Dimethyl 2,2'-azobis(2-methylpropionate)

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:

30.05.-30.06.16

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)

Version / remarks:

2011

Deviations:

yes

Remarks:

temperature range was exceeded and undercut during testing period. As normal growth was observed, deviation is considered uncritical.

GLP compliance:

yes (incl. QA statement)

Vehicle:

no

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name:
- Strain: 86.81
- Source (laboratory, culture collection): MBM Sciencebridge GmbH (Inst. für Pflanzenphysiologie, Universität Göttingen)
- Age of inoculum (at test initiation):
- Method of cultivation:

ACCLIMATION
- Acclimation period:
- Culturing media and conditions (same as test or not):
- Any deformed or abnormal cells observed:

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

20.4-24 °C

pH:

start: 7.7 ± 0.1
end: 8.5 ± 0.1 (except highest concentration: pH 7.6)

Nominal and measured concentrations:

nominal: 1; 3.2; 10; 32; 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: flasks
- Type (delete if not applicable): open (covered with perforated plastic foil)
- Material, size, headspace, fill volume: glass, 20 mL, 45 mL
- Aeration:
- Type of flow-through (e.g. peristaltic or proportional diluter): -
- Renewal rate of test solution (frequency/flow rate):-
- Initial cells density: 4.6E3 cells/mL
- Control end cells density: 5.9E5
- No. of organisms per vessel:-
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates):-

GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used:-

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine:
- Alkalinity:
- Ca/mg ratio:
- Conductivity:
- Culture medium different from test medium:
- Intervals of water quality measurement:

OTHER TEST CONDITIONS
- Sterile test conditions: yes/no
- Adjustment of pH:
- Photoperiod: 16/8
- Light intensity and quality: 5300 Lux
- Salinity (for marine algae):

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]
- Chlorophyll measurement:
- Other:

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations: for different reasons three tests in total were performed
- Results used to determine the conditions for the definitive study:

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 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.:

14 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.:

0.7 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

16 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.69 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.7 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape:-
- Colour differences: -
- Flocculation: -
- Adherence to test vessels: -
- Aggregation of algal cells: -
- Other: -
- Any stimulation of growth found in any treatment: -
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values:-
- Effect concentrations exceeding solubility of substance in test medium:-

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50: 0.91 mg/L (0.8-1.0 mg/L)
- Other:

Reported statistics and error estimates:

ToxRat Pro 3.21 was usd for calculations.

Validity criteria fulfilled:

yes

Executive summary:

Three valid experiments were performed.

 

The first experiment was performed using 5 concentrations ranging from 10 to 100 mg/L. Significant inhibition of algal growth in the lowest concentration of 10 mg/L was observed. Therefore, the first experiment was repeated.

 

The second and third experiments were performed using 5 concentrations ranging from 1 to 100 mg/L

In the second experiment the analytical samples at the end of the test (72 hours) were not measured due to a technical error.Therefore, the second experiment was repeated.

 

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 yield were determined from the cell number at the respective observation times.

 

The biological results were based on the results of the third experiment.

Significant inhibition of algal growth in the third experiment was observed at all concentrations with the exceptions of the lowest concentrated treatment.

 

At the beginning and every 24 hours during the course the study, the content of the test item in the test solution was determined using HPLC-UV-determination. The concentrations determined at the start of the third experiment were in the range of 85 % and 98 % of the nominal concentration. At the end of the third experiment the determined concentrations were in the range of 23 % and 52 % of the nominal concentration. In the lowest concentration of 1 mg/L nominal concentration the determined concentration was below the limit of quantification at the end of the test. Therefore, the determination of the biological results was based on the geometric mean of the measured concentrations.

 

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 itemdimethyl-2,2'-azobisisobutyratewere determined:

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	0.7 mg/L	1.9 mg/L	14 mg/L	> 100 mg/L
Yield	0.7 mg/L	1.9 mg/L	0.69 mg/L	16 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.

Key value for chemical safety assessment

EC50 for freshwater algae:

100 mg/L

EC10 or NOEC for freshwater algae:

14 mg/L