Tetrabutylammonium bromide

Endpoint summary

Description of key information

Acute oral toxicity: 

Acute oral toxicity dose (LD50) of test chemical was considered based on experimental study conducted on rats, the LD50 value was considered in between 300-2000 mg/kg bw in rats. Thus, comparing this range with the criteria of CLP regulation, the given test chemical can be classified in “Category 4” for acute oral toxicity.

Acute Inhalation toxicity: 

The acute inhalation toxicity study need not be conducted because exposure to humans via inhalation route is not likely taking into account the low vapour pressure of the test substance, which is reported as 0.000000000796 mm Hg. Thus, exposure to inhalable dust, mist and vapour of the chemical is highly unlikely. Therefore this study is considered for waiver.

Acute Dermal Toxicity:

Acute Dermal toxicity dose (LD50) for target chemical was considered based on experimental study conducted on rats, the value was considered to be >2000 mg/kg bw in rats. Thus, comparing this value with the criteria of CLP regulation, the given test chemical cannot be classified for acute dermal toxicity.

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:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Data is from experimental study report.

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

The aim of this study was to assess the toxicity potential of test chemical after single oral administration in rats.

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

no

Specific details on test material used for the study:

- Substance type:Organic
- Physical state:White Solid (Crystals)
- Analytical purity:99.9%
- Lot/batch No.:Lot 1/02
- Storage condition of test material:Stored in cool, dry place. Kept in closed container when not in use.
- Other: Handling and Disposal
Safety precautions: Avoided eye and skin contact. Avoided inhalation and spilling. Aprons, caps, mask, gloves and goggles were used to ensure the health and safety of the personnel.

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:Bharat Serum and Vaccines Limited, India.
- Age at study initiation:8- 11 weeks at the time of dosing.
- Health Status:Healthy young adult animals were used for the study. Females were nulliparous and non pregnant.
- Weight at study initiation:Minimum: 197 g and Maximum: 218 g (Individual body weights were within ± 4% prior to treatment after overnight fasting)
- Fasting period:The animals were fasted for minimum 16-18 hours prior to dosing and for 3-4 hours post dosing.
- Housing:The animals were housed individually in polycarbonate cages.
- Bedding: All cages were provided with corn cobs.
- Room Sanitation:The experimental room floor and work tops were swept and mopped with disinfectant solution every day.
- Cages and water bottle:All the cages and water bottles were changed at least twice every week.
- Diet (e.g. ad libitum):All animals were provided conventional laboratory rodent diet (Nutrivet Life Sciences, Pune) ad libitum.
- Water (e.g. ad libitum):Aqua guard filtered tap water was provided ad libitum via drinking bottles.
- Acclimation period:Animal nos. 1-3 were acclimatized for five days and 4-6 for seven days prior to administration of the test item.

ENVIRONMENTAL CONDITIONS
- Temperature (°C):Minimum: 20.40°C and Maximum: 23.10°C.
- Humidity (%):Minimum: 38.40 % and Maximum: 58.70 %.
- Air changes (per hr):More than 12 changes per hour.
- Photoperiod (hrs dark / hrs light):12:12

IN-LIFE DATES: From: February 05, 2014 To: February 26, 2014

Route of administration:

oral: unspecified

Vehicle:

other: distilled water

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 300 mg/kg and 2000 mg/kg
- Amount of vehicle (if gavage):10 ml

Doses:

G1 = 300 mg/kg bw
G2 = 2000 mg/kg bw

No. of animals per sex per dose:

9 female rats

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical Observation - After test item administration, individual animals were frequently observed at 30 minutes, 1, 2, 3 and 4 hours post dosing on day 0 (day of dosing). Subsequently, all the surviving animals were observed once a day during the 14 day observation period.
Body weight - All surviving rats were weighed on days 0 (prior to dosing), 7 and 14. Animals were weighed immediately after found dead.
Mortality - All surviving animals were observed twice daily (morning and evening) for morbidity and mortality, throughout the acclimatization and study period.
- Necropsy of survivors performed: yes, at the end of 14 day observation period, all the survived rats were euthanised by overdose of CO2 for external and internal observations.

Statistics:

not specified

Preliminary study:

not specified

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 300 - < 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: no other details available

Mortality:

No mortality was observed in the animals treated with 300 mg/kg dose throught out the 14 days observation period, whereas all three animals treated with 2000 mg/kg dose level were found dead on day 0 post dosing.

Clinical signs:

other: At 300 mg/kg, all the six animals were observed normal throughout the experiment period. At 2000 mg/kg, animal no. 7 was observed with mild tremors at 30 minutes, mild to moderate abdominal breathing at 30 minutes and 1 hour and sternal recumbency at 1 h

Gross pathology:

No external gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, animal no. 7 was observed with no abnormalities, whereas animal nos. 8 and 9 were observed with wet around mouth. No internal gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, all three animals were observed with severe red discoloration of all lobes lungs and test item was observed in stomach and intestine.

Other findings:

not specified

Table 1: Individual Animal Body Weight (g) andBody Weight Changes(%)

Sex:Female

Animal No.	Group/ Dose (mg/kg)	Body Weight (gram)				Body Weight Change (%)
		Day 0	Day 7	Day 14	Found Dead Animal	Day 0-7	Day 0-14
1	G1/ 300	197	219	227	-	11.17	15.23
2		206	238	258	-	15.53	25.24
3		207	230	246	-	11.11	18.84
4		200	205	210	-	2.50	5.00
5		197	218	212	-	10.66	7.61
6		198	220	223	-	11.11	12.63
7	G2/ 2000	218	-	-	214	-	-
8		215	-	-	217	-	-
9		212	-	-	211	-	-

Key:- = Not Applicable

 

Table 2: Summary of Animal Body Weight (g) and Body Weight Changes (%)

Sex:Female

Group/ Dose (mg/kg)		Rats Body Weight (g)			Body Weight Changes (%)
		Day 0	Day 7	Day 14	0-7	0-14
G1/ 300	Mean	200.83	221.67	229.33	10.35	14.09
	SD	4.54	11.29	19.08	4.25	7.42
	n	6	6	6	6	6
G2/ 2000	Mean	215.00	-	-	-	-
	SD	3.00	-	-	-	-
	n	3	-	-	-	-

Keys:- = Not Applicable, SD = Standard Deviation, n = Number of Animals

Table 3: Individual Animal Clinical Signs and Symptoms

Sex:Female

Animal No.	Group/ Dose (mg/kg)	Hours (Day 0)
		1/2	1	2	3	4
1	G1/ 300	1	1	1	1	1
2		1	1	1	1	1
3		1	1	1	1	1
4		1	1	1	1	1
5		1	1	1	1	1
6		1	1	1	1	1
7	G2/ 2000	166^+ 4^+	155 4^++ 2	-	-	-
8		166^+ 4^++ 155 145^++ 2	-	-	-	-
9		166^+ 4^++ 155 145^++ 2	-	-	-	-

 

Animal No.		Group/ Dose (mg/kg)		Days post dosing
			1		2		3		4		5		6		7		8		9		10		11		12		13		14
1		G1/ 300		1		1		1		1		1		1		1		1		1		1		1		1		1		1
2			1		1		1		1		1		1		1		1		1		1		1		1		1		1
3			1		1		1		1		1		1		1		1		1		1		1		1		1		1
4			1		1		1		1		1		1		1		1		1		1		1		1		1		1
5			1		1		1		1		1		1		1		1		1		1		1		1		1		1
6			1		1		1		1		1		1		1		1		1		1		1		1		1		1
7		G2/ 2000		-		-		-		-		-		-		-		-		-		-		-		-		-		-
8			-		-		-		-		-		-		-		-		-		-		-		-		-		-
9			-		-		-		-		-		-		-		-		-		-		-		-		-		-

Keys:- = Not applicable, 1 = Normal, 2 = Found dead, 4 = Abdominal breathing, 145 = Salivation,      155 = Sternal recumbency, 166 = Tremors, + = Mild, ++ = Moderate

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The acute oral LD50 value of test chemical was considered in between 300-2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that the given test chemical exhibits acute oral toxicity in “Category 4” LD50 >300 to ≤ 2000 mg/kg body weight.

Executive summary:

Acute oral toxicity study of test chemical was conducted as per OECD No. 423 in rats. Nine female Wistar rats were selected for acute oral toxicity study. The animals were fasted for minimum 16-18 hours prior to dosing and for 3-4 hours post dosing, with food withheld but drinking water providedad libitum. The time intervals between dosing were determined by the onset, duration and severity of toxic signs. Three rats of first group were dosed with starting dose of 300 mg/kg body weight and the animals did not show any mortality so another three animals of the same group were dosed with 300 mg/kg body weight and no mortality was observed. Hence three rats of second group were dosed with 2000 mg/kg weight. All the rats at 2000mg/kg were found dead on day 0 post dosing. Hence, further dosing was stopped. Body weights were re­corded on day 0 (prior to dosing), 7 and 14. Body weight gain was observed in all surviving animals treated with 300 mg/kg body weight, during the 14 day observation period, as compared to day 0. At 300 mg/kg, all the six animals were observed normal throughout the experiment period. At 2000 mg/kg, animal no. 7 was observed with mild tremors at 30 minutes, mild to moderate abdominal breathing at 30 minutes and 1 hour and sternal recumbency at 1 hour followed by found dead. Animal nos. 8 and 9 were observed with mild tremors, moderate abdominal breathing, sternal recumbency and moderate salivation at 30 minutes followed by found dead. No external gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, animal no. 7 was observed with no abnormalities, whereas animal nos. 8 and 9 were observed with wet around mouth. No internal gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, all three animals were observed with severe red discoloration of all lobes lungs and test item was observed in stomach and intestine. Under the conditions of this, acute oral toxicity dose LD50 value of test chemical was considered in between 300-2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that the given test chemical exhibits acute oral toxicity in “Category 4” LD50 >300 to ≤ 2000 mg/kg body weight.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

1 000 mg/kg bw

Quality of whole database:

Data is Klimisch 1 and from experimental study report.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

other justification

Justification for data waiving:

other:

Clinical signs:

other:

Endpoint conclusion

Quality of whole database:

Waiver

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Data is from experimental study report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Principles of method if other than guideline:

The objective of the study was to assess the dermal toxicity of the test chemical after single dose application by dermal route in rats.

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
- Source:In-House Bred at Sa-Ford, Animal Facility.
- Age at study initiation:Healthy young adult animals were used for the study.
- Females (if applicable) nulliparous and non-pregnant: yes
- Weight (Prior to Treatment):Male:Minimum: 217 g and Maximum: 251 g, Female:Minimum: 227 g and Maximum: 242 g
- Housing:The animals were housed individually in polycarbonate cages.
- Bedding : All cages were provided with corn cobs.
- Room Sanitation : The experimental room floor and work tops were swept and mopped with disinfectant solution every day.
- Cages and water bottle : All the cages and water bottles were changed at least twice every week.
- Diet (e.g. ad libitum):All animals were provided conventional laboratory rodent diet (Nutrivet Life Sciences, Pune) ad libitum.
- Water (e.g. ad libitum):Aqua guard filtered tap water was provided ad libitum via drinking bottles.
- Acclimation period:All animals were acclimatized to the test conditions for 5 days prior to administration of the test item.
- Randomization : Animals were selected manually. No computer generated randomization program was used.

ENVIRONMENTAL CONDITIONS
- Temperature (°C):Minimum: 20.40 °C and Maximum: 23.10 °C
- Humidity (%):Minimum: 38.40% and Maximum: 58.70%
- Air changes (per hr):More than 12 changes per hour
- Photoperiod (hrs dark / hrs light):12:12

IN-LIFE DATES: From: February 05, 2014 To: February 21, 2014

Type of coverage:

occlusive

Vehicle:

other: distilled water

Details on dermal exposure:

TEST SITE
- Area of exposure:The test item was applied uniformly over clipped dorsal area of rat skin.
- % coverage:Approximately 10% body surface area of rat.
- Type of wrap if used:The porous gauze dressing and non-irritating tape.

REMOVAL OF TEST SUBSTANCE
- Washing (if done):The residual test item was removed by using distilled water.
- Time after start of exposure:24-hour.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit):A limit dose of 2000 mg/ kg body weight of test item was applied.
- Constant volume or concentration used: yes
- For solids, paste formed: yes

VEHICLE
- Amount(s) applied (volume or weight with unit):0.2 ml distilled water

Duration of exposure:

24 hrs

Doses:

2000 mg/kg body weight.

No. of animals per sex per dose:

10 (Five per sex)

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:Daily
- Necropsy of survivors performed: yes
At the end of 14 day observation period, all the surviving rats were euthanised by overdose of CO2 and subjected to gross pathology examination, for external and internal observations.
- Other examinations performed:
- Clinical signs : After test item administration, individual animals were frequently observed at 1, 2, 3 and 4 hours post dosing on day 0 (day of dosing). Subsequently, all animals were observed once a day during the 14 day observation period.
- Body weight: All rats were weighed on days 0 (prior to dosing), 7 and 14.
other:
- Local Signs/Skin Reactions
All animals were observed once daily during days 1-14 (in common with clinical signs).
- Mortality
Animals were observed twice daily for any mortality during the experimental period.

Statistics:

No statistical analysis was performed since the study was terminated with limit test.

Preliminary study:

not specified

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: no other details available

Mortality:

No mortality was observed at limit dose of 2000 mg/kg body weight of test item during the 14 day observation period.

Clinical signs:

other: In males, animal nos. 1 and 2 were observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 2 and mild scaling on day 3 to 6. Animal no. 3 was observed with vocalization at 1 hour, normal at 2

Gross pathology:

The external and internal gross pathological observation of all terminally sacrificed animals did not show any pathological abnormality, except animal nos. 6 to 9 were observed externally with scab formation.

Other findings:

not specified

Table 1: Individual Animal Body Weight (g) andBody Weight Changes(%)

 

Dose:2000 mg/ kg bodyweight                                                                                                         

Animal No.	Sex	Body Weight (gram)			Body Weight Change (%)
		Day 0	Day 7	Day 14	Day 0-7	Day 0-14
1	Male	217	213	215	-1.84	-0.92
2		238	221	238	-7.14	0.00
3		235	223	252	-5.11	7.23
4		248	255	296	2.82	19.35
5		251	237	271	-5.58	7.97
6	Female	238	248	255	4.20	7.14
7		242	238	241	-1.65	-0.41
8		236	232	226	-1.69	-4.24
9		227	237	228	4.41	0.44
10		232	210	196	-9.48	-15.52

Keys:* = Based on day 0 body weight taken prior to dose application.

Table 2: Individual Animal Clinical Signs and Symptoms

 

Dose:2000 mg/kg body weight

Animal No.	Sex	Hour(s) - Day 0				Day
		1	2	3	4	1	2	3	4	5	6	7
1	Male	172	1	1	1	65^++	65^+	147^+	147^+	147^+	147^+	1
2		172	1	1	1	65^++	65^+	147^+	147^+	147^+	147^+	1
3		172	1	1	1	65^++	65^+	65^+ 147^+	147^++	147^++	147^+	1
4		172	1	1	1	65^++	65^+	65^+ 147^+	65^+ 147^+	147^+	147^+	1
5		172	1	1	1	65^+	1	1	1	1	1	1
6	Female	172	1	1	1	65^++	65^++	65^++	65^++	65^++	65^++	65^++
7		172	1	1	1	65^+++	65^+++	65^+++	65^+++	65^+++	65^++	146
8		172	1	1	1	65^+++	65^+++	65^+++	65^+++	65^+++	65^++	65^++ 146
9		172	1	1	1	65^+++	65^+++	65^+++	65^+++	65^+++	65^++	146
10		172	1	1	1	65^+	65^+	147^+	147^+	147^+	147^+	1

 

Animal No.	Sex	Day
		8	9	10	11	12	13	14
1	Male	1	1	1	1	1	1	1
2		1	1	1	1	1	1	1
3		1	1	1	1	1	1	1
4		1	1	1	1	1	1	1
5		1	1	1	1	1	1	1
6	Female	146	146	146	146	146	146	146
7		146	146	146	146	146	146	146
8		65^+ 146	146	146	146	146	146	146
9		146	146	146	146	146	146	146
10		1	1	1	1	1	1	1

Keys: 1 = Normal, 65 = Erythema, 146 = Scab, 147 = Scale, 172 = Vocalization, + =Mild (slight),                  ++ = Moderate, +++ = Severe

Table 3: Individual Animal Mortality Record

 

Dose:2000 mg/kg body weight

Animal No.	Sex	Days of Observation (0 to 14)
		Morning Observations	Evening Observations
1	Male	No mortality and morbidity	No mortality and morbidity
2		No mortality and morbidity	No mortality and morbidity
3		No mortality and morbidity	No mortality and morbidity
4		No mortality and morbidity	No mortality and morbidity
5		No mortality and morbidity	No mortality and morbidity
6	Female	No mortality and morbidity	No mortality and morbidity
7		No mortality and morbidity	No mortality and morbidity
8		No mortality and morbidity	No mortality and morbidity
9		No mortality and morbidity	No mortality and morbidity
10		No mortality and morbidity	No mortality and morbidity

Interpretation of results:

other: Not classified

Conclusions:

The acute dermal median lethal dose of the test chemical was considered to be >2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that the given test chemical does not exhibit acute dermal toxicity i.e it is acutely non toxic to animals.

Executive summary:

Acute dermal toxicity study of given test chemical was performed as per OECD No.402 in rats. Five male and five female healthy young adult rats were randomly selected and used for conducting acute dermal toxicity study. Rats free from injury and irritation of skin were selected for the study. Twenty four hours prior to dermal application of test item, approximately 10% of body surface area of each rat was clipped. A limit dose of 2000 mg/ kg body weight of test item moistened with 0.2 ml distilled water was applied by single dermal application and observed for 14 days after treatment. On test day 0, an amount of test item moistened with 0.2 ml distilled water was applied directly on the intact skin of clipped area of rats; the porous gauze dressing was put on to the intact skin of clipped area. This porous gauze dressing was covered with a non-irritating tape. The dressing was wrapped around the abdomen and anchored with non-irritating adhesive tape.After the 24-hour application period, the dressings were removed and theskin was gently wiped with distilled water. The skin reactions were assessed. The animals were observed daily for mortality and clinical signs, during the acclimatization period. All animals were observed for clinical signs at approximately 1, 2, 3 and 4 hours after treatment on day 0 and once daily during test days 1‑14. Mortality was recorded after application on test day 0 and twice daily during days 1-14 (at least once on the day of sacrifice). Local signs / Skin reactions were observed daily from test days 1-14 (in common with clinical signs). Body weights were re­corded on day 0 (prior to application) and on day 7 and 14. All animals were necropsied and examined macroscopically. No mortality was observed in any animal till the end of the experimental period. In males, animal nos. 1 and 2 were observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 2 and mild scaling on day 3 to 6. Animal no. 3 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 3 and mild to moderate scaling on day 3 to 6. Animal no. 4 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 3 and mild scaling on day 3 to 6.Animal no. 5 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 2 to 14 and mild erythema on day 1.  In females, animal no. 6 was observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 7 and scab from day 8 to 14. Animal nos. 7 and 9 were observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 6 and scab from day 7 to 14. Animal no. 8 was observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 8 and scab from day 8 to 14. Animal no. 10 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, mild erythema on day 1 to 2 and mild scaling from day 3 to 6. In males, mean body weight gain was observed with on day 14, whereas decline in mean body weight gain was observed on day 7 as compared to day 0. In females, declined mean body weight gain was observed with on day 7 and 14 as compared to day 0. The external and internal gross pathological observation of all terminally sacrificed animals did not show any pathological abnormality, except animal nos. 6 to 9 were observed externally with scab formation. Under the conditions of this, the acute dermal median lethal dose of test chemical was considered to be >2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that the given test chemical does not exhibit acute dermal toxicity i.e it is acutely non toxic to animals.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

Data is Klimisch 1 and from experimental study report.

Additional information

Acute oral toxicity:

Acute oral toxicity study of test chemical was conducted as per OECD No. 423 in rats. Nine female Wistar rats were selected for acute oral toxicity study. The animals were fasted for minimum 16-18 hours prior to dosing and for 3-4 hours post dosing, with food withheld but drinking water provided ad libitum. The time intervals between dosing were determined by the onset, duration and severity of toxic signs. Three rats of first group were dosed with starting dose of 300 mg/kg body weight and the animals did not show any mortality so another three animals of the same group were dosed with 300 mg/kg body weight and no mortality was observed. Hence three rats of second group were dosed with 2000 mg/kg weight. All the rats at 2000 mg/kg were found dead on day 0 post dosing. Hence, further dosing was stopped. Body weights were re­corded on day 0 (prior to dosing), 7 and 14. Body weight gain was observed in all surviving animals treated with 300 mg/kg body weight, during the 14 day observation period, as compared to day 0. At 300 mg/kg, all the six animals were observed normal throughout the experiment period. At 2000 mg/kg, animal no. 7 was observed with mild tremors at 30 minutes, mild to moderate abdominal breathing at 30 minutes and 1 hour and sternal recumbency at 1 hour followed by found dead. Animal nos. 8 and 9 were observed with mild tremors, moderate abdominal breathing, sternal recumbency and moderate salivation at 30 minutes followed by found dead. No external gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, animal no. 7 was observed with no abnormalities, whereas animal nos. 8 and 9 were observed with wet around mouth. No internal gross pathological changes were seen in all the six animals treated with 300 mg/kg body weight during terminal sacrifice. At 2000 mg/kg, all three animals were observed with severe red discoloration of all lobes lungs and test item was observed in stomach and intestine. Under the conditions of this, all rats were died at 2000 mg/kg bw, hence the LD100 was considered to be 2000 mg/kg bw. Considering this value, the LD50 value can be assumed to be 1000 mg/kg bw. Therefore, comparing this value with the criteria of CLP regulation, the given test chemical can be classified in “Category 4 (300 – ≤ 2000)” for acute oral toxicity.

Acute Inhalation toxicity: 

The acute inhalation toxicity study need not be conducted because exposure to humans via inhalation route is not likely taking into account the low vapour pressure of the test substance, which is reported as 0.000000000796 mm Hg. Thus, exposure to inhalable dust, mist and vapour of the chemical is highly unlikely. Therefore this study is considered for waiver.

Acute Dermal toxicity:

Acute dermal toxicity study of the given test chemical was performed as per OECD No.402 in rats. Five male and five female healthy young adult rats were randomly selected and used for conducting acute dermal toxicity study. Rats free from injury and irritation of skin were selected for the study. Twenty four hours prior to dermal application of test item, approximately 10% of body surface area of each rat was clipped. A limit dose of 2000 mg/ kg body weight of test item moistened with 0.2 ml distilled water was applied by single dermal application and observed for 14 days after treatment. On test day 0, an amount of test item moistened with 0.2 ml distilled water was applied directly on the intact skin of clipped area of rats; the porous gauze dressing was put on to the intact skin of clipped area. This porous gauze dressing was covered with a non-irritating tape. The dressing was wrapped around the abdomen and anchored with non-irritating adhesive tape. After the 24-hour application period, the dressings were removed and the skin was gently wiped with distilled water. The skin reactions were assessed. The animals were observed daily for mortality and clinical signs, during the acclimatization period. All animals were observed for clinical signs at approximately 1, 2, 3 and 4 hours after treatment on day 0 and once daily during test days 1‑14. Mortality was recorded after application on test day 0 and twice daily during days 1-14 (at least once on the day of sacrifice). Local signs / Skinreactions were observed daily from test days 1-14 (in common with clinical signs). Body weights were re­corded on day 0 (prior to application) and on day 7 and 14. All animals were necropsied and examined macroscopically. No mortality was observed in any animal till the end of the experimental period. In males, animal nos. 1 and 2 were observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 2 and mild scaling on day 3 to 6. Animal no. 3 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 3 and mild to moderate scaling on day 3 to 6. Animal no. 4 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, moderate to mild erythema on day 1 to 3 and mild scaling on day 3 to 6.Animal no. 5 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 2 to 14 and mild erythema on day 1.  In females, animal no. 6 was observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 7 and scab from day 8 to 14. Animal nos. 7 and 9 were observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 6 and scab from day 7 to 14. Animal no. 8 was observed with vocalization at 1 hour, normal at 2, 3 and 4 hours, severe to moderate erythema on day 1 to 8 and scab from day 8 to 14. Animal no. 10 was observed with vocalization at 1 hour, normal at 2, 3, 4 hours and on day 7 to 14, mild erythema on day 1 to 2 and mild scaling from day 3 to 6. In males, mean body weight gain was observed with on day 14, whereas decline in mean body weight gain was observed on day 7 as compared to day 0. In females, declined mean body weight gain was observed with on day 7 and 14 as compared to day 0. The external and internal gross pathological observation of all terminally sacrificed animals did not show any pathological abnormality, except animal nos. 6 to 9 were observed externally with scab formation. Under the conditions of this, the acute dermal median lethal dose of the given test chemical was considered to be >2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that the given test chemical does not exhibit acute dermal toxicity i.e. it is acutely non toxic to animals.

Justification for classification or non-classification

Based on the above experimental studies on test chemical, it can be concluded that LD50 value is between 300-2000 mg/kg bw, for acute oral toxicity and LD50 value is >2000 mg/kg bw for acute dermal toxicity. Thus, comparing these values with the criteria of CLP regulation, the given test chemical can be classified in “Category 4” for acute oral toxicity and cannot be classified for acute dermal toxicity.For acute inhalation toxicity wavier were added so, not possible to classify.