Registration Dossier

ACUTE ORAL
LD50 > 2000 mg/kg bw (rat); OECD 423, EU Method B.1 tris, EPA OPPTS 870.1100 and JMAFF 12 Nousan, Notification No. 8147
ACUTE DERMAL
LD50 > 2000 mg/kg bw (rat); OECD 402, EU Method B.3, EPA OPPTS 870.1200 and JMAFF 12 Nousan, Notification No. 8147

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 August 2014 to 02 September 2014

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

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF, 12 Nousan, Notification No 8147

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Strain: Wistar strain Crl:WI (Han) (outbred, SPF-Quality)
- Age at study initiation: Approximately 8 weeks old
- Weight at study initiation: 147 to 163 g. Body weight variation did not exceed ± 20 % of the sex mean.
- Fasting period before study: Animals were deprived of food overnight prior to dosing and until 3 to 4 hours after administration of the test material. Water was available ad libitum.
- Housing: Group housing of 3 animals per cage in labelled cages (height 18 cm) containing sterilised sawdust as bedding material and paper as cage-enrichment.
- Diet: Free access to pelleted rodent diet
- Water: Free access to tap water
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 18 to 24 °C
- Humidity: 40 to 70 % (relative)
- Air changes: at least 10 air changes/hour
- Photoperiod: 12-hour light/12-hour dark cycle

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

MAXIMUM DOSE VOLUME APPLIED: 2.215 mL/kg body weight. Dose volume was calculated as dose level (g/kg) / density (g/mL).

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

6 females; the test material was administered to two subsequent groups of three female rats

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed for mortality and viability twice daily. Body weights were recorded on Day 1 (pre-administration of the test material) and on Days 8 and 15. At periodic intervals on the day of dosing (Day 1) and once daily thereafter until Day 15, animals were observed for clinical signs. The signs were graded according to fixed scales and the time of onset, degree and duration were recorded.
- Necropsy of survivors performed: Yes. At the end of the observation period, all animals were sacrificed by oxygen/carbon dioxide procedure and subjected to necropsy.
- Other examinations performed: Descriptions of all internal macroscopic abnormalities were recorded.

Statistics:

No statistical analysis was performed as the method used is not intended to allow the calculation of a precise LD50 value.

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Hunched posture and piloerection were noted for all animals between Days 1 and 3.

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of the animals.

Table 1: Body Weight Data (g)

Dose Group	Animal Number	Day 1	Day 8	Day 15
First group dosed with 2000 mg/kg bw	1	154	178	181
	2	163	188	198
	3	157	182	196
	Mean (SD)	158 (5)	183 (5)	192 (9)
Second group dosed with 2000 mg/kg bw	4	148	182	198
	5	153	186	196
	6	147	180	194
	Mean (SD)	149 (3)	183 (3)	196 (2)

SD = Standard deviation

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

Executive summary:

The potential of the test material to cause acute oral toxicity in the rat was investigated in accordance with the standardised guidelines OECD 423, EU Method B.1 tris, EPA OPPTS 870.1100 and JMAFF 12 Nousan, Notification No. 8147 under GLP conditions.

The test material was administered by oral gavage to two subsequent groups of three fasted female Wistar rats at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was conducted after terminal sacrifice (Day 15).

No mortality occurred. Hunched posture and piloerection were noted for all animals between Days 1 and 3. The body weight gain shown by the animals over the study period was considered to be normal. No abnormalities were found at macroscopic post mortem examination of the animals.

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

The study was conducted in accordance with standardised guidelines and under GLP conditions. The study was awarded a reliability score of 1 in accordance with the principles for assessing data quality set forth by Klimisch et al. (1997). The quality of the database is therefore considered to be high.

Endpoint conclusion:

no study available

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21 January 2015 to 04 February 2015

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

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1200 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF 12 Nousan, Notification No 8147

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Strain: Wistar strain Crl:WI (Han) (outbred, SPF-Quality)
- Age at study initiation: Approximately 10 weeks old
- Weight at study initiation: Males 259 to 285 g; females 187 to 205 g. Body weight variation did not exceed ± 20 % of the sex mean.
- Fasting period before study: No
- Housing: During acclimatisation, animals were group housed (cage height 18 cm). During the study, animals were individually housed in labelled cages (height 18 cm) containing sterilised sawdust as bedding material and paper as cage-enrichment.
- Diet: Free access to pelleted rodent diet
- Water: Free access to tap water
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 18 to 24 °C
- Humidity: 40 to 70 % (relative)
- Air changes: at least 10 air changes/hour
- Photoperiod: 12-hour light/12-hour dark cycle

Type of coverage:

occlusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: One day before exposure (Day -1) an area of approximately 5 x 7 cm on the back of each animal was clipped.
- % coverage: The test material was applied on an area of approx. 10 % of the total body surface, i.e. approx. 25 cm² for males and 18 cm² for females.
- Type of wrap if used: The test material was held in contact with the skin with a dressing, consisting of a surgical gauze patch, successively covered with aluminium foil and elastic bandage. A piece of tape was additionally used for fixation of the bandages in females only.

REMOVAL OF TEST MATERIAL
- Washing (if done): Dressings were removed and the skin cleaned of residual test material using tap water.
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied: 2.215 mL/kg body weight. Dose volume calculated as dose level (g/kg) / density (g/mL).
- Other: The test material was stirred on a magnetic stirrer during application.

Duration of exposure:

24 hours

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

5 animals per sex per dose

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed for mortality and viability twice daily. Body weights were recorded on Day 1 (pre-administration of the test material) and on Days 8 and 15. At periodic intervals on the day of dosing (Day 1) and once daily thereafter until Day 15, animals were observed for clinical signs. The signs were graded according to fixed scales and the time of onset, degree and duration were recorded.
- Necropsy of survivors performed: Yes. At the end of the observation period, all animals were sacrificed by oxygen/carbon dioxide procedure and subjected to necropsy.
- Other examinations performed: Descriptions of all internal macroscopic abnormalities were recorded.

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Lethargy, flat posture, piloerection, shallow respiration, chromodacryorrhoea (snout) and/or ptosis were noted for the animals between Days 1 and 3. Additionally, hunched posture was noted for the animals between Days 1 and 15. General erythema, erythema

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of the animals.

Table 1: Body Weight Data (g)

Dose Group	Animal Number	Day 1	Day 8	Day 15
Males dosed with 2000 mg/kg bw	1	264	267	273
	2	285	286	313
	3	272	284	311
	4	284	280	308
	5	259	267	292
	Mean (SD)	273 (12)	277 (9)	299 (17)
Females dosed with 2000 mg/kg bw	6	191	185	204
	7	205	204	203
	8	191	194	205
	9	201	194	198
	10	187	188	192
	Mean (SD)	195 (8)	193 (7)	200 (5)

SD = Standard deviation

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

Executive summary:

The potential of the test material to cause acute dermal toxicity in the rat was investigated in accordance with the standardised guidelines OECD 402, EU Method B.3, EPA OPPTS 870.1200 and JMAFF 12 Nousan, Notification No. 8147 under GLP conditions.

The test material was administered to five Wistar rats of each sex by a single dermal application at 2000 mg/kg body weight for 24 hours. After the exposure period, dressings were removed and the skin cleaned of residual test material using tap water. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (Day 15).

No mortality occurred. Lethargy, flat posture, piloerection, shallow respiration, chromodacryorrhoea (snout) and/or ptosis were noted for the animals between Days 1 and 3. Additionally, hunched posture was noted for the animals between Days 1 and 15.

General erythema, erythema maculate, scales, scabs, wounds and/or thickened areas were seen in the treated skin, abdomen and/or flanks of the animals during the observation period.

The mean body weight gain during the observation period was within the range expected for rats used in this type of study.

No abnormalities were found at macroscopic post mortem examination of the animals.

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

The study was conducted in accordance with standardised guidelines and under GLP conditions. The study was awarded a reliability score of 1 in accordance with the principles for assessing data quality set forth by Klimisch et al. (1997). The quality of the database is therefore considered to be high.

Acute Oral Toxicity

In the key study, the potential of the test material to cause acute oral toxicity in the rat was investigated in accordance with the standardised guidelines OECD 423, EU Method B.1 tris, EPA OPPTS 870.1100 and JMAFF 12 Nousan, Notification No. 8147 under GLP conditions.

The test material was administered by oral gavage to two subsequent groups of three fasted female Wistar rats at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was conducted after terminal sacrifice (Day 15).

No mortality occurred. Hunched posture and piloerection were noted for all animals between Days 1 and 3. The body weight gain shown by the animals over the study period was considered to be normal. No abnormalities were found at macroscopic post mortem examination of the animals.

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

Acute Inhalation Toxicity

In accordance with Section 8.5.2. of Column 2 of Annex VIII of the REACH Regulation, the registrant proposes to waive the acute toxicity testing by the inhalation route on the grounds that exposure of humans via inhalation is not likely taking into account the vapour pressure of the substance and the possibility of exposure to aerosols, particles or droplets of an inhalable size.

Acute Dermal Toxicity

In the key study, the potential of the test material to cause acute dermal toxicity in the rat was investigated in accordance with the standardised guidelines OECD 402, EU Method B.3, EPA OPPTS 870.1200 and JMAFF 12 Nousan, Notification No. 8147 under GLP conditions.

The test material was administered to five Wistar rats of each sex by a single dermal application at 2000 mg/kg body weight for 24 hours. After the exposure period, dressings were removed and the skin cleaned of residual test material using tap water. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (Day 15).

No mortality occurred. Lethargy, flat posture, piloerection, shallow respiration, chromodacryorrhoea (snout) and/or ptosis were noted for the animals between Days 1 and 3. Additionally, hunched posture was noted for the animals between Days 1 and 15.

General erythema, erythema maculate, scales, scabs, wounds and/or thickened areas were seen in the treated skin, abdomen and/or flanks of the animals during the observation period.

The mean body weight gain during the observation period was within the range expected for rats used in this type of study.

No abnormalities were found at macroscopic post mortem examination of the animals.

Under the conditions of this study the LD50 value was found to exceed 2000 mg/kg bw.

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

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

In accordance with the criteria for classification as defined in Annex I, Regulation (EC) No. 1272/2008, the substance does not require classification with respect to acute toxicity.