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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:
27 November 2015 - 15 January 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Version / remarks:
2006
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Batch number - 1721964
Purity - 99.0%
Physical state - yellow liquid
Analytical monitoring:
yes
Details on sampling:
Samples were taken from the control and each test group from the bulk test preparation at 0 hours and from the pooled replicates at 72 hours for quantitative analysis. All samples were stored frozen prior to analysis. Duplicate samples were taken at 0 and 72 hours and stored frozen for further analysis if necessary.
Vehicle:
no
Details on test solutions:
A nominal amount of test item (100 mg) was dissolved in culture medium and the volume adjusted to 1 liter to give a 100 mg/L stock solution from which a series of dilutions was made to give further stock solutions of 10, 1.0 and 0.10 mg/L. An aliquot (900 mL) of each of the stock solutions was separately inoculated with algal suspension (7.9 mL) to give the required test concentrations of 0.10, 1.0, 10 and 100 mg/L.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
Strain CCAP 278/4, obtained from the Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Scottish Marine Institute, Oban, Argyll, Scotland.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Remarks on exposure duration:
Not applicable
Post exposure observation period:
Not applicable
Hardness:
Not applicable
Test temperature:
Temperatire was maintianed between 23 and 25°C
pH:
pH range at the start of the study was 7.8 - 8.3 and at the end of the 72 hour exposure period was 8.6 - 8.7.
Dissolved oxygen:
Not applicable
Salinity:
Not applicable
Conductivity:
Not applicable
Nominal and measured concentrations:
Nominal exposure concentrations were control (0), 1.0, 3.2, 10, 32 and 100 mg/L.
All measured concentrations were within ±20% of the nominal exposure concentration. Full details are presented in the results section.

Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL glass conical flasks
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: completely filled and sealed with ground glass stoppers

- Initial cells density: 5000 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes, but with additional sodium bicarbonate according to Herman et al (1990) to compensate for the restriction on gaseous exchange imposed by the use of sealed incubation vessels.


OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: approximately 7000 lux, warm white lighting, 380 - 730 nm

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]


TEST CONCENTRATIONS
- Range finding study - yes
- Test concentrations: 0.1, 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
A separate reference test was performed with potasssium dichromate.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
54 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
32 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
42 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
32 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Details on results:
The effects of 2-[[2-(Dimethylamino)ethyl]methylamino]ethanol [CAS# 2212-32-0] on the growth of the freshwater unicellular green alga Pseudokirchneriella subcapitata was determined in a test performed in accordance with OECD TG 201 (2006). The 72-hour ErC50 was 54mg/L and the EyC50 based on yield was 42 mg/L with 95% confidence limits of 39 to 46 mg/L. For both endpoints he corresponding 72 -hour NOEC was 32 mg/L.
Results with reference substance (positive control):
The 72 h ErC50 obtained with potassium dichromate was 1.5 mg/L (95% CL: 1.3 - 1.7 mg/L) and was within the normal response range for this substance.
Reported statistics and error estimates:
Determination of ECx values - For each individual test vessel (mean values for yield), percentage inhibition (arithmetic axis) was plotted against test concentration (logarithmic axis) and a line fitted by computerized interpolation using the Xlfit software package (IDBS). ECx values were then determined from the equation for the fitted line. Where appropriate 95% confidence limits for the EC50 values were calculated, using the simplified method of evaluating dose-effect experiments of Litchfield and Wilcoxon (1949).

One way analysis of variance incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf, 1981) and Dunnett's multiple comparison procedure for comparing several treatments with a control (Dunnett, 1955) was carried out on the growth rate and yield data after 72 hours for the control and all test concentrations to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package (SAS, 1999 - 2001).

Analytical results

Nominal exposure concentration (mg/)

Measured concentrations (mg/L)

0 hours (% of nominal)

72 hours (% of nominal)

Control

<LOQ

<LOQ

1.0

0.874 (87%)

0.888 (89%)

3.2

2.85 (89%)

2.81 (89%)

10

9.47 (95%)

9.44 (94%)

32

28.7 (90%)

29.4 (92%)

100

90.5 (90%)

92.7 (93%)

<LOQ = <Limit of Quantification = 0.0047 mg/L

Cell densities

Nominal exposure concentration (mg/)

Mean cell density (cells /mL)

24 hours

48 hours

72 hours

Control

1.70E+04

5.40 E+04

1.87E+05

1.0

1.88E+04

5.37+04

1.67E+05

3.2

1.44E+04

6.23+04

1.82E+05

10

2.23E+04

6.17+04

2.01E+05

32

1.93+04

5.63E+04

1.56E+05

100

8.53E+03

5.17E+03

6.27E+03

 

Inhibition of growth rate and yield

Nominal exposure concentration (mg/)

Growth rate (cells/mL/hour)

Yield (cells/mL)

0-72 h (SD)

% inhibition

0-72 h (SD)

% inhibition

Control

0.051 (0.002)

-

1.82E+05 (2.00E+04)

-

1.0

0.049 (0)

4

1.62E05 (2.37E+03)

11

3.2

0.050 (0.003)

3

1.77E+05 (3.25E+04)

3

10

0.051 (0.001)

+1

1.96E+05 (1.33E+04)

+8

32

0.048 (0.002)

7

1.51E+05 (2.50E+04)

17

100

0.002 (0.007)

96

1.27E+03 (2.66E+03)

99

 

Validity criteria fulfilled:
yes
Conclusions:
The effects of 2-[[2-(Dimethylamino)ethyl]methylamino]ethanol [CAS# 2212-32-0] on the growth of the freshwater unicellular green alga Pseudokirchneriella subcapitata was determined in a test performed in accordance with OECD TG 201 (2006). The 72-hour ErC50 was 54 mg/L and the EyC50 based on yield was 42 mg/L with 95% confidence limits of 39 to 46 mg/L. For both endpoints he corresponding 72 -hour NOEC was 32 mg/L.
Executive summary:

The effects of 2-[[2-(Dimethylamino)ethyl]methylamino]ethanol [CAS# 2212-32-0] on the growth of the freshwater unicellular green alga Pseudokirchneriella subcapitata were determined in a test performed in accordance with OECD TG 201 (2006). Algal cells were incubated for 72 hours under static conditions in vessels that were stoppered to limit volatilisation losses of the test substance and were exposed to nominal exposure concentrations of 0 (control), 1.0, 3.2, 10, 32 and 100 mg/L. Analysis of the test media at the start and end of the exposure periods demonstrated that the test material was stable for the duration of the exposure period.  The results of the study were therefore based on nominal exposure concentrations. The 72-hour ErC50 was 54 mg/L and the EyC50 based on yield was 42 mg/L with 95% confidence limits of 39 to 46 mg/L. For both endpoints he corresponding 72 -hour NOEC was 32 mg/L. The guideline validity criteria were fulfilled.

Description of key information

The effects of 2-[[2-(Dimethylamino)ethyl]methylamino]ethanol [CAS# 2212-32-0] on the growth of the freshwater unicellular green alga Pseudokirchneriella subcapitata was determined in a test performed in accordance with OECD TG 201 (2006). The 72-hour ErC50 was 54 mg/L and the EyC50 based on yield was 42 mg/L with 95% confidence limits of 39 to 46 mg/L. For both endpoints he corresponding 72 hour NOEC was 32 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:
54 mg/L
EC10 or NOEC for freshwater algae:
32 mg/L

Additional information

The effects of 2-[[2-(Dimethylamino)ethyl]methylamino]ethanol [CAS# 2212-32-0] on the growth of the freshwater unicellular green alga Pseudokirchneriella subcapitata were determined in a test performed in accordance with OECD TG 201 (2006). Algal cells were incubated for 72 hours under static conditions in vessels that were stoppered to limit volatilisation losses of the test substance and were exposed to nominal exposure concentrations of 0 (control), 1.0, 3.2, 10, 32 and 100 mg/L. Analysis of the test media at the start and end of the exposure periods demonstrated that the test material was stable for the duration of the exposure period.  The results of the study were therefore based on nominal exposure concentrations. The 72-hour ErC50 was 54mg/L and the EyC50 based on yield was 42 mg/L with 95% confidence limits of 39 to 46 mg/L. For both endpoints he corresponding 72 -hour NOEC was 32 mg/L. The guideline validity criteria were fulfilled.