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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted under GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report Date:
2005

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Dibutyl terephthalate
- Substance type: production sample
- Physical state: clear liquid
- Analytical purity: 98.7%
- Lot/batch No.: X-29455-010

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Samples of the control and test substance exposure solutions were analyzed at test start and test end to verify test substance concentration. The samples analyzed at test start were collccted from approximately mid-depth of the 22-L mixing vessels using a spigot-valve assembly. The samples analyzed at test end were surrogate exposure solutions collected at test start that were stoppered without headspace throughout the study. Samples were analyzed using gas chromatography with flame ionization detection (GC/FID).

Test solutions

Vehicle:
yes
Details on test solutions:
The solubility of the substance in the test medium was measured by the analytical laboratory. The solubility of the substance was determined to be 0.13 mg/L. Exposure solutions containing the substance at nominal concentrations of 0.025, 0.05, 0.1, 0.2, and 0.4 mg/L were tested. The exposure solutions were prepared by direct addition of a stock solution containing the substance to dilution water. Five stock solutions were prepared at nominal concentrations of 0.26,0.56, 1.1, 2.1, and 4.1 mg/mL by weighing the appropriate amounts of substance into volumetric flasks that were brought to volume with a solvent, N,N-Dimethylformamide (DMF). The stocks were sonicated and inverted several times to dissolve the substance, then 2 mL of each stock solution were added to 22-L vessels containing 20 liters of laboratory dilution water. Dilution water and carrier solvent control solutions were also prepared. Carrier solvent controls were prepared by pipetting 2 mL ofDMF into 20 liters of laboratory dilution water. The solutions were stirred with stir bars and stir plates for approximately 3 hours.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Daphnia magna organisms were cultured within the testing facility. An organism origination statement and taxonomic verification are located in a central file maintained by the Ecotoxicology Group, Room 2020, Building 320, Kodak Park. The test organisms are cultured in 100-L stainless steel tanks provided with a continuous flow of filtered-treated-tempered water. The water depth is maintained by use of an ovelflow pipe that discharges into a floor drain. The tanks are continuously aerated by passing oil-free filtered air through air stones. The culturing facility is maintained at a constant temperature of 20±1°C and is illuminated with fluorescent lighting for 16 hours followed by a 30-minute transition period leading to 8 hours of darkness. The daphnids are fed an ample amount of a spinach-yeast-fish food slurry that may be supplemented with a yeast-Cerophyll leaves-trout chow (YCT) mixture or green algae, such as Selenastrum capricomutum. The dietary components of the YCT are analyzed routinely to identify contaminants that could interfere with the outcome of the study. The brood stock used in this test were cultured in the water used for preparing the test substance
solutions. Approximately 24 hours before test start an ample number of gravid adult daphnids were transferred into 20-cm diameter bowls, containing laboratory culture water, and fed. The neonates produced in the following 24-hour period were used as the test organisms. The neonates were collected by pipette and transferred dircctly into the test vessels. Sequential randomization was accomplished by allocating to each vessel no more than 50% of anyone set of test organisms at a time.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Test temperature:
The temperature of the dilution water and carier solvent control solutions ranged from 19.6 to 20.4°C. The temperature of the test substance solutions ranged from 20.1 to 20.5°C.
pH:
The pH of the control and test substance solutions ranged from 8.0 to 8.3.
Dissolved oxygen:
The dissolved oxygen concentrations of the control solutions ranged from 7.9 to 9.2 mg/L, and the test substance solutions
ranged from 8.2 to 9.1 mg/L.
Nominal and measured concentrations:
The combined replicate exposure concentrations of the nominally prepared 0.025, 0.05, 0.1, 0.2 and 0.4-mg/L solutions were calculated to be 0.02, 0.04, 0.08, 0.16, and 0.34 mg/L. The loss of test substance concentration from the individual replicates ranged from 30.4% to -13.3% during the 48-hour exposure.
Details on test conditions:
The test was performed as a 48-hour static exposure. The exposure solutions were maintained at a temperature of 20 ± 1°C and illuminated with fluorescent lighting for 16 hours followed by a 30-minute transition period leading to 8 hours of darkness. The test vessels were 300-mL pyrex BOD bottles containing 300 mL of test solution with no headspace. The test vessels were stoppered during the study. They were not cleaned or aerated, nor were the test organisms fed. Two replicates were prepared for each exposure. At test start, the exposure solutions were prepared and transferred into their respective test vessels. The physical parameters of the solutions were measured and ten test organisms were introduced into each replicate. Observations for mobility and stress were made at test start and after 4, 24, and 48 hours of exposure. The appearances of the exposure solutions were observed and recorded at test start, after 4 hours of exposure, and at the end of each 24-hour period. The physical parameters of each exposure solution were measured again at test end.

Results and discussion

Effect concentrationsopen allclose all
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 0.34 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
>= 0.16 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
No organisms were adversely affected in the dilution water or carrier solvent controls, or in the 0.025 and 0.1-mg/L solutions. In the 0.05 and 0.2 mg/L solutions 5% immobility was observed. In the 0.4-mg/L solutions six organisms were immobile and one organism was not found and assumed to be immobile. Statistical analyses of the study data were performed using the TOXSTAT statistical software. All data were used in the statistical analyses. For the purpose of calculating or estimating the 48-hour EC50 value, immobility served as the requisite endpoint. The 48-hour EC50 value
was estimated by extrapolation to be 0.41 mg/L. However, since this exceeds the highest measured concentration the EC50 is reported here as > the highest measured concentration (0.34 mg/L). The highest tested concentration resulting in < or =10% immobility (NOEC) was determined to be > or = 0.16 mg/L. The results indicate that the 48-hour EC50 and NOEC values are greater than the measured aqueous solubility of the test substance.
Reported statistics and error estimates:
Probit EC50 95% confidence limits = 0.2923 to 0.5184
NOEC determined using Fisher's Exact Test (alpha level = 0.05)

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
Well conducted guideline study conducted under GLP.
Executive summary:

Daphnia magna were exposed to five concentrations of the test substance in a 48-hour, static, aquatic effects test. The solubility of the substance in the test medium was investigated and determined to be 0.13 mg/L. Exposure solutions containing the substance at nominal concentrations of 0.025, 0.05, 0.1, 0.2, and 0.4 mg/L were tested. The concentrations of test substance in the exposure solutions were analytically verified during the study. The average concentrations of the prepared solutions were calculated to be 0.02, 0.04, 0.08, 0.16. and 0.34 mg/L. The 48-hour EC50 value was estimated to be 0.41 mg/L by extrapolation. However, since this exceeds the highest measured concentration the EC50 is reported here as greater than the highest measured concentration (48 h EC50 >0.34 mg/L). The highest tested concentration resulting in < or =10% immobility (NOEC) was determined to be > or = 0.16 mg/L. The results indicate that the 48-hour EC50 and NOEC values are greater than the measured aqueous solubility of the test substance.