Registration Dossier

Administrative data

Description of key information

Oral LD50 (rat/female) > 2000 mg/kg bw (OECD 423, K, rel.1)

Dermal LD50 (rat/combined) > 2000 mg/kg bw (OECD 402, K, rel.1)

Inhalation LC50 (rat/combined) > 5.97 mg/L = 5970 mg/m3 (OECD 403, K, rel.1)

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 December 2013 to 03 January 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP study conducted according to OECD Guideline 423 without deviation.
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Qualifier:
according to guideline
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries, Test Data for Registration of Agricultural Chemicals, Acute oral toxicity (2-1-1), 12 Nousan No 8147, Agricultural Production Bureau, November 24, 2000.
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
UK GLP Compliance Programme (inspected on 20-22 November 2012 / signed on 24 April 2014)
Test type:
acute toxic class method
Limit test:
yes
Specific details on test material used for the study:
- Purity test date: 30 October 2013
- Storage Conditions: 4 °C in the dark under nitrogen
Species:
rat
Strain:
other: Crl:CD (SD)
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Ltd.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: ca. 8-10 weeks
- Weight at study initiation: 222-252 g
- Fasting period before study: Animals were fasted overnight prior to and approximately four hours after dosing.
- Housing: Animals were housed in groups of three rats of the same sex in solid bottomed polycarbonate cages with a stainless steel mesh lid.
- Diet: Rat and Mouse No. 1 Maintenance Diet, ad libitum
- Water: Potable water taken from the public supply, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 19-23 °C
- Humidity: 40-70 %
- Animal room was kept at positive pressure with respect to the outside by its own supply of filtered fresh air, which was passed to atmosphere and not re-circulated.
- Photoperiod: 12 h dark / 12 h light

IN-LIFE DATES: 12 December 2013 to 03 January 2014
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 200 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION:
- The test substance was formulated at a concentration of 200 mg/mL in the vehicle and administered at a volume of 10 mL/kg bw. The test substance formulation was prepared on the day of dosing.

CLASS METHOD
- Rationale for the selection of the starting dose: Dose level for the study was chosen in compliance with the study guidelines. Based on information (QSAR prediction, structural alert and toxicological information for the potential surrogates) provided by the Sponsor the initial dose was 2000 mg/kg bw.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
6 females/dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations:
Mortality: Cages of rats were checked at least twice daily for any mortalities.
Clinical signs: Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1. On subsequent days, animals were observed once in the morning and again at the end of the experimental day (with the exception of Day 15 - morning only). All animals were observed for 14 days after dosing.
- Frequency of weighing: The weight of each rat was recorded on Days 1 (prior to dosing), 8 and 15. Individual weekly body weight changes and group mean body weights were calculated.
- Necropsy of survivors performed: Yes; all animals were humanely killed on Day 15 by carbon dioxide asphyxiation and subjected to gross necropsy.
Statistics:
None
Preliminary study:
Not applicable
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: no mortality was observed
Mortality:
There were no deaths during the study.
Clinical signs:
Clinical signs of reaction to treatment comprised of under activity and irregular breathing seen in all females dosed at 2000 mg/kg bw, flat posture, partially closed eye lids, reduced body tone, piloerection, unsteady gait, splayed hind limbs, fast breathing, reduced body temperature and hunched posture. Clinical signs were first noted from approximately 15 minutes after dosing.
Recovery of animals, as judged by external appearance and behaviour, was complete by Days 2 or 3.
Body weight:
All animals were considered to have achieved satisfactory body weight gains throughout the study.
Gross pathology:
No abnormalities were noted in any animal at the macroscopic examination at study termination on Day 15.
Other findings:
None

None

Interpretation of results:
Category 5 based on GHS criteria
Conclusions:
Under the test conditions, the substance is::
- not classified according to the Annex VI of the Regulation EC No. 1272/2008 (CLP) as the oral LD50 is higher than 2000 mg/kg bw.
- classified as 'category 5' according to the GHS based on significant clinical signs observed at 2000 mg/kg bw.
Executive summary:

An acute oral toxicity study was performed according to OECD Guideline 423 and in compliance with GLP.

One group of three fasted female Crl:CD (SD) rats received a single oral gavage dose of the test substance, formulated in corn oil, at a dose level of 2000 mg/kg bw. As results at this dose level indicated the acute lethal oral dose of the test substance to be greater than 2000 mg/kg bw, in compliance with the study guidelines, a further group of three fasted females was similarly dosed at 2000 mg/kg bw to complete the study. Animals were then observed for mortality, clinical signs and bodyweights for 14 days and were all sacrificed for macroscopic examination.

There were no deaths during the study. Clinical signs of reaction to treatment comprised of under activity and irregular breathing, flat posture, partially closed eye lids, reduced body tone, piloerection, unsteady gait, splayed hind limbs, fast breathing, reduced body temperature and Hunched posture. Clinical signs were first noted from approximately 15 minutes after dosing. Recovery of animals, as judged by external appearance and behaviour, was complete by Days 2 or 3. All animals were considered to have achieved satisfactory body weight gains throughout the study. No abnormalities were noted in any animal at the macroscopic examination at study termination on Day 15.

Oral LD50 (female) > 2000 mg/kg bw.

Under the test conditions, the substance is::

- not classified according to the Annex VI of the Regulation EC No. 1272/2008 (CLP) as the oral LD50 is higher than 2000 mg/kg bw.

- classified as 'category 5' according to the GHS based on significant clinical signs observed at 2000 mg/kg bw.

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

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
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
Type of information:
experimental study
Adequacy of study:
key study
Study period:
17 July to 09 October 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP study conducted in compliance with OECD Guideline No. 403 without any deviation.
Qualifier:
according to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
UK GLP Compliance Programme (inspected on 12-14 March 2014 / signed on 12 May 2014)
Test type:
traditional method
Limit test:
yes
Specific details on test material used for the study:
- Purity test date: 30 October 2013
- Storage Conditions: ca. 4 °C in the dark under nitrogen
Species:
rat
Strain:
Wistar
Remarks:
RccHan™: WIST strain
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK Ltd, Oxon, UK.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: Approximately 8-12 weeks
- Weight at study initiation: 198-350 g
- Housing: Animals were housed in groups of up to five by sex in solid-floor polypropylene cages with stainless steel lids, furnished with softwood flakes.
- Diet: Food (Harlan 2014C Rodent Diet, Harlan Laboratories UK Ltd, Oxon, UK), ad libitum
- Water: Mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 19-25 °C
- Humidity: 30-70 %
- Air changes: 15 per hour
- Photoperiod: 12 h dark / 12 h light

N-LIFE DATES: 17 July to 09 October 2014
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Mass median aerodynamic diameter (MMAD):
2.63 µm
Geometric standard deviation (GSD):
2.57
Remark on MMAD/GSD:
Predicted amount less than 4 μm = 67.3 %
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Test item was aerosolized using a glass concentric jet nebulizer (Radleys, Saffron Walden, Essex, UK) located at the top of the exposure chamber.
- Exposure chamber volume: Cylindrical exposure chamber had a volume of approximately 30 L (dimensions: 28 cm diameter x 50 cm high).
- One day prior to the day of exposure each rat was acclimatized (for approximately 2 h) to a tapered polycarbonate restraining tube.
- Method of holding animals in test chamber: During the exposure period, each rat was individually held in a tapered, polycarbonate restraining tube fitted onto a single tier of the exposure chamber and sealed by means of a rubber ‘O’ ring. Only the nose of each animal was exposed to the test atmosphere.
- Source and rate of air: Chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.
- Method of particle size determination: The particle size of the generated atmosphere inside the exposure chamber was determined three times (33, 94 and 214 minutes of exposure respectively) during the exposure period using a Marple Personal Cascade Impactor (Westech IS Ltd, Beds., UK).
- Treatment of exhaust air: The extract from the exposure chamber passed through a ‘scrubber’ trap and was connected with a high efficiency filter to a metered exhaust system.
- Temperature and humidity in air chamber: Temperature and relative humidity inside the exposure chamber were measured by an electronic thermometer/humidity meter (Hanna Instruments Ltd, Beds., UK) located in a vacant port in the animals’ breathing zone of the chamber and recorded every thirty minutes throughout the four-hour exposure period. Temperature and humidity in air chamber were 21 °C and 37-39%, respectively.
- Exposure chamber oxygen concentration: Oxygen levels within the exposure chamber were measured by an electronic oxygen analyzer (Servomex (UK) Ltd, Crowborough, East Sussex) located in a port in the animals breathing zone during the four-hour exposure period. The test atmosphere was generated to contain at least 19% oxygen.
- Exposure Chamber Atmosphere Concentration: Prior to the start of the study, the non-volatile component of the test item was determined by adding a small, known amount of test item to glass fiber filters and recording their weights. The filters were then dried in a desiccator between 20 and 22 °C for approximately 24 hours and then weighed again. The difference in the two weights was taken as the volatile content of the test item and the non-volatile component was calculated as a percentage. The non-volatile component of the batch used during the study was found to be approximately 91.06 % (n=10). However, once a drying technique had been employed on filters sampled from the generated test atmosphere it was apparent that the concentration calculated was much lower than the results noted on the test filters prior to drying and as such it was concluded that the non-volatile content was being affected by the generation method. It was therefore considered that chemical analysis should be employed in order to determine test atmosphere concentrations. The test atmosphere was sampled nine times (5, 30, 60, 90, 120, 150, 180, 210 and 235 minutes of exposure respectively) during the exposure period. The sampling procedure involved 2 liters of test atmosphere being drawn through a series of two glass impingers containing Hexane (each made up to 80mL).
The nominal chamber concentration was calculated by dividing the mass of test item used by the total volume of air passed through the chamber.
The nominal concentration was 598 % of the actual mean achieved atmosphere concentration and shows that keeping the aerosol airborne was moderately difficult.
- Pretreatment: Prior to the start of the study, test item atmospheres were generated within the exposure chamber. During this characterization period test item input rates were varied to achieve the required atmospheric conditions.
- Following an appropriate equilibration period a single group of ten rats (five males and five females) was exposed to an atmosphere of the test item for a period of four hours. A target concentration of 5.0 mg/L was used for the exposure. As the mean achieved concentration was 119 % of target, the standard deviation of the test atmosphere concentrations was within acceptable limits and no deaths occurred, no further levels were required.

TEST ATMOSPHERE
- Brief description of analytical method used: Test item concentration in the test samples was determined by gas chromatography (GC) using an external standard technique.
- Samples taken from breathing zone: Yes
- Particle size distribution: See table 7.2.2/1
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): 2.63 μm / 2.57
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
Nominal concentration: 35.7 mg/L
Mean achieved atmosphere concentration: 5.97 mg/L
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: All animals were observed for clinical signs at hourly intervals during exposure, immediately on removal from the restraining tubes at the end of exposure, one hour after termination of exposure and subsequently once daily for up to fourteen days. Any deaths or evidence of overt toxicity were recorded at each observation.
- Frequency of weighing: Individual body weights were recorded on arrival, prior to treatment on the day of exposure and on Days 1, 3, 7 and 14 or at death.
- Necropsy of survivors performed: Yes, at the end of the 14 day observation period all animals were killed by intravenous overdose of sodium pentobarbitone. All animals, were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded. The respiratory tract was subjected to a detailed macroscopic examination for signs of irritancy or local toxicity.
Statistics:
None
Preliminary study:
Not applicable
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.97 mg/L air (nominal)
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
other: Only one death occurred in a group of ten rats
Mortality:
Only one death occurred in a group of ten rats at 5.97 mg/L air.
Clinical signs:
other: Signs of hunched posture and pilo-erection are commonly seen in animals for short periods on removal from the chamber following 4-Hour inhalation studies. Wet fur is commonly recorded both during and for a short period after exposure. These observations a
Body weight:
All animals exhibited body weight losses or showed no body weight gain on the first day post-exposure. Body weight gains were noted in all animals during the remainder of the recovery period, with the exception of one female animal (Animal No. 6) which showed no body weight gain from Days 1 to 3 post-exposure.
Gross pathology:
No macroscopic abnormalities were detected at necropsy amongst animals that survived until the end of the fourteen day recovery period, with the exception of one surviving male (Animal No. 4) which exhibited dark patches on the lungs.
The following macroscopic abnormalities were detected in the animal (Animal No. 8) that was humanely killed during the course of the study at necropsy:
Liver – pale;
Kidneys – pale.
Due to the observations noted it is considered that the death noted during the study may have been mainly attributable to systemic toxicity.
Other findings:
None

Exposure Chamber Concentration:

The actual concentration of the test item was measured off-line by high performance gas chromatography (GC). The test atmospheres were sampled after theoretical chamber equilibration and then at approximately half-hourly intervals during the exposure period.

The concentration of the test item was shown to be stable and the mean values obtained were:

Atmosphere Concentration

Mean Achieved (mg/L)

 

Standard Deviation

 

Nominal (mg/L)

 

5.97

0.25

35.7

 

The chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.

The theoretical chamber equilibration time (T99) was 3 minutes* (Silver, 1946).

Particle Size Distribution

The particle size analysis of the atmosphere drawn from the animals’ breathing zone, was as follows:

Mean Achieved Atmosphere Concentration (mg/L)

 

Mean Mass Median Aerodynamic Diameter (μm)

 

Inhalable Fraction

(% <4 μm)

Geometric Standard Deviation

 

5.97

2.63

67.3

2.57

 

Table 7.2.2/1: Exposure Chamber Atmosphere Concentrations

Duration of Exposure (minutes)

 

Volume of Air Sampled (L)

 

Chamber Flow Rate (L/min)

 

Atmosphere Concentration (mg/L)

 

5

2

60

5.51

3

2

60

5.86

60

2

60

5.82

90

2

60

6.11

120

2

60

6.36

150

2

60

6.03

180

2

60

6.21

210

2

60

5.93

235

2

60

5.86

Mean achieved atmosphere concentration (mg/L) = 5.97

Standard deviation = 0.25

Nominal concentration:

Test item used (g)

535

Air Flow (L/min)

60

Total Generation Time (mins)

250*

Nominal Concentration (mg/L)

35.7

  * = Test atmospheres were generated for a total of 10 minutes prior to animal insertion to ensure test item concentration was being achieved.

Table 7.2.2/2: Mortality Data

 

Mean Achieved Atmosphere Concentration (mg/L)

 

Sex

 

Deaths During Exposure

 

Deaths Post Exposure

(1 Hour)

Deaths During Day of Observation

 

Total Deaths

 

1

2

3

4

5

6

7

8-14

5.97

Male

 

0

0

0

0

0

0

0

0

0

0

1/10

Female

0

0

1*

0

0

0

0

0

0

0

* = Humanely killed as considered unlikely to survive

Interpretation of results:
Category 5 based on GHS criteria
Conclusions:
Under the test conditions, the substance is:
- not classified according to the Regulation EC No. 1272/2008 (CLP) as the LC50 is higher than 5.97 mg/L.
- classified as 'category 5' according to the GHS based on significant clinical signs observed at 5.97 mg/L.
Executive summary:

In an acute inhalation toxicity study performed according to OECD Guideline 403, groups (5/sex/dose) of RccHan: WIST strain rats were exposed (nose only) to aerosol atmosphere of ST 14 C 12 at concentration of 5.97 mg/L (mean achieved) for 4 h. Animals were observed for mortality, clinical signs and body weight for 14 days and at the end of the study the animals were sacrificed for macroscopic examination.

 

Observation changes noted during the study in the surviving animals included decreased respiratory rate, hunched posture, pilo-erection and wet fur. Frequent instances of increased respiratory rate and lethargy, occasional instances of labored respiration, noisy respiration, comatose and dehydration and isolated instances of ataxia, dry eyes (blinking less than usual with no blinking on contact with the eyes) and pallor of the extremities were also noted. Surviving animals recovered to appear normal on Day 8 post-exposure. Observations noted in the animal that was humanely killed during the study included decreased respiratory rate, labored respiration, noisy respiration, comatose, dehydration, pilo-erection and wet fur.

 

All animals exhibited body weight losses or showed no body weight gain on the first day post-exposure. Body weight gains were noted in all animals during the remainder of the recovery period, with the exception of one female animal which showed no body weight gain from Days 1 to 3 post-exposure.

No macroscopic changes were detected at necropsy amongst animals that survived until the end of the fourteen day recovery period, with the exception of one surviving male animal which exhibited dark patches on the lungs. The following macroscopic changes were detected in the animal that was humanely killed during the course of the study at necropsy: Liver and kidneys – pale. Due to the observations noted it is considered that the death noted during the study may have been mainly attributable to systemic toxicity.

 

Only one death occurred in a group of ten rats (five males and five females) exposed to a mean achieved atmosphere concentration of 5.97 mg/L for four hours. Therefore, the acute inhalation median lethal concentration (4 h LC50) of ST 14 C 12 was greater than 5.97 mg/L.

LC50(combined-rats) > 5.97 mg/L

Under the test conditions, the substance is:

- not classified according to the Regulation EC No. 1272/2008 (CLP) as the LC50 is higher than 5.97 mg/L.

- classified as 'category 5' according to the GHS based on significant clinical signs observed at 5.97 mg/L.

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

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
Value:
5 970 mg/m³
Quality of whole database:
The key study is GLP-compliant and of high quality (Klimisch score = 1)

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:
12 June to 03 July 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP study conducted according to OECD Guideline 402 without deviation.
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
UK GLP Compliance Programme (inspected on 12-14 March 2014 / signed on 12 May 2014)
Test type:
standard acute method
Limit test:
yes
Specific details on test material used for the study:
- Purity test date: 30 October 2013
- Storage Conditions: Stored at room temperature in the dark
Species:
rat
Strain:
Wistar
Remarks:
RccHan™: WIST strain
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK Limited, Bicester, Oxon, UK.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 249-289 g (males); 210-222 g (females)
- Housing: Animals were housed in suspended solid-floor polypropylene cages furnished with woodflakes. Animals were housed individually during the 24 h exposure period and in groups of five by sex for the remainder of the study.
- Diet: Food (2014C Teklad Global Rodent diet supplied by Harlan Laboratories UK Ltd., Oxon, UK), ad libitum
- Water: Mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 19-25 °C
- Humidity: 30-70 %
- Air changes: 15 changes per hour
- Photoperiod: 12 h dark / 12 h light

IN-LIFE DATES: 12 June to 03 July 2014
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Pretreatment: On the day before treatment the back and flanks of each animal were clipped free of hair.
- Area of exposure: Back and flank area
- % coverage: Approximately 10 % of the total body surface area.
- Application of test item: Test item was used as supplied. The test item was applied as evenly as possible to an area of shorn skin (approximately 10 % of the total body surface area) using a graduated syringe.
- Type of wrap if used: A piece of surgical gauze was placed over the treatment area and semi-occluded with a piece of self-adhesive bandage.

REMOVAL OF TEST SUBSTANCE
- Washing: After the 24 h contact period, the bandage was carefully removed and the treated skin and surrounding hair wiped with cotton wool moistened with arachis oil BP to remove any residual test substance.
- Time after start of exposure: 24 h

TEST MATERIAL
- For the purpose of the study the test item was used as supplied. The specific gravity was determined (1.100) and used to calculate the appropriate dose volume for the required dose level.
- Amount(s) applied (volume or weight with unit): 2000 mg/kg bw
- Constant volume or concentration used: Yes; 1.82 mL/kg bw
Duration of exposure:
24 h
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
not required
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: Animals were observed for mortality or clinical signs of toxicity at 0.5, 1, 2 and 4 h after dosing and subsequently once daily for 14 days. After removal of the dressings and subsequently once daily for fourteen days, the test sites were examined for evidence of primary irritation and scored according to Draize scale.
- Frequency of weighing: Individual bodyweights were recorded prior to application of the test substance on Day 0 and on Days 7 and 14.
- Necropsy of survivors performed: Yes; at the end of the study animals were killed by cervical dislocation and subjected to gross necropsy.
Statistics:
None
Preliminary study:
Not applicable
Key result
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 signs of systemic toxicity were noted during the observation period.
- Very slight erythema and crust formation were noted at the test sites of two females. Glossy skin was also noted at the test site of one of these females. There were no signs of dermal irritation noted at the test sites of all males and three females.
Body weight:
Animals showed expected gains in body weight, except for one female which showed no gain in body weight during the first week with expected gain in body weight during the second week. However, this was still considered to be within the historical range for this strain.
Gross pathology:
No abnormalities were noted at necropsy.
Other findings:
None

None

Interpretation of results:
GHS criteria not met
Conclusions:
Under the test conditions, the dermal LD50 of the test substance is higher than 2000 mg/kg bw in rats therefore it is not classified for acute dermal toxicity according to the criteria of the Annex VI of the Regulation (EC) N°1272/2008 (CLP) and to the GHS since there is no reliable evidence that indicates the LD50 to be in the range of Category 5 values.
Executive summary:

An acute dermal toxicity study (limit test) performed according to OECD Guideline No. 402 and in compliance with GLP. Initially, two animals (one male and one female) were given a single, 24 hour, semi-occluded dermal application of the undiluted test item to intact skin at a dose level of 2000 mg/kg bw. Based on the results of the initial test, a further group of eight animals (four males and four females) was similarly treated. Test sites were covered with a semi-occlusive dressing for 24 h. Animals were 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.

 

There were no deaths or signs of systemic toxicity. Very slight erythema, glossy skin and/or crust formation were noted at the test sites of two females. There were no signs of dermal irritation noted at the test sites of all males and three females. Animals showed expected gains in body weight, except for one female which showed no gain in body weight during the first week with expected gain in body weight during the second week. However, this was still considered to be within the historical range for this strain. No abnormalities were noted at necropsy.

 

Dermal LD50 Combined > 2000 mg/kg bw.

 

Under the test conditions, the test substance is not classified according to the Annex VI of Regulation (EC) No. 1272/2008 (CLP) and to the GHS.

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

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

Additional information

Acute toxicity: oral

A key study was identified (HLS, 2014, rel.1). In this limit acute oral toxicity study performed according to the OECD test guideline No. 423 and in compliance with GLP, 6 female rats were treated with a single oral gavage administration of 2000 mg/kg bw of test material in corn oil. The animals were observed for mortality, clinical signs and body weight for 14 days and then necropsied for macroscopic observations.

There were no deaths during the study. Clinical signs of reaction to treatment comprised of under activity and irregular breathing, flat posture, partially closed eye lids, reduced body tone, piloerection, unsteady gait, splayed hind limbs, fast breathing, reduced body temperature and Hunched posture. Clinical signs were first noted from approximately 15 minutes after dosing. Recovery of animals, as judged by external appearance and behaviour, was complete by Days 2 or 3. All animals were considered to have achieved satisfactory body weight gains throughout the study. No abnormalities were noted in any animal at the macroscopic examination at study termination on Day 15.

Oral LD50 (female) > 2000 mg/kg bw.

Evidence of significant adverse effects following acute exposure to the substance were observed in a 28-day toxicity study by oral route (gavage) to rats at doses of 30, 300 or 750/500 mg/kg bw/day (HLS, 2015, Rel.1). Administration of 750 mg/kg bw/day resulted in unforeseen clinical signs (unsteady gait, decreased activity, piloerection, partially closed eyelids, unresponsive and abnormal posture (swaying/flattened/prostrate)), one female was killed for humane reasons on Day 1 and further male on Day 5 due to persistent unresponsive behaviour. As a precautionary measure the high dose level was reduced to 500 mg/kg/day from Day 3 onwards. Administration of 750/500 mg/kg/day resulted in the death of 2 rats due to the severity of the clinical signs observed, findings associated with local irritation to the stomach.

Acute toxicity: inhalation

A key study was identified (Harlan, 2014, rel.1). In this limit acute inhalation toxicity study performed according to the OECD test guideline No. 403 and in compliance with GLP, groups (5/sex/dose) of RccHan: WIST strain rats were exposed (nose only) to aerosol atmosphere of ST 14 C 12 at concentration of 5.97 mg/L (mean achieved) for 4 h. Animals were observed for mortality, clinical signs and body weight for 14 days and at the end of the study the animals were sacrificed for macroscopic examination.

Only one death occurred in a group of ten rats (five males and five females) exposed to a mean achieved atmosphere concentration of 5.97 mg/L for four hours. Observations noted in the animal that was humanely killed during the study included decreased respiratory rate, labored respiration, noisy respiration, comatose, dehydration, pilo-erection and wet fur. The following macroscopic changes were detected in the animal that was humanely killed during the course of the study at necropsy: Liver and kidneys – pale. Due to the observations noted it is considered that the death noted during the study may have been mainly attributable to systemic toxicity. Observation changes noted during the study in the surviving animals included decreased respiratory rate, hunched posture, pilo-erection and wet fur. Frequent instances of increased respiratory rate and lethargy, occasional instances of labored respiration, noisy respiration, comatose and dehydration and isolated instances of ataxia, dry eyes (blinking less than usual with no blinking on contact with the eyes) and pallor of the extremities were also noted. Surviving animals recovered to appear normal on Day 8 post-exposure. All animals exhibited body weight losses or showed no body weight gain on the first day post-exposure. Body weight gains were noted in all animals during the remainder of the recovery period, with the exception of one female animal which showed no body weight gain from Days 1 to 3 post-exposure. No macroscopic changes were detected at necropsy amongst animals that survived until the end of the fourteen day recovery period, with the exception of one surviving male animal which exhibited dark patches on the lungs.

Inhalation LC50 (combined) > 5.97 mg/L

Acute toxicity: dermal

A key study was identified (Harlan, 2014, rel.1). In this limit acute dermal toxicity study performed according to the OECD test guideline No. 402 and in compliance with GLP, initially two animals (one male and one female) were given a single, 24 hour, semi-occluded dermal application of the undiluted test item to intact skin at a dose level of 2000 mg/kg body weight. Based on the results of the initial test, a further group of eight animals (four males and four females) was similarly treated. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

There were no deaths or signs of systemic toxicity. Very slight erythema, glossy skin and/or crust formation were noted at the test sites of two females. There were no signs of dermal irritation noted at the test sites of all males and three females. Animals showed expected gains in body weight, except for one female which showed no gain in body weight during the first week with expected gain in body weight during the second week. However, this was still considered to be within the historical range for this strain. No abnormalities were noted at necropsy.

Dermal LD50 Combined > 2000 mg/kg bw.

Justification for classification or non-classification

Harmonized classification:

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

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

- classified as 'category 5' according to the GHS based on significant clinical signs observed at 2000 mg/kg bw.

Acute toxicity (Inhalation):

Based on the available data, the substance is:

- not classified according to the Regulation EC No. 1272/2008 (CLP) as the LC50 is higher than 5.97 mg/L.

- classified as 'category 5' according to the GHS based on significant clinical signs observed at 5.97 mg/L.

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

- not classified according to the GHS since there is no reliable evidence that indicates the LD50 to be in the range of Category 5 values (GHS criteria not met)

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 reversible or irreversible adverse health effects were observed immediately or delayed after exposure and no effects were observed 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.

The criteria for Transient Organ effects (STOT-SE Category 3) according to Annex VI of the Regulation (EC) No. 1272/2008 are met since narcotic effects were observed in the acute oral toxicity study and in the repeated toxicity study.

Specific target organ toxicity: single exposure (Inhalation):

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 (inhalation, dust/mist) for a Category 1 classification (C≤ 1 mg/L/4h) and at the guidance value (inhalation, dust/mist) for a Category 2 classification (5 mg/L/4h ≥ C > 1 mg/L/4h). No classification is required.

The criteria for Transient Organ effects (STOT-SE Category 3 / H336) according to Annex VI of the Regulation (EC) No. 1272/2008 are met since narcotic effects were observed in the acute inhalation toxicity study.

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.

The criteria for Transient Organ effects (STOT-SE Category 3) according to Annex VI of the Regulation (EC) No. 1272/2008 are not met since narcotic effects were not observed in the acute dermal toxicity study.