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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2011-12-16 to 2011-12-22
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution of 1 mg/l was freshly prepared with dilution water and stirred for 48 h (1100rpm). Undissolved particles were removed by membrane filtration (membrane filter 0.45 µm, RC, MACHEREY-NAGEL). Centrifugation for 20 min at 10000 rpm was found to be insufficient.
- Eluate: Dilution water
- Differential loading: None, limit test item concentration
- Controls: Six replicates without test item were tested under the same test conditions as the test replicates.
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.
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Hardness:
Dilution water: 0.24 mmol Ca+mg/L.
Test temperature:
22.5 - 24.0 °C, mean value: 23.3 °C
pH:
Nominal concentration pH-value
[mg/L] Start; 0 hours End; 72 hours
1.00 7.89 7.94
Control 7.90 7.97
Dissolved oxygen:
Not measured
Nominal and measured concentrations:
Nominal test concentrations: control and 1 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type: 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: None
- Renewal rate of test solution (frequency/flow rate): One application at test start
- Initial cells density: Nominal: approximately 2 - 5 x 103 cells/mL, Actual: mean 4910 cells/mL
- Control end cells density: Mean 634285 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
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: No acute toxic effects within the range of solubility.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: No acute toxic effects within the range of solubility.
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
EC50 Values of the Reference Item
based on nominal concentrations (mg/L), (0-72 hours)
Current Study Valid Range (average ± 3 x SD)
Rate-related inhibition
ErC50 0.488 0.699 ± 0.471
95 % confidence interval 0.466 – 0.512
Yield inhibition
EyC50 0.291 0.318 ± 0.147
95 % confidence interval 0.274 – 0.328


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) and DUNNETT’s Test were 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.

NOEC, LOEC, EC - values of PV-Echtbraun RL (0-72 hours)

                   based on nominal concentration [mg/L]

                               

Rate-related Inhibition

NOEC

1.00

LOEC

> 1.00

ErC10

> 1.00

ErC20

> 1.00

ErC50

> 1.00

Inhibition of Yield

NOEC

1.00

LOEC

> 1.00

EyC10

> 1.00

EyC20

> 1.00

EyC50

> 1.00

Cell Densities

 

Nominal concentration

Replicate

Cell density [cells/mL]

[mg/L]

No.

0 hours

24 hours

48 hours

72 hours

 

1.00

1

4910

29017

165338

711305

 

2

4910

21642

135977

771998

 

3

4910

22620

135753

676405

 

4

4910

26607

173530

760929

 

5

4910

27315

163950

683717

 

6

4910

24681

134846

697494

 

Mean

4910

25314

151566

716975

 

Control

1

4910

24999

142808

651898

 

2

4910

23784

149659

608401

 

3

4910

22508

146782

693872

 

4

4910

22454

139897

566799

 

5

4910

25470

140194

624886

 

6

4910

26851

163111

659853

 

Mean

4910

24344

147075

634285

 

Evaluation after 72 hours

                   Statistically significant differences of growth rates and yield compared to

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

Nominal concentration

Replicate

Growth rate

Rate-related inhibition

Yield

Inhibition of yield

[mg/L]

No.

[d-1]

[%]

[cells/mL]

[%]

1.00

1

1.66

-2.40

706395

-12.2

2

1.69

-4.09

767088

-21.9

3

1.64

-1.37

671495

  -6.69

4

1.68

-3.79

756019

-20.1

5

1.65

-1.59

678807

  -7.85

6

1.65

-2.00

692584

-10.0

Mean

(+)*

1.66

-2.54

(+)*

712065

-13.1

Control

1

1.63

646988

2

1.61

603491

3

1.65

688962

4

1.58

561889

5

1.62

619976

6

1.63

654943

Mean

1.62

629375

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

1.63

1.74

1.52

1.63

0.112

  6.88

11.0

2

1.58

1.84

1.40

1.61

0.220

13.7

3

1.52

1.88

1.55

1.65

0.195

11.8

4

1.52

1.83

1.40

1.58

0.222

14.0

5

1.65

1.71

1.50

1.62

0.109

  6.74

6

1.70

1.80

1.40

1.63

0.211

12.9

Mean

1.62

 

SD ±

0.02

 

CV [%]

1.45

 

Validity criteria fulfilled:
yes
Conclusions:
No acute toxic effects occured within the range of solubility. The test substance is with high probability acutely not harmful to aquatic algae.

Description of key information

The substance is with high probability not harmful to aquatic algae. 
No toxic effects occur at the range of the substances solubility.
EC50 (72h) and EC10 (72h) >= 1 mg/L (nominal) for growth rate (OECD 201)

Key value for chemical safety assessment

Additional information

The toxicity on aquatic algae was investigated for two ‘red disazo condensation pigments’(CAS 68516-75-6 and CAS 71566-54-6) [DR.U.NOACK-LABORATORIEN, 2012; lnstitute of lndustrial Organic Chemistry, Branch Pszczyna, 2012]. The studies were conducted with the green algae species Desmodesmus subspicatus (CAS 68516-75-6) and Pseudokirchneriella subcapitata (CAS 71566-54-6). Based on the low water solubility of the test item, saturated stock solutions were prepared and the tests were conducted as limit tests with nominal concentrations of 1 mg/L (CAS 68516-75-6) and 10 mg/L (CAS 71566-54-6), respectively. After 72 hours of exposure no toxic effects on algae observed, neither in the study with Desmodesmus subspicatus nor in the study with Pseudokirchneriella subcapitata.