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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
Reliability:
1 (reliable without restriction)
Reference:
Composition 0
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes (incl. certificate)
Test material information:
Composition 1
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:
Culture Medium
NaNO3
25.5 mg/L
MgCl2.6H2O
12.16 mg/L
CaCl2.2H2O
4.41 mg/L
MgSO4.7H2O
14.6 mg/L
K2HPO4
1.044 mg/L
NaHCO3
15.0 mg/L
H3BO3
0.186 mg/L
MnCl2.4H2O
0.415 mg/L
ZnCl2
0.00327 mg/L
FeCl3.6H2O
0.160 mg/L
CoCl2.6H2O
0.00143 mg/L
Na2MoO4.2H2O
0.00726 mg/L
CuCl2.2H2O
0.000012 mg/L
Na2EDTA.2H2O
0.30 mg/L

The culture medium was prepared using reverse osmosis purified deionized water* and the pH adjusted to 7.5 with 0.1N NaOH or HCl.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
The test was carried out using Pseudokirchneriella subcapitata strain CCAP 278/4. Liquid cultures of Pseudokirchneriella subcapitata were obtained from the Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Scottish Marine Institute, Oban, Argyll, Scotland. Master cultures were maintained in the laboratory by the periodic replenishment of culture medium.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
See section Details on test solutions
Test temperature:
24 C
pH:
7.5
Nominal and measured concentrations:
Measured concentrations were within the range of 92-99% of nominal concentration.
Details on test conditions:
In the definitive test 250 mL glass conical flasks were used. Six flasks each containing 100 mL of test preparation were used for the control and three flasks each containing 100 mL were used for each treatment group.
The control group was maintained under identical conditions but not exposed to the test item.
Pre-culture conditions gave an algal suspension in log phase growth characterized by a cell density of 3.15 x 105 cells per mL. Inoculation of 500 mL of test medium with 7.9 mL of this algal suspension gave an initial nominal cell density of 5.00 x 103 cells per mL and had no significant dilution effect on the final test concentration.
The flasks were plugged with polyurethane foam bungs and incubated (INFORS Multitron Version 2 incubator) at 24 ± 1 °C under continuous illumination (intensity approximately 7000 lux) provided by warm white lighting (380 – 730 nm) and constantly shaken at approximately 150 rpm for 72 hours.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate.
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
53 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
59 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
70 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
42 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
47 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
28 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Details on results:
Range-finding Test
The cell densities and percentage inhibition of growth values results showed no significant effect on growth at the test concentrations of 0.10, 1.0, 10 and 100 mg/L.
Based on this information a single test concentration of six replicates, of 100 mg/L was selected for the initial experiments. This experimental design conforms to a "limit test" to confirm that at the maximum test concentration given in the OECD/EEC Test Guidelines no effect on growth was observed.
Chemical analysis of the 100 mg/L test preparations at 0 and 72 hours showed near nominal concentrations were obtained indicating that the test item was stable under test conditions.
Initial Experiments
Two initial experiments were conducted at a concentration of 100 mg/L.
In both instances significant inhibition of growth occurred and as such it was considered necessary to conduct the definitive test using a range of test concentrations to ensure that a No Observed Effect Concentration (NOEC) value was obtained.
Definitive Test
Following a preliminary range-finding test and initial experiments, Pseudokirchneriella subcapitata was exposed to an aqueous solution of the test item at concentrations of 6.25, 12.5, 25, 50 and 100 mg/L (three replicate flasks per concentration) for 72 hours, under constant illumination and shaking at a temperature of 24 ± 1 °C.
The inhibitory effects observed at 100 mg/L in the definitive test were more significant than those seen in the range-finding test and initial experiments. Examination of the data could find no obvious cause for this. Not only do the results of the chemical analyses indicate that the test system was correctly dosed, the corresponding chromatography matches that of the analytical standards suggesting there were no additional peaks attributable to contamination.
Growth rate (r) and yield (y) data of Pseudokirchneriella subcapitata (CCAP 278/4) were affected by the presence of the test item over the 72-Hour exposure period.
Accordingly the presented effect concentrations were calculated.
All test and control cultures were inspected microscopically at 72 hours. After 72 hours there were no abnormalities detected in the control or test cultures at 6.25, 12.5, 25 and 50 mg/L, however no intact cells were observed to be present in the test cultures at 100 mg/L.
Results with reference substance (positive control):
A positive control used potassium dichromate as the reference item at concentrations of 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L. The results from the positive control with potassium dichromate were within the normal ranges for this reference item.
Reported statistics and error estimates:
Statistical analysis of the growth rate data was carried out for the control and all test concentrations using 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). There were no statistically significant differences (P≥0.05), between the control, 6.25, 12.5, 25 and 50 mg/L test concentrations however the 100 mg/L test concentration was significantly different (P<0.05) and, therefore the "No Observed Effect Concentration" (NOEC) based on growth rate was 50 mg/L. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) based on growth rate was 100 mg/L.
Validity criteria fulfilled:
yes
Conclusions:
The effect of the Propan-1-ol, 2-amino-2-methyl-, 4-methylbenzenesulphonate on the growth of Pseudokirchneriella subcapitata has been investigated over a 72-Hour period and gave the following results - ErC50 (0 - 72 h) = 70 mg/L based on growth rate and EyC50 (0 - 72 h)=58 mg/L based on yeild inhibition.
Executive summary:

The effect of the Propan-1-ol, 2-amino-2-methyl-, 4-methylbenzenesulphonate on the growth of Pseudokirchneriella subcapitata has been investigated over a 72-Hour period and gave the following results - ErC50 (0 - 72 h) = 70 mg/L based on growth rate and EyC50 (0 - 72 h)=58 mg/L based on yeild inhibition.

Description of key information

The effect of the Propan-1-ol, 2-amino-2-methyl-, 4-methylbenzenesulphonate on the growth of Pseudokirchneriella subcapitata has been investigated over a 72-Hour period and gave the following results - ErC50 (0 - 72 h) = 70 mg/L based on growth rate and EyC50 (0 - 72 h)=58 mg/L based on yeild inhibition.

Key value for chemical safety assessment

EC50/LC50 for freshwater algae:
58 mg/L
EC10, LC10 or NOEC for freshwater algae:
42 mg/L

Additional information