Cyclopenta[c]cyclopropa[g][1,6]diazacyclotetradecine-12a(1H)-carboxylic acid, 2,3,3a,4,5,6,7,8,9,11a,12,13,14,14a-tetradecahydro-2-[[7-methoxy-8-methyl-2-[4-(1-methylethyl)-2-thiazolyl]-4-quinolinyl]oxy]-5-methyl-4,14-dioxo-, ethyl ester, (2R,3aR,10Z,11aS,12aR,14aR)-

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:

2015-12-09 to 2015-28-09

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult substances and mixtures OECD series on testing and assessment number 23 december 14 2000

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

ethyl (1R,4R,6S,7Z,15R,17R)-17-({7-methoxy-8-methyl-2-[4-(propan-2-yl)-1,3-thiazol-2-yl]quinolin-4-yl}oxy)-13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.0^{4,6}]octadec-7-ene-4-carboxylate

EC Number:

618-843-3

Cas Number:

923604-57-3

Molecular formula:

C37H46N4O6S

IUPAC Name:

ethyl (1R,4R,6S,7Z,15R,17R)-17-({7-methoxy-8-methyl-2-[4-(propan-2-yl)-1,3-thiazol-2-yl]quinolin-4-yl}oxy)-13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.0^{4,6}]octadec-7-ene-4-carboxylate

Test material form:

solid

Details on test material:

- Name of test material (as cited in study reports): JNJ-38970191-AAA (T003009)
- Physical state: solid
- Appearance white solid

Specific details on test material used for the study:

- Appearance: White solid (determined by WIL Research Europe at receipt of the test item)
- Batch: I15AB0148
- Purity/Composition: 97.8 % v/v
- Test item storage conditions: At room temperature
- Stable under storage conditions: until 21 January 2017 (retest date)

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: samples were taken before the start of the test, after 24 hours and after 72 hours from all test concentrations and from the control. 2 ml volume was taken. The filter used for preparation of the SS was kept for possible analysis of the residue. At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling. Reserve samples 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: The test medium of the highest nominal test item concentration of 100 mg/L was prepared applying 2 days of magnetic stirring in the dark. The obtained mixture was filtered through a 0.45 μm membrane (RC55, Whatman) to remove the undissolved fraction. The filter was pre-conditioned with a small volume of test solution that was discarded. The obtained clear and colourless saturated solution (SS) was used as the highest test concentration. Lower test concentrations were prepared by subsequent dilutions of the SS in test medium and were clear and colourless
- Controls: yes
- Evidence of undissolved material (e.g. precipitate, surface film, etc): final test solutions were clear and colourless

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:

21.3-22.4°C

pH:

7.7-8.1

Dissolved oxygen:

not reported

Salinity:

not applicable

Nominal and measured concentrations:

nominal test concentrations: 0, 1, 10, 100 % v/v saturated solution
measured concenrtation: 100 % v/v saturated solution: 0.31 µg/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: 205.5 x 10,000 cells/mL
- No. of vessels per concentration (replicates): 3
- 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= continuously, pH at the beginning and the end of the test.

OTHER TEST CONDITIONS
- Sterile test conditions: no information
- Adjustment of pH: none
- Photoperiod: continuous illumination
- Light intensity and quality: using TLD-lamps.

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.
- effect calculated parameters: specific growth rate and yield

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2

- Range finding study
- Test concentrations: 1.00, 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

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

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50: 72-h EC50 (growth rate) = 1.2 mg/L (95% C.L. 1.1 to 1.2 mg/L)

Reported statistics and error estimates:

NOEC and EC values were determined using a Williams Multiple Sequential t-test Procedure (alpha=0.05, one-sided, smaller).
Calculation of ECx values was based on probit analysis using linear max. likelihood regression.

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 wit the test substance T003009 according to the OECD guideline 201 (GLP conditions).
Under the test conditions with Pseudokirchneriella subcapitata, T003009 did not inhibit growth rate of this fresh water algae species significantly. The 72-h ECr50 was determined to be > 100% v/v saturated solution.
The results of the test can be considered reliable without restrictions.