5-[[4-chloro-6-[(3-sulphophenyl)amino]-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]naphthalene-2,7-disulphonic acid, sodium salt

Endpoint summary

Description of key information

The results of basic toxicity testing give no reason to anticipate unusual characteristics with regards to the toxicokinetics of Reactive Red 198. The data indicate that there is little or no dermal absorption. No signs of a significant systemic toxicity associated with absorption potential have been observed. Bioaccumulation of Reactive Red 198 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.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

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 K_OW, etc. and the results of basic toxicity testing of the test article. The assessment of the toxicokinetic properties of Reactive Red 198 given below is based on the results obtained for the following toxicological endpoints of the test substance and of its structural analogue (SA01) where stated:

 

·       Acute oral toxicity in rats

·       Acute dermal toxicity in rats (Structural Analogue 01)

·       In vivo skin irritation in rabbits

·       In vivo eye irritation in rabbits

·       Skin sensitisation in guinea pigs

·       Bacterial reverse mutation test

·       In-vitro mutagenicity assay in mammalian cells

·       In-vitro cytogenicity assay in mammalian cells

·       In-vivo cytogenicity assay in mammalian cells (Structural Analogue 01)

·       Subacute oral toxicity in rats (Structural Analogue 01)

Physico-chemical properties

Name:                                     Reactive Red 198

EC number:                           279-015-1

EC name:                                5-[[4-chloro-6-[(3-sulphophenyl)amino]-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]naphthalene-2,7-disulphonic acid, sodium salt

CAS number:                        78952-61-1

CAS name:                             2,7-Naphthalenedisulfonic acid, 5-[[4-chloro-6-[(3-sulfophenyl)amino]-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[2-[4-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]diazenyl]-, sodium salt (1:?)

Physical state:                        solid, orange odourless powder

Empirical formula:              C₂₇H₂₂ClN₇O₁₆S₅.xNa / C₂₇H₁₈ClN₇Na₄O₁₆S₅

Molecular weight:                895.0 — 984.189g/mol                        (<500 daltons = good absorption)

Water solubility:                   123 g/L                                                     (= soluble in water)

Partition coefficient:            log Kow = -2.43                                      (>-0.4 or <5.6 = good absorption)

Surface tension:                    52.8 mN/m                                              (<60 = surface active)

Vapour pressure:                  decomposition prior to melting         (not volatile)

Atom count (natoms):         56                                                               (<70 =good bioavailability)

H-bond acceptor (nON):   23                                                               (<10 = good bioavailability)

H-bond donor (nOHNH):  3                                                                 (<5 =good bioavailability)

Toxicological Profile

The acute oral toxicity testing of Reactive Red 198 in the female Wistar rat was carried out as a limit test at a dose level of 2000 mg/kg body weight. No deaths were observed during the study. The animals showed stilted gait, ataxia, squatting posture, irregular respiration, diarrhoea and red discoloured faeces. The test substance was excreted with faeces and urine. Development of body weight was not impaired. Based on the results obtained in this study the median lethal dose value (LD₅₀) of Reactive Red 198 was > 2000 mg/kg body weight. Dermal treatment with 2000 mg/kg body weight of the structural analogue (SA01) caused also neither mortality nor symptoms of toxicological relevance. Exclusively the surface of the skin was red discoloured. There were no macroscopically visible changes in all animals killed at study end.

No systemic toxicity was observed during testing for skin or eye irritating properties of Reactive Red 198 in rabbits. Reactive Red 198 proved not to be irritating to skin or eyes according to the classification criteria of Regulation (EC) No 1272/2008.

The toxicity of the structural analogue (SA01) administered once daily over a period of 28 days at dose levels up to 1000 mg/kg body weight was determined in male and female rats. The test substance did not cause any compound related mortality. Body weight development, haematological and clinical parameters as well as organ weights were unaffected. Faeces of animals of the 1000 mg/kg bw/day test group were red discoloured, from day 6 onwards. Urine was red-brown discoloured in all animals of the 1000 mg/kg bw/day test group. Necropsy revealed red discoloration of the kidneys in all animals of the 1000 mg/kg/day test group. Additionally the testes of the males were red discoloured. Microscopic examination revealed resorption vacuoles with a homogenous content in the tubular epithelial cells of the convoluted segment of the proximal tubulus of the renal cortex in two female animals of the highest dose group. This finding indicates a re-absorption of the dye from the primary urine and its intracellular lysosomal deposition. A deposition of substance in the kidneys of a kind as described was not detected in the remaining animals of this dose group. Likewise a deposition of the dye in the tissue of the testes was not detected.

Based on all findings the ‘No observed effect level’ (NOEL) was conservatively placed at 1000 mg/kg body weight/day for male and 250 mg/body weight/day for female animals. As the dye-re-absorption did not cause any structural of functional effects, the ‘No observed adverse effect level’ (NOAEL) was determined to be 1000 mg/kg bw/day in both genders.

The test substance was negative in two Ames tests with four different Salmonella strains, one additionally with Prival modification and the other with one additional E. coli strain. In addition, Reactive Red 198 was negative in the mutagenicity test in mammalian cells (HPRT test). Reactive Red 198 did not show any clastogenic effects in an in-vitro chromosome aberration test with metabolic activation system. However, it showed a slight clastogenic effect in this test in the absence of the metabolic activation system. This is in line with the result of many in-vitro clastogenicity tests for vinyl-sulphone dyes. It is well known that vinyl-sulphone compounds result in false positive test results in in-vitro tests for clastogenicity (Dearfield KL et al. (1991); Warra TJ et al. (1990)). This is due to the fact that these chemical agents react via the Michael addition reaction. Chemical reactivity via Michael addition is essential for many of the uses for which these compounds are important. As in the currently assessed dye, Reactive Red 198, vinyl sulphone moieties are used in fiber-reactive dyes (MacGregor et at. (1980)). These compounds are known to deplete glutathione in in‑vitro test systems, in which the concentration of phase II enzymes is very low. Glutathione plays a role in the detoxification of many compounds. Conjugation with glutathione via Michael addition and subsequent excretion is the most common bio-elimination route for these compounds. Since in-vitro systems have low levels of glutathione, the glutathione depletion leads to a positive result in the in-vitro test system, which is not the case in the in-vivo test system, where glutathione is present in adequate amount, as could be shown in the negative in-vivo micronucleus study with the structural analogue, which showed also a positive in-vitro result.

In conclusion, it can be stated that under the experimental conditions reported the test item did not induce genotoxic effects in bacterial and mammalian test systems. Therefore, Reactive Red 198 is considered to harvest no genotoxic properties.

Evaluation and Assessment

Based on all available data, Reactive Red 198 does not exhibit conspicuous toxicokinetic behaviour.

Reactive Red 198 is a solid at room temperature conditions. The degree of purity of the substance is ca. 58%. The substance decomposes prior to melting; therefore a significant inhalation exposure to vapours is not expected. In view of the low n-octanol/water partition coefficient (log Kow < ‑2.43 at 25°C), systemic bioavailability after dermal exposure is not anticipated.

Reactive Red 198 has a very low acute toxicity potential. The data of the dermal irritation test indicate low dermal permeability, owing to the fact that neither systemic nor irritating effects were observed. This is in accordance with the extremely good solubility of the test substance in water and with the molecular weight and number of H-bond acceptors, giving evidence of a poor systemic bioavailability.

According to its atom count and H-bond donors, Reactive Red 198 should be absorbed from the gastrointestinal tract to some extent, whereas the molecular weight, log Kow and number of H-bond acceptors indicate a low absorption of the test substance. Taking the results of the subacute oral toxicity study into account, Reactive Red 198 is absorbed from the gastrointestinal tract and is systemically available to some extent, as proved by the tissue and urine staining. The latter being a good indication of the bioelimination of absorbed Reactive Red 198 or its metabolites. According to the molecular weight, excretion of Reactive Red 198 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.

Due to its high water solubility and low log Kow, Reactive Red 198 is not bioaccumulative. This is confirmed by the results of the bioaccumulation modelling, excluding a significant bioaccumulation potential of Reactive Red 198. Additionally, Reactive Red 198 was also not genotoxic; 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 Reactive Red 198. The data indicate that there is little or no dermal absorption. No signs of a significant systemic toxicity associated with absorption potential have been observed. Bioaccumulation of Reactive Red 198 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.