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EC number: 224-073-5 | CAS number: 4193-55-9
The effects of the test article on the integrity and performance of male and female reproductive systems, including gonadal function, estrous cycle, mating behaviour, conception, gestation, parturition, lactation, weaning and growth and development of the spring were assessed testing rats, according to the EPA OPPTS 870.3800 (Reproduction and Fertility Effects) guideline.
Animals were exposed to the test articloe daily, via oral gavage at the dose levels of 0, 100, 300 and 1000 mg/kg bw, at a constant volume of 10 ml/kg. The duration of the whole study was approx. 9 months.
Based on the results, the No Observed Adverse Effect Level (NOAEL) for parental toxicity was 300 mg/kg day and for parental reproductive performance, the NOAEL was l000 mg/kg day. For offspring growth and development, the NOAEL was also 1000 mg/kg day.
A range-finding test (Turk A.P., 2000) on reproduction toxicity in Sprague-Dawley rats as dose range finding for the two -generation study is available (CAS 16470-24-9). 10 rats/sex/dose were dosed with 30, 100, 300 or 1000 mg/kg bw/day by oral gavage during premating, mating, gestation and lactation. Males were killed after mating and females and pups were killed on day 4 of lactation. No substance-related finding was noted in any of the parental animals or pups at any dose level, thus a NOAEL of 1000 mg/kg bw/day for parental and offspring toxicity was established.
In the definitive 2-generation rat study (Turk A.P., 2001), according to EPA Guideline OPPTS 870.3800 / OECD 416 and performed under GLP, 26 Sprague-Dawley rats per sex per group were administered 100, 300 or 1000 mg/kg bw/day by oral gavage. In parental animals, the only test substance-related effect noted was an increased kidney weight. In F0 animals, an increased kidney weight (absolute and relative to body and brain weight) was observed in females at 1000 mg/kg bw/day. In F1 parental animals, there was an increase in kidney weight in males (absolute and relative to body weight) and females (absolute and relative to body and brain weight) at 1000 mg/kg bw/day as well as an increase in kidney weight (relative to body weight) in females at 300 mg/kg bw/day. The statistical change in 300 mg/kg bw/day was considered to be spurious since no changes in absolute weight or kidney weight relative to brain weight were seen, and similar increases were not observed in 300 mg/kg bw/day males. There were no test substance-related effects on reproductive performance noted for either parental generation. No adverse, test substance-related changes in growth or development of offspring were observed in either the F1 or the F2 generations. Based on the results of this study, the NOAEL for parental toxicity was 300 mg/kg bw/day. For parental reproductive performance, the NOAEL was 1000 mg/kg bw/day. For offspring growth and development, the NOAEL was also 1000 mg/kg bw/day.
Both the tests were performed on a similar substance within the category of Stilbene Fluorescent Whitening Agents: the analogous dihydroxyethyl derivative tetrasulphonated sodium salt, CAS 16470-24-9. This substance has the same organic functionalities, with a higher sulphonation degree. The sulphonation degree has an impact on the solubility of the substance at neutral/basic pH, but both the analogous and the substance under registration can be considered highly soluble. Therefore, the substance CAS 16470-24-9 can be considered a good representative within the category for CAS 4193-55-9.
From a metabolic point of view the two substances share the same metabolic pathway. Based on the profiling proposed by the OECD Toolbox, the same systemic effects can be expected, being identical the organic functional groups and metabolic pathway. Impurity profile has no influence on this read across, since both substances contain about 10 % of organic by-products that are related to the similar production process: they are in fact di substitution of the dihydroxyethylamino on the same triazine moiety or of the aniline part. In all case the same metabolic pathway is expected for the impurities related to the two substances.
The review of all the available information on the category members seems provide enough evidences in order to avoid the performance of a specific reproduction test on the substance under registration, for sake of animal welfare.
It has to be noticed that in the described conditions of use the potential human exposure is negligible. In fact, the substance is used as fluorescent whitening agent in the detergency field, where no oral exposure is involved, neither inhalation exposure of worker or consumer due to the fact that the substance is a solid, with very low vapour pressure.
Based on the described uses, the only possible consumer contact scenario is identified in direct skin contact with the final consumer product through pre-treated clothes or hand-wash laundry. Indirect skin contact may occur via residual deposits on clothing or by inhalation of detergent dust during consumer product handling, and oral ingestion from residues on dishes or from accidental product ingestion.
Though toxicokinetic studies on rats indicate very low dermal or intestinal absorption rates of 0.1 % of the administered dose, for the consumer exposure estimates worst case absorption rates of 10 % were assumed for dermal absorption since no experimental data are available for repeated contact or application scenarios.
The maximum exposure was estimated occurring by skin contact during pre-treatment of clothes (spot pre-treatment) (HERA 2004 - Substance: Fluorescent Brightener FWA-1): 0.21 mg/Kg bw/day.
Two studies revealed no evidence of teratogenicity in rats and rabbits: rat: NOAEL = 1000 mg/kg bw/day (maternal and foetal toxicity); rabbit: NOAEL = 100 mg/kg bw/day (maternal and foetal toxicity). GLP-compliant OECD 414 studies (Turk 1999 and 2000).
Two studies on developmental toxicity and teratogenicity, according to EPA Guideline OPPTS 870.3700 and under GLP conditions, were performed in rats and rabbits with CAS 16470-24-9.
In the rat study (Turk, 1999), 30 pregnant Sprague-Dawley rats per group were dosed with 100, 400 or 1000 mg/kg bw/day by oral gavage on gestation days 6-19. The only substance-related effect observed was discoloured faeces at 400 and 1000 mg/kg bw/day. At skeletal examination of foetuses, the incidence of misaligned sternebra was slightly increased in all dose groups but was well within historical control range and not dose-related and therefore not considered to be test substance related. The incidence of rudimentary ribs was slightly above the historical control range at 100 and 1000 mg/kg bw/day. As the difference from the concurrent control group was not statistically significant and the increase was not dose-related, these findings were not considered biologically significant or test substance-related. The number of vertebral malformations at 1000 mg/kg bw/day (litter incidence 7.1 %) was very slightly above the historical control range (0 - 7 %) and not statistically different from the vehicle controls. Therefore, also this border finding was considered to be within normal variation and unrelated to test substance administration. As there were no adverse maternal or developmental effects seen at any dose level, the NOAEL for both maternal and foetal toxicity is the highest dose tested (1000 mg/kg bw/day).
In the rabbit study (Turk 2000), 7 pregnant New Zealand White rabbits per group were dosed with 100, 400 or 800 mg/kg bw/day by oral gavage on gestation days 7 - 28. The application of 800 mg/kg bw/day resulted in excessive maternal toxicity as exhibited by death, abortion, increased incidence of clinical and gross pathological findings, and a marked decrease in food consumption and body weight gain. As a consequence this group was terminated prior to study. Abortion or early delivery and soft stool and discoloured faeces also occurred in some dams at 400 mg/kg bw/day. The foetal body weights were lower in the 400 mg/kg bw/day group than compared to controls, which is considered to be secondary to maternal toxicity. At visceral examination of foetuses, the litter incidence of hemorrhagic iris at 400 mg/kg bw/day was slightly above the historical control range while the slightly increased incidences of gallbladder agenesis, hypoplasia of the gallbladder and azygous lobe of lung absent were within historical control range. Since all the above findings were within or slightly above historical control range, the findings were considered to be spontaneous in nature and unrelated to test substance. Also, no substance-related effects were noted at external and skeletal examinations. At a dose level of 100 mg/kg bw/day no substance-related effects were seen in dams or at foetal examinations. The NOEL for both maternal and foetal toxicity therefore was established as 100 mg/kg bw/day.
Both the tests were performed on a similar substance within the category of Stilbene Fluorescent Whitening Agents: the analogous dihydroxyethyl derivative tetrasulphonated sodium salt, CAS 16470-24-9. This substance has the same organic functionalities, with a higher sulphonation degree. The sulphonation degree has an impact on the solubility of the substance at neutral/basic pH, but both substances can be considered highly soluble. From a metabolic point of view the two substances share the same metabolic pathway, therefore it can be assumed that the test result will be representative of the substance under registration, too.
In order to select the most ‘biologically active’ surrogate for further reproduction and developmental toxicity testing and in order to generate data to help establish dosage levels, two pilot prenatal developmental toxicity studies were performed both in rabbits and rats on CAS 16470-24-9 and CAS 32466-49-6 (the acid form of 16090-02-1) administered via oral gavage. According to the agreed decision tree within SOCMA (Stilbene Whitener Task Force toxicology testing program) the most biologically active surrogate was selected and two main prenatal developmental toxicity studies performed in rabbits and rats as well as a 2-generation reproductive toxicity and fertility study in rats.
Based on the excessive maternal toxicity observed in rabbits treated with CAS 16470-24-9 at 1000 mg/kg bw/day, it was concluded to be more biologically active than 32466-49-6 under the employed experimental conditions and therefore it was selected for the definitive prenatal developmental toxicity studies in rats and rabbits. The appropriateness of this choice was confirmed by the U. S. Environmental Protection Agency (U. S. EPA) in its letter dated June 4, 1998.
It has to be taken into account the result of the study on rabbits presents some inconsistencies: rabbit is regarded as a too sensitive specie for repeated dose toxicity, and this is also confirmed by the number of mothers died for gavage mistakes. Since the observed effects on pups (weight gain loss) is reported to a maternal toxicity effect, it has to be evaluate if the test is appropriate to assess Reproductive toxicity for the category members.
According to the CLP Regulation (EC 1272/2008), 3.7 Reproductive toxicity section, reproductive toxicity includes adverse effects on sexual function and fertility in adult males and females, as well as developmental toxicity in the offspring.
The available experimental data are adequate for classification and labelling and the substance is not classified for reproductive toxicity according to the CLP Regulation (EC 1272/2008).
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
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