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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:
weight of evidence
Study period:
10 Sep - 13 Sep 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes (incl. QA statement)
Remarks:
Ministerium für Arbeit, Integration und Soziales des Landes Nordrhein-Westfalen, Germany
Analytical monitoring:
yes
Details on sampling:
- Concentrations: WAF of nominal 100 mg/L and test culture
- Sampling method: Samples of test media were taken directly from the filtered WAF and from the test culture after addition of the algae at test start (0 h). At test end (72 h) the samples were taken from the replicates and pooled. Samplings of the 'OECD growth medium' were done by the staff of the analytical laboratory using a 'research 5000' pipette. The aqueous samples of 5.0 mL were pipetted directly into the extraction (centrifuge) tubes; the pipette tips were rinsed multiple and the surface was wiped with a lint free paper towel prior taking the analytical sub-sample. To assess possible wall effects on the glass surface of the test vessels (250 mL conical glass flasks), a sub-sample of the prepared “stock WAF” and of the control WAF were filtered directly into an extraction tube; the volume was determined by differential weighing.
- Sample storage conditions before analysis: The samples were processed and measured directly after sampling without any time delay.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Due to the very low water solubility of the test item a Water Accommodated Fraction (WAF) of the test item will be prepared according to the OECD guidance No. 23. Before weighing the test item, which was delivered in pellets, was crushed using a mortar. 200.0 mg test item was transferred into a 2 L sterile brown glass flask (acetone washed and sterilised) that has a bottom draining port. Sterile growth medium (2 L) and an acetone washed, sterilised star shaped stirring bar were added to the bottle and stirred slowly to avoid bubble and foam formation for about 66 h at room temperature (about 20 °C). Another flask was filled with growth medium only for the controls replicates and stirred in the same way. All work was done under sterile conditions. The test solution (WAF) was filtered by using a 0.2 µm filter (e.g. Sartolab 150v, PES-membrane, Sartorius Stedim Biotech S.A., Aubagne Cedex, France) to separate insoluble parts from the aqueous phase.
- Eluate: no
- Differential loading: yes
- Controls: yes, test medium control
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: green algae
- Strain: 61.81 SAG
- Source: SAG, Culture Collection of Algae at Pflanzenphysiologisches Institut of the University at Göttingen, Albrecht von Haller Institut, Göttingen, Germany
- Method of cultivation: The stock cultures are maintained fulfilling the criteria of the OECD guideline. Prior to testing a pre-culture was established in standard OECD growth medium to obtain exponentially-growing algae for the test.

ACCLIMATION
- Culturing media and conditions (same as test or not): same as test
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Test temperature:
20 °C
pH:
8.03 - 8.18 (control)
8.07 - 8.55 (100 mg/L WAF)
Nominal and measured concentrations:
nominal: 0, 100 mg/L (WAF)
measured: < LOQ, 0.63 - 0.82 µg/L (0 h), 0.48 - 4.92 µg/L (72 h)
Details on test conditions:
TEST SYSTEM
Test vessel:
- Type: covered with air-permeable silicone-sponge caps
- Material, size: glass, 250 mL
- Aeration: continuously shaken on an orbital laboratory shaker (100-150 rpm)
- Initial cells density: approx. 10000 cells/mL
- Control end cells density: 102118E+04 cells/mL
- No. of vessels per concentration (replicates): 8
- No. of vessels per control (replicates): 8

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: purified water processed using an ELGA "PURELAB" system

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: 7038 - 7399 lux (OSRAM "day light")

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: The cell concentrations were determined in the inoculum culture prior to the addition to the test vessels at test start and after 24, 48 and 72 h in the test cultures. The cell density was measured using an electronic particle counter (CASY 1 TT, Roche Diagnostics GmbH).

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1, 10 and 100 mg/L
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 1.465 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
>= 1.465 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: yield
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: yield
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 1.465 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
>= 1.465 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield
Details on results:
- Exponential growth in the control (for algal test): yes
Results with reference substance (positive control):
- Results with reference substance valid? yes
- EC50: ErC50: 2.91 mg/L (95% confidence limit: 2.35 - 3.36 mg/L)

Table 1: Cell number (x 10E+04 to equal cells/mL) dependent on loading of the test item and time (CV: coefficient of variation). Cell number at test start: 10000 cells/mL.

 

24 h

48 h

72 h

Loading [mg/L]

Control

100 mg/L

Control

100 mg/L

Control

100 mg/L

 

4,54

3,318

22,240

18,290

93,550

83,230

 

4,636

3,045

24,920

21,740

129,700

93,110

 

4,265

4,083

24,250

23,990

117,200

117,800

 

3,856

3,623

22,740

24,600

114,000

118,700

 

3,763

3,238

21,270

23,310

98,780

81,500

 

4,303

2,551

25,080

23,350

114,500

107,000

 

4,43

3,176

26,380

20,760

121,500

103,700

 

4,698

3,906

27,360

24,070

140,800

111,900

Replicates

8

8

8

8

8

8

Mean

4,311

3,368

24,280

22,514

116,254

102,118

Std.Dev.

0

0

2

2

15,31

14,67

CV

8,0

14,6

8,6

9,5

13,2

14,4

Validity criteria fulfilled:
yes
Conclusions:
Based on the present study performed with tetradecyl myristate (CAS No 3234-85-3), a constituent of the registration substance, the EL50 (72 h) values for both, growth rate and yield of P. subcapitata, were derived at > 100 mg/L (nominal) or > 1.456 µg/L (measured/arithm. mean).
Executive summary:

The toxicity of tetradecyl myristate (CAS No 3234-85-3), a constituent of the registration substance, to P. subcapitata was determined according to OECD guideline 201. The aim of the study was to assess potential effects on growth rate and yiled over a period of 72 h. The study was conducted under static conditions with an initial cell density of approx. 10000 cells/mL. Due to the very low water solubility of the test item a Water Accommodated Fraction (WAF) of the test item was prepared.

The nominal and measured concentrations were as follows:

nominal: 0, 100 mg/L (WAF)

measured: < LOQ, 0.63 - 0.82 µg/L (0 h), 0.48 - 4.92 µg/L (72 h).

The concentration (test start and end (72 h)) was analytical monitored by GC/EI-MS.

Results:

NOEL (72 h; growth rate): >/= 100 mg/L (nominal); >/= 1.465 µg/L (measured)

EL50 (72 h; growth rate): > 100 mg/L (nominal); 1.465 µg/L (measured)

NOEL (72 h; yield): >/= 100 mg/L (nominal); >/= 1.465 µg/L (measured)

EL50 (72 h; yield): > 100 mg/L (nominal); 1.465 µg/L (measured)

Description of key information

No effects up to the limit of water solubility based on available data (OECD 201).

Key value for chemical safety assessment

EC50 for freshwater algae:
100 mg/L

Additional information

Two GLP Guideline studies conducted according to OECD 201 are available investigating potential toxic effects of tetradecyl myristate (CAS 3234-85-3) and dodecyl oleate (CAS No 36078-10-1),

constituents of the registration substance,

to freshwater algae P. subcapitata and D. subspicatus. The algae were exposed to a Water Accommodated Fraction (WAF) of nominal 100 mg/L for 72 h.

1. Result of study with tetradecyl myristate:

NOEL (72 h; growth rate): >/= 100 mg/L (nominal); >/= 1.465 µg/L (measured)

EL50 (72 h; growth rate): > 100 mg/L (nominal); 1.465 µg/L (measured)

NOEL (72 h; yield): >/= 100 mg/L (nominal); >/= 1.465 µg/L (measured)

EL50 (72 h; yield): > 100 mg/L (nominal); 1.465 µg/L (measured)

2. Result with dodecyl oleate:

NOEL (72 h; growth rate): >/= 100 mg/L (nominal)

EL50 (72 h; growth rate): > 100 mg/L (nominal)

NOEL (72 h; yield): >/= 100 mg/L (nominal)

EL50 (72 h; yield): > 100 mg/L (nominal)

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