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EC number: 627-025-5 | CAS number: 101541-04-2
- 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:
- 02 October 2018 - 05 October 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- The test was carried out according to GUIDELINE OECD 201 (2006, corrected 28 July 2011)
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
- Version / remarks:
- GUIDELINE OECD 201 (2006, corrected 28 July 2011)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Batch number :1700000153
- Expiration date of the lot/batch: 25 January 2020
- Purity test date: January 2017
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Stored in Darkness at a temperature of 20°C
- Solubility and stability of the test substance in the solvent/vehicle: the stability (non-GLP)
was checked for reference and test material stock solutions after 8 days at - 20°C +/- 5°C. Recoveries (n=2) were higher than 90%.
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Since the test item is poorly soluble under the test conditions and according to the OECD 23, the choice of a solvent was necessary to solubilize the test item, at the concentration lower than 0.1 ml.L-1. So, a stock solution of test item was prepared in dimethyl sulfoxide. Test solutions were prepared by dilution of this solution.
FORM AS APPLIED IN THE TEST (if different from that of starting material)
The test material was applied as liquid after solubilization in dimethyl sulfoxide (DMSO) - Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: limit test at 5 mg/L (nominal concentration) correspond to 2.56 mg/L (geometric mean concentration)
- Sampling method: Test item solution and control were sampled and immediately analysed at 0h, 24h, 48h, and 72h.
- Sample storage conditions before analysis: no storage before analysis - Vehicle:
- yes
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:Dilution of the stock solution prepared by dissolving 501 mg of test item in 10 mL of dimethyl sulfoxide(DMSO), concentration of stock solution = 50 g/L
- Chemical name of vehicle (organic solvent, emulsifier or dispersant):Dimethyl Sulfoxide
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): according to the OECD monograph 23, the solvent concentration was not exceeded 0.1 mL.L-1 in the test and control solutions. - Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: Freshwater green Algae
- Strain: Pseudokirchneriella subcapitata
- Source (laboratory, culture collection): CCAP (Culture Collection of Algae and Protozoa),
reference CCAP 1405/1.
- Method of cultivation: A culture was prepared from the stock culture and maintained in an exponential growth by successive transfers every 3 to 4 days.
ACCLIMATION
- Culturing media and conditions (same as test or not):
Medium: OECD medium
Temperature: 23 ± 2°C
Light: continuous illumination between 4440 and 8880 lux
pH: 7,80
Any deformed or abnormal cells observed: No abnormal appearance of the algae at the start and at the end of the test. - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- yes
- Total exposure duration:
- 72 h
- Post exposure observation period:
- Microscopic observation:
No abnormal appearance of the algae at the start and at the end of the test. - Test temperature:
- 0h (23.75 -24.25 (°C)) ; 24h (24.13 - 24.31 (°C)) ; 48h (23.06 - 24.31 (°C)) ; 72h ( 23.31 - 24.25 (°C))
- pH:
- pH : 0h Control (7.8) Solvent control (7.69) Test material at [0.1] (7.67)
pH : 72 h Control (8.67) Solvent control ( 8.66) Test material at [0.1] (8.63) - Nominal and measured concentrations:
- Limit test concentration: 5 mg/L in nominal concentration and 2.56 mg/L geometric mean concentration
- Details on test conditions:
- TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Type (delete if not applicable): open
- Material, size, headspace, fill volume:Glass Erlenmeyer flasks of 250 mL. .
- Initial cells density: 25 000 cells per millilitres.
- Control end cells density: 3 203 333 cells per millilitres.
- No. of organisms per vessel: 25 000 cells per millilitres.
- No. of vessels per concentration (replicates):6 replicates for each
treatment
GROWTH MEDIUM
- Standard medium used: yes (OECD test medium)
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: It was reconstituted with chemical reagents and ultra-pure water
OTHER TEST CONDITIONS
- Sterile test conditions: No
- Photoperiod: Continuous illumination
- Light intensity and quality: Between 4440 and 8880 lux
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Beckman coulter Multisizer 3
TEST CONCENTRATIONS
- Spacing factor for test concentrations: limit test
- Justification for using less concentrations than requested by guideline: The toxicity was observed under the limit of solubility during the range finding-test
- Range finding study: 16.33 µg/L; 285.87 µg/L and 2770.53 µg/L
- Test concentrations: 5 mg test item.L-1. - Reference substance (positive control):
- yes
- Remarks:
- Potassium dichromate results: Starting date :18 May 2018 CEt50 = 1,07( mg.L-1) with 95% confidence limits between 0,94 – 1,2 (mg.L-1)
- Key result
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- ca. 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- LOEC
- Effect conc.:
- > 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- ca. 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- biomass
- Remarks:
- Based on yield rate and on day by day geometric mean analysed concentrations, NOEC (72h) was 2.56 mg test item.L-1
- Key result
- Duration:
- 72 h
- Dose descriptor:
- LOEC
- Effect conc.:
- > 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- biomass
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 2.56 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- biomass
- Details on results:
- - Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): No abnormal appearance of the algae at the start and at the end of the test. - Results with reference substance (positive control):
- Potassium dichromate results:
Starting date :18 May 2018
The concentration of the reference substance dissolved in test medium that results in an 50% reduction in growth of the test organism within an exposure period of 50 h is 1,07 (mg.L-1) CEt50 with 95% confidence limits between 0,94 – 1,2 (mg.L-1) - Reported statistics and error estimates:
- The statistical evaluation (NOEC - No Observed Effect Concentration) was conducted with R
software. For 72-hour growth rate and yield, since data were consistent with a normal
distribution (Shapiro-Wilk test) and the variance homogeneity was confirmed (Bartlett test),
data were analysed for significant differences compared to the control using the parametric
ANOVA (p ≤ 0.05;) - Validity criteria fulfilled:
- yes
- Remarks:
- The growth multiplication factor in the control within the 72-hour test period was higher than 16 (actual value: 128), the coefficient of variation of the average specific growth rates in the control was less than 35% (actual value: 21.42% after 72h).
- Conclusions:
- The effects of DL-Isoleucine, N-(1-oxohexadecyl) diluted in OECD test medium at the rates of
5 mg test item.L-1 to Pseudokirchneriella subcapitata were investigated under laboratory
conditions.
The study is valid since the growth multiplication factor in the control within the 72-hour test
period was higher than 16 (actual value: 128), the coefficient of variation of the average specific
growth rates in the control was less than 7% (actual value: 1.45%) and the mean coefficient of
variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control was
less than 35% (actual value: 21.42% after 72h).
The geometric mean concentration at the tested concentration was higher than the solubility of the test item (1.5 mg.L-1 according the OECD 105).
Based on growth rate and on day by day geometric mean analysed concentrations, the
NOEC72h was 2.56 mg test item.L-1 and both the LOEC72h and the EC50-72h were > 2.56 mg test
item.L-1.
Based on yield rate and on day by day geometric mean analysed concentrations, NOEC72h
was 2.56 mg test item.L-1 and both the LOEC72h and the EC50-72h were > 2.56 mg test item.L-1. - Executive summary:
The objective of this study was to assess the effects of test item DL-Isoleucine, N-(1-
oxohexadecyl) on the growth of Pseudokirchneriella subcapitata over a period of 72 hours
according to OECD 201 Guideline. Pseudokirchneriella subcapitata is a freshwater green
algae that is recommended by the OECD Guideline 201.
Since the test item is poorly soluble under the test conditions and according to the OECD
monograph 23, the choice of a solvent was necessary to solubilize the test item, at the
concentration lower than 0.1 ml.L-1. So, a stock solution of test item was prepared in dimethyl
sulfoxide. Test solutions were prepared by dilution of this solution.
Test item quantification was performed according to a validated analytical method (final report
RRCo-000155_01) LC-MS/MS from Sciex (LC Sciex Exion AD and MS QTRAP 5500) as mass
spectrometry detection to measure DL-Isoleucine, N-(1-oxohexadecyl) concentrations at 0, 24,
48 and 72 hours.
According to the range-finding-test, no toxicity was observed under the limit of solubility. So, a
test limit at 5 mg.L-1 was performed.
The effects of DL-Isoleucine, N-(1-oxohexadecyl) diluted in OECD test medium at the rates of
5 mg test item.L-1 to Pseudokirchneriella subcapitata were investigated under laboratory
conditions.
The study is valid since the growth multiplication factor in the control within the 72-hour test
period was higher than 16 (actual value: 128), the coefficient of variation of the average specific
growth rates in the control was less than 7% (actual value: 1.45%) and the mean coefficient of
variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control was
less than 35% (actual value: 21.42% after 72h).
Based on growth rate and on day by day geometric mean analysed concentrations, the
NOEC72h was 2.56 mg test item.L-1 and both the LOEC72h and the EC50-72h were > 2.56 mg test
item.L-1.
Based on yield rate and on day by day geometric mean analysed concentrations, NOEC72h
was 2.56 mg test item.L-1 and both the LOEC72h and the EC50-72h were > 2.56 mg test item.L-1.
The geometric mean concentration at the tested concentration was higher than the solubility
of the test item (1.5 mg.L-1 according the OECD 105).
Reference
Range finding test results :
There were no significant differences between growth and yield rates in the solvant control and all test material treatement after 72h (ANOVA, p ≤ 0.05), thus, for both growth and yield rates, the NOEC was 2770.531 μg test material.L -1 and both the EC50 and the LOEC were estimated to be > 2770.531μg test material.L-1.Then a limit test at the highest rate tested for preliminary test was performed for the definitive test.
Details of growth algae results are available in the attached report.
Validity criteria:
After 72 hours:
Actual value | Reference value | Compliance | |
Growth multiplication factor in the control within the 72-hour test period | 128 | > 16 | Yes |
Coefficient of variation of the average specific growth rates in then control during the whole test period | 1.45 | < 7 | Yes |
Mean coefficient of variation for section-by-section specific growth rates (days 0 -1 , 1-2 and 2 -3) in the control | 21.42 | < 35 | Yes |
Validity criteria are met.
Quality Control :
Actual value | Reference value | Compliance | |
Recovery at 20 (%) | 101 | 70 -120 | Yes |
Recovery at 80 (%) | 100 | 70 -120 | Yes |
Within reproducibility relative standard deviation (recovery at 20 (%) | 7.4 | < 20 | Yes |
Within reproducibility relative standard deviation (recovery et 80(%) | 6.1 | < 20 | Yes |
Recoveries difference (%) between 20% and 80%) | 1 | < 20 | Yes |
The analytical series are valid since the measured values are included in the reference interval.
Analytical results:
Treatment ID | C01 | C02 | C03 | |
Target concentration [μg test item/L] | 0 (control) |
0 (solvent control) |
5000 | |
0h | <LOQ | <LOQ | 4966.8 | |
Deviation from target rate[%] | NA | NA | -0.7 | |
24h | <LOQ | <LOQ | 3814.0 | |
Analysed concntration[μg test item/L] | Deviation from target rate[%] | NA | NA | -23.7 |
48h | <LOQ | <LOQ | 1751.9 | |
Deviation from target rate [%] | NA | NA | -65.0 | |
72h | <LOQ | <LOQ | 1246.4 | |
Deviation from target rate[%] | NA | NA | -75.1 | |
Geometrical mean [μg test item/L | NA | NA | 2552.2 |
NA = not applicable. LOQ = limit of quantification = 0.5 µg active substance.L-1
Analysis of the results were based on the geometric mean concentrations after 72h. The geometric mean concentration at the tested concentration was higher than the solubility of the test item (1.5 mg.L-1 according the OECD 105).
Description of key information
The objective of this study was to assess the effects of test item DL-Isoleucine, N-(1-
oxohexadecyl) on the growth of Pseudokirchneriella subcapitata over a period of 72 hours
according to OECD 201 Guideline.Since the test item is poorly soluble under the test conditions and according to the OECD monograph 23,it was solubilized in dimethyl sulfoxide at the concentration lower than 0.1 ml.L-1
Based on growth rate and on day by day geometric mean analysed concentrations, the
NOEC72h was 2.56 mg test item.L-1 and both the LOEC72h and the EC5072h were > 2.56 mg test
item.L-1.
Based on yield rate and on day by day geometric mean analysed concentrations, NOEC72h
was 2.56 mg test item.L-1 and both the LOEC72h and the EC50-72h were > 2.56 mg test item.L-1
.
Key value for chemical safety assessment
- EC50 for freshwater algae:
- 2.56 mg/L
- EC10 or NOEC for freshwater algae:
- 2.56 mg/L
Additional information
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