UNITED STATES ENVIRONMENTAL PROTECTION AGENCY 

WASHINGTON, D.C. 20460 

OFFICE OF CHEMICAL SAFETY 

AND POLLUTION PREVENTION 

MEMORANDUM 

DATE: December 6, 2013 

SUBJECT: Ethoprop: Review of 4-Week Inhalation Rat Study. 

DP Barcode: D401438 

Registration No.: NA 

PC Code: 041101 

Decision No.: 464348 

Petition No.: NA 

Risk Assessment Type: Single Chemical 

TXR No.: 0056347 

Regulatory Action: Registration Review 

CaseNo.: NA 

MRID No.: 48779601 

CAS No.: 13194-48-4 

40 CFR: NA 

FROM: 

THRU: 

TO: 

John Liccione, Ph.D.,_ Toxicologist 

Risk Assessment Branch V 

Health Effects Division (7509P) I 

,r/ 

(,}f--:, ... Q:~--··- / 

Michael Metzger, Branch Chief / } J j ~ }.;1"- """ 

Risk Assessment_ ~r~ch VI J/{ l j'V:iv f ./1 - (/ -j Health Effects DivisiOn (7509P) { · I 

Monica Wait, Chemical Risk Manager 

RMIB3 

Pesticide Reevaluation Division (7508P) 

I. CONCLUSIONS 

A 4-week inhalation rat study with ethoprop has been reviewed and is attached. The study will 

be included in the hazard characterization and hazard identification for risk assessment. 

II. ACTION REQUESTED 

Review the submitted studies for the requested registration action and risk assessment. 

III. MRID Summary Table 

MRID Comments 

48779601 NewDER 
DATA EVALUATION RECORD 

ETHOPROP 

(Ethoprophos) 

STUDY TYPE: 4-WEEK INHALATION TOXICITY- RAT 

(OCSPP 870.3465) 

Primary Reviewer: 

MRID 48779601 

Prepared for 

Health Effects Division 

Office of Pesticide Programs 

U.S. Environmental Protection Agency 

One Potomac Yard 

2777 South Crystal Drive 

Arlington, VA 22202 

Prepared by 

Summitec Corporation 

9724 Kingston Pike, Suite 602 

Knoxville, Tennessee 37922 

Task Order No. 6-24 

Thomas C. Marshall, Ph.D., DABT Signature: 

Date: 

Secondary Reviewers: 

H.T. Borges, Ph.D., MT(ASCP), DABT Signature: 

Date: 

Robert Ross, M.S., Program Manager Signature: 

Date: 

Quality Assurance: 

Jennifer Goldberg, B.S. Signature: 

Date: 

Disclaimer 

.1bKMU~c ~4~~ AE 

y.y:DE~ti~ t+E 

~~.~ DEC 0 4 2012 

This review may have been altered subsequent to the contractor's signatures above. 

Summitec Corp. for the U.S. Environmental Protection Agency under Contract No.EP-W-11-014 
ETHOPROP TECHNICAL/041101 

EPA Reviewer: Kit Farwell 

Risk Assessment Branch VII, Health Effects Division (7509P) 

EPA Secondary Reviewer: ---"-J-"'-'"-" hn """.=.c cc"""'""o.!..On.=.c e _ ______ _ 

Health Effects Division (7509P) 

TXR#: 005634 7 

DATA EVALUATION RECORD 

STUDY TYPE: 4-Week Inhalation Toxicity - Rat OCSPP 870.3465. 

PC CODE: 041101 DP BARCODE: D401438 

TEST MATERIAL (PURITY): Ethoprop Technical (96.6%) 

SYNONYMS: Ethoprophos Technical 

CITATION: Phillips, L. (2012). Ethoprop Technical: Toxicity study by inhalation 

administration to rats for 4 weeks. Huntingdon Life Sciences, Ltd., England. 

Project ID # BDG0059, March 22, 2012. MRID 48779601. Unpublished. 

SPONSOR: AMV AC Chemical Corporation. 

EXECUTIVE SUMMARY: 

In a nose-only inhalation exposure exposure toxicity study (MRID 48779601) Ethoprop 

Technical (Lot No. PEA4000032; 96.6% purity) aerosol was administered to male and female 

Sprague-Dawley rats (1 0/sex/concentration) for 6 hours/day, 5 days/week for 4 weeks at 

exposure concentrations of 0, 0.5, 2, or 6 !lg/L. Actual exposure concentrations as measured by 

HPLC were 0.498, 2.05, and 6.36 !lg/L. 

All animals survived until scheduled sacrifice. There were no treatment-related clinical findings. 

There were no treatment-related effects on body weight, body weight gain, or in any of the organ 

weight data in any exposure group. There were no treatment-related effects on hematology or 

clinical chemistry parameters. There were no test substance-related macroscopic changes 

observed in any tissues. Ethoprop exposure resulted in port-of-entry effects at all exposure levels 

in the larynx consisting of squamous cell metaplasia and epithelial hyperplasia. Among 

observations in the testes (seminiferous tubular atrophy, tubular degenerative germ 

cells/vacuolation and in the epididymides, degenerative spermatogenic cells in the duct at 2 and 

6 !lg/L), the tubular atrophy observed at 6 !lg/L could not be ruled out as a substance-related 

effect since its incidence was above the historical background range and similar effects at a 

lower incidence were observed at 2 !lg/L. Erythrocyte cholinesterase activity was decreased 

relative to controls at all exposure concentrations by 9% to 41% (statistically significant only in 

males at the highest concentration). Brain cholinesterase activity was statistically decreased by 

13% to 37% at the two highest exposure concentrations. Brain cholinesterase activity inhibition 

at 10% or above is generally considered toxicologically relavant. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 3 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

Based on the effects seen in this study, the NOAEL in male and female Sprague-Dawley 

rats was 0.5 μg/L. The LOAEL in male and female rats was 2 μg/L based on brain 

cholinesterase activity inhibition. 

This inhalation toxicity study is classified as Acceptable (Guideline) and satisfies the guideline 

requirement for 4-week inhalation toxicity study in rats (870.3465). 

COMPLIANCE: Signed and dated GLP, Quality Assurance, and Data Confidentiality 

statements were provided. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 4 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

I. MATERIALS AND METHODS:

A. MATERIALS:

1. Test material: Ethoprop Technical

 Description: Oily yellow liquid 

 Lot/batch #: PEA4000032 

 Purity: 96.6% a.i. 

 CAS # of TGAI: 13194-48-4 

Structure: 

2. Vehicle: Filtered air 

3. Test animals: 

Species: Rat 

Strain: Crl:CD(SD) 

Age/weight at study initiation: 47-53 days/ Males 230-298 g; Females 181-239 g 

Source: Charles River Ltd., UK 

Housing: 5 of one sex/polycarbonate cage with sterilized wood based bedding changed weekly 

Diet: Certified Rodent LabDiet® 5002, PMI Nutrition International, LLC, ad libitum except during 

exposure. 

Water: Tap water, ad libitum

Environmental conditions: Temperature: 

Humidity: 

Air changes: 

Photoperiod: 

19-23 0

C 

40-70% 

Not reported 

12 hours light/dark

Acclimation period: 11 days 

B. STUDY DESIGN:

1. In life dates: Start: October 31, 2011; End: November 30, 2011 

2. Animal assignment and treatment: Rats were assigned to the experimental groups 

presented in Table 1, but the method of assignment was not specified. The exposure 

concentrations were selected based on the results of a one-week pilot study in which 

erythrocyte and brain acetylcholinesterase activity inhibition (AChEI) were observed at 

exposure concentrations of 5.42 to 66.6 μg/L, but not at 2.17 μg/L. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 5 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

TABLE 1. Study Design 

Experimental parameter (test relative to dosing) Ethoprop Concentrations (μg/L)a 

0 0.5 2 6 

Number of Rats 

Total no. of animals assigned 20 (10/sex) 20 (10/sex) 20 (10/sex) 20 (10/sex) 

 Blood AChEI 10/sex 10/sex 10/sex 10/sex 

Brain AChEI 10/sex 10/sex 10/sex 10/sex 

Gross and microscopic pathology 10/sex 10/sex 10/sex 10/sex 

 

a

 Actual exposure levels were 0.498, 2.05, and 6.36 μg/L. 

Data obtained from pages 14-22 (MRID 48779601) 

3. Generation of the test atmosphere / chamber description: Chamber atmospheres were 

generated by stainless steel concentric jet atomizers coupled with a distribution system that 

delivered a controlled amount of vapor/aerosol to the inlet of each aluminum alloy nose-only 

exposure chamber. A syringe pump (Precidor) was used to deliver the test substance to the 

generator and combined with filtered compressed air, both of which were metered to deliver 

the appropriate atmosphere concentration to the chamber. Nose-only exposure chambers (one 

for each exposure level) were fitted with air flow meters, pressure gauges, and other devices 

to measure temperature, humidity, and test article concentration. 

4. Test atmosphere concentration: The exposure concentrations were measured three times 

per day by drawing a known volume of the test atmosphere at a rate of 2L/minute through a 

glass sintered impinger trap containing acetonitrile/water/antifoam (80/20/0.02 v/v/v). 

Samples were taken from the breathing zone of the animals. Analyses were conducted using 

HPLC with UV detection. Results are in Table 2 and show that the achieved levels were 100, 

103, and106% of target values. Time to equilibrium was not reported. 

Table 2. Mean Exposure Concentrations during the 4-Week Exposure Regimen. 

Data obtained from page 210 (MRID 48779601)

5. Particle size determination: Particle size for the highest exposure concentration was 

determined using a Marple 290 Series cascade impactor with stainless steel substrates. The 

mass median aerodynamic diameter and geometric standard deviation were determined 

(Table 3). Sampling intervals and the number of samples was not provided. No particle size 

samples were taken for the two lower concentrations because preliminary trials showed that 

the atmospheres were predominately vapor with a negligible proportion of droplets. These 

preliminary data were not provided. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 6 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

Table 3. Mean Particle Size during the 4-Week Exposure Regimen.

Data obtained from page 211 (MRID 48779601)

6. Statistics: All analyses were conducted comparing each test substance-exposed group to the 

control group by sex. The overall level of significance for intergroup differences was p≤0.05. 

Body weight, body weight change, organ weight, clinical pathology, erythrocyte AChEI, and 

brain AChEI activity were subjected to a series of tests to determine intergroup differences. 

Bartlett’s test for variance homogeneity, if not significant (p<0.01), was followed by the F1

approximate test. If not significant the William’s test for monotonic trend was applied. If the 

F1 was significant, then Dunnett's test was used to compare the test substance-exposed 

groups to the control group. If Bartlett’s test was significant, then the nonparametric H1 test 

was applied, followed by either Shirley’s test for monotonic trend or Steel’s test. Clinical 

pathology, erythrocyte AChEI, and brain AChEI activity were further analyzed by Fisher’s 

Exact test if 75% of the data had the same c value. Analysis of covariance (ANCOVA) was 

used to analyze the effect of body weight on organ weight. 

The Reviewer considers the statistical analyses used to be appropriate. 

C. METHODS / OBSERVATIONS:

1. Mortality and clinical observations: Animals were observed twice daily for clinical signs, 

abnormal behavior, morbidity, and mortality. Detailed clinical observations were recorded 

weekly. 

2. Body weight: Animals were weighed weekly. 

3. Food consumption: Food consumption was estimated for each animal by recording weekly 

food usage by cage (5 rats) less spillage then calculating mean daily diet consumption as g 

food/rat/week. 

4. Hematology and clinical chemistry: Blood was collected from all surviving animals at the 

end of the treatment period for hematology and clinical chemistry; the animals were fasted 

overnight prior to blood collection. Blood was collected from the sublingual vein while the 

rats were under light isoflurane anesthesia. The CHECKED (X) parameters were examined. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 7 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

a. Hematology:

X Hematocrit (HCT)* X Leukocyte differential count* 

X Hemoglobin (HGB)* X Mean corpuscular HGB (MCH)* 

X Leukocyte count (WBC)* X Mean corpusc. HGB conc.(MCHC) 

X Erythrocyte count (RBC)* X Mean corpusc. volume (MCV) 

X Platelet count* X Reticulocyte count 

 Blood clotting measurements* 

X (Thromboplastin time) 

 (Clotting time) 

X (Prothrombin time) 

* Recommended for 28-day oral rodent studies based on Guideline 870.3050

b. Clinical chemistry:

X ELECTROLYTES X OTHER 

X Calcium X Albumin* 

X Chloride X Creatinine* 

 Magnesium X Urea nitrogen* 

X Phosphorus X Total Cholesterol* 

X Potassium* Globulins 

X Sodium* X Glucose* 

ENZYMES (more than 2 hepatic enzymes eg., *) X Total bilirubin 

X Alkaline phosphatase (ALK)* X Total protein (TP)* 

 Cholinesterase (ChE) X Triglycerides 

 Creatine phosphokinase Serum protein electrophoresis 

 Lactic acid dehydrogenase (LDH) X Albumin/Globulin ratio 

X Alanine aminotransferase (ALT/also SGPT)* 

X Aspartate aminotransferase (AST/also SGOT)* 

 Sorbitol dehydrogenase* 

X Gamma glutamyl transferase (GGT)* 

 Glutamate dehydrogenase 

* Recommended for 28-day oral rodent studies based on Guideline 870.3050

c. Cholinesterase determination: Blood samples were centrifuged and the plasma frozen 

immediately. The brain was separated at the midline after weighing during necropsy and 

the left side snap frozen for AChE determination. Erythrocyte and brain cholinesterase 

activities were determined using a modified Ellman assay. 

6. Urinalysis: Urine was not examined. 

7. Ophthalmoscopic examination: Eyes were examined by an ophthalmoscope prior to the 

first day of exposure and during week 4. 

8. Sacrifice and pathology: Rats were killed by i.p. injection of sodium pentobarbital followed 

by exsanguination before necropsy and collection of tissues for microscopic examination. 

Any gross lesions were recorded. Tissues from animals in the control and high dose groups 

were processed for microscopic evaluation. The larynx, epididymides, and testes were 

examined in animals from all exposure groups. The CHECKED (X) parameters were 

collected and examined. The (XX) organs were weighed. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 8 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

X DIGESTIVE SYSTEM X CARDIOVASC./HEMAT. X NEUROLOGIC 

 Tongue Aorta XX Brain 

 Salivary glands XX Heart Peripheral nerve 

 Esophagus Bone marrow Spinal cord (3 levels) 

 Stomach Lymph nodes Pituitary 

 Duodenum XX Spleen Eyes (optic nerve ) 

 Jejunum XX Thymus X GLANDULAR

 Ileum XX Adrenal gland 

 Cecum X UROGENITAL Lacrimal gland 

 Colon XX Kidneys X Parathyroid 

 Rectum Urinary bladder X Thyroid 

XX Liver XX Testes X OTHER

 Gall bladder (not rat) X Epididymides Bone (sternum and/or femur) 

 Bile duct (rat) Prostate Skeletal muscle 

 Pancreas Seminal vesicles Skin 

X RESPIRATORY X Ovaries X All gross lesions and masses 

X Trachea X Uterus 

XX Lungs Mammary gland 

X Nasal cavity X Cervix 

 Pharynx 

X Larynx 

II. RESULTS:

A. OBSERVATIONS:

1. Clinical signs and Mortality: All animals survived until scheduled sacrifice. There were no 

treatment-related clinical findings. There were no treatment-related effects on body weight, 

body weight gain, or in any of the organ weight data in any exposure group. There were no 

treatment-related effects on hematology or clinical chemistry parameters. There were no test 

substance-related macroscopic changes observed in any tissues. Ethoprop exposure resulted 

in minimal to mild portal-of-entry effects in the larynx at all exposure levels consisting of 

squamous cell metaplasia and epithelial hyperplasia. These were not significant 

toxicologically. Among observations in the testes (seminiferous tubular atrophy, tubular 

degenerative germ cells/vacuolation and in the epididymides, degenerative spermatogenic 

cells in the duct at 2 and 6 μg/L), the tubular atrophy observed at 6 μg/L could not be ruled 

out as a substance-related effect. Its incidence was above the historical background range and 

similar effects at a lower incidence were observed at 2 μg/L. Erythrocyte cholinesterase 

activity was decreased relative to controls at all exposure concentrations by 9% to 41% 

(statistically significant only in males at the highest concentration). Brain cholinesterase 

activity was statistically decreased by 13% to 37% at the two highest exposure 

concentrations. Brain cholinesterase activity inhibition at 10% or above is generally 

considered toxicologically relevant. 

B. BODY WEIGHT AND BODY WEIGHT GAIN: There were no treatment-related effects 

on body weight or body weight gain in any exposure group (Table 4). 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 9 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

Data obtained from pages 37-38 (MRID 48779601) 

C. FOOD CONSUMPTION: There were no treatment-related effects on food consumption. 

D. OPHTHALMOSCOPIC RESULTS: There were no treatment-related findings during the 

ophthalmoscopic examinations. 

E. BLOOD ANALYSES: 

1. Hematology: There were no treatment-related effects on hematology parameters.

2. Clinical chemistry: There were no treatment-related effects on clinical chemistry 

parameters. 

TABLE 4. 
4-Week Inhalation Toxicity Study (rat) (2012) / Page 10 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

3. Cholinesterase activity: Erythrocyte cholinesterase activity was decreased relative to 

controls at all exposure concentrations by 9% to 41% (statistically significant only in males 

at the highest concentration). Brain cholinesterase activity was statistically decreased by 13% 

to 37% at the two highest exposure concentrations (Table 5). 

Data obtained from page 30 (MRID 48779601) 

** p≤0.01

E. SACRIFICE AND PATHOLOGY: 

1. Gross pathology: There were no test substance-related macroscopic changes observed in 

any tissues evaluated on the day of scheduled sacrifice. 

2. Organ weight: There were no test substance-related effects observed in any of the organ 

weight data. 

3. Microscopic pathology: Microscopic alterations associated with 4-weeks of exposure 

included laryngeal ventrolateral squamous metaplasia in both males and females at all three 

ethoprop exposure concentrations (Table 6). Laryngeal arytenoid epithelial hyperplasia was 

also observed, but to a lesser extent than the squamous metaplasia. The squamous metaplasia 

and epithelial hyperplasia in the larynx were considered adaptive responses and not 

toxicologically significant. In the testes, seminiferous tubular atrophy and tubular 

degenerative germ cells/vacuolation and in the epididymides, degenerative spermatogenic 

cells in the duct were seen in males exposed to 2.05 and 6.36 μg/L at a higher incidence than 

controls. The findings were within historical background ranges except the tubular atrophy 

observed at 6.36 μg/L.

TABLE 5.
4-Week Inhalation Toxicity Study (rat) (2012) / Page 11 of 11 

ETHOPROP TECHNICAL/041101 OCSPP 870.3465 

III. DISCUSSION AND CONCLUSIONS: 

A. INVESTIGATORS’ CONCLUSIONS: The nose-only inhalation exposure of rats to 

ethoprop for 4 weeks caused reductions in erythrocyte and brain cholinesterase activities in 

both sexes at the two highest exposure levels (2.05 and 6.36 μg/L). There was a local irritant 

response of the larynx at all exposure levels. The no-observed-adverse-effect level (NOAEL) 

was considered to be 0.498 μg/L. 

B. REVIEWER COMMENTS: 

The reviewer generally agrees with the investigators’ conclusions. Inhalation exposure of rats 

to ethoprop at concentrations of 0, 0.5, 2, or 6 μg/L for 6 hours/day, 5 days/week for 4 weeks 

resulted in port-of-entry effects in the larynx consisting of squamous cell metaplasia and 

epithelial hyperplasia in the mid and high exposure groups. Among observations in the testes, 

the tubular atrophy observed at 6 μg/L could not be ruled out as a substance-related effect, 

since its incidence was above the historical background range and similar effects at a lower 

incidence were observed at 2 μg/L. Erythrocyte cholinesterase activity was decreased relative 

to controls at all exposure concentrations by 9% to 41% (statistically significant only in 

males at the highest concentration). Brain cholinesterase activity was statistically decreased 

by 13% to 37% at the two highest exposure concentrations. Brain cholinesterase activity 

inhibition at 10% or above is considered toxicologically significant. 

Based on the effects seen in this study, the NOAEL in male and female Sprague-Dawley 

rats was 0.5 μg/L. The LOAEL in male and female rats was 2 μg/L based on brain 

cholinesterase activity inhibition. 

This inhalation toxicity study is classified as Acceptable (Guideline) and satisfies the 

guideline requirement for 4-week inhalation toxicity study in rats (870.3465). 

C. STUDY DEFICIENCIES: A minor deficiency was the time to ethoprop equilibrium in the 

exposure chamber was not reported. 

TABLE 6.