Registration Dossier

Diss Factsheets

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Experimental start date 11 January 2017 - Experimental completion date 16 March 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
the study plan required clinical observations and body weights to be recorded for the spare mated females to the same schedule as the study animals until at least Day 6 after mating. Therefore, clinical observations should have been recorded on Day 5 after mating. For eight of the ten spare mated females, clinical observations were recorded on Day 7 or Day 8 after mating rather than on Day 5 after mating in error; body weights were recorded to the correct schedule. Since none of the spare mated females were required for assignment to study, this deviation from study plan had no impact upon the scientific integrity of the study.
Deviations:
yes
Remarks:
This deviation from study plan had no impact upon the scientific integrity of the study.
Qualifier:
according to guideline
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Version / remarks:
The study plan required clinical observations and body weights to be recorded for the spare mated females to the same schedule as the study animals until at least Day 6 after mating. Therefore, clinical observations should have been recorded on Day 5 after mating. For eight of the ten spare mated females, clinical observations were recorded on Day 7 or Day 8 after mating rather than on Day 5 after mating in error; body weights were recorded to the correct schedule. Since none of the spare mated females were required for assignment to study, this deviation from study plan had no impact upon the scientific integrity of the study.
Deviations:
yes
Remarks:
This deviation from study plan had no impact upon the scientific integrity of the study.
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Version / remarks:
The study plan required clinical observations and body weights to be recorded for the spare mated females to the same schedule as the study animals until at least Day 6 after mating. Therefore, clinical observations should have been recorded on Day 5 after mating. For eight of the ten spare mated females, clinical observations were recorded on Day 7 or Day 8 after mating rather than on Day 5 after mating in error; body weights were recorded to the correct schedule. Since none of the spare mated females were required for assignment to study, this deviation from study plan had no impact upon the scientific integrity of the study.
Deviations:
yes
Remarks:
This deviation from study plan had no impact upon the scientific integrity of the study.
Qualifier:
according to guideline
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries, Test Data for Registration of Agricultural Chemicals, 12 Nohsan No. 8147, Agricultural Production Bureau, November 24, 2000.
Version / remarks:
The study plan required clinical observations and body weights to be recorded for the spare mated females to the same schedule as the study animals until at least Day 6 after mating. Therefore, clinical observations should have been recorded on Day 5 after mating. For eight of the ten spare mated females, clinical observations were recorded on Day 7 or Day 8 after mating rather than on Day 5 after mating in error; body weights were recorded to the correct schedule. Since none of the spare mated females were required for assignment to study, this deviation from study plan had no impact upon the scientific integrity of the study.
Deviations:
yes
Remarks:
This deviation from study plan had no impact upon the scientific integrity of the study.
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2-ethylanthraquinone
EC Number:
201-535-4
EC Name:
2-ethylanthraquinone
Cas Number:
84-51-5
Molecular formula:
C16H12O2
IUPAC Name:
2-ethyl-9,10-anthraquinone
Details on test material:
-Test item: 2-Ethylanthraquinone.
-CAS number: 84-51-5.
-Storage conditions: At ambient temperature, in the dark.
-Batch number: 151127.
-Expiry date: 31 December 2017.
-Purity: 98.73%.

Test animals

Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at study initiation: 77 to 83 days old.
- Weight at study initiation: 178 to 218 g.
- Housing: Cages comprised of a polycarbonate body with a stainless steel mesh lid; Solid (polycarbonate) bottom cages were used during the acclimatization and gestation periods; Grid bottomed cages were used during pairing. Cages were suspended above absorbent paper which was changed daily during pairing.
- Diet (e.g. ad libitum): Pelleted diet, non-restricted.
- Water (e.g. ad libitum): Potable water from the public supply, non-restricted.
- Acclimation period: Five days before commencement of pairing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24ºC
- Humidity (%): 40-70%
- Air changes (per hr): Filtered fresh air which was passed to atmosphere and not recirculated.
- Photoperiod (hrs dark / hrs light): Artificial lighting, 12 hours light : 12 hours dark.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Finely ground test item was mixed with the vehicle to form a paste which was diluted to final volume with vehicle and mixed using a high shear homogenizer. A series of suspensions at the required concentrations were prepared in this way by dilution of individual weighings of the test item. Formulations were stirred using a magnetic stirrer before and throughout the dosing procedure

DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly.
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food: Refrigerated (2-8ºC).

VEHICLE
- Concentration in vehicle: 1; 3; 10.
- Amount of vehicle (if gavage): Vehicle at the same volume dose as treated groups.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Homogeneity and stability of the test material in the vehicle demonstrated that the test item at a concentration range of 1 to 25 mg/mL was homogenous and stable in the vehicle for one day following ambient storage (+15 to +25°C) and for 15 days following refrigerated storage (+2 to +8°C). Samples of each formulation prepared for administration on Days 6 and 19 after mating were analyzed for achieved concentration of the test item.
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1 with identified stock males
- Length of cohabitation: one day
- Proof of pregnancy:Ejected copulation plugs in cage tray and vaginal smears were checked for the presence of sperm. Only females showing at least two copulation plugs were allocated. Referred to as day 0 of pregnancy
Duration of treatment / exposure:
Day 6 to Day 19 (inclusive) after mating
Frequency of treatment:
once daily at approximately the same time each day
Duration of test:
20 days
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Volume dose: 5 mL/kg body weight.
Dose / conc.:
5 mg/kg bw/day (nominal)
Remarks:
Volume dose: 5 mL/kg body weight.
Dose / conc.:
15 mg/kg bw/day (nominal)
Remarks:
Volume dose: 5 mL/kg body weight.
Dose / conc.:
50 other: mg/kg/day
Remarks:
Volume dose: 5 mL/kg body weight.
No. of animals per sex per dose:
20
Control animals:
yes, concurrent vehicle
Details on study design:
Mating: Day 0
Treatment: Day 6 - Day 19
Necropsy: Day 20

Treated at: Constant doses in mg/kg/day.
Volume dose: 5 mL/kg body weight.
Individual dose volume: Calculated from the most recently recorded scheduled body weight.
Control (Group 1): Vehicle at the same volume dose as treated groups.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily
- Cage side observations: Cages were inspected daily for evidence of animal ill-health amongst the occupant(s).

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: recorded daily

BODY WEIGHT: Yes
- Time schedule for examinations: Days 0, 5, 12, 18 and 20 after mating

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption : Days 0-2, 3-5, 6-9, 10-13, 14-17 and 18-19 after mating inclusive

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: a full macroscopic examination of the tissues was performed
Ovaries and uterine content:
Uterus: Gravid uterine weight (including cervix and ovaries).
For each ovary/uterine horn Number of: Corpora lutea.
Implantation sites.
Resorption sites (classified as early or late).
Fetuses (live and dead).

Fetal examinations:
- Examinations of all viable fetuses and placentae: all
- Examination of nominally 50% of fetuses in each litter
- Fetal Pathology Examination
Statistics:
Body weight, using absolute values and gains over appropriate study periods
Gravid uterine weight and adjusted body weight
Food consumption, over appropriate study periods
Litter size and survival indices
Fetal, placental and litter weight
The following comparisons were performed: Group 1 vs 2, 3 and 4
Historical control data:
Yes

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Description (incidence and severity):
There were no test item-related macroscopic abnormalities detected at scheduled termination.
Mortality:
no mortality observed
Description (incidence):
There were no premature deaths.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The mean body weight gain of females receiving 5 mg/kg/day was unaffected by treatment.
Between Day 6 and Day 7 of gestation: Dose-dependent mean body weight loss was recorded after the first dose administration for females receiving 15 or 50 mg/kg/day, compared with slight mean weight gain of Controls. (p<0.01)
Days 6-20 of gestation: mean body weight gain in the 15 mg/kg/day group was 101% of Control. (p<0.01)
Days 6-20 of gestation: mean body weight gain in the 50 mg/kg/day group was 88% of Control. (p<0.01)
Day 20 of gestation: Net mean body weight gain was 50% lower than Control at 50 mg/kg/day (p<0.01). At 5 or 15 mg/kg/day, net mean body weight gain was unaffected by 2-Ethylanthraquinone.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
At 5 mg/kg/day, mean food consumption was unaffected by 2-Ethylanthraquinone administration.
Days 6-9 of gestation: dose-dependent decrease in mean food consumption was apparent (12% and 31% lower than Control at 15 and 50 mg/kg/day, respectively)
to Day 13 of gestation: A gradual increase in food intake at 15 mg/kg/day
to Day 17 of gestation: A gradual increase in food intake at 50 mg/kg/day
From Day 10 of gestation in each of these groups: food intake returned to pre-treatment levels (Day 0-2 of gestation)
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Day 20 of gestation: mean gravid uterine weight of females in all treated groups was similar to Control.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no test item-related macroscopic abnormalities detected at scheduled termination
Details on results:
At dose levels up to and including 50 mg/kg/day the test item was generally well tolerated, with no premature deaths, no changes in clinical condition or signs related to treatment observed and no test item-related macroscopic findings.

At 50 mg/kg/day moderate maternal toxicity resulting in an overall 12% reduction in mean body weight gain during the dosing period (Day 6-20 of gestation), a 50% reduction in adjusted mean body weight change when the contribution of the gravid uterine weight was taken into consideration and a 31% reduction in mean food intake during Days 6-9 of gestation. Although food consumption remained lower than Control to Day 17 of gestation, levels of food intake returned to pre-treatment levels (Day 0-2 of gestation) from Day 10 of gestation.

At 15 mg/kg/day, maternal effects were limited to slightly low mean food intake during Days 6-13 of gestation (maximum effect 12% lower than Control, p<0.01). Since these maternal effects did not induce any changes in general clinical condition, gravid uterine weights or the ability of the females to maintain pregnancy they were deemed to be not adverse.

Maternal developmental toxicity

Number of abortions:
no effects observed
Description (incidence and severity):
All females were pregnant with live young at scheduled termination on Day 20 of gestation.
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
The mean numbers of implantations, pre- and post-implantation losses, was considered to be unaffected by maternal treatment at all dose levels investigated. Sex ratio was essentially similar in all groups.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
The mean numbers of resorptions was considered to be unaffected by maternal treatment at all dose levels investigated. Sex ratio was essentially similar in all groups.
Early or late resorptions:
no effects observed
Description (incidence and severity):
The mean numbers of resorptions was considered to be unaffected by maternal treatment at all dose levels investigated. Sex ratio was essentially similar in all groups.
Dead fetuses:
no effects observed
Description (incidence and severity):
All females were pregnant with live young at scheduled termination on Day 20 of gestation.
Changes in pregnancy duration:
not examined
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not specified
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
There was no effect of maternal treatment with 2-Ethylanthraquinone on mean litter at any dose level investigated.
Description (incidence and severity):
.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
15 other: mg/kg/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
There was no effect of maternal treatment with the test item on mean fetal weights at any dose level investigated.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): There was no effect of maternal treatment with the test item on mean fetal weights at any dose level investigated
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
There was no effect of maternal treatment with test item on mean litter or fetal weights at any dose level investigated.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The incidence of major and minor fetal abnormalities and skeletal variants showed no relationship to maternal treatment with test item:
- At 50 mg/kg/day there was an increased fetal and litter incidence of partially fused jugal to maxilla compared to concurrent control and exceeding the Historical Control Data (HCD) range (highest previous incidence in the Control population being 2 fetuses in 2 litters). This finding is considered to be representative of precocious ossification but seen in isolation is deemed to be not adverse.
- There were no treatment-related findings observed at 5 or 15 mg/kg/day

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
50 other: mg/kg/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
reduction in number of live offspring
changes in sex ratio
fetal/pup body weight changes
changes in litter size and weights
changes in postnatal survival
skeletal malformations
Remarks on result:
not determinable due to absence of adverse toxic effects

Fetal abnormalities

Key result
Abnormalities:
effects observed, non-treatment-related
Localisation:
skeletal: skull
Description (incidence and severity):
At 50 mg/kg/day there was an increased fetal and litter incidence of partially fused jugal to
maxilla compared to concurrent control and exceeding the Historical Control Data (HCD) range. This finding is considered to be representative of precocious ossification but seen in isolation is deemed to be not adverse.
There were no treatment-related findings observed at 5 or 15 mg/kg/day.
Embryo-fetal survival, growth and development were unaffected by maternal treatment with 2-Ethylanthraquinone.

Overall developmental toxicity

Key result
Developmental effects observed:
no
Treatment related:
no
Dose response relationship:
yes
Relevant for humans:
yes

Any other information on results incl. tables

Embryo-fetal survival, growth and development were unaffected by maternal treatment with the test item.

 

Applicant's summary and conclusion

Conclusions:
Based on the results obtained in this study of embryo-fetal development, it was concluded that the dose level of 15 mg/kg/day represented the No Observed Adverse Effect Level (NOAEL) for maternal toxicity and the dose level of 50 mg/kg/day represented the NOAEL for embryo-fetal survival, growth and development.
Executive summary:

The effect of the test item on embryo-fetal survival and development when administered during the organogenesis and fetal growth phase of pregnancy was investigated in the Han Wistar rat in accordance with OECD TG 414.

Three groups of 20 females were dosed at 5, 15 or 50 mg/kg/day by oral gavage administration at a volume dose of 5 mL/kg body weight from Day 6 to Day 19 after mating.

Treatment of pregnant female rats with the test item at dose levels up to and including 50 mg/kg/day was generally well tolerated and there were no unscheduled deaths or test item-related signs observed. There were no test item-related maternal macroscopic abnormalities detected at scheduled termination on Day 20 of gestation, and all females were pregnant with live young. There was no effect of maternal treatment with the test item on litter data, as assessed by the number of implantations, resorptions, live young, sex ratio and pre- and post-implantation losses.

Placental, litter and fetal weights were similar in all groups. The incidence of major and minor fetal abnormalities and skeletal variants showed no relationship to maternal treatment with the test item.

 The dose level of 15 mg/kg/day represented the No Observed Adverse Effect Level (NOAEL) for maternal toxicity and the dose level of 50 mg/kg/day represented the NOAEL for embryo-fetal survival, growth and development.

Categories Display