Registration Dossier
Registration Dossier
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EC number: 268-919-1 | CAS number: 68154-72-3
Short-term toxicity to aquatic invertebrates
Administrative data
Link to relevant study record(s)
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: This study is carried out in accordance to OECD guideline and is GLP compliant.
- Justification for type of information:
- REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'. - Reason / purpose for cross-reference:
- read-across: supporting information
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
- Deviations:
- yes
- Remarks:
- In the range-finding test the Total Organic Carbon concentration was determined with the TOC instead of NPOC method. As this is an equivalent alternative it was not considered to affect the outcome of the test.
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.2 (Acute Toxicity for Daphnia)
- Deviations:
- yes
- Remarks:
- In the range-finding test the Total Organic Carbon concentration was determined with the TOC instead of NPOC method. As this is an equivalent alternative it was not considered to affect the outcome of the test.
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- Details on properties of test surrogate or analogue material (migrated information):
Not applicable - Analytical monitoring:
- yes
- Details on sampling:
- The confirmatory chemical analysis was based on the concentration of the fatty acid component (sebacate) and on the concentration of the lithium ion. The concentration of total organic carbon (TOC) was also measured.
Samples for analysis were taken from the control and the 100 mg/L treatment at the start of the test and after 48 hours of exposure and stored in a refrigerator (TOC) or freezer (lithium and sebacate) until analysis.
For the determination of the test substance based on sebacate, the samples were diluted in a 1:1 (v:v) ratio with 2% formic acid in acetonitrile and analysed. If necessary, the samples were further diluted with 1% formic acid in 50/50 (v/v) acetonitrile/ISO-medium to obtain concentrations within the calibration range.
For the determination of the test substance based on lithium, the samples were diluted in a 24:1 (v:v) ratio with HNO3 and analysed. If necessary, the samples were further diluted with 4% HNO3 in ISO-medium to obtain concentrations within the calibration range. - Vehicle:
- no
- Details on test solutions:
- - Solution: Adjusted ISO medium
- Solubility: The test item was completely soluble in test medium at the concentrations tested.
- Preparation of test solutions: A solution with a concentration of 100 mg/L was prepared using magnetic stirring for 34 minutes to 2.5 hours to completely dissolve test substance in test medium. The lower test concentrations were prepared by subsequent dilutions of the highest concentration in the test medium.
- Appearance: The solutions were clear and colourless at test start. - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- Source: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions from an in-house laboratory culture with a known history.
Batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress.
Age: Young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old. - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 48 h
- Post exposure observation period:
- None
- Hardness:
- Not reported. Breeding water: 180 mg CaCO3/L
- Test temperature:
- 20-21°C
- pH:
- pH 7.7-8.1
- Dissolved oxygen:
- 8.8-9.2 mg/L
- Salinity:
- Not applicable
- Nominal and measured concentrations:
- Nominal concentrations: 10, 18, 32, 56 and 100 mg/L.
The measured concentrations based on sebacate were at the level of 94-97% of nominal during the exposure period.
The measured concentrations based on lithium were at the level of 97-101% of nominal during the exposure period.
The control (0 mg/L) and highest (100 mg/L) were measured at 0 hours to be 0.050 and 55 mg TOC/L and at 48 hours they were not detected and 54 mg TOC/L, respectively. The TOC remained relatively stable throughout the test. - Details on test conditions:
- TEST SYSTEM
- Test vessel: 100 mL
- Material, fill volume: Glass, 80 mL
- Aeration: No aeration of test solutions
- No. of organisms per vessel: 5
- No. of daphnids per concentration: 20
-Introduction of daphnids: Within 32 minutes of preparation of the test solutions
OTHER TEST CONDITIONS
- Photoperiod: 16 hours
- Feeding: No feeding - Reference substance (positive control):
- yes
- Remarks:
- potassium dichromate (K2Cr2O7)
- Duration:
- 48 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- mobility
- Details on results:
- The 48h-EC50 exceeded the nominal concentration of 100 mg/L.
The first combined limit/range finding test showed no immobility in the control and concentrations up to and including 10 mg/L during exposure. In the highest concentration (100 mg/L), all daphnids were immobilised at the end of the test. It should be noted that a number of daphnids were observed to be trapped at the surface in the control treatment at 24 hours. Despite this, it was decided to continue testing and use the results as an indicator for the final test. A slight layer of floating material was observed in the highest concentration at 24 hours. Based on these results, the expected EC50 was between 10 and 100 mg/L.
In contrast to the combined limit/range finding study, the definitive test showed no immobilisation in any of the groups tested. In the first study, as daphnids were observed to be trapped at the surface of the control solution, the results of the test were only considered to be indicative and not definitive. Surface trapping was not observed in the final test and as the measured test concentrations were also confirmed to be equivalent to the anticipated nominal concentrations during the final test, the results obtained from the definitive test were considered to be the most reliable. - Results with reference substance (positive control):
- The actual responses in this reference test with K2Cr2O7 are within the ranges of the expected responses at the different concentrations, i.e. the 48h-EC50 was between 0.3 and 1.0 mg/L. Hence, the sensitivity of the Daphnia was within the range determined with the historical data collected at WIL Research Europe.
- Validity criteria fulfilled:
- yes
- Remarks:
- test conditions were constant, measured concentrations at least 80% of nominal, no daphnids in the control were immobilised, oxygen concentration was ≥3 mg/L
- Conclusions:
- No EC50 could be calculated because the test substance did not cause a significant difference to the mobility of the Daphnia magna (EC50 > 100 mg/L, the maximum concentration tested.
- Executive summary:
The effect of dilithium sebacate on the mobility of the Daphnia Magna was investigated according to an OECD 202 guideline and EC method C2. A 48 hour test at 10, 18, 32, 56 and 100 mg/L was conducted under static conditions. At the start of the test all solutions were clear and colourless. Analysis of the fatty acid (sebacate) was between 94 and 97% nominal and the lithium concentration was between 97 and 101% of nominal throughout the test. The total organic carbon (TOC) measured at the start of the test was 55 mg/L and at the end of the test was 54 mg/L for the highest test concentration. The test concentrations were considered to be maintained and the results are reported based on nominal concentrations. The study met all the validity criteria for the test. As no effects on the mobility of Daphnia Magna were observed in the test the EC50 is > 100 mg/L dilthium sebacate.
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: This study is carried out in accordance to OECD guideline and is GLP compliant.
- Justification for type of information:
- REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'. - Reason / purpose for cross-reference:
- read-across source
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
- Deviations:
- yes
- Remarks:
- In the range-finding test the Total Organic Carbon concentration was determined with the TOC instead of NPOC method. As this is an equivalent alternative it was not considered to affect the outcome of the test.
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.2 (Acute Toxicity for Daphnia)
- Deviations:
- yes
- Remarks:
- In the range-finding test the Total Organic Carbon concentration was determined with the TOC instead of NPOC method. As this is an equivalent alternative it was not considered to affect the outcome of the test.
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- Details on properties of test surrogate or analogue material (migrated information):
Not applicable - Analytical monitoring:
- yes
- Details on sampling:
- The confirmatory chemical analysis was based on the concentration of the fatty acid component (sebacate) and on the concentration of the lithium ion. The concentration of total organic carbon (TOC) was also measured.
Samples for analysis were taken from the control and the 100 mg/L treatment at the start of the test and after 48 hours of exposure and stored in a refrigerator (TOC) or freezer (lithium and sebacate) until analysis.
For the determination of the test substance based on sebacate, the samples were diluted in a 1:1 (v:v) ratio with 2% formic acid in acetonitrile and analysed. If necessary, the samples were further diluted with 1% formic acid in 50/50 (v/v) acetonitrile/ISO-medium to obtain concentrations within the calibration range.
For the determination of the test substance based on lithium, the samples were diluted in a 24:1 (v:v) ratio with HNO3 and analysed. If necessary, the samples were further diluted with 4% HNO3 in ISO-medium to obtain concentrations within the calibration range. - Vehicle:
- no
- Details on test solutions:
- - Solution: Adjusted ISO medium
- Solubility: The test item was completely soluble in test medium at the concentrations tested.
- Preparation of test solutions: A solution with a concentration of 100 mg/L was prepared using magnetic stirring for 34 minutes to 2.5 hours to completely dissolve test substance in test medium. The lower test concentrations were prepared by subsequent dilutions of the highest concentration in the test medium.
- Appearance: The solutions were clear and colourless at test start. - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- Source: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions from an in-house laboratory culture with a known history.
Batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress.
Age: Young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old. - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 48 h
- Post exposure observation period:
- None
- Hardness:
- Not reported. Breeding water: 180 mg CaCO3/L
- Test temperature:
- 20-21°C
- pH:
- pH 7.7-8.1
- Dissolved oxygen:
- 8.8-9.2 mg/L
- Salinity:
- Not applicable
- Nominal and measured concentrations:
- Nominal concentrations: 10, 18, 32, 56 and 100 mg/L.
The measured concentrations based on sebacate were at the level of 94-97% of nominal during the exposure period.
The measured concentrations based on lithium were at the level of 97-101% of nominal during the exposure period.
The control (0 mg/L) and highest (100 mg/L) were measured at 0 hours to be 0.050 and 55 mg TOC/L and at 48 hours they were not detected and 54 mg TOC/L, respectively. The TOC remained relatively stable throughout the test. - Details on test conditions:
- TEST SYSTEM
- Test vessel: 100 mL
- Material, fill volume: Glass, 80 mL
- Aeration: No aeration of test solutions
- No. of organisms per vessel: 5
- No. of daphnids per concentration: 20
-Introduction of daphnids: Within 32 minutes of preparation of the test solutions
OTHER TEST CONDITIONS
- Photoperiod: 16 hours
- Feeding: No feeding - Reference substance (positive control):
- yes
- Remarks:
- potassium dichromate (K2Cr2O7)
- Duration:
- 48 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- mobility
- Details on results:
- The 48h-EC50 exceeded the nominal concentration of 100 mg/L.
The first combined limit/range finding test showed no immobility in the control and concentrations up to and including 10 mg/L during exposure. In the highest concentration (100 mg/L), all daphnids were immobilised at the end of the test. It should be noted that a number of daphnids were observed to be trapped at the surface in the control treatment at 24 hours. Despite this, it was decided to continue testing and use the results as an indicator for the final test. A slight layer of floating material was observed in the highest concentration at 24 hours. Based on these results, the expected EC50 was between 10 and 100 mg/L.
In contrast to the combined limit/range finding study, the definitive test showed no immobilisation in any of the groups tested. In the first study, as daphnids were observed to be trapped at the surface of the control solution, the results of the test were only considered to be indicative and not definitive. Surface trapping was not observed in the final test and as the measured test concentrations were also confirmed to be equivalent to the anticipated nominal concentrations during the final test, the results obtained from the definitive test were considered to be the most reliable. - Results with reference substance (positive control):
- The actual responses in this reference test with K2Cr2O7 are within the ranges of the expected responses at the different concentrations, i.e. the 48h-EC50 was between 0.3 and 1.0 mg/L. Hence, the sensitivity of the Daphnia was within the range determined with the historical data collected at WIL Research Europe.
- Validity criteria fulfilled:
- yes
- Remarks:
- test conditions were constant, measured concentrations at least 80% of nominal, no daphnids in the control were immobilised, oxygen concentration was ≥3 mg/L
- Conclusions:
- No EC50 could be calculated because the test substance did not cause a significant difference to the mobility of the Daphnia magna (EC50 > 100 mg/L, the maximum concentration tested.
- Executive summary:
The effect of dilithium sebacate on the mobility of the Daphnia Magna was investigated according to an OECD 202 guideline and EC method C2. A 48 hour test at 10, 18, 32, 56 and 100 mg/L was conducted under static conditions. At the start of the test all solutions were clear and colourless. Analysis of the fatty acid (sebacate) was between 94 and 97% nominal and the lithium concentration was between 97 and 101% of nominal throughout the test. The total organic carbon (TOC) measured at the start of the test was 55 mg/L and at the end of the test was 54 mg/L for the highest test concentration. The test concentrations were considered to be maintained and the results are reported based on nominal concentrations. The study met all the validity criteria for the test. As no effects on the mobility of Daphnia Magna were observed in the test the EC50 is > 100 mg/L dilthium sebacate.
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- Between 16 March 2010 and 26 March 2010.
- 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 relevant results.
- Justification for type of information:
- REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'. - Reason / purpose for cross-reference:
- read-across: supporting information
- 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
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Date of inspection: 15th September 2009. Date of signature: 26th November 2009
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: Loading rate of 100 mg/l in definitive test
- Sampling method: Analysis of the WAF was carried out by Total Organic Carbon (TOC) analysis. 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.
- Sample storage conditions before analysis: Duplicate samples were taken and stored at approximately -20°C for further analysis if necessary. - Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
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 deionised reverse osmosis 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.
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.
Range-finding test
Amounts of test item (5.0, 50 and 500 mg) were separately added to the surface of 5 litres of reconstituted water to give the 1.0, 10 and 100 mg/l loading rates respectively. After the addition of the test item, 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. Microscopic observations made on the WAFs indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAFs by filtering through a glass wool plug (2-4 cm in length). 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. A glass wool plug was inserted into the opposite end of the tubing and the WAF 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 observations of the WAFs were performed after filtering and showed no undissolved test item to be present.
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
Due to the low aqueous solubility and complex nature of the test item for the purposes of the definitive test the test item was prepared as a Water Accommodated Fraction (WAF).
An amount of test item (500 mg) was added to the surface of 5 litres of reconstituted water to give the 100 mg/l loading rate. After the addition of the test item, 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. Microscopic observations made on the WAF indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAF by filtering through a glass wool plug (2-4 cm in length). 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. A glass wool plug was inserted into the opposite end of the tubing and the WAF removed by mid-depth siphoning (the first approximate 75-100 ml discarded) to give the 100 mg/l loading rate WAF. Microscopic observations of the WAF were performed after filtering and showed no micro-dispersions to be present.
Total Organic Carbon (TOC) analysis was performed on the test preparations at 0 and 48 hours - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- TEST ORGANISM
- Common name: Water flea
- Strain: not applicable
- Source: In-house laboratory culture
- Age at study initiation (mean and range, SD): 1st instar
- Weight at study initiation (mean and range, SD): not determined
- Length at study initiation (length definition, mean, range and SD): not determined
- Valve height at study initiation, for shell deposition study (mean and range, SD): not applicable
- Peripheral shell growth removed prior to test initiation: not applicable
- Method of breeding: Parthenogenesis
- Feeding during test: not fed during exposure period
ACCLIMATION
- Not applicable
QUARANTINE (wild caught)
- Not applicable - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- yes
- Total exposure duration:
- 48 h
- Post exposure observation period:
- Not applicable
- Hardness:
- The reconstituted water had an approximate theoretical total hardness of 250 mg/l as CaCO3.
- Test temperature:
- Recorded daily throughout the test using a Hanna Instruments HI 93510 digital thermometer.
Temperature maintained at 21°C to 22°C throughout the test.
Some of the temperatures were observed to be slightly in excess of the range given in the study plan of 20 ± 1°C. This deviation was considered not to have affected the outcome or the validity of the test as no adverse effects of exposure were observed throughout the duration of the test and that the temperature range was within guideline specification. - pH:
- Recorded at start and termination of the test using a WTW pH/Oxi 340I pH and dissolved oxygen meter.
Range: 7.5 to 8.1 - Dissolved oxygen:
- Recorded at start and termination of the test using a WTW pH/Oxi 340I pH and dissolved oxygen meter.
The oxygen concentration in some of the test vessels at was observed to have an air saturation value (ASV) in excess of 100%. This was considered to be due to the presence of microscopic air bubbles in the media super-saturating the diluent and was considered not to have had an impact on the outcome or integrity of the test as no adverse effects were observed. - Salinity:
- Not applicable, as freshwater study.
- Nominal and measured concentrations:
- Range-finder: 1.0, 10 and 100 mg/l
Definitive test: Nominal loading rate of 100 mg/l - Details on test conditions:
- TEST SYSTEM
- Test vessel:250 ml glass jars
- Type: closed
- Material, size, headspace, fill volume: 250 ml glass jars containing approximately 250 ml of test preparation
- Aeration: not aerated
- Type of flow-through (e.g. peristaltic or proportional diluter): not applicable
- Renewal rate of test solution (frequency/flow rate): The test preparations were not renewed during the exposure period
- No. of organisms per vessel: 5 Daphnids
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): not applicable
- Biomass loading rate: not stated in report
TEST MEDIUM / WATER PARAMETERS
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
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.
OTHER TEST CONDITIONS
- Adjustment of pH: not required
- Photoperiod: photoperiod of 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods.
- Light intensity: not stated in report
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : 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.
TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable, as definitive test was a limit test
- Justification for using less concentrations than requested by guideline: 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.
- Range finding study
- Test concentrations: 1.0, 10 and 100 mg/l loading rate
- Results used to determine the conditions for the definitive study: A single immobilised daphnid was observed at a test concentration of 1.0 mg/l after 24 hours exposure. This was considered to be due to natural causes rather than a toxic effect given that no other immobilisation was observed.
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. - Reference substance (positive control):
- yes
- Remarks:
- Potassium dichromate
- Duration:
- 48 h
- Dose descriptor:
- EL50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: nominal loading rate WAF
- Basis for effect:
- other: immbolisation
- Duration:
- 48 h
- Dose descriptor:
- NOELR
- Effect conc.:
- 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: nominal loading rate WAF
- Basis for effect:
- other: immobilisation
- Details on results:
- - Behavioural abnormalities: None observed
- Observations on body length and weight: Not observed
- Other biological observations: Immobilisation only observed.
- Mortality of control: immobilisation only observed. Mortality not assessed.
- Other adverse effects control: none stated in report
- Abnormal responses: none stated in report
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not applicable as WAFs used, so measured values not available
- Effect concentrations exceeding solubility of substance in test medium: No - Results with reference substance (positive control):
- - Results with reference substance valid? Yes.
- Mortality: Not applicable
- EC50/LC50: The No Observed Effect Concentrations after 24 and 48 hours was 0.32 mg/l. The No Observed Effect Concentration is based upon zero immobilisation at this concentration.
The slope and its standard error of the response curve at 24 hours was 7.7 (SE = 1.6). Due to the unsuitable nature of the data it was not possible to calculate the slope and error of response curve at 48 hours.
The results from the positive control with potassium dichromate were within the normal range for this reference item. The mean 48-Hour EC50 value calculated from all positive controls was 0.77 mg/l (sd = 0.20). - Reported statistics and error estimates:
- Analysis of the mortality data was by the probit method (Finney 1971) at 24 hours and the trimmed Spearman-Karber method (Hamilton et al 1977) at 48 hours using the ToxCalc computer software package (ToxCalc 1999).
Probit analysis is used when two or more partial responses to exposure are shown. - Validity criteria fulfilled:
- yes
- Conclusions:
- The 48-Hour EL*50 for the test item to Daphnia magna based 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.
- Executive summary:
Introduction.A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method followed that described in the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphnia sp, Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.
Methods. Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to a Water Accommodated Fraction (WAF) of the test item, at a single nominal loading rate of 100 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 24 and 48 hours.
A positive control conducted approximately every six months used potassium dichromate as the reference item. Daphnia magnawas exposed to an aqueous solution of the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/l for 48 hours at a temperature of approximately 20°C under static test conditions. Immobilisation and any adverse reactions to exposure were recorded after 24 and 48 hours.
Results. The 48-Hour EL*50for the test item to Daphnia magna based 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.
Samples of the control and 100 mg/l loading rate WAFs were taken at 0 (fresh media) and 48 hours (old media) for Total Organic Carbon (TOC) analysis. Given the background level of carbon in the control vessels and also the low level of carbon in the test vessels, it was considered that the results gave no evidence of the presence of test item in the WAF.
Therefore, given that the toxicity cannot be attributed to a single component or a mixture of components but to the test item as a whole, and the dissolved test item was below the quantifiable limit of the analytical method, the results were based on nominal loading rates only.
The 48-Hour EC50for the reference item (potassium dichromate) to Daphnia magna based on nominal concentrations was 0.65 mg/l with 95% confidence limits of 0.58 – 0.72 mg/l. The No Observed Effect Concentration was 0.32 mg/l.
*EL = Effective Loading rate
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Study period:
- Between 16 March 2010 and 26 March 2010.
- 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 relevant results.
- Justification for type of information:
- REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'. - Reason / purpose for cross-reference:
- read-across source
- 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
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Date of inspection: 15th September 2009. Date of signature: 26th November 2009
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: Loading rate of 100 mg/l in definitive test
- Sampling method: Analysis of the WAF was carried out by Total Organic Carbon (TOC) analysis. 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.
- Sample storage conditions before analysis: Duplicate samples were taken and stored at approximately -20°C for further analysis if necessary. - Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
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 deionised reverse osmosis 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.
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.
Range-finding test
Amounts of test item (5.0, 50 and 500 mg) were separately added to the surface of 5 litres of reconstituted water to give the 1.0, 10 and 100 mg/l loading rates respectively. After the addition of the test item, 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. Microscopic observations made on the WAFs indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAFs by filtering through a glass wool plug (2-4 cm in length). 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. A glass wool plug was inserted into the opposite end of the tubing and the WAF 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 observations of the WAFs were performed after filtering and showed no undissolved test item to be present.
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
Due to the low aqueous solubility and complex nature of the test item for the purposes of the definitive test the test item was prepared as a Water Accommodated Fraction (WAF).
An amount of test item (500 mg) was added to the surface of 5 litres of reconstituted water to give the 100 mg/l loading rate. After the addition of the test item, 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. Microscopic observations made on the WAF indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAF by filtering through a glass wool plug (2-4 cm in length). 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. A glass wool plug was inserted into the opposite end of the tubing and the WAF removed by mid-depth siphoning (the first approximate 75-100 ml discarded) to give the 100 mg/l loading rate WAF. Microscopic observations of the WAF were performed after filtering and showed no micro-dispersions to be present.
Total Organic Carbon (TOC) analysis was performed on the test preparations at 0 and 48 hours - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- TEST ORGANISM
- Common name: Water flea
- Strain: not applicable
- Source: In-house laboratory culture
- Age at study initiation (mean and range, SD): 1st instar
- Weight at study initiation (mean and range, SD): not determined
- Length at study initiation (length definition, mean, range and SD): not determined
- Valve height at study initiation, for shell deposition study (mean and range, SD): not applicable
- Peripheral shell growth removed prior to test initiation: not applicable
- Method of breeding: Parthenogenesis
- Feeding during test: not fed during exposure period
ACCLIMATION
- Not applicable
QUARANTINE (wild caught)
- Not applicable - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- yes
- Total exposure duration:
- 48 h
- Post exposure observation period:
- Not applicable
- Hardness:
- The reconstituted water had an approximate theoretical total hardness of 250 mg/l as CaCO3.
- Test temperature:
- Recorded daily throughout the test using a Hanna Instruments HI 93510 digital thermometer.
Temperature maintained at 21°C to 22°C throughout the test.
Some of the temperatures were observed to be slightly in excess of the range given in the study plan of 20 ± 1°C. This deviation was considered not to have affected the outcome or the validity of the test as no adverse effects of exposure were observed throughout the duration of the test and that the temperature range was within guideline specification. - pH:
- Recorded at start and termination of the test using a WTW pH/Oxi 340I pH and dissolved oxygen meter.
Range: 7.5 to 8.1 - Dissolved oxygen:
- Recorded at start and termination of the test using a WTW pH/Oxi 340I pH and dissolved oxygen meter.
The oxygen concentration in some of the test vessels at was observed to have an air saturation value (ASV) in excess of 100%. This was considered to be due to the presence of microscopic air bubbles in the media super-saturating the diluent and was considered not to have had an impact on the outcome or integrity of the test as no adverse effects were observed. - Salinity:
- Not applicable, as freshwater study.
- Nominal and measured concentrations:
- Range-finder: 1.0, 10 and 100 mg/l
Definitive test: Nominal loading rate of 100 mg/l - Details on test conditions:
- TEST SYSTEM
- Test vessel:250 ml glass jars
- Type: closed
- Material, size, headspace, fill volume: 250 ml glass jars containing approximately 250 ml of test preparation
- Aeration: not aerated
- Type of flow-through (e.g. peristaltic or proportional diluter): not applicable
- Renewal rate of test solution (frequency/flow rate): The test preparations were not renewed during the exposure period
- No. of organisms per vessel: 5 Daphnids
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): not applicable
- Biomass loading rate: not stated in report
TEST MEDIUM / WATER PARAMETERS
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
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.
OTHER TEST CONDITIONS
- Adjustment of pH: not required
- Photoperiod: photoperiod of 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods.
- Light intensity: not stated in report
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : 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.
TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable, as definitive test was a limit test
- Justification for using less concentrations than requested by guideline: 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.
- Range finding study
- Test concentrations: 1.0, 10 and 100 mg/l loading rate
- Results used to determine the conditions for the definitive study: A single immobilised daphnid was observed at a test concentration of 1.0 mg/l after 24 hours exposure. This was considered to be due to natural causes rather than a toxic effect given that no other immobilisation was observed.
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. - Reference substance (positive control):
- yes
- Remarks:
- Potassium dichromate
- Duration:
- 48 h
- Dose descriptor:
- EL50
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: nominal loading rate WAF
- Basis for effect:
- other: immbolisation
- Duration:
- 48 h
- Dose descriptor:
- NOELR
- Effect conc.:
- 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- other: nominal loading rate WAF
- Basis for effect:
- other: immobilisation
- Details on results:
- - Behavioural abnormalities: None observed
- Observations on body length and weight: Not observed
- Other biological observations: Immobilisation only observed.
- Mortality of control: immobilisation only observed. Mortality not assessed.
- Other adverse effects control: none stated in report
- Abnormal responses: none stated in report
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not applicable as WAFs used, so measured values not available
- Effect concentrations exceeding solubility of substance in test medium: No - Results with reference substance (positive control):
- - Results with reference substance valid? Yes.
- Mortality: Not applicable
- EC50/LC50: The No Observed Effect Concentrations after 24 and 48 hours was 0.32 mg/l. The No Observed Effect Concentration is based upon zero immobilisation at this concentration.
The slope and its standard error of the response curve at 24 hours was 7.7 (SE = 1.6). Due to the unsuitable nature of the data it was not possible to calculate the slope and error of response curve at 48 hours.
The results from the positive control with potassium dichromate were within the normal range for this reference item. The mean 48-Hour EC50 value calculated from all positive controls was 0.77 mg/l (sd = 0.20). - Reported statistics and error estimates:
- Analysis of the mortality data was by the probit method (Finney 1971) at 24 hours and the trimmed Spearman-Karber method (Hamilton et al 1977) at 48 hours using the ToxCalc computer software package (ToxCalc 1999).
Probit analysis is used when two or more partial responses to exposure are shown. - Validity criteria fulfilled:
- yes
- Conclusions:
- The 48-Hour EL*50 for the test item to Daphnia magna based 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.
- Executive summary:
Introduction.A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method followed that described in the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphnia sp, Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.
Methods. Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to a Water Accommodated Fraction (WAF) of the test item, at a single nominal loading rate of 100 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 24 and 48 hours.
A positive control conducted approximately every six months used potassium dichromate as the reference item. Daphnia magnawas exposed to an aqueous solution of the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/l for 48 hours at a temperature of approximately 20°C under static test conditions. Immobilisation and any adverse reactions to exposure were recorded after 24 and 48 hours.
Results. The 48-Hour EL*50for the test item to Daphnia magna based 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.
Samples of the control and 100 mg/l loading rate WAFs were taken at 0 (fresh media) and 48 hours (old media) for Total Organic Carbon (TOC) analysis. Given the background level of carbon in the control vessels and also the low level of carbon in the test vessels, it was considered that the results gave no evidence of the presence of test item in the WAF.
Therefore, given that the toxicity cannot be attributed to a single component or a mixture of components but to the test item as a whole, and the dissolved test item was below the quantifiable limit of the analytical method, the results were based on nominal loading rates only.
The 48-Hour EC50for the reference item (potassium dichromate) to Daphnia magna based on nominal concentrations was 0.65 mg/l with 95% confidence limits of 0.58 – 0.72 mg/l. The No Observed Effect Concentration was 0.32 mg/l.
*EL = Effective Loading rate
Referenceopen allclose all
Time (h) |
Replicate |
Nominal concentration Dilithium sebacate (mg/L) |
||||
Control |
0.10 |
1.0 |
10 |
100 |
||
0 |
A |
5 |
5 |
5 |
5 |
5 |
B |
5 |
5 |
5 |
5 |
5 |
|
C |
4 |
5 |
5 |
5 |
5 |
|
D |
5 |
5 |
5 |
5 |
5 |
|
Total introduced |
20 |
20 |
20 |
20 |
20 |
|
24 |
A |
0 (1) |
0 |
0 |
0 |
0 |
B |
0 |
0 |
0 |
0 |
0 |
|
C |
0 |
0 |
0 |
0 |
0 |
|
D |
0 |
0(1) |
0 |
0 |
0 |
|
Total immobilised |
0 |
0 |
0 |
0 |
0 |
|
Effect % |
0 |
0 |
0 |
0 |
0 |
|
|
|
|
|
|
||
48 |
A |
0 |
0 |
0 |
0 |
0 |
B |
0 |
0 |
0 |
0 |
0 |
|
C |
0 |
0 |
0 |
0 |
0 |
|
D |
0 |
0 |
0 |
0 |
1 |
|
Total immobilised |
1 |
0 |
0 |
0 |
1 |
|
Effect % |
5 |
0 |
0 |
0 |
5 |
|
Parameter |
Nominal concentration Dilithium sebacate (mg/L) |
24-48h-EC50 |
>100 |
Concentration K2Cr2O7 (mg/L) |
Number Exposed |
% immobile |
Expected response (%) After 48 hours1 |
|||
24h |
48h |
Minimal |
Maximal |
|||
control |
20 |
0 |
0 |
0 |
102 |
|
0.10 |
20 |
0 |
0 |
0 |
10 |
|
0.18 |
20 |
0 |
0 |
0 |
10 |
|
0.32 |
20 |
0 |
10 |
0 |
30 |
|
0.56 |
20 |
10 |
100 |
0 |
100 |
|
1.0 |
20 |
95 |
100 |
40 |
100 |
|
1.8 |
20 |
100 |
100 |
100 |
100 |
|
( ) between brackets: Number of Daphnia observed trapped at the surface of the test solutions. This organism was reimmersed into the respective solution before recording mobility.
1- Based on historical data of the previous years (n>60)
2 – A maximum response of 10% does not invalidate the results of the test
Time (h) |
Replicate |
Nominal concentration Dilithium sebacate (mg/L) |
||||
Control |
0.10 |
1.0 |
10 |
100 |
||
0 |
A |
5 |
5 |
5 |
5 |
5 |
B |
5 |
5 |
5 |
5 |
5 |
|
C |
4 |
5 |
5 |
5 |
5 |
|
D |
5 |
5 |
5 |
5 |
5 |
|
Total introduced |
20 |
20 |
20 |
20 |
20 |
|
24 |
A |
0 (1) |
0 |
0 |
0 |
0 |
B |
0 |
0 |
0 |
0 |
0 |
|
C |
0 |
0 |
0 |
0 |
0 |
|
D |
0 |
0(1) |
0 |
0 |
0 |
|
Total immobilised |
0 |
0 |
0 |
0 |
0 |
|
Effect % |
0 |
0 |
0 |
0 |
0 |
|
|
|
|
|
|
||
48 |
A |
0 |
0 |
0 |
0 |
0 |
B |
0 |
0 |
0 |
0 |
0 |
|
C |
0 |
0 |
0 |
0 |
0 |
|
D |
0 |
0 |
0 |
0 |
1 |
|
Total immobilised |
1 |
0 |
0 |
0 |
1 |
|
Effect % |
5 |
0 |
0 |
0 |
5 |
|
Parameter |
Nominal concentration Dilithium sebacate (mg/L) |
24-48h-EC50 |
>100 |
Concentration K2Cr2O7 (mg/L) |
Number Exposed |
% immobile |
Expected response (%) After 48 hours1 |
|||
24h |
48h |
Minimal |
Maximal |
|||
control |
20 |
0 |
0 |
0 |
102 |
|
0.10 |
20 |
0 |
0 |
0 |
10 |
|
0.18 |
20 |
0 |
0 |
0 |
10 |
|
0.32 |
20 |
0 |
10 |
0 |
30 |
|
0.56 |
20 |
10 |
100 |
0 |
100 |
|
1.0 |
20 |
95 |
100 |
40 |
100 |
|
1.8 |
20 |
100 |
100 |
100 |
100 |
|
( ) between brackets: Number of Daphnia observed trapped at the surface of the test solutions. This organism was reimmersed into the respective solution before recording mobility.
1- Based on historical data of the previous years (n>60)
2 – A maximum response of 10% does not invalidate the results of the test
Validation of Mixing Period
Pre-study work indicated that there was no significant increase in the amount of total organic carbon by extending the preparation period for longer than 24 hours. As such, for the purposes of testing, the WAF was prepared using a stirring period of 23 hours followed by a 1 -Hour standing period.
Range-finding Test
Cumulative immobilisation data from the exposure of Daphnia magna to the test item during the range-finding test are given inTable1.
No significant immobilisation was observed at the 1.0, 10 and 100 mg/l loading rate WAFs.
Immobilisation data
Cumulative immobilisation data from the exposure of Daphnia magna to the test item during the definitive test are given in Table 2.
There was no 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) |
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 zero immobilisation at this loading rate.
It was considered unnecessary and unrealistic to test at loading rates in excess of 100 mg/l.
*EL = Effective Loading rate
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.
Observations on test item solubility
Observations on the test media were carried out during the mixing and testing of the WAFs.
At the start of the mixing period the 100 mg/l loading rate was observed to be a clear, colourless water column, with test item floating on the surface. After 23 hours stirring and a 1-Hour standing period the 100 mg/l loading rate was observed to remain a clear, colourless water column with test item floating on the surface. Microscopic examination of the WAF showed micro-dispersions of test item to be present and therefore it was considered appropriate to remove the WAF by filtering through a glass wool plug (2-4 cm in length). Microscopic examination after filtering showed the glass wool plug had removed all the micro-dispersions of the test item. During the test the 100 mg/l loading rate was observed to be a clear, colourless solution.
Total organic carbon analysis
Total Organic Carbon (TOC) analysis of the test preparations was conducted at 0 and 48 hours. Given the background level of carbon in the control vessels and also the low level of carbon in the test vessels, it was considered that the results gave no evidence of the presence of test item in the WAF.
Given that toxicity cannot be attributed to a single component or mixture of components but to the test item as a whole and the dissolved test item was below the quantifiable limit of the analytical method, the results were based on nominal loading rates only.
Validation of Mixing Period
Pre-study work indicated that there was no significant increase in the amount of total organic carbon by extending the preparation period for longer than 24 hours. As such, for the purposes of testing, the WAF was prepared using a stirring period of 23 hours followed by a 1 -Hour standing period.
Range-finding Test
Cumulative immobilisation data from the exposure of Daphnia magna to the test item during the range-finding test are given inTable1.
No significant immobilisation was observed at the 1.0, 10 and 100 mg/l loading rate WAFs.
Immobilisation data
Cumulative immobilisation data from the exposure of Daphnia magna to the test item during the definitive test are given in Table 2.
There was no 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) |
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 zero immobilisation at this loading rate.
It was considered unnecessary and unrealistic to test at loading rates in excess of 100 mg/l.
*EL = Effective Loading rate
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.
Observations on test item solubility
Observations on the test media were carried out during the mixing and testing of the WAFs.
At the start of the mixing period the 100 mg/l loading rate was observed to be a clear, colourless water column, with test item floating on the surface. After 23 hours stirring and a 1-Hour standing period the 100 mg/l loading rate was observed to remain a clear, colourless water column with test item floating on the surface. Microscopic examination of the WAF showed micro-dispersions of test item to be present and therefore it was considered appropriate to remove the WAF by filtering through a glass wool plug (2-4 cm in length). Microscopic examination after filtering showed the glass wool plug had removed all the micro-dispersions of the test item. During the test the 100 mg/l loading rate was observed to be a clear, colourless solution.
Total organic carbon analysis
Total Organic Carbon (TOC) analysis of the test preparations was conducted at 0 and 48 hours. Given the background level of carbon in the control vessels and also the low level of carbon in the test vessels, it was considered that the results gave no evidence of the presence of test item in the WAF.
Given that toxicity cannot be attributed to a single component or mixture of components but to the test item as a whole and the dissolved test item was below the quantifiable limit of the analytical method, the results were based on nominal loading rates only.
Description of key information
This endpoint is completed using data read across from structurally similar substances, dilithium sebacate and fatty acids, lanolin.
The 48-h EC50 is > 100 mg/L for dilthium sebacate. For fatty acids lanolin the 48-hour EL50 is > 100 mg/L loading rate WAF.
Key value for chemical safety assessment
Fresh water invertebrates
Fresh water invertebrates
- Effect concentration:
- 100 mg/L
Additional information
No data are available for the substance fatty acids, lanolin, lithium salts. Read across from dilithium sebacate and fatty acids, lanolin is used to complete the short term toxicity to aquatic invertebrates endpoint.
Two key studies conducted with read across substances, dilithium sebacate and fatty acids lanolin are available for this endpoint.
In the study with dilithium sebacate, five daphnids per test vessel were exposed for 48 hours at five test concentrations 10, 18, 32, 56 and 100 mg/L under static conditions. No effects on the mobility of Daphnia magna were observed in the study, and the 48-h EC50 is > 100 mg/L for dilthium sebacate.
In the study with fatty acids, lanolin, twenty daphnids (4 replicates of 5 animals) were exposed to a Water Accommodated Fraction (WAF) of the test item, at a single nominal loading rate of 100 mg/L for 48 hours under static test conditions. Immobilisation and any adverse reactions to exposure were recorded after 24 and 48 hours. The 48-hour EL50 for Daphnia magna based on nominal loading rates was greater than 100 mg/L loading rate WAF.
The studies were conducted according to the OECD 202 guideline and EC method C.2 and are GLP-compliant, therefore are considered reliable and suitable for this endpoint.
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