Reaction products of 3-methyl-5-[(1S,4aS,8aS)-5,5,8a-trimethyl-2-methylenedecahydro-1-naphthalenyl]-1-penten-3-ol, cyclized

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 - 23 September 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study conducted in accordance with international guidelines and in accordance with GLP. All guideline validity criteria were met.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

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)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Methods for the Determination of Toxicity and other health effects, Guideline B.40 BIS: "In Vitro Skin Corrosion: Human Skin Model Test"

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

1

Reference substance name:

Reaction products of 3-methyl-5-[(1S,4aS,8aS)-5,5,8a-trimethyl-2-methylenedecahydro-1-naphthalenyl]-1-penten-3-ol, cyclized

Molecular formula:

Not applicable (a generic molecular formula cannot be provided for this specific UVCB substance)

IUPAC Name:

Reaction products of 3-methyl-5-[(1S,4aS,8aS)-5,5,8a-trimethyl-2-methylenedecahydro-1-naphthalenyl]-1-penten-3-ol, cyclized

Test material form:

liquid: viscous

Details on test material:

Appearance: Off white paste

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:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm Skin Model
- Tissue batch number(s): Lot no.: 24364 kits M and L
- Production date: 21 September 2016
- Shipping date: Not reported
- Delivery date: Not reported
- Date of initiation of testing: Within 24 h of recieving kit

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: All incubations, with the exception of the test item incubation of 3 minutes at room temperature, were carried out at RH: 80 - 100% (actual range 53 - 90%), containing 5.0 ± 0.5% CO2 in air in the dark at 37.0 ± 1.0°C (actual range 35.6 - 37.3°C).
- Temperature of post-treatment incubation (if applicable): See above.

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: Volume and number of steps not reported. PBS was used as a washng solution.
- Observable damage in the tissue due to washing: No
- Modifications to validated SOP: No

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: MTT concentrate (5 mg/ml) diluted (1:5) with MTT diluent (supplemented DMEM).
- Incubation time: 3 h
- Spectrophotometer: TECAN Infinite® M200 Pro Plate Reader
- Wavelength: 570 nm
- Filter: Not reported
- Filter bandwidth: Not reported
- Linear OD range of spectrophotometer: Not reported

NUMBER OF REPLICATE TISSUES:

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE (N/A as MTT interference was discounted through experimentation)
- N/A
- Procedure used to prepare the killed tissues (if applicable): N/A
- N. of replicates : N/A
- Method of calculation used: N/A

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- 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%.
- Justification for the selection of the cut-off point(s) if different than recommended in TG 431 and 439: N/A

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 50 µL
- Concentration (if solution): Undiluted

VEHICLE
- Amount(s) applied (volume or weight with unit): N/A
- Concentration (if solution): N/A
- Lot/batch no. (if required): N/A
- Purity: N/A

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): N/A

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): 8 N

Duration of treatment / exposure:

3 minutes (2 tissues) and 60 mins (2 tissues)

Duration of post-treatment incubation (if applicable):

Incubated in MTT-medium for 3 hours followed by a formazan extraction period overnight.

Number of replicates:

2 per test group

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 min exposure

Value:

104

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: no indication of corrosion

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 min exposure

Value:

89

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: no indication of corrosion

Any other information on results incl. tables

Table 1:       Mean adsorption (OD₅₇₀) in thein vitroskin corrosion test with the test item

	3-minute exposure			60-minute exposure
	Rep A	Rep B	Mean ± SD	Rep A	Rep B	Mean ± SD
Negative control	1.526	1.586	1.556 ± 0.042	1.770	1.588	1.679 ± 0.129
Test item	1.581	1.654	1.618 ± 0.052	1.700	1.277	1.488 ± 0.299
Positive control	0.180	0.155	0.167 ± 0.018	0.205	0.163	0.184 ± 0.030

 

Table 2:       Mean tissue viability in the in vitro skin corrosion test with the test item (%)

	3-minute exposure viability (percentage of control)	60-minute exposure viability (percentage of control)
Negative control	100	100
Test item	104	89
Positive control	11	11

 

Table 3:       Coefficient of variance between tissue replicates (%)

	3-minute exposure	60-minute exposure
Negative control	3.8	10
Test item	4.4	25
Positive control	14	21

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this summary.

Executive summary:

The skin corrosion potential of the test item was evaluated in accordance with the OECD 431 guidance document and GLP using the reconstructed human epidermis (RHE) skin model.

The test item was tested through topical application for 3 minutes and 1 hour on a human three dimensional epidermal model (EpiDerm (EPI-200)). The test item was heated up to 60°C and applied undiluted (50 µl) directly on top of the skin tissue.

The positive control had a mean relative tissue viability of 11% 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 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 ≤ 25%, 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 104% and 89%, 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.

It is concluded that this test is valid and that the test item is not corrosive in thein vitro skin corrosion test under the experimental conditions described in this summary.