1,4-Benzenedisulfonic acid, 2,2'-[1,2-ethenediylbis[(3-sulfo-4,1-phenylene)imino[6-[bis(2-hydroxypropyl)amino]-1,3,5-triazine-4,2-diyl]imino]]bis-, sodium salt (1:6)

Repeated dose toxicity: oral

Administrative data

Endpoint:

chronic toxicity: oral

Remarks:

combined repeated dose and carcinogenicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Basic data given: comparable to guidelines/standards

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1978

Report date:

1978

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1977

Report date:

1977

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 4,4'-bis[(4-anilino-6-morpholino-1,3,5-triazin-2-yl)amino]stilbene-2,2'-disulphonate

EC Number:

240-245-2

EC Name:

Disodium 4,4'-bis[(4-anilino-6-morpholino-1,3,5-triazin-2-yl)amino]stilbene-2,2'-disulphonate

Cas Number:

16090-02-1

IUPAC Name:

disodium 2,2'-ethene-1,2-diylbis{5-[(4-anilino-6-morpholin-4-yl-1,3,5-triazin-2-yl)amino]benzenesulfonate}

Details on test material:

- Analytical purity: 92%

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Winkelmann, Borchen
- Age at study initiation: 28 to 32 days
- Weight at study initiation: males: 53 g; females: 54 g (mean)
- Diet: Altromin R-Pulverfutter ( Altromin GmbH, Lage), ad libitum
- Water: tap water ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±3 °C

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

24 months

Frequency of treatment:

continously in food

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 100, 1000, 10000 ppm
Basis:
nominal in diet

No. of animals per sex per dose:

50

Control animals:

yes, plain diet

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily


DETAILED CLINICAL OBSERVATIONS: No data


BODY WEIGHT: Yes
- Time schedule for examinations: weekly until study week 27 and every second week thereafter


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes


OPHTHALMOSCOPIC EXAMINATION: No data


HAEMATOLOGY: Yes
- Time schedule for collection of blood: after 1, 3, 6, 12 and 24 month.
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: 5 male and 5 female rats after 1, 3, 6 and 12 month and from 10 male and 10 female rats after 24 month.
- Parameters examined: Erythrocyte count, Hemoglobin, Leukocyte count, Reticulocyte count, Thrombocyte count, differential counts, Hematocrit


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after 1, 3, 6, 12 and 24 month.
- Animals fasted: No data
- How many animals: 5 male and 5 female rats after 1, 3, 6 and 12 month and from 10 male and 10 female rats after 24 month.
- Parameters examined: Alkaline Phosphatase, Total Protein, Albumin, Glucose, Urea, Creatinine, Bilirubin, total Cholesterol, Aspartate aminotransferase, Alanine aminotransferase,


URINALYSIS: Yes
- Time schedule for collection of urine: after 1, 3, 6, 12 and 24 month.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- Parameters examined: Glucose, Blood, total Protein, pH, Bilirubin, Urobilirubin, Sedime


NEUROBEHAVIOURAL EXAMINATION: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, necropsies were performed on all animals
HISTOPATHOLOGY: Yes, adrenals, aorta, brain, epididymides, eyes, femur, heart, ichiatic nerve, intestine, kidneys, liver, lung, lymph nodes, muscle, esophagus, ovaries, pancreas, pituitary gland, prostate seminal vesicle, salivary gland, spleen, sternum, stomach, testes, trachea, thyroids, urinary bladder, uterus, as well as all gross pathological lesion were sampled and subjected to histopathological examination.

Other examinations:

The organ weights of adrenals, heart, kidneys, liver, lung, ovaries, spleen, testes and thyroid gland were recorded

Statistics:

Mann-Whitney and Wilcoxon test

Results and discussion

Results of examinations

Details on results:

CLINICAL SIGNS AND MORTALITY
Treatment with FWA-1 did not affect mortality, appearance or behavior of treated animals.

BODY WEIGHT AND WEIGHT GAIN
Body weight development of treated animals were similar to those of the control group.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Food consumption of treated animals were similar to those of the control group.

HAEMATOLOGY
The assessment of hematological data did not indicate any adverse effects in treated animals. The significantly and dose dependently increased number of thrombocytes in female rats after one month in all dose groups, was not considered adverse because there was no confirmation of these findings in the further course of the study and all values were within historical control ranges of this Wistar rat strain. The not dose-dependent but statistically significant decrease in the number of reticulocytes in males after 3 months was also considered not to be toxicologically relevant because of the same reasons

CLINICAL CHEMISTRY
The assessment of clinical biochemical data did not indicate treatment related disturbances. Slightly and not dose-dependently but statistically significant increased ALAT (GPT) activities were observed in males after 24 months at the end of the study in all dose groups. Slightly and not dose-dependently but statistically significant increased protein concentrations in blood serum were observed after 6 months in both sexes and all dose groups as well as after 24 months in males in all dose groups. These effects on ALAT and serum protein were considered not to be toxicologically relevant, but due to relative low control values as compared with normal historical data in this Wistar rat strain.

URINALYSIS
The assessment or urine analysis data did not indicate treatment related disturbances.

ORGAN WEIGHTS
Treatment at 10000 ppm slightly increased absolute liver and kidney weights in males to 111% and 106% of control, respectively. Absolute ovary weights were increased to 127% of control at 10000 ppm. The increased organ weights were considered not to be toxicologically relevant by the study authors, because there were no accompanying hematological, biochemical or histopathological changes.

GROSS PATHOLOGY
A number of benign and malignant neoplasms, including thyroid interstitial cell neoplasia, pituitary neoplasia, endometrial neoplasia, phaeochromocytoma, and testicular interstitial cell neoplasia, were noted in all dose groups including controls. However, statistical analysis of tumor incidences revealed no significant differences between control and treated groups. In addition, the tumor incidences were not organ or neoplastic class specific and therefore were regarded not to be biologically significant.

HISTOPATHOLOGY: NON-NEOPLASTIC
Histopathological investigations revealed no evidence for treatment related changes.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

10 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects, highest dose tested

2

Dose descriptor:

NOAEL

Effect level:

523.9 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: overall effects, highest dose tested

3

Dose descriptor:

NOAEL

Effect level:

790.6 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: overall effects, hightest dose tested

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Based on the outcomes of this 'Combined Oral Chronic Toxicity/Carcinogenicity' study in Wistar rats, FWA-1 is considered not to be toxic at dietary levels upt to 10000 ppm, corresponding with 524 mg/kg bw/day for males and with 791 mg/kg bw/day for females.