9-Anthracenecarboxylic acid, (triethoxysilyl)methyl ester

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2003-01-21 to 2003-05-08

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

a combined limit/range-finding test was performed using 4 concentrations in the range of 100% (saturated solution) and 0.1%

Qualifier:

according to guideline

Guideline:

ISO 6341 (Water quality - Determination of the Inhibition of the Mobility of Daphnia magna Straus (Cladocera, Crustacea))

Version / remarks:

1996-04-01

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

1984-04

GLP compliance:

yes (incl. QA statement)

Remarks:

signed 2003-05-08

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: In the freezer under nitrogen and protected from light.

Analytical monitoring:

yes

Remarks:

- All test solutions were protected from light as much as possible, due to the presumed light sensitivity of the test substance.

Details on sampling:

- Concentrations: Duplicate samples were taken from blank-control and the filtrate prepared at 100 mg/L.
- Sampling method: 10 mL from the approximate centre of the test vessel.
- Timing/Frequency of sampling: Once at the start of the test (t= 0 h).
- Sample storage conditions before analysis: Not applicable, samples were analysed on the day of sampling.
- Standard/Calibration solutions: Standard solutions of TESAC were prepared in hexane using glassware flushed with nitrogen gas. Calibration solutions in hexane were made up from two standard solutions. Both solutions were protected from light as much as possible.
- Further sampling: In order to demonstrate proper preparation of the test solutions, a sample from the filter used for preparation of the nominal 100 mg/L test solution was collected and analysed.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preparation started with a supersaturated solution at nominally 100 mg/L that was magnetically stirred for one day in the dark prior to testing. The resulting dispersion was then filtered through a paper filter (Schleicher and Schuell 604, ca. 5 µm). The final, clear colourless, solution was then used for preparation of the lower test concentrations (10%, 1.0% and 0.1%) by subsequent dilution in the test medium.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Age at study initiation: < 24 h
- Source: In-house laboratory culture
- Method of breeding: 250 organisms (< 3 days old) in 10 L of M7 medium
- Temperature of medium: 18-22°C (±1°C)
- Maximum age of breeding cultures: 4 weeks
- Renewal of breeding cultures: After 7 days of cultivation half of the medium twice a week.
- Age of parental stock: Parental daphnids were more than 2 weeks old and showed no signs of stress such as mortality > 20%, presence of males, ephippla or decolorization and delay in the production of the first brood.
- Food type: A suspension of fresh water algae
- Frequency: Daily

ACCLIMATION
- Test medium: ISO, prepared in milli-RO water
- Introduction of daphnids to the test medium: Within ¾ h after preparation of the test solutions.
- Feeding during test: No

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

250 mg/L expressed as CaCO3

Test temperature:

20.5 to 21.4°C

pH:

Blank control: 7.7 at the start (t= 0 h) and 7.9 at the end (t= 48 h)
100 mg/L test substance test group: 7.8 at the start (t= 0 h) and 7.9 at the end (t= 48 h)

Dissolved oxygen:

Blank control: 9.5 mg/L at the start (t= 0 h) and 8.7 mg/L at the end (t= 48 h)
100 mg/L test substance test group: 9.3 mg/L at the start (t= 0 h) and 8.7 mg/L at the end (t= 48 h)

Nominal and measured concentrations:

Nominal concentrations in the combined limit/range-finding test: 0.1%, 1.0%, 10% and 100% prepared from a 100 mg/L filtered stock solution of the test substance.

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL, all-glass
- Volume of solution: 80 mL medium
- Aeration: No
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 2 replicates in the case of 0.1%, 1.0%, 10% filtrate dilutions and 4 replicates in the case of the 100% filtrate concentration (100 mg/L test substance).
- No. of vessels per control (replicates): 4 replicates

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO- and M7-medium prepared in tap water purified by reverse osmosis (milli-RO, Millipore Corp., Bedford, Mass., USA) according to OECD guidline No.202/ ISO 6341.
- Culture medium different from test medium: ISO-medium was used under test conditions, instead of M7-medium which was used for the breeding of the organisms.

OTHER TEST CONDITIONS
- Light intensity: The study was performed in the dark, due to the presumed light sensitivity of the test substance

EFFECT PARAMETERS MEASURED: Immobility at 24 h and 48 h

RANGE-FINDING STUDY
- Test concentrations: 0.1%, 1.0% and 10% of a filtered solution prepared from a 100 mg/L stock of the test substance.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7, Art. 4864, batch no. K28974764, Merck, Darmstadt, Germany)

Key result

Duration:

48 h

Dose descriptor:

EC50

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Remarks on result:

other: 48h-EC(50) > solubility of test material in test medium

Details on results:

- Immobilisation of control: No immobilization in the blank-control.
- Effect concentrations exceeding solubility of substance in test medium: Water solubility of the test substance was below the limit of detection (0.003 mg/L).

Results with reference substance (positive control):

The 24h-EC50 was 1.0 mg/L with a 95% confidence interval between 0.94 and 1.2 mg/L. The 48h-EC50 was 0.74 mg/L with a 95% confidence interval between 0.66 and 0.84 mg/L.

Table 1: Results of the recovery samples

Concentration nominal [mg/L]	Concentration analysed* [mg/L]	Recovery [%]
0.00500	0.00502	100.5
0.0200	0.0199	99.7

^*Results of duplicate samples. The maximum deviation between the response (n=2) was calculated for each sample and was < 10%.

Table 2: Acute immobilization of daphnia after 24 h and 48 h in reference test with potassium dichromate

Concentration [mg/L]	Number Exposed	% Immobile		Expected response (%) After 48 h*
		24 h	48 h	Minimal	Maximal
Blank-control	20	0	0	0	10
0.10	20	0	0	0	10
0.18	20	0	0	0	30
0.32	20	0	0	0	100
0.56	20	0	5	0	100
1.0	20	35	95	40	100
1.8	20	100	100	100	100

*Based on historical data of the previous years (n>60)

Table 3: Acute immobilization of Daphnia after 24 and 48 hours in the combined limit/range-finding test

Test group* test substance	Vessel Number	Number Daphnia exposed	Response at 24 h		Response at 48 h
			number	Total %	number	Total %
Blank-control	A B C D	5 5 5 5	0 0 0 0	0	0 0 0 0	0
0.1%	A B	5 5	0 0	0	0 0	0
1.0%	A B	5 5	0 0	0	0 0	0
10%	A B	5 5	0 0	0	0 0	0
100%	A B C D	5 5 5 5	0 0 0 0	0	0 0 0 0	0

* Test groups represent percentages of the 5 µm filtered solution prepared at 100 mg/L

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test substance on Daphnia magna (age: <= 24 h) was assessed in a 48 h acute toxicity test conducted in freshwater in accordance with EN ISO 6341, following OECD 202 (1984). 4 concentrations (between 100% saturated solution and 27%) and blank were tested in a combined limit/range-finding test resulting in an EC50 (48h) of > solubility of the test item in test medium (dissolved test item).

Executive summary:

The aim of the study was to determine under laboratory conditions whether the test substance has acute toxicity effects on the aquatic invertebrate Daphnia magna. Therefore, a combined limit/range-finding test was performed according to the guidelines ISO 6341 and OECD No. 202, in which the organisms were exposed to a blank-control and different dilutions (4 concentrations in the range of 100% (saturated solution), 10%, 1%, 0.1%) of a filtrate of a supersaturated solution of the test substance (nominal: 100 mg/L). Analysis of samples taken during the test showed, that the measured concentrations in the samples were below the limit of detection (3 µg/L). Immobilized Daphnia were neither observed in the blank-control, nor at the different concentrations of the test substance tested. The results of the reference test with K2Cr2O7 as positive control, were in the expected ranges of the test method. All test conditions (pH, oxygen concentration and temperature) remained within the protocol limits. Due to the low water solubility of the test substance, the 48h-EC50 (daphnia magna) is > water solubility of the test substance in test medium.

Description of key information

The effect of the test substance on Daphnia magna (age: <= 24 h) was assessed in a 48 h acute toxicity test conducted in freshwater in accordance with EN ISO 6341, following OECD 202 (1984). 4 concentrations (between 100% saturated solution and 27%) and blank were tested in a combined limit/range-finding test resulting in an EC50 (48h) of > solubility of the test item in test medium (dissolved test item).