4,4-dimethyl-3,5,8-trioxabicyclo[5.1.0]octane

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Apr 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

92/69/EEC

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

1993

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: For the stock solution 127.5 mg of the test substance were dissolved in 1 L demineralized water. This solution was used for the highest test concentration of 100 mg/L (nominal concentration). To achieve the other test concentrations the stock solution was diluted.
- Controls: yes, blank control
- Test concentration separation factor: 10

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Source: Sammlung von Algenkulturen, University Göttingen
- Age of inoculum (at test initiation): pre-culture taken 72 hours before test initiation from an stock culture up to 7 days old.
- Method of cultivation: the stock culture was dissolved in a liquid nutrient solution and subsequently transferred approximately once weekly.

ACCLIMATION
- Acclimation period: The pre-culture was incubated for 72 hours in an incubator shaker apparatus at approximately 23 °C. 'The same culturing media was used for pre-culture and test inoculum (nutrient solution diluted with demineralized water, stirred under aeration for ca. 1 hour and filtered under sterile conditions).
- Culturing media and conditions: different nutrient media used for stock culture and pre-culture/test inoculum.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

20.4 to 23.4 °C

pH:

8.0 to 8.2

Nominal and measured concentrations:

0.1, 1, 10, 100 mg/L (nominal test concentration)

Details on test conditions:

TEST SYSTEM
- Test vessel: incubator shaker
- Material, size, headspace, fill volume: 100 mL
- Initial cells density: 10000 cells/mL
- Control end cells density: 924000 cells/mL
- No. of vessels per concentration (replicates): 3 replicates
- No. of vessels per control (replicates): 6 replicates

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: yes
- Intervals of water quality measurement: pH-value and temperature were measured at 0 and 72 hours of exposure.

OTHER TEST CONDITIONS
- Sterile test conditions: nutrient solution was filtered under sterile conditions before use
- Photoperiod: continuous light

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: 'Bürker'-counting chamber, electronic particle counting apparatus 'Coulter Counter', observation interval 24 hours.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Test concentrations: 0.1, 1, 10, 100 mg/L (nominal concentration)

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Details on results:

- Exponential growth in the control (for algal test): yes
- Any stimulation of growth found in any treatment: no

Validity criteria for the measurement of the algae toxicity

Target condition according to guideline:	Actual condition according to the study:	Validity criteria met:
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period. This corresponds to a specific growth rate of 0.92 day^-1. This criterion may not be met when species that grow slower than those listed in Annex 2 of OECD TG 201 are used. In this case, the test period should be extended to obtain at least a 16-fold growth in control cultures, while the growth has to be exponential throughout the test period. The test period may be shortened to at least 48 hours to maintain unlimited, exponential growth during the test as long as the minimum multiplication factor of 16 is reached.	During the test, the biomass in the control increased exponentially by a factor >16  and the specific growth rate was 1.51 day^-1 within the 72-hour test period.	Yes
The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures must not exceed 35%. This criterion applies to the mean value of coefficients of variation calculated for replicate control cultures.	The mean coefficent of variation for section-by-section specific growth rates in the control was <6%	yes
The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% in tests with Pseudokirchneriella subcapitata and Desmodesmus subspicatus. For other less frequently tested species, the value should not exceed 10%.	The coefficent of variation of average specific growth rates (0-72h) in the control was <1%	Yes
The cultures should be maintained at a temperature in the range of 21 to 24°C, controlled at ± 2°C.	During the test, cultures were maintained at a temperature range of 20.4 to 23.4°C. The temperature un the incubator chamber were slightly below the recommended range. However, as temperature were only 0.6°C below the recommended range and di not cary by mor thean +/- 2°C this deciation should not have had any influence on the test results.	Yes

Validity criteria fulfilled:

yes

Remarks:

See 'Any other information on results incl. tables'.

Conclusions:

The test substance had no inhibitory effect on the growth of Desmodesmus subspicatus (Scenedesmus subspicatus).

Executive summary:

A study was performed to determine the effects of the test substance on the growth of the green freshwater algae Desmodesmus subspicatus (Scenedesmus subspicatus). The study was conducted in accordance with the OECD Guideline for Testing of Chemicals No 201 'Alga, Growth Inhibition Test' and the EC guideline, EC Directive 92/69/EEC, Annex C3, the 'Algae inhibition test'. The test substance was incubated in an aqueous solution including nutrients for test duration of 72 hours. The test concentrations were 0, 0.1, 1, 10, and 100 mg/L (nominal concentrations). The cell densities were measured at 24 hour intervals. The calculated biomass and growth rate of each concentration were compared to those of the controls and the inhibition was calculated. According to the measurements, no growth inhibition was observed, and the test substance had no inhibitory effect on algae growth at test substance concentrations up to 100 mg/L (nominal concentration).

Description of key information

A study was performed to determine the effects of the test substance on the growth of the green freshwater algae Desmodesmus subspicatus (Scenedesmus subspicatus). The study was conducted in accordance with the OECD Guideline for Testing of Chemicals No 201 'Alga, Growth Inhibition Test' and the EC guideline, EC Directive 92/69/EEC, Annex C3, the 'Algae inhibition test'. The test substance was incubated in an aqueous solution including nutrients for test duration of 72 hours. The test concentrations were 0, 0.1, 1, 10, and 100 mg/L (nominal concentrations). The cell densities were measured at 24 hour intervals. The calculated biomass and growth rate of each concentration were compared to those of the controls and the inhibition was calculated. According to the measurements, no growth inhibition was observed, and the test substance had no inhibitory effect on algae growth at test substance concentrations up to 100 mg/L (nominal concentration).