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Description of key information

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 11- 16, 2012
Reliability:
1 (reliable without restriction)
Qualifier:
according to guideline
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Qualifier:
according to guideline
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Qualifier:
according to guideline
Guideline:
other: L’Oréal Standard Operating Procedure: “EpiSkin™ Skin Irritation Test Method“; - ECVAM Skin Irritation Validation Study - Validation of the EpiSkin™ Test Method15 min - 42 hours for the Prediction of Acute Skin Irritation of Chemicals. Version 1.8, 02-2009
Qualifier:
according to guideline
Guideline:
other: ECVAM Performance Standards for in vitro Skin Irritation Test Methods based on Reconstructed human Epidermis (RhE). Updated Version 24-Aug-2009
Qualifier:
according to guideline
Guideline:
other: ISO 10993-10, Annex D: 2010, “Biological evaluation of medical devices - Part 10: Tests for irritation and skin sensitization”
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Details on animal used as source of test system:
The test will be carried out with the reconstituted three-dimensional human skin model EPISKIN-SM™ (SkinEthic). This skin model consists of normal (non-cancerous), adult human-derived epidermal keratinocytes (NHEK) which have been cultured to form a multilayered, highly differentiated model of the human epidermis. The NHEK are cultured on chemically modified, collagen-coated cell culture inserts. A highly differentiated and stratified epidermis model is obtained after 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum
Species:
other: human skin model
Irritation / corrosion parameter:
% tissue viability
Value:
ca. 92
Vehicle controls validity:
valid
Negative controls validity:
not specified

The potential of the test item to induce skin irritation was analysed by using the three-dimensional human skin model EPISKIN-SM™ (SkinEthic) comprising a reconstructed epidermis with a functional stratum corneum.

In the present study Formaldehyde, reaction product with ethylenediamine was applied topically to the EPISKIN-SM™ tissue for 15 min followed by a 42 h post incubation period and immediate determination of cytotoxic effects via MTT reduction assay.

Irritant potential of the test item was predicted from the relative mean tissue viabilities obtained compared to the corresponding negative control tissues concurrently treated with PBS.

The test item showed no irritant effects. The mean relative tissue viability (% negative control) was > 50% (92%) after 15 min treatment and 42 h post incubation.

The controls confirmed the validity of the study. The mean OD550 of the six blank values was < 0.1 (92%) after 15 min treatment and 42 h post incubation. The controls confirmed the validity of the study. The mean OD550 of the six blank values was < 0.1. The mean absolute OD550 of the three negative control tissues was ≥ 0.6 and ≤ 1.5. The mean relative tissue viability (% negative control) of the positive control was ≤ 40% (7%). The maximum standard deviation of viability of replicate tissues of all dose groups was < 18% (3.4% - 9.8%)

Interpretation of results:
not irritating
Remarks:
Migrated information
Conclusions:
In this study under the given conditions the test item showed no irritant effects. The test item is therefore classified as “non-irritant” in accordance
with UN GHS and EU CLP “No Category”.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
May 30, 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)
Principles of method if other than guideline:
The Bovine Corneal Opacity and Permeability (BCOP) test method is an ex vivo test method that can be used to classify substances as “ocular corrosives and severe irritants”. The BCOP is recommended for use as part of a tiered-testing strategy for regulatory classification and labelling within a specific applicability domain. Test substances can be classified as ocular corrosives or severe irritants without further testing in rabbits.
To ensure test method integrity the data of the BCOP assay (Bovine Corneal Opacity and Permeability Assay), reference substances are tested in parallel to the test item.
GLP compliance:
yes (incl. QA statement)
Species:
cattle
Irritation parameter:
in vitro irritation score
Value:
ca. 29
Negative controls validity:
valid
Positive controls validity:
valid

The eye irritancy potential of Formaldehyde, reaction product with ethylenediamine was investigated in the bovine corneal opacity and permeability assay.

The following mean in vitro irritation score was calculated: 29.32

Therefore the test item was classified as moderate irritant.

The in vitro irritation score obtained with the positive control fell within the two standard deviations of the current historical mean and therefore this assay is considered to be valid.

OPACITY

Cornea No.

Test Item

Initial Opacity

Final Opacity

Change of Opacity Value

Corrected Opacity Value

1

Negative Control

3

7

4

 

2

3

3

0

 

3

3

4

1

 

MV

3.0

4.67

1.67

 

4

Positive

Control

4

101

97

95.33

5

4

59

55

53.33

6

4

69

65

63.33

MV

4.0

76.33

72.33

70.67

7

Test Item

2

35

33

31.33

8

2

32

30

28.33

9

2

32

30

28.33

MV

2.0

33.00

31.00

29.33

PERMEABILITY

Cornea No.

Test Item

OD490

Corrected OD490 Value

1

Negative Control

0.038

 

2

0.024

 

3

0.031

 

MV

0.031

 

4

Positive

Control

0.370

0.039

5

0.183

0.152

6

0.328

0.297

MV

0.294

0.263

7

Test Item

0.037

0.006

8

0.004

-0.027

9

0.049

0.018

MV

0.030

-0.001

IN VITRO IRRITATION SCORE

Cornea No.

Test Item

Corrected Opacity Value

Corrected OD490 Value

IVIS

1

Negative Control

4.00

0.038

 

2

0.00

0.024

 

3

1.00

0.031

 

MV

1.67

0.031

2.13

4

Positive

Control

95.33

0.039

 

5

53.33

0.152

 

6

63.33

0.297

 

MV

70.67

0.263

74.61

7

Test Item

31.33

0.006

 

8

28.33

-0.027

 

9

28.33

0.018

 

MV

29.33

-0.001

29.32
Interpretation of results:
Category 2 (irritating to eyes) based on GHS criteria
Conclusions:
Formaldehyde, reaction product with ethylenediamine is classified as Eye Irrit 2
Executive summary:

The test was considered valid to confirm classification as Eye Irrit 2 being needed. No further animal testing is justified.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

The results of the in vitro tests performed with formaldehyde, reaction products with ethylendiamine show neither corrosive nor irritation properties related to the test item. The ex vivo study performed to evaluate the irritation potential of the tet substance to the eyes, showed a moderate irritating potential.

Justification for selection of skin irritation / corrosion endpoint:

As in the skin corrosion study formaldehyde, reaction products with ethylendiamine resulted non corrosive, a skin irritation study has been carried out. Also this study gave negative results and therefore it has been choosen as endpoint selection study.

Effects on eye irritation: moderately irritating

Justification for classification or non-classification

According to the results of the available studies, formaldehyde, reaction products with ethylendiamine is classified as follows, according to Regulation 1272/2008/EC:

Eye Irrit. 2, H319.