N,N’-di[3-(p-toluene sulfonyl)oxy]phenyl urea

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

June 18, 2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

N,N’-di[3-(p-toluene sulfonyl)oxy]phenyl urea

EC Number:

876-151-9

Cas Number:

2292123-68-1

Molecular formula:

C27H24N2O7S2

IUPAC Name:

N,N’-di[3-(p-toluene sulfonyl)oxy]phenyl urea

Test material form:

solid: particulate/powder

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

Recommended in the test guideline.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: LabCyte EPI-MODEL24 - SIT
- Tissue batch number(s): LCE24-200727-A
- Expiry date: 31 July 2020
- Date of initiation of testing: 29 July 2020

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37°C 

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: rinsed 15 times or more with PBS
- Observable damage in the tissue due to washing: None

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 0.5mg/mL per well
- Incubation time: 180 +/- 5 minutes
- Spectrophotometer: FLUOstar Omega, BMG LABTECH
- Wavelength: 570 and 650nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD 1.1
- Barrier function: IC50 = 0.27%
- Morphology: Observation of multilayered epidermis with a stratum corneum


NUMBER OF REPLICATE TISSUES: 3

Control samples:

yes, concurrent negative control

Amount/concentration applied:

Negative control: Distilled water - 25µl
Positive control: 5% w/v SDS solution - 25µl
Test item: neat (triplicate) - 25 mg

Duration of treatment / exposure:

15 minutes +/- 3 minutes

Duration of post-treatment incubation (if applicable):

42 +/- 1 hr

Number of replicates:

3

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1

Value:

92.1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

2

Value:

90.2

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3

Value:

103.4

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: no
- Direct-MTT reduction: no
- Colour interference with MTT: no

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes - mean measurement OD was 0.737
- Acceptance criteria met for positive control: yes - mean cell viability was 2.8
- Acceptance criteria met for variability between replicate measurements: yes - SDs of cell viability were, 2.7%, 1.0% and 7.1%

Any other information on results incl. tables

Table 2: Result of skin irritation test

Group	Tissue no.	Measurement value		Cell viability^a)(%)		SD^c) (%)	Category
			Mean		Mean^d)
Negative control (Distilled water)	1	0.759	0.737	103.0	100.0	2.7	-
	2	0.721		97.8
	3	0.731		99.2
Positive control (5% w/w SDS solution)	1	0.014	0.021	1.9	2.8	1.0	-
	2	0.02		2.7
	3	0.029		3.9
Test Item^d)	1	0.679	0.702	92.1	95.3	7.1	Non-irritant
	2	0.665		90.2
	3	0.762		103.4

^{a) Cell viability in the negative control was regarded as 100%}

^{b) The mean cell viability was calculated from mean OD value of each group}

^{c) The SD was calculated from the cell viability (n=3) of each tissue insert}

^{d) The mean OD value was not corrected because the staining ratio was <5%}

 

Table 3: Results of tissue-binding test

Group	Tissue no.	Measurement value		Staining ratio^b) (%)
			Mean
Negative contol item^a)	1	0.000	0.0	-
	2	0.000
Test item^a)	1	0.000	0.0	0.0
	2	0.000

^{a) Medium without MTT was used instead of MTT medium}

^{b) staining ratio ([mean OD value of the test item (without MTT)] - [mean OD value of negative control (without MTT)] / [mean OD value of negative control (with MTT)]) x 100}

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The mean cell viability of the test item was 95.3% and therefore the substance is not classified as a skin irritant.