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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP Guideline Study
Qualifier:
according to guideline
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
no
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Test concentrations: at 0 and 48 hours, Control at 48hours
- Sampling method: from freshly prepared medium
- Sample storage conditions before analysis: The biological test solutions were routinely measured on the day of sampling. If this was exceptionally not possible, the samples were stored in a refrigerator at 4°C until the analysis was carried out. The biological test solutions were analysed in the same way as the calibration samples
Vehicle:
no
Details on test solutions:
Pre-treatment of test item and preparation of test item concentrations:
- 100.8 mg of PDI were added to 1 l of dilution water (Water Accommodated Fraction (WAF))
- stirred for 24 hour on a magnetic stirrer
- undissolved particles were removed by filtration using a folded filter with a pore size of 7-12 μm
- pH was measured to be 7.1
- for the loading 6 replicates were prepared
- 100 mL of the solution were taken and 0.526 mL of the algal inoculum was added to each replicate resulting in a final cell density of 5000 cells/mL
- for the test item loading 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:
- Strain No. 86.81 SAG
- stock culture: strain obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany)
- exponentially-growing stock cultures maintained under constant temperature conditions (21-24°C, +/- 2°C), light intensity (60 - 120 µE. x mE-2 x sE-1),measured in the range 400 to 700 nm using a spherical quantum flux meter)
- Cell density measurements were made using a microcell counter, Sysmex F300, Digitana
- growth medium: OECD medium of OECD TG 201, renewed once a week
- Pre-cultures: set up three days before test start, grown under identical exposure conditions as the stock cultures, except from the use of a different growth medium
- 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, corresponding to 22.5 mg/L CaCO3
Test temperature:
21 °C to 24 °C (+/- 2 °C)
pH:
7.9 at start, 8.0 after 72 h (Control)
7.3 at start, 8.2 after 72 H ( 100 mg/L test concentration)
Dissolved oxygen:
No data
Salinity:
n.a.
Nominal and measured concentrations:
Nominal concentration of test substance : 100 mg/L
Details on test conditions:
Exposure conditions:
- Test vessels: 300 mL Erlenmeyer flasks with cotton stoppers 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.
- Method of administration: direct weighing
- 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.
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
LOELR
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
DOC Value (100 mg/L PDI): 47.54 mg/L at start, 44.181 mg/L after 48 h
DOC Value (Control): 2.7054 mg/L after 48 h

NOEC and LOEC were determined according to STUDENT-t test for Homogeneous Variances.

Validity criteria fulfilled:
yes
Remarks:
(-The factor of biomass parameter 117.0 >16; -The mean of the replicate coefficients of variation in the section-by-section growth 13.1 % <35 %; -The coefficient of variation of the mean specific growth rate replicates in the control 1.1 % <7 %.)
Conclusions:
No toxic effects against algae were observed at the limit loading of 100 mg/L under exposure conditions.
Executive summary:

In order to test acute toxicity to algae of PDI, exponentially growing algal cells (Desmodesmus subspicatus) were exposed for a period of 72 hours to a limit test concentration of nominally 100 mg/L of PDI dissolved in dilution water. During the test a temperature range of 21 - 24 °C was maintained in the test vessels. The pH was measured at the beginning of the test and after 72 hours of exposure. PDI has not a definite or unique structure and consists of serveral components. Water Accommodated Fraction (WAF) was used to test effects at limit effective loading. The content of PDI during the exposure period was verified by DOC determination. 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. Growth rates were also used to calculate a No Observed Effective Loadings and a Lowest Observed Effective Loadings according to STUDENT-t test for Homogeneous Variances. The following values were determined: ErL 50 (0-72 h) : > 100; ErL 10 (0-72 h) : > 100; NOEL [r] (tα 0.05) : ≥ 100; LOEL [r] (tα 0.05) : > 100. No toxic effects against algae were observed at a limit test concentration of nominally 100 mg/L.

This toxicity study is classified as acceptable and satisfies the guideline requirements for the algae study.

Description of key information

No toxic effects against algae were observed at the limit loading of 100 mg/L after 72 hours under exposure conditions (Spoo-Klöppel, 2016).

Key value for chemical safety assessment

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

Additional information

"Should read: EL-50 > 100, NOEL >= 100"

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