Tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate

Endpoint summary

Description of key information

NOAEL was considered to be in the rang of 100-1300 mg/kg bw for tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate orally.

Thus, comparing this value with the criteria of CLP regulation tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (70161 -16 -9) is not likely to classify as repeated dose toxicant.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

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

Adequacy of study:

weight of evidence

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 and functionally similar read across chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Principles of method if other than guideline:

WoE derived based on the experimental data from structurally and functionally similar read across chemicals

GLP compliance:

not specified

Limit test:

no

Species:

other: 1, Mice, 2, pigs; 3 Rat

Strain:

other: 1. CFW, 2. Large White , 3. Osborne-Mendel

Sex:

male/female

Route of administration:

other: 1. feed, 2. feed, 3. Water

Vehicle:

other: 2. diet, 3. Distilled water

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

1. 80 weeks, 2. 90 days, 3. 20 days

Frequency of treatment:

Daily

Remarks:

1. 0.1, 0.25,0.5 or 1.0% (130,325,650,1300 mg/kg bw/d)

Remarks:

2. 0 (control), 100, 300 or 900 mg/kg/day

Remarks:

3. 0 and 0.2 % (260.2 mg/kg body weight/day )

No. of animals per sex per dose:

1. 30 male and 30 female mice2. 6 animal per sex per dose3. Totol: 550 mg/kg bw/day: 300 mg/kg bw/day: 25

Control animals:

yes, concurrent vehicle

Observations and examinations performed and frequency:

Study1:CAGE SIDE OBSERVATIONS: Yes - Time schedule: in groups of 15 in a roommaintained at 21 ± 1°C with a relative humidity of 50-60%- Cage side observations checked in table [No.?] were included.: No dataDETAILED CLINICAL OBSERVATIONS: No data- Time schedule: No dataBODY WEIGHT: Yes - Time schedule for examinations: start of the experiment, at wk 3 and then at intervals of 2 wk until wk 73 of the experimentFOOD 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: No data- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No dataFOOD EFFICIENCY:- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No dataWATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data- Time schedule for examinations: No dataOPHTHALMOSCOPIC EXAMINATION: No data - Time schedule for examinations: No data- Dose groups that were examined: No dataHAEMATOLOGY: Yes - Time schedule for collection of blood:At wk 28 and 55 from the caudal vein of ten males and ten females from the control group and from the groups of 0.5 and 1.0% dietary levels.At 80 wk, blood samples were collected from the aorta of all surviving mice during the autopsy.- Anaesthetic used for blood collection: No data- Animals fasted: No data- How many animals: 20 animals (10 male and 10 female)- Parameters were examined: counting the reticulocyte and leucocytes.CLINICAL CHEMISTRY: No data- Time schedule for collection of blood: No data- Animals fasted: No data- How many animals: No data- Parameters were examined: No dataURINALYSIS: Yes - Time schedule for collection of urine: At 28 wks at 6-hr period from three groups of five mice of each sex from the controls and the groups on the two highest dietary levels (0.5 and 1.0%) of Black PN.- Metabolism cages used for collection of urine: No data- Animals fasted: No data- Parameters were examined: reducing substances, bile salts and blood as well as for colour, pH and microscopic constituentsNEUROBEHAVIOURAL EXAMINATION: No data - Time schedule for examinations: No data- Dose groups that were examined: No data- Battery of functions tested: sensory activity / grip strength / motor activity / other: No dataStudy 2:CAGE SIDE OBSERVATIONS: No data available - Time schedule: No data available- Cage side observations checked in table [No.?] were included.: No data availableDETAILED CLINICAL OBSERVATIONS: Yes - Time schedule: No data availableBODY WEIGHT: Yes -Time schedule for examinations: WeaklyFOOD 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: No data available- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data availableFOOD EFFICIENCY:- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data available WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data available- Time schedule for examinations: No data availableOPHTHALMOSCOPIC EXAMINATION: No data available - Time schedule for examinations: No data available- Dose groups that were examined: No data availableHAEMATOLOGY: Yes - Time schedule for collection of blood: Haematological examinations were made at wk 6, using blood from an ear vein, and at wk 13 using blood collected from the anterior vena cava at autopsy. Haemoglobin concentrations, packed cell volumes and, in the terminal samples only, methaemoglobin concentrations were measured.- Anaesthetic used for blood collection: No data- Animals fasted: No data- How many animals: No data- Parameters examined: Counts were made of total erythrocytes, reticulocytes, erythrocytes containing Heinz bodies, and total and differential leucocytes.CLINICAL CHEMISTRY: Yes - Time schedule for collection of blood: No data available- Animals fasted: No data available - How many animals: No data available- Parameters examined: glutamic-oxalacetic and glutamic-pyruvic transaminases were estimated in blood collected at autopsy.URINALYSIS: Yes - Time schedule for collection of urine: 2-hr period during wk 6 and wk 13- Metabolism cages used for collection of urine: No data- Animals fasted: No data - Parameters examined: content of glucose, blood and bile were determined.NEUROBEHAVIOURAL EXAMINATION: No data available - Time schedule for examinations: No data available- Dose groups that were examined: No data available- Battery of functions tested: sensory activity / grip strength / motor activity / other: No data availableStudy 3:CAGE SIDE OBSERVATIONS: No data available - Time schedule: No data available- Cage side observations checked in table [No.?] were included. No data available DETAILED CLINICAL OBSERVATIONS: Yes - Time schedule: Daily BODY WEIGHT: Yes - Time schedule for examinations: Daily FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data available - Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data available - Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data available FOOD EFFICIENCY: No data available - Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data available WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes - Time schedule for examinations: Daily OPHTHALMOSCOPIC EXAMINATION: No data available - Time schedule for examinations: No data available - Dose groups that were examined: No data available HAEMATOLOGY: No data available - Time schedule for collection of blood: No data available - Anaesthetic used for blood collection: No data available - Animals fasted: No data available - How many animals: No data available - Parameters checked in table [No.?] were examined. No data available CLINICAL CHEMISTRY: No data available - Time schedule for collection of blood: No data available - Animals fasted: No data available - How many animals: No data available - Parameters checked in table [No.?] were examined. No data available URINALYSIS: No data available - Time schedule for collection of urine: No data available - Metabolism cages used for collection of urine: No data available - Animals fasted: No data available - Parameters checked in table [No.?] were examined. No data available NEUROBEHAVIOURAL EXAMINATION: No data available - Time schedule for examinations: No data available - Dose groups that were examined: No data available - Battery of functions tested: sensory activity / grip strength / motor activity / other: No data available OTHER: No data available

Sacrifice and pathology:

Study 1GROSS PATHOLOGY: no dataHISTOPATHOLOGY: YesStudy 2GROSS PATHOLOGY: Yes HISTOPATHOLOGY: Yes

Clinical signs:

no effects observed

Description (incidence and severity):

1. No effect on the condition or behavior of the animals. No effect on the condition or behavior of the animals. 2. Faeces of pigs fed Black PN were coloured black, but apart from this there were no abnormalities of appearance or behaviour.3. No external signs of toxicity and animals were appeared healthy and behaved normally in treated groups as compared to control. .

Mortality:

no mortality observed

Description (incidence):

1. There were no statistically significant differences between the number of deaths in the control mice and those given test chemical.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

1. The body weights of mice of both sexes were similar in all groups and No dose related effects on weight gain.2. No consistent or dose-related differences in body weight gain between the four groups3. No effect on maternal weight gain was observed in treated groups as compared to control.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

3. Daily consumption of test chemical as the 0.2 % solution was 260.2 mg/kg body weight.

Food efficiency:

not specified

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

no effects observed

Description (incidence and severity):

3. Significant decrease in water consumption was observed in treated rats as compare to control.

Haematological findings:

no effects observed

Description (incidence and severity):

2. Haematological examination after treatment for 13 week were similar in treated and control animals.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

1. There were no consistent or dose-related differences between treated and control groups.2. Haematological examination after treatment for 13 week were similar in treated and control animals.

Urinalysis findings:

no effects observed

Description (incidence and severity):

1. No abnormal constituents were detected in the urine from the control or treated mice.

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

1.Scattered differences in mean organ weights between treated and control animals.A lower brain weight in females, compared with the control value, was the only difference affecting animals fed 1% test chemical.The relative brain weight of females fed 0.25% was higher than the control figure. Liver weights of female but not of male mice fed 0.25% were lower than control values, but again this was an isolated finding and there were no significant differences in relative liver weights. Kidney weights of male animals only were significantly lower than control values at the two lowest levels of treatment (0.1 and 0.25%), but a significant difference in relative kidney weights of males occurred only at the 0.5% level, at which a higher value was recorded for the treated mice. The only other significant differences occurred in the relative heart weights, which were raised in male mice fed 0.5% and in females fed 0.25%.The changes in organ weight showed no dose related effects. Moreover the isolated changes seen at the lower levels were not found in both sexes and were not evident when expressed relative to body weight. It is considered that these random findings were not associated with treatment.2. There were no consistent or dose-related differences between treated and control groups.

Gross pathological findings:

no effects observed

Description (incidence and severity):

2. In one female at the 300 mg/kg/day dose level and in two of each sex at the highest dose level (900 mg/kg/day), hard black nodules, 1-2 mm in diameter, were present in the mucosa of the ileum.

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

1. Most of the tumours in the study occurred with either a comparable or a greater incidence in the control groups than in the treated mice. Several isolated tumours were identified in mice given the lower levels of test chemical, without comparable findings in the controls or in the highest dose group. Mammary fibroadenoma, a uterine fibromyoma, squamous-cell carcinoma were seen at different concentration.The frequency of histopathological findings in the treated animals did not differ significantly from those in the controls. Hence, no relationship was obvious between these findings and treatment with test chemical. Only in the case of adenomas of the lung and of the mammary tissue did more than one tumour of a given type occur among the treated animals of any group.

Histopathological findings: neoplastic:

not specified

Other effects:

no effects observed

Description (incidence and severity):

Estrous cycle:No significant changes in or treatment-related effects on the numbers of corpora lutea, implantations, early and late deaths and viable foetuses per litter or on the percentage of preimplantation loss were observed in treated group as compared to control. Reproductive performance:No significant effect on percentage of early resorptions and percentage of females with more than one and with more than two resorptions were observed in treated rats as compared to control.

Dose descriptor:

NOAEL

Effect level:

100 - 1 300 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

clinical signs

food consumption and compound intake

gross pathology

haematology

histopathology: non-neoplastic

mortality

organ weights and organ / body weight ratios

urinalysis

water consumption and compound intake

Remarks on result:

other: No effect 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

Conclusions:

NOAEL was considered to be in the rang of 100-1300 mg/kg bw for tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate orally.

Executive summary:

Repeated dose toxicity:

Data available for the read across chemicals was reviewed to determine the reproductive toxicity of tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (CAS no. 70161 -16 -9). The studies are as mentioned below:

Study 1:

Repeated dose toxicity test were performed on mice with test chemical in the concentrations of 0.1, 0.25, 0.5 or 1.0% (130,325,650,1300 mg/kg bw/d) for 80 wk. 30 males and 30 females was used for the treatment and group of 60 mice of each sex as control.

The general condition and behavior of the animals were observed frequently and any mouse that showed signs of ill-health was isolated, to be returned to its cage on recovery or to be killed if its condition deteriorated. The mice were weighed at the start of the experiment, at week 3 and then at intervals of 2 week until week 73 of the experiment. Blood sample were taken at week 28 and 55.At 80 week, blood samples were collected from the aorta of all surviving mice during the autopsy. For haematology blood samples were collected and for urine sample from 6-hr period from three groups of five mice of each sex from the controls and the groups on the two highest dietary levels of test chemical. Histopathology was also conducted.

There were no dose-related effects on body-weight gain, haematology or organ weights. The incidence of histopathological findings, including tumours, was not altered by the feeding of test chemical.

Therefore, the endpoint for the repeated dose toxicity was found to be NOAEL at 1 % (1300 mg/kg/day) concentration of test chemical to mice.

Study 2:

The repeated dose toxicity test was performed on pigs treated with test chemical at different concentration as 0 (control), 100, 300 and 900 mg/kg/day. Three litters each containing four male and four female 10-week-old Large White pigs from a minimal disease herd were used with diet Hi-lean Rearers Pencils fed in quantities of 0.5-1.5 lb/day according to the age of the animals. Water was available ad lib.

The pigs were weighed weekly. Haematological examinations, urine analysis, clinical chemistry, gross and histopathology were examined. There were no adverse effects on growth, haematology, urine analyses, serum transaminase activities or organ weights. Cysts containing mucus and fibrin were found in the mueosa of the ileum of four of the six pigs given the highest level of treatment and one of six given 300 mg/kg/day.

Therefore, theend point for the repeated dose toxicity was found to be NOEL at concentration 100 mg/kg/day when treated to pigs with test chemical.

Study 3:

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening, Osborne-Mendel female rats were treated with test chemical in the concentration of 260.2 mg/kg body weight /day in distilled water by oral drinking water. No external signs of toxicity and animals were appeared healthy and behaved normally in treated groups as compared to control. No effects on maternal weight gain and significant decrease in water consumption were observed in treated female rats as compared to control. No significant effect on numbers of corpora lutea, implantations, early and late deaths and viable foetuses per litter or on the percentage of preimplantation loss was observed in treated group as compared to control. Similarly, no significant effect on percentage of early resorptions and percentage of females with more than one and with more than two resorptions were observed in treated rats as compared to control. In addition, No effect on live or dead foetuses and mean foetal body weight and percentage of males and females per treatment group were observed as compared to control. No external variations were observed in litters of treated dams as compared to control. No effect on Crown-rump length and Sex distribution were observed in litters of treated dams as compared to control. Slight increase in number of affected litters, number of bipartite sternebrae and reduced ossification of the parietal bone were observed in litters of treated dams as compared to control. But, the observed increases did not appear to be dose-related. Therefore, NOAEL was considered to be 260.2 mg/kg body weight /day when Osborne-Mendel female rats were treated with test chemical orally in Drinking water for 20 days.

Thus, Based on the data available for the read across chemical, No Observed Adverse Effect Level (NOAEL) was considered to be above 100 -1300 mg/kg bw . Thus, comparing this value with the criteria of CLP regulation tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (70161 -16 -9) is not likely to classify as repeated dose toxicant.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 300 mg/kg bw/day

Study duration:

subchronic

Species:

mouse

Quality of whole database:

Data is Klimicsh 2 and from secondary source

Additional information

Repeated dose toxicity:

Data available for the read across chemicals was reviewed to determine the reproductive toxicity of tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (CAS no. 70161 -16 -9). The studies are as mentioned below:

Study 1:

Repeated dose toxicity test were performed on mice with test chemical in the concentrations of 0.1, 0.25, 0.5 or 1.0% (130,325,650,1300 mg/kg bw/d) for 80 wk. 30 males and 30 females was used for the treatment and group of 60 mice of each sex as control.

The general condition and behavior of the animals were observed frequently and any mouse that showed signs of ill-health was isolated, to be returned to its cage on recovery or to be killed if its condition deteriorated. The mice were weighed at the start of the experiment, at week 3 and then at intervals of 2 week until week 73 of the experiment. Blood sample were taken at week 28 and 55.At 80 week, blood samples were collected from the aorta of all surviving mice during the autopsy. For haematology blood samples were collected and for urine sample from 6-hr period from three groups of five mice of each sex from the controls and the groups on the two highest dietary levels of test chemical. Histopathology was also conducted.

There were no dose-related effects on body-weight gain, haematology or organ weights. The incidence of histopathological findings, including tumours, was not altered by the feeding of test chemical.

Therefore, the endpoint for the repeated dose toxicity was found to be NOAEL at 1 % (1300 mg/kg/day) concentration of test chemical to mice.

Study 2:

The repeated dose toxicity test was performed on pigs treated with test chemical at different concentration as 0 (control), 100, 300 and 900 mg/kg/day. Three litters each containing four male and four female 10-week-old Large White pigs from a minimal disease herd were used with diet Hi-lean Rearers Pencils fed in quantities of 0.5-1.5 lb/day according to the age of the animals. Water was available ad lib.

The pigs were weighed weekly. Haematological examinations, urine analysis, clinical chemistry, gross and histopathology were examined. There were no adverse effects on growth, haematology, urine analyses, serum transaminase activities or organ weights. Cysts containing mucus and fibrin were found in the mueosa of the ileum of four of the six pigs given the highest level of treatment and one of six given 300 mg/kg/day.

Therefore, theend point for the repeated dose toxicity was found to be NOEL at concentration 100 mg/kg/day when treated to pigs with test chemical.

Study 3:

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening, Osborne-Mendel female rats were treated with test chemical in the concentration of 260.2 mg/kg body weight /day in distilled water by oral drinking water. No external signs of toxicity and animals were appeared healthy and behaved normally in treated groups as compared to control. No effects on maternal weight gain and significant decrease in water consumption were observed in treated female rats as compared to control. No significant effect on numbers of corpora lutea, implantations, early and late deaths and viable foetuses per litter or on the percentage of preimplantation loss was observed in treated group as compared to control. Similarly, no significant effect on percentage of early resorptions and percentage of females with more than one and with more than two resorptions were observed in treated rats as compared to control. In addition, No effect on live or dead foetuses and mean foetal body weight and percentage of males and females per treatment group were observed as compared to control. No external variations were observed in litters of treated dams as compared to control. No effect on Crown-rump length and Sex distribution were observed in litters of treated dams as compared to control. Slight increase in number of affected litters, number of bipartite sternebrae and reduced ossification of the parietal bone were observed in litters of treated dams as compared to control. But, the observed increases did not appear to be dose-related. Therefore, NOAEL was considered to be 260.2 mg/kg body weight /day when Osborne-Mendel female rats were treated with test chemical orally in Drinking water for 20 days.

Thus, Based on the data available for the read across chemical, No Observed Adverse Effect Level (NOAEL) was considered to be above 100 -1300 mg/kg bw . Thus, comparing this value with the criteria of CLP regulation tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (70161 -16 -9) is not likely to classify as repeated dose toxicant.

Justification for classification or non-classification

Based on the data available for the read across chemical, No Observed Adverse Effect Level (NOAEL) was considered to be above 100 -1300 mg/kg bw . Thus, comparing this value with the criteria of CLP regulation tetrasodium 2-[[4-[[4-[(4-amino-6-chloro-1,3,5-triazin-2-yl)amino]-5-sulphonato-1-naphthyl]azo]-7-sulphonato-1-naphthyl]azo]benzene-1,4-disulphonate (70161 -16 -9) is not likely to classify as repeated dose toxicant.