Registration Dossier

Administrative data

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 report.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Principles of method if other than guideline:
Acute dermal toxicity study of the given test chemical was performed in rats.
GLP compliance:
yes
Test type:
other: not specified
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
SOURCE OF TEST MATERIAL
- Identity: Reaction mass of 4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one and -(2,6,6-trimethylcyclohex-1-ene-1-yl)-but-3-ene-2-one 100%
- EC no: 907-706-6
- Lot/batch No.of test material: 9000533813
- Expiration date of the lot/batch: April 14, 2004
- Purity: 85.1% (sum of 2 isomers)
- Colour: colourless to pale yellow

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:On the day of the experiment, the test item was dissolved in ethanol (purity > 99 %,MERCK, D-64293 Darmstadt).

Specific details on test material used for the study:
- Name of test material: Reaction mass of 4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one and -(2,6,6-trimethylcyclohex-1-ene-1-yl)-but-3-ene-2-one
- Molecular formula: C13H20O
- Molecular weight: 192.3 g/mole
- Smiles : C1([C@@H](C(=CCC1)C)\C=C\C(C)=O)(C)C
- Inchl: 1S/C13H20O/c1-10-6-5-9-13(3,4)12(10)8-7-11(2)14/h6-8,12H,5,9H2,1-4H3/b8-7+
- Substance type: Organic
- Physical state: Liquid

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: In-house animals, bred at Animal House, sa-FORD.CPCSEA Registration No. 1256/bc/09/CPCSEA.
- Females (if applicable) nulliparous and non-pregnant: Yes
- Weight at study initiation: Male:Minimum: 229 g and Maximum: 262 g
(Prior to Treatment) Female:Minimum: 203 g and Maximum: 241 g
- Fasting period before study: The food was withheld prior to dosing and 3-4 hours post dosing but drinking water was provided ad libitum.
- Housing: The animals were housed individually in polycarbonate cages.All cages were provided with corn cobs (Sparconn Life Sciences,Bangalore) SPAR – 24/ 2013.
- Diet (e.g. ad libitum): All animals were provided conventional laboratory rodent diet (Nutrivet Life Sciences, Pune) ad libitum. Batch No.: 400012.
- 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.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Room temperature was maintained at 21.10 to 24.40 degree centigrade.
- Humidity (%): Room humidity was maintained at 36.60% to 57.30%.
- Air changes (per hr): More than 12 changes per hour
- Photoperiod (hrs dark / hrs light): An artificial light and dark cycle of 12 hours each was provided to the room.

Administration / exposure

Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: over clipped dorsal area of rat skin. Approx. 10% of body surface area of rat
- % coverage: porous gauze dressing
- Type of wrap if used: The test site was further covered in a suitable manner to retain the gauze dressing

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, At the end of the exposure period, residual test item was removed by using distilled water.
- Time after start of exposure:24hr

TEST MATERIAL
- Concentration (if solution): 2000mg/kgbw
- Constant volume or concentration used: yes
Duration of exposure:
24hr
Doses:
2000mg/kg bw
No. of animals per sex per dose:
Total : 10
male : 5
female: 5
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
- 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.
Local Signs/Skin Reactions: All animals were observed once daily during days 1-14 (in common with clinical signs).
body weight:All rats were weighed on days 0 (prior to dosing), 7 and 14.
Mortality:Animals were observed twice daily for any mortality during the experimental period
Pathology: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.
Statistics:
No statistical analysis was performed since the study was terminated with limit test

Results and discussion

Preliminary study:
not specified
Effect levels
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: No mortality was observed
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:
No systemic or local signs of toxicity were observed at limit dose of 2000 mg/kg body weight of test item during the experimental period
Body weight:
The male and female animals were observed with body weight gain compared to day 0 throughout the experiment
Gross pathology:
The external and internal gross pathological observation of all terminally sacrificed animals did not show any pathological abnormality
Other findings:
not specified

Any other information on results incl. tables

Individual Animal Body Weight (g) andBody Weight Changes(%)

 

Dose:2000 mg/ kg bodyweight                                                                             

 Density:0.9407

Animal No.

Sex

Dose (ml) Applied*

Body Weight (gram)

Body Weight Change (%)

Day 0

Day 7

Day 14

Day 0-7

Day 0-14

1

Male

0.49

229

247

256

7.86

11.79

2

0.50

235

256

278

8.94

18.30

3

0.49

230

245

272

6.52

18.26

4

0.53

247

262

296

6.07

19.84

5

0.56

262

289

310

10.31

18.32

6

Female

0.51

241

240

258

-0.41

7.05

7

0.43

203

205

211

0.99

3.94

8

0.43

203

206

213

1.48

4.93

9

0.50

236

241

245

2.12

3.81

10

0.48

226

231

232

2.21

2.65

Key:* = Based on density of test item and day 0 body weight taken prior to dose application.

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

8

9

10

11

12

13

14

1

Male

1

1

1

1

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

1

1

1

1

3

1

1

1

1

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

1

1

1

1

5

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

6

Female

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

7

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

8

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

9

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

10

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

Key: 1 = Normal

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

Summaryof Animal Body Weight (g) and Body Weight Changes (%)

 

Dose:2000 mg/kg body weight

Sex

Body Weight (gram)

Body Weight Changes (%)

Day 0

Day 7

Day 14

Day 0-7

Day 0-14

Male

Mean

231.20

242.20

257.10

7.94

17.30

SD

18.12

25.01

33.03

1.74

3.15

n

10

10

10

5

5

Female

Mean

221.80

224.60

231.80

1.28

4.48

SD

17.99

17.87

20.29

1.07

1.65

n

5

5

5

5

5

Keys:SD= Standard deviation, n = Number of animals

Gross Necropsy Observation

 

 Dose:2000 mg/kg body weight                                               

Mode of Death:Terminal Sacrifice


Animal No.

Sex

Gross Observation

External

Internal

1

Male

No abnormalities detected

No abnormalities detected

2

No abnormalities detected

No abnormalities detected

3

No abnormalities detected

No abnormalities detected

4

No abnormalities detected

No abnormalities detected

5

No abnormalities detected

No abnormalities detected

6

Female

No abnormalities detected

No abnormalities detected

7

No abnormalities detected

No abnormalities detected

8

No abnormalities detected

No abnormalities detected

9

No abnormalities detected

No abnormalities detected

10

No abnormalities detected

No abnormalities detected


Applicant's summary and conclusion

Interpretation of results:
other: Not classified
Conclusions:
The LD50 value was considered to be >2000 mg/kg body weight, when female wistar rats were treated with the given test chemical by dermal application.
Executive summary:

Acute dermal toxicity study of the given test chemical was performed as per OECD No.402 in Wistar Rats. Five male and five female healthy young adult rats were randomly selected. 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 was applied by single dermal application and observed for 14 days after treatment. On test day 0, as such amount of test item, calculated based on density (0.9407) and body weight was applied directly on the intact skin of clipped area of rats; the surgical gauze patch was put on to the intact skin of clipped area. This gauze patch was covered with a semi-occlusive dressing. 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 / 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. No clinical signs and any skin reaction were observed throughout the experimental period in all treated animals. The male and female animals were observed with body weight gain compared to day 0 throughout the experiment. The external and internal gross pathological observation of all terminally sacrificed animals did not show any pathological abnormality. Hence the LD50 value was considered to be >2000 mg/kg body weight, when female wistar rats were treated with the given test chemical by dermal application.