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Ecotoxicological information

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:
Aug 28- 31, 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
Preparation and storage
Tue test medium (reconstituted water and test material) was freshly prepared. Therefore, the calibrated flask with test material and vehicle, reconstituted water, was treated in an ultrasonic device for 1 hour. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Afterwards, the formulation was given through a suction filter (pore size >6 - <10 μm) and the filtrate was used for the study.
Analytical monitoring:
no
Remarks:
Due to the low solubility of the test item, an analyitcal method was not developed.
Vehicle:
no
Details on test solutions:
Reconstituted water

Macro nutrients:
NaHCO3 50 mg/L
CaCl2 x 2 H2O 18 mg/L
NH4Cl 15 mg/L
MgSO4 x 7 H2O 15 mg/L
MgCI2 x 6 H2O 12 mg/L
KH2PO4 1.6 mg/L



Trace elements:

Na2EDTA x 2 H2O 100.00 µg/L
FeC13 x 6 H2O 80.00 µg/L
MnC12 x 4 H2O 415.00 µg/L
H3BO3 185.00 µg/L
Na2MoO4 x 2 H2O 7.00 µg/L
ZnC12 3.00 µg/L
CoCl2x 6 H2O 1.50 µg/L
CuC12 x 2 H2O 0.01 µg/L

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Desmodesmus subspicatus
- Strain: No. SAG 86.81
- Source (laboratory, culture collection): Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universitat Göttingen, Germany
- Age of inoculum (at test initiation): freshly prepared
- Method of cultivation: under standardised lab conditions

ACCLIMATION
- Acclimation period: Prior to start the study, the algae were grown in reconstituted water as preculture and the growth rate was determined.
- Culturing media and conditions (same as test or not): same
- Any deformed or abnormal cells observed: no
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
-
Hardness:
250 mg/L
Test temperature:
23 °C
pH:
7.6 - 10
Nominal and measured concentrations:
Nominal concentrations: 100.0 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks
- Type: open
- Material, size, headspace, fill volume: 300 mL Erlenmeyer glas flasks filled with 100 mL cell suspension
- Aeration: no
- Initial cells density: 10 000 cells per ml
- Control end cells density: 1 146 231 cells per ml
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water according to OECD TG 201
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Light intensity and quality: The mean light intensities were 7500 Lux to 7500 Lux prior to and at the end of the exposure period, respectively


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic particle counter

TEST CONCENTRATIONS
- Nominal test concentration: 100 mg/L
Reference substance (positive control):
not required
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 0.0114 mg/L (limit of water solubility)
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 0.0114 mg/L (limit of water solubility)
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 0.0114 mg/L (limit of water solubility)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 0.0114 mg/L (limit of water solubility)
Details on results:
Pretest

In a pretest under static conditions, inhibition of the growth rate of the algae, Desmodesmus subspicatus, was not observed at a concentration of 100 mg/I, nominally.

Main study

In the control group, the factor of cell growth was approx. 115.

The algae increased from 10,000 cells/mL at the start to 1,146,231 cells/mL after 72 hours.

The nominal test material concentrations and the growth inhibition data after 72 hours exposure are given below:


---------------------------------------------------------------------------------------------------
Nominal
conc. (mg!L) Cell density per mL Growth
inhibition
(0-72 hours)
0 hour 24 hours 48 hours 72 hours IA lµt
---------------------------------------------------------------------------------------------------
10,000 61,983 281,299 1,173,933
10,000 60,390 267,461 1,079,163
10,000 63,270 314,160 1,185,597
mean 10,000 61,881 287,640 1,146,231
10,000 48,717 263,721 1,128,897
100 10,000 51,660 275,791 1,165,023
10,000 52,947 313,990 1,314,063
mean 10,000 51,108 284,501 1,202,661

Inhibition IA 20.8 4.2 -3.8 -1.6 -1.0
---------------------------------------------------------------------------------------------------

IA = inhibition of cell growth in percent
Iµt= inhibition of growth rate in percent

negative values = the growth of the algae was better in the test medium than in the control group


The nominal test material concentrations and the growth inhibition data after 72 hours exposure are given below:

Nominal Concentration
(mg/L)
Mean cell density per mL
0 hour 24 hours 48 hours 72 hours
0 10000 61881 287640 1146231
100 10000 51108 284501 1202661
IA
20.8 4.2 -3.8

Comments:
The analysis revealed, that the solubility of the test material in reconstituted water was < 0.0114 mg/L. Because of this low water solubility and the fact that the study was performed as a limit test, the test material concentration
in the aqueous medium at the start and the end of this study was not quantified.

For the test material the following nominal EC50  values and no effect concentration for Desmodesmus subspicatus were determined:

72 h EbC50 >100 mg/L (nominal)

72 h ErC50 >100 mg/L (nominal)

NOEC >=100 mg/L (nominal)

Validity criteria fulfilled:
yes
Conclusions:
An aqueous preparation of 100 mg/L (nominal) of the test materail revealed no toxic effect in the test system. Therefore, the 72 hours EC50 could not be determined, because it exceeded the maximum solubility of the test material in reconstituted water (EC50 > 0.0114 mg/L).
Executive summary:

Purpose
The purpose of this assay was to identify the aquatic toxicity potential of the test item in algea to provide a rational basis for hazard estimation for the test item in aquatic environments.

Study Design
The influence of the test material on the growth and growth rate of the green algae species Desmodesmus subspicatus was tested. Therefore, a GLP compliant study according to OECD TG 201 was undertaken. The study design included one control and one test material group with three replicates, each containing 100 mL reconstituted water or test medium and about 10,000 cells/mL at the start of the experimental phase.
The study was performed as a limit test in an open static test system. The algae were exposed to a filtrate of nominal 100 mg/L.
The growth of the algae was calculated after 24, 48, and 72 hours exposure in the test medium.


Results
The analysis of the saturated aqueous preparation revealed that the water solubility of the test item in reconstituted water was < 0.0114 mg/L. An exact value could not be determined. The test material concentration in the aqueous medium could not be quantified since it was below the limit of quantification.

After exposure of Desmodesmus subspicatus for 72 hours the following results were obtained.

Nominal Concentration
(mg/L)
Mean cell density per mL
0 hour 24 hours 48 hours 72 hours
0 10000 61881 287640 1146231
100 10000 51108 284501 1202661
IA
20.8 4.2 -3.8



Growth Inhibition data (0-72 h). IA=-1.6; Iµt=-1.0

IA= inhibition of growth in percent
Iµt= inhibition of growth rate in percent


The test item had no significant inhibitory effect on the mean biomass and the mean growth rate after the exposure period of 72 hours at the nominal concentration of 100 mg/L.

For the test material, the following EC50 values and no effect concentration for Desmodesmus subspicatus were determined:
72 h EbC50    >    0.0114 mg/L (nominal >    100 mg/L)
72 h ErC50    >    0.0114 mg/L (nominal >    100 mg/L)
NOEC          >=    0.0114 mg/L (nominal >=   100 mg/L)


Conclusion
An aqueous preparation of 100 mg/L (nominal) of the test materail revealed no toxic effect in the test system. Therefore, the 72 hours EC50 could not be determined, because it exceeded the maximum solubility of the test material in reconstituted water (EC50 > 0.0114 mg/L).

Description of key information

No adverse effects on algae were observed up to the limit of water solubility. Thus, the 72 -h EC50 value and 72 -h EC10 value are greater than the limit of water solubility

of 0.0114 mg/L which corresponds to a nominal concentration of 100 mg/L.

OECD 201: EC50 (72 h) > 100 mg/L (nominal), NOEC (72 h) > 100 mg/L (0.0114 mg/L (limit of water solubility))

Key value for chemical safety assessment

Additional information

A study was conducted according to OECD Guideline 201 in accordance with good laboratory practice. Control experiments were run witth water without andy further additives. After 24 and 48 h the immobilized animals were determined. The test item had did not show any effect at a nominal concentration if 100 mg/L with a maximum water solubility of 0.0114 mg/L.

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