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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 2009-04-20 to 2009-07-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
reference to other study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
see below
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
yes
Remarks:
see below
Principles of method if other than guideline:
Deviations:
• The temperature in the positive control was higher than stated in the guidelines. As algal growth in the controls was normal, this deviation was stated as uncritical.
• The temperature in the main study was higher than stated in the guidelines. As algal growth in the controls was normal, this deviation was stated as uncritical.

GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
not indicated.
Analytical monitoring:
yes
Details on sampling:
Treatment and control were measured threefold at start and end of the study.
Vehicle:
no
Details on test solutions:
A stock solution containing 125 mg/L test item in deionised water was prepared. For each treatment, 40 mL of the respective test solution (1.25-fold concentrated) was prepared and mixed with 10 mL of the test culture.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
Specification
Unicellular freshwater green alga.
Genus, Species Desmodesmus subspicatus
Strain CHODAT
Family Chlorophyceae
Order Chlorococcales

Origin and Culture
The culture of Desmodesmus subspicatus was obtained in 18 Feb. 2008 by the "Collection of Alga“, Institut für Pflanzenphysiologie of Universität Göttingen. The alga are kept as stock culture on solid agar at 8 °C. From an aliquot of this stock culture the pre-culture was prepared.
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
no post exposure period.
Hardness:
not indicated
Test temperature:
Temperature range was 25.0 – 30.0 °C.
pH:
The pH values in the control ranged from 7.6 to 8.8.
Dissolved oxygen:
not indicated
Salinity:
freshwater was used.
Nominal and measured concentrations:
nominal: limit test: 0 and 100 mg/L
measured (after 72 h): 0 and 106.82 mg/L
The mean of the measured concentration after 72 hours was 109 % of the start concentration. The mean of the measured concentration at the beginning was 98 % and at the end 107 % of the nominal concentration. Therefore, the determination of the biological results was based on the nominal concentration.
Details on test conditions:
Algae
Three days before the start of each test, an aliquot of the stock culture containing a few cells was brought into pre-culture medium and incubated for 72 hours. The resulting culture is growing exponentially.
Before usage, the culture was checked on the absence of cell aggregates. After the adjustment to a cell concentration of about 10E04/mL through photometric measurement and addition of algal medium, the culture was usable for the test. The adjusted pre-culture was mixed with the same amount of 10-fold-nutrient solution (composition 10-fold-nutrient solution see page 11). This mixture was the test culture.
Preparations
On the day before the start of the test, a sufficient number of vessels were provided. Two hours before the start, the microgreeny® was turned on in order to warm up.
Performance of the Study
For each treatment, 40 mL of the respective test solution (1.25-fold concentrated) was prepared and mixed with 10 mL of the test culture. In this mixture, the pH-value was measured. A sample for the analytical determination was taken.
The test vessels were filled with 5 mL of the mixture and were incubated open for 72 hours stirred with magnetic stirrers. Before the start of incubation and every 24 hours, the cell number was determined and the values of the absorption of the cuvettes were recorded. After the test, the pH value in the treatments was measured once more.
At the end of the test, the treatments were observed microscopically to verify a normal and healthy appearance of the alga (e.g. caused by the exposure of the test item).
The content of the test item in the test vessels was measured at the start and at the end of the test.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate K2Cr2O7
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
Normal growth was observed.
The analytical determinations of the test item showed good correlation between nominal and measured concentrations and good recovery rates. Therefore, the test item sodium cyanate can be stated as “stable under the test conditions” and the determination of the results was based on the nominal concentrations. The recovery rate at the end of the test was more than 100 % of the start concentration. This might be caused by evaporation of dilution water and therefore higher measured concentrations at the end of the test.
The result of the test can be considered valid.
Results with reference substance (positive control):
The EC50s of potassium dichromate were determined in a current reference test. For the estimation of the EC50s of the positive control, the fits showed sufficient statistical correspondence of the data with the dose-response-equation. The values were within the normal range of the laboratory as well as in correspondence with the values from the international ring test which are stated in EU Method C.3.
Reported statistics and error estimates:
Determination of the NOEC
The difference between the treatment 100 mg/L and the control can be considered as not significant (level of significance: 97.5 %) for the endpoints growth rate and yield, as the calculated t-values were smaller than the limit of significance. Therefore the concentration 100 mg/L is stated as NOEC for growth rate and yield.
The endpoint biointegral (AUC) shows a small, but significant inhibition. Therefore, for this endpoint, no NOEC can be stated.

Determination of the LOEC
As the differences between the treatment 100 mg/L and the control are considered as not significant for the endpoints growth rate and yield, no LOEC can be stated for these end-points.
For the endpoint biointegral (AUC), LOEC is stated as 100 mg/L.
Validity criteria fulfilled:
yes
Conclusions:
In an acute toxicity test to Algae an EC50 value of > 100 mg/L was determined for sodium cyanate.
Executive summary:

The study was performed using one concentration containing 100 mg/L. The treatments were used to incubate the unicellular freshwater green alga Desmodesmus subspicatus for a period of 72 hours. The cell concentration of each replicate was determined by measuring the absorption of the cuvettes at 440 nm every 24 hours with a spectral photometer. With these measured values, the number of cells was calculated (linear correlation be-tween cell concentration and absorption given). Then the growth rate µ, the area under the growth curve (AUC ) and the Yield were determined.

At the beginning and at the end of the test, the content of the test item in the test solution was determined using a spectral photometer. The recovery after 72 hours was 109 % of the start concentration. The correlation between nominal and measured concentration was good (98% at the beginning.and.107 % at the end). Therefore, the determination of the biological results was based on nominal concentration.

The EC50s of potassium dichromate were tested in a current positive control test. The values lay within the normal range of the laboratory.

The following results for the test item sodium cyanate were determined:

Endpoint

NOEC

LOEC

EC50

Growth Rate

100 mg/L

>100 mg/L

>100 mg/L

AUC

100 mg/L

>100 mg/L

>100 mg/L

Yield

100 mg/L

>100 mg/L

>100 mg/L

Description of key information

In an acute toxicity test to Algae an EC50 value of > 100 mg/L was determined for sodium cyanate.

Key value for chemical safety assessment

EC50 for freshwater algae:
100 mg/L
EC10 or NOEC for freshwater algae:
100 mg/L

Additional information

Key study:

The study was performed using one concentration containing 100 mg/L. The treatments were used to incubate the unicellular freshwater green alga Desmodesmus subspicatus for a period of 72 hours. The cell concentration of each replicate was determined by measuring the absorption of the cuvettes at 440 nm every 24 hours with a spectral photometer. With these measured values, the number of cells was calculated (linear correlation between cell concentration and absorption given). Then the growth rate µ, the area under the growth curve (AUC ) and the Yield were determined.

At the beginning and at the end of the test, the content of the test item in the test solution was determined using a spectral photometer. The recovery after 72 hours was 109 % of the start concentration. The correlation between nominal and measured concentration was good (98% at the beginning.and 107 % at the end). Therefore, the determination of the biological results was based on nominal concentration.

The EC50s of potassium dichromate were tested in a current positive control test. The values lay within the normal range of the laboratory.

The following results for the test item sodium cyanate were determined:

Endpoint

NOEC

LOEC

EC50

Growth Rate

100 mg/L

>100 mg/L

>100 mg/L

AUC

100 mg/L

>100 mg/L

>100 mg/L

Yield

100 mg/L

>100 mg/L

>100 mg/L