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EC number: 843-143-1 | CAS number: 709647-81-4
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Toxicity to aquatic algae and cyanobacteria
Administrative data
Link to relevant study record(s)
- Endpoint:
- toxicity to aquatic algae and cyanobacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 18 February 2020 - XX August 2020
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- The study was conducted in accordance with GLP and to the relevant OECD Test Guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
- Version / remarks:
- 2006
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.3 (Algal Inhibition test)
- Deviations:
- no
- GLP compliance:
- yes
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: The concentration and stability of the test item in the test preparations (100 mg/L only in the full main study) were verified by chemical analysis at 0 and 72 h.
- Sampling method: Samples were taken from the control and test groups from the bulk test preparation at 0 h and from the pooled replicates at 72 h for immediate quantitative analysis. A set of duplicate samples were taken at 0 and 72 h and stored frozen for further analysis if necessary.
- Sample storage conditions before analysis: The 0 h test samples were analysed on the day of sampling and stored refrigerated at approx. 4 ºC prior to analysis. The 72 h test samples were analysed on the day of sampling. - Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Based on the range-finder results (see 'Details on test conditions'), a limit test was conducted. A nominal amount of test item (100 mg) was dissolved in culture medium and the volume adjusted to 1000 mL to give a 100 mg/L stock solution. An aliquot (1000 mL) of the stock solution was separately inoculated with algal suspension (3.1 mL) to give an initial nominal cell density of 5.00 x 10^3 cells/mL.
- Eluate: N/A
- Differential loading: N/A
- Controls: Yes: A positive control test using potassium dichromate as the reference item was performed twice in a 12 month period to demonstrate satisfactory conditions of the test. A concurrent negative control group was maintained under identical conditions but not exposed to the test item.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N/A
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): N/A
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): The stock solution prepared concentration was inverted several times to ensure adequate mixing and homogeneity. Preliminary solubility work showed that inversion was sufficient to allow the test item to fully dissolve.
- Other relevant information: N/A - Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: Green alga
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Cultures of R. subcapitata were obtained from the Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Scottish Marine Institute, Oban, Argyll, Scotland.
- Age of inoculum (at test initiation): Approx. 3 - 4 days old
- Method of cultivation: Approx.3 to 4 days before the test start, inoculum cultures of algae was set up at an initial cell density of approx. 10^3 cells/mL. The flasks were plugged with polyurethane foam stoppers and kept under constant agitation by orbital shaker (approximately 150 rpm) and constant illumination at 24 ºC until the algal cell density was approximately 10^5 to 10^6 cells/mL.
ACCLIMATION
- Acclimation period: N/A
- Culturing media and conditions (same as test or not): The culture medium used for both the range-finding and definitive tests was the same as that used to maintain the stock culture.
- Any deformed or abnormal cells observed: None reported - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- yes
- Total exposure duration:
- 72 h
- Remarks on exposure duration:
- Guideline specified
- Post exposure observation period:
- None - not required.
- Test temperature:
- 24 ºC
- pH:
- Control: 8.0 - 8.2
100 mg/L: 7.3 - 7.8 - Dissolved oxygen:
- Not reported
- Salinity:
- Not reported/not relevant
- Conductivity:
- Not reported
- Nominal and measured concentrations:
- Nominal: 100 mg/L; procedural recoveries 1 and 2: 105 %
Measured:
0 h: 99.6 mg/L; procedural recovery 1: 111 % and 2: 101 %;
72 h: 109 mg/L
The concentrations for the procedural recovery are the actual fortified concentrations. The corresponding percentages of nominal concentration values relate to the percentage of fortified concentration recovered. Analysis of the test preparations at 0 and 72 h showed measured test concentrations to be near nominal, therefore the results are based on the nominal concentrations. - Details on test conditions:
- TEST SYSTEM
- Test vessel: Conical flask
- Type: Closed - plugged with polyurethane foam bungs to reduce evaporation
- Material, size, headspace, fill volume: Glass, 250 mL, containing 100 mL test preparation
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): N/A
- Renewal rate of test solution (frequency/flow rate): None, static test.
- Initial cells density: 5.00 x 10^3 cells/mL (nominal)
- Control end cells density: The control group was maintained under identical conditions but not exposed to the test item. Mean cell density at 0 h: 5.59 x 10^3; at 72 h: 3.37 x 10^5
- No. of organisms per vessel: See "initial and end cell densities"
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): N/A
GROWTH MEDIUM
- Standard medium used: yes (AAP-medium, U.E. EPA). The culture medium was prepared using reverse osmosis purified deionized water (Elga Optima 15+ or Elga Purelab Option R-15 BP)
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Not specified
- Culture medium different from test medium: No, the culture medium used for both the range-finding and definitive tests was the same as that used to maintain the stock culture.
- Intervals of water quality measurement: pH was measured for the control and each test preparation at initiation of the test and after 72 h exposure. The pH was measured using a Hach HQ30d Flexi handheld meter. The temperature within the incubator was recorded daily. Appearance of the test media was also recorded daily.
OTHER TEST CONDITIONS
- Sterile test conditions: Not reported
- Adjustment of pH: pH was adjusted to 7.5 ± 0.1 with 0.1N NaOH or HCl.
- Photoperiod: Continuous illumination provided by warm white lighting (380 to 730 nm)
- Light intensity and quality: Approx. 7000 lux
The flasks were plugged with polyurethane foam bungs and incubated (INFORS Multitron incubator) at 24 °C and constantly shaken at approximately 150 rpm for 72 h.
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Samples of the algal populations were removed daily (at 24, 47 and 72 h) and cell concentrations determined for each control and treatment group, using a Coulter® Multisizer Particle Counter (v3.53). Three determinations were made for each sample. Growth rate and yield data were inspected and ECx values determined.
- Chlorophyll measurement: N/A
- Other: Shape and size of algal cells was inspected microscopically and any abnormalities recorded.
TEST CONCENTRATIONS
- Spacing factor for test concentrations: N/A
- Justification for using less concentrations than requested by guideline: N/A
- Range finding (R-F) study:
The range-finding test was conducted by exposing R. subcapitata cells to nominal test concentrations of 0.10, 1.0, 10 and 100 mg/L for a period of 72 h. The test was conducted in 250 mL glass conical flasks each containing 100 mL of test preparation and plugged with polyurethane foam bungs to reduce evaporation. Two replicate flasks were used for each control and test concentration. The test item was dissolved directly in culture medium. A nominal amount of test item (100 mg) was dissolved in culture medium and the volume adjusted to 1000 mL to give a 100 mg/L stock solution, from which a series of dilutions was made to give further stock solutions of 0.10, 1.0 and 10 mg/L. An aliquot (450 mL) of each of the stock solutions was separately inoculated with algal suspension (2.3 mL) to give an initial nominal cell density of 5.00 x 103 cells/mL. Prior to use, the test item was heated at 80 ºC to aid weighing. The stock solutions and each test concentration were inverted several times to ensure adequate mixing and homogeneity. Preliminary solubility work showed that inversion was sufficient to ensure that test item was fully dissolved. At the start of the range-finding test a sample of each test and control culture was removed and the cell density determined using a Coulter® Multisizer Particle Counter. The flasks were then plugged with polyurethane foam bungs and incubated (INFORS Multitron incubator) at 24 oC under continuous illumination (intensity approx. 7000 lux) provided by warm white lighting (380 to 730 nm) and constantly shaken at approximately 150 rpm for 72 h.
After 72 h the cell density of each flask was determined using a Coulter® Multisizer Particle Counter. A sample of each test concentration was taken for chemical analysis at 0 and 72 h in order to determine the stability of the test item under test conditions.
- R-F test concentrations: 0.10, 1.0, 10 and 100 mg/L (nominal)
- R-F results used to determine the conditions for the definitive study: The results showed no effect on growth at the test concentrations of 0.10, 1.0, 10 and
100 mg/L. Based on the result of the R-F test, a "limit test" was conducted at a concentration of 100 mg/L to confirm that at the maximum concentration given in the OECD/EC Test Guidelines, no effect on algal growth was observed. - Reference substance (positive control):
- yes
- Remarks:
- potassium dichormate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Key result
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- >= 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- >= 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Details on results:
- - Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): None observed
- Unusual cell shape: None observed
- Colour differences: After 72 h, all control and test cultures were pale green dispersions.
- Flocculation: Not reported
- Adherence to test vessels: Not reported
- Aggregation of algal cells: Not reported
- Other: From the results obtained, it is concluded that the growth rate (r) and yield (y) of R. subcapitata (CCAP 278/4) were not inhibited by the presence of the test item at a nominal test concentration of 100 mg/L over the 72 h exposure period.
- Any stimulation of growth found in any treatment: Yes
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No - Results with reference substance (positive control):
- - Results with reference substance valid? Yes
- ErC50: (0 to 72 h) 1.2 mg/L
- Other:
Exposure of Raphidocelis subcapitata (CCAP 278/4) to the reference item gave the following results:
ErC50 (0 to 72 hour) : 1.2 mg/L; 95% confidence limits 1.1 to 1.3 mg/L
EyC50 (0 to 72 hour) : 0.51 mg/L; 95% confidence limits 0.45 to 0.58 mg/L
No Observed Effect Concentration based on growth rate: 0.25 mg/L
No Observed Effect Concentration based on yield: 0.125 mg/L
Lowest Observed Effect Concentration based on growth rate: 0.50 mg/L
Lowest Observed Effect Concentration based on yield: 0.25 mg/L
The results from the positive control with potassium dichromate were within the normal ranges for this reference item based on in-house data. - Reported statistics and error estimates:
- A Students t-test was carried out on the growth rate and yield data after 72 h for the control and the 100 mg/L test concentration to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package v8.2 (SAS, 1999 - 2001).
ErCx was determined as the test concentration that reduced growth rate by x %. A significant difference was found between the control and the test group, however, this was due to an increase in growth and therefore the NOEC based on growth rate was 100 mg/L.
EyCx was determined as the test concentration that reduced yield by x %. A significant difference was found between the control and test group however, this was due to an increase in yield and therefore the NOEC based on yield was 100 mg/L. - Validity criteria fulfilled:
- yes
- Remarks:
- The control culture cell concentration increased by a factor of 70 after 72 h. The mean coefficient of variation for section by section specific growth rate for the control cultures was 8 %. The coefficient of variation for average specific growth rat
- Conclusions:
- Based on the conditions of the test, the test item ErC50 and EyC50 were determined to be > 100 mg/L and the NOEC was determined to be ≥ 100 mg/L.
- Executive summary:
A test was performed in accordance with OECD 201 (2006), in order to determine the aquatic toxicity of the test item to algae.
Following a preliminary range-finding test, Raphidocelis subcapitata (strain CCAP 278/4) was exposed to an aqueous solution of the test item at a nominal concentration of 100 mg/L (6 replicate flasks) for 72 h, under constant illumination and shaking at a temperature of 24 °C. Inoculation of 1000 mL of test medium with 3.1 mL of this algal suspension gave an initial nominal cell density of 5.00 x 103 cells per mL.
Samples of the algal populations were removed daily (24, 47 and 72 h) and cell concentrations determined for each control and treatment group, using a Coulter® Multisizer Particle Counter. The shape and size of the algal cells was inspected microscopically and any abnormalities recorded. pH was recorded at the start and end of the test.
A concurrent negative control was run in the test, and a positive control test using potassium dichromate as the reference item was performed twice in a 12 month period to demonstrate satisfactory conditions of the test.
Chemical analysis of the test preparations at 0 and 72 h showed measured test concentrations to be 100 and 105 % of nominal respectively and so the results are based on nominal test concentrations only.
The results of the test were considered valid based on fulfillment of the following performance criteria:
- The data demonstrated that the cell concentration of the control cultures increased by a factor of 70 after 72 h. This increase was in line with the OECD Guideline that states the increase must be at least by a factor of 16 after 72 h (nominal cell density of control at 0 h: 5.00 x 103 cells/mL; mean cell density of control at 72 h: 3.50 x 105 cells/mL).
- The mean coefficient of variation for section by section specific growth rate for the control cultures was 8 % (i.e. <= 35 % stated in OECD Guideline).
The coefficient of variation for average specific growth rate for the control cultures over the test period (0 to 72 h) was 3 % (i.e. <= 7 % stated in OECD Guideline).
Following microscopic inspection, no abnormalities were detected in any test and control cultures. Growth rate (r) and yield (y) of R. subcapitata were not affected by the presence of the test item at a nominal test concentration of 100 mg/L over the 72 h exposure period. EC50 values were based on the nominal test concentrations of greater than 100 mg/L. The NOEC was concluded to be ≥ 100 mg/L. The ErC50, EyC50, ErC10 and EyC10 were all concluded to be > 100 mg/L.
Under the conditions of this study, the test item does not cause toxicity to freshwater algae and would not be classified under CLP Regulation (EC) 1272/2008 based on freshwater algae alone.
Reference
Analysis of the test preparations at 0 and 72 hours showed measured test concentrations of 100 and 105 % of nominal respectively, therefore the results are based on the nominal concentrations.
The 72 h ErC10, ErC20, ErC50 (growth rate) were all concluded to be > 100 mg/L. The 72 h EyC10, EyC20, EyC50 (yield) were all concluded to be > 100 mg/L. The NOEC was concluded to be 100 mg/L.
Table 1. Cell densities and pH values in the definitive test
Nominal Concentration (mg/L) | pH | Cell Densities* (cells per mL) | pH |
Temperature ºC
| |||
0 h | 24 h | 47 h | 72 h | 72 h | 0 - 72 h | ||
Control | R1 | 8.0 | 1.61E+04 | 6.40E+04 | 3.10E+05 | 8.2 | 24 ºC |
R2 | 1.99E+04 | 8.38E+04 | 3.47E+05
| ||||
R3 | 1.93E+04 | 7.89E+04 | 3.91E+05 | ||||
R4 | 1.93E+04 | 7.44E+04 | 3.51E+05 | ||||
R5 | 1.83E+04 | 6.74E+04 | 3.12E+05 | ||||
R6 | 1.84E+04 | 8.82E+04 | 3.89E+05 | ||||
Mean | 1.85E+04 | 7.61E+04 | 3.50E+05 | ||||
100 | R1 | 7.3 | 2.35E+04 | 1.23E+05
| 6.90E+05 | 7.8 | 24 ºC |
R2 | 2.59E+04 | 1.21E+05 | 6.61E+05 | ||||
R3 | 2.47E+04 | 8.84E+04 | 4.47E+05 | ||||
R4 | 2.17E+04 | 8.76E+04 | 3.73E+05 | ||||
R5 | 2.34E+04 | 1.21E+05 | 5.68E+05 | ||||
R6 | 2.00E+04 | 1.04E+05 | 6.13E+05 | ||||
Mean | 2.32E+04 | 1.08E+05 | 5.59E+05 |
*Cell densities represent the mean number of cells per mL calculated from the mean of the cell counts from 3 counts for each of the replicate flasks
R = replicate
Table 2. Inhibition of Growth Rate and Yield in the Definitive Test
Nominal Concentration (mg/L) | Growth rate (cells/mL/h) | Yield (cells/mL) | |||
0 to 72 h
| % Inhibition
| 0 to 72 h
| % Inhibition*
| ||
Control | R1 | 0.057 | - | 3.05E+05 | - |
R2 | 0.059 | 3.42E+05 | |||
R3 | 0.061 | 3.86E+05 | |||
R4 | 0.059 | 3.46E+05 | |||
R5 | 0.057 | 3.07E+05 | |||
R6 | 0.060 | 3.84E+05 | |||
Mean | 0.059 | 3.45E+05 | |||
SD | 0.002 | 3.51E+04 | |||
100 | R1 | 0.068 | [15] | 6.85E+05 |
|
R2 | 0.068 | [15] | 6.56E+05 |
| |
R3 | 0.062 | [5] | 4.42E+05 |
| |
R4 | 0.060 | [2] | 3.68E+05 |
| |
R5 | 0.066 | [12] | 5.63E+05
|
| |
R6 | 0.067 | [14] | 6.08E+05 |
| |
Mean | 0.065 | [11] | 5.54E+05 | [61] | |
SD | 0.003 | - | 1.25E+05 |
|
*In accordance with the OECD test guideline only the mean value for yield for each test concentration is calculated
R = Replicate
SD = Standard Deviation
[] = Increase in growth compared to controls
- = N/A
Description of key information
OECD 201:
ErC50 > 100 mg/L, ErC10 > 100 mg/L and NOEC ≥ 100 mg/L
EyC50 > 100 mg/L, EyC10 > 100 mg/L and NOEC ≥ 100 mg/L
(Ablitt, 2020)
Key value for chemical safety assessment
Additional information
- The data demonstrated that the cell concentration of the control cultures increased by a factor of 70 after 72 h. This increase was in line with the OECD Guideline that states the increase must be at least by a factor of 16 after 72 h (nominal cell density of control at 0 h: 5.00 x 103 cells/mL; mean cell density of control at 72 h: 3.50 x 105 cells/mL).
- The mean coefficient of variation for section by section specific growth rate for the control cultures was 8 % (i.e. <= 35 % stated in OECD Guideline).
The coefficient of variation for average specific growth rate for the control cultures over the test period (0 to 72 h) was 3% (i.e. <= 7 % stated in OECD Guideline).
A test was performed in accordance with OECD 201 (2006), in order to determine the aquatic toxicity of the test item to algae.
Following a preliminary range-finding test, Raphidocelis subcapitata (strain CCAP 278/4) was exposed to an aqueous solution of the test item at a nominal concentration of 100 mg/L (6 replicate flasks) for 72 h. A concurent negative control was run and results from a seperate study was available for a positive control.
Samples of the algal populations were removed daily (24, 47 and 72 h) and cell concentrations determined for each control and treatment group, using a Coulter® Multisizer Particle Counter. The shape and size of the algal cells was inspected microscopically and any abnormalities recorded.
Chemical analysis of the test preparations at 0 and 72 h showed measured test concentrations to be 100 and 105 % of nominal, respectively, so the results are based on nominal test concentrations.
The results of the test were considered valid based on fulfillment of the following performance criteria as precribed by the OECD 201 (2006 Annex 5 corrected 2011):
Following microscopic inspection, no abnormalities were detected in any test and control cultures. Growth rate (r) and yield (y) of R. subcapitata were not affected by the presence of the test item at a nominal test concentration of 100 mg/L over the 72 h exposure period. The NOEC was concluded to be ≥ 100 mg/L. The ErC50, EyC50, ErC10 and EyC10 were all concluded to be > 100 mg/L.
Under the conditions of this study, no effects on growth or yield were observed and therefore it was concluded that the test item does not cause toxicity to freshwater algae. The test item would therefore not be classified under CLP Regulation (EC) 1272/2008 for aquatic toxicity, based on freshwater algae alone.
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