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Toxicity to aquatic algae and cyanobacteria

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toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference Type:
Toxicity testing of highly volatile chemicals with green algae
Brack W, Rottler H
Bibliographic source:
Environmental Science and Pollution Research 1(4), 223-228

Materials and methods

Principles of method if other than guideline:
A modified cell multiplication inhibition test was applied
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:

Sampling and analysis

Analytical monitoring:
Details on sampling:
Defined volumes (10 or 100 microlitre) were taken in duplicate from the test cultures with a gas-tight syringe through the septa and injected into 2 mL vials containing 1 mL hexane with trichlorobromomethane as internal standard. Measurement at the beginning and at the end of the assays showed no significant differences and all samples were taken at the end of the assays.

Test solutions

Details on test solutions:
Saturated solutions in growth medium were prepared. Defined aliquots of these solutions were added to the medium. The effective concentrations of the compounds in the alga culture were determined using GC/ECD analysis of extracts at the end of the assays.

Test organisms

Test organisms (species):
Chlamydomonas reinhardtii
Details on test organisms:
Unicellular freshwater green alga Chlamydomonas reinhardtii (strain number 11-32a SAG) from the University of Göttingen, Germany, were obtained. Precultures and test cultures were grown in the medium for unicellular algae according to Kuhl (1962). Incubation of all cultures were shaken permanently with a frequency of 120 rpm. They were illuminated from above with 130 uE/m2s without light-dark cycle. The photosynthetically effective light was determined with a Quantum Sensor (Licor, Inc.).

Study design

Test type:
Water media type:
Limit test:
Total exposure duration:
72 h

Test conditions

no data given
Test temperature:
20 +/- 1 °C
6.5 to 7.5
Dissolved oxygen:
no data given
no data given
Nominal and measured concentrations:
no data given
Details on test conditions:
All assays were performed in bipartite vessels. The lower flask contained 50 mL of the CO2 buffer with 2 mol K2CO3/2 mol KHCO3 (35/65), which provided an atmospheric concentration of 1.14 % CO2. The CO2 concentration in the gas phase of the vessel was determined using GC/TCD. 1.2 % were measured without algal growth. Algal growth reduced CO2 concentrations to a minimum of 0.4 %. CO2 deficiency was not expected and not observed. The upper part of the vessel contained 30 mL of the growth medium with the algae. Both liquids were interconnected by their gas phase. The whole system was sealed gas-tight against the atmosphere. The CO2 buffer and the contaminated medium were filled into the test vessel at least 15 hours before the assay was started by adding the algal inoculum. Continuously shaken and temperated to 20 °C, the system was allowed to equilibrate. The inoculum was prepared from 7 day old precultures at a concentration of 3e5 cells/mL. For the test 0.5 mL inoculum was added to 30 mL medium. The cell density in the test cultures amounted to 5e^3 cells/mL at the beginning of the assays.
Reference substance (positive control):
Cu 2+ (CuSO4)

Results and discussion

Effect concentrationsopen allclose all
72 h
Dose descriptor:
Effect conc.:
3.61 mg/L
Nominal / measured:
meas. (initial)
Basis for effect:
growth rate
Remarks on result:
other: 2.55-4.72
72 h
Dose descriptor:
Effect conc.:
13.3 mg/L
Nominal / measured:
meas. (initial)
Basis for effect:
growth rate
Remarks on result:
other: 11.0-15.77

Applicant's summary and conclusion

Validity criteria fulfilled:
The growth of the green alga Chlamydomonas reinhardtii was inhibited by 50 % at a chloroform concentration of 13.3 mg/L.
Executive summary:

The toxicity of chloroform to the green alga Chlamydomonas reinhardtii was tested in a closed static test during a period of 72 hours at a temperature of 20 +/- 1 °C. The criterion for toxicity was the inhibition of the algal growth rate. The algal growth was inhibited by 10 % at a chloroform concentration of 3.61 (2.55 -4.72) mg/L and by 50 % at a chloroform concentration of 13.3 (11.0 -15.77) mg/L. The EC50 value for algal growth inhibition thus is 13.3 mg chloroform/L.