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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2012-04-20 to 2012-04-26
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Performed according to relevant guidelines and compliant to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
according to German Chemikaliengesetzt and directive 88/320/EEC

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
no

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution of 100 mg/L was freshly prepared with dilution water. The stock solution was stirred for 72 hours (1100 rpm) with a magnetic stirrer. Undissolved particles were removed by centrifugation for 2 x 10 min at 10000 rpm in plastic centrifuge tubes.
- Eluate: Dilution water
- Differential loading: None, saturated soklution corresponding to a nominal test item concentration of 100 mg/L
- Controls: Six replicates (without test item) were tested under the same test conditions as the test replicates.

Test organisms

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Desmodesmus subspicatus Chodat
- Strain: SAG 86.81
- Source (laboratory, culture collection): SAG Pflanzenphysiologisches Institut der Universitaet Goettingen, Nikolausberger Weg 18, D-37073 Goettingen, Germany
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: Fresh stocks were prepared every month on Z-Agar. Light intensity amounted to 35 - 70 µE x m-2 x s-1 for 24 h per day.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
No

Test conditions

Hardness:
Dilution water: 0.24 mmol Ca+Mg/L.
Test temperature:
min.: 21.0 max.: 21.5, mean value: 21.3
pH:
pH-value
Start; 0 hours End; 72 hours
Saturated solution
prepared with 100 mg test item/L 7.91 8.01
Control 7.90 8.02
Dissolved oxygen:
Not measured
Salinity:
Not measured, freshwater conditions
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type (delete if not applicable): open, cotton wool plugs
- Material, size, headspace, fill volume: sterile 250 mL Erlenmeyer flasks, test volume 100 mL
- Aeration: Test containers were placed on a rotary shaker and oscillated at appr. 70 rpm
- Type of flow-through (e.g. peristaltic or proportional diluter): None
- Renewal rate of test solution (frequency/flow rate): One application at test start
- Initial cells density: Nominal: approximately 2 - 5 x 10^3 cells/mL, Actual: mean 4910 cells/mL
- Control end cells density: 979354 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6



GROWTH MEDIUM
- Standard medium used: yes



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dilution water, composition acc. to the guideline
- Hardness: 0.24 mmol Ca+Mg/L
- pH-value: 8.1 +/- 0.2
- Culture medium: Nutrient Medium Z acc. to LÜTTGE et al. (1994)


OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: 24 h
- Light intensity and quality: 60 - 120 µE x m-2 x s-1, mean value: 69.2



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter] fluorimeter, Cell density and self fluorescence was measured daily via Chlorophyll-a- fluorescence, excitation at 436 nm, emission at 685 nm. Dilution water was used as background signal.



TEST CONCENTRATIONS
- Range finding study:
No


Reference substance (positive control):
yes
Remarks:
Potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
100 other: mg/L (saturated solution)
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
& yield
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 other: mg/L (saturated solution)
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
& yield
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
& yield
Details on results:
- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test):
- Unusual cell shape: No
- Colour differences: No
- Flocculation: No
- Adherence to test vessels: No
- Aggregation of algal cells: No
- Other: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No
Results with reference substance (positive control):
- Results with reference substance valid? Yes
Current Study Valid Range (average ± 3 x SD)
Rate-related inhibition
ErC50 0.540 0.686 ± 0.470
95 % confidence interval 0.528 - 0.551
Yield inhibition
EyC50 0.324 0.319 ± 0.140
95 % confidence interval 0.304 - 0.347
SD = Standard deviation



Reported statistics and error estimates:
NOEC/LOEC was determined by calculation of statistical significance of growth rates and yield. As standard One Way Analysis of Variance (ANOVA) was used for NOEC/LOEC calculations. When running a One Way Analysis of Variance a Normality test and an Equal Variance test were done first. The SHAPIRO-WILK-Test was used to test for normally distributed populations. The Levene median test was used for equal variance. P-values for both Normality and Equal Variance test were 0.05. The a-value (acceptable probability of incorrectly concluding that there is a difference) is a=0.05.

Any other information on results incl. tables

NOEC, LOEC, EC - values of the test item (0-72 hours)

                   based on the saturated solution prepared with 100 mg test item/L

                               

Rate-related Inhibition

NOEC

100

LOEC

> 100

ErC10

> 100

ErC20

> 100

ErC50

> 100

Inhibition of Yield

NOEC

100

LOEC

> 100

EyC10

> 100

EyC20

> 100

EyC50

> 100

Cell Densities

 

Replicate

Cell density [cells/mL]

No.

0 hours

24 hours

48 hours

72 hours

 

Saturated solution prepared with 100 mg test item/L

1

4910

29190

195702

1088814

 

2

4910

24400

189406

1131566

 

3

4910

30896

202993

1133022

 

4

4910

29478

198944

945086

 

5

4910

26465

178127

872174

 

6

4910

25798

180605

1132852

 

Mean

4910

27705

190963

1050586

 

Control

1

4910

26117

161764

935439

 

2

4910

22278

159597

905211

 

3

4910

25477

191728

1191041

 

4

4910

23781

157519

880365

 

5

4910

28462

175087

1018981

 

6

4910

25138

152915

945086

 

Mean

4910

25209

166435

979354

 

Evaluation after 72 hours

                   Statistically significant differences of growth rates and yield compared to

                   control values are marked (+), not significant differences are marked (-).

Replicate

Growth rate

Rate-related inhibition

Yield

Inhibition of yield

No.

[d-1]

[%]

[cells/mL]

[%]

Saturated solution prepared with 100 mg test item/L

1

1.80

-2.10

1083904

-11.2

2

1.81

-2.83

1126656

-15.6

3

1.81

-2.85

1128112

-15.8

4

1.75

0.57

940176

3.52

5

1.73

2.09

867264

11.0

6

1.81

-2.85

1127942

-15.8

Mean

(-)

1.79

-1.33

(-)

1045676

-7.31

Control

1

1.75

930529

2

1.74

900301

3

1.83

1186131

4

1.73

875455

5

1.78

1014071

6

1.75

940176

Mean

1.76

974444

Section-by-Section and Average Specific Growth Rates of the Control Group
                   (0 – 72 hours)

Replicate No.

Specific growth rate [d-1]

Mean

(0 - 72 h)

SD

±

CV
[%]

Mean CV [%]

section-by-section                           

0 - 24 hours

24 - 48 hours

48 - 72 hours

Control

1

1.67

1.82

1.76

1.75

0.0762

4.36

7.42

2

1.51

1.97

1.74

1.74

0.228

13.1

3

1.65

2.02

1.83

1.83

0.186

10.2

4

1.58

1.89

1.72

1.73

0.157

9.06

5

1.76

1.82

1.76

1.78

0.0332

1.87

6

1.63

1.81

1.82

1.75

0.104

5.96

Mean

1.76

 

SD ±

0.04

 

CV [%]

2.08

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
In this study no inhibition on growth of the freshwater green alga Desmodesmus subspicatus was found after 72 hours exposure to the test item. The NOEC for inhibition of specific growth rate and yield after 72 hours was 100 mg/L. All effect levels are given based on the saturated solution prepared with nominal 100 mg test item/L. As the test item is highly insoluble in water, no suitable method for determination of the test item could be established.
Executive summary:

The toxicity of the test item to the unicellular freshwater green alga Desmodesmus subspicatus was determined according to the principles of OECD 201. The aim of the study was to assess the effects on growth rate and yield over a period of 72 hours.

The study was conducted under static conditions with an initial cell density of 4910 cells/mL. A saturated solution with a nominal concentration of 100 mg/L was tested. Six replicates were tested for the limit concentration and the control. Environmental conditions were determined to be within the acceptable limits.

No analytical determination was carried out since the test item is highly insoluble in water and no suitable method for determination of the test item could be established. All effect levels are given based on the saturated solution prepared with nominal 100 mg test item/L.

NOEC, LOEC, EC - values of test item (0-72 hours)

                   based on the saturated solution prepared with 100 mg test item/L

                               

Rate-related Inhibition

NOEC

100

LOEC

> 100

ErC10

> 100

ErC20

> 100

ErC50

> 100

Inhibition of Yield

NOEC

100

LOEC

> 100

EyC10

> 100

EyC20

> 100

EyC50

> 100