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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:
June 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: well documented guideline study in compliance with GLP
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
5 ml of acetonitrile was added to 5 ml of water sample (control sample, sample contaminated with test material or fortified with standard solution) in Erlenmeyer flask. Contents of the flask was mixed throughly and applied to the chromatographic column in volume of 20 µL. Solutions of Fe(III)IDHA in concentration below 2 mg/L were injected to the chromatographic column without dissolving in acetone.
Vehicle:
no
Details on test solutions:
The test material was used in one concentration (limit test) - 100 mg/L. Concentration of test material was prepared from stock solution in concentration 1000 mg/L. Stock solution was prepared by weighting 50 mg of test material and dissolving in 50 mL of ES growth medium. Thest concentration was prepared by adding of appropriate volume of stock solution to growth medium and inoculating with algae to obtain initial concentration 4 x 10 4 cell/mL. Six replicates were used for test concentration as well as for control. Content of test material in test solutions was confirmed by analytical measurement.
Test organisms (species):
other: Pseudokirchneriell subcapitata 61.81 SAG (formely Scenedesmus subspicantus)
Details on test organisms:
Alge culture of Pseudokirchneriella subcapitata 61.81 SAG was obtained on agar bevel on May 20, 2002 from the Alge Collection University of Gottingen, Germany.
Alge were transfered from vials to the fresh medium (ES Basal Medium) in 250 ml Erlenmeyer flask and incubated in 25oC in constant illumination (basic culture). Growth medium was preparaed based on sterile destilled water (fiter Barnstead EASYPure RF, USA) with reagent-grade chemicals (Sigma and Aldrich). The basic culture were inoculated to the new medium once a week with a sterile pipette.
Before the experiments, set up of the initial culture were started (inoculated from basic cultures) in thsame conditions as during the experiments. After attaining the exponential growth phase in initial cutures (about three days of incubation) algae were used for inoculation of the test cultures provided that the cells showed no abnormalities or any deformations (microscpic observation).

ALGAE CELL COUNTING
The algae cells number was counted in every replicate of concentration and control after 24, 48 and 72 hours following the beginning of the experiment. The cells number was determined in Bürker’s chamber using microscope (Olympus CX40 RF200) with magnification: ocular x 10, objective x 40.
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
not reported
Hardness:
not reported
Test temperature:
24.5 - 25oC
pH:
control t0 pH = 6.85
control t72 pH = 7.02
tested solution t0 pH = 6.58
tested solution t72 pH = 7.25
Dissolved oxygen:
not reported
Salinity:
not applicable
Nominal and measured concentrations:
The test material in test solution iFe(III)IDHA in test solution after 72 h was on 57% level of concentration measured at t0.
Details on test conditions:
The experiment was conducted in photothermostatic room with temeprature 24.5 - 25oC and illumination 5890 - 6620 lx (planar lux-meterSonopan L-50, Poland). The test flasks and contol flasks were arranged at random and during the test rearranged repeatedly as well as shaen manually three times per day. At the begining and the end of the test pH values were measured (inoLAB pH/Oxi-meter Level 3, WTW, Germany). The test duration was 72 h.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 72.2 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
The toxicity of test material to algae is expressed as its median concentration causing 50% in-hibition of average specific growth rate or yield (ErC50, EyC50) of algae culture in relation to control. In the experiment only one concentration of test material was used – 100 mg/L (limit test).
Because no growth inhibition effect in tested concentration in relation to control was observed population, concentration causing 50% inhibition of average specific growth rate of Pseu-dokirchneriella subcapitata culture (ErC50/72 h) as well as concentration causing 50% of yield inhibition of Pseudokirchneriella subcapitata culture (EyC50/72 h) are above value of used concentration – 100 mg/L. Because of only one conce ntration used in test there is no possi-bilities to NOEC and LOEC values estimation. NOEC and LOEC values are above 100 mg/L.

All validity criteria were met according to OECD Guideline 201:
- Increase of biomass in control during 72 h was 47 times (criterion – not less than 16 times)
- Coefficient of variation for specific growth rate for control in the whole period of test (t0-t72) is 6.08 % (criterion – not more than 7 %)
- pH value in control during the test was changed by +0.17 (criterion – change should be lower than ± 1.5).
Validity criteria fulfilled:
yes
Conclusions:
Based on obtained results no growth inhibition effect in tested concentration in relation to control population was observed. Moreover slight stimulation effect was observed in test con-centration. This stimulation was on level of 26.75 % above the control.

According to this, concentration causing 50% inhibition of average specific growth rate of

Pseudokirchneriella subcapitata culture ErC50/72 h is above 100 mg/L as well as concentra-tion causing 50% of yield inhibition of Pseudokirchneriella subcapitata culture EyC50/72 h is above 100 mg/L based on nominal concentration of test material.

Values ExC50/72 h and EyC50/72 h based on measured concentrations of test material is above 72.3 mg/L estimated on the base of geometrical mean of measures in the time t0 and t72.
Executive summary:

Growth inhibition assessment of Fe(III)IDHA was performed according to the OECD Guideline No 201, method C.3 and Principles of Good Laboratory Practice (GLP, OECD 1997).

 

The aim of the study was determination of growth inhibition of algae Pseudokirchneriella subcapitata after 24, 48 and 72 h exposure based on average specific growth rate inhibition(expressed as the ErCx in mg/L) and yield inhibition (expressed as the EyCx in mg/L). Also the aim of the study was assessment of NOEC and LOEC values. The test material was used in one concentration (limit test) – 100 mg/L.

 

Based on obtained results no growth inhibition effect in tested concentration in relation to control population was observed. According to this, concentration causing 50% inhibition of average specific growth rate ofPseudokirchneriella subcapitataculture ErC50/72 h is above 100 mg/L as well as concentration causing 50% of yield inhibition ofPseudokirchneriellasubcapitataculture EyC50/72 h is above 100 mg/L based on nominal concentration of test ma-terial.

 

Values ExC50/72 h and EyC50/72 h based on measured concentrations of test material is above 72.3 mg/L estimated on the base of geometrical mean of measures in the time t0 and t72.

Description of key information

The average specific growth rate of Pseudokirchneriella subcapitata culture ErC50/72 h

is above 100 mg/L as well as concentration causing 50% of yield inhibition of Pseudokirchneriella subcapitata culture EyC50/72 h is above 100 mg/L based on nominal concentration of test ma-terial.

Key value for chemical safety assessment

EC50 for freshwater algae:
100 mg/L

Additional information

Based on obtained results no growth inhibition effect in tested concentration in relation to control population was observed. According to this, concentration causing 50% inhibition of average specific growth rate ofPseudokirchneriella subcapitataculture ErC50/72 h is above 100 mg/L as well as concentration causing 50% of yield inhibition ofPseudokirchneriellasubcapitataculture EyC50/72 h is above 100 mg/L based on nominal concentration of test ma-terial.

 

Values ExC50/72 h and EyC50/72 h based on measured concentrations of test material is above 72.3 mg/L estimated on the base of geometrical mean of measures in the time t0 and t72.