12H-phthaloperin-12-one

Acute Toxicity: dermal

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 study report.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Principles of method if other than guideline:

The objective of this acute dermal toxicity study was to assess the toxicological profile of the test item on application as a single semi-occlusive dermal application to rats.

GLP compliance:

yes

Test type:

fixed dose procedure

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

12H-phthaloperin-12-one

EC Number:

230-049-5

EC Name:

12H-phthaloperin-12-one

Cas Number:

6925-69-5

Molecular formula:

C18H10N2O

IUPAC Name:

12H-phthaloperin-12-one

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material : 12H-phthaloperin-12-one
- Common name : C.I. Solvent Orange 60
- Molecular formula : C18H10N2O
- Molecular weight : 270.29 g/mol
- Smiles notation : O=C1N2c3c4c(cccc4N=C2c2ccccc12)ccc3
- InChl : 1S/C18H10N2O/c21-18-13-8-2-1-7-12(13)17-19-14-9-3-5-11-6-4-10-15(16(11)14)20(17)18/h1-10H
- Substance type : Organic
- Physical state : Solid

Specific details on test material used for the study:

- Purity as per Certificate of Analysis:97%
- Lot No.:20120720
- Manufactured date:DEC-2013
- pH:4.69 at 26.8°C
- Density:1.176 g/cm3 at 27°C
- Storage conditions:Ambient (+15 to +25°C)
- SAFETY PRECAUTIONS: Gloves, cap and face mask were used in addition to protective body garments and shoes, to ensure adequate personal health and safety and to avoid inhalation and skin contact with the test item.

Test animals

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Geniron Biolabs Pvt. Ltd. Bengaluru
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 10 to 12 weeks
- Weight at study initiation: Females: 235.18 to 238.33 g
- Identification:By rat accession number. Identification of individual rats is by cage card and crystal violet colour body markings. The temporary body marking during acclimatization period was done with crystal violet. The rat accession numbers were allotted during the course of the study and was included in raw data and reported.
- Housing: Animals were housed individually in standard polysulfone cages (Size: L 425 x B 266 x H 185 mm), with stainless steel top grill. Additionally, polycarbonate rat huts were placed inside the cage as enrichment objects and were changed along with the cage once a week. Bedding: Steam sterilized corn cob was used and changed once a week along with the cage.
- Diet (e.g. ad libitum): Hypro Rat & Mice pellet feed, ad libitum
- Water (e.g. ad libitum): Deep bore-well water passed through activated charcoal filter and exposed to UV rays in Aquaguard on-line water filter-cum-purifier, ad libitum
- Acclimation period: The rats were acclimatized for six, eight, twelve and fourteen days before treatment for dose range finding and main study respectively under standard laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 to 24°C
- Humidity (%): 65 to 67%
- Air changes (per hr): air conditioned with adequate fresh air supply (12.4 air changes/hour)
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark cycle

IN-LIFE DATES: From: 03 July 2018 To: 09 July 2018

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: clipped skin of dorsolateral thoracic surface
- % coverage: 10% of the body surface
- Type of wrap if used: The applied area was covered with cotton gauze (size: Females: 8 x 5 cm of 6 ply) and it was secured in position by adhesive tape wound around the torso.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The dressing was removed and the applied area was washed with deionized water and wiped dry using clean towel.
- Time after start of exposure:24 hours

-Preparation of test site for application
Approximately 24 hours before treatment, the hair on the dorsolateral thoracic surface of the skin was clipped (approximately 10 x 8 cm) with an electric clipper Care was taken to avoid abrading the skin, which could alter its permeability

Duration of exposure:

24 hours

Doses:

DRF G1 - 200 mg/kg
DRF G2 - 1000 mg/kg
DRF G3 - 2000 mg/kg
Main G3 - 2000 mg/kg

No. of animals per sex per dose:

DRF G1 - 200 mg/kg - 1
DRF G2 - 1000 mg/kg - 1
DRF G3 - 2000 mg/kg - 1
Main G3 - 2000 mg/kg - 2

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical examination and pre-terminal deaths - The animals were observed for clinical signs and pre-terminal deaths (mortality) once during first 30 minutes after application, and at hourly intervals for 6 hours after application on the day of treatment (day 1) and once daily during Days 2 to 15. In addition, the treatment site was observed for skin reactions at 24, 48 and 72 hours after removal of test chemical using the Draize criteria. All rats were observed for changes in skin and fur, eyes and mucous membranes, and also respiratory, circulatory, autonomic and central nervous systems and somatomotor activity and behaviour pattern. Attention was directed to observations of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
Body weights - Individual body weights of animals were recorded on test days 1(Pre-application), 8 (7 days post application), and 15 (14 days post application).
- Necropsy of survivors performed: yes, at the end of the observation period, all rats were euthanised and exsanguinated under isoflurane anesthesia and subjected to detailed necropsy by an experienced prosector and the findings were recorded.
- Other examinations performed: Microscopic examination was not carried out as no gross pathological changes were observed.

Statistics:

not specified

Results and discussion

Preliminary study:

not specified

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: no other details available

Mortality:

There were no pre-terminal deaths (mortality) observed during the study.

Clinical signs:

other: There were no clinical signs observed during the study. There were no skin reactions at the site of application at 24 and 48 hours after test patch removal (as per draize method).

Gross pathology:

No abnormality was detected at necropsy.

Other findings:

not specified

Any other information on results incl. tables

TABLE 1.            Individual body weight, body weight changes and pre-terminal deaths

Group and Dose (mg/kg body weight)	Rat No.	S e x	Body weight (g)					Pre-terminal deaths
			Initial (Day 1 - at treatment)	8^th   day	Weight change (day 8 – Initial)	15^th day	Weight change (day 15 – Initial)
G1 and 200 DRF	Rw283	F	235.18	242.68	7.50	248.19	13.01	0
G2 and 1000 DRF	Rw284	F	238.33	245.45	7.12	251.67	13.34	0
G3 and 2000 DRF	Rw285	F	238.16	244.73	6.57	249.14	10.98	0
G3 and 2000 Main study	Rw286	F	230.34	236.59	6.25	241.08	10.74	0
	Rw287	F	234.62	242.93	8.31	247.63	13.01	0

 DRF: Dose Range Finding   F: Female

APPENDIX 1.      Individual test item application, clinical signs, skin reactionand necropsy findings

Dose range finding study

 

Group & Dose (mg/kg body weight)

Date and time of application

Rat Number

S e x

Initial Bwt (g)

Quantity (mL) applied

Observations and skin reaction

Days

1

2

3

4

5

30 min

1 h #

2 h #

3 h #

4 h #

5 h #

6 h #

*

Er @

Ed @

*

Er @

Ed @

*

Er @

Ed @

G1 and 200 DRF

11 April 2018 and 10.55 AM

Rm8907

F

223.58

0.04

N

N

N

N

N

N

N

N

N

0

0

N

0

0

N

0

0

 

Group & Dose (mg/kg body weight)

Rat Number

S e x

Observation

Necropsy findings

Days

6

7

8

9

10

11

12

13

14

15

G1 and 200 DRF

Rm8907

F

N

N

N

N

N

N

N

N

N

N

NAD

F: Female             N: Normal          h: hour    min: minutes                      NAD: No abnormality detected      Er: Erythema                      Ed: Edema  

Score 0: No Erythema / Edema       

    

*: Clinical signs; @: Skin scoring as per Draize method (approximately 24, 48 and 72 hours) after test patch removal

 

 

APPENDIX 2 contd. Individual test item application, clinical signs, skin reaction and necropsy findings

 

Dose range finding study

Group & Dose (mg/kg body weight)

Date and time of application

Rat Number

S e x

Initial Bwt (g)

Quantity (mg) applied

Observations and skin reaction

Days

1

2

3

4

30  min

1 h

2 h

3 h

4 h

5 h

6 h

*

Er @

Ed @

*

Er @

Ed@

G1- DRF  and 200

03 July 2018 and 11.36 AM

Rw283

F

235.18

47

N

N

N

N

N

N

N

N

N

0

0

N

0

0

Group & Dose (mg/kg body weight)

Animal Number

S e x

Observation/ Days

Necropsy findings

5

*

Er @

Ed @

6

7

8

9

10

11

12

13

14

15

G1- DRF  and 200

Rw283

F

N

0

0

N

N

N

N

N

N

N

N

N

N

NAD

 

F: Female             N: Normal          h: hour    min: minutes                       NAD: No abnormality detected      Er: Erythema                       Ed: Edema  

Score 0: No Erythema / Edema       

    

*: Clinical signs; @: Skin scoring as per Draize method (approximately 24, 48 and 72 hours) after test patch removal

APPENDIX 2 contd. Individual test item application, clinical signs, skin reaction and necropsy findings

 

Dose range finding study

 

 

Group & Dose (mg/kg body weight)

Date and time of application

Rat Number

S e x

Initial Bwt (g)

Quantity (mg) applied

Observations and skin reaction

Days

1

2

3

4

30  min

1 h

2 h

3 h

4 h

5 h

6 h

*

Er @

Ed @

*

Er @

Ed @

G2- DRF  and 1000

05 July 2018 and 11.56 AM

Rw284

F

238.33

238

N

N

N

N

N

N

N

N

N

0

0

N

0

0

Group & Dose (mg/kg body weight)

Animal Number

S e x

Observation/ Days

Necropsy findings

5

*

Er @

Ed @

6

7

8

9

10

11

12

13

14

15

G2- DRF  and 1000

Rw284

F

N

0

0

N

N

N

N

N

N

N

N

N

N

NAD

F: Female             N: Normal          h: hour    min: minutes                       NAD: No abnormality detected      Er: Erythema                       Ed: Edema  

Score 0: No Erythema / Edema       

    

*: Clinical signs; @: Skin scoring as per Draize method (approximately 24, 48 and 72 hours) after test patch removal

 

 

APPENDIX 2 contd. Individual test item application, clinical signs, skin reaction and necropsy findings

 

Main study

 

Group & Dose (mg/kg body weight)

Date and time of application

Rat Number

S e x

Initial Bwt (g)

Quantity (mg) applied

Observations and skin reaction

Days

1

2

3

4

30 min

1 h

2 h

3 h

4 h

5 h

6 h

*

Er @

Ed @

*

Er @

Ed @

G3- Main and 2000

12 July 2018 and 11.41 AM to 11.42 AM

Rw286

F

230.34

461

N

N

N

N

N

N

N

N

N

0

0

N

0

0

Rw287

F

234.62

469

N

N

N

N

N

N

N

N

N

0

0

N

0

0

 

Group & Dose (mg/kg body weight)

Animal Number

S e x

Observation/ Days

Necropsy findings

5

*

Er @

Ed @

6

7

8

9

10

11

12

13

14

15

G3- Main  and 2000

Rw286

F

N

0

0

N

N

N

N

N

N

N

N

N

N

NAD

Rw287

F

N

0

0

N

N

N

N

N

N

N

N

N

N

F: Female             N: Normal          h: hour    min: minutes                       NAD: No abnormality detected      Er: Erythema                       Ed: Edema  

Score 0: No Erythema / Edema          

*: Clinical signs; @: Skin scoring as per Draize method (approximately 24, 48 and 72 hours) after test patch removal

 

Applicant's summary and conclusion

Interpretation of results:

other: Not classified

Conclusions:

Acute dermal toxicity study was conducted in order assess the toxicological profile of the test item on wistar rats. Based on the results, The LD50 of test chemical was determined to be >2000 mg/kg ,Thus, comparing this value with the criteria of CLP regulation, the given test chemical cannot be classified for acute dermal toxicity.

Executive summary:

The acute dermal toxicity of test chemical in Wistar rats was tested with 200 (G1-DRF), 1000 (G2-DRF) and 2000 (G3-DRF) mg/kg with 1 female each for the dose range finding study and 2 female for main study (G3).Based on the individual body weight, the test item at the dose of 200, 1000 and 2000 mg/kg body weight was weighed on an aluminum foil and made as a paste in Milli-Q water and applied directly to the clipped skin of the animal to cover about 10% of the body surface of the animal (semi-occlusive). The area of application was covered with cotton gauze (size: Females: 8 x 5 cm of 6 ply) and it was secured in position by adhesive tape wound around the torso. The test item contact period with the skin was for 24 hours.After the 24 hours contact period, the dressing was removed and the applied area was washed with deionized water and wiped dry using clean towels. All the rats were observed for clinical signs of toxicity and mortality for 14 days post application. In addition, the treatment site was observed at 24, 48 and 72 hours after removal of test item using the Draize criteria. There were no clinical signs of toxicity and mortality. There was no skin reaction observed at the site of application of test item. Body weights were measured on Days 1, 8 and 15 and growth was unaffected by test item. At the end of observation period (Day 15), all animals were euthanized and subjected to necropsy, and there were no gross lesions noted at the necropsy.Based on the present study results, the acute dermal LD50of test chemical is greater than 2000 mg/kg body weight in female Wistar rats.Thus, comparing this value with the criteria of CLP regulation, the given test chemical cannot be classified for acute dermal toxicity.