Registration Dossier

Administrative data

Description of key information

NOAEL (systemic toxicity) =  100 mg/kg/day); OECD 408; Cooper S. (2012)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
16 August 2011 - 23 May 2012
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:
1998
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
yes
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL

- Source and lot/batch No.of test material: JA01YX10
- Expiration date of the lot/batch: April 2012
- Purity test date: 95.8%



RADIOLABELLING INFORMATION (if applicable): N/A




STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL

- Storage condition of test material: Room temperature (ca 20°C), in the dark under Nitrogen
- Stability under test conditions: Stable

- Solubility and stability of the test substance in the solvent/vehicle: stable in the liquid matrix at concentrations of 2 and 200 mg/mL for 24 hours at ambient temperature and for up to 15 days when refrigerated (nominally 2 to 8C).

- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: N/A



TREATMENT OF TEST MATERIAL PRIOR TO TESTING

- Treatment of test material prior to testing: The test material was used as supplied. The Reaction product of 3,5,5-trimethyl-hexanoic acid and 2-metheylpropanoic acid and pentaerythritol was prepared for administration as a series of graded concentrations in the vehicle - corn oil.
- Preliminary purification step (if any): Dose range finder was conducted prior to the main experiment ignorer to determine suitable dose levels.

- Final dilution of a dissolved solid, stock liquid or gel: 5 mL/kg
- Final preparation of a solid: N/A



FORM AS APPLIED IN THE TEST (if different from that of starting material): N/A

Species:
rat
Strain:
Crj: CD(SD)
Remarks:
Crl:CD(SD) rats
Details on species / strain selection:
Crl:CD(SD) rats from Charles River (UK) Ltd.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Ltd.
- Females (if applicable) nulliparous and non-pregnant: [yes/no]: Yes
- Age at study initiation: 28 to 35 days
- Weight at study initiation: Weight range of 110 to 150 g for males and 100 to 140 g for females.
- Fasting period before study: Not Stated
- Housing: Animals were housed inside a barriered rodent facility (Building 8, Room 1826).T he animals were housed five of one sex per cage
- Diet (e.g. ad libitum): Standard rodent diet (Rat and Mouse No. 1 Maintenance Diet).
- Water (e.g. ad libitum): Polycarbonate bottles fitted with sipper tubes.
- Acclimation period: 5 days 


ENVIRONMENTAL CONDITIONS

- Temperature (°C): 19 to 23°C
- Humidity (%): 40 to 70%
- Air changes (per hr): Not stated

- Photoperiod (hrs dark / hrs light):12 hours continuous light and 12 hours continuous dark per 24 hours.


IN-LIFE DATES: From: To: 12 October 2011 - 20 Jan 2012
Route of administration:
oral: gavage
Details on route of administration:
The oral route of administration was chosen to simulate a condition of potential human exposure.
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
 
 DIET PREPARATION

- Rate of preparation of diet (frequency): N/A

- Mixing appropriate amounts with (Type of food): N/A

- Storage temperature of food: N/A
 


VEHICLE

- Justification for use and choice of vehicle (if other than water): The test item was insoluble in water hence corn oil was used

- Concentration in vehicle: 2 mg/mL and 200 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg bodyweight,
- Lot/batch no. (if required): Not stated

- Purity: Not stated
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Approximately 50 % of the final volume of vehicle was added to the test item and magnetically stirred until the test material had dispersed and homogenous. Specimen formulations were prepared at concentrations of 2 mg/mL and 200 mg/mL .

The stability was assessed following storage at ambient temperature (nominally 21 °C) for 0, 2 and 4 hours and 1 day, and refrigeration (nominally 2 to 8 ºC) for 1 day, 8 days and 15 days.

Single samples were taken for assay from the top, middle and bottom of the magnetically stirred formulation and homogeneity was determined by analysis of these samples. Stability was determined from the mean concentration of Tetraesters of pentaerythritol with 2- methylpropanoic acid and 3,5,5-trimethyl-hexanoic acid in the vehicle at each sampling point.

The results of the analysis confirmed that Tetraesters of pentaerythritol with 2- methylpropanoic acid and 3,5,5-trimethyl-hexanoic acid produced an homogenous suspension and was stable in the liquid matrix at concentrations of 2 and 200 mg/mL for 24 hours at ambient temperature and for up to 15 days when refrigerated (nominally 2 to 8 ºC).
Duration of treatment / exposure:
90 days
Frequency of treatment:
Once each day
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Group 1 Control
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
Group 2. Test item
Dose / conc.:
300 mg/kg bw/day (nominal)
Remarks:
Group 3. Test item
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
Group 4. Test item
No. of animals per sex per dose:
20
Control animals:
yes
Details on study design:
- Dose selection rationale: Dose selection was based on the result obtained from a 14 day dose range finder study.

- Rationale for animal assignment (if not random): Animal were assigned randomly

- Fasting period before blood sampling for clinical biochemistry: Not stated

- Rationale for selecting satellite groups: Not included

- Post-exposure recovery period in satellite groups: N/A

- Section schedule rationale (if not random): N/A

- Other: Not stated
Positive control:
No
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Twice daily


BODY WEIGHT: Yes
- Time schedule for examinations: The weight of each rat was recorded one week before treatment commenced (Week -1), on the day that treatment commenced (Week 0), weekly throughout the treatment period and before necropsy.


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):

- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes: The weight of food supplied to each cage, that remaining and an estimate of any spilled was recorded for each week throughout the treatment period. From these records the mean weekly consumption per animal (g/animal/week) was calculated for each cage.
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: N/A



FOOD EFFICIENCY:

- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Not specified



WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: Daily


OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: During Week 12
- Dose groups that were examined: Groups 1 (Control) and 4 (1000 mg/kg/day)


HAEMATOLOGY: Yes 

- Time schedule for collection of blood: During Week 13
- Anaesthetic used for blood collection: Yes (identity)
- Animals fasted: Yes (over night fasting)

- How many animals: All animals


CLINICAL CHEMISTRY: Yes 

- Time schedule for collection of blood: Not specified
- Animals fasted: Not specified

- How many animals: Not specified

URINALYSIS: Yes

- Time schedule for collection of urine: Not specified
- Metabolism cages used for collection of urine: No

- Animals fasted: Not specified


NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations:Not specified
- Dose groups that were examined:Not specified
- Battery of functions tested: sensory activity / grip strength / motor activity /: Yes
Sacrifice and pathology:
GROSS PATHOLOGY: Yes 


HISTOPATHOLOGY: Yes

Other examinations:
Not spacified
Statistics:
In-house statistical analysis packages; Quasar (version 1.3), StarTox (version 3.2) and SAS (version 9.1.3).
Clinical signs:
no effects observed
Description (incidence and severity):
Salivation and chin rubbing were seen shortly after test item administration at all dosages although chin rubbing was seen the Controlsgroups.
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
When compared with the controls, marginally low overall bodyweight gains (Week 0 to 13) were seen in males receiving 300 or 1000 mg/kg/day (-7% and -6% for males receiving 300 or 1000 mg/kg/day, respectively) and marginally high gains were seen in females receiving 300 or 1000 mg/kg/day (+5% and +3% for females receiving 300 or 1000 mg/kg/day, respectively). For both sexes the greatest degree of change was seen in animals receiving 300 mg/kg/day and, therefore, given that there was no clear dose response, it is considered that these bodyweight fluctuations were likely to be due to normal biological variation and not an effect of treatment.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Fmales receiving 300 mg/kg/day and males and females receiving 1000 mg/kg/day ate slightly more food than the Controls. Also, males receiving 1000 mg/kg/day only the animals in Cage 2 ate substantially more food than controls and in the females one cage of animals in each of Groups 3 and 4 (cages 11 and 9, respectively) ate more food than the Controls.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
effects observed, non-treatment-related
Description (incidence and severity):
Measurements in Week 9/10 indicated that one cage of males and one cage of females at 1000 mg/kg/day drank more water than the controls.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no ophthalmic lesions that were attributed to treatment.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Low haematocrit and haemoglobin concentration in females receiving100 and 300 mg/kg/day and in males and females receiving 1000 mg/kg/day. In addition, mean cell haemoglobin concentration was marginally high in females receiving 1000 mg/kg/day. Activated partial thromboplastin time was reduced in males receiving 1000 mg/kg/day (statistical significance attained, p<0.05) but there was no dose response in the degree of change. In addition, a statistically significant reduction in group mean platelet numbers was seen in females receiving 1000 mg/kg/day
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Statistically significantly high plasma enzyme activities (alkaline phosphatase, alanine amino-transferase and asparate amino-transferase activities) were seen in females receiving 300 or 1000 mg/kg/day. Urea and blood urea nitrogen concentrations were high in males and females receiving 300 or 1000 mg/kg/day, glucose concentrations were low in females receiving 300 mg/kg/day and in males and females receiving 1000 mg/kg/day and plasma cholesterol concentration was low in females receiving 1000 mg/kg/day. Creatinine concentrations were high in males receiving 100 mg/kg/day and in males and females receiving 300 or 1000 mg/kg/day and, although there was no clear dose-relationship, the differences from Control were of sufficient magnitude to be considered treatment-related.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
Forelimb grip strength values for males receiving 300 or 1000 mg/kg/day were low (p<0.05) compared with those of Controls during Week 12 of treatment but there was no dose- relationship and all group mean values were within the Historical Control Data range. These differences were therefore attributed to natural variation and grip strength values for females were unaffected.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Mean absolute and adjusted kidney and spleen weights were high for all groups of test item treated males and liver weights were high for males given 300 or 1000 mg/kg/day. In females there was an increase in kidney weight in animals given 1000 mg/kg/day and an increase in liver weight in animals given 300 or 1000 mg/kg/day. In addition, adrenal weight was increased in females given 1000 mg/kg/day and heart weight was slightly increased in all groups of test item treated females.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
The liver was noted as pale in three males and four females given 100 mg/kg/day, six males and seven females given 300 mg/kg/day, and six males and nine females given 1000 mg/kg/day. Enlargement and irregular surface of the kidneys were seen in male at all dose groups. The nature and incidence of all other findings were consistent with the commonly seen background of macroscopic changes.
Neuropathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Changes related to treatment with test item were seen in the liver, thyroid and kidneys of males and in the liver and thyroid of females. Periportal hepatocyte vacuolation was observed in males given 1000 mg/kg/day and in all female groups but with a dose related increase in the incidence and severity in treated groups. Minimal or slight centrilobular hepatocyte hypertrophy was seen in males given 300 or 1000 mg/kg/day and minimal centrilobular hypertrophy was seen in females given 1000 mg/kg/day.

Minimal to moderate hyaline droplets and granular cast accumulation were seen in the cortical tubules and chronic progressive nephropathy was seen in the majority of male animals in all treated groups. The number of animals affected and/or the severity of the changes was dose-related.

An increased incidence of follicular cell hypertrophy was seen in the thyroids of males and females given 1000 mg/kg/day and in males given 300 mg/kg/day.

An increased incidence of inflammatory cell infiltration was seen in the zona reticularis of 1000 mg/kg/day treated female.
Histopathological findings: neoplastic:
not specified
Other effects:
no effects observed
Details on results:
No other relevant observation other the ones stated above.
Key result
Dose descriptor:
NOAEL
Effect level:
ca. 100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
gross pathology
organ weights and organ / body weight ratios
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
300 mg/kg bw/day (nominal)
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
yes
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
300 mg/kg bw/day (nominal)
System:
urinary
Organ:
kidney
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
no

Tables containing raw data are attached in full study report.

Conclusions:
Sub-chronic exposure of rats at oral dose levels up to 1000 mg/kg/day was well tolerated. However, the terminal investigations indicated the liver, thyroids and kidneys as target organs. Collectively, the findings in these organs, with associated findings, were considered to be adverse in nature at dose levels of 300 and 1000 mg/kg bw/day. Therefore the no-observed-adverse-effect-level (NOAEL) was considered to be 100 mg/kg bw/day.
Executive summary:

In an OECD 408 - 2012, the test item was administered to Crl:CD (SD) rats via oral gavage at doses of 100, 300 and 1000 mg/kg bw/day up to 13 weeks. While no satellite group was not included in the study, control groups were administered vehicle (corn oil) only.

The test item was well-tolerated by the animals in the in-life phase of the study with no animals dying prematurely, no clinical signs related to treatment or of toxicological importance and no effects seen on sensory reactivity, grip strength or motor activity. Bodyweight and food consumption were also unaffected and there were no treatment-related ophthalmic lesions reported. Histopathological examination identified the liver, thyroids and kidneys as target organs.

In the liver, periportal hepatocyte vacuolation and centrilobular hepatocyte hypertrophy were observed at mid to high dose groups. Periportal hepatocyte vacuolation (fatty change)was characterised by cytoplasmic fat accumulation mainly in female rat and only in high dose in male in hepatocytes with associated loss of glycogen. The centrilobular hepatocyte hypertrophy was considered and adaptive response commonly associated with hepatic enzyme inducers . This is usually accompanied with decrease in thyroid hormones leading to secondary changes (follicular cell hypertrophy) in the thyroids and such a change as observed in this study. The centrilobular hepatocyte hypertrophy and the associated changes in the thyroid are not considered to be of any toxicological significance to man because in man the hormone-binding profiles differ to that in rodents and the half-life of thyroxine in rats is approximately 12-18 hours, compared with approximately five to nine days in man. Serum/plasma thyroid stimulating hormone (TSH) levels in the rat is also approximately 25-times greater than in man and the rat has enhanced thyroid hormone elimination. As a consequence, the rodent thyroid is many times more responsive to hormonal imbalance than the human thyroid. Other observations included pale liver, increased liver weights, increases in plasma enzyme activities of alkaline phosphatase, alanine aminotransferase and aspartate amino transferase activities and the decrease of glucose or cholesterol levels.

In the kidneys hyaline droplet nephropathy with early induction of chronic progressive nephropathy was seen in all treated male groups.The hyaline droplet accumulation in the cortical tubules, accompanied by the accumulation of granular casts in the corticomedullary junction is a consequence of  test compound forms complexes with a rat specific protein in the glomerular filtrate. Proteins of the glomerular filtrate are taken up by the tubular epithelium of proximal tubules where they normally hydrolyse. Complex formation hinders protein metabolism and the complex accumulates.  Hyaline droplet nephropathy regularly triggers the early development of chronic progressive nephropathy, an age related spontaneous change in both sexes of rats. Increased urea (and blood urea nitrogen), and creatinine levels correlated with the kidney changes seen in males given 300 or 1000 mg/kg/day and at the lowest dose group for creatinine only. However, no histopathological changes were seen in the females to account for the increased kidney weight seen at 1000 mg/kg/day, the pale kidneys seen at 300 or 1000 mg/kg/day and the increased urea, (and blood urea nitrogen) and creatinine levels reported at 300 or 1000 mg/kg/day. Urinalysis was not performed on this study but increased urination was inferred given that animals receiving 1000 mg/kg/day did drink more water than the controls during the study. The sllightly reduced haematocrit seen in the high dose animals indicates possible haemodilution. These changes are most probably associated with the slightly impaired kidney function. These changes are rat-specific and are not relevant to man.

Incidental changes such as inflammatory cell infiltration in the zona reticularis of the adrenals was observed in females given 1000 mg/kg/day. There was  slight increase in spleen weight was reported for all treated male groups and slightly increased heart weights were seen in the test item treated female groups but did not correlate with any histopathological findings.

There were also small dose-related changes apparent in several of the haematology parameters measured in Week 13 of treatment (changes seen in the red cell parameters in females at 300 mg/kg/day and in animals at 1000 mg/kg/day and in the white cell parameters (decreases in lymphocytes, eosinophils and basophils) in all groups of treated females). These changes were not considered to be of sufficient magnitude to have compromised the general physiology of the animals and, as such, are of only minor toxicological importance.

Treatment-related changes seen at 100 mg/kg/day were a small increase in mean cell haemoglobin concentration and a decrease in circulating lymphocytes, eosinophils and basophils in females, a slight increase in creatinine concentration in males, an increase in heart weight in females and increases in kidney and spleen weights in males. At necropsy pale livers were reported in several animals and one male had irregular surface of the liver. Histopathological examination revealed periportal hepatocyte vacuolation in the liver of one male and seven females. Hyaline droplets, granular casts and chronic progressive nephropathy were seen in the kidneys of males. Treatment at the low dose did not influence bodyweight, food consumption or sensory and motor activity and the fatty change in the liver was not seen in the low dose males and in the females was not accompanied with abnormal clinical chemistry findings indicating that functional impairment of the liver did not occur. In addition, the kidney changes that were seen are rat specific.

Therefore, it is considered that under the conditions of this study, the no-observed-adverse-effect-level (NOAEL) on this study is 100 mg/kg/day  based on the terminal investigations indicated the liver, thyroids and kidneys as target organs which were considered to be adverse in nature at dose levels of 300 and 1000 mg/kg/day.

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
March 8th 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Remarks:
Although its a non-guideline study, it was conducted under GLP
Qualifier:
no guideline followed
Principles of method if other than guideline:
This a none guideline study conducted under GLP conditions.
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: JA01YX10
- Expiration date of the lot/batch: April 2012
- Purity test date: 95.8 %



RADIOLABELLING INFORMATION (if applicable): N/A




STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL

- Storage condition of test material: Room temperature (ca 20°C), in the dark under Nitrogen
- Stability under test conditions: Not stated

- Solubility and stability of the test substance in the solvent/vehicle: Not stated

- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: Not stated



TREATMENT OF TEST MATERIAL PRIOR TO TESTING

- Treatment of test material prior to testing: Suspensions of the test substance were prepared in corn oil obtained from Sigma batch number MKBF6012V.
- Preliminary purification step (if any): N/A

- Final dilution of a dissolved solid, stock liquid or gel: 5 mL/kg/day
- Final preparation of a solid: N/A



FORM AS APPLIED IN THE TEST (if different from that of starting material): N/A

Species:
rat
Strain:
CD-1
Sex:
male/female
Details on test animals or test system and environmental conditions:
No detail was reported
Route of administration:
oral: gavage
Details on route of administration:
oral gavage as a suspension in corn oil
Vehicle:
corn oil
Details on oral exposure:
Reaction product of 3,5,5-trimethyl-hexanoic acid and 2-metheylpropanoic acid and pentaerythritol was administered by oral gavage as a suspension in corn oil, at a dose volume of 5 mL/kg/day, once each day for 14 days.
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
A dose volume of 5 mL/kg/day was used
Duration of treatment / exposure:
14 days
Frequency of treatment:
Once a day
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
Group 1 (Test item)
Dose / conc.:
300 mg/kg bw/day (nominal)
Remarks:
Group 2 (Test item)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
Group 3 (Test item)
No. of animals per sex per dose:
6
Control animals:
no
Details on study design:
Tetraesters of pentaerythritol with 2- methylpropanoic acid and 3,5,5-trimethyl-hexanoic acid was administered by oral gavage as a suspension in corn oil, at a dose volume of 5 mL/kg/day, once each day for 14 days.
Positive control:
No
Sacrifice and pathology:
Clinical signs, bodyweight, food consumption and water consumption (by visual assessment) were examined at regular intervals throughout the study. Terminal investigations included macroscopic pathology and organ weight measurement.
Statistics:
None
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 specified
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Liver weights were slightly high for females given 300 or 1000 mg/kg/day but no similar effect was present in the males.
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
There were no macroscopic findings at necropsy that were attributed to treatment with Test item.
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
There were no macroscopic findings at necropsy that were attributed to treatment with Test item.
Other effects:
not specified
Key result
Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effect observed
Key result
Critical effects observed:
no

Tables containing raw data are attached in full study report.

Conclusions:
Under the condition of the study, it was concluded that a high dose of 1000 mg/kg/day would be appropriate for the associated 13-week oral toxicity study (OWH0019). Consequently, proposed low and intermediate doses are 100 and 300 mg/kg/day, respectively.
Executive summary:

In a non guideline dose range finding study conducted under GLP, Tetraesters of pentaerythritol with 2-methylpropanoic acid and 3,5,5-trimethyl-hexanoic acid was administered by oral gavage as a suspension in corn oil, at a dose volume of 5 mL/kg/day, once each day for 14 days.

Clinical signs, bodyweight, food consumption and water consumption (by visual assessment) were examined at regular intervals throughout the study. Terminal investigations included macroscopic pathology and organ weight measurement.

There were no deaths during the treatment period and the general appearance and behaviour of the animals were not affected by treatment.

Food and water consumption, body weight gains were unaffected. Liver weights were slightly high for females given 300 or 1000 mg/kg/day but no similar effect was present in the males. Macroscopic examination did not reveal any findings related to treatment.

Under the condition of the study, it was concluded that a high dose of 1000 mg/kg/day would be appropriate for the associated 13-week oral toxicity study (OWH0019). Consequently, proposed low and intermediate doses are 100 and 300 mg/kg/day, respectively.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
100 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The endpoint is concluded based on a single key study with a Klimish rating of 1.
System:
hepatobiliary
Organ:
kidney
liver

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:
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
In accordance with Annex VIII, Section 8.6.1, Column 2 and supporting ECHA Guidance, concerning repeated dose toxicity testing, the oral route is the default one because it is assumed to maximise systemic availability (internal dose). There was no evidence available to support the use of an alternative route for this study.
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:
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
In accordance with Annex VIII, Section 8.6.1, Column 2 and supporting ECHA Guidance, concerning repeated dose toxicity testing, the oral route is the default one because it is assumed to maximise systemic availability (internal dose). There was no evidence available to support the use of an alternative route for this study.
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:
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
In accordance with Annex VIII, Section 8.6.1, Column 2 and supporting ECHA Guidance, concerning repeated dose toxicity testing, the oral route is the default one because it is assumed to maximise systemic availability (internal dose). There was no evidence available to support the use of an alternative route for this study.
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:
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
In accordance with Annex VIII, Section 8.6.1, Column 2 and supporting ECHA Guidance, concerning repeated dose toxicity testing, the oral route is the default one because it is assumed to maximise systemic availability (internal dose). There was no evidence available to support the use of an alternative route for this study.
Endpoint conclusion
Endpoint conclusion:
no study available

Mode of Action Analysis / Human Relevance Framework

Mode of toxicity is mainly through hepatotoxicity and nephrotoxicity: In this case however, the hepatotoxicity observed i.e. centrilobular hepatocyte hypertrophy is considered an adaptive response as it correlates to accumulation of fat droplets especially with the test item being a lubricant. The centrilobular hepatocyte hypertrophy and the associated changes in the thyroid are not considered to be of any toxicological significance to man because in man the hormone-binding profiles differ to that in rodents and the half-life of thyroxine in rats is approximately 12-18 hours, compared with approximately five to nine days in man.  However, the hepatotoxicity observed in terms of liver periportal hepatocyte vacuolation in males given 1000 mg/kg/day and all treated female groups with the severity of effects more apparent as the dose of the substance increases cannot be considered an adoptive respond.   This is because the effect is accompanied by pale liver, increased liver weights, increases in plasma enzyme activities of alkaline phosphatase, alanine aminotransferase and aspartate amino transferase activities and the decrease of glucose or cholesterol levels. These enzymes are similar marker used to assess hepatotoxicity profile in human and as such the potential for similar hepatotoxicity profile cannot be rule out in man.

Nephrotoxicity in the form of hyaline droplet nephropathy were observed in the kidneys of males. The hyaline droplet accumulation in the cortical tubules, accompanied by the accumulation of granular casts in the corticomedullary junction is a consequence of test compound forming complexes with a rat specific protein in the glomerular filtrate. Proteins of the glomerular filtrate are taken up by the tubular epithelium of proximal tubules where they normally hydrolyse. Complex formation hinders protein metabolism and the complex accumulates.  Hyaline droplet nephropathy regularly triggers the early development of chronic progressive nephropathy, an age related spontaneous change in both sexes of rats. However, this profile of nephrotoxicity is only common in rats and it’s relevant to human can be ruled out.

Additional information

In an OECD 408 - 2012, the test item was administered to Crl:CD (SD) rats via oral gavage at doses of 100, 300 and 1000 mg/kg bw/day up to 13 weeks. While no satellite group was not included in the study, control groups were administered vehicle (corn oil) only.

The test item was well-tolerated by the animals in the in-life phase of the study with no animals dying prematurely, no clinical signs related to treatment or of toxicological importance and no effects seen on sensory reactivity, grip strength or motor activity. Bodyweight and food consumption were also unaffected and there were no treatment-related ophthalmic lesions reported. Histopathological examination identified the liver, thyroids and kidneys as target organs.

In the liver, periportal hepatocyte vacuolation and centrilobular hepatocyte hypertrophy were observed at mid to high dose groups. Periportal hepatocyte vacuolation (fatty change) was characterised by cytoplasmic fat accumulation mainly in female rat and only in high dose in male in hepatocytes with associated loss of glycogen. The centrilobular hepatocyte hypertrophy was considered and adaptive response commonly associated with hepatic enzyme inducers. This is usually accompanied with decrease in thyroid hormones leading to secondary changes (follicular cell hypertrophy) in the thyroids and such a change as observed in this study. The centrilobular hepatocyte hypertrophy and the associated changes in the thyroid are not considered to be of any toxicological significance to man because in man the hormone-binding profiles differ to that in rodents and the half-life of thyroxine in rats is approximately 12-18 hours, compared with approximately five to nine days in man. Serum/plasma thyroid stimulating hormone (TSH) levels in the rat is also approximately 25-times greater than in man and the rat has enhanced thyroid hormone elimination. As a consequence, the rodent thyroid is many times more responsive to hormonal imbalance than the human thyroid. Other observations included pale liver, increased liver weights, increases in plasma enzyme activities of alkaline phosphatase, alanine aminotransferase and aspartate amino transferase activities and the decrease of glucose or cholesterol levels.

In the kidneys, hyaline droplet nephropathy with early induction of chronic progressive nephropathy was seen in all treated male groups. The hyaline droplet accumulation in the cortical tubules, accompanied by the accumulation of granular casts in the corticomedullary junction is a consequence of test compound forms complexes with a rat specific protein in the glomerular filtrate. Proteins of the glomerular filtrate are taken up by the tubular epithelium of proximal tubules where they normally hydrolyse. Complex formation hinders protein metabolism and the complex accumulates.  Hyaline droplet nephropathy regularly triggers the early development of chronic progressive nephropathy, an age related spontaneous change in both sexes of rats. Increased urea (and blood urea nitrogen), and creatinine levels correlated with the kidney changes seen in males given 300 or 1000 mg/kg/day and at the lowest dose group for creatinine only. However, no histopathological changes were seen in the females to account for the increased kidney weight seen at 1000 mg/kg/day, the pale kidneys seen at 300 or 1000 mg/kg/day and the increased urea, (and blood urea nitrogen) and creatinine levels reported at 300 or 1000 mg/kg/day. Urinalysis was not performed on this study but increased urination was inferred given that animals receiving 1000 mg/kg/day did drink more water than the controls during the study. The slightly reduced haematocrit seen in the high dose animals indicates possible haemodilution. These changes are most probably associated with the slightly impaired kidney function. These changes are rat-specific and are not relevant to man.

Incidental changes such as inflammatory cell infiltration in the zona reticularis of the adrenals was observed in females given 1000 mg/kg/day. There was slight increase in spleen weight was reported for all treated male groups and slightly increased heart weights were seen in the test item treated female groups but did not correlate with any histopathological findings.

There were also small dose-related changes apparent in several of the haematology parameters measured in Week 13 of treatment (changes seen in the red cell parameters in females at 300 mg/kg/day and in animals at 1000 mg/kg/day and in the white cell parameters (decreases in lymphocytes, eosinophils and basophils) in all groups of treated females). These changes were not considered to be of sufficient magnitude to have compromised the general physiology of the animals and, as such, are of only minor toxicological importance.

Treatment-related changes seen at 100 mg/kg/day were a small increase in mean cell haemoglobin concentration and a decrease in circulating lymphocytes, eosinophils and basophils in females, a slight increase in creatinine concentration in males, an increase in heart weight in females and increases in kidney and spleen weights in males. At necropsy pale livers were reported in several animals and one male had irregular surface of the liver. Histopathological examination revealed periportal hepatocyte vacuolation in the liver of one male and seven females. Hyaline droplets, granular casts and chronic progressive nephropathy were seen in the kidneys of males. Treatment at the low dose did not influence bodyweight, food consumption or sensory and motor activity and the fatty change in the liver was not seen in the low dose males and in the females was not accompanied with abnormal clinical chemistry findings indicating that functional impairment of the liver did not occur. In addition, the kidney changes that were seen are rat specific.

Therefore, it is considered that under the conditions of this study, the no-observed-adverse-effect-level (NOAEL) on this study is 100 mg/kg/day based on the terminal investigations indicated the liver, thyroids and kidneys as target organs which were considered to be adverse in nature at dose levels of 300 and 1000 mg/kg/day.

Justification for classification or non-classification

The substance meets the criteria for classification for specific target organ toxicity - repeated exposure (STOT-RE) in accordance with Regulation (EC) No 1272/2008 (CLP).