Registration Dossier

Toxicological information

Skin irritation / corrosion

Currently viewing:

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2011-08-31 to 2011-10-12
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD guideline No. 439 and EU method B.46 in a GLP certified testing facility.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Details on test material:
Name: 400160
Batch no.: KL-11-038
Appearance: yellow, clear viscous liquid
Composition: UVCB, 2-Ethylhexyl esters of dimerised fatty acids, C18-unsaturated
Purity: 100% (GPC)
Production date: 24 March 2011
Expiry date: 24 March 2016
Storage: room temperature (20 ± 5 °C)

Test system

Vehicle:
unchanged (no vehicle)
Controls:
not required
Amount / concentration applied:
30 µL test item were applied on the tissue surface
Duration of treatment / exposure:
60 min
Observation period:
not applicable (cell viability was determined after the end of treatment period)
Number of animals:
not applicable (this is an in vitro test)
Details on study design:
The human skin model test was performed with commercially available Epi-200-SIT-Kit. The EpiDermTM tissues were produced from MatTek Corporation in Ashland, USA. The day of delivery was 01 July 2010 batch: 13657

Negative control: Dulbecco’s Phosphate Buffered Saline (DPBS) without CaCl2 and MgCl2
One plate (three tissues) was used as negative control; each tissue was treated with 30 µL DPBS buffer
Positive control: Sodium dodecyl sulphate (SDS), CAS No. 151-21-3, solution in deionised H2O, 5%
One plate (three tissues) was used as positive control; each tissue was treated with 30 µL SDS-solution
Test item: BYK-400160
One plate (three tissues) was used for treatment with the test item

Experimental performance:
After dosing the tissues with negative control, positive control and test item, they were incubated at 37°C for 35 min. After 60 min a rinsing step and two incubation periods at 37°C over a total period of 42 h followed.
After incubation, a MTT assay was performed and formazan production was measured by a spectral photometer at 570 nm.

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
other: other: formazan production in comparison to the negative control
Value:
100.9
Remarks on result:
other:
Remarks:
Time point: 60 min. Max. score: 102.5. Remarks: Result: not irritant. (migrated information)

In vivo

Irritant / corrosive response data:
After the treatment with the test item, the mean relative absorbance values were increased to 100.9 % in comparison to the negative control. This value is well above the threshold for irritation potential (50%).
Other effects:
no other effects observed

Any other information on results incl. tables

Evaluation and calculation:

The photometric absorption of the negative controls was considered as 100%. For each replicates of the test item and positive control, formazan production was calculated as % photometric absorption compared with the negative control.

% Formazan production = (OD test item resp. positive control / OD mean negative control) x 100

Table 1      Absorption values of the negative control, test item and positive control (OD at 570 nm)     

Designation

Measurement

Negative Control

400160

Positive Control

Tissue 1 

1

1.734

1.595

0.145

2

1.713

1.582

0.149

Tissue 2 

1

1.504

1.654

0.143

2

1.513

1.626

0.144

Tissue 3 

1

1.562

1.602

0.147

2

1.577

1.629

0.149

 

From the measured absorption values, the mean of each tissue was calculated and the mean absorption of isopropanol (0.046) was subtracted.

Table 2      Mean Absorption Values

Designation

Negative Control

400160

Positive Control

Mean – blank (Tissue 1)

1.678

1.543

0.101

Mean – blank (Tissue 2)

1.463

1.594

0.098

Mean – blank (Tissue 3) 

1.524

1.570

0.102

Mean of the three Tissues

1.555

1.569

0.100

Relative Standard Deviation
of the three tissues

7.1%

1.6%

2.1%

Comparison of formazan production:

For the test item and the positive control, the following percentage values of formazan production were calculated in comparison to the negative control:

Table 3     % Formazan Production

Designation

400160

Positive Control

% Formazan production (Tissue 1)

99.2%

6.5%

% Formazan production (Tissue 2)

102.5%

6.3%

% Formazan production (Tissue 3)

101.0%

6.6%

% Formazan production Mean

100.9%

6.5%

All validity criteria were fulfilled.

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the experimental conditions reported, 400160 did not exhibit any skin irritating potential.
Executive summary:

This in-vitro human skin model test (EU method B.46 resp. OECD 439) was performed in order to evaluate the potential of 400160 to evoke skin irritation in a human-skin-model.The study was performed under GLP compliance.

400160 was spread on the surface of three sections of an EpiDermTM tissue and incubated for 60 min. The negative control consisting of Dulbecco’s phosphate buffered saline was applied to another three tissue sections. As positive control a SDS solution (50 g/L) was used for three other tissue sections. After rinsing and two incubation steps of altogether 42 h an enzyme assay to determine cell viability was performed. The tissues were incubated with MTT-reagent for 3 h. The tissues were washed with PBS, dried and transferred to a 24 -well-plate. After incubating with isopropanol at room temperature over night, the developed formazan concentration was measured in a spectral photometer at 570 nm.

The mean percentage values of formazan production were calculated in comparison to the negative control. The relative absorbance values were increased to 100.9% after the treatment with 400160. This value is well above the threshold for irritation (50%). The positive control showed a reduction of formazan production of 6.5%. All validity criteria were fulfilled

In conclusion, it can be stated that the test item 400160 is considered as non skin irritant under the experimental conditions used in this study.