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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:
1997-09-27 to 1997-10-17
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
The study was conducted according to an appropriate OECD test guideline, with acceptable restrictions. The analytical method used was Dissolved Organic Carbon, which could not distinguish between the parent test substance and its hydrolysis products.
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
see below for details.
Principles of method if other than guideline:
In the algal medium the concentration of NaHCO3 was twice that indicated in the Guideline and the quantification of the test substance was performed using DOC/TOC determination method. According to the test report a test substance was stable in test solutions.
An initial cell concentration of 10^5 cells/ml was used instead of the recommended 10^4 cells/ml.
GLP compliance:
yes (incl. certificate)
Test material information:
Composition 1
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All concentrations tested.

- Sampling method: stock solutions were analysed for TOC determinations (Total Organic Carbom/Dissolved Organic Carbon) to ensure that the test substance was accurately diluted. After completion of the test the test solutions were analysed for DOC.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: The sock solution was adjusted to pH ~8. One part (5 parallels for each concentration) was diluted and inoculated with the algal suspension. The second part was diluted, the test algal medium was added and used as blanks for measuring test concentrations.

- Controls: test medium
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM

- Strain: no. 86.81

- Source (laboratory, culture collection): Sammlung von Algenkulturen, Pflanzen-physiologisches Institut der Universitat Gottingen [SAG]

- Age of inoculum (at test initiation): 3 days

ACCLIMATION

- Acclimation period: 3 days

- Culturing media and conditions (same as test or not): same as test
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
25 degC +/- 1 degC, once the test temperature was recorded at 27.6 degC, it was not thought to have interfered with the results by the authors.
pH:
The pH of the test was about 8 in the beginning of the test and increased in the end of the test.
The increase of pH is caused by the algal growth in the test vessels. In common with other species of freshwater algae, Desmodesmus subspicatus gains CO2 from HCO3-, leading to a release of hydroxide ions, that alkalise the medium.

The shift in pH of >1 unit is indicative of good algal growth and is not considered to invalidate the test result.
Nominal and measured concentrations:
Nominal concentrations: 312.5, 625, 1250, 2500, 5000 and 10 000 mg/L.
Details on test conditions:
TEST SYSTEM

- Test vessel:

- Type: closed with a sterilised plexiglass lid.

- Material, size, headspace, fill volume: 50 ml glass cylinders filled with 50 ml test solution.

- Aeration: no aeration, but in otder to assure input of CO2 the test solutions were stirred with a magnetic stirrer for 15 min per day.

- Initial cells density: ca. 10^5 cells/ml

- No. of vessels per concentration (replicates): 5

- No. of vessels per control (replicates): 7



GROWTH MEDIUM

- Standard medium used: no

- Detailed composition if non-standard medium was used: see Table 1 for details.


TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: ultrapure water (MilliQ, Millipore)

- Culture medium different from test medium: no

- Intervals of water quality measurement: temperature - daily,


OTHER TEST CONDITIONS

- Sterile test conditions: yes

- Adjustment of pH: yes, the stock solution was adjusted to pH 8 (from 4.1) with 1M NaOH and then sterilised.

- Light intensity and quality: ~8000 lux


EFFECT PARAMETERS MEASURED

- Determination of cell concentrations: cell density was measured by a spectophotometer at 578 nm. Measurements were taken at time 0, 24, 48 and 71 +/- 0.5 h.



TEST CONCENTRATIONS

- Spacing factor for test concentrations: 2

- Range finding study

- Test concentrations: 100, 1000 and 10 000 mg/L  

- Results used to determine the conditions for the definitive study: NOEC < 100 mg/L
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
3 200 mg/L
Nominal / measured:
nominal
Conc. based on:
not specified
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 10 mg/L
Nominal / measured:
nominal
Conc. based on:
not specified
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
360 mg/L
Nominal / measured:
nominal
Conc. based on:
not specified
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
not specified
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
< 100 mg/L
Nominal / measured:
nominal
Conc. based on:
not specified
Basis for effect:
biomass
Remarks on result:
other: based on preliminary study
Details on results:
- Exponential growth in the control (for algal test): yes
Reported statistics and error estimates:
The inhibition of the growth rates was calculated by extrapolation of the concentration/response curve and according to the guideline. The inhibition of biomass production and the growth rate were calculated using the commercial computer program EXCEL, v. 5.0.

Table 1. Summary of findings.

 

 Nominal concentration (mg/L)

 Number of cells (/ml)

 Biomass inhibition (%)

 Growth rate inhibition (%)

 24h

 48h

 72h

 Control

 3.64 x 10^5 

 1.75 x 10^6

 3.7 x 10^6

 0

 0

 10,000

 2.75 x 10^5

 6.74 x 10^5

 1.85 x 10^6

 55

 17

 5,000

 3.19 x 10^5

 1.19 x 10^6

 2.11 x 10^6

 37

 13

 2,500

 2.99 x 10^4

 1.26 x 10^6

 2.56 x 10^6

 30

 9

 1,250

 3.61 x 10^5

 1.3 x 10^6

 3.02 x 10^6

 21

 5

 625

 3.81 x 10^5

 1.54 x 10^6

 3.01 x 10^6

 14

 5

 312.5

 3.74 x 10^5

 1.71 x 10^6

 3.14 x 10^6

 8

 5

 

Initial cell concentration is reported as 7.38 x 10^4 cell/ml in the control and ~8 x 10^4 in the other treatment levels.

 

Validity criteria fulfilled:
no
Remarks:
pH shifted by more than 1 unit.
Conclusions:
A 72 h ErC50 value of >10,000 mg/L has been determined for the effects of the test substance on growth rate of S. subspicatus. The pH value in the controls and the low treatment levels had varied by more than 1 unit. This is thought to not compromise the validity of the test since the controls appear to be unaffected, the cell numbers grew exponentially by a factor of >16 in 72 h.

Description of key information

A 72 h ErC50 value of >10,000 mg/L and a 72 h ECr10 of 3200 mg/L (nominal) have been determined for the effects of the test substance on growth rate of Scenedesmus subspicatus. The study was conducted acording to the OECD guideline 201.

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater algae:
3 200 mg/L

Additional information