trans-1-butyl-4-[trans-4-[(E)-prop-1-enyl]cyclohexyl]cyclohexane

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in chemico

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

June 02 - Nov 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

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Direct Peptide Reactivity Assay (DPRA) for Skin Sensitization Testing, DB-ALM Protocol n°154, January 12, 2013

GLP compliance:

yes (incl. QA statement)

Remarks:

Behörde für Gesundheit und Verbraucherschutz, Freie Handelsstadt Hamburg

Type of study:

direct peptide reactivity assay (DPRA)

Test material

Test material information

Constituent 1

Reference substance name:

trans-1-butyl-4-[trans-4-[(E)-prop-1-enyl]cyclohexyl]cyclohexane

EC Number:

821-932-1

Cas Number:

694510-10-6

Molecular formula:

C19 H34

IUPAC Name:

trans-1-butyl-4-[trans-4-[(E)-prop-1-enyl]cyclohexyl]cyclohexane

Test material form:

solid

In chemico test system

Details on the study design:

The in chemico direct peptide reactivity assay (DPRA) enables detection of the sensitising potential of a test item by quantifying the reactivity of test chemicals towards synthetic peptides containing either lysine or cysteine.

Results and discussion

Positive control results:

The 100 mM stock solution of the positive control (cinnamic aldehyde) showed high reactivity towards the synthetic peptides. The mean depletion of both peptides was 68.60%.

In vitro / in chemico

Results

Results 1

Key result

Run / experiment:

other: cysteine run

Parameter:

other: mean peptide depletion [%]

Value:

0

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Key result

Run / experiment:

other: lysine run

Parameter:

other: mean peptide depletion [%]

Value:

0

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Acceptance Criteria

The following criteria must be met for a run to be considered valid:
a) The standard calibration curve should have an r2 > 0.99.
b) The mean percent peptide depletion value of the three replicates for the positive control cinnamic aldehyde should be between 60.8% and 100% for the cysteine peptide and between 40.2% and 69.0% for the lysine peptide and the maximum standard deviation (SD) for the positive control replicates should be < 14.9% for the percent cysteine depletion and < 11.6% for the percent lysine depletion.
c) The mean peptide concentration of reference controls A should be 0.50 ± 0.05 mM and the coefficient of variation (CV) of peptide peak areas for the nine reference controls B and C in acetonitrile should be <15.0%.
If one or more of these criteria is not met, the run would have been repeated.

The following criteria must be met for a test item’s results to be considered valid:
a) The maximum standard deviation for the test item replicates should be < 14.9% for the percent cysteine depletion and < 11.6% for the percent lysine depletion.
b) The mean peptide concentration of the three reference controls C in the appropriate solvent should be 0.50 ± 0.05 mM.

If these criteria were not met, the data would have been rejected and the run have been repeated for that specific test item.

Any other information on results incl. tables

Peptide Depletion in %

1) Cystein Depletion

Sample	Peak Area	Actual Peptide [mM]	Peak Are Ref Control C	Peptide Depletion [%]	Corr. peptide Depletion [%]
1	50.9758	0.503	50.8281	-0.03	0.00
2	51.5350	0.509	51.0861	-1.13	0.00
3	51.5140	0.508	50.9661	-1.09	0.00
Mean	51.342	0.507	50.9601	-0.75	0.00
Evaluation Reactivity Class. Negative

1) Lysine Depletion

Sample	Peak Area	Actual Peptide [mM]	Peak Are Ref Control C	Peptide Depletion [%]	Corr. peptide Depletion [%]
1	36.3497	0.464	35.0532	-2.95	0.00
2	36.4728	0.465	35.3563	-3.29	0.00
3	36.5649	0.466	35.5197	-3.55	0.00
Mean	36.462	0.465	35.3097	-3.26	0.00
Evaluation Reactivity Class. Negative

Applicant's summary and conclusion

Interpretation of results:

other: The result has to be considered in an integrated approach.

Conclusions:

The test material revealed a mean cysteine and lysine peptide depletion of 0.0% and, hence, the test item is considered negative and predicted to be a non-sensitiser (no or minimal reactivity) in the Direct Peptide Reactivity Assay (DPRA).

Executive summary:

Pupose

The purpose of this study was to determine the sensitising potential of the test material in a Direct Peptide Reactivity Assay (DPRA).

Methods

The study was performed according to OECD guideline 442C. The DPRA is an in chemico method which quantifies the remaining concentration of cysteine- or lysine-containing peptide following incubation with the test item. Cysteine and lysine peptide percent depletion values are then calculated and used in a prediction model, which allows assigning the test item to one of four reactivity classes used to support the discrimination between sensitisers and nonsensitisers.

Results

The test item was dissolved at a concentration of 100 mM in acetone. No precipitate in the reaction mixture at the end of the incubation time and no co-elution were observed.

The test material-treated samples revealed a cysteine peptide depletion of 0.0% and a lysine peptide depletion of 0.0% (mean peptide depletion of 0.0%) and, hence, were well

below 6.38%. The test material is considered negative and predicted to be a nonsensitiser (no or minimal reactivity) in the Direct Peptide Reactivity Assay (DPRA).

Cinnamic aldehyde was used as positive control at a concentration of 100 mM in acetone. Treatment with the positive control item revealed a cysteine and lysine peptide depletion of

68.65% for cysteine and 59.73% for lysine peptide. These values are within the required range. Therefore, the study can be regarded as valid.

Conclusion

The test material revealed a mean cysteine and lysine peptide depletion of 0.0% and, hence, the test item is considered negative and predicted to be a non-sensitiser (no or

minimal reactivity) in the Direct Peptide Reactivity Assay (DPRA).