase assessments

made 6 hours after the last

dose. Single

- dosing study:

Inhibition of erythrocyte ChE in males and females at 20 mg/kg after 24 hours.

Repeated

- dosing study:

I nhibition of erythrocyte and brain ChE at 5/mg/kg/day in males and females.

Comparative

Cholinesterase Studies (PND 11 rats single dosing study

- time to peak effect)

(oral gavage) 48709902 (2011)

Acceptable/Non

- guideline

Single oral dose of 50 mg/kg Time to peak effect = 6

- 8 hours based on erythrocyte

and brain ChE inhibition ; brain ChE inhibition

decreased by 48 hours.

Comparative

Cholinesterase Studies (adult and pup rats single dosing study)

(oral gavage) 48709904 (2011)

Acceptable/Non

- guideline

Pups: 0, 2, 10 or 50 mg/kg

Adults (females only): 0, 2, 10, or 80 mg/kg

Cholinesterase assessments

made at 24 hours post

- dosing in adults and 6 hours post

dosing in pups Pups:

NOAEL: Not established

LOAEL: 2 mg/kg based on RBC ChE inhibition

Adu l ts:

NOAEL: 10 mg/kg

LOAEL: 80 mg/kg based on brain and RBC ChE

inhibition

BMD 10 = 1.41 mg/kg/day

BMDL 10 = 1.05 mg/ kg/day based on RBC ChE inhibition in male pups

Comparative

Cholinesterase Studies (adult and

pup rats

- repeated dosing study)

(oral gavage) 48709905 (2011)

Acceptable/Non

- guideline

0, 0.1, 1.0, or 5.0 mg/kg/day for 11 days

(females only for adult group)

Cholinesterase assessments made at 24 hours post

- dosing in adults and 6 hours post

dosing in pups Pups:

NOAEL: 1.0 mg/kg/day

LOAEL: 5.0 mg/kg/day based on RBC and brain

ChE inhibition

Adu l ts:

NOAEL: 0.1 mg/kg/day

LOAEL: 1.0 mg/kg/day based on RBC ChE

inhibition

Gestational

Cholinesterase

inhibition 48709906 (2011)

Acceptable/Non

- guideline Maternal systemic NOAEL = 7 mg/kg/day

Maternal systemic LOAEL = 28 mg/kg/day based on decreased food consumption

.Tribufos Human Health Risk Assessment DP No. D357537

Page 61 of 84 Table A. 3. 2.2 Subchronic, Chronic and Other Toxicity Profile

- Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

rats

(oral gavage) 0, 1 (adjusted 0.3

- 0.8

mg/kg/day), 7, or 28 mg/kg/day from GD 6

- 19

Cholinesterase assessments made at 24 hours post

dosing. Maternal c holinesterase Inhibition NOAEL = 0.3

- 0.8 mg/kg/day

Maternal c holinesterase Inhibition LOAEL = 7 mg/kg/day based on decre ases in brai n ChE activity.

Offspring systemic NOAEL = 7 mg/kg/day

Offspring systemic LOAEL = 28 mg/kg/day based on decreased body weight in males

Offspring c holinesterase Inhibition NOAEL 28 mg/kg/day

Offspring c holinesterase Inhibition LOAEL = Not

established

ChE = chol inesterase ; HED = human equivalent dose; RBC = red blood cell.

PoD Conversion Calculations

As summarized in Section 4.5.4, inhalation PoDs were derived using the following conversion

calculations:

Inhalation BMDL 10

The BMDL 10 of 0.79 mg/m 3 (male RBC ChE result from BMD modeling summarized in appendix 1) is first duration

- adjusted as follows:

PoD adj = (BMDL 10 in mg/L) (daily duration adjustment) (weekly duration adjustment)

PoD adj = (0.00079 mg/L) (6h rat exposure/8h expected human) (5 day rat/5 day expected human)

PoD adj = 0.00059 mg/L

The PoD adj is converted to an HEC as follows:

HEC = (regional deposited dose ratio or RDDR) (PoD adj )

HEC = (2.926) (0.00059 mg/L) = 0.0017 mg/L

The HEC is then converted to an HED using the following formula:

HED = HEC (mg/L) x CF(L/hr/kg bw) x D (hr) x AF X A

AF is the ratio of t he exposure specific scenario ventilation rate (8.3 L/min) and the default value

of 13.8 L/min

CF is the default ventilation rate of 13.8 L/min converted to hours and normaliz ed to a body weight of 70 kgTribufos Human Health Risk Assessment DP No. D357537

Page 62 of 84 D is the human daily duration (8 hrs)

A is the absorption ratio as compared to the oral route (assumed to be 1)

Therefore,

HED = (0.0017) (11.8) (8) (0.6) (1)

HED = 0.10 mg/kg/day

Inhalation BMD

Similarly, the BMD of 1.00 mg/m 3 (male RBC ChE result from BMD modeling summarized in appendix 1) is converted to an HED as follows:

PoD adj = (0.001 mg/L) (6h rat exposure/8h expected human) (5 day rat/5 day expected human)

PoD adj = 0.00075 mg/L

HEC = (2.926) (0.00075 mg/L) = 0.0022 mg/ L

HED = (0.0022) (11.8) (8) (0.6) (1) = 0.12 mg/kg/day

A. 3. 3 Hazard Identification and Endpoint Selection

A. 3. 3.1 Acute Dietary Reference Dose (aRfD)

– All Populations

Study Selected: Acute Comparative Cholinesterase (CCA) Study in Rats

- Single Dose

MRID No.: 48707704 Dose and Endpoint for Risk Assessment: A BMDL 10 of 1.05 mg/kg/day associated with RBC

ChE inhibition in PND 11 rat male pups was selected as a suitable P o D for the acute dietary (all popul ations, including females 13

- 49 ) exposure scenari o. The corresponding BMD 10 was 1.41 mg/kg /day

. Comments about Study/Endpoint/Uncertainty Factors: RBC ChE inhibition was selected

for the P o D since the RBCs were more sensiti ve than the brain, and the RBC ChE data were

more robust than brain ChE data . Data from the PND 11 pups represent the highly exposed sub

populations (infants and young children) and thus are appropriate for P o D derivation. Additionally, RBC ChE data were the most sensitive endpoint and indi cated a quantitative susceptibility of the pups compar ed to adult rats in an acute exposure scenario . An uncertainty factor of 100X (10X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10 to obtain an aRfD of 0.01 mg/kg /day . Based on the FQ PA safety

factor of 1 0 X, the acute pop ulation adjusted dose (aPAD) is 0.0 0 1 mg/kg /day

.

Ac ute RfD (All Populations) = (BMDL 10 ) 1.05 mg/kg /day = 0.01 mg/kg /day (UF) 100

aPAD = aRfD = 0.01 mg/kg/day = 0.001 mg/kg/day FQPASF 10

A .3. 3.2 Steady State Dietary Reference Dose (ssRfD)

– All PopulationsTribufos Human Health Risk Assessment DP No. D357537

Page 63 of 84 Study Selected: Subchronic Oral Neurotoxicity Study in Rats

MRID No.: 453 69101 Dose and Endpoint for Risk Assessment: A BMDL 10 of 0.19 mg/kg/day associated with RBC

ChE inhibition in adult female rats was selected since the RBCs were more sensiti ve than the brain, and the RBC ChE data were more robust than brain ChE data. It is, therefore, considered

as a suitable P o D for the steady

- state dietary (all populations) exposure scenario. The corresponding BMD 10 was 0.27 mg/kg/day. Comments about Study/Endpoint/Uncertainty Factors: Although pups are more sensitive than adult rats to ChE inhibition in acute studies, in repeated

- dosing studies ChE is similarly inhibited in pup and adult rats (see table A.5). This is likely due to maturation of enzymes in the pups over the study duration (PND 11

- PND21). Therefore, t he available ChE data at durations

o f 4 weeks in adult female rats provide an endpoint that is protective of highly exposed sub

populations (infants and young children) and, thus, are appropriate for P o D derivation. An uncertainty factor of 100X (10X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10 to obtain a n ssRfD 17 of 0.0019 mg/kg/day. Based on the FQPA safety factor of 1 0 X, the steady

- state population adjusted dose (ssPAD) is

0.00 0 2 mg/kg/day.

ssRfD (All Populations)= ssPAD = (BMDL 10 ) 0.19 mg/kg/day = 0.002 mg/kg/day (UF) 100

ssPAD = ss RfD = 0.002 mg/kg/day = 0.0002 mg/kg/day

FQPASF 10

A. 3. 3.3 Incidental Oral Exposure (Steady State)

Study Selected: Subchronic Oral Neurotoxicity Study in Rats

MRID No.: 453 69101 Dose and Endpoint for Risk Assessment: A BMDL 10 of 0.19 mg/kg/day associated with RBC

ChE inhibition in adult female rats was selected since the RBCs were more sensiti ve than the brain, and th e RBC ChE data were more robust than brain ChE data. It is, therefore, considered

as a suitable P o D for the steady

- state dietary (all populations) exposure scenario. The corresponding BMD 10 was 0.27 mg/kg/day. Comments about Study/Endpoint/Uncertainty F actors: Although pups are more sensitive than adult rats to ChE inhibition in acute studies, in repeated

- dosing studies ChE is similarly inhibited in pup and adult rats (see table A.5). This is likely due to maturation of enzymes in the pups over the stud y duration (PND 11

- PND21). Therefore, t he available ChE data at durations

o f 4 weeks in adult female rats provide an endpoint that is protective of highly exposed sub

populations (infants and young children) and, thus, are appropriate for P o D derivation. An uncertainty factor of 100 0 X (10X to account for interspecies extrapolation and 10X for intra

species variation and 10X FQPASF) is applied.

17 ssRfD: Reference dose derived from steady state durationTribufos Human Health Risk Assessment DP No. D357537

Page 64 of 84 A.3 .3 .4 Dermal Exposure ( Steady State)

Study Selected: 21

- Day Dermal Study in Rabbits

MRID No.: 42007201 Dose and Endpoint for Risk Assessment: A BMDL 10 of 0.16 mg/kg/day associated with RBC ChE inhibition in adult female rats, following dermal exposure, was selected as a suitable P o D for assessing the potential risk associated with dermal exposure (s hort

- term intermediate ). The corresponding BMD 10 was 0.22 mg/kg/day. Comments about Study/Endpoint/Uncertainty Factors: This is the only dermal study

available for tribu fos and wa s an acceptable/ guideline study. RBC ChE inhibition was selected

for the P o D since the RBCs were more sensiti ve than the brain through the dermal route of

exposure. An uncertainty factor of 100X (10X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10.

A.3.3 .5 Inhalati on Expos ure ( Steady State)

Study Selected: 13

- Week Inhalation Toxicity Study in Rats

MRID No.: 42399801 Dose and Endpoint for Risk Assessment: A BMDL 10 of 0.79 mg/m 3 (HED = 0.10 mg/kg/day)

associated with RBC ChE inhibition in adu lt male rats following inhalation exposure was selected as a suitable P o D for assessing the potential risk associated with inhalation exposure (s hort

- term intermediate ). The corresponding BMD 10 was 1.00 mg/m 3 (HED = 0.12 mg/kg/day)

. Comments about Study/Endpoint/Uncertainty Fa ctors: The particle size distribution from this study suggests that most particles are within the respirable range for an aerosol (1

- 3 μm),

thus the BMD and BMD 10 were converted to human equivalent concentrations (HECs) followed by conversion to human eq uivalent doses (HEDs). An uncertainty factor of 30X (3 X to account for interspecies extrapolation and 10X for intraspecies variation) is applied to the BMDL 10.

A. 3. 4 Executive Summaries

A. 3. 4.1 Subchronic Toxicity

870.3100 90

- Day Oral Toxicity

– Rat

See Subchronic Neurotoxicity study 870.6200b [MRID 45369101 ]

870.3150 90

- Day Oral Toxicity

– Non

- Rodent

Subchronic oral toxicity studies in non

- rodent species are not available. However, these studies are not required because acceptable chronic stud ies are available in the dog.

870.3200 21/28

- Day Dermal Toxicity

– Rabbit

In a 21

- day dermal toxicity study (MRID 42007201), groups of New Zealand White rabbitsTribufos Human Health Risk Assessment DP No. D357537

Page 65 of 84 (10/sex/dose) received repeated dermal applications of tribuf os at doses of 0, 2, 10 or 25 mg/kg/day nominal (0, 2, 11 or 29 mg/kg/day actual), six hours per day, five da ys per week over a period of 21 day s

. Polyethylene glycol 400 was used as the solvent and applied neat to the control animals. The application sit e in the clipped back of the rabbits approximated 240 cm 2

. The day after the last treatment, 5 animals/sex/group were sacrificed, the remaining animals were sacrificed 14 days after the last treatment. Animals were observed daily for clinical signs and the application site graded for signs of irritation. Body weight and food consumption were determined weekly. Ophthalmic examination was performed on both eyes of the survivors on week 3 of treatment. Blood samples were collected 24 hours after the last dose and at sacrifice.

No mortality occu rred at 2 or 11 mg/kg/day where as one male and four females died or were

sacrificed in extremis at 29 mg/kg/day. Mild to moderate dermal irritation was observed at 11 and 29 mg/kg/day in both sexes. Signs of dose

- rela ted toxicity (muscle fasciculations, anal stain) were observed in both sexes at 11 and 29 mg/kg/day, with a greater effect at the higher dose. At termination, dose

- related inhibition of plasma, erythrocyte (RBC) and brain cholinesterase ( ChE ) activity was observed in both sexes at all dose levels. Statistically

- significant (p<0.05) inhibition

of ChE was observed in plasma (males) and RBC (females) at 2 mg/kg/day and in all compartments (plasma, RBC and brain) at 11 and 29 mg/kg/day in both sexes. No reco very was observed in RBC and brain ChE activity at 33 to 34 days (14 days post

- dose).

The dermal and systemic NOAEL for tribufos in ra bbits is 2 mg/kg/day and the LOAEL is 11 mg/kg/day based on mild to moderate dermal irritation, fasciculations and anal stain in

both males and females. The NOAEL for cholinesterase inhibition is not identified and the LOAEL for cholinesterase inhibition is 2 mg/kg/day based on decreased plasma (males) and RBC (females) cholinesterase activity.

This study is classified as Acceptable/Guideline and does satisfy the guideline requirements for a repeated

- dose dermal study [OPPTS 870.3200 (§82

- 2)] in ra bbits

.

870.3465 Subchronic Inhalation

– Rat

In a subchronic toxicity study (MRID 42399801), groups of Wistar rats (10/sex/concentration) were exposed via inhalation to an aerosol of tribufos at concentrations of 0, 0.93, 2.43, 12.2 or 59.5 mg/m 3 actual (0, 0.3, 0.9, 4.5, 22 mg/kg/day), six hours per day, five days per week for 90 days. Rats were weighed pr ior to the first exposure and weekly during the study. Appearance and behavior were evaluated daily. Blood samples were collected monthly and at necropsy for hematological and clinical chemistry tests. Electroretinographic (ERG) tests were performed during week 10 and prior to necropsy on 5 males and 5 females each from the control and high dose group.

No ChE inhibition was observed in either sex at 0.93 or 2.43 mg/m 3 . Plasma ChE inhibition was

observed in males at 12 .2 and 59.5 mg/m 3 and in females at 59.5 mg/m 3 . RBC ChE inhibition

was observed at 12 .2 and 59.5 mg/m 3 in both sexes. Brain ChE inhibition was seen at 59.5 mg/m 3 both sexes. The adrenals showed cortical fat deposition at 59.5 mg/m 3 in both sexes.

Electroretin ogram (ERG ) was depressed (a

- and b

- waves) at 59.5 mg/m 3 in both sexes indicative

of a toxic effect on t he rods and cones of the retina.Tribufos Human Health Risk Assessment DP No. D357537

Page 66 of 84 The NOAEL is 2.43 mg/m 3 (0.9 mg/kg/day) and the LOAEL is 12 .2 mg/m 3 (4.5 mg/kg/day)

based on RBC cholinesterase inhibition.

This subchronic inhalation toxicity study in the rat is Acceptable/Guideline and does satisfy the guideline requirements for a subchronic inhalation study OPPTS 870.3465; OECD 413 in the rat.

A. 3. 4.2 Prenatal Developmental Toxicity

870.3700a Prenatal Developmental Toxicity Study

– Rat

In a developmental toxicity study (MRID 40190601), pregnant Crl:COBS

- CD (SD) rats received oral doses of tribufos at 0, 1, 7 or 28 mg/ kg/day during gestation days (GD) 6 thr ough 16. Animals were observed daily and weighed on GD 0, 6, 8, 10, 12, 15, and 20. Food consumption recorded on GD 1, 6, 8, 12, 15, and 20. Dams were sacrificed on GD16 (5 dams/dose) or GD 20 (10 dams/dose), and blood and brain were collected for determination of cholinesterase activity.

Da ms were subjected to gross necropsy, and all fetuses examined externally. Approximately

one

- half of each litter was processed for visceral examination and the remaining one

- half was processed for skeletal examination. Fetal brain cholinesterase was measur ed in 20 fetuses from each dose group.

All animals survived to terminal sacrifice. No clinical signs of toxicity were observed, except for salivation in two high dose animals on GD 9 and 12. Maternal body weights and food consumption were similar betwee n the treated and control groups throughout the study. On GD16, plasma and RBC ChE were inhibited at 7 and 28 mg/kg/day, while brain ChE was inhibited at 28 mg/kg/day. On GD 20, RBC ChE was inhibited at 7 and 28 mg/kg/day, while

brain ChE was inhibited at 28 mg/kg/day. Maternal necropsy was unremarkable. Therefore, the maternal toxicity NOAEL is 1 mg/kg/day and the maternal toxicity LOAEL is 7 mg/kg/day,

based on inhibiti on of plasma and RBC cholinesterase activity

.

No differences were observed between the treated and control groups for number of corpora lutea, number of implantation sites, live fetuses/dam, pre

- and post

- implantation losses, fetal body weights, or fetal sex ratios. No treatment

- related external, visceral, or skeletal

malformations/variatio ns were observed in any fetus from any group. Therefore, the developmental toxicity NOAEL is 28 mg/kg/day and the developmental toxicity LOAEL

was not identified.

This study is classified as Acceptable/Guideline and satisfies the requirements for a

develo pmental toxicity study [OECD 870.3700 (§83

- 3a)] in rats.

870.3700b Prenatal Developmental Toxicity Study

– Rabbit

In a developmental toxicity study (MIRD 40190602) , pregnant American Dutch rabbits (17/dose) were administered tribufos via oral gavage at 1, 3, or 9 mg/kg/day during gestation days (GD) seven through 19. Does were observed daily for morbidity and mortality. BodyTribufos Human Health Risk Assessment DP No. D357537

Page 67 of 84 weights were obtained on GDs 0, 7, 10, 14, 19, 21 and 28. Food consumption was measured on GDs 1, 6, 8, 12, 15, 20, 23, and 28. Bl ood was obtained for RBC and plasma ChE determinations on GD 20 (24 hours after administering the last dose) and on GD 28 (prior to sacrifice). Half the

brain of each doe was collected at termination for brain ChE measurements. On GD 28, all does

were sacr ificed and the uteri were weighed and examined. All fetuses were dissected, sexed and

eviscerated. All fetuses were examined for visceral and bone anomalies.

No treatment

- related clinical signs were observed in does at any dose. Two low dose does and

on e high dose doe died of respiratory disease. One low dose doe and one high dose doe aborted during the study. No treatment

- related effects were observed on food consumption. Statistically significant decreases in body weight gain were observed in does from the high

- dose group. Plasma and RBC ChE activities were decreased in does from all dose groups at GD 20. RBC ChE activity remained decreased at GD 28. No treatment

- related changes in brain ChE were reported. Therefore , the maternal toxicity LOAEL i s 1 mg/kg/day based on inhibition of plasma and RBC ChE activity; a NOAEL i s not established

.

No developmental toxicity was observed

. Therefore, the developmental toxicity NOAEL for tribufos in rabbits is 9 mg/kg/day. The developmental toxicity LOAEL is not established.

This study is classified as Acceptable/Guideline and does satisfy the guideline requirements for a developmental toxicity study [OECD 870.3700 (§83

- 3b)] in rabbits.

A. 3. 4.3 Reproductive Toxicity

870.3800 Reproduction and Fertility Effects

– Rat

In a two

- generation reproduction study (MRID 42040101), Sprague

- Dawley rats were fed diets containing tribufos at 0, 4, 32 or 260 ppm (0, 0.2, 1.7, or 15 mg/kg/day) for two successive generations. Thirty animals/sex/dose (F0) were fed tribu fos in the diet for 10 weeks and then bred within their test groups. Thirty males and thirty females were selected from the weanlings of each test group to parent the second generation (F1). After 10 weeks on the test diet, F1

animals were bred within the ir test group. Animals were observed daily for morbidity, mortality and clinical signs. Body weight and food consumption were measured weekly for each male and female. During gestation, female body weight was determined on days 0, 6, 13, and 20 of gestatio n and days 1, 4, 7, 14, and 21 of lactation. Food consumption in females was measured weekly during gestation, twice during week one of lactation and once a week during weeks 2 and

3 of lactation. F0 and F1 males were sacrificed after the last F1 and F2 li tters, respectively, were delivered or the dams had gone past day 24 of gestation. F0 and F1 adult females were sacrificed after each dam’s pups were weaned/died or when day 24 of gestation was reached. All adult animals were subjected to a complete gross necropsy.

Live and still born pups and sex distribution were determined for each litter. Littler counts were performed daily and pup weights obtained within 24 hours of birth and on days 4, 7, 14, and 21. Each litter was culled on day 4, with 8 pups (4 ma les, 4 females) selected for sacrifice and necropsy. Plasma and RBC ChE were determined on F0 and F1 adults during week 8 and justTribufos Human Health Risk Assessment DP No. D357537

Page 68 of 84 prior to sacrifice while brain ChE was determined at termination. Plasma, RBC, and brain ChE

were determined for one male an d one female pup each from 10 litters at culling (day 4) and at weaning (day 21 post

- partum).

No treatment

- related effect on mortality was observed in adult rats. In F0 dams in the high

- dose group, body weight was decreased on day 20 of gestation and throughout the lactation period while food consumption was decreased during lactation. F1 dams in t he high

- dose group exhibited decreased body weight throughout the lactation period and decreased food consumption on days 14 and 21 of lactation. Birth index, live birth index, viability index and litter size were significantly reduced in F0 dams in the hi gh

- dose group. Birth index, live birth index, fertility index, and viability index were significantly decreased in F1 dams in the high

dose group. Plasma, RBC and brain ChE activity was significantly decreased in F0 and F1 adults

in the mid

- and high

- dose groups. Plasma ChE activity was also significantly decreased in F0

females in the low

- dose group while RBC ChE activity was decreased in F0 females as well as

F1 adults in the low

- dose group. Therefore, the parental systemic toxicity LOAEL is 4 ppm (0.2 mg /kg/day), the lowest dose tested, based on inhibition of plasma (F0 females) and RBC (F0 females, adult F1 males/females) cholinesterase activity. A parental/systemic toxicity NOAEL is not established.

The number of still births in F1 and F2 litters as well as the gestation length for F2 litters was significantly increased in the high

- dose group. Mean body weights for F1 and F2 pups in the high

- dose group were significantly reduced. Although brain and RBC ChE activity were significantly inhibited in F1 f emale pups at day 4 and F2 pups at day 21, no clear dose

- response was observed. Plasma ChE was significantly inhibited on day 21 in F1 females in the mid

- and high

- dose groups. However, in accordance with current policy inhibition of plasma

cholinesterase activity alone in the absence of clinica l signs is not considered appropriate for use

as the basis for a LOAEL. There was no evidence of increased susceptibility of pups over the

adults. For reproductive toxicity, the NOAEL was 32 ppm (1.7 mg/kg/day) and the LOAEL was 260 ppm (15 mg/kg/day) based on a significant increase in the number of litters with still born pups and pup death (including cannibalism) through lactation; decrease in F1 and F2 pup body weights ; and significant increase in the gestation pe riod for F2 litters

.

This study is classified as Acceptable/Guideline and satisfies the guideline requirement for a reproduction study (OPPTS 870.3800 §83

- 4) in rats.

A. 3. 4.4 Chronic Toxicity

870.4100a (870.4300) Chronic Toxicity

– Rat

See Carcinogenicity section below.

870.4100b Chronic Toxicity

- Dog

In a chronic toxicity study (MRID 42007203), groups of four male and four female Beagle dogsTribufos Human Health Risk Assessment DP No. D357537

Page 69 of 84 were fed diets containing tribufos at doses of 0, 4, 16 or 64 ppm (equivalent to 0, 0. 1, 0.4, or 1.7 mg/k g/day in males and 0, 0.1, 0.4, or 2.0 mg/kg/day in females, respectively) for 52 weeks. Individual body weights and feed consumption were recorded weekly and daily, respectively. Animals were observed twice daily for morbidity and mortality. Det ailed physical examinations and body temperatures were taken weekly. Ophthalmic examinations were performed before dosing and prior to sacrifice. Clinical neurological examinations, clinical chemistry, hematology and urinalysis were performed on all dogs b efore dosing and at 3, 6, 9, and 12 months. Gross pathological examinations were performed on all animals at termination.

No abnormalities were observed during clinical, neurological or ophthalmological examinations. Therefore, the systemic NOAEL is 64 pp m (1.7 mg/kg/day in males and 2.0 mg/kg/day ) . A

LOAEL is not established.

Inhibition of plasma ChE activity was observed in both sexes at 16 ppm. However, in accordance

with current policy inhibition of plasma cholinesterase activity alone in the absence of clinica l signs is not considered appropriate for use as the basis for a LOAEL. Inhibition of erythrocyte ChE activity was observed in both sexes at 64 ppm. A possible decrease in the number of erythrocytes at 64 ppm was observed in both sexes. No oth er toxic effects were observed. Therefore, the ChE inhibition NOAEL is 16 ppm (0.4 mg/kg/day) and the LOAEL is 64

ppm (1.7 mg/kg/day in males; 2.0 mg/kg in females) based on decreased erythrocyte ChE activity.

This chronic oral toxicity study in dogs is classified as Acceptable/Guideline and satisfies the guideline requirement for a chronic oral toxicity study [OPPTS: 870.4100b (§83

- 1b )] in dogs.

A. 3. 4.5 Carcinogenicity

870.4200a Carcinogenicity Study

- Rat

In a combined chronic toxicity/carcinogenicity study (MRID 4255360) , Fischer rats (50/sex/dose) received diets containing tribufos at doses of 0, 4, 40 or 320 ppm (equivalent to 0.0, 0.2, 1.8 and 16.8 mg/kg/day in males and 0.0, 0.2, 2.3 and 21.1 mg/kg/day in females, respectiv ely) for 24 months

. Body weights and food consumption and body temperature were determined weekly. Clinical observations were recorded daily and a detailed physical examination performed weekly. Prior to dosing and just prior to sacrifice, ophthalmic exam s were conducted on all animals in the study. Blood was drawn for clinical pathology determinations at 3, 6, 12, 18 and 24 months of the study.

Complete bilateral retinal atrophy (obliteration) was observed at 12 mont hs at the high dose (320 ppm). At 24 months statistically

- significant ocular damage at the high dose included cataract, lens opacity, corneal opacity, corneal neovascularization and bilateral retinal atrophy (obliteration). At d oses of 4 and 40 ppm
, terminal retinal atrophy was generally uni lateral and histopathologically different from that seen at the high dose. Other t reatment

- related effects

observed included: at 4 ppm decreased plasma ChE was observed in both sexes; at 40 ppm decreased weight gain, cholesterol and calcium were observed in males; and decreased RBCTribufos Human Health Risk Assessment DP No. D357537

Page 70 of 84 ChE, RBC count, hemoglobin, and hematocrit were obser ved in both sexes. At 320 ppm, decreased weight gain was observed in the females. In addition, both sexes of rats at 320 ppm exhibited the following effects: increased food consumption, cataract, lens and corneal opacity, corneal neovascularization, iritis/uveitis; decreased total protein, globulin, cholesterol, calcium; increased blood urea nitrogen (BUN); decreased brain ChE adrenals; vacuolar degeneration (12 month); reti nal atrophy (12 month); autolysis, vacuolar degeneration in the small intestines (12 and 24 months); retinal atrophy, uveitis, cataract, neovascularization (24 month); atrophy of the optic nerve (24 month); vacuolar degeneration, hyperplasia of the small i ntestines (24 months). There was no evidence of carcinogenicity in rats.

For chronic toxicity, the LOAEL is 320 ppm (16.8 mg/kg/day for males and 21.1 mg/kg/day for females), based on ocular damage in both sexes. The NOAEL is 40 ppm (1.8 mg/kg/day

for males and 2.3 mg/kg/day for females). For ChE inhibition, the N OAEL is 4 ppm ( 0.2 mg/kg/day for males and females), and the LOAEL is 40 ppm (1.8 mg/kg/day for males and 2.3 mg/kg/day for females) based on decreased RBC ChE activity

.

The chronic toxicity/carcinogenicity study in the rat is A cceptable/ G uideline and satisfies the guideline requirements for a chronic toxicity /carcinogenicity oral study (OPPTS 870.4200a ; §

83

- 1) in rats.

870 .4200b Carcinogenicity Study

– Mouse

In a carci nogenicity study (MRID 41171001)

, CD

- 1 mice (50/sex/dose) were fed diets containing tribufos at doses of 0, 10, 50 or 250 ppm for 90 weeks. These doses were equivalent to 0, 1.64, 8.28 or 48.02 mg/kg/day in males and 0, 2.08, 11.14 or 63.4 mg/kg/day in fe males. Complete blood counts were performed in 10 mice/sex/dose at 12 months, 18 months and at

termination or when animals were sacrificed in extremis . Plasma and RBC ChE activity was determined in 10 mice/sex/dose at 12 months, 18 month and at terminatio n. Brain ChE activity was measured in the same mice at termination of the study. Gross pathology was performed on all mice found dead, sacrificed in extremis and at termination.

At 78 weeks, males in the 10 ppm group showed decreased mean corpuscular volu me (MCV) and mean corpuscular hemoglobin (MCH) and, at week 90 females in the 10 ppm group showe d

decreased hematocrit. A n increased number of males in the 50 ppm group showed paleness and

hunched b acks was well as decreased MCV and MCH at week 78 and decreased MCH at week

90 . At week 90 females in the 50 ppm group showed decreased RBC count, hemoglobin and

hematocrit.

Histopathology of males in the 50 ppm group showed: adrenals amyloid, epididymis hyperspermatogenensis, small intestine amyloid and vacuolar degeneration epithelium, and spleen hematopoiesis. At 250 ppm loose stools were observed in females, enlarged abdomen in both sexes, increased mortality/decreased life span in both sexes, and increased food consumption and body weight in both sex es. Decreased RBC count, hemoglobin, hematocrit, MCV and MCH was observed in males and decreased RBC count, hemoglobin and hematocrit in

females. There was evidence of carcinogenicity in mice only at the highest dose tested ( 250 ppm

or 48.02 mg/kg/day in males and 63.4 mg/kg/day in females)

. Histopathology of males in the 250Tribufos Human Health Risk Assessment DP No. D357537

Page 71 of 84 ppm group showed: adrenals degeneration, liver hemangiosarcoma, rectum acute inflammation, necrosis aid ulcer, small intestine adenocarcinoma, dilated/distended and mucosal hyperpla sia.

In females in the 250 ppm group , histopathology showed: adrenals calcification and degeneration/pigmentation, caecum edema, liver hypertrophy, lung alveolar/bronchiolar adenoma, mesenteric lymph node congestion, rectum acute inflammation, necrosis a nd ulcer, and small intestine adenocarcinoma dilated/distended, mucosal hyperplasia). Statistically

- significant decreases in plasma , RBC and brain ChE activity were observed in both sexes at all dose levels.

The LOAEL is 10 ppm (1.64 mg/kg/day in males and 2.08 mg/kg/day in females) based on decreased MCV/MCH in males, decreased hematocrit in females as well as decreased plasma, erythrocyte and brain cholinesterase in both sexes. A NOAEL is not established.

The study is classified as A cceptable/ G uideli ne and satisfies the guideline requirements for a carcinogenicity toxicity study in mice (OPPTS 870.4200 ; §83

- 5 ).

A. 3. 4.6 Mutagenicity

Mutagenicity Studies with Tribufos

Guideline

No. Study Type MRID No. (year)/

Classification /Doses Results

870.5100 Gene Mutation

-

Salmonella 41459101 (1990)

Acceptable/Guideline

TA98, TA100, TA1537 and

TA1538 strains

667, 1000, 3333, 6667, and

10000  g/plate Not mutagenic with or without microsomal activation at concentrations up to 10,000  g/plate.

870.5300 Gene Mutation

Mammalian cells This study is not available but the HED Carcinogenicity Peer Review Committee

evaluated tribufos carcinogenicity data (TRX#0012242) and concluded that there is no apparent concern for mutagenicity. Therefore, this s tudy is not needed.

870.5375 Chr omosome

A berrations

Chinese Hamster Ovary cells 41459103 (1990)

Acceptable/Guideline

0.0005

- 50  l/mL Negative for chromosomal aberrations with or

without microsomal activation . Doses tested without activation showed toxicity at 0.025  l/mL and 0.05

 g/mL. Doses tested with activation showed toxicity

at 0.05  l/mL and 0.1  l/mL.

870.5397 Mammalian

Erythrocyte

Micronucleus This study is not available but the HED Carcinogenicity Peer Review Committee

evaluated tribufos carcinogenicity data (TRX#0012242) and concluded that there is no apparent concern for mutagenicity. Therefore, this study is not needed.

870.5500 Unscheduled DNA

Synthesis

Primary rat hepatocytes 41459102 (1990)

Acceptable/Guideline

0.0003

- 10  l/mL Negative in primary rat hepatocytes at

concentrations of 0.0001

- 0.006  l/mL. Higher

concentrations were toxic.

A. 3. 4.7 Neurotoxicity

870.6100b Delayed Neurotoxicity Study

– HenTribufos Human Health Risk Assessment DP No. D357537

Page 72 of 84 In a delayed n eurotoxicity study (MRID 42007202)

, t ribufos was applied to the comb of 12 hens at doses of 0, 2.6, 11, or 42 mg/kg/day for 13 weeks (a total of 65 applications) . Triortho

cresolphosphate (TOCP) was utilized as a positive contro l at 18 mg/kg/day. Hens were observed daily d uring treatment for clinical signs and weighed on day 0 and weekly thereafter. Whole blood cholinesterase activity was determined prior to initiation of the study, at one week and at one, two and three months of exposure. All surviving hens were sacrificed 10

- 13 days after the last treatment to accommodate observation of possible recovery.

Body weight was significantly decreased in the 42 mg/kg/day dose group and in the TOCP positive control group. Histopathology indicative of organophosphate induced neuropathy (OPIDN) was observed primarily in the brain and spinal cord of hens at the highest dose tested (42 mg/kg/day). Whole blood cholinesterase inhibition was observed at the lowest dose tested (2. 6 mg/kg/day ). Therefore, the OPIDN NOAEL is 11 mg/kg/day and the LOAEL is 42 mg/kg/day based on delayed neuropathy observed in the brain and spinal cord of hens. For Cholinesterase inhibition, the LOAEL is 2.6 mg/kg/day based on whole blood

cholinesteras e inhibition; a NOAEL is not established.

870.6200a Acute Neurotoxicity Screening Battery

– Rat

In an a cute neurotoxicity study ( MRID No.: 45194401), four groups of 18/sex Wistar s train rats

were dosed with 0 , 2, 20 or 100 mg/kg of tribufos (98% purity in corn oil at 5 mL/kg). Twelve rats/sex per group were assessed for clinical signs, body weight, FOB and motor activity at pretest, estimated time for peak effects (six hours post dosing based on preliminary studies), and on days 7 and 14 postdosing. At termination the 6 rats/sex/group were perfused and assessed histologically for neuropathological effects. The remaining 6 rats/sex/group were assessed for cholinesterase at pretest and the time of peak effect.

N o lesions in the nervous system were attributed to tribufos treatment. At 20 mg/k g statistically significant decre ased motor and locomotor activity was evident in males. In females statistical significance was not attained but decreases in motor activity and in locomotor activity were

appare nt. At 100 mg/kg the decreases in motor and locomotor activity were larger to indicate a dose response and the locomotor activity decrease in females was statistically significant. There was also decreased rearing and arousal. Body temperature was decre ased 1.9  C and 2.8  C in

males and females

, respectively. Two deaths among the females on days 2 and 4 were attributed

to tribufos. The LOAEL is 20 mg/kg based on decreased motor activity. The NOAEL is 2 mg/kg.

Plasma ChE was statist ically signi ficantly inhibited at 20 mg/kg and at 100 mg/kg when assess ed 6 hours after dosing. RBC ChE was also statis tically significantly inhibited at 20 mg/kg and 100 mg/kg. Brain ChE was statistically significantly inhibited in the 100 mg/kg female dose group only. The LOAEL for cholinesterase inhibition is 20 mg/kg based on inhibition of both plasma and RBC ChE. The NOAEL is 2 mg/kg. This acute neurotoxicity study is classified Acceptable/Guideline and satisfies the guideline requirement for an acute neu rotoxicity study [OPPTS 870.6200a (§ 81

- 8) ] in rats.Tribufos Human Health Risk Assessment DP No. D357537

Page 73 of 84 870.6200b Subchronic Neurotoxicity Screening Battery

– Rat

In a subchr onic neurotoxicity study ( MRID 45369101), Tribufos (98. 0

- 98.1% purity ) was administered to 18 young adult Wistar rats/sex at dietary dose levels of 0, 2, 40, or 500 ppm (equivalent to mean doses of 0, 0.14, 2.89, or 36.8 mg/kg/day for males and 0, 0.17, 3.54, or 42.6 mg/kg/day for females, respectively) for 13 weeks. Neuro behavioral assessment (functional observational battery (FOB) and motor activity testing) was performed in 12 animals/sex/group at pretest and at weeks 4, 8 and 13. Cholinesterase was assessed in 6 rats/sex/dose for plasma and erythrocytes at pre

- treatmen t and at weeks 4 and 13, and in whole brain at week 13. At study termination, 6 control and high dose animals/sex/group were perfused in situ for neuropathological examination of brain and peripheral nervous system tissues.

M ean body weights and food co nsumption were decreased for females in the 40 ppm group and

for males and females in the 500 ppm group . Treatment

- related clinical signs were limited to red nasal staining in one high

- dose male on day 4, perianal staining in one high

- dose male on day 11, and urine staining in two high

- dose females from days 53

- 88. FOB effects included sluggish/minimal movement and decreased approach response in the open field in high

- dose males and a decreased number of rearing counts in the open field was noted in high

- dose females, both at week 4 only. Slight yellow urine staining was noted for 1/12 high

- dose females at week 8 and for 2/12 high

- dose females at week 13. Although statistical significance was not attained, these effects are considered treatment

- related. Motor activity was decreased in high

dose males and females during weeks 4, 8, and 13 . Locomotor act ivity was significantly decreased in high

- dose males and females at weeks 4, 8, and 13, with decreases averaging 32

- 55% for males and 48

- 53% for females . Neuropathology indicated retinal degeneration (mostly bilateral) was noted in 5/6 high

- dose males and 6/6 high

- dose females but was not seen in the

controls or the mid dose. The LOAEL in females i s 40 ppm (3.54 mg/kg/day) based on decreased body weight and decreased food consumption. The NOAEL i s 2 ppm (0.17 mg/kg/day). The LOAEL for males i s 500 ppm (36.8 mg/kg/day) based on decreased body weight, decreased food consumption, and motor activity decrease and retinal degeneration.

The NOAEL i s 40 ppm (2.89 mg/kg bw/day).

RBC and plasma ChE were significantly inhibited at 40 ppm and 500 ppm in both males and

females . Brain cholinesterase activity was decreased in f emales at 40 ppm and a t 500 ppm in

males and females

. The NOAEL for ChE inhibition is 2 ppm (0.14 mg/kg/day in males and 0.17 mg/kg/day in females) and the LOAEL is 40 ppm (2.89 ppm in males and 3.54 mg/kg/day in females) based on ChE inhibition in plasma and RBC (both sexes) and brain (females only).

The study is classified as Acceptable/Guideline and satisfies the guideline requirement for a subchronic neurotoxicity study in rats (870.6200b).

870.6300 Developmental Neurotoxicity

– RatTribufos Human Health Risk Assessment DP No. D357537

Page 74 of 84 In a developmental neurotoxicity study (MRID 45499501), Tribufos (98.0

- 98.1 % a.i. ) was

adm inistered to parent female Wistar rats in the diet at concentrations of 0, 4, 40 or 200 ppm from gestation day 0 through postnatal day 21. The average daily intake of Tribufos was 0, 0.4,

3.4

- 3.5, and 16.4

- 18.2 mg/kg bw/day during gestation and 0, 0.6

- 1.0 , 6.1

- 9.9, and 33.5

- 55.4 mg/kg/day during lactation, for the 0, 4, 40, and 200 ppm groups, respectively. Two studies,

with a two

- week difference in start date, were conducted under similar conditions. The main study (00

- D72

- AG) included 20 parent females /dose level, and the satellite study (00

- D2

- AS) included 10 parent females/dose level and was utilized to provide samples to assess cholinesterase activity. A Functional Operational Battery (FOB) was performed on 10 dams/dose from the main study on gestat ion days 6, 13, and 20 and on and lactation days 4, 11, and 21. On postnatal day 4, litters were culled to yield four males and four females (as closely as possible). Offspring from the main study, representing at least 20 litters/dose, were allocated fo r detailed clinical observations (abbreviated FOB), assessment of motor activity, assessment of auditory startle response habituation, assessment of learning and memory, electroretinography, and neuropathology at study termination (day 75 of age). Animal s from the satellite study were used to provide blood and brain samples for the offspring (postnatal days 11 and 21) and dams (lactation day 21) for measureme nt of cholinesterase (ChE) activity

.

No treatment

- related effects were seen on survival, food con sumption or reproductive parameters. Dams exhibi ted clinical signs of toxicity (slight tremors) at parturition which are likely associated with the observed ChE inhibition. Maternal body weight during lactation was decreased at each time point recorded fo r the high

- dose group, with average differences of 8

- 12% compared to the control.

The maternal systemic LOAEL is 200 ppm (16.4 mg/kg/day) based on clinical signs (tremor) and decreased body weight during lactation. The maternal systemic NOAEL is 40

ppm (3.4 mg/kg/day).

Plasma, brain and RBC ChE activity was decreased in maternal animals at 40 and 200 ppm. Therefore, t he maternal ChE LOAEL is 40 ppm (3.4 mg/kg/day) based on decreases in plasma, RBC and brain ChE activity. The maternal ChE NOAEL is 4 ppm (0.4 mg/kg bw/day).

Treatment had no adverse effects on offspring survival. No treatment

- related effects on body weight or body weight gains were seen at 4 or 40 ppm pups. At the high d ose (200 ppm), pup body weights were significantly d ecreased on PNDs 4, 1 1 and 21 . Preputial separation for high

dose males was delayed relative to controls. Additionally, the development of a surface

- righting reflex in high do se animals was significantly delayed. Motor activity was decreased in males

and females at the high dose on PND 13. Conversely, motor activity was increased in males and

females at the high dose on PND 17 . There were no apparent effects on motor activity in males or females on PND 21 and PND 60. Startle response amplitude for high

- d ose males and females was decreased compared to controls on day 21, but not on subsequent test days. No treatment

related effects were seen on learning and memory. Absolute brain weights of high

- dose males and females were decreased compared to controls on days 11 and 21

. The anterior to posterior lengths of the cerebrum were marginally decreased for high

- dose males and females compared to controls on PND 11. The anterior to posterior length for the cerebellum was also marginallyTribufos Human Health Risk Assessment DP No. D357537

Page 75 of 84 decreased in high

- dose males and females, although the effect did not reach statistical significance

The offspring systemic LOAEL is 200 ppm (16.4 mg/kg/day), based on decreased body weight, body weight gain, delay in preputial separation, delayed surface righting reflex,

decre ased motor activity, decreased startle response amplitude, decreased absolute brain weights, and changes in the brain morphometrics (cerebrum and cerebellum). The offspring systemic NOAEL is 40 ppm (3.4 mg/kg/day).

For the day 21 offspring, the only eff ect was a 21

- 22% decrease in plasma cholinesterase in high

- dose males and female. RBC and brain cholinesterase activity was not adversely affected in the offspring. In accordance with the current policy inhibition of plasma cholinesterase activity alone i n the absence of clinical signs is not considered to be appropriate for use as the

basis for a LOAEL. Therefore, t he offspring cholinesterase NOAEL is 200 ppm (16.4 mg/kg/day), the highest dose tested. An offspring cholinesterase NOAEL was not

established

.

This study is classified Acceptable/Non Guideline and may be used for regulatory purposes, however it does not satisfy the guideline requirement for a developmental neurotoxicity study in rats (OPPTS 870.6300, §83

- 6)

.

A. 3. 4.8 Metabolism

870.7485 Metabolism

– Rat

In a rat metabolism study (MRID 42034501)

, [1

- C 14 ] tribufos (97 % ai) was administered to male

and female Sprague

- Dawley rats (5 animals/sex/dose) by gavage as a single dose at levels of 5 or

100 mg/kg, or as a single dose at 5 mg/kg foll owing a 14

- day pretreatment with unlabeled

tribufos at 5 mg/kg/day. Five additional rats/sex received a single 5 mg/kg int ravenous dose

.

Fifty

- five to 80% of tribufos was absorbed, with >90% excreted in 72 hours. The majority of the

radioactive dose was excreted in urine (55

- 80%), followed by feces (15

- 42%), with <1% in the expired air. Females showed more radioactive compound in the urine than males for each dose regimen. There was no significant tissue residue . Only two excreted compounds could be

id entified, unmetabolized tribufos in the feces and butyl

- gamma

- glutamylcysteinylglycine disulfide in the urine. The data indicate complete metabolic breakdown of absorbed tribufos with

extensive breakdown of the S

- butyl moieties, probably to two carbon frac tions. No evidence of sulfonation or glucuronidation was observed following treatment with sulfatase or b

glucuronidase.

This metabolism study in the rat is classified A cceptable /Guideline and satisfies the guideline requirement for a metabolism study [OPPTS 870.7485 ( § 85

- 1) ] in the rat.

A. 3. 4.9 ImmunotoxicityTribufos Human Health Risk Assessment DP No. D357537

Page 76 of 84 870.7800 Immunotoxicity

In an immunotoxicity study (MRID #48709907 ), Tribufos was administered to female Han Wistar rats (10 /dose) in the diet at dose levels of 0, 5, 5 0, or 50 0 ppm (equivalent to 0, 0.43, 4.32, or 44.62 mg/kg/day, respectively) for 4 weeks. T he positive control group (8 females) was

administered 50 mg/kg bw/day of cyclophosphamide (CP) by intraperitoneal injection on study Day 27. During the study, clinical conditi on, bodyweight, food and water consumption, organ weight, brain cholinesterase, and macroscopic pathology were evaluated. Four days before

sacrifice , all animals in all grou ps received a single intravenous dose of sheep red blood cells 1.0 mL/animal (2 x 10 8 SRBC/mL) in 0.9% saline. At sacrifice (Day29) , selected organs were removed and weighed ( spleen , brain and thymus ). Brain and blood cholinesterase levels were

determined. The anti

- SRBC IgM concentration was measured with Plaque Forming Cell (PFC)

A ssay

.

There were no premature deaths and no treatment

- related clinical signs. At 500 ppm there was significant reduction in weight gain during the first 11 days of treatment and at week 4. Food and water consumption were reduced only in week one. Erythro cyte cholinesterase (ChE) activity was significantly reduced at 50 ppm and 500 ppm (66% and 91%, respectively) in week 4. At 500 ppm, brain ChE activity was reduced 78%. There was increased spleen weight at 500 ppm. Gross pathology showed splenomegaly onl y in one animal at 500 ppm; there were presence of pale contents in jejunum in the majority of animals at 500 ppm. The systemic LOAEL i s 50 ppm (equivalent to 4.32 mg/kg/day) based on significant decreased erythrocyte ChE activity . The systemic NOAEL is 5 ppm ( equivalent to 0.43 mg /kg/day )

.

There were no stati stically significant decreases in the numbers of PFC/10 6 viable cells or PFC/spleen in treated groups when compare d to the vehicle control group. High dose group receiving 500 ppm had significantl y increased cells/spleen. High inter

- individual variability was noted in all the treatment groups as well as in the control group. Evaluation of the individual animal data of this study did not show any trend or distribution that would demonstrate signifi cant suppression of anti

- SRBC antibody response. The p ositive control group had a statistically significant decrease in the PFC response

. This confirmed the ability of the test system to detect immuno

- suppressive effects and confirmed the validity of the s tudy design.

The Natural Killer (NK) cells activity was not evaluated in this study. The toxicology database for Tribufos technical does not reveal any evidence of immunotoxicity. The overall weight of evidence suggests that the chemical does not directly target the immune system. Under HED guidance a NK cell activity assay is not required at this time. The NOAEL for immunotoxicity

i s 500 ppm (equivalent to 44.62 mg/kg/day) , the highest dose tested

. The immunotoxicity

LOAEL i s not established.

This immunotoxicity s tudy is classified acceptable /guideline and satisfies the guideline requirement for an immunotox icity study (OPPTS 870.7800) in rats

.

A. 3. 4.10 Special StudiesTribufos Human Health Risk Assessment DP No. D357537

Page 77 of 84 Comparative Cholinesterase Studies

Thi s series of non

- guideline c holinesterase (ChE) inhibition studies (MRIDs 48 7099 01

- 48 709906 ) was undertaken to evaluate differences between PND

- 11 through PND

- 21 pups of both sexes and young adult female rats, and between dams and their fetuses, with regard to erythrocyte (RBC) or brain ChE following a dministration of tribu fos . All studies used Crl: CD Sprague Dawley rats and the same batch of tribufos (Batch No. NX81AX0015 ; 98.2% a.i. )

delivered in corn oil . All doses were administered by oral gavage, and the a ctual doses were quantified by gas chromat ography using electron capture detection. In addition to assessing ChE

inhibition , most of the studies also included determinations of clinical signs, mortality, body weight, food consumption, and necropsy with macroscopic examinations.

Acute and Repeat Dose

– Range Finding Studies in PND 11 Rat Pups

In pilot dose range

- finding studies (MRID 48709901 ) neonate 11 day old rat pups were administered acute (0, 20, 40 or 50 mg/kg/day) or repeated (11 consecutive doses of 0, 5, 10, 15, or 20 mg/kg/day) gavag e doses of Tribufos

. The small number of animals per dose precludes making firm conclusions. However some reported effects included decreased b od y weights and weight gains, appearance of being underactive, cold to touch, increased body tone, unsteady gait, body tremor and irregu lar breathing or vocalization. The signs were reported in both acute and repeat dosing with only flat posture and splaye d hindlimbs in the repeat dosing phase.

RBC and brain (but not plasma) ChE assessments were made at 24 and 48 hours following a single dose of Tribufos

. RBC ChE was inhibited in males and females at 20 and 50 mg/kg/day

after 24 and 48 hours. Brain ChE w as inhibited at 20 mg/kg/day in females but not in males after

24 hours. B rain ChE was inhibited in both males and females at 40 and 50 mg/kg/day after 48

hours. In the repeated dose study, assessments for RBC and brain ChE were made about 6 hours

after t he last dose. RBC and brain ChE were significantly inhibited in both males and females starting at 5 mg/kg/day.

Overall conclusion: This study supports a maximum tolerated dose of 50 mg/kg tribufos for a single oral dose and 15 mg/kg/day for repeat do sing in an 11 day multiple dosing study for 11

d ay old Sprague

- Dawley rat pups.

This study is classified as acceptable/non

- guideline and provides information for doses for definitive acute and repeat dosing comparative cholinesterase studies

. It is not ed that assessment for plasma ChE inhibition was not included.

Acute Oral Dosing in Young Rats to Determine Time to Peak Effect

In an acute study (MRID 48709902 ) to determine the time to peak effect on clinical signs and cholinesterase (ChE) inhibition , groups of 15/sex neonate rats (Sprague

- Dawley, aged 11 days)

, were given a single oral dose of 50 mg/kg bw Tribufos. Three pups/sex were scheduled for observation and sacrificed at 2, 4, 6, 8, 24 and 48 hours post dosing. ChE was determined in red blood cells (RBC) and brains

.Tribufos Human Health Risk Assessment DP No. D357537

Page 78 of 84 Two pups appearing in poor condition were sacrificed. Clinical signs reported (i.e underactivity, reduced body temperature) do not provide data that can delineate the time to peak of effect. Body weights trended t o be reduced at 24 and 48 hours in both sexes. Brain weight and gross pathology were not affected.

RBC ChE was inhibited in both sexes at all times. Brain ChE was inhibited in both sexes after 4

hours and maximum inhibition was noted in males at 6 hou rs and in females at both 6 and 8 hours postdosing. Brain ChE inhibition declined to about 50% by 48 hours but RBC ChE

remained at about 88%.

Conclusion: Based on observed RBC and brain ChE inhibition, 6

- 8 hours is a reasonable

selection for the time to peak effect for the definitive study.

This study is classified as acceptable/non

- guideline and provides information on the time

course for ChE inhibition.

Acute Oral Dosing to Young Adult Rats to Determine Time to Peak Effect

In an acute oral toxicity study (MRID 48709903 ) , groups of 24 young adult female (aged 48

- 5 2 days) SD rats were given a single oral dose of Tribufos at doses of 0 or 80 mg/kg b w to determine the time to a peak effect for systemic toxicity and cholinesterase inhibition . The study was conducted in 2 phases (Phase I , 24 hrs. used 15 animals/group) (Phase II , 48 hrs. used 9 animals/group). Clinical observations were made prior to sacrifice and at 2, 4, 6, 8, and 24 hours (3 per time) post dosing in Phase I, and at 24, 36, and 48 hours post dosing in Phase II. ChE was determined in red blood cells (RBCs) and brains.

There were no outstanding clinical signs observed, or effects on mortality, body weight and gain, brain weight, or gross pathology. RBCs ChE was significantly reduced by 90% in Phase I at 24

hours and in Phase II by 83% and 89% between 24 and 36 hours respectively. Brain ChE

inhibition in Phase I was 15% at 24 hours, while in Phase II inhibition was 1 5% at 24 hours and

20% at 36 hours.

Conclusion: The brain an d RBC inhibition time to peak effect appears to be between 24 and 36

hours.

This study is classified as acceptable/non

- guideline as a pilot study. The study is considered to

have provided information on time frames in which to search for the values of t he time to peak

effect for RBC and brain ChE inhibition.

Acute Oral Dosing Adult and Pup Comparative ChE

In a comparative cholinesterase inhibition study (MRID 4870990 4 ) , groups of 8 young adult SD rats( female only, aged 48

- 52 days) and pups (11 days old male and female) were administered

Tribufos at doses of 0, 2, 10 or 5 0 (pups only) or 80 (adults only) mg/kg bw

. Clinical

observati ons were made prior to dosing and at 3 and 6 hours and 24 (adults only). RBC andTribufos Human Health Risk Assessment DP No. D357537

Page 79 of 84 brain cholinesterase (ChE), brain weights and gross pathology were determined at times of sacrifice or at 6 hours for the pups and 24 hours for the adults.

Clinical signs were not noted in adult females. The 10 mg/kg neonates exhibited under activity at 6 hours post dos ing and at 50 mg/kg under activity and uncoordinated gait was noted as early as 3 hours and signs of being unsteady at 6 hours. There were no effects on mortality, brain weight, or gross pathology. Body weights of male and female pups were slightly lower (males 6.7 to 11% and females 7.3 to 8.9%) for all doses without a dose response and were not definitely related to treatment and will not be considered in the LOAEL statement. The LOAEL for systemic effects in pups is 10 mg/kg based on under activity. The NOAEL for pups is 2 mg/kg. The LOAEL for systemic effects in adults is not established since no effects were noted. The NOAEL is > 80 mg/kg.

RBC ChE was inhibited for the 2, 10 and 50 mg/kg female pup dose groups and for the 10 and 50 mg/kg male p up dose groups. In female adults there was inhibition at 24 hours in the 80 mg/kg dose group only. Brain ChE was inhibited in male and female pups at 10 and 50 mg/kg. Adult female brain ChE was inhibited only at 80 mg/kg. The LOAEL for ChE inhibition i n pups is < 2 mg/kg based on RBC inhibition in females. The NOAEL is not established. The LOAEL for adults for both RBC and brain ChE inhibition is 80 mg/kg. The NOAEL is 10 mg/kg.

Conclusion: Pups are more sensitive to the inhibitory effects of tri bufos than adults.

This comparative cholinesterase study is classified as acceptable/non

- guideline and established that pups are more sensitive to both systemic and ChE inhibitory effects of tribufos.

Repeat Dose Comparative Sensitivity Study in Young Adult Female and 11 Day Old Neonatal CD Rats by Oral Gavage

In a repeat dose comparative sensitivity study (MRID 48709905) in SD rats , tribufos was administered via gavage at dose levels of 0 , 0.1, 1.0 or 5.0 mg/kg bw/day to 8 young adult

female s /group and 8 male and 8 female neonatal pups (11 days old) /group for 11 consecutive days . Clinical signs of toxicity, mortality, body weights, cholinesterase (ChE) measurements [erythrocyte (RBC) and brain determinations], brain weights and macroscopic pat hology data

were obtained.

In adults, s alivation was noted in 1 female in the 1.0 and one in the 5. 0 mg/kg/day 1

- 2 h after dosing. However, salivation was not reported as an adverse effect in the subchronic neurotoxicity study (MRID No.: 45369101) at diet ary doses up to 4.6 mg/kg/day. No other effects were noted

in the adult females. In pups , reactions were noted at 5 mg/kg/day 1

- 2 hours postdosing including f

our males and 5 females with unsteady muscle rea ction and one male and five females with prostra te posture as well as under activity. At t erminal sacrifice, pup male body weight was statistically signific antly decreased in the 5.0 mg/kg/day group. Female pup body weight at termination was statistically significantly (p<0.05) decre ased in the 0.1, 1 .0 and 5.0 mg/kg/day

grou p. There was, however, no dose response to allow for a firm conclusion that body weightTribufos Human Health Risk Assessment DP No. D357537

Page 80 of 84 was affected in female pups. The LOAEL for systemic toxicity in young adult females was not established. The NOAEL is > 5 mg/kg/day. The LOA EL for systemic toxicity in pups is 5 mg/kg/day based on unsteady muscle reaction, prostrate appearance and underactivity. The NOAEL is 1 mg/kg/day.

RBC ChE was statistically signifi cantly inhibited in adults in th e 1.0 and 5.0 mg/kg/day group. RBC ChE was statistically significantly inhib ited only i n the 5.0 mg/kg/day group in pup males and females. Thus, the adults are more sensitive than the pups to inhibition of RBC ChE by

tribufos. Brain Ch E was not inhibited in adults. Brain ChE was sta tistically sign ificantly

inhibited in the 5.0 mg/kg/ day group in pup males and females. Thus, the pups are more sensitive than the adults to the inhibition of brain ChE by tribufos. The ChE LO AEL is 1.0 mg/kg /day based on RBC ChE inhibition in young adu lt females. The NOAEL is 0.1 mg/kg/day. The ChE LOAEL for male and female pups is 5.0 m g/kg /day based on

inhibition of RBC and brain ChE. The NOAEL is 1.0 mg/kg /day.

This repeat dose comparative sensitivity study in young adult fe male and 11 day old ne onatal

rats is acceptable/non

- guideline

.

Table A.5. Comparison of Tribufos RBC ChE inhibition in rat female adults vs female pups following repeated

- dosing a Dose (mg/kg/day) % ChE inhibition

Female adults % ChE inhibition Female Pups (PND11)

0

- - 0.1 10 12

1.0 18 18

5.0 64 69

a MIRD 48709905

Gestational ChE Inhibition

- Rat

In a gestational cholinesterase (ChE) inhibition study (MRID 48709906 ), tribufos was administered to pregnant SD rats (10/dose) by gavage at targeted doses of 0 , 1 (nominal), 7, or 28 mg/kg /day on gestation days (GD) 6

- 19. Because of low test substance concentra tions measured in the 1 mg/kg /day formulations, the actual time

- weighted dose was between 0.3 and 0.8 mg/kg /day. On GD 20, blood samples were collected from the dams and from the umbilical cord of all litters for the analysis of erythrocyte ChE activity. At necropsy on GD 20, each dam’s

uterus was removed via cesarean section and its contents examined. Brains were removed from

the dams and fetuses for the determinati on of brain ChE activity. Fetuses were examined for

external malformations.

A statistically significant decrease in mean body weight gain (not adjusted for gravid uterine weight) was observed in the 28 mg/kg /day group over the treatment period. Mean food consumption in the 28 mg/kg /day group was significantly lower during GD 6

- 9 and GD 18

- 19. There were no treatment

- related effects on maternal mortality or clinical signs, or on reproductive parameters, includ ing mean number of corpora lutea, implantations, resorptions,

pre

- and post

- implantation losses, number of live young, and sex ratio

. The LOAEL forTribufos Human Health Risk Assessment DP No. D357537

Page 81 of 84 maternal toxicity in rats is 28 mg/kg /day based on decreased body weight gain and

decreased food consu mpt ion. The NOAEL is 7 mg/kg /day

.

There were no observed fetal external abnormalities. Mean male, female, and overall fetal weights, and mean l itter weights in the 28 mg/kg /day group were slightly decreased (approx. 6% each), reaching statistical significance for male fetal weight only. The LOAEL for offspring toxicity in rats is 28 mg/kg /day based on decreased body weight in males. The NOAEL is 7 mg/kg /day.

Maternal eryt hrocyte and brain ChE were significantl y decreased at 7 and 28 mg/kg/day. At 28 mg/kg /day, fetal erythrocyte (from the umbilical cord) and brain ChE activities were lower than

the control but did not reach statistical significance. The LOAEL for maternal brain and erythroc yte ChE inhibition is 7 mg/kg /day. The NOAEL is between 0.3 and 0.8 mg/kg /day. The LOAEL for offspring erythrocyte and brain ChE inhibition is not determined. The NOAEL is 28 mg/kg/day, the highest dose tested.

This gestational expos ure cholinesterase inhibition study is classified acceptable/non

- guideline.Tribufos Human Health Risk Assessment DP No. D357537

Page 82 of 84 Appendix 3. IRL Sheet

International Residue Limits

Summary of US and International Tolerances and Maximum Residue Limits

US Canada Mexico 1 Codex

Residue Definition:

40CFR 180.272 Tribuphos (S,S,S

tributylphosphorotrithioate) None None

Commodity Tolerance (ppm) /Maximum Residue Limit (mg/kg) Commodity US Canada Mexico 1 Codex
Cattle, fat 0.15 None None None
Cattle, meat 0.02 Cattle, meat byproducts 0.02 Cotton, gin byproducts 40
Cotton, undelinted seed 4.0
Goat, fat 0.15
Goat, meat 0.02 Goat, meat byproducts 0.02 Hog, fat 0.15 Hog, meat 0.02 Hog, meat byproducts 0.02
Horse, fat 0.15
Horse, meat 0.02
Horse, meat byproducts 0.02

Milk 0.01 Sheep, fat 0.15 Sheep, meat 0.02 Sheep, meat byproducts 0.02 Completed by: M. Negussie; 06/19/15

1 Mexico adopts US tolerances and/or Codex MRLs for its export purposes.Tribufos Human Health Risk Assessment DP No. D357537

Page 83 of 84 Appendix 5

. Summary of Steady State Non Cancer Occupational Handler Dermal Risk Estimates for Tribufos

Appendix 5 Table A. 5. 1.: Tribufos Steady State Non Cancer Occupational Handler Dermal Risk Estimates on Cotton.

Exposure Scenario Application

Rate 2 Area

Treated 3 Dermal Unit Exposures (ug/lb ai) 1 Dermal Dose (mg/kg/day) 4 Dermal MOE 5

Worker Activity Formulation Application Equipment Value Units Value Unit SL/No

G SL/G DL/G EC SL/No

G SL/G DL/G EC SL/No

G SL/G DL/G EC

M/L L/SC/EC Aerial 1.875

lb

ai/A 1200

A 220 37.6 29.1 8.6 7.170 1.230 0.949 0.281 0.022 0.13 0.17 0.57

1.125 220 37.6 29.1 8.6 4.300 0.736 0.570 0.168 0.037 0.22 0.28 0.95

Groundboom 1.875
200 220 37.6 29.1 8.6 1.200 0.204 0.158 0.047 0.13 0.78 1 3.4

1.125 220 37.6 29.1 8.6 0.717 0.123 0.095 0.028 0.22 1.3 1.7 5.7

Applicator Spray Aerial 1.875

lb

ai/A 1200

A No

Data No

Data No

Data 2.08 No

Data No

Data No

Data 0.068 No

Data No

Data No

Data 2.4

1.125 No

Data No

Data No

Data 2.08 No

Data No

Data No

Data 0.041 No

Data No

Data No

Data 3.9

Groundboom 1.875
200 78.6 16.1 12.6 5.1 0.428 0.088 0.069 0.028 0.37 1.8 2.3 5.8

1.125 78.6 16.1 12.6 5.1 0.257 0.053 0.041 0.017 0.62 3 3.9 9.6

Flagger Spray Aerial 1.875 lb

ai/A 350 A 11 12 10.6 No

Data 0.105 0.114 0.101 No

Data 1.5 1.4 1.6 No

Data

1.125 11 12 10.6 No

Data 0.063 0.069 0.060 No

Data 2.5 2.3 2.6 No

Data
1 PHED/ORETF (Eng. Controls [EC]).

2

. Based on registered label (e.g. Reg. No. 5481

- 504, 5481

- 903 or EPA Reg. No. 264

- 730, 34704

- 867, and 85678

- 15). 3. Exposure Science Advisory Council Policy #9.1. 4. Dermal Dose = Dermal Unit Exposure (ug/lb ai) x Conversion Factor (0.001 mg/ug) x Application Rate (lb ai/acre or gal) x Area Treated or Amount Handled (A) / BW (69 kg). 5. Dermal MOE = Dermal POD (0.16 mg/kg/day)/Dermal Dose (mg/kg/day). Bolded numbers are risk estimates of concern. (MOE<1,000) . Hig hlighted risk estimates are current label PPE mitigation.Tribufos Human Health Risk Assessment DP No. D357537

Page 84 of 84 Appendix 5: Table A. 5. 2: Tribufos Steady State Non Cancer Occupational Handler Inhalation Risk Estimates on Cotton.

Exposure Scenario Application

Rate 2 Amount

Handled /

Area

Treated 3 Inhalation Unit Exposures (ug/lb ai) 1 Inhalation Dose (mg/kg

- day) 4 Inhalation MOE 5

Worker Activity Formulation Application Equipment Value Units Value Unit No

- R PF5

R PF10

R EC No

- R PF5

R PF10

R EC No

R PF5

R PF10

R EC

M/L L/SC/EC Aerial 1.875

lb ai/A 1200

A 0.219 0.044 0.022 0.083 0.007 0.001 0.001 0.003 28 140 280 130

1.125 0.219 0.044 0.022 0.083 0.004 0.001 0.000 0.002 47 230 470 210

Groundboom 1.875

200 0.219 0.044 0.022 0.083 0.001 0.000 0.000 0.000 170 840 1700 750

1.125 0.219 0.044 0.022 0.083 0.001 0.000 0.000 0.000 280 1400 2800 1300

Applicator Spray Aerial 1.875

lb ai/A 1200

A No

Data No

Data No

Data 0.005 No

Data No

Data No

Data 0.000 No

Data No

Data No

Data 630

1.125 No

Data No

Data No

Data 0.005 No

Data No

Data No

Data 0.000 No

Data No

Data No

Data 1000

Groundboom 1.875

200 0.340 0.068 0.034 0.043 0.002 0.000 0.000 0.000 54 270 540 1500

1.125 0.340 0.068 0.034 0.043 0.001 0.000 0.000 0.000 90 450 900 2400

Flagger Spray Aerial 1.875

lb ai/A 350 A 0.350 0.070 0.035 No

Data 0.003 0.001 0.000 No

Data 100 510 1000 No

Data

1.125 0.350 0.070 0.035 No

Data 0.002 0.000 0.000 No

Data 170 850 1700 No

Data

1. PHED/ORETF (Eng. Controls [EC]).

2. Based on registered label (e.g. Reg. No. 5481

- 504, 5481

- 903 or EPA Reg. No. 264

- 730, 34704

- 867, and 85678

- 15). 3. Exposure Science Advisory Council Policy #9.1. 4. Inhalation Dose = Inhalation Unit Exposure (ug/lb ai) x Conversion Factor (0.001 mg/ug) x Application Rate (lb ai/acre or gal) x Area Treated or Amount Handled (A) / BW (69 kg). 5. Inhalation MOE = Inhalation HED (0.10 mg/kg/day)/ Inhalation Dose (mg/kg/day). Bolded numbers are risk estimates of concern (MOE<300). ). Highlighted risk estimates are current label PPE mitigation.