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Developmental toxicity / teratogenicity

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Endpoint:
developmental toxicity
Type of information:
experimental study
Remarks:
Experimental study based on read across substances
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
[Please provide information for all of the points below. Indicate if further information is included as attachment to the same record, or elsewhere in the dataset (insert links in 'Cross-reference' table)]

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
[Describe why the read-across can be performed (e.g. common functional group(s), common precursor(s)/breakdown product(s) or common mechanism(s) of action]

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
[Provide here, if relevant, additional information to that included in the Test material section of the source and target records]

3. ANALOGUE APPROACH JUSTIFICATION
[Summarise here based on available experimental data how these results verify that the read-across is justified]

4. DATA MATRIX
Cross-referenceopen allclose all
Reason / purpose for cross-reference:
read-across source
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Remarks:
Read-across
Adequacy of study:
weight of evidence
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
secondary literature
Justification for type of information:
Data is from the US EPA.
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Version / remarks:
1997
GLP compliance:
not specified
Limit test:
no
Species:
rabbit
Strain:
New Zealand White
Details on test animals or test system and environmental conditions:
The age of the animals at the beginning of the study was approximately 7 months.
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5% aqueous carboxymethylcellulose
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
Time mated New Zealand White Rabbits were used in the study.
Duration of treatment / exposure:
Gestational days 7-28
Frequency of treatment:
Once daily
Dose / conc.:
800 mg/kg bw/day (nominal)
Dose / conc.:
400 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Vehicle control group
No. of animals per sex per dose:
25 animals per sex per dose
Control animals:
yes, concurrent vehicle
Details on study design:
No Data Available
Maternal examinations:
Mortality, clinical signs, body weight and food intake.
Ovaries and uterine content:
Gravid uterine weight, number of corpora lutea, number of implantations, live fetuses, resorptions, pre- and post-implantation loss.
Fetal examinations:
Live fetuses, sex ratio, fetal weight, total number of fetuses, and gross, skeletal, and visceral malformations/abnormalities.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Clinical signs of convulsions, decreased defecation, soft stool, discolored feces and reddish fluid in refuse pan were observed at the highest dose tested. Slight increases in reduced, soft or discolored feces were noted at 400 mg/kg bw.
Dermal irritation (if dermal study):
not specified
Mortality:
mortality observed, treatment-related
Description (incidence):
At the dose level of 800 mg/kg bw/day, excessive mortality and maternal toxicity were observed (8/25 found dead, 1/25 euthanized due to morbidity). At the dose level of 400 mg/kg bw/day, 1/25 dams died from mechanical injury from gavage.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Significant decreases in body weight gain and food consumption was observed. No changes in body weight, body weight gain or food consumption were noted at 400 mg/kg bw.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Significant decreases in body weight gain and food consumption was observed. No changes in body weight, body weight gain or food consumption were noted at 400 mg/kg bw.
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:
effects observed, treatment-related
Description (incidence and severity):
Necropsy findings included discoloration of the liver, edematous and/or discolored stomach, red discolored and/or edematous intestines, bloody and/or mucoid contents in the intestines at the highest dose group tested. At necropsy the dam that aborted showed an edematous stomach and lquid, bloody contents in the intestines. There were no further substance-related findings at necropsy at other dose groups.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Number of abortions:
effects observed, treatment-related
Description (incidence and severity):
Abortion occurred in 7/25 animals at the highest dose groups tested. Abortion occurred in 1/25 and early delivery in 2/25 at 400 mg/kg bw which was considered to be treatment-related.
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
not specified
Changes in pregnancy duration:
effects observed, treatment-related
Description (incidence and severity):
Early delivery in 2/25 at 400 mg/kg bw was observed which was considered to be treatment-related.
Details on maternal toxic effects:
No Data Available
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
changes in pregnancy duration
clinical signs
early or late resorptions
effects on pregnancy duration
food consumption and compound intake
gross pathology
maternal abnormalities
mortality
number of abortions
pre and post implantation loss
total litter losses by resorption
Abnormalities:
not specified
Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Fetal body weights were significantly lower when compared to controls which was considered to be secondary to the maternal toxicity and not an indicator of developmental toxicity.
Reduction in number of live offspring:
not specified
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
At visceral examination of fetuses, the litter incidence of hemorrhagic iris at 400 mg/kg bw/day was slightly above the historical range while the incidences of gallbladder agenesis, hypoplasia of the gallbladder and azygous lobe of lung absent were slightly increased but well within historical control range.
Dose descriptor:
NOAEL
Effect level:
100 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
external malformations
skeletal malformations
visceral malformations
Abnormalities:
not specified
Developmental effects observed:
no
Treatment related:
no
Conclusions:
Based on all the available observations and results, it was concluded that NOAEL as per the maternal toxicity and fetal parameters was found to be 100 mg/kg bw/day.
Executive summary:

A teratology study was conducted using the test chemical was conducted to analyze the effects on the maternal and fetal health. In this study, four groups each of 25 mated female New Zealand white rabbits per group were treated once per day via oral gavage either with 0.5% carboxymethylcellulose (vehicle) alone (one control group) or with test chemical each at dose levels of 100, 400 and 800 mg/kg bw/day in a dosing volume of 10 ml/kg bw/day. Dosing was initiated on day 5 of gestation and continued to and included day 29 of gestation.  At the dose level of 800 mg/kg bw/day, excessive mortality and maternal toxicity were observed (8/25 found dead, 1/25 euthanized) with clinical signs of convulsions, decreased defecation, soft stool, discolored feces and reddish fluid in refuse pan and with significant decreases in body weight gain and food consumption. Abortion occurred in 7/25 animals. Necropsy findings included discoloration of the liver, edematous and/or discolored stomach, red discolored and/or edematous intestines, and bloody and/or mucoid contents in the intestines. As a result this group was terminated prior to completion of study. At the dose level of 400 mg/kg bw/day, 1/25 dams died from mechanical injury from gavage. Slight increases in reduced, soft or discolored feces were noted. Abortion occurred in 1/25 and early delivery in 2/25 which was considered to be treatment-related. No changes in body weight, body weight gain or food consumption were noted. At necropsy the dam that aborted showed an edematous stomach and liquid, bloody contents in the intestines. There were no further substance-related findings at necropsy. No effects on the following uterine parameters were observed: number of corpora lutea, implantation, live fetuses, resorption, uterine weight and adjusted body weight gain. Fetal body weights were significantly lower when compared to controls which was considered to be secondary to the maternal toxicity and not an indicator of developmental toxicity. At visceral examination of fetuses, the litter incidence of hemorrhagic iris at 400 mg/kg bw/day was slightly above the historical range while the incidences of gallbladder agenesis, hypoplasia of the gallbladder and a zygous lobe of lung absent were slightly increased but well within historical control range. Since all the above findings were within or only slightly above the historical control range, the findings were considered to be spontaneous in nature and unrelated to test substance. Also, no significant treatment-related effects were noted at external and skeletal examinations. At the dose level of 100 mg/kg bw/day there was no mortality observed; abortion occurred in 1/25 animal having mechanical injuries consistent with gavage errors; no changes in body weight, body weight gain or food consumption; no findings at necropsy; no effects on uterine parameters and fetal examinations. Control group: 2/25 animals died from mechanical injury from gavage. Based on all the available observations and results, it was concluded that NOAEL as per the maternal toxicity and fetal parameters was found to be 100 mg/kg bw/day.

Reason / purpose for cross-reference:
read-across source
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Remarks:
Read across
Adequacy of study:
key study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
secondary literature
Justification for type of information:
Data is from the US EPA.
Qualifier:
equivalent or similar to guideline
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Version / remarks:
1997
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
Age at the beginning of treatment: About 8 weeks.
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5%
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
Time mated females were used in this study.
Duration of treatment / exposure:
14 days (GD 6-19).
Frequency of treatment:
Once daily
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Dose / conc.:
400 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Vehicle control
No. of animals per sex per dose:
30 presumed pregnant rats per dose level.
Control animals:
yes, concurrent vehicle
Maternal examinations:
Mortality, clinical signs, body weight and food intake.
Ovaries and uterine content:
Gravid uterine weight, number of corpora lutea, number of implantations, live fetuses, resorptions, pre- and post-implantation loss.
Fetal examinations:
Live fetuses, sex ratio, fetal weight, total number of fetuses, and gross, skeletal, and visceral malformations/abnormalities.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry 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):
No significant effects on gravid uterine weight were observed.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No gross pathological alterations were noted at necropsy from any animal on test.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Number of abortions:
no effects observed
Description (incidence and severity):
No abortions were observed.
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
Pre- and post-implantation losses were similar between the groups.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
No significant changes on resorptions were observed.
Early or late resorptions:
no effects observed
Description (incidence and severity):
No significant changes on early or late resorptions were observed.
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
The numbers of pregnant animals were 30, 29, 28 and 28 and 0, 100, 400 and 1000 mg/kg, respectively.
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
clinical signs
dead fetuses
early or late resorptions
food consumption and compound intake
gross pathology
mortality
pre and post implantation loss
total litter losses by resorption
other: Please see 'remarks'
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
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):
The total numbers of foetuses were 383, 357, 363 and 348 at 0, 100, 400 and 1000 mg/kg, respectively. No significant effects were observed between the groups.
Anogenital distance of all rodent fetuses:
not examined
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One foetus at 1000 mg/kg showed omphalocele and a second foetus from a different litter lacked tail and anal atresia. Because only one foetus was affected in each case, the findings were considered to be spontaneous in nature.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Skeletal abnormalities were limited to an increased incidence of vertebral malformations at 1000 mg/kg (7.1% of litters) compared to the control (0% of the litters); increased incidences of misaligned sternebra at 100 (10.3% of litters) and 1000 mg/kg (10.7% of litters) compared to control (3.3% of litters); and increased incidences of rudimentary ribs at 100 (75.9% of litters) and 1000 mg/kg (78.6% of litters) compared to control (63.3% of litters). The incidence of vertebral malformation at 1000 mg/kg was very slightly above the historical control range (0 to 7%) and was not statistically different from the control group. The incidences of misaligned sternebra at 100 and 1000 mg/kg were within the historical control data (0 to 27% of litters). The incidences of rudimentary ribs at 100 and 1000 mg/kg were above the historical control data (0 to 70%) but at no dose level statistically different from the control group. No clear dose dependency was observed for this effect since the incidence of rudimentary ribs at 400 mg/kg (57.1% of litters) was lower compared to the control group (63.3% of litters).
Visceral malformations:
no effects observed
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/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
external malformations
skeletal malformations
visceral malformations
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Conclusions:
The study-derived NOAEL for maternal and developmental toxicity was 1000 mg/kg bw/day.
Executive summary:

The chemical was given by oral gavage to 30 presumed pregnant rats per dose level at 0 (vehicle control), 100, 400 and 1000 mg/kg bw/day from day 6 to 19 of gestation. All animals survived to planned death. There was no compound-related effect on body weight, body weight gain or food intake. The numbers of corpora lutea, implantations, live foetuses and resorptions, foetal weight, and sex ratio were similar between the groups. Pre- and post-implantation losses were similar between the groups, as was gravid uterine weight. The numbers of pregnant animals were 30, 29, 28 and 28 and 0, 100, 400 and 1000 mg/kg, respectively. No abortions were observed. The total numbers of foetuses were 383, 357, 363 and 348 at 0, 100, 400 and 1000 mg/kg, respectively. No dead foetuses were observed. One foetus at 1000 mg/kg showed omphalocele and a second foetus from a different litter lacked tail and anal atresia. Because only one foetus was affected in each case, the findings were considered to be spontaneous in nature. Visceral findings were unremarkable. Skeletal abnormalities were limited to an increased incidence of vertebral malformations at 1000 mg/kg (7.1% of litters) compared to the control (0% of the litters); increased incidences of misaligned sternebra at 100 (10.3% of litters) and 1000 mg/kg (10.7% of litters) compared to control (3.3% of litters); and increased incidences of rudimentary ribs at 100 (75.9% of litters) and 1000 mg/kg (78.6% of litters) compared to control (63.3% of litters). The incidence of vertebral malformation at 1000 mg/kg was very slightly above the historical control range (0 to 7%) and was not statistically different from the control group. The incidences of misaligned sternebra at 100 and 1000 mg/kg were within the historical control data (0 to 27% of litters). The incidences of rudimentary ribs at 100 and 1000 mg/kg were above the historical control data (0 to 70%) but at no dose level statistically different from the control group. No clear dose dependency was observed for this effect since the incidence of rudimentary ribs at 400 mg/kg (57.1% of litters) was lower compared to the control group (63.3% of litters). The study-derived NOAEL for maternal and developmental toxicity was 1000 mg/kg bw/day. The study was performed according to GLP. 

Data source

Reference
Reference Type:
review article or handbook
Title:
Developmental toxicity study.
Author:
Global Product Compliance (Europe) AB
Year:
2023
Bibliographic source:
WoE report, Global Product Compliance (Europe) AB, 2023.

Materials and methods

Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
Developmental toxicity study has been carried out on test species.
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
Disodium 4,4'-bis[6-anilino-[4-[bis(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate
EC Number:
224-073-5
EC Name:
Disodium 4,4'-bis[6-anilino-[4-[bis(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate
Cas Number:
4193-55-9
Molecular formula:
C40H44N12O10S2.2Na
IUPAC Name:
disodium 2-[(1E)-2-[4-({4-[bis(2-hydroxyethyl)amino]-6-(phenylamino)-1,3,5-triazin-2-yl}amino)-2-sulfonatophenyl]ethenyl]-5-({4-[bis(2-hydroxyethyl)amino]-6-[(cyclohexa-1,3-dien-1-yl)amino]-1,3,5-triazin-2-yl}amino)benzene-1-sulfonate
Test material form:
solid: particulate/powder
Details on test material:
Reference substance name:
Disodium 4,4'-bis[6-anilino-[4-[bis(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate
EC Number:
224-073-5
EC Name:
Disodium 4,4'-bis[6-anilino-[4-[bis(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate
Cas Number:
4193-55-9
Molecular formula:
C40H42N12Na2O10S2
IUPAC Name:
disodium 4,4'-bis[6-anilino-[4-[bis(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate

Test animals

Species:
other: Rat and rabbit

Administration / exposure

Route of administration:
oral: gavage

Results and discussion

Results: maternal animals

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain

Results (fetuses)

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
changes in sex ratio

Overall developmental toxicity

Developmental effects observed:
no
Treatment related:
no

Applicant's summary and conclusion