Ethoxycarbonylmethyl ethyl phthalate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in chemico

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07. Feb. 2019 - 09. Apr. 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 442C (In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA))

Deviations:

no

GLP compliance:

yes

Type of study:

direct peptide reactivity assay (DPRA)

Justification for non-LLNA method:

This in chemico test method was conducted to investigate skin sensitisation potential of the test item. The information needed for the classification or risk assessment of a substance has to be obtained through non-animal methods as a first step.
Non-animal methods are the default requirement.

Test material

Test material information

Constituent 1

Reference substance name:

Ethoxycarbonylmethyl ethyl phthalate

EC Number:

201-555-3

EC Name:

Ethoxycarbonylmethyl ethyl phthalate

Cas Number:

84-72-0

Molecular formula:

C14H16O6

IUPAC Name:

ethoxycarbonylmethyl ethyl phthalate

Test material form:

liquid

In chemico test system

Details on the study design:

To quantify the sensitisation potential, the depletion of the cysteine and lysine containing peptides caused by known amounts of the test item was measured using HPLC. The test item solution in acetonitrile and the respective peptide is incubated 22 h at 25 °C together with Cys- and Lys-peptides, respectively. The peptide concentration after the in-cubation is measured using HPLC-UV.

Three replicates are prepared using 1:10 and 1:50 molar ratio of the test item with the Cys- and Lys-peptide, respectively. Triplicate samples of the solvent without test item are incubated and measured simultaneously.

Results and discussion

Positive control results:

The mean peptide depletion with 100.0 % and the standard deviation of 0.00 % of the three replicates of the positive control cinnamaldehyde were in the acceptable range of 60.8 – 100.0 % and < 14.9 %, respectively, for the Cys-peptide.
As cinnamaldehyde mixed with the lysine peptide turned turbid in all experiments performed during the implementation phase, it was considered unsuitable as positive control. Instead, the proficiency chemical 2,3-Butanedione is used as positive control showing mid-range depletion for the lysine peptide.
The percent mean area ratio at 220/258 nm of the positive control 2,3-Butanedione in the Lys-peptide assay was in the given range of 90-110 %.
The mean peptide depletion with 16.56 % and the standard deviation of 0.77 % of the three replicates of the positive control 2,3-Butanedione were in the acceptable range of 10.0 – 45.0 % and < 11.6 %, respectively, for the Lys-peptide.

In vitro / in chemico

Results

Results 1

Key result

Run / experiment:

other: 1

Parameter:

other: mean peptide depletion [%]

Value:

0.59

Vehicle controls validity:

valid

Positive controls validity:

valid

Results 2

Run / experiment:

other: 1

Parameter:

other: Cys-peptide depletion [%]

Value:

0.34

Vehicle controls validity:

valid

Positive controls validity:

valid

Results 3

Run / experiment:

other: 1

Parameter:

other: Lys-peptide depletion [%]

Value:

0.84

Vehicle controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

All acceptance criteria were fulfilled.

Acceptance criteria:
The r² of linear calibration should be > 0.99.
The mean peptide concentration of solvent control samples of sets A and C should be 0.50 ± 0.05 mM
The variation coefficient (relative standard deviation, RSD) of measured values of the nine samples from sets B1, B2 and C should be < 15 %
The mean peptide depletion value for the positive control cinnamaldehyde should be 60.8 % - 100.0 % with a maximum standard deviation (SD) of < 14.9 % for the Cys-peptide.
The mean peptide depletion value for the positive control 2,3-butanedione should be 10.0 % - 45.0 % with a maximum standard deviation < 11.6 % for the Lys-peptide.
The standard deviation for the test item replicates should be < 14.9 % for the percent cysteine depletion and < 11.6 % for the percent lysine depletion.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The DPRA prediction is “negative” with no or minimal reactivity according to the Cysteine 1:10/Lysine 1:50 prediction model. It can be stated that in this study and under the experimental conditions reported, the test item Ethylphthalyl ethyl glycolate possesses no or minimal skin sensitisation potential.

Executive summary:

The study was performed in order to evaluate the reactivity of the test item Ethylphthalyl ethyl glycolate towards cysteine (Cys-) and lysine (Lys-) containing peptides. A test item solution in acetonitrile was incubated 22 h at 25 °C together with Cys- and Lys-peptides, respectively. The peptide concentration after the incubation was measured using HPLC-UV.

Three replicates were prepared using 1:10 and 1:50 molar ratio of the test item with the Cys- and Lys-peptide, respectively. Triplicate samples of the solvent without test item were incubated and measured simultaneously.

One experiment was performed.

Experiment 1 was valid for both peptide assays and the results are reported here.

The percent mean area ratio at 220/258 nm of the positive control 2,3-Butanedione in the Lys-peptide assay was in the given range of 90-110 %.

The mean peptide depletion [%] after incubation was 0.59%.

In conclusion, the DPRA prediction is “negative” with no or minimal reactivity according to the Cysteine 1:10/Lysine 1:50 prediction model. It can be stated that in this study and under the experimental conditions reported, the test item Ethylphthalyl ethyl glycolate possesses no or minimal skin sensitisation potential.