Disodium [2,2'-bi-1H-indole]-3,3'-diolate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1971/1972

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Acceptable, well-documented study report which meets basic scientific principles. The registered substance is an aqueous solution of the sodium salt of leucoindigo. It oxidizes in the presence of water and air-borne oxygen rapidly to Indigo (CAS No. 482-89-3) and sodium hydroxide. 

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1972

Report date:

1972

Reference 2

Reference Type:

publication

Title:

Toxicology of Indigo - a review

Author:

Ferber KH

Year:

1987

Bibliographic source:

J. Environ. Pathol. Toxicol. Oncol. 7, 73 -84 (1987)

Reference 3

Reference Type:

publication

Title:

Teratogenic Studies with Certified Colors in Rats and Rabbits.

Author:

Burnett CM et al.

Year:

1974

Bibliographic source:

Toxicol  Appl Pharmacol. 1974;29:121-122

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

no maternal toxicity observed

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Indigo

IUPAC Name:

Indigo

Details on test material:

- Name of test material (as cited in study report): D&C Blue No. 6
- Molecular formula (if other than submission substance): C16H10N2O2
- Molecular weight (if other than submission substance): 262.2628
- Smiles notation (if other than submission substance): O=C3c4ccccc4NC3=C2Nc1ccccc1C2=O
- InChl (if other than submission substance): InChI=1/C16H10N2O2/c19-15-9-5-1-3-7-11(9)17-13(15)14-16(20)10-6-2-4-8-12(10)18-14/h1-8,17-18H
- Structural formula attached as image file (if other than submission substance): see Fig.
- Substance type: active compound
- Physical state: solid
- Analytical purity: 96%
- Lot/batch No.: Y-2206 and Y-2678
- FDA certification no.: Y-PMA-1

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (obtained as weanlings)
- Age at study initiation: satisfactory age for mating (about 8 weeks)
- Weight at study initiation: -
- Fasting period before study: -
- Housing: single
- Diet: Purina Laboratory Chow ad libitum
- Water: tap ad libitum
- Acclimation period: ca. 5 weeks

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: methylcellulose 0.5%

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
suspension in 0.5% methylcellulose


VEHICLE
- Dow Methocel 90 HG, 15000 CPS
- Justification for use and choice of vehicle (if other than water): TS is insoluble in water
- Concentration in vehicle: 50 mg/mL
- Amount of vehicle: 10 mL/kg (control and high dose), 3.2 mL/kg, 1 mL/kg
- Lot/batch no.: MM 101012K

POSITIVE CONTROL
- Acetylsalicylic acid (Aspirin)
- Supplier: Mallinckrodt

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

- Impregnation procedure: cohoused
- 1 M/1 F per cage
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy

Duration of treatment / exposure:

Day 6 through 15 of gestation

Frequency of treatment:

daily

Duration of test:

21 days

Doses / concentrations

Remarks:

Doses / Concentrations:
50, 160, 500 mg/kg/day
Basis:
other: nominal in vehicle

No. of animals per sex per dose:

20 pregnant females

Control animals:

yes, concurrent vehicle

other: positive control: aspirin

Details on study design:

3 dosage levels were run concurrently with 2 negative (vehicle) and one positive (Aspirin) control groups

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule: -

BODY WEIGHT: Yes
- Time schedule for examinations: Days 0, 6, 15, 20 of gestation

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: uterus

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: No data
- Number of implantations: Yes
- Number of resorptions: Yes
- Number viable and dead fetuses: Yes
- Distribution by sex: Yes
- Mean litter size: Yes

Fetal examinations:

- Mean fetal weight
- Determination of sex
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: one-third per litter
- Skeletal examinations: Yes: two-third per litter

Statistics:

Frequency of occurrence and seriousness of anomaly compared to control groups

Indices:

no data

Historical control data:

no data

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
-

Effect levels (maternal animals)

1

Dose descriptor:

NOEL

Effect level:

500 mg/kg bw/day

Basis for effect level:

other: developmental toxicity

2

Dose descriptor:

NOEL

Effect level:

500 mg/kg bw/day (nominal)

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
-

Effect levels (fetuses)

Dose descriptor:

NOEL

Effect level:

500 mg/kg bw/day (nominal)

Basis for effect level:

other: teratogenicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

D&C Blue No. 6 administered by gavage to groups of 20 female rats each during days 6 through 15 of gestation at dose levels of 50, 160, and 500 mg/kg/day was without effect on reproductive performance, maternal weight gain, and fetal development.

Executive summary:

Groups of 20 pregnant female Charles River CD rats received suspensions of D&C Blue No. 6 by stomach tube at doses of 500, 160 and 50 mg/kg/day from day 6 through day 15 of gestation. Comparable groups received either the methylcellulose suspending vehicle as negative controls or aspirin, 250 mg/kg/day as positive controls. On day 20 of gestation Caesarean section was performed, the uterus examined and fetuses processed for subsequent examination for skeletal or visceral anomalies.

On the basis of numbers of viable and dead fetuses, resorption sites, mean fetal weight, distribution by sex, mean litter size, frequency of skeletal and visceral or structural anomalies; weight gain of pregnant females: Indigo was without effect on reproductive performance, maternal weight gain and fetal development.