Disodium 7-[[4,6-bis[(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[[4-[(4-sulphonatophenyl)azo]phenyl]azo]naphthalene-2-sulphonate

Endpoint summary

Description of key information

Oral LD50 > 2000 mg/kg bw

Key value for chemical safety assessment

Acute toxicity: via oral route

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

ACUTE TOXICITY - ORAL ROUTE

The acute oral toxicity potential of the test substance was tested on rats. Unfortunately, only a short sheet of toxicological data is currently available, thus reliability cannot be assigned; test procedures and study results are not detailed. The oral LD50 indicated is higher than 5000 mg/kg.

The result is confirmed in a second investigation performed on the substance salificated by sodium and triethanolammonium salt (50:50) instead of sodium salt alone. Also in this case only a brief sheet is available; the LD50 reported is greater than 5000 mg/kg.

Despite the reliability of the available information cannot be judged, Direct Red 253 is expected to be not acutely harmful/toxic, by oral route.

The substance has been tested in a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test, by oral route. During the dose-range finding experiment with Direct Red 253 no animal died during the application period. Clinical observation did not detect the impact of the test substance on the health condition of animals at all dose levels, the administration of the test substance did not induce treatment-related changes of red and white blood component in both sexes and no serious macroscopical changes were observed during necropsy of treated animals up to the highest tested dose of 1000 mg/kg bw/day.

The main test confirmed the outcomes of the range-finding, in fact no significant signs of systemic toxicity were recorded up to 1000 mg/kg bw/day, which was established to be the No Observed Adverse Effect Level; no signs of reproduction/developmental injuries were recorded and the NOAEL was stated 1000 mg/kg bw/day (Koryntová, 2016; for details IUCLID sections 7.5 and 7.8).

The relationship between LD50 and NOAEL has been investigated by Bulgheroni et al. on 2009. Following the initiatives undertaken in the pharmaceutical industry to waive the acute oral toxicity testing before going to clinical studies by using information from other in vivo studies, the authors proposed an approach to identify non-toxic compounds (LD50 > 2000 mg/kg) using information from 28 days repeated dose toxicity studies. Taking into account the high prevalence of non-toxic substances (87 %) in the New Chemicals Database, it was concluded that NOAEL values obtained from studies in rat could be useful to estimate acute oral toxicity. Indeed, on the basis of NOAEL values ≥ 200 mg/kg obtained from 28 days repeated dose studies in rat it was possible to correctly identify ‘‘non-toxic” substances (LD50 > 2000 mg/kg b.w.). This approach would miss <1 % of the substances which are harmful classifying them as ‘‘non-toxic”. Notably, this misclassification was only within one category and none of the toxic substances would have been missed (Bulgheroni et al, 2009).

The approach for the prediction of acute oral toxicity based on the results of a sub-acute oral toxicity study, has been evaluated as proposed in the ECHA R.7a Endpoint specific guidance, currently under update, Draft Version 5.0 (Public) (August 2016). In the draft guidance the suitability of the approach is confirmed, however a different threshold is setted: the non-classification for oral acute toxicity can be correctly predicted based on the results of oral sub-acute studies, when the NOAEL is at or above 1000 mg/kg bw.

In conclusion, the substance can be considered as not harmful/toxic for acute toxicity by oral route.

ACUTE TOXICITY - INHALATION ROUTE

No acute toxicity studies by inhalation route are available on Direct Red 253.

Nevertheless, because of the physical state and the trade forms of the substance under registration, inhalation is not an appropriate route of exposure.

ACUTE TOXICITY - DERMAL ROUTE

The inhalation and the skin contact of Direct Red 253 are unlikely. Furthermore, because of the physical/chemical properties of the substance the dermal absorption is expected as negligible.

According to the Commission Regulation (EU) 2016/863, amending Annexes VII and VIII to REACH Regulation (EC 1907/2006) as regards skin corrosion/irritation, serious eye damage/eye irritation and acute toxicity, testing by the dermal route does not need to be conducted if no systemic effects have been observed in in vivo studies with dermal exposure (e.g. skin irritation, skin sensitisation). Furthermore, it is explained that recent scientific analysis of available data from in vivo acute toxicity studies have shown that substances that are not toxic via the oral route may be expected with high certainty to be also non-toxic via the dermal route. Therefore, testing those substances via the dermal route does not provide essential information for their safety assessment.

 

The amendment is the consequence of studies and scientific debates. In particular, the 15^th Meeting of Competent Authorities for REACH and CLP (CARACAL, 2014) concluded that an adaptation of point 8.5.3 of Annex VIII to REACH is justified in order to not require information on acute dermal toxicity for substances that have shown no toxicity in acute oral toxicity test up to the limit dose of 2000 mg/kg bw.

The skin sensitisation assay has been performed on the Direct Red 253 salificated by sodium and triethanolammonium salt (50:50) instead of sodium salt alone; the difference in salification does not impact the systemic toxicity potential of the substance. The allergenic potential of the substance was examined in a modified Buehler test, according to OECD test guideline 406. No mortality occurred and bodyweight gain of all animals was unaffected during the study. The application area around the epidermal induction sites showed staining around day 2 to day 26. No systemic symptoms were observed in any animal. No skin reactions were observed in the irritation screen for induction and irritation screen for challenge (Ullmann, 1990; see IUCLID section 7.4.1).

 

In conclusion, because of the dermal absorption and the systemic toxicity potentials, no further investigation is proposed.

REFERENCE

Bulgheroni A., Kinsner-Ovaskainen A., Hoffmann S., Hartung T., Prieto P. (2009). Estimation of acute oral toxicity using the No Observed Adverse Effect Level (NOAEL) from the 28 day repeated dose toxicity studies in rats. Regulatory Toxicology and Pharmacology 53, 16–19.

CARACAL, 2014. 15th Meeting of Competent Authorities for REACH and CLP (CARACAL), 8 – 9 July 2014. Charlemagne building, Brussels, Belgium. Brussels, 26 July 2014. Doc. CA/61/2014. Stakeholder proposal to modify REACH standard information requirements for acute toxicity (REACH Annex VIII, point 8.5).

ECHA (2016). Guidance on Information Requirements and Chemical Safety Assessment Chapter R.7a: Endpoint specific guidance Draft Version 5.0 August 2016. Available: https://echa.europa.eu/documents/10162/13643/ir_csa_r7a_r7-4_caracal_en.pdf/1f97ae8b-052c-4713-959a-5fd742b63b03

Justification for classification or non-classification

According to the CLP Regulation (EC 1272/2008), 3.1 Acute toxicity section, substances can be allocated to one of four toxicity categories based on acute toxicity by the oral, dermal or inhalation route according to the numeric criteria. Acute toxicity values are expressed as (approximate) LD50 (oral, dermal) or LC50 (inhalation) values or as acute toxicity estimates (ATE).

The oral LD50 value was established to be greater than 2000 mg/kg, therefore the test substance is out of any classification limit for acute oral toxicity (oral acute toxicity category 4: 300 < ATE ≤ 2000 mg/kg bw).

Dermal and inhalation are considered as negligible exposure routes, thus no acute toxicity tests have been conducted and no further investigations are required.

In conclusion, the test substance does not meet the criteria to be classified for oral acute toxicity, according to the CLP Regulation (EC 1272/2008).