N-phenylprop-2-enamide

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

March 23, 2006

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

December 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Version / remarks:

01 October 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling.
- Sampling method:
Frequency at t=0 h, t= 24h and t= 48h
Volume 1.5 mL
- Sample storage conditions before analysis: Samples were stored in a freezer until analysis at the analytical laboratory of the Test Facility.

Compliance with the Quality criteria regarding maintenance of actual concentrations was checked by running a test vessel at the highest test item concentration but without algae and samples for analysis were taken at the start, after 24h and at the end of the test period.

Vehicle:

no

Details on test solutions:

The batch of CH03543 tested was a white powder with a purity of 99.3% which was completely soluble in test medium at the concentrations tested.

Preparation of test solutions started with the highest concentration of 100 mg/L applying a 10‑minute treatment with ultrasonic waves followed by maximum 18 minutes of magnetic stirring to accelerate dissolution of the test item in medium. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure.

After preparation, volumes of 50 mL were added to each replicate of the respective test concentration. Subsequently, 1 mL of an algal suspension was added to each replicate providing a cell density of 10^4 cells/mL.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Strain: NIVA CHL 1
- Source (laboratory, culture collection): In-house laboratory culture.
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light in a climate room at a temperature of 21-24°C.

ACCLIMATION
3 days before the start of the test, cells from the algal stock culture were inoculated in culture medium at a cell density of 1 x 10^4 cells/mL. The pre-culture was maintained under the same conditions as used in the test. The cell density was measured immediately before use.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test temperature:

21.5°C-23.7°C

pH:

7.9-8.2

Nominal and measured concentrations:

Analysis of the samples taken during the final test showed that measured concentrations were stable and in agreement with nominal during the test period (98-110%).
nominal concentrations: 0, 010, 18, 32, 56, 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL, all-glass, containing 50 mL of test solution
- Static
- aeration: continuous
- Initial cells density: 1 x 10^4 cells/mL
- Control end cells density: 55.5 x 10^4 cells/mL
- No. of organisms per vessel: 1 x 10^4 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): not applicable

GROWTH MEDIUM
- Standard medium used: yes, M2

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuous (24h)
- Light intensity and quality: Cool-whie TLD-lamps of 30W with a light intensity within the range of 86 to 92 µm

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
pH At the beginning and at the end of the test.
Temperature of medium Continuously in a temperature control vessel.
Appearance of the cells At the end of the final test, microscopic observations were performed on the highest test concentration to observe for any abnormal appearance of the algae.
At the beginning of the test, cells were counted using a microscope and a counting chamber. Thereafter, cell densities were determined by spectrophotometric measurement of samples at 680 nm using a spectrophotometer with immersion probe (pathlength =20 mm for the combined limit/range-finding test; pathlength=10 mm for the final test). Algal medium was used as blank.


TEST CONCENTRATIONS
- Range finding study: 0.1, 1, 10 and 100 mg/L
- Test concentrations: Six replicates of exponentially growing algae were exposed to a control and a concentration of 100 mg/L.
- Results used to determine the conditions for the definitive study: yes, 1.0, 3.2, 10, 32, 56 and 100 mg/L in the final test

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

< 10 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

< 10 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

58 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Remarks on result:

other: 54-61mg/L

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

30 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Remarks on result:

other: 22-40mg/L

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

28 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Remarks on result:

other: 27-30mg/L

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

15 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Remarks on result:

other: 11-20mg/L

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50:
growth rate: EC50 = 2.1 mg/L (1.5-3.0 mg/L)
yield inhibition: EC50 = 0.85 mg/L (0.5-1.5 mg/L)
- Other:

Validity criteria fulfilled:

yes

Conclusions:

A OECD 201 study with Pseudokirchneriella subcapitata was performed for CH03543. The EC50 for growth rate was 58 mg/L (54-61 mg/L); the EC50 for yield inhibition was 30 mg/L (22-40 mg/L).

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

See attached Read-across document.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

March 23, 2006

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

December 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Version / remarks:

01 October 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling.
- Sampling method:
Frequency at t=0 h, t= 24h and t= 48h
Volume 1.5 mL
- Sample storage conditions before analysis: Samples were stored in a freezer until analysis at the analytical laboratory of the Test Facility.

Compliance with the Quality criteria regarding maintenance of actual concentrations was checked by running a test vessel at the highest test item concentration but without algae and samples for analysis were taken at the start, after 24h and at the end of the test period.

Vehicle:

no

Details on test solutions:

The batch of CH03543 tested was a white powder with a purity of 99.3% which was completely soluble in test medium at the concentrations tested.

Preparation of test solutions started with the highest concentration of 100 mg/L applying a 10‑minute treatment with ultrasonic waves followed by maximum 18 minutes of magnetic stirring to accelerate dissolution of the test item in medium. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure.

After preparation, volumes of 50 mL were added to each replicate of the respective test concentration. Subsequently, 1 mL of an algal suspension was added to each replicate providing a cell density of 10^4 cells/mL.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Strain: NIVA CHL 1
- Source (laboratory, culture collection): In-house laboratory culture.
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light in a climate room at a temperature of 21-24°C.

ACCLIMATION
3 days before the start of the test, cells from the algal stock culture were inoculated in culture medium at a cell density of 1 x 10^4 cells/mL. The pre-culture was maintained under the same conditions as used in the test. The cell density was measured immediately before use.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test temperature:

21.5°C-23.7°C

pH:

7.9-8.2

Nominal and measured concentrations:

Analysis of the samples taken during the final test showed that measured concentrations were stable and in agreement with nominal during the test period (98-110%).
nominal concentrations: 0, 010, 18, 32, 56, 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL, all-glass, containing 50 mL of test solution
- Static
- aeration: continuous
- Initial cells density: 1 x 10^4 cells/mL
- Control end cells density: 55.5 x 10^4 cells/mL
- No. of organisms per vessel: 1 x 10^4 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): not applicable

GROWTH MEDIUM
- Standard medium used: yes, M2

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuous (24h)
- Light intensity and quality: Cool-whie TLD-lamps of 30W with a light intensity within the range of 86 to 92 µm

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
pH At the beginning and at the end of the test.
Temperature of medium Continuously in a temperature control vessel.
Appearance of the cells At the end of the final test, microscopic observations were performed on the highest test concentration to observe for any abnormal appearance of the algae.
At the beginning of the test, cells were counted using a microscope and a counting chamber. Thereafter, cell densities were determined by spectrophotometric measurement of samples at 680 nm using a spectrophotometer with immersion probe (pathlength =20 mm for the combined limit/range-finding test; pathlength=10 mm for the final test). Algal medium was used as blank.


TEST CONCENTRATIONS
- Range finding study: 0.1, 1, 10 and 100 mg/L
- Test concentrations: Six replicates of exponentially growing algae were exposed to a control and a concentration of 100 mg/L.
- Results used to determine the conditions for the definitive study: yes, 1.0, 3.2, 10, 32, 56 and 100 mg/L in the final test

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

< 9 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

< 9 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

53 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Remarks on result:

other: 49-56 mg/L

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

27 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Remarks on result:

other: 20-37 mg/L

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

26 mg/L

Nominal / measured:

nominal

Basis for effect:

growth rate

Remarks on result:

other: 25-27 mg/L

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

14 mg/L

Nominal / measured:

nominal

Basis for effect:

biomass

Remarks on result:

other: 10-18 mg/L

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50:
growth rate: EC50 = 2.1 mg/L (1.5-3.0 mg/L)
yield inhibition: EC50 = 0.85 mg/L (0.5-1.5 mg/L)
- Other:

Validity criteria fulfilled:

yes

Remarks:

ba

Conclusions:

Based on the read-across from a structural analogue (CH03543), the EC50 (growth rate) for CH03859 was 53 mg/L

Description of key information

Read-across was performed based on a structural analogue.

Key value for chemical safety assessment

EC50 for freshwater algae:

53 mg/L

EC10 or NOEC for freshwater algae:

26 mg/L