[(2,4-dibromophenoxy)methyl]oxirane

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:

June - August 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: CG272
- Expiration date of the lot/batch: 11. July 2018
- Purity test date: not stated

RADIOLABELLING INFORMATION (if applicable)
not applicable

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- Stability under test conditions: assumed stable
- Solubility and stability of the test substance in the solvent/vehicle: 68.3 mg/L solubility, assumed stable
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not applicable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
none

Analytical monitoring:

yes

Details on sampling:

Sample Preparation
Twofold enrichment
20 mL of a solution in matrix was mixed with 3 g NaCl, then the solution was extracted 2
times with the solvent i-hexane (2 x 4 mL) and the organic phase was collected into a
10 mL flask, after drying with Na2SO4. The flask was filled up to 10 mL with i-hexane after
addition of 500 μL ISTD stock solution (1,2,4-trichlorbenzol 1000 mg/L in i-hexane) and the
solution was measured via GC/FID. Twofold enrichment was achieved during extraction.
This procedure was performed for 32 mg/L and 100 mg/L nominal concentration.
Tenfold enrichment
100 mL of a solution in matrix was mixed with 6 g NaCl, then the solution was extracted
two times with the solvent i-hexane (2 x 4 mL) and the organic phase was collected into a
10 mL flask, after drying with Na2SO4. The flask was filled up to 10 mL with i-hexane after
addition of 500 μL ISTD stock solution (1,2,4-trichlorbenzol 1000 mg/L in i-hexane) and the
solution was measured via GC/FID. Tenfold enrichment was achieved during extraction.
This procedure was performed for the Blank control, 1.0 mg/L, 3.2 mg/L and 10 mg/L nominal
concentration.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
A saturated solution was prepared for the test. This was done by mixing the nominal load
of 100.1 mg/L with the corresponding amount of algal medium (demineralised water enriched
with minerals but without algae) and shaking vigorously for 24 hours. The resulting
solution was filtrated through 0.45 μm PTFE filters.
The lower treatments were prepared by dilution of this saturated solution with nutrient medium.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Genus: Desmodesmus
- Species: subspicatus
- SAG Strain Number: 86.81
- Taxonomic position: Chlorophyta - Chlorophyceae
- Source (laboratory, culture collection): MBM Sciencebridge
GmbH (Institut für Pflanzenphysiologie of Universität Göttingen)
- Age of inoculum (at test initiation): ca. 18 month
- Method of cultivation: The algae are
kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture
was prepared. From an aliquot of the permanent culture, the pre-culture was prepared.

ACCLIMATION
not applicable

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

none

Post exposure observation period:

none

Hardness:

not stated

Test temperature:

21.3 – 22.9 °C

pH:

7.7 - 8.7

Dissolved oxygen:

not stated

Salinity:

not stated

Conductivity:

not stated

Nominal and measured concentrations:

Nominal Concentration Test Item Geometric Mean
mg/L mg/L
Blank control --
1 0.49*
3.2 1.70
10 5.22
32 16.74
100 55.24

* postulated concentration at 72 hours (based on LOD)

Details on test conditions:

TEST SYSTEM
- Test vessel: glass flasks total volume 65 mL
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: see above
- Aeration: see above
- Type of flow-through (e.g. peristaltic or proportional diluter): not applicable
- Renewal rate of test solution (frequency/flow rate): not applicable
- Initial cells density: 2*10^3 cells/mL
- Control end cells density: 534010
- No. of organisms per vessel: not applicable
- No. of vessels per concentration (replicates):3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): not stated

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionised water
- Total organic carbon: not stated
- Particulate matter: not stated
- Metals: not stated
- Pesticides: not stated
- Chlorine: according to Guideline
- Alkalinity: according to Guideline
- Ca/mg ratio: according to Guideline
- Conductivity: according to Guideline
- Culture medium different from test medium: no
- Intervals of water quality measurement: not stated

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: contineous
- Light intensity and quality: 7500 Lux, constant
- Salinity (for marine algae): not applicable

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic particle counter
- Chlorophyll measurement: no
- Other: daily measurment

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Justification for using less concentrations than requested by guideline: not applicable
- Range finding study not performed

Reference substance (positive control):

yes

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

< 0.49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

< 0.49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

<= 0.49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

<= 0.49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

ca. 0.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

ca. 0.31 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

ca. 2.65 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

ca. 0.83 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
- Observation of abnormalities (for algal test): no
- Unusual cell shape: no
- Colour differences: no
- Flocculation: no
- Adherence to test vessels: no
- Aggregation of algal cells: no
- Other: no cells observed in high doses
- 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: no

Results with reference substance (positive control):

Parameter Value 95% confidence interval
72h ErC50 0.94 mg/L 0.90 - 0.98 mg/L
72h EyC50 0.42 mg/L 0.39 - 0.45 mg/L

Reported statistics and error estimates:

Calculation of results was performed with the help of validated software. Calculation was
performed with Microsoft Excel® and ToxRat® Professional, version 3.2.1. see attached document

Validity criteria fulfilled:

yes

Conclusions:

The following results for the test item DENACOL EX-147 were determined:
Endpoint              NOEC       LOEC              EC10       EC50
Growth Rate       < 0.49 mg/L ≤ 0.49 mg/L    0.80 mg/L 2.65 mg/L
Yield                   < 0.49 mg/L ≤ 0.49 mg/L 0.31 mg/L 0.83 mg/L

Executive summary:

One valid experiment was performed.

The study was performed using 5 concentrations ranging from 1 to 100 mg/L. Incubation

time (test system Desmodesmus subspicatus) was 72 hours. The cell concentration of

each replicate was determined by measuring the cell numbers every 24 hours with an

electronic particle counter. Growth rate μ and the yield1 were determined from the cell

number at the respective observation times.

Significant inhibition of algal growth was observed in each tested concentration.

At the start and at the end of the test, the content of the test item in the test solutions was

determined using GC/FID. The measured concentrations lay between 45 % and 89 % of

the nominal concentrations. For the lowest concentration of 1 mg/L no test item was detectable

at the end of the exposure period. According to the OECD guidance document no.

23, GUIDANCE DOCUMENT ON AQUATIC TOXICITY TESTING OF DIFFICULT SUBSTANCES

AND MIXTURES, the absent test item concentration was replaced by the limit

of detection (LOD).

The measured concentrations in the replicates without algal cells were slightly higher than

in the replicates with algal cells. Nevertheless the concentrations were in the same range.

Therefore, the determination of the results was based on the geometric mean of the

measured and partly postulated (see above) concentrations in the replicates including algal

cells.

The 72h-EC50s of potassium dichromate (K2Cr2O7, CAS No. 7778-50-9) were determined

in a separate reference test. The values lay within the range of the laboratory (growth rate

0.73 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

The following results for the test item DENACOL EX-147 were determined:

Results of the test item

Endpoint              NOEC       LOEC              EC10       EC50

Growth Rate       < 0.49 mg/L ≤ 0.49 mg/L    0.80 mg/L 2.65 mg/L

Yield                   < 0.49 mg/L ≤ 0.49 mg/L 0.31 mg/L 0.83 mg/L

Note: According to the guideline, NOEC is determined by comparing of the respective treatment with

the blank control. Statistically insignificant variation is considered as “no observed effect”, although

the EC10 which is read from the graph toxicity vs. concentration may lie lower.

Description of key information

one Guideline study available.

For chemical assessment, the Effect concentrations on Yield are used as key values.

Key value for chemical safety assessment

EC50 for freshwater algae:

0.83 mg/L

EC10 or NOEC for freshwater algae:

0.31 mg/L