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EC number: 215-947-7 | CAS number: 1458-18-0
- 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
- Endpoint:
- toxicity to aquatic algae and cyanobacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 02 April 2013 - 25 April 2013
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 013
- Report date:
- 2013
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Alga, Growth Inhibition Test)
- Version / remarks:
- 2011
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000.
- Deviations:
- no
- GLP compliance:
- yes
Test material
- Reference substance name:
- Methyl 3-amino-5,6-dichloropyrazine-2-carboxylate
- EC Number:
- 215-947-7
- EC Name:
- Methyl 3-amino-5,6-dichloropyrazine-2-carboxylate
- Cas Number:
- 1458-18-0
- Molecular formula:
- C6H5Cl2N3O2
- IUPAC Name:
- methyl 3-amino-5,6-dichloropyrazine-2-carboxylate
- Test material form:
- solid: particulate/powder
- Details on test material:
- - Name of test material (as cited in study report): Amiloride Compound 7
- Description: Tan powder
- Storage condition of test material: In refrigerator protected from light, desiccated
- Other: Sample is hygroscopic and reactive to light and oxygen
Constituent 1
- Specific details on test material used for the study:
- Details on properties of test surrogate or analogue material (migrated information):
Purity/composition correction factor required: No
Hygroscopic: Yes, store desiccated
Volatile: No
Reactivity: Reactive to light, moisture and oxygen
Test substance handling: No specific handling required for moisture and oxygen, Use amber-coloured glassware or wrap container in aluminium-foil Stability in water: Unknown
Solubility in water: Insoluble
Sampling and analysis
- Analytical monitoring:
- yes
- Details on sampling:
- Samples for possible analysis were taken from all test concentrations and the control according to the schedule below. In addition, the filter containing undissolved residue was kept for possible analysis.
Frequency:at t=0 h, t=24 h and t=72 h
Volume: 2.0 ml
Storage : Samples were stored in a freezer until analysis.
At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling.
Compliance with the Quality criteria regarding maintenance of actual concentrations was demonstrated by running a test vessel at the highest substance concentration but without algae and samples for analysis were taken at the start, after 24 hours of exposure and at the end of the test period.
Additionally, reserve samples of 2.0 ml were taken from all test solutions for possible analysis. If not already used, these samples were stored in a freezer for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.
Test solutions
- Vehicle:
- no
- Details on test solutions:
- The batch of Amiloride Compound 7 tested was a tan powder with a purity of 99.7 area% and not completely soluble in test medium at the loading rate initially prepared.
Preparation of test solutions started with a loading rate of 100 mg/l applying a two-day period of magnetic stirring to ensure maximum dissolution in the test medium. The resulting dispersion was subsequently filtered through a 0.45 µm membrane filter (Whatman, RC55) to remove the undissolved fraction of test substance. The test concentrations were prepared by subsequent dilutions of the filtrate in test medium. The final test solutions were all clear and colourless.
Note that due to light sensitivity of the test substance all handlings were performed under dimmed or yellow light and the glassware covered in aluminium foil.
After preparation, volumes of 50 ml were added to each replicate of the respective test concentration. Subsequently, 1 ml of an algal suspension was added to each replicate providing a cell density of 10E4 cells/ml.
Test organisms
- Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: NIVA CHL 1.
- Source (laboratory, culture collection): In-house laboratory culture
- Age of inoculum (at test initiation):3 days before the start of the test, cells from the algal stock culture were inoculated in culture medium at a cell density of 1 x 10E4 cells/ml.
- Method of cultivation: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light (60 to 120 µE/m2/s when measured in the photosynthetically effective wavelength range of 400 to 700 nm. in a climate room at a temperature of 21-24°C.
Study design
- Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 72 h
- Post exposure observation period:
- no
Test conditions
- Hardness:
- 0.24 mmol(24 mg CaCO3/L)
- Test temperature:
- between 21.7 and 22.9°C
- pH:
- T=0 h between 8.1-8.2
T=72 h between 7.9-8.1 - Nominal and measured concentrations:
- Nominal concentrations: control and 0.32, 1.0, 3.2, 10 and 32% of a 0.45 μm filtered solution prepared at a loading rate of 100 mg/l
Measured concentrations:The actual test concentrations at the start of the test were 0.028, 0.072, 0.22, 0.64 and 2.1 mg/l in the solutions containing 0.32, 1.0, 3.2, 10 and 32% of the filtrate, respectively. The actual concentrations in solutions containing 3.2 to 32% of the filtrate decreased to 19-54% of initial at the end of the test. The measured concentration in the remaining test groups were below the limit of detection of the analytical method (LOD=0.0044 mg/l) at the end of the test. Calculated Time Weight Average concentrations were 0.028, 0.072, 0.22, 0.64 and 2.1 mg/l. - Details on test conditions:
- TEST SYSTEM
- Type: open
- Material, size, headspace, fill volume: 100 ml, normal headspace, 50 ml
- Aeration: no
- Initial cells density: 10000 cells/ml
- Control end cells density: 2336000 cells/ml
Replicates:
6 replicates of the control,
3 replicates of each test concentration,
2 replicates of the undiluted filtrate without algae,
1 replicate of each test concentration for sampling purposes
GROWTH MEDIUM
- Standard medium used: yes
OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: Continuously using TLD-lamps of the type ‘Cool-white’ of 30 Watt, with a light intensity within the range of 91 to 99 µE.m-2.s-1
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
72 h NOErC, 72 h NOEyC, 72 h ErC10, 72 h EyC10, 72 h ErC50, 72 h EyC50
TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Test concentrations: control and 0.32, 1.0, 3.2, 10 and 32% of a 0.45 μm filtered solution prepared at a loading rate of 100 mg/l. - Reference substance (positive control):
- yes
- Remarks:
- potassium dichromate (K2Cr2O7)
Results and discussion
Effect concentrationsopen allclose all
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- 0.028 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: -
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 0.048 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: 95% CI: 0.015 -0.15 mg/l TWA
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- 0.3 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: 95% CI: 0.10 - 0.91 mg/l TWA
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- 0.028 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: -
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 0.025 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: 95% CI: 0.019- 0.034 mg/l TWA
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- 0.092 mg/L
- Nominal / measured:
- meas. (TWA)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: 95% CI:0.068 - 0.12 mg/l TWA
- Details on results:
- Exponential growth in the control (for algal test): yes
Obeservation of abnormalities (for algal test): no abnormalities observed.
Unusual cell shape: no
Colour differences: no - Results with reference substance (positive control):
- Validity criteria met: yes
The EC50 for growth rate reduction (ERC50: 0-72h) was 0.97 mg/l with a 95% confidence interval ranging from 0.62 to 1.5 mg/l. The historical ranges for growth rate reduction lie between 0.82 and 2.3 mg/l. Hence, the ERC50: 0-72h for the algal culture tested corresponds with this range.
- Reported statistics and error estimates:
- For determination of the NOEC and the EC50 the approaches recommended in the OECD guideline 201 were used. An effect was considered to be significant if statistical analysis of the data obtained for the test concentrations compared with those obtained in the negative control revealed significant reduction of growth rate or inhibition of yield (ANOVA, Bonferroni t-test, TOXSTAT Release 3.5, 1996, D.D. Gulley, A.M. Boelter, H.L. Bergman). Additionally, the EC10 was determined to meet the recommendations as put down in "A Review of Statistical Data Analysis and Experimental Design in OECD Aquatic Toxicology Test Guidelines" by S. Pack, August 1993.
Calculation of the EC50 and EC10 values was based on log-linear regression analysis of the percentages of growth rate reduction and the percentages of yield inhibition versus the logarithms of the corresponding TWA concentrations of the test substance.
Applicant's summary and conclusion
- Validity criteria fulfilled:
- yes
- Conclusions:
Amiloride Compound 7 reduced growth rate and inhibited the yield of this fresh water algae species significantly at a TWA concentration of 0.12 mg/l and higher.
The EC50 for growth rate reduction (72h-ERC50), based on TWA concentrations, was 0.30 mg/l with a 95% confidence interval ranging from 0.10 to 0.91 mg/l.
The EC50 for yield inhibition (72h-EYC50), based on TWA concentrations, was 0.092 mg/l with a 95% confidence interval ranging from 0.068 to 0.12 mg/l.
The 72h-NOEC for both growth rate reduction and yield inhibition was, based on the TWA concentration, 0.028 mg/l.
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