Registration Dossier

Administrative data

Description of key information

An acute oral, an acute dermal and an acute inhalation toxicity studies were performed in rat. The LD50 were > 2000  mg/kg bw in the acute oral and dermal studies and the LC50 was > 5.6 mg/L air in the inhalation study.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 14 Nov 2006 to 08 Mar 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was performed according to an international test guideline and to GLP.
Qualifier:
according to
Guideline:
OECD Guideline 425 (Acute Oral Toxicity: Up-and-Down Procedure)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
up-and-down procedure
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: RCC Ltd, Laboratory Animal Services, CH-4414 Füllinsdorf, Switzerland
- Age at study initiation: 11 weeks
- Weight at study initiation: 179.7 to 196.3 g
- Fasting period before study: No
- Housing: Individually in Makrolon type-3 cages with standard softwood bedding ("Lignocel", Schill AG, CH-4132 Muttenz) during treatment and observation.
- Diet (e.g. ad libitum): Pelleted standard Provimi Kliba 3433 rat/mouse maintenance diet, batch no. 48/06 (Provimi Kliba AG, CH-4303 Kaiseraughst/Switzerland) ad libitum. Results of analyses for contaminants are archived at RCC Ltd.
- Water (e.g. ad libitum): Community tap water from Füllinsdorf ad libitum. Results of bacteriological, chemical and contaminant analyses are archived at RCC Ltd.
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3°C
- Humidity (%): 30-70%
- Air changes (per hr):10-15
- Photoperiod (hrs dark / hrs light):12/12 (music during the daytime light period)

IN-LIFE DATES: From: 22 Nov 2006 to 19 Dec 2006
Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: 1.90 mL/kg

Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 females
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: daily during the acclimatization period, during the first 30 minutes and at approximately 1, 2, 3 and 5 hours after adminitration on test day 1 (with the clinical signs) and twice daily during days 2-15.
- Frequency of weighing: On test day-1 (prior to removal of food), on test days 1 (prior to administration), 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs: daily during the acclimatization period, during the first 30 minutes and at approximately 1, 2, 3 and 5 hours after adminitration on test day 1. Once daily during days 2-15. All abnormalities were recorded.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
Female: 2000 mg/kg bw; Number of animals: 5; Number of deaths: 0
Clinical signs:
All animals showed slightly ruffled fur from the 30-minute, 1-hour or 2-hour reading until the 5-hour observation timepoint. This sign persisted in four animals until test day 2, 3 or 4. Hunched posture was also observed in 4 animals from the 30- minute or 2-hour reading until the 3-hour, 5-hour reading or test day 3. Slight sedation was also noted in 2 animals from the 3- to the 5-hour reading.
Body weight:

All the animals gained body weight during the study period. The body weight of the animals was within the range commonly observed for this strain and age.
Gross pathology:
Effects on organs:
No macroscopic findings were recorded at the scheduled necropsy.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material is not harmul by the oral route.
Executive summary:

In an acute oral toxicity study (RCC 2007 study n° B05657), 5 Wistar female rats were given a single oral dose of undiluted Dimethyl 2-methyl glutarate at the dose of 2000 mg/kg bw and observed for 14 days.

 

Oral LD50Females > 2000  mg/kg bw.

 

Dimethyl 2-methyl glutarate is not classified based on the LD50 in females.             

 

No mortality was observed. The clinical signs consisted of slightly ruffled fur, hunched posture and slight sedation in most animals. There was no effect on body weight gain.

 

This acute oral study is classified as acceptable.  It does satisfy the guideline requirement for an acute oral study (OECD425) in the rat.

 

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
Value:
2 000 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 27 May 2008 to 20 Feb 2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was performed according to international test guidelines and to GLP.
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1300 (Acute inhalation toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories Ltd, Laboratory Animals Services, Wölferstrasse 4, 4414 Füllinsdorf, Switzerland
- Age at study initiation: 9 weeks for males and 10 weeks for females
- Weight at study initiation: 241.9 to 264.1 g for males and 197.1 to 214.7g for females
- Fasting period before study: no
- Housing:In groups of five in Makrolon type-4 cages with wire mesh tops and sterilized standard softwood bedding (‘Lignocel’ Schill AG, 4132 Muttenz / Switzerland).
- Diet (e.g. ad libitum):Pelleted standard Kliba Nafag 3433 (batch no. 12/08) rat maintenance diet (Provimi Kliba SA, 4303 Kaiseraugst / Switzerland) was available ad libitum except during the period when the animals were restrained in exposure tubes. The feed batch was analyzed for contaminants.
- Water (e.g. ad libitum):Tap-water was available ad libitum in water bottles except during the period when the animals were restrained in exposure tubes. Results of bacteriological assay, chemical and contaminant analyses of representative samples are archived at Harlan Laboratories Ltd.
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light):12/12

IN-LIFE DATES: From 20 Aug 2008 to 03 Sep 2008
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus:Inhalation exposure was performed using a system similar to that originally described by Sachsse et al..
- Method of holding animals in test chamber: The animals were confined separately in restrained tubes which were positioned radially around the flow-past, nose-only exposure chamber as described by Cannon et al.. The design of this chamber is based upon the fluid dynamic modeling of the test aerosol flow.
- Source and rate of air: The exposure system ensured a uniform distribution and provided a constant flow of test material to each exposure tube. The flow of air at each tube was 1 L/min, which was sufficient to minimize re-breathing of the test atmosphere as it was more than twice the respiratory minute volume of a rat.
- System of generating particulates/aerosols: The test aerosol was generated using a plastic nebuliser connected to a syringe pump. The
polyethylene injector inside model was replaced by a stainless steel injector.
- Method of particle size determination: The cumulative particle size distribution of the test atmosphere was determined using a 7-stage Mercer cascade impactor. The test atmosphere was impacted at each stage onto an appropriate medium and the particle size distribution of the test item in the generated atmosphere was measured by gravimetrically analyzing the test item deposited on each stage of the cascade impactor.
- Temperature, humidity, pressure in air chamber: The relative humidity and temperature in the chamber were measured continuously during each
exposure using a calibrated device. The results are reported at 30 minute intervals from the start of exposure during the whole treatment.

TEST ATMOSPHERE
- Brief description of analytical method used:Test atmosphere samples were collected on Millipore® durapore filters, Type HVLP, (Polyvinylidenedifluoride membrane, pore size 0.45 μm) loaded in a 47 mm in-line stainless steel filter sampling device. The duration of sampling was sufficient to ensure reliable results. The weighing of these filters was delayed for 30 seconds to 3 minutes after sampling, in order to attain stable filter weights
- Samples taken from breathing zone: yes

TEST ATMOSPHERE: see details in the table below.
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
5.6 mg / L air
No. of animals per sex per dose:
- 5 animals/sex in the control group
- 10 animals/sex in the treatment group
Control animals:
yes
Details on study design:
- Duration of observation period following administration: 1 or 15 days
- Frequency of observations: Once before exposure on the day of exposure (test day 1), once per hour during exposure, once after exposure on test day 1 and twice daily during the observation period.
- Frequency of weighing:
- Main study animals (sacrificed after a 15-day observation period): On test days 1 (before exposure), 4, 8 and 15 (before necropsy)
- Interim sacrifice animals (sacrificed after a 1-day observation period): On test days 1 (before exposure) and 2 (before necropsy).
- Necropsy of survivors performed: yes
- Other examinations performed:
- clinical signs: Once per hour during exposure (only grossly abnormal signs as animals were restrained), once
after exposure on test day 1 and once daily during the observation period.
- histopathology:The nasal cavities of all animals (except of one animal of group 1) were processed, embedded in paraffin, cut at a nominal thickness of 2-4 micrometers, stained with hematoxylin & eosin (H&E) and examined by light microscope.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.6 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
No mortality occured during the study.
Clinical signs:
other: Slight to moderate salivation was recorded in all animals given the test item during exposure only. Ruffled fur was noted in all animals on test day 1 after the end of exposure, lasting up to test day 3. From test day 4 onwards, the animals were free from
Body weight:
No treatment-related effects on body weight were noted in animals given the test item.
Gross pathology:
No macroscopical findings were noted at scheduled necropsy.
Other findings:
- Histopathology:
Different degrees of necrosis of the olfactory epithelium were recorded one day after exposure in the nasal cavities of males and females receiving the test item, mainly at levels II-IV. These findings were sometimes noted along with luminal hemorrhages. After an observation period of 15 days, there was no necrosis at any level and signs of degeneration/regeneration of the olfactory epithelium and findings were limited to levels II and III.
No findings were noted in the nasal cavities of control animals.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material is not harmful by the inhalation route.
Executive summary:

In a acute inhalation toxicity study (Harlan Laboratories Ltd 2009 study no. B90527) one group of 10 male and 10 female Wistar rats were exposed by nose-only, flow past inhalation, for a single 4-hour period, to an aqueus dilution of Dimethyl 2-methylglutarate at a chemically determined mean aerosol concentration of 5.6 mg/L air. Half of the animals was necropsied after a 1-day observation period, the other half was necropsied after a 15-day observation period. A further group of 5 male and 5 female rats served as air control (exposed to air only) and were sacrificed after a 15-day observation period.

Acute inhalation LC50> 5.6 mg/L air.

 

Dimethyl 2-methylglutarate is not classified based on the LC50 in rats.

 

No mortality was observed. The clinical signs consisted of slight to moderate salivation recorded in all animals given the test item during exposure only. Ruffled fur was noted in all animals on test day 1 after the end of exposure, lasting up to test day 3. From test day 4 onwards, the animals were free from clinical signs.

There was no treatment-related effect on body weight.

 Histopathological examination showed different degrees of necrosis of the olfactory epithelium which were recorded one day after exposure in the nasal cavities of males and females receiving the test item, mainly at levels II-IV. These findings were sometimes noted along with luminal hemorrhages. After an observation period of 15 days, there was no necrosis at any level and signs of degeneration/regeneration of the olfactory epithelium and findings were limited to levels II and III.

 

This acute inhalation study is classified as acceptable. It does satisfy the guideline requirement for an acute inhalation toxicity study (OECD 403) in the rat.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating conc.
Value:
5.6 mg/m³

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 12 Sep 2007 to 29 Nov 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was performed according to an international test guideline and to GLP.
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: RCC Ltd, Laboratory Animal Services, Wölferstrasse 4, 4414 Füllinsdorf, Switzerland
- Age at study initiation: 8 weeks for males and 11 weeks for females
- Weight at study initiation: approximately 250 g for males and 200 for females
- Fasting period before study: No
- Housing: During acclimatization in groups of five per sex in Makrolon type-4 cages with standard softwood bedding. Individually in Makrolon type-3 cages with standard softwood bedding ("Lignocel", Schill AG, CH-4132 Muttenz) during treatment and observation.
- Diet (e.g. ad libitum):Pelleted standard Provimi Kliba 3433 rat/mouse maintenance diet (Provimi Kliba AG, CH-4303 Kaiseraugst/ Switzerland) ad libitum. Results of analyses for contaminants will be archived at RCC Ltd.
- Water (e.g. ad libitum):Community tap-water, from Füllinsdorf ad libitum. Results of bacteriological, chemical and contaminant analyses will be archived at RCC Ltd.
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12 (music during the daytime light period)

IN-LIFE DATES: From 20 sep 2007 to 04 Oct 2007
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: back of the animals
- % coverage: 10% of the total body surface
- Type of wrap if used: The semi-occlusive dressing was wrapped around the abdomen and fixed with an elastic adhesive bandage.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): with lukewarm tap water and dried with disposable paper towels.
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1.90 mL/kg bw
- Concentration (if solution): 1.0553 g/mL
- Constant volume or concentration used: yes
- For solids, paste formed: not applicable
Duration of exposure:
24 h
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 animals/sex/dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: Daily during the acclimatization period, at approximately 1, 2, 3 and 5 hours after administration on test day 1 (with the clinical signs) and twice daily during days 2-15.
- Frequency of weighing:On test days 1 (prior to administration), 8 and 15.
- Necropsy of survivors performed: yes/no
- Other examinations performed: clinical signs: Daily during acclimatization and at approximately 1, 2, 3 and 5 hours after administration on test day 1. Once daily during days 2-15. All abnormalities were recorded. Local signs: Once daily during days 2-15. All abnormalities were recorded.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No deaths occured during the study.
Clinical signs:
No clinical signs were observed during the observation period.
Body weight:
All animals gained weight during the study period. The body weight of the animals was within the range commonly recorded for this strain and age.
Gross pathology:
Effects on organs: No macroscopic findings were recorded at the scheduled necropsy.
Other findings:
Signs of toxicity (local):
At removal of the patch, no local signs were observed in any animal. However, on test day 5, one female exhibited a slight local erythema. Additionally, slight scaling was recorded at the same observation time point up to day 8 (in the same female). Otherwise, no local reactions were observed in any other animal up to the end of the observation period.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material is not harmul by the dermal route.
Executive summary:

In an acute dermal toxicity study ((RCC 2007 study n° B31195), 5 male and 5 female Wistar rats were dermally exposed to undiluted Dimethyl 2-methyl glutarate for 24 hours to 10% of body surface area at the dose of 2000 mg/kg bw.  Animals were then observed for 14 days.

 

Dermal Combined LD50> 2000  mg/kg bw.

Dimethyl 2-methyl glutarate is not classified based on the LD50.

 

No mortality and no clinical signs were noted during the study. No local signs were observed in any animal except for one female which exhibited a slight local erythema on day 5 and a slight scaling from days 5 to 8. There was no effect on body weight gain.

 

This acute dermal study is classified as acceptable.  It does satisfy the guideline requirement for an acute dermal study (OECD402) in the rat.

 

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
Value:
2 000 mg/kg bw

Additional information

Oral:

On study of reliability 1 according to Klimisch cotation critera, is available for oral route (Arcelin G., 2007) and was selected as a key study. In this study, 5 Wistar female rats were given a single oral dose of undiluted Dimethyl 2-methyl glutarate at the dose of 2000 mg/kg bw and observed for 14days. No mortality was observed. The clinical signs consisted of slightly ruffled fur, hunched posture and slight sedation in most animals. There was no effect on body weight gain. Based on this study, the LD50 for oral route in rats was higher than 2000 mg/kg bw. No classification for acute oral toxicity is therefore warranted on the basis on these in vivo data.

 

Inhalation:

On study of reliability 1 according to Klimisch cotation critera, is available for inhalation route (Schuler D., 2009) and was selected as a key study. In this study, one group of 10 male and 10 female Wistar rats were exposed by nose-only, flow past inhalation, for a single 4-hour period, to an aqueus dilution of Dimethyl 2-methylglutarate at a chemically determined mean aerosol concentration of 5.6 mg/L air. Half of the animals was necropsied after a 1-day observation period, the other half was necropsied after a 15-day observation period. A further group of 5 male and 5 female rats served as air control (exposed to air only) and were sacrificed after a 15-day observation period.

No mortality was observed. The clinical signs consisted of slight to moderate salivation recorded in all animals given the test item during exposure only. Ruffled fur was noted in all animals on test day 1 after the end of exposure, lasting up to test day 3. From test day 4 onwards, the animals were free from clinical signs. There was no treatment-related effect on body weight. Histopathological examination showed different degrees of necrosis of the olfactory epithelium which were recorded one day after exposure in the nasal cavities of males and females receiving the test item, mainly at levels II-IV. These findings were sometimes noted along with luminal hemorrhages. After an observation period of 15 days, there was no necrosis at any level and signs of degeneration/regeneration of the olfactory epithelium and findings were limited to levels II and III.

Based on this study, the LC50 for inhalation route in rats was higher than 5.6 mg/L air. No classification for acute oral toxicity is therefore warranted on the basis on these in vivo data.

Based on the classification criteria of Annex VI Directive 67/548/EECand EU Regulation 1272/2008 (CLP), no R37 or 'STOT – single exposure' classification, respectively, is warranted because no clinical symptoms or histological changes indicative of serious respiratory tract irritation/dysfunction were observed following a single exposure to the test item in rats. Furthermore, the toxicological relevance of rat findings to humans is limited based on well-known interspecies differences in the anatomy and physiology of the nasal and olfactory epithelia between rats and humans which support a lower susceptibility of humans to olfactory changes.

 

Dermal:

On study of reliability 1 according to Klimisch cotation critera, is available for dermal route (Simon C., 2007) and was selected as a key study. In this study, 5 male and 5 female Wistar rats were dermally exposed to undiluted Dimethyl 2-methyl glutarate for 24 hours to 10% of body surface area at the dose of 2000 mg/kg bw.  Animals were then observed for 14days. No mortality and no clinical signs were noted during the study. No local signs were observed in any animal except for one female which exhibited a slight local erythema on day 5 and a slight scaling from days 5 to 8. There was no effect on body weight gain.

Based on this study, the LD50 for dermal route in rats was higher than 2000 mg/kg bw. No classification for acute oral toxicity is therefore warranted on the basis on these in vivo data.


Justification for selection of acute toxicity – oral endpoint
only one study available (GLP and OECD 425 compliant)

Justification for selection of acute toxicity – inhalation endpoint
only one study available (GLP and OECD 403 compliant)

Justification for selection of acute toxicity – dermal endpoint
only one study available (GLP and OECD 402 compliant)

Justification for classification or non-classification

Due to the absence of mortality following exposure of rats to limit test doses or concentrations and a 2-week observation period whatever the route of administration, no classification for acute toxicity is warranted according to the criteria of Annex VI Directive 67/548/EEC or EU Regulation 1272/2008 (CLP).