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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:
From 2000-02-04 to 2000-05-29
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
This study was performed according to international guidelines with GLP statement. Validity criteria were not fulfilled: the mean CV for section-by-section specific growth rates in the cultures exceed 35% (41.52%). Furthermore, the pH of the control medium increase by more than 1.5 units during the test. These deviations are not critical and show only that the results should be taken with care.
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
. The pH of the control medium increase by more than 1.5 units during the test.
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
yes
Remarks:
. The pH of the control medium increase by more than 1.5 units during the test.
Principles of method if other than guideline:
Not applicable.
GLP compliance:
yes (incl. certificate)
Remarks:
1999-09-22
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.01, 0.1, 1 mg/L
- Sampling method: Three groups of 2 replicate flasks were exposed to nominal concentrations of the test substance under the same conditions of light and temperature. AT T0, T24, T48, T72, and T96 hours, cell numbers were determined using Mallassez cell counter.
- Sample storage conditions before analysis: no data
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Test solutions were prepared by further dilution of the stock solution with water to provide a geometric series of concentrations: 0, 0.01, 0.1, 1, 10 mg/L. Glass test vessels containing algae and test concentrations were filled directly from the test solution containers immediately after preparation and remained unchanged throughout the study.
- Eluate: Water
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: green algae
- Strain: CCAP 276/20
- Source (laboratory, culture collection): Culture Collection of Algae and Protozoa (Cumbria, UK)
- Age of inoculum (at test initiation): no data
- Method of cultivation: a primary stock solution is made up every month from which a final culture medium is prepared. This solution is autoclaved at which point it has a stock life of one week.

ACCLIMATION
- Acclimation period: algae are cultivated for 7 days before harvesting for use in a new culture.
- Culturing media and conditions (same as test or not): yes
- Any deformed or abnormal cells observed: none
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
no data
Hardness:
34 +/- 17 mg/L as CaCO3
Test temperature:
23.4-23.6°C
pH:
At 0 mg/L (control): T0 = 7.4; T72 = 8.49 and T96 = 9.04.
At 10 mg/L (nominal): T0 = 7.39; T72 = 8.25 and T96 = 8.99.
The pH of the control medium increase by more than 1.5 units during the test.
Dissolved oxygen:
no data
Salinity:
not applicable.
Nominal and measured concentrations:
- nominal conc.: 0, 0.01, 0.1, 1, and 10 mg/L
- measured conc.: See table 6.1.5/3 in "Any other information on results incl. tables".
Details on test conditions:
TEST SYSTEM
- Test vessel: conical flasks
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 250 mL
- Aeration: no forced aeration but the cultures were constantly agitated.
- Initial cells density: 1 x 10^4 cell/mL
- Control end cells density: 226.83 x 10^4 cell/mL
- No. of organisms per vessel:
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: none
- Photoperiod: continuous
- Light intensity and quality: 6600-7320 lux
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 4.19 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
and biomass
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 3.73 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
and biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
>= 4.19 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
and biomass
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
>= 3.73 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
and biomass
Details on results:
See tables in "Any other information on results incl. tables".
Results with reference substance (positive control):
not applicable
Reported statistics and error estimates:
no data

Table 6.1.5/1: number of cells at each observation time and calculation of the area under the growth curve (biomass) at each concentration after 72h (10^4 cells/mL x hour).

 

 

Total number of cells

Growth

% inhibition

Nominal concentration (mg/L)

Rep.

At 24 h

At 48 h

At 72 h

0 to 24 h

24 to 48 h

48 to 72 h

0 to 72 h

Average (0-72 h)

 

0

1

1.40

8.30

53.70

4.80

92.40

720.00

817.20

933.20

 

2

2.50

10.60

59.00

18.00

133.20

811.20

962.40

 

 

3

1.30

9.60

64.00

3.60

106.80

859.20

969.60

 

 

4

1.40

8.20

58.10

4.80

91.20

771.60

867.60

 

 

5

2.10

10.00

67.00

13.20

121.20

900.00

1034.40

 

 

6

2.80

10.10

58.20

21.60

130.80

795.60

948.00

 

 

0.01

1

2.00

10.80

72.10

12.00

129.60

970.80

112.40

996.00

-6.73

2

1.80

9.70

55.30

9.60

114.00

756.00

879.60

 

 

0.1

1

2.40

11.00

67.20

16.80

136.80

914.40

1068.00

1023.60

-9.69

2

1.70

10.10

63.00

8.40

117.60

853.20

979.20

 

 

1

1

1.60

9.40

60.00

7.20

108.00

808.80

924.00

1015.20

-8.79

2

2.20

11.20

70.40

14.40

136.80

955.20

1106.40

 

 

10

1

2.20

9.40

50.90

14.40

115.20

699.60

829.20

840.00

9.99

2

1.80

8.20

50.70

9.60

96.00

682.80

788.40

 

 

3

2.10

10.20

55.60

13.20

123.60

765.60

902.40

 

 

Table 6.1.5/2: number of cells at each observation time and calculation of the area under the growth curve (biomass) at each concentration after 96h (10^4 cells/mL x hour).

 

 

Total number of cells

Growth

% inhibition

Nominal concentration (mg/L)

Rep.

At 24 h

At 48 h

At 72 h

At 96 h

0 to 24 h

24 to 48 h

48 to 72 h

72 to 96 h

0 to 96 h

Average (0-96 h)

 

0

1

1.40

8.30

53.70

202.0

4.80

92.40

720.00

3044.40

3861.60

4351.20

 

2

2.50

10.60

59.00

245.0

18.00

133.20

811.20

3624.00

4586.40

 

 

3

1.30

9.60

64.00

199.0

3.60

106.80

859.20

3132.00

4101.60

 

 

4

1.40

8.20

58.10

225.0

4.80

91.20

771.60

3373.20

4240.80

 

 

5

2.10

10.00

67.00

243.0

13.20

121.20

900.00

3696.00

4730.40

 

 

6

2.80

10.10

58.20

247.0

21.60

130.80

795.60

3638.40

4586.40

 

 

0.01

1

2.00

10.80

72.10

256.0

12.00

129.60

970.80

3913.20

5025.60

4850.40

-11.47

2

1.80

9.70

55.30

263.0

9.60

114.00

756.00

3795.60

4675.20

 

 

0.1

1

2.40

11.00

67.20

296.0

16.80

136.80

914.40

4334.40

5402.40

5152.80

-18.42

2

1.70

10.10

63.00

266.0

8.40

117.60

853.20

3924.00

4903.20

 

 

1

1

1.60

9.40

60.00

248.0

7.20

108.00

808.80

3672.00

4596.00

4539.60

-4.33

2

2.20

11.20

70.40

213.0

14.40

136.80

955.20

3376.80

4483.20

 

 

10

1

2.20

9.40

50.90

205.0

14.40

115.20

699.60

3046.80

3876.00

3860.80

11.27

2

1.80

8.20

50.70

172.0

9.60

96.00

682.80

2648.40

3436.80

 

 

3

2.10

10.20

55.60

227.0

13.20

123.60

765.60

3367.20

4269.60

 

 

Table 6.1.5/3: Analytical measurements

Nominal (mg/L)

Measured (mg/L)

Geometric mean (mg/L), without algae

0h, without algae

24h, with algae

24h, without algae

48h, with algae

48h, without algae

72h, with algae

72h, without algae

96h, with algae

96h, without algae

0-72h

0-96h

10

4.74

3.62

4.66

2.56

3.88

2.15

3.60

2.04

2.35

4.19

3.73

Validity criteria fulfilled:
no
Remarks:
The mean CV for section-by-section specific growth rates in the cultures exceed 35% (41.52%)
Conclusions:
Under experimental conditions, the following results were obtained:
- 72h-EC50 > 4.19 mg/L (measured) based on the specific growth rate and biomass data.
- 72h-NOEC >= 4.19 mg/L (measured) based on the specific growth rate and biomass data.
- 96h-EC50 > 3.73 mg/L (measured) based on the specific growth rate and biomass data.
- 96h-NOEC >= 3.73 mg/L (measured) based on the specific growth rate and biomass data.
Executive summary:

The experiment was conducted to assess the effects of the test substance on algae, Scenedesmus subspicatus, using a 96 -h static test according to international guidelines, under GLP.

Three groups of 2 replicates flasks were exposed to nominal concentrations of 0, 0.01, 0.1, 1, and 10 mg/L of the test substance under the same conditions of light and temperature.

Concentrations of the test substance after 96 -h, corresponded to 43 -50% of the initial value. Furthermore, measured concentrations in the solution without algae were systematically higher than those in the solution with algae. This indicates that the test substance may be adhering to the surface of the algae of being degraded by it. According, all effects are based on geometric means of measured concentrations on the solution at 10 mL (nominal) without algae, calculated as 4.19 mg/L for the 0 -72 hour period and 3.73 mg/L for 0 -96 hour period.

Under experimental conditions, the following results were obtained:

- 72h-EC50 > 4.19 mg/L (measured) based on the specific growth rate and biomass data.

- 72h-NOEC >= 4.19 mg/L (measured) based on the specific growth rate and biomass data.

- 96h-EC50 > 3.73 mg/L (measured) based on the specific growth rate and biomass data.

- 96h-NOEC >= 3.73 mg/L (measured) based on the specific growth rate and biomass data.

Description of key information

OECD Guideline 201, EU Method C.3, GLP, key study, validity 2:

96h-ErC50 and NOECr (Scenedesmus subspicatus) were >= 3.73 mg/L, based on geometric mean measured concentrations.

Key value for chemical safety assessment

Additional information

One key study, performed by CIT laboratory, according to international guidelines with GLP standards is available on the growth of the green alga Scenedesmus subspicatus, using a 96h static test.

Three groups of 2 replicates flasks were exposed to nominal concentrations of 0, 0.01, 0.1, 1, and 10 mg/L of the test substance. Concentrations of the test substance after 96 -h, corresponded to 43 -50% of the initial value. Furthermore, measured concentrations in the solution without algae were systematically higher than those in the solution with algae. This indicates that the test substance may be adhering to the surface of the algae of being degraded by it. According, all effects are based on geometric means of measured concentrations on the solution at 10 mL (nominal) without algae, calculated as 4.19 mg/L for the 0 -72 hour period and 3.73 mg/L for 0 -96 hour period.

Under experimental conditions, the following results were obtained:

- 72h-EC50 > 4.19 mg/L (measured) based on the specific growth rate and biomass data.

- 72h-NOEC >= 4.19 mg/L (measured) based on the specific growth rate and biomass data.

- 96h-EC50 > 3.73 mg/L (measured) based on the specific growth rate and biomass data.

- 96h-NOEC >= 3.73 mg/L (measured) based on the specific growth rate and biomass data.