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EC number: 700-347-4 | CAS number: -
- 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
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Short-term toxicity to aquatic invertebrates
Administrative data
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 7 September 2009 and 5 November 2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 009
- Report date:
- 2009
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.2 (Acute Toxicity for Daphnia)
- Deviations:
- no
- Principles of method if other than guideline:
- In view of the difficulties associated with the evaluation of aquatic toxicity of poorly water soluble test materials, a modification of the standard method for the preparation of aqueous media was performed. An approach endorsed by several important regulatory authorities in the EU and elsewhere (ECETOC 1996, OECD 2000 and Singer et al 2000), is to expose organisms to a Water Accommodated Fraction (WAF) of the test material in cases where the test material is a complex mixture and is poorly soluble in water and in the permitted auxiliary solvents and surfactants. Using this approach, aqueous media are prepared by mixing the test material with water for a prolonged period. Pre-study work showed that a preparation period of 23 hours was sufficient to ensure equilibration between the test material and water phase. At the completion of mixing, the test material phase is separated by siphon and the test organisms exposed to the aqueous phase or WAF (which may contain dissolved test material and/or leachates from the test material). Exposures are expressed in terms of the original concentration of test material in water at the start of the mixing period (loading rate) irrespective of the actual concentration of test material in the WAF
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Date of GLP inspection: 19 August 2008 Date of Signature on GLP certificate:04 March 2009
Test material
- Reference substance name:
- E0286P/040A
- IUPAC Name:
- E0286P/040A
- Details on test material:
- Sponsor's identification :E0286P/040A
Description : :amber coloured viscous liquid
Batch number : :E0289P/040A/01
Date received : :20 July 2009
Storage conditions :room temperature on the dark
The integrity of supplied data relating to the identity, purity and stability of the test material is the responsibility of the Sponsor.
Constituent 1
Sampling and analysis
- Analytical monitoring:
- yes
- Details on sampling:
- Water samples were taken from the control (replicates R1 – R4 pooled) and the 100 mg/l loading rate WAF test group (replicates R1 – R2 and R3 – R4 pooled) at 0 and 48 hours for quantitative analysis.
The method of analysis, recovery and test preparation analyses are described in Appendix 3.
Test solutions
- Vehicle:
- no
- Details on test solutions:
- Procedure
Validation of mixing period
Pre-study work was carried out to determine whether stirring for a prolonged period produced significantly higher levels of total organic carbon, as an indicator of soluble organic substances in the WAF. A WAF of nominal loading rate of 100 mg/l was prepared, in duplicate, in reconstituted water. One loading rate was stirred for a period of 23 hours and the other for a period of 95 hours. After a 1-Hour standing period the mixtures were then removed by siphon and samples taken for Total Organic Carbon analysis (see Appendix 2).
Range-finding test
Due to the low aqueous solubility and complex nature of the test material for the purposes of the test the test material was prepared as a Water Accommodated Fraction (WAF).
The loading rate to be used in the definitive test was determined by a preliminary range-finding test.
In the range-finding test Daphnia magna were exposed to a series of nominal loading rates of 1.0, 10 and 100 mg/l.
Amounts of test material (20, 200 and 2000 mg) were each separately added to the surface of 20 litres of reconstituted water to give the 1.0, 10 and 100 mg/l loading rates respectively. After the addition of the test material, the reconstituted water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixtures allowed to stand for 1 hour. A wide bore glass tube, covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed through the Nescofilm seal. The aqueous phase or WAF was removed by mid-depth siphoning (the first approximate 75-100 ml discarded) to give the 1.0, 10 and 100 mg/l loading rate WAFs. Microscopic inspection of the WAFs showed no micro-dispersions or undissolved test material to be present.
In the range-finding test 10 daphnids were placed in each test and control vessel and maintained in a temperature controlled room at 21C to 22ºC with a photoperiod of 16 hours light and 8 hours darkness for a period of 48 hours with 20 minute dawn and dusk transition periods. Each 250 ml test and control vessel contained 200 ml of test media and was covered to reduce evaporation. After 24 and 48 hours the number of immobilised Daphnia magna were recorded.
The control group was maintained under identical conditions but not exposed to the test material.
At both 0 and 24 hours some temperatures were measured to be slightly in excess of the 20 ± 1°C given in the Protocol. This was considered not to have affected the outcome or validity of the test as no adverse effects of exposure were observed in the daphnids throughout the duration of the test and that the temperature was within test guideline specification.
Definitive test
Based on the results of the range-finding test a "Limit test" was conducted at a single loading rate of 100 mg/l to confirm that no immobilisation or adverse reactions to exposure were observed.
Experimental preparation
An amount of test material (2000 mg) was added to the surface of 20 litres of reconstituted water to give the 100 mg/l loading rate. After the addition of the test material, the reconstituted water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixture allowed to stand for 1 hour. A wide bore glass tube, covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed through the Nescofilm seal. The aqueous phase or WAF was removed by mid-depth siphoning (the first approximate 75-100 ml discarded) to give the 100 mg/l loading rate WAF. Microscopic inspection of the WAF showed no micro-dispersions or undissolved test material to be present.
The concentration and stability of neodecanoic acid in the test preparations were verified by chemical analysis at 0 and 48 hours (see Appendix 3).
Positive Control
A positive control (Harlan Laboratories Ltd Project No: 0039/1094) conducted approximately every six months used potassium dichromate as the reference material at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/l.
An amount of reference material (100 mg) was dissolved in reconstituted water and the volume adjusted to 1 litre to give a 100 mg/l stock solution. An aliquot (50 ml) of this stock solution was diluted in reconstituted water and the volume adjusted to 500 ml to give a 10 mg/l stock solution. Aliquots (16, 28, 50, 90 and 160 ml) of the 10 mg/l stock solution were each separately dispersed in a final volume of 500 ml of reconstituted water to give the test series of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/l respectively.
Each stock solution and prepared concentration was inverted several times to ensure adequate mixing and homogeneity.
Exposure conditions for the positive control were similar to those used in the definitive test.
Test organisms
- Test organisms (species):
- Daphnia magna
- Details on test organisms:
- Daphnia magna is a freshwater invertebrate representative of a wide variety of natural habitats, and can therefore be considered as an important non-target organism in freshwater ecosystems.
Test Species
The test was carried out using 1st instar Daphnia magna derived from in-house laboratory cultures.
Adult Daphnia were maintained in polypropylene vessels containing approximately 2 litres of reconstituted water in a temperature controlled room at approximately 20C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a suspension of algae (Chlorella sp.). Culture conditions ensured that reproduction was by parthenogenesis. Gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing. The diet and diluent water are considered not to contain any contaminant that would affect the integrity or outcome of the study.
Test Water
The reconstituted water used for both the range-finding and definitive tests was the same as that used to maintain the stock animals.
Study design
- Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 48 h
- Post exposure observation period:
- Exposure conditions
As in the range-finding test 250 ml glass jars containing approximately 200 ml of test preparation were used. At the start of the test 5 daphnids were placed in each test and control vessel at random, in the test preparations. Four replicate test and control vessels were prepared. The test vessels were then covered to reduce evaporation and maintained in a temperature controlled room at 20C to 21ºC with a photoperiod of 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods. The daphnids were not individually identified, received no food during exposure and the test vessels were not aerated.
The control group was maintained under identical conditions but not exposed to the test material.
The test preparations were not renewed during the exposure period. Any immobilisation or adverse reactions to exposure were recorded at 24 and 48 hours after the start of exposure. The criterion of effect used was that Daphnia were considered to be immobilised if they were unable to swim for approximately 15 seconds after gentle agitation.
Physico-chemical measurements
Water temperature was recorded daily throughout the test. Dissolved oxygen concentrations and pH were recorded at the start and termination of the test. The pH and dissolved oxygen concentration were measured using a WTW pH/Oxi 340I pH and dissolved oxygen meter and the temperature was measured using a Hanna Instruments HI 93510 digital thermometer.
Vortex depth measurements
The vortex depth was recorded at the start and end of the mixing period
Test conditions
- Hardness:
- EXAMPLE
The reconstituted water had an approximate theoretical total hardness of 250 mg/l as CaCO3.
EXAMPLE - Test temperature:
- Temperature was maintained at 20 deg C to 21 deg C throughout the test.
Some of the temperatures were measured to be slightly in excess of the 20 ± 1°C given in the protocol. This was considered not to affect the results of the test as no adverse effects of exposure were observed in the control daphnids throughout the duration of the test and that the temperatures were within the test guideline specification.
The temperature was measured using a Hanna Instruments HI 93510 digital thermometer. - pH:
- The reconstituted water had a pH of 7.8 ± 0.2 adjusted (if necessary) with NaOH or HCl.
The pH was measured using a WTW pH/Oxi 340I pH meter.
There were no treatment related differences for pH.
See Appendix 4 for results. - Dissolved oxygen:
- The reconstituted water was aerated until the dissolved oxygen concentration was approximately air-saturation value.
Dissolved oxygen concentrations were recorded at the start and termination of the test. The dissolved oxygen concentration was measured using a dissolved oxygen meter.
See Appendix 4 for results. - Salinity:
- Freshwater used.
- Nominal and measured concentrations:
- In the range-finding test Daphnia magna were exposed to a series of nominal loading rates of 1.0, 10 and 100 mg/l.
Cumulative immobilisation data from the exposure of Daphnia magna to the test material during the range-finding test are given in Table 1.
No immobilisation was observed at 1.0, 10 and 100 mg/l loading rate WAF.
Based on this information, a single loading rate of four replicates, of 100 mg/l, using a stirring period of 23 hours followed by a 1-Hour standing period, was selected for the definitive test. This experimental design conforms to a "Limit test" to confirm that no immobilisation or adverse reactions to exposure were observed - Details on test conditions:
- Reconstituted Water
i) Stock Solutions
a) CaCl2.2H2O 11.76 g/l
b) MgSO4.7H2O 4.93 g/l
c) NaHCO3 2.59 g/l
d) KCl 0.23 g/l
ii) Preparation
An aliquot (25 ml) of each of solutions a-d was added to each litre (final volume) of deionised water with a conductivity of <5 µS cm-1. The reconstituted water had a pH of 7.8 ± 0.2 adjusted (if necessary) with NaOH or HCl and was aerated until the dissolved oxygen concentration was approximately air-saturation value.
The reconstituted water had an approximate theoretical total hardness of 250 mg/l as CaCO3.
Validation of Mixing Period
Pre-study work was carried out to determine whether stirring for a prolonged period produced significantly higher levels of total organic carbon, as an indicator of soluble organic materials, in the WAF. A WAF of a nominal loading rate of 100 mg/l was prepared in duplicate in reconstituted water and stirred using a stirring rate such that a vortex was formed to give a slight dimple at the water surface. One loading rate was stirred for a period of 23 hours and the other for a period of 95 hours. After a 1-Hour standing period the mixtures were then removed by siphon and samples taken for Total Organic Carbon analysis.
The results are summarised as follows:
Nominal
Loading Rate
(mg/l) Time (Hours)
24 96
mg C/l mg C/l Corrected for Control mg C/l mg C/l Corrected for Control
Control100 30.98 30.98 39.95 39.95
It is evident from this work that increasing the stirring period did not significantly increase the amount of carbon in the WAF and so preparation of the WAF was maintained at 24 hours. - Reference substance (positive control):
- yes
- Remarks:
- Potassium dichromate
Results and discussion
Effect concentrationsopen allclose all
- Duration:
- 48 h
- Dose descriptor:
- EL50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: immobilisation
- Remarks on result:
- other: 95% CL of 10 - 16 mg/l.
- Duration:
- 48 h
- Dose descriptor:
- NOELR
- Remarks:
- EXAMPLE
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Remarks:
- EXAMPLE
- Basis for effect:
- other: immobilisation
- Remarks on result:
- other: 95% CL not stated
- Details on results:
- RESULTS
Validation of Mixing Period
Pre-study work (see Appendix 2) indicated that there was no significant increase in the amount of total organic carbon by extending the preparation period for longer than 24 hours. Therefore it was considered appropriate to use a 24-Hour preparation period.
Range-finding Test
Cumulative immobilisation data from the exposure of Daphnia magna to the test material during the range-finding test are given in Table 1.
No immobilisation was observed at 1.0, 10 and 100 mg/l loading rate WAF.
Based on this information, a single loading rate of four replicates, of 100 mg/l, using a stirring period of 23 hours followed by a 1-Hour standing period, was selected for the definitive test. This experimental design conforms to a "Limit test" to confirm that no immobilisation or adverse reactions to exposure were observed.
Definitive Test
Immobilisation data
Cumulative immobilisation data from the exposure of Daphnia magna to the test material during the definitive test are given in Table 2.
There was no significant immobilisation in 20 daphnids exposed to a 100 mg/l loading rate WAF for a period of 48 hours. Inspection of the immobilisation data gave the following results:
Time (h) EL*50 (mg/l Loading Rate WAF)
24 >100
48 >100
The No Observed Effect Loading rate after 24 and 48 hours exposure was 100 mg/l loading rate WAF. The No Observed Effect Loading rate is based upon no significant immobilisation at this loading rate.
It was considered unnecessary and unrealistic to test at loading rates in excess of 100 mg/l.
Physico-chemical measurements
The results of the physico-chemical measurements are given in Appendix 4. Temperature was maintained at 20C to 21ºC throughout the test, while there were no treatment related differences for oxygen concentration or pH.
Vortex depth measurements
The vortex depth was recorded at the start and end of the mixing period and was observed to be a dimple at the water surface on each occasion (see Table 3).
Observations on test material solubility
Observations on the test media were carried out during the mixing and testing of the WAF.
At the start of the mixing period the 100 mg/l loading rate was observed to have formed a clear colourless water column with test material floating at the media surface. After 23 hours stirring and a 1-Hour standing period the 100 mg/l loading rate was observed to have formed a clear colourless water column with a thin layer of test material across the water surface. Microscopic inspection of the WAF showed no micro-dispersions or undissolved test material to be present.
After siphoning and for the duration of the test, the 100 mg/l loading rate was observed to be a clear colourless solution.
Chemical analysis of test loading rates
The test material is a mixture of titanium salts (80 %) and alkyl acids (20%). Method development work conducted indicated that at the test concentrations to be employed in the definitive test analysis of the test samples for titanium by GC-MS was unreliable possibly due to interference from other components of the test material. Analysis by HPLC-MS did however give a good response at m/z 171 which corresponds to the neodecanoic acid present in the test material. At the request of the Sponsor analysis of the test samples for the presence of neodecanoic acid was to be performed to provide an indication of the presence of dissolved hydrocarbon constituents.
Analysis of the 100 mg/l loading rate WAF at 0 and 48 hours (se Appendix 3) showed measured test concentrations of neodecanoic acid to range from 30 to 32 mg/l.
Given that the toxicity of a complex UVCB chemical that has limited water solubility cannot be attributed to a single, or combination of, constituents the results are expressed based on nominal loading rates for the whole material. - Results with reference substance (positive control):
- Positive Control
Cumulative immobilisation data from the exposure of Daphnia magna to the reference material (Harlan Laboratories Ltd Project No: 0039/1094) during the positive control are given in Table 4. The relationship between percentage immobilisation and concentration at 24 and 48 hours is given in Figures 1 and 2.
Inspection of the immobilisation data at 3 hours and analysis of the immobilisation data by the probit method (Finney 1971) at 24 and 48 hours based on the nominal test concentrations gave the following results:
Time (h)
EC50 (mg/l) 95% Confidence limits
(mg/l)
3 > 3.2 -
24 1.0 0.90 - 1.2
48 0.78 0.68 - 0.88
The No Observed Effect Concentrations after 24 and 48 hours were 0.56 and 0.32 mg/l respectively. The No Observed Effect Concentration is based upon zero immobilisation at this concentration.
The slopes and their standard errors of the response curves at 24 and 48 hours were 7.8 (SE = 1.7) and 12 (SE = 2.4) respectively.
The results from the positive control with potassium dichromate were within the normal range for this reference material. The mean 48-Hour EC50 value calculated from all positive controls was 0.78 mg/l (sd = 0.20). - Reported statistics and error estimates:
- An estimate of the EC50 value at 3 hours was given by inspection of the immobilisation data.
The EC50 values and associated confidence limits at 24 and 48 hours and the slope of the response curves and standard errors were calculated by the maximum-likelihood probit method (Finney 1971) using the ToxCalc computer software package (Toxcalc 1999).
Probit analysis is used when two or more partial responses to exposure are shown
Any other information on results incl. tables
Applicant's summary and conclusion
- Validity criteria fulfilled:
- yes
- Conclusions:
- CONCLUSION
The acute toxicity of the test material to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-Hour EL*50 value of greater than 100 mg/l loading rate WAF. Correspondingly the No Observed Effect Loading rate was 100 mg/l loading rate WAF. - Executive summary:
Introduction.A study was performed to assess the acute toxicity of the test material toDaphnia magna. The method followed that described in the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphniasp, Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.
Methods. Information provided by the Sponsor indicated that the test material was a mixture of titanium salts and alkyl acids. Given this and also the poor solubility of the test material in water the most appropriate method of preparation for this material was as a Water Accommodated Fraction (WAF).
Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to a Water Accommodated Fraction (WAF) of the test material, at a single nominal loading rate of 100 mg/l for 48 hours at a temperature of approximately 21°C under static test conditions. Immobilisation and any adverse reactions to exposure were recorded after 24 and 48 hours.
A positive control conducted approximately every six months used potassium dichromate as the reference material. Daphnia magnawas exposed to an aqueous solution of the reference material at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/l for 48 hours at a temperature of 21°C to 22°C under static test conditions. Immobilisation and any adverse reactions to exposure were recorded after 3, 24 and 48 hours.
Results. The 48-Hour EL*50for the test material toDaphnia magnabased on nominal loading rates was greater than 100 mg/l loading rate WAF and correspondingly the No Observed Effect Loading rate was 100 mg/l loading rate WAF.
It was considered unnecessary and unrealistic to test at loading rates in excess of 100 mg/l.
The test material is a mixture of titanium salts (80 %) and alkyl acids (20%). Method development work conducted indicated that at the test concentrations to be employed in the definitive test analysis of the test samples for titanium by GC-MS was unreliable possibly due to interference from other components of the test material. Analysis by HPLC-MS did however give a good response at m/z 171 which corresponds to the neodecanoic acid present in the test material. At the request of the Sponsor analysis of the test samples for the presence of neodecanoic acid was to be performedto provide an indication of the dissolved hydrocarbon content.
Analysis of the 100 mg/l loading rate WAF at 0 and 48 hours showed measured test concentrations of neodecanoic acid to range from 30 to 32 mg/l.
Given that the toxicity of a complex UVCB chemical that has limited water solubility cannot be attributed to a single, or combination of, constituents the results are expressed based on nominal loading rates for the whole material.
The 48-Hour EC50for the reference material toDaphnia magnabased on nominal concentrations was 0.78 mg/l with 95% confidence limits of 0.68 – 0.88 mg/l. The No Observed Effect Concentration was 0.32 mg/l.
*EL = Effective Loading Rate
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