Bisbenzimidazo[2,1-b:1',2'-j]benzo[lmn][3,8]phenanthroline-6,9-dione

Endpoint summary

Description of key information

Acute oral toxicity:

The test item (C.I. Pigment Red 194) did not cause any mortality or clinical signs after single oral gavage administration to 10 female rats at 15 000 mg/kg bw in a valid pre-GLP guideline study according to FDA-guideline, similar to OECD 401.

 

Acute dermal toxicity:

Study was waived and classification for this endpoint is considered unwarranted, because the substance is not likely to become systemically available after dermal exposure and there was no evidence of toxicity observed in any of the endpoints tested.

 

Acute inhalation toxicity:

In an OECD 433 followed study the test item (C.I. Pigment Red 194) did not cause any mortality or clinical signs in rats after single inhalation of 5.02 mg/L air exposure of male rats. Hence the LC50 of the test item is >5.02 mg/L of air.  

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Test performed prior to the implementation of the current acknowledged testing and GLP guidelines . The test conduct however was in principle very similar to the OECD TG 401 as adopted in 1981. Important aspects (e.g. 14 day-postobservation time) were considered.

Qualifier:

according to guideline

Guideline:

other: Company guideline similar to OECD 401

Deviations:

no

GLP compliance:

no

Remarks:

pre-guideline study

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Strain specifics: Hoe: WISKf (SPF71)
- Source: Hoechst AG, breeding colony, SPF-breed
- Weight at study initiation: female 110 g - 120 g (mean 114 g)
- Fasting period before study: approximately 16 hours before until 2 hours after treatment, access to water permitted
- Housing: in plastic cage with standard softwood bedding
- Diet (e.g. ad libitum): standard rat diet (Altromin 1324) ad libitum
- Water (e.g. ad libitum): tap water in plastic bottles ad libitum
- Acclimation period: not necessary (breeding at identical conditions)

Route of administration:

oral: gavage

Vehicle:

other: sesame oil

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 12.5 % suspension

Doses:

15 000 mg/kg bw; 2 applications of 7 500 mg/kg bw respectively within 1 hour.

No. of animals per sex per dose:

10 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of weighing: weekly
- Necropsy of survivors performed: yes

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 15 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: no lethality within the 14 day observation period

Mortality:

- no deaths occurred

Clinical signs:

other: - post application faeces was stained red - diarrhea, squatting posture

Gross pathology:

No macroscopic findings at scheduled necropsy.

Interpretation of results:

GHS criteria not met

Remarks:

the test item has not to be classified for acute oral toxicity according to Regulation (EC) No 1272/2008

Conclusions:

Single application of 15 000 mg test item per kg bw did not cause lethality in female rats during the 14 days observation period, resulting in a LD50 > 15 000 mg/kg bw.

Executive summary:

Female rats were subjected to test acute oral toxicity according to a protocol similar to OECD TG 401 (limit test). The test item was administered at one dose level of 15 000 mg/kg bw to10 female rats. During the 14 days observation period no animals died and there were no abnormalities found in necropsy, thus leading to an LD50 > 15 000 mg/kg bw.

Therefore, the test item has not to be classified for acute oral toxicity according to Regulation (EC) No 1272/2008.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

15 000 mg/kg bw

Quality of whole database:

reliable

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:

19 September 2022 to 07 November 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 433 (Acute Inhalation Toxicity: Fixed Concentration Procedure)

Version / remarks:

adopted on 25 June 2018

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed concentration procedure

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: In-house bred animals
- Females nulliparous and non-pregnant: yes
- Age at study initiation: Sighting study: 9 to 11 weeks; Main study: 9 weeks
- Weight at study initiation: Sighting Study: Males: 183.54 g to 185.38 g; Females: 173.42 g to 174.59 g
Main Study: Males: 185.38 g to 192.86 g
- Housing: polycabonate cage (size: L 430 × B 280 × H 210 mm) with stainless steel mesh top grill
- Diet (e.g. ad libitum): Altromin maintenance diet 1324 for rats and mice (manufactured by Altromin Spezialfutter GmbH & Co. KG)
- Water (e.g. ad libitum): Deep bore-well water passed through Reverse osmosis unit
- Acclimation period: 19 September 2022 to 23 October 2022

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.8°C to 22.7°C
- Humidity (%): 43% to 66%
- Air changes (per hr): 12 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light and 12 hours dark cycle

IN-LIFE DATES: From: 19 September 2022 To: 07 November 2022

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

clean air

Mass median aerodynamic diameter (MMAD):

> 2.95 - <= 3.2 µm

Geometric standard deviation (GSD):

> 2.74 - <= 2.89

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: flow-past, nose-only dynamic inhalation exposure system
- Exposure chamber volume: 0.76
- Method of holding animals in test chamber: Restrainer
- Source and rate of air (airflow): 20 liters/min
- Method of conditioning air: Compressor air
- System of generating particulates/aerosols: RBG
- Method of particle size determination: Cascade impactor
- Treatment of exhaust air: NaOH
- Temperature, humidity, pressure in air chamber: 22.5 to 22.6°C, 55.7 to 56.4% & 60Psi

TEST ATMOSPHERE
- Samples taken from breathing zone: yes
- Time needed for equilibrium of exposure concentration before animal exposure: 0.11 min

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

> 0 - <= 4 h

Concentrations:

5 mg/L of air

No. of animals per sex per dose:

Sighting Study: 1 Male + 1 Female
Main Study: 5 Males

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: once daily thereafter for clinical signs and twice daily for mortality and body weight on day 1, 2, 4, 8 and 15
- Necropsy of survivors performed: yes

Preliminary study:

A sighting study was performed with 1 male and 1 female, and the starting concentration for the sighting study was selected from the fixed concentration levels as per Annexure 3 i.e., 1 mg/L. No mortality was observed at concentration 1.05 mg/L of air in sighting study I and 5.03 mg/L of air in sighting study II.

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.02 mg/L air (analytical)

Based on:

test mat.

Mortality:

No treatment related clinical signs and no mortalities were observed in sighting study I, II and main study at the limit concentration of 1.05, 5.03 and 5.02 mg/L of air

Clinical signs:

other: No treatment related clinical signs and no mortalities were observed in sighting study I, II and main study at the limit concentration of 1.05, 5.03 and 5.02 mg/L of air

Body weight:

The overall body weights of the animals were unaffected by treatment.
Slight decrease in body weight was noted in main study animals on day 2 after to exposure. However, all animals showed increase in body weight on days 4, 8 and 15

Gross pathology:

No treatment related gross pathological findings were noted in sighting study I, II and main study

CLINICAL SIGNS AND MORTALITY RECORD

Group  & Concentration (mg/L of air)

Animal No.

Sex

Day 1

Days

During Exposure

Post exposure

1hr*

2hrs*

3hrs*

4hrs*

30-40 min

1hr*

2

3

4

5

6

7

8

Sighting Study-I & 1.05

Rh5201

M

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5202

F

N

N

N

N

N

N

N

N

N

N

N

N

N

Sighting Study-II & 5.03

Rh5203

M

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5204

F

N

N

N

N

N

N

N

N

N

N

N

N

N

                                         *: ± 10 minutes; N: Normal; M: Male; F: Female; min: minute; hr(s): hour(s): Post exposure test item color was retained in snout region.

 

TABLE 1 (Contd…).            CLINICAL SIGNS AND MORTALITY RECORD

Group  & Concentration (mg/L of Air)

Animal No.

Sex

Day 1

Days

During Exposure

Post exposure

1 hr*

2 hrs*

3 hrs*

4 hrs*

30-40 min

1 hr*

2

3

4

5

6

7

8

9

10

11

12

13

14

15

Main study & 5.02

Rh5205

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5206

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5207

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5208

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh5209

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

            *: ± 10 minutes; N: Normal; M: Male; F: Female; min: minute; hr(s): hour(s); Post exposure test item color was retained in snout region.

 

• BODY WEIGHT (g) AND PERCENT CHANGE IN BODY WEIGHT WITH RESPECT TO DAY 1

Group &    Concentration (mg/L of air)	Animal No.	Sex	Body Weight (g) on Days						Percent Change in Body Weight with Respect to Day 1
			1^#	8	-	-	-		1-8	-	-	-
Sighting study I & 1.05	Rh5201	M	201.32	224.56	-	-	-		11.54	-	-	-
	Rh5202	F	186.56	200.05	-	-	-		7.23	-	-	-
Sighting study II & 5.03	Rh5203	M	205.52	228.14	-	-	-		11.01	-	-	-
	Rh5204	F	186.65	201.42	-	-	-		7.91	-	-	-

#: Prior to exposure; M: Male; F: Female; SD: Standard Deviation; n: Number of animals; -: Not applicable.

 

TABLE 2 (Contd…). BODY WEIGHT (g) AND PERCENT CHANGE IN BODY WEIGHT WITH RESPECT TO DAY 1

Group &    Concentration (mg/L of air)	Animal No.	Sex	Body Weight (g) on Days						Percent Change in Body Weight with Respect to Day 1
			1^#	2	4	8	15		1- 2	1-4	1-8	1-15
Main study & 5.02	Rh5205	M	210.42	207.15	213.62	228.63	252.63		-1.55	1.52	8.65	20.06
	Rh5206	M	214.92	210.46	217.12	232.44	255.91		-2.08	1.02	8.15	19.07
	Rh5207	M	211.64	209.34	215.74	229.86	254.42		-1.09	1.94	8.61	20.21
	Rh5208	M	209.34	206.05	212.28	224.82	247.85		-1.57	1.40	7.39	18.40
	Rh5209	M	215.77	212.38	218.44	233.53	258.90		-1.57	1.24	8.23	19.99
	Mean		212.42	209.08	215.44	229.86	253.94		-1.57	1.42	8.21	19.55
	(±) SD		2.81	2.54	2.51	3.43	4.11		0.35	0.34	0.51	0.78
	n		5	5	5	5	5		5	5	5	5

#: Prior to exposure; M: Male; SD: Standard Deviation; n: Number of animals.

 

• BREATHING ZONE CONCENTRATION (ACTUAL TEST ITEM CONCENTRATION)

Technical Pre-test I:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	16	600	349.61	351.17	1.56	1.52	1	1.03	1.04
2	16	600	351.95	353.54	1.59	1.52	1	1.05

 Sighting study I:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	16	600	361.71	363.31	1.60	1.52	1	1.05	1.05
2	16	600	349.07	350.64	1.57	1.52	1	1.03
3	16	600	356.94	358.58	1.64	1.52	1	1.08

BZC: Breathing Zone Concentration; Sampled volume: 1.52 L/min; Sampling time: 1 minute

BZC (f) =	Mass of test item collected on the filter paper (c)
	Volume of air passed through the filter paper (d) X time (e)

In technical pre-test, no irregular sampling (Individual sampling was within the ±20% range, when compared with mean breathing zone concentration) was noted, hence sampling was performed three times during sighting study I.

 

 

TABLE 3 (Cont…). BREATHING ZONE CONCENTRATION (ACTUAL TEST ITEM CONCENTRATION)

 Technical Pre-test II:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	50	600	354.79	362.41	7.62	1.52	1	5.01	5.02
2	50	600	353.27	360.92	7.65	1.52	1	5.03

 Sighting study II:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	50	600	351.75	359.42	7.67	1.52	1	5.05	5.03
2	50	600	348.63	356.24	7.61	1.52	1	5.01
3	50	600	352.16	359.80	7.64	1.52	1	5.03

BZC: Breathing Zone Concentration; Sampled volume: 1.52 L/min; Sampling time: 1 minute

BZC (i) =	Mass of test item collected on the filter paper (f)
	volume of air passed through the filter paper (g) X time (h)

In technical pre-test, no irregular sampling (Individual sampling was within the ±20% range, when compared with mean breathing zone concentration) was noted, hence sampling was performed three times during sighting study II.

TABLE 3 (Cont…). BREATHING ZONE CONCENTRATION (ACTUAL TEST ITEM CONCENTRATION)

  Main study:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	50	600	353.75	361.40	7.65	1.52	1	5.03	5.02
2	50	600	346.05	353.67	7.62	1.52	1	5.01
3	50	600	355.01	362.65	7.64	1.52	1	5.03

BZC: Breathing Zone Concentration; Sampled volume: 1.52 L/min; Sampling time: 1 minute

BZC (i) =	Mass of test item collected on the filter paper (f)
	volume of air passed through the filter paper (g) X time (h)

In technical pre-test, no irregular sampling (Individual sampling was within the ±20% range, when compared with mean breathing zone concentration) was noted, hence sampling was performed three times during Main study.

 

Example for calculation of Individual actual concentration variability (%) from the mean concentration.

 

Individual actual conc. variability (%) of each sample from the mean conc.	=	[Mean BZC (mg/L) –BZC (mg/L) of the sample] x 100 (%)
		Mean BZC (mg/L)

 

 

Individual actual conc. variability (%) of each sample from the mean conc. (S.I. No. 1)	=	[5.02 (mg/L) – 5.03 (mg/L)] x 100 (%)	= -0.20
		5.02 (mg/L)

 

 

 

• CHAMBER (EXPOSURE) CONDITIONS

Technical Pre-test I:

Concentration               (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Total Air flow rate (L/min) (c) = (a) + (b)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)
1.04	1	16	600	20	22.7	54.6	20.4	619
	2	16	600	20	22.6	56.7	20.5	621

Sighting study I:

Concentration (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Total Air flow rate (L/min) (c) = (a) + (b)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)
1.05	1	16	600	20	22.5	57.5	20.5	618
	2	16	600	20	22.7	55.6	20.3	619
	3	16	600	20	22.4	56.8	20.5	622
	4	16	600	20	22.6	55.6	20.4	625

Note: 1% Carbon dioxide = 10000 ppm

 

 

TABLE 4 (Cont…). CHAMBER (EXPOSURE) CONDITIONS

Technical Pre-test II:

Concentration               (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Total Air flow rate (L/min) (c) = (a) + (b)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)
5.02	1	50	600	20	22.7	55.1	20.6	620
	2	50	600	20	22.5	56.8	20.3	611

Sighting study II:

Concentration (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Total Air flow rate (L/min) (c) = (a) + (b)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)
5.03	1	50	600	20	22.5	53.9	20.4	611
	2	50	600	20	22.7	55.7	20.3	619
	3	50	600	20	22.4	56.1	20.6	621
	4	50	600	20	22.5	57.3	20.3	625

Note: 1% Carbon dioxide = 10000 ppm

 

 

TABLE 4 (Cont…). CHAMBER (EXPOSURE) CONDITIONS

Main study:

Concentration (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Total Air flow rate (L/min) (c) = (a) + (b)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)
5.02	1	50	600	20	22.5	55.4	20.4	616
	2	50	600	20	22.6	56.5	20.3	625
	3	50	600	20	22.3	57	20.5	630
	4	50	600	20	22.7	53.9	20.2	625

Note: 1% Carbon dioxide = 10000 ppm

 

• SUMMARY OF CHAMBER (EXPOSURE) CONDITIONS

Technical Pre Test I
Chamber Conditions	Range
Temperature (°C)	22.6 to 22.7
Relative humidity (%)	54.6 to 56.7
Oxygen concentration (%)	20.4 to 20.5
Carbon dioxide concentration (ppm)	619 to 621
Air Flow (L/min)*	20
BZC (mg/L)	1.03 to 1.05
MMAD (µm)	2.76 ^#
GSD	2.78 ^#
Sighting study I
Temperature (°C)	22.4 to 22.7
Relative humidity (%)	55.6 to 57.5
Oxygen concentration (%)	20.3 to 20.5
Carbon dioxide concentration (ppm)	618 to 625
Air Flow (L/min)*	20
BZC (mg/L)	1.03 to 1.08
MMAD (µm)	2.87 to 2.99
GSD	2.88 to 2.92

*: Values were constant throughout the exposure; ^#_: Individual value; BZC: Breathing Zone Concentration (Actual Concentration); MMAD: Mass Median Aerodynamic Diameter; GSD: Geometric Standard Deviation.

 

 

TABLE 7 (Cont…). SUMMARY OF CHAMBER (EXPOSURE) CONDITIONS

Technical Pre Test II
Chamber Conditions	Range
Temperature (°C)	22.5 to 22.7
Relative humidity (%)	55.1 to 56.8
Oxygen concentration (%)	20.3 to 20.6
Carbon dioxide concentration (ppm)	611 to 620
Air Flow (L/min)*	20
BZC (mg/L)	5.01 to 5.03
MMAD (µm)	3.50 ^#
GSD	2.87 ^#
Sighting study II
Temperature (°C)	22.4 to 22.7
Relative humidity (%)	53.9 to 57.3
Oxygen concentration (%)	20.3 to 20.6
Carbon dioxide concentration (ppm)	611 to 625
Air Flow (L/min)*	20
BZC (mg/L)	5.01 to 5.05
MMAD (µm)	3.17 to 3.37
GSD	2.72 to 2.85

*: Values were constant throughout the exposure; ^#_: Individual value; BZC: Breathing Zone Concentration (Actual Concentration); MMAD: Mass Median Aerodynamic Diameter; GSD: Geometric Standard Deviation.

  

TABLE 7 (Cont…). SUMMARY OF CHAMBER (EXPOSURE) CONDITIONS

Main study
Chamber Conditions	Range
Temperature (°C)	22.3 to 22.7
Relative humidity (%)	53.9 to 57.0
Oxygen concentration (%)	20.2 to 20.5
Carbon dioxide concentration (ppm)	616 to 630
Air inlet (L/min)*	20
BZC (mg/L)	5.01 to 5.03
MMAD (µm)	3.17 to 3.21
GSD	2.71 to 2.78

*: Values were constant throughout the exposure; BZC: Breathing Zone Concentration (Actual Concentration); MMAD: Mass Median Aerodynamic Diameter; GSD: Geometric Standard Deviation.

 

• GROSS PATHOLOGY FINDINGS

Group & Concentration (mg/L of Air)	Animal No.	Sex	Fate	Gross Pathological Findings
				External	Internal
Sighting Study-I & 1.05	Rh5201	M	TS	NAD	NAD
	Rh5202	F	TS	NAD	NAD
Sighting Study-II & 5.03	Rh5203	M	TS	NAD	NAD
	Rh5204	F	TS	NAD	NAD
Main Study & 5.02	Rh5205	M	TS	NAD	NAD
	Rh5206	M	TS	NAD	NAD
	Rh5207	M	TS	NAD	NAD
	Rh5208	M	TS	NAD	NAD
	Rh5209	M	TS	NAD	NAD

NAD: No Abnormality Detected; M: Male; F: Female; TS: Terminal Sacrifice

Interpretation of results:

GHS criteria not met

Remarks:

Test item has not to be classified as acutely toxic by inhalation route according to Regulation (EC) No 1272/2008.

Conclusions:

Under the experimental conditions employed and based on the above results of this study, there were no clinical signs and no mortalities observed at the mean maximum achievable concentration of 5.02 mg/L of air. Hence, the LC50 of the test item is > 5.02 mg/L of air.

Executive summary:

The test item was evaluated for acute inhalation toxicity in Sprague Dawley rats.

The objective of the study was to assess the toxic potential and to determine the LC₅₀ of the test item, when administered by inhalation route through flow-past nose-only dynamic inhalation equipment for a single 4 hours exposure to rats and also to classify for risk assessment of chemical according to globally harmonized system of classification and labelling of chemicals. One male and one female rat was used for each sighting study and five male rats were used to perform the main study.

As such the test item was used during technical pre-test, sighting study I, II and main study to generate the dust aerosols through rotating brush generator. The technical pre-test was carried out without animals. During, technical pre-test of sighting study -I and technical pre-test of sighting study -II, the limit concentration was achieved 1.04 and 5.02 mg/L of air at the feed rate of 16 and 50 mm/hour with a rotation of 600 RPM, hence the same speed was selected for sighting study and main study to generate the limit concentration of test item.

The mean chamber conditions like temperature, relative humidity, oxygen concentration, carbon dioxide concentration, MMAD and GSD were observed during the exposure and were found within the acceptable range.

All the animals were observed for clinical signs of toxicity and mortality during exposure and post exposure on day 1 and once daily thereafter for clinical signs and twice daily for mortality for 7 days for sighting study I, II and 14 days for main study post exposure period. Individual animal body weight was recorded on the day of dosing, and on days 1 and 8 for the sighting studies and on days 1, 2, 4, 8 and 15 for the main study. All rats were euthanized after 7 or 14 days post exposure by intraperitoneal administration of sodium thiopentone and they were subjected to gross necropsy.

In sighting study I and II, the animals were exposed at mean concentration of 1.05 and 5.03 mg/L of air, determined by gravimetric method. The Rotating Brush Generator was used to generate the dust particles (aerosols) into an airborne state.

In the main study five male animals were exposed at mean concentration of 5.02 mg/L of air determined by gravimetric method. During main study the target concentration of 5.02 mg/L of air was achieved at the feed rate of 50 mm/hour with a rotation of 600 RPM.

No treatment related clinical signs of toxicity and mortalities were observed in the sighting study I, II and main study. Slight decrease in body weight was noted in main study on day 2 due to exposure. All animals showed increase in body weight on days 4, 8 and 15.

No treatment related gross pathological findings were noted in any of the animals in the sighting study I, II and the main study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LC50

Value:

> 5.02 mg/L air

Physical form:

inhalation: aerosol

Quality of whole database:

reliable without restriction

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

The LD50 of the test item after a single oral administration to female rats, observed over a period of 14 days, was greater than 15 000 mg/kg body weight. In addition, the LC50 of the test item after inhalation exposure of male rats was >5.02 mg/L air. 

Therefore, the test item has not to be classified as acutely toxic (including STOT SE) according to Regulation (EC) No 1272/2008.