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Administrative data

Description of key information

Oral toxicity:The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was found to be > 2000 mg/kg bw (OECD 423, EU Method B.1 Tris and OPPTS 870.1100).

 

Dermal toxicity:The acute dermal median lethal dose (LD50) of the test item was found to be > 2000 mg/kg bw in male and female Sprague Dawley rats(OECD Guideline 402, EU Method B.3 and OPPTS 870.1200).

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:
12 May 2016 to 02 June 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
EXPERIMENTAL ANIMALS
- Species and strain: Sprague Dawley rats
- Source: Charles River Laboratories, Research Models and Services GmbH, Sandhofer Weg 7, D-97633 Sulzfeld, Germany.
- Sex: Female, nulliparous and non-pregnant.
- Age of animals at dosing: Young healthy adult rats, 8 weeks old.
- Body weight at treatment: 189 to 200 g
- Acclimatisation period: 6 or 7 days.

HUSBANDRY
- Animal health: Only healthy animals were used for the test. The veterinarian certified health status.
- Number of animal room: 522/12.
- Housing: Three animals per cage.
- Cage type: Type II polypropylene/polycarbonate.
- Bedding: Lignocel 3/4-S Hygienic Animal Bedding produced by J. Rettenmaier & Söhne GmbH & Co.KG (D-73494 Rosenberg, Germany) was available to animals during the study.
A copy of the Certificate of Analysis is retained in the archive at CiToxLAB Hungary Ltd.
- Lighting period: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
- Temperature: 20.1 to 23.7 °C.
- Relative humidity: 36 to 70 %.
- Temperature and relative humidity were measured continuously and recorded twice daily during the study.
- Ventilation: 15 to 20 air exchanges/hour.
- Enrichment: Animals were housed by group to allow social interaction and with deep wood sawdust bedding to allow digging and other normal rodent activities.

ANIMAL IDENTIFICATION
- Animals were individually identified using numbers written on the tail with an indelible
marker pen. The numbers were given on the basis of CiToxLAB Hungary Ltd.'s Master File, for each animal allocated to the treatment groups.
- Cages were identified by cards, with information about study code, sex, dose group, cage number and individual animal numbers.
Route of administration:
oral: gavage
Vehicle:
peanut oil
Remarks:
Sigma Aldrich (batch MKBQ0728V; expiry date 30 September 2016; room temperature storage)
Details on oral exposure:
FORMULATION
- The test item was freshly formulated at a concentration of 200 mg/mL in the vehicle, in the Pharmacy of CiToxLAB Hungary Ltd. on the day of administration.
- The formulation container was stirred continuously until treatment was completed.

Doses:
2000 mg/kg bw
No. of animals per sex per dose:
Two groups of three animals
Control animals:
no
Details on study design:
FOOD AND WATER SUPPLY
- Animals received ssniff SM R/M "Autoclavable complete diet for rats and mice – breeding and maintenance" produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany (Batch No.: 278 5652, expiry date: November 2016), ad libitum, and tap water from the municipal supply, as for human consumption from a 500 ml bottles, ad libitum. The food was considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
- Water quality control analysis is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József, A.u.36., Hungary). The quality control results are retained in the archives at CiToxLAB Hungary Ltd.

TEST ITEM ADMINISTRATION
- The initial dose level was selected by the study director to be that which is most likely to produce mortality in some of the dosed animals. In the lack of any preliminary toxicological information, 2000 mg/kg bw was selected to be the starting dose.
- Initially, three female animals (Group 1) were treated with 2000 mg/kg bw of test item. No mortality was observed, therefore further 3 animals were treated at the dose level of 2000 mg/kg bw. As no mortality was observed in this second dose group, further testing was not required according to the test guidelines (OECD 423, Commission Regulation (EC) NO 440/2008 of 30 May 2008, B.1.Tris).
- A single oral gavage administration was followed by a fourteen-day observation period. On the night before treatment, the animals were fasted. The food but not water was withheld during an overnight period. Animals were weighed just before treatment. The test item was administered by oral gavage in the morning. Food was returned three hours after treatment.

CLINICAL OBSERVATIONS
- Clinical observations were performed on all surviving animals at 0.5, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter.
- Individual observations were performed on the skin, fur, eyes, mucous membranes, respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

BODY WEIGHT MEASUREMENT
- The body weights of the animals were recorded on the day before treatment (Day -1), on the day of the treatment (Day 0) and weekly thereafter.

NECROPSY
- Macroscopic examination was performed on all animals. The animals were sacrificed by exsanguination under pentobarbital anaesthesia (Euthanimal 40%; Lot No.: 1409236-06, Expiry Date: September 2017, produced by: AlfasanNederland BV, Woerden, Netherlands).
- After examination of the external appearance, the cranial, thoracic and the abdominal cavities were opened and the organs and the tissues were observed. Macroscopic abnormalities were recorded.
Statistics:
EVALUATION OF TEST RESULTS
- The method used was not intended to allow the calculation of a precise LD50 value.
- Clinical signs, body weight, body weight gain and gross macroscopic data were tabulated.
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
- The test item did not cause mortality at a dose level of 2000 mg/kg bw.
Clinical signs:
- Treatment with the test item at a dose level of 2000 mg/kg bw caused decreased activity (4/6), hunched back (6/6), piloerection (6/6), and increased salivation (1/6) (see Appendix 1, attached).
Body weight:
- Body weight gains during the study showed no indication of a test item-related effect (see Appendix 2, attached).
Gross pathology:
- There was no evidence of the macroscopic effects at a dose level of 2000 mg/kg bw (see Appendices 3 and 4, attached).
Interpretation of results:
GHS criteria not met
Conclusions:
The acute oral median lethal dose (LD50) of the test item in the female Sprague Dawley rat was found to be > 2000 mg/kg bw.
Executive summary:

GUIDELINE

The single-dose oral toxicity of the test item was investigated according to the acute toxic class method (OECD 423, Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris and OPPTS 870.1100) in Sprague Dawley rats.

 

METHODS

Two groups of three female Sprague Dawley rats were treated with the test item at a dose level of 2000 mg/kg bw (Group 1 and Group 2). A single oral treatment was carried out by gavage for each animal after withdrawal of food overnight. Food was made available again 3 hours after the treatment. The test item was administered formulated in peanut oil at a concentration of 200 mg/mL at a dose volume of 10 mL/kg bw. Initially, three female animals (Group 1) were treated with 2000 mg/kg bw of test item. No mortality was observed, therefore a further three animals were treated at the dose level of 2000 mg/kg bw. No mortality was observed in the confirmatory group; therefore no further testing was required according to the test guidelines. Clinical observations were performed at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter. Body weight was measured on Days -1, 0 and 7 and before necropsy. All animals were subjected to a necropsy and a macroscopic examination.

 

RESULTS

The test itemdid not cause mortality at a dose level of 2000 mg/kg bw. Clinical observations at the dose level of 2000 mg/kg bw were reported as decreased activity (4/6), hunched back (6/6), piloerection (6/6), and increased salivation (1/6). Body weight gains of treated animals during the study did not indicate a test item-related effect. No macroscopic observations were reported at 2000 mg/kg bw.

 

CONCLUSION

The acute oral median lethal dose (LD50) of the test item in the female Sprague Dawley rat was found to be > 2000 mg/kg bw.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Endpoint conclusion
Endpoint conclusion:
no study available

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:
16 June 2016 to 05 July 2016
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
yes
Remarks:
clinical observations conducted at 2 and 5 hours after application with no impact on results or integrity of the study
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Version / remarks:
L 142
Deviations:
yes
Remarks:
clinical observations conducted at 2 and 5 hours after application with no impact on results or integrity of the study
Qualifier:
according to
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Version / remarks:
EPA 712-C-98-192, August 1998
Deviations:
yes
Remarks:
clinical observations conducted at 2 and 5 hours after application with no impact on results or integrity of the study
GLP compliance:
yes (incl. certificate)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
EXPERIMENTAL ANIMALS
- Species and strain: Sprague Dawley Rat (Crl:CD(SD))
- Source: Charles River Laboratories, Research Models and Services GmbH, Sandhofer Weg 7, D-97633 Sulzfeld, Germany.
- Sex: Male and female, female rats were nulliparous and non-pregnant.
- Age of animals at study start: Young adult rats
- Body weight range at dosing: 215 to 247 g
- Acclimatisation period: 5 days

HUSBANDRY
- Animal health: Only healthy animals were used for the study. The veterinarian certified the health status.
- Room-Box: 245/9
- Housing: Individual caging
- Cage type: Type II. polypropylene/polycarbonate
- Bedding: Lignocel ¾S Hygienic Animal Bedding (produced by J. Rettenmaier & Söhne GmbH + Co.KG, Germany) was available to animals during the study. A copy of the Certificate of Analysis is retained in the archives at CiToxLAB Hungary Ltd.
- Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
- Temperature: 20.7 to 24.6 °C
- Relative humidity: 39 to 70 %
- Temperature and relative humidity were recorded twice daily during the study.
- Ventilation: 15 to 20 air exchanges/hour
- Enrichment: Rodents were housed with deep wood sawdust bedding to allow digging and other normal rodent activities.

ANIMAL IDENTIFICATION
- The individual identification was performed using numbers written on the tail with a marker pen. The numbers were given on the basis of CiToxLAB Hungary Ltd.' s Master File for each animal allocated to the treatment groups.
- Cages were identified by cards containing information about study code, sex, dose group, cage number and individual animal number.
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST ITEM ADMINISTRATION
- The test item was not expected to be lethal at 2000 mg/kg bw and a limit test was therefore performed.
- A single dermal application was made and was followed by a fourteen-day observation period (male animals cages 1 to 5; female animals cages 6 to 10).

PROCEDURE
- The back of each animal was shaved (approximately 10 % area of the total body surface) approximately 24 hours prior to treatment.
- Test item was applied as a single dose to the shaved skin and remained in contact with the skin for the 24-hour exposure period.
- Sterile gauze pads were placed on the skin of rats to cover the test item. These gauze pads were kept in contact with the skin using a patch with adhesive hypoallergenic plaster. The entire trunk of the animal was then wrapped with semi occlusive plastic wrap for 24 hours.
- At the end of the exposure period, the area of skin treated with the test item was washed with water of body temperature.
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 males and 5 females
Control animals:
no
Details on study design:
FOOD AND WATER SUPPLY
- Animals received ssniff SM R/M "Autoclavable complete diet for rats and mice – breeding and maintenance" produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany (Batch No.: 278 5652, expiry date: 30 November 2016), ad libitum, and tap water from the municipal supply, as for human consumption from a 500 ml bottles, ad libitum. The food was considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study and the water was considered fit for human consumption.
The supplier provided an analytical certificate for the batch used and copy certificates were archived with the raw data.
- Water quality control analysis is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A. u. 36., Hungary). The quality control results are retained in the archives at CiToxLAB Hungary Ltd.

CLINICAL OBSERVATIONS
- Clinical observations were performed on the day of treatment at 2 and 5 hours after application of the test item and once each day for 14 days thereafter.
- Observations included the skin and fur, eyes and mucous membranes, the respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

SKIN IRRITATION
- Adverse skin reactions at the site of application were recorded daily following the removal of the dressing.

MEASUREMENT OF BODY WEIGHT
- Body weights were recorded on Day 0 (before test item administration) and on Days 7 and 14 just before necropsy.

NECROPSY
- Macroscopic examination was performed on all animals. All animals were anaesthetised with pentobarbital sodium and exsanguinated.
- After examination of the external appearance, the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed. All macroscopic changes were recorded.
Statistics:
EVALUATION OF DATA
- The method used was not intended to allow the calculation of a precise Ld50 value.
- Clinical signs, body weight, body weight gain and gross macroscopic data were tabulated.
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
- The test item did not cause mortality at the dose level of 2000 mg/kg bw.
Clinical signs:
- There were no systemic clinical signs noted in any animal throughout the study (see Table 1, attached).
Body weight:
- There were no treatment related effects on body weight or body weight gain during the observation period (see Table 2, attached).
Gross pathology:
- No macroscopic observations were noted at a dose level of 2000 mg/kg bw (see Table 3 and Appendix 1, attached).
Other findings:
LOCAL DERMAL EFFECTS
- Treatment with test item at the dose level of 2000 mg/kg bw caused oedema (oedema score: 1 or 2) (10/10), erythema (erythema score: 1 or 2) (8/10) and crust (1/10) in rats (see Table 1, attached).
- All rats were symptom free from Day 2 (3/10) or Day 3 (3/10) or Day 5 (4/10).
Interpretation of results:
GHS criteria not met
Conclusions:
The acute dermal median lethal dose (LD50) of the test item was found to be > 2000 mg/kg body weight in male and female Sprague Dawley rats.
Executive summary:

GUIDELINE

An acute dermal toxicity study was performed with test item in Sprague Dawley rats, in compliance with OECD Guideline No 402, EU Method B.3 and OPPTS 870.1200.

 

METHODS

A limit test was carried out at 2000 mg/kg body weight (bw) in both sexes (5 rats/sex). The test item was applied as a single dermal 24-hour exposure followed by a 14-day observation period. Clinical observations were performed on all animals at 2 and 5 hours after dosing and daily for 14 days thereafter. Body weight was measured prior to dosing on Day 0 and on Days 7 and 14. Gross macroscopic examination was performed on all animals at the end of the 2-week observation period (Day 14).

 

RESULTS

The test item did not cause mortality at the dose level of 2000 mg/kg bw and no systemic clinical signs noted in any animal throughout the study. Treatment at a dose level of 2000 mg/kg bw caused oedema (oedema score: 1 or 2) (10/10), erythema (erythema score: 1 or 2) (8/10) and crust (1/10) in rats. All rats were symptom free from Day 2 (3/10) or Day 3 (3/10) or Day 5 (4/10). There were no treatment related effects on body weight or body weight gain during the observation period and no macroscopic observations were noted at a dose level of 2000 mg/kg bw.

 

CONCLUSION

The acute dermal median lethal dose (LD50) of the test item was found to be > 2000 mg/kg body weight in male and female Sprague Dawley rats.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Additional information

Oral route

Single-dose oral toxicity of the test item was investigated according to the acute toxic class method (OECD 423, Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris and OPPTS 870.1100) in Sprague Dawley rats.

 

Two groups of three female Sprague Dawley rats were treated with the test item at a dose level of 2000 mg/kg bw (Group 1 and Group 2). A single oral treatment was carried out by gavage for each animal after withdrawal of food overnight. Food was made available again 3 hours after the treatment. The test item was administered formulated in peanut oil at a concentration of 200 mg/mL at a dose volume of 10 mL/kg bw. Initially, three female animals (Group 1) were treated with 2000 mg/kg bw of test item. No mortality was observed, therefore a further three animals were treated at the dose level of 2000 mg/kg bw. No mortality was observed in the confirmatory group; therefore no further testing was required according to the test guidelines. Clinical observations were performed at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter. Body weight was measured on Days -1, 0 and 7 and before necropsy. All animals were subjected to a necropsy and a macroscopic examination.

 

The test itemdid not cause mortality at a dose level of 2000 mg/kg bw. Clinical observations at the dose level of 2000 mg/kg bw were reported as decreased activity (4/6), hunched back (6/6), piloerection (6/6), and increased salivation (1/6). Body weight gains of treated animals during the study did not indicate a test item-related effect. No macroscopic observations were reported at 2000 mg/kg bw. The acute oral median lethal dose (LD50) of the test item in the female Sprague Dawley rat was found to be > 2000 mg/kg bw.

Inhalation route

The substance has been shown to have a high onset boiling point (self-reaction from approximately 118 °C at 102 kPa followed by partial boiling (approximately 75%) from approximately 227 °C) and the vapour pressure has been determined to be 0.1 Pa at 25 °C. It is therefore expected that inhalation exposure will be low under general use conditions at ambient temperature. Lack of systemic toxicity when the substance is administered via the oral and dermal routes suggests that the determined vapour pressure of 15.0 Pa at 80°C also gives no cause for concern. The inhalation route is therefore not the most applicable method of investigating acute toxicity.

Dermal route

An acute dermal toxicity study was performed with the test item in Sprague Dawley rats, in compliance with OECD Guideline No 402, EU Method B.3 and OPPTS 870.1200.

 

A limit test was carried out at 2000 mg/kg body weight (bw) in both sexes (5 rats/sex). The test item was applied as a single dermal 24-hour exposure followed by a 14-day observation period. Clinical observations were performed on all animals at 2 and 5 hours after dosing and daily for 14 days thereafter. Body weight was measured prior to dosing on Day 0 and on Days 7 and 14. Gross macroscopic examination was performed on all animals at the end of the 2-week observation period (Day 14).

 

The test item did not cause mortality at the dose level of 2000 mg/kg bw and no systemic clinical signs noted in any animal throughout the study. Treatment at a dose level of 2000 mg/kg bw caused oedema (oedema score: 1 or 2) (10/10), erythema (erythema score: 1 or 2) (8/10) and crust (1/10) in rats. All rats were symptom free from Day 2 (3/10) or Day 3 (3/10) or Day 5 (4/10). There were no treatment related effects on body weight or body weight gain during the observation period and no macroscopic observations were noted at a dose level of 2000 mg/kg bw. The acute dermal median lethal dose (LD50) of the test item was found to be > 2000 mg/kg body weight in male and female Sprague Dawley rats.

Justification for classification or non-classification

The acute median lethal dose (LD50) of the test item in the Sprague Dawley rat was reported as > 2000 mg/kg bw following oral and dermal administration. Classification for acute oral or dermal toxicity is therefore unnecessary under the terms of Regulation (EC) No 1272/2008.