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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report date:
2013

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:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Details on test material:
- Purity: 100%

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Duplicate samples were collected from each batch of prepared test solution at the beginning of the test, and from two of the four replicate test chambers at 48 hours (± 1 hour) to measure concentrations of the test substance. All samples were collected at mid-depth from the batch solutions or the test chambers, placed in glass scintillation vials, and acidified with 2 drops of 10% phosphoric acid. One set of samples was processed immediately for analysis while the second set was stored refrigerated for possible future analysis.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION:
The highest concentration test solution was prepared by mixing 0.2400 g of test substance in 2 L of UV sterilized dilution water at a nominal concentration of 120 mg a.s./L. The test solution was adjusted to 100% active ingredient during preparation, based on the test substance purity (100%), and also served as the primary stock solution for preparing the remaining test solutions. The 120 mg a.s./L test solution was sonicated for 40 minutes, stirred on a stir plate for 2 hours, and inverted to mix. Aliquots of the 120 mg a.s./L test solution were proportionally diluted with dilution water to prepare four additional test solutions at nominal concentrations of 7.5, 15, 30 and 60 mg a.s./L. The test solutions were stirred on a stir plate for 60 minutes, and the 7.5, 15, and 30 mg a.s./L test solutions appeared clear and colourless. The 60 and 120 mg a.s./L test solutions were clear and colourless, with an oily sheen on the surface. After mixing, 220 mL of solution was placed in each of four replicate test chambers per treatment group. The negative control solution was dilution water only. At test termination the 7.5, 15, 30, and 60 mg a.s./L test solutions appeared clear and colourless. The 120 mg a.s./L test solution was clear and colourless, with an oily sheen on the surface.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM:
- Strain: Daphnia magna from cultures maintained by Wildlife International
- Age at study initiation: Daphnid neonates used in the test were less than 24 hours old
- Feeding during test: None

ACCLIMATION
- Acclimation period: The four adult daphnids used to supply neonates for the test were held for 21 days prior to collection of the juveniles for testing and had each produced at least three previous broods.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Hardness:
136 (mg/L as CaCO3)
Test temperature:
20 ± 1 °C
pH:
8.1 - 8.4
Dissolved oxygen:
≥8.0 mg/L (≥89% of saturation)
Salinity:
freshwater
Nominal and measured concentrations:
Nominal concentrations: 0, 7.5, 15, 30, 60 and 120 mg a.s./L
Measured concentrations:
Details on test conditions:
TEST SYSTEM
- Test vessel: 250-mL glass beakers containing 220 mL of test water
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4


TEST MEDIUM / WATER PARAMETERS:
Temperature was measured in each test chamber at the beginning and end of the test and at approximately 24 hours during the test using a liquid-in-glass thermometer. Temperature also was monitored continuously in one representative test chamber using an AmegaView centralized monitoring system. Dissolved oxygen and pH were measured in each test chamber at the beginning and end of the test and at approximately 24 hours during the test. Hardness, alkalinity, specific conductance and total organic carbon (TOC) were measured in the dilution water at the beginning of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: A photoperiod of 16 hours of light and 8 hours of darkness was controlled with an automatic timer. A 30-minute transition period of low light intensity was provided when lights went on and off to avoid sudden changes in lighting.
- Light intensity: 557 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
All organisms were observed periodically to determine the number of immobile organisms in each treatment group. Immobility was defined as a lack of movement by the organism except for minor activity of the appendages. The numbers of individuals exhibiting signs of toxicity or abnormal behaviour also were evaluated. Observations were made approximately 2, 24 and 48 hours after test initiation.

Results and discussion

Effect concentrations
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 111 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
The highest mean measured test concentration causing no immobility at test end was 27 mg a.s./L. The lowest mean measured test concentration causing 100% immobility at test end was greater than 111 mg a.s./L.
Reported statistics and error estimates:
There was less than 50% immobility for daphnids in any of the test substance treatment groups during the test, which precluded the statistical calculation of EC50 values at 24 and 48 hours. Therefore, the EC50 values were estimated to be greater than the highest concentration tested. The highest mean measured test concentration causing no immobility at test end and the lowest mean measured test concentration causing 100% immobility at test end were assessed by visual observation of the immobility and observation data.

Any other information on results incl. tables

Table 2. Immobility

 

 

~2 Hours

24 Hours

48 Hours

Percent

Immobile

Mean Measured

Concentration

(mg a.s./L)

Daphnia/

Replicate

Number Immobile1

Number Immobile1

Number Immobile1

 

Negative Control

5

5

5

5

 

0

0

0

0

 

0

0

0

0

 

0

0

0

0

 

0

 

7.1

5

5

5

5

 

0

0

0

0

 

0

0

0

0

 

0

0

0

0

 

0

14

5

5

5

5

 

0

0

0

0

 

0

0

0

0

 

0

0

0

0

 

0

27

5

5

5

5

 

0

0

0

0

 

0

0

0

0

 

0

0

0

0

 

0

55

5

5

5

5

 

0

0

0

0

 

0

0

0

0

 

0

0

1

2

15

111

5

5

5

5

0

0

0

0

 

0

0

0

0

 

2

1

2

2

35

1 Cumulative number of immobile daphnids.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
48-hour EC50 > 111 a.s./L
Executive summary:

Daphnids were exposed to a geometric series of five test concentrations and a negative control (dilution water) for 48 hours under static conditions. Four replicate test chambers were maintained in each treatment and control group, with five daphnids in each test chamber for a total of 20 daphnids per concentration. Nominal test concentrations were selected in consultation with the Sponsor and were based on the results of exploratory range finding toxicity data. Nominal test concentrations selected were 7.5, 15, 30, 60 and 120 mg a.s./L. The mean measured concentrations were 7.1, 14, 27, 55, and 111 mg a.s./L.

The highest mean measured test concentration causing no immobility at test end was 27 mg a.s./L. The lowest mean measured test concentration causing 100% immobility at test end was greater than 111 mg a.s./L. The 48-hour EC50 value was greater than 111 mg a.s./L based on mean measured test concentrations.