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EC number: 216-341-5 | CAS number: 1561-92-8
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Endpoint summary
Administrative data
Description of key information
90 day oral toxicity study in rats showed no effects at a top dose of 1000 mg/kg/day
Key value for chemical safety assessment
Repeated dose toxicity: via oral route - systemic effects
Link to relevant study records
- Endpoint:
- sub-chronic toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: OECD protocol, GLP-accredited lab, no major deviations from protocol.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
- Species:
- rat
- Strain:
- Wistar
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- Wistar HSD/WIN:WU;
- Route of administration:
- oral: drinking water
- Vehicle:
- water
- Details on oral exposure:
- dosing through drinking water "at lib". Water consumption in each cage was measured weekly. Mean achieved dosages (mg/kg bodyweight) of the test material were calculated by water consumption. Concentrations were 0, 500, 2000 and 7000 ppm, expected to result in dose levels of 0, 50, 250 and 1000 mg/kg bw respectively.
- Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- A 20% standard solution of the test material in aqua bidest. was prepared every second week.
To achieve the target concentrations, the calculated amount of aqueous solution were weekly prepared by dilution of the standard solution with the appropriate amount of tap water. To achieve homogeneity, the solution was stirred by a magnetic stirring device.
A sample of all freshly prepared standard solutions except the last one was analyzed for concentration and homogeneity. A limited number of samples was analysed for stability.
Analysis was also performed on the dosing solutions of the first four weeks and the 11th week of dosing to determine the actual concentrations, uniformity of distribution (homogeneity) and stability of the formulations. - Duration of treatment / exposure:
- 90 days
- Frequency of treatment:
- drinking water at lib.
- Remarks:
- Doses / Concentrations:
0 ppm
Basis:
nominal in water - Remarks:
- Doses / Concentrations:
500 ppm
Basis:
nominal in water - Remarks:
- Doses / Concentrations:
2000 ppm
Basis:
nominal in water - Remarks:
- Doses / Concentrations:
7000 ppm
Basis:
nominal in water - No. of animals per sex per dose:
- 10
- Control animals:
- yes, concurrent vehicle
- other: 7000 ppm, 14 days recovery period
- Details on study design:
- 6 groups, each of 20 rats (ten males and ten females) were dosed as follows:
Group
Number Dose Level ppm Concentration
% Treatment
Period
0 0 0 90
1 500 0,05 90
2 2000 0,20 90
3 7000 0,70 90
4 0 0 90
5 7000 0,70 90
rationale for 7000 ppm satellite group: 14 days recovery to assess reversibility of effects
Doses were chosen on the base of a range finding study, the concentrations of 7000, 2000 and 500 ppm were estimated to result in a mean test substance intake of 1000, 250 and 50 mg/kg/day, respectively.
Administration:
Treatment period:
Recovery period:
2.4 Observations
2.4.1 Clinical observation
All animals were examined twice daily for general health state, behavioural changes, and toxicosis as well as for mortality. At weekends the animals were observed once a day. In addition, each animal was given a detailed clinical examination once a week. Any observed changes were recorded.
Ophthalmoscopy: In the week of acclimatization and prior terminal bleeding, the eyes of the rats in the control and the high dose groups were examined using an ophthalmoscope. One eye of each animal was treated with Mydriaticum Stulln® prior to investigation.
2.4.2 Bodyweight
Individual bodyweights were recorded at time of the allocation of animals to groups, prior to dosing on the day of commencement of treatment and subsequently at weekly intervals throughout the study. Last recordings of bodyweights were performed at the day of necropsy.
2.4.3 Food consumption
Food consumption in each cage was measured in weekly intervals throughout the study. Mean food intake per rat (g/rat/week) was calculated from the amount of food consumed in each cage and the number of rats in each cage. - Observations and examinations performed and frequency:
- Observations
Clinical observation
All animals were examined twice daily for general health state, behavioural changes, and toxicosis as well as for mortality. At weekends the animals were observed once a day. In addition, each animal was given a detailed clinical examination once a week. Any observed changes were recorded.
Ophthalmoscopy: In the week of acclimatization and prior terminal bleeding, the eyes of the rats in the control and the high dose groups were examined using an ophthalmoscope. One eye of each animal was treated with Mydriaticum Stulln® prior to investigation.
Bodyweight
Individual bodyweights were recorded at time of the allocation of animals to groups, prior to dosing on the day of commencement of treatment and subsequently at weekly intervals throughout the study. Last recordings of bodyweights were performed at the day of necropsy.
Food consumption
Food consumption in each cage was measured in weekly intervals throughout the study. Mean food intake per rat (g/rat/week) was calculated from the amount of food consumed in each cage and the number of rats in each cage.
Water consumption
Water consumption in each, cage was measured weekly. Mean achieved dosages (mg/kg bodyweight) of the test material were calculated by water consumption.
Monitoring of environment
Relative humidity and temperature of the animal rooms was monitored and recorded throughout the study. - Sacrifice and pathology:
- On completion of dosing period or recovery period, all animals were killed by C02 asphyxiation and exsanguination. Animals were fasted the night prior to necropsy. A complete autopsy including a macropathological examination was performed.
Organ weights:
The following organs from animals killed at the end of the study, dissected free from fat, were weighed before fixation:
Pathohistologv
Samples from the following tissues were removed from all animals and preserved in 10% formalin. Eyes of groups 0, 1, 2 and 3 were preserved in Davidson solution. The underlined organs from all high dose and control group animals were prepared as paraffin blocks, sectioned and stained for subsequent microscopic examination.
Adrenals Aorta (thoracic)
Anus
Brain
Bone
Caecum
Coagulation gland Colon
Concha (tattooed) Duodenum Epididymidis Eyes
Exorbital lacrimal glands
Gross lesions
Heart
Ileum
Jaw (upper)
Jejunum
Kidneys
Larynx
Liver
Lungs
Lymphnodes (skin, cervi¬cal & mesenteric) Mammary gland Muscle (skeletal)
Ovaries
Oesophagus
Pancreas
Pituitary
Prostate
Rectum
Salivary glands Sciatic nerve Seminal vesicles Skin
Spinal cord (cervical, mid thoracic, lumbal)
Spleen
Stomach
Testes
Thymus
Thyroid/Parathyroid
Tongue
Trachea
Urinary bladder
Uterus
Vagina - Other examinations:
- All animals were submitted to a terminal blood sampling for serum chemical and haematological investigations. Animals were not fasted prior to blood sampling. Blood samples were collected by orbital sinus puncture under light ether anaesthesia
Haematology:
The following parameters were measured on blood collected into containers supplied with EDTA:
Red blood cell count (RBC) 1012/1 = Tera/1
Total white blood cell count (WBC) 109/1 = Giga/1
Platelet count (PLT) 109/1 = Giga/1
Haemoglobin (HGB) mmol/1
Haematocrit (HCT) VI
Erythrocyte indices:
- mean corpuscular volume (MCV) fl
- mean corpuscular haemoglobin (MCH) mmol/1
- mean corpuscular haemoglobin concentration (MCHC) fmol
Differential white blood cell count %
Differential count evaluated for percentage of the different leucocytes:
Neutrophils (NEUTRO): metamyelocytes, band, segmented Eosinophils (EOS)
Basophils (BASO)
Lymphocytes (LYMF)
Monocytes (MONO)
Plasma cells (PLAS)
Unclassifiable cells (UNCLC)
Clinical chemistry of serum:
The following parameters were measured on serum from blood collected into containers supplied with a separating gel and subsequently separated by centrifugation:
Sodium (NA) mmol/1
Potassium (K) mmol/i
Calcium (CA) mmol/1
Aspartate aminotransferase (AST) U/l
Alanine aminotransferase (ALT) U/l
Alkaline phosphatase (AP) U/l
Glucose (GLUC) mmol/1
Cholesterol (CHOL) mmol/1
Triglycerides (TRIG) mmol/1
Total bilirubin (TBIL) fimol/1
Blood urea nitrogen (BUN) mmol/1
Creatinine (CREA) jxmol/1
Total protein (TPROT) g/1
Albumin (ALB) g/1
Urine analysis
For terminal urine sampling, animals were placed in metabolism cages for six hours. If no urine was discharged in this time, animals were forced to urinate by slight pressure on bladder. The following parameters were investigated:
Volume (VOL) ml
Specific gravity (SPGR) pH (pH)
The following parameters were investigated by a semi quantitative method:
Protein (PROT) minimum 0, maximum 3
Glucose (GLUC) minimum 0, maximum 4
Keton (KETON) minimum 0, maximum 3
Urobilinogen (UBG) minimum 0, maximum 4
Blood ingredients (BLOOD) minimum 0, maximum 3
A urine sediment analysis was performed on all samples.
The sediment was evaluated by a semi quantitative method where a minimum was 0, a maximum was 4:
Leucocytes (LEUCO)
Erythrocytes (ERY)
Bacteria (BAC)
Epithelial cells - squamous (ECSQ) Epithelial cells - renal (ECRE) Oxalate (OXA)
Triple phosphate (TRIP)
Carbonate (CARB)
Urate (URA)
Phosphate (PHOSP) - Statistics:
- Comparisons were performed between vehicle control (group 0) and dose groups 1, 2 and 3.
All data (bodyweight and organ weight, haematological, urine analysis and serum clinical chemistry data) were analysed by a one way analysis of variance (ANOVA) incorporating a Bartlett’s test for homogeneity of variance. Data showing heterogenous variances were analysed using a Kruskal Wallis test. In case of significance, a pair wise comparison between negative control and dose groups was performed using a Wilcoxon, Mann, and Whitney U-Test. In case of a not significant ANOVA, no further analysis was performed. In case of a significant ANOVA, group means were compared by a Scheffe Test (Gad, S.C.; Weil, C.S.; 1989). Only the significant differences to the control group are indicated in the mean tables.
Food and water consumption were measured cage wise. Mean food consumption and water consumption were calculated per animal and week. Data are expressed as absolute values and as percentages in difference from the control group. By further calculations with the data of water consumption, the mean test substance uptake [mg/kg/day] was calculated. Means of these data were calculated for every four weeks of any of the dose groups.
Significance levels are indicated as follows: * p < 0.05** p <0.01
Group means and standard deviation were calculated where appropriate. All results are presented in group mean tables and with the individual data. The results of the statistical calculations are presented in the mean tables and with the individual data. Individual data are part of the appendices.
Food conversion rate was calculated by the formula mean food consumption/week * 100 mean bodyweight gain/week
As shown in chapter 3, data evaluation and pathohistology did not reveal any major substance related changed in either dose group. Therefore, only a limited data evaluation was performed on the data of recovery group animals. - Clinical signs:
- no effects observed
- Mortality:
- no mortality observed
- Body weight and weight changes:
- no effects observed
- Food consumption and compound intake (if feeding study):
- no effects observed
- Food efficiency:
- no effects observed
- Water consumption and compound intake (if drinking water study):
- effects observed, treatment-related
- Description (incidence and severity):
- A clear and dose dependent increase in water consumption was observed in intermediate and high dose groups. Low dose group animals equally revealed a slight increase, however the difference to control animals was very small.
- Ophthalmological findings:
- no effects observed
- Haematological findings:
- no effects observed
- Clinical biochemistry findings:
- effects observed, treatment-related
- Description (incidence and severity):
- Statistically significant decreases were observed in serum sodium concentration in high dose group, intermediate dose males equally revealed a slight but not significant decrease.
- Urinalysis findings:
- no effects observed
- Behaviour (functional findings):
- no effects observed
- Organ weight findings including organ / body weight ratios:
- no effects observed
- Gross pathological findings:
- no effects observed
- Histopathological findings: non-neoplastic:
- no effects observed
- Histopathological findings: neoplastic:
- no effects observed
- Details on results:
- all effects noted were within historical control ranges or never reached clinically significant individual levels (e.g.serium sodium)
- Dose descriptor:
- LOAEL
- Effect level:
- >= 1 000 mg/kg bw/day (actual dose received)
- Based on:
- dissolved
- Sex:
- male/female
- Basis for effect level:
- other: No adverse effects were seen in males and females of teh top dose group (7000 ppm in drinking water). In these animals, the actual ingested dose, based on measure mean water consumption, was calculated to be around 1000 mg/kg bw.d
- Dose descriptor:
- LOAEL
- Effect level:
- >= 7 000 mg/L drinking water
- Based on:
- dissolved
- Sex:
- male/female
- Basis for effect level:
- other: No adverse effects were seen in males and females of teh top dose group (7000 ppm in drinking water). In these animals, the actual ingested dose, based on measure mean water consumption, was calculated to be around 1000 mg/kg bw.d
- Critical effects observed:
- not specified
- Conclusions:
- No findings were observed in the low dose (500 ppm) group except for a slight increase in water consumption. The intermediate (2000 ppm) dose group revealed a clear increase in water consumption. Urine examination revealed the presence of renal epithelial cells in urine sediment. High (7000 ppm) dose animals showed a clear increase in water consumption and the presence of renal epithelial cells and associated protein increase in urine. High dose females revealed a significantly increased relative liver weight, which however may be due to chance as no accompanying changes were observed. After a recovery period of 28 days, water consumption had declined to the level of the control group. Urine protein had the same level as in the control group. Presence of renal epithelial cells was equally reduced. No differences were observed concerning liver weight. Continuous intake of sodium with the test substance is thought to cause the increase in water consumption and the changes in renal parameters. These findings are regarded as adaptive responses and not as adverse effects, especially since the macroscopic and histological examination did not show any substance-related lesions.
Oral administration of the test material NATRIUM-METHALLYLSULFONAT to rats for a period of 90 days at a maximum dose level of 7000 ppm (about 1000 mg/kg) did not result in a clear pattern of clinical findings or the identification of a target organ for adverse effects. No mortalities occurred, and bodyweight did not differ between dose and control animals. - Executive summary:
NATRIUM-METHALLYLSULFONATwas administered by drinkingwaterat three dose levels to groups of ten male and ten female Wistar rats, Hsd/Win:WU strain. Administration was performed for 90 days, at dose levels of 500, 2000, and 7000 ppm corresponding to approximately 50, 250 and 1000 mg/kg/day. A control group of ten males and ten females was dosed with tap water alone.
A control and a high dose recovery group each consisting of 10 males and 10 females, respectively, were observed for a period of 28 days after the end of the dosing period.
Clinical signs, bodyweight, food and water consumption were monitored during the study. Blood chemistry and haematology as well as urine analysis were evaluated for all animals at the end of the study. Ophthalmological examinations were performed on all animals prior to the start and at the end of the study.
All animals were subjected to a gross necropsy examination and a limited pathohistological evaluation of tissues from high dose and controlanimalswas performed.
The results are summarised as follows:
Mortality data
No mortalities were observed during the dosing period.
Clinical observations
There were no signs observed in either dose group that could reasonably be attributed to test substance treatment.
Bodvweight
No differences in bodyweight development were observed between control and dose groups.Food consumption
No major differences were observed in mean food consumption between control and dose groups.Water consumption/test substance uptake
A clear and dose dependent increase in water consumption was observed in the intermediate and high dose groups. Low dose group animals equally revealed a slight increase, however, the difference to control animals was very small. With the ending of dosing, water consumption in the high (7000 ppm) dose recovery group declined to the level of the control group.
Solving of the test substance in water results in a corresponding amount of sodium ions. The reason for the increased water consumption is supposed to be the high sodium uptake in high and intermediate dose groups.
The concentrations of 7000, 2000 and 500 ppm resulted in a test substance uptake of about 1000, 250 and 50 mg/kg bodyweight/day, with females taking up a slightly higher amount and with decreasing test substance uptake in both sexes towards the end of the study.
Clinical chemistry
A statistical significant decrease in serum sodium was observed in the males of the high dose group. However, a hyponatremia can not be stated, since all data fell within the range of historical control data. A slight increase in sodium in high dose females equally was not associated with any other change which could reasonably be attributed to a clear disease state. In summary, it can be stated that the continuous intake of sodium with the test substance did not result in a clear pattern of changes of the serum sodium concentration.
Significantbut only slight alanine amino transferase (ALT) decreases were observed in low dose males. Due to the missing dose dependency this finding was not considered to be of biological significance. The statistically significant decreases of ALT in females of either dose group were also regarded as biologically not significant as the data were withinthe range of historical control data.
Other statistically significant differences calculated between control and treatment groups concerning creatinine (decrease in low and intermediate dose animals of either sex) and total protein (increase in high dose males) were considered of no toxicological concern, due to lacking dose dependency or missing biologically detrimental consequence.
Haematology
No significant differences were observed in males of either dose group, group means fell within the range of the historical control data.
A statistically significant increase of mean corpuscular volume (MCV) was observed in low dose females. As group mean and all individual values were within the range of the historical control data, this finding was not considered of biological significance.
Other minor haematological changes were observed in several individuals, however, as no dose dependency occurred and as these findings were observed in both, dose and control groups, they were not considered to be test substance related.
Urine analysis
Renal epithelial cells were observed in the urine sediment from several intermediate and high dose animals. Probably related to this finding is an increase in urine protein observed in high dose females. These findings might be interpreted as indicative of an active renal disease. However, such an interpretation is not confirmed by the histological examination. In addition, the number of animals concerned as well as the severity of the findings decreased in the recovery group. Therefore, the presence of renal epithelial cells in the urine sediment and the increase in urine protein are considered as expression of adaptive responses to the high sodium load rather than as adverse effects. Other observed urine analysis findings or calculated differences were not considered to be test substance related due to lacking dose dependency or because all individual values were within the range of the historical control data.
Organ weights
In high dose females, a statistically significant increase in relative liver weight was observed. Thisfinding was not accompanied by any corresponding pathohistological findings. In addition, no overt changes were observed in clinical chemistry parameters usually related to liver cell damage, e.g. Alanine aminotransferase or Aspartate aminotransferase. The reason for this increase therefore remains unclear, and may be due to chance. In the female recovery groups, no statistically significant or otherwise striking differences concerning absolute or bodyweight adjusted liver weight were observed between control and high (7000 ppm) dose groups. No other differences in organ weights were observed at the end of the dosing period.
Macroscopical and histological examination
No macroscopic lesions considered to be related to treatment were observed. Changes which were recorded were considered part of the normal background in the rat.
The pathohistological findings consisted of spontaneous lesions in the male and female animals of all groups such as hydrometriosis of the uterus, calcification of the Peyer’s patches, focal histiocytes in the liver, and multifocal lymphocytes in the lung. There was also pigmentation in the lymphnodes cervicales caused by tattooing ears.
Extramedulary haematopoiesis of the liver was observed in animals of all groups examined (groups
0, 3; male and female animals) which is considered as a normal finding in this age.
Summarizing, no substance-related lesions were observed by the macroscopical and pathohisto- logical examination of high dose male and female animals in comparison to the control animals.
Conclusions
No findings were observed in the low dose (500 ppm) group except for a slight increase in water consumption. The intermediate (2000 ppm) dose group revealed a clear increase in water consumption. Urine examination revealed the presence of renal epithelial cells in urine sediment. High (7000 ppm) dose animals showed a clear increase in water consumption and the presence of renal epithelial cells and associated protein increase in urine. High dose females revealed a significantly increased relative liver weight, which however may be due to chance as no accompanying changes were observed. After a recovery period of 28 days, water consumption had declined to the level of the control group. Urine protein had the same level as in the control group. Presence of renal epithelial cells was equally reduced. No differences were observed concerning liver weight. Continuous intake of sodium with the test substance is thought to cause the increase in water consumption and the changes in renal parameters. These findings are regarded as adaptive responses and not as adverse effects, especially since the macroscopic and histological examination did not show any substance-related lesions.
Oral administration of the test material NATRIUM-METHALLYLSULFONAT to rats for a period of 90 days at a maximum dose level of 7000 ppm (about 1000 mg/kg) did not result in a clear pattern of clinical findings or the identification of a target organ for adverse effects. No mortalities occurred, and bodyweight did not differ between dose and control animals. Pathohistological evaluation did not reveal any test substance related findings in either kidneys or in liver, or in any other organ, respectively. No toxicologically significant effects were demonstrated in animals treated with 7000 ppm. Therefore, the "No Observed Adverse Effect Level" (NOAEL) is considered to be 7000 ppm.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed
- Dose descriptor:
- NOAEL
- 1 000 mg/kg bw/day
- Study duration:
- subchronic
- Species:
- rat
- Quality of whole database:
- good
Repeated dose toxicity: inhalation - systemic effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: inhalation - local effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: dermal - systemic effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: dermal - local effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Additional information
A well-conducted oral subchronic (90 days) study in rats did not show any effects on any organ at a dose of 1000 mg/kg bw.
a subchronic inhalation study is not scientifically justifiable since a) the substance does not contain any particulate matter with a MMAD < 10 micron, therefore inhalation exposure is not an issue and b) the oral subchronic study did not show any effects at 1000 mg/kg bw; therefore inhaled particles that are trapped in the mucociliary elevator and subsequently swallowed, will not cause appreciable toxicity since a dose higher than 1000 mg/kg bw cannot be administrated in this fashion
A subschronic dermal study is not indicated since a) dermal exposure does not occur and b) the substance is not a skin irritant and does not show any effects in an acute dermal toxicity study at a dose of 1000 mg /kg bw
Justification for classification or non-classification
substance need not be classified since no effects were observed at 1000 mg/kg in an oral 90 -day study and consequently no target organs could be identified.
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