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REACH

Bumetrizole

EC number: 223-445-4 | CAS number: 3896-11-5

Life Cycle description
Uses advised against

Toxicological information

Endpoint summary

Administrative data

Description of key information

Subacute studies: NOAEL(rat) = ca. 1000 (male and female) mg/kg bw /d (highest dose tested)
Subchronic studies: NOAEL (rat) = 637 (male), 740 (female) mg/kg bw/d (highest dose tested)
Subchronic studies: NOAEL (dog) = 153 (female), 168 (male) mg/kg bw/d (highest dose tested)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference 1

Endpoint:: short-term repeated dose toxicity: oral
Type of information:: experimental study
Adequacy of study:: other information
Reliability:: 3 (not reliable)
Rationale for reliability incl. deficiencies:: significant methodological deficiencies
Remarks:: Approximately one-half of the animals in this experiment showed evidence of chronic infection with round cell infiltrations and, in part, granulomas in liver and kidneys.
Principles of method if other than guideline:: 12 rats/sex/group were administered the test substance admixed in diet or plain diet (control group) over a period of 4 weeks. Daily food consumption per animal was calculated and weight was checked once a week. After the end of the experiment, the animals were turned over to Experimental Pathology for macroscopic and microspoic examination.
GLP compliance:: no
Specific details on test material used for the study:: - Name of test substance (as cited in study report): Tinuvin 326, cryst.
- Substance No.: GP 38771
Species:: rat
Strain:: other: Wistar-CF
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Weight at study initiation: 128 - 140 g
- Housing: 3/cage
- Diet: Nafag rat food No. 185 (meal, unpelleted); ad libitum
- Water: ad libitum
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Analytical verification of doses or concentrations:: no
Duration of treatment / exposure:: 4 weeks
Frequency of treatment:: daily intake with feed
Dose / conc.:: 500 ppm
Remarks:: nominal in diet. Equivalent to 0.05%.
No. of animals per sex per dose:: 12
Control animals:: yes, plain diet
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: Yes
- Time schedule: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE: yes, but no details in study report

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: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:: GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Details on results:: - Mortality:
No mortality occurred during the study period.

- Clinical signs:
No signs of toxicity were observed in any animal.

- Body weight:
180 - 345 g; mean: 255 g and 187 - 340 g; mean 250 g (control and test group, respectively)
Dose descriptor:: NOAEL
Effect level:: 500 ppm
Based on:: test mat.
Sex:: male/female
Critical effects observed:: no

Reference 2

Endpoint:: sub-chronic toxicity: oral
Type of information:: experimental study
Adequacy of study:: key study
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: comparable to guideline study with acceptable restrictions
Qualifier:: equivalent or similar to guideline
Guideline:: OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Deviations:: yes
Remarks:: (no neurobehavioural assessment and no detailed clinical examinations, no ophthalmoscopic examinations)
GLP compliance:: no
Limit test:: no
Specific details on test material used for the study:: - Name of test material (as cited in study report): CH 3504
Species:: rat
Strain:: Wistar
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Central Instititute for Nutrition and Food Research colony
- Age: Weanling rats
- Weight at study initiation: males, 45 - 64 g; females 45 - 62 g
- Housing: Wire screen cages (5 to a cage)
- Diet: ad libitum
- Water: ad libitum
- Acclimation: no data

ENVIRONMENTAL CONDITIONS: no data
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: DIET PREPARATION
- Rate of preparation of diet (frequency): every 2 weeks
- Mixing appropriate amounts with stock diet (see table 1 for composition)
Analytical verification of doses or concentrations:: no
Duration of treatment / exposure:: 13 weeks
Frequency of treatment:: daily
Dose / conc.:: 25 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 400 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 62 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 153.9 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 2500 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 637.4 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 10,00 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 28.9 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 400 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 70.6 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 176 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 2500 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
Dose / conc.:: 740.1 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 10,00 ppm nominal concentration. Calculated by average food consumption and average body weights after 13 weeks)
No. of animals per sex per dose:: 10
Control animals:: yes, plain diet
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: Yes; mortality and abnormalities in condition and behaviour

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: once per week

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes, but as g food/rat/day
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes
- Time schedule for examinations: in the fisrt 4 weeks and then in week 11 and 12

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: 12th week
- Anaesthetic used for blood collection: no data
- Animals fasted: no data
- How many animals: all animals
- Parameters checked in table No. 2 were examined.

CLINICAL CHEMISTRY: Yes
- How many animals: 5 animals/sex/dose
- Parameters checked: glutamic pyruvic transaminase (SGPT), glutamic oxalo-acetic transaminase (SGOT) and alkaline phosphatase (SAP), glucose-6-phosphatase (G6Pase) and glucose-6- phosphate dehydrogenase (G6PD).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:: GROSS PATHOLOGY: Yes
- Organ weight: heart, liver, kidney, spleen, testicle/ovary, thymus, pituitary, thyroid and adrenal

HISTOPATHOLOGY: Yes
- How many animals: all animals of the control and high dose groups
- Parameters checked: heart, *liver, *kidney, spleen, testicle/ovary, thymus, pituitary, *thyroid and adrenal, lung, salivary glands (sub-maxillary, sub-lingual and parotid), prostate, epididermis, coagulating gland, seminal vesicle, trachea, oesophagus, thoracic aorta, spinal cord, femoral nerve, skeletal muscle, uterus, preputial gland, external lacrimal gland, fore- and glandular stomach, duodenum, ileum, caecum, mandibular and mesenteric lymph nodes, pancreas, urinary bladder, skin and bone marrow (sternum).

*checked in the 400, 1000 and 2500 ppm dose groups
Details on results:: CLINICAL SIGNS AND MORTALITY: No deaths occurred and no abnormalities in condition or behaviour were observed.

BODY WEIGHT AND WEIGHT GAIN: There were no significant differences between the treated and control rats.

FOOD CONSUMPTION AND COMPOUND INTAKE: There was no compound effect at any of the dosing intervals.

FOOD EFFICIENCY: There was no compound effect at any of the dosing intervals.

HAEMATOLOGY: The haemoglobin content and packed cell volume of males were slightly, though significantly (p < 0.01) decreased at the 1000, 2500, and 10000 ppm feeding levels by 6, 6.6 and 5.4% compared to controls for haemaglobin content and at 2500 and 10000 ppm feeding levels by 5.6 and 5.1% compared to control for packed cell content. At the respective lower feeding levels these effects were not observed. These small differences did not show a dose relationship. In females, such effects were not seen. Therefore the low values in males are not explained as compound effects but rather as the result of relatively high values in male controls except for a slight decrease in haemoglobin content at 0.1% females, however, no distinct differences in haematological data between the treated and control rats were noticed.

CLINICAL CHEMISTRY: The alkaline phosphatase activity of females was slightly decreased at the highest feeding level. A trend towards lower SAP activity with increasing levels of the compound was not observed. A slight increase in G6Pase activity at the two highest levels in either sex was noticed.

ORGAN WEIGHTS: None of the organs showed considerable differences between groups. Relative weights of liver and kidney in percent of body weight were slighly increased at the highest feeding level. The difference was significant in females only. Kidney increased by 6% conpared to controls (p < 0.05) and liver increased by 10.7% compared to controls (p < 0.01).

GROSS PATHOLOGY: Gross autopsy findings were essentially negative.

HISTOPATHOLOGY: NON-NEOPLASTIC: No changes were fond in spleen, brain, testicle, epididymis, ovary, thymus, pituitary, submaxillary and partotid gland, seminal vesicle, coagulating gland, aorta, spinal cord, femoral nerve, oesophagus, stomach, duodenum, ileum, manibular and mesenteric lymph nodes, uterus, skin and bone marrow. The abnormalities that were found in liver, kidneys, thyroid, lungs, trachea, heart, bladder, caecum, colon, sublingual muscle, preputial gland and prostate are common findings in the strain of rat used. Some of the abnormalities, e.g. nephrocalcinosis in kidneys, activated appearance of the thyroid gland and interstitial pneumonitis (lung) occurred more frequently and to a distinctly higher degree in one or several test groups than in the control group. However, the incidence of all three phenomena in the control group was strikingly low and, moreover, their severity in treated animals was not dose-dependent. Therefore, none of the histopathological changes observed may be ascribed to the ingestion of the test substance.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: ca. 637 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male
Basis for effect level:: other: corresponding to 10000 ppm in diet. No adverse effects seen in any parameter at the highest dose level tested.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: ca. 740 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: female
Basis for effect level:: other: corresponding to 10000 ppm in diet. No adverse effects seen in any parameter at the highest dose level tested.
Critical effects observed:: no

Reference 3

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
Reason / purpose for cross-reference:: reference to same study
Qualifier:: according to guideline
Guideline:: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:: no
GLP compliance:: yes
Limit test:: no
Specific details on test material used for the study:: - Name of test material (as cited in study report): bumetrizole and 2-(2'-Hydroxy-3-tert -butyl-5’-methyphenyl) -5-chlorobenzotriazole
- Appearance/physical state: light yellow/powder
- Analytical purity: 99.9% w/w
- Lot/batch No.: 01721IW4
- Stability under test conditions: Stable
- Storage condition of test material: Stored sealed in a cabinet at 20.0 - 25.3° C
- Supplier: Ciba Specialty Chemicals, Osaka, Japan
Species:: rat
Strain:: Crj: CD(SD)
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Charles river Laboratories Japan (Hino Breeding Center)
- Age at study initiation/start of treatment: 7 weeks/10 weeks
- Weight at study initiation: Test group males 230 - 249 g; Test group females 176 - 202 g; Recovery group females 179 - 198 g
- Fasting period before study: Day before administration
- Housing: Individually
- Diet: CRF-1 (Oriental Yeast Co.), ad libitum
- Water: Tap water, ad libitum
- Acclimation period: 5 days quarantine, 16 days acclimation (17 days for recovery group)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 - 25
- Humidity (%): 40 - 56 (on 5 occasions, deviations below 40% humdity were recorded. As humidity deviation was slight (36 - 39.8%) and humidity for other period was within allowable limits, it is considered that this deviations had no influence to the test results.
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From May 13th, 2005 to July 13-15th (main group) and July 27-28th, 2005 (recovery group)
Route of administration:: oral: gavage
Vehicle:: methylcellulose
Remarks:: 0.5 % w/v solution
Analytical verification of doses or concentrations:: yes
Details on analytical verification of doses or concentrations:: The concentration of test material in the dosing material used for administration was determined by HPLC on the first and last days of administration.
Duration of treatment / exposure:: Males: Total of 42 days (from 14 days before mating to 28 days after mating)
Females: Total of 44 - 56 days (from 14 days before mating, during pregnancy and up to lactation day 6)
Frequency of treatment:: Daily
Dose / conc.:: 62.5 mg/kg bw/day (actual dose received)
Dose / conc.:: 250 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:: Test group (male): 12/dose (6 were assigned to the recovery group)
Test group (female): 12/dose
Recovery group females: 6/group at control, 250 or 1000 mg/kg bw/day
Control animals:: yes, concurrent vehicle
Details on study design:: - Dose selection rationale: Doses selected on the basis of a preliminary repeat dose study (no deaths or clinical signs at 1000 mg/kg bw/day).
- Rationale for animal assignment: random
- Post-exposure recovery period: Males - half the animals in each dose group were allowed a recovery period of 14 days after the administration period; Recovery group females - 14 day recovery period after a 42 day administration period.
Observations and examinations performed and frequency:: DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Males - before and after administration on each administration day, once every day during the recovery period and once on the termination day; Females - before and after administration on each administration day and once on the termination day; before and after administration on each administration day, once during the recovery period and once on the termination day.

BODY WEIGHT: Yes
- Time schedule for examinations: Males - measured twice per week on administration days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32, 36, 39, 42 and 43 (recovery day 1), recovery day 4, 8, 11, 14 and 15; Females - measured twice a week for 14 days before start of mating and during mating period on administration day 1, 4, 8, 11, 15, 18, 22 and 25, during pregnancy period on days 0, 7, 14 and 21 and during lactation period on days 0, 4, 6 and 7; recovery group females - measured twice a week on adminstration day 1, 4, 8, 11, 15, 18 ,22, 25, 29, 32, 36, 39 and 42, recovery day 1, 4, 8, 11, 14 and 15.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Males - measured twice a week for 14 days before the start of breeding and after the mating period on administration days 2, 5, 9, 12, 30, 33, 37 and 40, recovery day 2, 5, 9 and 12; Females - twice a week for 14 days before the start of mating on administration days 2, 5, 9 and 12, during pregnancy on days 2, 9, 16 and 20 and during lactation on day 2; Recovery group females - food consumption was measured twice a week on administration days 2, 5, 9, 12, 16, 19, 23, 26, 30, 33, 37 and 40 and recovery days 2, 5, 9 and 12.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Males - day after final administration and after the recovery period; Females - lactation day 7; Recovery group females - end of the recovery period.
- Anaesthetic used for blood collection: Yes (pentobarbital sodium, 40 mg/kg bw)
- Animals fasted: No data
- How many animals: six animals/sex/group
- Parameters checked in table 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Males - day after final administration and after the recovery period; Females - lactation day 7; Recovery group females - end of the recovery period.
- Animals fasted: No data
- How many animals: six animals/sex/group
- Parameters checked in table 2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: Males - Administration day 37 and recovery day 9 after fasting. Then collected for 24 hours with food and water avaliable; Females - Lactation day 5 after fasting, then collected for 24 hours with food and water available; Recovery group females - Recovery day 9 after fasting, then collected for 24 hours with food and water available.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Refer to time schedule
- Parameters checked in table 3 were examined.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations:
Males: FOB - day 0, 7, 14, 21, 28, 35 and 41 of administration; Sensory activity - administration day 41; grip strength - administration day 41; motor activity - administration day 40.
Females: FOB - day 0, 8 and 15 of administration, pregnancy day 1, 8 and 15, lactation day 3; Sensory activity - lactation day 3; grip strength - lactation day 3; motor activity - lactation day 4.
Recovery group females: FOB - day 0, 8, 15, 22, 29, 36 and 42 of administration.
- Dose groups that were examined: All dose groups
- Battery of functions tested: sensory activity / grip strength / motor activity/ Functional Observational Battery (FOB)
Sacrifice and pathology:: GROSS PATHOLOGY: Yes (see table 4)
HISTOPATHOLOGY: Yes (see table 5)
Statistics:: The test of significance was performed between control groups and each test group and recorded separately at p<0.05 and p<0.01. Unimpregnated females after copulation were excluded from the counting of general signs, body weight, food consumption and FOB.

Averages and standard deviations in each group were calculated for body weights, food consumption, frequency of rearing and grooming for FOB, grip strength, spontaneous motor activity, urinary volume, urinary specific gravity, records of haematological/blood chemical examination, absolute and relative weights of organs. Subsequently, the test of distribution uniformity by Bartlett's test was performed and the Dunett test was performed when the distribution was uniform and Dunett test using rank was performed when the homoscedastic was not recognised. For the FOB (excluding frequency of rearing and grooming) and sensory response, the average and range in each group were calculated and the Dunett test using rank performed.
Details on results:: CLINICAL SIGNS AND MORTALITY
Males/females/recovery group females: No dead or moribund animals in any group. No abnormality in clinical signs in any group.

BODY WEIGHT AND WEIGHT GAIN
Males/females/recovery group females: No significant differences in body weight in any group compared to controls on any measurement day.

FOOD CONSUMPTION
Males: During the administration period, no significant difference was observed in food consumption in any group compared to controls on any measurement day. During the recovery period, no significant difference was observed in food consumption in the 1000 mg/kg bw/day group compared to controls on any measurement day. Significant low values were seen on recovery day 5 in the 250 mg/kg bw/day group compared to controls, but as the effect was not dose-related it was not considered to be due to administration of the test material.

Females: Before the start of mating, during pregnancy and during lactation, no significant difference was observed in food consumption compared to controls on any measurement day.

Recovery group females: During the administration and recovery periods, no significant difference was observed in food consumption in any dose group compared to controls on any measurement day.

HAEMATOLOGY
Termination of administration period:
Males: For each measured parameter, no significant difference was observed in any group compared to controls.
Females: No significant difference observed in each measured parameter in the 1000 mg/kg bw/day group compared to controls. Significantly lower values of platelet were seen in the 250 and 62.5 mg/kg bw/day groups compared to controls, but as the effect was not dose-related it was not considered to be due to administration of the test material.

Termination of recovery period:
Males: For each measured parameter, no significant difference was observed in any group compared to controls.
Recovery group female rats: Significant low value of lymphocyte and neutrophil was seen in the 1000 mg/kg bw/day group compared to controls. However, the data were within the range of historical control values from the test laboratory and the effects are not considered to be due to administration of the substance. In the 250 mg/kg bw/day group, significant high value of MCHC was seen compared to controls, but as the effect was not dose-related it was not considered to be due to administration of the test material.

CLINICAL CHEMISTRY
Termination of administration period:
Males: A significantly lower value of total bilirubin was seen in 1000 mg/kg bw/day group compared to controls. As this was a slight variance, it was not considered to be of toxicological significance. In the 250 mg/kg bw/day group no significant difference compared to controls in each measured parameter was observed. In the 62.5 mg/kg bw/day group, a significantly higher value for Cl, and significantly lower levels of total protein and total bilirubin were noted compared to controls. But as the effects were not dose-related they were not considered to be due to administration of the test material.
Females: For each measured parameter, no significant difference was observed in any group compared to controls.

Termination of recovery period:
Males: For each measured parameter, no significant difference was observed in any group compared to controls.
Recovery group females: For each measured parameter, no significant difference was observed in any group compared to controls.

URINALYSIS
Before termination of administration period:
Males/females: For the urinary volume and the urinary specific gravity, no significant difference was observed in any dose group compared to controls. Colour tone, pH, protein, glucose, ketone body, bilirubin, occult blood, urobilinogen and deposit were nearly equal to controls in each dose group.

Before termination of recovery period:
Males/Recovery group females: For the urinary volume and the urinary specific gravity, no significant difference was observed in any dose group compared to controls. Colour tone, pH, protein, glucose, ketone body, bilirubin, occult blood, urobilinogen and deposit were nearly equal to controls in each dose group.

NEUROBEHAVIOUR
Functional Observational Battery (FOB)
Males/females: No abnormality was observed in parameters in any group on any examination day.
Recovery group females: No abnormality was observed in parameters in the 1000 mg/kg bw/day group compared to controls on any examination day. Significantly higher values of rearing frequency were seen on administration day 36 in the 250 mg/kg bw/day group compared to controls, but as the effect was not dose-related it was not considered to be due to administration of the test material. Also, in 250 mg/kg bw/day group, a significantly higher incidence of rearing frequency were observed before the administration period.

Sensory Response:
Males/females: No abnormality of parameters was observed in any administration group.

Grip Strength:
Males/females: No significant difference was observed in grip strength of foreleg and hind leg in dose groups compared to controls.

Spontaneous Motor Activity:
Males: In the 1000 and 62.5 mg/kg bw/day groups, no significant difference was observed in parameters compared to controls. Significantly lower values of ambulatory activity and rearing frequency were observed at 60 minutes after the administration in the 250 mg/kg bw/day group compared to controls, but as the effect was not dose-related it was not considered to be due to administration of the test material.
Females: For each measurement parameter, no significant difference was observed in dose groups compared to controls.

ORGAN WEIGHTS
Termination of administration period:
Males: No significant difference was observed in the body weights of the necropsy day in dose groups compared to controls. No significant difference was observed in the absolute and relative weight of organs in dose groups compared to controls.
Females: No significant difference was observed in the body weights on the necropsy day in dose groups compared to controls. No significant difference was observed in the absolute and relative weight of organs in 1000 and 250 mg/kg bw/day groups compared to the control group. Significant high values for relative weight of thymus observed in the 62.5 mg/kg bw/day group in comparison to controls was not considered to be an effect of the administration as the variance was not relevant to the administration dose.

Termination of recovery period:
Males: For the body weights of the necropsy day, no significant difference was observed in dose groups compared to controls. No significant difference was observed in the absolute and relative weight of organs in dose groups compared to controls.
Recovery group females: For the body weights on necropsy day, no significant difference was observed in dose groups compared to controls. For organ weights, no significant difference was observed in absolute and relative organ weight in 1000 mg/kg bw/day group compared to controls. Significantly lower relative weight of the ovary was seen in 250 mg/kg bw/day group compared to controls. Because the variance was not related to the administration dose, it was not considered to be an effect of the administration.

GROSS PATHOLOGY
Termination of administration period:
Males: In the 1000, 250 and 62.5 mg/kg bw/day groups, no abnormality was observed. In the control group, there was 1 case of pale yellow tuber on the left epididymis caudal portion observed.
Females: No abnormality was observed in any group.

End of recovery period:
Males: No abnormality was observed in any group.
Recovery group females: No abnormality was observed in any group.

HISTOPATHOLOGY: NON-NEOPLASTIC
Termination of the administration period:
Males:
Lung: A case of foam cell gregarine was seen in 1000 mg/kg bw/day and 2 cases in the controls.
Intestinum jejunum: A case of mineral deposition of Peyer's patch was seen in the 1000 mg/kg bw/day group.
Epididymis: A case of lymphocyte infiltration and unilateral spermatozoa granuloma was seen in the controls.
Prostate: A case of lymphocyte infiltration was seen in 1000 mg/kg bw/day group.

These variances were present even in the control group. In addition, the grade is quite slight. In view of this and because the occurrence frequency in the 1000 mg/kg bw/day was not much different from that of the control group, the effects are considered to be contingent variance.

Otherwise, in 1000 mg/kg bw/day and control groups, no abnormality was observed in heart, bronchus, liver, pancreas, salivary gland, mandibular gland, oesophagus, stomach, duodena, ileum, caecum, colon, rectum, thymus, spleen, mandibular lymph node, mesenteric lymph nodes, kidney, bladder, testicle, vesicula, hypophysis, adrenal, thyroid, parathyroid, cerebrum, cerebellum, medullary, spinal marrow, ischiadic nerve, eyeball, harderian gland, bone marrow (sternum and femur) and bone (sternum and femur).

Females:
Intestinum jejunum: A case of mineral deposition of Peyer's patch was seen in the 1000 mg/kg bw/day group.
Spleen: A case of extramedullary haematopoiesis was seen in the 1000 mg/kg bw/day and 3 cases in the controls.
Eyeball: A case of right retinal dysplasia was observed in the 1000 mg/kg bw/day group.

These variances were present even in the control group. In addition, the grade is quite slight. In view of this and because the occurrence frequency in the 1000 mg/kg bw/day was not much different from that of the control group, the effects are considered to be contingent variance.

Otherwise, in the 1000 mg/kg bw/day and control groups, no abnormality was observed in heart, lung, bronchus, liver, pancreas, salivary gland, mandibular gland, oesophagus, stomach, duodena, ileum, caecum, colon, rectum, thymus, mandibular lymph node, mesenteric lymph nodes, kidney, bladder, ovary, uterine, vagina, hypophysis, adrenal, thyroid, parathyroid, cerebrum, cerebellum, medullary, spinal marrow, ischiadic nerve, harderian gland, bone marrow (sternum and femur), bone(sternum and femur), and mammary gland.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: 1 000 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male/female
Basis for effect level:: other: No adverse effects seen in any parameter at the highest dose level tested.
Critical effects observed:: no

Reference 4

Endpoint:: chronic toxicity: oral
Remarks:: combined repeated dose and carcinogenicity
Type of information:: experimental study
Adequacy of study:: supporting study
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: comparable to guideline study with acceptable restrictions
Reason / purpose for cross-reference:: reference to same study
Qualifier:: equivalent or similar to guideline
Guideline:: OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
GLP compliance:: no
Limit test:: no
Specific details on test material used for the study:: - Name of test material (as cited in study report): TK 10 048
- Lot/batch No.: 26580
Species:: rat
Strain:: Sprague-Dawley
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Anglia Laboratory Animals, Alconbury, UK
- Age at study initiation: 28 ± 1 day
- Weight at study initiation: 70 - 90 g; weights of the rats per group differed by no more than -2.5 g.
- Housing: 5 animals/cage (wire mesh floors)
- Diet: ad libitum; Spratt's Laboratory Diet 2
- Water: ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2
- Humidity (%): 50 ± 5
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with Spratt's Laboratory Diet 2
- Storage temperature of food: in heat sealed, opaque polythene bags
Analytical verification of doses or concentrations:: yes
Details on analytical verification of doses or concentrations:: Performed every 13 weeks. Samples were extracted with distilled chloroform and determined spectrophometrically.
Recovery
- 1000 ppm: 95 - 105%
- 3000 ppm: 98.3 - 101.6%
- 10000 ppm: 90 -103%
Duration of treatment / exposure:: 104 weeks
Frequency of treatment:: Daily
Dose / conc.:: 37.7 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm nominal concentration)
Dose / conc.:: 113.2 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 3000 ppm nominal concentration)
Dose / conc.:: 382.6 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 10,000 ppm nominal concentration)
Dose / conc.:: 50.4 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm nominal concentration)
Dose / conc.:: 147.7 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 3000 ppm nominal concentration)
Dose / conc.:: 501.9 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 10,000 ppm nominal concentration)
Dose / conc.:: 37.7 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm nominal concentration. Calculated by food intake and body weight)
Dose / conc.:: 113.2 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 3000 ppm nominal concentration. Calculated by food intake and body weight)
Dose / conc.:: 382.6 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 10,000 ppm nominal concentration. Calculated by food intake and body weight)
Dose / conc.:: 50.4 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm nominal concentration. Calculated by food intake and body weight)
Dose / conc.:: 147.7 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 3000 ppm nominal concentration. Calculated by food intake and body weight)
Dose / conc.:: 501.9 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 10,000 ppm nominal concentration. Calculated by food intake and body weight)
No. of animals per sex per dose:: 50
Control animals:: yes, plain diet
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes
- Time schedule for examinations: weekly for the first eight weeks then fortnightly

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined: Yes
- Mean daily diet consumption calculated as g food/rat/week: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: 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: Yes
- Time schedule for examinations: During weeks 6, 12 and 26
- Dose groups: control and 10000 ppm groups

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: weeks 0, 12, 26, 52, 78 and 103
- Dose groups that were examined: control and 10000 ppm groups

HAEMATOLOGY: Yes
- Time schedule for collection of blood: weeks 0, 6, 12, 25, 77 and 104
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes, overnight
- How many animals: 10/sex control and 10000 ppm group. The investigations were extended at week 25 to 10 females from the mid dose group (3000 ppm), at week 52 to 10 males and 10 females of the low and mid dose groups, at week 79 to 10 males of the low and mid dose groups and at week 102 to 10 males and 10 females for the low and mid dose groups for packed cell volume, haemoglobin and red cell count.
- Parameters in table 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: weeks 6, 12, 25, 52, 77 and 102
- Animals fasted: Yes, overnight
- How many animals: 10/sex control and 10000 ppm group. The investigations were extended at weeks 52 to the 1000 and 3000 ppm groups for measurement of plasma glucose only and at week 85 for total serum protein and serum protein electrophoresis.
- Parameters in table 2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: weeks 6, 12, 26, 52, 78 and 103
- Metabolism cages used for collection of urine: No data
- How many animals: 10/sex control and 10000 ppm group
- Animals fasted: Yes, overnight
- Parameters checked in table 3 were examined.

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:: GROSS PATHOLOGY: Yes (see table 4)
HISTOPATHOLOGY: Yes (see table 5)
Statistics:: Although it is clear from the study report that statistical methods were used, there is no description of the actual methods
Details on results:: CLINICAL SIGNS AND MORTALITY
There were no visible signs of reaction to treatments. No marked differences between the distribution of mortalities in control and treated groups were observed

BODY WEIGHT AND WEIGHT GAIN
A marginally lower rate of bodyweight gain was recorded during the first 52 weeks of the study among males receiving 10000 ppm. Thereafter, body weight change was comparable to that of control animals.

FOOD CONSUMPTION
A transient reduction of food intake was recorded between weeks 6 and 78 for males receiving 10000 ppm. From week 79 till the study end, food intake was similar to that of controls.

COMPOUND INTAKE:
1000 ppm: 37.7 mg/kg bw (male); 50.4 mg/kg bw (female)
3000 ppm 113.2 mg/kg bw (male); 147.7 mg/kg bw (female)
10000 ppm 382.6 mg/kg bw (male); 501.9 mg/kg bw (female)

WATER CONSUMPTION
Water consumption was not influenced as recorded in the controls and high dose animals during weeks 6, 12 and 26.

FOOD EFFICIENCY
Food efficiency was unimpaired

OPHTHALMOSCOPIC EXAMINATION
No abnormalities of the eyes were detected that were considered to be treatment related.

HAEMATOLOGY
Minimally lower values for the red cell parameters were consistently recorded at weeks 6, 12, 25 and 52 for animals receiving 10000 ppm. At week 25, lower values were also recorded in females receiving 3000 ppm.
During week 77 a more marked reduction in red cell parameters was recorded in males receiving 10000 ppm although this could be accounted for by the low values recorded in 3 out of 10 of the rats examined.
Other small differences between values for control and treated rats although in some cases attaining a level of statistical significance were considered to be of no toxicological significance, as all values were within the accepted range for rats of the age and strain employed.

CLINICAL CHEMISTRY
A marginally higher SAP (Serum alkaline phosphatase) value was recorded in week 12 for males receiving 10000 ppm and in week 102 for females of the same dose group. As this finding was not seen consistently in either males or females, it was considered to be of doubtful toxicological significance.
Higher glucose values were recorded during week 52 in some rats (particularly males) receiving 10000 ppm. This was however not considered a finding of toxicological significance since no dose dependent relationship could be seen when rats of the low and mid dose groups were subjected to examination. In addition, subsequent recordings of the glucose levels indicated no difference to control animals.
During week 77, a marked reduction in albumin levels was recorded in all male rats receiving 10000 ppm. Because later examinations (week 85) in all males of treated groups did not indicate any differences from controls, this abnormal findings at week 77 were considered to be of no toxicological significance.
All other parameters measured revealed similar values for controls and rats receiving 10000 ppm

URINALYSIS
There were no differences in urinary parameters among controls or treatment groups.

ORGAN WEIGHTS
A slight but significant increase in relative liver weight was recorded in males receiving 1000 or 10000 ppm and in females receiving 3000 ppm. These changes were not dosage related and no significant increase in absolute liver weight was recorded. In adddition, in the absence of any histopathological changes, these findings were considered unrelated to treatment.

GROSS PATHOLOGY
No treatment related changes were observed.

HISTOPATHOLOGY (NON-NEOPLASTIC):
There was no evidence that treatment with test substance induced any adverse effects.

HISTOPATHOLOGY (NEOPLASTIC):
There was no evidence to suggest that the administration of the substance was associated with the disturbance of the spontaneous tumour profile in the strain of rat employed.

A summary of the effects seen follows:

Endocrine glands:
The largest incidence of tumours in this category occurred in the pituitary gland, and this tumour was seen in rats from both control and treated groups. Pituitary gland tumours are commonly seen in laboratory rats of this age, and there was no evidence of a treatment-related change. The majority of tumours seen were pituitary gland adenomata. The incidence of tumours in other endocrine glands showed no evidence of a treatment-related change.

Cutaneous and subcutaneous tissue:
There was no evidence of a treatment-related effect in the incidence of tumours in cutaneous and subcutaneous locations.

Lymphoreticular system:
The low incidence of tumours of lymphoreticular origin showed no evidence to suggest a treatment-related effect.

Liver:
One malignant and five benign liver cell tumours were recorded. This was within the expected range.

Reproductive system:
The only neoplasms recorded in male rats were testicular interstitial cell tumours in three control rats and in one treated rat. Among female rats, the tumours recorded were four ovarian adenomas, one luteinoma, one uterine adenocarcinoma and one uterine fibrosarcoma. These tumours of the reproductive system are not uncommon in the laboratory rat and were considered to be of no toxicological significance.

Mammary tissue:
No treatment-related effect was seen in the incidence of the fibroadenomas, adenomas and adenocarcinomas recorded in this study.

Miscellaneous tumours:
The individual tumour types and the incidence of these neoplasms were within the normal range for this strain of rats used in our Iaboratories.

Total tumour incidence:
The total number of rats bearing neoplasms was similar in treated and control groups.
Dose descriptor:: NOEL
Remarks:: (systemic)
Effect level:: ca. 113.2 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male
Basis for effect level:: other: corresponding to 3000 ppm based on overall effects seen in the 10000 ppm dose group; clinical signs; body weight; clinical chemistry; haematology
Dose descriptor:: NOEL
Remarks:: (systemic)
Effect level:: 147.7 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: female
Basis for effect level:: other: corresponding to 3000 ppm based on overall effects seen in the 10000 ppm dose group; clinical signs; body weight; clinical chemistry; haematology
Critical effects observed:: no

Reference 5

Endpoint:: sub-chronic toxicity: oral
Type of information:: experimental study
Adequacy of study:: supporting study
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: comparable to guideline study with acceptable restrictions
Qualifier:: equivalent or similar to guideline
Guideline:: OECD Guideline 409 (Repeated Dose 90-Day Oral Toxicity Study in Non-Rodents)
Deviations:: yes
Remarks:: (at initiation, dogs were slightly older than 9 months, satellite group composed of only 1 animal/sex in the control and high dose groups.)
GLP compliance:: yes
Limit test:: no
Specific details on test material used for the study:: - Name of test material (as cited in study report): TK 10048
- Lot/batch No.: 108947
- Physical state: solid (powder)
- Colour: yellow
- Storage condition of test material: In the dark at room temperature
- Stability at storage conditions: stability of up to 4 weeks at room temperature was ascertained
Species:: dog
Strain:: Beagle
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Beaulong Ltd, Harewood Park, Herefordshire, England
- Age at study initiation: 45 - 55 weeks (males); 46 - 53 weeks (females)
- Weight at study initiation: 10.3 - 13.2 kg (males); 9.3-14.0 kg (females)
- Fasting period before study: none
- Housing: individually in kennels (6 ft x 2 ft with 5 ft dividing partitions)
- Diet: Spratts Dog Diet No. 2, 400 g each morning
- Water: ad libitum
- Acclimation period: 7 weeks
During the acclimatisation period the dogs were re-inoculated against distemper, hepatitis and leptospirosis (Maxovoc: Hoechst Pharmaceuticals), and received a course of treatment (3 doses at 10-day intervals), with the anthelmintic piperazine (Coopone: Burroughs Wellcome).
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly
- Mixing appropriate amounts with (Type of food): powdered standard dry diet (Spratts Dog Diet No.2)
- Storage temperature of food: Room temperature
Analytical verification of doses or concentrations:: yes
Details on analytical verification of doses or concentrations:: Samples of prepared diet were collected from the mixes prepared for feeding during weeks 1, 6 and 13. All samples of diet were sent to the Sponsor for analysis. Test substance was extracted from samples with distilled chloroform for 2 hours in a Soxhlet apparatus. Test substance was determined spectrophotometrically using a double beam spectrophotometer at 354 nm.
Recovery:
200 ppm: 100 - 110%
1000 ppm: 95 - 100%
5000 ppm: 95% - 102%
Duration of treatment / exposure:: 13 weeks
Frequency of treatment:: once per day, 7 days per week
Dose / conc.:: 6.2 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 200 ppm nominal concentration)
Dose / conc.:: 29.6 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm nominal concentration)
Dose / conc.:: 168 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 5000 ppm nominal concentration)
Dose / conc.:: 6.5 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 200 ppm nominal concentration)
Dose / conc.:: 32.2 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm nominal concentration)
Dose / conc.:: 153 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 5000 ppm nominal concentration)
Dose / conc.:: 6.2 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 200 ppm)
Dose / conc.:: 29.6 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm)
Dose / conc.:: 168 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 5000 ppm)
Dose / conc.:: 6.5 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 200 ppm)
Dose / conc.:: 32.2 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm)
Dose / conc.:: 153 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 5000 ppm)
No. of animals per sex per dose:: test group: 4
recovery (control and high dose only): 1
Control animals:: yes, plain diet
Details on study design:: - Dose selection rationale: no data
- Rationale for selecting satellite groups: test for reversiblity or capturing later effects
- Post-exposure recovery period in satellite groups: 4 weeks
- No. of animals in satellite group: 1/sex at the control and high dose groups.
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: Yes
- Time schedule: All animals were checked regularly throughout the working day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: prior to feeding, once a week throughout pre-dosing, dosing and observation periods

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated at weekly intervals from the consumption and individual bodyweight at the end of the week.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: during the 4 weeks prior to commencement of dosing and during weeks 1 - 4 and 9 - 12 of the dosing period and weeks 1 - 4 of the recovery period

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: once before commencement of dosing and again during weeks 6 and 11
- Dose groups that were examined: all

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Once before the commencement of dosing and again during weeks 5, 9 and 13 of dosing and week 4 of the observation period.
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes (16 hours)
- How many animals: all
- Parameters checked in table [No. 1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Once before the commencement of dosing and again during weeks 5, 9 and 13 of dosing and week 4 of the observation period.
- Animals fasted: Yes (16 hours)
- How many animals: all
- Parameters checked in table [No. 2] were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: Once before the commencement of dosing and again during weeks 5, 9 and 13 of dosing and week 4 of the observation period.
- Metabolism cages used for collection of urine: No
- Animals fasted: Yes (water withdrawn 5 hours before urine collection commenced)
- Parameters checked in table [No. 3] were examined.

NEUROBEHAVIOURAL EXAMINATION: No

OTHER:
HEARING TESTS:
- Time schedule for test: Twice before the commencement of dosing and again during weeks 5, 9 and 13 of dosing and week 4 of the observation period.
- Parameters checked: response to Galton whistle set to 10 kHz
Sacrifice and pathology:: GROSS PATHOLOGY: Yes (see table 4)
HISTOPATHOLOGY: Yes (see table 5)
Statistics:: Whenever it was necessary to determine whether significant differences existed between mean values relating to test and control animals, the method routinely used was Analysis of Variance followed by Student’s ‘t’ test. The results being expressed as the least significant difference (LSD).

When examining the organ weights, analysis of variance was carried out adjusting for final bodyweight as covariant where the regression coefficient describing the linear relationship between argon weight and bodyweight was significantly different from zero at the 10% level. Where there was no such relationship analysis of variance was carried out on the unadjusted values.

If heterogeneity of variance was present at the 1% level of significance (Bartlett’s test) a logarithmic transformation was performed on the data in order to stabilize the variance.

The group means (adjusted where appropriate) were compared using Williams test for contrasting increasing dose levels of a compound with a control. Significance testing was carried out at the 5% and 1% levels.

The only other statistical procedure adopted was that used to describe the range of results obtained during the pre-dosing investigation. These results have been described by the grand mean and 95% range, the Iatter phase being used to describe the mean +/- (t x standard deviation).
Details on results:: CLINICAL SIGNS AND MORTALITY
One female in the 1000 ppm group was killed for humane reasons on day 79, having shown a marked loss of appetite and a pain reaction on palpitation of the abdomen. Macroscopic post mortem examination revealed areas of petechial haemorrhage throughout the stomach antrum and a pitted area 2 mm in diameter in the mucosal lining. Histological examination revealed that the condition of the animal was due possibly to a myocardial lesion. No other animals died during the study.
There were no clinical signs that could be related to dosage with the test substance.

BODY WEIGHT AND WEIGHT GAIN
Female animals receiving 5000 ppm showed a significant (p<0.01) weight loss of -440 g in comparison to initial weights and by ca. 7% (- 800 g) when compared to control animals. In the 2 recovery animals of the high dose group, there were no marked bodyweight changes during the 4-week observation period in comparison to recovery control animals. One male receiving 5000 ppm lost a total of 1900 g during the study, but it is considered that this was unrelated to treatment, as this animal also lost 1100 g during pre-dosing weeks 1 & 2 but consumed nearly all the food offered.

FOOD CONSUMPTION AND COMPOUND INTAKE
With the exception of one female in the control and one female in the 5000 ppm group, which consumed relatively small quantities of food during the dosing period, there were no statistically significant differences in food consumption between control and treated animals. The female animal of the high dose group showed a marked improvement during the recovery period becoming maximal from the second week onwards.
The maximum, minimum and average dosages in mg/kg bw/day achieved during the dosing period are shown in Table 6 below.

WATER CONSUMPTION
With the exception of one female in the 5000 ppm group, which consumed relatively small quantities of water during the dosing period but showed a marked improvement during the recovery period, there was no adverse effect on water consumption due to treatment with the test substance.

OPHTHALMOSCOPIC EXAMINATION
No abnormalities were detected that could be related to dosage with the test substance.

HAEMATOLOGY
Although haematological parameters were strictly within normal limits, red blood cell values for some animals in the 5000 ppm group were on occasion lower than might be expected for adult animals. It was, on occasion, possible to show that mean values relating to PCV, Hb and RBC for this group were significantly lower than the control values (see table 7). It should be noted that the differences were slight and there was no evidence of a reticulocyte response in any animal.

CLINICAL CHEMISTRY
No effects due to treatment with the test substance noted.

URINALYSIS
No effects due to treatment with the test substance noted.

ORGAN WEIGHTS
After 13 weeks, the mean liver weights for males and females in the 5000 ppm group were significantly higher than in controls by 4%. Relative liver weights to body weight for both sexes of the 5000 ppm dose group and for females that received 1000 or 200 ppm were also significantly higher than in controls. However liver weights in recovery animals were within normal limits. In view of the observed reversibility coupled with the fact that there were no histopathological findings in the liver, the increased liver weights may reflect an adaptation to functional load. The mean heart weights for all groups of females receiving the test substance were significantly greater than controls. It was also posssible to demonstrate that the mean pituitary weight for females that received 5000 ppm significantly greater than the control values. Because there were no histopathological findings associated with the weight increase, this effect is regarded as a non-adverse, physiological and reversible adaptation to the treatment. After the 4 week treatment free period; liver and pituitary weights were within normal limits. The relative heart weight for the male dog of the high dose group was however, higher than expected.

GROSS PATHOLOGY
There were no abnormalities that could be related to dosage with the test substance.

HISTOPATHOLOGY: NON-NEOPLASTIC
No abnormalities or variation from the normal was encountered in the tissues examined which was attributable to treatment with the test substance. The increased liver weights reported macroscopically may reflect an adaptation to functional load. The effects seen in the lungs, livers, and kidney were of comparable incidence across all groups hence were rather of spontaneous origin than of from treatment with the test substance.

OTHER FINDINGS
HEARING TESTS
It is considered that there was no effect on the ability of animals to respond to a 10 kHz whistle.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: ca. 168 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male
Basis for effect level:: other: corresponding to 5000 ppm in diet
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: ca. 153 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: female
Basis for effect level:: other: corresponding to 5000 ppm in diet
Critical effects observed:: no

Reference 6

Endpoint:: short-term repeated dose toxicity: oral
Type of information:: experimental study
Adequacy of study:: supporting study
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: comparable to guideline study with acceptable restrictions
Qualifier:: equivalent or similar to guideline
Guideline:: OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:: yes
Remarks:: (no neurobehavioural evaluation, no haematology, clinical chemistry, ophthalmoscopic evaluations)
GLP compliance:: no
Limit test:: no
Specific details on test material used for the study:: - Name of test substance (as cited in study report): CH 3504
An infra-red spectrum was taken. No other information was present in report
Species:: rat
Strain:: Wistar
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Central Institute for Nutrition and Food Research's colony
- Age: Weanling rats
- Weight at study initiation: Males, 47.8 - 48.8 g, females, 45.8 - 46.2 g
- Fasting period before study: no
- Housing: metal wire screen cages (5 to a cage)
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: no

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24
no further details.
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: DIET PREPARATION
- Rate of preparation of diet (frequency): once per fortnight
- Mixing appropriate amounts with: stock diet (see table 1 for composition)
Analytical verification of doses or concentrations:: no
Duration of treatment / exposure:: 28 days
Frequency of treatment:: daily
Dose / conc.:: 36.9 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 400 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 100.2 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 1000 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 235.4 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 2500 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 996.7 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 10,00 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 41.5 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 400 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 96.7 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 1000 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 252.5 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 2500 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
Dose / conc.:: 1 014.8 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 10,00 ppm nominal concentration. Calculated by food consumption and body weights after 4 weeks)
No. of animals per sex per dose:: 5
Control animals:: yes, plain diet
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: Yes

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: one week intervals

FOOD CONSUMPTION AND COMPOUND INTAKE:
- 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:
- 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

OPHTHALMOSCOPIC EXAMINATION: No

ORGAN WEIGHTS: Yes (liver and kidney of all animals)

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:: GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes (liver and kidney)
Details on results:: CLINICAL SIGNS AND MORTALITY: No mortality occurred during the study and no signs of toxicity were observed.

BODY WEIGHT AND WEIGHT GAIN: There were no significant differences in body weights between treated and control animals over the 4 week dosing period.
(control, 400, 1000, 2500 and 10000 ppm)
Average weights after 1 week: 78.0, 79.8, 79.8, 77.6 and 73 g for males
Average weights after 1 week: 75.4, 73.2, 70.2, 69.8 and 70.4 g for females
Average weights after 2 weeks: 111.6, 117.8, 112.2, 109.8, and 104.4 g for males
Average weights after 2 weeks: 100.8, 100.4, 92.2, 92.0, and 93.6 g for females
Average weights after 3 weeks: 147.8, 154.8, 150.8, 140.2 and 136.4 g for males
Average weights after 3 weeks: 122.4, 120.0, 112.47, 110.6 and 113.4 for females
Average weights after 4 weeks: 184.4, 189.8, 184.2, 173.6 and 167.8 g for males
Average weights after 4 weeks: 137.6, 130.6, 126.2 and 128.8 for females

FOOD CONSUMPTION AND COMPOUND INTAKE: Food consumption was comparable to those of the controls in both sexes.
(control, 400, 1000, 2500 and 10000 ppm)
Average weights after 1 week: 8.3, 9.0, 8.7, 8.6 and 7.8 g/rat/day for males
Average weights after 1 week: 8.3, 8.1, 7.5, 7.3 and 7.5 g/rat/day for females
Average weights after 2 weeks: 11.9, 12.9, 12.0, 12.1, 11.2 g//rat/day for males
Average weights after 2 weeks: 10.9, 11.6, 9.8, 10.3 and 10.3 g/rat/day for females
Average weights after 3 weeks: 14.2, 15.4, 15.3, 13.8 and 13.5 g/rat/day for males
Average weights after 3 weeks: 11.9, 12.6, 10.6, 11.4 and 11.9 g/rat/day for females
Average weights after 4 weeks: 15.7, 17.0, 16.7, 12.6 and 15.3 g/rat/day for males
Average weights after 4 weeks: 11.9, 12.2, 11.0, 11.3 and 11.8 g/rat/day for females

FOOD EFFICIENCY: comparable to those of the controls in both sexes.
(control, 400, 1000, 2500 and 10000 ppm)
After 1 - 2 weeks: 0.45, 0.46, 0.44, 0.43 and 0.42 g gain per g food for males
After 1 - 2 weeks: 0.41, 0.39, 0.38, 0.37 and 0.38 g gain per g food for females
After 3 - 4 weeks: 0.35, 0.32, 0.32, 0.35 and 0.32 g gain per g food for males
After 3 - 4 weeks: 0.24, 0.17, 0.23, 0.20 and 0.21 g gain per g food for females

ORGAN WEIGHTS: Considerable differences between groups did not occur although some slight differences in liver weights of females were statistically significant. These differences showed no distinct dose-relationship. In addition, no correlating effects were seen in histopathology. There were no significant differences in kidney weight between treated and control animals.

GROSS PATHOLOGY: No abnormalities attributable to the ingestion.

HISTOPATHOLOGY: NON-NEOPLASTIC: No abnormalities attributable to the ingestion.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: ca. 1 005.7 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male/female
Basis for effect level:: other: Corresponding to 10000 ppm in diet. No adverse effects seen in any parameter at the highest dose level tested.
Critical effects observed:: no

Reference 7

Endpoint:: chronic toxicity: oral
Type of information:: experimental study
Adequacy of study:: supporting study
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: comparable to guideline study with acceptable restrictions
Reason / purpose for cross-reference:: reference to same study
Qualifier:: equivalent or similar to guideline
Guideline:: other: OECD Guideline 451 (Carcinogenicity Studies)
Deviations:: yes
Remarks:: (photoperiod (hrs dark / hrs light): 14 dark/10 light)
GLP compliance:: no
Limit test:: no
Specific details on test material used for the study:: - Name of test substance (as cited in study report): TK 10048
- Batch number : EN 26590/74
- Description: solid
Species:: mouse
Strain:: other: Tif: MAGf (SPF)
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source: Animal Production Stein, Ciba Geigy Ltd.
- Age at study initiation: approx. 4 weeks old
- Weight at study initiation: 24.0 - 24.5 g (male); 21.5 - 21.7 g (female)
- Housing: Groups of 5 in Macrolon cages type 3
- Diet: Nafag No. 890; ad libitum
- Water: as libitum
- Acclimation period: 7 days

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

IN-LIFE DATES: From October 1st, 1978: To: Oct 1st, 1980
Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: DIET PREPARATION
- Rate of preparation of diet (frequency): not stated
- Mixing appropriate amounts with (Type of food): TK 10048 was weighed on a calibrated balance. The pulverised food was then homogenously mixed with the appropriate concentrations of the compound and 30% water was added before pelleting to ensure the necessary pellet quality. The pellets were subsequently air-dried.
- Storage temperature of food: not stated
Analytical verification of doses or concentrations:: yes
Details on analytical verification of doses or concentrations:: Prior to the initiation of the study pretest feed samples were analysed for concentration and stability of the test material. The same was undertaken with the food batches applied during the test. These analyses were carried out in the Analytical Laboratories of the Plastics and Additives Division of Ciba Geigy Ltd, Basle/Switzerland.
Duration of treatment / exposure:: 24 months
Frequency of treatment:: daily
Dose / conc.:: 0.7 mg/kg bw/day (actual dose received)
Remarks:: males and females (dose equivalent to 5 mg/kg feed nominal concentration. Calculated by food consumption and body weight)
Dose / conc.:: 6 mg/kg bw/day (actual dose received)
Remarks:: males and females (dose equivalent to 50 mg/kg feed nominal concentration. Calculated by food consumption and body weight)
Dose / conc.:: 62 mg/kg bw/day (actual dose received)
Remarks:: males (dose equivalent to 500 mg/kg feed nominal concentration. Calculated by food consumption and body weight)
Dose / conc.:: 59 mg/kg bw/day (actual dose received)
Remarks:: females (dose equivalent to 500 mg/kg feed nominal concentration. Calculated by food consumption and body weight)
No. of animals per sex per dose:: 50
Control animals:: yes, plain diet
Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly, after 3 months: monthly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: 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: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

OTHER:
ORGAN WEIGHTS: Liver, adrenals, brain, heart, kidneys and gonads were weighed.
FOOD CONVERSION: Measured as g food consumed/kg body weight/day
Sacrifice and pathology:: GROSS PATHOLOGY: Yes (see Table 1)
HISTOPATHOLOGY: Yes (see Table 1)
Statistics:: For each time point and parameter a uni-variate statistical analysis was conducted. Due to the routine manner of the analysis system, parameter free methods were applied. Each treated group was compared to the control group in respect of dispersion and displacement. In addition a trend test was applied considering all groups.

Survival analysis was performed by the generalised Wilcoxon Test (Breslow 1970) and the generalised Savage Test (Mantel-Cox 1966). The Mantel-Cox and Breslow Tests differ in the way they weight observations. The Breslow Test gives greater weight to early observations, and is less sensitive to late events which occur when few animals on the study remain alive. Both tests are valid in large samples whether the censoring patterns (moribund sacrifice or interim sacrifice) are equal or unequal.

Statistical analysis is performed to draw attention to distinct values. A statistically significant difference between two values does not necessarily imply biological relevance of that deviation and is not conclusive for a treatment related effect.
Details on results:: CLINICAL SIGNS AND MORTALITY
No clinical signs and no signs of local and/or systemic toxicity were observed. Statistical analysis showed significant intergroup and intragroup differences in survival time. These differences were not dose dependent and therefore attributed to spontaneous variation rather than to the treatment (see Table 2 & 3).

BODY WEIGHT AND WEIGHT GAIN
The mean body weight gain of all treated male and female groups was similar to the controls.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Specific food consumption in relation to body weight of all treated groups was similar to the controls. Based on feed intake, the mean daily intake of test substance was calculated to be 0, 0.7, 6 or 62 mg/kg bw/day in males and 0, 0.7, 6 or 59 mg/kg bw/day in females.

FOOD CONVERSION
Statistical analysis of food conversion data was not performed. Specific food consumption in relation to body weight of all treated groups was similar to that of controls.

ORGAN WEIGHTS
Organ weights and ratios showed marked inter- and intra-group differences. The analysis of organ weights and ratios revealed a slight increase in brain, heart, kidney and adrenal weights in males, and a slight increase in heart, adrenal and ovary weights in females in both the high and intermediate dosage groups (50 and 500 mg/kg feed). However, as such weight variations are not unusual in aged mice, and since the histopathological findings in these organs did not indicate any difference between the controls and the treated animals, no experimental significance is attributed to these fluctuations.

GROSS PATHOLOGY
The macroscopical appearances of the treated animals were comparable to those of the controls. All gross changes seen in some treated and control mice are regarded as incidental in nature.

HISTOPATHOLOGY: NON-NEOPLASTIC
Microscopical lesions and changes found in some control and test animals and described as congenital, degenerative or inflammatory in nature are attributed to naturally occurring diseases which are common in aged mice of this breeding colony.
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: 59 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: male
Basis for effect level:: other: corresponding to 500 ppm
Dose descriptor:: NOAEL
Remarks:: (systemic)
Effect level:: 62 mg/kg bw/day (actual dose received)
Based on:: test mat.
Sex:: female
Basis for effect level:: other: corresponding to 500 ppm
Critical effects observed:: no

Endpoint conclusion

Endpoint conclusion:: no adverse effect observed
Dose descriptor:: NOAEL
: 113 mg/kg bw/day
Study duration:: chronic
Species:: rat

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

Subacute Studies

In a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test Crj: CD (SD) rats (both sexes) were treated with the test substance (99.9% pure) at dose levels of 62.5, 250 or 1000 mg/kg bw/day by gavage. Control animals received the vehicle (0.5% w/w methylcellulose). The males (12 per dose) were dosed beginning 14 days before mating and continuing until 28 days after mating (a total of 42 days). The females (12 per dose) were dosed beginning 14 days before mating, during pregnancy and up to lactation day 6 (a total of 44 - 56 days). After treatment, 6 of the 12 treated males were allowed a recovery period of 14 days. For the females, a satellite group of 6 animals was maintained for additional 14 days after treatment for controls and the mid and high dose levels.

The test substance was well tolerated by the test animals at levels up to 1000 mg/kg/day based on clinical observations and body weights. There were no test substance related effects on food consumption, functional observational battery (FOB), sensory response examination, grip strength, spontaneous motor activity measurements, clinical chemistry, macroscopic and microscopic examinations including organ weight. The NOAEL is 1000 mg/kg bw (Nihon Bioresearch Center Inc. 2007).

The study is acceptable for assessment as it was performed under GLP. Moreover, it satisfies the requirement for a Combined Repeated Dose Toxicity Study with Reproduction/developmental Toxicity Screening Test as defined by the OECD 422.

In a range finding study for a subchronic 90 day study, the test substance (no data on purity) was administered to 5 Wistar rats per sex per dose in diet at dose levels of 400, 1000, 2500 and 10000 ppm. Control animals were fed plain diets. Diets were presently ad libitum for 13 weeks. The concentrations of the test substance in diet amounted to doses of ca. 36.9, 100.2, 235.4 and 996.7 for males and 41.5, 96.7, 252.5 and 1014.8 mg/kg bw/d for females; (calculated from food consumption and body weights after 4 weeks).

There were no compound related effects in mortality, clinical signs, body weight, food consumption, organ weights (liver and kidney), or gross and histologic pathology. The NOAEL is 996.7 mg/kg bw/d and 1014.8 mg/kg bw for male and females, respectively (NOAEL combined = 1005.7 mg/kg bw).

The study is acceptable for assessment as it was range finding study in which only limited parameters were evaluated. It does not fully satisfy the requirement for a subacute oral study (OECD 407) in rodents as haematology and clinical chemistry examinations, neurobehavioural examinations and ophthalmoscopic evaluations were not performed.

Subchronic studies

In a subchronic toxicity rat study, the test substance (no data on purity) was administered to 10 Wistar rats per sex per dose in diet at dose levels of 400, 1000, 2500 and 10000 ppm. Control animals were fed plain diets. Diets were presently ad libitum for 13 weeks. The concentrations of the test substance in diet amounted to doses ofca. 25, 62, 153.9, 637.4 (males) and 28.9, 70.6, 176, 740.1 (females); (calculated from average food consumption and average body weights after 13 weeks).

There were no compound related effects in mortality, clinical signs, body weight, food consumption, haematology, clinical chemistry, organ weights, or gross and histologic pathology. The NOAEL is ca. 637.4 mg/kg bw/d and 740.1 mg/kg bw/d for males and females, respectively.

This subchronic toxicity study in the rat is acceptable but does not fully satisfy the guideline requirement for a subchronic oral study (OECD 408) in rodents as no neurobehavioural assessments, no detailed clinical examinations and no ophthalmoscopic examinations were performed.

In a subchronic toxicity dog study, the test substance (no data on purity) was administered to 4 beagle dogs per sex per dose in diet at dose levels of 0, 200, 1000 or 5000 ppm. Control animals were fed plain diets. Diets were presented once daily, 7 days per week for the duration of 13 weeks. A sentinel group of 1 dog per sex per dose was maintained post treatment for another 4 weeks on plain diet for the control and high dose groups. The concentrations of the test substance in diet amounted to doses of ca. 6.2, 29.6, 168 mg/kg bw for males and 6.5, 32.2, 153 mg/kg bw for females (calculated at weekly intervals from the consumption and individual bodyweight at the end of the week).

There were no compound related effects in mortality, clinical signs, water consumption, haematology, clinical chemistry, urinalysis, ophthalmoscopic examinations, hearing or gross and histologic pathology. Female animals receiving 5000 ppm showed a significant (p<0.01) mean weight loss of - 440 g in comparison to initial weights and by ca. 7% (- 800 g) when compared to control animals. No test substance dependent effect was seen in the weights of the male animals. In the 2 recovery animals of the high dose group, there were no marked bodyweight changes during the 4-week observation period in comparison to recovery control animals.

With the exception of one female in the control and one female in the 5000 ppm group, which consumed relatively small quantities of food during the dosing period, there were no statistically significant differences in food consumption between control and treated animals. The female animal of the high dose group showed a marked improvement during the recovery period becoming maximal from the second week onwards.

After 13 weeks, the mean liver weights for males and females in the 5000 ppm group were significantly higher than in controls by 4%. Relative liver weights to body weight for both sexes of the 5000 ppm dose group and for females that received 1000 or 200 ppm were also significantly higher than in controls. However liver weights in recovery animals were within normal limits. In view of the observed reversibility coupled with the fact that there were no histopathological findings in the liver related to treatment, the increased liver weights may reflect an adaptation to functional load. The mean heart weights for all groups of females receiving the test substance were significantly greater than controls. It was also posssible to demonstrate that the mean pituitary weight for females that received 5000 ppm significantly greater than the control values. Because there were no histopathological findings associated with the weight increase, this effect is regarded as a non-adverse, physiological and reversible adaptation to the treatment. After the 4 week treatment free period; liver and pituitary weights were within normal limits. The relative heart weight for the male dog of the high dose group was however, higher than expected. The NOAEL is ca. 168 mg/kg bw/d and 153 mg/kg bw/d for males and females, respectively.

This subchronic toxicity study in the rat is acceptable as it was performed under GLP but it does not fully satisfy the guideline requirement for a subchronic oral study (OECD 409) in non rodents as the age of the animals as initiation was higher than 9 months. In addition, the satellite groups destined to monitor reversibility or persistence of any toxic effects consisted of only 1 animal per sex at the control and high dose groups.

Chronic studies

Refer to the chapter on carcinogenicity.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. There were no significant toxic effects at doses of less than 300 mg/kg bw upon subacute oral exposure in rats. As a result, the substance is not classified for repeated dose toxicity under Regulation (EC) No. 1272/2008, as amended for the thirteenth time in Regulation (EC) No. 2018/1480.

Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.

		Week
		0	2	4	6	8	10	12
	Concentration
Male	0	52.6	111.6	179.2	235.4	271.6	292.6	311.6
	400	52.8	113.8	182.6	239.3	274.6	291.3	310.3
	1000	52.6	117.8	193	254.3	287.5	309.1	331
	2500	52.9	115.1	184.6	249.2	284.7	304.6	328.1
	10000	53	111	176.9	235	272.6	293.9	313.2

Female	0	51.2	97.3	129.2	152.8	170.6	182.3	186.1
	400	51.1	100.3	131.2	154.7	170	182.7	189.6
	1000	51	96.9	126.6	151	169.7	181.7	187.9
	2500	51	98.2	130.5	154.4	172.2	184.3	192
	10000	50.9	94.8	130.5	158.3	175	185.4	193.7

Test Preparation Administration Period		Male June 1, 2005 - July 12, 2005 [total 42 days]. Female (test group): June 1, 2005 - July 26, 2005 [total 44-56 days]. Female (recovery group): June 2, 2005 – July 13, 2005 [total 42 days]
Laboratory Animal Species, Genealogy		Rat(s) Crl: CD (SD)				Number of animals per group Control Group, Low, Medium, High Dose Group: male each 12 animals, female each 12 animals (Recovery Group, Control, Medium, High Dose Group: male each 6 animals, female each 6 animals)
Administration Route		Forced Oral Administration
Purity of Test Preparation 99.9%	Administration Dose	Control Group		Low Dose Group		Medium Dose Group		High Dose Group		Recovery Group
	(mg/kg)	0		62.5		250		1000		0		62.5		250		1000
	Sex	M	F	M	F	M	F	M	F	M	F	M	F	M	F	M	F
Body Weights Transition				-	-	-	-	-	-			-	*	-	-	-	-
Food Consumption				-	-	-	-	-	-			-	*	^DDa)	-	-	-
General Signs		-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-
Functional Observation Findings (N) FOB Rearing		12	12	12	12	12	12	12	12	*	6	*	*	*	6	*	6
Functional Observation Findings (N) FOB Rearing				-	-	-	-	-	-	*		*	*	*	^Ha)	*	-
Sensory Response				-	-	-	-	-	-	*	*	*	*	*	*	*	*
Grip Strength				-	-	-	-	-	-	*	*	*	*	*	*	*	*
Spontaneous Motor Activity				-	-	^DDa)	-	-	-	*	*	*	*	*	*	*	*
Urinary Findings				-	-	-	-	-	-	-	-	-	*	-	-	-	-
Hematological Findings (N)		6	6	6	6	6	6	5	6	6	6	6	*	6	6	6	6
Average RBC Hemoglobin Concentration				-	-	-	-	-	-			-	*	-	^Ha)	-	-
Platelet				-	^Da)	-	^DDa)	-	-			-	*	-	-	-	-
Lymphocytes				-	-	-	-	-	-			-	*	-	-	-	^HHa)
Neutrophil				-	-	-	-	-	-			-	*	-	-	-	^DDa)
Blood Chemical Findings (N)		6	6	6	6	6	6	6	6	6	6	6	*	6	6	6	6
TP				^DDa)	-	-	-	-	-			-	*	-	-	-	-
T-Bil				^DDa)	-	-	-	^DD ^a)	-			-	*	-	-	-	-
Cl				^HHa)	-	-	-	-	-			-	*	-	-	-	-
Macroscopic Findings
Number of Cases (N)		6	12	6	12	6	12	6	12	6	6	6	*	6	6	6	6
Epididymis Caudal Portion
PaleYellow Tuber		1	-	-	-	-	-	-	-	-	-	-	*	-	-	-	-
Organ Weights Transition (N)		6	11	6	12	6	12	6	11	6	6	6	*	6	6	6	6
Absolute Weights Ovary		-	-	-	-	-	-	-	-	-	-	-	*	-	^DDa)	-	-
Relative Weights Thymus Gland		-	-	-	^HHa)	-	-	-	-	-	-	-	*	-	-	-	-
Ovary		-	-	-	-	-	-	-	-	-	-	-	*	-	^DDa)	-	-

Administration Dose		Control Group			Low Dose Group			Medium Dose Group			High Dose Group
(mg/kg)		0			62.5			250			1000
Sex		M			M			M			M
Grade		-	±	+~	-	±	+~	-	±	+~	-	±	+~
Pathological findings		.	.	.	.	.	.	.	.	.	.	.	.
On termination of administration period		.	.	.	.	.	.	.	.	.	.	.	.
Lung	Foam cell gregarine (N)	4	2	0	*	*	*	*	*	*	5	1^a)	0
Intestinum jejunum	Mineraldeposition of Peyer's patch (N)	6	0	0	*	*	*	*	*	*	5	1^a)	0
Epididymis	Lymphocyte infiltration and (N)	5	1	0	*	*	*	*	*	*	6	0	0
.	Unilateral spermatozoa granuloma (N)	5	1	0	*	*	*	*	*	*	6	0	0
Prostate	Lymphocyte infiltration (N)	6	0	0	*	*	*	*	*	*	5	1^a)	0
Sex	.	F			F			F			F
Grade	.	-	±	+~	-	±	+~	-	±	+~	-	±	+~
Pathological Findings		.	.	.	.	.	.	.	.	.	.	.	.
On termination of administration period		.	.	.	.	.	.	.	.	.	.	.	.
Intestinum jejunum	Mineraldeposition of Peyer's patch (N)	6	0	0	*	*	*	*	*	*	5	1^a)	0
Spleen	Extramedullary hematopoiesis (N)	3	3	0	*	*	*	*	*	*	5	1^a)	0
Eyeball	Right retinal dysplasia (N)	6	0	0	*	*	*	*	*	*	5	1^a)	0
NOAEL		Male: 1000 mg/kg/day. Female: 1000 mg/kg/day.
Grounds for choice of NOAEL		Because no effect was observed in any parameter at 1000 mg/kg administration for male rats. Because no effect was observed in any parameter at 1000 mg/kg administration for female rats.

Organ	Dietary concentrations (ppm)	Organ weight in grams		As percentage of body weights		Adjusted for final body weight		William’s test
Organ	Dietary concentrations (ppm)	Male	Female	Male	Female	Male	Female	Male	Female
Pituitary	Control	73	68	0.0006	0.0006	-	63	N.S.	N.S
	200	68	82	0.0005*	0.0007	-	81	N.S.	N.S
	1000	83	67	0.0006	0.0006	-	70	N.S.	N.S
	5000	71	98*	0.0007*	0.0009***	-	100***	N.S.	**
Heart	Control	99.3	86.6	0.78	0.74	97.6	82.1	N.S.
	200	99.1	92.3	0.75	0.82*	94.2	91.8*	N.S.	*
	1000	104.5	91.4	0.79	0.86**	98.9	95*	N.S.	*
	5000	93.6	96.1	0.85	0.89***	105.7	98.5**	N.S.	**
Liver	Control	392.0	342.7	3.08	2.94	385	327.8	N.S.
	200	400.8	371.4	3.05	3.34*	380.4	369.7*	N.S.	*
	1000	477.3	392.1	3.62	3.66***	453.4	403.9**	N.S.	**
	5000	504.3*	453.7**	4.55***	4.19***	555.5***	461.5***	**	**

	Livers (g/100 g bw)		Kidney (g/100 g bw)
	Males	Females	Males	Females
Control	4.4	3.85	0.79	0.78
400 ppm	4.51	4.15	0.79	0.80
1000 ppm	4.40	4.10*	0.76	0.80
2500 ppm	4.80	4.28	0.80	0.87
10000 ppm	4.85	4.39**	0.81	0.83

	Average food consumption							Food efficiency
	Week							weeks
		1	2	3	4	11	12	1 to 4	11 + 12
Male	0	8.3	11.8	14.3	16.5	16.2	16.5	0.35	0.08
	400	8.3	13	14.9	17.4	15.8	18.7	0.35	0.08
	1000	8.8	12.8	15.8	18.2	16.4	17.9	0.36	0.09
	2500	8.7	12.5	15.4	17.4	16.4	17.3	0.35	0.10
	10000	8.3	12.4	14.8	16.8	17	17.6	0.34	0.08

Female	0	7.7	10.2	11	11.3	10.6	10.5	0.28	0.03
	400	8.6	11.1	12.1	12.2	11.3	11.7	0.26	0.04
	1000	8	10.6	11.4	11.7	11.9	10.9	0.26	0.04
	2500	7.9	11.1	11.9	12	11.1	11.6	0.27	0.05
	10000	7.7	11	12.9	13	12.3	12.5	0.25	0.05

Dose in feed (ppm)	Dose in mg/kg bw/day
	Male		Female
	Range	Mean	Range	Mean
200	3.99-7.08	6.2	4.43-8.99	6.5
1000	20.7-33.3	29.6	22.8-36.4	32.2
5000	113-238	168	94 -215	153

PCV %	Dietary con. ppm				Weeks dosed
					Pre dose	5	9		13	Recovery
	Control		Male		48.0	44.0	48.6		53.0	45.0
			Female		47.8	45.2	48.2		51.6	48.0
	200		Male		45.7	44.8	47.0		53.3
			Female		47.5	42.7	44.5*		49.7
	1000		Male		46.0	41.8	43.3		47.3
			Female		47.5	46.3	48.3		53.3
	5000		Male		46.2	39.4.	41.0*		45.4	37.0
			Female		47.8	39.4*	42.3**		46.2	42.0
LSD						Male	female
	5% *				4.7	5.7	3.4
	1% **				6.6	7.9	4.8
	0.1% ***				9.1	11.0	6.8
Hbg (%)	Dietary con. ppm				Weeks dosed
					Pre dose	5	9		13	Recovery
	Control	Male			15.6	14.3	15.5		16.1	15.2
		Female			15.6	14.9	15.8		15.9	16.5
	200	Male			15.6	14.9	14.9		16.4
		Female			16.1	14.0	14.0*		15.4
	1000	Male			15.2	13.8	14.3		14.8
		Female			16.0	15.3	16.2		16.5
	5000	Male			15.6	13.2	13.4*		14.2*	14.2
		Female			16.6	13.4	14.3*		14.4	16.4
LSD	5% *						Male	female
							1.9	1.4	1.7
	1% **						2.7	1.9	2.4
	0.1% ***						3.7	2.7	3.3
RBC mill/cmm	Dietary con. ppm				Weeks dosed
					Pre dose	5	9		13	Recovery
	Control			Male	6.5	5.9	6.2		6.3	6.4
				Female	6.5	6.1	6.3		6.1	6.3
	200			Male	6.5	6.1	6.2		6.5
				Female	6.6	5.7	5.7		6.1
	1000			Male	6.2	5.6	5.8		5.8
				Female	6.6	6.2	6.5		6.6
	5000			Male	6.3	5.3	5.4		5.6*	5.8
				Female	6.5	5.2*	5.4*		5.6	6.2
LSD	5% *					0.7	0.8		0.6
	1% **					1.0	1.2		0.8
	0.1% ***					1.4	1.7		1.1

Registration Dossier

Registration Dossier

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Diss Factsheets

Bumetrizole

Endpoint summary

Administrative data

Description of key information

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

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Reference 1

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Reference 6

Reference 7

Endpoint conclusion

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Additional information

Justification for classification or non-classification

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