Registration Dossier

Administrative data

Description of key information

Oral (OECD 423), rat: LD50 > 2000 mg/kg bw (limit test)
Inhalation (OECD 403), rat: LC50 > 2.1 mg/L air (maximum attainable concentration, limit test)
Dermal (OECD 402), rat: LD50 > 2000 mg/kg bw (limit test)

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:
31 July - 22 Aug 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-Guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
2002
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No. 8147, April 2011; including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
other: Crl:WI (Han) (outbred, SPF-Quality)
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approx. 8-10 weeks
- Weight at study initiation: 159-196 g
- Fasting period before study: Animals were fasted overnight prior to administration and until approx. 1 hour and 10 minutes (first group at 2000 mg/kg bw) or approx. 30 minutes (second group at 2000 mg/kg bw) after the second administration of the test substance. Water was available ad libitum.
- Housing: groups of 3 animals in Makrolon cages (MIV type; height 18 cm), sterilized sawdust was used as bedding material, paper as cage-enrichment
- Diet: SM R/M-Z (SSNIFF Spezialdiäten GmbH, Soest, Germany), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24
- Humidity (%): 40-70
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 31 July 2014 To: 22 Aug 2014
Route of administration:
oral: gavage
Vehicle:
propylene glycol
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

FREQUENCY: Due to a maximum concentration of 10% test substance formulations two dosages within 24 hours on Day 1 were applied. The first on t=0 hours and the second on t=3 hours.
Doses:
2000 mg/kg bw (by administration of two times 1000 mg/kg bw)
No. of animals per sex per dose:
6 (3 per step)
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed at periodic intervals on the day of dosing (Day 1) and once daily thereafter until Day 15. Individual body weights were determined on Day 1 before administration of the test substance and thereafter in weekly intervals. Observations on mortality were made twice daily.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: LD50 cut-off 5000 mg/kg bw according to OECD 423
Mortality:
No mortality occurred during the study period.
Clinical signs:
Hunched posture and piloerection were noted for all animals between Days 1 and 3. Uncoordinated movements were noted for one animal on Day 1.
Body weight:
All animals gained the expected body weight over the study period.
Gross pathology:
No abnormalities were noted at necropsy of animals.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
CLP: not classified
DSD: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

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:
20 Nov - 04 Dec 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-Guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Version / remarks:
2009
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1300 (Acute inhalation toxicity)
Version / remarks:
1998
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No. 8147, April 2011; including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
Health Care Inspectorate, Ministry of Health, Welfare and Sport, Den Haag, The Netherlands
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: Crl:WI(Han) (outbred, SPF-Quality)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approx. 9 weeks old
- Weight at study initiation: 246-300 g (males), 180-196 g (females)
- Housing: groups of 5 animals per sex per cage in Makrolon cages (type IV; height 18 cm), sterilized sawdust was used as bedding material, paper as cage-enrichment
- Diet: SM R/M-Z (SSNIFF Spezialdiäten GmbH, Soest, Germany), ad libitum except during exposure period
- Water: tap water, ad libitum except during exposure period
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24
- Humidity (%): 40-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 20 Nov 2014 To: 04 Dec 2014
Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The design of the exposure chamber is based on the flow past nose-only inhalation chamber. The chamber consisted of three animal sections with eight animal ports each. Each animal port had its own atmosphere inlet and exhaust outlet.
- Method of holding animals in test chamber: Animals were placed in restraining tubes and connected to the animal ports.
- Source and rate of air: 1 L/min
- System of generating particulates/aerosols: The test substance was fed to a stream of pressurized air (mean air flow 22 L/min) by means of a spiral feeder and a vertically placed air mover. Vibrators were used to keep the material moving. The dust was passed through two vertical elutriators, allowing larger particles to settle, and subsequently let through the exposure chamber. The rotation speed of the feeder and air flow were varied to obtain the desired exposure concentration. The generation was interrupted eight times to remove test substance deposits from the system or to change the set-up. The generation time was elongated with 27 minutes in order to achieve an actual exposure time of 240 minutes.
- Method of particle size determination: The particle size distribution was characterized twice during the exposure period. The samples were drawn (2 L/min) from the test atmosphere through a tube mounted in one of the free animal ports of the middle section of the exposure chamber. The samples were collected with an 8 stage Marple personal cascade impactor containing fiber glass filters and a fiber glass back-up filter. Amounts of test substance collected were measured gravimetrically. Subsequently the Mass Median Aerodynamic Diameter (MMAD) and the Geometric Standard Deviation (GSD) were determined.
- Treatment of exhaust air: From the exposure chamber the test atmosphere was passed through a filter before it was released to the exhaust of the fume hood.
- Temperature, humidity in air chamber: 20.0-20.2 °C, 24-28%

TEST SUBSTANCE PREPARATION: The test substance was sieved (wire mesh sieve 500 µm) and dried in a desiccator overnight prior to use.

TEST ATMOSPHERE
- Brief description of analytical method used: The actual concentration was determined sixteen times during the exposure period. Samples were drawn from the test atmosphere through a tube mounted in one of the free animal ports of the middle section of the exposure chamber. Samples were drawn through a glass fiber filter. The collected amount of the test substance in the air sample was measured gravimetrically. Sample volumes were measured by means of a dry gas meter. Subsequently the time-weighted mean concentration with the standard deviation was calculated.

TEST ATMOSPHERE
- Particle size distribution: Please refer to Table 1.
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): 4.0 µm / 2.4 µm (in both determination)
Analytical verification of test atmosphere concentrations:
yes
Remarks:
gravimetric
Duration of exposure:
4 h
Concentrations:
2.1 ± 0.06 mg/L (mean actual concentration)
38 mg/L (nominal concentration)
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 observed three times during exposure for mortality, behavioural signs of distress and effects on respiration. After exposure animals were observed on Day 1, one and three hours after exposure and once daily thereafter until Day 15. Individual body weights were determined before administration of the test substance (Day 1) and thereafter on Days 2,4,8 and 15. Observations on mortality were made twice daily.
- Necropsy of survivors performed: yes

- Other examinations performed: Clinical signs: The symptoms were graded according to fixed scales and the time of onset, degree and duration were recorded.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 2.1 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
other: maximum attainable concentration
Mortality:
No mortality occurred during the study period.
Clinical signs:
other: During exposure, no clinical signs were seen. After exposure, one female showed lethargy, hunched posture, laboured respiration and chromodacryorrhoea (eye left and right). The animal had recovered from the signs by Day 4. No clinical signs of systemic to
Body weight:
All animals gained the expected body weight over the study period.
Gross pathology:
Macroscopic post mortem examination revealed reddish discoloration of the thymus for one female. No abnormalities were noted at necropsy of the other animals that were killed at the end of the study.

Table 1. Aerodynamic particle size distribution in the test atmosphere.

Measurement 1

Stage

Cut point (µm)

Mass sampled (mg)

Relative mass (%)

Cumulative mass
(% of total sampled)

1

21

0.03

1.46

98.54

2

15

0.04

1.94

96.6

3

10

0.36

17.48

79.13

4

6

0.34

16.5

62.62

5

3.5

0.36

17.48

45.15

6

2

0.41

19.9

25.24

7

0.9

0.44

21.36

3.88

8

0.5

0.08

3.88

0

Back up

0.25

0

0

0

Measurement 2

1

21

0.23

4.22

95.78

2

15

0.11

2.02

93.76

3

10

0.41

7.52

86.24

4

6

0.97

17.8

68.44

5

3.5

0.73

13.39

55.05

6

2

1.49

27.34

27.71

7

0.9

1.16

21.28

6.42

8

0.5

0.34

6.24

0.18

Back up

0.25

0.01

0.18

0

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
CLP: not classified
DSD: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

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:
14 - 28 Aug 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-Guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Version / remarks:
1987
Deviations:
yes
Remarks:
occlusive patch was used
Qualifier:
according to guideline
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Version / remarks:
1998
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No. 8147, April 2011; including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: Crl:WI (Han) (outbred, SPF-Quality)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approx. 11 weeks old
- Weight at study initiation: 279-292 g (males), 198-203 g (females)
- Housing: Animals were housed individually in Makrolon cages (MIII type, height 18 cm), sterilized sawdust was used as bedding material, paper as cage-enrichment.
- Diet: SM R/M-Z (SSNIFF Spezialdiäten GmbH, Soest, Germany), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24
- Humidity (%): 40-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 18 Aug 2014 To: 28 Aug 2014
Type of coverage:
occlusive
Vehicle:
propylene glycol
Details on dermal exposure:
TEST SITE
- Area of exposure: clipped skin of back (25 cm² for males and 18 cm² for females)
- % coverage: 10%
- Type of wrap if used: The treated skin was covered with a dressing, consisting of a surgical gauze patch, successively covered with aluminum foil and Coban elastic bandage. A piece of Micropore tape was additionally used for fixation of the bandages in females only.

REMOVAL OF TEST SUBSTANCE
- Washing: Residual test material was removed with tap water.
- Time after start of exposure: 24 h

TEST MATERIAL
- Amount(s) applied: 20 mL/kg bw
- Constant volume or concentration used: yes
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 and weighing: Animals were observed at periodic intervals on the day of dosing (Day 1) and once daily thereafter until Day 15. Individual body weights were determined before administration of the test substance and thereafter in weekly intervals. Observations on mortality were made twice daily.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality occurred during the study period.
Clinical signs:
No clinical signs of systemic toxicity were observed up to the end of the 14-day observation period. Scales and/or scabs were seen in the treated skin-area of three animals from Day 8 onwards.
Body weight:
All animals gained the expected body weight over the study period.
Gross pathology:
No abnormalities were noted at necropsy of animals.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
CLP: not classified
DSD: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

Additional information

Oral

The acute oral toxicity of the test substance was assessed in a limit test according to OECD Guideline 423 and in compliance with GLP (Latour, 2014). A total dose of 2000 mg/kg bw of the test substance in propylene glycol was administered to two subsequent groups of three female Wistar rats. The total dose was administered as two dosages of 1000 mg/kg bw each (the first on t=0 hours and the second on t=3 hours), due to a maximum concentration of 10% test substance formulations. Animals were observed daily for clinical signs and body weight was determined weekly. Macroscopic examination was performed at the end of the 14-day observation period at terminal sacrifice. There were no mortalities during the observation period and body weight gain of all animals was considered to be normal. Uncoordinated movements were noted for one animal on Day 1 and hunched posture and piloerection were noted for all animals between Days 1 and 3. Macroscopic post mortem examination did not reveal any abnormalities. Based on the results of this study, the LD50 value was determined to be > 2000 mg/kg bw in rats. In accordance with OECD Guideline 423, Annex 2d, a cut-off value of>5000 mg/kg bw was derived, since no mortality occurred at 2000 mg/kg bw.

 

Inhalation

The acute inhalation toxicity of the test substance was assessed in a limit test performed according to OECD Guideline 403 and in compliance with GLP (van Huygevoort, 2015). The test substance was administered as a dust by nose-only inhalation for 4 hours to one group of 5 male and 5 female Wistar rats to the technically maximum attainable concentration. The time-weighted mean actual concentration was 2.1 mg/L and the nominal concentration was 38 mg/L. The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined 4.0 µm (gsd 2.4) in two measurements during the exposure period. Animals were subjected to daily observations and determination of body weights on Day 1, 2, 4, 8 and 15. Macroscopic examination was performed after terminal sacrifice on Day 15. There was no mortality and no effects on body weight gain during the 14-day observation period. During exposure, no clinical signs were seen. After exposure, one female showed lethargy, hunched posture, laboured respiration and chromodacryorrhoea (eye left and right). The animal had recovered from the signs by Day 4. No clinical signs were seen for the other animals. Macroscopic post mortem examination revealed reddish discoloration of the thymus for one female. No abnormalities were found at macroscopic post mortem examination of the other animals. The LC50 value for inhalation toxicity is considered to be > 2.1 mg/L air.

 

Dermal

The acute dermal toxicity of the test substance was assessed in a limit test performed in 5 male and 5 female Wistar rats according to OECD Guideline 402 and in compliance with GLP (Latour, 2014). A single dose of 2000 mg/kg bw of the test substance in propylene glycol was applied to the clipped skin of rats under occlusive conditions for 24 hours. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice on Day 15. There was no mortality and no effects on body weight gain during the 14-day observation period. Scales and/or scabs were seen in the treated skin-area of 3 animals from Day 8 onwards. No abnormalities were found at macroscopic post mortem examination of the animals. The LD50 value for dermal toxicity is considered to be > 2000 mg/kg bw.


Justification for selection of acute toxicity – oral endpoint
The reliable GLP compliant OECD Guideline study was chosen.

Justification for selection of acute toxicity – inhalation endpoint
The reliable GLP compliant OECD Guideline study was chosen.

Justification for selection of acute toxicity – dermal endpoint
The reliable GLP compliant OECD Guideline study was chosen.

Justification for classification or non-classification

The available data on acute oral, inhalation and dermal toxicity of the test substance do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.