Registration Dossier

Administrative data

Description of key information

Acute toxicity: oral: LD50 > 2000 mg/kg bw  (EU B.1 in rats, GLP, K, rel.1);
Acute toxicity: dermal: LD50 Combined = 2900 mg/kg bw (similar to OECD 402, rats, read-across, K, rel. 2);
Acute toxicity: inhalation: waiver.

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 April 30 to May 28, 1986
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study conducted according to EU Method B.1 (Acute Toxicity (Oral)).
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study
Qualifier:
according to
Guideline:
EU Method B.1 (Acute Toxicity (Oral))
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: A. Smith, Warlingham, Surrey.
- Weight at study initiation: Males: 268-322 g; females: 198-220 g
- Fasting period before study: Animals were fasted overnight prior dosing.
- Housing: Animals were housed in groups of 5/sex in grid bottomed polypropylene cages.
- Diet: Pelleted rodent diet (modified 41B supplied by Pilsbury’s Limited, Birmingham), ad libitum
- Water: Mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20-24 °C
- Humidity: 49-67 %
- Photoperiod: 12 h dark/12 h light
Route of administration:
oral: gavage
Vehicle:
other: 0.25 % aqueous solution of gum tragacanth
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION (if unusual): Test material was suspended in a 0.25 % aqueous solution of gum tragacanth.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were examined for signs of toxicity or other effects frequently after dosing and then daily for 14 days. Animals were weighed at weekly intervals.
- Necropsy of survivors performed: yes; on completion of the observation period, all surviving animals were sacrificed by carbon dioxide asphyxiation and subjected to gross necropsy. Animals which died during the course of the study were also subjected to gross necropsy.
Statistics:
None
Preliminary study:
Not applicable
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: No mortality was observed.
Mortality:
- No mortality was observed.
Clinical signs:
- No clinical signs were observed.
Body weight:
- All animals showed expected gains in bodyweight over the 14 day study period.
Gross pathology:
- No abnormalities were noted at necropsy.
Other findings:
None

None

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Oral LD50 Combined > 2000 mg/kg bw
Executive summary:

In an acute oral toxicity study (limit test) performed according to the E.U Test Method B.1 and in compliance with GLP, a group of Sprague Dawley rats (5/sex) was administered a single oral dose of test material at 2000 mg/kg bw by gavage. Animals were then observed for mortality, clinical signs and bodyweights for 14 days and at the end of the study the surviving animals were sacrificed for macroscopic examination.

No mortality or clinical signs were observed. Animals showed expected gains in bodyweight during the study. No abnormalities were noted at necropsy.

Oral LD50 Combined > 2000 mg/kg bw

Under the test conditions, the test material is not classified according to the annex VI of the Regulation EC No. 1272/2008 (CLP).

This study is considered as acceptable and satisfies the requirement for acute oral toxicity endpoint.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
2 000 mg/kg bw
Quality of whole database:
The key study is GLP compliant and of high quality (Klimisch score = 1)

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Data waiving:
other justification
Justification for data waiving:
other:
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From December 19, 1978 to April 26, 1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: This study was performed prior to the OECD test guideline No. 402 but the protocol is similar to that guidance. The supporting substance is considered adequate for read-across purpose (see Iuclid section 13 for justification).
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
yes
Remarks:
environmental conditions not reported
Principles of method if other than guideline:
Not applicable
GLP compliance:
no
Remarks:
pre-GLP
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Wilmington, Massachusetts.
- Weight at study initiation: 127-222 g (range-finding study); 140-234 g (main study)
- Housing: Animals were housed individually in suspended stainless steel cages.
- Diet: Purina Rodent Laboratory Chow®#5001, ad libitum
- Water: Water, ad libitum
- Acclimation period: 7 days
Type of coverage:
open
Vehicle:
other: alcohol
Details on dermal exposure:
TEST SITE
- Area of exposure: Dorsal area

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Any excess test material was wiped off with a cloth dampened with water.
- Time after start of exposure: 24 h

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 6 mL/mg bw
- Constant volume or concentration used: yes
- Based on a specific gravity of 0.8682, appropriate amounts of test material were admixed with 78-058 alcohol (vehicle) to achieve required dose levels.
Duration of exposure:
24 h
Doses:
1670, 2150, 2780, 3600 and 4640 mg/kg bw
No. of animals per sex per dose:
5/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
Animals were observed for mortality and signs of toxicity at 1, 3 and 24 h after dosing and daily thereafter for 14 days. Individual body weights were recorded prior to dosing and at the time of death or sacrifice on Day 14.
- Necropsy of survivors performed: Yes; all surviving animals were sacrificed using T-61® Euthanasia Solution (National Laboratories Corporation, Somerville, N. J.) on Day 14. Necropsies were performed on all animals at the time of death or sacrifice.
Statistics:
Litchfield and Wilcoxon (1949).
Preliminary study:
Not applicable
Sex:
male
Dose descriptor:
LD50
Effect level:
3 600 - 4 640 mg/kg bw
Based on:
test mat.
Remarks on result:
other: 5/5 males died at 4640 mg/kg bw.
Sex:
female
Dose descriptor:
LD50
Effect level:
2 150 - 2 780 mg/kg bw
Based on:
test mat.
Remarks on result:
other: 3/5, 5/5 and 5/5 females died at 2780, 3600 and 4640 mg/kg bw, respectively.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
2 900 mg/kg bw
Based on:
test mat.
95% CL:
2 164 - 3 886
Remarks on result:
other: 3/10, 5/10 and 10/10 animals died at 2780, 3600 and 4640 mg/kg bw, respectively.
Mortality:
3/10 (0 male + 3 females), 5/10 (0 male + 5 females) and 10/10 (5 males + 5 females) animals died at 2780, 3600 and 4640 mg/kg bw, respectively. No mortality was observed at 1670 or 2150 mg/kg bw.
Clinical signs:
Soft feces, mucous discharge or morbidity were observed ante mortem in animals receiving 2780, 3600 or 4640 mg/kg bw. Animals in all dosed groups were noted to have crusts around the eyes and/or nose between 1 h post-dosing and Day 3. However, this finding was attributed to the animals being collared during the first 24 h. Desquamation was observed between Days 7 and 14 in animals exposed to 1670, 2150, 2780 and 3600 mg/kg bw. The frequency at which this finding was observed increased with dose level. No dermal irritation was observed in animals treated with 4640 mg/kg bw (all died by Day 4) or vehicle control.
Body weight:
- The mean body weight gain was slightly lower in males at 1670, 2150, 2780 and 3600 mg/kg bw and moderately lower in females at 2150 mg/kg bw when compared to vehicle control. Mean body weight gains for the females receiving 1670 and 2780 mg/kg bw were comparable to the controls.
- The change in mean body weight in males at 4640 mg/kg bw and females at 3600 or 4640 mg/kg bw could not be calculated due to the mortalities in these groups.
Gross pathology:
No gross tissue alterations attributable to treatment were observed in any of the animals. Gross pathology findings were similar in all of the found-dead animals. In all but one of these animals the stomach was filled with food or bile and in all but three the cecum was full. Eleven animals were found to have dark lungs, and nine had dark areas on the liver. These signs were attributed to autolysis. Crust around the eyes and/or nose was also frequently observed. However, no dose-related differences were noted in gross pathology and these findings were not believed to be compound related. Sacrifice of the remaining animals on Day 14 revealed only mottled lungs which is considered to be an artifact of T-61® euthanasia.
Other findings:
None

Range-finding study:

No mortality or signs of compound induced toxicity were observed at 0, 50, 160 or 500 mg/kg bw. At 5000 mg/kg bw, mortality was observed in two females (antemortem: hunched appearance in one of these animals) and one male (antemortem: soft feces) on Day 2 and 3 post-dosing, respectively. The surviving male appeared morbid on Day 3 post-dosing. Necropsy of the found dead animals revealed only red or darkened lungs in the male and in one of the females. Crust was observed around the eyes and/or nose of most of the animals 24 h after dosing and is attributed to the animals being collared during this time.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Dermal LD50 Combined = 2900 mg/kg bw (95 % confidence limits of 2164-3886 mg/kg bw)
Executive summary:

In an acute dermal toxicity study, groups of CD® Sprague-Dawley outbred albino rats (5/sex/dose) were given single dermal application of test material diluted in alcohol at 1670, 2150, 2780, 3600 and 4640 mg/kg bw at a constant volume of 6 mL/mg bw. Test material was applied topically to the previously shaven dorsal area of the animal and remained in contact for 24 h. A vehicle control group was treated with undiluted alcohol in the same manner. Animals were then observed for mortality, clinical signs and bodyweights for 14 days and at the end of the study the surviving animals were sacrificed for macroscopic examination.

Range finding study was conducted at the dose levels of 50, 160, 500, 1600 and 5000 mg/kg bw (2/sex/dose) to determine the dose levels for main study. In the range-finding study, no mortality or signs of compound induced toxicity were observed at 0, 50, 160 or 500 mg/kg bw. Mortality was observed in one male and two females at 5000 mg/kg bw.

In the main study, no mortality was observed at 1670 or 2150 mg/kg bw. 3/10 (0 male + 3 females), 5/10 (0 male + 5 females) and 10/10 (5 males + 5 females) animals died at 2780, 3600 and 4640 mg/kg bw, respectively. Soft feces, mucous discharge or morbidity were observed ante mortem in animals receiving 2780, 3600 or 4640 mg/kg bw.

No mortality and no dermal irritation was observed in the vehicle control animals.

Desquamation was observed between Days 7 and 14 in animals exposed to 1670, 2150, 2780 and 3600 mg/kg bw. The mean body weight gain was slightly lower in males at 1670, 2150, 2780 and 3600 mg/kg bw and moderately lower in females at 2150 mg/kg bw when compared to vehicle control. Mean body weight gains for the females receiving 1670 and 2780 mg/kg bw were comparable to the controls. No gross tissue alterations attributable to treatment were observed in any of the animals.

Dermal LD50 Combined = 2900 mg/kg bw (95 % confidence limits of 2164-3886 mg/kg bw).

Under the test conditions, the test material is not classified according to the annex VI of the Regulation EC No. 1272/2008 (CLP).

This study is acceptable and satisfies the requirement for acute dermal toxicity endpoint.

The supporting substance is considered adequate for read-across purpose (see Iuclid section 13 for justification).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
2 900 mg/kg bw
Quality of whole database:
The key study is of good quality (Klimisch score = 2).

Additional information

Acute toxicity: oral

A key study was available (Toxicol 1986, Rel.1). In this acute oral toxicity study (limit test) performed according to the E.U. Test Method B.1 and in compliance with GLP, a group of Sprague Dawley rats (5/sex) was administered a single oral dose of test material at 2000 mg/kg bw by gavage. Animals were then observed for mortality, clinical signs and bodyweights for 14 days and at the end of the study the surviving animals were sacrificed for macroscopic examination. No mortality or clinical signs were observed. Animals showed expected gains in bodyweight during the study. No abnormalities were noted at necropsy.

Oral LD50 Combined > 2000 mg/kg bw.  

Acute toxicity: dermal

A study was available on the substance itself (Biosearch, 1979) but was not sufficient to conclude on the classification of the substance. Indeed, the maximum dose tested was 2000 mg/kg bw of a 50% dilution of test material. This is equivalent to a maximum dose of 1000 mg/kg bw. Moreover only three animals per sex were used. It should be noted that, even under the worst-case conditions employed (abraded skin and occlusive dressings), there were no mortality nor clinical signs up to the end of the study.

A read-across approach was used to conclude on acute dermal classification. The supporting substance, (2E)-1-(2,6,6-trimethyl-2-cyclohexen-1-yl)-2-buten-1-one, is considered adequate for read-across purposes (see Iuclid section 13 for additional justification).

A key study was available on the supporting substance (Borriston 1979, Rel.2). This acute dermal toxicity study was performed prior to OECD test guideline No. 402 but the protocol is similar to that guidance. Groups of CD® Sprague-Dawley outbred albino rats (5/sex/dose) were given single dermal application of test material diluted in alcohol at 1670, 2150, 2780, 3600 and 4640 mg/kg bw at a constant volume of 6 mL/mg bw. Test material was applied topically to the previously shaved dorsal area of the animal and remained in contact for 24 h. A vehicle control group was treated with undiluted alcohol in the same manner.Animals were then observed for mortality, clinical signs and bodyweights for 14 days and at the end of the study the surviving animals were sacrificed for macroscopic examination. A range finding study was conducted at the dose levels of 50, 160, 500, 1600 and 5000 mg/kg bw (2/sex/dose) to determine the dose levels for main study.

In the range-finding study, no mortality or signs of compound induced toxicity were observed at 0, 50, 160 or 500 mg/kg bw. Mortality was observed in one male and two females at 5000 mg/kg bw.

In the main study, no mortality was observed at 1670 or 2150 mg/kg bw.3/10 (0 male + 3 females), 5/10 (0 male + 5 females) and 10/10 (5 males + 5 females) animals died at 2780, 3600 and 4640 mg/kg bw, respectively. Soft feces, mucous discharge or morbidity were observed ante mortem in animals receiving 2780, 3600 or 4640 mg/kg bw.

Desquamation was observed between Days 7 and 14 in animals exposed to 1670, 2150, 2780 and 3600 mg/kg bw. The mean body weight gain was slightly lower in males at 1670, 2150, 2780 and 3600 mg/kg bw and moderately lower in females at 2150 mg/kg bw when compared to vehicle control. Mean body weight gains for the females receiving 1670 and 2780 mg/kg bw were comparable to the controls. No gross tissue alterations attributable to treatment were observed in any of the animals.

Dermal LD50 Combined = 2900 mg/kg bw (95 % confidence limits of 2164-3886 mg/kg bw).


Justification for selection of acute toxicity – oral endpoint
Only one study available

Justification for selection of acute toxicity – inhalation endpoint
In accordance with Column 2 of REACH Annex VIII, in addition to the oral route (8.5.1), for substances other than gases, the information mentioned under 8.5.2 (acute toxicity by inhalation) and 8.5.3 (acute toxicity by the dermal route) shall be provided for at least one other route. The choice for the second route will depend on the nature of the substance and the likely route of human exposure
In the present case, inhalation exposure will be less than dermal exposure because the registered substance has a low vapour pressure (3.0 Pa at 25°C) and a low melting point (1.8°C), so the potential for the generation of inhalable forms is low. Dermal exposure is the more likely route of exposure based on physico-chemical properties (Log Kow = 3.4, WS = 188 mg/L) and based on skin irritation properties that may enhance penetration.

Justification for selection of acute toxicity – dermal endpoint
The only study with the test substance itself was performed only up to 1000 mg/kg bw, therefore a read-across approach was used. The supporting substance is considered adequate for read-across purpose (see Iuclid section 13 for additional justification).

Justification for classification or non-classification

Harmonized classification:

The substance has no harmonized classification according to the Regulation (EC) No. 1272/2008 including ATP6.

Self classification:

Acute toxicity via Oral route:

Based on the available data, the substance is not classified according to the Regulation (EC) No. 1272/2008 as the LD50 is greater than 2000 mg/kg bw.

Acute toxicity via Dermal route:

Based on the available data, the substance is not classified according to the Regulation (EC) No. 1272/2008 as the LD50 is greater than 2000 mg/kg bw.

Acute toxicity (Inhalation):

No data was available.

Specific target organ toxicity: single exposure (Oral):

The classification criteria according to the Annex VI of the Regulation (EC) No. 1272/2008 as specific target organ toxicant (STOT) – single exposure, oral are not met since no significant health effects were observed immediately or delayed after exposure at the guidance value (oral) for a Category 1 classification (C ≤ 300 mg/kg bw) and at the guidance value (oral) for a Category 2 classification (2000 mg/kg bw≥C > 300 mg/kg bw). No classification is required.

Specific target organ toxicity: single exposure (Dermal):

The classification criteria according to the Annex VI of the Regulation (EC) No 1272/2008 as specific target organ toxicant (STOT) – single exposure, dermal are not met since no reversible or irreversible adverse health effects were observed immediately or delayed after exposure and no effects were observed at the guidance value (dermal) for a Category 1 classification (C≤ 1000 mg/kg bw) and at the guidance value (dermal) for a Category 2 classification (2000 mg/kg bw ≥ C > 1000 mg/kg bw). No classification is required.

Specific target organ toxicity: single exposure (Inhalation):

No data was available.