2,3-dihydrothieno[3,4-b][1,4]dioxine

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:

26.2.-11.3.2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was carried out in accordance with internationally valid GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes

Test material

Reference

Name:

Unnamed

Type:

Constituent

Details on test material:

- Name of test material (as cited in study report): 3,4-Ethylenedioxythiophene
- Comercial name. EDOT
- Substance type: pure organic substance
- Physical state: liquid near colourless to pale yellow
- Analytical purity: > 99%
- Lot/batch No.: 091201
- Expiration date of the lot/batch: 21.1.2011
- Storage condition of test material: Keep in dark at room temperature. Stored in amber glass bottles under the argon atmosphere. Minimalize time for handling with substance without protective argon atmosphere as possible. Maximal time without protective atmosphere is 30 min.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 14, 100 mg/L
- Sample storage conditions before analysis: The samples for analysis (0 hours) were prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. An analysis of samples was started immediately after delivery. The samples for analysis at the end of the test (72 hours) were delivered to analytical laboratory immediately after the end of testing. An analysis of samples was started immediately after delivery. All samples were stored at laboratory temperature.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The stock solution of the test substance was prepared in the test medium. 100 mg of the test substance was weighed into 1000 mL of the test medium for the preliminary test and the full test. The stock solution was stirred for 30 min. on a magnetic stirrer before the using. The concentrations of solutions used in the preliminary and the full test were obtained by dilution of the stock solution test medium.

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Strain: Brinkmann 1953/SAG 86.81
- Source (laboratory, culture collection): the collection of autotrophic organisms of The Botanic Institute of the Czech Academy of Science, Třeboň
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: The strain culture was preinoculated from the stock solution and cultivated in flasks with the test medium on indirect daylight at laboratory temperature.

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions: same as test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Test temperature:

23.5-24.5°C

pH:

7.7

Nominal and measured concentrations:

Nominal concentration: 100, 71, 51, 36, 26, 19, 14 mg/L
Measured concentration: See Table 2

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks with stoppers venting air
- Type: open
- Aeration: no
- Initial cells density: 10000 cells in 1 mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The water with conductivity smaller than 5 μS.cm-1 was used for the preparation of solutions. According to Method C.2 – Daphnia sp. Acute Immobilisation Test were prepared four basic stock solutions. The test medium was prepared by admixing of basic stock solution.

OTHER TEST CONDITIONS
- Lighting: continuous, min 6000 lux, max. 10000 lux


EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: The density of algae culture was evaluated microscopically at 24, 48 and 72 hours. The cell density was measured by direct counting of living cells in Burker´s counting chamber.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.4

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

other: EbC50

Effect conc.:

41.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 21.9-60.3 mg/L

Effect concentrations 2

Duration:

72 h

Dose descriptor:

other: ErC50

Effect conc.:

64 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 53.2-74.8 mg/L

Effect concentrations 3

Duration:

72 h

Dose descriptor:

other: NOECb

Effect conc.:

36 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 4

Duration:

72 h

Dose descriptor:

other: NOECr

Effect conc.:

36 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Results with reference substance (positive control):

- Results with reference substance valid: yes. The inhibition of test organisms in the reference substance solution meets criteria determined by the interlaboratory tests

72 hour – ErC50 = 0.86 mg.L-1 (95% confidence limit: 0.84 – 0.88 mg.L-1)
72 hour – EbC50 = 0.32 mg.L-1 (95% confidence limit: 0.31 – 0.33 mg.L-1)

Any other information on results incl. tables

TEST EVALUATION

The values of of EbC50 and ErC50 were calculated by means of software “Toxicita”(product of Water Research Institute, Ostrava, Czech Republic).

The determination of NOEC values was done by ANOVA (Analysis of Variance) analysis. Used ANOVA method is the part of statistical software QC.Expert 2.5 © 1998-2000 (TriloByte, Czech Republic).

For the evaluation EC and NOEC values the nominal concentrations were used.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Executive summary:

The test substance, 3,4-Ethylenedioxythiophene, was tested for growth inhibition on algae Desmodesmus subspicatus.

The test was performed according to method C.3 - Algal Inhibition Test, Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008.

The preliminary test was performed in range of test substance nominal concentrations 1 – 100 mg.L-1. The analytical results showed, that the test substance 3,4 Ethylenedioxythiophene was stable in the test medium in conditions of the test. Based on the toxicity of the test substance found in preliminary test, the full test was performed in appropriate concentration range.

The full test was performed in range of test substance nominal concentrations 14 – 100 mg.L-1. Samples for analytical determination were taken at the beginning and at the end of the test. The analytical results showed that the test substance 3,4 Ethylenedioxythiophene was stable in test medium in conditions of the test, therefore the nominal concentrations were used for all evaluations and results.

Test results:

72 hour - EbC50 = 41.1 mg.L-1 (nominal concentration) (95% confidence limit: 21.9 – 60.3 mg.L-1)

72 hour - ErC50 = 64.0 mg.L-1 (nominal concentration) (95% confidence limit: 53.2 – 74.8 mg.L-1)

72 hour - NOECb = 36 mg.L-1 (nominal concentration)

72 hour - NOECr = 36 mg.L-1 (nominal concentration)