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EC number: 460-490-0 | CAS number: 477218-42-1
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Repeated dose toxicity: oral
Administrative data
- Endpoint:
- sub-chronic toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 13-05-2004 to 26-10-2004
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Guideline study performed under GLP. All relevant validity criteria were met.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 004
- Report date:
- 2004
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: Japanese Guidelines for Screening, Toxicity Testing of Chemicals: Testing Methods for New Substances, (July 13, 1974, amended December 5, 1986)
- Deviations:
- no
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- other: US EPA OPPTS 870.3050, Repeated Dose 28-Day Oral Toxicity Study in Rodents (July 2000)
- Deviations:
- no
- GLP compliance:
- yes
- Limit test:
- yes
Test material
- Reference substance name:
- -
- EC Number:
- 460-490-0
- EC Name:
- -
- Cas Number:
- 477218-42-1
- Molecular formula:
- C18H32O3
- IUPAC Name:
- 2-[(1S)-1-[(1R)-3,3-dimethylcyclohexyl]ethoxy]-2-methylpropyl cyclopropanecarboxylate; 2-[(1S)-1-[(1S)-3,3-dimethylcyclohexyl]ethoxy]-2-methylpropyl cyclopropanecarboxylate
- Test material form:
- liquid
- Details on test material:
- Physical state: Liquid
- Storage condition of test material: room temperature (17-23°C) away from direct sunlight in original container
Constituent 1
Test animals
- Species:
- rat
- Strain:
- other: HanBrl:WIST (SPF)
- Details on species / strain selection:
- The species and strain was selected in accordance with the OECD TG 407 and the other relevant guidelines.
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Recognised supplier (reported in the full study report)
- Females (if applicable) nulliparous and non-pregnant: Yes.
- Age at study initiation: approximately 6 weeks
- Weight at study initiation: males 132.6 – 154.2 g and females 116.2 – 131.5 g (at acclimatisation); individuals were randomly allocated to treatment groups.
- Fasting period before study: None
- Housing: Makrolon type-4 cages with wire mesh tops and standardized softwood bedding. Cage distribution within the holding rack was randomized.
- Diet: Rodent Pelleted Maintenance Diet (certified supplier), ad libitum (removed overnight before blood sampling for haematology or blood chemistry and during the period of urine collection).
- Water (e.g. ad libitum): ad libitum
- Acclimation period: > 5 days.
DETAILS OF FOOD AND WATER QUALITY: Feed: Rodent Pelleted Maintenance Diet (certified supplier) – batch numbers and certificates of analysis provided in the full study report. The diet, drinking water, bedding and environmental enrichment were considered not to contain any contaminant at a level that might have affected the purpose or integrity of the study.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23
- Humidity (%): 30-70%
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12 h light / 12 h dark
IN-LIFE DATES: From: 2004-05-20 To: 2004-07-01
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- propylene glycol
- Remarks:
- PEG 300
- Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS:
The dose formulations were prepared weekly. The test item was prepared at the appropriate concentrations in PEG 300 vehicle. The required amounts of test material were weighed out into a suitable container. The formulation was magnetically stirred until all test material was thoroughly mixed. The required volume was made up with vehicle and the formulation returned to the container and mixed using a magnetic stirrer until homogenous.
DIET PREPARATION
- Rate of preparation of diet (frequency): Not applicable.
- Mixing appropriate amounts with (Type of food): Not applicable.
- Storage temperature of food: Not applicable.
VEHICLE
- Justification for use and choice of vehicle (if other than water): Applicant assessment indicates: Aqueous vehicle was not applicable due to limited solubility. PEG 300 was considered as appropriate based on test item solubility. The stability and homogeneity of the test item formulations were determined during the study. Samples for analysis of concentration and homogeneity were stored deep-frozen (about -20 °C) until analysis. Storage stability samples were collected after storage (2 hours and 7 days under storage conditions) and then delivered to the analytical laboratory where they were stored deep frozen until analysis. Results show the formulations to be homogeneous and stable for at least seven days when stored refrigerated. Stability was confirmed at concentrations of 30, 150 and 750 mg/kg or 6, 30 and 150 mg/mL following storage. Formulations were therefore prepared weekly during the treatment period, divided into daily aliquots and stored.
- Concentration in vehicle: Samples of the test item formulations were taken and analysed for concentration of test item (method of analysis provided in full study report). The results indicate that the prepared formulations were within ±6% of the nominal concentration. Formulations was assessed and confirmed at nominal concentrations, during refrigerated storage. The test item concentrations for each group are indicated in table 1.
- Amount of vehicle (if gavage): Treatment volume was 5 mL/kg for control (negative, untreated group) and all treatment groups with applicable test item concentrations per group. For further information see 'Doses / concentrations'.
- Other: Dose-formulations were analysed during the study and were reported as with ± 10 % applied limits. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- - The stability and homogeneity of the test item formulations were determined during the study. Samples for analysis of concentration and homogeneity were stored deep-frozen (about -20 °C) until analysis. Storage stability samples were collected after storage (2 hours and 7 days under storage conditions) and then delivered to the analytical laboratory where they were stored deep frozen until analysis. Results show the formulations to be homogeneous and stable for at least seven days when stored refrigerated. Stability was confirmed at concentrations of 30, 150 and 750 mg/kg or 6, 30 and 150 mg/mL following storage. Formulations were therefore prepared weekly during the treatment period, divided into daily aliquots and stored.
- The analysis consisted of GC FID analysis with internal calibration (within a dedicated formulation analysis report attached to the full study report). Delivered samples were dissolved in an appropriate volume (about 3/4 of target volume) of cyclohexane using an ultrasonic bath. Then, the 100-mL volumetric flasks were filled to the
mark with cyclohexane. Depending on the dose group, the latter sample solutions were further diluted with cyclohexane to yield concentrations within the calibration range. The concentration of test item in the final solution was quantified by GC using FID detection as detailed in the chromatographic section. The analytical method was validated (details available within the full study report).
- Mean concentrations of dose-formulations analysed during the study were within ± 10% applied limits and % difference from mean were within 6% nominal confirming accurate test item/vehicle formulation. - Duration of treatment / exposure:
- Minimum period 28 days followed by a 14 day recovery period (treatment free). The last dose was administered on Day 28.
- Frequency of treatment:
- Once daily at approximately the same time each day.
Doses / concentrationsopen allclose all
- Dose / conc.:
- 0 mg/kg bw/day (nominal)
- Remarks:
- Control
- Dose / conc.:
- 0 mg/kg bw/day (nominal)
- Remarks:
- Recovery control group
- Dose / conc.:
- 30 mg/kg bw/day (nominal)
- Remarks:
- Low - Group
- Dose / conc.:
- 150 mg/kg bw/day (nominal)
- Remarks:
- Intermediate - Group
- Dose / conc.:
- 750 mg/kg bw/day (nominal)
- Remarks:
- High - Group
- Dose / conc.:
- 750 mg/kg bw/day (nominal)
- Remarks:
- Recovery High – Group
- No. of animals per sex per dose:
- 5 per sex per dose (5 male / 5 female); 5 per sex per dose for recovery phase groups
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale: Dose levels were based on the results of a previously conducted 5-day sighting study (Report number attached to and cited in the full study report). Basis: other: nominal in vehicle (PEG 300).
- Rationale for animal assignment (if not random): Randomly assigned
- Post-exposure recovery period in satellite groups: No satellite groups.
Examinations
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily up to day 3, then once daily.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All individuals were observed in their home cages, outside their home cages in a standard arena and in the hand. These observations were performed in random sequence once before commencement of administration and once weekly (weeks 1-3) thereafter. Additional functional observations were made as ‘additional evaluations’. During week 4, relevant Functional Observational parameters from a modified lrwin screen test, grip strength, locomotor activity and of functional/behavioural toxicity were evaluated in all individuals.
BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded prior to dosing on Day 1 and at weekly intervals thereafter. Body weights were also performed prior to termination.
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: Not applicable.
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Not applicable.
- Other: Food consumption was recorded for each cage group at weekly intervals throughout the study.
FOOD EFFICIENCY: No.
WATER CONSUMPTION: No.
- Time schedule for examinations: Typically, daily. Water intake can be observed daily, for each cage group, by visual inspection. If any abnormalities are noted then further quantitative investigation is performed.
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes
- Time schedule for collection of blood: End of treatment period (day 29) for all test and control group individuals. End of recovery period (day 15; recovery phase) for all recovery group individuals.
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes, overnight/18 hours.
- How many animals: All main study and recovery.
- Parameters checked: Erythrocyte count, Hemoglobin, Hematocrit, Mean corpuscular volume, Red cell volume distribution width, Mean corpuscular haemoglobin, Mean corpuscular hemoglobin concentration, Hemoglobin concentration distribution width, Platelet (thrombocyte) count, Reticulocyte count, Reticulocyte maturity index, Methemoglobin, Totalleukocyte count, Differential leukocyte count, Coagulation: Thromboplastin time, Activated partial thromboplastin time
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: End of treatment period (day 29) for all test and control group individuals. End of recovery period (day 15; recovery phase) for all recovery group individuals
- Animals fasted: Yes, overnight/18 hours.
- How many animals: All main study and recovery.
- Parameters checked: Glucose, Urea, Creatinine, Bilirubin, total, Cholesterol, total, Triglycerides, Phospholipids, Aspartate aminotransferase, Alanine aminotransferase, Lactate dehydrogenase, Glutamate dehydrogenase, Creatine kinase, Alkaline phosphatase, Gamma-glutamyl-transferase, Sodium, Potassium, Chloride, Calcium, Phosphorus inorganic, Protein, total, Albumin, Globulin, Albumin/Globulin ratio
URINALYSIS: Yes
- Time schedule for collection of urine: Urinalytical investigations were performed on all test and control group animals during day 29 fasting period and on all during fasting in the recovery.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes (food withheld during time of urine collection; overnight/18 hours)
- Parameters checked: Volume (18 hours), Specific gravity (relative density), Colour, Appearance, pH, Nitrite, Protein, Glucose, Ketones, Urobilinogen, Bilirubin, Erythrocytes, Leukocytes
NEUROBEHAVIOURAL EXAMINATION: Yes. Was conducted as part of ‘special evaluations’
- Time schedule for examinations: Additional functional observations were made as ‘additional evaluations’. During week 4, relevant Functional Observational parameters from a modified lrwin screen test, grip strength, locomotor activity and of functional/behavioural toxicity were evaluated in all individuals.
- Dose groups that were examined: All.
- Battery of functions tested: sensory activity / grip strength / motor activity
IMMUNOLOGY: No
OTHER: Additional post-termination observations were made at necropsy. - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
- organs weighed: Adrenal glands
Aorta, Bone (sternum, femur including joint), Bone marrow (femur), Brain (4 levels), Cecum, Colon, Duodenum, Epididymides (fixed in Bouin's solution), Esophagus, Eyes with optic nerve (fixed in Davidson's solution), Harderian gland (fixed in Davidson's solution), Heart, Ileum, with Peyer's patches, Jejunum with Peyer's patches, Kidneys, Larynx, Lacrimal gland (exorbital), Liver, Lungs (infused with formalin at necropsy), Lymph nodes (mesenteric, mandibular), Mammary gland area, Nasal cavity, Ovaries, Pancreas, Pituitary gland, Prostate gland (incl. coagulating gland), Rectum, Salivary glands (mandibular, sublingual), Sciatic nerve, Seminal vesicles, Skeletal muscle, Skin, Spinal cord (cervical, midthoracic, lumbar), Spleen, Stomach, Testes (fixed in Bouin's solution), Thymus, Thyroid (incl. parathyroid gland), Tongue, Trachea, Urinary bladder (infused with formalin at necropsy), Uterus, Vagina, Gross lesions
HISTOPATHOLOGY: Yes
- Organs and tissues preserved in neutral buffered 4% formalin (unless otherwise indicated):
Adrenal glands, Bone marrow (femur), Brain (4 levels), Cecum, Colon, Duodenum, Epididymides (fixed in Bouin's solution), Heart, Ileum, with Peyer's patches, Jejunum with Peyer's patches, Kidneys, Liver, Lungs (infused with formalin at necropsy), Lymph nodes (mesenteric, mandibular), Ovaries, Prostate gland (incl. coagulating gland), Rectum, Sciatic nerve, Seminal vesicles, Spinal cord (cervical, midthoracic, lumbar), Spleen, Stomach, Testes (fixed in Bouin's solution), Thymus, Thyroid (incl. parathyroid gland), Trachea, Urinary bladder (infused with formalin at necropsy), Uterus, Vagina, Gross lesions. Microscopic analysis was conducted thereof. Any macroscopically observed lesions were also processed. - Statistics:
- The following statistical methods were used to analyze the grip strength, locomotor activity, body weight, organ weights and ratios:
The Dunnett-test (many to one t-test) based on a pooled variance estimate were applied if the variables could be assumed to follow a normal distribution for the comparison of the treated groups and the control groups for each sex. The Steel-test (many-one rank test) were applied instead of the Dunnett-test when the data cannot be assumed to follow a normal distribution. Fisher's exact-test were applied to the macroscopic findings.
The following statistical methods were used for statistical analysis of clinical laboratory data:
Quantitative data were analyzed by a one-way analysis of variance (ANOVA) when the variances are considered homogeneous according to Bartlett. Alternatively, if the variances are considered to be heterogenous (p < or = 0.05), a non-parametric Kruskai-Wallis test was used. Treated groups were compared to the control groups using Dunnett's test if the ANOVA was significant at the 5% level and by Dunn's test in the case of a significant Kruskai-Wallis test (p < or = 0.05).
Results and discussion
Results of examinations
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: episodes of sedation were noted intermittently in males (days 17-18 and 24-26) and persistently in females (days 16-28). Emaciation was recorded in one female treated with 750 mg/kg bw/day and in two females during the first week of recovery.
No other findings were considered of toxicological relevance. - Mortality:
- no mortality observed
- Description (incidence):
- There was no test item related mortality.
One control male (#10) was found dead on treatment day 12. - Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: mean body weights of females were slightly lower (-5%) during the treatment period than those of the control females and were considered to be test item related. During recovery no effects attributed to treatment were noted.
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: females treated indicated mean daily food consumption values were slightly lower than those of the controls. During recovery no differences were noted.
- Food efficiency:
- no effects observed
- Description (incidence and severity):
- See body weight and weight changes sections.
- Water consumption and compound intake (if drinking water study):
- not examined
- Description (incidence and severity):
- No effects on water consumption were reported nor in related clinical signs/functional observations or urinalysis investigations were anything significant noted.
- Ophthalmological findings:
- not examined
- Description (incidence and severity):
- There were no toxicologically significant reported effects to the eyes (in life or post termination) in the parameters examined.
- Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: mean absolute and relative reticulocyte counts of the females were reduced, after four weeks of treatment. Both values exceeded the lower limit of the historical control values. The mean reticulocyte maturity indices were shifted towards cells of low fluorescence (increased) from middle fluorescence (reduced) and high fluorescence (reduced), which generally indicates retention of older cells and slower/impaired replacement of younger cells. This finding was considered to be test item related, but was largely reversible after 2 weeks of recovery.
After two weeks recovery all other differences were within the ranges of historical controls. - Clinical biochemistry findings:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: Aspartate aminotransferase activity was increased in males/females and lactate dehydrogenase activity was elevated in females when compared with the controls. These values exceeded the ranges of the historical control data and were considered related to metabolic activation caused by the test item. These findings were, however, reversible during the recovery period and therefore considered to be non-adverse. After two weeks recovery all differences were within the ranges of historical controls.
- Urinalysis findings:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: Elevated ketone noted in the urinalysis parameters of males/females after 4 weeks of treatment, was considered possibly related to metabolism of the test item. After 2 weeks of recovery, this finding was no longer observed. Indicating recovery.
- Behaviour (functional findings):
- effects observed, treatment-related
- Description (incidence and severity):
- No functional observation findings were considered of toxicological relevance.
At 750 mg/kg bw/day: reduced mean hindlimb grip strength was noted in males whereas females had reduced mean fore- and hind-limb grip strength. These differences were considered to be test item related. Reductions of locomotor activity were noted in males and females. - Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: males had elevated absolute and relative liver weights, as well as elevated kidney to body weight ratios. Females had elevated absolute and relative liver weights, elevated absolute and relative kidney weights and reduced absolute and relative spleen weights. Additionally, elevated heart-to-body weight ratio when compared with controls. After two weeks recovery, no test item-related changes were noted in males or females previously treated with 750 mg/kg bw/day indicating recovery.
At 150 mg/kg bw/day: males had elevated liver-to-body weight ratios and elevated kidney-to-body weight ratios noted after four weeks of treatment. Females had elevated absolute and relative liver weights, and reduced absolute spleen weights. - Gross pathological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: item-related macroscopic findings were recorded in the Harderian Glands (black foci: one female after treatment, and five recovery females).
At 150 mg/kg bw/day: item-related macroscopic findings were recorded at the liver (clay-coloured: 4 females). - Neuropathological findings:
- no effects observed
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- At 750 mg/kg bw/day: The test item induced histopathological changes in the heart, skeletal muscle, kidneys, Harderian gland, liver, and thyroid.
HEART : minimal to severe sarcoplasmic vacuolation, minimal to moderate single cell necrosis, increased incidence and mean grade of mononuclear foci, and minimal myocardial (interstitial) fibrosis were recorded in main study males/females treated with 750 mg/kg/day or 150 mg/kg/day, reflecting a cardiotoxic potential of the test item. After a 14-day treatment-free recovery period, minimal myocardial fibrosis was recorded in two females previously treated with 750 mg/kg/day. These changes were considered adverse effects.
SKELETAL MUSCLE: minimal to slight mixed-cellular, interstitial inflammation, accompanied by minimal to slight interstitial edema, minimal single fiber necrosis, and partly minimal muscle cell regeneration was recorded in some main study females treated with 750 mg/kg/day. These changes were considered adverse effects.
KIDNEYS: increased incidence and mean grade of hyaline droplets and of tubular basophilia was recorded in males treated with 750 mg/kg/day and 150 mg/kg/day.
The incidence and mean grade of tubular basophilia was still increased in recovery males. In rats, these changes were adverse effects, whose relevance for human health risk assessment is unclear. Applicant assessment indicates that this may be: cortical hyaline droplets that could be considered to represent alpha2uglobulin, which is a male rat-specific lesion, supported by absence of these kidney related effects in females. This is suggestive of this effect not being relevant to human health. However, further evidence may be required to support that hypothesis.
HARDERIAN GLAND: minimal acinar degeneration, minimal to slight acinar hyperplasia, and increased porphyrin deposition was recorded in females treated with 750 mg/kg/day or 150 mg/kg/day. Although these findings were considered adverse effects in the rat, their relevance for human health risk assessment are unclear.
LIVER: increased incidence and mean grade of liver fatty change was recorded in main study females treated with 750 mg/kg/day or 150 mg/kg/day. After recovery this finding showed tendency to regression. The toxicological relevance of this finding is unclear. Additionally an increased incidence and mean grade of hepatocellular hypertrophy was recorded. This was absent from recovery males/females. This finding was not an adverse effect.
THYROID: an increased incidence of minimal follicular hypertrophy in males. This was absent from recovery males/females. This finding was not considered an adverse effect.
At 150 mg/kg bw/day: see mentioned sections above (HEART, KIDNEYS, HARDERIAN GLAND and LIVER) - Histopathological findings: neoplastic:
- no effects observed
- Description (incidence and severity):
- There were no treatment-related abnormalities detected.
- Other effects:
- no effects observed
Effect levels
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 30 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- body weight and weight gain
- clinical signs
- gross pathology
- histopathology: non-neoplastic
Target system / organ toxicity
- Critical effects observed:
- no
Applicant's summary and conclusion
- Conclusions:
- Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for males and females is defined as 30 mg/kg body weight per day in males and females.
- Executive summary:
The study was performed according the requirements of OECD TG 407, EU method B.7 and Japanese Guidelines for Screening, Toxicity Testing of Chemicals: Testing Methods for New Substances guidelines under GLP conditions. Following a previously conducted 5-day sighting study, the systemic toxic potential of the test item was assessed orally in a 28 day gavage study in HanBrl:WIST (SPF) rats. Recovery from any effects was evaluated during a subsequent 14 day recovery period. Three groups, each comprising five male and five female rats, received test item at doses of 30, 150 or 750 mg/kg bw/day. A control group of five males and five females was dosed with vehicle alone (PEG 300) at a dose volume of 5 mL/kg. Two recovery groups, each of five males and five females, were treated with the high dose (750 mg/kg bw/day) or the vehicle alone for twenty-eight consecutive days and then maintained without treatment for a further fourteen days. Clinical signs, functional observations including sensory reactivity, grip strength and motor activity were performed, body weight change, food consumption were monitored during the study. Hematology, blood chemistry and urinalysis were evaluated for all non-recovery group animals at the end of the treatment period and for all recovery group animals at the end of the treatment free period. All individuals were subjected to gross necropsy examination and at termination. Histopathological examination of selected tissues was performed. There was no test item related effect on mortalities. At 750 mg/kg bw/day: episodes of sedation were noted intermittently in males (days 17-18 and 24-26) and persistently in females (days 16-28). Emaciation was recorded in one female treated with 750 mg/kg bw/day and in two females during the first week of recovery.No other findings were considered of toxicological relevance. In functional observations, reduced mean hindlimb grip strength was noted in males whereas females had reduced mean fore- and hind-limb grip strength. These differences were considered to be test item related. Reductions of locomotor activity were noted in both sexes. Females treated indicated mean daily food consumption values were slightly lower than those of the controls. During recovery no differences were noted.Females treated indicated mean daily food consumption values were slightly lower than those of the controls. During recovery no differences were noted.Aspartate aminotransferase activity was increased in males/females and lactate dehydrogenase activity was elevated in females when compared with the controls. These values exceeded the ranges of the historical control data and were considered related to metabolic activation caused by the test item. These findings were, however, reversible during the recovery period and therefore considered to be non-adverse. After two weeks recovery all differences were within the ranges of historical controls.In urinalysis at 750 mg/kg bw/day: elevated ketone noted in males/females after 4 weeks of treatment, was considered possibly related to metabolism of the test item. After 2 weeks of recovery, this finding was no longer observed. Indicating recovery. Males had elevated absolute and relative liver weights, as well as elevated kidney to body weight ratios. Females had elevated absolute and relative liver weights, elevated absolute and relative kidney weights and reduced absolute and relative spleen weights. Additionally, elevated heart-to-body weight ratio when compared with controls. After two weeks recovery, no test item-related changes were noted in males or females previously treated with 750 mg/kg bw/day indicating recovery. At 150 mg/kg bw/day: males had elevated liver-to-body weight ratios and elevated kidney-to-body weight ratios noted after four weeks of treatment. Females had elevated absolute and relative liver weights, and reduced absolute spleen weights.At 750 mg/kg bw/day: item-related macroscopic findings were recorded in the Harderian Glands (black foci: one female after treatment, and five recovery females, whereas at 150 mg/kg bw/day: item-related macroscopic findings were recorded at the liver (clay-coloured: 4 females).At 750 mg/kg bw/day: The test item induced histopathological changes in the heart, skeletal muscle, kidneys, Harderian gland, liver, and thyroid. In the heart : minimal to severe sarcoplasmic vacuolation, minimal to moderate single cell necrosis, increased incidence and mean grade of mononuclear foci, and minimal myocardial (interstitial) fibrosis were recorded in main study males/females treated with 750 mg/kg/day or 150 mg/kg/day, reflecting a cardiotoxic potential of the test item. After a 14-day treatment-free recovery period, minimal myocardial fibrosis was recorded in two females previously treated with 750 mg/kg/day. These changes were considered adverse effects. In skeletal muscle: minimal to slight mixed-cellular, interstitial inflammation, accompanied by minimal to slight interstitial edema, minimal single fiber necrosis, and partly minimal muscle cell regeneration was recorded in some main study females treated with 750 mg/kg/day. These changes were considered adverse effects. In kidneys: increased incidence and mean grade of hyaline droplets and of tubular basophilia was recorded in males treated with 750 mg/kg/day and 150 mg/kg/day. The incidence and mean grade of tubular basophilia was still increased in recovery males. In the Harderian gland: minimal acinar degeneration, minimal to slight acinar hyperplasia, and increased porphyrin deposition was recorded in females treated with 750 mg/kg/day or 150 mg/kg/day. Although these findings were considered adverse effects in the rat, their relevance for human health risk assessment are unclear. In the liver: increased incidence and mean grade of liver fatty change was recorded in main study females treated with 750 mg/kg/day or 150 mg/kg/day. After recovery this finding showed tendency to regression. The toxicological relevance of this finding is unclear. Additionally an increased incidence and mean grade of hepatocellular hypertrophy was recorded. This was absent from recovery males/females. This finding was not an adverse effect. In the thyroid: an increased incidence of minimal follicular hypertrophy in males. This was absent from recovery males/females. This finding was not considered an adverse effect. The oral (gavage) administration of the test item to males/females at dose levels of 30, 150 or 750 mg/kg bw/day resulted indicated that, the No-Observed-Effect-Leve (NOEL) was 30 mg/kg bw/day and the No-Observed-Adverse-Effect-Level (NOAEL) was regarded to be 30 mg/kg bw/day for males/females.
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