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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:
12 - 15 Jul 1993
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: EU-Guideline 92/69 EWG
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: all concentrations
- Sampling method: samples were taken at 0 and 72 h.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: a stock solution was prepared by dissolving the substance in purified water. Concentration of the stock solution : 0.143 g/L
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Strain: 86.81 SAG
- Source (laboratory, culture collection): originally obtained from Institut für Wasser-, Boden- und Lufthygiene, Berlin. In-house culture
- Age of inoculum (at test initiation): 3 days
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
24 °C +/- 2 °C
pH:
Test start: 7.9 - 8.1
Test end: 7.2 - 8.8
Nominal and measured concentrations:
nominal concentrations: 0.7, 1.5, 3, 6, 12 and 24 mg/L based on nominal anionic surfactants concentration
Measured concentrations (initial.): 0.93, 1.8, 3.2, 6.3, 13.8 and 24.5 mg/L based on measured anionic surfactants concentration (MBAS)
Details on test conditions:
TEST SYSTEM
- Test vessel:Erlenmeyer flasks
- Material, fill volume: glass
- Aeration: yes
- Initial cells density: 20000 cells/mL
- Control end cells density: 730000 cells/mL
- No. of vessels per concentration (replicates): 5
- No. of vessels per control (replicates): 8


GROWTH MEDIUM
- Standard medium used: yes according to EU-Guideline 92/69/EWG
Composition of medium according to guideline (final concentration of nutrients in the test solution)
Macronutrients
NH4Cl: 15 mg/l
MgCl2.6H2O: 12mg/l
CaCl2.2H2O: 18 mg/l
MgSO4.7H2O: 15 mg/l
KH2PO4: 1.6 mg/l

Fe -EDTA
FeCl3.6H2O: 0.08 mg/l
Na2EDTA.2H2O: 0.1 mg/l

Trace elements
H3BO3: 0,185 mg/l
MnCl2.4H2O: 0,415 mg/l
ZnCl2: 3 x 10-3 mg/l
CoCl2.6H2O: 1.5 x 10-3 mg/l
CuCl2.2H2O: 10-5 mg/l
Na2MoO4.2H2O: 7 x 10-3mg/l
Stock solution 4: NaHCO3
NaHCO3: 50 mg/l


OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Photoperiod: continuous illumination
- Light intensity and quality: approximately 8000 Lux (white)


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: spectrophotometer (wave length: 685 nm), determined at 0, 24, 48 and 72 h



Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
27.7 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: corresponds to a measured concentration of 27 mg/L based on anionic surfactants concentration (MBAS)
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
4.4 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: corresponds to a measured concentration of 4.3 mg/L based on anionic surfactants concentration (MBAS)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.95 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: corresponds to a measured concentration of 0.93 mg/L based on anionic surfactants concentration (MBAS)
Details on results:
- Exponential growth in the control (for algal test): yes
Reported statistics and error estimates:
The EC-values were determined by probit transformation of percental growth inhibition values using regression analysis. Based on these values an additional probit analysis according to Cavalli-Sforza were conducted.

Table 1: Results

Concentration [mg/L]

Cell density (x 104cells/mL)

Test duration

0 h

24 h

48 h

72 h

Control

2

5

18

73

0.93

2

5

19

73

1.8

2

5

17

58

3.2

2

5

16

59

6.3

2

5

16

53

13.8

2

4

12

19

24.5

2

4

13

14

Table 2: Measured concentrations based on methylene blue active substance concentration (MBAS)

Nominal concentration [mg/L] Measured concentration [mg/L]  
0 h 48 h
stock solution 105  -- 
control 0.3* 0.12*
0.7 0.93 0.94
1.5 1.75 1.85
3 3.15 3.4
6 6.25 7.1
12 13.8 13
24 24.5 24.8

* concentrations < 0.5 mg/L are not considered

Table 3: Validity criteria for OECD 201.

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.

37

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%

29.2

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%.

1.27

Yes

Validity criteria fulfilled:
yes
Remarks:
The validity crietria according to OECD 201 are fulfilled (for details see table 3 under "any information on results")

Description of key information

In the key-study an EC50-value for algae (growth rate) of 27.7 mg a.s./L and a NOEC of 0.95 mg a.s./L could be determined after 72 h of exposure. 

Key value for chemical safety assessment

Additional information

A reliable algae studies is available for AES (C12-14, 1-2.5EO) Na (CAS 68891-38-3). The study was conducted according to the guidelines OECD 201 and EG Guideline 92/69 C.1 with the green algae Scenedesmus subspicatus as test organism. The algae were exposed to nominal test substance concentrations of 0.7, 1.5, 3, 6, 12 and 24 mg/L based on nominal anionic surfactants concentration. The test concentrations were analytically verified using the MBAS assay (methylene blue active substances assay) as analytic test method. The measured concentrations were 0.93, 1.8, 3.2, 6.3, 13.8 and 24.5 mg/L based on measured anionic surfactants concentration (MBAS) at test start. The GLP-study resulted in an ErC50 (72 h) of 27.7 mg a.s./L (corresponding to 27. mg/L based on measured anionic surfactants concentration (MBAS)). The test was conducted with standard medium according to EU-Guideline 92/69/EWG.