Registration Dossier

Administrative data

Description of key information

A Subchronic oral toxicity study was designed to investigate the systemic toxicity of the test item and is compatible with the following regulatory guidelines:

·        The OECD Guidelines for Testing of Chemicals No. 408 "Subchronic Oral Toxicity - Rodent: 90 Day Study” (Adopted 21 September 1998).

This study was also designed to be compatible with Commission Regulation (EC) No 440/2008 of 30 May 2008, laying down test methods pursuant to Regulation (EC) No 1907/2006 of the European Parliament and of the Council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).The No Observed Adverse Effect Level (NOAEL) for 2,2-dimethylpropane-1,3-diyl cyclohex-4-ene-1,2-dicarboxylate over a ninety-day treatment period was considered to be 100 mg/kg bw/day, due to adverse histopathological kidney changes for both sexes at the high dosage of 750 mg/kg bw/day.

Key value for chemical safety assessment

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:
18 January 2018 - 28 Jan 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Batch No.of test material: Colkb03
- Expiration date of the lot/batch: 01 February 2021
- Purity : > 85 %
- Physical state/Appearance: White solid

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Ambient temperature over silica gel in darkness; used/ formulated at ambient humidity in light
Species:
rat
Strain:
Wistar
Details on species / strain selection:
Wistar Han™:RccHan™:WIST strain rat
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Females (if applicable) nulliparous and non-pregnant: [yes/no]
- Age at study initiation: six to eight weeks old
- Weight at study initiation: the males weighed 191 to 221g, the females weighed 136 to 173g.
- Fasting period before study:
- Housing: The animals were initially housed in groups of three or four by sex in solid floor polypropylene cages with stainless steel mesh lids and softwood flake bedding.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: Males - eight days, Females - nine days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 50 ± 20
- Air changes (per hr): At least fifteen air changes
- Photoperiod (hrs dark / hrs light): twelve hours continuous light and twelve hours darkness

IN-LIFE DATES: From: 14 June 2018 To:13 September 2018
Route of administration:
oral: gavage
Details on route of administration:
Oral, by gavage using a rubber dosing catheter attached to a disposable plastic syringe.
Vehicle:
propylene glycol
Details on oral exposure:
The test item was prepared at the appropriate concentrations in Propylene Glycol. The stability and homogeneity of the test item formulations were determined. Results from the previous study show the formulations to be stable for at least twenty six days. Formulations were therefore prepared on in batches covering up to two weeks, at least one day ahead of the day of intended use, divided into daily aliquots and with each aliquot stored at approximately 4°C in the dark until the day of use.
Stability and homogeneity of test item formulations in the vehicle were determined as part of this study.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The samples of each test item formulation were taken and analyzed for concentration of 2,2-dimethylpropane-1,3-diyl cyclohex-4-ene-1,2-dicarboxylate. The results indicate that the prepared formulations were approximately 97-110% of the nominal concentration confirming the suitability of the formulation procedure.
Duration of treatment / exposure:
Ninety consecutive days
Frequency of treatment:
Once daily
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
10 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
750 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
Animals were allocated to treatment groups as follows: Dose levels were initally selected, in collaboration with the Sponsor based on available toxicity data including a twenty-day oral range finding study dose range finding study in the rat. In the range finding study, a dosage of 1000 mg/kg bw/day was considered to be a No Observed adverse Effect Level (NOAEL), therefore dosages of 0 (Control), 10, 100 and 1000 mg/kg bw/day, following the recommendations of the Global Harmonisation Scheme, were selected for use on this study. However, following results from a Preliminary Oral (Gavage) Pre-Natal Development Toxicity Study in the Rat, and a Fourteen Day Repeated Dose Oral (Gavage) Range-Finding Toxicity Study in the Rat, it was decided that a dosage of 1000 mg/kg bw/day would be unsuitable for further investigation of toxicity. In view of this, dose levels for use on this study were amended, in collaboration with the Sponsor, to 0 (Control), 10, 100 and 750 mg/kg bw/day utilizing a dosage volume of 5 mL/kg.

The test item was administered daily, for up to ninety consecutive days, by gavage using a rubber dosing catheter attached to a disposable plastic syringe. Control animals were treated in an identical manner with 5 mL/kg of Propylene Glycol.

The volume of test and control item administered to each animal was based on the most recent scheduled body weight and was adjusted at weekly intervals.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Not specified

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Immediately before dosing, up to thirty minutes post dosing and one hour after dosing

BODY WEIGHT: Yes
- Time schedule for examinations: Day 1 (prior to dosing) and at weekly intervals thereafter.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Pretreatment, Week 12
- Dose groups that were examined: All control and high dose animals

HAEMATOLOGY: Yes
- Anaesthetic used for blood collection: Not specified
- Time schedule for examinations: Day 90
- Animals fasted: No
- How many animals: All surviving animals
- Parameters checked in table [No.1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Day 90
- Animals fasted: No
- How many animals: All surviving animals
- Parameters checked in table [No.2] were examined.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Week 12
- Dose groups that were examined: All animals
- Battery of functions tested: sensory activity, grip strength, motor activity
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 3)

HISTOPATHOLOGY: Yes (see table 4)
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, Body Weight Change, Hematology, Blood Chemistry, Absolute Organ Weights, Body Weight-Relative Organ Weights.
Data were generally 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 non-homogeneity 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):
At 750 mg/kg bw/day, treatment was associated with incidences of increased post-dosing salivation for all surviving animals, throughout the study. However, at lower dosages, this finding was restricted to just one female at 10 mg/kg bw/day and two females at 100 mg/kg bw/day, on a single occasion (Day 5) during the study. Increased salivation is frequently observed when unpalatable or slightly irritant formulations are dosed via the oral gavage route and this finding was considered not to indicate any systemic effect of treatment.
At 750 mg/kg bw/day, treatment was also associated with occasional incidences of noisy respiration for both sexes throughout the study. Occasional incidences of noisy respiration were apparent for three males and four females at 10 mg/kg bw/day and two males and two females at 100 mg/kg bw/day. At the high dosage, the incidence of noisy respiration probably reflect the increased salivation apparent at this dosage and the irritant nature of the Test Item. The lower incidences at the lower dosages probably reflect occasional difficulties in dosing isolated animals during the study. Microscopic evaluation of the lungs did not indicate any histopathology change and this finding was considered not to indicate any systemic effect of treatment.
At 750 mg/kg bw/day, one male (number 62) showed hunched posture and decreased respiration rate on Day 1 and decreased respiration rate again on Days 8 and 9. However, subsequent clinical signs for the remainder of the study were restricted to instances of increased post-dosing salivation and noisy respiration. Additionally, at this dosage, one female (number 74) showed pallor of the extremities on Days 78 and 79.
Mortality:
mortality observed, treatment-related
Description (incidence):
Female number 70 was killed on Day 7 of the study after showing decreased respiration rate and laboured respiration, apparent hypothermia, piloerection, pallor of the skin, hunched posture and an open mass located on thoracic ventral region. The mass, which had been present from Day 1, had only become open on Day 7 and was considered to be the underlying cause for the decline in the physical condition of the animal. Necropsy confirmed the presence of the mass and also revealed dark coloured content in the stomach, jejunum, duodenum and ileum (considered to be ingested blood from the wound), pale brain, liver; lungs and ovaries, enlarged spleen and red patches on the glandular region of the stomach.
Male number 63 was found dead on Day 86 of the study. Clinical signs for this animal were unremarkable, being restricted to occasional incidences of increased post-dosing salivation during the study, although a notable body weight loss had been observed the previous day. Necropsy revealed enlarged kidneys, small spleen, red discoloration of lungs, dark patch of right liver lobe, dark discoloration of the thymus and pancreas, sloughing of the stomach and raised limiting ridge and thin appearance and red discoloration of the caecum. Microscopic examination of the tissues revealed ulceration of the non-glandular stomach, nephropathy and notable cellular depletion in the spleen, thymus and other lymphoid tissues. The cause of death was considered likely to be due to the ulceration of the stomach; this would represent a local irritant effect of the test item rather than indicating any underlying systemic toxicity.
Body weight and weight changes:
no effects observed
Food efficiency:
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
At 750 mg/kg bw/day, mean absolute and body weight relative kidney weights for both sexes were statistically significantly higher than control. For this organ, relative values are generally considered the better indicator of toxicological effect and for these treated animals, five male and nine female individual relative values exceeded the respective historical control range compared to two male and three females individual relative values for the control group.
For males at 100 mg/kg bw/day and both sexes at 750 mg/kg bw/day, mean absolute and body weight relative liver weights were statistically significantly higher than control. For this organ, relative values are generally considered the better indicator of toxicological effect and for males, nine individual relative values exceeded the historical control range at each dosage compared to only two individual control values for the control group. For females at 750 mg/kg bw/day, five individual relative values exceeded the historical control range whilst, one relative control value was below and another relative control value exceeded this historical range. Whilst the increase in relative organ weight indicated an effect of treatment, in the absence of any histopathological change, this was considered not to be adverse.
For females at all dosages, mean absolute and body weight relative spleen weights were statistically significantly higher than control, however, whilst the highest mean spleen weights were apparent at 750 mg/kg bw/day, mean values showed no consistent dosage relationship at lower dosages. At 750 mg/kg bw/day, four individual absolute values and five body weight relative values exceeded the respective historical control range while one absolute value and one body weight relative value were below the respective historical control range. At lower dosages, one absolute and one body weight relative value at 10 mg/kg bw/day and three absolute and two body weight relative values at 100 mg/kg bw/day exceeded the historical control range. In the absence of any convincing supporting evidence from blood analyses or microscopic evaluation of the tissues, this finding was considered not to represent an adverse effect.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
At 750 mg/kg bw/day, necropsy of surviving animals revealed a raised limited ridge within the stomach for seven males and six females. Additionally, two of these males showed sloughing of the stomach or non-glandular region of the stomach and a mass on the non-glandular region. Stomach findings at necropsy for animals receiving 100 mg/kg bw/day were restricted to raised limiting ridge for one male and one female. At 10 mg/kg bw/day, one male also showed a raised limiting ridge within the stomach and the stomach of one female had a thin appearance; this female also showed red discoloration of the stomach, jejunum, duodenum and ileum. Collectively these findings were considered to indicate a local irritant effect of the Test item rather than any systemic effect.
At 750 mg/kg bw/day, the two males with the more pronounced stomach findings also showed a hard kidney with mottled appearance, a dark area on the liver and red discoloration of the lungs. A further male showed dark liver and another male showed mottled appearance of the kidneys.
At 100 mg/kg bw/day, two males showed a mottled appearance of the kidneys, with one also showing a dark liver. A further male, at this dosage, showed a mottled appearance to the liver and another male showed red discoloration of the lungs.
At 10 mg/kg bw/day, one female had a fluid filled sac on the right ovary and another female had red discoloration of the lungs.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Female 77 was killed in extremis on Day 7. It had an open mass and showed pallor in several organs and an enlarged spleen. The animal had a malignant basal cell tumour which was ulcerated through the skin and this is considered to be the cause of death. Other changes were mainly related to hematopoiesis in some tissues. The death is unrelated to the administration of the test item.
Male 63 was found dead on Day 86. It had ulceration of the non-glandular stomach, nephropathy and notable cellular depletion in the spleen, thymus and other lymphoid tissues. The cause of death is considered to be due to the ulceration in the stomach and is likely to be related to the administration of the test item.
Histopathological examination of surviving animals at 750 mg/kg bw/day revealed the following:
Kidney
Nephropathy was recorded when two or more of the following changes were present: basophilic tubules, tubular dilation, interstitial cellular infiltration/fibrosis, granular casts.
At 750 mg/kg bwday, multifocal nephropathy was present in 5/9 males with severity minimal to moderate; a diffuse, marked change was present in one other. Multifocal basophilic tubules (mild) was present in one male. In females, multifocal nephropathy was present in 5/9 animals at a minimal or mild severity at this dosage. The nephropathy in the kidneys is considered to be a direct effect of the test item. This correlates with the increase in kidney weight for both sexes at this dosage.
No changes were present in any animals receiving 10 or 100 g/kg bw/day which could be related to the administration of the test item.
Adrenal Glands
Hypertrophy of the zona glomerulosa was present in one control male, one male receiving 10 mg/kg bw/day and in 7/9 males and 3/9 females receiving 750 mg/kg bw/day. The change in the zona glomerulosa of the adrenal gland at 750 mg/kg bw/day is likely to be an adaptive response. The zona glomerulosa is the site of synthesis of aldosterone, a mineralocorticoid which is mainly involved in the control of salt and water balance in the body (Domenici Lombardo, 1990; Greaves 2007). This change may be secondary to, or linked with, the previously discussed kidney changes at this dosage.
Stomach
Hyperplasia of the non-glandular epithelium at the limiting ridge was present in 4/9 males and 5/9 females receiving 750 mg/kg bw/day. The changes in the non-glandular stomach are likely to reflect an irritant effect of the test item on the non-glandular mucosa where there is prolonged exposure due to the unique structure of the rodent stomach. This is not generally considered to be significant to man as the corresponding anatomical area is not present and is due to irritation at point of contact and is not an indicator of systemic toxicity.
No changes were present in any animals receiving 10 or 100 g/kg bw/day which could be related to the administration of the test item.
Key result
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
750 mg/kg bw/day (actual dose received)
System:
urinary
Organ:
kidney
Treatment related:
yes
Conclusions:
The No Observed Adverse Effect Level (NOAEL) for 2,2-dimethylpropane-1,3-diyl cyclohex-4-ene-1,2-dicarboxylate over a ninety-day treatment period was considered to be 100 mg/kg bw/day, due to adverse histopathological kidney changes for both sexes at the high dosage of 750 mg/kg bw/day.
Executive summary:

Introduction

The study was designed to investigate the systemic toxicity of the test item and is compatible with the following regulatory guidelines:

·        The OECD Guidelines for Testing of Chemicals No. 408 "Subchronic Oral Toxicity - Rodent: 90 Day Study” (Adopted 21 September 1998).

This study was also designed to be compatible with Commission Regulation (EC) No 440/2008 of 30 May 2008, laying down test methods pursuant to Regulation (EC) No 1907/2006 of the European Parliament and of the Council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).

Methods

The test item was administered by gavage to three groups, initially each of ten male and ten female Wistar Han™:RccHan™:WIST strain rats, for up to ninety consecutive days, at dose levels of 10, 100 and 750 mg/kg bw/day. A control group of ten males and ten females was dosed with vehicle alone (Propylene Glycol) over the same treatment period. One female receiving 100 mg/kg bw/day escaped from its cage and subsequently mated and became pregnant. This female was killed early and, as far as possible, removed from the study.

Clinical signs, functional observations, body weight change, dietary intake and water consumption were monitored during the study. Hematology and blood chemistry were evaluated for all animals at the end of the study. Ophthalmoscopic examination was also performed on control group and high dose animals.

All animals were subjected to gross necropsy examination and histopathological evaluation of selected tissues from high dose and control animals was performed.

Results….

Mortality

There were two unscheduled deaths at 750 mg/kg bw/day.

One female was killed on Day 7 of the study after showing adverse clinical signs including an open mass on the thoracic ventral region. This mass was considered to be the underlying cause for the decline in the physical condition of the animal. Necropsy confirmed the presence of the mass and also revealed dark coloured content (considered to be ingested blood from the wound) in the stomach, jejunum, duodenum and ileum, pale brain, liver; lungs and ovaries, enlarged spleen and red patches on the glandular region of the stomach.

One male was found dead on Day 86 of the study. Clinical signs for this animal were unremarkable, but a notable body weight loss had been apparent the day prior to death.  Necropsy revealed enlarged kidneys, small spleen, red discoloration of lungs, dark patch of right liver lobe, dark discoloration of the thymus and pancreas, sloughing of the stomach and raised limiting ridge and thin appearance and red discoloration of the caecum. Microscopic examination of the tissues revealed ulceration of the non-glandular stomach, nephropathy and notable cellular depletion in the spleen, thymus and other lymphoid tissues. The cause of death was considered to be the ulceration of the stomach, representing a local irritant effect of the test item rather than indicating underlying systemic toxicity.   

Clinical Observations

At 750 mg/kg bw/day, one male showed hunched posture and decreased respiration rate on Day 1 and decreased respiration rate again on Days 8 and 9 and one female showed pallor of the extremities on Days 78 and 79.     

At 750 mg/kg bw/day, treatment was associated with incidences of increased post-dosing salivation for all surviving animals, throughout the study. This finding was consistent with the oral gavage dosing of unpalatable or slightly irritant formulations and was considered not to indicate any systemic effect of treatment. Treatment at this dosage was also associated with occasional incidences of noisy respiration for both sexes throughout the study but this finding was considered to reflect the irritant nature of the Test Item and the increased salivation apparent at this dosage, rather than any systemic effect of treatment. 

At 10 or 100 mg/kg bw/day, occasional incidences of increased post-dosing salivation or noisy respiration were observed but these findings were considered to be incidental and of no toxicological significance. 

Behavioral Assessment

There were no effects of treatment apparent during behavioral assessments for either sex at 10, 100 or 750 mg/kg bw/day.

Functional Performance Tests

There were no effects of treatment apparent during functional performance tests for either sex at 10, 100 or 750 mg/kg bw/day.

Sensory Reactivity Assessments

There were no effects of treatment apparent during sensory reactivity assessments for either sex at 10, 100 or 750 mg/kg bw/day.

Body Weight

There were no adverse effects of treatment on body weight gain for either sex at 10, 100 or 750 mg/kg bw/day.

Food Consumption

There was no effect of treatment on food consumption for either sex at 10, 100 or 750 mg/kg bw/day.

Food Conversion Efficiency

There were no adverse effects of treatment on food conversion efficiency for either sex at 10, 100 or 750 mg/kg bw/day.

Water Consumption

Visual assessment of water intake did not indicate any effect on water consumption for either sex at 10, 100 or 750 mg/kg bw/day.

Ophthalmoscopy

There were no effects of treatment apparent during ophthalmoscopic examination of the eyes for either sex at 10, 100 or 750 mg/kg bw/day.

Hematology

There were no adverse effects of treatment on hematology parameters for either sex at 10, 100 or 750 mg/kg bw/day.

Blood Chemistry

There were no adverse effects of treatment on blood chemistry parameters for either sex at 10, 100 or 750 mg/kg bw/day.

Necropsy

At 750 mg/kg bw/day, seven males and six females showed a raised limited ridge within the stomach with two of these males also showed sloughing of the stomach or non-glandular region of the stomach and a mass on the non-glandular region. These two males also showed a hard kidney with mottled appearance, a dark area on the liver and red discoloration of the lungs. One male also showed a dark liver and another male showed mottled appearance of the kidneys. The findings for the stomach indicated a local irritant effect of the Test item rather than any systemic effect of treatment.       

At 100 mg/kg bw/day, one male and one female showed a raised limited ridge within the stomach, two males showed a mottled appearance of the kidneys, with one also showing a dark liver. Another male also showed a mottled appearance to the liver and one male showed red discoloration of the lungs.

At 10 mg/kg bw/day, one male also showed a raised limiting ridge within the stomach. The stomach of one female had a thin appearance and this female also showed red discoloration of the stomach, jejunum, duodenum and ileum. One female had a fluid filled sac on the right ovary and another female had red discoloration of the lungs.

Organ Weights

At 750 mg/kg bw/day, mean absolute and body weight relative kidney weights for both sexes were statistically significantly higher than control.

For males at 100 mg/kg bw/day and both sexes at 750 mg/kg bw/day, mean absolute and body weight relative liver weights were statistically significantly higher than control, but, in the absence of any histopathological change, this finding was considered not to be adverse. 

For females at all dosages, mean absolute and body weight relative spleen weights were statistically significantly higher than control, with the highest mean spleen weights occurring at 750 mg/kg bw/day. In the absence of any convincing supporting evidence from blood analyses or microscopic evaluation of the tissues, this finding at 750 mg/kg bw/day was considered not to represent an adverse effect. At the lower dosages, there was no dosage relationship and this finding appeared incidental and unrelated to treatment.

Histopathology

At 750 mg/kg bw/day, multifocal nephropathy was present in the kidneys of 5/9 males (minimal to moderate severity) and 5/9 females (minimal or mild severity); a diffuse, marked change was present in one other male. Mild multifocal basophilic tubules were also present in the kidneys in one male. 

No changes were present in any animals receiving 10 or 100 g/kg bw/day which could be related to the administration of the test item.

At 750 mg/kg bw/day, hypertrophy of the zona glomerulosa of the adrenal glands was present in 7/9 males and 3/9 females.  This change is likely to be an adaptive response and be secondary to, or linked with, kidney changes at this dosage.

Hypertrophy of the zona glomerulosa of the adrenal glands was present in one control male and one male receiving 10 mg/kg bw/day.

At 750 mg/kg bw/day, hyperplasia of the non-glandular epithelium at the limiting ridge of the stomach was present in 4/9 males and 5/9 females and were considered to reflect an irritant effect of the test item.

No changes were present in the stomach of any animals receiving 10 or 100 g/kg bw/day which could be related to the administration of the test item.

Conclusion

The No Observed Adverse Effect Level (NOAEL) for 2,2-dimethylpropane-1,3-diyl cyclohex-4-ene-1,2-dicarboxylate over a ninety-day treatment period was considered to be 100 mg/kg bw/day, due to adverse histopathological kidney changes for both sexes at the high dosage of 750 mg/kg bw/day.

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

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

In a 28 -day oral range finding study performed accoring to OECD no. 407 guideline and under GLP condistions Wistar rats were treated for 28 consecutive days by oral gavage at dose levels up to 1000 mg/kg.

There were no changes at determination of body weight and food consumption measurements, or alterations during clinical laboratory investigations, macroscopic and microscopic examination that were considered to be an effect of treatment.

 The clinical signs, such as rales, hunched posture and piloerection, were considered not toxicologically relevant based on the low incidence and low severity of the symptoms and the occurrence in also the control males. 

 The slightly higher liver weights as noted in females at 500 and 1000 mg/kg were not accompanied by any related clinical biochemistry parameter or morphological changes. Therefore no toxicological significance was ascribed to this finding.

Based on the absence of functional or morphological disturbances supporting the change in liver weight noted a No Observed Adverse Effect Level (NOAEL) of 1000 mg/kg was established.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Only existing study.

Justification for classification or non-classification

Specific Target Organ Toxicity – Repeated Exposure: The notifiable substance did not exhibit significant toxic effects arising from a repeated exposure. As a result, the substance does not meet the criteria for classification according to Regulation (EC) No 1272/2008, Annex I section 3.9