3-(4,4-dimethylcyclohex-1-en-1-yl)propanal

Endpoint summary

Description of key information

Oral: NOAEL (rat (male/female) = 150 mg/kg bw/day, (oral gavage to rats; corn oil vehicle : 0 (control), 75, 150, and 300mg/kg/day), Repeated dose 28-day oral study with 14-day recovery, OECD TG 407, GLP. 2010

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30-07-2009 to 17-09-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Guideline study performed under GLP. All relevant validity criteria were met.

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:

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 (incl. QA statement)

Remarks:

inspected: July 2007 ; signature: November 2007

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

Crl:WU

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 7 to 8 weeks
- Weight at study initiation: males 216.9 – 247.9 g and females 151.5 – 181.2 g; individuals were randomly allocated to treatment groups.
- Fasting period before study: None
- Housing: Randomly assigned into four groups consisting of 6 or 12 animals per sex in each group (12 rats for recovery investigations in group 1 and 4). Makrolon (polycarbonate) cages type III, two rats per cage, and were maintained under conventional laboratory conditions. Cages and absorbing softwood
bedding material (certified) were changed twice a week or more often as required. Cage distribution within the holding rack was randomized.
- Diet: Rodent 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 (except during urine collection)
- Acclimation period: at least 7 days.

DETAILS OF FOOD AND WATER QUALITY: Feed: Rodent Maintenance Diet – 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): 20-24 (i.e. 22 ± 2°C)
- Humidity (%): 40-70% (i.e. 55 ± 15%
- Air changes (per hr): Filtered fresh air which was passed to atmosphere and not recirculated.
- Photoperiod (hrs dark / hrs light): 12 h light / 12 h dark

IN-LIFE DATES: From: 14-12-2009 To: 27-01-2010

Route of administration:

oral: gavage

Details on route of administration:

In accordance with the OECD TG 407 guideline.

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The dose formulations were prepared twice weekly (specifically up to 3 days in advance of dosing). During the 4-week test item administration period, analyses of the test item formulations were done after preparation of stock emulsions of the test item in corn oil. On day 0, 7, 14 and 21 (day 1: first day of test item administration) aliquots were taken from the stock emulsions and subjected to an analysis. Test item stability in the corn oil formulation (vehicle) for at least 3 days was confirmed by analysis. No comments as to inhomogeneity were made in the study report. During testing the actual/nominal concentration (%) was range 101.0% to 104.7% with mean 103.5% in vehicle corn oil, for all groups. Test item formulations were stored in the dark in a refrigerator (< 10°C) until administration. The test item was formulated in an appropriate vehicle (i.e. corn oil) resulting in a concentration of 10, 20, and 40 mg/mL (low, mid, high dose). 1.5 mL per individual (200 gram body weight) of this formulation was administered by gavage to the individuals amounting to a dose of 75, 150, and 300 mg/kg (exact volumes were calculated based on actual body weights). Individuals of group 1 received the same amount of the vehicle (corn oil) daily by oral gavage. During the course of the study, the concentration/homogeneity of the test item in the prepared corn oil formulation were checked for all dose groups (group 2 - 4) once a week. No comments as to checked homogeneity or the concentrations were made in the study report. All doses were based on nominal concentrations that were apparently analytically confirmed.

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. Corn oil was considered as appropriate based on test item solubility. The stability and homogeneity of the test item formulations were determined during the study. Test item stability in the corn oil formulation (vehicle) for at least 3 days was confirmed by analysis. No comments as to inhomogeneity were made in the study report. During testing the actual/nominal concentration (%) was range 101.0% to 104.7% with mean 103.5% in vehicle corn oil, for all groups. During the course of the study, the concentration/homogeneity of the test item in the prepared corn oil formulation were checked for all dose groups (group 2 - 4) once a week. No comments as to checked homogeneity or the concentrations were made in the study report. All doses were based on nominal concentrations that were apparently analytically confirmed.
- 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. Corn oil 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 7.5 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 homogeneity and stability was confirmed in corn oil formulations. The dose formulations were prepared twice weekly (specifically up to 3 days in advance of dosing). During the 4-week test item administration period, analyses of the test item formulations were done after preparation of stock emulsions of the test item in corn oil.
- Test item stability in the corn oil formulation (vehicle) for at least 3 days was confirmed by analysis. No comments as to inhomogeneity were made in the study report. During testing the actual/nominal concentration (%) was range 101.0% to 104.7% with mean 103.5% in vehicle corn oil, for all groups.
- The analysis consisted of NMR analysis (within a dedicated formulation analysis report attached to the full study report).
During the course of the study, the concentration/homogeneity of the test item in the prepared corn oil formulation were checked for all dose groups (group 2 - 4) once a week. No comments as to checked homogeneity or the concentrations were made in the study report. All doses were based on nominal concentrations that were apparently analytically confirmed. 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 5% 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.

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Control

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Recovery control group

Dose / conc.:

75 mg/kg bw/day (nominal)

Remarks:

Low - Group

Dose / conc.:

150 mg/kg bw/day (nominal)

Remarks:

Intermediate - Group

Dose / conc.:

300 mg/kg bw/day (nominal)

Remarks:

High - Group

Dose / conc.:

300 mg/kg bw/day (nominal)

Remarks:

Recovery High - Group

No. of animals per sex per dose:

3 per sex per dose (3 male / 3 female)
Additional 6 animals per sex and group (control and high dose group only) were treated for 28 days and then allowed a 14-day treatment-free recovery period.

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels were based on the results of a previously conducted studies (acute oral toxicity indicating toxicity between 300 mg/kg bw and 2000 mg/kg bw) and all available information. The dose range was 0 (control), 75, 150, and 300 mg/kg bw/day in group 1, 2, 3 and 4 respectively. Basis: other: nominal in vehicle (Corn Oil)
- Rationale for animal assignment (if not random): Randomly assigned
- Post-exposure recovery period in satellite groups: 14 days (six individuals: 3 males and 3 females in group 1 (0 mg/kg bw/day) control and group 4 (300 mg/kg bw/day) high groups, respectively.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily. (Including general condition, fur, grooming activity, visible mucous membranes, behaviour and locomotor activity (lethargy, coma, convulsions, diarrhoea, salivation), central nervous symptoms, breathing pattern) See “Detailed Clinical Observations” and/or “Neurobehavioural Examination” for further information.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All individuals were examined for overt signs of toxicity, ill-health or behavioural change immediately before dosing, up to thirty minutes post dosing and one to two hours after dosing. Additionally, animals were observed daily throughout the study. All observations were recorded. Additional functional observations were made as ‘additional evaluations’ consisting of Functional observation battery (FOB) and Motor activity assessment (MOTI), one day during week 4 (all groups) and/or one day during week 6 (recovery groups).

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 and, in the case of recovery group animals 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 at weekly intervals throughout the study.

FOOD EFFICIENCY: No.

WATER CONSUMPTION: Yes
- Time schedule for examinations: Daily. Water intake was recorded at weekly intervals throughout the study.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: End of treatment period (day 29) for all non-recovery 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 (ca. 16 hours)
- How many animals: All main study and recovery.
- Parameters checked: Hemoglobin (Hb), Erythrocyte count (RBC), Hematocrit (Hct), Erythrocyte indices – including: mean corpuscular hemoglobin (MCH), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), Total leukocyte count (WBC), Differential leukocyte count – including: neutrophils (Neut), lymphocytes (Lymph), monocytes (Mono), eosinophils (Eos), basophils (Bas), Platelet count (PLT). Morphology: Anisocytosis, Macrocytosis, Microcytosis, , Hypochromasia, Hyperchromasia, would also typically be checked.
Additionally: Prothrombin time (PT) was assessed and Activated partial thromboplastin time (APTT) was assessed.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: End of treatment period (day 29) for all non-recovery test and control group individuals. End of recovery period (day 15; recovery phase) for all recovery group individuals.
- Animals fasted: Yes, overnight ca. 16 hours.
- How many animals: All main study and recovery.
- Parameters checked: Alkaline phosphatase (ALP), Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Gamma-glutamyl transferase (gGT), Total bilirubin (Bili), Total bile acids (Bi Ac), Urea, Urea nitrogen (BUN), Creatinine (Creat), Glucose (Gluc), Total cholesterol (Chol), Triglycerides (Trig), Sodium (Na), Potassium (K), Chloride (Cl), Calcium (Ca), Inorganic phosphorus (Phos), Total protein (Total Prot), Albumin (Alb), Albumin/globulin ratio (A/G Ratio) was calculated

URINALYSIS: Yes
- Time schedule for collection of urine: Urinalytical investigations were performed on all non-recovery test and control group animals during day 29 and on all recovery group animals during days 15.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes (food withheld during time of urine collection; overnight ca. 16 hours)
- Parameters appearance, volume, specific gravity, pH, protein, glucose and blood were measured semi quantitatively

NEUROBEHAVIOURAL EXAMINATION: Yes. Was conducted as part of ‘special evaluations’
- Time schedule for examinations: Consisting of Functional observation battery (FOB) and Motor activity assessment (MOTI), one day during week 4 (all groups) and/or one day during week 6 (recovery groups).
- 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: the lung (including 2/3 of trachea), liver, adrenals, kidneys, testes, epididymides, uterus, ovaries, thymus, spleen, brain and heart (paired organs weighted separately). Terminal body weight, and relative organ weight data were also computed.

HISTOPATHOLOGY: Yes
- Organs and tissues preserved in neutral buffered 10% formalin: brain, pituitary, tongue, eyes, lachrymal glands, harderian glands, nasal and paranasal
cavities, larynx, pharynx, trachea, thyroid, parathyroids, lungs, thymus, heart, aorta, lung associated lymph nodes, salivary glands, mandibular lymph nodes, liver, pancreas, spleen, kidneys, adrenals, esophagus, forestomach, glandular stomach, duodenum, jejunum, ileum, caecum, colon, rectum, mesenterium and lymph nodes, urinary bladder, testes, epididymides, prostate, seminal vesicles, ovaries, uterus (including cervix), vagina, mammary glands, skeletal muscle, femur with bone marrow and joint, spinal cord, skin, peripheral nerve (N. ischiadicus), sternum with bone marrow. Tissues for histological examination were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned at 3-4 μm, and stained with hematoxylin and eosin. Bones were decalcified prior to embedding. For fixation of the testes, a modified Davidson's fluid (15% ethanol, 5% glacial acetic acid and 30% formalin) were used. Microscopic analysis was conducted thereof. Any macroscopically observed lesions were also processed.

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. Body weight and food/water consumption data were analysed using analysis of variance as a global test. Pair wise comparison of the means of the treatment groups with the means of the corresponding control group was performed per sex using Dunnett's
modification of the t-test. Thus the experiment wise error rate was controlled in this multiple testing procedure. For comparisons between two treatment groups, the two-sided t-test at a level of p = 0.05 was used. If applicable, other statistical tests were used. Statistical evaluation for histopathological findings was as follows: Significance of differences of the frequencies was evaluated as pair wise comparison between vehicle control and treatment groups using
Fisher‘s exact test. These tests were performed at the local significance level of p = 0.05.

Clinical signs:

no effects observed

Description (incidence and severity):

There were no test item related clinical signs in relation to treatment.

Activity levels were reduced (short-term lethargy), however, grooming activity started directly after treatment. Some individuals presented signs of minor discomfort returning to normal within 0.5 – 1.0 hour. It was considered that these signs were from test item administration rather than from toxicity.

Mortality:

no mortality observed

Description (incidence):

There was no test item related mortality.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

There was no test item related effect in bodyweight gain.

The statistical significance observed in the female high dose group started already at the beginning of the study and was due to an incidental imbalance in body weights. As the standard deviation of the concurrent control is small a continuous significance was observed. This was not considered a treatment related effect within the study.

Food consumption and compound intake (if feeding study):

not examined

Description (incidence and severity):

There was no test item related effect in food consumption.

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):

effects observed, treatment-related

Description (incidence and severity):

At 300 mg/kg bw/day dose level: males/females water consumption was increased relative to controls. This returned to normal levels in the 14-day recovery period. This observation is considered as an adaptative response, related to test item administration.

Ophthalmological findings:

not examined

Description (incidence and severity):

There were no substance related and 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):

For red blood cells, hemoglobin and hematocrit : statistically significant decreases were observed in the male groups and partially in the female groups. Although these were considered as test-item related alterations (without dose-response) - the alterations were very slight and the values are still in the range usually observed for this species, strain, sex and age of rat (historical data of test laboratory). No treatment related adverse effects were detected in other haematology parameters at day 29. On day 42 (14 days after end of treatment) no statistically significant effects were observed, therefore the mentioned significant decreases on day 29 showed full reversibility and are therefore adaptive responses.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

There were no substance related and toxicologically significant reported effects in the parameters examined.

Urinalysis findings:

no effects observed

Description (incidence and severity):

There were no substance related and toxicologically significant reported effects in the parameters examined.

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

There was no test item related effect on sensory reactivity or grip strength.

Some statistically significant findings, e.g. for the male high dose group in the recovery period were considered as incidental findings.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Statistically significant and dose-dependent increases were observed in both sexes for liver and kidney weights (effects more pronounced in males). The effect in livers recovered within 14 days and was not accompanied by a morphological correlate. Therefore, this finding is considered as an adaptative response (possibly caused by the uptake of water). Increased water consumption correlated with increased kidney weights and an increased urine volume. As no adverse histopathological findings were detected in kidneys the effects are considered as treatment-related, adaptative effects.

Specially, absolute and relative liver and kidney weights showed a dose-dependent increase in males on day 29, however, these effects recovered during the 14-day post observation period (left kidney weights still increased tot 5% level). Females principally showed the same tendency in liver and kidney weights, however, with statistical significance in the high dose group only (300 mg/kg bw/day) on day 29 and full recovery on day 42. All other organ data were in the range expected for this species, strain, sex and age and therefore not considered relevant.

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no substance related and toxicologically significant reported effects in the parameters examined.

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

At 300 mg/kg bw/day only : histopathological test item-related changes (without statistical significance) were observed in the colon and rectum of 3 males (2 out of 6 on day 29; 1 out of 6 on day 42). The findings were chronic inflammation in the tunica muscularis or chronic ulceration of the colon or a severe focal ulceration of the rectum. These effects are considered as treatment related.

Two out of six males of the 29-day subset at 300 mg/kg bw/day showed ulcerative lesions of the colon/rectum, one of them also revealed inflammatory changes in the forestomach. The morphological appearance of the ulceration indicates a long standing focal inflammatory process. In the recovery group (42-day subset at 300 mg/kg bw/day) one male rat revealed a chronic inflammatory process in the colon wall which can be interpreted as recovery of a preceding ulcerative lesion.

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

There were no treatment-related abnormalities detected.

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

150 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

Remarks on result:

not determinable due to adverse toxic effects at highest dose / concentration tested

Remarks:

All other observations werre considered either adaptive effects and/or have no relevance to human health.

Key result

Critical effects observed:

no

Conclusions:

Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for males and females is defined as 150 mg/kg body weight per day in males and females; due to histopathological findings in the gastrointestinal tract (forestomach, colon/rectum) caused by inflammation. All other observations were considered either adaptive effects or have no relevance to human health.

Executive summary:

The study was performed according to the requirements of OECD TG 407 and EU method B.7 guidelines under GLP conditions. The systemic toxic potential of the test item was assessed orally in a 28 day gavage study in Wistar Crl:WU rats. Recovery from any effects was evaluated during a subsequent 14 day recovery period. Three groups, each comprising three male and three female rats, received test item at doses of 75, 150 or 300 mg/kg bw/day. A control group of five males and five females was dosed with vehicle alone (Corn Oil) at a dose volume of 7.5 mL/kg bw/day. Two recovery groups, each of three males and three females, were treated with the high dose (300 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 and water consumption were monitored during the study. Haematology, 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, clinical signs, effect on sensory reactivity, grip strength or motor activity, bodyweight gain or food consumption. Water consumption was increased in both sexes (more pronounced in females), with statistical significance in the high dose groups. In the recovery period the values returned to normal. The increased water consumption correlated with increased kidney weights and an increased urine volume. As no adverse histopathological findings were detected in kidneys the effects are considered as treatment-related adaptive effects. Examination of haematological parameters did not reveal any substantial treatment-related changes. Examination of clinical chemistry and urinalysis parameters did not reveal relevant treatment-related changes. Statistically significant and dose-dependent increases were observed in both sexes for liver and kidney weights (effects more pronounced in males). The effect in livers recovered within 14 days and was not accompanied by a morphological correlate. Therefore, this finding was considered as an adaptative effect (possibly caused by the uptake of water). Histopathological test item-related changes (without statistical significance) were observed at 300 mg/kg bw/day in the colon and rectum of 3 males (2 out of 6 on day 29; 1 out of 6 on day 42). The findings were chronic inflammation in the tunica muscularis or chronic ulceration of the colon or a severe focal ulceration of the rectum. The morphological appearance of the ulceration indicates a long standing focal inflammatory process. In the recovery group (42-day subset at 300 mg/kg bw/day) one male rat revealed a chronic inflammatory process in the colon wall which can be interpreted as recovery of a preceding ulcerative lesion. There were no other adverse effects determined within the study. Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for males and females is defined as 150 mg/kg body weight per day in males and females; due to histopathological findings in the gastrointestinal tract (forestomach, colon/rectum) caused by inflammation. All other observations were considered either adaptive effects or have no relevance to human health.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

150 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

The key study is GLP compliant and of a high quality (Klimisch 1); The available information as a whole meets the tonnage driven information requirements of REACH.

System:

gastrointestinal tract

Organ:

colon

rectum

stomach

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

Repeated dose - Oral:

Key study : OECD TG 407, 2012 : The study was performed according to the requirements of OECD TG 407 and EU method B.7 guidelines under GLP conditions. The systemic toxic potential of the test item was assessed orally in a 28 day gavage study in Wistar Crl:WU rats. Recovery from any effects was evaluated during a subsequent 14 day recovery period. Three groups, each comprising three male and three female rats, received test item at doses of 75, 150 or 300 mg/kg bw/day. A control group of five males and five females was dosed with vehicle alone (Corn Oil) at a dose volume of 7.5 mL/kg bw/day. Two recovery groups, each of three males and three females, were treated with the high dose (300 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 and water consumption were monitored during the study. Haematology, 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, clinical signs, effect on sensory reactivity, grip strength or motor activity, bodyweight gain or food consumption. Water consumption was increased in both sexes (more pronounced in females), with statistical significance in the high dose groups. In the recovery period the values returned to normal. The increased water consumption correlated with increased kidney weights and an increased urine volume. As no adverse histopathological findings were detected in kidneys the effects are considered as treatment-related adaptive effects. Examination of haematological parameters did not reveal any substantial treatment-related changes. Examination of clinical chemistry and urinalysis parameters did not reveal relevant treatment-related changes. Statistically significant and dose-dependent increases were observed in both sexes for liver and kidney weights (effects more pronounced in males). The effect in livers recovered within 14 days and was not accompanied by a morphological correlate. Therefore, this finding was considered as an adaptative effect (possibly caused by the uptake of water). Histopathological test item-related changes (without statistical significance) were observed at 300 mg/kg bw/day in the colon and rectum of 3 males (2 out of 6 on day 29; 1 out of 6 on day 42). The findings were chronic inflammation in the tunica muscularis or chronic ulceration of the colon or a severe focal ulceration of the rectum. The morphological appearance of the ulceration indicates a long standing focal inflammatory process. In the recovery group (42-day subset at 300 mg/kg bw/day) one male rat revealed a chronic inflammatory process in the colon wall which can be interpreted as recovery of a preceding ulcerative lesion. There were no other adverse effects determined within the study. Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for males and females is defined as 150 mg/kg body weight per day in males and females; due to histopathological findings in the gastrointestinal tract (forestomach, colon/rectum) caused by inflammation. All other observations were considered either adaptive effects or have no relevance to human health.

Justification for classification or non-classification

The substance does not meet classification criteria under Regulation (EC) No 1272/2008 for specific organ toxicity repeated exposure (STOT-RE).

 

Specifically, the observed pathological findings in the gastrointestinal tract were not “consistent” (occurred in a minority of individuals at the top dose level 300 mg/kg bw/day only, without achieving statistical significance) and were not correlated with significant functional changes and/or marked organ disfunction meeting the classification criteria (e.g. correlates to bodyweight declines or clinical signs). The single individual in the equivalent recovery group indicated recovery. Additionally, such effects were not seen at the next lower dose level which was identified as the NOAEL at 150 mg/kg bw/day. At this dose level pathological changes were not seen and all observed changes were adaptive effects in nature.

 

References:

1. ECHA Guidance on Application on the CLP Criteria, (v5.0, July 2017), Section 3.9.2 : Table 3.16 - Equivalent guidance values for 28-day and 90-day studies