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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
06 - 23 Apr 2009
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Due to the nature of the test item, analytical measurement is not possible.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Version / remarks:
2006
Deviations:
yes
Remarks:
temperature exceeded 23 ±2 °C
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Qualifier:
according to
Guideline:
other: No.23-Guidance document on aquatic toxicity testing of difflcult substances and mixtures
GLP compliance:
yes (incl. certificate)
Remarks:
THE DEPARTMENT OF HEALTH OF THE GOVERNMENT OF THE UNITED KlNGDOM, UK GLP Monitoring Authority

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
no
Remarks:
Due to the nature of the test item, analytical measurement is not possible.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Differential loading: For each nominal concentration the required amount of homogenised sample was added to 500 mL of culture medium in an aspirator. The aspirators were mixed for 48 h at 21 to 23 °C and then allowed to separate for four hours. The aqueous phase, avoiding all settled and floating material, was drawn off and filtered using Whatman No. 1 filter paper. The resultant solution was used as the test exposure solution.
- Controls: test medium without test material

Test organisms

Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Green algae
- Strain: CCAP 278/4
- Source: Culture Collection of Algae and Protozoa, lnstitute of Freshwater Ecology Windermere Laboratory
- Age of inoculum (at test initiation): Exponential phase
- Method of cultivation: The tests and culture were performed in deionised water with added nutrients. Before use the water was sterilised by autoclaving at 120 °C for 30 min. Sterile nutrient stock solutions were then added and the pH value adjusted to 8.0 ± 0.2 to obtain the culture medium.

ACCLIMATION
- Culturing media and conditions: Temperature: 23 ± 2 °C, Illumination: 6000 - 10000 lux continous white light, Shaking: 200 rpm

Study design

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

Test conditions

Test temperature:
23.3 - 26.5 °C (26.5 °C was measured at test end in test and control treatments)
temperature exceeded 23 ± 2 °C
pH:
7.1 - 7.9 (Control)
7.1 - 7.6 (Test item concentrations)
Nominal and measured concentrations:
Control, 10, 18, 32, 56, 100 mg/L (nominal loading rate)
Details on test conditions:
TEST SYSTEM
Test vessel:
- Type: open
- Material, size, headspace, fill volume: 250 mL conical glass flask; 100 mL test solution, 150 mL headspace
- Initial cells density: 1 x 10E+04 cells/mL
- Control end cells density: 141.1 x 10E+04 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: De-ionised,flltered and sterilised by autoclaving at 120 °C for 30 min.
- Culture medium different from test medium: Same medium
- Intervals of water quality measurement: At test start and test end.

OTHER TEST CONDITIONS
- Photoperiod: Continously
- Light intensity and quality: 6000 to 10000 lux white light

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: Cell density was determined after 24, 48 and 72 h exposure periods. The cell counts were made using a haemocytometer and microscope. Cells, which showed abnormalities were included in the count but also reported in the observation chapter.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
Range finding study
- Test concentrations: Control, 0.1, 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: Percent of inhibition by growth rate 4, 3, 3 and 21% at 0.1, 1, 10 and 100 mg/L.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EL10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not possible to calculate 95% confidence limits
Duration:
72 h
Dose descriptor:
EL20
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not possible to calculate 95% confidence limits
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not possible to calculate 95% confidence limits
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): All cells appeared normal, no cells were observed that showed any morphological abnormalities.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- EC50 (72 h): 0.84 mg/L for growth rate (historical data: ErC50 (72 h): 0.86 mg/L)
Reported statistics and error estimates:
The statistical evaluation was caclulated with the program ToxCalcTMVersion 5.0 “Comprehensive Toxicity Data Analysis and Database software.

Any other information on results incl. tables

Biological Results

Table 1: Cell numbers (mean initial cell density: 1 x 10E+04 cells/mL)

Loading rate

Replicate

Cell density measurements (cells/mL x 10E+04)

[mg/L]

24 hours

48 hours

72 hours

0

1

5.0

21.3

130.7

2

5.7

26.7

139.0

3

5.0

17.7

126.0

4

4.3

21.3

154.0

5

7.3

29.7

159.7

6

6.0

18.3

137.0

Mean

5.6

22.5

141.1

10

1

4.3

17.0

95.0

2

5.3

17.7

153.3

3

5.3

15.0

88.7

Mean

5.0

16.6

112.3

18

1

4.0

15.3

92.7

2

7.0

19.0

90.0

3

5.3

15.3

74.0

Mean

5.4

16.5

85.6

32

1

7.7

17.0

80.7

2

6.7

17.0

139.7

3

4.0

16.7

68.7

Mean

6.1

16.9

96.4

56

1

5.3

11.0

116.3

2

5.0

15.0

89.7

3

4.3

10.7

72.7

Mean

4.9

12.2

92.9

100

1

4.0

16.0

109.0

2

3.3

16.3

90.7

3

5.3

12.3

88.3

Mean

4.2

14.9

96.0

 Table 2: Inhibition of growth rate

Loading rate

Percent inhibition by growth rate

[mg/L]

0 - 48 h

0 – 72 h

10

9

5

18

10

10

32

9

9

56

20

9

100

13

8

 

Table 3: Endpoints ELx and NOELR values by growth rate (based on the nominal loading rate)

Periode of exposure

ErLx value

[h]

ErL10

ErL20

ErL50

0 to 48 h

>100 mg/L*

>100 mg/L*

>100 mg/L*

0 to 72 h

>100 mg/L*

>100 mg/L*

>100 mg/L*

NOELR

100 mg/L (0 – 72 hours)

* Not possible to calculate 95% confidence limits

Table 4: Validity criteria

Criterion from the guideline

Outcome

Validity criterion fulfilled

The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period.

141

Yes

The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures must not exceed 35%

16.78

Yes

The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% in tests with Pseudokirchneriella subcapitata and Desmodesmus subspicatus. For other less frequently tested species, the value should not exceed 10%.

1.94

Yes

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
For further details please refer to “Any other information on results incl. tables”.