Trisodium 8-hydroxypyrene-1,3,6-trisulphonate

Developmental toxicity / teratogenicity

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

Justification for type of information:

Data from peer reviewed journal

Data source

Reference

Reference Type:

publication

Title:

Reproductive toxicity of test material was assessed in Osborne-Mendel rats in 20 days study

Author:

COLLINS,et.al

Year:

1989

Bibliographic source:

Fd Chem. Toxic.

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

other: As mentioned below

Principles of method if other than guideline:

The teratogenic toxicity study of test material was performed on rat

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate

EC Number:

247-368-0

EC Name:

Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate

Cas Number:

25956-17-6

Molecular formula:

C18H16N2O8S2.2Na

IUPAC Name:

disodium 6-hydroxy-5-[(2-methoxy-3-methyl-4-sulfonatophenyl)diazenyl]naphthalene-2-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report):FD & C Red NO. 40 or Allura Red AC
- IUPAC name: disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate
- Molecular formula :C18H16N2O8S2.2Na
- Molecular weight:496.4266 g/mole
- Substance type:Organic
- Physical State: Solid

Specific details on test material used for the study:

No data available

Test animals

Species:

rat

Strain:

Osborne-Mendel

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: FDA rat breeding colony
- Age at study initiation: female: 12 - 21 wk
- Weight at study initiation: female: 220 - 270 g
- Fasting period before study: no data
- Housing: Stainless-steel hanging cages.
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): Purina Laboratory Chow ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 25 °C
- Humidity (%): 30 - 63%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12-hr light/dark cycle (8 am 8 pro).

IN-LIFE DATES: From: To:

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
test material , Certified Batch No. AA-4181 was dissolved in distilled water (w/v). Fresh solutions were prepared daily

DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): No data available
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0, 30, 75, 150, 300, 600 or 1000mg/kg/day.
- Amount of vehicle (if gavage): 1ml/100g body weight
- 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: 1 day
- Proof of pregnancy: sperm in vaginal smear was considered day 0 of gestation
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility: no data
- Further matings after two unsuccessful attempts: no data
- After successful mating each pregnant female was caged (how): no data
- Any other deviations from standard protocol: no data

Duration of treatment / exposure:

19 days (Gestation day 0 to day 19)

Frequency of treatment:

Daily

Duration of test:

19 days

Doses / concentrations

Remarks:

0, 30, 75, 150, 300, 600 or 1000mg/kg/day.

No. of animals per sex per dose:

Total: 294-301
0 mg/kg/bw/day : 42-43 females
30mg/kg/bw/day : 42-43 females
75mg/kg/bw/day : 42-43 females
150 mg/kg/bw/day : 42-43 females
300mg/kg/bw/day : 42-43 females
600mg/kg/bw/day : 42-43 females
1000 mg/kg/bw/day : 42-43 females

Control animals:

yes, concurrent vehicle

Details on study design:

No data available

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule:
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS:No data
- Time schedule:

BODY WEIGHT: Yes
- Time schedule for examinations:The rats were weighed daily

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): yes ,food consumption was measured weekly.
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Water intake was not measured.
- Time schedule for examinations:

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
- Other:

Fetal examinations:

- External examinations: Yes: [all per litter ]
- Soft tissue examinations: Yes:The remaining half of the foetuses were fixed in Bouin's solution, serially sectioned by razor blade and examined under a dissecting microscope for internal variations of the soft tissues.
- Skeletal examinations: Yes:Approximately one-half of the foetuses were fixed in alcohol, stained with Alizarin Red S and examined under a dissecting microscope for all skeletal variations
- Head examinations: No data

Statistics:

All data analyses were performed by the Division of Mathematics at the FDA. Data on maternal initial body weights and food consumption were analysed by straight analysis of variance (ANOVA) and two-tailed t-test, and a regression analysis. The number of dams affected was analysed by a Fisher's exact test. Data on maternal weight gain were submitted to an analysis of covariance (ANOCOVA) and a two-tailed t-test. Data on the numbers of implants, corpora lutca, total viable young and viable males and females were analysed by ANOVA followed by a one-tailed t-test. Data on implantation efficiency, early deaths, late deaths and total resorptions (early and late deaths) were transformed by using the Freeman-Tukey arc-sine transformation (Freeman and Tukey, 1950) and then analysed by ANOVA and a one-tailed t-test. Data on litters having one or more or two or more resorptions were analysed by a Fisher's exact test. Similar tests were applied to the number of runts per litter. Data on foetal body weights, crown-!o-rump lengths and foetal ossified vertebrae were analysed by nested ANOVA and a one-tailed t-test. The ANOVA included a correction for unequal sample size (Sokal and Rohlf, 1981). Data on the average number of foetuses per litter with skeletal, sternebral, combined missing plus incomplete plus bipartite sternebrae or soft-tissue variations were transformed by using the Frceman-Tukey arc-sine transformation and then analysed by ANOVA and a one-tailed t-test. Litters with foetuses with at least one, at least two, etc. skeletal, sternebral, combined missing plus incomplete plus bipartite sternebrae, or soft-tissue variations, and specific variations were analysed by Fisher's exact test.

Indices:

No data available

Historical control data:

No data available

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Description (incidence and severity):

No unusual behaviours were observed in the animals during the study.The external maternal findings were unremarkabke.

Dermal irritation (if dermal study):

not specified

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One female inthe group given 300mg/kg died at day 12 of gestation as a result of gavage difficulties unrelated to dosage

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Initial body weight at day 0 and maternal body-weight gain during gestation were similar in all groups

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Mean food consumption on days 0-7, 7-14, 14 -20 and 0-20 by the treated animals was similar to that of the control animals

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:

not specified

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:

no effects observed

Description (incidence and severity):

No litters were totally resorbed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

The mean numbers of corpora lutea and implants per female were similar in all groups

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

The percentage of females with at least one resorption was similar in all groups. The percentage of females given 600mg/kg that had at least two resorptions was significantly greater than the percentage in the control females, but there was no dose related effect in the percentage of females with at least two resorptions.

Early or late resorptions:

no effects observed

Description (incidence and severity):

The number of early deaths per litter and the number of early plus late deaths per litter were greatest in the 600 mg/kg group, but these appeared to be random occurrences. The percentage of females with at least one resorption was similar in all groups. The percentage of females given 600mg/kg that had at least two resorptions was significantly greater than the percentage in the control females, but there was no dose related effect in the percentage of females with at least two resorptions.

Dead fetuses:

no effects observed

Description (incidence and severity):

The mean number of viable foetuses per female was similar in all groups. The number of viable male foetuses was increased over the control value in the groups given 30 and 1000mg/kg, but because of the lack of relation to dosage, these increases were considered to be random. The number of viable female foetuses was not affected in any group.

Changes in pregnancy duration:

not specified

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed

Changes in number of pregnant:

no effects observed

Description (incidence and severity):

The pregnancy rate ranged from 85.71 to 95.35% with no evidence of dose correlation.

Other effects:

not specified

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

changes in number of pregnant

clinical signs

dead fetuses

early or late resorptions

food consumption and compound intake

mortality

number of abortions

pre and post implantation loss

total litter losses by resorption

Remarks on result:

other: No maternal toxic effects on was observed

Maternal abnormalities

Abnormalities:

not specified

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Mean foetal weights of males and females and crown rump lengths were similar in all groups. The number of litters with runts was similar in all groups.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not specified

Reduction in number of live offspring:

not specified

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):

Aside from haemorrhages present in foetuses in all groups in similar numbers, there were no other external variations in any of the dosed groups. Two foetuses had multiple anomalies: one foetus from the control group had a club foot, a short tail and four digits on the hind legs, and one foetus from the 150-mg/kg group had exencephalus and hydrocephalus. Three foetuses in a single litter of a female given 75 mg/kg were oedematous

Skeletal malformations:

no effects observed

Description (incidence and severity):

No dose-related increase in sternebral variations was seen among the foetuses with reduced ossification, bipartite, missing, malaligned or fused sternebrae,nor in the numbers of litters containing these foetuses. The incidence of specific skeletal variations was similar in all groups of foetuses, except for a significant increase in the incidence of 14th rib bud in foetuses from the 300 mg/kg group which was considered to be a random occurrence.

Visceral malformations:

no effects observed

Description (incidence and severity):

The numbers of soft-tissue variations in foetuses and their incidences per litter were similar in all groups except for a significant increase in the 150-mg/kg group in the number of litters with at least one variation; this increase was considered a random occurrence

Other effects:

not specified

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

external malformations

skeletal malformations

visceral malformations

Remarks on result:

other: No developmental toxic effects was observed

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Treatment related:

not specified

Relation to maternal toxicity:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Applicant's summary and conclusion

Conclusions:

In developmental toxicity study, The NOAEL was considered to be 1000mg/kg bw as no effects on development of fetus was observed , when Osborne-Mendel male and female rats were treated with test material orally in 20 days

Executive summary:

The developmental toxicity study of test material was performed on male and female Osborne-Mendel rats. The test material was dissolved in distilled water (w/v). Fresh solutions were prepared daily. The dose concentration of 0, 30, 75, 150, 300, 600 or 1000mg/kg/day was administered by oral gavage route in volume 1ml/100g body weight .On mating days , two females were randomly mated with one male at approximately 4.30pm. The following morning, a vaginal smear was obtained from each female to determine whether copulation had taken place. Sperm positive dams were considered to be at day 0 of gestation. 42-43 female/dose groups were used. All the animals were observed for signs of toxicity. The rats were weighed daily and food consumption was measured weekly. Water intake was not measured. On day 20 of gestation, the females were examined for gross abnormalities for the last time before being killed by CO, asphyxiation. Caesarean sections were performed. Corpora lutea were counted. The uterus was opened and examined in situ. The uterine positions of all implantation sites were noted and their condition (early or late resorptions, living or dead foetuses) was determined. Each live foetus was promptly weighed, sexed and examined for gross external malformations, and the crown-rump length was measured. Any foetus that weighed less than 70% of the average weight of the concurrent male or female controls was considered to be a runt. Approximately one-half of the foetuses were fixed in alcohol, stained with Alizarin Red S and examined under a dissecting microscope for all skeletal variations. The remaining half of the foetuses were fixed in Bouin's solution, serially sectioned by razor blade and examined under a dissecting microscope for internal variations of the soft tissues.

No unusual behaviours were observed in the animals during the study. The external maternal findings were unremarkable. One female in the group given 300mg/kg died at day 12 of gestation as a result of gavage difficulties unrelated to dosage. Initial body weight at day 0 and maternal body-weight gain during gestation were similar in all groups. Mean food consumption on days 0-7, 7-14, 14 -20 and 0-20 by the treated animals was similar to that of the control animals. The pregnancy rate ranged from 85.71 to 95.35% with no evidence of dose correlation.

The mean numbers of corpora lutea and implants per female were similar in all groups. No litters were totally resorbed. The mean number of viable foetuses per female was similar in all groups. The number of viable male foetuses was increased over the control value in the groups given 30 and 1000mg/kg, but because of the lack of relation to dosage, these increases were considered to be random. The number of viable female foetuses was not affected in any group. The number of early deaths per litter and the number of early plus late deaths per litter were greatest in the 600 mg/kg group, but these appeared to be random occurrences. The percentage of females with at least one resorption was similar in all groups. The percentage of females given 600mg/kg that had at least two resorptions was significantly greater than the percentage in the control females, but there was no dose related effect in the percentage of females with at least two resorptions.Mean foetal weights of males and females and crown rump lengths were similar in all groups. The number of litters with runts was similar in all groups.Aside from haemorrhages present in foetuses in all groups in similar numbers, there were no other external variations in any of the dosed groups. Two foetuses had multiple anomalies: one foetus from the control group had a club foot, a short tail and four digits on the hind legs, and one foetus from the 150-mg/kg group had exencephalus and hydrocephalus. Three foetuses in a single litter of a female given 75 mg/kg were oedematous. No dose-related increase in sternebral variations was seen among the foetuses with reduced ossification, bipartite, missing, malaligned or fused sternebrae, nor in the numbers of litters containing these foetuses. The incidence of specific skeletal variations was similar in all groups of foetuses, except for a significant increase in the incidence of 14th rib bud in foetuses from the 300 mg/kg group which was considered to be a random occurrence. Hence the NOAEL was considered to be 1000mg/kg bw as no effects on development of fetus was observed, when Osborne-Mendel male and female rats were treated with test material orally in 20 days.