6H-Benzofuro[3a,3,2-ef][2]benzazepin-6-one, 1-bromo-4a,5,9,10,11,12-hexahydro-3-methoxy-11-methyl-, (4aR,8aR)-rel-

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001-11-16 to 2002-02-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

439-750-2

EC Name:

-

Cas Number:

183487-56-1

Molecular formula:

Hill formula: C17H19BrClNO3 CAS formula: C17H19BrClNO3

IUPAC Name:

(4aR*,8aR*)-1-bromo-4a,5,9,10,11,12- hexahydro-3-methoxy-11-methyl-6H-ben zofuro[3a,3,2-ef][2]benzazepin-6-one hydrochloride

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: RT002113G3A231
- Expiration date of the lot/batch: 2001-12-01 (retest date)
- Purity test date: no data

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature at about 20°C, away from direct sunlight
- Stability under test conditions: Determined in RCC study 825265
- Solubility and stability of the test substance in the solvent/vehicle: in water 120 g/L (non-GLP, sponsor's information; determined in RCC study 825030)

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: samples were taken before the start of the test and after 72 hours from each test medium and from the control (all without algae). The concentration of the test item T002113 was analyzed in the duplicate test medium samples. From the control samples only one of the duplicate samples was analyzed.
- Sample storage conditions before analysis: To prevent degradation of the test item after sampling and until analysis, the samples were acidified to about pH 4-5 immediately after sampling using a 0.1 M phosphoric acid solution. The samples were analyzed immediately after sampling without prior storage.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: the highest nominal test item concentration of 100 mg/L was prepared by dissolving 60 mg T002113 in 600 mL of test water by intense stirring (5 minutes at room temperature). The lower test item concentrations were prepared by dilution. The test media were prepared just before addition of algae.
- Controls: yes

Test organisms

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Strain:n° 61.81 SAG
- Source (laboratory, culture collection): Collection of Algal Cultures (SAG, Institute for Plant Physiology, University of Göttingen, D-37073 Göttingen, Germany)
- Method of cultivation: in the RCC laboratories under standardized conditions according to the test guideline (OECD 201). The algae were cultivated and tested in synthetic test water, prepared according to the test
guidelines. Analytical grade salts were dissolved in sterile purified water to obtain the following final nominal concentrations:
* Macro-nutrients: NaHCO3 50.0 mg/L
CaCI2 x 2 H2O 18.0 mg/L
NH4CI 15.0 mg/L
MgS04 x 7 H2O 15.0 mg/L
MgCl2 X 6 H2O 12.0 mg/L
KH2P04 1.6 mg/L
* Trace elements: Na2EDT A x 2 H2O 100.0 µg/L
FeCl3 x 6 H2O 80.0 µg/L
MnCI2 x 4 H2O 415.0 µg/L
H3BO3 185.0 µg/L
Na2Mo04 x 2 H2O 7.0 µg/L
ZnCl2 3.0 µg/L
CoCl2 x 6 H2O 1 .5 µg/L
CuCI2 x 2 H2O 0.01 µg/L

ACCLIMATION
- Acclimation period: not relevant (except pre-culture 3 days before start of the test)
- Culturing media and conditions (same as test or not): same as the test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Hardness:

24 mg CaCO3/L

Test temperature:

23°C

pH:

At the start of the test: 7.5-7.9
At the end of the test: 7.7 - 8.2

Dissolved oxygen:

not reported

Salinity:

not applicable

Nominal and measured concentrations:

nominal test concentrations: control, 4.6, 10, 22, 46, 100 mg/L
day 0: mean measured concentrations: -, 4.52, 10.2, 22.9, 46.7, 96.7 mg/L
day 3: mean measured concentrations: -, 2.36, 5.48, 12.3, 25.2, 48.4 mg/L

The mean measured concentrations (calculated as the average over all measurements per test concentration) ranged from 73 to 80% of nominal.

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Type (delete if not applicable): covered with glass dishes
- Material, size, headspace, fill volume: 50 mL Erlenmeyer flasks filled with 15 mL algal suspension
- Initial cells density: 10,000 cells/mL
- Control end cells density: 116 x 10,000 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes (OECD 201)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: synthetic test water
- Culture medium different from test medium: no
- Intervals of water quality measurement: temperature: daily, pH at the begin and the end of the test

OTHER TEST CONDITIONS
- Sterile test conditions: no information
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: mean of 9100 lux (range: 8460-9510 lux) by fluorescent tubes (Philips TLD 36W/840)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic particle counter
- effect calculated parameters: specific growth rate and biomass (area under the growth curve)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2 to 2.2
- Range finding study
- Test concentrations:data not shown
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

potassium dichromate (test once a year)

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

42 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 23-158

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.4 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

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: clear test medium
- EC50 (biomass) = 10.4mg T002113 /L.
- EC10 (growth rate) = 6.7 mg T002113 /L.
- EC10 (biomass) = 3.2 mg T002113 / L.
- EC90 (growth rate) > 73 mg T002113 / L.
- EC90 (biomass) = 33 mg T002113 / L.
- LOEC (growth rate and biomass) = 7.9 mg T002113 / L.
- NOEC (growth rate and biomass) = 3.4 mg T002113 / L.

Results with reference substance (positive control):

- Results with reference substance valid? no data in the report

Reported statistics and error estimates:

EC50 (biomass and growth rate) and corresponding EC10, EC90 values and their 95% confidence intervals were calculated by Probit analysis.
NOEC and LOEC values were determined using a Dunnett-test

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

A 72-h growth inhibition test with the unicellular green alga Pseudokirchnerielle subcapitata was performed with the test substance T 2113 according to the OECD 201 guideline. The results were based on mean measured concentrations. The 72-h EC50 (growth rate) was determined to be 42 mg T002113/L. The 72-h NOEC was dtermined to be 3.4 mg T002113/L. The results of the test can be considered reliable without restrictions.