Registration Dossier

Administrative data

Description of key information

The key study for repeated oral toxicity was performed similar to OECD TG 408 (BASF SE, 1982). Young adult male and female rats were treated with the test substance orally via the drinking water. A LOAEL of 7.5 mg/kg bw/d was determined based on changes in the thyroid in male and female animals at the lowest dose level (slight stimulation of the thyroidal follicular epithelium). Target organ systems were liver, thyroid and the urinary system. The analysis of hematology parameters revealed treatment-related anemia in both sexes. The histopathological findings in the liver were accompanied by elevated serum levels of liver specific enzymes and other parameters related to hepatotoxicity.

The test report is supported by two studies (NTP, 1983) performed similar to OECD TG 408 in young adult rats (male/female) or adult mice, respectively. After 90 d repeated dosing a NOAEL of 7.1 mg/kg bw/d was determined in rats and a NOAEL of 11.4 mg/kg bw/d was determined in mice. In both species, target organs were the hepatobiliary system and the thyroid system, the latter being more pronounced in the rat.

Chronic toxicity study data in rats and mice are summarized in endpoint section 7.7 (Carcinogenicity). These data revealed histopathological findings in the liver and thyroid of both species, with rats being more sensitive than mice (NTP, 1983). Additional study information is available for repeated oral dosing in rats (Gohlke et al., 1978; Tullner et al., 1966; Dow Chemical Company, 1955; Miyamoto et al., 1977; Pludro et al., 1969; Fukushima et al., 1979), cats (Hofmann et al., 1966) and dogs (Deichmann et al., 1978; Dow Chemical Company, 1955). The outcome of these subacute and subchronic studies corroborate the findings of the key study.

For inhalative toxicity only a short-term toxicity study in male guinea-pigs was available with limited reliability (only one concentration was applied) and confirming the systemic toxicity of the test substance (Leong et al., 1987). All animals showed degeneration of the retina. No treatment related alterations in liver and kidney tissue were observed.

The key study for repeated dermal toxicity was performed similar to OECD TG 411 (Huntsman Polyurethanes, 1998). Young adult male and female rats were treated with the test substance under occlusive conditions. A NO(A)EL of 90 mg/kg bw/d was determined. No treatment related systemic toxicity and no target organ systems were spotted.

The outcome of the key study was supported by the 21 d repeated dermal dosing study in rats (Huntsman Polyurethanes, 1997). Additional study information for repeated dermal dosing in other species is available. The 14 d dermal dosing of mice (Oak Ridge National Laboratory, 1987) showed test substance related mortality and changes in organ weights (spleen, liver). Repeated dermal dosing of rabbits (10 days) did not reveal any relevant changes in clinical chemistry parameter, organ weights or histopathology (Haskell Laboratory, 1975).

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
not specified
GLP compliance:
no
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Lot/batch No.of test material: 607212
- Physical state: solid
- Analytical purity: > 98 %
- Purity test date: July 24, 1980

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: stable


Species:
rat
Strain:
other: RAIf (F3-hybrid of RII 1/Tif x RII 2/Tif) (Specified pathogen free)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Animal Production, Ciba-Geigy LTD., 4332 Stein, Switzerland
- Age at study initiation: 4 weeks
- Weight at study initiation: 88 - 91 g (males); 90 - 91g (females)
- Fasting period before study: not specified
- Housing: Animals housed in groups of 5 in Macrolon cages type 4.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 10
- Air changes (per hr): 15 -17
- Photoperiod (hrs dark / hrs light): 14/10

IN-LIFE DATES: From: August 11, 1980 To: December 15, 1980
Route of administration:
oral: drinking water
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:

- Rate of preparation (frequency): daily
- Mixing appropriate amounts with: water
- Storage temperature of food: ambient
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Pretest samples were analysed for concentration and stability of the test material. Samples taken at random from the prepared dosing solutions were analysed for concentration in the Central Analytical Laboratories of Ciba-Geigy LTD., Basle, Switzerland.
Duration of treatment / exposure:
90 days
Frequency of treatment:
continuous
Dose / conc.:
0 mg/L drinking water
Dose / conc.:
80 mg/L drinking water
Remarks:
Due to the small variation between nominal and actual measured concentrations, nominal values were used in calculation of test material intake.
Basis: nominal in water
Dose / conc.:
400 mg/L drinking water
Remarks:
Due to the small variation between nominal and actual measured concentrations, nominal values were used in calculation of test material intake.
Basis: nominal in water
Dose / conc.:
800 mg/L drinking water
Remarks:
Due to the small variation between nominal and actual measured concentrations, nominal values were used in calculation of test material intake.
Basis: nominal in water
No. of animals per sex per dose:
80 males and 80 females per group
Control animals:
yes, concurrent vehicle
Details on study design:
- Rationale for animal assignment: random
- Rationale for selecting satellite groups: random
- Post-exposure recovery period in satellite groups: 4 weeks
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: daily

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: week 15 (after 30 days exposure) and after 1 month recovery
- Anaesthetic used for blood collection: No
- Animals fasted: Yes (20 hours fasting)
- How many animals: 10 rats/sex/group
- Parameters checked: Erythrocytes, Leukocytes, Haemoglobin, Haematocrit, Reticulocytes, Thrombocytes, MCV, MCH, Differential, prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: week 15 (after 30 days exposure) and after 1 month recovery
- Animals fasted: Yes (20 hours fasting)
- How many animals: 10 rats/sex/group
- Parameters checked: Glucose, urea nitrogen, sodium, potassium, calcium, ASP aminotransferase (GOT), ALT aminotransferase (GPT), alkaline phosphatase, creatinine, bilirubin, cholesterol, total proteins, protein electrophoresis

URINALYSIS: Yes
- Time schedule for collection of urine: week 15 (after 30 days exposure) and after 1 month recovery
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked: Specific gravity, pH, chemical findings test

NEUROBEHAVIOURAL EXAMINATION: No

OTHER: Hearing tests
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Statistics:
For each time point and parameter, a uni-variate statistical analysis was conducted. Each treated group was compared to control group in respect of dispersion and displacement. In addition, a trend test was applied considering all groups.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
effects observed, treatment-related
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
No clinical symptoms and no signs of local and/or systemic toxicity were observed, except that the animals of groups 3 and 4 (400 and 800 ppm) failed to gain weight.

BODY WEIGHT AND WEIGHT GAIN
The mean body weight of male and female groups 3 and 4 (400 and 800 ppm) was markedly depressed, whereas the mean body weight of male and female group 2 (80 ppm) was similar to that of the respective controls. After replacement of the medicated water with ordinary drinking water male and female groups 3 and 4 (400 and 800 ppm) did not recover essentially from this weight depression during a 4 weeks recovery period.

WATER CONSUMPTION AND COMPOUND INTAKE
The mean water consumption of male and female groups 3 and 4 (400 and 800 ppm) was markedly depressed, whereas the mean water consumption of male and female group 2 (80 ppm) was similar to that of the respective control groups. After replacement of the medicated drinking water the water consumption of male and female groups 3 and 4 (400 and 800 ppm) increased slowly to the amount consumed by the respective control groups towards the end of the 4 weeks recovery period.

HAEMATOLOGY
The males and females of group 3 and 4 (400 and 800 ppm) showed reduced numbers of erythrocytes, a reduced haemoglobin concentration and a lower haematocrit still present at the end of the recovery period.
The mean cell haemoglobin was increased in males of group 3 and 4 (400 and 800 ppm) at the end of the recovery period.
The mean cell volume was increased in the males of group 3 and 4 (400 and 800 ppm) at the end of the recovery period.
The number of reticulocytes in females of group 3 and 4 (400 and 800 ppm) was increased. The number of reticulocytes was increased in the males of group 3 and 4 (400 and 800 ppm) and in females of group 4 (800 ppm) at the end of the recovery period.
The number of leucocytes was increased in the males and females of group 4 (800 ppm). A slight increase of the number of the segmented neutrophils (relative) was observed in the males and females of group 4 (800 ppm) and in the males of group 3 and 4 (400 and 800 ppm) at the end of the recovery period. A decrease in the number of lymphocytes (relative) was observed in the females of group 4 (800 ppm). The prothrombin time was prolonged in the males and females of group 4 (800 ppm).

CLINICAL CHEMISTRY
The activity of the alkaline phosphatase was increased in the males and females of group 3 and 4 (400 and 800 ppm) at the end of the treatment period as well as at the end of the recovery period.
The alanine aminotransferase (GPT) activity was slightly increased in males and females of group 3 and 4 (400 and 800 ppm). A similar increase of the alanine aminotransferase (GPT) activity was observed in the males of group 4 (800 ppm) at the end of the recovery period.
The aspartate aminotransferase (GOT) activity was increased in the males and females of group 3 and 4 (400 and 800 ppm) and in the males of group 3 and 4 (400 and 800 ppm) at the end of the recovery period.
The urea-nitrogen-concentration was increased in the males and females of group 3 and 4 (400 and 800 ppm) and in males of group 4 (800 ppm) at the end of the recovery period.
The bilirubin and cholesterin concentrations were increased in the males and females of group 3 and 4 (400 and 800 ppm).
The total protein concentration was slightly increased in the males of group 3 and 4 (400 and 800 ppm). The albumin fraction (relative) was increased in the females of group 2 and 4 (80 and 800 ppm). The A1 globulin fraction (relative) was decreased in the males of group 3 and 4 (400 and 800 ppm). The beta-globulin fraction (relative) was increased in the males of group 3 and 4 (400 and 800 ppm). The beta-globulin fraction (relative) was decreased in the females of group 2 and 4 (80 and 800 ppm). The gamma-globulin fraction (relative) was decreased in the females of group 4 (800 ppm).

URINALYSIS
The findings in the urine were generally unremarkable and comparable to those of the control animals.

ORGAN WEIGHTS
Corresponding to the lower body weight in male and female groups 3 and 4 (400 and 800 ppm) the absolute organ weights of the animals in these groups were lower than in the respective control groups at the end of the treatment period. This influences also the organ weight ratios in that in male and female groups 3 and 4 (400 and 800 ppm) the organ to brain weight ratio was lower and the organ to body weight ratio was higher than in the respective control groups. No marked improvement in absolute and relative organ weights of males and females of group 3 and 4 (400 and 800 ppm) was observed at the end of the recovery period.

HISTOPATHOLOGY: NON-NEOPLASTIC
Histopathological examination revealed the following findings in animals of group 4 (800 ppm):
In all animals marked biliary liver lesions and goitrogeneous effects on the thyroid were apparent. All histologically examined treated male and female rats showed moderate or marked hyperplasia of small biliary ducts with initial fibrosis in the peripheral parts of the liver lobules. These liver changes persisted also after the recovery period. Clear fluid in the abdominal cavity was found in one male and in 4 females. Slight or moderate oedema in the interstitium of the pancreas was seen microscopically in 5 males and 3 females. In nearly all males (18/20) and in all females slight, moderate or marked hypertrophy of the thyroid follicular epithelial cells and diffuse hyperplasia of the glandular structures with marked colloid depletion was noted. In one male rat also focal nodular hyperplasia with reaccumulation of the colloid in some distended follicles developed. After the recovery period the thyroid stimulation was much less pronounced. Only in 3/10 males and 2/10 females slight stimulation of the follicular epithelium was still evident.

Histopathological examination revealed the following findings in animals of group 3 (400 ppm):
All the males and the majority of females (16/20) showed slight or moderate hyperplasia of small biliary ducts. "The severity of these microscopical liver lesions was similar in rats sacrificed after the recovery period. In 4/20 males and in 17/20 females slight or moderate stimulation of the follicular epithelium in the thyroids was noted. In one male and one female also focal nodular hyperplasia of the thyroid parenchyma with colloid reaccumulation in some follicles developed. No compound related thyroid changes were noted after the recovery period, however.

Histopathological examination revealed the following findings in animals of group 2 (80 ppm):
No compound related liver lesions were noted. However in 2/20 males and in 2/20 females slight stimulation of the follicular epithelium in the thyroids was observed upon histopathological examination.
OTHER FINDINGS
Whereas nephrocalcinosis was evident in all females of treatment and control groups, mineralisation was seen in all males of the mid dose group and in 21 of 30 males of the high dose groups. One male of the 80 ppm group and none of the control males showed kidney mineralisation.

Simple hearing tests performed at the beginning of the treatment period and towards the end of the recovery period revealed no treatment related effects on the auditory perception.
Key result
Dose descriptor:
LOAEL
Effect level:
7.5 mg/kg bw/day (nominal)
Sex:
male
Basis for effect level:
histopathology: non-neoplastic
Remarks on result:
other: slight stimulation of the thyroidal follicular epithelium
Dose descriptor:
LOAEL
Effect level:
8 mg/kg bw/day (nominal)
Sex:
female
Basis for effect level:
histopathology: non-neoplastic
Remarks on result:
other: slight stimulation of the thyroidal follicular epithelium
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
22 mg/kg bw/day (nominal)
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
22 mg/kg bw/day (nominal)
System:
endocrine system
Organ:
thyroid gland
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
22 mg/kg bw/day (nominal)
System:
urinary
Organ:
kidney
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LOAEL
7.5 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
similar to OECD TG 408
Organ:
kidney
liver
thyroid gland

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

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
4th November 1997 - 22nd May 1998
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)
Version / remarks:
1981
Qualifier:
according to
Guideline:
EU Method B.28 (Sub-Chronic Dermal Toxicity Test: 90-Day Repeated Dermal Dose Study Using Rodent Species)
Version / remarks:
1988
Qualifier:
according to
Guideline:
other: TSCA guideline reference CFR 798.2250
Version / remarks:
draft June 1996
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: Ex Fluka, 352776-1796
- Purity: 97.4%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Ambient temperature in the dark
Species:
rat
Strain:
other: Alpk:APrSD
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Rodent Breeding Unit, Zeneca Pharmaceuticals, Alderley Park
- Weight at study initiation: 120 - 145 g (Males), 100 - 125 g (Females) upon arrival at CTL
- Fasting period before study: No
- Housing: The rats were individually housed in multiple rat racks suitable for animals of this strain and weight range expected during the course of the study.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 11 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): At least 15
- Photoperiod (hrs dark / hrs light): Artificial light 12/12
Type of coverage:
occlusive
Vehicle:
ethanol
Details on exposure:
TEST SITE
- Area of exposure: at least 7 cm x 7 cm of the dorso-Iumbar skin.
- % coverage: ca. 10 %
- Type of wrap if used: Each dressing consisted of a 4 ply gauze patch (approximately 7 cm x 7 cm) to cover the treated area. This patch was covered by a patch of plastic film held in position by an adhesive bandage (approximately 25 cm x 5 or 7.5 cm) around which were wrapped 2 pieces of PVC tape.
- Time intervals for shavings or clipplings: Sixteen to thirty-two hours before the first application of the test substance

REMOVAL OF TEST SUBSTANCE
- Washing: warm water
- Time after start of exposure: 6 hours

TEST MATERIAL
- Amount applied: 2 mL/kg bodyweight
- Constant volume or concentration used: no
- For solids, paste formed: no

VEHICLE
- Lot/batch No: Y00332/005

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The dose preparations were analysed to confIrm satisfactory achieved concentration at all dose levels. This analysis was performed twice during the study. The stability of the test substance in the vehicle at room temperature was evaluated for the low and high dose solutions.
Duration of treatment / exposure:
The animals were dosed dermally, for a total of 50 applications (4-6 applications per 7 day period), over a period of 70 days.
Frequency of treatment:
4 - 6 applications per 7 day period
Dose / conc.:
0 mg/kg bw/day
Remarks:
Group 1
Dose / conc.:
3 mg/kg bw/day
Remarks:
Group 2
Dose / conc.:
30 mg/kg bw/day
Remarks:
Group 3
Dose / conc.:
60 mg/kg bw/day
Remarks:
Group 4
Dose / conc.:
90 mg/kg bw/day
Remarks:
Group 5
No. of animals per sex per dose:
10 males and 10 females
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: at the time of dosing and after decontamination on each application day

DERMAL IRRITATION: Yes
- Time schedule for examinations: at the time of dosing and after decontamination on each application day

BODY WEIGHT: Yes
- Time schedule for examinations: each day of dosing throughout the study

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Yes

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: The eyes of all animals were examined pre-experimentally and those of the control and high dose group animals during the week of termination.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At study termination
- Anaesthetic used for blood collection: No
- Animals fasted: No
- How many animals: all animals
- Parameters examined: haemoglobin, red cell count, mean cell volume, mean cell haemoglobin concentration, red cell distribution width, total and differential white blood cell count, platelet count, blood cell morphology, mean cell haemoglobin, haematocrit

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At study termination
- Animals fasted: No
- How many animals: all animals
- Parameters examined: urea, creatinine, glucose, total protein, triglycerides, potassium, calcium, total bilirubin, creatine kinase activity, alkaline phosphatase activity, alanine aminotransferase activity, albumin, albumin/globulin ratio, cholesterol, sodium, chloride, phosphorus (as phosphate), gamma-glutamyl transferase activity, aspartate aminotransferase activity

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals were subjected to a full examination post mortem. This involved an external observation and a careful internal examination of all organs and structures.

HISTOPATHOLOGY: Yes
Abnormal tissues, liver, thyroid and skin (treated and untreated) were processed from all animals. The kidneys and lungs were processed from the control and high dose group animals only.
Statistics:
All data were evaluated using the GLM procedure in SAS (1989).
Clinical signs:
effects observed, non-treatment-related
Dermal irritation:
effects observed, treatment-related
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, non-treatment-related
Urinalysis findings:
not specified
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Gross pathological findings:
effects observed, treatment-related
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
CLINICAL SIGNS AND MORTALITY
It was noted that some animals on some days of dosing exhibited signs of distress whilst bandaged for 6 hours. Consequently, alternative types of dressings, bandages and methods to retain the test substance were evaluated to try and alleviate the distress of the animals. As the alternative methods did not significantly reduce the stress experienced by some of the animals, the study was terminated early on day 71.
There were four early terminations unrelated to treatment with the test item. In addition, one male rat (30 mg test item/kg body weight/day group) was killed for humane reasons on day 11 since necrotic skin was noted on the area of application. With regards to general clinical observations, there were no toxicologically relevant findings at any dose level in either males or females treated with the test item.

BODY WEIGHT AND WEIGHT GAIN
Treatment of male and female rats had no significant effects on bodyweights throughout the study.

FOOD CONSUMPTION
Food consumption was slightly lower in female rats in the 90 mg /kg bodyweight/day group, and the maximum effect was 7 % below control values. This small effect is considered to be of little toxicological relevance.

FOOD EFFICIENCY
There was no effect of treatment on food utilisation.

OPHTHALMOSCOPIC EXAMINATION
The administration of 90 mg /kg bodyweight/day to both male and female rats for 5 days per week for 70 days had no effect on the appearance of the eyes as evaluated by ophthalmoscopy

HAEMATOLOGY
Overall, there were no treatment related findings on any haematological parameters evaluated.

CLINICAL CHEMISTRY
In the high dose group, plasma cholesterol was increased in male rats, plasma albumin to globulin ratio was decreased in males, and plasma levels of glucose, albumin and total protein were significantly decreased in female rats which may reflect the lower food consumption in this group. All of these changes are minimal and are considered to be of no toxicological significance. Certain other parameters were also statistically significantly changed in either male or female rats, but as they only occurred in the 2 lowest groups, it is considered that they are unrelated to treatment.

ORGAN WEIGHTS
Kidney weights for the males in the high dose group were slightly lower than the control values. However, in the absence of pathological findings, this small decrease is considered to be of little toxicological significance. There were no other effects of treatment on organ weights in either male or female rats.

GROSS PATHOLOGY
There were treatment-related macroscopic findings in the treated skin. There was an increased incidence of scabs in males and females treated with 30, 60 or 90 mg/kg bodyweight/day dose groups, and discoloration of the skin in males from all treatment groups and in females treated with 30, 60 or 90 mg/kg bodyweight/day dose groups. In addition, a small number of animals from treated groups had scabs in the untreated skin. There were no other treatment-related macroscopic findings.

HISTOPATHOLOGY: NON-NEOPLASTIC
There were treatment-related microscopic findings in the treated skin of rats killed intercurrently and at termination. There was an increased incidence of lesions which were considered to represent dermatitis in males and females treated with 30, 60 or 90 mg/kg bodyweight/day.
No treatment-related effects were seen in any of the other tissues examined.
Key result
Dose descriptor:
NOEL
Remarks:
systemic toxicity
Effect level:
90 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: highest test dose
Dose descriptor:
NOEL
Remarks:
skin lesions
Effect level:
3 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: scabs and discoloration of the skin
Key result
Critical effects observed:
no
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
90 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
similar to OECD TG 411

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
adverse effect observed

Additional information

Oral Toxicity

Chronic Toxicity

Further to the below summarized data on subchronic and short-term toxicity, chronic toxicity data are available for rats and mice (NTP, 1983). The thyroid and liver were the most relevant target organs after repeated administration, rats being more sensitive than mice. Please refer to the carcinogenicity section 7.7 for details.

Subchronic Toxicity

The key study was performed similar to OECD TG 408. Young adult (4 weeks old) RAIf rats (male/female) were treated orally with MDA via drinking water for 90 d (80, 400, or 800 ppm). The determined LOAEL of 7.5 mg/kg bw/d (male) is based on slight stimulation of the thyroidal follicular epithelium. Animals of the high and mid dose group showed statistically significant decreases in body weight and no essential recovery was observed during the 4 week recovery period. In the high and mid dose groups significant changes in hematology parameter were observed, i.e. decrease in red blood cells, reduction in hemoglobin and hematocrit and correspondingly increases in MCV, MCH and reticulocytes, as well as a increased number of neutrophils. In addition, a decrease in lymphocytes and a prolonged prothrombin time was found at the highest dose level. The clinical chemistry parameter were significantly changed in the high and mid dose group as follows: elevated serum concentrations of alkaline phosphatase, ALAT, ASAT, urea, bilirubin, cholesterol. After the recovery period alkaline phosphatase, ASAT (males) and urea remained at higher levels. Male animals showed higher concentrations of total protein with a decreasing trend after recovery. The potassium levels were decreased in males at the end of treatment and increased in females after the recovery period.

The changes in organ weight observed in nearly all organs in the high and mid dose group were considered to be related to the marked body weight depression. In the high and mid dose group, histopathology revealed changes in the hepatobiliary system, i.e. hyperplasia of biliary ducts, being also evident after the recovery period. Hypertrophy of thyroid follicular epithelial cells and hyperplasia were observed at the high and mid dose group. The observed findings were still evident after the recovery period at the high dose group, but less in severity. At the mid dose group none of the thyroid changes were observed after the recovery period. Nephrocalcinosis and kidney mineralization were observed in female and male rats, respectively, at the high and mid dose group.

(BASF SE, Report No TK 10504, 1982)

The result of the key study is supported by a study similar to OECD TG 408 in young adult (7 weeks) F344 rats (male/female) and adult (8 weeks) B6C3F1 mice (male/female). The test substance was administered orally via the drinking water for 90 d (rats: 50, 100, 200, 400, or 800 ppm; mice: 25, 50, 100, 200, 400 ppm). The study did not include the evaluation of hematology and clinical biochemistry parameters. A NOAEL of 100 ppm was determined in both species, being equivalent to 7.1 mg/kg bw/d was in rats (male) and 11.4 mg/kg bw/d in mice (male), respectively. The hepatobiliary system was found to be a target organ in both species (bile duct hyperplasia). Additionally, the thyroid system was a target organ in rats (follicular hyperplasia).

(National Toxicology Program (NTP), Report No NTIS/PB83-238824, 1983)

Short-term toxicity

In a short term toxicity study young adult (10 weeks) F344 rats or young adult (8 weeks) B6C3F1 mice (male and female) were treated orally via drinking water with the test substance for 14 days (200, 400, 800, 1600, or 3200 ppm). No NOAEL was determinable in rats. In mice, a NOAEL of 57 mg/kg bw/d (female) was determined.

(National Toxicology Program (NTP), Report No NTIS/PB83-238824, 1983)

A repeated dose toxicity study was performed in Sprague-Dawley rats. 4,4'-Diaminodiphenylmethan (MDA) was administered to the animals via oral gavage at doses of 0, 25 and 50 mg/kg bw/day over a period of 14 days (5 days per week). No animal died during the course of the study. Body weight development and food consumption was comparable to the untreated control group. In the 50 mg/kg bw/day dose group an increase in calcium, total lipid, bilirubin GTP, AP and leucocytes was observed in males and females. An increase in albumin was observed in males only. Decease in hemoglobin, erythrocytes and hematocrit was observed in males and females. In the 25 mg/kg bw/day dose group total lipids and AP were increased in males and females. Urinalysis revealed an increase in degenerated kidney epithelia and cylinder in the sediment in both treatment groups. Gross pathology showed hepatomegalia and splenomegaly at the highest dose group. Relative organ weights of liver, kidney, spleen and thyroid were increased in the highest dose group in males and females. Histopathological examination of the rats receiving 50 mg/kg bw/day revealed treatment-related changes in liver, cholangiofibrosis, proliferation of bile ducts and necrosis in liver. These changes were also detected in some animals of the lower dose group. Dose-dependent effects in spleen (extramedullary hematopoiesis), thyroid (hyperplasia) and kidney (cell desquamation) were reported. Application of 25 and 50 mg/kg bw/day caused adverse effects in both male and female rats.

(BASF SE, 1977a).

A repeated dose toxicity study similar to OECD TG 407 was performed in Sprague-Dawley rats. Animals were treated via feed with 0, 50 and 200 ppm 4,4'-Diaminodiphenylmethan (MDA) over a period of 28 days (doses were not reported as mg/kg bw/d) . No clinical signs were detected and no animal died during the course of the study. Body weight development and food consumption was comparable to the untreated control group. Histopathological examination of the rats receiving 50 ppm or 200 ppm revealed clear signs of a glomerulonephritis. In the 200 ppm dose group epithelial activation and parenchymatous hyperplasia of the thyroid was detected. In the liver mesenchymal activation was detected in the highest dose group. An increase in absolute and relative liver weight in female animals of the highest dose group compared to the untreated control was observed. The organ weights of liver, heart and kidneys were not changed in the 50 ppm dose group and in the 200 ppm dose group compared to the control. In the highest dose group an increase in urea, total lipids and cholesterol compared to the untreated control group was observed. A NOAEL could not be determined since animals of the lowest dose group showed glomerulonephritis.

(BASF SE, 1977b).

 

Other information

The studies summarized below did not follow a validated testing guideline (e.g. OECD TG 408) for repeated dose toxicity. In most of the studies only one dose level was used and duration or frequency differed from validated guideline. Although the documentation of the studies was rather poor the outcome of the studies supported the findings of the relevant key and supporting studies.

In a combined subacute to subchronic oral toxicity study young adult (80 d) male albino rats (strain not specified) were treated via gavage (Vehicle: peanut oil) with the test substance for 2 or 6 weeks (8 mg/kg bw/d). After 2 weeks they showed increased liver mitosis. After 6 weeks predominantly large and macronuclear hepatocytes with nuclear membrane hyperchromicity and large vesicular nuclei in the bile duct epithelium were observed. These observations were transient, sinceanimals fed with the test substance for 6 weeks and sacrificed after a post-exposure period of 2 weeks did not differ from the control animals.

(Gohlke R. et al., Z. Ges. Hyg.Grenzgebiete 24 (3): 159-62, 1978)

In a short term toxicity study young adult female castrated Sprague Dawley rats were treated via gavage (Vehicle: water) with the test substance for 5 to 14 d (150 mg/kg bw/d). There was marked adrenal, thyroid, and uterine hypertrophy. The zona fasciculata and zona reticularis of the adrenal glands showed extensive lipid accumulation. Thyroid follicles contained little or no colloid. The endometrium presented an atypical folliculoid response.

(Tullner W. et al., Endocrinology 66 Mar: 470-4, 1966)

In a combined subacute to subchronic oral toxicity study young adult (80 d) male albino rats (strain not specified) were treated via food (Vehicle: peanut oil) with the test substance for 2, 6 or 16 weeks (8 or 20 mg/kg bw/d). The animals receiving 8 mg/kg bw/d test substance showed increased liver cell and bile duct mitosis. The livers of those receiving 20 mg/kg bw/d had hyperplastic nodules, bile duct proliferation, and connective tissue changes. There was also a reduced life-span. The incidence of tumours was not increased. Based on these results, a LOAEL of 8 mg/kg bw/d was determined.

(Gohlke R. et al., Z. Ges. Hyg.Grenzgebiete 24 (3): 159-62, 1978)

In a subchronic feeding study in young (30 d) Wistar rats (male/female) the test substance was administered via feed for 90 d (30 or 100 mg/kg bw/d). The mortality was not related to the test substance dosage. Urine analyses gave normal values for sugar and protein. Periodic blood samples showed normal haematological values. The rats receiving 100 mg test substance/kg bw/d exhibited growth retardation, but their body weight gain per gram of food was practically the same as for the control group as they ate less. They also had increased liver weights, and a striking proliferation of the bile ducts. At the end of the 90 days blood analysis elevated icteric indices and bilirubin content were observed. Based on the observed effects, a NOAEL of 30 mg/kg bw/d was determined.

(Dow Chemical Company, Report No: NTIS Order No. OTS0205950, 1955)

In a subchronic feeding study in young (5 weeks) HLA-Wistar rats (male) the test substance was administered via feed for 12 weeks (1000 ppm). Treated animals showed decreased body weight gain and augmented activity of glutamic-oxalacetic transaminase, glutamic-pyruvic transaminase, and alkaline phosphatase was observed. The cholesterol and bilirubin concentrations were significantly higher than that of the controls. Histopathological examinations revealed extensive bile duct proliferation, accompanied by fibrosis, and infiltration of oval cells and inflammatory cells. Dilation and increase of smooth endoplasmic reticulum, and partial deformation of mitochondria were observed, however, these structural changes tended to disappear on cessation of the exposure.

(Miyamoto et al., J. Pesticide Sci. 2: 257-69, 1977)

In a subchronic oral study in Wistar rats (male/female) the test substance was administered in propylene glycol (vehicle) for 84 d (8.3 or 83 mg/kg bw/d). Histopathological examination revealed degeneration of the liver and spleen, and parenchymatous liver damage. Based on these results, a LOAEL of 8.3 mg/kg bw/d was determined.

(Pludro et al., Acta Polon. Pharm. XXVI (4): 353-8, 1969)

In a subchronic oral study in Wistar rats (male) the test substance was administered in corn oil (vehicle) via feed for 8, 16, 24, 32, or 40 weeks (1000 ppm). There was reduced growth in the rats during the test substance administration. The rats fed the test substance had a higher liver to body weight ratio than the control rats, and the ratio increased with the treatment period. Furthermore, transient changes in various serum enzymes were observed, but returned to (nearly) normal values after recovery. Macroscopically the liver was essentially normal for 16 weeks. After 24 weeks of feeding with the test substance the liver surface was rough and granular. There were fibrotic changes, and a marked resistance to cutting. The changes were more pronounced with longer the test substance administration. The test substance caused proliferation of bile ductular and oval cells in the liver. The hepatic parenchyma was replaced by proliferating bile ducts and cirrhosis-like inflammatory oval cell infiltration was most prominent in week 40.

(Fukushima et al., Toxicol. Appl. Pharmacol. 48: 145-55, 1979)

In a subchronic oral study in cats the test substance was administered up to 37 weeks (3 or 10 mg/kg bw/d). Treated animals exhibited liver damage with the first signs of cirrhosis as well as blood damage with anaemia. Kidney damage was observed at the high dose, but was not evident in the low dose. In some cats repeated oral doses of 10 mg/kg bw/d were fatal after 61 or 81 doses.

(Hofmann et al., Proc. Int.Kongress Arbeitsmedizin, Wien, 19-24 September 1966, 336-7, 1966)

In a chronic feeding study in young (5 – 6 months) beagle dogs (female) the test substance was administered via gelatin capsules for 47 – 86 months (70 mg/d, 3x per week; total dose 4 - 6.26 g/kg bw). The test substance was administered as purified or crude form in corn oil (vehicle). Both forms produced similar effects. Moderate to severe gross and histopathological changes in the liver were observed. The lesions ranged from enlarged liver cells and slight structural changes to degeneration, portal fibrosis, liver cell necrosis, haemosiderosis, cell infiltration of the portal areas, dilated bile ducts and thickened bile. Less severe effects were seen in the kidneys and spleen.

(Deichmann et al., Toxicology ll: 185-8, 1978)

In a subacute feeding study in dogs the test substance was administered via capsules for 47 d (0.65, 1.25, 2.5, 5, 8, or 12 mg/kg bw/d). Pathological changes were limited to the liver (cholangitis). Inflammatory changes in the tissue were observed in the absence of stasis of bile or evidence of obstruction.

(Dow Chemical Company, EPA/OTS Doc. 878211081, 1955)

Inhalative Toxicity

For inhalative repeated dose toxicity only a short-term toxicity study is available with limited reliability.

Short-term toxicity

In a short-term repeated dose toxicity study, male guinea pigs (Albino Hartley and pigmented animals) were nose-only exposed to test substance aerosol with polyethylene glycol (PEG) 200 as vehicle for 14 d (10 exposures, 4 h/d; 5 d/week) at a concentration of 0.44 mg/L air. The aerosol was generated by atomizing a solution of 200 mg test substance/mL PEG into an air elutriator. The average aerosol particle size was 2.4 µm. No visible symptoms in guinea pigs (albino and pigmented) after exposure were observed. The most prominent histopathological findings were degeneration of the inner and outer segments of the photoreceptor cells and the pigmented epithelial cell layer of the retina in both kinds of guinea pig. No histopathological changes in liver and kidney were observed.

(Leong et al., Fund. Appl. Toxicol. 9: 645-58, 1987)

Dermal toxicity

Subchronic Toxicity

The key study is a subchronic repeated dose toxicity study according to OECD TG 411. Adult Alpk:APrSD rats (male/female) were dermally exposed to the test substance (vehicle: ethanol) for 70 days (50 applications, 6 h/d) at concentrations of 3, 30, 60, or 90 mg/kg bw/d under occlusive conditions. Clinical observations (systemic including ophthalmoscopy), bodyweights, food consumption, haematological parameters, blood clinical chemistry, organ weights and histopathology of the liver, kidney, thyroid and lungs were all unaffected in both sexes by treatment with the test substance at doses up to and including 90 mg/kg bw/d. Systemically there were no treatment-related findings and the NOEL was 90 mg/kg bw/d.

(Huntsman Polyurethanes, Report No: CTL/P/5936, 1998)

A supporting dose range finding dermal toxicity study in rats corroborates the absence of effects in rats after repeated dermal exposure. Male and female animals were dosed dermally with 0, 0.5, 5 or 50 mg/kg bw/d of the test substance 5 days per week over a period of 21 days (15 applications in total). No treatment related effects on body weight, bodyweigt gain, food consumption, haematological parameters, clinical chemistry and liver pathology were observed. Only minor skin damage was noted in some animals at the two highest dose groups indicating test substance induced irritation. Based on these findings, the NOAEL for systemic toxicity in this study is 50 mg/kg bw/d, and the NOAEL for local toxicity is 0.5 mg/kg bw/d.

(Huntsman Polyurethanes, Report No: CTL/L/8010, 1997)

Other information

The studies summarized below did not follow a validated testing guideline for repeated dose toxicity and were documented rather poorly. In these studies duration or frequency differed from validated guidelines. The study duration was 14 days in mice (only reduced application frequency of 5 days/week) and 10 days in rabbits. In the dermal toxicity study in mice mortality occurred at least at the highest dose level.

In a 14 d short-term dermal repeated dose toxicity study young adult C3Hf/Bd mice (male/female) were exposed to the test substance (vehicle: methanol or acetone) for 5 d/week at concentrations of 2.5, 5 or 10 % (w/v) on a basis of 50 µL/animal/d (equivalent to approximately 50, 100 and 200 mg/kg bw/d). When using methanol as the vehicle 4/9 females and 1/9 males died at the high dose, and 1/9 males after the mid dose. When using acetone as the vehicle 3/10 females and 3/10 males died at the high dose. The changes in body weight did not allow any clear conclusions. There was no sign of skin irritation. There was dose-related increased spleen weight in males and females, and some indication of increased liver weight. Histopathology was not performed in this study. Based on the results, a NOAEL could not be determined.

(Oak Ridge National Laboratory, Report No: ORNL/TM-10472, 1987)

In a short-term dermal repeated dose toxicity study mature albino rabbits (male) were exposed to the test substance (vehicle: water) for 10 d (6 h/d) at concentrations of 1000 or 2000 mg/kg bw/d under occlusive conditions. No group showed any unusual clinical signs. The blood plasma values for the treated rabbits did not differ from those of the controls. There were no statistically significant differences in organ weights, or organ/body weight ratios between the test and control groups. The skin test sites showed a dose-related increased incidence of slight acanthosis. There was minimal irritation, but no hyperkeratosis, necrosis or ulceration. Based on these results a LOAEL of 1000 mg/kg bw/d was determined.

(Haskell Laboratory, EPA/OTS: Doc #878220288, Report No: 55-75, 1975)

Justification for classification or non-classification

The available experimental information is reliable and suitable for classification according to Regulation 1272/2008. The classification is based on the results of a valid oral repeated dose toxicity study according to OECD TG 408. The test substance is considered to be classified for specific target organ toxicity - repeated exposure (liver; UN GHS STOT RE Category 2, H373). The classification is based on persistent histopathological findings in the liver (hyperplasia of biliary ducts) which corresponded to elevations of clinical chemistry parameter indicative of hepatotoxicity at doses > 10 mg/kg bw/d. A classification of the substance for repeated toxicity UN GHS Category 1 based on the findings in the thyroid is not warranted due to only slight stimulation of the thyroidal follicular epithelium in a low number of male and female rats, which does not indicate a significant organ damage in the absence of any supporting findings.