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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:
2008-09-22 to 2008-09-25
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
Adopted 23 March 2006
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.3 (Algal Inhibition test)
Version / remarks:
December 1992
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: all concentrations after 24 and 72 h of exposure
- Sampling method: 2 mL were taken from the test vessels for analysis
- Sample storage conditions before analysis: not applicable
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Separate stock solutions were prepared. The solutions were stirred for 2 days and subsequent dispersions were then filtered through a 0.45 µm membrane filter (Schleicher & Schuell RC55).
- Eluate: no
- Differential loading: no
- Controls: yes, test medium control
Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: green algae
- Strain: NIVA CHL 1
- Source (laboratory, culture collection): in-house laboratory culture
- Method of cultivation: Cultivated in M1 medium

ACCLIMATION
- Acclimation period: 3 or 4 days before test start cells from the algal stock culture were inoculated in culture medium (M2).
- Culturing media and conditions (same as test or not): same as in test
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
22.5 - 23.4 °C
pH:
7.7 - 8.0 (test start)
7.7 - 7.9 (test end)
Nominal and measured concentrations:
Nominal tests substance concentrations: 2.2, 4.6, 10, 22, 46 and 100 mg/L
Time weighted average (TWA): 0.007, 0.065, 0.17, 0.5, 1.3, 3.5 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: glass vessels (nominal volume 100 mL)
- Fill volume: 50 mL
- Headspace: 50 mL
- Aeration: no
- Initial cells density: 10000 cells/mL
- Control end cells density: 197.67 E+04 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: M2 medium according to guideline
- Intervals of water quality measurement: pH at test start and test end, temperatue continously

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continous illumination
- Light intensity and quality: 79 - 86 µE/m²s (cool white)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: spectrophotometer after 24, 48, and 72 h

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
- Range finding study: Comined limit/range-finding
- Test concentrations: 1.0, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: LC50 (72 h) >= 10 <= 100 mg/L
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
0.66 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: confidence interval: 0.36 to 1.2 mg/L
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.065 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
1.8 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: confidence interval: 1.0 to 3.3 mg/L
Details on results:
- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape: no
Results with reference substance (positive control):
.- Results with reference substance valid? Yes
- EC50 (0-72 h): 1.7 mg/L with a 95% confidence interval ranging from 1.1 to 2.7 mg/L.
Reported statistics and error estimates:
An effect was considered to be significant if statistical analysis of the data obtained for the test concentrations compared with those obtained in the negative control revealed significant reduction of growth rate or inhibition of yield (ANOVA, Tukey test, Bonferroni t-test, TOXSTAT Release 3.5, 1996, D.D. Gulley, AM. Boelter, H.L. Bergman). Calculation of the EC50 and EC10 values was based on log-linear regression analysis of the percentages of growth rate reduction and the percentages of yield inhibition versus the logarithms of the corresponding average exposure concentrations of the test substance.
Validity criteria fulfilled:
yes
Conclusions:
The study resulted in an ErC50 (72 h) of 1.8 mg/L and an ErC10 (72 h) of 0.66 mg/L based on TWA measured concentrations.

Description of key information

ErC50 (72 h) 1.8 mg/L (TWA, OECD 201)
ErC10 (72 h) 0.66 mg/L (TWA, OECD 201)

Key value for chemical safety assessment

EC50 for freshwater algae:
1.8 mg/L
EC10 or NOEC for freshwater algae:
0.66 mg/L

Additional information

The effect on inhibition of the growth rate of green algae was investigated following OECD guideline 201 and in compliance with GLP. The green algae Pseudokirchneriella subcapitata was exposed to nominal tests substance concentrations of 2.2, 4.6, 10, 22, 46 and 100 mg/L. Analytical dose verification performed at 24 and 72 h determined measured concentrations clearly below nominal concentrations and decreased during the test period. Consequently, time weighted average (TWA) exposure concentrations were calculated. The ErC50 (72 h) based on TWA concentrations was 1.8 mg/L. The ErC10 (72 h) was 0.66 mg/L.