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Description of key information

Repeated dose toxicity - oral


No adverse effects were observed in male and female rats in a 28 days and a 90 days repeated dose toxicity test conducted according to OECD 407 and OECD 408 respectively (Calvert Laboratories, Inc., 2011 and Envigo Research Limited, 2018) in which animals were exposed orally (gavage) to 0, 100, 500 or 1000 mg/kg bw/d (28 days study) and to 0, 10, 100 or 1000 mg/kg bw/d (90 days study). In both studies, an NOAEL of 1000 mg/kg bw was determined.
Repeated dose toxicity - dermal


A key study is available for the oral route of exposure. According to the REACH Regulation, only one route of exposure should be tested for repeated dose toxicity (column 2, annex VIII, section 8.6.1). The oral route of administration is considered to result in a higher systemic exposure and therefore be most predictive of the potential hazard. Therefore, it is not necessary to perform a repeated dose toxicity study via the dermal route of exposure .
Repeated dose toxicity - inhalation


A key study is available for the oral route of exposure. According to the REACH Regulation, only one route of exposure should be tested for repeated dose toxicity (column 2, annex VIII, section 8.6.1). The oral route of administration is considered to result in a higher systemic exposure and therefore be most predictive of the potential hazard. Therefore, it is not necessary to perform a repeated dose toxicity study via the inhalation route of exposure.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-10-25 to 2010-11-22
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
GLP study performed similar to OECD 407. No analyses of the dose formulations was performed, so no information is available on stability of the test substance in the dose formulations and the actual exposure concentrations.
Qualifier:
according to guideline
Guideline:
other: ICH Guidance for industry: M3 (R2) Non clinical Safety Studies for the Conduct of Human Clinical Trials and Marketing Authorization for Pharmaceuticals
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
yes
Remarks:
no analysis of the dose formulation samples was performed.
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Amine C-8
- Physical state: dark liquid
- Analytical purity: no data
- Impurities (identity and concentrations): no data
- Lot/batch No.: not provided
- Expiration date of the lot/batch: no data
- Stability under test conditions: no data
- Storage condition of test material: room temperature, 10 to 25°C
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan
- Age at study initiation: ca. 7 weeks at start of dosing
- Weight at study initiation: 231-264 g for males and 166-189 g for females
- Fasting period before study: no
- Housing: Animals were group housed by sex upon receipt and individually housed upon assignment to study in compliance with National Research Council "Guide for the care and use of laboratory animals". The room in which the animals are kept is documented in the study records. No other species are kept in the same room.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: Study animals are acclimated to their housing for a minimum of 7 days prior to their first day of dosing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 16-22°C
- Humidity (%): 14-76%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 hours light/12 hours dark, except when room lights will be turned on during the dark cycle to accomodate blood sampling or other study procedures.

Route of administration:
oral: gavage
Vehicle:
water
Remarks:
deionized water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: the test article and vehicle control preparations are prepared weekly or additionaly as needed by diluting the test article in vehicle (w/v) to reach the proper concentrations.

Dose formulation samples: On the first and last day of dosing, duplicate 1-mL samples will be obtained from top, middle, and bottom of each formulation, including the vehicle control, to determine the concentration and homogeneity of the test article in vehicle. These samples are stored at room temperature, approximately 10 to 30°C.

Dose levels: 0, 100, 500, 1000 mg/kg bw/d
Concentration: 0, 10, 50, 100 mg/ml
Dose volume: 10 ml/kg
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
28 days
Frequency of treatment:
once daily
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
actual ingested
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
actual ingested
Dose / conc.:
500 mg/kg bw/day (nominal)
Remarks:
actual ingested
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
actual ingested
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected by the sponsor in consultation with the Study Director
- Rationale for animal assignment: at random
Positive control:
No
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily, once prior to scheduled sacrifices
- Cage side observations: each animal observed for evidence of death or impending death

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: prior to each dose administration, at approximately 1-2 hours post dose, and additionally as needed; once prior to scheduled sacrifice

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded at the time of randomization/selection, prior to dose administration on Day 1 and weekly thereafter. A fasted body weight will be recorded before all scheduled sacrifices.

FOOD CONSUMPTION:
- Frequency: Full feeder weights and/or feeder weigh backs were recorded weekly for determination of food consumption.

HAEMATOLOGY: Yes / No / No data
- Time schedule for collection of blood: prior to terminal sacrifice on Day 29
- Anaesthetic used for blood collection: Yes (CO2 inhalation)
- Animals fasted: Yes (Animals will be fasted overnight (approximately 12-24 hours)
- Parameters checked: red blood cell count and morphology, white blood cell count, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean corpuscular volume, platelet count, hematocrit, hemoglobin, reticulocyte count, coagulation: activated partial thromboplastin time and prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: prior to terminal sacrifice on Day 29
- Anaesthetic used for blood collection: Yes (CO2 inhalation)
- Animals fasted: Yes (Animals will be fasted overnight (approximately 12-24 hours)
- Parameters checked: alanine aminotransferase, albumin, albumin/globulin ratio (calculated), alkaline phosphatase, aspartate aminotransferase, calcium, chloride, cholesterol, creatinine, globulin (calculated), glucose, phosphorus, potassium, sodium, total bilirubin, total protein, triglycerides, urea nitrogen


Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All surviving animals will be euthanized by CO2 asphyxiation following terminal blood collection on Day 29

Gross necropsy:
A complete gross necropsy was performed on all animals that were sacrificed or found dead during the study. The necropsy included examination of:
- the external body surface
- all orifices
- the cranial, thoracid and abdominal cavities and their contents

Organ weights: adrenals, brain, heart, kidneys, liver, testes, ovaries, spleen, thyroids/parathyroids

HISTOPATHOLOGY: Yes
For all animals necropsied, tissues were preserved in 10% neutral buffered formalin (except for the eyes, which were preserved in Davidson's fixative and the testes that were preserved in Bouin's fixed for optimum fixation). Tissues collected:
cardiovascular: aorta, heart;
digestive: salivary gland(s), tongue, esophagues, stomach, small intestine: duodenum, jejunum, ileum; large intestine: cecum, colon, rectum; pancreas, liver;
respiratory: trachea, larynx, lung with mainstem bronchus;
lymphoid/hematopoietic: sternum with bone marrow, thymus, spleen, lymph nodes, mandibular, mesenteric;
urogenital: kidneys, urinary bladder, ovaries, uterus, cervix, vagina, testes, epididymides, prostate, seminal vesicles;
endocrine: adrenals, pituitary, thyroid/parathyroid;
skin/musculoskeletal: skin, mammary gland, skeletal muscle (thigh), femur with articular surface;
nervous/special sense: eye with optic nerve, sciatic nerve, brain, spinal cord - cervical, spinal cord-midthoracic, spinal cord - lumbar, lacrimal glands
Statistics:
Statistical analysis was performed on in-life data, clinical pathology, and necropsy data when 3 or more animals are present in 2 or more dose group. Statistical analysis were be performed if N< 3 animals per group. For in-life and clinical pathology parameters, the homogeneity of the data was determined by Bartlett's Test. If the data was homogeneous, a one-way analysis of variance was performed to assess statistical significance. If statistically significant differences between the means were found, Dunnett's test was used to determine the degree of significance for the control means (p<0.05, p<0.01), the Mann-Whitney U-Test was used to determine the degree of significance from the control means (p<0.05 and p<0.01). If only 2 dose groups were present for evaluation, the Mann-Whitney U-test was used to assess statistical significance between the 2 groups. For necropsy organ weight data, the evaluation of the equality of means was made by a one-way analysis of variance using the F distribution to assess statistical significance. If statistically significant differences between the means were found, Dunnett's test was used to determine the degree of significance from the control means (p<0.05 and p<0.01).
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Statistically significant increases in absolute kidney weights (mid- and high-dose females), kidney-to-body weight ratio (high-dose females) and kidney-to-brain weight ratio (all dosed females).
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
no
Conclusions:
Rats (5 per sex per dose) were exposed once daily to 0, 100, 500, 1000 mg/kg bw/d of test substance for 28 consecutive days. The only statistically significant effects in absolute and relative female kidney weights could not be related to any macroscopic or microscopic findings or changes in clinical chemistry parameters. There is thus no indication of kidney toxicity. The NOAEL was therefore considered to be 1000 mg/kg bw/d.
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-11-06 to 2018-06-13
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
adopted 21 September 1998
Deviations:
no
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: LL150693
- Expiration date of the lot/batch: 30 December 2018
- No correction for purity was made

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark
- Solubility and stability of the test substance in the solvent/vehicle: Initially formulations were found to be stable for at least ten days and therefore formulations were prepared weekly and stored at approximately 4 °C in the dark. Further stability at twenty-one days was confirmed during the later stage of the study.

Species:
rat
Strain:
Wistar
Remarks:
RccHan(TM):WIST
Details on species / strain selection:
The rat was selected for this study as it is a readily available rodent species historically used in safety evaluation studies and is acceptable to appropriate regulatory authorities.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS (UK) Ltd., Oxon, UK
- Females (if applicable) nulliparous and non-pregnant: no data
- Age at study initiation: Approximately 7 weeks old
- Weight at study initiation: Males 182-226 g, Females 133 -168 g
- Fasting period before study: no data
- Housing: housed in groups of three or four by sex in solid floor polypropylene cages with stainless steel mesh lids and furnished with softwood flake bedding
- Diet (e.g. ad libitum): ad libitum, Rodent 2014C Teklad Global Certified Pelleted Diet
- Water (e.g. ad libitum): ad libitum, tap water
- Acclimation period: nine days

DETAILS OF FOOD AND WATER QUALITY: The diet and drinking water are considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 30 to 70%
- Air changes (per hr): At least fifteen air changes per hour
- Photoperiod (hrs dark / hrs light): twelve hours continuous light and twelve hours darkness
Route of administration:
oral: gavage
Details on route of administration:
Once daily, by gavage, using a stainless steel dosing cannula attached to a disposable plastic syringe for ninety consecutive days. The oral route was selected as the most appropriate route of exposure, based on the physical properties of the test item, and the results of the study are believed to be of value in predicting the likely toxicity of the test item to man.
Vehicle:
water
Remarks:
distilled water
Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS:
For the purpose of this study the test item was prepared at the appropriate concentrations as a solution in Distilled water. The stability and homogeneity of the test item formulations were determined by Envigo Research Limited, Shardlow, UK, Analytical Services.
Initially formulations were found to be stable for at least ten days and therefore formulations were prepared weekly and stored at approximately 4 °C in the dark. Further stability at twenty-one days was confirmed during the later stage of the study.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of each test item formulation were taken and analyzed on two occasions for concentration of the test substance at Envigo Research Limited, Shardlow, UK, Analytical Services.
The homogeneity and stability, determined with respect to the concentration of Test Item in Water formulations at nominal concentrations of 2 mg/mL and 200 mg/mL. The test item concentration in the test samples was determined by high performance liquid chromatography with mass spectrometry (HPLC-MS) using an external standard technique. The test item gave a chromatographic profile consisting of a single peak. The analytical procedure was successfully validated with respect to specificity of chromatographic analysis, linearity of detector response, method accurracy and precision. The homogeneity and stability was performed for Test Item in Distilled Water formulations at nomincal concentrations of 2 mg/mL and 200 mg/mL when stored refrigerated for 21 days and at room temperature in the light for 24 hours. The mean concentrations of Test Item in test formulations analyzed for the study were within ± 10% of nominal concentrations, confirming accurate formulation.
Duration of treatment / exposure:
90 days
Frequency of treatment:
daily
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Control (vehicle only - distilled water)
Dose / conc.:
10 mg/kg bw/day (nominal)
Remarks:
Low dose group
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
Intermediate dose group
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
High dose group
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on available toxicity data including a14 day repeated dose oral (gavage) range-finding toxicity study in rats
Positive control:
no
Observations and examinations performed and frequency:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Individual clinical observations were performed immediately before dosing, up to 30 minutes after dosing and one hour after dosing.
- Parameters: overt signs of toxicity, ill-health or behavioral change

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded on Day 1 (prior to dosing) and at weekly intervals thereafter. Individual body weights were also recorded at terminal kill.

FOOD CONSUMPTION AND COMPOUND INTAKE: yes
Food consumption was recorded for each cage group at weekly intervals throughout the study.

FOOD EFFICIENCY:
- Food conversion efficiency was calculated retrospectively.

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Water intake was observed daily, for each cage group, by visual inspection of the water bottles for any overt changes.

OPHTHALMOSCOPIC EXAMINATION: Yes
- The eyes of all Groups 1 to 4 animals were examined pre-treatment. During Week 12, the eyes of all control and high dose animals (Groups 1 and 4, respectively) were examined.
Examinations included observation of the anterior structures of the eye and following pupil dilation with 0.5% Tropicamide solution (Mydriacyl® 0.5%, Alcon Laboratories (UK) Ltd., Pentagon Park, Boundary Way, Hemel Hampstead, Hertfordshire), detailed examination of the internal structure of the eye using an ophthalmoscope was performed.

HAEMATOLOGY: Yes
Hematological investigations were performed on all animals from each test and control group at the end of the study (Day 90). Blood samples were obtained from the lateral tail vein. Where necessary repeat samples were obtained by cardiac puncture prior to necropsy on Day 91. Animals were not fasted prior to sampling.
- Parameters checked: The following parameters were measured on plasma from blood collected into tubes containing potassium EDTA anti-coagulant: hemoglobin, erythrocyte count, hematocrit, erythrocyte indices (mean corpuscular hemoglobin, volume and hemoglobin concentration), total leukocyte count, differential leukocyte count (neutrophyls, lymphocytes, monocytes, eosinophils, basophils), platelet count, reticulocyte count (methylene blue stained slides were prepared but reticulocytes were not assessed), prothrombin time was assessed by 'Innovin' and activated partial thromboplastin time was assessed by 'Actin FS' using samples collected into sodium citrate solution (0.11 mol/L).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood chemical investigations were performed on all animals from each test and control group at the end of the study (Day 90). Blood samples were obtained from the lateral tail vein. Where necessary repeat samples were obtained by cardiac puncture prior to necropsy on day 91.
- Animals fasted: No
- How many animals: all animals
- Parameters checked: urea, glucose, total protein, albumin, albumin/globulin ratio (by calculation), sodium, potassium, chloride, calcium, inorganic phosphorus, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, creatinine, total cholesterol, total bilirubin, bile acids

FUNCTIONAL OBSERVATIONS
Prior to the start of treatment and at weekly intervals thereafter, all animals were observed for signs of functional/behavioral toxicity. During Week 12 functional performances tests were also performed on all animals together with an assessment of sensory reactivity to different stimuli.
- Behavioral assessment: Detailed individual clinical observations were performed for each animal using a purpose built arena. The following parameters were observed: gait, tremors, twitches, convulsions, bizarre/abnormal/stereotypic behavior, salivation, pilo-erection, exophthalmia, lachrymation, hyper/hypothermia, skin color, respiration, palpebral closure, urination, defecation, transfer arousal, tail elevation. This test was developed from the methods used by Irwin (1968) and Moser et al (1988). The scoring system used is outlined in The Key to Scoring System and Explanation for Behavioral Assessments and Sensory reactivity Tests.
- Functional performance tests:
- motor activity : 20 purpose built 44 infra-red beam automated activity monitors were used to assess motor activity. Animals of one sex were tested at each occasion and were randomly allocated to the activity monitors. The tests were performed at approximately the same time each occasion (at least 2 hours after dosing), under similar laboratory conditions. The evaluation period was one hour for each animal. The time in seconds each animal was active and mobile was recorded for the overall one hour period and also during the final 20% of the period (considered to be the asymptotic period, reiter and Macphail, 1979).
- forelimb/hindlimb grip strength: An automated grip strength meter was used. Each animal was allowed to grip the proximal metal bar of the meter with its forepaws. The animal was pulled by the base of the tail until its grip was broken. The animal was drawn along the trough of the meter by the tail until its hind paws gripped the distal metal bar. A record of the force required to break the grip for each animal was made. Three consecutive trials were performed for each animal. The assessment was developed from the method employed by Meyer et al (1979).
- Sensory reactivity: Each animal was individually assessed for sensory reactivity to auditory, visual and proprioceptive stimuli. This assessment was developed from the methods employed by Irwin (1968) and moser et al (1988). The scoring system used is outlined in The Key to Scoring System and Explanation for behavioral Assessments and Sensory Reactivity Tests. The following parameters were observed: gasp response, vocalization, toe pinch, tail pinch, fingers approach, touch escape, pupil reflex, blink reflex, startle reflex
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
On completion of the dosing period all surviving animals were killed by intravenous overdose of a suitable barbiturate agent followed by exsanguination.
All animals were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded.

ORGAN WEIGHTS:
The following organs, removed from animals that were killed at the end of the study, were dissected free from fat and weighed before fixation:
adrenals, brain, epididymides, heart, kidneys, liver, ovaries, spleen, testes, thymus, uterus

HISTOPATHOLOGY: Yes
Samples of the following tissues were removed from all animals and preserved in buffered 10% formalin, except where stated:
adrenals, aorta (thoracic), bone & bone marrow (femur including stifle joint) - retained only and not processed, bone & bone marrow (sternum), brain (including cerebrum, cerebellum and pons), caecum, colon, duodenum, epididymides - preserved in Modified Davidson's fluid, esophagus, eyes - fixed in Davidson's fluid, gross lesions, heart, ileum (including Peyer's patches), jejunum, ovaries, pancreas, pituitary, prostate, rectum, salivary glands (submaxillary), sciatic nerve, seminal vesicles, skin, spinal cord (cervical, mid-thoracic and lumbar), spleen, stomach, testes - preserved in Modified Davidson's fluid, thymus, kidneys, liver, lungs (with bronchi) - lungs were inflated to approximately normal inspiratory volume with buffered 10% formaline before immersion in fixative, lymph nodes (mandibular and mesenteric), mammary glands, muscle (skeletal), thyroid/parathyroid, tongue - retained only and not processed, trachea, urinary bladder, uterus (with cervix), vagina
All tissues were dispatched to the Test Site (Propath UK Ltd., Willow Court, Netherwood Road, Rotherwas, Hereford, HR2 6JU) for processing (Principal Investigator: N Fower). All tissues from control and 1000 mg/kg bw/day dose group animals were prepared as paraffin blocks, sectioned at a nominal thickness of 5 μm and stained with Hematoxylin and Eosin for subsequent microscopic examination. Any macroscopically observed lesions were also processed.
Other examinations:
No data
Statistics:
Where considered appropriate, quantitative data was subjected to statistical analysis to detect the significance of intergroup differences from control; statistical significance was achieved at a level of p<0.05. Statistical analysis was performed on the following parameters:
Grip Strength, Motor Activity, Body Weight Change, Hematology, Blood Chemistry, Absolute Organ Weights, Body Weight-Relative Organ Weights.
Data were analyzed using the decision tree from the ProvantisTM Tables and Statistics Module as detailed as follows:
Where appropriate, data transformations were performed using the most suitable method.
The homogeneity of variance from mean values was analyzed using Bartlett’s test.
Intergroup variances were assessed using suitable ANOVA, or if required, ANCOVA with appropriate covariates. Any transformed data were analyzed to find the lowest treatment level that showed a significant effect using the Williams Test for parametric data or the Shirley Test for non-parametric data. If no dose response was found but the data shows nonhomogeneity of means, the data were analyzed by a stepwise Dunnett’s (parametric) or Steel (non-parametric) test to determine significant difference from the control group. Where the data were unsuitable for these analyses, pair-wise tests was performed using the Student t-test(parametric) or the Mann-Whitney U test (non-parametric).
Probability values (p) are presented as follows:
p<0.01 **
p<0.05 *
p>0.05 (not significant)
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Increased salivation was evident in animals of either sex treated with 1000 mg/kg bw/day from Day 19 (males) or Day 21 (females) throughout the majority of the treatment period. Observations such as increased post-dose salivation are often observed when animals are dosed via the oral gavage route and indicates unpalatability or slight irritancy of the dosing formulations, rather than evidence of true systemic toxicity.
Isolated instances of noisy respiration were noted in two males from each treatment group and one female each treated with 100 or 1000 mg/kg bw/day. At this low frequency, this is likely to indicate difficulty in dosing these animals on isolated occasions, rather than evidence of true systemic toxicity.
Incidental observations considered to be unrelated to treatment included one animal of either sex treated with 10 mg/kg bw/day exhibiting isolated occurrences of generalized fur loss, and two males and one female treated with 10 mg/kg bw/day showing staining of the fur/head for short periods of time.
Mortality:
no mortality observed
Description (incidence):
There were no treatment-related deaths on the study.
One female treated with 10 mg/kg bw/day was killed in extremis on Day 81 due to the severity of observations noted, these included ptosis, pilo-erection, lethargy, hypothermia, hunched posture, pallor of the extremities, dehydration, red fluid from the snout and staining around the snout. Necropsy showed a pale brain, dark colored contents throughout the gastro-intestinal tract, a raised limiting ridge in the stomach and a pale uterus. There were no histopathological changes to account for the early death.
One male treated with 10 mg/kg bw/day was found dead on Day 90 during the blood sampling procedure; no clinical signs were observed prior to death, and no abnormalities were seen at necropsy. There was tracheal necrosis at histopathology, suggesting possible dosing trauma or reflux.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Body weight:
There was no adverse effect detected for body weight development at 10, 100 or 1000 mg/kg bw/day.

Body weight gain:
Males from all treatment groups showed a statistically significant reduction (p<0.05) in body weight gain during the first week of treatment. Improvement was evident thereafter, however, body weight gain was again statistically significantly reduced (p<0.01) during Week 11 for males treated with 1000 and 100 mg/kg bw/day. During Week 12 for these males, body weight gain exceeded controls and during Week 13, body weight gain was comparable to controls. As a consequence of the slight reductions in body weight gain at 1000 mg/kg bw/day, overall body weight gain was slightly lower than controls. Overall, the fluctuations in body weight gain were not considered to represent an adverse effect based on improvement evident throughout the majority of the treatment period. No such effects were detected in any treated female animals. For more details, please refer to Table 1 in the section 'Additional information on results'.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There was no adverse effect of treatment on food consumption for either sex at 10, 100 or 1000 mg/kg bw/day.
Food efficiency:
no effects observed
Description (incidence and severity):
There was no adverse effect of treatment on food conversion efficiency for either sex at 10, 100 or 1000 mg/kg bw/day.
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
Daily visual assessment of water consumption did not reveal any significant intergroup differences.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Ophthalmoscopic examination of animals of both sexes from the control and 1000 mg/kg bw/day dose groups during Week 12 of the treatment period did not indicate any treatment-related differences.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
There were no toxicologically significant effects on the hematological parameters measured for either sex at 10, 100 or 1000 mg/kg bw/day.
* Platelet count: Males treated with 1000 mg/kg bw/day showed a statistically significant increase (p<0.01) in platelet count in relation to controls. All of the individual values were within the background control ranges (Table 2b), and in the absence of any histopathological correlates, this finding was considered to be of no toxicological significance.
* Neutrophils: Females treated with 1000 mg/kg bw/day showed a statistically significant reduction in neutrophil count in relation to controls. All of the individual values were within the background control ranges, and 5/10 individual control values actually exceeded the background control range. In isolation, and in the absence of any histopathological correlates, these findings were considered to be the result of normal biological variation.
For more details, please refer to ection 'Additional information on results'.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Assessment of blood chemistry parameters did not indicate any obvious effect of treatment for either sex at 10, 100 or 1000 mg/kg bw/day.
* Aspartate aminotransferase: Males treated with 100 or 1000 mg/kg bw/day showed a statistically significant reduction (p<0.05) in aspartate aminotransferase (ASAT); with the exception of one animal per dose group, all of the individual values were within the background control range. However, it was noted that the majority of the control values were at the higher end of the background control range. A reduction in ASAT is generally considered not an adverse effect, and therefore of no toxicological significance.
* Urea: Males treated with 1000 mg/kg bw/day also had a statistically significant reduction (p<0.05) in urea compared to the controls. The majority of the individual values were within the background control range, and without any histopathological correlates, this finding was considered to be incidental and of no toxicological significance.
* Calcium: Females from all treatment groups exhibited statistically significant increases in calcium (p<0.05); however a true dose relationship was not apparent for either parameter. The individual values for all treated females were within background control ranges, and this finding is likely due to normal biological variation.
* Bilirubin: Females from all treatment groups exhibited statistically significant increases in bilirubin (p<0.05 or p<0.01); however a true dose relationship was not apparent for either parameter. The individual values for all treated females were within background control ranges, and this finding is likely due to normal biological variation.
* Chloride: Females treated with 1000 mg/kg bw/day exhibited a statistically significant reduction (p<0.05) in chloride concentration, with 5/10 values below the background control ranges. In the absence of any corresponding histopathological changes, this reduction is considered to be a result of normal biological variation and of no toxicological significance.
For more details, please refer to Table 3a and 3b in section 'Additional information on results'.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
Behavioral assessments: There were no treatment-related changes in behavioral parameters observed
Functional performance tests: There were no treatment-related changes in functional performance. The final 20% of activity was statistically significantly lower (p<0.05) than controls for all treated males. A true dose related response was not evident, there was no effect on overall activity or any corresponding effect seen in females, and no clinical signs of neurotoxicity were evident. The intergroup differences were therefore considered not to be of toxicological significance.
Sensory reactivity assessments: There were no treatment-related changes in sensory reactivity.
For more details, please refer to Table 4 in section 'additional information on results'
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Liver weight: Both males and females treated with 100 or 1000 mg/kg bw/day showed a statistically significant increase (p<0.05-0.01) in liver weight; both absolute and relative to terminal body weight, in a dose related manner. The majority of the individual values were within the background control ranges, however, the increased weights correlate with the histopathological changes of increase rarefaction in the liver.
Kidney weight: Females treated with 100 or 1000 mg/kg bw/day showed a statistically significant increase (p<0.05 or p<0.01, respectively) in kidney weights; both absolute and relative to terminal body weight in a dose related manner. The majority of the individual absolute values were within the background control ranges; with the exception of two absolute values at 1000 mg/kg bw/day which were above these ranges. The majority of relative to body weight values were within the background control ranges, with the exception of two and four values above the background control ranges (100 and 1000 mg/kg bw/day, respectively). As there were no histopathological correlates the intergroup differences were considered to be of no toxicological significance.
For more details, please refer to Table 5a and 5b in section 'Additional information on results'.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Premature deaths: The male treated with 10 mg/kg bw/day that was found dead on Day 90 and did not show any macroscopic abnormalities. One female treated with 10 mg/kg bw/day that was killed in extremis on Day 81 exhibited a pale brain, dark colored contents throughout the gastro-intestinal tract, a raised limiting ridge in the stomach and a pale uterus.
Terminal kill: There were no toxicologically significant abnormalities detected in animals of either sex treated up to 1000 mg/kg bw/day.
The following macroscopic abnormalities detected were without treatment-related histopathogical correlates or showed a true dose-related response and were considered not to be of toxicological significance.
* One male treated with 1000 mg/kg bw/day exhibited an enlarged liver and reddened lungs.
* One further male treated with 1000 mg/kg bw/day had a raised limiting ridge in the stomach.
* One male treated with 100 mg/kg bw/day had a raised limiting ridge in the stomach.
* One female treated with 100 mg/kg bw/day exhibited a pale and mottled liver.
* A further female treated with 100 mg/kg bw/day had reddened lungs.
* One male treated with 10 mg/kg bw/day had small epididymides, prostate, seminal vesicles and testes.
* One female treated with 10 mg/kg bw/day had reddened lungs.
No macroscopic abnormalities were detected in female animals treated with 1000 mg/kg bw/day.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Premature Decedents:
There were two premature decedents during the study, both deaths were considered to be unrelated to test item administration.
* Male 30 receiving the test item at 10 mg/kg bw/day was found dead during the bleeding procedure on Day 90. At histopathology there was tracheal necrosis suggesting a possible dosing trauma or reflux.
* Female 32 receiving the test item at 10 mg/kg bw/day was killed in extremis on Day 81. There was dark colored contents in the gastrointestinal tract and several pale tissues at necropsy. No changes were noted at histopathology to account for the early death.

Terminal Kill:
Liver: Increased rarefaction was present in both males and females at 1000 mg/kg bw/day. This was considered to be associated with minor metabolic changes, and correlated with the increased liver weights at necropsy. This finding is generally considered to be non-adverse.
Histopathological findings: neoplastic:
not examined
Other effects:
not specified
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse treatment related effects observed
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
no

Table 1: Body weight gain

Groups

(sex) 

 From Day

To Day

1

8

8

15 

15

22 

22

29

29

36

36

43

43

50

50

57

57

64 

64

71 

71

78

78

85 

85

91

Abs

Gain

1

91 

 %

Gain

1

91

control

(M)

 Mean

SD

38.3

4.9

29.9

4.6

26.0

4.9

22.7

4.6

14.8

5.5

12.4

4.7

14.0

4.2

10.0

3.4 

10.5

4.8

9.2

2.4

10.9

3.0

5.8

4.5 

5.7

3.9

210.2

27.8 

99.9

11.7

10 mg/kg bw

(M)

Mean

SD

34.4*

5.5 

28.5

4.5 

23.8

3.5 

21.3

3.3 

16.2

7.2 

12.4

2.9 

14.7

3.3 

11.7

5.9

11.1

3.8 

9.6

4.6 

9.1

3.9

8.8

3.8

3.4

3.4

203.1

30.1

97.8

12.7

100 mg/kg bw

(M)

Mean

SD

34.5*

4.4

29.3

5.2

24.2

5.6 

22.1

4.9

14.1

2.1

11.2

2.5 

15.4

3.2 

10.3

27.5 

10.0

30.3

10.8

2.3 

7.4**

2.6

7.8

2.8

3.9

3.7

201.0

29.0

95.0

11.2

1000 mg/kg bw

(M)

Mean

SD

33.4*

4.7

27.3

5.4

23.9

5.3

19.6

3.2

14.9

6.0 

10.9

3.2 

14.5

5.0

8.9

3.8

8.3

4.2 

10.1

3.4

6.2**

1.6

7.7

3.0

5.3

4.2 

191.0

34.6

93.2

13.2

control

(F)

Mean

SD

13.1

3.0

13.4

4.1

9.8

4.4

11.1

6.2 

5.5

3.9

6.6

3.9

2.9

4.9

5.8

4.0

3.9

2.3

1.6

2.6

3.7

4.5 

5.4

3.2 

-1.2

3.6

81.6

7.0

52.5

3.1 

10 mg/kg bw

(F)

Mean

SD

13.1

6.0 

13.8

5.1 

8.5

4.7 

11.1

7.1 

2.8

3.7

10.3

4.1

0.5

3.5 

3.4

4.3

3.3

3.4 

4.2

4.2

1.0

4.7 

3.8

4.5 

3.9

4.0

80.4

16.5 

52.4

8.6

100 mg/kg bw

(F) 

Mean

SD

14.2

7.8 

10.2

4.7

10.6

3.8

12.4

5.9 

4.4

4.6 

5.9

3.4

5.0

4.7 

4.0

4.2

3.6

3.5

3.1

3.4 

2.2

4.3

7.3

5.0 

0.1

5.1 

 83.0

16.7

54.0

9.5

1000 mg/kg bw

(F) 

Mean

SD

15.1

5.0

10.8

2.9 

9.5

5.6

11.2

4.3 

4.9

5.4

5.8

5.5 

2.7

5.6

5.4

3.8 

3.8

3.7 

1.3

3.7

2.2

6.3 

5.3

4.1 

-0.6

5.0

77.4

11.3

50.7

7

Probability values (p) are presented as follows: p<0.01 ** - very significant; p<0.05 * - significant; p>0.05 - not significant

abbreviations: M males; F females; SD standard deviation

Table 2a: Hematological values (platelet count and neutrophils)

Groups

Sex

control

M

10 mg/kg bw

M

100 mg/kg bw

M

1000 mg/kg bw

M

control

F

10 mg/kg bw

F

100 mg/kg bw

F

1000 mg/kg bw

F

 Neut (10^9/l) (mean +/-SD)

 2.0929

+/-0.509

 1.876

+/- 0.428

 1.488

+/-0.299

 1.728

+/-0.605

 1.180

+/-0.302

 1.452

+/-0.350

1.230

+/-0.360 

 0.742**

+/-0.295

 PLT  (10^9/l) (mean +/-SD)

 507.4

+/-118.2

 567.1

+/-126.0

 580.5

+/-96.0

 659.8**

+/-89.4

541.9

+/-104.7 

 579.8

+/-142.7

 513.1

+/-179.0

 498.2

+/-213.5

Probability values (p) are presented as follows: p<0.01 ** - very significant; p<0.05 * - significant; p>0.05 - not significant

Abbreviations: Neut: neutrophils, PLT: platelet count

Table 2b: Normal Ranges for Hematological parameters in the Wistar Han™:RccHan™:WIST Strain Rat (appropriate strain and age)

 Hematology Male ranges*    No of animals  Female ranges*   No of animals 
 Neut (10^9/l)

 0.20 - 2.05

(1.12) [0.46]

 173

 0.11 -1.18

(0.65) [0.27]

 173
 PLT (10^9/l)

 473 - 732

(603) [65]

 171

 446 - 785

(615) [85]

 172

* Range = mean ± 2 standard deviations values in brackets indicate (group mean) and [standard deviation]  

Table 3a: Group Mean Blood Chemical Values

Group

(sex)

control

(M)

10 mg/kg bw

(M)

100 mg/kg bw

(M)

1000 mg/kg bw

(M)

control

(F)

10 mg/kg bw

(F)

100 mg/kg bw

(F)

1000 mg/kg bw

(F)

 Urea (mg/dl)

 44.2

+/-4.7

 45.8

+/-8.7

41.3

+/-15.5 

 32.3*

+/-11.6

 43.8

+/-7.4

 44.2

+/-4.2

54.6

+/-24.5 

41.5

+/-7.9 

 Cl- (mmol/l)

 101.5

+/-6.9

104.2

+/-7.7

102.9

+/-7.7

104.5

+/-5.3

 101.4

+/-2.1

 100.7

+/-4.0

100.4

+/-4.4 

97.3*

+/-4.9 

 Ca++ (mmol/l)

2.698

+/-0.119

2.357

+/-0.893 

2.710

+/-0.086 

2.424

+/-0.858 

 2.536

+/-0.224

 2.761*

+/-0.118

2.718*

+/-0.199 

 2.702*

+/-0.185

 ASAT (IU/l)

110 .7

+/-28.5

 99.0

+/-62.8

 85.4*

+/-21.7

 82.6*

+/-28.6

 103.3

+/-42.3

 90.4

+/-38.0

105.2

+/-28.9 

82.9

+/-19.1 

 Bili (mg/dl)

 0.847

+/-2.433

 0.130

+/-0.030

0.105

+/-0.044 

0.117

+/-0.049 

 0.063

+/-0.052

 0.114*

+/-0.025

0.082*

+/-0.054 

0.118**

0.034 

Probability values (p) are presented as follows: p<0.01 ** - very significant; p<0.05 * - significant; p>0.05 - not significant

Table 3b Normal Ranges for Blood Chemical Values in the Wistar Han™:RccHan™:WIST Strain Rat

 Blood chemistry  Male  No of animals   Female  No of animals
 Urea (mg/dl)

 30 - 55

(43) [6]

176

 30 - 62

(46) [8]

 172

 Cl- (mmol/l)

 99 - 107

(103) [2]

176 

 99 - 108

(104) [2]

 171
 Ca++ (mmol/l)

 1.24 - 3.12

(2.18) [0.47]

176

 1.22 - 3.17

(2.20) [0.49]

 177
 ASAT (IU/l)

 45 - 122

(83) [19]

 174

 21 - 173

(97) [38]

178 
 Bili (mg/dl)

0.05 - 0.16

(0.11) [0.03] 

 173

 0.04 - 0.13

(0.09) [0.02]

158

* Range = mean ± 2 standard deviations values in brackets indicate (group mean) and [standard deviation]  

Table 4: Group Mean Functional Test Values and standard Deviations (Mean +/-SD)

Group

(Sex)

control

(M)

10 mg/kg bw

(M)

100 mg/kg bw

(M)  

1000 mg/kg bw

(M)

control

(F) 

10 mg/kg bw

(F)

100 mg/kg bw

(F) 

1000 mg/kg bw

(F) 

 last 20% activity

 89.8

+/-54.3

36.8*

+/-71.5 

 34.9*

+/-48.5

38.4*

+/-44.5 

 60.1

+/-57.8

61.6

+/-49.2 

 100.7

+/-39.2

44.6

+/-47.6 

Probability values (p) are presented as follows: p<0.01 ** - very significant; p<0.05 * - significant; p>0.05 - not significant

Table 5a: Group Mean (+/- SD) Organ Weights with Corresponding Relative (% of Body Weight)

 Group

(sex)

control

(M)

10 mg/kg bw 

(M)

100 mg/kg bw 

(M)  

1000 mg/kg bw 

(M)

control

(F) 

10 mg/kg bw 

(F) 

100 mg/kg bw 

(F)   

1000 mg/kg bw 

(F) 

Kidneys (g)

 2.30695

+/-0.28602

 2.34176

+/-0.29851

2.31172

+/-0.22578 

 2.35556

+/-0.32921

1.45316

+/-0.10376 

1.45564

+/-0.11038

1.53995*

+/-0.14362

 1.60262**

+/-0.15012

Kidneys (%)

0.549

+/-0.056

0.571

+/-0.064

0.562

+/-0.045 

0.595

+/-0.037

0.614

+/-0.030 

0.626

+/-0.048

0.654*

+/-0.048 

 0.697**

+/-0.051

 Liver (g)

 13.6783

+/-1.97583

13.0220

+/-2.22944 

 14.2141*

+/-1.73269

14.4790**

+/-1.86953 

7.85257

+/-0.82498 

8.11848

+/-1.23007 

8.41874*

+/-1.17860

 8.48672*

+/-0.93354

 Liver (%)

3.245

+/-0.311

3.156

+/-0.327 

 3.444*

+/-0.228

 3.660**

+/-0.204

3.314

+/-0.302 

3.474

+/-0.313

3.559*

+/-0.296

 3.688*

+/-0.331

Probability values (p) are presented as follows: p<0.01 ** - very significant; p<0.05 * - significant; p>0.05 - not significant

Table 5b Normal Ranges for Absolute and Body Weight-Relative Organ Weight Values in the Wistar Han™:RccHan™: WIST Strain Rat

 Organ weight  Male  No of animals  Female  No of animals
 Kidney absolute weight (g)

 1.7255 - 2.8352

(2.2804) [0.2774]

 162

1.2110 - 1.8014

(1.5062) [0.1476] 

 164
 Kidney relative weight (%)

 0.4589 - 0.6654

(0.5622) [0.0516]

 165

 0.5233 - 0.7051

(0.6142) [0.0454]

 160
 Liver absolute weight (g)

 10.0828 - 16.2974

(13.1901) [1.5537]

 160

 6.5535 - 10.3234

(8.4384) [0.9425]

 162
 Liver relative weight (%)

 2.7896 - 3.7163

(3.2529) [0.2317]

161 

 2.8487 - 4.0584

(3.4536) [0.3024]

162 

* Range = mean ± 2 standard deviations values in brackets indicate (group mean) and [standard deviation]

Conclusions:
The oral administration of the test substance to rats by gavage, at dose levels of 10, 100 and 1000 mg/kg bw/day, did not result in any toxicologically significant effects for body weight development, food intake, clinical observations, hematological or blood chemistry parameters, or necropsy findings. The histopathological changes of increased rarefaction in the liver at 1000 mg/kg bw/day was considered to be non adverse, therefore the No Observed Adverse Effect Level (NOAEL) for either sex was considered to be 1000 mg/kg bw/day. The substance is therefore not to be classified as STOT RE according to criteria laid down in the CLP regulation.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
guideline studies

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated dose toxicity - oral


Two reliable repeated dose toxicity studies with the test substance are available, namely a 28 days and a 90 days study.


* 28 days repeated dose toxicity study:


Rats (5 per sex per dose) were exposed once daily to 0, 100, 500, 1000 mg test substance/kg bw/day for 28 consecutive days (Walz, 2011; Klimisch 2, equivalent to OECD guideline 407). No clinical signs of toxicity or mortality was observed. There were no treatment-related effects on body weight, food consumption, hematology or clinical chemistry. The only statistically significant effects in absolute and relative female kidney weights could not be related to any macroscopic or microscopic findings or changes in clinical chemistry parameters. There is thus no indication of kidney toxicity. The NOAEL was therefore considered to be 1000 mg/kg bw/day.


 


*90 days repeated dose toxicity study:


Rats (10 per sex per dose) were exposed once daily to 0, 10, 100, 1000 mg test substance/kg bw/day for 90 consecutive days (Edwards, 2018; Klimisch 1, according to OECD guideline 408).


The oral administration of the test substance to rats by gavage did not result in any toxicologically significant effects for body weight development, food intake, clinical observations, hematological or blood chemistry parameters, or necropsy findings, at any dose level. The histopathological changes of increased rarefaction in the liver at 1000 mg/kg bw/day was considered to be non adverse, therefore the NOAEL for either sex can be considered to be 1000 mg/kg bw/day.


 


Repeated dose toxicity - dermal


No reliable data were available for this exposure route. This type of study does not need to be performed, since a 90 -day repeated dose toxicity study is available via oral route of administration (REACH Regulation, Annex IX, Column 2 adaptation). The oral route of administration is considered to result in a higher systemic exposure and therefore be most predictive of the potential hazard. Moreover, the dermal route is not expected to be the main route of exposure.


 


Repeated dose toxicity - inhalation


No reliable data were available for this exposure route. This type of study does not need to be performed, since a 90 -day repeated dose toxicity study is available via oral route of administration (REACH Regulation, Annex IX, Column 2 adaptation). The oral route of administration is considered to result in a higher systemic exposure and therefore be most predictive of the potential hazard. Moreover, the inhalation route is not expected to be the main route of exposure.

Justification for classification or non-classification

Based on the available data and according to the criteria of the CLP Regulation, the substance should not be classified for STOT repeated exposure via the oral route.