Registration Dossier

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Experimental starting: 2009-06-16 - experimental completion: 2009-06-18
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study according to international guideline

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
TEST ITEM: Ketoprofen I CFPPN
Batch number: F-PPR-080059
CAS RN: 42872-29-7
Chemical name: 3-(1-Cyanoethyl)benzoyl chloride
Content: 95.0%
Appearance: Brown liquid
Sum formula: C10H8ClNO
Molecular weight: 193.63 g/mol
Water solubility: Hydrolyses at 20 °C
Expiry date: 2009-08-12
Recommended storage: Refrigerator, 7 ± 2 °C, protected from moisture and light
Storage at test facility: 6 ± 2 °C, protected from moisture and light
Retention of test item: At least 1 g has been retained.

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: Stock solutions of 100 mg/L of each standard were prepared in methanol. A mixed standard of 4.0 mg/L was prepared from these
stock solutions in mobile phase, diluted with mobile phase (6 concentrations (Diluted level of the saturated concentration: 1:1 -1:2-1:4-1:8-1:16)), and used for calibration.
- Sampling method: Analytical evaluation of various concentrations of Ketoprofen, active ingredient of Ketoprofen / CFPPN, and its metabolite Capril was carried out via HPLC on a reversed-phase column using external standards. Analysis was performed in gradient mode with a Diode Array Detector.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until start of analysis, if necessary.

Test solutions

Vehicle:
yes
Details on test solutions:
Stock solution: The stock solution (100 mg/L test item was weighed out) was prepared with dilution water one day prior to application.
Dispersion treatment: The stock solution was shaken with 20 rpm for 24 h (rotating shaker 3040, GFL). Undissolved particles were removed by centrifugation with 3000 rpm for 20 min at room temperature.
Test dilutions of the saturated solution: 1:1 -1:2-1:4-1:8-1:16 (dilution factor: 2), corresponding to the nominal test item concentrations of 100 - 50.0 - 25.0 - 12.5 - 6.25 mg test item/L.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Origin: lnstitut fOr Wasser-, Boden- und Lufthygiene (WaBolu)
Breeder: DR.U.NOACK-LABORATORIEN, Kathe-Paulus-Str. 1, D-31157 Sarstedt, Germany
Culture: In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 ± 2 °C, in an incubator, 16 h illumination, illumination strength max. 20 µE.m^-2/s .
Culture medium: Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaC03/L.
Feeding: At least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata, Desmodesmus subspicatus and Chlorella vulgaris, with an algae cell density of > 10^6 cells/ml.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
24 and 48h

Test conditions

Hardness:
Hardness of the dilution water: 265 mg CaC03/L
Test temperature:
18 - 22 °C, ± 1 °C constant
pH:
Additionally a pH-control was tested. Therefore, 4 replicates of the highest test concentration was adjusted to a pH in the range of 7.0 ± 0.2 by addition of 1 M NaOH.
Dissolved oxygen:
The dissolved oxygen concentration was 3 mg/L.
Nominal and measured concentrations:
The nominal test item concentrations: 100 - 50.0 - 25.0 - 12.5 - 6.25 mg/l
Details on test conditions:
Test vessels: Glass beakers (4 cm ID x 7 cm H), 50 ml capacity
Test volume per vessel: 20 ml
Number of study organisms: 20 animals, divided into 4 parallel samples, each with 5 animals per test concentration, pH control and control
Age of test organisms: 2 to 24 h old daphnids were used for the definitive study. They were obtained by removing the mother animals twice within 22 h.
Acclimatization: At least 2 h in dilution water
Illumination: Diffuse light, illumination strength max. 20 IJE/m/s
Photoperiod: 16/8 h light/dark cycle
Feeding: The daphnids were not fed during the study.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate p.a.

Results and discussion

Effect concentrationsopen allclose all
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
70.7 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: CI 50.0- 100 mg/L
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
70.7 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Cl: 50.0- 100 mg/L
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
50 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC10
Effect conc.:
67.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
67.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC100
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC100
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Results with reference substance (positive control):
The percentage immobility for the reference item was determined after 24h. The EC50-value with 95 % confidence interval (Cl) was calculated by sigmoidal dose-response regression to be:
EC50: 1.66 mg/L (Cl 1.50- 1.83 mg/L)
The EC50-value of the reference item potassium dichromate after 24 h is within the prescribed concentration range of 1.0 - 2.5 mg/L of quality criteria according to AQS P 9/2 (05/1996) for daphnids clone 5 cultured in Elendt M4 medium. The EC50-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202.
Reported statistics and error estimates:
The EC100-values after 24 and 48 h, the NOEC and the LOEC were deduced directly from the dose-response-relationships without mathematical calculation.
EC10-values were calculated by sigmoidal dose-response regression. Since only 0 % and 100 % immobilization occurred, the EC50-value was determined by calculating the geometric mean of the highest concentration causing no immobilization and the lowest concentration causing 100 % immobilization. As confidence limits for the EC50 the highest concentration causing no immobility and the lowest concentration causing 100 % immobility were used, because only 0% and 100 % mortality occurred within the tested concentration range. The concentration effect relationships are shown graphically.
The EC50 for the of the reference test were calculated by sigmoidal dose- response regression. The confidence interval for the EC50 of the reference test was calculated from the best-fit values, the standard error and the t-distribution with the software GraphPad prism.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
In this study Ketoprofen/CFPPN was found to be toxic to Daphnia magna after 48h at concentrations of 100 mg/L (LOEC48 h). The NOEC after 48 hours was 50.0 mg/L. The EC50 (48h) was calculated to be 70.7 mg/L (Cl: 50.0- 100 mg/L). All effect values are given based on the nominal concentrations of the test item Ketoprofen/CFPPN.