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Long-term toxicity to aquatic invertebrates

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Reference
Endpoint:
long-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:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Identification: Ethylene dibenzoate CAS 94-49-5
Physical state/Appearance: Off white solid flakes
Batch: 100000005282
Purity: 99.4%
Analytical monitoring:
yes
Vehicle:
no
Details on test solutions:
The test item solutions were prepared by stirring an excess (100 mg/L) of test item in culture medium using a propeller stirrer at approximately 1500 rpm for 48 hours. After the stirring period any undissolved test item was removed by filtration (0.2 µm Sartorius Sartopore filter, first approximate 2 liters discarded in order to pre condition the filter) to produce a 100% v/v saturated solution of the test item. This saturated solution was then further diluted as necessary, to provide the remaining test groups.
Test organisms (species):
Daphnia magna
Details on test organisms:
The daphnia were fed daily with a mixture of algal suspension and Tetramin® flake food suspension.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Nominal and measured concentrations:
Only the test concentrations where effects were found were analyzed, therefore only for the two highest concentrations measured concentrations are available.
Nominal concentrations: 6.25, 12.5, 25, 50 and 100% v/v saturated solution
The geometric mean measured concentrations: 0.65 and 1.4 mg/L.
Reference substance (positive control):
not specified
Key result
Duration:
21 d
Dose descriptor:
EC10
Effect conc.:
0.79 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
1.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.65 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
1.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
EC10
Effect conc.:
> 1.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
immobilisation
Details on results:
Analytical results:
A decline in measured test concentration of the aged test preparations on Days 1, 6, 13 and 21 was observed to between 0.31 and 1.6 mg/Land hence it was considered appropriate to calculate the results based on the geometric mean measured test concentrations only in order to give a “worst case” analysis of the data.
Validity criteria fulfilled:
yes
Conclusions:
The study is valid because it meets all validity criteria. The lowest EC10 is 0.79 mg/L measured geometric mean.
Executive summary:

Introduction

A study was performed to assess the chronic toxicity of the test item to Daphnia magna.  The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (2012) No 211, "Daphniamagna Reproduction Test" referenced as Method C.20 of Commission Regulation (EC) No. 440/2008.

Methods

Preliminary solubility work conducted indicated that it was not possible to obtain a testable solution of the test item using traditional methods of preparation e.g. ultrasonication and high shear mixing.

A preliminary media preparation trial indicated that a saturated solution method of preparation was most appropriate for this test item.

Based on the results of a preliminary range‑finding test, Daphnia magna were exposed (10 replicates of a single daphnid per group) to solutions of the test item at nominal concentrations of 6.25, 12.5, 25, 50 and 100% v/v saturated solution for a period of 21 days.  The test solutions were renewed daily throughout the test.  The test item solutions were prepared by stirring an excess (100 mg/L) of test item in culture medium using a propeller stirrer at approximately 1500 rpm for 48 hours.  After the stirring period any undissolved test item was removed by filtration (0.2 µm Sartorius Sartopore filter, first approximate 2 liters discarded in order to pre‑condition the filter) to produce a 100% v/v saturated solution of the test item.  This saturated solution was then further diluted as necessary, to provide the remaining test groups.

The numbers of live and dead adult daphnia and young daphnids (live and dead) were determined daily.  The daphnia were fed daily with a mixture of algal suspension and Tetramin®flake food suspension.

Results

Analysis of the fresh 50 and 100% v/v saturated solution test preparations on Days 0, 5, 12 and 21 showed measured test concentrations to range from 0.56 to 2.0 mg/L.  A decline in measured test concentration of the aged test preparations on Days 1, 6, 13 and 21 was observed to between 0.31 and 1.6 mg/Land hence it was considered appropriate to calculate the results based on the geometric mean measured test concentrations only in order to give a “worst case” analysis of the data.  The geometric mean measured concentrations were 0.65 and 1.4 mg/L.

Exposure of Daphnia magna to the test item gave the following results based on the geometric mean measured test concentrations:

 

Endpoint

Geometric Mean Measured Concentration (mg/L)

Immobilization

EC10

>1.4

 

EC50

>1.4

Reproduction

EC10

0.79

 

95% confidence limits

0.73 – 0.84

 

EC50

1.4

 

95% confidence limits

Not possible to determine due to the nature of the data

No Observed Effect Concentration

0.65

Lowest Observed Effect Concentration

1.4

Description of key information

A long-term study according to OECD 211 guideline under GLP was conducted with Daphnia magna.

The test was performed with a semi-static system, and test solutions were renewed daily.

The geometric mean exposure concentration were calculated and used for endpoint calculations.

 

Endpoint

Geometric Mean Measured Concentration (mg/L)

Immobilization

EC10

>1.4

 

EC50

>1.4

Reproduction

EC10

0.79

 

95% confidence limits

0.73 – 0.84

 

EC50

1.4

 

95% confidence limits

Not possible to determine due to the nature of the data

No Observed Effect Concentration

0.65

Lowest Observed Effect Concentration

1.4

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater invertebrates:
0.79 mg/L

Additional information