1-ethyl-3-methyl-3H-imidazolium dicyanidoamide(1-)

Endpoint summary

Description of key information

The test substance was not irritant or corrosive to the skin in a GLP-compliant OECD 431 and 439 study.

The test substance was not irritant to the eyes in a GLP-compliant OECD 492 and 437 study.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

GLP compliance:

yes

Specific details on test material used for the study:

- Name of the test substance (as cited in the study report): EMIM Dicyanamid
- Purity: 97.4g / 100 g
- pH value: ca. 5 (undiluted test substance)

Test system:

human skin model

Source species:

human

Amount/concentration applied:

30 µL (irritation test); 50 µL (corrosion test)

Duration of treatment / exposure:

- Irritation test: 1 hour followed by a 42-hours post-incubation period
- Corrosion test: 3 minutes and 1 hour, respectively

Number of replicates:

- Irritation test: 3 tissues
- Corrosion test: 2 tissues

Vehicle:

unchanged (no vehicle)

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean

Run / experiment:

1 h exposure, 42 h post-incubation (Irritation test)

Value:

102

Negative controls validity:

valid

Remarks:

PBS

Positive controls validity:

valid

Remarks:

SDS

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean

Run / experiment:

3 min exposure (Corrosion test)

Value:

88

Negative controls validity:

valid

Remarks:

H2O

Positive controls validity:

valid

Remarks:

KOH

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean

Run / experiment:

1 h exposure (Corrosion test)

Value:

73

Negative controls validity:

valid

Remarks:

H2O

Positive controls validity:

valid

Remarks:

KOH

Remarks on result:

no indication of irritation

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Version / remarks:

July, 2015

Principles of method if other than guideline:

according to:
- MatTek Corporation, Ashland, MA 01721, USA: EpiOcularTM human cell construct: Procedure details, Version 3.1a of February 10, 2010.
- Harbell J.W. et al. (2009): COLIPA Program on Optimization of Existing In Vitro Eye Irritation Assays for Entry into Formal Validation: Technology Transfer and Intra/Inter Laboratory Evaluation of EpiOcular Assay for Chemicals. Poster # 378, Society of Toxicology, March 2009.

GLP compliance:

yes

Specific details on test material used for the study:

- Name of the test substance (as cited in study report): EMIM Dicyanamid
- Purity: 97.4 g/100 g
- pH value: ca. 5 (undiluted test substance)

Species:

other: reconstructed human cornea model

Vehicle:

unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

50 uL

Duration of treatment / exposure:

30 min

Duration of post- treatment incubation (in vitro):

2-hour post-incubation period

Number of animals or in vitro replicates:

Two tissues (test group)

Details on study design:

To remove the test substance, the tissues were washed with sterile PBS. For this purpose the tissues were immersed and swiveled three times in each of three beakers filled with PBS. Washed tissues were immediately immersed into 12-well plates, pre-filled with 5 mL/well prewarmed medium (post-soak immersion) in order to remove residual test substance. Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the test substance treated epidermal tissues is compared to that of negative control tissues. The quotient of the values indicates the relative tissue viability. After 12 minutes of post-soak immersion, each tissue was dried on absorbent paper and transferred to fresh 6-well plates filled with 1 mL/well pre-warmed medium. Subsequently, the tissues were incubated at standard culture conditions for 2 hours (liquids) (postincubation period). The irritation potential of the test materials is predicted from the mean relative tissue viabilities compared to the negative control tissues concurrently treated with sterile water. A chemical is considered as "irritant", if the mean relative tissue viability with a test material is less than or equal to 50%. According to the new OECD guideline 492 (July 28th, 2015), the percentage tissue viability cut-off value distinguishing classified from non-classified test chemicals is 60%.

Irritation parameter:

other: mean viability in %

Value:

69

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

H2O

Positive controls validity:

valid

Remarks:

Methyl acetate

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

The test substance is not able to reduce MTT directly.

Interpretation of results:

GHS criteria not met

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

GLP compliance:

yes

Specific details on test material used for the study:

- Name of the test substance(as cited in the study report): EMIM Dicyanamid
- Purity: 97.4 g/100 g
- pH value: ca. 5 (undiluted test substance)

Species:

other: isolated bovine cornea

Details on test animals or tissues and environmental conditions:

Bovine eyes are obtained as a by-product of freshly slaughtered cattle (age of the animals: minimum 12 months, maximum 60 months).

Vehicle:

unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

750 uL of the undiluted liquid test substance was applied into the anterior chamber using a pipette

Duration of treatment / exposure:

10 min

Observation period (in vivo):

2-hour post-incubation period

Number of animals or in vitro replicates:

Three bovine corneas

Details on study design:

Corneas free of defects (opacity, scratches, pigmentation etc.) were dissected with a 2 to 3 mm rim of sclera. Isolated corneas were mounted in specially designed corneal holders that consists of anterior and posterior chambers. Both chambers were filled to excess with prewarmed Eagles’s MEM (without phenol red) and then equilibrated in a vertical position at about 32°C for at least 1 hour. After the equilibration period the medium in both chambers was replaced with fresh prewarmed medium and initial corneal opacity readings were taken for each cornea with an opacitometer. Any corneas that show macroscopic tissue damage or an opacity value < 507 opacity units were discarded. Corneas with opacity values close to the median value of all corneas were selected as negative control (NC). The remaining corneas were then distributed into treatment and positive control groups. Each corneal holder was uniquely identified with a number on the chambers. Corneal opacity was measured quantitatively as the amount of light transmission through the cornea. Permeability was measured quantitatively as the amount of sodium fluorescein dye that passes across the full thickness of the cornea. Both measurements were used to calculate an In Vitro Irritancy Score of the test substance relative to the control corneas.

Irritation parameter:

in vitro irritation score

Value:

7.3

Negative controls validity:

valid

Remarks:

H2O

Positive controls validity:

valid

Remarks:

NaOH

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

- Mean opacity value: 5.0
- Mean permeability value: 0.158
- In vitro irritancy score: 7.3
A substance is considered to be a risk of serious damage to the eyes, if IVIS > 55).

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

The skin irritation/corrosion potential of the test substance was assessed in an OECD 431 and OECD 439 study (EpiDerm SCT and SIT), performed in compliance with GLP criteria. In this study, amounts of 30 µL (irritation test) and 50 µL (corrosion test) of undiluted test substance were applied to 3 and 2 tissues of human skin model (EpiDermskin), respectively. In the irritation test, the model was exposed for 1 hour followed by a 42-hours post-incubation period and in the corrosion test the exposure time was 1 and 42 hours (no post-incubation period). After the exposure times the viability was determined. The viability was determined to be 102% in the irritation test and 88 and 73% in the corrosion test after 3 min. and 1 hour exposure, respectively. Based on these results, it was concluded that the test substance does not show a skin irritation potential in the EpiDerm skin corrosion/irritation test under the test conditions chosen.

Additionally, the potential of the test substance to induce eye irritation was determined in an in vitro eye irritation assay according to OECD 492 (EpiOcular), which was GLP-compliant. In this study, 50 µL of undiluted test substance was applied to a reconstructed human cornea model. Exposure time was 30 minutes followed by a 2 -hour post-incubation period. To remove the test substance, the tissues were washed with sterile PBS. Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The mean viability of the model was 69%, meaning the test substance is not able to reduce MTT directly. Therefore, it was concluded, that the test substance does not show an eye irritation potential in the EpiOcular eye irritation test under the test conditions chosen.

Regarding the eye irritation potential of the test substance, supporting information is available from an OECD 437 study (BCOP) performed in compliance with GLP-criteria. In this study, 3 isolated bovine cornea were exposed to 750 µL of undiluted test substance for a period of 10 minutes, followed by a 2-hour post-incubation period. Subsequently, the corneal opacity and the pemeability were assessed. Corneal opacity was measured quantitatively as the amount of light transmission through the cornea and permeability was measured quantitatively as the amount of sodium fluorescein dye that passes across the full thickness of the cornea. Both measurements were used to calculate an In Vitro Irritancy Score of the test substance relative to the control corneas. All controls were valid. The in vitro irritation score was determined to be 7.3, based on an opacity value of 5.0 and a mean permeability value of 0.158, respectively. Based on the observed results it was concluded, that the test material does not cause serious eye damage in the Bovine Corneal Opacity and Permeability Test (BCOP Test) under the test conditions chosen.

Justification for classification or non-classification

Based on the available information classification for skin or eye irritation is not warranted in accordance with EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation No. 1272/2008.