Registration Dossier

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013-07-30 to 2013-09-04
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: well documented study according to a OECD guideline and GLP
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report):FC-131
- Physical state: yellow crystalline powder
- Analytical purity: 99.8% (area HPLC)
- Impurities (identity and concentrations): about 0.1% 4-diethylamino-2-methoxy-benzaldehyde
- Isomers composition:No data
- Purity test date:03.07.2013
- Lot/batch No.:30610045
- Expiration date of the lot/batch:12.06.2015
- Homogeneity: yes
- Storage condition of test material: Room temperature (20+-5 °C), no humidity
- Production date: Jun. 2013

Test animals

Species:
other: in vitro test
Strain:
other: EpiDerm TM tissue
Details on test animals and environmental conditions:
Commercially available Epi-200-SIT-Kit:

The EpiDermTM tissue consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multi-layered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in vivo. The EpiDermTM tissues are cultured on specially prepared cell cultures inserts.

Origin:
- Epi-200 tissues were procured from MatTek In Vitro Life Science Laboratories, Bratislava
- Day of delivery: 20. Aug. 2013
- Batch: 18361

Test system

Type of coverage:
other: not applicable
Preparation of test site:
other: not applicable
Vehicle:
unchanged (no vehicle)
Controls:
other: positive and negative controls included
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit):
1: 25.7 mg FC-131
2: 26.3 mg FC-131
3: 25.0 mg FC-131

direct exposure of substance to Epi-200 tissues in 6-well-plates
Duration of treatment / exposure:
60 min
Observation period:
24 hours (after rinsing of the plates)
Number of animals:
in vitro method applied
Details on study design:
- preincubation of tissues in 6-well-plates containing 0.9 ml assay medium
- tissues wetted with 25 µL DPBS buffer before applying the test item
- after dosing, tissues rinsed immediatly
- transfer into new 6-well-plates
- evaluation of tissue surfaces with stereo microscope
- incubation for 24h

Control details:
- one plate: (three wells) negative control (Dulbecco’s Phosphate Buffered Saline)
- one plate: (three wells) positive control (5% SDS solution in deionised H20)

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
other:
Value:
ca. 109.2
Remarks on result:
other:
Remarks:
Basis: mean. Time point: 24h. Reversibility: other: not applicable. Remarks: in vitro test, score is related to relative absorbance (indicator for cell viability); the threshold of relative absorbance for irritation potential is 50 %; the observed value of 109% is well above this treshold. (migrated information)

In vivo

Irritant / corrosive response data:
After treatment, relative absorbance values and thus cell viability values were not reduced compared to the untreated negative controls (see Table 1 and 2). Relative absorbance in a mean was about 109% (see Table 3) and well above the treshold for irritation. Hence, no skin irritation potential of FC-131 was observed with the applied test conditions.

Any other information on results incl. tables

Table 1: Absorption Values negative control, test item and positive control (OD at 570 nm)        

Designation

Measurement

Negative Control

FC-131

Positive Control

Tissue 1 

1

2.004

2.151

0.093

2

2.116

2.335

0.087

Tissue 2 

1

2.131

2.397

0.100

2

2.127

2.367

0.094

Tissue 3 

1

2.250

2.403

0.084

2

2.226

2.362

0.086

Table 2: Mean absorption values

Designation

Negative Control

FC-131

Positive Control

Mean – blank (Tissue 1)

2.022

2.205

0.052

Mean – blank (Tissue 2)

2.091

2.344

0.059

Mean – blank (Tissue 3) 

2.200

2.345

0.047

Mean of the three Tissues

2.104

2.298

0.053

Relative Standard Deviation
of the three tissues

4.3 %

3.5 %

11.4 %

Table 3: Absoprtion in % related to untreated control for FC-131 treated cells and cells treated with the positive control (values related to formazan production)

Designation

FC-131

Positive Control

% Formazan production (Tissue 1)

104.8 %

2.5 %

% Formazan production (Tissue 2)

111.4%

2.8 %

% Formazan production (Tissue 3)

111.5 %

2.2 %

% Formazan production Mean

109.2 %

2.5 %

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Using the in vitro human skin model test (OECD 439) for testing of the skin irritation potential of FC-131, no irritation potential for the test item could be observed under the test conditions applied.
Executive summary:

The skin irritation potential of FC-131 was determined using the in vitro human skin model test in accordance with the OECD guideline 439 and the EU-Method B.46 under GLP. As the relative absorption value (correlating with the cell viability) with 109% was well above the treshold for irritation potential (50%), FC-131 is considered as not irritant in the human skin model test.