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EC number: 232-589-7 | CAS number: 9001-22-3
- 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:
- 28 March 2011 - 12 March 2012
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- Reliability 1
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 012
- Report date:
- 2012
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Alga, Growth Inhibition Test)
- Deviations:
- yes
- Remarks:
- Deviations are considered not to have affected the validity and integrity of the study.
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- certificate included in report
Test material
- Reference substance name:
- Active enzyme protein of Cellulase (EC no. 232-734-4, CAS no. 9012-54-8, EC name: Cellulase, Enzyme Class no. 3.2.1.4)
- Molecular formula:
- Not applicable, see remarks.
- IUPAC Name:
- Active enzyme protein of Cellulase (EC no. 232-734-4, CAS no. 9012-54-8, EC name: Cellulase, Enzyme Class no. 3.2.1.4)
- Reference substance name:
- Protein as a constituent of enzyme deriving from the fermentation or extraction process
- Molecular formula:
- Not available. See remarks.
- IUPAC Name:
- Protein as a constituent of enzyme deriving from the fermentation or extraction process
- Reference substance name:
- Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process
- Molecular formula:
- Not available. See remarks.
- IUPAC Name:
- Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process
- Reference substance name:
- Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.
- Molecular formula:
- Not available. See remarks.
- IUPAC Name:
- Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.
- Reference substance name:
- Lipids as a constituent of enzyme deriving from the fermentation or extraction process
- Molecular formula:
- Not available. See remarks.
- IUPAC Name:
- Lipids as a constituent of enzyme deriving from the fermentation or extraction process
- Test material form:
- liquid
- Details on test material:
- - Lot/batch No.: PPC31776
- Expiration date of the lot/batch: 07 December 2020
- Stability under test conditions: Stable for at least 24 hours at room temperature.
- Storage condition of test material: Frozen
Constituent 1
Constituent 2
Constituent 3
Constituent 4
Constituent 5
Sampling and analysis
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: 0 and 100 mg TOS/L
- Sampling method: At the start of the definitive test, three samples (15 mL) were taken from the freshly-prepared control and test media. After 72 hours, the contents of the replicate flasks for each group were pooled and further samples taken for analysis. Additional samples were also taken from a flask containingCellulase at a nominal concentration of 100 mg TOS/L but with no algal cells, in order to obtain information on the extent of adsorption/absorption of the test substance by the algal cells.
- Sample storage conditions before analysis: frozen
Test solutions
- Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test substance (878 mg) was added directly to algal culture medium (1 L) to provide the test medium at a nominal concentration of 100 mg TOS/L. An aliquot (8.6 mL) of the algal inoculum was added to a portion (800 mL) of the test medium to give an initial cell density of 1 x 104 cells/mL. An aliquot (100 mL) of the appropriate inoculated test medium was added to each of the test vessels.
- Controls: Medium
Test organisms
- Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: green alga Pseudokirchneriella subcapitata
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd., Dunstaffnage Marine Laboratory, Dunbeg, Oban, Argyll, Scotland
- Method of cultivation: The liquid slope cultures were stored in an illuminated refrigerator. Sterile algal nutrient medium was inoculated with cells aseptically removed from the slope culture; these primary liquid cultures (100 mL) were incubated for approximately three days in an orbital incubator under continuous illumination at nominal temperatures in the range 21 to 25°C. Subsequently, appropriate volumes of these primary cultures were aseptically transferred to fresh sterile algal nutrient medium to prepare secondary liquid cultures; these cultures were incubated, as stated above, for a further three days to provide an inoculum in the log phase of growth, characterised by a cell density of 9.3 x 10^5 cells/mL.
Study design
- Test type:
- static
- Water media type:
- freshwater
- Limit test:
- yes
- Total exposure duration:
- 72 h
Test conditions
- Test temperature:
- 21.8-24.1°C
- pH:
- 7.54-7.96
- Nominal and measured concentrations:
- nominal: 0 and 100 mg TOS/L
At the start of the test, enzyme recovery was 104% of the nominal value. After 72 hours, the recovery decreased to 90% of nominal. - Details on test conditions:
- TEST SYSTEM
- Test vessel: conical flask
- Type (delete if not applicable): Flasks were loosely plugged with with foam bungs.
- Material, size, fill volume: glass, 250 mL, 100 mL
- Initial cells density: 1 x 10^4 cells/mL
- Control end cells density: 1372083 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6
GROWTH MEDIUM
- Standard medium used: yes (sterile algal nutrient medium as recommended in Official Journal No. L383A Part C.3 and OECD Procedure 201)
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Filtered, dechlorinated tap water which had been softened and treated by reverse osmosis, before microfiltration and purification (resistivity of 18 Megohm/cm).
OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: nominally 4440 to 8880 lux provided by 6 x 30 W "cool white" 1 metre fluorescent tubes
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Samples were taken at 24, 48 and 72 hours and the cell densities measured using a haemocytometer. The estimate of cell numbers in each sample was based on the mean of four consecutive counts. The presence of any abnormal cell numbers in each sample was also noted during screening of each test level.
TEST CONCENTRATIONS
- Range finding study
- Test concentrations:1, 10, 100 mg TOS/L
- Results used to determine the conditions for the definitive study: yes - Reference substance (positive control):
- no
Results and discussion
Effect concentrationsopen allclose all
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- >= 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: TOS (Total Organic Solids)
- Basis for effect:
- other: growth rate, biomass and yield
- Key result
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- >= 52.1 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- act. ingr.
- Remarks:
- aep (active enzyme protein)
- Basis for effect:
- other: growth rate, biomass and yield
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- >= 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: TOS
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 52.1 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- act. ingr.
- Remarks:
- aep
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: TOS
- Basis for effect:
- biomass
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 52.1 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- act. ingr.
- Remarks:
- aep
- Basis for effect:
- biomass
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: TOS
- Basis for effect:
- other: yield
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 52.1 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- act. ingr.
- Remarks:
- aep
- Basis for effect:
- other: yield
- Details on results:
- - Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Any stimulation of growth found in any treatment: at 100 mg TOS/L there was a significant stimulation at 24 to 48h, but no significant stimulation after 72h of exposure. - Reported statistics and error estimates:
- The mean coefficient of variation (CoV) for daily growth rates in control cultures ranged between 5.87 and 8.18 during the definitive test and the CoV for the average specific growth rates of the control culture was 1.46 during the 72h period.
Applicant's summary and conclusion
- Validity criteria fulfilled:
- yes
- Conclusions:
- Under the conditions of the test, Cellulase was not found to be acutely toxic to Pseudokirchneriella subcapitata at a nominal concentration of 100 mg TOS/L equivalent to 52.1 mg aep/L.
Consequently, the 72-hour EbC50, ErC50 and EyC50 values for Cellulase could not be calculated but must be >100 mg TOS/L equivalent to 52.1 mg aep/L and the “no observed effect concentration” was ≥ 100 mg TOS/L equivalent to 52.1 mg aep/L. - Executive summary:
The effect of Cellulase on the growth of the unicellular green alga Pseudokirchneriella subcapitata was assessed under non-axenic conditions.
The study was conducted in accordance with EC Methods for Determination of Ecotoxicity, Annex IV to Commission Regulation (EC) No 761/2009 (O.J. No. L220/36, 2009) Part C, Method 3 “Freshwater Algae and Cyanobacteria, Growth Inhibition Test” and Procedure 201 of the “Guidelines for Testing of Chemicals” of the Organisation for Economic Co-operation and Development: Freshwater Alga and Cyanobacteria, Growth Inhibition Test” (2006).
Six algal cultures, with an initial nominal cell density of 1 x 10^4 cells/mL, were exposed to Cellulase at a nominal concentration of 100 mg TOS (Total Organic Solids)/L or 52.1 mg active enzyme protein (aep)/L. The test medium was prepared in OECD medium by the direct addition of the test substance to the
dilution medium. The cultures were incubated in an orbital incubator under continuous illumination at temperatures ranging from 21.8 to 24.1 °C for 72 hours.
At the request of the Sponsor, the test concentration was verified by analysis of the enzyme concentration, which was performed at the Sponsor’s laboratory.
Cell numbers were counted daily to monitor growth. The test results are expressed in terms of the area under the growth curve, growth rate and yield.
The following values were derived from the data:
Nominal Cellulase concentration
(mg TOS/L)
Nominal Cellulase concentration
(mg aep/L)
Area under the growth curve
EbC50 (72 h)
>100 >52.1 Average specific growth rate
ErC50 (0 - 72 h)
>100 >52.1
Yield
EyC50 (0 - 72 h)
>100 >52.1 “No observed effect concentration”
≥ 100 ≥ 52.1
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