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EC number: 296-117-1 | CAS number: 92257-28-8
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- 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
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- Toxicological Summary
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- Acute Toxicity
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- Exposure related observations in humans
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Endpoint summary
Administrative data
Description of key information
No Observed Adverse Effect Level (NOAEL) of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 1000 mg/kg body weight in male and female animals.
Thus, comparing this effect with the criteria of CLP regulation, 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) can be not classified for repeated dose oral toxicity.
Key value for chemical safety assessment
Repeated dose toxicity: via oral route - systemic effects
Link to relevant study records
- 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:
- guideline study
- Justification for type of information:
- Data is from study report
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
- Principles of method if other than guideline:
- To assess toxicological profile of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8), to determine target organ of toxicity, its reversibility and No Observed Adverse Effect Level (NOAEL) in the rat after 28 consecutive days of oral administration.
- GLP compliance:
- yes
- Limit test:
- yes
- Specific details on test material used for the study:
- - Name of test material : 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar' and ar''-Me derivs.
- IUPAC name : N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine
- Molecular formula : C30H33N3
- Molecular weight : 435.60 g/mol
- Substance type : Organic
- Physical state : Liquid - Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source:National Institute of Biosciences, Pune
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) x wks; (F1) x wks: 6 to 8 weeks
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g: Male - 169.52 g (= 100 %) and Female - 162.35 g (= 100 %)
- Fasting period before study:
- Housing:The rats were housed in polycarbonate cages with paddy as bedding.
After allocation to respective dose groups rats were housed 2/sex/cage.
identified by the picric acid marking. A group of animals in one cage was additionally identified by the label affixed to each cage. A label according to groups identified the cage and each label contained information on cage and study number. It also bear species, strain, sex and identification numbers of rats within it.
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): Rodent feed, provided ad libitum from individual feeders on cage top
- Water (e.g. ad libitum): Water was from a local source and passed through the reverse osmosis membrane before use, ad libitum
- Acclimation period:5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°C (actual range, 19.3 °C to 22.5 °C)
- Humidity (%):30% to 70% (actual range, 45.1% to 58.7%).
- Air changes (per hr): at least ten air changes per hour of 100% fresh air that has been passed through the HEPA filters.
- Photoperiod (hrs dark / hrs light):An artificial light and dark cycle of 12 hours each was provided to the room.
IN-LIFE DATES:
From: 06-12-2017
To: 02-03-2018 - Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS: The test item was diluted with corn oil for preparation of solution(s).
The solution(s) of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl] azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) were made at volumes suitable for daily use for 28 days. The solution(s) were prepared at concentrations of 0, 25, 50 and 100 mg/ml such that dosage of 0 (vehicle), 250, 500 and 1000 mg/kg body weight respectively were administered.
DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:
VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil
- Concentration in vehicle: 0 (vehicle), 250, 500 and 1000 mg/kg body weight
- Amount of vehicle (if gavage): upto 10 ml/kg body weight
- Lot/batch no. (if required):
- Purity: - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Stability determined by UV
- Duration of treatment / exposure:
- 28 days
- Frequency of treatment:
- Daily
- Dose / conc.:
- 0 mg/kg bw/day (actual dose received)
- Dose / conc.:
- 250 mg/kg bw/day (actual dose received)
- Dose / conc.:
- 500 mg/kg bw/day (actual dose received)
- Dose / conc.:
- 1 000 mg/kg bw/day (actual dose received)
- No. of animals per sex per dose:
- Total: 72
0 mg/kg bw: 6 male, 6 female
250 mg/kg bw: 6 male, 6 female
500 mg/kg bw: 6 male, 6 female
1000 mg/kg bw: 6 male, 6 female
Reversal group
0 mg/kg bw: 6 male, 6 female
1000 mg/kg bw: 6 male, 6 female - Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale:
- Rationale for animal assignment (if not random):The animals of uniform body weight were selected. The individual body weights of the animals did not exceed ± 20% of group mean body weight. The group means body weights of all the groups were approximately equal.
- Other: - Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule:twice daily
- Cage side observations checked in table [No.?] were included. Viability were observed
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:daily
BODY WEIGHT: Yes
- Time schedule for examinations:Body weights were recorded on the day of randomization, day of first dosing, weekly thereafter and a fasting body weight at scheduled sacrifice on day 29 and day 43.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- 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
FOOD EFFICIENCY:Not specified
- 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: Not specified
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not specified
- Time schedule for examinations:Not specified
OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to the initiation of the dosing and at scheduled sacrifice.
- Dose groups that were examined: All dose group were examined.
HAEMATOLOGY: Yes
- Time schedule for collection of blood:At the end of dosing period on day 29 and at termination of recovery period on day 43.
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Yes, overnight
- How many animals: All 72 animals were examined.
- Parameters checked in table [No.?] were examined.Hemoglobin (g/dL), Red Blood Corpuscles (x 106 /µL), Hematocrit (%), Mean Corpuscular Volume (fL), Mean Corpuscular Hemoglobin (pg), Mean Corpuscular Hemoglobin Concentration (g/dL), Platelets (x 103 /µL), White Blood Corpuscles (x 103 /µL), Reticulocytes (%), Neutrophils (%), Lymphocytes (%), Eosinophils (%), Monocytes (%), Basophil (%) and Prothrombin time (sec.) were examined.
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:At the end of dosing period on day 29 and at termination of recovery period on day 43.
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Yes, overnight
- How many animals: All 72 animals were examined.
- Parameters checked in table [No.?] were examined.: Total Protein (g/dL), Blood Urea Nitrogen (mg/dL), Urea Nitrogen (mg/dL) Calculated, Alanine Aminotransferase (U/L), Aspartate Aminotransferase (U/L), Alkaline Phosphatase (U/L), Gamma Glutamyl Transferase (U/L), Glucose (mg/dL), Calcium (mmol/L), Phosphorous (mg/dL), Albumin (g/dL), Total Bilirubin (mg/dL), Creatinine (mmol/L), Total Cholesterol (mg/dL), Triglycerides (mg/dL), Globulin (g/dL) Calculated, Sodium (mmol/L), Potassium (mmol/L), Chloride (mmol/L) and Bile acid (mmol/L) were examined.
URINALYSIS: Yes
- Time schedule for collection of urine: during the last week of dosing period and on reversal group rats at termination of recovery period on day 43.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Not specified
- Parameters checked in table [No.?] were examined.: Volume, Appearance, Colour, pH, Specific Gravity, Proteins, Glucose, Ketones, Bilirubin, Urobililogen, Occult Blood and Nitrite were examined.
NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Towards the end of the exposure period of 28 days and towards the end of the recovery period on day 42
- Dose groups that were examined:All dose group were examined.
- Battery of functions tested: sensory activity, grip strength, motor activity, Visual Placing Response were examined.
IMMUNOLOGY: Not specified
- Time schedule for examinations:Not specified
- How many animals:Not specified
- Dose groups that were examined:Not specified
- Parameters checked in table [No.?] were examined.Not specified
OTHER:Organ Weights were examined. - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
after 28 consecutive days of oral administration, all surviving study rats were sacrificed on day 29 (Group I, III, IV, V). In addition all rats from reversal groups were sacrificed on day 43 (Group II and VI).
HISTOPATHOLOGY: Yes
From each rat, samples or the whole of the tissues listed below were preserved. All tissues were fixed in 10% neutral buffered formalin except, eyes and testes of all animals were preserved in Davidson’s solution for 24 hours and transferred to 10% neutral buffered formalin. - Statistics:
- Raw data was processed and analyzed for reporting group means and standard deviations with significance between the controls and treated groups, using SYSTAT 13 validated statistical software supplied by Starcom Information Technology Limited, Bangalore developed by Systat Software, Inc. USA. All the parameters characterized by continuous data such as body weight, feed consumption (calculated as gram per animal), organ weight, relative organ weight, haematological and clinical chemistry data were subjected to Bartlett’s test to meet the homogeneity of variance before conducting Analysis of Variance (ANOVA) and Dunnett’s t-test. Where the data was not meet the homogeneity of variance, Student’s t-test were performed to calculate significance.
Significance was calculated at 5% level and indicated in the summary tables as follows:
* = Significant than control at 95% level of confidence (p<0.05). - Clinical signs:
- no effects observed
- Description (incidence and severity):
- Male -
Group I (Control, 0 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.1 to 6).
Group II (Control, 0 mg/kg, Reversal): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days and during the post-dosing recovery period (animal nos.13 to 18).
Group III (250 mg/kg): Test item coloured faeces were observed in all animals (animal nos.25 to 30, with onset from day 2) during the dosing period of 28 days.
Group IV (500 mg/kg): Test item coloured faeces were observed in all animals (animal nos.37 to 42, with onset from day 2) during the dosing period of 28 days.
Group V (1000 mg/kg): Test item coloured faeces were observed in all animals (animal nos.49 to 54, with onset from day 2) during the dosing period of 28 days.
Group VI (1000 mg/kg, Reversal): Test item coloured faeces were observed in all animals (animal nos.61 to 66, with onset from day 2) throughout the dosing period of 28 days and during the post-dosing recovery period.
Female -
Group I (Control, 0 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.7 to 12).
Group II (Control, 0 mg/kg, Reversal): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days and during the post-dosing recovery period (animal nos.19 to 24).
Group III (250 mg/kg): Test item coloured faeces were observed in all animals (animal nos.31 to 36, with onset from day 2) during the dosing period of 28 days
Group IV (500 mg/kg): Test item coloured faeces were observed in all animals (animal nos.43 to 48, with onset from day 2) during the dosing period of 28 days
Group V (1000 mg/kg): Test item coloured faeces were observed in all animals (animal nos.55 to 60, with onset from day 2) during the dosing period of 28 days
Group VI (1000 mg/kg, Reversal): Test item coloured faeces were observed in all animals (animal nos.67 to 70, with onset from day 2) throughout the dosing period of 28 days and during the post-dosing recovery period
Before commencement of treatment:
In home cage observation, rat from different dose groups and control group revealed normal behavior, alterations, vocalization, respiration and palpebral closer.
During handling observation, handling of rats did not reveal any abnormality from different dose groups and control group.
In the open field observation, rat did not reveal any abnormality from different dose groups and control group.
During treatment:
In home cage observation, rat from different dose groups and control group revealed normal behavior, alterations, vocalization, respiration and palpebral closer.
During handling observation, handling of rats did not reveal any abnormality from different dose groups and control group.
In the open field observation, rat did not reveal any abnormality from different dose groups and control group.
Detailed clinical observation did not reveal any abnormality in all groups during the dosing period of 28 days and during the post-dosing recovery period. - Mortality:
- no mortality observed
- Description (incidence):
- All animals from control and different dose groups survived throughout the dosing period of 28 days and the post-dosing recovery period of 14 days.
- Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- Male -
Animals from control and different dose groups exhibited normal body weight gain throughout the dosing period of 28 days. Reduced body weight gain of 5.48% was observed in male animals from 1000 mg/kg dose group and this effect was attributed to the biological variation within the test system and considered to be of no toxicological importance.
During the post-dosing recovery period, animals from 1000 mg/kg reversal group exhibited normal body weight gain when compared with that of control animals.
Female -
Animals from control and different dose groups exhibited normal body weight gain throughout the dosing period of 28 days.
During the post-dosing recovery period, animals from 1000 mg/kg reversal group exhibited normal body weight gain when compared with that of control animals. - Food consumption and compound intake (if feeding study):
- no effects observed
- Description (incidence and severity):
- Male and Female -
Animals from control and different dose groups exhibited normal feed consumption at the end of the dosing period of 28 days.
Animals from control reversal and high reversal dose groups exhibited normal feed consumption at the end of the recovery period of 14 days. - Food efficiency:
- not specified
- Water consumption and compound intake (if drinking water study):
- not specified
- Ophthalmological findings:
- no effects observed
- Description (incidence and severity):
- No ocular abnormalities were observed on ophthalmological examination in the animals during pre-exposure and at the end of the respective termination.
- Haematological findings:
- no effects observed
- Description (incidence and severity):
- Male and Female -
Haematological investigations conducted at the end of dosing period on day 29 and at the end of recovery period on day 43, revealed following significant changes in the values of different parameters studied when compared with that of respective controls, however the increase/decrease in the values obtained was within normal biological and laboratory limits or the effect was not dose dependent.
Male :
MCHC : Increased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05) and
HCT : Decreased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05).
Female :
Platelets : Increased values were obtained for animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Hb, HCT and MCHC : Decreased values were obtained for animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Total RBC : Decreased values were obtained for animals from 500 mg/kg and 1000 mg/kg dose groups, sacrificed on day 29 (p<0.05) and
Total RBC : Decreased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05). - Clinical biochemistry findings:
- no effects observed
- Description (incidence and severity):
- Male and Female -
Biochemical investigations conducted at the end of dosing period on day 29 and at the end of recovery period on day 43, revealed following significant changes in the values of different parameters studied when compared with that of respective controls, however the increase/decreased in the values obtained was within normal biological and laboratory limits or the effect was not dose dependent.
Male :
Total Bilirubin : Elevated levels were observed in animals from 1000 mg/kg dose group, sacrificed on day 29 (p<0.05),
Creatinine : Elevated levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05),
Glucose : Decreased levels were observed in animals from 500 mg/kg dose group, sacrificed on day 29 (p<0.05),
Calcium : Decreased levels were observed in animals from 500 mg/kg and 1000 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Globulin and Bile Acid : Decreased levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05),
Total Cholesterol : Elevated levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05) and
Glucose : Decreased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05).
Female :
Total Protein and Aspartate Aminotransferase : Elevated levels were observed in animals from 250 mg/kg and 500 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Total Bilirubin : Elevated levels were observed in animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Albumin : Elevated levels were observed in animals from 500 mg/kg dose group, sacrificed on day 29 (p<0.05),
Calcium : Decreased levels were observed in animals from 250 mg/kg and 500 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Triglycerides : Decreased levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05) and
Total Protein : Elevated levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05). - Urinalysis findings:
- no effects observed
- Description (incidence and severity):
- Male and Female -
No statistically significant variation was observed in the urine analyses conducted at the end of the dosing period in week 4 and 6 (on day 23, 24, 25, 26 and 43) in male and female animals of different dose groups as compared to control group animals, except for higher volume of urine was observed in female animals from 250 mg/kg dose group (p<0.05). This higher volume of urine analyses were considered to be incidental and of no toxicological importance. - Behaviour (functional findings):
- no effects observed
- Description (incidence and severity):
- Sensory Reactivity Observations:
All animals from control and different dose groups showed normal arousal level, visual response, touch response, auditory response, tail pinch response and visual placing response. Normal air righting reflex was observed in all animals from control and different dose groups in week 4.
Grip Strength:
Grip strength values observed in male and female animals for control and different dose groups were comparable.
Motor Activity:
Motor activity values observed in male and female animals for control and different dose groups were comparable. - Immunological findings:
- no effects observed
- Organ weight findings including organ / body weight ratios:
- no effects observed
- Description (incidence and severity):
- Male -
In comparison with controls at the end of dosing on day 29, organ weight data of animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups revealed increased relative weights of liver (p<0.05). Increased relative weights of kidneys (p<0.05) were observed in animals from 250 mg/kg dose group. In addition, decreased relative weights of kidneys and heart (p<0.05) were observed in animals from 1000 mg/kg dose group. Decreased relative weights of adrenals (p<0.05) were observed in animals from 250 mg/kg and 1000 mg/kg dose groups. Decreased relative weights of thymus (p<0.05) were observed in animals from 500 mg/kg and 1000 mg/kg dose groups.
In comparison with controls at the end of post-dosing recovery period on day 43, organ weight data of animals from 1000 mg/kg reversal group revealed decreased relative weights of kidneys and prostate + seminal vesicle with coagulation gland as whole (p<0.05).
Female -
In comparison with controls at the end of dosing on day 29, organ weight data of animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups revealed increased relative weights of liver (p<0.05).
In comparison with controls at the end of post-dosing recovery period on day 43, organ weight data of animals from 1000 mg/kg reversal group was found to be comparable.
Although significant changes in the values of organ weight were observed in male and female animals from different dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance. - Gross pathological findings:
- no effects observed
- Description (incidence and severity):
- Gross pathological examination revealed test item coloured perianal region externally and test item coloured stomach mucosa in male and female animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups.
Gross pathological examination in male and female animals from control, control reversal and 1000 mg/kg reversal dose groups did not reveal any abnormality. - Neuropathological findings:
- not specified
- Histopathological findings: non-neoplastic:
- no effects observed
- Description (incidence and severity):
- No treatment related histopathological changes were evident in male and female animals from control and high dose groups.
Incidental, physiological and congenital histopathological changes which were covered in the background historical data of the pathology from control and high dose groups includes minimal, focal to multifocal periportal mononuclear cells infiltration in liver; minimal, multifocal tubular eosinophilic luminal secretion in the kidneys; minimal, multifocal brown pigmentation in spleen; minimal, diffuse dilatation of zona reticularis and/or minimal, multifocal vacuolation in zona fasciculata in the adrenals; minimal, luminal seminal coagulum in urinary bladder; minimal, luminal dilatation in the uterus; minimal, multifocal dilatation of Brunner’s gland in the duodenum; presence of persistent Rathke’s pouch in the pituitary in male and female animals from control and high dose group. - Histopathological findings: neoplastic:
- not specified
- Other effects:
- not specified
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- behaviour (functional findings)
- body weight and weight gain
- clinical biochemistry
- clinical signs
- food consumption and compound intake
- gross pathology
- haematology
- histopathology: non-neoplastic
- mortality
- ophthalmological examination
- organ weights and organ / body weight ratios
- urinalysis
- Remarks on result:
- other: No effect observed
- Critical effects observed:
- not specified
- System:
- other:
- Organ:
- not specified
- Treatment related:
- not specified
- Dose response relationship:
- not specified
- Relevant for humans:
- not specified
- Conclusions:
- No Observed Adverse Effect Level (NOAEL) of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 1000 mg/kg body weight in male and female animals.
- Executive summary:
In a Repeated Dose 28-day Oral Toxicity study, Sprague Dawley male and female rats were treated with 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the concentration of 0, 250 mg/kg, 500 mg/kg and 1000 mg/kg body weight orally by gavage in corn oil. Two additional dose levels were added to the study as control 0 mg/kg (Rev.) and test item 1000 mg/kg (Rev.), in order to study the reversibility or delayed occurrence of symptoms, if any.
No effect on survival, clinical sign, body weight, Feed intake, Ophthalmoscopic examination and functional observation battery of treated male and female rats at the end of dosing period of 28 days and the recovery period of 14 days. Similarly, in male animals at the end of the recovery period on day 43, revealed statistically significant increase in the values of MCHC and statistically significant decrease in the values of HCT at 1000 mg/kg and in female on day 29, revealed statistically significant increase in the values of Platelets at 250 mg/kg, 500 mg/kg and 1000 mg/kg, and statistically significant decrease in the values of Hb, HCT and MCHC at 250 mg/kg, 500 mg/kg and 1000 mg/kg, and Total RBC at 500 mg/kg and 1000 mg/kg, Haematological analysis in female animals conducted at the end of the recovery period on day 43, revealed statistically significant decrease in the values of Total RBC at 1000 mg/kg. In male and female animals at the end of the dosing period on day 29, revealed statistically significant increase in the values of Total Bilirubin at 1000 mg/kg and Creatinine at 250 mg/kg, in male, Total Protein and Aspartate Aminotransferase at 250 mg/kg and 500 mg/kg, Total Bilirubin at 250, 500 and 1000 mg/kg, and Albumin at 500 mg/kg in female rat. In addition, statistically significant decrease was observed in the values of Glucose at 500 mg/kg, Calcium at 500 mg/kg and 1000 mg/kg, Globulin and Bile Acid at 250 mg/kg in male rat and, Calcium at 250 and 500 mg/kg, and Triglycerides at 250 mg/kg in female rat. At the end of the recovery period on day 43 (Reversal groups) statistically significant increase were observed in the values of Total Cholesterol at 1000 mg/kg in male rat and Total Protein at 1000 mg/kg in female rat. In addition, statistically significant decrease was observed in the values of Glucose at 1000 mg/kg in male rat. The increase/decrease in the values of various parameters was marginal and within the normal biological and laboratory limits for hematology and clinical chemistry parameters. In Urine analysis conducted during 4th and 6th week of dosing period (on day 23, 24, 25, 26 and 43), revealed no abnormality attributable to the treatment except for higher volume of urine was observed in female animals from 250 mg/kg dose group and considered to be incidental and of no toxicological importance. In addition, at termination of dosing on day 29, male animals from 250, 500 and 1000 mg/kg dose groups revealed increased relative weights of liver when compared with that of controls. In addition, increased relative weights of kidneys were observed in male animals from 250 mg/kg dose group, when compared with that of controls. Decreased relative weights of kidneys and heart weight at 1000 mg/kg in male as compared to controls. In addition, decreased relative weights of adrenals were observed in male animals from 250 and 1000 mg/kg dose groups when compared with that of controls. Decreased relative weights of thymus were observed in male animals from 500 mg/kg and 1000 mg/kg dose groups when compared with that of controls. Organ weight data of male animals sacrificed on day 43 from 1000 mg/kg reversal group, revealed decreased relative weights of kidneys and prostate + seminal vesicle with coagulation gland as whole when compared with that of controls. At termination of dosing on day 29, female animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups revealed increased relative weights of liver. Organ weight data of female animals sacrificed on day 43 from 1000 mg/kg reversal group, was found to be comparable with that of controls. Although significant changes in the values of organ weight were observed in male and female animals from different dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance. Test item coloured perianal region externally and test item coloured stomach mucosa in male and female animals at 250, 500 and 1000 mg/kg dose groups. Gross pathological examination in male and female animals from control, control reversal and 1000 mg/kg reversal dose groups did not reveal any abnormality. Histopathological examination did not reveal any abnormality attributable. Therefore, No Observed Adverse Effect Level (NOAEL) of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 1000 mg/kg body weight in male and female animals.
Reference
VIABILITY
|
Dose (mg/kg) |
Mortality |
||||
Group |
Males |
Females |
||||
Number |
Male |
Female |
Absolute |
Relative % |
Absolute |
Relative % |
I |
0 |
0 |
0/6 |
0 |
0/6 |
0 |
II |
0 (Reversal) |
0 (Reversal) |
0/6 |
0 |
0/6 |
0 |
III |
250 |
250 |
0/6 |
0 |
0/6 |
0 |
IV |
500 |
500 |
0/6 |
0 |
0/6 |
0 |
V |
1000 |
1000 |
0/6 |
0 |
0/6 |
0 |
VI |
1000 (Reversal) |
1000 (Reversal) |
0/6 |
0 |
0/6 |
0 |
GROUP MEAN BODY WEIGHT (g)
Sex - Male
Group |
Dose (mg/kg) |
|
Weeks |
||||
Number |
|
Day 0 |
Day 1 |
1 |
2 |
3 |
|
I |
0 |
Mean |
169.17 |
170.12 |
200.23 |
228.53 |
254.67 |
±SD |
8.50 |
8.68 |
8.97 |
8.90 |
8.24 |
||
II |
0 (Reversal) |
Mean |
168.87 |
169.73 |
197.90 |
225.98 |
251.90 |
±SD |
8.94 |
8.82 |
9.33 |
8.34 |
7.82 |
||
III |
250 |
Mean |
169.37 |
170.28 |
200.75 |
226.62 |
253.32 |
±SD |
8.04 |
7.66 |
8.06 |
8.49 |
8.30 |
||
IV |
500 |
Mean |
169.67 |
170.67 |
199.43 |
227.15 |
253.32 |
±SD |
7.37 |
7.12 |
7.50 |
8.52 |
7.45 |
||
V |
1000 |
Mean |
170.12 |
170.92 |
197.05 |
219.97 |
242.32 |
±SD |
6.55 |
6.31 |
4.84 |
3.52 |
3.64 |
||
VI |
1000 (Reversal) |
Mean |
169.93 |
171.15 |
199.50 |
224.07 |
248.75 |
±SD |
5.65 |
5.71 |
6.66 |
5.96 |
6.05 |
Group |
Dose (mg/kg) |
|
Weeks |
||
Number |
|
4 |
5 |
6 |
|
I |
0 |
Mean |
279.80 |
|
|
±SD |
9.06 |
||||
II |
0 (Reversal) |
Mean |
278.05 |
295.50 |
314.10 |
±SD |
9.40 |
11.63 |
9.91 |
||
III |
250 |
Mean |
277.17 |
|
|
±SD |
9.28 |
||||
IV |
500 |
Mean |
279.68 |
||
±SD |
6.85 |
||||
V |
1000 |
Mean |
264.47* |
||
±SD |
4.66 |
||||
VI |
1000 (Reversal) |
Mean |
271.88 |
294.02 |
317.88 |
±SD |
7.01 |
6.14 |
6.80 |
* = Significant at 95% level of confidence (p<0.05)
Sex - Female
Group |
Dose (mg/kg) |
|
Weeks |
||||
Number |
|
Day 0 |
Day 1 |
1 |
2 |
3 |
|
I |
0 |
Mean |
162.00 |
163.50 |
182.68 |
195.25 |
200.42 |
±SD |
9.85 |
9.92 |
12.09 |
13.39 |
14.17 |
||
II |
0 (Reversal) |
Mean |
161.78 |
163.08 |
179.25 |
188.37 |
196.62 |
±SD |
10.42 |
10.09 |
17.80 |
20.61 |
24.43 |
||
III |
250 |
Mean |
161.60 |
162.28 |
179.13 |
196.15 |
208.33 |
±SD |
10.64 |
10.98 |
6.35 |
9.89 |
15.63 |
||
IV |
500 |
Mean |
163.15 |
163.93 |
178.10 |
191.40 |
202.67 |
±SD |
9.72 |
9.62 |
9.14 |
8.09 |
9.01 |
||
V |
1000 |
Mean |
162.98 |
163.78 |
180.02 |
198.17 |
207.32 |
±SD |
8.90 |
8.97 |
8.69 |
13.79 |
12.42 |
||
VI |
1000 (Reversal) |
Mean |
162.57 |
163.35 |
178.02 |
196.28 |
206.73 |
±SD |
7.22 |
7.24 |
9.57 |
13.81 |
17.55 |
Group |
Dose (mg/kg) |
|
Weeks |
||
Number |
|
4 |
5 |
6 |
|
I |
0 |
Mean |
208.07 |
|
|
±SD |
12.94 |
||||
II |
0 (Reversal) |
Mean |
204.57 |
213.80 |
223.95 |
±SD |
23.71 |
23.21 |
21.65 |
||
III |
250 |
Mean |
215.58 |
|
|
±SD |
14.03 |
||||
IV |
500 |
Mean |
210.32 |
||
±SD |
9.17 |
||||
V |
1000 |
Mean |
216.13 |
||
±SD |
11.36 |
||||
VI |
1000 (Reversal) |
Mean |
213.83 |
221.57 |
229.22 |
±SD |
15.90 |
15.25 |
11.54 |
GROUP MEAN FEED CONSUMPTION (g/animal/day)
Sex - Male
Group |
Dose (mg/kg) |
|
Day |
||||
Number |
|
1 |
8 |
15 |
22 |
28 |
|
I |
0 |
Mean |
18.88 |
20.18 |
21.30 |
22.80 |
23.65 |
II |
0 (Reversal) |
Mean |
18.07 |
19.65 |
20.78 |
22.23 |
23.25 |
III |
250 |
Mean |
17.45 |
19.02 |
20.10 |
21.45 |
22.70 |
IV |
500 |
Mean |
19.20 |
20.62 |
21.60 |
22.92 |
23.80 |
V |
1000 |
Mean |
18.15 |
19.73 |
20.67 |
21.90 |
22.48 |
VI |
1000 (Reversal) |
Mean |
18.12 |
19.88 |
20.88 |
22.02 |
22.80 |
Group |
Dose (mg/kg) |
|
Day |
|
Number |
|
36 |
42 |
|
I |
0 |
Mean |
|
|
II |
0 (Reversal) |
Mean |
24.00 |
25.35 |
III |
250 |
Mean |
|
|
IV |
500 |
Mean |
||
V |
1000 |
Mean |
||
VI |
1000 (Reversal) |
Mean |
23.30 |
24.50 |
Sex - Female
Group |
Dose (mg/kg) |
|
Day |
||||
Number |
|
1 |
8 |
15 |
22 |
28 |
|
I |
0 |
Mean |
15.33 |
16.60 |
16.99 |
18.63 |
19.52 |
II |
0 (Reversal) |
Mean |
15.53 |
16.77 |
18.10 |
19.27 |
20.17 |
III |
250 |
Mean |
14.27 |
15.57 |
16.80 |
18.13 |
18.90 |
IV |
500 |
Mean |
16.02 |
17.45 |
18.87 |
19.78 |
20.70 |
V |
1000 |
Mean |
14.65 |
16.20 |
17.45 |
18.53 |
19.28 |
VI |
1000 (Reversal) |
Mean |
14.38 |
15.93 |
17.07 |
18.27 |
19.27 |
Group |
Dose (mg/kg) |
|
Day |
|
Number |
|
36 |
42 |
|
I |
0 |
Mean |
|
|
II |
0 (Reversal) |
Mean |
20.83 |
21.50 |
III |
250 |
Mean |
|
|
IV |
500 |
Mean |
||
V |
1000 |
Mean |
||
VI |
1000 (Reversal) |
Mean |
20.07 |
20.78 |
OPHTHALMOSCOPIC EXAMINATION
Sex : Male
Day : 0
Group Number |
Dose mg/kg |
Ophthalmoscopic Finding |
Total Number of Animals |
Animal Number |
I |
0 |
No abnormality detected |
6 |
1 - 6 |
II |
0 (Rev.) |
No abnormality detected |
6 |
13 - 18 |
III |
250 |
No abnormality detected |
6 |
25 - 30 |
IV |
500 |
No abnormality detected |
6 |
37 - 42 |
V |
1000 |
No abnormality detected |
6 |
49 - 54 |
VI |
1000 (Rev.) |
No abnormality detected |
6 |
61 - 66 |
Sex : Female
Day : 0
Group Number |
Dose mg/kg |
Ophthalmoscopic Finding |
Total Number of Animals |
Animal Number |
I |
0 |
No abnormality detected |
6 |
7 - 12 |
II |
0 (Rev.) |
No abnormality detected |
6 |
19 - 24 |
III |
250 |
No abnormality detected |
6 |
31 - 36 |
IV |
500 |
No abnormality detected |
6 |
43 - 48 |
V |
1000 |
No abnormality detected |
6 |
55 - 60 |
VI |
1000 (Rev.) |
No abnormality detected |
6 |
67 - 72 |
Sex : Male
Week: 4 and 6
Group Number |
Dose mg/kg |
Ophthalmoscopic Finding |
Total Number of Animals |
Animal Number |
I |
0 |
No abnormality detected |
6 |
1 - 6 |
II |
0 (Rev.) |
No abnormality detected |
6 |
13 - 18 |
III |
250 |
No abnormality detected |
6 |
25 - 30 |
IV |
500 |
No abnormality detected |
6 |
37 - 42 |
V |
1000 |
No abnormality detected |
6 |
49 - 54 |
VI |
1000 (Rev.) |
No abnormality detected |
6 |
61 - 66 |
Sex : Female
Week: 4 and 6
Group Number |
Dose mg/kg |
Ophthalmoscopic Finding |
Total Number of Animals |
Animal Number |
I |
0 |
No abnormality detected |
6 |
7 - 12 |
II |
0 (Rev.) |
No abnormality detected |
6 |
19 - 24 |
III |
250 |
No abnormality detected |
6 |
31 - 36 |
IV |
500 |
No abnormality detected |
6 |
43 - 48 |
V |
1000 |
No abnormality detected |
6 |
55 - 60 |
VI |
1000 (Rev.) |
No abnormality detected |
6 |
67 - 72 |
Rev. = Reversal
SUMMARY OF CLINICAL OBSERVATIONS AND GENERAL APPEARANCE
Sex : Male
Group Number |
Dose (mg/kg) |
Observed Signs |
Total Number of Animals |
Animal Nos. |
Period of signs in days from - to |
Mortality |
I |
0 |
Nil |
6 |
1 - 6 |
1 - 28 |
0/6 |
II |
0 (Reversal) |
Nil |
6 |
13 - 18 |
1 - 42 |
0/6 |
III |
250 |
Test Item coloured faces |
6 |
25 - 30 |
2 - 28 |
0/6 |
IV |
500 |
Test Item coloured faces |
6 |
37 - 42 |
2 - 28 |
0/6 |
V |
1000 |
Test Item coloured faces |
6 |
49 - 54 |
2 - 28 |
0/6 |
VI |
1000 (Reversal) |
Test Item coloured faces |
6 |
61,66 62,64 63 65 |
2 - 33 2 - 32 2 - 34 2 - 31 |
0/6 |
Sex : Female
Group Number |
Dose (mg/kg) |
Observed Signs |
Total Number of Animals |
Animal Nos. |
Period of signs in days from - to |
Mortality |
I |
0 |
Nil |
6 |
7 - 12 |
1 - 28 |
0/6 |
II |
0 (Reversal) |
Nil |
6 |
19 - 24 |
1 - 42 |
0/6 |
III |
250 |
Test Item coloured faces |
6 |
31 - 36 |
2 - 28 |
0/6 |
IV |
500 |
Test Item coloured faces |
6 |
43 - 48 |
2 - 28 |
0/6 |
V |
1000 |
Test Item coloured faces |
6 |
55 - 60 |
2 - 28 |
0/6 |
VI |
1000 (Reversal) |
Test Item coloured faces |
6 |
67 - 72 |
2 - 33 |
0/6 |
SUMMARY OF FUNCTIONAL OBSERVATIONAL BATTERY
Sex : Male |
|
Day : 28 and 42 |
|
Group Number |
I |
II |
III |
IV |
V |
VI |
|
Dose |
0 mg/kg |
0 mg/kg Reversal |
250 mg/kg |
500 mg/kg |
1000 mg/kg |
1000 mg/kg Reversal |
|
Number of animals observed |
6 |
6 |
6 |
6 |
6 |
6 |
|
Number of animals within normal limit |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Number of animals with significant deviation |
0/6 |
0/6 |
0/6 |
0/6 |
0/6 |
0/6 |
|
Parameters |
|
|
|
|
|
|
|
Arousal level : Apparently normal |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Visual response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Touch response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Auditory response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Tail pinch response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Visual placing response : Early extension of forelimbs to reach for the screen |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Air righting response : Lands with all feet on ground |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Grip Strength (kg) - Mean ± SD |
0.902 ± 0.032 |
0.893 ±0.050 |
0.866 ± 0.045 |
0.917 ± 0.045 |
0.863 ± 0.035 |
0.853 ±0.065 |
|
Motor Activity - |
Interval ‘1’ |
534.17 ±94.79 |
648.00 ±200.76 |
598.00 ±65.24 |
527.33 ±84.91 |
557.67 ±129.09 |
657.00 ±173.42 |
Mean ± SD |
Interval ‘2’ |
402.67 ±81.27 |
428.67 ±64.15 |
443.17 ±91.57 |
422.50 ±54.27 |
462.33 ±65.58 |
443.33 ±116.99 |
|
Interval ‘3’ |
313.83 ±73.96 |
293.17 ±81.21 |
338.83 ±71.97 |
348.83 ±42.85 |
362.83 ±91.58 |
223.33 ±76.16 |
Sex : Female |
|
Day : 28 and 42 |
|
Group Number |
I |
II |
III |
IV |
V |
VI |
|
Dose |
0 mg/kg |
0 mg/kg Reversal |
250 mg/kg |
500 mg/kg |
1000 mg/kg |
1000 mg/kg Reversal |
|
Number of animals observed |
6 |
6 |
6 |
6 |
6 |
6 |
|
Number of animals within normal limit |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Number of animals with significant deviation |
0/6 |
0/6 |
0/6 |
0/6 |
0/6 |
0/6 |
|
Parameters |
|
|
|
|
|
|
|
Arousal level : Apparently normal |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Visual response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Touch response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Auditory response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Tail pinch response : Orienting response |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Visual placing response : Early extension of forelimbs to reach for the screen |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Air righting response : Lands with all feet on ground |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
6/6 |
|
Grip Strength (kg) - Mean ± SD |
0.793 ± 0.031 |
0.860 ±0.069 |
0.801 ± 0.038 |
0.796 ± 0.034 |
0.768 ± 0.028 |
0.834 ±0.030 |
|
Motor Activity - |
Interval ‘1’ |
450.17 ±180.70 |
651.67 ±96.29 |
551.17 ±71.54 |
578.50 ±150.27 |
588.33 ±98.26 |
604.33 ±44.37 |
Mean ± SD |
Interval ‘2’ |
267.33 ±150.34 |
488.50 ±145.77 |
317.17 ±94.30 |
416.17 ±107.29 |
373.50 ±171.44 |
440.00 ±87.34 |
|
Interval ‘3’ |
208.33 ±114.31 |
375.17 ±114.19 |
221.50 ±81.29 |
331.17 ±101.84 |
237.17 ±103.36 |
312.33 ±110.57 |
GROUP MEAN HAEMATOLOGY
Sex : Male
Day : 29 and 43
Group |
Dose (mg/kg) |
|
Hb |
Total RBC |
Rt |
HCT |
MCV |
MCH |
MCHC |
Number |
|
(g/dL) |
(6/µL) |
(%) |
(%) |
(fL) |
(pg) |
(g/dL) |
|
I |
0 |
Mean |
15.10 |
8.25 |
4.70 |
47.13 |
57.23 |
18.33 |
32.03 |
±SD |
1.00 |
0.69 |
0.84 |
2.78 |
2.06 |
0.48 |
0.61 |
||
II |
0 (Reversal) |
Mean |
15.03 |
8.38 |
4.70 |
47.27 |
56.53 |
18.00 |
31.85 |
±SD |
1.16 |
0.76 |
0.54 |
3.57 |
2.90 |
0.88 |
0.23 |
||
III |
250 |
Mean |
15.12 |
8.24 |
5.03 |
47.43 |
57.60 |
18.37 |
31.93 |
±SD |
0.46 |
0.20 |
0.62 |
1.10 |
1.17 |
0.38 |
0.31 |
||
IV |
500 |
Mean |
14.88 |
8.09 |
4.58 |
46.47 |
57.43 |
18.42 |
32.02 |
±SD |
0.95 |
0.50 |
0.85 |
2.25 |
1.11 |
0.48 |
0.71 |
||
V |
1000 |
Mean |
14.78 |
7.96 |
4.67 |
46.42 |
58.37 |
18.60 |
31.87 |
±SD |
0.71 |
0.49 |
0.88 |
2.33 |
0.87 |
0.33 |
0.37 |
||
VI |
1000 (Reversal) |
Mean |
14.47 |
7.69 |
4.30 |
41.95* |
54.70 |
18.87 |
34.52* |
±SD |
0.96 |
0.68 |
0.67 |
2.60 |
2.50 |
0.78 |
0.44 |
Group |
Dose (mg/kg) |
|
Platelets |
Total WBC |
Differential % |
Pt. |
||||
Number |
|
(3/ µL) |
(3/µL) |
N |
L |
E |
M |
B |
(Sec.) |
|
I |
0 |
Mean |
364.17 |
12.18 |
17.33 |
81.83 |
0.33 |
0.50 |
0.00 |
19.00 |
±SD |
77.50 |
4.24 |
2.58 |
3.19 |
0.52 |
0.84 |
0.00 |
4.20 |
||
II |
0 (Reversal) |
Mean |
374.00 |
13.68 |
15.83 |
82.67 |
0.83 |
0.67 |
0.00 |
20.50 |
±SD |
62.86 |
2.73 |
2.86 |
2.16 |
0.75 |
0.82 |
0.00 |
3.08 |
||
III |
250 |
Mean |
388.50 |
13.75 |
18.17 |
80.67 |
0.83 |
0.33 |
0.00 |
20.00 |
±SD |
54.96 |
1.84 |
2.14 |
1.86 |
0.75 |
0.52 |
0.00 |
6.96 |
||
IV |
500 |
Mean |
391.50 |
12.95 |
18.17 |
79.83 |
0.83 |
1.17 |
0.00 |
20.17 |
±SD |
31.63 |
2.40 |
3.31 |
2.79 |
0.75 |
0.75 |
0.00 |
6.11 |
||
V |
1000 |
Mean |
417.67 |
14.28 |
16.83 |
82.33 |
0.67 |
0.17 |
0.00 |
17.67 |
±SD |
38.88 |
1.90 |
3.31 |
3.27 |
0.82 |
0.41 |
0.00 |
4.63 |
||
VI |
1000 (Reversal) |
Mean |
396.00 |
13.50 |
16.83 |
81.83 |
0.67 |
0.67 |
0.00 |
19.83 |
±SD |
59.64 |
3.22 |
3.06 |
3.60 |
0.82 |
1.21 |
0.00 |
4.71 |
Sex : Female
Day : 29 and 43
Group |
Dose (mg/kg) |
|
Hb |
Total RBC |
Rt |
HCT |
MCV |
MCH |
MCHC |
Number |
|
(g/dL) |
(6/µL) |
(%) |
(%) |
(fL) |
(pg) |
(g/dL) |
|
I |
0 |
Mean |
15.38 |
7.83 |
4.40 |
46.60 |
59.60 |
19.70 |
33.05 |
±SD |
0.69 |
0.53 |
0.87 |
2.54 |
3.18 |
1.01 |
0.34 |
||
II |
0 (Reversal) |
Mean |
14.02 |
8.46 |
4.55 |
44.10 |
52.20 |
16.60 |
31.82 |
±SD |
0.74 |
0.51 |
0.94 |
2.36 |
1.75 |
0.52 |
0.50 |
||
III |
250 |
Mean |
13.63* |
7.37 |
4.63 |
42.30* |
57.40 |
18.47 |
32.20* |
±SD |
0.95 |
0.45 |
0.94 |
2.68 |
1.76 |
0.59 |
0.24 |
||
IV |
500 |
Mean |
13.25* |
6.90* |
4.17 |
41.25* |
59.90 |
19.27 |
32.13* |
±SD |
0.68 |
0.44 |
0.88 |
2.17 |
3.75 |
1.35 |
0.48 |
||
V |
1000 |
Mean |
12.73* |
6.90* |
4.72 |
40.23* |
58.43 |
18.48 |
31.63* |
±SD |
0.89 |
0.60 |
0.63 |
2.97 |
1.95 |
0.44 |
0.58 |
||
VI |
1000 (Reversal) |
Mean |
13.42 |
7.75* |
4.53 |
42.75 |
55.28 |
17.33 |
31.37 |
±SD |
0.71 |
0.51 |
0.90 |
2.06 |
2.80 |
0.75 |
0.40 |
Group |
Dose (mg/kg) |
|
Platelets |
Total WBC |
Differential % |
Pt. |
||||
Number |
|
(3/ µL) |
(3/µL) |
N |
L |
E |
M |
B |
(Sec.) |
|
I |
0 |
Mean |
360.17 |
11.92 |
15.67 |
82.67 |
1.17 |
0.50 |
0.00 |
20.00 |
±SD |
50.76 |
3.33 |
2.73 |
2.16 |
1.17 |
0.84 |
0.00 |
6.36 |
||
II |
0 (Reversal) |
Mean |
329.67 |
9.23 |
15.33 |
82.50 |
1.00 |
1.17 |
0.00 |
21.17 |
±SD |
92.46 |
2.35 |
2.16 |
2.07 |
0.89 |
0.75 |
0.00 |
3.87 |
||
III |
250 |
Mean |
447.00* |
11.63 |
18.00 |
80.83 |
0.83 |
0.33 |
0.00 |
20.33 |
±SD |
70.63 |
2.32 |
3.46 |
3.82 |
0.75 |
0.52 |
0.00 |
6.74 |
||
IV |
500 |
Mean |
509.17* |
10.42 |
17.67 |
80.83 |
0.83 |
0.67 |
0.00 |
18.67 |
±SD |
54.25 |
1.46 |
2.58 |
2.56 |
0.75 |
0.82 |
0.00 |
6.44 |
||
V |
1000 |
Mean |
480.83* |
13.43 |
16.50 |
82.00 |
1.00 |
0.50 |
0.00 |
18.67 |
±SD |
48.62 |
3.21 |
3.27 |
4.10 |
0.63 |
0.84 |
0.00 |
4.32 |
||
VI |
1000 (Reversal) |
Mean |
393.83 |
8.18 |
15.83 |
82.50 |
0.83 |
0.83 |
0.00 |
21.50 |
±SD |
72.71 |
3.84 |
3.19 |
2.74 |
0.98 |
0.75 |
0.00 |
3.73 |
GROUP MEAN CLINICAL BIOCHEMISTRY
Sex : Male
Day : 29 and 43
Group Number |
Dose (mg/kg) |
|
TotalProtein (g/dL) |
BUN (mg/dL) |
Urea (mg/dL) |
ALT (U/L) |
AST (U/L) |
ALP (U/L) |
Glucose (mg/dL) |
I |
0 |
Mean |
6.44 |
14.50 |
31.61 |
52.67 |
84.67 |
179.83 |
91.50 |
±SD |
0.25 |
1.52 |
3.31 |
8.94 |
7.26 |
80.63 |
3.83 |
||
II |
0 (Reversal) |
Mean |
6.73 |
14.67 |
31.97 |
52.83 |
126.50 |
78.33 |
107.50 |
±SD |
0.09 |
1.51 |
3.28 |
4.07 |
10.75 |
11.54 |
9.81 |
||
III |
250 |
Mean |
6.11 |
14.00 |
30.52 |
39.33 |
81.00 |
119.50 |
85.33 |
±SD |
0.38 |
1.67 |
3.65 |
4.72 |
7.77 |
30.75 |
15.65 |
||
IV |
500 |
Mean |
6.19 |
14.67 |
31.97 |
41.50 |
86.83 |
133.33 |
73.17* |
±SD |
0.32 |
1.37 |
2.98 |
11.61 |
6.43 |
36.81 |
6.74 |
||
V |
1000 |
Mean |
6.37 |
14.17 |
30.88 |
43.17 |
88.67 |
133.00 |
95.00 |
±SD |
0.39 |
2.14 |
4.66 |
11.51 |
13.00 |
50.51 |
6.99 |
||
VI |
1000 (Reversal) |
Mean |
6.83 |
16.83 |
36.70 |
44.50 |
122.83 |
65.17 |
89.83* |
±SD |
0.22 |
2.86 |
6.23 |
8.43 |
18.88 |
16.03 |
7.08 |
Group Number |
Dose (mg/kg) |
|
Calcium(mmol/L) |
Phospho-rous (mg/dL) |
GGT (U/L) |
Total Bilirubin (mg/dL) |
Albumin (g/dL) |
Globulin (g/dL) |
Creatinine (mg/dL) |
I |
0 |
Mean |
3.43 |
7.88 |
5.67 |
0.22 |
1.04 |
5.38 |
0.45 |
±SD |
0.23 |
0.99 |
0.52 |
0.02 |
0.08 |
0.17 |
0.04 |
||
II |
0 (Reversal) |
Mean |
4.18 |
9.18 |
6.50 |
0.14 |
1.12 |
5.60 |
0.47 |
±SD |
0.03 |
0.86 |
1.64 |
0.04 |
0.09 |
0.15 |
0.07 |
||
III |
250 |
Mean |
3.16 |
7.95 |
5.67 |
0.17 |
1.18 |
4.90* |
0.51* |
±SD |
0.17 |
0.58 |
0.52 |
0.03 |
0.09 |
0.37 |
0.04 |
||
IV |
500 |
Mean |
3.13* |
8.50 |
5.17 |
0.19 |
1.13 |
5.05 |
0.48 |
±SD |
0.21 |
0.57 |
0.75 |
0.03 |
0.14 |
0.21 |
0.03 |
||
V |
1000 |
Mean |
3.00* |
7.18 |
6.67 |
0.30* |
0.90 |
5.45 |
0.42 |
±SD |
0.17 |
0.56 |
1.03 |
0.04 |
0.10 |
0.40 |
0.02 |
||
VI |
1000 (Reversal) |
Mean |
4.17 |
7.97 |
5.83 |
0.14 |
1.04 |
5.78 |
0.49 |
±SD |
0.18 |
1.04 |
1.72 |
0.04 |
0.07 |
0.22 |
0.08 |
Sex : Male
Day : 29 and 43
Group Number |
Dose (mg/kg)
|
|
Sodium (mmol/L) |
Potassium (mmol/L) |
Chloride (mmol/L) |
Total Cholesterol (mg/dL) |
Triglycerides (mg/dL) |
Bile Acids (µmol/L) |
I |
0 |
Mean |
148.85 |
4.53 |
108.92 |
55.50 |
81.00 |
21.42 |
±SD |
0.61 |
0.26 |
1.72 |
7.89 |
21.89 |
12.81 |
||
II |
0 (Reversal) |
Mean |
147.50 |
4.51 |
101.51 |
57.50 |
73.83 |
12.26 |
±SD |
0.91 |
0.40 |
1.29 |
3.83 |
24.47 |
5.79 |
||
III |
250 |
Mean |
149.39 |
4.42 |
107.58 |
59.33 |
80.00 |
7.24* |
±SD |
1.61 |
0.22 |
1.21 |
11.93 |
26.82 |
2.41 |
||
IV |
500 |
Mean |
149.92 |
4.41 |
107.52 |
53.67 |
65.00 |
19.52 |
±SD |
1.16 |
0.23 |
1.71 |
8.80 |
14.72 |
16.87 |
||
V |
1000 |
Mean |
150.13 |
4.46 |
107.72 |
69.83 |
76.00 |
11.33 |
±SD |
1.43 |
0.36 |
2.59 |
20.67 |
27.53 |
6.41 |
||
VI |
1000 (Reversal) |
Mean |
147.81 |
4.30 |
101.30 |
70.67* |
83.83 |
16.23 |
±SD |
1.54 |
0.39 |
2.79 |
10.11 |
22.94 |
10.92 |
Sex : Female
Day : 29 and 43
Group Number |
Dose (mg/kg) |
|
TotalProtein (g/dL) |
BUN (mg/dL) |
Urea (mg/dL) |
ALT (U/L) |
AST (U/L) |
ALP (U/L) |
Glucose (mg/dL) |
I |
0 |
Mean |
6.73 |
17.50 |
38.15 |
51.00 |
102.83 |
113.67 |
96.83 |
±SD |
0.35 |
1.87 |
4.08 |
18.42 |
16.74 |
52.38 |
24.16 |
||
II |
0 (Reversal) |
Mean |
6.88 |
17.17 |
37.42 |
39.50 |
114.00 |
66.33 |
98.83 |
±SD |
0.03 |
2.32 |
5.05 |
5.75 |
10.77 |
25.94 |
13.86 |
||
III |
250 |
Mean |
7.16* |
19.17 |
41.78 |
40.50 |
128.33* |
82.17 |
90.17 |
±SD |
0.18 |
4.07 |
8.87 |
8.36 |
11.57 |
24.34 |
11.55 |
||
IV |
500 |
Mean |
7.29* |
20.17 |
43.96 |
46.83 |
129.33* |
79.83 |
81.67 |
±SD |
0.18 |
1.72 |
3.75 |
5.46 |
12.66 |
16.96 |
6.74 |
||
V |
1000 |
Mean |
6.68 |
19.00 |
41.42 |
41.67 |
111.83 |
125.67 |
86.50 |
±SD |
0.35 |
2.10 |
4.57 |
9.20 |
15.26 |
26.77 |
13.72 |
||
VI |
1000 (Reversal) |
Mean |
7.12* |
17.67 |
38.51 |
38.67 |
111.17 |
45.83 |
91.00 |
±SD |
0.24 |
3.14 |
6.85 |
8.62 |
16.29 |
10.55 |
10.49 |
Group Number |
Dose (mg/kg) |
|
Calcium(mmol/L) |
Phospho-rous (mg/dL) |
GGT (U/L) |
Total Bilirubin (mg/dL) |
Albumin (g/dL) |
Globulin (g/dL) |
Creatinine (mg/dL) |
I |
0 |
Mean |
4.02 |
7.15 |
6.33 |
0.15 |
1.29 |
5.43 |
0.66 |
±SD |
0.04 |
0.76 |
0.82 |
0.02 |
0.17 |
0.45 |
0.05 |
||
II |
0 (Reversal) |
Mean |
3.85 |
7.32 |
6.83 |
0.17 |
1.06 |
5.83 |
0.60 |
±SD |
0.07 |
0.57 |
0.75 |
0.02 |
0.11 |
0.14 |
0.08 |
||
III |
250 |
Mean |
3.93* |
7.83 |
7.33 |
0.19* |
1.43 |
5.72 |
0.65 |
±SD |
0.05 |
0.31 |
0.82 |
0.01 |
0.11 |
0.23 |
0.03 |
||
IV |
500 |
Mean |
3.83* |
7.53 |
7.33 |
0.23* |
1.52* |
5.78 |
0.66 |
±SD |
0.11 |
0.50 |
0.82 |
0.03 |
0.10 |
0.13 |
0.03 |
||
V |
1000 |
Mean |
3.93 |
6.97 |
6.33 |
0.30* |
1.29 |
5.38 |
0.59 |
±SD |
0.14 |
0.56 |
1.03 |
0.03 |
0.14 |
0.28 |
0.06 |
||
VI |
1000 (Reversal) |
Mean |
3.84 |
6.98 |
5.83 |
0.17 |
1.10 |
6.00 |
0.56 |
±SD |
0.11 |
0.91 |
0.98 |
0.02 |
0.06 |
0.22 |
0.06 |
Sex : Female
Day : 29 and 43
Group Number |
Dose (mg/kg)
|
|
Sodium (mmol/L) |
Potassium (mmol/L) |
Chloride (mmol/L) |
Total Cholesterol (mg/dL) |
Triglycerides (mg/dL) |
Bile Acids (µmol/L) |
I |
0 |
Mean |
144.83 |
3.96 |
107.46 |
70.67 |
81.50 |
15.40 |
±SD |
1.37 |
0.54 |
2.74 |
11.98 |
19.26 |
5.06 |
||
II |
0 (Reversal) |
Mean |
148.12 |
3.89 |
107.03 |
65.67 |
54.00 |
14.40 |
±SD |
1.54 |
0.29 |
1.82 |
14.75 |
20.83 |
12.69 |
||
III |
250 |
Mean |
145.83 |
4.47 |
107.25 |
81.17 |
57.00* |
15.92 |
±SD |
1.24 |
0.39 |
0.88 |
12.78 |
13.02 |
6.41 |
||
IV |
500 |
Mean |
146.12 |
4.28 |
108.20 |
81.67 |
72.83 |
25.22 |
±SD |
1.06 |
0.36 |
0.95 |
5.68 |
9.87 |
10.30 |
||
V |
1000 |
Mean |
144.53 |
4.54 |
108.99 |
86.17 |
89.17 |
10.48 |
±SD |
0.99 |
0.32 |
1.70 |
9.41 |
16.68 |
3.60 |
||
VI |
1000 (Reversal) |
Mean |
146.46 |
4.10 |
107.97 |
79.00 |
85.17 |
11.73 |
±SD |
1.85 |
0.44 |
2.28 |
4.90 |
41.24 |
6.23 |
* = Significant at 95% level of confidence (p<0.05)
GROUP MEAN URINE ANALYSES
Sex : Male
Day : 23, 24 and 43
Group |
Dose |
|
Volume |
Glucose |
Bilirubin |
Ketones |
Sp.Gr. |
Occult Blood |
Number |
mg/kg |
|
(ml) |
(mmol/L) |
(mmol/L) |
(mmol/L) |
(g/L) |
(caCELLS/µL) |
I |
0 |
Mean |
5.483 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
1.143 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
II |
0 (Rev.) |
Mean |
5.083 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.906 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
III |
250 |
Mean |
6.433 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.873 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
IV |
500 |
Mean |
6.383 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.711 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
V |
1000 |
Mean |
5.700 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.522 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
VI |
1000 (Rev.) |
Mean |
5.367 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
1.513 |
-ve |
-ve |
-ve |
0.003 |
-ve |
Group |
Dose |
|
pH |
Urobilinogen |
Nitrite |
Number |
mg/kg |
|
|
(mmol/L) |
|
I |
0 |
Mean |
7.250 |
-ve |
-ve |
±SD |
0.274 |
-ve |
-ve |
||
II |
0 (Rev.) |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
||
III |
250 |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
||
IV |
500 |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
||
V |
1000 |
Mean |
7.250 |
-ve |
-ve |
±SD |
0.274 |
-ve |
-ve |
||
VI |
1000 (Rev.) |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
Sex : Female
Day : 24, 25 and 43
Group |
Dose |
|
Volume |
Glucose |
Bilirubin |
Ketones |
Sp.Gr. |
Occult Blood |
Number |
mg/kg |
|
(ml) |
(mmol/L) |
(mmol/L) |
(mmol/L) |
(g/L) |
(caCELLS/µL) |
I |
0 |
Mean |
5.150 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.677 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
II |
0 (Rev.) |
Mean |
5.783 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.519 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
III |
250 |
Mean |
6.383* |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.828 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
IV |
500 |
Mean |
5.533 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.753 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
V |
1000 |
Mean |
5.317 |
-ve |
-ve |
-ve |
1.017 |
-ve |
±SD |
0.725 |
-ve |
-ve |
-ve |
0.003 |
-ve |
||
VI |
1000 (Rev.) |
Mean |
5.567 |
-ve |
-ve |
-ve |
1.018 |
-ve |
±SD |
0.802 |
-ve |
-ve |
-ve |
0.003 |
-ve |
Group |
Dose |
|
pH |
Urobilinogen |
Nitrite |
Number |
mg/kg |
|
|
(mmol/L) |
|
I |
0 |
Mean |
7.167 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
||
II |
0 (Rev.) |
Mean |
7.250 |
-ve |
-ve |
±SD |
0.274 |
-ve |
-ve |
||
III |
250 |
Mean |
7.250 |
-ve |
-ve |
±SD |
0.274 |
-ve |
-ve |
||
IV |
500 |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
||
V |
1000 |
Mean |
7.250 |
-ve |
-ve |
±SD |
0.274 |
-ve |
-ve |
||
VI |
1000 (Rev.) |
Mean |
7.333 |
-ve |
-ve |
±SD |
0.258 |
-ve |
-ve |
Sp.Gr. : Specific gravity +ve : Positive -ve : Negative
Qualitative Absent = 0 Trace = + Small amount of analyte = ++ Moderate amount of analyte = +++ Large amount of analyte = ++++ |
|
Rev. = Reversal
* = Significant at 95% level of confidence (p<0.05)
GROUP MEAN ABSOLUTE ORGAN WEIGHTS (g)
Sex : Male
Day : 29 and 43
GroupNumber |
Dose (mg/kg) |
|
Body Weight (g) |
Brain |
Liver |
Kidneys |
Adrenals |
Testes |
Prostate + Seminal Vesicle with Coagulation gland |
I |
0 |
Mean |
258.267 |
1.837 |
9.101 |
1.971 |
0.045 |
2.852 |
1.649 |
±SD |
11.271 |
0.098 |
0.664 |
0.205 |
0.006 |
0.129 |
0.464 |
||
II |
0 (Reversal) |
Mean |
294.200 |
1.879 |
11.420 |
2.480 |
0.050 |
2.882 |
1.784 |
±SD |
11.583 |
0.127 |
1.541 |
0.319 |
0.005 |
0.283 |
0.210 |
||
III |
250 |
Mean |
255.883 |
1.777 |
14.287 |
2.198 |
0.037 |
2.866 |
1.768 |
±SD |
9.429 |
0.105 |
1.364 |
0.126 |
0.005 |
0.160 |
0.209 |
||
IV |
500 |
Mean |
260.067 |
1.722 |
11.482 |
2.069 |
0.040 |
2.783 |
1.595 |
±SD |
5.421 |
0.137 |
1.318 |
0.152 |
0.005 |
0.226 |
0.164 |
||
V |
1000 |
Mean |
246.467 |
1.728 |
10.302 |
1.648 |
0.032 |
2.569 |
1.283 |
±SD |
2.295 |
0.065 |
1.134 |
0.169 |
0.005 |
0.323 |
0.067 |
||
VI |
1000 (Reversal) |
Mean |
301.067 |
1.812 |
10.614 |
1.890 |
0.048 |
2.612 |
1.485 |
±SD |
7.816 |
0.037 |
0.800 |
0.360 |
0.008 |
0.509 |
0.170 |
GroupNumber |
Dose (mg/kg) |
|
Heart |
Spleen |
Thymus |
Epididymides |
I |
0 |
Mean |
1.101 |
1.428 |
0.296 |
0.926 |
±SD |
0.277 |
0.521 |
0.073 |
0.114 |
||
II |
0 (Reversal) |
Mean |
1.320 |
1.159 |
0.356 |
0.933 |
±SD |
0.309 |
0.204 |
0.085 |
0.052 |
||
III |
250 |
Mean |
1.100 |
1.259 |
0.284 |
0.946 |
±SD |
0.040 |
0.169 |
0.035 |
0.118 |
||
IV |
500 |
Mean |
1.132 |
1.119 |
0.209 |
0.952 |
±SD |
0.075 |
0.147 |
0.034 |
0.132 |
||
V |
1000 |
Mean |
0.807 |
1.065 |
0.096 |
0.764 |
±SD |
0.073 |
0.370 |
0.019 |
0.095 |
||
VI |
1000 (Reversal) |
Mean |
1.112 |
1.022 |
0.414 |
1.042 |
±SD |
0.104 |
0.132 |
0.115 |
0.100 |
Sex : Female
Day : 29 and 43
GroupNumber |
Dose (mg/kg) |
|
Body Weight (g) |
Brain |
Liver
|
Kidneys
|
Adrenals
|
Ovaries
|
I |
0 |
Mean |
196.583 |
1.778 |
5.758 |
1.273 |
0.055 |
0.087 |
±SD |
13.034 |
0.058 |
0.251 |
0.092 |
0.008 |
0.021 |
||
II |
0 (Reversal) |
Mean |
210.850 |
1.715 |
7.461 |
1.497 |
0.047 |
0.097 |
±SD |
22.274 |
0.054 |
1.994 |
0.291 |
0.012 |
0.012 |
||
III |
250 |
Mean |
202.283 |
1.702 |
8.790 |
1.515 |
0.053 |
0.101 |
±SD |
15.398 |
0.121 |
1.055 |
0.147 |
0.010 |
0.016 |
||
IV |
500 |
Mean |
195.050 |
1.634 |
9.273 |
1.421 |
0.051 |
0.086 |
±SD |
9.819 |
0.096 |
0.984 |
0.147 |
0.006 |
0.021 |
||
V |
1000 |
Mean |
199.367 |
1.667 |
8.977 |
1.363 |
0.049 |
0.090 |
±SD |
13.093 |
0.085 |
1.103 |
0.230 |
0.009 |
0.018 |
||
VI |
1000 (Reversal) |
Mean |
214.050 |
1.893 |
8.813 |
1.784 |
0.063 |
0.124 |
±SD |
11.406 |
0.089 |
0.904 |
0.132 |
0.014 |
0.020 |
Group No. |
Dose (mg/kg) |
|
Heart |
Spleen |
Thymus |
Uterus |
I |
0 |
Mean |
0.719 |
0.932 |
0.238 |
0.425 |
±SD |
0.085 |
0.170 |
0.066 |
0.188 |
||
II |
0 (Reversal) |
Mean |
0.860 |
0.964 |
0.313 |
0.514 |
±SD |
0.078 |
0.270 |
0.060 |
0.087 |
||
III |
250 |
Mean |
0.798 |
0.950 |
0.215 |
0.379 |
±SD |
0.086 |
0.182 |
0.042 |
0.036 |
||
IV |
500 |
Mean |
0.832 |
0.824 |
0.216 |
0.317 |
±SD |
0.067 |
0.229 |
0.102 |
0.031 |
||
V |
1000 |
Mean |
0.737 |
0.991 |
0.358 |
0.370 |
±SD |
0.113 |
0.246 |
0.410 |
0.100 |
||
VI |
1000 (Reversal) |
Mean |
0.975 |
0.931 |
0.388 |
0.488 |
±SD |
0.057 |
0.254 |
0.049 |
0.056 |
GROUP MEAN RELATIVE ORGAN WEIGHTS (%)
Sex : Male
Day : 29 and 43
GroupNumber |
Dose (mg/kg) |
|
Body Weight (g) |
Brain |
Liver |
Kidneys |
Adrenals |
Testes |
Prostate + Seminal Vesicle with Coagulation gland |
I |
0 |
Mean |
258.267 |
0.712 |
3.524 |
0.762 |
0.017 |
1.105 |
0.640 |
±SD |
11.271 |
0.031 |
0.198 |
0.060 |
0.002 |
0.052 |
0.188 |
||
II |
0 (Reversal) |
Mean |
294.200 |
0.639 |
3.879 |
0.843 |
0.017 |
0.980 |
0.608 |
±SD |
11.583 |
0.054 |
0.472 |
0.107 |
0.002 |
0.097 |
0.085 |
||
III |
250 |
Mean |
255.883 |
0.694 |
5.576* |
0.859* |
0.014* |
1.121 |
0.690 |
±SD |
9.429 |
0.026 |
0.396 |
0.031 |
0.002 |
0.073 |
0.073 |
||
IV |
500 |
Mean |
260.067 |
0.662 |
4.416* |
0.796 |
0.015 |
1.071 |
0.613 |
±SD |
5.421 |
0.056 |
0.497 |
0.071 |
0.002 |
0.098 |
0.062 |
||
V |
1000 |
Mean |
246.467 |
0.701 |
4.180* |
0.669* |
0.013* |
1.044 |
0.520 |
±SD |
2.295 |
0.022 |
0.457 |
0.066 |
0.002 |
0.138 |
0.027 |
||
VI |
1000 (Reversal) |
Mean |
301.067 |
0.602 |
3.526 |
0.629* |
0.016 |
0.871 |
0.493* |
±SD |
7.816 |
0.023 |
0.258 |
0.126 |
0.002 |
0.188 |
0.054 |
GroupNumber |
Dose (mg/kg) |
|
Heart |
Spleen |
Thymus |
Epididymides |
I |
0 |
Mean |
0.425 |
0.551 |
0.114 |
0.359 |
±SD |
0.094 |
0.192 |
0.026 |
0.049 |
||
II |
0 (Reversal) |
Mean |
0.447 |
0.395 |
0.121 |
0.317 |
±SD |
0.093 |
0.071 |
0.028 |
0.015 |
||
III |
250 |
Mean |
0.430 |
0.491 |
0.111 |
0.369 |
±SD |
0.019 |
0.055 |
0.014 |
0.042 |
||
IV |
500 |
Mean |
0.436 |
0.430 |
0.080* |
0.367 |
±SD |
0.033 |
0.053 |
0.012 |
0.057 |
||
V |
1000 |
Mean |
0.327* |
0.432 |
0.039* |
0.310 |
±SD |
0.029 |
0.151 |
0.008 |
0.039 |
||
VI |
1000 (Reversal) |
Mean |
0.370 |
0.339 |
0.137 |
0.346 |
±SD |
0.035 |
0.040 |
0.036 |
0.029 |
Sex : Female
Day : 29 and 43
GroupNumber |
Dose (mg/kg) |
|
Body Weight (g) |
Brain |
Liver
|
Kidneys
|
Adrenals
|
Ovaries
|
I |
0 |
Mean |
196.583 |
0.907 |
2.935 |
0.650 |
0.028 |
0.044 |
±SD |
13.034 |
0.067 |
0.157 |
0.062 |
0.005 |
0.012 |
||
II |
0 (Reversal) |
Mean |
210.850 |
0.822 |
3.548 |
0.720 |
0.023 |
0.047 |
±SD |
22.274 |
0.101 |
0.894 |
0.179 |
0.008 |
0.009 |
||
III |
250 |
Mean |
202.283 |
0.848 |
4.386* |
0.755 |
0.026 |
0.050 |
±SD |
15.398 |
0.111 |
0.745 |
0.111 |
0.006 |
0.011 |
||
IV |
500 |
Mean |
195.050 |
0.839 |
4.756* |
0.730 |
0.026 |
0.044 |
±SD |
9.819 |
0.060 |
0.458 |
0.080 |
0.002 |
0.011 |
||
V |
1000 |
Mean |
199.367 |
0.838 |
4.494* |
0.680 |
0.024 |
0.045 |
±SD |
13.093 |
0.056 |
0.379 |
0.077 |
0.004 |
0.008 |
||
VI |
1000 (Reversal) |
Mean |
214.050 |
0.886 |
4.131 |
0.837 |
0.029 |
0.058 |
±SD |
11.406 |
0.062 |
0.513 |
0.095 |
0.007 |
0.012 |
Group No. |
Dose (mg/kg) |
|
Heart |
Spleen |
Thymus |
Uterus |
I |
0 |
Mean |
0.366 |
0.473 |
0.122 |
0.222 |
±SD |
0.036 |
0.075 |
0.036 |
0.113 |
||
II |
0 (Reversal) |
Mean |
0.411 |
0.468 |
0.150 |
0.245 |
±SD |
0.048 |
0.167 |
0.037 |
0.041 |
||
III |
250 |
Mean |
0.398 |
0.474 |
0.107 |
0.189 |
±SD |
0.062 |
0.109 |
0.021 |
0.030 |
||
IV |
500 |
Mean |
0.427 |
0.421 |
0.110 |
0.163 |
±SD |
0.041 |
0.109 |
0.050 |
0.014 |
||
V |
1000 |
Mean |
0.368 |
0.497 |
0.180 |
0.186 |
±SD |
0.037 |
0.126 |
0.206 |
0.050 |
||
VI |
1000 (Reversal) |
Mean |
0.457 |
0.441 |
0.181 |
0.229 |
±SD |
0.045 |
0.147 |
0.022 |
0.027 |
* = Significant at 95% level of confidence (p<0.05)
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed
- Dose descriptor:
- NOAEL
- 1 000 mg/kg bw/day
- Study duration:
- subacute
- Species:
- rat
- Quality of whole database:
- Data is Klimisch 1 and from study report
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: via oral route;
Study report was reviewed to determine the toxic nature of target substance N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8)IUPAC Name; N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine upon repeated exposure by oral route. The study is as mentioned below:
Repeated dose oral toxicity study was conducted by Sustainability Support Services (Europe) AB (SPL/122/130, 2018) to determine the oral toxic nature of N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) . In a Repeated Dose 28-day Oral Toxicity study, Sprague Dawley male and female rats were treated with 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the concentration of 0, 250 mg/kg, 500 mg/kg and 1000 mg/kg body weight orally by gavage in corn oil. Two additional dose levels were added to the study as control 0 mg/kg (Rev.) and test item 1000 mg/kg (Rev.), in order to study the reversibility or delayed occurrence of symptoms, if any. No effect on survival, clinical sign, body weight, Feed intake, Ophthalmoscopic examination and functional observation battery of treated male and female rats at the end of dosing period of 28 days and the recovery period of 14 days. Similarly, in male animals at the end of the recovery period on day 43, revealed statistically significant increase in the values of MCHC and statistically significant decrease in the values of HCT at 1000 mg/kg and in female on day 29, revealed statistically significant increase in the values of Platelets at 250 mg/kg, 500 mg/kg and 1000 mg/kg, and statistically significant decrease in the values of Hb, HCT and MCHC at 250 mg/kg, 500 mg/kg and 1000 mg/kg, and Total RBC at 500 mg/kg and 1000 mg/kg, Haematological analysis in female animals conducted at the end of the recovery period on day 43, revealed statistically significant decrease in the values of Total RBC at 1000 mg/kg. In male and female animals at the end of the dosing period on day 29, revealed statistically significant increase in the values of Total Bilirubin at 1000 mg/kg and Creatinine at 250 mg/kg, in male, Total Protein and Aspartate Aminotransferase at 250 mg/kg and 500 mg/kg, Total Bilirubin at 250, 500 and 1000 mg/kg, and Albumin at 500 mg/kg in female rat. In addition, statistically significant decrease was observed in the values of Glucose at 500 mg/kg, Calcium at 500 mg/kg and 1000 mg/kg, Globulin and Bile Acid at 250 mg/kg in male rat and, Calcium at 250 and 500 mg/kg, and Triglycerides at 250 mg/kg in female rat. At the end of the recovery period on day 43 (Reversal groups) statistically significant increase were observed in the values of Total Cholesterol at 1000 mg/kg in male rat and Total Protein at 1000 mg/kg in female rat. In addition, statistically significant decrease was observed in the values of Glucose at 1000 mg/kg in male rat. The increase/decrease in the values of various parameters was marginal and within the normal biological and laboratory limits for hematology and clinical chemistry parameters. In Urine analysis conducted during 4th and 6th week of dosing period (on day 23, 24, 25, 26 and 43), revealed no abnormality attributable to the treatment except for higher volume of urine was observed in female animals from 250 mg/kg dose group and considered to be incidental and of no toxicological importance. In addition, at termination of dosing on day 29, male animals from 250, 500 and 1000 mg/kg dose groups revealed increased relative weights of liver when compared with that of controls. In addition, increased relative weights of kidneys were observed in male animals from 250 mg/kg dose group, when compared with that of controls. Decreased relative weights of kidneys and heart weight at 1000 mg/kg in male as compared to controls. In addition, decreased relative weights of adrenals were observed in male animals from 250 and 1000 mg/kg dose groups when compared with that of controls. Decreased relative weights of thymus were observed in male animals from 500 mg/kg and 1000 mg/kg dose groups when compared with that of controls. Organ weight data of male animals sacrificed on day 43 from 1000 mg/kg reversal group, revealed decreased relative weights of kidneys and prostate + seminal vesicle with coagulation gland as whole when compared with that of controls. At termination of dosing on day 29, female animals from 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups revealed increased relative weights of liver. Organ weight data of female animals sacrificed on day 43 from 1000 mg/kg reversal group, was found to be comparable with that of controls. Although significant changes in the values of organ weight were observed in male and female animals from different dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance. Test item coloured perianal region externally and test item coloured stomach mucosa in male and female animals at 250, 500 and 1000 mg/kg dose groups. Gross pathological examination in male and female animals from control, control reversal and 1000 mg/kg reversal dose groups did not reveal any abnormality. Histopathological examination did not reveal any abnormality attributable. Therefore, No Observed Adverse Effect Level (NOAEL) of 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 1000 mg/kg body weight in male and female animals.
Repeated inhalation study:
According to Annex IX of the REACH regulation, testing by the inhalation route is appropriate only if exposure of humans via inhalation is likely. Taking into account the low vapour pressure of the substance N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8)IUPAC Name; N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine which is reported as 1.117592e-12 mmHg at 25 C. Thus, exposure to inhalable dust, mist and vapour of the chemical N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine is highly unlikely. Therefore this study is considered for waiver.
Repeated dermal study
The acute toxicity value for N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8)IUPAC Name; N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine (as provided in section 7.2.3) is in range of >2000 – 7100 mg/kg body weight. Also, given the use of the chemical; repeated exposure by the dermal route is unlikely since the use of gloves is common practice in industries. Thus, it is expected that N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that N-(2-ethylhexyl)-1-[4-(2-phenyldiazen-1-yl)phenyl]naphthalen-2-amine exhibit repeated dose toxicity by the dermal route. Hence this end point was considered for waiver.
Based on the data available for the target chemical and its prediction, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8)does not exhibit toxic nature upon repeated exposure by oral, inhalation and dermal route of exposure and hence is not likely to classify as per the criteria mentioned in CLP regulation.
Justification for classification or non-classification
Based on the above studies on 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8), it can be concluded that 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) is nontoxic to repeated dose orally at 1000 mg/kg bw. Thus, comparing this effect with the criteria of CLP regulation, 2-Naphthalenamine, N-(2-ethylhexyl)-1-[[4-(phenylazo)phenyl]azo]-, ar’ and ar’’-Me derivs (CAS No. 92257-28-8) can be not classified for repeated dose oral toxicity.
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