Dimethyl heptanedioate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 Sepember 2017 - 08 December 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study conducted according to OECD test Guideline No. 492 without any deviation.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

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:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

French GLP Compliance Programme for chemical products (inspected on 30-31 January 2017 / signed on 27 April 2017)

Test material

Test material information

Constituent 1

Reference substance name:

Dimethyl heptanedioate

EC Number:

217-057-4

EC Name:

Dimethyl heptanedioate

Cas Number:

1732-08-7

Molecular formula:

C9H16O4

IUPAC Name:

1,7-dimethyl heptanedioate

Test material form:

liquid

Details on test material:

Name: Dimethyl Pimelate
Batch no.: 1601070000
Appearance: Oily liquid, colorless
CAS No.: 1732-08-7
EINECS-No.: 217-057-4
Molecular formula: C9H16O4
Purity: 98.54 % w/w
Production date: 04 Jul. 2016
Expiry date: 01. Aug. 2019
Storage: Fridge (2-8°C), keep away from humidity

Test animals / tissue source

Species:

other: Reconstructed human cornea-like epithelium tissues (EpiOcular™ tissue model)

Strain:

other: Not applicable

Details on test animals or tissues and environmental conditions:

- Justification of the test method and considerations regarding applicability: The EpiOcular™ Eye Irritation Test (EIT) was validated by the European Union Reference Laboratory for Alternatives to Animal Testing (EURL ECVAM) and Cosmetics Europe between 2008 and 2013. This test guideline is applicable to liquids and semi-solids, so is considered to be applicable to the test item.

CELL SYSTEM:
- EpiOcular™ OCL-212-ver2.0, supplied by MatTek Corporation (Bratislava, Slovakia).
- Lot No.: 27006
- Keratinocyte strain: 4F1188
- Analysis for potential biological contaminants: none detected (HIV-1, Hep. B and Hep. C virus; Bacteria, yeast and other fungi)
- Tissue viability: 1.534, within the acceptance criteria (1.1-3.0)
- Barrier function: 17.77 min, within the acceptance criteria (12.2-37.5)
- Sterility: Sterile
- Transport: Not specified
- Storage: Not specified

Test system

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): not applicable

VEHICLE: Not applicable

Duration of treatment / exposure:

30 minutes

Observation period (in vivo):

Not applicable

Duration of post- treatment incubation (in vitro):

- 12-minute immersion period at room temperature
- 2 hour and 14 minutes post-exposure incubation at 37°C, 5% CO2

Number of animals or in vitro replicates:

2 replicates

Details on study design:

- Details of the test procedure used : procedure for liquids
* Pre-treatment: after an overnight incubation, tissues were pre-wetted with 20 µL of Ca2+Mg2+Free-DPBS. Then tissues were incubated for 30 minutes at standard culture conditions.
* Treatment: 50 µL of test item, positive or negative control was applied to the entire surface of 2 living RhCE tissue replicates during 30 minutes at standard culture conditions.
* Post-exposure incubation period: after treatment, tissues were carefully washed by extensive rinsing with Ca2+Mg2+Free-DPBS. Rinsed tissues were checked for any coloration and for comparable colour with negative control treated tissues (whitish). The rinsing step was followed by a 12-minute immersion period at room temperature in 5 mL of fresh medium to remove any test item absorbed into the tissue. Then the RhCE constructs were incubated for 2 hours and 14 minutes post-exposure incubation at standard culture conditions in 1 mL of fresh medium at 37°C, 5% CO2.

- Doses of test chemical and control substances used : 50 µL

- Duration and temperature of pre-treatment (30 minutes), exposure (30 minutes), post-exposure immersion (12 minutes) and post-exposure incubation (2 hour and 14 minutes)

- Description of any modifications to the test procedure : None

- Indication of controls used for direct MTT-reducers and/or colouring test chemicals (if applicable) : Since the test item did not interact with MTT, none killed control tissue models were added to the study. The test item did not show any coloration potential in presence of water or isopropanol, thus no non-specific coloration control was added to the study.

- Number of tissue replicates used per test chemical and controls (positive control, negative control): 2

- Description of evaluation criteria used including the justification for the selection of the cut-off point for the prediction model :
If the test item-treated tissue viability is > 60% relative to the negative control treated tissue viability, the test item is labeled non-irritant.
If the test item-treated tissue viability is ≤ 60% relative to negative control treated tissue viability, the test item is identified potentially requiring classification and labelling according to UN GHS (Category 2 or Category 1).
According to OECD guideline 492 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, attached to the study report.
* Historical negative control ranges 83.17 - 116.83%
* Historical positive control ranges 6.50 - 45.11%

- Complete supporting information for the specific RhCE tissue construct used : Yes, attached to the study report.

- Reference to historical data of the RhCE tissue construct : No

- Demonstration of proficiency in performing the test method before routine use by testing of the proficiency chemicals : Yes, attached to the study report.

- Positive and negative control means and acceptance ranges based on historical data : Yes

- Acceptable variability between tissue replicates for positive and negative controls: Yes

- Acceptable variability between tissue replicates for the test chemical: Yes

Results and discussion

In vitro

Results

Irritation parameter:

other: % Tissue viability

Remarks:

mean corrected percent viability

Run / experiment:

1

Value:

68.16

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

100.00%

Positive controls validity:

valid

Remarks:

43.31%

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: Not specified

DEMONSTRATION OF TECHNICAL PROFICIENCY: Yes

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes, OD values included between 0.848 and 1.169
- Acceptance criteria met for positive control: % viability= 43.31%
- Range of historical values if different from the ones specified in the test guideline:
- Negative control: OD values [0.645-1.563]
- Positive control: % viability [6.50-45.11]

Any other information on results incl. tables

Table 7.3.2/1: Mean OD₅₇₀Values and Percentage Viabilities for the Negative Control Item, Positive Control Item and Test Item

	Tissue	OD	Mean OD / disc (#)	Mean OD / product	Viability %	Mean viability %	Difference of viability %	Conclusion
Negative control	1	0.849 0.848 0.856	0.851	1.002	84.93	100.00	30.14
	2	1.169 1.135 1.157	1.153		115.07
Positive control	3	0.405 0.399 0.399	0.401	0.434	40.02	43.31	6.59	UN GHS Category 2 or 1
	4	0.467 0.463 0.471	0.467		46.61
Test item	5	0.707 0.684 0.688	0.693	0.683	69.16	68.16	2.00	No Category
	6	0.667 0.623 0.729	0.673		67.17

Note: the optical density was measured after a 1:2 dilution of the formazan extracts in isopropanol.

 

 

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

OD: optical density

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the experimental conditions adopted and in accordance with Regulation EC No. 1272/2008, test item, Dimethyl Pimelate, does not require classification for eye irritation or serious eye damage according to CLP and UN-GHS regulations.

Executive summary:

A study was performed to assess the eye irritation potential of the test item by means of the Human Cornea Model Test. The study was conducted according to the OECD guideline No. 492 and in compliance with GLP.

The test item, Dimethyl Pimelate, was applied as supplied at the dose of 50 µL to 2 living DPBS pre-treated RhCE (EpiOcular^TM 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 and 14 minutes post-exposure incubation at standard culture conditions. The tissue viability was measured by performing an MTT assay.

The mean percent tissue viability of the RhCE replicates treated with the test item Dimethyl Pimelate was 68.16 %, versus 43.31% in the positive control (Methyl acetate).

Under the experimental conditions adopted and in accordance with Regulation EC No. 1272/2008, test item Dimethyl Pimelate does not require classification for eye irritation or serious eye damage according to CLP and UN-GHS regulations.