2H-1-Benzopyran-2-methanol, alpha,alpha'-[[(phenylmethyl)imino]bis(methylene)]bis[6-fluoro-3,4-dihydro-, (alphaR,alpha'R,2R,2'S)-rel-

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:

2015-10-23 to 2015-10-31

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2H-1-Benzopyran-2-methanol, alpha,alpha'-[[(phenylmethyl)imino]bis(methylene)]bis[6-fluoro-3,4-dihydro-, (alphaR,alpha'R,2R,2'S)-rel-

EC Number:

618-043-4

Cas Number:

876666-07-8

Molecular formula:

C29H31F2NO4

IUPAC Name:

2H-1-Benzopyran-2-methanol, alpha,alpha'-[[(phenylmethyl)imino]bis(methylene)]bis[6-fluoro-3,4-dihydro-, (alphaR,alpha'R,2R,2'S)-rel-

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): JNJ-17303806-AAA (T001586)
- Physical state: solid, powder
- Appearance: white, beige

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: I15EB1984
- Expiration date of the lot/batch: 2016-11-08 (retest date)
- Purity test date: 2015-05-19

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature
- Stability under test conditions: not indicated
- Solubility and stability of the test substance in the solvent/vehicle: solubility in water <0.01 g/L

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: samples were taken before the start of the test and after 24 and 72 hours from all test concentrations and from the control. 3.0 mL of the control was taken. At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling. Reserve samples of 3.0 mL were taken from all test solutions for possible analysis.
- Sample storage conditions before analysis: stored in a freezer (< -15°C) until analysis.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preperation of test solutions started with a loading rate of 100 mg/L applying a 10 minuteultrasonication treatment followed by a two-day period of magnetic stirring at room temperature, mostly in the dark. The obtained mixture was filtered through a preconditioned 0.45 µm membrane (RC55, Whatman) to remove the undissolved fraction. The obtained SS was used as the highest test concentration. Lower test concentrations were prepared by subsequent dilutions of the SS in test medium.
All aqueous solutions were clear and colourless.
- Controls: yes

Test organisms

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
- 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
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not): same as the test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Hardness:

24 mg CaCO3/L

Test temperature:

22-23°C

pH:

t=0h: 8.0
t=72h: 7.8

Dissolved oxygen:

not reported

Salinity:

not applicable

Nominal and measured concentrations:

nominal test concentrations final test: 0.10, 1.0, 10 and 100 % SS prepared at 100 mg/L
nominal concentration analysed (mg/L): 100, 100 (without algae)
measured test concentration final test t= 0 h: 0.0105, 0.0110 mg/L
measured test concentration final test t= 24h: <0.00800, <0.00800 mg/L
measured test concentration final test t= 72h: <0.00800, <0.00800 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL all-glass flasks
- Type (delete if not applicable): capped vessels
- Material, size, headspace, fill volume: 100 mL all-glass flasks filled with 50 mL test solution
- Initial cells density: 10,000 cells/mL
- Control end cells density: 66.6 x 10,000 cells/mL (mean)
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: M2 according to the OECD 201 guideline, formulated using Milli-RO water
- Culture medium different from test medium: no
- Intervals of water quality measurement: temperature is continuously measured, pH at the beginning in all concentrations and the end of the test in the control and the limit concentration.

OTHER TEST CONDITIONS
- Sterile test conditions: no information
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: using TLD-lamps (no light intensity reported)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: At the beginning, cells were counted using a microscope and a counting chamber. thereafter, cell densities were determined by spectrophotometric measurement of samples at 720 nm using a spectrophotometer with immersion probe (pathlength = 20mm).
- effect calculated parameters: specific growth rate and yield

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Range finding study
- Test concentrations: 0.10, 1.0, 10 and 100% of the SS
- Results used to determine the conditions for the definitive study: yes.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 other: % v/v saturated solution

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: not reported

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 other: % v/v saturated solution

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Any stimulation of growth found in any treatment: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: clear test solution
- EC50 (yield) = >0.0048 mg T001586 /L
- EC10 (growth rate) = >0.0048 mg T001586 / L
- EC10 (yield) = >0.0048 mg T001586
- NOEC (yield) = 0.0048 mg T001586 / L.

Results with reference substance (positive control):

- Results with reference substance valid? yes (should be between 0.82 and 2.3 mg/L)
- EC50: 72-h EC50 (growth rate) = 1.5 mg/L (95% C.L. 1.4 to 1.6 mg/L)
- EC50: 72-h EC50 (yield decrease) = 0.51 mg/L (95% 0.49 to 0.53 mg/L) (should lie between 0.43 and 1.1 mg/L)

Reported statistics and error estimates:

No EC50-values could be calculated because the test item proved to be non-toxic (EC50 > maximum concentration tested)

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

A 72-h growth inhibition test with the unicellular green alga Pseudokirchneriella subcapitata was performed with the test substance T001586 according to the OECD guideline 201.
The test item T001586 had no inhibitory effect on the growth of the algae after the exposure period of 72 hours up to an including the highest test concentration of 100 v/v % saturated solution.
The results of the test can be considered reliable without restrictions.