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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Remarks:
Experimental data from various test chemicals
Justification for type of information:
Data for the target chemical is summarized based on data from various test chemicals
Cross-referenceopen allclose all
Reason / purpose:
read-across source
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed journal
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
Test chemical was evaluated for developmental toxicity in presumed pregnant Sprague-Dawley rats.
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
other: Crl:CD® (SD) IGS BR VAF®/Plus
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Raleigh, NC
- Age at study initiation: 6-8 weeks old
- Weight at study initiation: Females: 213 to 241 g,
- Fasting period before study: No data available
- Housing: Animals were housed individually in cage.
- Diet (e.g. ad libitum): Certified Rodent DietR 5002 (PMI Nutrition International, St. Louis, MO), ad libitum
- Water (e.g. ad libitum): reverse osmosis deionized water (with chlorine added to the processed water as a bacteriostat), ad libitum
- Acclimation period: acclimatizated for short period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17.7 ± 26.11°C
- Humidity (%): 30% to 70%
- Air changes (per hr):at least 10 changes per hour of 100% fresh air passed through 99.97% HEPA filters.
- Photoperiod (hrs dark / hrs light): 12:12-h light-dark lighting cycle was used

IN-LIFE DATES: From: To: No data available
Route of administration:
oral: gavage
Type of inhalation exposure (if applicable):
not specified
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Dosing formulations were prepared daily from bulk materials. Samples from each concentration of the dosing suspensions (first and last days of treatment) were analyzed for the test chemical content.3 mg/ml conc.level was found to be stable for 10 days when stored at ambient conditions and protected from light.

DIET PREPARATION
- Rate of preparation of diet (frequency):Not applicable
- Mixing appropriate amounts with (Type of food): Not applicable
- Storage temperature of food: Not applicable

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil
- Concentration in vehicle: 0, 3, 10 and 30 mg/kg/day
- Amount of vehicle (if gavage): 10 ml/kg
- Lot/batch no. (if required): 103K0107 and 074K0025
- Purity: Not available
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
- M/F ratio per cage:1:1
- Length of cohabitation:5 days
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts:Not available
- Verification of same strain and source of both sexes:Not available
- Proof of pregnancy: The presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0).
- Any other deviations from standard protocol:Not available

Duration of treatment / exposure:
11 days (gestational days 7 to 17)
Frequency of treatment:
Daily
Duration of test:
gestational day 21
Remarks:
Doses / Concentrations:
0, 3, 10, or 30 mg/kg/day
Basis:
no data
No. of animals per sex per dose:
Total:100
Control(0)-25 female pregnant rats
3 mg/kg/day-25 female pregnant rats
10 mg/kg/day-25 female pregnant rats
30 mg/kg/day-25 female pregnant rats
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dosages were selected on the basis of range-finding study. dosages of 0, 1.25, 2.5, 5, or 10 mg/kg/day were administered daily to 8 rats/group on GDs 7 to 17. No adverse test chemical related clinical signs and food consumption observed. During the post dosage period, the body weight gains remained increased. All rats were pregnant and all caesarean sectioning and litter observations were comparable among the five groups. No fetal gross external alterations were observed. Based on these data, a dosage greater than 10 mg/kg/day AIM was recommended for the definitive developmental toxicity study in rats.
- Rationale for animal assignment (if not random): The healthy, mated female rats, weighing 213 to 241 g, were assigned to four dosage groups, 25 rats/group, using a computer-generated (weight-ordered) randomization procedure based on body weights recorded on initiation of study.
- Rationale for selecting satellite groups:No data avaiable
- Post-exposure recovery period in satellite groups: No data avaiable
- Section schedule rationale (if not random):No data avaiable
Maternal examinations:
CAGE SIDE OBSERVATIONS: yes, survival observed. Animals were observed twice daily for viability and examined for abnormal clinical signs
DETAILED CLINICAL OBSERVATIONS: Yes
BODY WEIGHT: Yes, Body weights were recorded prior to the start of the study and daily during the dosage and postdosage periods.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, Feed consumption was recorded on GDs 0, 7, 10, 12, 15, 18, and 21
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
POST-MORTEM EXAMINATIONS: Yes
POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice on gestation day # On gestational day 21.
- Organs examined: Yes, Organ weight and gross pathology were examined.
OTHER: abortions and premature deliveries-
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes
- Head examinations: No data
Statistics:
Continuous data were analyzed by using Bartlett’s test of homogeneity of variances and the analysis of variance. Dunnett’s test was used to identify statistical significance of individual groups. If the analysis of variance was not appropriate, the Kruskal-Wallis test or Dunn’s method of multiple comparisons was used to identify the statistical significance of the individual groups. If there were greater than 75% ties, Fisher’s exact test was used to analyze the data.
Indices:
Dead or resorbed conceptuses indice and live male fetuses indice were examined.
Historical control data:
No data avaialble
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.
Dermal irritation (if dermal study):
not specified
Mortality:
no mortality observed
Description (incidence):
No effect was observed on survival of treated female rats as compared to control.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group.
No effect was observed on food consumption of treated female rats as compared to control.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Description (incidence and severity):
No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Number of abortions:
not specified
Pre- and post-implantation loss:
not specified
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
No significant effect were observed litter sizes, live fetuses, resorptions
Early or late resorptions:
no effects observed
Description (incidence and severity):
No significant effect were observed
Dead fetuses:
no effects observed
Description (incidence and severity):
No significant effect were observed percentage of live male fetuses of treated female rats as compared to control.
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
not specified
Other effects:
not specified
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Mortality: No effect was observed on survival of treated female rats as compared to control.

Clinical sing: Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats.

None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.

Body weight and body weight gain: No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).

Food consumption: Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group.

No effect was observed on food consumption of treated female rats as compared to control.

Reproductive performence: No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses of treated female rats as compared to control.

Gross pathology: No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 30 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food efficiency
gross pathology
maternal abnormalities
Remarks on result:
other: No toxic effects were observed
Abnormalities:
not specified
Localisation:
not specified
Fetal body weight changes:
no effects observed
Description (incidence and severity):
No effect was observed on fetus weight in treated groups as compared to control.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
No effect was observed on number of live fetuses in treated groups as compared to control.
Changes in sex ratio:
not specified
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not specified
External malformations:
not specified
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.
Visceral malformations:
not specified
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Mortality: No effect was observed on number of live fetuses in treated groups as compared to control.

Fetus weight: No effect was observed on fetus weight in treated groups as compared to control.

Gross pathology: No gross external fetal alterations were observed in treated rats as compared to control.

Soft tissue malformations:
Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.

Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day

Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed.

All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material
because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility.
Dose descriptor:
NOAEL
Effect level:
>= 30 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
reduction in number of live offspring
fetal/pup body weight changes
changes in litter size and weights
skeletal malformations
Remarks on result:
other: All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material
Key result
Abnormalities:
no effects observed
Localisation:
other: not specified
Developmental effects observed:
not specified
Treatment related:
not specified
Conclusions:
The no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when Crl:CD® (SD) IGS BR VAF®/Plus female rats treated with test material.
Executive summary:

Developmental study was initiated to study the toxicity of test material . In this study, Crl:CD® (SD) IGS BR VAF®/Plus male and female rats were mated for 5 days and the presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0). The test material in a corn oil vehicle was administered orally via gavage to four groups of presumed pregnant rats on GDs 7 to 17 at dosages of 0, 3, 10, or 30 mg/kg/day. No effects were observed on survival, body weight and body weight gain and food consumption of treated female rats as compared to control. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups. Similarly, No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses and no alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control. In addition, no effects were observed on number of live fetuses, fetus weight and on gross external fetal alterations in treated rats as compared to control. Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7thcervical vertebra in four foetuses were observed in 30 mg/kg/day and in control. Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day and Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed. All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to treatment because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility. Therefore, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when Crl: CD® (SD) IGS BR VAF®/Plus female rats were treated with test material orally by gavage for 11 days.

Reason / purpose:
read-across source
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed publication
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
The reproductive and embryotoxic effect of test material was evaluated in pregnant Wistar rats.
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Center for Breeding of Laboratory Animals, Oswaldo Cruz Foundation
- Age at study initiation: No data available
- Weight at study initiation: approx. 220 g (200-250 g)
- Fasting period before study: No data available
- Housing: Animals were housed in standard plastic cages with stainless steel cover lids and wood shavings as bedding in a controlled environment.
- Diet (e.g. ad libitum): Nuvital diet, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimatization period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±1°C
- Humidity (%):70%
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12-hr dark/12-hr light

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test chemical was mixed with corn oil at dose level of 0, 500, 1000 and 1500 mg/kg body wt
DIET PREPARATION-Not available
VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil
- Concentration in vehicle: 0,250, 500, 750 or 1000 mg/kg/day
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
- M/F ratio per cage: 1:2
- Length of cohabitation: 2 h
- Proof of pregnancy: Confirmed by the presence of sperm in the vaginal smear.The day on which spermatozoa were found in the smear was designated as day 0 ofpregnancy.
- After … days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts: [no / yes (explain)]: Not available
- After successful mating each pregnant female was caged (how): singly
- Any other deviations from standard protocol: Not available
Duration of treatment / exposure:
Once (on Day 11 of pregnancy)
Frequency of treatment:
Once
Duration of test:
21 days
Remarks:
Doses / Concentrations:
0, 250, 500, 750 or 1000 mg/kg bw
Basis:
actual ingested
No. of animals per sex per dose:
Total: 65
0 mg/kg bw : 22 pregnant females
250 mg/kg/day: 12 pregnant females
500 mg/kg/day: 9 pregnant females
750 mg/kg/day: 12 pregnant females
1000 mg/kg/day: 10 pregnant females
Control animals:
yes, concurrent vehicle
Details on study design:
Mating performed (8:00-10:00 h) for 2h.The rats were given single dose of beta-Ionone only on day 11 of pregnancy and on day 21 of gestation dams were anesthesized and killed by CO2 inhalation.
Maternal examinations:
Body weight gain: Yes
The rats were weighed on days 0, 11 and 21 of pregnancy.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
-Fetal body weight: Yes
- External examinations (incl. anomalies): Yes
- Head examinations: Yes
The living fetuses were weighed, numbered with a marker pen, examined for externally visible anomalies under a stereomicroscope and fixed in a 5% formalin solution.
The externally visible skull defects were scored for severity.
Statistics:
The data were analysed by the one-way analysis of variance (ANOVA) followed by the Scheffe’s test for multiple comparisons or, alternatively, by the Kruskal-Wallis test followed by the Mann-Whitney U-test whenever the variable did not fit a normal distribution. Proportions were analyzed by the chi square -test or, alternatively, by the Fisher’s exact test.
Indices:
Not available
Historical control data:
Not available
Clinical signs:
no effects observed
Description (incidence and severity):
No overt signs of maternal toxicity were observed as compared to control.
Dermal irritation (if dermal study):
not specified
Mortality:
not specified
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No dose-related reduction of pregnancy weight gain were observed in treated rats.
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.
Gross pathological findings:
not specified
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Number of abortions:
not specified
Pre- and post-implantation loss:
not specified
Total litter losses by resorption:
not specified
Early or late resorptions:
effects observed, treatment-related
Description (incidence and severity):
When treated with 1000 mg/kg/day of test material the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased
Dead fetuses:
not specified
Changes in pregnancy duration:
not specified
Changes in number of pregnant:
not specified
Other effects:
not specified
Details on maternal toxic effects:
Maternal toxic effects:no effects. Remark: No dose related effects observed

Details on maternal toxic effects:
Clinical sign: No overt signs of maternal toxicity were observed as compared to control.

Body weight gain: No dose-related reduction of pregnancy weight gain were observed in treated rats.

Reproductive performance-When treated with 1000 mg/kg/day of BI the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased, while the ratio of live fetuses per implantations per litter was drastically decreased.

Organ weights-Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.
Dose descriptor:
NOAEL
Effect level:
750 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Remarks on result:
other: No toxic effects were observed
Abnormalities:
not specified
Localisation:
not specified
Fetal body weight changes:
not specified
Reduction in number of live offspring:
effects observed, treatment-related
Description (incidence and severity):
When treated with 1000 mg/kg/day of BI the ratio of live fetuses per implantations per litter was drastically decreased.
Changes in sex ratio:
not specified
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not specified
External malformations:
no effects observed
Description (incidence and severity):
No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control.

Skeletal malformations:
not specified
Visceral malformations:
not specified
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Gross pathology:
No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control.

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: embryotoxicity
Remarks on result:
other: No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control.
Abnormalities:
not specified
Developmental effects observed:
not specified
Treatment related:
not specified

Table 1:Maternal section data and caesarean section data of rats treated orally with test material only:

Sr.No.

Parameters

                                        Dose (mg/kg bw)

 

 

0

250

500

750

1000

1.

Pregnant females (N)

22   

12

9

12

10

2.

Resorption of the whole litter (N)

0

0

1

1

9

3.

Gravid uterus weight (g )

66.99+-10.8

83 89+-11.9

61.69+-30.2

70.29+-30.0

4.69+-12.5

4.

Maternal weight gain

(Days 0 -21(minus uterus wt)

45.99+-16.8

40.49+-14.4

64.99+-11.1

39.09+-19.2

49.09+-16.1

5.

Implantation sites

 

 

 

 

 

 

Total (N)

267

173

121

159

120

 

Per litter (mean+-SD)

12.19+-1.9

14.49+-1.4

13.49+-2.3

13.39+-3.6

12.09+-1.9

6.

Resorptions

 

 

 

 

 

 

Total (N)

35  

16

36

31

114

 

Per implantation sites (%)

13.1

9.2

29.8

19.5

95.0

 

Per implantation per litter (%)

9.2 

6.4

11.1

11.8

100.0

7.

Live fetuses Per implantation sites (%)

86.9

90.8

70.2

80.5

5.0

8.

Fetal body weight Per litter

4.889+-0.36

 

4.829+-0.23

4.889+-0.18

 

4.969+-0.27

-

 

Conclusions:
The no observed adverse effect level (NOAEL) was considered to be 750 mg/kg bw for P generation and 1000 mg/kg bw for F1 generation when Wistar female rats were treated with test material once orally by gavage in corn oil.
Executive summary:

In a teratogenicity study, Wistar female rats were treated with test material in the concentration of 250, 500, 750 and 1000 mg/kg body weight orally by gavage in corn oil at day 11 of pregnancy. No overt signs of maternal toxicity and effect on body weight gain were observed in treated rats as compared to control. Substantially increased in ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter, while the ratio of live fetuses per implantations per litter was drastically decreased at 1000 mg/kg bw in treated rats as compared to control. Similarly, Decreased in gravid uterus weight were observed in dams treated with 1000 mg/kg/day. In addition, no significant effect on externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 750 mg/kg bw for P generation and 1000 mg/kg bw for F1 generation when Wistar female rats were treated with test material once orally by gavage in corn oil at day 11 of pregnancy.

Data source

Reference
Reference Type:
review article or handbook
Title:
WoE of developmental toxicity study for CAS no 127-41-3
Author:
Sustainability Support Services (Europe) AB
Year:
2019
Bibliographic source:
WoE report, Sustainability Support Services (Europe) AB, 2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
WoE for the target CAS is summarized based on data from various test chemicals
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: Alpha-iso-methylionone
- Molecular formula: C14H22O
- Molecular weight: 206.328 g/mol
- Substance type: Organic
- Physical state: Liquid

Test animals

Species:
rat
Strain:
other: 2. Crl:CD® (SD) IGS BR VAF®/Plus; 3. Wistar
Details on test animals and environmental conditions:
2. TEST ANIMALS
- Source: Charles River Laboratories, Raleigh, NC
- Age at study initiation: 6-8 weeks old
- Weight at study initiation: Females: 213 to 241 g,
- Fasting period before study: No data available
- Housing: Animals were housed individually in cage.
- Diet (e.g. ad libitum): Certified Rodent DietR 5002 (PMI Nutrition International, St. Louis, MO), ad libitum
- Water (e.g. ad libitum): reverse osmosis deionized water (with chlorine added to the processed water as a bacteriostat), ad libitum
- Acclimation period: acclimatizated for short period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17.7 ± 26.11°C
- Humidity (%): 30% to 70%
- Air changes (per hr):at least 10 changes per hour of 100% fresh air passed through 99.97% HEPA filters.
- Photoperiod (hrs dark / hrs light): 12:12-h light-dark lighting cycle was used

IN-LIFE DATES: From: To: No data available

3. TEST ANIMALS
- Source: Center for Breeding of Laboratory Animals, Oswaldo Cruz Foundation
- Age at study initiation: No data available
- Weight at study initiation: approx. 220 g (200-250 g)
- Fasting period before study: No data available
- Housing: Animals were housed in standard plastic cages with stainless steel cover lids and wood shavings as bedding in a controlled environment.
- Diet (e.g. ad libitum): Nuvital diet, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimatization period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±1°C
- Humidity (%):70%
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12-hr dark/12-hr light

Administration / exposure

Route of administration:
oral: gavage
Type of inhalation exposure (if applicable):
not specified
Vehicle:
corn oil
Details on exposure:
2. PREPARATION OF DOSING SOLUTIONS:
Dosing formulations were prepared daily from bulk materials. Samples from each concentration of the dosing suspensions (first and last days of treatment) were analyzed for the test chemical content.3 mg/ml conc.level was found to be stable for 10 days when stored at ambient conditions and protected from light.

DIET PREPARATION
- Rate of preparation of diet (frequency):Not applicable
- Mixing appropriate amounts with (Type of food): Not applicable
- Storage temperature of food: Not applicable

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil
- Concentration in vehicle: 0, 3, 10 and 30 mg/kg/day
- Amount of vehicle (if gavage): 10 ml/kg
- Lot/batch no. (if required): 103K0107 and 074K0025
- Purity: Not available

3. PREPARATION OF DOSING SOLUTIONS: The test chemical was mixed with corn oil at dose level of 0, 500, 1000 and 1500 mg/kg body wt
DIET PREPARATION-Not available
VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil
- Concentration in vehicle: 0,250, 500, 750 or 1000 mg/kg/day
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
2. - M/F ratio per cage:1:1
- Length of cohabitation:5 days
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts:Not available
- Verification of same strain and source of both sexes:Not available
- Proof of pregnancy: The presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0).
- Any other deviations from standard protocol:Not available

3. - M/F ratio per cage: 1:2
- Length of cohabitation: 2 h
- Proof of pregnancy: Confirmed by the presence of sperm in the vaginal smear.The day on which spermatozoa were found in the smear was designated as day 0 of pregnancy.
- After … days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts: [no / yes (explain)]: Not available
- After successful mating each pregnant female was caged (how): singly
- Any other deviations from standard protocol: Not available
Duration of treatment / exposure:
2. 11 days (gestational days 7 to 17)
3. Once (on Day 11 of pregnancy)
Frequency of treatment:
2. Daily
3. Once
Duration of test:
2. gestational day 21
3. 21 days
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations: / 2
0, 3, 10, or 30 mg/kg/day
Basis:
no data
Remarks:
Doses / Concentrations: / 3
0, 250, 500, 750 or 1000 mg/kg bw
Basis:
actual ingested
No. of animals per sex per dose:
2. Total:100
Control(0)-25 female pregnant rats
3 mg/kg/day-25 female pregnant rats
10 mg/kg/day-25 female pregnant rats
30 mg/kg/day-25 female pregnant rats

3. Total: 65
0 mg/kg bw : 22 pregnant females
250 mg/kg/day: 12 pregnant females
500 mg/kg/day: 9 pregnant females
750 mg/kg/day: 12 pregnant females
1000 mg/kg/day: 10 pregnant females
Control animals:
yes, concurrent vehicle
Details on study design:
2. - Dose selection rationale: Dosages were selected on the basis of range-finding study. dosages of 0, 1.25, 2.5, 5, or 10 mg/kg/day were administered daily to 8 rats/group on GDs 7 to 17. No adverse test chemical related clinical signs and food consumption observed. During the post dosage period, the body weight gains remained increased. All rats were pregnant and all caesarean sectioning and litter observations were comparable among the five groups. No fetal gross external alterations were observed. Based on these data, a dosage greater than 10 mg/kg/day AIM was recommended for the definitive developmental toxicity study in rats.
- Rationale for animal assignment (if not random): The healthy, mated female rats, weighing 213 to 241 g, were assigned to four dosage groups, 25 rats/group, using a computer-generated (weight-ordered) randomization procedure based on body weights recorded on initiation of study.
- Rationale for selecting satellite groups:No data avaiable
- Post-exposure recovery period in satellite groups: No data avaiable
- Section schedule rationale (if not random):No data avaiable

3. Mating performed (8:00-10:00 h) for 2h.The rats were given single dose of beta-Ionone only on day 11 of pregnancy and on day 21 of gestation dams were anesthesized and killed by CO2 inhalation.

Examinations

Maternal examinations:
2. CAGE SIDE OBSERVATIONS: yes, survival observed. Animals were observed twice daily for viability and examined for abnormal clinical signs
DETAILED CLINICAL OBSERVATIONS: Yes
BODY WEIGHT: Yes, Body weights were recorded prior to the start of the study and daily during the dosage and postdosage periods.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, Feed consumption was recorded on GDs 0, 7, 10, 12, 15, 18, and 21
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
POST-MORTEM EXAMINATIONS: Yes
POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice on gestation day # On gestational day 21.
- Organs examined: Yes, Organ weight and gross pathology were examined.
OTHER: abortions and premature deliveries-

3. Body weight gain: Yes
The rats were weighed on days 0, 11 and 21 of pregnancy.
Ovaries and uterine content:
2. The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

3. The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:
2. - External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes
- Head examinations: No data

3. -Fetal body weight: Yes
- External examinations (incl. anomalies): Yes
- Head examinations: Yes
The living fetuses were weighed, numbered with a marker pen, examined for externally visible anomalies under a stereomicroscope and fixed in a 5% formalin solution.
The externally visible skull defects were scored for severity.
Statistics:
2. Continuous data were analyzed by using Bartlett’s test of homogeneity of variances and the analysis of variance. Dunnett’s test was used to identify statistical significance of individual groups. If the analysis of variance was not appropriate, the Kruskal-Wallis test or Dunn’s method of multiple comparisons was used to identify the statistical significance of the individual groups. If there were greater than 75% ties, Fisher’s exact test was used to analyze the data.

3. The data were analysed by the one-way analysis of variance (ANOVA) followed by the Scheffe’s test for multiple comparisons or, alternatively, by the Kruskal-Wallis test followed by the Mann-Whitney U-test whenever the variable did not fit a normal distribution. Proportions were analyzed by the chi square -test or, alternatively, by the Fisher’s exact test.
Indices:
2. Dead or resorbed conceptuses indice and live male fetuses indice were examined.
3. Not available
Historical control data:
No data avaialble

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
2. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.

3. No overt signs of maternal toxicity were observed as compared to control.
Dermal irritation (if dermal study):
not specified
Mortality:
no mortality observed
Description (incidence):
No effect was observed on survival of treated female rats as compared to control.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
2. No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).
3. No dose-related reduction of pregnancy weight gain were observed in treated rats.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
2. Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group. No effect was observed on food consumption of treated female rats as compared to control.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
2. No data
3. Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.
Gross pathological findings:
no effects observed
Description (incidence and severity):
2. No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified

Maternal developmental toxicity

Number of abortions:
not specified
Pre- and post-implantation loss:
not specified
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
No significant effect were observed litter sizes, live fetuses, resorptions
Early or late resorptions:
no effects observed
Description (incidence and severity):
2. No significant effect were observed
3. When treated with 1000 mg/kg/day of test material the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased
Dead fetuses:
no effects observed
Description (incidence and severity):
No significant effect were observed percentage of live male fetuses of treated female rats as compared to control.
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
not specified
Other effects:
not specified
Details on maternal toxic effects:
2. Maternal toxic effects:no effects

Details on maternal toxic effects:
Mortality: No effect was observed on survival of treated female rats as compared to control.

Clinical sing: Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats.

None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.

Body weight and body weight gain: No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).

Food consumption: Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group.

No effect was observed on food consumption of treated female rats as compared to control.

Reproductive performence: No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses of treated female rats as compared to control.

Gross pathology: No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.

3. Maternal toxic effects:no effects. Remark: No dose related effects observed

Details on maternal toxic effects:
Clinical sign: No overt signs of maternal toxicity were observed as compared to control.
Body weight gain: No dose-related reduction of pregnancy weight gain were observed in treated rats.
Reproductive performance-When treated with 1000 mg/kg/day of BI the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased, while the ratio of live fetuses per implantations per litter was drastically decreased.
Organ weights-Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Remarks:
2
Effect level:
>= 30 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food efficiency
gross pathology
maternal abnormalities
Remarks on result:
other: No toxic effects were observed
Dose descriptor:
NOAEL
Remarks:
3
Effect level:
750 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity: No effects on Maternal weight gain, reproductive performance and organ weight

Maternal abnormalities

Abnormalities:
not specified
Localisation:
not specified

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
2. No effect was observed on fetus weight in treated groups as compared to control.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
2. No effect was observed on number of live fetuses in treated groups as compared to control.
3. When treated with 1000 mg/kg/day of BI the ratio of live fetuses per implantations per litter was drastically decreased.
Changes in sex ratio:
not specified
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not specified
External malformations:
not specified
Description (incidence and severity):
2. No data
3. No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control.
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.
Visceral malformations:
not specified
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
2. Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Mortality: No effect was observed on number of live fetuses in treated groups as compared to control.

Fetus weight: No effect was observed on fetus weight in treated groups as compared to control.

Gross pathology: No gross external fetal alterations were observed in treated rats as compared to control.

Soft tissue malformations:
Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.

Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day

Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed.

All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material
because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility.

3. Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Gross pathology:
No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control

Effect levels (fetuses)

open allclose all
Dose descriptor:
NOAEL
Remarks:
2
Effect level:
>= 30 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
reduction in number of live offspring
fetal/pup body weight changes
changes in litter size and weights
skeletal malformations
Remarks on result:
other: All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material
Dose descriptor:
NOAEL
Remarks:
3
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: embryotoxicity: No effect on Fetal body weight, gross pathology and histopathology

Fetal abnormalities

Key result
Abnormalities:
no effects observed
Localisation:
other: not specified

Overall developmental toxicity

Developmental effects observed:
not specified
Treatment related:
not specified

Applicant's summary and conclusion

Conclusions:
The no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure.
Executive summary:

Data available for various test chemicals was reviewed to determine the developmental toxic nature of the test chemical. The studies are as mentioned below:

Developmental study was initiated to study the toxicity of test material . In this study, Crl:CD® (SD) IGS BR VAF®/Plus male and female rats were mated for 5 days and the presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0). The test material in a corn oil vehicle was administered orally via gavage to four groups of presumed pregnant rats on GDs 7 to 17 at dosages of 0, 3, 10, or 30 mg/kg/day. No effects were observed on survival, body weight and body weight gain and food consumption of treated female rats as compared to control. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups. Similarly, No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses and no alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control. In addition, no effects were observed on number of live fetuses, fetus weight and on gross external fetal alterations in treated rats as compared to control. Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7thcervical vertebra in four foetuses were observed in 30 mg/kg/day and in control. Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day and Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed. All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to treatment because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility. Therefore, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when Crl: CD® (SD) IGS BR VAF®/Plus female rats were treated with test material orally by gavage for 11 days.

In another teratogenicity study, Wistar female rats were treated with test material in the concentration of 250,500, 750 and 1000 mg/kg body weight orally by gavage in corn oil at day 11 of pregnancy.No overt signs of maternal toxicity and effect on body weight gain were observed in treated rats as compared to control. Substantially increased in ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter, while the ratio of live fetuses per implantations per litter was drastically decreased at 1000 mg/kg bw in treated rats as compared to control. Similarly, Decreased in gravid uterus weight were observed in dams treated with 1000 mg/kg/day. In addition, no significant effect on externally visible anomaly and skull defects were observed in fetuses of treated rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 750 mg/kg bw for P generation and 1000 mg/kg bw for F1 generation when Wistar female rats were treated with test material once orally by gavage in corn oil at day 11 of pregnancy.

Based on the details of the study, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure.