[No public or meaningful name is available]

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:

Experiment start date - 02 March 2010; Experiment completion date - 23 March 2010; Study completion date - 17 May 2010.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Identification: FAT 40851/A TE
Batch Number: TZ 5891 / BOP 02-09
Purity: 69.9 % all coloured components
Appearance: Orange powder
Expiry Date: July 31, 2014
Storage Conditions: At room temperature at about 20 °C

Analytical monitoring:

yes

Details on sampling:

For the determination of the actual test item concentrations, duplicate samples were taken from each treatment at the start and at the end of the test after 48 hours. For the 48-hour stability samples, the contents of the respective replicate test vessels were combined prior to sampling. All samples were stored deep-frozen (at about -20 °C) immediately after sampling. The test item proved to be stable under these storage conditions (pretest, non-GLP).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The test medium was prepared by dissolving 100.9 mg of test item completely in 1000 mL of test water using ultrasonic treatment for 15 minutes and intense stirring for 5 minutes at room temperature. The test medium was prepared just before introduction of the daphnids (i.e. start of the test).

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Strain: Defined by the supplier as clone 5
- Source: Originally supplied by the University of Sheffield / UK in 1992.
- Age at study initiation: At the start of the test, the organisms used in the test were 6-24 hours old and were not first brood progeny.
- Method of breeding: Since 1992, the clone has been bred at Harlan Laboratories in reconstituted water of the quality identical to the water quality used in the tests (in respect to pH, main ions, and total hardness) and under temperature and light conditions identical to those of the tests. During breeding, daphnids are generally fed three times a week with an algal suspension of the green algae Desmodesmus subspicatus CHODAT, Strain No. 86.81 SAG, supplied by the Collection of Algal Cultures (SAG, Institute for Plant Physiology, University of Göttingen, 37073 Göttingen / Germany) and cultivated at Harlan Laboratories under standardized conditions or a mixture of this algal suspension and a commercial fish diet (Tetra Min® Hauptfutter, supplied by TETRA-Werke, 49304 Melle / Germany).
- Feeding during test: No

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

2.5 mmol/L (250 mg/L as CaCO3)

Test temperature:

The test was performed in a temperature-controlled room with continuous monitoring of the room temperature. The water temperature was maintained at 20 °C.

pH:

The pH values of the test medium and control were between 7.8 and 7.9.

Dissolved oxygen:

At the beginning and end of the test period, the dissolved oxygen concentration in the test medium and control was at least 8.6 mg/L.

Salinity:

See table "Test Water" below

Nominal and measured concentrations:

100 mg/L nominal:
The concentration of the test item was analytically measured in the duplicate test medium samples of the single test concentration taken at the start and end of the test. From the control, only one of the duplicate samples was analyzed per sampling time. The analytically determined concentration of the test item in the test medium at the start and end of the test was 104 and 103% of the nominal value, respectively. Thus, the test item was stable under the test conditions during the test period of 48 hours and all reported results were related to the nominal concentration of the test item.

Details on test conditions:

TEST SYSTEM
- Test vessel:
Type: closed (covered with glass plates)
Material, size, headspace, fill volume: 100 mL glass beakers filled with 50 mL of test medium.
Aeration: The test water was aerated prior to the start of the study until oxygen saturation was reached. During the test period, the test water was not aerated.
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: The volume of test solution provided for each daphnia was 10 mL. Thus, the requirement of the test guidelines for the minimum volume of 2 mL test medium per daphnia was fulfilled.
The daphnids were randomly distributed to the test vessels at initiation of the test.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water according to ISO 6341 was used in the study. It consisted of analytical grade salts dissolved in purified water at the nominal concentrations as given in the table "Test Water" below.
- Alkalinity: 0.8 mmol/L
- Ca/Mg ratio: 4:1 (based on molarity)
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: A 16-hour light to 8-hour dark cycle with a 30-minute transition period was used.
- Light intensity: Light intensity during the light period was approximately between 520 and 680 Lux.

EFFECT PARAMETERS MEASURED:
- The immobility of the daphnids was determined by visual inspection after 24 and 48 hours of exposure.
- The NOEC and EC0 were determined directly from the raw data.
- The EC50 and the EC100 could not be calculated due to the absence of a toxic effect of the test item.
- At the start and end of the test, the pH values, dissolved oxygen concentrations and water temperature were determined of each treatment.
- The appearance of the test media was visually recorded at the start of the test and after 24 and 48 hours.

TEST CONCENTRATIONS
- Range finding study: yes
The limit test was based on the results of a range-finding test (non-GLP).

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Mortality of control: none
- No remarkable observations were made concerning the appearance of the test medium.
- No mortality was observed in limit test.

Results with reference substance (positive control):

For evaluation of the quality of the daphnia clone and the experimental conditions, potassium dichromate is tested as a positive control twice a year. The result of the latest positive control test in September 2009 were valid.
- EC50/LC50: 48-hour EC50: 1.1 mg/L, study C63680

Reported statistics and error estimates:

The R² fit of the calibration curve used was 1.0000.

Table 1: Effect of test substance on the Mobility of daphnids (Daphnia magna)

Nominal test item concentration [mg/L]	No. of daphnids tested	Immobilized daphnids
		after 24 hours		after 48 hours
		No.	%	No.	%
Control	20	0	0	0	0
100	20	0	0	0	0

Table 2: Dissolved Oxygen Concentrations, pH Values and Temperature in the Treatments.

Nominal test item concentration [mg/L]	Start (0 hours)			End (48 hours)
	pH	Oxygen [mg/L]	Temperature [°C]	pH	Oxygen [mg/L]	Temperature [°C]
Control	7.8	8.7	20	7.9	8.6	20
100	7.8	8.7	20	7.9	8.7	20

 

Validity criteria fulfilled:

yes

Conclusions:

The EC50 for Daphnia was found to be greater than 100 mg/L.

Executive summary:

The acute toxicity of the test substance to daphnids (Daphnia magna) was determined in a 48-hour static test under GLP according to the EU Commission Directive 92/69/EEC, Part C.2, the Commission Regulation (EC) No 440/2008, Part C.2 and the OECD Guideline for Testing of Chemicals, No. 202 (2004). A limit test was performed in accordance with the test guidelines to demonstrate that the test item has no toxic effect on the test organisms up to and including a nominal concentration of 100 mg/L. Thus, the only nominal concentration tested was 100 mg/L. Additionally a control group was tested in parallel. The analytically determined concentration of the test item in the test medium at the start and end of the test was 104 and 103 % of the nominal value, respectively. Thus, the test item test substance was stable under the test conditions during the test period of 48 hours and all reported results were related to the nominal concentration of the test item. In the control and at the test item concentration of 100 mg/L, no immobilized test organisms were observed during the test period of 48 hours. Therefore, the 48-hour NOEC and EC₀ of test substance to the test animals (Daphnia magna) were determined to be at least 100 mg/L. The 48-hour NOEC and EC₀ might even be higher, but concentrations above 100 mg/L were not tested, in accordance with the test guidelines. The 48-hour EC₅₀ and EC₁₀₀ were clearly higher than 100 mg/L. These values could not be quantified due to the absence of toxicity of test substance at the test concentration of 100 mg/L. No remarkable observations were made concerning the appearance of the test medium. The test medium was a clear solution coloured by the test item throughout the whole test duration. At the beginning and end of the test period, the dissolved oxygen concentration in the test medium and control was at least 8.6 mg/L. The pH values of the test medium and control were between 7.8 and 7.9. The water temperature during the test was 20 °C. The test is considered to be valid, as the guideline requirement that not more than 10 % of the daphnids in the control show immobilization or other signs of disease or stress (e.g., discoloration or unusual behaviour such as trapping at the surface water) was fulfilled. Furthermore, the dissolved oxygen concentration at the end of the test was ≥3 mg/L in the control and test vessels. In conclusion, the test substance had no acute toxic effects on daphnids (Daphnia magna) up to the nominal concentration of 100 mg/L under the test conditions.

Description of key information

The substance was found to be not acutely toxic to Daphnia magna at a nominal concentration of 100 mg/L in a static aquatic toxicity test performed for 48 hours.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

The acute toxicity of the test substance to daphnids (Daphnia magna) was determined in a 48-hour static test under GLP according to the EU Commission Directive 92/69/EEC, Part C.2, the Commission Regulation (EC) No 440/2008, Part C.2 and the OECD Guideline for Testing of Chemicals, No. 202 (2004). A limit test was performed in accordance with the test guidelines to demonstrate that the test item has no toxic effect on the test organisms up to and including a nominal concentration of 100 mg/L. Thus, the only nominal concentration tested was 100 mg/L. Additionally a control group was tested in parallel. The analytically determined concentration of the test item in the test medium at the start and end of the test was 104 and 103 % of the nominal value, respectively. Thus, the test item test substance was stable under the test conditions during the test period of 48 hours and all reported results were related to the nominal concentration of the test item. In the control and at the test item concentration of 100 mg/L, no immobilized test organisms were observed during the test period of 48 hours. Therefore, the 48-hour NOEC and EC₀of test substance to the test animals (Daphnia magna) were determined to be at least 100 mg/L. The 48-hour NOEC and EC₀might even be higher, but concentrations above 100 mg/L were not tested, in accordance with the test guidelines. The 48-hour EC₅₀ and EC₁₀₀ were clearly higher than 100 mg/L. These values could not be quantified due to the absence of toxicity of test substance at the test concentration of 100 mg/L. No remarkable observations were made concerning the appearance of the test medium. The test medium was a clear solution coloured by the test item throughout the whole test duration. At the beginning and end of the test period, the dissolved oxygen concentration in the test medium and control was at least 8.6 mg/L. The pH values of the test medium and control were between 7.8 and 7.9. The water temperature during the test was 20 °C. The test is considered to be valid, as the guideline requirement that not more than 10 % of the daphnids in the control show immobilization or other signs of disease or stress (e.g., discoloration or unusual behaviour such as trapping at the surface water) was fulfilled. Furthermore, the dissolved oxygen concentration at the end of the test was ≥3 mg/L in the control and test vessels. In conclusion, the test substance had no acute toxic effects on daphnids (Daphnia magna) up to the nominal concentration of 100 mg/L under the test conditions.