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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-04-20 till 2010-05-27
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline and GLP-compliant study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
yes
Remarks:
To avoid evaporation of the test item, the Erlenmeyer flasks were closed tightly with glass stoppers. Therefore the pH value decreased more than 1.5 pH units. This minor deviation is not considered to have any impact on the putcome of the study.
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Details on test material:
Partie number - Q101
Sample number - 1152
Purity - not stated
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not applicable

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Samples were taken for analysis from each test treatment and the control at the start of the study and at 24 hour intervals until the end of the study.

Test solutions

Vehicle:
no
Details on test solutions:
A stock solution was prepared daily,271.2 uL of the test substance was added to 2 L of dilution water. Undissolved particles were removed by filtration through a 7 - 12 um pore size filter paper. The pH of the stock solution was 8.0. The test media were prepared by addition of appropriate volumes of the stock solution to test vessels and dilution to 100 mL with water. Three replicates were prepared for each test treatment and control. Test vessels were filled to reduce loss of test material due to volatisation.

A volume of 0.667 mL of algal inoculum was added to each replicate vessel resulting in a final volume of 5000 cells/mL.

Test organisms

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
Desmodesmus subspicatus (formerly Scenedesmus subspicatus) Strain No. 86.81 SAG

Strain of the test species obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany).

Exponentially-growing stock cultures were maintained in the test facility under constant temperature conditions (21-24°C with a maximum fluctuation of +/- 2°C) at a light intensity in the range 60 – 120 µE. x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter). The growth medium (according to BRINGMANN & KÜHN (1977) was renewed once a week. Cell density measurements were made using a microcell- counter, Sysmex F300, Digitana.

Pre-cultures were set up three days before the start of a test. They were grown under identical exposure conditions as the stock cultures, except from the use of a different growth medium

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Post exposure observation period:
Not applicable

Test conditions

Hardness:
Not applicable
Test temperature:
Temperature range was 21 - 24°C
pH:
See table in the results section for details.
Dissolved oxygen:
Not applicable
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal exposure concentrations: 0 (control), 5.0, 10, 20, 40, 80 and 160 mg/L. Geometric mean measured concentrations were 0.422, 0.888, 1.464, 2.869, 5.855 and 17.182mg/L.
Details on test conditions:
At the average of the test solutions, a light intensity in the range 60 to 120 µE. x m-2 x s-1, or an equivalent range of 4000 to 8000 lx, was used. Light intensity was checked before start of the study.

Light chamber in which a temperature in the range 21°C to 24°C was maintained at +/- 2°C, and continuous uniform illumination was provided in the spectral range 400 to 700 nm. Temperature was measured and recorded daily in a water filled flask which was incubated under the same conditions as the test flasks.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
4.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 3.8 - 5.0
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
2.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 1.7 - 2.9
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
<= 0.422 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
2.9 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 2.7 - 3.2
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
1.6 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 1.3 - 1.9
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.422 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Results with reference substance (positive control):
Not applicable
Reported statistics and error estimates:
All calculations were perfofmed using the statistical programme ToxRatPro Version 2.10 (relaeased 20.02.2010). EC50 values were calcuated by probit analysis. NOEC and LOEC values were determined by multisampling comparison, using Williams Multiple Sequential t-Test.

Any other information on results incl. tables

 Measured 2,3-dichloro-1,3-butadiene concentrations

Test item
concentration
[m/L]

GC values
[mg/L]

0 h

24 h

48 h

72 h

Geometric mean

Control

< 0.629

< 0.629

< 0.629

< 0.629

-

5.0

0.793

0.399*

0.405*

0.247*

0.422

10

1.578

0.869

0.917

0.495*

0.888

20

2.694

1.406

1.398

0.868

1.462

40

5.199

2.765

2.900

1.625

2.869

80

10.562

5.540

5.872

3.420

5.855

160

29.598

16.587

15.603

11.378

17.182

*Values below the limit of quantification (0.629 mg/L) were calculated on basis of the mean recovery rate of the remaining concentrations at 24, 48 and 72 hour

pH values at start and end of the study

Test item
concentration
[mg/L]

pH values at 0 h and 72 h

0 h

72 h

Control

7.5

10.4

5.0

7.7

10.5

10

7.8

10.5

20

7.8

10.5

40

7.8

9.4

80

7.8

8.0

160

7.8

7.9

Cell numbers during the study

Mean measured concentrations (mg/L)

Mean cell number (initial value 0.5 X 104cells/mL)

24 hours

48 hours

72 hours

Control

2.0

7.7

20.0

0.422

2.0

8.0

19.0

0.888

2.0

7.7

18.0

1.464

1.7

6.3

18.7

2.869

1.0

3.0

10.7

5.855

1.0

1.0

1.3

17.182

1.0

1.0

1.0

 

 

Effects over the 72 hour exposure period:

Mean measured concentrations (mg/L)

72 h growth rate
Mean ± CV %

 

% inhibition compared to the control

72 h yield
Mean ± CV %

 

% inhibition compared to the control

Control

1.229 ±0.03

-

19.5 ± 2.0

-

0.422

1.212 ±0.02

1.3

18.5 ± 1.0

5.1

0.888

1.195 ±0

2.8

17.5 ± 0

10

1.464

1.206 ±0.03

1.8

18.2 ±1.5

6.8

2.869

1.202 ± 0.02

17

10.2 ± 0.58

48

5.855

0.308 ± 0.13

75

0.83 ± 0.58

96

17.182

0.231 ±0

81

0.5 ± 0

97

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
Based on mean measured concentrations the 72 hour ErC50 value for Scenedesmus subspicatus exposed to 2,3-dichlorobutadiene-1,3 was calculated to be 4.4 mg/L (with 95% confidence intervals of 3.8 - 5.0 mg/L), the corresponding no-observed effect concentration (NOEC) was 0.888 mg/L. The 72 hour EyC50 value was calculated to be 2.9 mg/L (with 95% confidence intervals of 2.7 – 3.2 mg/L), the corresponding NOEC was 0.422 mg/L.
Executive summary:

The alga Scenedesmus subspicatus were exposed to nominal 2,3-dichlorobutadiene-1,3 concentrations of 0 (control), 5.0, 10, 20, 40, 80 and 160 mg/L for 72 hours according to EU Method C.3 Freshwater Alga and Cyanobacteria, Growth Inhibition test (2009).  To reduce volatilisation of 2,3-dichlorobutadiene-1,3 the study was conducted in sealed test vessels.

Geometric mean measured concentrations were 0.422, 0.888, 1.464, 2.869, 5.855 and 17.182 mg/L in the nominal 5.0, 10, 20, 40, 80 and 160 mg/L treatments. Based on mean measured concentrations the 72 hour ErC50value was calculated to be 4.4 mg/L (with 95% confidence intervals of 3.8 - 5.0 mg/L), the corresponding no-observed effect concentration (NOEC) was0.888 mg/L. The 72 hour EyC50value was calculated to be 2.9 mg/L (with 95% confidence intervals of 2.7 – 3.2 mg/L), the corresponding NOEC was 0.422 mg/L.