Caramel (color)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vitro

Remarks:

Keratinosens

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)

Deviations:

yes

Remarks:

without impact on the study conclusion (experimentation was redone due to a technical error which made results uninterpretable).

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: ECVAM DB-ALM protocol 155: KeratinoSensTM

GLP compliance:

yes

Type of study:

activation of keratinocytes

Justification for non-LLNA method:

Alternative methods are recommended first before performing the LLNA test.

Test material

Test material information

Constituent 1

Reference substance name:

Caramel (color)

EC Number:

232-435-9

EC Name:

Caramel (color)

Cas Number:

8028-89-5

Molecular formula:

C6H12OH

IUPAC Name:

Caramel

In vitro test system

Details on the study design:

Cells (KeratinoSens™) are cultured in maintenance medium at 37°C, 5% CO2. Cells are exempt of mycoplasma. Cells were used at passage 17 in repetition 1 and passage 19 in repetition 2.

Test item
The test item was tested at 12 concentrations according to a geometric progression of ratio 2 from 0.2 μg/ml to 400 μg/ml.

Reference item
Negative control: 6 wells of solvent control (1% DMSO in treatment medium) with cells and 1 well of solvent control without cell by culture plate.
Positive control: 5 concentrations of cinnamaldehyde on each culture plate. The concentration varies from 4 to 64 μM according to a geometric progression of ratio 2.

The study was composed of two independent repetitions. For each repetition the test item and the reference items were replicated on three independent plates for the measurement of induction and two plates for the measurement of cytotoxicity. Each repetition was performed on a different day with fresh stock solution.

Results and discussion

Positive control results:

The positive control (cinnamaldehyde) gave the following results:
-EC1.5 = 7.19 (geometric mean)
-Imax = 6.36

In vitro / in chemico

Results

Results 1

Key result

Run / experiment:

other: repetition 1

Parameter:

other: EC1.5 (ug/mL)

Value:

200

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of skin sensitisation

Results 2

Key result

Run / experiment:

other: repetition 2

Parameter:

other: EC1.5 (ug/mL)

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

not measured/tested

Remarks:

EC1.5 was not calculated since Imax was lower than 1.5

Other effects / acceptance of results:

The positive control (cinnamaldehyde) fulfilled the acceptability criteria of the study:
-the gene induction was statistically significant above the threshold of 1.5 in at least one dose
-the EC1.5 value was between historical data of the laboratory: mean EC1.5 value ± 2 SD
-the average induction, in each repetition, for cinnamaldehyde at 64 μM was between 2 and 8.

The negative control (control solvent) fulfilled the acceptability criteria of the study:
For each repetition, the coefficient of variation (CV) of the solvent controls (3 x 6 wells) was less than 20% (Repetition 1: CV of 16.1 and Repetition 2: CV of 17.3)

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the retained experimental conditions, Caramel E150 C may be classified as not skin sensitizer.
The test method KeratinoSensTM is considered scientifically valid to be used as part of an integrated approaches to testing and assessment, to support the identification of the sensitization potential of test item for hazard classification and labeling purposes.