Fatty acids, C18-unsatd., phosphates

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation, other

Remarks:

expert report

Type of information:

other: expert report

Adequacy of study:

supporting study

Study period:

01 July 2017 - 06 July 2017

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

other:

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

GLP compliance:

no

Type of study:

other:

Test material

Test material information

Constituent 1

Reference substance name:

9-(dimethoxyphosphoryl)octadecanoic acid 10-(dimethoxyphosphoryl)octadecanoic acid

Molecular formula:

C20H41O5P

IUPAC Name:

9-(dimethoxyphosphoryl)octadecanoic acid 10-(dimethoxyphosphoryl)octadecanoic acid

Constituent 2

Reference substance name:

9,12-bis(dimethoxyphosphoryl)octadecanoic acid 9,13-bis(dimethoxyphosphoryl)octadecanoic acid 10,12-bis(dimethoxyphosphoryl)octadecanoic acid 10,13-bis(dimethoxyphosphoryl)octadecanoic acid

Molecular formula:

C22H46O8P2

IUPAC Name:

9,12-bis(dimethoxyphosphoryl)octadecanoic acid 9,13-bis(dimethoxyphosphoryl)octadecanoic acid 10,12-bis(dimethoxyphosphoryl)octadecanoic acid 10,13-bis(dimethoxyphosphoryl)octadecanoic acid

Constituent 3

Reference substance name:

Oleic acid

EC Number:

204-007-1

EC Name:

Oleic acid

Cas Number:

112-80-1

Molecular formula:

C18H34O2

IUPAC Name:

octadec-9-enoic acid

Constituent 4

Reference substance name:

Linoleic acid

EC Number:

200-470-9

EC Name:

Linoleic acid

Cas Number:

60-33-3

Molecular formula:

C18H32O2

IUPAC Name:

octadeca-9,12-dienoic acid

Constituent 5

Reference substance name:

Dimethyl (2-oxopropyl)phosphonate

EC Number:

224-110-5

EC Name:

Dimethyl (2-oxopropyl)phosphonate

Cas Number:

4202-14-6

Molecular formula:

C5H11O4P

IUPAC Name:

dimethyl (2-oxopropyl)phosphonate

Constituent 6

Reference substance name:

Dimethyl propylphosphonate

EC Number:

242-555-3

EC Name:

Dimethyl propylphosphonate

Cas Number:

18755-43-6

Molecular formula:

C5H13O3P

IUPAC Name:

dimethyl propylphosphonate

Constituent 7

Reference substance name:

Dimethyl methylphosphonate

EC Number:

212-052-3

EC Name:

Dimethyl methylphosphonate

Cas Number:

756-79-6

Molecular formula:

C3H9O3P

IUPAC Name:

dimethyl methylphosphonate

Test material form:

liquid

Specific details on test material used for the study:

Identification: Fatty acids, C18-unsatd., phosphates.
Appearance: Yellow liquid
Batch: 0101891886
Purity/Composition: 100% UVCB
Test item storage: At room temperature
Stable under storage conditions until: 02 November 2017 (retest date)
Test Facility test item number: 208073/A
Chemical name (IUPAC, synonym or trade name: Phosphorylated fatty acid
CAS number: 68604-99-9
pH (1% in water, indicative range): 2.28 – 2.21 (determined by Charles RiverDen Bosch)
Specific gravity / density: 1.04

Results and discussion

Any other information on results incl. tables

A valid KeratinoSensTM assay was performed according to OECD 442D and GLP (Westerink, 2017). For the KeratinoSensTM assay Fatty acids, C18-unsatd., phosphates was dissolved in DMSO to a final concentration of 40 mg/mL. The 100-fold dilution in DMEM of 40 mg/mL formed a non-homogeneous solution and was therefore not suitable to test. The 100-fold dilution of the 20 mg/mL DMSO stock formed a homogeneous solution (slight precipitation). This concentration was selected as highest concentration for the assays. No precipitate was observed in the assays at the start or end of the treatment period. Due to toxicity, the test concentrations used for the second experiment were in the range of0.008 – 15.63 μM.

Three independent experiments were performed. The test item showed toxicity with an IC30 of 73.7 μg/mL and an IC50 of 107.8 μg/mL in experiment 1, an IC30 of 69.8 μg/mL and IC50 value of 79.1 μg/mL in experiment 2, and an IC30 of 41.0 μg/mL and IC50 value of 54.1 μg/mL in experiment 3. In the first experiment, no induction of the luciferase activity (no EC1.5 value) was measured. The maximum luciferase activity induction (Imax) was 1.26 -fold.

In the second experiment, a statistically significant induction of the luciferase activity (EC1.5 value 26.7 μg/mL; p<0.001 Student’s t test) was measured. The maximum luciferase activity

induction (Imax) was 4.01-fold at 50 μg/mL. The test item induced the luciferase activity very close to cytotoxic dose levels. Therefore, the test item was retested with more narrow doseresponse

analysis using a lower dilution factor (1.25 fold dilution between wells), to determine if induction has occurred at cytotoxic levels or not.

In the third experiment, a statistically significant induction of the luciferase activity (EC1.5 value 38.5 μg/mL; p<0.001 Student’s t test) was measured. The luciferase activity inductions were 1.70- and 1.90 fold at 41.9 μg/mL and 52.4 μg/mL, respectively. However, these inductions are observed at cytotoxic dose levels with a cellular viability < 70% (69.6% and 55.0% at 41.9 μg/mL and 52.4 μg/mL, respectively). Therefore, no real positive effect was measured at any of the tested concentrations. Based on the EC1.5 (38.5 μg/mL) and IC30 (41.0 μg/mL) values, the test item is positive in the KeratinoSensTM assay, although the values are very close together.

Overall, the results of the third experiment are not clearly positive or negative and therefore inconclusive. Three experiments were performed all with different conclusions. Due to the fact that induction of the luciferase activity was very close to the cytotoxic levels, it is not possible to make a final conclusion.

Applicant's summary and conclusion

Conclusions:

Fatty acids, C18-unsatd., phosphates was inconclusive in the KeratinoSensTM assay and no conclusion can be drawn from an alternative testing strategy for skin sensitization. Therefore, Fatty acids, C18-unsatd., phosphates was tested in vivo to determine skin sensitization potential. The guinea pig Maximization test was selected since the test item is a fatty acid and the Local Lymph Node Assay as preferred alternative has shown to provide false positive results for fatty acids (Reference 1).

Ref 1. Comparison of the skin sensitizing potential of unsaturated compounds as assessed by the murine local lymph node assay (LLNA) and the guinea pig maximization test (GPMT), Food and chemical Toxicology 46 (2008) 1896–1904

Executive summary:

The objective of this study was to reach an overall conclusion on the endpoint skin sensitization based on the available relevant information, including alternative testing strategy data.

As Fatty acids, C18-unsatd., phosphates is a UVCB, a KeratinoSensTM assay was performed in accordance with the strategy presented in ECHA Guidance on information requirements and chemical safety assessment Chapter R.7a. Since the KeratinoSensTM assay was inconclusive, an in vitro test for key event 3 does not need to be performed, as one in vitro test with a clear negative or positive outcome is not sufficient to decide on skin sensitization of Fatty acids, C18-unsatd., phosphates. Moreover, in vitro testing does not give an indication of skin sensitization potential. In the absence of a reliable and acceptable in vitro test for determination of skin sensitising potential, the potency needs to be determined by means of an in vivo test.

Based on the above, no conclusion can be drawn based on the alternative strategy for skin sensitization of Fatty acids, C18 -unsatd., phosphates and the test item will be tested in vivo.