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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19-22 May 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study in compliance with EU-Method B.46 resp. OECD guideline 439

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
The following information concerning identity and composition of the test item is provided by the sponsor.
Name SABOSTAT A 300
Batch no. 20085596
Appearance whitish pastilles
Composition Reaction product of stearyl-diethanol-amine with C16-C18 saturated fatty acids
CAS No. not applicable (CAS No. of main compound 2-[(2-hydroxyethyl)octadecylamino]ethyl stearate: 52497-24-2)
EINECS-No. not stated
Molecular formula not applicable
Molecular weight 668 g/mol (calculated based on average UVCB composi-tion)
Purity 100%
Homogeneity homogeneous
Vapour pressure 5.85E-21 Torr Calculated using Advanced Chemistry De-velopment (ACD/Labs) Software V11.02 (© 1994-2015 ACD/Labs) h Pa at 25°C
Stability H2O; EtOH; acetone; CH3CN; DMSO: unknown
Solubility H2O : not soluble DMSO: soluble at 50°C; EtOH; acetone; CH3CN;
Production date 16. Apr. 2015.
Expiry date 10. Apr. 2016.
Storage Room temperature (20 ± 5 °C), keep away from light/ hu-midity

Test animals

Species:
human
Strain:
other: human skin model EpiDermTM

Test system

Amount / concentration applied:
25 mg on average
Duration of treatment / exposure:
60 minutes
Number of animals:
3 epidermic tissue
Details on study design:
This in-vitro study was performed in order to evaluate the potential of SABOSTAT A 300 to evoke skin irritation in a human-skin-model.
The test consists of a topical exposure of the neat test item to a human reconstructed epi-dermis model followed by a cell viability test. Cell viability is measured by dehydrogenase conversion of MTT (= 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide), pre-sent in cell mitochondria, into a blue formazan salt that is quantitatively measured after extraction from tissues. The percentage reduction of cell viability in comparison of untreat-ed negative controls is used to predict skin irritation potential.

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
other: other: Optical Density percentage with respect to negative control
Value:
119.6
Remarks on result:
other:
Remarks:
Basis: mean. (migrated information)

Any other information on results incl. tables

Findings and Results

 Measured Values

As blank, the optical density of isopropanol was measured in eight wells of the 96-well-plate. The measured values and their mean are given in the following table:

   Absorbance values blank isopropanol (OD at 570 nm)

Replicate

1

2

3

4

5

6

7

8

Mean

Absorbance

0.038

0.037

0.037

0.038

0.036

0.037

0.037

0.037

0.037

 

The absorbance values of negative control, test item and positive control are given in the following table:

    Absorbance Values negative control, test item and positive control (OD at 570 nm)

Designation

Measurement

Negative Control

SABOSTAT A 300

Positive Control

Tissue 1 

1

1.828

2.320

0.088

2

1.794

2.250

0.085

Tissue 2 

1

1.786

1.999

0.083

2

1.780

1.980

0.083

Tissue 3 

1

1.877

2.235

0.089

2

1.840

2.215

0.089

 

From the measured absorbances, the mean of each tissue was calculated, subtracting the mean absorbance of isopropanol as given in table above. Mean and relative standard deviation (comparison of the 3 tissues) were also calculated.

  Mean Absorbance Values

Designation

Negative Control

SABOSTAT A 300

Positive Control

Mean – blank (tissue 1)

1.774

2.248

0.050

Mean – blank (tissue 2)

1.746

1.953

0.046

Mean – blank (tissue 3) 

1.822

2.188

0.052

Mean of the 3 tissues

1.781

2.130

0.049

Relative standard deviation
of the 3 tissues

2.2%

7.3%

6.2%

 


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:

    % Formazan Production

Designation

SABOSTAT A 300

Positive Control

% Formazan production (tissue 1)

126.2%

2.8%

% Formazan production (tissue 2)

109.7%

2.6%

% Formazan production (tissue 3)

122.9%

2.9%

% Formazan production (mean)

119.6%

2.8%

 

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test item is considered as not skin irritant.
After the treatment, the relative absorbance values were increased to 119.6%. This value is well above the threshold for skin irritation (50%).
The optical density of the negative control (mean OD = 1.8) was well within the required acceptability criterion of 0.8 ≤ mean OD ≤ 2.8. The positive control induced a decrease in the relative absorbance as compared to the negative control to 2.8 % (required:  20%) for thus ensuring the validity of the test system. Variation within replicates was within the accepted range for negative control, positive control and test item.
For these reasons, the result of the test is considered valid.
Executive summary:

3 tissues of the human skin model EpiDermTMwere treated with the substance for 60 min. On average, 25 mg of the solid test item (wetted with 25 µL DPBS-buffer) were applied to each tissue and spread to match the tissue size (0.63 cm2; as indicated by supplier). DPBS-buffer was used as negative control, 5 % SDS solution was used as positive control. After treatment with the negative control, the absorbance values were within the required acceptability criterion of 0.8 ≤ mean OD ≤ 2.8, OD was 1.8. The positive control showed clear irritating effects. Relative absorbance was reduced to 2.8 %. Variation within tissues was acceptable (< 18%).   After the treatment with the test item, the relative absorbance values were increased to 119.6%. This value is well above the threshold for irritation potential (50%).   Therefore,the substance is considered as not skin irritant in the Human Skin Model Test.