Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl]amino]-2,4,6-trimethyl-5-sulphonatophenyl]amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate

Endpoint summary

Description of key information

Repeated dose toxicity: Oral

The no observed adverse effect level (NOAEL) for Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl]amino] -2,4,6- trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate is considered to be 1300 mg/kg/day. Hence the test chemical is not likely to be toxic as per the criteria mentioned in CLP regulation.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

repeated dose toxicity: oral

Remarks:

combined repeated dose and carcinogenicity

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 similar read across chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

WoE derived based on the experimental data from read across from four similar read across chemicals

GLP compliance:

not specified

Limit test:

no

Species:

mouse

Strain:

other: 1. CFW strain, 2. Charles River CD

Details on species / strain selection:

No data

Sex:

male/female

Details on test animals or test system and environmental conditions:

1. TEST ANIMALS- Source: From a specified-pathogen-free colony- Age at study initiation: No data available- Weight at study initiation: The mice were weighed at the start of the experiment (exact weight not mentioned)- Fasting period before study: No - Housing: They were caged in groups of 15 in a room- Diet (e.g. ad libitum): Oxoid pasteurized breeding diet,ad libitum- Water (e.g. ad libitum): ad libitum- Acclimation period: No data availableENVIRONMENTAL CONDITIONS- Temperature (°C): 21±1°C- Humidity (%): 50-60%- Air changes (per hr): No data available- Photoperiod (hrs dark / hrs light): No data available2. TEST ANIMALS- Source: SPF breeding colony- Weight at study initiation: Male- 21-30 g and female - 17-25 g- Fasting period before study: no - Housing: housed in cages of 15- Diet (e.g. ad libitum): ground Oxoid pasteurized breeding diet; ad libitum- Water (e.g. ad libitum): water ad libitumENVIRONMENTAL CONDITIONS- Temperature (°C): 21± 1°C- Humidity (%):50-60%

Route of administration:

oral: feed

Details on route of administration:

No data

Vehicle:

other: 1. No data

Details on oral exposure:

1. PREPARATION OF DOSING SOLUTIONS: The test chemical was mixed with feed at dose level of 0, 0.1, 0.25, 0.5 or 1.0% (0, 130, 325, 650, 1300 mg/kg bw/d) DIET PREPARATION - Rate of preparation of diet (frequency): No data - Mixing appropriate amounts with (Type of food): No data - Storage temperature of food: No data VEHICLE - Justification for use and choice of vehicle (if other than water): Oxoid pasteurized breeding diet - Concentration in vehicle: 0, 0.1, 0.25, 0.5 or 1.0% (0, 130, 325, 650, 1300 mg/kg bw/d) - Amount of vehicle (if gavage): No data - Lot/batch no. (if required): No data - Purity: No data2. PREPARATION OF DOSING SOLUTIONS: The test chemical was mixed with feed at dose levels of 0, 100, 200, 400, 800 mg/l (0, 0.2, 0.4, 0.8 or 1.6%) DIET PREPARATION - Rate of preparation of diet (frequency): No data - Mixing appropriate amounts with (Type of food): Oxoid pasteurized breeding diet - Storage temperature of food: No data VEHICLE - Justification for use and choice of vehicle (if other than water): Oxoid pasteurized breeding diet - Concentration in vehicle: 0, 100, 200, 400, 800 mg/l (0, 0.2, 0.4, 0.8 or 1.6%) - Amount of vehicle (if gavage): No data - Lot/batch no. (if required): No data - Purity: No data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

1. No data

Duration of treatment / exposure:

1. 80 wk2. 80 weeks

Frequency of treatment:

1. Daily2. 1 wk before mating, throughout the 3-wk breeding period, and during the gestation and lactation periods3. Daily

Remarks:

0, 0.1, 0.25, 0.5 or 1.0% (0, 130, 325, 650, 1300 mg/kg bw/d) / 1

Remarks:

Doses / Concentrations:0, 100, 200, 400, 800 mg/l (0, 0.2, 0.4, 0.8 or 1.6%) / 2Basis:nominal in diet

No. of animals per sex per dose:

1. Total: 180 male and 180 female mice0 mg/kg bw/d: 60 male and 60 female mice130 mg/kg bw/d: 30 male and 30 female mice325 mg/kg bw/d: 30 male and 30 female mice650 mg/kg bw/d: 30 male and 30 female mice1300 mg/kg bw/d: 30 male and 30 female mice2. Control group:60 male and 60 femalesTest group: 100 mg/Kg: 30 male and 30 females200 mg/Kg: 30 male and 30 females400 mg/Kg: 30 male and 30 females800 mg/Kg: 30 male and 30 females

Control animals:

yes, concurrent vehicle

Details on study design:

1. No data available

Positive control:

1. No data available

Observations and examinations performed and frequency:

1. CAGE SIDE OBSERVATIONS: Yes - Time schedule: Frequently dring the study period- Cage side observations checked in table [No.?] were included.: general condition and behaviour, the animals were also observed for ill healthDETAILED 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: haemoglobin concentration and packed cell volume, as well as for counts of erythrocytes and leucocytes. In addition, the methaemoglobin concentrations were determined in the samples collected at 80 wk. 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: protein, 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 data2. CAGE SIDE OBSERVATIONS: Yes- Time schedule: Continued surveillance- Cage side observations checked in table [No.?] were included. Any abnormalities in condition and behaviorDETAILED CLINICAL OBSERVATIONS: No data- Time schedule: No dataBODY WEIGHT: Yes- Time schedule for examinations: At 4 weeks interval throughout the studyFOOD 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: wk 13, 26 and 52 and from all surviving mice at wk 80- Anaesthetic used for blood collection: No data- Animals fasted: No data- How many animals: 10/sex from control group and animals fed 400 and 800 mg/Kg- Parameters checked in table [No.?] were examined. Haemoglobin concentration and packed cell volume and counts were made of erythrocytes and total leucocytes. Differential leucocyte counts and reticulocyte counts were made on the blood from the mice fed on the control diet and from those given diet containing 1.6% test chemical at wk 26, 52 and 80, and reticulocyte counts were also carried out for these groups at wk 13.CLINICAL CHEMISTRY: No data- Time schedule for collection of blood: No data- Animals fasted: No data - How many animals: No data- Parameters checked in table [No.?] were examined. No dataURINALYSIS: No data- Time schedule for collection of urine: No data- Metabolism cages used for collection of urine: No data- Animals fasted: No data- Parameters checked in table [No.?] were examined. No dataNEUROBEHAVIOURAL 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 dataIMMUNOLOGY: Yes / No / Not specified- Time schedule for examinations:- How many animals:- Dose groups that were examined:- Parameters checked in table [No.?] were examined.OTHER:

Sacrifice and pathology:

1. GROSS PATHOLOGY: Yes, The animals were killed by exsanguination from the aorta under sodium pentobarbitone anaesthesia following an overnight period without food. At autopsy, macroscopic abnormalities were recorded and the brain, heart, liver, spleen, kidneys, adrenal glands and gonads were weighed.HISTOPATHOLOGY: Yes, samples of the brain, heart, liver, spleen, kidneys, adrenal glands and gonads and of salivary glands, pituitary, thyroid, thymus, various lymph nodes, pancreas, urinary bladder, lungs, stomach, duodenum, ileum, colon, caecum, rectum, striped muscle (hind limb), spinal cord, uterus, aortic arch and any other tissue that appeared abnormal were preserved in 10% buffered formalin. Paraffin-wax sections of these tissues were stained with haematoxylin and eosin. All tissues from the control mice and from those fed diet containing 1% Black PN were examined histologically. At the lower dose levels, the examination was confined to the liver, kidney and any tissues seen to be abnormal at autopsy.2. GROSS PATHOLOGY: Yes, At autopsy, all macroscopic abnormalities were noted and the brain, heart, liver, kidneys, spleen, stomach, small intestine, caecum and testes were weighed.HISTOPATHOLOGY: Yes, the brain, heart, liver, kidneys, spleen, stomach, small intestine, caecum and testes together with salivary gland, thyroid, adrenals, lymphnodes, pancreas, pituitary, ovaries, uterus, urinary bladder, lungs, colon, rectum, spinal cord, skeletal muscle and any other tissue that appeared abnormal were preserved in 10% buffered formalin. Paraffin-wax sections of these tissues were stained with haemotoxylin and eosin. All tissues from the control mice and those fed 800 mg/Kg the test chemical were examined microscopically, while at the lower dietary levels examination was confined to the liver and kidney together with any tissue seen to be abnormal at autopsy.

Other examinations:

1. No data2. Behavior and were weighed at 4-wk intervals throughout the study. During the first half of the study it was noticed that there was a tendency for the male mice to fight. Bite lesions of the anogenital region were particularly frequent and these were associated with obstructions of the urinary tract. To avoid further fighting, all the mice were caged individually from month 8.

Statistics:

1. chi-square test, Student's t test.2. No data

Clinical signs:

no effects observed

Description (incidence and severity):

1. The ingestion of Black PN had no effect on the condition or behaviour of the animals.

Mortality:

no mortality observed

Description (incidence):

1. No statistically significant differences between the number of deaths in the control mice and those given the test chemical were noted.2. There were deaths in all groups during the study and there was no relationship between the number of deaths at any time and the dietary intake of the test chemical

Body weight and weight changes:

no effects observed

Description (incidence and severity):

1. Throughout the study the body weights of mice of both sexes were similar in all groups2. The terminal body weights and the body-weight gains were similar in all groups.

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:

effects observed, treatment-related

Description (incidence and severity):

1. The haematological examinations revealed only inconsistent isolated changes of statistical significance. At wk 28 the haemoglobin concentrations and red blood cell counts were lower in female mice fed diet containing if 0.5 or 1.0% than in the controls. There were no comparable findings in the males. The total white cell count of the male animals fed 1.0% in the diet was higher than that of the controls at this time, but there was no comparable change in this measurement in the females, or in either sex at any other time. At wk 55, the haemoglobin concentrations of male animals fed 0.5% of the coiouring in the diet were significantly (P < 0.05) lower than the control values. However, the corresponding value at the higher dietary level was not affected and there were no differences from the control value in the females. Also at wk 55, the erythrocyte counts of females fed 1.0% were lower (P < 0.01) than those of the controls, but this finding was again isolated. There were no statistically significant differences between the control and test samples taken at wk 80.2. No evidence of any haematological adverse effect due to the administration of the test chemical

Clinical biochemistry findings:

not specified

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:

effects observed, treatment-related

Description (incidence and severity):

1. There were only 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%. This difference, which did not occur in the males, was only marginally significant and there was no significant difference when the weights were expressed relative to body weight By contrast, 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%.2. No significant differences between the organ weights or relative organ weights of the test groups and the corresponding controls.

Gross pathological findings:

not specified

Description (incidence and severity):

2. Distension of the bladder, frequently associated with hydroureter and hydronephosis was noted. Pus and hard protein deposits were found in the bladders of many of the mice and in a number of cases it was possible to show that an obstruction had occurred in the urethra in the region of the bulbo-urethral complex. After the male mice were put into individual cages, there was a marked reduction in the number showing this syndrome, although isolated cases of urinary retention were seen throughout the study.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

1. The incidence of histological findings was similar in all groups of mice, including the controls 2. No differences between treated and control mice in the incidence or severity of the lesions seen. There as an unusually high incidence of pyelonephritis and chronic inflammation of the bladder and urethra, but these lesions occurred mainly in the male mice killed because of urinary retention in the early stages of the study.

Histopathological findings: neoplastic:

not specified

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 the test chemical, without comparable findings in the controls or in the highest dose group. They were a mammary fibroadenoma (in a female on 0.1%), a uterine fibromyoma (0.19%) and a squamous-cell carcinoma of the skin (female, 0.5%). The only tumour found at the highest dietary level without comparable control findings was a squamous- cell carcinoma of the skin in a male mouse fed 1%.2. Of the malignant tumours found, lymphomas and reticulum-cell neoplasms occurred in treated and control mice. In addition a splenic haemangiosarcoma and a mammary carcinoma were found in the female mice fed on the diet containing the lowest level (100 mg/Kg). A single tumour of the Harderian gland was found in a female from the group given the 1.6% level. The commonest benign tumours were pulmonary adenomas and benign cysts in the ovary, found with a similar frequency in treated and control mice. The only other tumour occurring in treated mice without a similar finding in the controls was a granulosa-cell tumour of the ovary in a mouse given 800 mg/Kg test chemical

Other effects:

not specified

Dose descriptor:

NOAEL

Remarks:

1

Effect level:

1 300 other: mg/kg/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Body weight, weight gain, organ weight and histopathology.

Dose descriptor:

NOAEL

Remarks:

2

Effect level:

800 mg/kg diet

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No significant effects were noted at the mentioned dose level

Critical effects observed:

not specified

Conclusions:

The no observed adverse effect level (NOAEL) for Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl]amino]-2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate is considered to be 1300 mg/kg/day.

Executive summary:

Repeated dose oral toxicity studies were reviewed to determine the toxic nature of Trisodium 1-amino-4-[[3-[ [4-chloro-6-[(sulphonatophenyl)amino]- 1,3,5-triazin-2-yl] amino]-2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate. The studies are as summarized below:

Study 1:

Repeated dose toxicity test were performed on mice with different concentrations of the test chemical at dose levels 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. During the study period the animals were observed for clinical signs, mortality, hematology, urine analysis was performed and the animals were subjected to gross and histopathology. The general condition and behaviour 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 wk 3 and then at intervals of 2 wk until wk 73 of the experiment. Blood sample were taken at wk 28 and 55. At 80 wk, 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 Black PN. Histopathology was also conducted. The ingestion of the test chemical had no effect on the condition or behaviour and mortality of the animals. The haematological examinations revealed only inconsistent isolated changes of statistical significance. At wk 28 the haemoglobin concentrations and red blood cell counts were lower in female mice fed diet containing if 0.5 or 1.0% test chemical than in the controls. There were no comparable findings in the males. The total white cell count of the male animals fed 1.0% test chemical in the diet was higher than that of the controls at this time, but there was no comparable change in this measurement in the females, or in either sex at any other time. At wk 55, the haemoglobin concentrations of male animals fed 0.5% of the coiouring in the diet were significantly (P < 0.05) lower than the control values. However, the corresponding value at the higher dietary level was not affected and there were no differences from the control value in the females. Also at wk 55, the erythrocyte counts of females fed 1.0% test chemical were lower (P < 0.01) than those of the controls, but this finding was again isolated. There were no statistically significant differences between the control and test samples taken at wk 80. There were only 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% the test chemical. This difference, which did not occur in the males, was only marginally significant and there was no significant difference when the weights were expressed relative to body weight By contrast, the relative brain weight of females fed 0.25% Black PN was higher than the control figure. Liver weights of female but not of male mice fed 0.25% test chemical 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% test chemical. The incidence of histological findings was similar in all groups of mice, including the controls. 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, without comparable findings in the controls or in the highest dose group. They were a mammary fibroadenoma (in a female on 0.1%), a uterine fibromyoma (0.19%) and a squamous-cell carcinoma of the skin (female, 0.5%). The only tumour found at the highest dietary level without comparable control findings was a squamous- cell carcinoma of the skin in a male mouse fed 1%. Based on these considerations, the no observed adverse effect level (NOAEL) for the test chemical in mice is considered to be 1 % (1300 mg/kg/day).

Study 2:

Repeated oral toxicity of the test chemical was determined by performing a 80 weeks repeated dose toxicity study using Charles River CD mouse (male and female) at dose levels of 0, 100, 200, 400 or 800 mg/Kg (0.2, 0.4, 0.8 or 1.6% respectively) by oral diet route. The animals were observed for clinical signs, mortality, body weight changes, hematology and were subjected to gross and histopathology. The feeding of the test chemical did not adversely affect the death rate within the groups, the rate of body-weight gain, the organ weights or the haematological findings. The incidence and severity of the histopathological findings were similar in treated and control mice and there was no evidence of an increased incidence of tumours in the mice given Sunset Yellow FCF. Distension of the bladder, frequently associated with hydroureter and hydronephosis was noted. Pus and hard protein deposits were found in the bladders of many of the mice and in a number of cases it was possible to show that an obstruction had occurred in the urethra in the region of the bulbo-urethral complex. After the male mice were put into individual cages, there was a marked reduction in the number showing this syndrome, although isolated cases of urinary retention were seen throughout the study. Of the malignant tumours found, lymphomas and reticulum-cell neoplasms occurred in treated and control mice. In addition a splenic haemangiosarcoma and a mammary carcinoma were found in the female mice fed on the diet containing the lowest level (100 mg/Kg). A single tumour of the Harderian gland was found in a female from the group given the 1.6% level. The commonest benign tumours were pulmonary adenomas and benign cysts in the ovary, found with a similar frequency in treated and control mice. The only other tumour occurring in treated mice without a similar finding in the controls was a granulosa-cell tumour of the ovary in a mouse given 800 mg/Kg test chemical. Thus, on the basis of above results the NOAEL (no observed adverse effect level) for the test chemical was considered to be 800 mg/kg diet.

The no observed adverse effect level (NOAEL) for Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl] amino]- 2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate is considered to be 1300 mg/kg/day.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 300 mg/kg bw/day

Study duration:

chronic

Species:

mouse

Quality of whole database:

Data is from K2 publication

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Repeated dose toxicity: Oral

Repeated dose oral toxicity studies were reviewed to determine the toxic nature of Trisodium 1-amino-4-[[3-[ [4-chloro-6-[(sulphonatophenyl)amino]- 1,3,5-triazin-2-yl] amino]-2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate (CAS no 72214 -18 -7). The studies are as summarized below:

Study 1:

Repeated dose toxicity test were performed on mice with different concentrations of the test chemical at dose levels 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. During the study period the animals were observed for clinical signs, mortality, hematology, urine analysis was performed and the animals were subjected to gross and histopathology. The general condition and behaviour 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 wk 3 and then at intervals of 2 wk until wk 73 of the experiment. Blood sample were taken at wk 28 and 55. At 80 wk, 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 Black PN. Histopathology was also conducted. The ingestion of the test chemical had no effect on the condition or behaviour and mortality of the animals. The haematological examinations revealed only inconsistent isolated changes of statistical significance. At wk 28 the haemoglobin concentrations and red blood cell counts were lower in female mice fed diet containing if 0.5 or 1.0% test chemical than in the controls. There were no comparable findings in the males. The total white cell count of the male animals fed 1.0% test chemical in the diet was higher than that of the controls at this time, but there was no comparable change in this measurement in the females, or in either sex at any other time. At wk 55, the haemoglobin concentrations of male animals fed 0.5% of the coiouring in the diet were significantly (P < 0.05) lower than the control values. However, the corresponding value at the higher dietary level was not affected and there were no differences from the control value in the females. Also at wk 55, the erythrocyte counts of females fed 1.0% test chemical were lower (P < 0.01) than those of the controls, but this finding was again isolated. There were no statistically significant differences between the control and test samples taken at wk 80. There were only 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% the test chemical. This difference, which did not occur in the males, was only marginally significant and there was no significant difference when the weights were expressed relative to body weight By contrast, the relative brain weight of females fed 0.25% Black PN was higher than the control figure. Liver weights of female but not of male mice fed 0.25% test chemical 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% test chemical. The incidence of histological findings was similar in all groups of mice, including the controls. 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, without comparable findings in the controls or in the highest dose group. They were a mammary fibroadenoma (in a female on 0.1%), a uterine fibromyoma (0.19%) and a squamous-cell carcinoma of the skin (female, 0.5%). The only tumour found at the highest dietary level without comparable control findings was a squamous- cell carcinoma of the skin in a male mouse fed 1%. Based on these considerations, the no observed adverse effect level (NOAEL) for the test chemical in mice is considered to be 1 % (1300 mg/kg/day).

Study 2:

Repeated oral toxicity of the test chemical was determined by performing a 80 weeks repeated dose toxicity study using Charles River CD mouse (male and female) at dose levels of 0, 100, 200, 400 or 800 mg/Kg (0.2, 0.4, 0.8 or 1.6% respectively) by oral diet route. The animals were observed for clinical signs, mortality, body weight changes, hematology and were subjected to gross and histopathology. The feeding of the test chemical did not adversely affect the death rate within the groups, the rate of body-weight gain, the organ weights or the haematological findings. The incidence and severity of the histopathological findings were similar in treated and control mice and there was no evidence of an increased incidence of tumours in the mice given Sunset Yellow FCF. Distension of the bladder, frequently associated with hydroureter and hydronephosis was noted. Pus and hard protein deposits were found in the bladders of many of the mice and in a number of cases it was possible to show that an obstruction had occurred in the urethra in the region of the bulbo-urethral complex. After the male mice were put into individual cages, there was a marked reduction in the number showing this syndrome, although isolated cases of urinary retention were seen throughout the study. Of the malignant tumours found, lymphomas and reticulum-cell neoplasms occurred in treated and control mice. In addition a splenic haemangiosarcoma and a mammary carcinoma were found in the female mice fed on the diet containing the lowest level (100 mg/Kg). A single tumour of the Harderian gland was found in a female from the group given the 1.6% level. The commonest benign tumours were pulmonary adenomas and benign cysts in the ovary, found with a similar frequency in treated and control mice. The only other tumour occurring in treated mice without a similar finding in the controls was a granulosa-cell tumour of the ovary in a mouse given 800 mg/Kg test chemical. Thus, on the basis of above results the NOAEL (no observed adverse effect level) for the test chemical was considered to be 800 mg/kg diet.

The no observed adverse effect level (NOAEL) for Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl] amino]- 2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate is considered to be 1300 mg/kg/day.

Justification for classification or non-classification

Based on the data available for the test chemicals, Trisodium 1-amino-4-[[3-[[4-chloro-6-[(sulphonatophenyl)amino]-1,3,5-triazin-2-yl] amino]-2,4,6-trimethyl-5-sulphonatophenyl] amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate (CAS no 72214 -18 -7) is not likely to be toxic as per the criteria mentioned in CLP regulation.