[2-[[4-[(2-chloro-4-nitrophenyl)azo]phenyl]ethylamino]ethyl](2-hydroxypropyl)dimethylammonium acetate

Endpoint summary

Description of key information

Based on test data and physico-chemical properties, there is little or no dermal absorption of Basic Red 18:1, but a good oral bioavailability. Bioaccumulation of Basic Red 18:1 is unlikely based on available data. Based on the results of genotoxicity assays, a metabolisation towards genotoxic metabolites can also be excluded for mammalian species.

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 Basic Red 18:1 given below is based on the results obtained for the following toxicological endpoints:

• Acute oral toxicity in rats
• Acute dermal toxicity in rats
• In vivo skin irritation in rabbits
• In vivo eye irritation in rabbits
• Skin sensitization in mice
• Bacterial reverse mutation test
• In vitro mutagenicity assay in mammalian cells
• In vivo micronucleus test in mice
• Subacute oral toxicity in rats
• Reproductive screening study in rats

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

Substance identity and physico-chemical properties

Name:                                     Basic Red 18:1Acetate

CAS number:                         82205-20-7

CAS name:                             1-Propanaminium, N-[2-[[4-[2-(2-chloro-4-nitrophenyl)diazenyl]phenyl] ethylamino]ethyl]-2-hydroxy-N,N-dimethyl-, acetate (1:1)

Physical state:                        red solid in aqueous solution - Basic Red 18:1 Acetate is only stable and put on the market in an aqueous acetic acid solution

Empirical formula:              C₂₁H₂₉ClN₅O₃.C₂H₃O₂

Molecular weight:                493.98 g/mol                                                    (< 500 daltons=good absorption)

Water solubility:                   > 400 g/L                                                           (= soluble in water)

Partition coefficient:            log Kow < 0.3                                                   (>-0.4 or <5.6 = good absorption)

Atom count (n atoms):        30                                                                        (<70 = good bioavailability)

H-bond acceptor (nON):    8                                                                          (<10 = good bioavailability)

H-bond donor (nOHNH):  1                                                                          (<5 = good bioavailability)

 

 

Name:                                     Basic Red 18:1Chloride

CAS number:                        54229-13-9

CAS name:                             1-Propanaminium, N-[2-[[4-[2-(2-chloro-4-nitrophenyl)diazenyl]phenyl] ethylamino]ethyl]-2-hydroxy-N,N-dimethyl-, chloride (1:1)

Physical state:                        solid, red powder

Empirical formula:              C₂₁H₂₉ClN₅O₃.Cl

Molecular weight:                470.4 g/mol                                                      (<500 daltons=good absorption)

Water solubility:                   132 g/L                                                              (= soluble in water)

Partition coefficient:            log Kow = 0.3                                                   (>-0.4 or <5.6 = good absorption)

Surface tension:                    < 57 mN/m                                                       (<60 = surface active)

Melting range:                       185 – 190 °C                                                     (= not volatile)

Vapor pressure:                    3.4E-7 hPa at 20°C                                          (= not volatile)

Atom count (natoms):        30                                                                        (<70 = good bioavailability)

H-bond acceptor (nON):   8                                                                          (<10 = good bioavailability)

H-bond donor (nOHNH): 1                                                                          (<5 = good bioavailability)

Toxicological profile

Acute oral toxicity of Basic Red 18:1 was evaluated in rats in studies one with the acetate, the chloride and the methylsulfate form. Adverse effects observed at dose levels > 1000 mg/kg body weight consisted in bad general condition, diarrhea, sedation, lateral and prone position. All signs of intoxication occurred within ca. 10 min and were reversible within a maximum of 4 days. The oral median lethal dose varied between 1500 mg/kg body weight (Cl), 2500 mg/kg body weight (Acetate) and > 3500 mg/kg body weight (Cl/MeSO4) based on 100% active ingredient for the different salt forms. No mortality or adverse effects were seen in the dermal toxicity study at the limit dose of 200 mg/kg body weight. Furthermore, no systemic toxicity was observed during testing for skin sensitising and skin or eye irritating properties of Basic Red 18:1 in mice and rabbits, respectively. Basic Red 18:1 was not irritating to skin, but caused irreversible damage to eyes and proved to be skin sensitising according to the classification criteria of Regulation (EC) No 1272/2008.

The reproductive and repeated dose toxicity of the test substance (Basic Red 18:1 Chloride) 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. In the preliminary dose-range finding study, dose levels of 300, 600, and 1000 mg/kg body weight/day were administered to four rats/sex/dose group for 14 days. No animals died. Treated animals of both sexes receiving 600 and 1000 mg/kg body weight/day showed salivation as the major clinical sign. As a consequence of the colour of the test item, orange staining was observed inside the cage of all treated animals of both sexes starting from Day 3 of the study until termination. Loss of body weight and lower body weight compared to controls was noted on Days 8 and 15 of the study in females and males dosed with 600 and 1000 mg/kg body weight/day, respectively. Lower body weight and lower body weight gain, compared to control, was also observed in low-dose females on Day 8 and males and females on Day 15. All treated animals of both sexes showed a decrease in food consumption on Days 8 and 15 of the study, when compared to the control group. Changes were more evident in females than in males and more prominent on Day 8 that Day 15 for both gender. Terminal body weight was lower in males dosed at 600 and 1000 mg/kg body weight/day and in all treated females compared to the control group. Macroscopically, swollen and/or enlarged livers were seen in of most males and in a few females dosed at ≥ 300 mg/kg body weight/day. Besides red or brown discolouration of the non glandular region of the stomach, the muzzle, skin, tail and forelimbs due to the test substance colour, no staining of inner organs/tissues were observed. Based on the results of the preliminary study, rats were administered the test substance orally by gavage at dose levels of 40, 100, and 250 mg/kg body weight/day in the main study. 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. Males were treated for a total of 36 or 37 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 41 d). Recovery group animals were dosed once a day, 7 days a week, for 4 consecutive weeks. No treatment was given during the recovery period. No adverse effects of the test item were found up to the dose level of 250 mg/kg body weight/day. No mortality occurred in the control or in any of the dose groups during the treatment period of this study. There were no clinical signs of toxicological relevance in the dose groups when compared to the control group; no staining in the cage tray was observed. The neurotoxicity assessment (motor activity, grip strength and sensory activity) did not reveal any changes attributable to the test item. There were changes of minimal severity in body weight and body weight gain in both male and female animals, which were considered irrelevant. There were no adverse effects on food consumption of males and females of the dosing groups compared to control animals during the study period. Furthermore, no adverse changes were measured for haematology and clinical chemistry parameters. Additionally, no adverse effects or staining of organs/tissues were seen during necropsy. At microscopic observation, no adverse effects were noted; the only treatment-related changes recorded in the liver of the animals of both sexes were considered a hepatic adaptive change due to hepatic metabolism of the test substance. The NOAEL in this study is considered to be 250 mg/kg body weight/day.

The test substance was tested in a reverse gene mutation assay in bacteria (equivalent to OECD 471), in strains TA 1535, TA 100, TA 1537 and TA 98 of Salmonella typhimurium. The test item was tested up to the limit concentration (5000 µg/plate). Based on bacteriotoxic effects, dose groups above 400 µg per plate were not used for assessment purposes. In Salmonella typhimurium strains TA 100, TA 1537 and TA 98 a mutagenic effect was observed at non-cytotoxic doses. In the Salmonella typhimurium strain TA 1535 no mutagenic effect was observed. Basic Red 18:1 was examined for mutagenic activity by assaying for the induction of 5 trifluorothymidine resistant mutants in mouse lymphoma L5178Y cells. Based on the results obtained in the preliminary trial revealing cytotoxicity in higher dose levels, two independent assays for mutation at the TK locus were performed using the dose levels between 3.13 and100 µg/mL. Adequate levels of cytotoxicity, covering a range from the maximum to slight or no toxicity, were observed in all treatment series. No relevant increases in mutant frequencies were observed following treatment with the test item, in the absence or presence of S9 metabolism. Basic Red 18:1 was further tested in a NMRI mouse bone marrow micronucleus test at dose levels of 0 and 50 mg/kg body weight. After test substance administration, the animals showed apathy, roughened fur, staggering gait, spasm, twitching, difficulty in breathing and orange discoloured urine; proving sufficient exposure to the test substance. There was not a significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow after any treatment time (16, 24 and 48 hours).

Evaluation and Assessment

Basic Red 18:1 acetate is only stable in presence of significant concentrations of free acetic acid; it cannot be isolated as such. The test substance is an aqueous solution in acetic acid and therefore no inhalable dust is formed. Basic Red 18:1 as chloride salt has a low volatility (3.4E-7 hPa at 20°C) and should therefore not be released into air. Hence, no uptake of the substance via inhalation is expected.

Basic Red 18:1 acetate has a very high solubility in acetic acid solution; the commercial form is a ca 40 % preparation (400 g/L) of Basic Red 18:1 acetate. Basic Red 18:1 chloride has a solubility of about 132 g/L, which means that the solubility of Basic Red 18:1 acetate is likely to decrease slightly with dissolution of the acetic acid solution.

Based on physico-chemical data and the results of toxicity studies, Basic Red 18:1 has a good oral, however only limited dermal bioavailability. According to its low log Kow, Basic Red 18:1 acetate does not show any potential for bioaccumulation. This is confirmed by the results of the bioaccumulation modelling (BCFBAF v3.01), excluding a significant bioaccumulation potential of Basic Red 18:1 and the lack of staining in organs/tissues in the repeat dose toxicity studies.

According to the molecular weight, excretion of Basic Red 18:1 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.

Basic Red 18:1 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 Basic Red 18:1. 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. Based on physico-chemical data and the results of oral toxicity studies, Basic Red 18:1 has a good oral bioavailability. Bioaccumulation of Basic Red 18:1 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.

The substance is therefore not considered to be of concern for ADME related effects.