2-{(E)-[6-({4-chloro-6-[(4-{[2-(sulfooxy)ethyl]sulfonyl}phenyl)amino]-1,3,5-triazin-2-yl}amino)-1-hydroxy-3-sulfonaphthalen-2-yl]diazenyl}naphthalene-1,5-disulfonic acid

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Reproductive toxicity study

Based on the data available from different studies, NOAEL for test material was considered to be in range of  500-1500mg/kg bw/day for F0, F1 and F2 generation. When male and female CD rats were treated with test material orally. Thus, comparing this value with the criteria of CLP regulation test material is not likely to classify as reproductive toxicant.

 

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

Experimental data of read across substances

Justification for type of information:

Data for the target chemical is summarized based on the structurally similar read across chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

other: As mentioned below

Principles of method if other than guideline:

WoE report is based on two reproductive toxicity studies on rats
1.The reproductive effects of test material on male and female Charles River CD rats by oral (Diet) route at different dose levels was examined.
2.Multigeneration toxicity study of test material was performed on male and female wistar rats.

GLP compliance:

not specified

Limit test:

no

Justification for study design:

No data available

Species:

rat

Strain:

other: 1.Charles River CD 2.Wistar

Details on species / strain selection:

No data available

Sex:

male/female

Details on test animals or test system and environmental conditions:

1. TEST ANIMALS
- Source:No data available
- Females (if applicable) nulliparous and non-pregnant: [yes/no]
- Age at study initiation: (P) x wks; (F1) x wksNo data available
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x gNo data available
- Fasting period before study:No data available
- Housing:Animals were housed individually
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum):the test diet ad libitum
- Water (e.g. ad libitum):No data available
- Acclimation period:No data available
ENVIRONMENTAL CONDITIONS
- Temperature (°C):No data available
- Humidity (%):No data available
- Air changes (per hr):No data available
- Photoperiod (hrs dark / hrs light):
IN-LIFE DATES: From: To:No data available

Route of administration:

oral: feed

Type of inhalation exposure (if applicable):

not specified

Vehicle:

not specified

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: test material mixed with food

Details on mating procedure:

No data available

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

1. Pre-Mating Exposure / Males and Females: 60 days and test material adminstered upto three generation
2.From F1 to F3 generation

Frequency of treatment:

Daily

Details on study schedule:

No data available

Remarks:

Doses / Concentrations:
Study 1.
5, 50, 150 or 500 mg/kg bw/day
Study 2.
0, 0.1, 1.0 or 3% (0, 50, 500,1500 mg/kg bw per day)
Basis:

No. of animals per sex per dose:

Study1.
Total :150
0mg/kg bw/day : 10 males and 20 females
5 mg/kg bw/day : 10 males and 20 females
50 mg/kg bw/day :10 males and 20 females
150 mg/kg bw/day :10 males and 20 females
500 mg/kg bw/day : 10 males and 20 females


Study2.
Total:360
0 mg/kg bw: 60 male and 60 female
50 mg/kg bw : 40male and 40 female
500 mg/kg bw : 40male and 40 female
1500 mg/kg bw : 40male and 40 female
After a nine-week test period, 24 males and 24 females from the control group, and 14 males and 14 females from each test group were used for teratogenicity studies; the remainder were used for the reproduction study.

Control animals:

yes

Details on study design:

No data available

Positive control:

No data available

Parental animals: Observations and examinations:

Study1.
Parental animals observation and examinations
CAGE SIDE OBSERVATIONS: yes
Clinical observations were recorded twice daily with at least 5 hours between observations

DETAILED CLINICAL OBSERVATIONS: Yes

Time schedule: Detailed physical examinations and palpation for masses were performed weekly.

BODY WEIGHT: Yes
Time schedule for examinations: weekly for the first fourteen weeks, bi-weekly for the next 12 weeks and every 4 weeks thereafter until the end of the study.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes weekly for the first fourteen weeks, bi-weekly for the next 12 weeks and every 4 weeks thereafter until the end of the study..

Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes ,

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
Time schedule for examinations:

OTHER: Haematology tests, including hemoglobin, hematocrit, erythrocyte and total and differential leukocyte counts, and erythrocyte morphology, were conducted on ten randomly selected animals at months 3, 6, 12, 18 and 24 of the study.

Study2.
CAGE SIDE OBSERVATIONS: yes
- Time schedule:
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS: Yes / No / No data
- Time schedule:

BODY WEIGHT: Yes
- Time schedule for examinations:

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

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
- Time schedule for examinations:

OTHER:

Oestrous cyclicity (parental animals):

No data available

Sperm parameters (parental animals):

No data available

Litter observations:

2.STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: [yes/no]
- If yes, maximum of [...] pups/litter ([...]/sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, were observed

GROSS EXAMINATION OF DEAD PUPS:yes


ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY:

ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY:

Postmortem examinations (parental animals):

1.SACRIFICE
- Male animals: All surviving animals [describe when, e.g. as soon as possible after the last litters in each generation were produced.]
- Maternal animals: All surviving animals [describe when, e.g. after the last litter of each generation was weaned.]

GROSS NECROPSY
Necropsies were conducted on all animals dying prior to study termination, killed in a moribund condition or killed on schedule,Tissues examined included adrenal glands, aorta, blood smear, brain, cecum, colon, duodenum, epididymus or uterus, esophagus, eyes, femur including marrow, tissue masses, gallbladder, heart, ileum, jejunum, duodenum, kidneys, liver, lungs and bronchi, mammary gland, nerves (sciatic), ovaries, lymph nodes, pancreas, parathyroids, pituitary gland, prostrate, rectum, skin, spleen, seminal vesicles, skeletal muscle, testes with ep ididymides, stomach, thymus, thyroid gland including parathyroid, trachea, urinary bladder, uterus


HISTOPATHOLOGY / ORGAN WEIGHTS
Histological examinations were conducted on all animals from both control groups, the highest dose group (2.0 or 5.0%) from each study and also on 10 rats randomly selected from each group for an interim sacrifice at 12 months. Histology was also performed on any animal with gross lesions or masses


Study 2.
Postmortem examinations (Parent Animal)
SACRIFICE
Yes, Autopsy did of parent rats

GROSS NECROPSY:No data available

HISTOPATHOLOGY / ORGAN WEIGHTS: No data available

Postmortem examinations (offspring):

2.Postmortem examinations (offspring)
SACRIFICE
F1,F2, F3 pups were sacrificed.

GROSS NECROPSY
Yes


HISTOPATHOLOGY / ORGAN WEIGTHS
Yes

Statistics:

No data available

Reproductive indices:

No data available

Offspring viability indices:

No data available

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

1.No treatment-related effects were reported on survival

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on haematological finding

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on clinical chemistry

Urinalysis findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on urinalysis.

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

1.no compound related lesions in any tissue examined histologically, including kidneys and adrenal glands from parental rats.

Histopathological findings: neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

1.Histological evaluation revealed a variety of lesions, including neoplasms, present at similar incidences in control and treated animals.The authors considered the lesions to be spontaneous and not related to administration of the test material.

Other effects:

not specified

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Description (incidence and severity):

1.There were no compound related effects on fertility, gestation, pup viability or lactation indices, on reproductive organs of females, or on organ weights among parents.
2.No treatment-related effects on reproductive function was observed

Dose descriptor:

NOAEL

Effect level:

> 500 - < 1 500 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

haematology

clinical biochemistry

urinalysis

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

histopathology: neoplastic

reproductive performance

other: see 'Remark'

Remarks on result:

other: No effects on reproductive performance was observed

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not specified

Mortality / viability:

no mortality observed

Description (incidence and severity):

1&2 .no compound related effects on pup viability or lactation indices.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

2.No effects on body weight was observed

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

1.Food consumption was similar for control and treated animals at the lower dietary levels, but was slightly higher in the 500 mg/kg bw dose group although not statistically significant.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Description (incidence and severity):

2.No treatment-related effects were reported on Haematological finding

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on clinical chemistry

Urinalysis findings:

not specified

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

1&2.no compound related effects on organ weights

Gross pathological findings:

no effects observed

Description (incidence and severity):

1.Necropsies did not reveal any treatment-related gross or microscopic changes.

Histopathological findings:

no effects observed

Description (incidence and severity):

1.There were no compound related lesions in any tissue examined histologically, including kidneys and adrenal glands from offspring.

Other effects:

no effects observed

Description (incidence and severity):

1.There were no compound related effects on fertility, gestation, pup viability or lactation indices, on reproductive organs of females, or on organ weights among offspring.

Behaviour (functional findings):

not specified

Developmental immunotoxicity:

not specified

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

> 500 - < 1 500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

not specified

Basis for effect level:

viability

mortality

food consumption and compound intake

haematology

clinical biochemistry

urinalysis

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

Remarks on result:

other: Overall no developmental toxic effects were observed

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not specified

Mortality / viability:

no mortality observed

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

1.Food consumption was similar for control and treated animals at the lower dietary levels, but was slightly higher in the 500 mg/kg bw dose group although not statistically significant.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on Haematological finding

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on clinical chemistry

Urinalysis findings:

no effects observed

Description (incidence and severity):

1.No treatment-related effects were reported on urinalysis

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

1&2.no compound related effects on organ weights

Gross pathological findings:

no effects observed

Description (incidence and severity):

1.Necropsies did not reveal any treatment-related gross or microscopic changes.

Histopathological findings:

no effects observed

Description (incidence and severity):

1.There were no compound related lesions in any tissue examined histologically, including kidneys and adrenal glands from offspring.

Other effects:

no effects observed

Description (incidence and severity):

1.There were no compound related effects on fertility, gestation, pup viability or lactation indices, on reproductive organs of females, or on organ weights among offspring.

Behaviour (functional findings):

not specified

Developmental immunotoxicity:

not specified

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

> 500 - < 1 500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

not specified

Basis for effect level:

viability

mortality

food consumption and compound intake

haematology

clinical biochemistry

urinalysis

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

Remarks on result:

other: overall no developmental effects were observed

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Reproductive effects observed:

not specified

Treatment related:

not specified

Relation to other toxic effects:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Conclusions:

NOAEL was considered to be in range of 500-1500mg/kg bw/day for F0, F1 and F2 generation. When male and female CD rats were treated with test material orally.

Executive summary:

Data available for the read across chemicals was reviewed to determine the reproductive toxicity of test material  .The studies are as mentioned below:

Study 1

Three generation reproductive study of test material was performed on male and female Charles River CD rats. Animals were housed individually and fed the test diet ad libitum. The test material mixed with food in dose concentration 0, 5, 50, 150 or 500 mg/kg bw/day and 10 males and 20 females/group/generation were used .Pre-Mating Exposure / Males and Females were 60 days. Clinical observations were recorded twice daily with at least 5 hours between observations. Detailed physical examinations and palpation for masses were performed weekly. Body weights and food consumption were determined weekly for the first fourteen weeks, bi-weekly for the next 12 weeks and every 4 weeks thereafter until the end of the study. The intake of the test substance was determined from body weight, food consumption and dietary concentration. Haematology tests, including haemoglobin, haematocrit, erythrocyte and total and differential leukocyte counts, and erythrocyte morphology, were conducted on ten randomly selected animals at months 3, 6, 12, 18 and 24 of the study. Necropsies were conducted on all animals dying prior to study termination, killed in a moribund condition or killed on schedule. Histological examinations were conducted on all animals from both control groups, the highest dose group (2.0 or 5.0%) from each study and also on 10 rats randomly selected from each group for an interim sacrifice at 12 months. Histology was also performed on any animal with gross lesions or masses. Also tissues examined included adrenal glands, aorta, blood smear, brain, cecum, colon, duodenum, epididymus or uterus, esophagus, eyes, femur including marrow, tissue masses, gallbladder, heart, ileum, jejunum, duodenum, kidneys, liver, lungs and bronchi, mammary gland, nerves (sciatic), ovaries, lymph nodes, pancreas, parathyroids, pituitary gland, prostrate, rectum, skin, spleen, seminal vesicles, skeletal muscle, testes with epididymides, stomach, thymus, thyroid gland including parathyroid, trachea, urinary bladder, uterus.

 At the end of study, no treatment-related effects were reported on survival. No treatment-related changes were reported at gross necropsy. Histological evaluation revealed a variety of lesions, including neoplasms, present at similar incidences in control and treated animals. The authors considered the lesions to be spontaneous and not related to administration of the test material. There were no compound related effects on fertility, gestation, pup viability or lactation indices, on reproductive organs of females, or on organ weights among parents and offspring. There were no compound related lesions in any tissue examined histologically, including kidneys and adrenal glands from parental rats or from offspring. There was no toxicity to either the F1 or F2 generation. Food consumption was similar for control and treated animals at the lower dietary levels, but was slightly higher in the high-dose study, although not statistically significant. Haematological, clinical chemistry and urinalysis parameters did not differ significantly from the controls. Necropsies at one year did not reveal any treatment-related gross or microscopic changes. Hence NOAEL was considered to be 500mg/kg bw/day for F0, F1 and F2 generation. When male and female Charles River CD rats were treated with test material orally.

Study 2

A multigeneration reproductive toxicity study of test material was performed on male and female wistar rats. The test material mixed with feed in dose concentration 0, 0.1, 1.0 or 3% (0, 50, 500, 1500 mg/kg bw per day) and administered orally for three successive generations. Each generation having 60 animals of each sex in the control and 40 animals of each sex in test animals. No adverse effects were observed with respect to fertility, litter size and weight, general condition, male/female ratio, growth during lactation, survival or maturation. Autopsy of parent rats and pups at weaning did not reveal any treatment related changes in organ weights other than caecal enlargement in the 3% dose group. Gross and microscopic examination of the F3 generation at weaning did not reveal any abnormalities due to treatment and no adverse effects were seen in the teratology study. It was concluded that Brilliant Black PN did not exert any adverse effects on reproductive function of Wistar rats when fed at dietary levels up to 3% (1500 mg/kg bw per day) for three successive generations. Therefore, NOAEL was considered to be at 3% (1500 mg/kg bw per day) for F0 F1 and F2 generation .When male and female wistar rats were treated with test material orally.

 

 Thus, Based on the data available for the read across chemical, No Observed Adverse Effect Level (NOAEL) was considered to be above 500 -1500mg/kg bw . Thus, comparing this value with the criteria of CLP regulation test material is not likely to classify as reproductive toxicant.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

500 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

Data is Klimicsh 2 and from authoritative database

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Reproductive toxicity

Data available for the read across chemicals was reviewed to determine the reproductive toxicity of test material .The studies are as mentioned below:

 Study 1

Three generation reproductive study of test material was performed on male and female Charles River CD rats. Animals were housed individually and fed the test diet ad libitum. The test material mixed with food in dose concentration 0, 5, 50, 150 or 500 mg/kg bw/day and 10 males and 20 females/group/generation were used .Pre-Mating Exposure / Males and Females were 60 days. Clinical observations were recorded twice daily with at least 5 hours between observations. Detailed physical examinations and palpation for masses were performed weekly. Body weights and food consumption were determined weekly for the first fourteen weeks, bi-weekly for the next 12 weeks and every 4 weeks thereafter until the end of the study. The intake of the test substance was determined from body weight, food consumption and dietary concentration. Haematology tests, including haemoglobin, haematocrit, erythrocyte and total and differential leukocyte counts, and erythrocyte morphology, were conducted on ten randomly selected animals at months 3, 6, 12, 18 and 24 of the study. Necropsies were conducted on all animals dying prior to study termination, killed in a moribund condition or killed on schedule. Histological examinations were conducted on all animals from both control groups, the highest dose group (2.0 or 5.0%) from each study and also on 10 rats randomly selected from each group for an interim sacrifice at 12 months. Histology was also performed on any animal with gross lesions or masses. Also tissues examined included adrenal glands, aorta, blood smear, brain, cecum, colon, duodenum, epididymus or uterus, esophagus, eyes, femur including marrow, tissue masses, gallbladder, heart, ileum, jejunum, duodenum, kidneys, liver, lungs and bronchi, mammary gland, nerves (sciatic), ovaries, lymph nodes, pancreas, parathyroids, pituitary gland, prostrate, rectum, skin, spleen, seminal vesicles, skeletal muscle, testes with epididymides, stomach, thymus, thyroid gland including parathyroid, trachea, urinary bladder, uterus.

 At the end of study, no treatment-related effects were reported on survival. No treatment-related changes were reported at gross necropsy. Histological evaluation revealed a variety of lesions, including neoplasms, present at similar incidences in control and treated animals. The authors considered the lesions to be spontaneous and not related to administration of the test material. There were no compound related effects on fertility, gestation, pup viability or lactation indices, on reproductive organs of females, or on organ weights among parents and offspring. There were no compound related lesions in any tissue examined histologically, including kidneys and adrenal glands from parental rats or from offspring. There was no toxicity to either the F1 or F2 generation. Food consumption was similar for control and treated animals at the lower dietary levels, but was slightly higher in the high-dose study, although not statistically significant. Haematological, clinical chemistry and urinalysis parameters did not differ significantly from the controls. Necropsies at one year did not reveal any treatment-related gross or microscopic changes. Hence NOAEL was considered to be 500mg/kg bw/day for F0, F1 and F2 generation. When male and female Charles River CD rats were treated with test material orally.

Study 2

A multigeneration reproductive toxicity study of test material was performed on male and female wistar rats. The test material mixed with feed in dose concentration 0, 0.1, 1.0 or 3% (0, 50, 500, 1500 mg/kg bw per day) and administered orally for three successive generations. Each generation having 60 animals of each sex in the control and 40 animals of each sex in test animals. No adverse effects were observed with respect to fertility, litter size and weight, general condition, male/female ratio, growth during lactation, survival or maturation. Autopsy of parent rats and pups at weaning did not reveal any treatment related changes in organ weights other than caecal enlargement in the 3% dose group. Gross and microscopic examination of the F3 generation at weaning did not reveal any abnormalities due to treatment and no adverse effects were seen in the teratology study. It was concluded that Brilliant Black PN did not exert any adverse effects on reproductive function of Wistar rats when fed at dietary levels up to 3% (1500 mg/kg bw per day) for three successive generations. Therefore, NOAEL was considered to be at 3% (1500 mg/kg bw per day) for F0 F1 and F2 generation .When male and female wistar rats were treated with test material orally.

 

Thus, Based on the data available form different studies , No Observed Adverse Effect Level (NOAEL) was considered to be above 500 -1500mg/kg bw . Thus, comparing this value with the criteria of CLP regulation test material is not likely to classify as reproductive toxicant

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

Thus, comparing this value with the criteria of CLP regulation test material is not likely to classify as reproductive toxicant.