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Acute Toxicity: dermal

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Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 01 MAR 2012 to 04 MAY 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (OECD 402) and according to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
4-[(2,5-dichlorophenyl)azo]-N-(2,3-dihydro-2-oxo-1H-benzimidazol-5-yl)-3-hydroxynaphthalene-2-carboxamide
EC Number:
230-258-1
EC Name:
4-[(2,5-dichlorophenyl)azo]-N-(2,3-dihydro-2-oxo-1H-benzimidazol-5-yl)-3-hydroxynaphthalene-2-carboxamide
Cas Number:
6992-11-6
Molecular formula:
C24H15Cl2N5O3
IUPAC Name:
4-[(2,5-dichlorophenyl)diazenyl]-3-hydroxy-N-(2-oxo-2,3-dihydro-1H-benzimidazol-5-yl)-2-naphthamide
Test material form:
solid: nanoform, no surface treatment

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann, 33178 Borchen, Germany
- Age at study initiation: males: 9 to 10 weeks; females 12 to 13 weeks
- Weight at study initiation: males: 229 to 241 g; females: 211-224 g
- Housing: The animals were kept individually in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding (lot no. 110811)
- Diet: Altromin 1324 maintenance diet for rats and mice (lot no. 1114), ad libitum
- Water: tap water, sulphur acidified to a pH value of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals), ad libitum
- Acclimation period: Adequate acclimatisation period (at least five days) under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 55 +/- 10
- Air changes (per hr): 10 x / hour
- Photoperiod: 12 hours light, 12 hours dark
- Full barrier in an air-conditioned room
- Certificates of food, water and bedding are filed at BSL BIOSERVICE

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
other: cottonseed oil
Details on dermal exposure:
Preparation of the Animals:
The animals were marked for individual identification by tail painting.
Approximately 24 hours before the test, the fur was removed from the dorsal area of the trunk using an electric clipper.
Care was taken to avoid abrading the skin, and only animals with healthy intact skin were used.
No less than 10% of the body surface was cleared for the application.
Prior to the application a detailed clinical observation was made of all animals.

TEST SITE
The test item was applied at a single dose, uniformly over an area which was approximately 10% of the total body surface.
The test item was held in contact with the skin by a dressing throughout a 24-hour period.
- Type of wrap if used: The dressing consisted of a gauze-dressing and non-irritating tape and was fixed with an additional dressing in a suitable manner

REMOVAL OF TEST SUBSTANCE
- Washing (if done): residual test item was removed using the vehicle cottonseed oil
- Time after start of exposure: 24 hours (i.e. at the end of the exposure period)

VEHICLE
- Justification: The vehicle was chosen due to its minimal potential influence on irritation of the skin.
- Lot/batch no. (if required): MKBG0088V (Sigma Aldrich, expiry date: 2012-05)
Duration of exposure:
24 hours period
Doses:
2000 mg/kg body weight
No. of animals per sex per dose:
5
Control animals:
not required
Details on study design:
Observation period:
All animals were observed for 14 days after dosing

Weight Assessment:
The animals were weighed on day 1 (prior to the application) and on days 8 and 15.

Clinical Examination:
careful clinical examination was made several times on the day of dosing (at least once during the first 30 minutes
and with special attention given during the first 4 hours post-dose). As soon as symptoms were noticed they were recorded.
Thereafter, the animals were observed for clinical signs once daily until the end of the observation period. All abnormalities were recorded.
Cageside observations included changes in the skin and fur, eyes and mucous membranes. Also respiratory,
circulatory, autonomic and central nervous systems and somatomotor activity and behaviour pattern were examined.
Attention was directed to observations of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

Pathology:
At the end of the observation period the surviving animals were sacrificed with an overdosage of pentobarbital injected
intraperitoneally (Narcoren®, Merial) at the dosage of approximately 8 mL/kg bw. All animals were subjected to gross necropsy.
All gross pathological changes were recorded and in case of findings the tissues were preserved for a possible histopathological evaluation.
The preserved tissues of which no histopathological evaluation was made will be discarded 3 months after the release of the final report
unless otherwise agreed upon with the sponsor.

Evaluation of Results:
Individual reactions of each animal were recorded at each time of observation.
Toxic response data were recorded by sex and dose level.
Nature, severity and duration of clinical observations were described.
The body weight changes were summarised in a tabular form.
Necropsy findings were described.
Statistics:
According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results,
a statistical evaluation of the results is not regarded as necessary.

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: no animal died during the 14-day observation period
Mortality:
No mortality was observed
Clinical signs:
No treatment-related effects were observed.






Body weight:
A slight weight loss was recorded for 2 out of 5 female animals during the first week, but all of the female animals showed
weight gain during the second week. The effects on weight development might be secondary to the dressing, and toxicological
relevance of this finding cannot clearly be concluded.
The male animals showed weight gain during the first and the second week of the observation.
Gross pathology:
No treatment-related effects were observed.
Other findings:
No erythema or oedema was observed.

Any other information on results incl. tables

Table Absolute Body Weights in g and Body Weight Gain in %:

Dose: 2000 mg/kg body weight

Animal No. / Sex

g
Day 1

g
Day 8

g
Day 15

%
Day 1-15

21 / male

233

257

279

20

22 / male

231

251

281

22

23 / male

234

240

262

12

24 / male

241

261

285

18

25 / male

229

237

289

26

26 / female

211

212

221

5

27 / female

217

222

223

3

28 / female

221

219

225

2

29 / female

219

225

232

6

30 / female

224

223

228

2

Table LD50:

Dose (Unit)

 

Number of Animals Investigated

Number of Intercurrent Deaths

LD50

2000 mg/kg bw

5 males

0

> 2000 mg/kg bw

2000mg/kg bw

5 females

0

> 2000 mg/kg bw

bw = body weight

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of the present study, single dermal application of the test item to rats at a dose of 2000 mg/kg body weight was associated with no mortality and neither signs of toxicity nor signs of irritation. The dermal LD50 was determined to be > 2000 mg/ kg body weight.
Executive summary:

To test acute dermal toxicity 5 male and 5 female Wistar rats were subjected to a test according to OECD test guideline 402. The test material was applied once in cottonseed oil at a dose of 2000 mg/kg bw for 24 hours under semi-occlusive conditions to the skin of rats. During the 14 -day observation period none of the tested animals died, showed specific signs of systemic toxicity or revealed signs of dermal irritation. No findings were made at the macroscopic examination performed at the end of the observation period. The rather low weight gain (2 -6 % during the observation period) was not accounted for as adverse as there were no effects seen at the clinical and pathological examinations. Study authors mentioned that they might be secondary to the dressing. Under the conditions of the present study the dermal LD50 was determined to be > 2000 mg/kg body weight.

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