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Diss Factsheets

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: well performed GLP and OECD guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report date:
1992

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2,5-dimethoxyaniline
EC Number:
203-040-9
EC Name:
2,5-dimethoxyaniline
Cas Number:
102-56-7
Molecular formula:
C8H11NO2
IUPAC Name:
2,5-dimethoxyaniline
Constituent 2
Reference substance name:
2,5-Dimethoyaniline
IUPAC Name:
2,5-Dimethoyaniline
Details on test material:
- Name of test material (as cited in study report): Aminohydrochinondimethylether

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: HOECHST AG breeding colony
- Age at study initiation: approx. 6 weeks
- Weight at study initiation: day 1, male, mean 130.8 g
- Weight at study initiation: day 1, female, mean 130.2 g
- Fasting period before/during study: only for the period in which the animals were kept in diuresis cages
- Housing: in fully air-conditioned rooms in makrolon cages on soft wood granulate in groups of 5 animals
- Diet (e.g. ad libitum): Altromin 1324 rat diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: approx. 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +-3
- Humidity (%): 55 +-20
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: sesame oil (Oleum sesami, Ph. Eur. III)
Details on oral exposure:
Group 1 dose 0.0 mg/kg bw/d (5 males and 5 females)
Group 2 dose 10.8 mg/kg bw/d (5 males and 5 females)
Group 3 dose 54.0 mg/kg bw/d (5 males and 5 females)
Group 4 dose 270.0 mg/kg bw/d (5 males and 5 females)

28 applications within 29 days, 7 days/week
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
recovery (found concentration in % of nominal): 86.3 - 108
Duration of treatment / exposure:
28 administrations within 29 days, necropsy at day 29
Frequency of treatment:
oral gavage daily for 28 days (necropsy at day 29)
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
Dose: 0.0 mg/kg bw/d, Concentration: 0.0 % (w/v)
Basis:
other: nominal in sesame oil
Remarks:
Doses / Concentrations:
Dose: 10.08 mg/kg bw/d, Concentration: 0.216 (w/v)
Basis:
other: nominal in sesame oil
Remarks:
Doses / Concentrations:
Dose: 54.0 mg/kg bw/d, Concentration: 1.080 % (w/v)
Basis:
other: nominal in sesame oil
Remarks:
Doses / Concentrations:
Dose: 270.0 mg/kg bw/d, Concentration: 5.400 % (w/v)
Basis:
other: nominal in sesame oil
No. of animals per sex per dose:
5 males and 5 females per dose (0.0, 10.8, 54.0 and 270.0 mg/kg bw/d)
Control animals:
yes, concurrent vehicle
Positive control:
no

Examinations

Observations and examinations performed and frequency:
Behaviour and state of health
The behaviour and general health condition of the animals were observed twice daily, and at weekends and on public holidays once daily. The animals were examined weekly for neurological disturbances, opacity of the refracting media of the eyes, damage to the oral mucosa and impairment of dental growth.

Body weight
The body weights of all animals were determined at the start of the study and then twice weekly throughout the study.

Food and water consumption
Food consumption was determined continuously (2 times per week). Values were calculated for food consumption per 100 g body weight over a 24 hour period. Water consumption was determined once weekly over a period of 16 hours and is given in the results as water consumption / animal / 16 h (from 3:15 p.m. to 7:15 a.m.)

Haematological Investigations
At the termination of the study, haematological examinations were performed on all animals without previous withdrawal of food. Blood samples were taken from the retroorbital venous plexus in narcosis (intraperitoneal injection of 50 - 100 mg Ketanest® / kg body weight). In order to prevent systematic errors, blood sampling was conducted in a randomised order.
Haematological parameters determined: Erythrocyte count, Haemoglobin, Haematocrlt, Mean cellular volume (MCV), Mean cellular haemoglobin (MCH), Mean cellular haemoglobin concentr. (MCHC), Leucocyte count, Thrombocyte count, Differential leucocyte count and red cell morphology, Reticulocyte count*, Heinz bodies*, Coagulation time (* only in the animals of the control and high dose group).

Clinical Chemistry
After blood sampling for haematological testing, the animals were killed by cutting of the vena cava cranialis.
Serum values determined: Sodium, Potassium, Anorganic phosphorus, Uric acid, Bilirubin total, Bilirubin direct, Creatinine, Serum-glucose, Urea nitrogen, Calcium, Chloride, Aspartate aminotransferase (ASAT/GOT), Alanine aminotransferase (ALAT/GPT), Alkaline phosphatase (AP), Gamma-glutamyltransferase (GGT), Total protein, Albumin

Urine analysis
Urine analysis was performed on all animals a few days before termination of the study.
Parameters checked: Appearance, Colour , pH-Value, Haemoglobin, Protein, Glucose, Ketone bodies, Bilirubin, Urobilinogen, Specific Weight, Sediment, Volume

Sacrifice and pathology:
Necropsy and macroscopic examination
After exsanguination, all animals were necropsied and checked for macroscopically visible abnormalities. The autopsy included macroscopic examination of the skin, orifices, eyes, teeth, oral mucosa and internal organs. All abnormal findings were recorded.

Organ weights
The following organs were weighed and the organ to body weight ratio calculated: Heart, Lung, Liver, Kidneys, Adrenals, Spleen, Brain, Thymus, Testes, Epididymis, Ovaries

Histopathology
The following tissues or organs (or pieces of them) were preserved and processed for histopathological investigations: Heart, Liver, Kidneys, Adrenals, Spleen, Lung, Brain, Thymus, Ovaries, Testes, Epididymides, Trachea, Stomach, Jejunum, Colon, Urinary bladder, Uterus, Prostata, Seminal vesicles, Skeletal muscle, N. ischiadicus, Femur with bone marrow, Spinal cord (cervical, thoracal and lumbal), Lymph nodes (cervical and iliacal)
Statistics:
The following parameters were compared statistically with the control group values at the level of significance p = 0.05:

Body weights at the designated measurement times
Haematological data
Clinical chemistry parameters
Urinary parameters (volume, pH-value and specific weight)
Absolute organ weights and organ to body weight ratios

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No mortality. In the high dose group: stilted gait, squatting posture and increased salivation
Mortality:
mortality observed, treatment-related
Description (incidence):
No mortality. In the high dose group: stilted gait, squatting posture and increased salivation
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
slightly increased in the high dose group
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
high dose group: decreases in erythrocyte counts, haemoglobin and haematocrit values; reticulocyte counts were slightly increased in males
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
high dose group: Increases in urea levels and decreases in phosphate values in males, additionally increases in bilirubin values in females
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
Intermediate dose group: Urine was discoloured brownish-yellow to brownish. High dose group: brownish discolouration of the urine. Additionally epithelial cells were detected by examination of the urine sediment.
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
High dose group: Liver to body weight ratios were slightly increased
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
acute renal tubular necrosis in the high dose group and in two animals of the intermediate dose group, the severity of the change being dose-dependent. No changes in the other organs.
Histopathological findings: neoplastic:
not specified

Effect levels

Dose descriptor:
NOEL
Effect level:
10.8 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: kidney damage, epithelial cells in urine sediment, anaemia

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In conclusion repeated oral application of 270 mg Aminohydrochinondimethyl ether/ kg body weight caused kidney damage in male and female Wistar rats, reflected in histopathological alterations as well as frequent occurrence of epithelial cells in the urine sediment. Increases in urea levels and decreases in phosphate values might be interpreted in connection with kidney damage. Additionally the application of the test substance caused anaemia in both sexes.
After repeated application of 54 mg Aminohydrochinondimethylether / kg body weight kidney damage was still observed in two animals by histopathological examination. Aminohydrochinondimethylether did not cause any changes in normal, not previously altered kidneys or any other effect when administered at the dose of 10.8 mg/kg body weight per day for a period of 28 times during 29 days. With regard to this, in the present study the "no observed effect level" (NOEL) is 10.8 mg/kg body weight per day.
Executive summary:

Groups of 5 male and 5 female Wistar rats received Aminohydrochinondimethylether by oral gavage at dose levels of 0.0, 10.8, 54.0 or 270.0 mg/kg body weight per day for 28 days and were necropsied at day 29. Behaviour and state of health were observed daily in all groups. Body weights and food consumption were recorded twice weekly, and water consumption once weekly. Haematological examinations and clinical chemistry were carried out at the termination of the study. Urine analysis was also performed at the end of the study. During necropsy the animals were examined for macroscopically visible abnormalities, the main organs were weighed and the organ to body weight ratios were calculated. A range of organs and tissues were processed for histopathological examination and checked for microscopically visible changes. Body weights, haematological and clinical chemistry data, urine data (volume, specific weight and pH-value), absolute and relative organ weights were analysed with the aid of a statistical program to show differences compared with the controls.

Behaviour, state of health, body weight development, food and water consumption remained unaffected by the administration of the test compound in the low and intermediate dose group. Clinical signs were observed in the high dose group, consisting of stilted gait, squatting posture and increased salivation. Body weight development and food consumption were normal in the high dose group. Relative water consumption was slightly increased in both sexes of the high dose group.

Haematological examinations revealed decreases in erythrocyte counts, haemoglobin and haematocrit values in both sexes of the high dose group. Additionally, reticulocyte counts were slightly increased in males of the high dose group. Clinical chemistry investigations revealed increases in urea levels and decreases in phosphate values in males of the high dose group Additionally increases in bilirubin values were observed in females of the high dose group.

Urine was discoloured brownish-yellow to brownish in both sexes of the intermediate dose group. Animals of the high dose group showed intensively brownish discolouration of the urine. Additionally epithelial cells were detected frequently by examination of the urine sediment in the latter group.

Liver to body weight ratios were slightly increased in both sexes of the high dose group.

No compound related changes were observed at necropsy.

Histopathological examinations revealed acute renal tubular necrosis in nine animals of the high dose group and In two animals of the intermediate dose group, the severity of the change being dose-dependent. This renal change was also observed in one animal of the low dose group, but differed from those of the other treatment groups as far as only one kidney, exhibiting a preexisting spontaneous change, was involved. Possibly the nephrotoxic threshold of the test compound was lowered in this previously altered organ. No compound-related changes were observed in the other organs.