Mixture of hexa(sodium,lithium) 4-{[1,7-disulfonato-5-hydroxy-6-({4-[(6-methoxy-5-sulfonatobenzoheteromonocycle-2-yl)diazenyl]-5-alkyl-2-(3-sulfonatopropoxy)phenyl}diazenyl)naphthalen-2-yl]diazenyl}-5-hydroxy-1-(4-sulfonatophenyl)heteromonocycle-3-carboxylate and hexa(sodium,lithium) 4-{[1,7-disulfonato-5-hydroxy-6-({4-[(6-methoxy-7-sulfonatobenzoheteromonocycle-2-yl)diazenyl]-5-alkyl-2-(3-sulfonatopropoxy)phenyl}diazenyl)naphthalen-2-yl]diazenyl}-5-hydroxy-1-(4-sulfonatophenyl)heteromonocycle-3-carboxylate

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20-24/12/2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed according to OECD guidelines and following GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

other: Alga growth inhibition test, Daphnia Sp. acute immobilisation test, and fish acute toxicity test"(Yakushokuhatsu No. 1121002, Heisei 15.11.13, Seikyoku No. 2, Kanpokihatsu No.031121002, November 21, 2003; the latest revision, November 20, 2006

GLP compliance:

yes

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material (as cited in study report): E-BK105
- Substance type: black powder
- Physical state: solid
- Lot/batch No.: MB-1
- Stability under test conditions: stable
- Storage condition of test material: the test substance was stored in a desiccator (room temperature, shaded, air-tight)

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

no

Details on test solutions:

The test solutions were prepared by dissolving 500 mg substance in 5000 ml dechlorinated water. The solution was mixed by stirring with a magnetic stirrer for 5 min. New test solutions were prepared at 0 and 48 h

Test organisms

Test organisms (species):

Oryzias latipes

Details on test organisms:

TEST ORGANISM
- Common name: Medaka
- Strain: Oryzias latipes
- Source: Tsunashima Fishing Co.
- Age at study initiation (mean and range, SD): less than 1/2 year after hatching
- Length: 2.02 cm (1.77-2.30 cm), n=10 (measurements for the control fish at the end of exposure)
- Weight: 0.069 g (0.046-0.097 g), n=10 (measurements for the control fish at the end of exposure)
- Feeding during test: none


ACCLIMATION
- Acclimation period: 13-20 December, 2010
- Acclimation conditions (same as test or not): yes
- Type and amount of food: TetraMin, TetraWerke
- Feeding frequency: daily until 24 h before exposure
- Health during acclimation (any mortality observed): less than 5%

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Test conditions

Hardness:

48 mg CaCO3/L

Test temperature:

23.1-23.6

pH:

7.5-7.9

Dissolved oxygen:

7.1-8.3 mg/L

Nominal and measured concentrations:

Nominal: 100 mg/L
Measured: 99 mg/L (mean)

Details on test conditions:

TEST SYSTEM
- Test vessel: 5 L glass
- Type: closed
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): batchwise renewal after 48 h exposure
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 10 fish per concentration

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:dechlorinated water processed by YOKOHAMA city tap water
- Total organic carbon: not detected
- Metals: only Al measured at 0.05 mg/L
- Pesticides: not detected
- Chlorine: not detected
- Alkalinity: at pH 4.8, 37 mg CaCO3/L
- Ca/mg ratio: 14/3 mg/L
- Culture medium different from test medium: no
- Intervals of water quality measurement: every six months

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light/ 8 h dark
- Light intensity: 1000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : median lethal concentration, every 24 h

TEST CONCENTRATIONS
- Spacing factor for test concentrations: only 0 and 100 mg/L tested
- Range finding study: yes
- Test concentrations: 0; 1; 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: for all test concentrations there were no mortality or toxicological symptoms of living fish observed

Reference substance (positive control):

yes

Remarks:

copper sulfate

Results and discussion

Effect concentrations

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

measured concentration was within 20% of nominal

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: no
- Observations on body length and weight: not reported
- Other biological observations: not reported
- Mortality of control: no
- Other adverse effects control: no
- Abnormal responses: 1 fish was deadafte 96 h at 100 mg/L
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no floating solids or precipitations observed
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

- Results with reference substance valid? yes
- LC50: 0.57 mg/L (95% C.I. 0.39-0.80 mg/L). The test was performed on December 6, 2010. The average LC in this laboratory ( n=59, since 1995) 0.81 +/- 0.27 mg/L

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the study, E-BK 105 did not cause mortality in fish up to 100 mg/L.