Commiphora myrrha, ext.

Endpoint summary

Description of key information

In an in vitro skin irritation (OECD 439) test item was not irritant to skin (GLP, Rel. K1)

In an in vitro eye irritation study (OECD 492) test item was considered non irritant to the eyes (GLP, Rel K1)

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The experiment was performed between 09 November 2017 and 16 November 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study conducted in compliance with OECD Guideline No. 439 without any deviation.

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Version 28 July 2015

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

27 April 2017

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
Batch No.: 0021835
Date received: 17 October 2017
Storage: room temperature, darkness
Form: liquid
Colour: amber-yellow to green-yellow
Production date: June 2017
Best before date: October 2019

Test system:

human skin model

Source species:

other:

Cell type:

other: reconstructed epidermis

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- The 0.50 cm² reconstructed epidermises (Episkin SA, RHE/S/17 Batch No. 17-RHE-116) were received on 13 November 2017. The same day, the insert (filter + epidermis) was gently removed
from the agarose while avoiding leaving agarose on the polycarbonate filter. The inserts were placed in 6 wells culture plate which had been previously filled with 1 mL of growth medium (Episkin SA, batch No. 17 SGM 033) during 2 hours and 05 minutes. Then just before treatment, the inserts were placed in 24 wells culture plate which had been previously filled with 300 μL of maintenance medium (Episkin SA, batch No. 17 SMM 022). The two killed Reconstructed Human epidermis control tissue models, supplied by Episkin (Batch No. 17-RHE-021) were defrost on 21 February 2017, re-freeze on 21 February 2017 and defrost to be used on 28 February 2017.

TREATMENT
- The test item was applied, as supplied, at the dose of 16 µL, to the epidermal surface of 3 living human skin models and 2 killed Reconstructed Human epidermis (for non-specific MTT reduction control) during 42 minutes at room temperature. To ensure a good contact with the epidermises, during all the treatment period, the test item was recovered with a nylon mesh provided by Episkin SA.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: Test item was applied for 42 minutes at room temperature
- Temperature of post-treatment incubation: 41 hours and 50 minutes post-treatment incubation period in fresh medium at 37 °C, 5% CO2

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 16 µL
- Concentration (if solution): Undiluted

Duration of treatment / exposure:

42 minutes at room temperature.

Duration of post-treatment incubation (if applicable):

41 hours and 50 minutes post-incubation period at 37 °C, 5% CO2

Number of replicates:

3 living human skin models and 2 killed Reconstructed Human epidermis (for non-specific MTT reduction control)

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Main test (42 minutes)

Value:

92.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

VIABILITY
- The mean corrected percent viability of the treated tissues was 92.5%), versus 1.2% in the positive control (5% Sodium Dodecyl Sulfate).

Table 7.3.1/1: Assessment of the skin irritation - individual and average values of OD after 42 minutes exposure

 

Items	Skin	OD	Mean OD/disc#	Mean OD/product	Viability%	Mean viability%	SD	Conclusion
Negative control	1	0.958	1.073	1.001	107.2	100.0	8.7	-
		1.129
		1.133
	2	1.009	1.1026		102.5
		1.018
		1.052
	3	0.885	0.904		90.3
		0.912
		0.917
Positive control	4	0.010	0.010	0.012	1.0	1.2	0.2	Irritant
		0.011
		0.011
	5	0.011	0.011		1.1
		0.011
		0.011
	6	0.014	0.014		1.4
		0.015
		0.014
Test item	7	1.012	1.062	0.926	106.1	92.5	16.7	Non irritant
		1.128
		1.046
	8	0.771	0.739		73.8
		0.733
		0.713
	9	1.065	0.976		97.5
		0.765
		1.098

 

≠: mean of 3 values (triplicate of the same extract)

OD: optical density

Acceptance criteria: SD ≤ 18%

Negative control: OD value of the 3 replicates in the range ≥ 0.8 and ≤ 3.0.

The optical density was measured after a 1:2 dilution of the formazan extracts in isopropanol; the acceptability criteria should be in the range ≥ 0.4 and ≤1.5 for the negative control.

Interpretation of results:

GHS criteria not met

Conclusions:

Under these experimental conditions and in accordance with the Regulation (CE) No.1272/2008 and in absence of information on a skin corrosion, the test item has not to be classified.

Executive summary:

The aim was to evaluate the possible irritating effects of the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle after topical application on in vitro human reconstructed epidermis (SkinEthic RHE® model).

The test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle was applied as supplied at the dose of 16 μL, to 3 living Reconstructed Human epidermis (SkinEthic RHE® model) during 42 minutes. The application was followed by a rinse with 25 mL of DPBS and a 41 hours and 50 minutes post-incubation period at 37°C, 5% CO2. Cell viability was then measured by enzymatic conversion of the vital dye MTT into a blue formazan salt that was quantitatively measured after extraction from tissues. The experimental protocol was established in accordance with O.E.C.D. Test Guideline No. 439 adopted 28 July 2015 and the Test method B.46 of Council regulation No. 761/2009 dated 23 July 2009 (EU Journal L220) - ATP Council regulation No. 440/2008 of 30

May 2008 (E.U. Journal L142).

The mean percent viability of the treated tissues was 92.5% versus 1.2% in the positive control (5% Sodium Dodecyl Sulfate).

In accordance with the Regulation EC No. 1272/2008, the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle has to be considered as Non-irritant to skin. It corresponds to UN GHS No Category.

No hazard statement or signal word is required.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The experiment was performed between 19 December 2017 and 21 December 2017,

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study performed according to OECD Guideline 492 without any deviation.

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Version / remarks:

Adopted 28 July 2015

Deviations:

yes

Remarks:

Difference of viability between the 2 tissues treated with the positive control was 31.46%, instead of 20%. Considering the fact that these values are conformed to our historical data, this deviation is without impact on the conclusion of the study.

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

27 April 2017

Specific details on test material used for the study:

SURCE OF THE TEST MATERIAL
Batch No.: 0021835
Date received: 17 October 2017
Storage: room temperature, darkness
Form: liquid
Colour: amber-yellow to green-yellow
Production date: June 2017
Best before date: October 2019

Species:

other: reconstructed human cornea-like epithelium tissues

Details on test animals or tissues and environmental conditions:

The 0.60 cm² Reconstructed human Cornea-like Epithelia (EpiOcularTM OCL-212-ver2.0, supplied by MatTek Corporation, batch No. 27018) were received on 19 December 2017.
The same day, the tissues in their well shipping container were equilibrated at room temperature for 15 minutes. Then the inserts (filter + epithelium) were gently removed from the agarose while
avoiding leaving agarose on the polycarbonate filter. They were placed in 6 wells culture plate which had been previously filled with 1 mL of 37°C pre-warmed assay medium (MatTek Corporation, batch No. 121817ISA) and incubated during 19 hours and 55 minutes at standard culture conditions.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

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

Duration of treatment / exposure:

30 minutes at 37 °C, 5% CO2, 95% humidity (standard culture conditions).

Duration of post- treatment incubation (in vitro):

After 30 minutes of exposure, extensive rinsing with DPBS at room temperature, a 12 minutes post-exposure immersion period at room temperature and a 2 hours and 03 minutes post-exposure incubation at standard culture conditions.

Number of animals or in vitro replicates:

Test item, negative and positive controls were applied on duplicate tissues (2 living RhCE tissue replicates)

Details on study design:

The 0.60 cm² Reconstructed human Cornea-like Epithelia (EpiOcularTM OCL-212-ver2.0, supplied by MatTek Corporation, batch No. 27018) were received on 19 December 2017.
The same day, the tissues in their well shipping container were equilibrated at room temperature for 15 minutes. Then the inserts (filter + epithelium) were gently removed from the agarose while
avoiding leaving agarose on the polycarbonate filter. They were placed in 6 wells culture plate which had been previously filled with 1 mL of 37°C pre-warmed assay medium (MatTek Corporation, batch No. 121817ISA) and incubated during 19 hours and 55 minutes at standard culture conditions.

Irritation parameter:

other: %mean viability

Run / experiment:

Main test - 30 minutes

Value:

97.37

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

MEAN PERCENT TISSUE VIABILITIES
- The mean corrected percent tissue viability of the RhCE replicates treated with the test item was 97.37 %, versus 18.36% in the positive control (Methyl acetate).
ACCEPTANCE OF RESULTS:
- The mean percent tissue viabilities obtained with the positive control and negative controls are within the range of historical data and therefore validate the experiment.

The difference of viability between the 2 tissues treated with the positive control was 31.46%, instead of 20% at the maximum as initially scheduled.
Considering the results obtained and the fact that these values are conformed to our historical data, this deviation is considered as without impact on the conclusion of the
study.

Table 7.3.2/1: Assessment of the eye irritation potential individual and average values of OD after 30 minutes exposure

 

Items	Tissues	OD	Mean OD/disc#	Mean OD / product	Viability%	Mean viability%	Difference of viability%	Conclusion
Negative control	1	0.920	0.950	0.951	99.95	100.00	0.11	-
		0.952
		0.978
	2	0.933	0.951		100.05
		0.954
		0.967
Positive control	3	0.023	0.025	0.175	2.63	18.36	31.46	UN GHS category 2 or 1
		0.026
		0.027
	4	0.318	0.324		34.09
		0.321
		0.334
Test item	5	0.907	0.945	0.926	99.42	97.37	4.10	-
		0.974
		0.955
	6	0.897	0.906		95.32
		0.916
		0.905

#: mean of 3 values (triplicate of the same extract)

OD: optical density

Interpretation of results:

GHS criteria not met

Conclusions:

Under the experimental conditions adopted and in accordance with the Regulation EC No. 1272/2008, the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle does not require classification for eye irritation or serious eye damage. It corresponds to the UN GHS No Category.
No hazard statement and the signal word are required.

Executive summary:

The aim of the study was to evaluate the eye hazard potential of the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle after topical administration on in vitro reconstructed human cornea-like epithelium tissues (EpiOcularTM tissue model).

The test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle was applied as supplied, at the dose of 50 μL, to 2 living DPBS pre-treated RhCE (EpiOcularTM tissue model) during 30 minutes at 37°C, 5% CO2, 95% humidity (standard culture conditions). The exposure period was followed by extensive rinsing with DPBS at room temperature, a 12 minutes post-exposure immersion period at room temperature and a 2 hours post-exposure incubation at standard culture conditions. The tissue viability was measured by performing an MTT assay. The experimental protocol was established in accordance with O.E.C.D. Test Guideline No. 492 adopted 28 July 2015.

The mean percent tissue viability of the RhCE replicates treated with the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle was 97.37%, versus 18.36% in the positive control (Methyl acetate). In conclusion, under the experimental conditions adopted and in accordance with the Regulation EC No. 1272/2008, the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle does not require classification for eye irritation or serious eye damage. It corresponds to the UN GHS No Category.

No hazard statement and the signal word are required.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation:

The mean percent viability of the treated tissues was 92.5% versus 1.2% in the positive control (5% Sodium Dodecyl Sulfate).

In accordance with the Regulation EC No. 1272/2008, the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle has to be considered as Non-irritant to skin. It corresponds to UN GHS No Category.

No hazard statement or signal word is required.

Eye irritation:

The mean percent tissue viability of the RhCE replicates treated with the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle was 97.37%, versus 18.36% in the positive control (Methyl acetate). In conclusion, under the experimental conditions adopted and in accordance with the Regulation EC No. 1272/2008, the test item HUILE ESSENTIELLE MYRRHE SOMALIE 100% pure et naturelle does not require classification for eye irritation or serious eye damage. It corresponds to the UN GHS No Category.

No hazard statement and the signal word are required.

Justification for classification or non-classification

Self-classification:

Based on the available information (OECD 492 test), the substance is not classified for eye irritation/corrosion according to the Regulation (EC) No. 1272/2008.

Based on the available information (OECD 439 test), the substance is not classified for eye irritation/corrosion according to the Regulation (EC) No. 1272/2008.