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Description of key information

Skin corrosion in vitro (3D-skin model with EST-1000, GLP, OECD TG 431 guideline draft, EU Method B.40): not corrosive [Bayer AG, Report No. PH-34412, 2006-04-20]



Skin irritation in vivo (Rabbit-NZW, GLP, OECD TG 404, EU Method B.4): not irritating to the skin [Bayer AG, Report No. PH-34402, 2006-04-10]



Eye irritation in vivo (Rabbit-NZW, GLP, OECD TG 405, EU Method B.5): not irritating to the eye [Bayer AG, Report No. PH-34401, 2006-04-10]


 

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Jan 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
other: OECD 431 guideline draft
Version / remarks:
1996/ 1998
Qualifier:
according to guideline
Guideline:
EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
purity 99.8%
Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
Corrosion of the skin is defined as an irreversible necrotic alteration of the tissue induced by a chemical. While the testing for corrosives usually involves the rabbit skin as a predictive in vivo test method, substances may be classified according to their corrosive potential by the determination of their cytotoxic effects on an in vitro reconstructed human epidermis. The test principle is based on the MTT assay reflecting the cell viability after short term exposure of the epidermal equivalent to topically applied Oxaphthalimid.
Vehicle:
physiological saline
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: The experiment was carried out on a reconstructed human epidermis EST-1000 (CellSystems, St. Katharinen, Germany).

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: RT: 3 min; 37 ± 2°C: 60 min
- Temperature of post-treatment incubation (if applicable):

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: 3 washing steps, volume not reported
- Observable damage in the tissue due to washing: No

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 2 h
- Spectrophotometer: (EL808, Bio-Tek; 96 well format, 200 µL)
- Wavelength: 570 nm


NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues / killed tissues: not reported
- Procedure used to prepare the killed tissues (if applicable): not reported
- N. of replicates : not reported
- Method of calculation used: not reported

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 2

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.
- Justification for the selection of the cut-off point(s) if different than recommended in TG 431 and 439: None
Control samples:
yes, concurrent negative control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg


NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
Duration of treatment / exposure:
3 min and 60 min
Number of replicates:
3
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1/ 60 min
Value:
95.27
Vehicle controls validity:
valid
Positive controls validity:
not examined
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1 / 3 min
Value:
97.26
Vehicle controls validity:
valid
Positive controls validity:
not examined

The MTT method has determined the following values of viability:

after 3 min of incubation: 97.36%

after 60 min of incubation: 95.27%

Interpretation of results:
GHS criteria not met
Conclusions:
the test item is not corrosive to skin
Executive summary:

The study on a reconstructed human epidermis EST-1000 was carried out for detection of topically applied skin corrosives with the test item Oxaphthalimid.

A 100% concentration was tested on the skin/epidermal equivalents in triplets. For the determination of time related cytotoxic effects the incubation periods were 3 minutes (room temperature) and 60 minutes (incubator), respectively. Using the MTT (methylthiazoletetrazolium) method the cell viability after 3 minutes or after 60 minutes of incubation was determined to be 97.26% and 95.27%, respectively. Thus, Oxaphthalimid was not characterised by a significant impact on cell viability and it therefore showed no corrosive properties.

Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Mar 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Version / remarks:
2002
Qualifier:
according to guideline
Guideline:
EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
purity 99.8%
Species:
rabbit
Strain:
New Zealand White
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, 88353 Kißlegg, Germany, 69592
- Age at study initiation: young adult animals
- Weight at study initiation: 2.6 kg — 2.8 kg
- Housing: The animals were housed individually in cage units Metall/Noryl by EBECO. Excrement trays below the cages contained low dust wood granulate bedding (J. Rettenmaier & Söhne, 73494 Rosenberg, Germany).
- Diet (e.g. ad libitum): The animals received the standard diet “SsniffK-Z” 4mm (manufacturer: Ssniff Spezialdiäten GmbH, 59494 Soest, Germany), approximately 100 g per animal per day
- Water (e.g. ad libitum): Tap water ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20±3
- Humidity (%): 50±25
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:
semiocclusive
Preparation of test site:
shaved
Vehicle:
water
Remarks:
pulverized test item moistened with water
Controls:
yes, concurrent no treatment
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit):0.5 g
Duration of treatment / exposure:
one animal: first patch removed after 3 minutes, second patch (because no serious skin reactions were observed after the first patch) removed after 1 hour and third patch removed after 4 hours
two more animals: exposed for 4 hours
Observation period:
72 hours
Number of animals:
3 females
Details on study design:
TEST SITE
- Area of exposure: 2.5 x 2.5 cm
- Type of wrap if used: 0.5 g of the pulverized test substance moistened with Aqua p.i. (to ensure good contact with the skin) was applied to the skin of the animal under a gauze patch. The patch was placed on the dorso-lateral areas of the trunk of each animal and was held in place with non-irritating tape for the duration of the exposure period.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The exposed skin area was carefully washed with water without altering the existing response, or the integrity of the epidermis.
- Time after start of exposure: 4 h

OBSERVATION TIME POINTS
(indicate if minutes, hours or days): 1, 24, 48, 72 h

SCORING SYSTEM: The degree of erythema/eschar formation and oedema formation was recorded as specified by DRAIZE
Irritation parameter:
erythema score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Irritation parameter:
edema score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Other effects:
There were no systemic intolerance reactions.

Results of the study:

  Irritant Effects (Scores)
  24 h   48 h      72 h         Mean scores
 animal   erythema   edema   erythema   edema   erythema   edema   erythema  edema 
 1
 2 0 0
 3 0
Interpretation of results:
GHS criteria not met
Conclusions:
the test item is not irritating to skin
Executive summary:

500 mg of the test substance was semi-occlusively applied to the shaved intact skin of 3 female rabbits for an exposure period of 4 h. Evaluation of skin irritation was made according to Draize.

In this skin irritation study Oxaphthalimid was not irritating to the skin.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Mar 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
2002
Qualifier:
according to guideline
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
purity 99.8%
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: Charles River, 88353 Kißlegg, Germany, 69592
- Age at study initiation: young adult animals
- Weight at study initiation: 2.5 kg — 2.8 kg
- Housing: The animals were housed individually in cage units Metall/Noryl by EBECO. Excrement trays below the cages contained low dust wood granulate bedding (J. Rettenmaier & Söhne, 73494 Rosenberg, Germany).
- Diet (e.g. ad libitum): The animals received the standard diet “SsniffK-Z” 4mm (manufacturer: Ssniff Spezialdiäten GmbH, 59494 Soest, Germany), approximately 100 g per animal per day
- Water (e.g. ad libitum): Tap water ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20±3
- Humidity (%): 50±25
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0,1 g
Duration of treatment / exposure:
24 hours
Observation period (in vivo):
72 hours
Number of animals or in vitro replicates:
3 females
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Time after start of exposure: 24h

SCORING SYSTEM: According to DRAIZE

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein: not reported
Irritation parameter:
cornea opacity score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Irritation parameter:
iris score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
2
Irritation parameter:
conjunctivae score
Basis:
animal #1
Time point:
24/48/72 h
Score:
0.3
Max. score:
3
Reversibility:
fully reversible within: 2 days
Irritation parameter:
conjunctivae score
Basis:
animal #2
Time point:
24/48/72 h
Score:
0.3
Max. score:
3
Reversibility:
fully reversible within: 2 days
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
24/48/72 h
Score:
0
Max. score:
3
Irritation parameter:
chemosis score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Other effects:
There were no systemic intolerance reactions.

Results of the study

      Irritant Effects (Score)         
 Animal    24 h 48 h   72 h  Mean scores
 1  Corneal Opacity  0 0.0
   Iritis  0  0  0  0.0
   Redness conjunctivae  1  0  0  0.3
   Chemosis conjunctivae  0  0  0  0.0
 2   Corneal Opacity  0  0  0  0.0
    Iritis  0  0  0  0.0
    Redness conjunctivae  1  0  0  0.3
    Chemosis conjunctivae  0  0  0  0.0
 3   Corneal Opacity  0  0  0  0.0
    Iritis  0  0  0  0.0
    Redness conjunctivae  0  0  0  0.0
    Chemosis conjunctivae  0  0  0  0.0

All animals showed reddening of the conjunctivae 1 h after administration.

Interpretation of results:
GHS criteria not met
Conclusions:
the test item is not an irritant to eyes
Executive summary:

0.1 g of the test substance was administered into the conjunctival sac of in each case one rabbit eye of 3 females and after 24 h the treated eyes were rinsed with physiologic saline.

Evaluation of eye irritation was made according to Draize.

According to the classification criteria Oxaphthalimid is not irritating to eyes.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Additional information

In the skin corrosion test in vitro with reconstructed human epidermis EST-1000 no corrosive property of the test item could be determined.

In the skin irritation test in all 3 animals no signs of skin irritation could be observed. There were no systemic intolerance reactions.

In the eye irritation test all animals showed reddening of the conjunctivae 1 h after instillation, only 2 of them had slight irritation signs at the conjunctivae after 24 h. All effects were fully reversible within 2 days. There were no systemic intolerance reactions.

Justification for classification or non-classification

Based on the study results a classification according to Directive 67/548/EEC and Regulation (EC) No. 1272/2008 (CLP) is not required.