3-[[4-[(2,6-dibromo-4-nitrophenyl)azo]phenyl]ethylamino]propiononitrile

Endpoint summary

Description of key information

Skin sensitiser.

Key value for chemical safety assessment

Skin sensitisation

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

As no data on the substance in itself was available, a read across approach was used to assess its skin sensitising potential. In particular, the assessment relied on an study on Similar Substance 03 along with evidences/studies on Similar Substance 01, Similar Substance 02 as well as on the group, as a whole, of azo dyes having the phenyl-azo-phenyl stucture, functionalised by nitro and amino groups, halogen atoms, etc.

Details on the read across approach are available in section 13.

An available study on Similar Substance 03 was used as key study. Similar Substance 03 was tested in the Buheler test, according to OECD guideline 406.

Twenty female animals of test group were treated topically with test substance at 50 % in bi-distilled water once a week for a 3-week induction phase. Two weeks after the final induction application, animals were challenged with test substance at 50 % concentration in bi-distilled water. Ten animals of the control group were not treated during the induction but were treated once at challenge with test substance at 50 % in bi-distilled water.

After challenge, 40 % of animals of test group showed skin reactions, while no skin reactions were observed in the control group. Therefore, test substance was considered as skin sensitiser.

Available data on Similar Substance 01 and Similar Substance 02 were also taken into account.

Similar Substance 01 was tested in mixture with 3 other dyes in a repeated insult patch test to evaluate dermal effects in humans. Occlusive application was continued for 4 weeks from Monday to Friday; on the second Monday, following last week of application, challenge was done with a contact period of 48 h duration. After removal, sites were examined, then re-examnined after 24 and 48 h. At challenge 2 out of 200 individuals showed positive responses. Upon testing with separate components of the mixture, Similar Substance 01 induced a positive reaction in one of the 2 sensitised individual.

Based on such result, Similar Substance 01 was associated to a potential hazard upon prolonged skin contact. However, as no data on Similar Substance 01 as such was available, but only data upon application of a mixture of 4 dyes, possible interefering effects could not be excluded. Accordingly, these findings were only used as supporting evidence to findings in the key study.

Conflicting results were available on Similar Substance 02, which was found to be negative in 2 LLNA studies and positive in a GPMT study.

A LLNA study (2002) was conducted according to OECD guideline 429. Male mice were exposed to test substance at concentrations of 0, 3, 10 and 30 % in acetone. Isotope (3H-methyl thymidine) incorporation was measured in lymph nodes. The isotope incorporation was increased by less than a threefold at all concentrations. Negative and positive controls were valid.

Modified protocols of LLNA, i.e. sensitisation and sensitisation-challenge protocols, were used in a study of 2006. Female mice were exposed to test substance at concentrations of 10 and 30 % in DMSO. In the sensitisation protocol, animals received 25 µl of test solution on the dorsum of the left ear for 3 consecutive days. The right ears were treated with equal volume of DMSO. In the sensitisation-challenge protocol, animals were shaved over a surface of approximately 2 cm² on their back and this surface was treated once daily on days 1 -3 with 50 µl of test solution. Mice remained untreated on days 4 -14 and were then challenged with 25 µl of test solution on days 15 -17. In both protocols, observations performed 48 h after the last exposure showed no changes regarding lymph node weight and cellularity, ear thickness, or lymphocytes subpopulations, compared to vehicle control.

A GPMT study of 1997 was available. Based on pre-test results, females Guines pigs were treated with: a 5 % test substance in bi-distilled water and in FAC / physiological saline emulsion for intradermal induction; 50 % test substance in bi-distilled water for epidermal induction under occlusion (clipped, shaved skin) upon pretreatment of a 10 % SLS solution in paraffinum perliquidum; 50 % in bi-distilled water under occlusive dressing for epidermal challenge two weeks after induction. Cutaneous reactions, i.e. erythema and eschar, as well as oedema formation were evaluated at 24 and 48 h after removal of the dressing. Positive reactions were observed in all animals when treated with the test substance at 50 % in bi-distilled water.

Lastly, it should be noted that most of the azo dyes, having the phenyl-azo-phenyl structure with nitro and amino groups along with halogen atoms as functional groups, was found to be skin sensitising in in vivo studies. Therefore, a potential for skin sensitisation could not be reasonably excluded in the target substance.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Category 1

Substances shall be classified as skin sensitizers in category 1 where data are not sufficient for sub-categorisation in accordance with the following criteria:

(a) if there is evidence in humans that the substance can lead to sensitisation by skin contact in a substantial number of persons; or

(b) if there are positive results from an appropriate animal test.

As for Buehler test, a response of at least 15 % of animals is considered positive.

Subcategorisation is done as follows in case of a Buehler assay:

Sub-category 1A

Substances showing a high frequency of occurrence in humans and/or a high potency in animals can be presumed to have the potential to produce significant sensitisation in humans. Severity of reaction may also be considered.

Specific criteria: ≥ 15 % responding at ≤ 0.2 % topical induction dose or ≥ 60 % responding at > 0.2 % to ≤ 20 % topical induction dose

Sub-category 1B

Substances showing a low to moderate frequency of occurrence in humans and/or a low to moderate potency in animals can be presumed to have the potential to produce sensitisation in humans. Severity of reaction may also be considered.

Specific criteria: ≥ 15 % to < 60 % responding at > 0.2 % to ≤ 20 % topical induction dose or ≥ 15 % responding at > 20 % topical induction dose.

Based on data derived from a Buehler assay, i.e. 40 % of test animals responding at 50 % topical induction dose, the substance is classified in category 1B of the CLP Regulation (EC 1272/2008)