[1,1'-Biphenyl]-2-carbonitrile, 4'-[(2-butyl-4-oxo-1,3-diazaspiro[4.4]non-1-en-3-yl)methyl]-

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:

16 July 1996 to 21 November 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study; GLP compliance

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

4'-((2-butyl-4-oxo-1,3-diazaspiro[4.4]non-1-ene-3-yl)methyl)(1,1'-biphenyl)-2-carbonitrile

EC Number:

604-075-6

Cas Number:

138401-24-8

Molecular formula:

C25H27N3O

IUPAC Name:

4'-((2-butyl-4-oxo-1,3-diazaspiro[4.4]non-1-ene-3-yl)methyl)(1,1'-biphenyl)-2-carbonitrile

Details on test material:

- Code of test material (as cited in study report): SR47563
- Chemical name:4’-[(2-butyl-4-oxo-1,3-diaza-spiro[4.4]non-1-en-3-yl)methyl]- biphenyl]-2-carbonitrile
- Physical state: white-beige powder
- Lot/batch No.: 6R00003
- Solubility: 1.3E-3 g/L at 23˚C
- Stability: stable under normal conditions of use
- Moll mass: 385.5
- Molecular formular: C25H27N3O
- Melting point: 97˚C

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 90 - 45 - 22.5 - 11.25 - 5.63 - 2.81% correspondant à des concentrations réelles d'exposition respectivement de 0.94 - 0.47 - 0.24 - 0.116 - 0.06 - 0.03 mg/l

Test solutions

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Substance is dissolved in 50/50 water/acetonitrile

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Raphidocelis subcapitaa
- Strain: Chloroccales (chlorophytes chlorophycées)
- Source (laboratory, culture collection): The culture Center of Algae and Protozoa (Cambridge, United Kingdom) ref 278/4

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Hardness:

34 mg/l of CaCO3

Test temperature:

23 +/- 2°C

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL erlenmeyers
- Material, size, headspace, fill volume: 6 erlenmeyers filled with 100mL
- Renewal rate of test solution (frequency/flow rate):
- Initial cells density:
- Control end cells density:
- No. of organisms per vessel:
- No. of vessels per concentration (replicates):
- No. of vessels per control (replicates):

GROWTH MEDIUM
- Standard medium used: yes/no
- 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:
- Light intensity and quality:
- 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:
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations:
- Results used to determine the conditions for the definitive study:

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.98 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 0.76-1.45

Effect concentrations 2

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.47 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

other: biomass and growth rate

Remarks on result:

other: See the conclusion for discussion of the reliability of the test results.

Any other information on results incl. tables

Analyses of the samples taken from the highest concentration showed that the initial concentration was 1.08 mg/l, which decreased to 0.82 mg/l after 72 h when the test was ended. The decrease was 24% and it was assumed that a similar average decrease would be valid for the lower concentrations. Based on these calculated concentrations, the estimated exposure range was 0.03, 0.06, 0.116, 0.24, 0.47, 0.94 mg/l.

Statistical analyses of the data showed that only the effect induced at the highest concentration was significant. For both biomass and growth rate the NOEC was determined at 0.47 mg/l (Bonferroni t-test).

Applicant's summary and conclusion

Validity criteria fulfilled:

no

Remarks:

The criterion with respect to the stability of the test concentrations (decrease not > 20%) was not fulfilled.

Conclusions:

Under the conditions of this study, it is not unequivocally clear that the observed effect on algal growth is directly related to the presence of the test substance. Therefore, the values for EC50 and NOEC reported here may not reflect the actual toxicity of the substance with respect to algal growth. The effects observed may have been caused by other conditions than the presence of the substance..