Sodium 4-[[3-(acetylamino)phenyl]amino]-1-amino-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14. Sep to 25 Nov 1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

yes

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-(3-(4-amino-9,10-dihydro-3-sulpho-9,10-dioxoanthracen-4-yl)aminobenzenesulphonyl)vinyl) disodium sulphate

EC Number:

219-949-9

EC Name:

2-(3-(4-amino-9,10-dihydro-3-sulpho-9,10-dioxoanthracen-4-yl)aminobenzenesulphonyl)vinyl) disodium sulphate

Cas Number:

2580-78-1

Molecular formula:

C22H18N2O11S3.2Na C22H18N2Na2O11S3

IUPAC Name:

disodium 1-amino-9,10-dioxo-4-[(3-{[2-(sulfonatooxy)ethyl]sulfonyl}phenyl)amino]-9,10-dihydroanthracene-2-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): C.I. Reactive Blue 19
- Physical state: solid
- Appearence: dark powder

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Hoechst AG
- Age at study initiation: 65 to 75 days
- Weight at study initiation: 193.2 +/- 14 g
- Fasting period before study: NA
- Housing: single
- Diet: Altromin 1310 ad libitum
- Water: tap water ad libitum
- Acclimation period: at least 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 22°C
- Humidity (%): 48 to 61%
- Air changes (per hr): 16 to 20
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 14. Sep To: 25. Nov 1993

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

- Frequency of preparation: daily
- Administration: within 3 hours after preparation


VEHICLE
- Concentration in vehicle: 200 mg/kg nominal
- Amount of vehicle (if gavage): 5 ml/kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

homogeneity and stability of test formulations were analytically verified

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: 1:1
- Length of cohabitation: over night (3:30 pm to 7:30 am next day)
- Verification of same strain and source of both sexes: yes - own breeding facility
- Proof of pregnancy: sperm in vaginal smear, referred to as day 1 of pregnancy

Duration of treatment / exposure:

7. - 16. day of pregnancy

Frequency of treatment:

daily

Duration of test:

cesarean section on Day 21 of pregnancy

Doses / concentrations

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

23 to 24 mated females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: test item was tolerated in the acute and subacute studies without adverse effects
The study compound was suspended fresh daily in a concentration of 200 g/L in distilled water and administered daily to all the animals within three hours of preparation.
All animals received the same volume of liquid, 5 ml/kg body weight. After each weighing of the animals, the compound doses were adjusted according to body weight. The stability and homogeneity of the soiutions were guaranteed over a period of 4 hours

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE : Yes
- Time schedule for examinations: weekly
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/100 g body weight/day: Yes


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: all organs examined macroscopically
uterus - live and dead fetuses, resorption sites, placentas
ovaries - corpora lutea


OTHER:
- diameter of conceptuses undergoing resorption
- placenta weights
- presence of iron in uterus walls with ammonium sulphide to detect invisible implantation sites

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Placenta 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: all per litter
- Body weight: Yes: all per litter
- Crown-rump length: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No data

Statistics:

comparison to actual control group and historical controls
MANOVA: body weight development, fetal weight, placental weight
PURI&SEN rank order test: food intake
Mantel-Haenszel's chi-squared test: live fetuses, intrauterine fetal death, number of implants, number of corpora lutea
multivariate analysis of variance: litter means of fetal weights, crown-rump length, placental weights
Fisher test: autopsy findings, body cross-sections, skelettal examination

Dams which had no live fetuses were excluded from the calculation of mean values and statistical evaluation

Indices:

No data

Historical control data:

Yes

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food efficiency:

no effects observed

Gross pathological findings:

no effects observed

Maternal developmental toxicity

Number of abortions:

no effects observed

Description (incidence and severity):

20 dams in the compound group and 19 dams in the control group carried live conceptuses to term and were delivered by caesarean section

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:

no effects observed

Changes in number of pregnant:

no effects observed

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
black discoloration of faeces and red discoloration of urine due to excretion of dye

Effect levels (maternal animals)

Key result

Dose descriptor:

NOEL

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 efficiency

gross pathology

maternal abnormalities

mortality

necropsy findings

number of abortions

pre and post implantation loss

total litter losses by resorption

Maternal abnormalities

Abnormalities:

no effects observed

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): 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

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Visceral malformations:

no effects observed

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
only minor anomalies or variation within the historical range of spontaneous findings were observed

Effect levels (fetuses)

Key result

Dose descriptor:

NOEL

Effect level:

> 1 000 mg/kg bw/day (nominal)

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

Fetal abnormalities

Abnormalities:

no effects observed

Overall developmental toxicity

Developmental effects observed:

no

Applicant's summary and conclusion

Conclusions:

No adverse effects were observed after daily administration of 1000 mg/kg/day Reactive Blue 19 in dams or their fetuses.
Maternal NOEL: 1000 mg/kg/day
Fetal NOEL: 1000 mg/kg/day

Executive summary:

In this limit test, the test substance, dissolved in distilled water, was administered orally by stomach tube in a single daily dose of 1000 mg/kg body weight to a group of 20 pregnant female Wistar rats from the 7th - 16th day of pregnancy. A simultaneous control group of the same size received the vehicle without test compound. On the 21st day of pregnancy, the dams were killed and delivered by caesarean section. The foetuses were then examined morphologically for developmental disorders.

The studies showed that the repeated oral administration of of the test substance, at a dose level of 1000 mg/kg body weight in the sensitive phase of organogenesis for the conceptuses, did not lead to any impairment of the general physical condition of the dams or impaired intrauterine development of conceptuses.

The morphological examination of the foetuses with regard to stage of development, outwardly detectable anomalies as well as anomalies of the internal (organs and the skeleton showed no Indication of an embryotoxic or teratogenic effect of the compound. The findings observed are to be regarded as spontaneous in origin.

On the basis of the results of this limit test, the "no observed adverse effect level" for the test substance in rats following oral administration lies at 1000 mg/kg body weight with regard to maternal and embryofoetal toxicity and teratogenicity. No teratogenic effect was observed.