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Diss Factsheets

Administrative data

developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well documented, according to accepted guidelines

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes (incl. QA statement)
MPI Research 54943 North Main Street Mattawan, Ml 49071-9399 U.S.A.
Limit test:

Test material

Constituent 1
Reference substance name:
Tetrasodium 4,4'-bis[[4-[bis(2-hydroxyethyl)amino]-6-(4-sulphonatoanilino)-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate]
EC Number:
EC Name:
Tetrasodium 4,4'-bis[[4-[bis(2-hydroxyethyl)amino]-6-(4-sulphonatoanilino)-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate]
Cas Number:
tetrasodium 2,2'-ethene-1,2-diylbis[5-({4-[bis(2-hydroxyethyl)amino]-6-[(4-sulfonatophenyl)amino]-1,3,5-triazin-2-yl}amino)benzenesulfonate]
Details on test material:
- Physical state: Pale yellow powder
- Analytical purity: Not reported
- Lot/batch No.: DSR97 10004.006
- Expiration: October 2, 1999
- Storage conditions: Room temperature; protected from light; dessicant added to storage container on July 30, 1999

Test animals

Details on test animals or test system and environmental conditions:
- Source: Charles River Laboratories, Portage, Michigan
- Age at study initiation: 8 weeks
- Weight at study initiation: 170 to 223 g
- Fasting period before study: None
- Housing: Animals were individually housed in suspended, stainless steel, wire-mesh cages
- Diet (e.g. ad libitum): Certified Rodent Chow #5002, PMI Feeds, Inc., St. Louis, Missouri; available ad libitum
- Water (e.g. ad libitum): Tap water; available ad libitum
- Acclimation period: 5 days

- Temperature: 68 to 72°F (20 to 22.2°C)
- Humidity (%): 48 to 71%
- Air changes (per hr): Not reported
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
CMC (carboxymethyl cellulose)
Details on exposure:
The test article was added to the vehicle to achieve the required concentrations, and mixed using a magnetic stir bar. The prepared test article solutions were transferred to amber glass containers by syringe and stored under refrigerated conditions.
Analytical verification of doses or concentrations:
Details on analytical verification of doses or concentrations:
The test article was characterized using a combination of HPLC and 1H and 13C NMR
Details on mating procedure:
- Impregnation procedure: [purchased timed pregnant] Females were time mated upon delivery, and the day on which evidence of copulation was observed was designated gestational day 0
- Proof of pregnancy: Not reported
Duration of treatment / exposure:
From gestational day 6 to 19
Frequency of treatment:
Duration of test:
day 6 of gestation until day 20 of gestation
Doses / concentrations
Doses / Concentrations:
100, 400, and 1000 mg/kg bw
actual ingested
No. of animals per sex per dose:
30 females/group
Control animals:
yes, concurrent vehicle


Maternal examinations:
- Time schedule: At least twice per day during the study period

- Time schedule: Once daily from gestational day 6 through 20

- Time schedule for examinations: Body weights recorded on gestational days 0, 6, 9, 12, 15, 18, and 20. Body weight change was recorded for gestational days 0 to 6, 6 to 9, 9 to 12, 12 to 15, 15 to 18, 18 to 20, 6 to 20, and 0 to 20.

- Food consumption reported as g food consumed/animal/day for gestational days 0 to 6, 6 to 9, 9 to 12, 12 to 15, 15 to 18, 18 to 20, 6 to 20, and 0 to 20.

- Sacrifice on gestation day 20
- Organs examined: Uterus, any other organ showing gross lesions
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: Position of cervix, gross examination of placentae, location of viable and non-viable fetuses, and location of early and late resorptions
Fetal examinations:
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [all per litter]
- Skeletal examinations: Yes: [all per litter]
- Head examinations: No
Differences between groups were assessed using Levene’s test, Dunnett’s test, Welch’s test with a Bonferroni correction, Fischer’s exact test with a Bonferroni correction, Chi-square test, Kruskal-Wallis test, Mann-Whitney U-test, and Pearson Chi-square test.
None reported
Historical control 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:
Discoloured feces was observed in 77 and 100% of the animals in the 400 and 1000 mg/kg groups, respectively. Although this observation was considered to be treatment-related, it was likely due to the presence of the test article in the feces, and its toxicological significance is unknown.

One dam in the 1000 mg/kg group had a liver adhesion and a nodule on the right kidney, and 1 dam in the 100 mg/kg group had granular material on the surface of the right kidney. These findings were considered to be spontaneous and not treatment-related.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
Effect level:
1 000 mg/kg bw (total dose)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
Effect level:
1 000 mg/kg bw (total dose)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
One fetus in the 1000 mg/kg group had omphalocele, and another fetus in the 1000 mg/kg group (from a different litter) had an absent tail and anal atresia. These findings were considered to be spontaneous and not treatment-related.

Vertebral malformation was observed in 7.1% of fetuses in the 1000 mg/kg group, compared to 0% in the control group. Slight increases in the incidence of rudimentary ribs and misaligned sternebra were noted in the 100 and 1000 mg/kg groups compared to the control. The incidence of misaligned sternebra and vertebral malformations were considered to be within historical control ranges, and were therefore not considered biologically relevant or treatment-related. Although the incidence of rudimentary ribs was slightly above the historical control range, a dose-related pattern was not observed, and there was no significant difference in incidence compared to the control group. Thus, the incidence of rudimentary ribs was not considered to be biologically relevant or treatment-related.

Fetal abnormalities

not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

As no adverse maternal or fetal effects were observed following the administration of 100, 400, or 1000 mg/kg/day to rats between gestational days 6 and 19, the results of this study indicate that test substance is not teratogenic to rats.

Applicant's summary and conclusion