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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:
23 Oct - 26 Oct 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
1984
GLP compliance:
yes
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: in a refridgerator
- Stability under test conditions: Test material was stable. Stability was confirmed before and after exposure by using IR spectrum.
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.005, 0.016, 0.05, 0.16, 0.5, 1.6, 5.0 mg/L
- Sampling method: Before exposure, the test water was sampled from the vessel which was separately prepared for analysis. At the end of the exposure, the same volume was sampled from three vessels and mixed.
- Sample storage conditions before analysis: Not stored
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 50 mg/L stock solution was prepared and sterilized. This stock solution was diluted in next stock solutions (0.05, 0.16, 0.5, 1.6, 5, 16, and 50 mg/L). 10 mL of these stock solutions were mixed with 90 mL of test medium.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: green algae
- Source (laboratory, culture collection): American Type Culture Collection
- Age of inoculum (at test initiation): 5 d

ACCLIMATION
- Acclimation period: 96 h
- Culturing media and conditions (same as test or not): Alga was cultured in C medium in the same condition
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
23.0 °C
pH:
8.7 - 10.1 (control), 7.8 - 8.9 (test item)
Nominal and measured concentrations:
0 (control), 0.005, 0.016, 0.05, 0.16, 0.5, 1.6, and 5.0 mg/L (nominal)
Details on test conditions:
TEST SYSTEM
- Test vessel: Erlenmeyer flask
- Fill volume: 300 mL
- Initial cells density: 1 x 10^4 cells/mL
- Control end cells density: 55.42 x 10^4 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to guideline
- Culture medium different from test medium: yes
- Intervals of water quality measurement: pH was measured at the beginning and at the end of the test

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Light intensity and quality: 4000 - 5000 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: by counting chamber at the beginning of the incubation and every 24 h

TEST CONCENTRATIONS
- Range finding study was performed but the detailed information is not available.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
0.039 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.005 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
0.225 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.05 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
- Exponential growth in the control (for algal test): yes, increase by a factor of 55
Results with reference substance (positive control):
- Results with reference substance valid: yes
- EbC50 (72 h): 0.82 mg/L (0.50 - 1.02 mg/L, n =8)
Reported statistics and error estimates:
Dunett test

Table 1. Measured concentrations of the test substance in test water

Nominal concentration (mg/L)

Measured concentration (mg/L)

0 hour

Percent of nominal

72 hours

Percent of nominal

Control

< 0.003

-

< 0.003

-

0.005

0.006

120

0.005

100

0.016

0.016

100

0.013

81

0.05

0.045

90

0.044

88

0.16

0.147

92

0.121

76

0.5

0.509

102

0.450

90

1.6

1.60

100

1.58

99

5.0

5.37

107

5.07

101

 

Table 2. Percent growth inhibition of Selenastrum capricornutum

Nominal concentration

Area under the growth curves

Growth rate

Inhibition (%)

IA (0-72 h)

Inhibition (%)

Im (24-48 h)

Inhibition (%)

Im (24-72 h)

Control

-

-

-

0.005

22.61

7.59

3.94

0.016

34.15*

20.55

21.02

0.05

44.51*

-22.69

-10.27

0.16

77.08*

57.84*

43.84*

0.50

89.88*

53.30

78.96*

1.6

94.39*

94.54*

85.01*

5.0

97.55**

81.15**

81.09**

Values are the percent inhibition relative to the control.

* Significant difference p<0.05, ** Significant difference p<0.01

Table 3. Calculated EC50 and NOEC

Areas under growth curve (0 -72 h)

EbC50 (0-72) (mg/L)

95% confidence limits (mg/L)

NOECb (0 - 72)

(mg/L)

0.039

0.032 – 0.047

0.005

Growth rates (24 – 48 h)

ErC50 (24-48) (mg/L)

95% confidence limits (mg/L)

NOECr (24 - 48)

(mg/L)

0.199

0.166 – 0.239

0.05

Growth rates (24 – 72 h)

ErC50 (24-72) (mg/L)

95% confidence limits (mg/L)

NOECr (24 - 72)

(mg/L)

0.225

0.189 – 0.269

0.05

Description of key information

ErC50 (72 h) = 0.225 mg/L (nominal, OECD 201, P. subcapitata)

NOEC (72 h) = 0.05 mg/L (nominal, OECD 201, P. subcapitata)

Key value for chemical safety assessment

EC50 for freshwater algae:
0.225 mg/L
EC10 or NOEC for freshwater algae:
0.05 mg/L

Additional information

There is one GLP study available investigating the effects of the substance on the growth of freshwater microalgae according to the OECD guideline 201.

In a static test, P. subcapitata was exposed to nominal concentrations of 0.005, 0.016, 0.05, 0.16, 0.5, 1.6, and 5.0 mg/L test item for 72 h under controlled conditions. Negative and positive controls were run in parallel. Cell concentration was measured at the beginning of incubation and then every 24 h. Test item concentrations were analytically verified by HPLC at the beginning (0 h) and end (72 h) of exposure.

The measured concentrations were largely within ± 20% of the nominal concentrations, with exception of the 0.16 mg/L concentration level, which was 76% of nominal. The analytical results indicate that the substance was fully dissolved and stable over the exposure period. Effect concentrations were based on nominal concentrations. The calculated ErC50 was 0.225 mg/L (95% confidence interval 0.189 – 0.269 mg/L ) and the NOErC (72 h) was 0.05 mg/L.