Registration Dossier

Ecotoxicological information

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:
3/10/2017 to 31/10/2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
OECD Chemicals Testing Guideline No.202 Daphnia sp. Acute Immobilisation Test (adopted April 13 2004)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Specific details on test material used for the study:
Test Substance Name: Bernel Ester TCC

Chemical Name: Tris(2-ethylhexyl) 2-hydroxypropane-1,2,3-tricarboxylate
CAS Number: 7147-34-4
Batch Number: P7803
Purity: 94%
Receipt Date: 12 June 2017
Expiry Date: 23 January 2019
Storage on Receipt: Room temperature (15 - 30°C)

Test substance details were supplied by the Sponsor. The Certificate of Analysis is presented in Appendix 4 (attached).

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Juvenile Daphnia magna (Straus), less than 24 hours old, were taken from healthy parental laboratory cultures, held and maintained at the test facility. The culture was originally obtained from Microbiotest, Belgium. Details of the Daphnia magna culture maintenance are presented in Appendix 1 (attached).

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h

Test conditions

Test temperature:
Control 19.2 °C (0 hours) to 18.6 °C (48 hours)
100 19.4 °C (0 hours) to 18.6 °C (48 hours)
pH:
Control 7.80 (0 hours) to 7.76 (48 hours)
100 7.87 (0 hours) to 7.84 (48 hours)
Dissolved oxygen:
Control 8.40 mg/l (0 hours) to 9.38 mg/l (48 hours)
100 8.49 mg/l (0 hours) to 9.34 mg/l (48 hours)
Reference substance (positive control):
yes
Remarks:
Potassium dichromate, Batch No. 16F014128, Purity: 100%, expiry date: 31 Jan 2021

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 0.185 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
ca. 0.185 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility

Any other information on results incl. tables

Range-finding Test

The results of the range-finding test are summarised below:

Nominal
concentration
(% saturated
solution)
Cumulative immobilisedDaphnia magna*
24 hours 48 hours % immobility
R1 R2 R1 R2  
Control  0  0  0  0
1 0 0 0 0 0
10 0 0 0 0 0
100 0 0 0 0 0

* Initial population of Daphnia magna: five per replicate

R Replicate

Chemical analysis was not conducted during the range-finding test.

The results of the range-finding test suggested that toxicity would not be observed at the highest concentration of 100% saturated solution.

Media Trial

Media trial results are summarised below:

Nominal
concentration
(% saturated
solution)
Measured concentrations (mg/l)
0 hours 48 hours
100 0.389* 0.144

LOQ (limit of quantification) = 0.01 mg/L

* Initial batch failed to meet acceptance criteria, due to results being above the calibration line. Samples re-run with 48 hour samples.

Results showed a decline in concentration over the 48-hour test period. Therefore, the definitive phase was run semi-static.

Definitive Test

Chemical Analysis

The results of the chemical analysis are presented in Table 1 (attached).

Example chromatograms of test samples, standard solution and a typical calibration line are presented in Appendix 2 (attached).

The limit of quantification (LOQ) for Bernel Ester TCC in Elendt M4 medium using this method was 0.01 mg/L.

Analysis of the test media samples was conducted on fresh media at 0 and 24 hours, analysis of the corresponding old media was conducted at 24 and 48 hours.

Chemical analysis showed concentrations of Bernel Ester TCC declined over each 24-hour renewal period. Therefore, results were based on time-weighted mean measured concentrations. This was calculated to be 0.185 mg/L

Water Quality

Water quality determinations for pH, temperature and dissolved oxygen (% ASV and mg/L) during the definitive test are presented in Table 2 (attached).

Test Media Descriptions

The test preparations were observed to be colourless solutions throughout the duration of the test.

Toxicity to Daphnia magna

The numbers of immobilised Daphnia magna at 24 and 48 hours, following exposure are presented in Table 3 and Table 4 (attached). The percentage immobility data is summarised below.

Nominal
concentration
(% saturated
solution)
Cumulative immobilisedDaphnia magna* (initial population: 5Daphniaper replicate)
24 hours 48 hours
R1 R2 R3 R4 % immobility R1 R2 R3 R4 % immobility
Control 0 0 0 0 0 0 0 0 0 0
1 0 0 0 0 0 0 0 0 0 0
10 0 0 0 0 0 0 0 0 0 0
100 0 0 0 0 0 0 0 0 0 0

R Replicate

* All daphnia in 100% saturated solution test group were observed to be at the surface of the test media at 48 hours, one daphnia in the 100% saturated solution test group was observed to be at the surface of the test media at 24 hours.

No significant toxicity (>10%) was observed during the test. Due to the decline in test substance concentration observed over each renewal period results were based on time-weighted mean measured concentration. This was calculated to be 0.185 mg/L.

Statistical Analysis

The 3EC50 values, the corresponding 4NOEC and 3LOEC values are presented in the following table.

  Nominal concentration (mg/l) Statistical test
  24 hour 48 hour  
EC50 >0.185 mg/l >0.185 mg/l  
NOEC 0.185 mg/l 0.185 mg/l Derived Empirically
LOEC >0.185 mg/l >0.185 mg/l  

3 Concentration resulting in 50% immobility of exposed Daphnia magna

4 No observed effect concentration

3 Lowest observed effect concentration

Based on time-weighted mean measured concentrations, the 48-hour EC50 was calculated to be >0.185 mg/L.

The corresponding NOEC values was calculated to be >0.185 mg/L.

The validity criteria for control immobility (≤10%) and dissolved oxygen (≥3 mg/L) were both satisfied. The test is therefore considered valid.

Validity Criteria

The validity criteria for control immobility (≤10%) and dissolved oxygen (≥3 mg/L) were both satisfied. The test is therefore considered valid.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The 48-hour acute toxicity of Bernel Ester TCC to the freshwater planktonic crustacean, Daphnia magna, was determined in accordance with the requirements of OECD Chemicals Testing Guideline No. 202 Daphnia sp. Acute Immobilisation Test (adopted April 13 2004).
The validity criteria for control immobility (≤10%) and dissolved oxygen (≥3 mg/L) were both satisfied. The test is therefore considered valid.
Based on time-weighted mean measured concentration, the 48-hour EC50 value was determined to be >0.185 mg/L. The corresponding No Observed Effect Concentration (NOEC) was considered to be 0.185 mg/L.
Executive summary:

The objective of the study was to determine the 48-hour acute toxicity of Bernel Ester TCC against the mobility of the freshwater planktonic crustacean, Daphnia magna.

The study was conducted in accordance with the requirements of OECD Chemicals Testing Guideline No. 202 Daphnia sp. Acute Immobilisation Test (adopted April 13 2004).

Based on the results of a range-finding test, for which the key results only have been reported, a definitive limit test was conducted at a single concertation of 100% saturated solution. A control group was also included. The test was conducted under semi-static test conditions with media preparation at 0 and 24 hours.

At the start of the test, five juvenile Daphnia magna were added to each test vessel. The Daphnia magna in each test vessel were observed at 24 and 48 hours. The Daphnia magna were not fed during the test.

Chemical analysis of the 100% saturated solution test concentration and control was conducted on fresh media at 0 and 24 hours, analysis was conducted on the corresponding old media at 24 and 48 hours. Analysis of the test samples at 0, 24 and 48 hours are summarised in the table below.

Nominal
concentration
(% saturated
solution)
Measured concentration (mg/l)
0 hours (New) 24 hours (Old) 24 hours (New) 48 hours (Old)
R1 R2 R1 R2
Control - - - -
100 0.183 0.0673 0.341 0.183

- None found above LOQ (0.01 mg/L)

24-hour renewal period. Therefore, results were based on time-weighted mean measured concentrations. This was calculated to be 0.185 mg/L.

The1EC50values, the corresponding 2NOEC and 3LOEC values are presented in the following table.

Nominal concentration (mg/l) Statistical test
  24 hour 48 hour  
EC50 >0.185 mg/l >0.185 mg/l  
NOEC 0.185 mg/l 0.185 mg/l Derived Empirically
LOEC >0.185 mg/l >0.185 mg/l  

1Concentration resulting in 50% immobility of exposed Daphnia magna

2No observed effect concentration

3Lowest observed effect concentration

Based on time-weighted mean measured concentrations, the 48-hour EC50 was calculated to be >0.185 mg/L.

The corresponding NOEC values was calculated to be >0.185 mg/L.

The validity criteria for control immobility (≤10%) and dissolved oxygen (≥3 mg/L) were both satisfied. The test is therefore considered valid.