Paraffin waxes (Fischer-Tropsch), isomerization

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Remarks:

Skin Irritation Test

Adequacy of study:

key study

Study period:

2022-02-10 until 2022-03-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Title:

Unnamed

Year:

2022

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: • MatTek Corporation Protocol: In vitro EpiDermTM Skin Irritation Test (EPI-200-SIT) For use with MatTek Corporation’s Reconstructed Human Epidermal Model EpiDerm (EPI-200-SIT); Version 12/04/2020. • MatTek Corporation Protocol: EpiOcularTM Eye Irritation

Version / remarks:

Note:
The Pre-Test (Assessment of Colored or Staining Materials and Assessment of Direct Test Article Reduction by MTT) was performed according to the MatTek Protocol “EpiOcularTM Eye Irritation Test.
Reason:
The OECD TG 439 requires the evaluation of colored or staining materials spectrophotometrically but does not state the evaluation criteria. These criteria are missing in the MatTek protocol “In vitro EpiDermTM Skin Irritation Test”, too. But the identical scientific hypothesis is to clarify in the Eye Irritation Test and there the requirements are described.

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: UN GHS

Version / remarks:

published 2003, last (8th revision) 2019

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Reconstructed Human Epidermis Test Methods (14 June 2021), and as described in detail in the Protocol for:
In vitro EpiDermTM Skin Irritation Test (EPI-200-SIT) for use with MatTek Corporation’s Reconstructed Human Epidermal Model EpiDerm (EPI-200).

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

06 July 2012

Deviations:

no

GLP compliance:

yes

Remarks:

Date of GLP Certificate: 2019-10-23

Test material

Test material information

Constituent 1

Reference substance name:

Paraffin waxes (Fischer-Tropsch), isomerization

Cas Number:

2658498-20-3

Molecular formula:

C25H52 - C150H302

IUPAC Name:

Paraffin waxes (Fischer-Tropsch), isomerization

Details on test material:

- Purity: 100%
- Molecular weight: Not applicable (UVCB)
- Physical state / Appearance: Solid / white
- Expiry Date / Retest Date: Not indicated
- Storage Conditions: Room temperature
- Stability in Solvent: Not indicated

Specific details on test material used for the study:

Appearance: White, solid (waxy)
Storage Conditions: At room temperature
Stability in Solvent: Not indicated

In vitro test system

Test system:

human skin model

Source species:

other: Standard Assay Kit and MTT-100 kit were purchased from MatTek Corporation (82105 Bratislava, Slovakia, Lot No.: 36129).

Justification for test system used:

Dermal irritation is generally defined as "the production of reversible inflammatory changes in the skin". The potential for chemical induced skin irritation is an important consideration in establishing procedures for the safe handling, packing and transport of chemicals. It is usually determined in the in vivo Draize rabbit skin irritation test as described in OECD guideline 404. Because systemic reactions play a minor role in modulating local skin toxicity potential of chemicals, skin irritation potential may be predicted by in vitro systems, provided they are sufficiently complex to mimic human skin barrier and cell reactivity. In an international prevalidation study performed by ECVAM, the in vitro skin irritation test using the human skin model EpiDerm™ and EpiSkin™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for skin irritancy potential.

Vehicle:

unchanged (no vehicle)

Details on test system:

In vitro Skin Irritation Test (OECD 439)

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit):25 +/- 2 mg (39.7 mg/cm2 according to guideline)

NEGATIVE CONTROL
- Concentration (if solution): 30 µL DPBS

POSITIVE CONTROL
- Concentration (if solution): 5% SDS solution in deionised water (MatTek)

Duration of treatment / exposure:

60 minutes

Number of replicates:

Three tissues

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Test Item

Value:

100.08

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Remarks:

according to UN GHS and EU CLP regulations

Any other information on results incl. tables

Main Experiment: Results after treatment with test item and controls

Treatment Group	Tissue No.	Well 1 OD 570 nm	Well 2 OD 570 nm	Well 3 OD 570 nm	Mean OD of 3 Wells	Mean OD of 3 Wells blank corrected	Mean OD of 3 tissues	Rel. Viability [%] Tissue 1, 2 + 3	Standard Deviation	Mean Rel. Viability [%]
Treatment Group	Tissue No.	0.037
Blank		1.580	0.037	0.037	0.037
Negative Control	1	1.625	1.537	1.561	1.559	1.522		97.658
	2	1.658	1.596	1.590	1.604	1.567	1.559	100.530	2.1	100.0
	3	0.125	1.606	1.607	1.624	1.587		101.813
Positive Control	1	0.100	0.110	0.109	0.115	0.078		5.004
	2	0.106	0.094	0.092	0.095	0.058	0.066	3.742	0.7	4.25
	3	1.575	0.097	0.094	0.099	0.062		3.997
Test Item	1	1.626	1.567	1.565	1.569	1.532		98.297
	2	1.607	1.609	1.611	1.615	1.579	1.560	101.272	1.6	100.08
	3				1.606	1.569		100.669

^{OD: optical density}

Applicant's summary and conclusion

Interpretation of results:

other: EU-GHS criteria not met

Conclusions:

In conclusion, it can be stated that in this study and under the experimental conditions reported, the registration substance is non-irritant to skin according to UN GHS and EU CLP regulation.

Executive summary:

This in vitro study was performed to assess the skin irritation potential of the registration substance 'Paraffin waxes (Fischer-Tropsch), isomerization' by means of the Human Skin Model Test.

The test item did not prove to be a MTT reducer in the MTT interference pre-experiment. Also, its intrinsic color was not intensive and the OD of the test item in deionised water or isopropanol at 570 nm after blank correction was < 0.08. Therefore, additional tests with freeze-killed tissues or viable tissues (without MTT addition) did not have to be performed.

Each three tissues of the human skin model EpiDerm^™ were treated with the test item, the negative control (DPBS) or the positive control (5% SDS) for 60 minutes.

After treatment with the negative control the absorbance values were well within the required range of the acceptability criterion of mean OD 0.8 and ≤ 2.8 for the 60 minutes treatment interval, thus assuring the quality of the tissues.

Treatment with the positive control induced a decrease in the viability compared with the negative control to 4.25%, thus ensuring the validity of the test system.

The standard deviations between the three tissue percentage viability values of each group (test item, positive and negative controls) in the main test were below 18 (threshold according to OECD 439: ≤ 18), thus ensuring the validity of the study.

After treatment with the test item the mean viability value was 100.08% compared to the relative absorbance value of the negative control. This value is above the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential.