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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in chemico
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
13 Jun 2014 - 10 Oct 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: Direct peptide binding assay (DPRA) performed as described in Bauch C. et al. (2011), Toxicology in Vitro 25, 1162 – 1168.
Qualifier:
according to
Guideline:
other: OECD: Draft Proposal for a New Guideline on In Vitro Skin Sensitization: Direct Peptide Reactivity Assay (DPRA), assessed on 13 Nov 2013 at http://www.oecd.org
Principles of method if other than guideline:
In the DPRA the reactivity of a test item (=test substance) towards synthetic cysteine (C)- or lysine (K)-containing peptides is evaluated. For this purpose the test substance is incubated with synthetic peptides for 24 hours at room temperature and the remaining non-depleted peptide concentration is determined thereafter by high performance liquid chromatography with gradient elution and UV-detection at 220 nm. The peptide depletion of test-substance incubated samples is compared to the peptide depletion of the NC samples and expressed as relative peptide depletion.
GLP compliance:
yes (incl. certificate)
Type of study:
other: Direct Peptide Reactivity Assay (DPRA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Dichlone
- Physical state: Solid/ yellow
- Storage condition of test material: Room temperature

In chemico test system

Details on study design:
synthetic peptides (Cysteine- (C-) containing peptide: Ac-RFAACAA-COOH; Lysine- (K-) containing peptide: Ac-RFAAKAA-COOH)

Results and discussion

Positive control results:
The peptide depletion by the positive control was 38.86%.

In vitro / in chemico

Results
Parameter:
other: mean peptide depletion in %
Value:
75.68

Any other information on results incl. tables

The mean Cysteine-peptide depletion, caused by the test substance was determined to be 100 %. The mean Lysine-peptide depletion, caused by the test substance was determined to be 51.36 %. Thus, the mean peptide depletion was calculated to be 75.68%. No co-elution of test substance and peptides was noticed. Based on the observed results and applying the prediction model proposed in Gerberick et. al (2007) it was concluded that Dichlone shows a high chemical reactivity in the DPRA under the test conditions chosen.

Applicant's summary and conclusion

Interpretation of results:
other: high chemical reactivity
Executive summary:

The reactivity of Dichlone towards synthetic cysteine (C)- or lysine (K)-containing peptides was evaluated in the Direct Peptide Reactivity Assay (DPRA). For this purpose the test substance was incubated with synthetic peptides for ca. 24 hours at room temperature and the remaining non-depleted peptide concentrations were determined by high performance liquid chromatography (HPLC) with gradient elution and UV-detection at 220 nm. The test substance was prepared at a 100 mM concentration in acetonitrile. Three samples of the test substance were incubated with each peptide in ratios of 1:10 (for C-peptide) or 1:50 (for K-peptide). Additionally triplicates of the concurrent vehicle control (= VC) were incubated with the peptides. Further, a co-elution control was performed in order to detect possible interference of the test substance with the peptides. The samples consisted of the test substance, vehicle and the respective peptide buffer but without peptide. Moreover the samples were analyzed by measuring UV absorbance at 258 nm and the area ratio 220 nm / 258 nm was calculated as a measure of peak purity. The following results were obtained in the DPRA: The test substance was not soluble in one of the vehicles used for the assay. Thus a homogeneous suspension in acetonitrile was used for sample preparation. The samples of the test substance with the peptides were suspensions directly after preparation and after 24 hours incubation time. Thus the samples of both peptides and the co-elution control were centrifuged prior to HPLC analysis. The mean C-peptide depletion, caused by the test substance was determined to be 100.00%. The mean K-peptide depletion, caused by the test substance was determined to be 51.36%. Thus, the mean peptide depletion was calculated to be 75.68%. No co-elution of test substance and peptides was noticed. Based on the observed results and applying the prediction model proposed in Gerberick et. al (2007) it was concluded that Dichlone shows a high chemical reactivity in the DPRA under the test conditions chosen.