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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:
2014-11-03 till 2015-01-19
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD guideline study without restrition.
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Version / remarks:
(2009)
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
(2006)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Sampling schedule: Concentration of 10.0 mg/L was measured at 0 and 72 hours; and concentration of 0 mg/L at 72 hour only.
- Sample storage conditions before analysis: Routinely, the samples were analysed immediately. Only in exceptional cases, they were stored overnight deep frozen and protected from light.
Vehicle:
no
Details on test solutions:
To produce the only concentration 20.4 mg of 14H-benz[4,5]isoquino[2,1-a]perimidin-14-one was added to 2 litres of dilution water, treated for 1 h in an ultrasonic bath and stirred for 24 h on a magnetic stirrer. Undissolved particles of 14H-benz[4,5]isoquino[2,1-a]perimidin-14-onem were removed by filtration using a folded filter with a pore size of 7-12 μm. The pH was measured to be 7.3.
100 mL of the solution were taken and 0.521 mL of the algal inoculum was added to each replicate resulting in a final cell density of 5000 cells/mL. For each test item concentration and the control 6 replicates were prepared. All flasks were sealed with cotton stoppers.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
- Name: Desmodesmus subspicatus (formerly Scene-desmus subspicatus) Strain No. 86.81 SAG
- Source: Strain of the test species obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany).
- Maintenance and Acclimatisation: 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 to 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.
- Preparation of pre cultures : 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.
- Test cultures: The algal inocula for the test were taken from an exponentially growing pre culture and were mixed with the growth medium to make up to a final cell density of about 5000 cells per millilitre in the test medium
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Hardness:
1.3 °dH (22.5 mg CaCO3/L)
Test temperature:
21 - 24 °C
pH:
Control: 7.7 at o h and 8.9 at 72 h
10.0 mg/L: 7.7 at 0 h and 8.4 at 72 h
Nominal and measured concentrations:
10.0 mg/L (nominal) plus contol
The results are expressed in terms of measured initial concentrations. Effective concentration corresponds to 8.2 % of nominal values at 0 hours, and corresponds to 7.3 % of nominal values at 72 hours. Based on the initial concentration the recovery was 88.7 % at the end of the incubation period.
Details on test conditions:
- Test vessels: 300 mL Erlenmeyer flasks with cotton, test volume: 100 mL
- Culturing apparatus : Shaking incubator 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.
- Light intensity: A light intensity ranging from 60 to 120 μE x m-2 x s-1, or an equivalent range of 4000 to 8000 lux, was measured. The light intensity was checked before the start of the study.
- Cell density measurements: Cell densities were measured in a microcell counter (Sysmex F300, Digitana) by taking small aliquots from each test flask, which were not replaced.
- Experimental design: 1 test concentration plus 1 control 6 replicates per concentration, 6 replicates per control Initial cell density in the test cultures approximately 5000 cells per millilitre. Additionally test concentration 10 mg/L without algae.
- Test item concentration: 10.0 mg/L (nominal)
- Method of administration: direct weighing
- Duration of exposure: 72 hours
- Criteria of effects: The criteria of adverse effects used in this study were the item-induced inhibition of yield [y] and growth rate [r] of the algal population.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
other: ErC10
Effect conc.:
> 0.824 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
other: ErC50
Effect conc.:
> 0.824 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
other: NOEC[r]
Effect conc.:
>= 0.824 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
other: LOEC[r]
Effect conc.:
> 0.824 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate

No toxic effects against algae were observed at the limit of water solubility under exposure conditions.

The results are expressed in terms of measured initial concentrations. Effective concentration corresponds to 8.2 % of nominal values at 0 hours, and corresponds to 7.3 % of nominal values at 72 hours. Based on the initial concentration the recovery was 88.7 % at the end of the incubation period.

Validity criteria fulfilled:
yes
Remarks:
(-The factor of biomass parameter 114.0 > 16; - The mean of the replicate coefficients of variation in the section-by-section growth rate 12.6 % < 35 %; -The coefficient of variation of the mean specific growth rate replicates in the control 0.5 % < 7 %.)
Conclusions:
No toxic effects against algae were observed at the limit of water solubility under exposure conditions.
Executive summary:

The acute toxicity to algae of 14H-benz[4,5]isoquino[2,1-a]perimidin-14-one was determined according to EU method C.3 "Freshwater Alga and Cyanobacteria, Growth inhibition test" (2009) which is equivalent to OECD Guideline for Testing of Chemicals No. 201 (2006). Desmodesmus subspicatus was exposed to the test solution of one nominal concentration of 14H-benz[4,5]isoquino[2,1-a]perimidin-14-one (10.0 mg/L) and black control solution for a period of 72 hours under static conditions. The cell densities were measured at 24 hour intervals. Inhibition of the algal population was measured as reduction in growth rate (index r), relative to control cultures grown under identical conditions. The results are expressed in terms of measured initial concentrations. Effective concentration corresponds to 8.2 % of nominal values at 0 hours, and corresponds to 7.3 % of nominal values at 72 hours. Based on the initial concentration the recovery was 88.7 % at the end of the incubation period. No toxic effects against algae were observed at the limit of water solubility under exposure conditions.

An ErC10 and an ErC50 of > 0.824 mg/L were measured and a NOEC [r] of ≥ 0.824 mg/L and a LOEC [r] of > 0.824 mg/L were calculated.

Description of key information

No toxic effects against algae were observed at the limit of water solubility under exposure conditions.

Key value for chemical safety assessment

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

Additional information

The key values should read as EC50 > 0.824 mg/L and NOEC ≥ 0.824 mg/L as there was no effect up to the limit of water solubility.