4-formyl-2-methoxyphenyl isobutyrate

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:

17 July - 22 December 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Name of test material (as cited in study report): Isobutavan
- Physical state: colourless to pale yellow liquid
- Lot/batch No.: SC00010048
- Expiration date of the lot/batch: December 17, 2015
- Storage condition of test material: ambient temperature, dark

Analytical monitoring:

yes

Details on sampling:

In the main test samples were taken from all concentrations and from the control at test start (day 0) and after 24, 48 and 72 hours.

Vehicle:

no

Details on test solutions:

Based on the results of the non-GLP range finding test, a static test with nominal test item concentrations of 0, 0.954, 3.05, 9.77, 31.3 and 100 mg/L was performed.The necessary amount of test item for preparing the stock solution (S1) was weighed on a weighing scoop and transferred to a volumetric flask. Test medium was added up to the bench mark and the solution was homogenised by shaking. The necessary amount of test item for preparing the stock solution (S1) was weighed on a weighing scoop and transferred to a volumetric flask. Test medium was added up to the bench mark and the solution was homogenised by shaking. The further concentrations were made by diluting the appropriate solutions to give the required test concentrations.

Test organisms (species):

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

Details on test organisms:

The test organism was Pseudokirchneriella subcapitata Hindák, strain SAG 61.81. The algae used in this study were purchased from MBM Sciencebridge GmbH, Göttingen, Germany.
The algae were grown semi-continuously in the laboratory in liquid cultures under permanent illumination. Old medium was periodically replaced by fresh mineral solution in order to keep the algae in an exponential growth state. Stock cultures were ordered regularly from the culture collection. Cultures were kept at 21-24oC and illuminated at 60-120 μE/m2/s.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

20.7 – 22.8 °C

pH:

7.31 – 8.31

Nominal and measured concentrations:

Nominal test item concentrations were 0, 0.954, 3.05, 9.77, 31.3 and 100 mg/L. The initial measured concentration in the test solution ranged from 44 to 59 % of nominal. The measured concentration in the aged samples after 72 h ranged from 9 % to 27 % of nominal. Since the measured concentrations of the test item were outside the recommended range of 80 % and 120 %, the toxicological endpoints were evaluated using the actual concentrations based on the geometric mean of each test item concentration (Measured concentrations were 0, 0.294, 0.882, 3.49, 11.4, and 40 mg/L).

Details on test conditions:

Six replicates were used for the control and three for each test item concentration. The test was performed in 100 mL Erlenmeyer flasks under continuous illumination at 87 – 113 μE×m-2×s-1 at cell culture level and continuously shaken at 105 rpm. The final volume in each test vessel was identical in each test vessel (in the range 40-60 mL). The temperature of the test was within a range of 20.7 to 22.8 °C.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 40 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

10.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.49 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

Effects based on yield were also reported (see below and executive summary). However, the preferred observational endpoint in the algal inhibition study is growth rate because it is not dependent on the test design (ECHA guidance Chapter R.7b v1.1). The EU CLP regulation (No 1272/2008 and its adaption 286/2011) also states that classification should be based on the ErC50. Thus only the effects based on growth rate are presented in the above "effects concentration" table.

Statistically significant inhibitory effects of the test item were observed at 31.3 and 100 mg/L (nominal) for growth rate and yield. Based on the actual test item concentrations (calculated with the geometric mean of each test item concentration) statistically significant effects were evaluated at 11.4 and 40.0 mg/L. For growth rate and yield the overall LOEC was determined to be at 31.3 mg/L (nominal) and at 11.4 mg/L (actual). The overall NOEC was 9.77 mg/L (nominal) and 3.49 mg/L (actual) for growth rate and yield. The 72h-EC50 value for growth rate (ErC50) was >100 mg/L (nominal; 120 mg/L extrapolated value). Based on the actual concentrations (calculated with the geometric mean of each test item concentration) the 72h- EC50 value for growth rate (ErC50) was determined to be at > 40.0 mg/L (48.5mg/L extrapolated value). The 72h-EC50 value for yield (EyC50) was 39.6 mg/L (nominal) and 14.9 mg/L (actual). The 72h-EC10 value for growth rate (ErC50) was 27.5 mg/L (nominal) and 10.1 mg/L (actual). The 72h-EC10 for yield (EyC10) was 7.29 mg/L (nominal) and 2.49 mg/L (actual).

Results with reference substance (positive control):

The overall NOEC for potassium dichromate was determined to be 0.128 mg/L. The EC50 values for growth rate, yield, and biomass were calculated to be 1.34 mg/L, 0.588 mg/L and 0.678 mg/L respectively.

Validity criteria fulfilled:

yes

Conclusions:

Statistically significant inhibitory effects of the test item were observed at 31.3 and 100 mg/L (nominal) for growth rate and yield. Based on the actual test item concentrations (calculated with the geometric mean of each test item concentration) statistically significant effects were evaluated at 11.4 and 40.0 mg/L. For growth rate and yield the overall LOEC was determined to be at 31.3 mg/L (nominal) and at 11.4 mg/L (actual). The overall NOEC was 9.77 mg/L (nominal) and 3.49 mg/L (actual) for growth rate and yield. The 72h-EC50 value for growth rate (ErC50) was >100 mg/L (nominal; 120 mg/L extrapolated value). Based on the actual concentrations (calculated with the geometric mean of each test item concentration) the 72h-EC50 value for growth rate (ErC50) was determined to be at > 40.0 mg/L (48.5 mg/L extrapolated value). The 72h-EC50 value for yield (EyC50) was 39.6 mg/L (nominal) and 14.9 mg/L (actual). The 72h-EC10 value for growth rate (ErC50) was 27.5 mg/L (nominal) and 10.1 mg/L (actual). The 72h-EC10 for yield (EyC10) was 7.29 mg/L (nominal) and 2.49 mg/L (actual).

The preferred observational end point in the algal growth inhibition test is growth rate because it is not dependent on the test design (ECHA guidance Chapter R.7b v1.1). The EU CLP regulation (No 1272/2008 and its adaption 286/2011) also states that classification should be based on the ErC50. The preferred observational endpoint in long-term studies is the EC10 value because it is derived from the dose response curve. In contrast the NOEC strongly depends on the experiment design (e.g. the concentrations used in the test). Thus the 72-h EC50 and EC10 based on growth rate are used for classification purposes, which were determined in this study to be > 40 mg/L and 10.1 mg/L respectively.

Executive summary:

An OECD 201 algal inhibition test was conducted with the test substance Isobutavan. The test species used wasPseudokirchneriella subcapitata. Based on the results of a non-GLP range finding test, a static test with nominal test item concentrations of 0, 0.954, 3.05, 9.77, 31.3 and 100 mg/L was performed. Six replicates were used for the control and three for each test item concentration. The test was performed in 100 mL Erlenmeyer flasks under continuous illumination at 87 – 113 μE×m^-2×s^-1at cell culture level and continuously shaken at 105 rpm. The final volume in each test vessel was identical in each test vessel (in the range 40-60 mL). The temperature of the test was within a range of 20.7 to 22.8 °C. After 1, 2 and 3 days, the cell density was determined by fluorescence detection.

Statistically significant inhibitory effects of the test item were observed at 31.3 and 100 mg/L (nominal) for growth rate and yield. Based on the actual test item concentrations (calculated with the geometric mean of each test item concentration) statistically significant effects were evaluated at 11.4 and 40.0 mg/L. For growth rate and yield the overall LOEC was determined to be at 31.3 mg/L (nominal) and at 11.4 mg/L (actual). The overall NOEC was 9.77 mg/L (nominal) and 3.49 mg/L (actual) for growth rate and yield. The 72h-EC₅₀value for growth rate (ErC₅₀) was >100 mg/L (nominal; 120 mg/L extrapolated value). Based on the actual concentrations (calculated with the geometric mean of each test item concentration) the 72h-EC₅₀value for growth rate (ErC₅₀) was determined to be at > 40.0 mg/L (48.5 mg/L extrapolated value). The 72h-EC₅₀value for yield (EyC₅₀) was 39.6 mg/L (nominal) and 14.9 mg/L (actual). The 72h-EC10 value for growth rate (ErC50) was 27.5 mg/L (nominal) and 10.2 mg/L (actual). The 72h-EC10 for yield (EyC10) was 7.29 mg/L (nominal) and 2.49 mg/L (actual).

Description of key information

ErC₅₀ > 40.0 mg/L (growth rate), 48.5 mg/L extrapolated value

ErC10 = 10.1 mg/L (growth rate)

Key value for chemical safety assessment

EC50 for freshwater algae:

48.5 mg/L

EC10 or NOEC for freshwater algae:

10.1 mg/L

Additional information

An OECD 201 algal inhibition test was conducted with the test substance Isobutavan. The test species used wasPseudokirchneriella subcapitata. Based on the results of a non-GLP range finding test, a static test with nominal test item concentrations of 0, 0.954, 3.05, 9.77, 31.3 and 100 mg/L was performed. Six replicates were used for the control and three for each test item concentration. The test was performed in 100 mL Erlenmeyer flasks under continuous illumination at 87 – 113 μE×m^-2×s^-1at cell culture level and continuously shaken at 105 rpm. The final volume in each test vessel was identical in each test vessel (in the range 40-60 mL). The temperature of the test was within a range of 20.7 to 22.8 °C. After 1, 2 and 3 days, the cell density was determined by fluorescence detection.

Statistically significant inhibitory effects of the test item were observed at 31.3 and 100 mg/L (nominal) for growth rate and yield. Based on the actual test item concentrations (calculated with the geometric mean of each test item concentration) statistically significant effects were evaluated at 11.4 and 40.0 mg/L. For growth rate and yield the overall LOEC was determined to be at 31.3 mg/L (nominal) and at 11.4 mg/L (actual). The overall NOEC was 9.77 mg/L (nominal) and 3.49 mg/L (actual) for growth rate and yield. The 72h-EC₅₀value for growth rate (ErC₅₀) was >100 mg/L (nominal; 120 mg/L extrapolated value). Based on the actual concentrations (calculated with the geometric mean of each test item concentration) the 72h-EC₅₀value for growth rate (ErC₅₀) was determined to be at > 40.0 mg/L (48.5 mg/L extrapolated value). The 72h-EC₅₀value for yield (EyC₅₀) was 39.6 mg/L (nominal) and 14.9 mg/L (actual). The 72h-EC10 value for growth rate (ErC50) was 27.5 mg/L (nominal) and 10.1 mg/L (actual). The 72h-EC10 for yield (EyC10) was 7.29 mg/L (nominal) and 2.49 mg/L (actual).

The preferred observational end point in the algal growth inhibition test is growth rate because it is not dependent on the test design (ECHA guidance Chapter R.7b v1.1). The EU CLP regulation (No 1272/2008 and its adaption 286/2011) also states that classification should be based on the ErC50. The preferred observational endpoint in long-term studies is the EC10 value because it is derived from the dose response curve. In contrast the NOEC strongly depends on the experiment design (e.g. the concentrations used in the test). Thus the 72-h EC50 and EC10 based on growth rate are used for classification purposes, which were determined in this study to be > 40 mg/L (48.5 mg/L extrapolated value) and 10.1 mg/L respectively.