4-Oxazolidinone, 3-cyclohexyl-2-(cyclohexylimino)-5-[[4-(diethylamino)-2-methoxyphenyl]methylene]-

Skin irritation / corrosion

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 for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

FC-131

IUPAC Name:

FC-131

Constituent 2

Reference substance name:

-

EC Number:

457-890-2

EC Name:

-

Cas Number:

860399-11-7

Molecular formula:

C27H39N3O3

IUPAC Name:

(2E,5Z)-3-cyclohexyl-2-(cyclohexylimino)-5-{[4-(diethylamino)-2-methoxyphenyl]methylidene}-1,3-oxazolidin-4-one

Constituent 3

Reference substance name:

453.62 g/mol

IUPAC Name:

453.62 g/mol

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 or test system 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: other: cell viability

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.