Myristica fragrans, ext.

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

September 01, 2010 - September 06, 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test was performed according to OECD Guideline 439 and under GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: OECD Guideline for the Testing of Chemicals 439: In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method (Original Guideline adopted July 22, 2010).

Deviations:

yes

Remarks:

updated guideline

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: "Commission Regulation (EC) No. 440/2008 B 46".

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: ECVAM international validation study on in vitro tests for acute skin irritation: Report on the validity of the EpiSkin™™ and EpiDerm assays and on the Skin Integrity Function Test (Altern Lab Anim. 2007 Dec; 35 (6): 559-601).

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Remarks:

Statement of Compliance

Test material

Test material information

Constituent 1

Reference substance name:

Nutmeg Oil - Myristica Fragrance Oil

IUPAC Name:

Nutmeg Oil - Myristica Fragrance Oil

Details on test material:

- Name of test material (as cited in study report): Nutmeg Oil - Myristica Fragrance Oil
- Analytical purity: Confidential
- Lot/batch No.: Confidential
- Stability in Solvent: Not relevant
- Storage condition of test material: At room temperature

Test animals

Species:

human

Details on test animals or test system and environmental conditions:

Human skin model EpiSkin™

Test system

Type of coverage:

other: not applicable

Preparation of test site:

other: not applicable

Vehicle:

unchanged (no vehicle)

Controls:

other: Deionised water

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 10 µL of the test item were applied to each of triplicate tissues.
- Concentration (if solution): 100%

Positive and negative controls: 10 µL were dosed per tissue.

Duration of treatment / exposure:

15 ± 1 min

Observation period:

Not applicable

Number of animals:

Three tissues of the human skin model EpiSkin™ were treated with the test item, the negative or the positive control.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Using a wash bottle the tissues were gently rinsed with PBS to remove any residual test material.
- Time after start of exposure: 15 ± 1 min

After further incubation for about 42 hours the tissues were treated with the MTT [(3-4,5-dimethyl thiazole 2-yl) 2,5-diphenyl-tetrazoliumbromide]solution for 3 hours following approx. 71 hours extraction of the colorant from the cells. The amount of extracted colorant was determined photometrically at 570 nm.

SCORING SYSTEM: The mean Optical Density (OD 570 nm ) of the 3 negative control tissues was calculated. This value corresponds to 100% tissue viability in the current test. For each individual tissue treated with the test item or the positive control the individual relative tissue viability is calculated. For the test item and the positive control the mean relative viability +/- standard deviation of the 3 individual tissues are calculated and used for classification. For the current test, an irritation potential of a test item according to EU classification Irritant (I), R38 (according to Directive 67/548/EEC), and H315 (category 2) (according to Regulation (EC) 1272/2008) is recommended if the mean relative tissue viability of 3 individual tissues < 50% of the negative control.

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

other: other: Relative Absorbance (% of Negative Control)

Value:

100

Remarks on result:

other:

Remarks:

Basis: other: Tissue 1, 2 and 3 - Negative control. Reversibility: other: Not applicable. Remarks: non-skin irritant. (migrated information)

Results 2

Irritation / corrosion parameter:

other: other: Relative Absorbance (% of Negative Control)

Value:

12.5

Remarks on result:

other:

Remarks:

Basis: other: Tissue 1, 2 and 3 - Positive control. Reversibility: other: Not applicable. Remarks: skin irritant. (migrated information)

Results 3

Irritation / corrosion parameter:

other: other: Relative Absorbance (% of Negative Control)

Value:

21.7

Remarks on result:

other:

Remarks:

Basis: other: Tissue 1, 2 and 3 - Test substance. Reversibility: other: Not applicable. Remarks: skin irritant (I), H315 (category 2). (migrated information)

In vivo

Irritant / corrosive response data:

After treatment with the test item Nutmeg Oil - Myristica Fragrance Oil the relative absorbance values decreased to 21.7%. This value is below the threshold for irritancy of ≤ 50%. Therefore, the test item is considered to possess an irritant potential.

Any other information on results incl. tables

After treatment with the negative control the absorbance values were within the required acceptability criterion of mean OD ≥ 0.6 till ≤1.5 for the 15 minutes treatment interval thus showing the quality of the tissues.

Treatment with the positive control induced a decrease in the relative absorbance as compared to the negative control to 12.5%, thus ensuring the validity of the test system.

The standard deviations between the % variabilities of the test item, the positive and negative controls were below 7% (threshold of the OECD Guideline 439: 18%), thus ensuring the validity of the study.

Applicant's summary and conclusion

Interpretation of results:

irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

After treatment with the test item Nutmeg Oil - Myristica Fragrance Oil the relative absorbance values decreased to 21.7%. This value is below the threshold for irritancy of ≤ 50%. Therefore, the test item is considered to possess an irritant potential. In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item Nutmeg Oil - Myristica Fragrance Oil is irritant to skin and according to EU classification needs to be classified as Irritant (I), R38 (according to Directive 67/548/EEC), and H315 (category 2) (according to Regulation (EC) 1272/2008).

Executive summary:

This in vitro study was performed according to OECD 439 to assess the irritation potential of Nutmeg Oil - Myristica Fragrance Oil by means of the Human Skin Model Test. Three tissues of the human skin model EpiSkin™ were treated with the test item, the negative or the positive control for 15 minutes. 10 µL of the test item were applied to each tissue, spread to match the tissue size. The test item and the positive and negative controls were washed off the skin tissues after 15 minutes treatment. After further incubation for about 42 hours the tissues were treated with the MTT [(3-4,5-dimethyl thiazole 2-yl) 2,5-diphenyl-tetrazoliumbromide] solution for 3 hours following approx. 71 hours extraction of the colorant from the cells. The amount of extracted colorant was determined photometrically at 570 nm. 10 µL of either the negative control (deionised water) or the positive control (5% Sodium lauryl sulfate) were applied to each tissue. After treatment with the negative control the absorbance values were within the required acceptability criterion of mean OD ≥ 0.6 till ≤1.5 for the 15 minutes treatment interval, thus showing the quality of the tissues. Treatment with the positive control induced a decrease in the relative absorbance as compared to the negative control to 12.5%, thus ensuring the validity of the test system. The standard deviations between the % variabilities of the test item, the positive and negative controls were below 7% (threshold of the OECD Guideline 439: 18%), thus ensuring the validity of the study. After treatment with the test item Nutmeg Oil - Myristica Fragrance Oil the relative absorbance values decreased to 21.7%. This value is below the threshold for irritancy of ≤ 50%. Therefore, the test item is considered to possess an irritant potential. In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item Nutmeg Oil - Myristica Fragrance Oil is irritant to skin and according to EU classification needs to be classified as Irritant (I), R38 (according to Directive 67/548/EEC), and H315 (category 2) (according to Regulation (EC) 1272/2008).