Fluocortolone

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:

15 Mar - 15 Jun 2016

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

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

not applicable

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.88, 9.4 and 100 mg/L
- Sampling method: Direct sampling
- Sample storage conditions before analysis: no storage, immediate measurement

Vehicle:

no

Details on test solutions:

PRE-TREATMENT OF THE TEST SUBSTANCE:
- 100.1 mg of the test item were added to 1 litre of dilution water and treated for 1 h in an ultrasonic bath and stirred for 24 h on a magnetic stirrer
- Undissolved particles of the test item were removed by filtration using a folded filter with a pore size of 7-12 μm
- The pH was measured to be 7.7
- Appropriate amounts of the stock solution were diluted with dilution water to a volume of 100 mL to produce the different test item concentrations
- Appropriate amounts of the stock solution was diluted with dilution water to a volume of 100 mL and 0.532 mL of the algal inoculum was added to each replicate resulting in a final cell density of 5000 cells/mL
- For each test item concentration and the control 3 replicates were prepared

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST SPECIES:
- Source: Strain of the test species obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany)
- Maintenance and Acclimatisation: Exponentially growing stock cultures maintained in the test facility under constant temperature conditions (21-24 °C with a maximum fluctuation of +/- 2 °C)
- Light intensity: in the range 60 to 120 µE x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter)
- Renewal of the growth medium: once a week
- Cell density measurements: microcell counter, Sysmex F300, Digitana
- Preparation of pre cultures: Pre cultures were set up three days before the start of a test and grown under identical exposure conditions as the stock cultures, except from the use of a different growth medium
- Test cultures: The algal inocula for the test were taken from an exponentially growing pre culture and were mixed with the growth medium
- Final cell density: about 5000 cells per millilitre in the test medium

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

22.5 mg/L CaCO3 (1.3 °dH)

Test temperature:

21 - 24 °C

pH:

7.9 -8.1

Nominal and measured concentrations:

Effective concentrations ranged from 98 % to 100 % of nominal values at 0 hours, and from 95 % to 101 % of nominal values at 72 hours.

Details on test conditions:

TEST DESIGN

Exposure conditions:
- Test vessels: 300 mL Erlenmeyer flasks with cotton stoppers, test volume: 100 mL
- Culturing apparatus: Shaking incubator, temperature in the range 21 °C to 24 °C (+/- 2 °C); continuous uniform illumination in the spectral range 400 to 700 nm; temperature was measured and recorded daily
- Light intensity: 60 to 120 µE x m-2 x s-1, or an equivalent range of 4000 to 8000 lux, was measured
- Cell density measurements: measured in a microcell counter (Sysmex F300, Digitana) by taking small aliquots from each test flask, which were not replaced
- Experimental design: 7 test concentrations plus 1 control
- Replicates: 3 per test concentration, 3 replicates per control
- Initial cell density in the test cultures: approx. 5000 cells/mL
- Test item concentration/s: 7 (0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L)
- Method of administration: stock solution
- Criteria of effects: item-induced inhibition of yield and growth rate of the algal population

Calculations:
- Statistics programme: ToxRatPro Version 2.10 (released 2010-09-10)
- For the calculations all algae counts were divided by a factor of 10000

Reference substance (positive control):

not required

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

78 mg/L

95% CI:

>= 60 - <= 115

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

16 mg/L

95% CI:

>= 6.4 - <= 23.7

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

21 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

45 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

21 mg/L

95% CI:

>= 11 - <= 40

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

7.7 mg/L

95% CI:

>= 0.28 - <= 13

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

9.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

21 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

Effective concentrations ranged from 99 % to 101 % of nominal values at 0 hours, and from 100 % to 104 % of nominal values at 72 hours.

Validity criteria for the measurement of the algae toxicity

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.	Test organisms: Scenedesmus subspicatus Test period: 72 h	Yes
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period.	Growth in the control flasks after 3 days showed a reproduction rate that was greater than a factor of 16. Areas ("biomass integrals") under the growth curves of Scenedesmus subspicatus also showed a minimum of 16-fold increase after 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. For test substances showing a flat concentration response curve a higher factor may be justified. The concentration series should preferably cover the range causing 5-75 % inhibition of algal growth rate.	Test concentration no.: 7 Factor of geometric series of test conc.: 2 Conc. series cover the range causing following inhibition of algal growth rate after 72 h: -14.6 - 95.0 %	Yes
The cultures should be maintained at a temperature in the range of 21 to 24°C, controlled at ± 2°C.	Incubation was at 23 ± 2°C	Yes
The pH of the control medium should not increase by more than 1.5 units during the test.	Solvent control medium pH was 8.05 at the beginning of the test and 10.10 after 72 h.	Comment in the report: At the start of the test, the solvent control cultures had a pH of 8.05. At the end of the test (72 h), the rapid growth of the algal cells had changed the pH to 10.10 units. Although slightly higher (0.55 units) than suggested in the Guidelines, it did not interfere with interpretation of the test, or determination of the toxicity values.   Yes

 

Validity criteria fulfilled:

yes

Remarks:

for details please refer to "Any other information on results incl. tables"

Conclusions:

After 72 hours of exposure NOECr, LOECr, ErC10 and ErC50 were determined as 21 mg/L, 45 mg/L, 16 mg/L and 78 mg/L, respectively. All results are expressed in terms of nominal concentrations.

Executive summary:

The toxicity to algae (Desmodesmus subspicatus) was investigated in a static test according to EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (O.J. No. L383A, 29.12.92) Part C, Method 3 "Algal inhibition test". One range finding tests preceded the main test and provided information about the range of concentrations which were used in the main test. In the main test, the algae were exposed to the test item added to dilution water at a range of concentrations for a period of 72 hours. Defined concentrations of the test item led to a certain inhibition of algal yield and growth rate at the end of the 72 hour study period. Cell densities were recorded at 24 hour intervals. The 72 hour EC 10 and EC 50 were calculated by probit analysis. During the test a temperature range of 21 - 24 °C was maintained in the test vessels. The pH was measured at the beginning of the test and after 72 hours of exposure. The maintenance of the test item concentrations was proved by analytical measurements. In order to avoid an impairment of the test system, an additional replicate was used for analysis and pH measurement at the beginning of the test. Chemical analysis and pH measurement at the end of the test. After 72 hours of exposure NOECr, LOECr, ErC10 and ErC50 were determined as 21 mg/L, 45 mg/L, 16 mg/L and 78 mg/L, respectively. All results are expressed in terms of nominal concentrations. Effective concentrations ranged from 98 % to 100 % of nominal values at 0 hours, and from 95 % to 101 % of nominal values at 72 hours. The validity criteria of the test were met and therefore this toxicity study is classified as acceptable and satisfies the guideline requirements for the Algae study.

Description of key information

After 72 hours of exposure NOECr, LOECr, ErC10 and ErC50 were determined as 21 mg/L, 45 mg/L, 16 mg/L and 78 mg/L, respectively. All results are expressed in terms of nominal concentrations.

Key value for chemical safety assessment

EC50 for freshwater algae:

78 mg/L

EC10 or NOEC for freshwater algae:

21 mg/L

Additional information

Effective concentrations ranged from 98 % to 100 % of nominal values at 0 hours, and from 95 % to 101 % of nominal values at 72 hours.