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Diss Factsheets

Toxicological information

Skin sensitisation

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Administrative data

skin sensitisation: in vitro
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 442C (In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA))
GLP compliance:
yes (incl. QA statement)
Type of study:
direct peptide reactivity assay (DPRA)
Justification for non-LLNA method:
The direct peptide reactivity assay (DPRA) is an in chemico assay to quantify the reactivity of the test item towards cysteine and lysine containing peptides. This reactivity is related to the skin sensitisation potential.
To quantify the sensitisation potential, the depletion of the cysteine and lysine containing peptides caused by known amounts of the test item is measured using HPLC.
The assay is used for supporting the discrimination between skin sensitizers (i.e. UN GHS Category 1) and non-sensitizers in accordance with the UN GHS. A categorization in the sub-categories 1 A and 1 B is not possible.

Test material

Constituent 1
Chemical structure
Reference substance name:
Lithium tetrafluoroborate, anhydrous
EC Number:
EC Name:
Lithium tetrafluoroborate, anhydrous
Cas Number:
Molecular formula:
lithium tetrafluoroborate, anhydrous
Test material form:
solid: particulate/powder

In vitro test system

Details on the study design:
6.2.1 HPLC system
Designation: HPLC_4
Components: Degasser G1322A
Quaternary pump G1311A
Autosampler G1313A
Column compartment G1316A
UV/VIS-Detector DAD G1315A
Manufacturer: Agilent Technologies
Software: CHROMELEON 6.80 SR15b Build 4981
Usage and calibration followed the corresponding SOP 114 00 526 in the current edition.
6.2.2 Column
An ACE Excel SuperC18 150x3 mm column with 3 µm particles and pre-column Phe-nomenex SecurityGuard C18, 4x3 mm was used. This column was selected because it delivers substantially better peak shape for the peptides than the Agilent Zorbax SB-C18 column recommended in the OECD 442C guideline.
6.2.3 HPLC program
Eluent A H2O + 0.1 % TFA
Eluent B Acetonitrile + 0.085 % TFA
Gradient time (min) % A % B
0 90 10
10 75 25
10.5 10 90
12 10 90
13 90 10
20 90 10
Flow rate 0.55 mL/min
Injection volume 7 µL
Column temperature 30 °C
Wavelength 1 220 nm
Wavelength 2 258 nm
6.3 Test System
Peptides with ≥ 95 % purity, synthesized by Genecust, Dudelange, Luxemburg, were used.
6.3.1 Cys-Peptide (Cysteine)
Sequence: Ac-RFAACAA-COOH (MW = 750.9 g/mol)
Batch no.:P170415-2-LR569638
Purity: 98.10%
6.3.2 Lys-Peptide (Lysine)
Sequence: Ac-RFAAKAA-COOH (MW = 775.9 g/mol)
Batch no.: P170415-2-LR569640
Purity: 98.85%

Results and discussion

Positive control results:
Historical data of Lys-peptide depletion values of valid tests support the acceptance crite-ria for the proficiency chemical 2,3-butanedione defined in the guideline (mean depletion of the Lys peptide of 10 – 45 %). Therefore, the usage as positive control with the same acceptance criteria is reasonable.

In vitro / in chemico

Resultsopen allclose all
Run / experiment:
other: 1
other: Peptide depletion[%]
Vehicle controls validity:
not specified
Negative controls validity:
not specified
Positive controls validity:
Run / experiment:
other: 2
other: Peptide depletion[%]
Vehicle controls validity:
not specified
Negative controls validity:
not specified
Positive controls validity:
Other effects / acceptance of results:
DPRA Prediction: Negative
Reactivity class: Minimal

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
All acceptance criteria were fulfilled.Therefore, the test was considered valid. The DPRA prediction for the test item Lithium Tetrafluoroborate was negative with reactivity class minimal according to the Cysteine 1:10/Lysine 1:50 prediction model. The result was verified in a second experiment.