1,2,3,4,4a,5,6,7-octahydro-2,5,5-trimethyl-2-naphthol

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

3 June 2020 - 5 June 2020

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

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

18 June 2019

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EC Guideline No. 440/2008. Part B: Methods for the Determination of Toxicity and other health effects, Guideline B.40 BIS: "In Vitro Skin Corrosion: Human Skin Model Test". Official Journal of the European Union No. L142

Version / remarks:

31 May 2008

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1,2,3,4,4a,5,6,7-octahydro-2,5,5-trimethyl-2-naphthol

EC Number:

255-256-8

EC Name:

1,2,3,4,4a,5,6,7-octahydro-2,5,5-trimethyl-2-naphthol

Cas Number:

41199-19-3

Molecular formula:

C13H22O

IUPAC Name:

2,5,5-trimethyl-1,2,3,4,4a,5,6,7-octahydronaphthalen-2-ol

Test material form:

liquid

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

Recommended test system in international guidelines (OECD and EC).

Vehicle:

unchanged (no vehicle)

Details on test system:

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37.0 ± 1.0ºC
- Temperature of post-treatment incubation: 37°C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: 1

DYE BINDING METHOD
- Dye used in the dye-binding assay: MTT
- Spectrophotometer: TECAN Infinite® M200 Pro Plate Reader
- Wavelength: 570 nm

NUMBER OF INDEPENDENT TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1 run (duplicates were used per exposure period)

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be corrosive to skin if the relative mean tissue viability obtained after 3-minute treatment compared to the negative control tissues is decreased below 50%. In addition, a test item considered non-corrosive (viability ≥ 50%) after the 3-minute treatment is considered corrosive if the relative tissue viability after 1-hour treatment with the test item is decreased below 15%.
- The test substance is considered to be non-corrosive to skin if the relative mean tissue viability obtained after the 3-minute treatment compared to the negative control tissues is not decreased below 50%. In addition, the relative tissue viability after the 1-hour treatment is not decreased below 15%.

ACCEPTABILITY CRITERIA
The in vitro skin corrosion test is considered acceptable if it meets the following criteria:
a) The absolute mean OD570 of the two tissues of the negative control should reasonably be within the laboratory historical control data range and the acceptance limits of OECD431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8).
b) The mean relative tissue viability following 1-hour exposure to the positive control should be <15 %.
c) In the range 20 - 100% viability, the Coefficient of Variation (CV) between tissue replicates should be ≤ 30%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 50 μL
- Concentration: undiluted

NEGATIVE CONTROL
- Amount applied: 50 μL

POSITIVE CONTROL
- Amount applied: 50 μL

Duration of treatment / exposure:

3-minute exposure or 1-hour exposure

Duration of post-treatment incubation (if applicable):

Incubation with 300 μL MTT-medium for 3 hours

Number of replicates:

2

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Remarks:

after 3-minute exposure

Value:

100

Negative controls validity:

valid

Remarks:

Mean (OD570): 2.008 ± 0.048 (HCD OD570: 1.258 – 2.414)

Positive controls validity:

valid

Remarks:

6.8%; Mean (OD570): 0.137 ± 0.002 (HCD OD570: 0.080 – 0.671)

Results 2

Irritation / corrosion parameter:

% tissue viability

Remarks:

after 1-hour exposure

Value:

47

Negative controls validity:

valid

Remarks:

Mean (OD570): 1.891 ± 0.199 (HCD OD570: 1.317 – 2.371)

Positive controls validity:

valid

Remarks:

7.2%; Mean (OD570): 0.137 ± 0.019 (HCD OD570: 0.032 – 0.319)

Other effects / acceptance of results:

The test item was checked for color interference in aqueous conditions. Addition of the test item to Milli-Q and isopropanol resulted after subtraction of the blank in an OD of -0.001 and 0.001, respectively. Therefore it was concluded that the test item did not induce color interference. In addition, because no color change was observed in the presence of MTT it was concluded that the test item did not interact with the MTT endpoint.

The relative mean tissue viability obtained after the 3-minute and 1-hour treatments with the test item compared to the negative control tissues was 100% and 47% respectively. Because the mean relative tissue viability for the test item was not below 50% after 3 minutes treatment and not below 15% after 1 hour treatment the test item is considered to be not corrosive.

ACCEPTIBILITY CRITERIA
a) The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range
b) The mean relative tissue viability following the 1-hour exposure to the positive control was 7.2%.
c) In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤ 14%, except for the 1-hour test item treated tissues, which showed a Coefficient of Variation of 36%. However, since the results of the individual treated tissues were within the same category, this deviation did not affect the study integrity indicating that the test system functioned properly.

Any other information on results incl. tables

Table 1: Mean Absorption in the in vitro Skin Corrosion Test with Ambrinol

	3-minute application (OD570)				1-hour application (OD570)
	A	B	Mean	SD	A	B	Mean	SD
Negative control	1.974	2.041	2.008 ±	0.048	1.750	2.032	1.891	0.199
Test item	2.094	1.910	2.002 ±	0.130	1.075	0.689	0.882	0.273
Positive control	0.135	0.139	0.137 ±	0.002	0.123	0.151	0.137	0.019

Applicant's summary and conclusion

Interpretation of results:

other: Not corrosive to the skin

Remarks:

in accordance with EU CLP (EC no 1272/2008 and its amendments)

Conclusions:

The substance is not corrosive to the skin.

Executive summary:

The corrosive potential of the test item was tested through topical application for 3 minutes and 1 hour. The study procedures described in this report were according to OECD TG 431 and GLP principles. The test item (a liquid) was applied undiluted (50 μL) directly on top of the skin tissue. The positive control had a mean relative tissue viability of 7.2% after the 1-hour exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD TG 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range. In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤ 14%, except for the 1-hour test item treated tissues, which showed a Coefficient of Variation of 36%. However, since both individual values were in the same category, this minor deviation did not affect the study integrity indicating that the test system functioned properly. The relative mean tissue viability obtained after 3-minute and 1-hour treatments with the test item compared to the negative control tissues was 100% and 47%, respectively. Because the mean relative tissue viability for the test item was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment the test item is considered to be not corrosive. In conclusion, the test item is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.