Disodium 4,4'-methylenebis[3-hydroxy-2-naphthoate]

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:

24 OCT 2013 - 5 FEB 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2012

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium für Umwelt, Energie, Landwirtschaft and Verbraucherschutz, Mainzer Straße 80, 65189 Wiesbaden (1 Feb 2013)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all
- Sampling method: For analysis, samples of control and test media were taken at the start and the end of exposure for both exposure periods (0-24 hour, 24-48 hour). The samples were diluted 1:4 with methanol:buffer pH 9.00 (1:1, v:v) and filtered.
- Sample storage conditions before analysis: no

Vehicle:

yes

Remarks:

reconstituted water

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The test medium (reconstituted water and test item) was prepared freshly. For this purpose, the test item was weighed into freshly reconstituted water and then shortly stirred with a mini shaker

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus
- Age at study initiation (mean and range, SD): < 24 h
- Method of breeding: Parent daphnids were used for reproduction until they were about 6 weeks old. Then they were replaced by newborn offspring. They were kept in individual 100 mL test vessels containing approximately 60 mL reconstituted water.
- Source: Daphnia magna Straus of the parent generation was originally bred by IBACON GmbH (Roßdorf, Germany). The clone was bred in the laboratories of Merck KGaA at a water temperature of 20 ± 2°C
- Feeding during test : No

ACCLIMATION
- Acclimation period: To avoid an acclimation phase before start of the study, reconstituted water was used during culture as well as during toxicity studies.
- Acclimation conditions (same as test or not): same
- Type and amount of food: The daphnids were fed ad libitum with unicellular green algae Desmodesmus subspicatus
- Feeding frequency: three times per week
- Health during acclimation (any mortality observed): The daphnids were visually inspected for motility. If daphnids did not make any swimming movements within 15 seconds after their containers have been gently agitated, they were considered to be immobile and were not used for the study.

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES, INCLUDING CULTURING CONDITIONS:
When the study was planned, all offspring were separated from their parents less than 24 hours before start of the experimental part, so on the following morning when exposure began, all offspring were not older than 24 hours. Before start of the experimental phase, the young daphnids were separated from their different parent animals and put into a collective test vessel containing reconstituted water. At the start of the experimental phase the daphnids were cautiously removed using a pipette, separated from the reconstituted water using a fine-mesh sieve, and placed into the test vessels containing test media (test item group) or reconstituted water (control).

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

2.5 mmol/L (250 mg CaCO3/L)

Test temperature:

20.8 - 21.5°C

pH:

7.48 - 7.65

Dissolved oxygen:

7.41 - 8.82 mg O2/L

Nominal and measured concentrations:

nominal: 0, 100 mg/L
measured (0 h): < 0.094 (LOD), 100.489 mg/L (test solution I)
measured (24 h): < 0.094 (LOD), 90.450 mg/L (test solution I)
measured (24 h): < 0.094 (LOD), 98.064 mg/L (test solution II)
measured (48 h): < 0.094 (LOD), 90.562 mg/L (test solution II)

The concentration of the test material has been satisfactorily maintained within ± 20% of the nominal concentration throughout the test and, therefore, the biological results can be based on the nominal value.

Details on test conditions:

TEST SYSTEM
- Test vessel: glass vessel
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: fill volume 20 mL
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): after 24 h
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per vehicle control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
The water was composed according to ELENDT (1990). The reconstituted water contained the following chemicals (analytical grade), which were dissolved in fully demineralized water to obtain the following concentrations:
Main components mg/L
CaCl2 * 2H2O 293.80
MgSO4 * 7H2O 123.30
NaHCO3 64.80
KCl 5.80
Na2SiO3 * 9H2O 10.00
NaNO3 0.27
KH2PO4 0.14
K2HPO4 0.18

Trace elements
B 0.5000
Fe 0.2000
Mn 0.1000
Li, Rb, and Sr 0.0500
Mo 0.0250
Br 0.0125
Cu and Zn 0.0063
Co and I 0.0025
Se 0.0010
V 0.0003
Na2EDTA * 2H2O 2.5000

Vitamins
Thiamin 0.07500
B12 0.00100
Biotin 0.00075

After preparation the reconstituted water was aerated for 24 hours.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hour light and 8 hour dark
- Light intensity: 646 Lux and 664 Lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : 24 and 48 hours after the daphnids had been placed into the reconstituted water (control) or respective test medium (test item group), they were visually examined for their ability to swim. Those daphnids that did not make any swimming movements within 15 seconds after the test vessel had been gently agitated, were regarded as immobilized.

VEHICLE CONTROL PERFORMED: yes

RANGE-FINDING STUDY
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: In a semistatic pretest no immobilization was observed at a concentration of 100 mg/L under open conditions. Therefore, the solution of a nominal test item concentration of 100 mg/L was tested in an open semistatic system.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Mortality of control: no (0/20)
- Immobilisation of control: no (0/20)

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50 (24 h) = 1.200 mg/L; EC50 (48 h) = 0.819 mg/L
Under the given experimental conditions, the test item Potassium dichromate showed a 24h EC50 value which was within the range of the published data of 0.6 to 1.7 mg/L (Council Regulation (EC) No. 440/2008) and 0.6 to 2.1 mg/L (OECD Guideline No. 202).

Validity criteria fulfilled:

yes

Remarks:

In the control group 0% of the introduced daphnids died. The dissolved oxygen concentration at the end of the test was ≥ 3 mg/L. Thus, the validity criteria were fulfilled.

Conclusions:

The test item solved in reconstituted water was tested in an open semistatic system. Under the conditions of the present study, an aqueous solution of 100 mg/L revealed no aquatic toxicity in the test system. The EC50 (48 h) was > 100 mg/L.

Executive summary:

The GLP compliant study was performed according to OECD TG 202 and EU Method C.2. The objective of this study was to evaluate the influence of the test item on the immobilization of Daphnia magna. For this purpose, juvenile daphnids were exposed to a nominal test item concentration of 100 mg/L (limit test) over 48 hours, in an open semistatic system. The daphnids were observed for immobilization 24 and 48 hours after placing in the aqueous test item solution. Due to the results of a stability determination before start of study, a semistatic test system was chosen and the test media was renewed after 24 hours. The study comprised of four test vessels in the test item group containing five daphnids each, i.e. 20 daphnids in total. Additionally, one control group (20 daphnids) was used. The analytical recovery of the first preparation (0-24h) was 100.5 and 90.5% after 0 and 24 hours, respectively. The recovery of the second preparation (24-48h) was 98.1 and 90.6% after 0 and 24 hours, respectively. As the stability of the test material in the aqueous solution was ≥ 80%, the EC₅₀ values were calculated with the nominal concentrations. Daphnids exposed to an aqueous preparation of a nominal concentration of 100 mg/L were not affected. Thus, the following nominal EC₅₀ values for daphnids were determined: EC₅₀ (24 h) > 100 mg/L EC₅₀ (48 h) > 100 mg/L.

Description of key information

Key, immobilisation, Daphnia magna, semistatic, open, OECD 202, GLP: EC₅₀(48 h) > 100 mg/L (nominal)

Additional information

The GLP compliant study was performed according to OECD TG 202 and EU Method C.2. The objective of this study was to evaluate the influence of the test item on the immobilisation of Daphnia magna. For this purpose, juvenile daphnids were exposed to a nominal test item concentration of 100 mg/L (limit test) over 48 hours, in an open semistatic system. The daphnids were observed for immobilization 24 and 48 hours after placing in the aqueous test item solution. Due to the results of a stability determination before start of study, a semistatic test system was chosen and the test media was renewed after 24 hours. The study comprised of four test vessels in the test item group containing five daphnids each, i.e. 20 daphnids in total. Additionally, one control group (20 daphnids) was used. The analytical recovery of the first preparation (0-24h) was 100.5 and 90.5% after 0 and 24 hours, respectively. The recovery of the second preparation (24-48h) was 98.1 and 90.6% after 0 and 24 hours, respectively. As the stability of the test material in the aqueous solution was ≥ 80%, the EC₅₀ values were calculated with the nominal concentrations. Daphnids exposed to an aqueous preparation of a nominal concentration of 100 mg/L were not affected. Thus, the following nominal EC₅₀ values for daphnids were determined: EC₅₀ (24 h) > 100 mg/L, EC₅₀ (48 h) > 100 mg/L.