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Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
09 February 2021 to 15 March 2021
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2021
Report date:
2021

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2-phenoxyethyl isobutyrate
EC Number:
203-127-1
EC Name:
2-phenoxyethyl isobutyrate
Cas Number:
103-60-6
Molecular formula:
C12H16O3
IUPAC Name:
2-phenoxyethyl 2-methylpropanoate
Test material form:
liquid
Remarks:
Colourless, clear liquid
Details on test material:
- Storage Conditions: At room temperature.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Remarks:
Crl:CD(SD), SPF
Details on species / strain selection:
Sprague-Dawley rats are extensively used in toxicity studies and are selected because of the abundance of historical control data.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at study initiation: Males and females were 5 weeks old at receipt, 6 weeks old at start of administration.
- Weight at study initiation: Males were 114.4 – 137.0 g and females were 104.1 – 116.9 g at receipt. Males were 172.6 - 204.5 g and females were 142.5 - 168.6 g at the start of administration.
- Fasting period before study: No
- Housing: Animals were housed in stainless wire mesh cages, 260 W × 350 D × 210 H (mm). One animal was housed per cage during the quarantine acclimation period and the dosing period.
- Diet: Powder feed rodent chow was placed in feeders and provided ad libitum.
- Water: Public tap water was filtered and irradiated by a UV water steriliser and then provided ad libitum.
- Acclimation period: Six days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.4 – 25.1 °C.
- Humidity (%): 40.7 – 55.4 % (relative)
- Air changes (per hr): 10 – 15 clean, fresh, filtered air changes per hour.
- Photoperiod (hrs dark / hrs light): 12 hour light/dark cycle (7 AM to 7 PM via automated timer), 150 - 300 Lux.

IN-LIFE DATES:
From: 09 February 2021
To: 02 March 2021

Administration / exposure

Route of administration:
oral: feed
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The required amount of the test material was weighed and placed in a container. The required amount of vehicle was added and mixed using a vortex mixer until dissolved.
The required amount of powder feed except for the amount of the test material was weighed. The required amount of the test material formulation and a small amount of powder feed were mixed in a container. The mixture was then placed in a ball mill and residual powder feed was added and mixed using the ball mill for approximately 5 – 10 minutes to yield the desired concentration.
The required amounts of corn oil and powder feed were weighed on an electronic balance and mixed using the ball mill for approximately 5 – 10 minutes for the control group.

DIET PREPARATION
- Storage temperature of food: The dosing feed was stored under refrigeration conditions for use within 14 days and used at room temperature (1 – 30 °C) for 6 days.

VEHICLE
- Concentration in vehicle: 10 mL of vehicle for 1 kg of powder feed containing test substance
Analytical verification of doses or concentrations:
no
Remarks:
Analysis for concentrations of dosing formulations was not performed.
Details on analytical verification of doses or concentrations:
The homogeneity and stability at room temperature (1 – 30 °C) for 3 and 6 days and under refrigeration for 8 and 14 days of the dosing formulations comprising the dose levels of 0.04 and 1.5 % were confirmed.
Duration of treatment / exposure:
14 days
Frequency of treatment:
Mixed test substance formulation and powder feed were placed in feeders and provided ad libitum
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
0 %
Dose / conc.:
182.9 mg/kg bw/day (nominal)
Remarks:
0.17 % males
Dose / conc.:
543.7 mg/kg bw/day (nominal)
Remarks:
0.5 % males
Dose / conc.:
1 512.4 mg/kg bw/day (nominal)
Remarks:
1.5 % males
Dose / conc.:
191.7 mg/kg bw/day (nominal)
Remarks:
0.17 % femlaes
Dose / conc.:
531.3 mg/kg bw/day (nominal)
Remarks:
0.5 % females
Dose / conc.:
1 512.4 mg/kg bw/day (nominal)
Remarks:
1.5 % females
No. of animals per sex per dose:
5 animals per sex per dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Based on discussions with the Sponsor, 1.5 % was selected as the high dose of this study. The mid and low doses were selected to be 0.5 and 0.17 %, respectively, by applying a geometric ratio of about 3.
- Rationale for animal assignment: On the last day of the quarantine-acclimation period (day of group assignment), 20 males and 20 females with body weights close to mean body weight were selected and assigned to 4 groups (5 animals/sex/group). Selected animals were randomly assigned to achieve even distribution of mean body weight of each group.
- Fasting period before blood sampling for clinical biochemistry: Yes. All animals were fasted overnight for approximately 18 hours.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE AND CLINICAL OBSERVATIONS: Yes
- Time schedule: All animals were observed once daily for clinical signs and twice daily for mortality and moribundity during the observation period.

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded on the day of start of dosing (Day 1, prior to dosing), twice a week during the dosing period and on the day of necropsy. However, body weight data recorded on the day of necropsy were not included in the evaluation of body weights since these data are fasted body weights. Body weights of animals found dead or moribund were recorded before necropsy during the observation period.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: Food consumption was recorded once from the day of group assignment to the first day of dosing.
- Food consumption: During the dosing period, the mean daily food consumption was calculated using the total amount of food consumed for 2 days twice a week.
- Test material intake: The test material consumption was calculated as follows and designated by mg/kg/day:
Test substance intake (mg/kg/day) = [Food consumption (mg/day) × Concentration (%)] / Body weight (kg)

FOOD EFFICIENCY: No

WATER CONSUMPTION AND COMPOUND INTAKE: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood samples were collected from the abdominal aorta on Day 15 at necropsy. Blood samples (approximately 1 mL) for haematology were placed in a vacutainer containing EDTA.
- Anaesthetic used for blood collection: Yes. Prior to taking blood samples, the animals were anaesthetised with isoflurane.
- Animals fasted: Yes. All animals were fasted overnight for approximately 18 hours prior to necropsy.
- How many animals: All animals.
- Parameters examined: The following parameters were analysed using an autoanalyser: Total erythrocyte count (RBC), Haemoglobin concentration (HGB), Haematocrit (HCT), Mean corpuscular volume (MCV), Mean corpuscular haemoglobin (MCH), Mean corpuscular haemoglobin concentration (MCHC), Platelets (PLT) and Leukocyte count (WBC).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood samples were collected from the abdominal aorta on Day 15 at necropsy. The blood samples from the abdominal aorta (except for blood samples for haematology) were centrifuged at 3 000 rpm for 10 minutes to obtain serum.
- Animals fasted: Yes. All animals were fasted overnight for approximately 18 hours prior to necropsy.
- How many animals: All animals.
- Parameters examined: The following parameters were analysed using an autoanalyser: Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (ALP), Blood urea nitrogen (BUN), Creatinine (Crea), Total protein (TP), Albumin (Alb), Albumin/Globulin ratio (A/G), Total cholesterol (T-Chol), Triglycerides (TG) and Glucose (Glu).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals were sacrificed by exsanguination from the abdominal aorta under isoflurane anaesthesia on the day of necropsy. Complete gross post-mortem examinations were performed on all animals including the external surface and internal organs. All grossly visible abnormalities were recorded.
For all animals, the following organs of terminal sacrifice animals were weighed individually and organ to body weight ratios were calculated. Paired organs were weighed together: Brain, Heart, Liver, Spleen, Kidney.

HISTOPATHOLOGY: Yes
At necropsy, the following organs and tissues were harvested and preserved in 10 % neutral buffered formalin except for the testes which were fixed in Davidson fixative, and then preserved in 10 % neutral buffered formalin: Brain, Thymus, Thyroid gland, Lung with bronchus, Parathyroid gland, Liver, Heart, Kidney, Spleen, Stomach, Adrenal gland, Jejunum, Duodenum, Cecum, Ileum with Peyer’s patch, Rectum, Colon, Testis, Pancreas, Ovary, Epididymis, Organ/tissue with gross lesions, Uterus with cervix.
Statistics:
Statistical analysis was performed on the data of body weight, food consumption, haematology, clinical chemistry and organ weights using SAS program (version 9.3, SAS Institute Inc., U.S.A.).
The data was analysed utilising Bartlett’s test for homogeneity of variance (significance level: 0.05). One-way analysis of variance (ANOVA) was employed on homogeneous data (significance level: 0.05); then, if significant, Dunnett's test was applied for multiple comparisons (significance levels: 0.05 and 0.01, two-tailed). Kruskal-Wallis test was employed on heterogeneous data (significance level: 0.05).

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
During the observation period, there were no abnormalities or clinical signs in both sexes in the control group, 0.17, 0.5, and 1.5 % dosing groups.
Mortality:
no mortality observed
Description (incidence):
During the observation period, there were no deaths of animals in both sexes in the control group, 0.17, 0.5, and 1.5 % dosing groups.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
There were no statistically significant differences in body weights and body weight gains in both sexes in the 0.17, 0.5 and 1.5 % dosing groups when compared to the control groups.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There was no statistically significant difference in food consumption in both sexes in the 0.17, 0.5 and 1.5 % groups when compared to the control group.
The mean total test material intakes at 0.17, 0.5 and 1.5 % were 182.9, 543.7 and 1 687.1 mg/kg/day for males, and 191.7, 531.3 and 1 512.4 mg/kg/day for females, respectively.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
Test material-related changes were not observed in both sexes in the 0.17, 0.5 and 1.5 % dosing groups.
The other changes with statistically significant difference were considered of no toxicological significance since they were minor changes and not accompanied by other related changes.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
Test material-related changes were not observed in both sexes in the 0.17, 0.5 and 1.5 % dosing groups.
The other changes with statistically significant difference were considered to be of no toxicological significance since they were slight changes and not accompanied by other related changes and abnormal changes in macroscopic findings.

Endocrine findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no statistically significant differences in body weights and body weight gains in both sexes in the 0.17, 0.5 and 1.5 % dosing groups when compared to the control groups.
Gross pathological findings:
no effects observed
Description (incidence and severity):
They were no abnormalities in necropsy in the control group, 0.17, 0.5 and 1.5 % dosing groups.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified

Effect levels

Dose descriptor:
NOAEL
Remarks on result:
not measured/tested
Remarks:
Dose range-finding study not designed to determine NOAEL

Target system / organ toxicity

Critical effects observed:
no

Any other information on results incl. tables

Summary of Clinical Signs


























































































































































































Group



No. of Animals



Clinical Sign



Day



1



2



3



4



5



6



7



8



9



10



11



12



13



14



Males



G1



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G2



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G3



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G4



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



Females



G1



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G2



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G3



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



G4



5



NOA



5



5



5



5



5



5



5



5



5



5



5



5



5



5



NOA: No Observable Abnormality.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Body Weights (g)































































































































































































































Group



Day



1



4



8



11



14



Males



G1



Mean



187.6



213.6



245.2



268.6



288.9



S.D.



11.7



14.1



17.3



19.0



23.8



N



5



5



5



5



5



G2



Mean



189.2



215.4



249.7



272.2



293.0



S.D.



6.1



10.1



15.8



19.3



26.3



N



5



5



5



5



5



G3



Mean



188.4



213.5



246.9



269.7



293.4



S.D.



6.5



7.6



8.5



9.1



11.6



N



5



5



5



5



5



G4



Mean



189.0



214.4



245.7



269.1



282.8



S.D.



5.9



8.6



14.7



13.6



22.2



N



5



5



5



5



5



Females



G1



Mean



155.7



170.5



186.1



197..7



208.2



S.D.



10.7



10.8



9.7



10.6



10.2



N



5



5



5



5



5



G2



Mean



156.2



168.0



180.2



192.3



204.7



S.D.



6.5



4.6



4.5



3.0



4.8



N



5



5



5



5



5



G3



Mean



157.6



169.4



186.1



196.7



210.1



S.D.



4.3



4.7



5.1



10.2



11.5



N



5



5



5



5



5



G4



Mean



156.6



168.1



180.1



191.8



200.4



S.D.



9.6



14.2



17.6



22.3



26.4



N



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Body Weight Gains (g)






































































































































































































Group



Day



4



8



11



14



Males



G1



Mean



25.9



31.8



23.4



20.3



S.D.



3.2



5.0



2.6



6.4



N



5



5



5



5



G2



Mean



26.2



34.4



22.5



20.7



S.D.



5.0



7.5



4.6



8.8



N



5



5



5



5



G3



Mean



25.1



33.4



22.9



23.7



S.D.



1.6



5.6



5.0



6.2



N



5



5



5



5



G4



Mean



25.4



31.3



23.4



13.7



S.D.



3.9



7.0



2.4



13.2



N



5



5



5



5



Females



G1



Mean



14.8



15.6



11.5



10.5



S.D.



4.9



2.1



2.0



3.8



N



5



5



5



5



G2



Mean



11.7



12.3



12.1



12.4



S.D.



5.1



2.0



2.8



2.5



N



5



5



5



5



G3



Mean



11.8



16.7



10.6



13.4



S.D.



2.6



2.3



5.8



4.0



N



5



5



5



5



G4



Mean



11.5



12.0



11.7



8.6



S.D.



5.2



46



6.5



4.6



N



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Food Consumption (g/animal/day)































































































































































































































Group



Day



1



4



8



11



14



Males



G1



Mean



22.3



24.3



26.2



25.6



26.7



S.D.



2.3



2.0



3.2



1.6



2.5



N



5



5



5



5



5



G2



Mean



22.8



25.0



27.7



26.5



27.6



S.D.



2.0



2.3



5.2



2.6



3.9



N



5



5



5



5



5



G3



Mean



22.8



24.8



26.9



27.1



28.3



S.D.



2.4



2.1



3.5



0.9



2.3



N



5



5



5



5



5



G4



Mean



23.2



25.3



28.0



26.7



30.4



S.D.



1.0



1.7



2.0



1.1



7.5



N



5



5



5



5



5



Females



G1



Mean



17.3



20.2



22.6



21.6



22.2



S.D.



2.3



1.2



1.7



1.6



1.5



N



5



5



5



5



5



G2



Mean



18.4



19.4



21.7



20.4



21.5



S.D.



3.8



1.1



4.2



2.2



2.2



N



5



5



5



5



5



G3



Mean



18.6



18.8



18.5



19.6



21.7



S.D.



1.1



1.4



1.2



2.3



1.8



N



5



5



5



5



5



G4



Mean



17.2



18.0



17.6



18.6*



8.9



S.D.



2.5



2.4



2.0



2.8



4.0



N



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean relative Food Consumption (g/kg bodyweight/day)































































































































































































































Group



Day



1



4



8



11



14



Males



G1



Mean



118.7



113.9



106.7



95.5



92.4



S.D.



4.8



3.0



5.8



2.8



2.8



N



5



5



5



5



5



G2



Mean



120.5



115.8



110.4



97.4



94.0



S.D.



8.2



6.7



15.5



2.9



6.6



N



5



5



5



5



5



G3



Mean



121.2



116.4



108.8



100.7



96.6



S.D.



12.8



9.4



12.8



3.2



7.7



N



5



5



5



5



5



G4



Mean



123.0



117.9



114.2



99.4



107.8



S.D.



3.8



3.6



8.6



2.3



26.9



N



5



5



5



5



5



Females



G1



Mean



111.1



118.6



121.8



109.4



106.7



S.D.



8.3



5.3



11.0



6.2



7.6



N



5



5



5



5



5



G2



Mean



117.3



115.5



120.4



105.9



104.8



S.D.



19.8



4.3



23.4



9.8



9.2



N



5



5



5



5



5



G3



Mean



118.1



111.2



99.6



99.2



103.2



S.D.



7.6



6.7



5.2



7.4



4.6



N



5



5



5



5



5



G4



Mean



109.3



106.6



97.9



97.0



93.3



S.D.



9.8



6.5



5.9



6.3



7.8



N



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Test Material Intake (mg/kg/day)













































































































































































Group



Day



Total Mean



4



8



11



14



Males



G2



Mean



196.8



187.6



165.5



159.9



182.9



S.D.



11.4



26.3



4.9



11.1



10.1



N



5



5



5



5



5



G3



Mean



581.9



544.1



503.3



483.2



543.7



S.D.



47.0



64.1



15.9



38.3



41.2



N



5



5



5



5



5



G4



Mean



1 768.8



1 716.5



1 491.6



1 616.9



1 687.1



S.D.



54.1



129.5



34.6



403.5



81.7



N



5



5



5



5



5



Females



G2



Mean



196.3



204.7



180.0



178.1



191.7



S.D.



7.2



39.8



16.7



15.7



18.6



N



5



5



5



5



5



G3



Mean



555.8



498.1



496.1



516.1



531.3



S.D.



33.6



25.9



37.0



22.9



27.7



N



5



5



5



5



5



G4



Mean



1 599.5



1 468.6



1 455.3



1 399.6



1 512.4



S.D.



97.7



88.7



94.2



117.3



70.3



N



5



5



5



5



5



G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Haematological Parameters







































































































































































































































































































Group



RBC


(x10^6/μL)



HGB


(g/dL)



HCT


(%)



MCV


(fL)



MCH


(pg)



MCHC


(g/dL)



PLT


(x10^3/μL)



WBC


(x10^3/μL)



Males



G1



Mean



7.22



15.1



42.9



59.5



21.0



35.3



1 249



11.5



S.D.



0.42



0.4



1.4



1.8



0.7



0.3



131



2.61



N



5



5



5



5



5



5



5



5



G2



Mean



6.99



14.7



41.7



59.7



21.0



35.2



1 014**



9.45



S.D.



0.15



0.3



0.5



1.2



0.3



0.4



60



2.19



N



5



5



5



5



5



5



5



5



G3



Mean



6.79



14.3*



10.9*



60.2



21.1



350



1 053*



10.44



S.D.



0.20



0.3



0.8



1.5



0.8



0.6



120



4.25



N



5



5



5



5



5



5



5



5



G4



Mean



7.26



15.4



43.6



60.1



21.2



35.2



1 079



9.06



S.D.



0.31



0.6



1.7



0.7



0.4



0.3



103



0.72



N



5



5



5



5



5



5



5



5



Females



G1



Mean



6.85



14.5



40.6



59.2



21.2



35.7



1 098



6.62



S.D.



0.45



0.4



1.3



2.0



0.7



0.1



71



1.80



N



5



5



5



5



5



5



5



5



G2



Mean



7.07



14.7



41.1



58.1



20.8



35.9



1 123



6.36



S.D.



0.20



0.4



1.3



1.2



0.4



0.2



123



2.06



N



5



5



5



5



5



5



5



5



G3



Mean



7.14



15.0



41.8



58.5



21.0



35.9



1 235



6.06



S.D.



0.35



0.6



1.4



1.4



0.5



0.4



141



1.09



N



5



5



5



5



5



5



5



5



G4



Mean



7.20



14.9



41.7



58.0



20.7



35.7



1 130



4.84



S.D.



0.34



0.4



1.0



1.8



0.5



0.3



45



1.19



N



5



5



5



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Clinical Chemistry


















































































































































































































































































































































































Group



ALT


(U/L)



AST


(U/L)



ALP


(U/L)



Glu


(mg/dL)



BUN


(mg/dL)



Crea


(mg/dL)



T-chol


(mg/dL)



TG


(mg/dL)



TP


(g/dL)



Alb


(g/dL)



A/G Ratio



Males



G1



Mean



23.8



84.7



739.5



133



12.2



0.39



65



44



5.0



2.3



0.86



S.D.



3.0



13.7



144.3



15



1.1



0.02



13



11



0.1



0.1



0.04



N



5



5



5



5



5



5



5



5



5



5



5



G2



Mean



22.7



82.5



764.1



126



12.0



0.37



56



43



5.0



2.3



0.83



S.D.



1.3



8.3



186.1



8



1.5



0.01



3



7



0.2



0.0



0.08



N



5



5



5



5



5



5



5



5



5



5



5



G3



Mean



24.1



92.8



831.5



115



11.2



0.38



49*



29



4.9



2.2



0.83



S.D.



7.5



13.9



218.4



15



2.3



0.05



8



13



0.1



0.1



0.10



N



5



5



5



5



5



5



5



5



5



5



5



G4



Mean



25.4



87.2



718.5



126



11.3



0.36



5.*



38



4.9



2.2



0.81



S.D.



3.6



8.1



143.3



8



2.3



0.03



8



16



0.2



0.1



0.06



N



5



5



5



5



5



5



5



5



5



5



5



Females



G1



Mean



19.2



82.9



423.9



120



12.3



0.39



60



13



4.9



2.4



0.92



S.D.



2.2



14.8



35.5



9



0.7



0.02



10



6



0.3



0.2



0.05



N



5



5



5



5



5



5



5



5



5



5



5



G2



Mean



18.0



85.6



412.8



120



11.2



0.39



46



14



5.3



2.5*



0.88



S.D.



3.6



17.4



111.7



18



1.5



0.02



11



4



0.2



0.2



0.09



N



5



5



5



5



5



5



5



5



5



5



5



G3



Mean



20.0



91.4



454.4



125



12.0



0.39



54



18



5.1



2.4



0.87



S.D.



2.6



14.3



47.1



11



0.7



0.03



14



8



0.1



0.1



0.03



N



5



5



5



5



5



5



5



5



5



5



5



G4



Mean



20.7



86.2



409.4



112



10.8



0.39



69



23



5.4



2.5**



0.86



S.D.



3.7



8.8



44.7



6



1.4



0.03



16



13



0.2



0.1



0.00



N



5



5



5



5



5



5



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 


Mean Organ Weights


























































































































































































































































































































































































Group



B.W.


(g)



Brain



Heart



Kidney



Liver



Spleen



 



Absolute


(g)



Relative


(%)



Absolute


(g)



Relative


(%)



Absolute


(g)



Relative


(%)



Absolute


(g)



Relative


(%)



Absolute


(g)



Relative


(%)



Males



G1



Mean



257.3



1.90



0.7403



1.04



0.4046



2.23



0.8682



8.26



3.2064



0.54



0.2070



S.D.



19.5



0.09



0.0564



0.10



0.0326



0.16



0.0376



0.91



0.1569



0.10



0.0257



N



5



5



5



5



5



5



5



5



5



5



5



G2



Mean



266.1



1.84



0.6939



1.10



0.4122



2.39



0.8956



8.48



3.1864



0.82



0.2958



S.D.



22.1



0.08



0.0429



0.15



0.0214



0.33



0.0535



0.82



0.1297



0.62



0.1915



N



5



5



5



5



5



5



5



5



5



5



5



G3



Mean



264.5



1.95



0.7385



1.08



0.4091



2.37



0.8984



8.22



3.1057



0.62



0.2359



S.D.



8.4



0.08



0.0381



0.11



0.0376



0.05



0.0403



0.63



0.2101



0.09



0.0346



N



5



5



5



5



5



5



5



5



5



5



5



G4



Mean



256.6



1.86



0.7268



1.08



0.4209



2.42



0.9424



8.98



3.4980*



0.57



0.2207



S.D.



16.0



0.05



0.0415



0.05



0.0196



0.20



0.0672



0.72



0.1400



0.08



0.0214



N



5



5



5



5



5



5



5



5



5



5



5



Females



G1



Mean



188.8



1.79



0.9467



0.79



0.4166



1.78



0.9446



6.00



3.1816



0.45



0.2387



S.D.



8.5



0.08



0.0403



0.03



0.0119



0.11



0.0167



0.25



0.0755



0.07



0.0290



N



5



5



5



5



5



5



5



5



5



5



5



G2



Mean



181.2



1.79



0.9853



0.80



0.04392



0.71



0.904



6.15



3.3938



0.44



0.2445



S.D.



3.9



0.10



0.0446



0.03



0.0148



0.14



0.0632



0.62



0.3275



0.09



0.0444



N



5



5



5



5



5



5



5



5



5



5



5



G3



Mean



186.0



1.8



0.9679



0.80



0.4294



1.72



0.9256



6.26



3.3686



0.43



0.2316



S.D.



9.0



0.05



0.0599



0.08



0.0423



0.13



0.0465



0.34



0.0985



0.07



0.0287



N



5



5



5



5



5



5



5



5



5



5



5



G4



Mean



183.8



1.82



0.9968



0.80



0.4373



1.70



0.9243



6.21



3.3708



0.41



0.2220



S.D.



20.2



0.06



0.1086



0.10



0.0205



0.19



0.0367



0.82



0.0833



0.08



0.0268



N



5



5



5



5



5



5



5



5



5



5



5



Significantly different from control by Dunnett's t-test: * p<0.05, ** p><0.01. Significantly different from control by <0.05, ** p<0.01.


Significantly different from control by Steel’s test: # p<0.05, ** p><0.01. Significantly different from control by <0.05, ## p<0.01.


G1: 0 %


G2: 0.17 %


G3: 0.5 %


G4: 1.5 %


 

Applicant's summary and conclusion

Conclusions:
Under the conditions of the study, the high dose level for the subsequent DRF prenatal development toxicity study should be selected at 1.5 %.
Executive summary:

This study was conducted according to OECD Test Guideline 407 and in compliance with GLP to evaluate the potential toxicity and to determine the dose levels for repeated dose study of the test material to Sprague-Dawley rats of both sexes for 2 weeks.


Test groups consisted of a dose group at a dose level of 0.17, 0.5 and 1.5 % groups and a control group (10 mL of Corn oil for 1 kg of powder feed containing test material) with 5 animals of each sex per group. All animals were dosed daily for 2 weeks by orally (dietary).


During the observation period, observation of clinical signs, measurement of body weights and food consumption were performed, and after the observation period, haematology and clinical chemistry, organ weight, gross post mortem examinations were performed.


No abnormal clinical signs or mortality were observed during the duration of the study.


There were no test material-related differences in body weight, food consumption, haematology, clinical chemistry, organ weights and necropsy in the animals of both sexes in the 0.17, 0.5 and 1.5 % dosing groups.


The mean total test material intakes at 0.17, 0.5 and 1.5% were 182.9, 543.7 and 1 687.1 mg/kg/day for males, and 191.7, 531.3 and 1 512.4 mg/kg/day for females, respectively.


Under the conditions of the study, the high dose level for the subsequent DRF prenatal development toxicity study should be selected at 1.5 %.