Registration Dossier

Administrative data

Description of key information

ORAL
LD50 > 2000 mg/kg, male/female rat, OECD 401, CIT (1994).
INHALATION
LC50 > 5.09 mg/L air, male/female rat, OECD 436, Harlan (2012).
DERMAL
No study available, a data waiver has been submitted.

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:
2 March 1994 - 16 March 1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted to GLP in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.
Qualifier:
according to
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Deviations:
no
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
- Strain: Sprague-Dawley ICO:OFA-SD (IOPS Caw)
- Age at study initiation: approximately 6 weeks
- Weight at study initiation (means): 164 ± 4g for the males, 161 ± 5 g for the females
- Fasting period before study: 18 hours before treatment; food was replaced approximately 4 hours after treatment
- Housing: animals were housed in groups of 4 to 7 animals of the same sex in polycarbonate cages (48 x 27 x 20 cm) covered with a stainless steel lid during the acclimatisation period and groups of 5 animals of the same sex during the study. Each cage contained graded, dust-free sawdust.
- Food consumption (e.g. ad libitum): ad libitum
- Water consumption (e.g. ad libitum): ad libitum filtered water contained in bottles
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 50 ± 20 % relative humidity
- Air changes (per hr): 13 cycles/hour of non-recycled and filtered air
- Photoperiod (hrs dark / hrs light): 12hr/12hr

In-life dates: From 02 to 16 March 1994
Route of administration:
oral: gavage
Vehicle:
other: aqueous solution of methylcellulose at 0.5 %
Details on oral exposure:
DOSAGE PREPARATION:
On the day of the treatment, the test material was ground using a mortar and pestle, then was suspended in the vehicle.

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg


Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 animals per sex per dose
Control animals:
other: The body weight of the animals treated with the test material was compared to laboratory historical control data for animals dosed by the oral route.
Details on study design:
DETAILS ON STUDY DESIGN
- Duration of observation period following administration: 14 days
- Frequency of observations: the animals were observed frequently after administration of the test material, for detection of possible treatment-related clinical signs and for mortality or signs of morbidity. Observations of the animals were made at least once a day for clinical signs and at least twice a day for mortality.
- Frequency of weighing: Animals were weighed just before administration of the test material and then on days 5, 8 and 15.
- Necropsy of survivors performed: yes (day 15). Animals were sacrificed by excess CO2 inhalation.
- Other examinations performed: macroscopic examination at necropsy (digestive tract, heart, kidneys, liver, lungs, pancreas, spleen and any other organ with obvious abnormalities).
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Mortality:
No deaths occured during the observation period.
Clinical signs:
No clinical signs were observed during the study.
Body weight:
The body weight gain of the animals was normal.
Gross pathology:
The macroscopic examination of the main organs of the animals sacrified at the end of the study revealed no apparent abnormalities.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the experimental conditions of this study, the oral LD50 of the test material was > 2000 mg/kg in rats. No signs of toxicity were observed at this dose; the test material therefore requires no classification in accordance with EU criteria.
Executive summary:

The acute oral toxicity of the test material was evaluated in a limit test which was conducted in accordance with the standardised guideline OECD 401.

Groups of fasted, 6 week old Sprague-Dawley rats (5 per sex) were given a single oral dose of the test material in an aqueous solution of methylcellulose at 0.5 % at a dose of 2000 mg/kg bw (dose volume 10 mL/kg) and observed for 14 days.

No mortality and no clinical signs were observed during the study. The body weight gains of the treated rats were normal. No gross abnormalities were observed at necropsy.

The oral LD50 (males and females) was >2000 mg/kg bw and therefore the test material is not classified for acute oral toxicity in accordance with EU criteria.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
One key and one supporting study are available to address this endpoint. The key study was a GLP study which was conducted to a standardised guideline with a high level of reporting. The supporting study was not a GLP study but was conducted to sound scientific principles. The key study was assigned a reliability score of 1 according to the criteria of Klimisch (1997) and the supporting study was assigned a reliability score of 2. Overall the quality of the database is good.

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:
21 August 2012 - 20 September 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted to GLP in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.
Qualifier:
according to
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Strain: RccHan:WIST
- Age at study initiation: approximately 8 - 12 weeks
- Weight at study initiation: 200 - 350 g
- Fasting period before study: no
- Housing: The animals were housed in groups of up to three by sex in solid-floor polypropylene cages with stainless steel lids, furnished with softwood flakes and provided with environmental enrichment items: wooden chew blocks and cardboard “fun tunnels”.
- Diet: With the exception of the exposure period, free access to food was allowed.
- Water: ad libitum with the exception of the exposure period.
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25 °C
- Humidity (%): 30 - 70 %
- Air changes (per hr): at least 15 per hour
- Photoperiod (hrs dark / hrs light): the lighting was controlled to give twelve hours continuous light and twelve hours darkness.
Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
clean air
Details on inhalation exposure:
ATMOSPHERE GENERATION
In order to facilitate aerosolisation and reduce particle size, the test material was ground using a small amount of diethyl ether in a Retsch Planetary Ball Mill (Retsch (UK) Ltd., Leeds, UK). The solvent was removed via evaporation prior to use.
A dust atmosphere was produced from the test material using an SAG 410 Solid Aerosol Generator (TOPAS GmbH, Dresden, Germany) located adjacent to the exposure chamber. The SAG 410 was connected to a metered compressed air supply.
Compressed air was supplied by means of an oil free compressor and passed through a water trap and respiratory quality filters before it was introduced to the SAG 410.
The cylindrical exposure chamber had a volume of approximately 30 litres (dimensions: 28 cm diameter x 50 cm high). The concentration within the chamber was controlled by adjusting the test material feed rate from the SAG 410. The extract from the exposure chamber passed through a ‘scrubber’ trap and was connected with a high efficiency filter to a metered exhaust system. The chamber was maintained under negative pressure.

Prior to the start of the study, test material atmospheres were generated within the exposure chamber. During this characterisation period test material input rates and the generation system were varied in order to achieve the required atmospheric conditions.


EXPOSURE PROCEDURE
Prior to the day of exposure each rat was acclimatised (for approximately 2 hours) to a tapered polycarbonate restraining tube. During the day of exposure, 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.
A target concentration of 5.0 mg/L was used for the exposure. The mean achieved concentration was 102 % of target.


EXPOSURE CHAMBER TEMPERATURE AND RELATIVE HUMIDITY
The 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.


EXPOSURE CHAMBER OXYGEN CONCENTRATION
Oxygen levels within the exposure chamber were measured by an electronic oxygen analyser (Servomex (UK) Ltd, Crowborough, East Sussex) located in a port in the animals’ breathing zone. The test atmosphere was generated to contain at least 19 % oxygen.


EXPOSURE CHAMBER ATMOSPHERE CONCENTRATION
The actual chamber concentration was measured at regular intervals during the exposure period. The gravimetric method used glass fibre filters placed in a filter holder. The holder was temporarily sealed in a vacant port in the exposure chamber in the animals’ breathing zone and a suitable, known volume of exposure chamber air was drawn through the filter using a vacuum pump.
Each filter was weighed before and after sampling in order to calculate the weight of collected test material. The difference in the two weights, divided by the volume of atmosphere sampled, gave the actual chamber concentration.
The nominal chamber concentration was calculated by dividing the mass of test material used by the total volume of air passed through the chamber.
The nominal concentration is 401 % of the actual mean achieved atmosphere concentration and shows that keeping the aerosol airborne was moderately difficult.


PARTICLE SIZE DISTRIBUTION
The particle size of the generated atmosphere inside the exposure chamber was determined three times during the exposure period using a Marple Personal Cascade Impactor (Westech IS Ltd, Beds., UK). This device consisted of six impactor stages (8.6, 5.5, 3.8, 1.7, 0.86 and 0.41 µm cut points) with stainless steel collection substrates and a back up glass fibre filter, housed in an aluminium sampler. The sampler was temporarily sealed in a sampling port in the animals’ breathing zone and a suitable, known volume of exposure chamber air was drawn through it using a vacuum pump.
The collection substrates and backup filter were weighed before and after sampling and the weight of the test material, collected at each stage, calculated by the difference.
The mean amount for each stage was used to determine the cumulative amount below each cut-off point size. In this way, the proportion (%) of aerosol less than 8.6, 5.5, 3.8, 1.7, 0.86 and 0.41 µm was calculated.
The resulting values were converted to probits and plotted against Log10 cut-point size. From this plot, the Mass Median Aerodynamic Diameter (MMAD) was determined (as the 50 % point) and the geometric standard deviation was calculated. In addition the proportion (%) of aerosol less than 4 µm (considered to be the inhalable fraction) was determined.

The MMAD was 3.19 µm, resulting in an inhalable fraction (< 4 µm) of 62.0 %. The geometric standard deviation was 2.11.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
The test atmosphere was sampled seventeen times during the exposure period and the actual concentration of the test material calculated.
Duration of exposure:
4 h
Concentrations:
5.09 mg/L
No. of animals per sex per dose:
3 animals per sex per dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 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 14 days. Any evidence of overt toxicity was recorded at each observation.
- Frequency of observations and weighing: Individual bodyweights were recorded on arrival, prior to treatment on the day of exposure and on Days 1, 3, 7 and 14.
- Necropsy of survivors performed: yes. At the end of the 14 day observation period the 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.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.09 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Mortality:
There was no mortality during the study.
Clinical signs:
other: In addition to the observations considered to be due to the restraint procedure (such as hunched posture, pilo-erection and wet fur), increased respiratory rate was noted in all animals during exposure, on removal from the chamber and one hour post-exposu
Body weight:
All males and one female exhibited bodyweight losses or showed no bodyweight gain on the first day post-exposure. All male animals exhibited a reasonable bodyweight gain during the remainder of the observation period. In contrast, all female animals exhibited slight bodyweight losses from Days 1 to 3 post-exposure. Throughout the remainder of the observation period, all female animals then exhibited reasonable bodyweight gains.
Gross pathology:
No macroscopic findings were detected amongst animals at necropsy.

Table 1 Exposure Chamber Atmosphere Concentrations

Duration of Exposure (minutes)

Net Weight of Sample (mg)

Volume of Air Sampled (L)

Chamber Flow Rate (L/min)

Atmosphere Concentration (mg/L)

6

9.66

2

60

4.83

15

10.13

2

60

5.07

30

11.01

2

60

5.51

45

10.03

2

60

5.02

60

10.79

2

60

5.40

74

10.58

2

60

5.29

90

9.71

2

60

4.86

105

10.46

2

60

5.23

120

9.78

2

60

4.89

135

10.08

2

60

5.04

151

9.53

2

60

4.77

165

11.08

2

60

5.54

180

10.04

2

60

5.02

195

10.39

2

60

5.20

210

9.73

2

60

4.87

225

10.24

2

60

5.12

237

9.60

2

60

4.80

Mean achieved atmosphere concentration: 5.09 mg/L (standard deviation 0.24)

 

Nominal Concentration

-Test material used: 312 g

-Air flow: 60 L/min

-Total generation time: 255 minutes*

-Nominal concentration: 20.4 mg/L

 

*Test atmospheres were generated for a total of 15 minutes prior to animal insertion to ensure that the test material concentration was being achieved.

 

Table 2 Particle Size Distribution - Cascade Impactor Data

Impactor Stage Number

Cut Point (µm)

Amount Collected (mg) Per Sample Number

Mean Amount Collected (mg)

1

2

3

3

8.6

0.17

0.13

0.11

0.14

4

5.5

0.86

0.43

0.33

0.54

5

3.8

0.95

0.51

0.48

0.65

6

1.7

1.08

0.53

0.58

0.73

7

0.86

0.56

0.28

0.29

0.38

8

0.41

0.19

0.03

0.08

0.10

Back-up filter

<0.41

0.01

0.00

0.01

0.01

Total mean amount of test material collected: 2.55 mg

 

Table 3 Particle Size Distribution - Calculation

Cut Point (µm)

Log10 Cut Point

Mean Cumulative Amount Less Than Cut Point

(mg)

(%)

Probit

8.6

0.935

2.41

94.5

6.60

5.5

0.740

1.87

73.3

5.62

3.8

0.580

1.22

47.8

4.95

1.7

0.230

0.49

19.2

4.13

0.86

-0.066

0.11

4.31

3.29

0.41

-0.387

0.01

0.392

2.34

MMAD: 3.19 µm

Geometric standard deviation: 2.11

Predicted amount <4 µm: 62.0 %

Table 4 Individual Bodyweights

Animal Number and Sex

Bodyweight (g) on Day:

Increment (g) During Days:

-8

0

1

3

7

14

-8 to 0

0 to 1

1 to 3

3 to 7

7 to 14

1 Male

238

277

273

278

298

313

39

-4

5

20

15

2 Male

242

272

270

271

285

301

30

-2

1

14

16

3 Male

234

273

266

271

283

298

39

-7

5

12

15

4 Female

204

227

229

225

231

239

23

2

-4

6

8

5 Female

192

211

212

208

210

221

19

1

-4

2

11

6 Female

192

215

215

212

219

225

23

0

-3

7

6

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The 4 hour LC50 was determined to be >5.09 mg/L, therefore the test material requires no classification under the conditions of this study in accordance with EU criteria.
Executive summary:

The acute inhalation toxicity of the test material was determined in a study which was conducted in accordance with the standardised guideline OECD 436.

During the study male and female RccHan:WIST strain rats (3 per sex) were exposed (nose only) for 4 hours to a dust atmosphere containing the test material at a mean concentration of 5.09 mg/L, with a MMAD of 3.19 µm.

Following exposure, the animals were observed for 14 days for signs of mortality and toxicity. At the end of the observation period, all animals were subjected to necropsy.

No deaths and no clinical signs occurred throughout the study and the 4 hour LC50 was therefore determined to be > 5.09 mg/L. The test material requires no classification under the conditions of this study in accordance with EU criteria.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The study was performed to GLP and conducted in accordance with the standardised guideline OECD 436 with a high level of reporting. The study was thereby assigned a reliability score of 1 according to the criteria of Klimisch (1997). The overall quality of the database is good.

Acute toxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

ORAL

In the key study, the acute oral toxicity of the test material was evaluated in a limit test which was conducted in accordance with the standardised guideline OECD 401. During the study groups of fasted, 6 week old Sprague-Dawley rats (5 per sex) were given a single oral dose of the test material in an aqueous solution of methylcellulose at 0.5 % at a dose of 2000 mg/kg bw (dose volume 10 mL/kg) and observed for 14 days. No mortality and no clinical signs were observed during the study. The body weight gains of the treated rats were normal. No gross abnormalities were observed at necropsy. The oral LD50 (males and females) was >2000 mg/kg bw and therefore the test material is not classified for acute oral toxicity in accordance with EU criteria.

In the supporting study, the acute oral toxicity of the test material was investigated in a limit test which was conducted broadly in accordance with the FHSA guideline 16 CFR 1500.3. During the study groups of fasted Sprague-Dawley rats (5 per sex) were given a single oral dose of the test material in distilled water at a dose of 5000 mg/kg bw and observed for 14 days. No mortality was observed, all rats gained weight and there were no signs of adverse pharmacologic effects, abnormal behaviour or gross toxicity. The oral LD50 was > 5000 mg/kg bw and therefore the substance is not classified for acute oral toxicity according to EU criteria on the basis of this study.

INHALATION

In the key study, the acute inhalation toxicity of the test material was determined in a study which was conducted in accordance with the standardised guideline OECD 436. During the study male and female RccHan:WIST strain rats (3 per sex) were exposed (nose only) for 4 hours to a dust atmosphere containing the test material at a mean concentration of 5.09 mg/L, with a MMAD of 3.19 µm. Following exposure, the animals were observed for 14 days for signs of mortality and toxicity. At the end of the observation period, all animals were subjected to necropsy. No deaths and no clinical signs occurred throughout the study and the 4 hour LC50 was therefore determined to be > 5.09 mg/L. The test material requires no classification under the conditions of this study in accordance with EU criteria.

DERMAL

In accordance with the column 2 adaptation of REACH Annex VIII, the acute dermal toxicity study (required in section 8.5.3) is not considered scientifically justified as the oral and inhalation exposure routes are the most appropriate to assess the acute toxicity hazard presented by the substance based on its physico-chemical characteristics and use pattern. Dermal exposure of the substance is considered unlikely.


Justification for selection of acute toxicity – oral endpoint
The key study was conducted in line with GLP and in accordance with the standardised guideline OECD 401. It was well reported and adhered to sound scientific principles. It was awarded a reliability score of 1 in accordance with the criteria of Klimisch (1997).

Justification for selection of acute toxicity – inhalation endpoint
Only one study is available.

Justification for selection of acute toxicity – dermal endpoint
A data waiver has been submitted to address this endpoint.

Justification for classification or non-classification

Oral

In accordance with the criteria for classification as defined in Annex I, Regulation 1272/2008, the test material does not meet the criteria for classification for acute oral toxicity.

Inhalation

In accordance with the criteria for classification as defined in Annex I, Regulation 1272/2008, the test material does not meet the criteria for classification for acute inhalation toxicity.