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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:
1998
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
(1984)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material: not reported
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.63, 1.3, 2.5 and 5.0 mg/L (nominal), measured: 0.56, 1.2, 2.4 and 4.8 mg/L
Vehicle:
yes
Details on test solutions:
2,2'-methylenebis(6-tert-butyl-4-methylphenol) was mixed with DMF/Castor oil HCO-40 (3/1(w/w)) and dissolved into test medium to prepare the nominal concentrations described above (No precipitations as well as emulsions were observed in all the exposure levels). To set the
dispersant concentrations equal (i.e. 100 µL/L) between the nominal concentrations, DMF/Castor oil HCO-40 solution was also used for the preparations. Due to low water solubility of 2,2'-methylenebis(6-tert-butyl-4-methylphenol), homogenous solution could be attained only at (and less than) the nominal concentration of 5.0 mg/L with the maximum allowable dispersant concentration of 100 µL/L.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
Green algae (Selenastrum capricornutum), the strain ATCC22662 obtained from American Type Culture Collection, and subcultured at the testing facility.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Remarks on exposure duration:
no remarks
Post exposure observation period:
not reported
Hardness:
not data
Test temperature:
23.5 to 23.7 °C measured at each vessel at the beginning and the end of the test.
pH:
7.6 to 7.7 measured at each vessel at the beginning of the test.
7.4 - 7.5 measured at each vessel at the end of the test.
Dissolved oxygen:
no data
Salinity:
no data
Nominal and measured concentrations:
Nominal Concentrations: 0.63, 1.3, 2.5, 5.0 mg/L, solvent control and control
Measured concentrations: 0.56, 1.2, 2.4 and 4.8 mg/L at the beginning (0h) of the test.
Measured concentrations: 0.47, 0.91, 2.1 and 4.2 mg/L at the end (72h) of the test.
Details on test conditions:
TEST SYSTEM
- Test vessel: 500 mL glas Erlenmeyer flask (silicon plugs breathable)
- Test volume: 100 mL
- Initial cell density: 10000 cells/mL
-Light conditions: 4000-5000 lx, continuous
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 1
- No. of vessels per vehicle control (replicates): 1

GROWTH MEDIUM
- OECD medium used: yes

- Determination of cell concentrations: ISOTON II from Beckman Coulter, Inc.

- Range finding study: 0.5 and 5.0 mg/L
- Test concentrations: 0.63, 1.3, 2.5 and 5.0 mg/L
Reference substance (positive control):
yes
Remarks:
Potassium dichromate, lot No.ACQ2610 from Wako Pure Chemical Industries, Ltd
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: actual concentration measured and greater than 80% of the nominal
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.63 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: actual concentration measured and greater than 80% of the nominal
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: actual concentration measured and greater than 80% of the nominal
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
1.3 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: actual concentration measured and greater than 80% of the nominal
Details on results:
The determined 72 hours EC50 values were >5.0 mg/L based on the nominal concentrations. The NOEC´s for biomass and growth rate were evaluated to be 0.63 mg/L and 1.3 mg/L, respectively.
Results with reference substance (positive control):
The 72 h-EC50 value (Biomass) determined was 0.44 mg/L.
Reported statistics and error estimates:
The NOEC’s described above were defined as the maximum nominal concentrations at which biomass and growth rate were not significantly different from the control (p = 5%).

Cell density measured during the experiment:

 

NominalConcentration (mg/L)

Mean Cell Density (x104cells/mL) +/- SD

0 h

24 h

48 h

72 h

Control

I.+/-0

7.67 +/-0.32

45.9 +/-0.9

278 +/-7

Solvent Control

II. +/-0

7.81 +/-0.21

44.1 +/-1.7

280 +/-26

0.63

III. +/-0

7.27 +/-0.56

42.9 +/-1.2

271 +/-3

1.3

IV. +/-0

7.32 +/-0.32

42.4 +/-0.3

242 +/-7

2.5

V. +/-0

7.52 +/-0.25

41.0 +/-1.7

226 +/-12

5.0

1.00 +/-0

7.45 +/-0.14

37.6 +/-1.5

185 +/-8

 

Growth rate curve (Area under the curve and the slope evaluated)

 

NominalConcentration (mg/L)

0 – 72 h

24 – 48 h

24 – 72 h

Mean Area

Inhibition %

Mean Rate

Inhibition %

Mean Rate

Inhibition %

Control

45569440

0

0.0745

0

0.0748

0

Solvent Control

45426080

0.3

0.0721

3.2

0.0745

0.4

0.63

43989280

3.5

0.0740

0.7

0.0754

-0.9

1.3

40351360

11.5

0.0732

1.7

0.0729

2.5

2.5

38121920

16.3

0.0706

5.3

0.0708

5.3

5.0

32365760

29.0

0.0674

9.5

0.0669

10.6

 

Solvent Control: DMF/HCO-40=3/1(100μL/L)

Validity criteria fulfilled:
yes
Remarks:
The test was valid since the cell density in control increased by 278 times at the termination of experiment.
Conclusions:
The determined 48 h-EC50 value was more than 5.0 mg/L (for biomass and growth rate) based on the nominal concentrations. The NOEC value for biomass (0-72 h) was evaluated to be 0.63 mg/L. The NOEC’s for growth rate were 2.5 mg/L (24-48 h) and 1.3 mg/L (24-72 h).
Executive summary:

In order to test acute toxicity to algae, Selenastrum capricornutum, of 2,2'-methylenebis(6 -tert-butyl-4 -methylphenol) was exposed to the test solution of four nominal concentrations (0.63, 1.3, 2.5 and 5.0 mg/L) of 2,2'-methylenebis(6 -tert-butyl-4 -methylphenol), solvent control and control for a period of 72 hours under static conditions. The cell densities were measured at 24 hour intervals. Inhibition of the algal population was measured as reduction in growth rate (index r), relative to control cultures grown under identical conditions. The measured concentrations confirmed that deviation from the nominal concentrations was less than 20 %( measured concentrations were in the range of 70 - 84 % of the nominal concentrations.

Due to low water solubility of the substance, maximum dispersable concentration was limited to be 5.0 mg/L (nominal).

The determined 48 h-EC50 value was more than 5.0 mg/L (for biomass and growth rate) based on the nominal concentrations. The NOEC value for biomass (0-72 h) was evaluated to be 0.63 mg/L. The NOEC’s for growth rate were 2.5 mg/L (24-48 h) and 1.3 mg/L (24-72 h).

Experimental designs and results were well documented and prescribed conditions in the guideline were well satisfies.

Description of key information

The 48 hr-EC50 was > 5.0 mg/L (for biomass and growth rate) based on nominal concentrations.
The NOEC value for biomass (0-72 hr) was evaluated to be 0.63 mg/L, while the NOEC values for growth rate was 1.3 mg/L. As the reported NOECs are above the water solubility no PNEC can be derived based on these results.

Key value for chemical safety assessment

Additional information

The above study is assessed in the OECD SIDS Report (2001) and the PBT Report (2009) of the European Chemical Bureau (ECB).

However the relevant test was performed with a solubilizer in order to overcome the low water solubility of 0.007 mg/L. As a consequence, no effects have been observed up to the limit of the water solubility.