2-[[(butylamino)carbonyl]oxy]ethyl acrylate

Endpoint summary

Description of key information

Oral (OECD 423), rat: LD50 > 2000- 5000 mg/kg bw
Inhalation (OECD 436), rat: LC50 > 0.5 - 1 mg/L air (test substance containing > 83.6% of the main compound)

Inhalation (OECD 436), rat: LC50 > 1 - 5 mg/L air (test substance containing >=80<84 %  of the main compound)

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:

03 - 20 Feb 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

adopted in 2001

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Gyógyszerészeti és Egészségügyi Minőség- és Szervezetfejlesztési Intézet (National Institute for Quality- and Organizational Development in Healthcare and Medicines), Budapest, Hungary

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

other: Crl:WI BR

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: TOXI COOP ZRT., Budapest, Hungary
- Age at study initiation: 11 weeks old
- Weight at study initiation: 207-228 g
- Fasting period before study: The day before treatment the animals were fasted.
- Housing: group caging (3 animals/cage), laboratory bedding, type II polypropylene/polycarbonate cages
- Diet: ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, Soest Germany
- Water: tap water from municipal supply; ad libitum
- Acclimation period: 26-28 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: Sunflower oil (Helianthi annui oleum raffinatum)

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 200 mg/mL and 500 mg/mL

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION: Formulations were prepared just before the administration and stirred continuously during the treatment.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Starting dose was selected on the basis of the available information about the test item.
A full test was performed. The acute toxic class method was carried out involving a stepwise procedure with the use of 5000 mg/kg bw as the starting dose in one female rat. This animal died and then dosing was proceeded at 2000 mg/kg in three females in accordance with the criteria of Annex 3 of OECD Guideline No. 423. No animal died in the second step at 2000 mg/kg bw dose level, so treatment with 2000 mg/kg bw was repeated on further three female rats. One animal died in the third step, only, so the test was finished, the stopping criteria of Annex 2d of OECD Guideline No. 423 was met.

Doses:

2000 and 5000 mg/kg bw

No. of animals per sex per dose:

5000 mg/kg bw (1st step): one female
2000 mg/kg bw (2nd step): 3 females
2000 mg/kg bw (3rd step): 3 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed individually after dosing at least once during the first 30 minutes, then 1 h, 2 h, 3 h, 4 h after the treatment and twice each day for 14 days thereafter. The body weights were recorded on day 0 (just before the treatment), on day 1, on day 7 and on day 15.
- Necropsy of survivors performed: yes
- Other examinations performed: Individual observations were performed on the skin and fur, eyes and mucous membranes and also respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 - < 5 000 mg/kg bw

Based on:

test mat.

Mortality:

5000 mg/kg bw (1st step): 1/1 female died on Day 1
2000 mg/kg bw (2nd step): 0/3 females died
2000 mg/kg bw (3rd step): 1/3 females died on Day 7

Clinical signs:

other: 5000 mg/kg bw (1st step): clinical signs of reaction comprised of decreased activity (4 cases of 5 observations), abnormal gait (4/5), decreased righting reflex (3/5), decreased grip- and limb tone (4/5), decreased body tone (3/5), decreased abdominal ton

Gross pathology:

5000 mg/kg bw (1st step): no external findings; autolysis was observed
2000 mg/kg bw (2nd step): no external findings; in 2/3 females slight hydrometra and in 1/3 animals severe hydrometra was detected
2000 mg/kg bw (3rd step): no external findings; in the female which died on Day 7 autolysis was observed, the two survivors showed moderate hydrometra

Table 1. Acute oral toxicity

Dose [mg/kg bw]	Mortality	Clinical signs
	N*	N*
Females
5000 (1st step)	1/1	1/1
2000 (2nd step	0/3	0/3
2000 (3nd step)	1/3	0/3

*N= Number of animals/ number of animals used

 

 

Interpretation of results:

other: CLP/GHS criteria not met; no classification required according to Regulation (EC) No. 1272/2008

Conclusions:

CLP: not classified

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

The available information comprises an adequate and reliable study, 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

Reference 1

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 Oct 2015 - 20 Jan 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Version / remarks:

adopted in 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Behörde für Gesundheit und Verbraucherschutz, Hamburg, Germany

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Research Models and Services Germany GmbH, Sulzfeld, Germany
- Age at study initiation: 8 weeks (males), 9 weeks (females)
- Weight at study initiation: 239 - 270 g (males) and 218 - 250 g (females)
- Fasting period before study: animals were fasted approximately 16 hours prior to administration.
- Housing: Enter as given in the study report. in groups of 2-3 animals of the same sex per cage in MAKROLON cages (type III plus), Granulated textured wood (Granulat A2, J. Brandenburg, Goldenstedt, Germany), cages were changed and cleaned twice a week.
- Diet: Commercial diet, ssniff® R/M-H V1534 (ssniff Spezialdiäten GmbH, Soest, Germany)(analysis was performed); ad libitum (except for fasting period)
- Water: tap water; ad libitum (analysis was performed)
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Example: 22 ± 3
- Humidity (%): 40-70
- Photoperiod (hrs dark / hrs light): 12/12

(IN-LIFE DATES: From: 04 Nov 2015 To: 20 Jan 2016)

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

other: no vehicle (unchanged test item was used)

Mass median aerodynamic diameter (MMAD):

>= 3.373 - <= 3.629 µm

Geometric standard deviation (GSD):

>= 3.05 - <= 3.19

Remark on MMAD/GSD:

No smaller GSD values could be obtained with the test item supplied.

Details on inhalation exposure:

EXPOSURE SYSTEM
- dynamic inhalation chamber (air changes/h (≥ 12 times))
- nose-only exposure according to KIMMERLE & TEPPER
- The apparatus consists of a cylindrical exposure chamber (volume 40 L) which holds the animals in pyrex tubes at the edge of the chamber in a radial position.
- Actual dimensions of the Inhalation Chamber: Inner Diameter: 28.2 cm; Height: 64.6 cm; Volume: 40.3 L

ADMINISTRATION:
- The aerosol of the test item was generated using a spray-jet.
- The spray-jet was fed with compressed air (5.0 bar) from a compressor and with the test item using an infusion pump.
- At the bottom of the exposure chamber, the air was sucked off at a lower rate than created by the spray-jet in order to produce a homogenous distribution and a positive pressure in the exposure chamber (inflow 900 L/h, outflow 800 L/h).
- Control of the constant supply of compressed air and the exhaust via a manometer or an air-flow meter, respectively.
- The oxygen content in the inhalation chamber was 21% v/v.
- Particle size distribution (determined via a cascade impactor): See Table 1 under "any other information on materials and methods incl. tables"

CLASS METHOD
- Rationale for the selection of the starting concentration: Maximum recommended concentration

Analytical verification of test atmosphere concentrations:

yes

Remarks:

gravimetrical

Duration of exposure:

4 h

Concentrations:

0.52, 1.05 or 5.06 mg/L air

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: animals were observed daily until all symptoms subsided, thereafter each working day, and individual body weights were determined before the exposure on test day 1 and on test days 2, 4, 8 and 15 and at the time of death.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs (tremor, convulsions, salivation, diarrhoea, lethargy, sleep and coma, changes in the skin and fur, eyes, mucous membranes, respiratory, circulatory, autonomic and central nervous system, as well as somatomotor activity and behaviour pattern); determination of lung weights

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 0.5 - < 1 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

5.06 mg/L air: 3/3 males and 3/3 females died
1.05 mg/L air: 3/3 males and 3/3 females died
0.52 mg/L air: No mortality.
For details refer to Table 2 under "any other information on results".

Clinical signs:

other: 5.06 and 1.05 mg/L air: moderately reduced motility, moderate ataxia, slight tremor, severe dyspnoea, lacrimation and vocalisation in 3/3 males and 3/3 females 0.52 mg/L air: slightly reduced motility, slight ataxia, slight tremor, moderate dyspnoea, lacr

Body weight:

5.06 and 1.05 mg/L air: all animals died prematurely
0.52 mg/L air: No inhibition of body weight gain in 3/3 males and 3/3 females.

Gross pathology:

5.06 and 1.05 mg/L air: Necropsy revealed oedematous or emphysematous lungs in 3/3 males and 3/3 females (refer to Table 3 under "any other information on results").
0.52 mg/L air: No change at necropsy.

Other findings:

Organ weights (lung tissue) were determined. Refer to Table 3 under "any other information on results".

Table 2: Table for acute inhalation toxicity

Target Concentration [mg/L air]	Mortality	Clinical Signs
	N*	N*
Males
0	0/5	0/5
1.337	0/5	0/5
2.871	0/5	0/5
Females
0	0/5	0/5
1.337	0/5	0/5
2.871	0/5	0/5

*N= Number of animals/ number of animals used

Table 3: Necropsy findings

Target concentration [mg/L air]	Animal (sex/number)	Tissue	Finding	Organ Weight (lung) [g]
5.06	m / 1	lung	oedematous	1.73
	m / 2	lung	oedematous	1.92
	m / 3	lung	oedematous	2.53
	f / 1	lung	emphysematous	4.02
	f / 2	lung	oedematous	1.78
	f / 3	lung	emphysematous	4.33
1.05	m / 1	lung	oedematous	2.18
	m / 2	lung	oedematous	1.90
	m / 3	lung	oedematous	3.78
	f / 1	lung	oedematous	2.62
	f / 2	lung	oedematous	2.11
	f / 3	lung	oedematous	2.70
0.52	m / 1	lung	none	1.56
	m / 2	lung	none	1.72
	m / 3	lung	none	1.44
	f / 1	lung	none	1.19
	f / 2	lung	none	1.23
	f / 3	lung	none	1.21
m = male; f = female

 

Interpretation of results:

other: CLP/EU GHS Category 3 (H331) according to Regulation (EC) No 1272/2008

Conclusions:

CLP: Acute Inhal. 3, H331

Reference 2

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 Nov 2016 - 03 Jan 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Version / remarks:

adopted in 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Behörde für Gesundheit und Verbraucherschutz, Hamburg, Germany

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Research Models and Services Germany GmbH, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: approx. 8 weeks (males), approx. 9 weeks (females)
- Weight at study initiation: 240 - 290 g (males), 226 - 240 g (females)
- Fasting period before study: 16 h before exposure
- Housing: individual in Makrolon cages (type II plus), granulated textured wood bedding
- Diet: ssniff® R/M-H V1534 (ssniff Spezialdiäten GmbH, Germany), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 3.841 - <= 3.856 µm

Geometric standard deviation (GSD):

>= 3.06 - <= 3.09

Remark on MMAD/GSD:

No smaller GSD values could be obtained with the test item. The slight elevated GSD was not judged to have an influence on the respirability of the test item.

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: dynamic inhalation apparatus with cylindrical exposure chamber
- Exposure chamber volume: 28.5 L
- Method of holding animals in test chamber: restraining pyrex tubes at the edge of the chamber (radial position) not imposing undue physical, thermical or immobilisation stress on the animals
- Source and rate of air: spray jet dust generator at 900 L/h (inflow) and 800 L/h (outflow)
- System of generating particulates/aerosols: The spray jet was fed with with compressed air (5.0 bar) and with the test item using an infusion pump.
- Method of particle size determination: Analysis of the particle size distribution was carried out twice during the exposure period using a cascade impactor. The air sample is drawn through a cascade of progressively finer nozzles at a constant flow rate (5 L/min). The air jets from from this impact on pre-weighed plane sampling surfaces (slides). Each stage represents an aerodynamic size range and collects finer particles than its predecessor. Each successive stage represents a special aerodynamic cut off diameter. The slides were weighed and deltas of slides´weight were determined.
- Treatment of exhaust air: The exhaust air was drawn through gas wash-bottles.
- Temperature, humidity, pressure in air chamber: 20.5 ± 0.1 °C and 58.5 ± 0.2% at the lower concentration, 20.5 ± 0.2 °C and 58.5 ± 0.3% at the higher concentration, positive pressure at both concentrations

TEST ATMOSPHERE
- Brief description of analytical method used: gravimetrically with an air sample filter controlled by a rotatometer
- Samples taken from breathing zone: yes

TEST ATMOSPHERE
see Table 1 under "Any other information on materials and methods incl. tables"

CLASS METHOD
- Rationale for the selection of the starting concentration: Before initiating the study with the animals, a pre-test was carried out with the exposure system in order to verify that under the experimental settings chosen, the limit concentration of 5 mg/L air could be achieved by gravimetric analysis.

Analytical verification of test atmosphere concentrations:

yes

Remarks:

gravimetric

Duration of exposure:

4 h

Concentrations:

13.33 and 27.78 µL/L air (nominal concentration)
1.06 and 5.12 mg/L air (actual concentration)

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Careful clinical examinations were made at least once daily until all symptoms subsided, thereafter each working day. Observations on mortality were made at least once daily. Body weights were determined once during the acclimatisation period, before the exposure on test day 1, on test days 2, 4, 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, lung weights

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 1 - < 5 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

1.06 mg/L air: 0/3 males and 0/3 females died
5.12 mg/L air: 3/3 males (within 24 h post-dose) died, 1/3 females (during inhalation) and 2/3 females (within 24 h post-dose) died

Clinical signs:

other: 1.06 mg/L air: slight dyspnoea in 3/3 males and 3/3 females 5.12 mg/L air: moderately reduced motility, moderate ataxia, slight tremor, severe dyspnoea, abdominal position, lacrimation and vocalisation in 3/3 males and 2/3 females.

Body weight:

No effect on body weight was noted.

Gross pathology:

1.06 mg/L air: oedematous lungs in 3/3 males and 3/3 females
5.12 mg/L air: oedematous and haemorrhagic lungs in 3/3 males and 3/3 females, haemorrhage in the thorax in 1/3 males

Table 2: Results

Target concentration [mg/L air]	Mortality	Clinical Signs
	N*	N*
Males
1.06	0/3	3/3
5.12	3/3	3/3
Females
1.06	0/3	3/3
5.12	3/3	3/3

*N = Number of animals/ number of animals used

Interpretation of results:

other: CLP/EU GHS Category 4 (H332) according to Regulation (EC) No 1272/2008

Conclusions:

CLP: Acute Inhal 4, H332

Reference 3

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07 Apr - 24 May 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Version / remarks:

adopted in 2009

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.52 (Acute Inhalation Toxicity - Acute Toxic Class Method)

Version / remarks:

adopted in 2014

GLP compliance:

yes (incl. QA statement)

Remarks:

Behörde für Gesundheit und Verbraucherschutz, Hamburg, Germany

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

other: Crl: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Research Models and Services Germany GmbH, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: approx. 8 weeks (males), approx. 9 weeks (females)
- Weight at study initiation: 253 - 284 g (males), 222 - 242 g (females)
- Fasting period before study: 16 h before exposure
- Housing: individual in Makrolon cages (type II plus), granulated textured wood bedding (granulat A2, Goldenstedt, Germany)
- Diet: ssniff® R/M-H V1534 (ssniff Spezialdiäten GmbH, Soest, Germany), ad libitum
- Water: tap water, ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 2.569 - <= 2.774 µm

Geometric standard deviation (GSD):

>= 2.63 - <= 2.82

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: dynamic inhalation apparatus with cylindrical exposure chamber
- Exposure chamber volume: 28.5 L
- Method of holding animals in test chamber: restraining pyrex tubes at the edge of the chamber (radial position) not imposing undue physical, thermical or immobilisation stress on the animals
- Source and rate of air: spray jet dust generator at 900 L/h (entrance) and 800 L/h (exit)
- System of generating particulates/aerosols: The spray jet was fed with with compressed air (5.0 bar) and with the test item using an infusion pump.
- Method of particle size determination: Analysis of the particle size distribution was carried out twice during the exposure period using a cascade impactor. The air sample is drawn through a cascade of progressively finer nozzles at a constant flow rate (5 L/min). The air jets from from this impact on pre-weighed plane sampling surfaces (slides). Each stage represents an aerodynamic size range and collects finer particles than its predecessor. Each successive stage represents a special aerodynamic cut off diameter. The slides were weighed and deltas of slides´weight were determined.
- Treatment of exhaust air: The exhaust air was drawn through gas wash-bottles.
- Temperature, humidity, pressure in air chamber: 20.5 ± 0.1 °C and 58.5 ± 0.2%, 20.5 ± 0.2 °C and 58.5 ± 0.3%, positive pressure at both concentrations

TEST ATMOSPHERE
- Brief description of analytical method used: gravimetrically with an air sample filter controlled by a rotatometer
- Samples taken from breathing zone: yes

TEST ATMOSPHERE
see Table 1 under "Any other information on materials and methods incl. tables"

CLASS METHOD
- Rationale for the selection of the starting concentration: Before initiating the study with the animals, a pre-test was carried out with the exposure system in order to verify that under the experimental settings chosen, the limit concentration of 1 mg/L air could be achieved by gravimetric analysis.

Analytical verification of test atmosphere concentrations:

yes

Remarks:

gravimetric

Duration of exposure:

4 h

Concentrations:

4.44 and 8.89 µL/L air (nominal concentration)
0.54 and 1.08 mg/L air (actual concentration)

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Careful clinical examinations were made at least once daily until all symptoms subsided, thereafter each working day. Observations on mortality were made at least once daily. Body weights were determined once during the acclimatisation period, before the exposure on test day 1, on test days 2, 4, 8 and 15 and at the time of death if survival was exceeding day 1.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, lung weights

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 0.5 - < 1 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

0.54 mg/L air: 0/3 males and 0/3 females died
1.08 mg/L air: 3/3 females (within 24 h post-dose) died, 1/3 male (during inhalation) and 2/3 males (within 24 h post-dose) died

Clinical signs:

other: 0.54 mg/L air: slight reduced motility, slight ataxia in 3/3 males and 3/3 females immediately to 3 h after administration; slight dyspnoea in 3/3 males and 2/3 females immediately after administration until test day 3 1.08 mg/L air: moderately reduced mo

Body weight:

No effect on body weight was noted.

Gross pathology:

0.54 mg/L air: Necropsy revealed no pathological findings.
1.08 mg/L air: oedematous lungs in 3/3 males and 3/3 females

Table 2 Results

Symptoms/Criteria	Test substance
	0.54 mg/L air		1.08 mg/L air
	males (n = 3)	females (n = 3)	males (n = 3)	females (n = 3)
Clinical signs
reduced motility	+	+	++	++
	0’ – 3 h	0’ – 3 h	0’ – 3 h	0’ – 3 h
	(3)	(3)	(2)	(3)
ataxia	+	+	++	++
	0’ – 3 h	0’ – 3 h	0’ – 3 h	0’ – 3 h
	(3)	(3)	(2)	(3)
dyspnoea	+	+	++ - +++	+++
	0’ – TD 3	0’ – TD 3	0’ – 3 h	0’ – 3 h
	(3)	(3)	(2)	(3)
lacrimation	none	none	+	+
			0’ – 3 h	0’ – 3 h
			(2)	(3)
vocalisation	none	none	+	+
			0’ – 3 h	0’ – 3 h
			(2)	(3)
Mortality
within 3 h	0	0	1	0
within 24 h	0	0	3	3
within 7 d	0	0	3	3
within 14 d	0	0	3	3
Mean body weight (g)
start	257.3	231.3	277.0	239.7
after 7 days (TD 8)	318.7 [+ 23.8]	240.7 [+ 4.1]	-	-
after 14 days (TD 15)	370.0 [+43.8]	260.3 [+12.6]	-	-
inhibition of body weight	none	none	-	-
Necropsy findings	none	none	3 of 3	3 of 3

() number of animals affected, [] body weight gain in % compared to the start value

‘ : minutes, h: hours, TD: test days

0’: immediately after exposure

+: slight/observed, ++: moderate, +++: severe

- : all animals died prematurely

Interpretation of results:

other: CLP/EU GHS Category 3 (H331) according to Regulation (EC) No 1272/2008

Conclusions:

CLP: Acute Inhal 3, H331

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LC50

Value:

0.5 mg/m³ air

Quality of whole database:

The available information comprises adequate, reliable (Klimisch score 1) and consistent studies, 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 dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Reliable studies regarding acute oral, inhalation and dermal toxicity are available for the test substance.

Oral:

The acute oral toxicity of the test substance was determined in female rats according to OECD Guideline 423 and in compliance with GLP (Mácsai Kuthy, 2015). The test substance was formulated in sunflower oil and a starting dose of 5000 mg/kg bw was used. In a first step, 1 female received the test substance in a single oral administration by gavage. Since the animal died on Day 1, a second test with a dose level of 2000 mg/kg bw was performed in three females. No animals died at 2000 mg/kg bw and the experiment was repeated in three additional females. In the third step, 1/3 animals was found dead on Day 7. At the dose level of 5000 mg/kg bw decreased activity, disturbance of coordination (abnormal gait), decreased righting- and plantary reflex, decreased muscular tension (grip- and limb tone, body tone, abdominal tone) and disturbance of the autonomic functions (piloerection) were observed between 1 and 4 hours after administration. Macroscopic examinations at necropsy did not reveal any treatment-related abnormalities in the high dose female. A dose level of 2000 mg/kg bw was tolerated by females without clinical signs, effects on weight gain and abnormal pathological findings, however one animal died in the third step. Based on the outcome of the stepwise procedure of OECD Guideline 423, the LD50 value is considered to be > 2000 – 5000 mg/kg bw.

Inhalation:

The acute inhalation toxicity of the test substance (> 83.6% main compound) was determined in groups of 6 rats (3 males and 3 females) according to OECD 436 via nose only treatment using an aerosol generated from the undiluted test substance (Haferkorn, 2016). The mass median aerodynamic diameter (MMAD) of the particles used was between 3.373 – 3.629 µm. The animals were exposed to test substance concentrations of 0.52, 1.05 and 5.06 mg/L air for 4 hours and observed for a period of 14 days following administration. At concentrations of 5.06 and 1.05 mg/L air clinical signs such as moderately reduced motility, moderate ataxia, slight tremor, severe dyspnoea, lacrimation and vocalisation were observed in all males and females. All of these animals died prematurely within 24 hours after administration. For all animals treated with a concentration of 0.52 mg/L air the following clinical signs were observed: slightly reduced motility, slight ataxia, slight tremor, moderate dyspnoea, lacrimation and vocalisation. Necropsy revealed effects to the target (oedematous or emphysematous lungs) in all test substance treated animals. In conclusion, since all animals treated with either 5.06 or 1.05 mg/L air died but none of the animals treated with 0.52 mg/L air the LC50 value (4 hour exposure) for males and females is considered to be > 0.5 - 1 mg/L air.

In addition the acute inhalation toxicity of the test substance containing >=80<84% of the main compound was determined in groups of 6 rats (3 males and 3 females) according to OECD 436 via nose only treatment using an aerosol generated from the undiluted test substance (Haferkorn, 2017a). The mass median aerodynamic diameter (MMAD) of the particles used was between 3.841 – 3.856 µm. The animals were exposed to test substance concentrations of 1.06 and 5.12 mg/L air for 4 hours and observed for a period of 14 days following administration. At concentrations of 5.12 mg/L air clinical signs such as moderately reduced motility, moderate ataxia, slight tremor, severe dyspnoea, abdominal position, lacrimation and vocalisation were observed in all males and 2/3 females. One female died during inhalation. All further animals died prematurely within 24 hours after administration. For all animals treated with a concentration of 1.06 mg/L air slight dyspnea was observed. No mortality occurred at this concentration. Necropsy revealed oedematous and haemorrhagic lungs in all animals and haemorrhage in the thorax in 1/3 males at 5.12 mg/L air. Oedematous lungs were observed in all animals at 1.06 mg/L air. In conclusion, since all animals treated with 5.12 mg/L air but none of the animals treated with 1.06 mg/L air died the LC50 value (4 hour exposure) for males and females is considered to be > 1 - 5 mg/L air. 

In a further approach the determination of the acute inhalation toxicity of the test substance containing > 83.6% of the main compound was repeated in groups of 6 rats (3 males and 3 females) according to OECD 436 via nose only treatment using an aerosol generated from the undiluted test substance (Haferkorn, 2017b). The mass median aerodynamic diameter (MMAD) of the particles used was between 2.569 – 2.774 µm. The animals were exposed to test substance concentrations of 0.54 and 1.08 mg/L air for 4 hours and observed for a period of 14 days following administration. At concentrations of 1.08 mg/L air clinical signs such as moderately reduced motility, moderate ataxia, moderate to severe dyspnoea, lacrimation and vocalisation were observed in all females and 2/3 males immediately to 3 hours after administration. One male died during inhalation. All further animals died prematurely within 24 hours after administration. For all animals treated with a concentration of 0.54 mg/L air slightly reduced motility and slight ataxia was observed immediately and up to 3 hours after administration and slight dyspnoea was observed immediately after administration until test day 3 in all male and female rats. No mortality occurred at this concentration. Necropsy revealed oedematous lungs in all animals at 1.08 mg/L air. No pathological findings were observed in all animals at 0.54 mg/L air. In conclusion, since all animals treated with 1.08 mg/L air but none of the animals treated with 0.54 mg/L air died the LC50 value (4 hour exposure) for males and females is considered to be > 0.5 - 1 mg/L air.

Justification for classification or non-classification

The available data on acute oral toxicity do not meet the criteria for classification according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.

The available data on acute inhalation toxicity of the test substance containing > 83.6% of the main compound meet the criteria for classification according to Regulation (EC) 1272/2008. The test substance is classified as Acute Tox., Category 3, H331: Toxic if inhaled. The available data on acute inhalation toxicity of the test substance containing >=80<84% of the main compound meet the criteria for classification according to Regulation (EC) 1272/2008. The test substance is classified as Acute Tox., Category 4, H332: Harmful if inhaled.