3,7-dimethyloct-1-en-3-yl acetate

Endpoint summary

Description of key information

Based on experimental data, DMOE-Acetate was found to be non-irritating to skin according to the key in vitro study with EpiDerm™ tissue (OECD 439) and non-irritating to the eye according to a weight of evidence approach based on in vitro studies (OECD 492 and 437).

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:

03 June 2015 to 29 June 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

July 26, 2013

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: EpiDerm™ tissue

Source strain:

other: human-derived epidermal keratinocytes

Justification for test system used:

The OECD Guideline 439 assay is based on reconstructed human epidermis (RhE).

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ tissue
- Tissue batch number: Lot No.: 21677
- Delivery date: June 23, 2015
- Date of initiation of testing: On day of receipt the preincubation phase of the EpiDerm™ tissues started.


TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 ± 1.5 °C
- Temperature of post-treatment incubation: 42 h

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps:
After the end of the treatment interval the inserts were removed immediately from the 6-well plate and tissues were gently rinsed with DPBS at least 15 times in order to remove any residual test material. After the rinsing the inserts were submerged in DPBS at least three times. Afterwards the inserts were once again rinsed with sterile DPBS from the inside and the outside. Excess DPBS was removed by gently shaking the inserts and blotting the bottom with sterile blotting paper.


MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 h
- Spectrophotometer: Versamax® Molecular Devices, Softmax Pro, version 4.7.1
- Wavelength: 570 ± 1 nm


NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
not required: The colour of test item/water mixture did not change during the incubation period compared with the colour of the pure test item. Therefore, the measurement of the OD of the test item in water at 570 nm was not required and consequently not performed.

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

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant or corrosive to skin if the mean tissue viability is less than or equal to 50%.

Control samples:

yes, concurrent negative control

Amount/concentration applied:

TEST MATERIAL
30 µL (47 µL/cm2 according to guideline) of the undiluted test item was dispensed directly atop the EpiDerm™ tissue and spread to match the surface of the tissue for a complete treatment time of 60 minutes.

NEGATIVE CONTROL
30 µL DPBS (MatTek) was used as negative control per tissue.

POSITIVE CONTROL
30 µL of a 5% SLS solution in deionised water (MatTek) was used a positive control per tissue.

Duration of treatment / exposure:

60 min

Duration of post-treatment incubation (if applicable):

42 h

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1

Value:

112.5

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:

Criterion 1 (negative control): The absolute OD 570 nm of the negative control tissues in the MTT test is an indicator of tissue viability obtained after the shipping and storing procedure and under specific conditions of the assay. Tissue viability is meeting the acceptance criterion if the mean OD570 of the negative control tissues is ≥ 0.8 and ≤ 2.8.
Criterion 2 (positive control): An assay is meeting the acceptance criterion if mean relative tissue viability of the positive control is ≤ 20%.
Criterion 3: Rel. standard deviation: The SD of 3 identical replicates should be < 18%. OD values should not be below historically established boundaries. Historical data and the quality certificate of the supplier of the test kit demonstrated the robustness of the test system or rather of the test kit.

Table 1: Results after treatment (60 min) with Dimethyl Octenyl Acetate (DMOE-Ac) and the controls

	Mean Absorbance of 3 Tissues	Relative Standard Deviation [%]	Mean Rel. Absorbance [% of Negative Control]
Negative control	1.792	4.1	100
Positive control	0.096	13.7	5.4
Test item	2.016	16.8	112.5

 

Interpretation of results:

GHS criteria not met

Conclusions:

Dimethyl Octenyl Acetate (DMOE-Ac) is not irritant to skin according to UN GHS.

Executive summary:

This in vitro study was performed to assess the irritation potential of Dimethyl Octenyl Acetate (DMOE-Ac) by means of the Human Skin Model Test. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Also its intrinsic colour was not intensive. Consequently, additional tests with freeze-killed or viable tissues were not necessary. Each three tissues of the human skin model EpiDerm™ were treated with the test item, the negative or the positive control for 60 minutes. 30 µL of the test item were applied to each tissue, and spread to match the surface of the tissue. 30 µL of either the negative control (DPBS) or the positive control (5% SLS) were applied to each tissue. After treatment with the negative control the absorbance values were well in 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 sufficient decrease in the relative absorbance as compared to the negative control for the 60 minutes treatment interval, and thus assuring the validity of the test system. After treatment with the test item the mean relative absorbance value was 112.5% 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. In conclusion, it can be stated that in this study and under the experimental conditions reported, Dimethyl Octenyl Acetate (DMOE-Ac) is not irritant to skin.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2015-05-15 to 2015-05-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

Version / remarks:

April, 1997

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Version / remarks:

July, 2013

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: Schlachthof Bensheim, 64625 Bensheim, Germany
- Number of animals: not specified
- Characteristics of donor animals: at least 9 month old donor cattle
- Storage, temperature and transport conditions of ocular tissue:
The isolated eyes were transported to the laboratory in HBSS at ambient temperature. The corneae were isolated on the same day after delivery of the eyes. The corneae were directly used in the BCOP test on the same day.
- Time interval prior to initiating testing:
For equilibration, the corneae in the holder were incubated in a vertical position for about one hour at 32 ± 1 °C in a water-bath. At the end of the incubation period, the basal opacity was determined (t0).
- Selection and preparation of corneas:
The cornea was carefully removed from the eye using scalpel and rounded scissors. A rim of about 2 mm of tissue (sclera) was left for stability and handling of the isolated cornea.
Each isolated cornea was mounted in a specially designed cornea holder according to the description given in OECD guideline 437, that consists of anterior and posterior compartments, which interface with the epithelial and endothelial sides of the cornea, respectively. The endothelial side of the cornea was positioned against the sealing ring (O-ring) of the posterior part of the holder. The cornea was gently flattened over the O-ring but stretching was avoided. The anterior part of the holder was positioned on top of the cornea and fixed in place with screws. Both compartments of the holder were filled with incubation medium. The posterior compartment was filled first to return the cornea to its natural convex position. Care was taken to assure no air bubbles were present within the compartments.

- Quality check of the isolated corneas: All eyes were carefully examined macroscopically for defects. Those presenting defects such as vascularization, pigmentation, opacity and scratches were discarded. The basal opacity of all corneae was recorded. Each corneae with a value of the basal opacity > 7 was discarded.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

0.75 mL on the surface of the corneae

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

2 hours after rinsing with saline

Number of animals or in vitro replicates:

3 (test item and controls)

Details on study design:

NUMBER OF REPLICATES: 3

NEGATIVE CONTROL USED: yes, see above

POSITIVE CONTROL USED: yes, see above

APPLICATION DOSE AND EXPOSURE TIME: see above

POST-INCUBATION PERIOD: yes, 2 hours after rinsing with saline and before opacity measurement, at 32 ± 1 °C in incubation medium

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: via opacitometer OP_KiT opacitometer (Electro Design, 63-Riom France), measured before treatment (t0) and after 10 min treatment with test item or controls and 3 hours of post exposure incubation (t130)
- Corneal permeability: passage of sodium fluorescein dye (1 mL of a 0.4% (w/v) sodium fluorescein solution) measured after a 90 minutes incubation in a water-bath at 32 ± 1 °C with the aid of spectrophotometry via Versamax® Molecular Devices at OD490

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)
IVIS(negative control) = opacity value + (15 x OD490 value)
IVIS(positive control or test item) = (opacity value – opacity value mean negative control) + (15 x corrected OD490 value)

DECISION CRITERIA:
1. IVIS ≤ 3: No Category (according to GHS)
2. IVIS > 3; ≤ 55: No prediction can be made
3. IVIS > 55: Serious eye damage according to CLP/EPA/GHS (Cat 1)

The test will be acceptable if
• the positive control gives an IVIS that falls within two standard deviations of the current historical mean (updated every three months), and if
• the negative control responses result in opacity and permeability values that are less than the established upper limits for background opacity and permeability values for bovine corneae treated with the respective negative control

Irritation parameter:

in vitro irritation score

Run / experiment:

1

Value:

3.85

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

not determinable

Remarks:

No prediction can be made; threshold for serious eye damage: IVIS ≥ 55

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: No

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: With the negative control (saline) neither an increase of opacity nor permeability of the corneae could be observed (mean IVIS = 1.06).
- Acceptance criteria met for positive control: The positive control (2-Ethoxyethanol) was tested undiluted and showed a clear increase in opacity and distinctive increased permeability of the corneae (mean IVIS = 56.25) corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1))

Test Group	Opacity value = Difference (t130-t0) of Opacity	Permeability at 490 nm (OD490)	Mean IVIS	Proposed in vitro Irritancy Score
Negative Control	0;0;0 Mean: 0.00	0.073; 0.066; 0.072 Mean: 0.070	1.10; 0.99; 1.08 Mean: 1.06	Not categorized
Positive Control	58.0; 52.0; 26.0	0.672; 0.784; 0.729	68.08; 63.76; 36.93 Mean: 56.25	Category 1
Test Item	4.00; 4.00; 4.00	-0.009; -0.017; -0.004	3.86; 3.74; 3.94 Mean: 3.85	No prediction can be made

Interpretation of results:

study cannot be used for classification

Conclusions:

In conclusion, according to the current study and under the experimental conditions reported, Dimethyl Octenyl Acetate (DMOE-Ac) is not serious eye damaging (CLP/EPA/GHS (Cat 1) but a prediction for the damage hazard cannot be made (GHS).

Executive summary:

This in vitro study was performed to assess the corneal damage potential of Dimethyl Octenyl Acetate (DMOE-Ac) by means of the BCOP assay using fresh bovine corneae.
After a first opacity measurement of the fresh bovine corneae (t0), the neat test item, the positive, and the negative controls were applied to corneae fixed in an incubation chamber in horizontal position for 10 minutes at 32 ± 1 °C. The posterior chamber contained incubation medium. After the incubation phase the test item, the positive, and the negative controls were each rinsed from the corneae. Further, the corneae were incubated for another 120 minutes at 32 ± 1 °C in a vertical position, while the anterior chamber contain incubation medium as well. Afterwards, opacity was measured a second time (t130).
After the opacity measurements permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein after incubation in a horizontal position for 90 minutes at 32 ± 1 °C.
With the negative control (0.9% (w/v) NaCl solution in deionised water) neither an increase of opacity nor permeability of the corneae could be observed.
The positive control (2-Ethoxyethanol) showed a clear increase in opacity and distinctive increased permeability of the corneae corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)).
Relative to the negative control, the test item Dimethyl Octenyl Acetate (DMOE-Ac) caused a slight increase of the corneal opacity but no relevant changes in permeability. The calculated mean in vitro irritancy score was 3.85. According to OECD 437 (see table in chapter 3.10.3) the test item is not classified as serious eye damaging (CLP/EPA/GHS (Cat 1) but the test item’s hazard for eye damaging cannot be predicted.

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2020-11-10 to 2020-12-10

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: MatTek Corporation Protocol: EpiOcular™ Eye Irritation Test (OCL-200-EIT) for the prediction of acute ocular irritation of chemicals; for use with MatTek Corporation’s Reconstructed Human EpiOcular™ Model; MK-24-007-0055

Version / remarks:

2015-05-29

Deviations:

no

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:

2019-06-18

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

human

Details on test animals or tissues and environmental conditions:

- Justification of the test method and considerations regarding applicability:
The EpiOcular™ Eye Irritation Test (EIT) is an in vitro procedure allowing the identification of chemicals (substances and mixtures) not requiring classification and labelling for eye irritation or serious eye damage in accordance with UN GHS. It makes use of reconstructed human cornea-like epithelium (RhCE) which closely mimics the histological, morphological, biochemical and physiological properties of the human corneal epithelium. In a pre-validation study performed by Avon Products Inc. and MatTek Corporation, the in vitro eye test using the human cornea model EpiOcular™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for eye irritancy potential. The EIT was validated by the European Union Reference laboratory for Alternatives to Animal Testing (EURL ECVAM) and cosmetics Europe between 2008 and 2013. A limitation of the test method following OECD 492 is that it does not allow discrimination between eye irritation/reversible effects on the eye (Category 2) and serious eye damage/irreversible effects on the eye (Category 1), nor between eye irritants (optional Category 2A) and mild eye irritants (optional Category 2B), as defined by UN GHS. For these purposes further testing with other suitable test methods is required.
- Description of the cell system used, incl. certificate of authenticity and the mycoplasma status of the cell live:
The EpiOcular tissue construct is a non-keratinized epithelium prepared from normal human keratinocytes. It models the cornea epithelium with progressively stratified, but not cornified cells. These cells are not transformed or transfected with genes to induce an extended life span in culture. The “tissue” is prepared in inserts (MILLICELL, 10 mm ∅) with a porous membrane through which the nutrients pass to the cells. A cell suspension is seeded into the insert in specialized medium. After an initial period of submerged culture, the medium is removed from the top of the tissue so that the epithelial surface is in direct contact with the air (surface 0.6 cm²). This allows the test item to be directly applied to the epithelial surface in a fashion similar to how the corneal epithelium would be exposed in vivo.
- RhCE tissue or hCE cell construct used, including batch number: MatTek Corporation (82105 Bratislava, Slovakia, Lot No.: 30688)

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

50 µL

Duration of treatment / exposure:

30 minutes

Number of animals or in vitro replicates:

duplicate tissues

Details on study design:

- Details of the test procedure used:
After pre-warming of the EpiOcular™ tissues was completed, and prior to application of the test item respectively the controls, all tissues were pre-wetted with 20 µL Ca2+Mg2+ free DPBS and incubated for 30 min. According to OECD guideline 492 a minimum of 83.3 μL/cm2 ≙ 50 µL for the liquid test item were used for treatment. Concurrent negative, positive control and the test item were applied in duplicate tissues at a volume of 50 µL atop the tissue surface and incubated for 30 min. Afterwards all tissues were rinsed several times and incubated for 12 min in 5 ml assay medium in a 12-well plate at room temperature (post exposure immersion). At the end of this incubation the tissues were transferred into a 6-well plate with 1 ml assay medium and incubated for a post-treatment incubation for 120 min at standard conditions.

- Temperature of exposure:
Each incubation of the tissues was performed under 37 ± 1.5°C and 5 ± 0.5% CO2 in DMEM Medium.

- Indication of controls used for direct MTT-reducers and/or colouring test chemicals:
1. Color Interference Test:
50 µL of the test item was added to 1 mL of deionised water and mixed. 1 mL of deionised water was used as control (blank). Both were incubated for 60 min under standard conditions. In parallel, 50 µL of the test item was added to 2 mL of isopropanol and mixed. A control (2 mL of isopropanol, blank) was run concurrently. Both were incubated for 3 hours at room temperature. After incubation the presence of the staining was evaluated by OD570 nm measurement.

2. The MTT Interference Test:
To test if a test item directly reduces MTT, 1 ml of a MTT solution (1 mg/mL) including 50 µL of the test item was incubated for 180 min under standard conditions. 50 µL deionised water in MTT solution was used as negative control. After incubation the change of color was determined by the unaided eye.

- Number of tissue replicates used per test chemical and controls (positive control, negative control, NSMTT, NSCliving and NSCkilled):
Duplicate tissues were used for all test groups (positive control, negative control, test item).
Since the OD of the test item in deionised water or isopropanol at 570 nm after blank correction was < 0.08 in the first and did not interfere with MTT in the second pre-experiment, no additional tissues were necessary.

- Description of the method used to quantify MTT formazan, if applicable:
The tissues were extracted from both, the top and the bottom of the tissues. For the MTT-Assay, tissues were incubated for 180 min in 300 µl MTT solution. Each tissue was extracted with isopropanol within 4-72 h at 2-8°C without shaking. To mix the extract, the plates were placed on an orbital plate shaker and shaken for 2-3 hours at room temperature. Then, the extracts were mixed and two 200 µL aliquots were transferred to a 96- well plate for OD measurement. 200 µL of isopropanol were added to the wells designated as blanks for 96-well plate. The optical density (OD570nm) was determined spectrophotometrically in duplicates by a microplate reader (Versamax® Molecular Devices). The absorbance values were determined using the software SoftMax Pro Enterprise.
- Description of evaluation criteria used including the justification for the selection of the cut-off point for the prediction model:
1. If the test item-treated tissue viability is > 60% relative to the negative control treated tissue viability, the test item is identified as not requiring classification and labelling according to UN GHS (No Category).
2. If the test item-treated tissue viability is ≤ 60% relative to negative control treated tissue viability, no prediction can be made for this test item.
3. A single test composed of at least two tissue replicates should be sufficient for a test chemical, when the result is unequivocal. However, in cases of borderline results, such as non-concordant replicate measurements and/or mean percent tissue viability equal to 60±5%, a second test should be considered, as well as a third one in case of discordant results between the first two tests.

- Reference to historical positive and negative control results demonstrating suitable run acceptance criteria: yes

- Demonstration of proficiency in performing the test method before routine use by testing of the proficiency chemicals:
Yes, Certificate of Proficiency certifies that the testing facility has demonstrated proficiency in the EpiOcular Eye Irritation Test (OECD TG 492) by correct prediction of the proficiency chemicals as defined by the OECD TG 492.
Acceptance criteria:
1. Positive and negative control means and acceptance ranges based on historical data: OD is > 0.8 and < 2.8.
2. Acceptable variability between tissue replicates for positive and negative controls: The mean relative viability of the positive control is below 50% of the mean negative control viability
3. Acceptable variability between tissue replicates for the test chemical: < 20 percentage points (p.p.) in the same run

Irritation parameter:

percent tissue viability 

Run / experiment:

1.

Value:

86.93

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: No

DEMONSTRATION OF TECHNICAL PROFICIENCY: Yes

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes, OD was with 2.328 >0.8 and <2.8
- Acceptance criteria met for positive control: yes, decrease below 50% with 22.28%
- Range of historical values if different from the ones specified in the test guideline: difference of relative viability between the two relating tissues was < 20 p.p. (values
between 1.01 p.p. and 8.13 p.p.) in the same run (for test item tissues, positive and negative control tissues).

Tab. 1 Results after treatment with the test item and the controls for 30 minutes:

Test Group	Tissue No.	Well 1 [OD570 ]	Well 2 [OD570 ]	Mean [OD570]	Mean [OD570] blank corr. (Well 1 and well 2)	Mean [OD570] of T1 and T2	Tissue viabil. [%]	Viabil. of T1 and T2 [%]	Diff. of viabil. between T1 and T2 [p.p.]
Blank		0.037	0.037	0.037
Negative Control	1	2.478 2.333	2.400 2.249	2.439 2.291	2.402 2.254	2.328	100.0	103.2 96.8	6.37
Positive Control	2	0.550 0.564	0.538 0.571	0.544 0.567	0.507 0.531	0.51	22.28	21.8 22.8	1.01
Test Item	1	2.155 1.973	2.156 1.959	2.155 1.966	2.118 1.929	2.024	86.9	91.0 82.9	8.13

[p.p]=Percentage Points

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, it can be stated that in this study and under the experimental conditions reported, Dimethyl Octenyl Acetate (DMOE-Ac) does not need to be classified according UN GHS.

Executive summary:

This in vitro study was performed to assess the eye irritation potential of Dimethyl Octenyl Acetate (DMOE-Ac, CAS: 68345-17-5) by means of the Human Cornea Model Test. The study was performed according to OECD 492 guideline under GLP. 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 50 µL of the test item, the negative control (deionised water) or the positive control (methyl acetate) were applied to each of duplicate tissue for 30 minutes. The mean OD of the tissue replicates treated with the negative control was > 0.8 and < 2.8, thus showing the quality of the tissues. Treatment with the positive control induced a decrease below 50% viability compared with the negative control value in the relative absorbance, thus ensuring the validity of the test system. The difference of relative viability between the two relating tissues was < 20 p.p. in the same run (for test item tissues, positive and negative control tissues). After treatment with the test item Dimethyl Octenyl Acetate (DMOE-Ac) the mean relative viability value decreased to 86.93% compared to the mean value of the negative control. This value is above the threshold for irritancy of ≤ 60%. Therefore, the test item does not need to be classified according UN GHS. In conclusion, it can be stated that in this study and under the experimental conditions reported, Dimethyl Octenyl Acetate (DMOE-Ac) does not need to be classified according UN GHS

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

This in vitro study according to OECD Guideline 439 was performed to assess the irritation potential of Dimethyl Octenyl Acetate (DMOE-Ac) by means of the Human Skin Model Test. The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Also its intrinsic colour was not intensive. Consequently, additional tests with freeze-killed or viable tissues were not necessary. Each three tissues of the human skin model EpiDerm™ were treated with the test item, the negative or the positive control for 60 minutes. 30 µL of the test item were applied to each tissue, and spread to match the surface of the tissue. 30 µL of either the negative control (DPBS) or the positive control (5% SLS) were applied to each tissue. After treatment with the negative control the absorbance values were well in 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 sufficient decrease in the relative absorbance as compared to the negative control for the 60 minutes treatment interval, and thus assuring the validity of the test system. After treatment with the test item the mean relative absorbance value was 112.5% 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. In conclusion, it can be stated that in this study and under the experimental conditions reported, Dimethyl Octenyl Acetate (DMOE-Ac) is not irritant to skin.

 

 

Eye irritation

Eye irritation potential of Dimethyl Octenyl Acetate (DMOE-Ac) was evaluated by means of a weight of evidence approach based on two in vitro assays.

 

The first in vitro study was performed to assess the corneal damage potential of DMOE-Ac by means of the BCOP assay according to 437 using fresh bovine corneae.
After a first opacity measurement of the fresh bovine corneae (t0), the neat test item, the positive, and the negative controls were applied to corneae fixed in an incubation chamber in horizontal position for 10 minutes at 32 ± 1 °C. The posterior chamber contained incubation medium. After the incubation phase the test item, the positive, and the negative controls were each rinsed from the corneae. Further, the corneae were incubated for another 120 minutes at 32 ± 1 °C in a vertical position, while the anterior chamber contain incubation medium as well. Afterwards, opacity was measured a second time (t130).
After the opacity measurements permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein after incubation in a horizontal position for 90 minutes at 32 ± 1 °C.
With the negative control (0.9% (w/v) NaCl solution in deionised water) neither an increase of opacity nor permeability of the corneae could be observed.
The positive control (2-Ethoxyethanol) showed a clear increase in opacity and distinctive increased permeability of the corneae corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)).
Relative to the negative control, the test item DMOE-Ac caused a slight increase of the corneal opacity but no relevant changes in permeability. The calculated mean in vitro irritancy score was 3.85. According to OECD 437 the test item is not classified as serious eye damaging (CLP/EPA/GHS (Cat 1) but the test item’s hazard for eye damaging cannot be predicted.

As a mean in vitro irritancy score ≤ 3 and a consequent classification to "No Category (according to GHS)" was not achieved, by means of a tiered approach a second in vitro assay was conducted to attain a classification for eye irration.

 

The second in vitro assay was conducted by means of the Human Cornea Model Test. The study was performed according to OECD 492 guideline under GLP. 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 50 µL of the test item, the negative control (deionised water) or the positive control (methyl acetate) were applied to each of duplicate tissue for 30 minutes. The mean OD of the tissue replicates treated with the negative control was > 0.8 and < 2.8, thus showing the quality of the tissues. Treatment with the positive control induced a decrease below 50% viability compared with the negative control value in the relative absorbance, thus ensuring the validity of the test system. The difference of relative viability between the two relating tissues was < 20 p.p. in the same run (for test item tissues, positive and negative control tissues). After treatment with DMOE-Ac the mean relative viability value decreased to 86.93% compared to the mean value of the negative control. This value is above the threshold for irritancy of ≤ 60%. Therefore, the test item does not need to be classified according UN GHS. In conclusion, it can be stated that in this study and under the experimental conditions reported, DMOE-Ac does not need to be classified according to CLP.

 

In conclusion and based on the experimental results outlined above, DMOE-Ac is expected not to pose eye irritation hazard and does not need to be classified according to CLP.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008

Based on the results obtained from an in vitro skin irritation study according to OECD 439, DMOE-Acetate is not classified for skin irritation according to Regulation (EC) No 1272/2008, as amended for the seventeenth time in Regulation (EU) No 2021/849.

 

Also, DMOE-Acetate was tested for its potential to cause eye irrtation in two in vitro studies according to OECD 492 and 437 and did not cause a positive response. Therefore, it does not need to be classified according to Regulation (EC) No 1272/2008, as amended for the seventeenth time in Regulation (EU) No 2021/849.