4,6-Dichloro-5-fluoropyrimidine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 Aug - 14 Sep 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.078, 0.156, 0.313, 0.625, 1.25, 2.5, 5 and 10 mg/L as nominal concentrations.
- Sampling method: Aliquots of the samples from the biological test were directly analysed by HPLC and UV/VIS-detection (range of the injection volume: 1-100 µL, depending on the expected concentration).
- Sample storage conditions before analysis: Routinely, the samples are analysed immediately. Only in exceptional cases, they are stored overnight deeply frozen and protected from light.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution was prepared to give the desired series of test concentrations. To achieve this 12.6 mg of the test item were added to 1 L of dilution water and treated for 1 h in an ultrasonic bath and stirred for 24 hours on a magnetic stirrer.
- Controls: 1 control, six replicates
- Test concentration separation factor: 2
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): None reported

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Source (laboratory, culture collection): obtained from The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Gottingen (Germany)
- Age of inoculum (at test initiation): 72 h
- Method of cultivation: Pre-cultures are set up three days before the start of a test. They are grown under identical exposure conditions as the stock cultures, except from the use of a different nutrient medium

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not): Different medium, other conditions identical
- Any deformed or abnormal cells observed: None

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

23 +/- 1°C

pH:

8.2 - 10.1 (0 and 72 h)

Nominal and measured concentrations:

0.078, 0.156, 0.313, 0.625, 1.25, 2.5, 5 and 10 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks with cotton stoppers
- Type (delete if not applicable): closed.
- Initial cells density: 10^4 cells per mL of medium
- Control end cells density: 936111 cells per mL
- No. of vessels per concentration (replicates): 3 replicates per concentration
- No. of vessels per control (replicates): 6 replicates of control

GROWTH MEDIUM
- Standard medium used: Yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to guideline
- Culture medium different from test medium: Yes

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: Continuous illumination
- Light intensity and quality: At the average of the test solutions, a light intensity in the range 60 to 120 µE. x m"2 x s"1, or an equivalent range of 4000 to 8000 Iux, throughout test

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Cell densities are measured in a microcell counter or, alternatively, are determined by means of a microscopic counting chamber.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2

CULTURING APPARATUS
-Details on culturing apparatus used: Light chamber

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.617 mg/L

95% CI:

>= 0.526 - <= 0.72

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.19 mg/L

95% CI:

>= 0.155 - <= 0.232

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.04 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.04 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control (for algal test): Yes
- Any stimulation of growth found in any treatment: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: None

Validity criteria for the measurement of the toxicity to algae:

Target condition according to guideline:	Actual condition according to the study:	Validity criteria met:
Exponentially growing test organisms are exposed to the test substance in batch cultures over a period of normally 72 hours.	Desmodesmus subspicatus was used as a test organisms. Exponential growth of the algae was confirmed. The test duration was 72 h.	Yes
For the final definitive test at least five concentrations, arranged in a geometric series with a factor not exceeding 3.2, should be selected.	Eight concentrations were used. The test concentration separation factor was 2.	Yes
The cultures should be maintained at a temperature in the range of 21 to 24°C, controlled at ± 2°C.	The test temperature was 23 +/- 1°C.	Yes

 

Validity criteria fulfilled:

yes

Remarks:

See 'Any other information including tables'.

Conclusions:

The following values were determined: EC50 (growth rate) = 0.617 mg/L, EC50 (biomass) = 0.19 mg/L, NOEC (biomass) = 0.04 mg/L, NOEC (growth rate) = 0.04 mg/L. All results are expressed in terms of geometric mean measured concentrations.

Executive summary:

Exponentially growing algal cells were exposed for a period of 72 hours to a range ofconcentrations, nominally 0.078, 0.156, 0.313, 0.625, 1.25, 2.5, 5 and 10 mg/L of the test substance dissolved in water. Auxiliaries used to prepare the test media were an ultrasonic bath and magnetic stirrer.The cell densities were measured at 24 hour intervals. Inhibition of the algal population wasmeasured as reduction in growth (index b) and growth rate (index r), relative to control cultures grown under identical conditions. Growth and growth rates were used to calculate a No Observed Effect Concentration and a Lowest Observed Effect Concentration according to DUNNETT (1955, 1964). The following values were determined: EC50 (growth rate) = 0.617 mg/L, EC50 (biomass) = 0.19 mg/L, NOEC (biomass) = 0.04 mg/L, NOEC (growth rate) = 0.04 mg/L. All results are expressed in terms of geometric mean measured concentrations. Recovery rates ranged from 80.3 - 91.7% of nominal values at 0 hours, and from 7.7 - 54.3% of nominal values at 72 hours, respectively.

Description of key information

Exponentially growing algal cells were exposed for a period of 72 hours to a range ofconcentrations, nominally 0.078, 0.156, 0.313, 0.625, 1.25, 2.5, 5 and 10 mg/L of the test substance dissolved in water. Auxiliaries used to prepare the test media were an ultrasonic bath and magnetic stirrer.The cell densities were measured at 24 hour intervals. Inhibition of the algal population wasmeasured as reduction in growth (index b) and growth rate (index r), relative to controlcultures grown under identical conditions. Growth and growth rates were used to calculate a No Observed Effect Concentration and a Lowest Observed Effect Concentration according to DUNNETT (1955, 1964). The following values were determined: EC50 (growth rate) = 0.617 mg/L, EC50 (biomass) = 0.19 mg/L, NOEC (biomass) = 0.04 mg/L, NOEC (growth rate) = 0.04 mg/L. All results are expressed in terms of geometric mean measured concentrations. Recovery rates ranged from 80.3 - 91.7% of nominal values at 0 hours, and from 7.7 - 54.3% of nominal values at 72 hours, respectively.

Key value for chemical safety assessment

EC50 for freshwater algae:

0.617 mg/L

EC10 or NOEC for freshwater algae:

0.04 mg/L