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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
Study period:
12 November 2013 - 14 July 2014
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 201 (Alga, Growth Inhibition Test)
according to guideline
EU Method C.3 (Algal Inhibition test)
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
- Physical State: Pale yellow powder
- Source: Sponsor
- Batch Number: OF1211
- Purity: 99.9%
- Storage: Room temperature in the dark
- Expiry Date: 30 September 2014
Details on sampling:
- Concentrations: nominal loading rates of 0.298, 0.954, 3.05, 9.77, 31.3 and 100 mg/mL
- Sampling method: As the test material was insoluble in water (<0.004 mg/L), it was not possible to develop a suitable method of analysis with sufficient sensitivity to monitor the exposure concentrations employed in the test, consequently, no supporting analysis was undertaken.
- Sample storage conditions before analysis: No samples takem fr analysis as detailed above.
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Batches of the test medium were prepared by adding the test substance (500, 156.5, 48.9, 15.3 or 9.54 mg) to dilution medium (ca. 4.5 or 9.5 L) in an aspirator (5 L for 500, 156.5, 48.9 and 15.3 mg/L and 10 L for 9.54 mg/L), after which the contents of the aspirators were adjusted to volume whilst being stirred. Each bulk preparation was stirred for a minimum of 24 hours in the dark and then a syphon was added to each preparation vessel and the media left to stand for at least 24 hours in the dark. An aliquot (1 L) of the WAF was removed via the syphon tube and discarded; the mid-vessel content (1 L) was syphoned to give water accommodated fractions (WAF) of test media with nominal loading rates of 100, 31.3, 9.77,
3.05 and 0.954 mg/L as T-1620L.
Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
- Strain: CCAP 276/4
- Source (laboratory, culture collection):Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd., Dunstaffnage Marine Laboratory, Dunbeg, Oban, Argyll, Scotland
- Age of inoculum (at test initiation):N/A
- Method of cultivation: The liquid slope cultures were stored in an illuminated refrigerator. Sterile algal nutrient medium was inoculated with cells aseptically removed from the slope culture; these primary liquid cultures (100 mL) were incubated for approximately three days in an orbital incubator under continuous illumination at nominal temperatures in the range 21 to 24°C. Subsequently, appropriate volumes of these primary cultures were aseptically
transferred to fresh sterile algal nutrient medium to prepare secondary liquid cultures; these cultures were incubated, as stated above, for a further three days to provide an inoculum in the log phase of growth, characterised by a cell density of 1.82 x 10^6 cells/mL.

- Acclimation period: Approx 6 Days
- Culturing media and conditions (same as test or not): Yes
- Any deformed or abnormal cells observed: No
Test type:
Water media type:
Limit test:
Total exposure duration:
72 h
Post exposure observation period:
Test temperature:
7.72 - 8.22
Dissolved oxygen:
Nominal and measured concentrations:
Study end-points have been expressed in terms of nominal loading rates of T-1620L. Nominal concentrations were 0.298, 0.954, 3.05, 9.77, 31.3 and 100 mg/L
Details on test conditions:
- Test vessel: conical flask
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 250mL glass flasks with 100mL control or test culture
- Aeration: orbital shaker
- Initial cells density: 1 x 10^4 cells/mL
- No. of organisms per vessel: 1.82 x 10^6 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

- Standard medium used: yes
- Detailed composition if non-standard medium was used: N/A

- Sterile test conditions: yes
- Adjustment of pH: 21-24°C
- Photoperiod: continuous
- Light intensity and quality:nominally 4440 to 8880 lux.

- Spacing factor for test concentrations: factor of 3
- Test concentrations:water accommodated fractions (WAF) with nominal loading rates of 1, 10 and 100 mg/L as T-1620L
Reference substance (positive control):
72 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
growth rate
Validity criteria fulfilled:
Based on nominal loading rates, there was insufficient inhibition for the calculation of the 72-hour EL10 and EL50 values for growth rate and yield. The ‘no-observed effect loading rate’ (NOELR) for growth rate and yield was 100 mg/L, the highest loading rate employed in the test.

Description of key information

Inhibition of Growth Rate and Yield:

NOELR: 100 mg/L

Key value for chemical safety assessment

Additional information