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Long-term toxicity to aquatic invertebrates

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Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009-04-07 to 2009-04-29
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Sampling method: Duplicate samples were taken from the freshly prepared test media of the loading rate of 100 mg/L and the control at the start of every test interval. In addition, duplicate samples were taken from the freshly prepared (Days 0, 2, 5 and 7) and aged test media (Days 2, 5 and 7) of all test concentrations and from the control at three test intervals. The aged samples were stored without food and test animals, but were incubated during the renewal periods under test conditions.

- Sample storage conditions before analysis: All samples were analyzed immediately after the sampling.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: A dispersion of the test item with the loading rate of 100 mg/L was prepared by adding 300 mg of the test item (dosed by pipetting 300 μL of the test item) into a beaker partially filled with test water under intense stirring. No auxiliary solvent or emulsifier was used. After addition of the test item, the beaker was filled up with test water to the volume of 3000 mL and the test item was mixed into the test water as homogeneously as possible using ultrasonic treatment (15 minutes). The pH was adjusted from 3.0 to 7.9 (± 0.3). Thereafter, the dispersion was stirred on a magnetic stirrer at room temperature over 24 hours in the dark. The stirring period of the dispersion was chosen according to the results of a pre-experiment.

Following the stirring period of 24 hours, the test medium was incubated in the dark for an equilibration period of one hour to allow any undissolved reaction products to float at the surface of the test solution or to settle on the bottom. The test water (containing the water dissolvable hydrolysis products) was taken from the middle of the water column to avoid any undissolved test item/hydrolysis products in the test water.

This test water was used as the highest concentrated test medium and as the stock solution for lower concentrated test media. Requisite aliquots were diluted with test water to obtain the additional test concentrations.

The test media were freshly prepared before the start of the test and before each test medium renewal.

- Controls: Dilution water
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM

- Strain/clone: Clone 5

- Source: University of Sheffield/UK

- Age of parental stock (mean and range, SD): The daphnids used for the test originated from parental daphnids that were at least 14 days old but not older than four weeks and were not first brood progeny. At the start of the test, the test animals were less than 24 hours old.

- Feeding during test: The test animals were fed daily with a food mixture containing a suspension of green algae of the species Scenedesmus subspicatus (freshly grown at Harlan Laboratories) and a fish food suspension.

The fish food suspension was prepared by dispersing 10 g of powdered commercial fish diet (TETRA MIN Hauptfutter, obtained from TETRA-Werke, 49304 Melle / Germany) in 500 mL of test water. The suspension was allowed to stand for 4 hours. Then, 400 mL of the supernatant were taken, diluted 1:1 with test water and boiled. The suspension was stored deep frozen in small quantities until use.

The carbon contents of the algal and fish food suspensions were determined using a Shimadzu TOC 5000A Analyzer. The food amount (based on the measured concentrations of the total organic carbon (TOC) in the food suspensions) was 0.20 mg TOC per Daphnia and day.


ACCLIMATION

- Acclimation conditions (same as test or not): The temperature and light conditions were identical to those of the test.

- Feeding frequency: The animals were fed normally three times a week with green algae of the species Scenedesmus subspicatus and with a fish food suspension.

- Health during acclimation (any mortality observed): No signs of stress were observed and the brood stock was healthy.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Hardness:
250 mg/L as CaCO3
Test temperature:
20-21°C
pH:
7.5-8.0
Dissolved oxygen:
≥7.2 mg/L
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal concentrations: 0 (Control), 100 mg/L and dilutions of 1:2.16, 1:4.7, 1:10 and 1:22.

Mean initial measured concentrations in the treated media: 65, 32, 14, 6.7 and 3.1 mg/L

At the start of the renewal periods the concentrations of the test item measured in the test medium of all tested concentrations (based on the TOC) were between 41 and 76% of the nominal concentrations. At the end of the three measured renewal periods the concentrations were between 62 and 85% (test media without food). Thus, the TOC content of the tested concentrations was sufficiently stable during the test medium renewal periods of two and three days. All reported biological results are related to the initially mean measured concentrations of the test item.
Details on test conditions:
TEST SYSTEM

- Test vessel: The test was performed in 100-mL glass beakers containing 80 mL of test medium. The test vessels were covered with glass plates to reduce the loss of water by evaporation and to avoid the entry of dust into the solutions.

- Replication: The study was started with 10 daphnids per treatment. Each test animal was kept individually in a glass beaker. The test animals were randomly distributed to the test vessels. The test duration was 21 days.

- Test medium renewal: The test media of all treatments were renewed on Days 2, 5, 7, 9, 12, 14, 16, and 19 of the test period (every Monday, Wednesday, and Friday). At these dates, the surviving test animals were carefully transferred by means of glass tubes into test vessels containing freshly prepared test medium.

- Lighting: A 16-hour light to 8-hour dark cycle with a 30 minute transition period was used. Light intensity during the light period was between approximately 520 and 680 Lux.

- Observations: The test replicates were observed for mortality of adults on Days 0-2 and thereafter three times per week before renewal of the test media. On the same dates, the test replicates were observed for live and dead offspring and for the presence of aborted eggs.

- Calculations: The reproduction rate was calculated as the total number of living offspring produced per parent female surviving until the end of the test.

- Water quality: At the beginning and end of each test medium renewal period, the pH and dissolved oxygen concentrations were measured in one replicate of each test concentration and of the control. At the same time, the water temperature was measured in one of the control replicates. Additionally, the room temperature was continuously monitored. The appearance of the test media was visually inspected and recorded at the beginning and end of each test medium renewal period.

- Selection of test concentrations: test concentrations were selected on the basis of the results of a non-GLP range-finding test (results not reported)
Reference substance (positive control):
no
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
32 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Mean initial measured concentration of Total Organic Carbon expressed as equivalent test substance concentration
Basis for effect:
mortality
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
65 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Mean initial measured concentration of Total Organic Carbon expressed as equivalent test substance concentration
Basis for effect:
mortality
Details on results:
- Control mortality: 0%

- Other biological observations: With the exception of the reported mortality, no visible abnormalities were observed at the test animals during the test. No EC values for the inhibition of the reproduction rate could be calculated since no effect was determined on the reproduction of the daphnids at all test concentrations with surviving parent animals.
Reported statistics and error estimates:
The mean reproduction rates of the daphnids at the test concentrations were compared to the control by multiple Williams’ tests

Table 1. Results of analysis of test media

 Nominal treatment  Nominal concentration (mg/L)  Mean measured initial concentration (mg/L)  Percentage of nominal  n
 0 (Control) 0  -  -  -
 1:22 dilution of stock  4.5  3.1  68  4
 1:10 dilution of stock  10 6.7   67  4
 1:4.7 dilution of stock  21  14  68  4
 1:2.16 dilution of stock  46  32  69  4
 100 mg/L stock  100  65  65  9

Table 2. Test results

 Mean initial measured concentration (mg/L)  Percentage mortality after 21 days  Day of first offspring release  Mean reproduction rate (live young per surviving adult) Mean reproduction rate as a percentage of control 
 0 (Control)  0  9  89.8  -
 3.1  20  7  86.1  96
 6.7  10  7  92.4  103
 14  10  9  84.3  94
 32  20  9  82.5  92
 65  100  7  Not applicable (all parents dead)
Validity criteria fulfilled:
yes
Conclusions:
A 21-day NOEC of 32 mg/L and a LOEC of 65 mg/L have been determined for the effects of the test substance on mortality of Daphnia magna. No effects on reproduction were determined in test concentrations below the LOEC for mortality. The report authors note that it cannot be excluded that the effects of the test item were at least partly caused by undissolved test item, which could not be removed by the preparation method used during the
test period. It is likely that the test organism were primarily exposed to the hydrolysis products of the substance.
Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
Please refer to the attached justification for grouping of substances provided in IUCLID Section 13.
Reason / purpose:
read-across source
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
32 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Mean initial measured concentration of Total Organic Carbon expressed as equivalent test substance concentration
Basis for effect:
mortality

Description of key information

21-d NOEC of 32 mg/l and LOEC of 65 mg/l; mobility of Daphnia magna (OECD 211), RL1

Key value for chemical safety assessment

Additional information

Measured data was not available for trimethoxy(2,4,4-trimethylpentyl)silane (CAS 034396-03-7) data was therefore read across from the comparable substance, trichloro(2,4,4-trimethylpentyl)silane (CAS 18379-25-4).

The influence of the test item on the mobility of Daphnia magna was investigated in a 21-d semi-static test according to the OECD 211 and GLP (Harlan, 2009c).

A 21-day NOEC of 32 mg/l and a LOEC of 65 mg/l have been determined for the effects of the test item on mortality of Daphnia magna. No effects on reproduction were determined in test concentrations below the LOEC for mortality. The report authors note that it cannot be excluded that the effects of the test item were at least partly caused by undissolved test item, which could not be removed by the preparation method used during the test period. It is likely that the test organism were primarily exposed to the hydrolysis products of the substance.