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Long-term toxicity to aquatic invertebrates

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Reference
Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
according to
Guideline:
other: Provisional Procedure: extended toxicology test with Daphnia magna, Recommendation of the Federal Environmental Agency on the Performance of Testing, Section 5, para 1, No. 3 of the Regulation on Application Documents and Evidence under the Chemicals Act
Version / remarks:
1984
GLP compliance:
no
Analytical monitoring:
yes
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Before preparing the dilution series, the substances to be tested were fully dissolved (both quantitatively and optically) in dilution water using magnetic stirrers (stock solution). In the case of substances of low water solubility, efforts were made to dissolve them by means of up to 24 h stirring. Solutions which were not optically clear after dissolution were filtered over fibre-glass filters and the filtrate was quantified chemically. From the stock solution of the substance to be tested, graduated dilutions with dilution water were produced in the concentration range in which effects were to be expected.
- Differential loading: No
- Controls: Yes
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain/clone: IRCHA
- Source: in-house culture
- Age of parental stock at test begin: 24 h
- Feeding during test: Yes
- Food type: dry algae (Scenedesmus)
- Amount: 2 mL of a solution containing 9 g feed in 1 L
- Frequency: Only on the days when the test media were replaced in the case of closed test vessels (bottles with ground-glass stoppers).

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES:
In order to obtain 24 h-old animals on the potential preparation days in a 21 d test series - Wednesdays or Fridays - it was necessary to remove the offspring from the cultivation beakers on Tuesday and/or Thursday. The daphnia which were at most 24 h old were removed by pipette and concentrated on a 0.25 mm DIN sieve, placed in as small an amount of dilution water as possible and used as test organisms.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Hardness:
16 °dH
Test temperature:
25 ± 1 °C
pH:
7.6 - 7.7 (at test begin)
Dissolved oxygen:
min. 58% (bottles) - 69% (beakers) saturation (at the end of the test)
Nominal and measured concentrations:
nominal: 0.063 - 8 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: wide-neck bottles (in the case of volatile or strongly smelling substances)
- Type: closed with ground-glass stoppers
- Material, size, headspace, fill volume: Size = 400 mL (beakers), 250 mL (bottles); Fill volume = 250 mL
- Renewal rate of test solution (frequency/flow rate): 3 x/week: Mondays, Wednesdays, and Fridays
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 1 daphnid/50 mL

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionized water and tap water
- Culture medium different from test medium: yes. Daphnia were cultured in dechlorinated and oxygen-saturated tap water. In the test and control preparations, an artifical medium (synthetic fresh water) was used.
- Intervals of water quality measurement: The pH and the oxygen concentration were measured when test and control media were changed.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 9 h light; 15 h dark
- Light intensity: fluorescent lamps, Philips TL 65/33W

EFFECT PARAMETERS MEASURED:
- Mortality of the parent animals: daily
- Reproduction rate: daily
- Apperance of the first offspring during the test: daily

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1:2 ratio or if it was necessary to use a concentration range of 3-4 logs in the test, then the dilution series was set up with a ratio of 1:10E-02.
- Range finding study: yes
Reference substance (positive control):
not specified
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.3 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: appearance of first offspring
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.3 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
other: appearance of first offspring
Reported statistics and error estimates:
The Student´s t-test and the U-test were the statistical methods used to calculate the the NOEC based on the mortality of the parent animals and the reproduction rate. The U-test did not require the normal distribution of the test results and was therefore more suitable for evaluating the 21 d Daphnia reproduction test. The highest concentration level, which did not differ (NOEC) from the control, and the lowest, which did differ from the control in respect of mortality of the parent animals and of the reproduction rate, could be ascertained from the calculations. Moreover, in order to determine the NOEC, observations and evaluations were also made of the appearance of the first offspring in the test preparations in comparison with the control preparations.

Description of key information

NOEC (21 d): 0.5 mg/L (nominal, based on reproduction of D. magna);

NOEC (21 d): 0.3 mg/L (minimum value, i.e. lowest analysed concentration, based on reproduction of D. magna)

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater invertebrates:
0.3 mg/L

Additional information

There is one scientific study available investigating the long-term toxicity to aquatic invertebrates. D. magna were exposed under semi-static conditions to nominal concentrations of 0.063 - 8 mg/L for 21 d in closed wide-neck bottles due to volatility of the substance. Test concentrations were verified by analytical measurements. The derived NOEC (21 d) was 0.5 mg/L (nominal) and 0.3 mg/L (measured) based on reproduction of D. magna.