Prop-2-yn-1-yl 1H-imidazole-1-carboxylate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in chemico

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Report date:

2021

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))

Version / remarks:

adopted 18. Jun. 2019

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EURL ECVAM (European Union Reference Laboratory for alternatives to animal test-ing): “DB-ALM Protocol n° 154: Direct Peptide Reactivity Assay (DPRA) for Skin Sen-sitisation Testing.”

Version / remarks:

29. Jun. 2015

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Commission Regulation (EU) 2017/735, EU-Method B.59 “In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA)”

Version / remarks:

adopted 14 February 2017

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

direct peptide reactivity assay (DPRA)

Test material

Test material information

Constituent 1

Reference substance name:

prop-2-yn-1-yl 1H-imidazole-1-carboxylate

EC Number:

695-595-2

Cas Number:

83395-38-4

Molecular formula:

C7H6N2O2

IUPAC Name:

prop-2-yn-1-yl 1H-imidazole-1-carboxylate

Test material form:

solid

Specific details on test material used for the study:

Name: 2-propyn-1-yl 1H-imidazole-1-carboxylate
Batch no.: 0802-2#
CAS no.: 83395-38-4
Composition: 2-propyn-1-yl 1H-imidazole-1-carboxylate
Storage: fridge (2 - 8 °C) under inert gas
Expiry date: 11. Jul. 2021
Stability : stable under storage conditions
Appearance: white solid
Purity: 99.1 %
Homogeneity: homogeneous
Production date: 13. Oct. 2020
EC no.: 695-595-2
Molecular formula: C7H6N2O2
Molecular weight: 150.13 g/mol

In chemico test system

Details on the study design:

Evaluate method: Evaluate the reactivity of the test item 2-propyn-1-yl 1H-imidazole-1-carboxylate towards cysteine (Cys-) and lysine (Lys-) containing peptides.
Incubate condition: 22 h at 25 °C for the Cys-peptide; 22 h at 25 °C for the Lys-peptide.
The peptide concentration measure method: HPLC-UV.
Replicate: 3
Test item samples: Samples were prepared in triplicate for each peptide. The Cys-peptide samples were pre-pared in 1:10 molar ratio (0.5 mM peptide: 5 mM test item solution), the Lys-peptide sam-ples in 1:50 molar ratio (0.5 mM peptide and 25 mM test item solution) using the stock solutions.
Per sample volume: 1 mL

Results and discussion

Positive control results:

a) The mean peptide depletion with 72.83% and a standard deviation of 0.63% of the three replicates of the positive control cinnamaldehyde were in the acceptable range of 60.8 – 100.0 % and < 14.9 %, respectively, for the Cys-peptide.
b) The mean peptide depletion with 21.45% and a standard deviation of 0.20% of three replicates of the positive control 2,3-Butanedione were in the acceptable range of 10.0 – 45.0 % and < 11.6 %, respectively, for the Lys-peptide.

In vitro / in chemico

Results

Results 1

Key result

Run / experiment:

other: mean of 3 repetitions

Parameter:

other: Cys-peptide depletion [%]

Value:

99.2 %

Vehicle controls validity:

valid

Positive controls validity:

valid

Results 2

Key result

Run / experiment:

other: Mean of 3 repetitions

Parameter:

other: Lys-peptide depletion [%]

Value:

24.96 %

Vehicle controls validity:

valid

Positive controls validity:

valid

Results 3

Key result

Run / experiment:

other: Mean of lysine and cysteine peptides

Parameter:

other: Mean peptide depletion [%]

Value:

62.08 %

Vehicle controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: no

DEMONSTRATION OF TECHNICAL PROFICIENCY: Yes. All 10 proficiency chemicals were correctly predicted.
For the Cys-peptide the depletion values for 9 out of the 10 substances, for the Lys-peptide assay all ten values fall within the respective reference range.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for calibration curve: Yes, coefficient r2 was higher than 0.99 for lysine and cysteine.
- Acceptance criteria met for positive control: Yes, SD of the 3 repetitions for each peptide was 0.63% for Cys-peptide and 0.2% for Lys-peptide which are lower than 14.9% and 11.6% for Cys-peptide and Lys-peptide respectively. The depletion mean rate was 72.83% for Cys-peptide and 21.45% for Lys-peptide which are between 60.8% and 100% for the cysteine and between 10.0% and 45.0% for the Lys-peptide.
- Acceptance criteria met for test item replicates: Yes. The standard deviation for the test item replicates with 0.06% was < 14.9 % for the percent cysteine depletion for the test item. The standard deviation for the test item replicates with 0.15% was < 11.6 % for the percent lysine depletion for the test item.

Any other information on results incl. tables

Table1. Calculated peptide depletion values for the Cys-Peptide

Sample name	Depletion [%]
	Single	Mean	SD
Positive control Rep. 1	72.51	72.83	0.63
Positive control Rep. 2	72.44
Positive control Rep. 3	73.56
Test item Rep. 1	99.13	99.20	0.06
Test item Rep. 2	99.22
Test item Rep. 3	99.24

Table 2. Calculated peptide depletion values for the Lys-Peptide

Sample name	Depletion [%]
	Single	Mean	SD
Positive control Rep. 1	21.24	21.45	0.20
Positive control Rep. 2	21.46
Positive control Rep. 3	21.65
Test item Rep. 1	24.92	24.96	0.15
Test item Rep. 2	24.83
Test item Rep. 3	25.13

Applicant's summary and conclusion

Interpretation of results:

Category 1 (skin sensitising) based on GHS criteria

Conclusions:

The DPRA prediction is ”positive” according to the Cysteine 1:10/Lysine 1:50 predic-tion model. Thus, under the experimental conditions reported, the test item 2-propyn-1-yl 1H-imidazole-1-carboxylate shows a reactivity towards the two model synthetic peptides.
This assignment supports the discrimination between skin sensitisers and non-sensitisers in the framework of an integrated approach (IATA).
For sensitising potency assessment within an IATA, the test item 2-propyn-1-yl 1H-imidazole-1-carboxylate could be assigned to the reactivity class that covers high reactivity under the conditions of this study.

Executive summary:

A DPRA skin sensitization test was performed for 2-propyn-1-yl 1H-imidazole-1-carboxylate as part of an integrated approach to testing and assessment (IATA) in accordance to OECD guideline 442C, under GLP conditions.

This in chemicos tudy was performed in order to evaluate the reactivity of the test item 2-propyn-1-yl 1H-imidazole-1-carboxylate towards cysteine (Cys-) and lysine (Lys-) containing peptides.

The calculated peptide depletion could be usedto support the discrimination between skin sensitisers and non-sensitisers. The DPRA is part of a tiered strategy for the evaluation of skin sensitisation potential in the context ofan integrated approach to testing and assessment (IATA).

 

The test item was incubated for 22 h at 25 °C together with Cys- and Lys-peptides, respectively. The peptide concentration after the incubation period was measured using HPLC-UV.

Three replicates were prepared using 1:10 and 1:50 molar ratio of the test item with the Cys- and Lys-peptides, respectively. Triplicate samples of the solvent without test item were incubated and measured simultaneously.

Under the conditions of the test, the test item 2-propyn-1-yl 1H-imidazole-1-carboxylate could be assigned to the reactivity class that covers high reactivity under the conditions of this study.