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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)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:

Water samples of each concentration (2 x 100 ml volume each) were taken from the freshly prepared test solutions at the start of exposure. At test termination (48 h samples of 50 ml were taken from 2 replicates of each concentration and on beaker was taken from each concentration (100 ml).
All samples were kept at -18 °C to -22 °C until analysis.

The specimens were thawed in a microwave oven at ambient temperature. After intensively shaking, the measured specimens were diluted with
acetonitrile in the ratio 1:1. The final volume was measured to calculate the dilution factor.

Test solutions

Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

Reference solution:
20.45 and 41.04mg CA 2218 A were weighed, dissolved and made up to 25.0 ml with acetonitrile. From these stock solutions, reference solutions in the range of the test concentrations (related to and extended to about 10 % below and above the expected concentration) were diluted with
acetonitrile/test water = 1/1 (v/v). So references with concentrations between 0.103 - 8.21 mg/L CA 2218 A were used.

Test solution:
The measured specimens were diluted with acetonitrile in the ratio 1:1. The final volume was measured to calculate the dilution factor.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna STRAUS
- Strain: Clone 5
- Source: In house culture
- Age at study initiation: < 24 hours

Study design

Test type:
static
Limit test:
yes
Total exposure duration:
48 h

Test conditions

Hardness:
272 mg CaCO3/L
Test temperature:
21 ± 1 °C
pH:
Start of exposure: 8.5 - 8.6
End of exposure: 8.4 - 8.5
Dissolved oxygen:
Oxygen saturation at start of exposure: 98 - 100 %
Oxygen saturation at end of exposure: 84 - 88 %
Nominal and measured concentrations:
NOMINAL CONCENTRATIONS:
0.50, 1.0, 2.0, 4.0, 8.0 and 16.0 mg test item/L.

MEASURED CONCENTRATIONS:
Start of exposure: 0.28, 0.56, 1.2, 2.2, 3.9 and 7.5 mg test item/L.
End of exposure: 0.30, 0.75, 1.6, 2.5, 4.8 and 7.1 mg test item/L.
Details on test conditions:
TEST PARAMETERS

- Procedure: Static system.
- Duration: 48 hours.
- Test vessels: 150 ml glass beakers containing 100 ml of test solution or blank test medium.
- Replicates: 4 test vessels for each concentration and the control.
- Animals/concentration: 20 for each concentration and the control.
- Animals/test vessel: 5 exposed per test vessel.

TEST MEDIUM / WATER PARAMETERS

- Test medium: Reconstituted water (M4-medium)
- Control Blank ( M4-medium)
- Temperature: 21 ± 1 °C
- Photoperiod: Light/dark cycle - 16/8 hours with 30 min transistion period.
- Light intensity: Fluorescent light (about 1130 lux)

TEST CONCENTRATIONS

STOCK SOLUTIONS

Preparation of stock solutions and test solutions:

100 mg of CA 2218 A were mixed and made up to 1000 ml with M4-medium (SL1). 160, 80, 40, 20, 10 and 0.5 ml of SL1 were mixed and made up to 1000 ml with M4-medium to prepare test solutions with test concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16 mg/L, respectively.

TEST CONCENTRATIONS

Range-finding Test:

109 mg of CA 2218 A were prepared and made up to 1000 ml with M-4 medium to prepare the stock solution. 25, 2.5 and 0.25 ml of this stock
solution were mixed and made up to 250 ml with M4-medium to prepare test solutions with test concentrations of 10, 1.0 and 0.1 mg/L.

Remarks:
Calculated amounts of the stock solution to produce the desired test concentrations were given into the reconstituted water and were homogenously distributed. Small parts of the test item were swimming on the surface of the test solutions at all test concentrations at test start.


Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
ca. 3.4 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % CL
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
ca. 0.89 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % CL
Details on results:
No immobilization of daphnids was observed in the control and at nominal test concentrations of 0.5 mg/L. After exposure for 48 hours 25, 85, 100, 100 and 100 % of daphnids were immobilized at nominal test concentrations of 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L.

No immobilization of daphnids was observed in the control (blank). In addition, during the biological phase of the study ,no unexpected phenomena were recorded. Therefore, the validity criteria regarding the biology of the test were fulfilled.
Reported statistics and error estimates:
A computer program according to the maximum liklihood method, the probit mdel (Finney, D.J., 1971) was run by using SAS Version 6.11 (SAS/STAT Users Guide, Vol 2, SAS Institute Inc.) to estimate EC50 values.

Any other information on results incl. tables

 

Exposure period

EC 50

95 % CL

Slope

No effect level (observed) mg/L

24

48

3.4

0.89

2.8 – 4.2

0.72 – 1.06

1.99

2.50

0.66

0.29

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The EC50 (48h) of CA 2218 A was determined to be 0.89 mg/L based on actual mean concentrations. The highest concentrations with no observed effects after 48 hours was 0.29 mg/L based on actual mean concentration of CA 2218 A.
Executive summary:

Test concentrations of nominal 0.50, 1.0, 2.0, 4.0, 8.0 and 16 mg/L of CA 2218 A were selected based on results of a range-finding test. Exposure started when the daphnids were placed in the test vessels containing the test solutions. Twenty daphnids were exposed to each of the desired test concentrations under static conditions in M-4 medium. For the test concentration and the control, four replicates were performed. Each replicated consisted of five specimens in 100 ml test solution. The daphnids were observed for immobilization and behavioural changes after 24 h and 48 h of exposure. 

Water temperature was maintained constant throughout the duration of the test at 21 ± 1 °C in all test conditions. The oxygen saturation was measured to be 98 – 100 % and 84 – 88 % at the start and at the end of exposure, respectively. The pH was 8.5 – 8.6 and 8.4 – 8.5 at the start and end of the exposure, respectively. 

The measured concentrations of CA 2218 A were 0.28, 0.56, 1.2, 2.2, 3.9 and 7.5 mg/L at the start of exposure and 0.30, 0.75, 1.6, 2.5, 4.8 and 7.1 mg/L corresponding for 104, 118, 117, 107, 112 and 97 % of measured concentrations at the start of exposure, respectively. 

No immobilization of daphnids was observed in the control and at nominal test concentrations of 0.5 mg/L. After exposure for 48 hours 25, 85, 100, 100 and 100 % of daphnids were immobilized at nominal test concentrations of 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L. 

Following values are based on actual mean concentrations. 

 

Exposure period

EC 50

95 % CL

Slope

No effect level (observed) mg/L

24

48

3.4

0.89

2.8 – 4.2

0.72 – 1.06

1.99

2.50

0.66

0.29

 

Conclusion:

The EC50 (48h) of CA 2218 A was determined to be 0.89 mg/L based on actual mean concentrations. The highest concentrations with no observed effects after 48 hours was 0.29 mg/L based on actual mean concentration of CA 2218 A.