Registration Dossier

Ecotoxicological information

Long-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
4 June 1996 to 25 June 1996
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP Guideline Study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: Study report cites OECD Guideline 202, Daphnia sp., 14-day Reproduction Test (OECD, 1984) but with modification to 21-day exposure the study is equivalent to the current OECD Guideline 211, Daphnia magna Reproduction Test.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): TXIB Plasticizer
- Physical state: clear liquid
- Analytical purity: 99.0%
- Purity test date: 1 July 1994
- Lot/batch No.: EDC 22998 No. 10
- Storage condition of test material: Room temperature in dark, ventilated cabinet.

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: Control, Solvent Control, 0.13, 0.25, 0.50, 1.0, 2.0, QC 1, 2 and 3
- Sampling method: During the in-life phase of the definitive study, water samples were removed and analyzed for test substance on test days 0, 1, 7, 14 and 21. Samples were removed from alternate test vessels (A/B or C/D) at each sampling interval. All samples were removed from the approximate mid-point of the vessels by gravimetric siphoning. In addition, three quality control (QC) samples were prepared at each sampling interval and remained with the set of exposure solution samples throughout the analytical process.
- Sample storage conditions before analysis: Samples transferred directly to separatory funnels for extraction.

Test solutions

Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A 22 mg a.i./L stock solution was prepared on days -6, 1, 8 and 15 by dissolving 1.1110 g (1.1000g as active ingredient) with acetone to volume in a 50 mL volumetric flask. In addition, a 0.72 mL/mL solvent stock solution was prepared by diluting 36 mL of acetone with distilled water to a volume of 50 mL in a volumetric flask. The diluter was calibrated to deliver 0.035 mL of the test substance stock solution (22 mg A.I./mL) into the mixing chamber which also received 0.382 L of dilution water per cycle. The solution in the mixing chamber represented the highest nominal treatment level (2 mg A.I./L) and was subsequently diluted (50%) to provide the remaining nominal test concentrations: 1.0, 0.50, 0.25 and 0.13 mg A.I./L. A similar system was used to deliver the solvent stock solution to the solvent control vessels at the maximum amount of solvent contained in any test solution (i.e., 0.092 mL/L)
- Controls: Dilution water and solvent controls
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Evidence of undissolved material (e.g. precipitate, surface film, etc): None observed

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Source: Laboatory culture
- Feeding during test
- Food type: Suspension of green algae + YCT (yeast, cereal leaves, and digested flaked fish food)
- Amount: 3.0 mL algae (4x10e7 cells/mL) + 1.0 mL YCT per test vessel
- Frequency: 3X daily on weekdays and 2X daily on weekends




Study design

Test type:
flow-through
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d

Test conditions

Hardness:
160-180 mg/L as calcium carbonate
Test temperature:
20 + or - 1 degree C
pH:
7.8 - 8.3
Dissolved oxygen:
7.9 - 8.8 mg/L
Salinity:
NA
Nominal and measured concentrations:
Nominal concentration: Mean measured concentration (units mg A.I./L)
0.13 : 0.11
0.25 : 0.14
0.5 : 0.37
1.0 : 0.70
2.0 : 1.3
See Table 1 below for greater detail
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 1.6 L glass battery jars with drains 15 cm from bottom to hold volume of 1.4 L
- Aeration: No aeration
- Type of flow-through (e.g. peristaltic or proportional diluter): proportional diluter
- Renewal rate of test solution (frequency/flow rate): 6.0 test chamber volumes per 24 hours - 90% solution replacement at ~9 hours
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Fortified well water
- Metals: Periodic analyses on representative samples conducted but results not reported for study.
- Pesticides: Periodic analyses on representative samples conducted but results not reported for study.
- Alkalinity: 110-120 mg/L
- Conductivity: 500-550 micromhos per centimeter



OTHER TEST CONDITIONS
- Photoperiod: 16 hours light and 8 hours dark per day
- Light intensity: 14-42 footcandles


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Survival (immobilization) and abnormal behavior recorded at test initiation and on days 1, 2, 4, 7, 10, 13, 14, 17, 20 and 21. Reproduction (offspring production) determined on day 7 and 3x/week through day 21.


VEHICLE CONTROL PERFORMED: yes


RANGE-FINDING STUDY
- Test concentrations: 14 day preliminary rangefinding study at 0.13, 0.25, 0.50 1.0 and 2.0 mg A.I./L
- Results used to determine the conditions for the definitive study: Water solubility (~ 2.0 mg/L) and preliminary test confirmed selection of
concentrations for definitive test.
Reference substance (positive control):
not required

Results and discussion

Effect concentrationsopen allclose all
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.7 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
1.3 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
other: MATC
Effect conc.:
> 0.7 - < 1.3 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
>= 1.3 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
immobilisation
Details on results:
- Mortality of parent animals:
Pooled Control = 1%
0.11 mg/L = 5%
0.14 mg/L = 5%
0.37 mg/L = 2%
0.70 mg/L = 0%
1.3 mg/L = 0%
- Mean Cumulative No. of offspring produced per female:
Pooled Control = 145
0.11 mg/L = 151
0.14 mg/L = 151
0.37 mg/L = 136
0.70 mg/L = 140
1.3 mg/L = 122

Results with reference substance (positive control):
NA
Reported statistics and error estimates:
EC50 for survival estimated emerically due to no concentration exhibiting 50% mortality.
Student's t-Test used to compare survival and reproduction in control and solvent control. No significant difference found and controls were pooled for comparisons to determine treatment effects.
Shapiro-Wilks Tests used to test for normality at 99% level of certainty. Data determined to be normally distributed.
Bartlett's Test used to determine homogeneity of variance at 99% level of certainty. Data determined to be homogenious.
Williams Test used to detemine treatment effects on survival and reproduction at a 95% level of certainty.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
Well conducted GLP study using methods similar to current OECD TG #211 in a continuous flow-through system. Study was designed to evaluate
chronic effects of the test material at concentrations up to what was believed to be the water solubility of the substance. No effects on survival (immobilization) were observed. A slight, but statistically significant effect on reproduction was observed at the highest test concentration but not in the remaining test concentrations.