2-[N-(2-cyanoethyl)-4-[(2,6-dichloro-4-nitrophenyl)azo]anilino]ethyl acetate

Endpoint summary

Additional information

Introduction

Toxicokinetic parameters such as uptake, distribution, metabolism and excretion form the essential toxicological profile of a substance. An approximate indication of the toxicokinetic pattern can be gained from the physico-chemical properties taking into account the molecular weight, the number of atoms (hydrogen bond donors and acceptors), the solubility in solvents, log Kow, etc. and the results of basic toxicity testing of the test article. The assessment of the toxicokinetic properties of Disperse Orange 30 given below is based on the results obtained for the following toxicological endpoints of the test substance:

• Acute oral toxicity in rats
• In vivoskin irritation in rabbits
• In vivoeye irritation in rabbits
• Skin sensitisation in guinea pigs and mice
• Bacterial reverse mutation test
• In vitromutagenicity assay in mammalian cells
• In vivomicronucleus assay in mammalian cells
• Subacute oral toxicity in rats
• Reproductive screening study in rats

All studies were carried out according to the principles or in the spirit of Good Laboratory Practice and met the requirements of the OECD and EU-Guideline for the Testing of Chemicals.

Physico-chemical properties

Name:                             Disperse Orange 30

EC number:                    226 -070 -4

EC name:                        2 -[N-(2-cyanoethyl)-4-[(2,6-dichloro-4-nitrophenyl)azo]anilino]ethyl acetate

CAS number:                  5261 -31 -4

CAS name:                     Propanenitrile, 3-[[2-(acetyloxy)-ethyl][4-[2-(2,6-dichloro-4nitrophenyl)diazenyl]phenyl]amino]-

Physical state:                Brown solid

Empirical formula:        C₁₉H₁₇Cl₂N₅O₄

Molecular weight:         450.3 g/mol                                                        (<500 daltons = good absorption)

Water solubility:           0.04 mg/L                                                                         (= low solubility in water)

Partition coefficient:    log Kow = 4.1                                                    (>-0.4 or <5.6 = good absorption)

Vapour pressure             <1 x 10E-5 Pa                                                                                        (not volatile)

 

Toxicological profile

Acute oral toxicity of Disperse Orange 30 was evaluated in rats in two studies. No adverse treatment-related effects were observed in either of the studies and the oral LD50 was greater than 2000 mg/kg bw.

No systemic toxicity was observed during testing for skin sensitising and skin or eye irritating properties of Disperse Orange 30 in guinea pigs, mice and rabbits, respectively. Disperse Orange 30 was not irritating to skin or eyes according to the classification criteria of Regulation (EC) No 1272/2008. Further, based on weight of evidence from the available studies, the test substance was not considered to be sensitising to the skin.

The reproductive and repeated dose toxicity of the test substance was determined in male and female rats in a combined repeated dose and reproductive-developmental screening study according to OECD Guideline 422, in compliance with GLP. Rats were administered the test substance orally by gavage at dose levels of 62.5, 250, 500, 1000 and 1000/500 mg/kg bw/day. Males animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing, through the mating period and thereafter through the day before necropsy (Days 38 and 39 of study). Males were treated for a total of 37 or 38 days. Female animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter during pairing, post-coitum and post-partum periods until Day 3 post-partum (for at least 40 d). Recovery groups (0 or 1000, 1000/500 mg/kg bw/day test substance) animals were dosed once a day, 7 days a week, for 4 consecutive weeks. No treatment was given during the recovery period. No treatment-related signs of toxicity were observed at doses lower than the highest dose of 500/1000 mg/kg bw/day. Staining due to test substance was observed in the internal organs in all test substance treated animals, confirming absorption of the test substance (also substantiated by the high partition coefficient value). Based on the results, the NOAEL of the test substance for the study was considered to be 250 mg/kg bw/day for general toxicity and reproductive and developmental toxicity in males and females. As no malformations were observed in the offspring, the NOEL for teratogenic effects considered to be 1000/500 mg/kg bw/day

The test substance was tested positive in the Ames test in Salmonella typhimurium TA 1535, TA 1537, TA 98 and TA 100. In addition, Disperse Orange 30 was negative in the forward mutation assay in HPRT locus of Chinese hamster V79 cells. It also did not show any clastogenic or aneugenic effects in an in vivo micronucleus study in rats. This positive effect in the bacterial mutation assay is a bacteria-specific effect due to bacterial nitro-reductases, which are highly effective in these bacterial strains, but not in mammalian cells.

Evaluation and Assessment

Examination of the data of the dermal irritation and skin sensitization studies gave no indication of there being any evidence for skin penetration of the test item. Oral bioavailability of Disperse Orange 30 is plausible due to the log Kow of 4.1, since substances with a log Kow > 0.5 might be significantly absorbed. Bioavailability of Disperse Orange 30 after oral administration was demonstrated in the repeated-dose toxicity study at all dose levels (discolouration of urine and tissues/organs). The discolouration of urine observed during treatment in high-dose rats (red staining of cage tray) was no longer present after cessation of treatment during the 14-day recovery period. Likewise, the discolouration of the tissues/organs was completely diminished in recovery animals. Hence, there was no indication of bioaccumulation.

According to the molecular weight, excretion of Disperse Orange 30 is most likely predominantly eliminated via intestine, as substances with a molecular weight above 300 g/mol are preferentially excreted via the faeces in rats. However, the test results showed that the test compound is at least partly eliminated via kidneys/urine, too.

Disperse Orange 30 did only show mutagen effects in bacteria, but was not was not genotoxic in mammalian cells or in in vivo-tests; therefore, metabolisation towards genotoxic structures by mammalian species can most probably be excluded.

Summary

The results of basic toxicity testing give no reason to anticipate unusual characteristics with regards to the toxicokinetics of Disperse Orange 30. The data indicate that there is little or no dermal absorption. No signs of a significant systemic toxicity associated with absorption through skin have been observed. Disperse Orange 30 is to some extent absorbed through the gastro-intestinal system when given orally by gavage and excreted predominantly via faeces and to a lesser extent via urine. Bioaccumulation of Disperse Orange 30 can most probably be excluded due to the available data. Based on the results of genotoxicity assays, a metabolisation towards genotoxic metabolites can also be excluded for mammalian species.

On the basis of the results, it is anticipated that the substance does not undergo significant metabolic activity; rather it is metabolised for excretion with little subsequent toxicity. The substance is therefore not considered to be of concern for ADME related effects.