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EC number: 263-974-8 | CAS number: 63157-72-2
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Repeated dose toxicity: oral
Administrative data
- Endpoint:
- short-term repeated dose toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 010
- Report date:
- 2010
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
Test material
- Reference substance name:
- Tris[4-(diethylamino)phenyl]methylium acetate
- EC Number:
- 263-974-8
- EC Name:
- Tris[4-(diethylamino)phenyl]methylium acetate
- Cas Number:
- 63157-72-2
- Molecular formula:
- C31H42N3.C2H3O2
- IUPAC Name:
- tris[4-(diethylamino)phenyl]methylium acetate
- Details on test material:
- - Name of test material: Ethylviolet Acetate
- Purity: 94.6 g/100 g
- Test substance no.: 09/0236-001
- Batch identification: GV-35268/084
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Wistar
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Sulzfeld, Germany
- Age at study initiation: 41-43 days
- Housing: 5 animals per cage in H-Temp polysulfonate cages supplied by TECNIPLAST, Hohenpeißenberg, Germany (floor area about 2065 cm2). Bedding in the cages were Type Lignocel PS 14, dust-free bedding, supplied by SSNIFF, Soest, Germany.
- Diet: ground Kliba maintenance diet mouse/rat “GLP” meal available ad libitum, supplied by Provimi Kliba SA, Kaiseraugst, Switzerland
- Water: ad libitum
- Acclimation period: 9 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 / 12
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- other: Drinking water containing 1% Carboxylmethylcellulose and Cremophor EL® (5 mg/100 mL)
- Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS:
The test substance was applied as a suspension. To prepare this suspension, an aliquot of test substance was pestled and 10% of drinking water was filled up into an Erlenmeyer flask. The pestled test substance was weighed out into the drinking water and mixed by high speed with an Ultraturrax (maximum 24,000 rpm). Then 1% Carboxylmethylcellulose in drinking water with Cremophor EL (5 mg/100 mL) was filled up to the desired volume. During administration of the test substance, preparations were kept homogeneous by stirring with a magnetic stirrer. The test-substance preparations were produced at least twice weekly. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- The analyses of the test-substance preparations were carried out at the Competence Center Analytics, BASF SE as a separate GLP study under the responsibility of a study director of this testing facility.
The stability of the test substance in drinking water containing 1% Carboxylmethylcellulose and Cremophor EL® (5 mg/100 mL) at room temperature for a period of 4 days was proven before the start of the administration period.
Homogeneity and concentration control analyses of the test-substance preparations were performed in samples of all concentrations at the start of the administration period and on study days 7, 11, and 14. - Duration of treatment / exposure:
- 29 days
- Frequency of treatment:
- Once daily
Doses / concentrations
- Remarks:
- Doses / Concentrations:
10, 30 (and 20), 100 (and 60) mg/kg bw/d
Basis:
actual ingested
- No. of animals per sex per dose:
- 5
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- Due to severe clinical signs and premature deaths, the dose level for test group 3 was decreased from 100 to 60 mg/kg bw/d from study day 4 onwards and for test group 2 from 30 to 20 mg/kg bw/d from study day 12 onwards.
- Positive control:
- None
Examinations
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS:
- Time schedule: Twice daily on working days and once daily on Saturdays, Sundays and public holidays
CLINICAL OBSERVATIONS:
- Time schedule: Daily before the administration as well as within 2 hours and within 5 hours after the administration
DETAILED CLINICAL OBSERVATIONS:
- Time schedule: Prior to the administration period and thereafter at weekly intervals
BODY WEIGHT:
- Time schedule for examinations: Before the start of the administration period, on day 0 (start of the administration period), study day 5 and thereafter at weekly intervals
FOOD CONSUMPTION:
- Time schedule for examinations: Weekly over a period of 1 day and calculated as mean food consumption grams per animal and day
WATER CONSUMPTION:
- Time schedule for examinations: Daily visual inspection of the water bottles for any changes in volume
HAEMATOLOGY:
- Time schedule for collection of blood: The morning after the last exposure
- Anaesthetic used for blood collection: Yes (isoflurane (Isoba®, Essex GmbH Munich, Germany))
- Animals fasted: Yes
- How many animals: 5 per test group and sex
- Parameters examined: Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration (MCHC), Platelet count (PLT), Differential blood count, Reticulocytes, Prothrombin time (Hepato Quick’s test; HQT). Blood smears were prepared and stained according to WRIGHT without being evaluated, because of non-ambiguous results of the differential blood cell counts measured by the automated instrument.
CLINICAL CHEMISTRY:
- Time schedule for collection of blood: The morning after the last exposure
- Animals fasted: Yes
- How many animals: 5 per test group and sex
- Parameters examined: Alanine aminotransferase (ALT; L-alanine: 2-oxoglutarate aminotransferase), Aspartate aminotransferase (AST; (L-aspartate: 2-oxoglutarate aminotransferase), Alkaline phosphatase (ALP; orthophosphoric acid monoester phosphohydrolase), γ-Glutamyltransferase (GGT; γ-glutamyl) peptide: aminoacid-γ-glutamyl-transferase), Sodium (NA), Potassium (K), Chloride (CL), Inorganic phosphate (INP), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Albumin (ALB), Globulins (GLOB), Triglycerides (TRIG), Cholesterol (CHOL), Magnesium (MG), Bile acids (BILE).
URINALYSIS:
- Time schedule for collection of urine: Overnight after the last exposure
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters examined: pH, Protein, Glucose, Ketones, Urobilinogen, Bilirubin, Blood, Specific gravity, Sediment, Color (turbidity), Volume.
NEUROBEHAVIOURAL EXAMINATION:
- Time schedule for examinations: At the end of the administration period
- Dose groups that were examined: All animals
- Battery of functions tested: Sensory activity, grip strength, motor activity - Sacrifice and pathology:
- ORGAN WEIGHTS
The following weights were determined: Anesthetized animals, Adrenal glands, Brain, Epididymides, Heart, Kidneys, Liver, Ovaries, Prostate, Seminal vesicles with coagulation glands, Spleen, Testes, Thymus, Thyroid glands, Uterus with cervix.
ORGAN/TISSUE FIXATION
The following organs/tissues were preserved in neutral buffered 4% formaldehyde or modified Davidson’s solution: All gross lesions, Adrenal glands, Aorta, Bone marrow (femur), Brain, Cecum, Cervix, Colon, Duodenum, Epididymides, Esophagus, Extraorbital lacrimal glands, Eyes with optic nerve, Femur with knee joint, Heart, Ileum, Jejunum, Kidneys, Larynx, Liver, Lungs, Lymph nodes (mesenteric and axillary lymph nodes), Mammary gland (males and females), Nose (nasal cavity), Ovaries, Oviducts, Pancreas, Parathyroid glands, Pharynx, Pituitary gland, Prostate, Rectum, Salivary glands (mandibular and sublingual glands), Seminal vesicle with coagulation glands, Sciatic nerve, Skeletal muscle, Skin, Spinal cord (cervical, thoracic and lumbar cords), Spleen, Sternum with marrow, Stomach (forestomach and glandular stomach), Testes (modified Davidson’s solution), Thymus, Thyroid glands, Trachea, Urinary bladder, Uterus, Vagina.
HISTOPATHOLOGY
The following organs were examined: All gross lesions, Adrenal glands, Bone marrow (femur), Brain, Cecum, Cervix, Colon, Duodenum, Epididymides, Eyes with optic nerve, Heart, Ileum, Jejunum (with Peyer’s patches), Kidneys, Liver, Lung, Lymph nodes (mesenteric and axillary lymph nodes), Ovaries (with oviducts), Pituitary gland, Prostate, Rectum, Sciatic nerve, Spinal cord (cervical, thoracic and lumbar cords), Spleen, Seminal vesicle with coagulation glands, Skeletal muscle, Sternum with marrow, Stomach (forestomach and glandular stomach), Testes, Thymus, Thyroid glands (with parathyroid glands), Trachea, Urinary bladder, Uterus , Vagina. - Other examinations:
- The immunorelevant organs and tissues were evaluated according to the following parameters:
Thymus:
- Increased/decreased grade of cortico-medullar ratio (related only to area)
- Increase of starry sky cells
- Changes of cellular density in the cortex
- Changes of cellular density in the medulla
Spleen:
- Changes of the cellularity of PALS, lymphoid follicles, marginal zone, red pulp
- Altered cellular composition of follicles
- Altered number of germinal centers
Lymph nodes (mesenteric and axillary lymph nodes):
- Changes in the cellularity of follicles, interfollicular area, paracortical area, medulla
- Altered cellular composition of paracortex
- Altered number of germinal centers
- Hyperplasia of high endothelial venules
Peyer's patches (of the jejunum):
- Changes of the cellularity of follicles (including mantle zone and germinal centers)
- Changes of the cellularity of interfollicular area
Bone marrow:
- Changes of the cellularity
- Changes of the myeloid/erythroid ratio - Statistics:
- Clinical pathology parameters, urine volume, urine specific gravity: Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (two-sided) for the equal medians.
Urinalysis, except color, turbidity, volume and specific gravity: Pairwise comparison of each dose group with the control group using FISHER's exact test for the hypothesis of equal proportions.
Weight parameters: Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (two-sided) for the equal medians.
Results and discussion
Results of examinations
- Details on results:
- CLINICAL SIGNS AND MORTALITY
In test group 3 (100 and 60 mg/kg bw/d) 2 female animals were found dead on study day 4 and another one on study day 5. One male animal was found dead on study day 6 and 1 male on study day 7. In addition, 2 female animals were sacrificed moribund on study 5, 1 male animal on study day 6 and the remaining 2 male animals on study day 7.
In test group 2 (30 and 20 mg/kg bw/d) 1 female animal was found dead on study day 11 and 1 male and 1 female were sacrificed moribund on study day 12.
In test group 3 (100 and 60 mg/kg bw/d) severe signs of general toxicity were observed. Poor general condition was observed in 3 male on study day 4 and 4 female animals on study day 5. Piloerection was observed in all animals, beginning on study day 4 in males and on study day 3 in females. Semiclosed eyelid was observed in 1 male (study day 5) and 2 females (study day 4). Diarrhea was observed in 1 male animal on study day 6. Blackish discolored feces were observed in all animals since study day 4. Discolored blue skin was observed in all animals, starting on study day 4. Reddish discolored fur, mouth and nose region was seen in 1 male animal and bluish discolored fur in the anogenital region was observed in 4 male animals starting on study day 5. Salivation after treatment was observed in all males on study days 5 and 6 and 1 female on study day 4. All findings were assessed as being related to treatment.
In test group 2 (30 and 20 mg/kg bw/d) poor general condition was observed for all animals from study day 10 to 13. Piloerection was seen in 2 male and 2 female animals on study day 11 and 12, 1 male on study day 20 and 21 and 1 female on study day 14. Semiclosed eyelid was observed in 1 male animal on study days 11 and 12. Discolored blue skin was observed in all animals from study day 10 to 14. Reddish discolored fur, mouth and nose region was seen in 1 female from study day 10 to 12. Blackish discolored feces were observed in all animals from study day 4 onwards. Test substance-like discolored fur at the anogenital region was observed in 1 female animal on study day 12. Salivation after treatment was observed in all animals on several days during the study starting on study day 8. All mentioned findings were assessed as being related to treatment. Respiration sounds were observed in 1 male animal on study day 21 and 22. Alopecia was observed in 1 male animal and 1 female animal from study day 26 on. These findings were assessed as being incidental and not related to treatment.
In test group 1 (10 mg/kg bw/d) only blackish discolored feces were observed in all animals from study day 5 on. Salivation after treatment was observed in all animals on several days during the study starting on study day 8. All findings were assessed as being related to treatment.
BODY WEIGHT AND WEIGHT GAIN
Because of the severe findings in test group 3 (100 and 60 mg/kg bw/d) body weights were additionally determined on study day 5.
Mean body weight in males of test group 3 (100 and 60 mg/kg bw/d) was significantly decreased by -23.7% on study day 5 and -35.1% on study day 7. Also, in females of test group 3 (100 and 60 mg/kg bw/d) loss of body weight was detected and resulted in 19.4% less when compared to the control (study day 5). Mean body weight in males of test group 2 (30 and 20 mg/kg bw/d) was significantly lower (–7.9%) on study day 7 and in females on study day 5 (-7.2%). No changes in body weights were observed for animals of test group 1 (10 mg/kg bw/d).
As the consequence of body weight loss, body weight change was significantly decreased in males of test group 3 (100 and 60 mg/kg bw/d), i.e. on study day 5 (-149.5%) and on study day 7 (-162.4%), as well as in females on study day 5 (-186.5%). Body weight change of test group 2 (30 and 20 mg/kg bw/d) was significantly decreased in males on study days 5 (-43.8%) and 7 (-42.3%) and also in females although not significantly altered. No treatment-related effects were observed for test group 1 (10 mg/kg bw/d).
FOOD CONSUMPTION
Food consumption was reduced for males of test group 3 (100 and 60 mg/kg bw/d) what is clearly related to the fact that only 2 male animals were alive up to study day 7 but in a poor general condition. For the females of this test group, no food consumption was measured, due to premature deaths.
In test group 2 (30 and 20 mg/kg bw/d), food consumption was clearly reduced in male animals between study days 6-7, 13-14 and 20-21. In females only between study days 6-7 impairment of food consumption was observed.
No test substance-related effects on food consumption were obtained in test group 1 (10 mg/kg bw/d).
WATER CONSUMPTION
During the daily visually inspection of the water bottles for any changes in volume no test substance-related findings were observed.
HAEMATOLOGY
No treatment-related changes among hematological parameters were measured.
In female rats of test group 1 (10 mg/kg bw/d) the relative reticulocyte counts were higher compared to controls. This change was not dose dependent and the values were within the historical control range. Therefore, this alteration was regarded as incidental and not treatment-related.
In males of test group 2 (30 and 20 mg/kg bw/d) the prothrombin time (Hepato Quick’s test) was reduced compared to controls. The values were within the historical control range, and, therefore, this alteration was regarded as incidental and not treatment-related.
CLINICAL CHEMISTRY
No treatment-related changes among clinical chemistry parameters were measured.
In females of test group 1 (10 mg/kg bw/d) the alkaline phosphatase (ALP) activity was higher compared to controls, but the values were within the historical control range and they were not changed dose-dependently.
In females of test group 2 (30 and 20 mg/kg bw/d) the alanine aminotransferase (ALT) activity as well as the magnesium values were higher compared to controls, but the values were within the historical control ranges.
In males of test group 2 (30 and 20 mg/kg bw/d) the globulin values were lower compared to controls, but the values were within the historical control range.
Therefore, all mentioned altered parameter values in clinical chemistry were regarded as incidental and not treatment-related.
URINALYSIS
No treatment-related changes among urinalyses parameters were measured.
NEUROBEHAVIOUR
No FOB was performed for test group 3 (100 and 60 mg/kg bw/d).
In male and female animals of test groups 0, 1 and 2 no test substance-related findings were observed except for discoloration of feces in test group 1 (10 mg/kg bw/d) and 2 (30 and 20 mg/kg bw/d), which was assessed as toxicologically not relevant. Any deviations from "zero values" were equally distributed between test substance-treated groups and controls or occurred in single animals only. Therefore, these observations were considered as being incidental.
With regard to the overall motor activity as well as single intervals 2 and 3 a significant decrease was measured in males of test group 2 (30 and 20 mg/kg bw/d). This effect was assessed as being related to the impaired general condition of the animals. No impairment was found in female animals of this test group.
No test substance-related changes were observed for animals of test group 1 (10 mg/kg bw/d). The significantly increased motor activity of females in interval 7 was assessed as spontaneous finding and not treatment-related.
ORGAN WEIGHTS
Significant absolute mean weight decreases were noted for the terminal body weight and thymus of males of test group 2 (30 and 20 mg/kg bw/d). Since no histopathological correlate was found in the thymus the weight decrease was regarded as secondary to the terminal body weight decrease. In females of test group 2 (30 and 20 mg/kg bw/d), weight changes without significance were found in the liver (minimal increase) and uterus (slight decrease).
All other mean absolute weight parameters did not show significant differences when compared to the control group.
The significant increase in the mean relative liver weight in males of test group 2 (30 and 20 mg/kg bw/d) was considered to be in relation with the terminal body weight decrease. In females of test group 2 (30 and 20 mg/kg bw/d), slight changes in mean relative weights without statistical significance were observed in the liver (increase) and uterus (decrease) when compared to control animals. No histopathological correlate was found for the liver. However, a treatment-related effect cannot be completely ruled out. No histomorphological abnormalities were detected in the genital organs that could explain the absolute and relative uterus weight decrease. All females showed vaginal characteristics of metestrus/diestrus in comparison to the control females which mostly showed characteristics of proestrus/estrus (3 of 5 control females). The difference in the cyclus phase most likely accounted for the decreased uterus weight in females of test group 2 (30 and 20 mg/kg bw/d) and was not regarded as treatment-related.
All other mean relative weight parameters did not show significant differences when compared to the control group.
GROSS PATHOLOGY
All male and female rats of test group 3 (100 and 60 mg/kg bw/d) showed a blue discoloration of the carcass, organs and gastrointestinal contents with exception of brain, spinal cord, lungs and fat tissue. In animals of test group 2 (30 and 20 mg/kg bw/d) and 1 (10 mg/kg bw/d) the blue discoloration was less generalized and intense and was mostly found in the gastrointestinal contents. Moreover, thickening of the duodenal wall was seen in 3 of 5 males and 2 of 5 females of test group 2 (30 and 20 mg/kg bw/d) as well as 1 of 5 males of test group 1 (10 mg/kg bw/d).
All other gross lesions observed in test animals occurred singularly. They were considered to be spontaneous in origin and were not related to treatment.
HISTOPATHOLOGY: NON-NEOPLASTIC
In the proximal duodenum, the villi showed subepithelial edema (minimal to slight) and crypt cell hyperplasia (minimal) in 2 of 4 males and 1 of 3 females of test group 2 (30 and 20 mg/kg bw/d) that were sacrificed at schedule. The crypt cell hyperplasia was characterized by minimal increase in the depth of the crypts in relation to the villi length. These findings most probably accounted for the “thickening of wall” observed at gross pathology. In animals of test group 1 (10 mg/kg bw/d) minimal to slight edema was seen in the villi. The findings in the proximal duodenum were considered to be treatment-related.
All other findings were either single observations, or were biologically equally distributed between control and treated animals. All of them were considered to be incidental and/or spontaneous in origin.
Animals of test group 2 (30 and 20 mg/kg bw/d) that died or were sacrificed revealed following findings: moderate to severe decreased cellularity in bone marrow (myeloid and erythroid cells); slight to moderate decreased cellularity in lymph nodes (cortical and paracortical) and reduced size (with normal histomorphology) or atrophy of thymus (reduced cellularity in cortex and medulla). A treatment-related effect on hemolymphatic organs could not be completely ruled out and most probably caused the death or moribund state of these animals. In the proximal duodenum, minimal crypt cell hyperplasia and villi edema (subepithelial, minimal to slight) was seen. In addition, atrophy was observed in genital organs of one male (prostate, seminal vesicles and coagulating glands) and one female (vagina with attenuated stratum germinativum, covered by a single layer of columnar mucous cells; hypertrophy/hyperplasia of interstitial stromal cells in the ovaries; atrophic endometrium and myometrium in the uterus). The atrophy of genital organs was regarded as a secondary effect and was attributed to the poor general conditions of the test animals rather than directly to treatment.
Overall, the study did not reveal a clear mode of action which might explain the observed toxicological effects.
Effect levels
- Dose descriptor:
- NOAEL
- Effect level:
- 10 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: severe signs of toxicity including death, significantly decreased body weight and body weight change, poor general condition.
Target system / organ toxicity
- Critical effects observed:
- not specified
Applicant's summary and conclusion
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