reaction mass of sodium 1-amino-4-[[3,5-bis[[(chloroacetyl)amino]methyl]-2,4,6-trimethylphenyl]amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate and sodium 1-amino-4-[[3-[[(chloroacetyl)amino]methyl]-2,4,6-trimethylphenyl]amino]-9,10-dihydro-9,10-dioxoanthracene-2-sulphonate

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

November from 17 to 21, 2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

adopted 17 July, 1992

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

adopted 31 July, 1992

Principles of method if other than guideline:

The study was conducted in accordance with ISO/IEC 17025.

GLP compliance:

no

Analytical monitoring:

no

Details on test solutions:

The individual test concentrations were prepared by dilution of a 200 mg/l stock solution.

Test organisms (species):

Poecilia reticulata

Details on test organisms:

TEST ORGANISM
- Common name: guppy.
- Length at study initiation: 1-3 cm .
- Validity: during 7 days before the definitive test, mortality in the batch of which fish were used for the test was less than 5 %.

BREEDING
- Breeding: parentaland young fish population was held in two 250 I glass aquaria.
- lllumination: 14 hours a day.
- Temperature: 23 ± 2 °C
- Medium: continuously aerated and filtered tap water from Schlieren. The bottom of the aquaria was covered with gravel (<1.5 cm). Every month, one third of the total volume of the aquaria was replaced by fresh tap water.
- Feed: Sera micropan, Sera vipan (Sera apanristic GmbH, D-52525 Heinsberg, Germany) daily. The fish were last fed 24 h betore the start of the definitive test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

21 ± 1 °C

pH:

Test vessels: 8.0 - 8.6
Control: 8.4 - 8.6

Dissolved oxygen:

Test vessels: 6.8 - 7.3 mg/l
Control: 6.7 - 7.8 mg/l

Nominal and measured concentrations:

Nominal 1, 10 and 100 mg/

Details on test conditions:

TEST SYSTEM
- Test vessel: 1 I aquaria, all-glass.
- No. of organisms per concentration: 7 fish per test concentration.
- No. of vessels per concentration: 1 replicate.
- No. of vessels per control: 1 replicate.

TEST MEDIUM / WATER PARAMETERS
- Preparation of dilution water: tap water from Schlieren, aerated. At least 1 h before the test the test vessels were filled with the medium to allow equilibration of pH, oxygen and temperature.

OTHER TEST CONDITIONS
- Adjustment of pH: pH was not adjusted.
- Photoperiod: 16 h photoperiod a day, supplied by overhead white fluorescent tubes.

EFFECT PARAMETERS MEASURED
Fish were considered to be dead when they showed no reaction after touching the caudal fin or no visible breathing movements.

PARAMETERS MEASURED
- Oxygen, pH, temperature: prior to the addition of fish to the test medium and after 2, 24, 48, 72 and 96 h of exposure.
- Observations: made after 2, 24, 48,72 and 96 h of exposure. Mortality of the fish was recorded at the same time intervals. Dead fish were removed immediately from the test medium and weight and length recorded.

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

10 - 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

All fish were dead at the nominal concentration of 100 mg/l after 2 hours of exposure. No mortality was observed at 10 and 1 mg/l and in the control after 96 h of exposure.
Other toxic effects than mortality, e.g. loss of coordination, hypoactivity and swimming on the back, were not observed.

Sublethal observations / clinical signs:

Incidence of mortality observed.

Nominal concentration (mg/l)	Number of dead fish observed at different points in time after start of exposure
	2 hrs	24 hrs	48 hrs	72 hrs	96 hrs
Control	0	0	0	0	0
Test item 1 mg/l	0	0	0	0	0
Test item 10 mg/l	0	0	0	0	0
Test item 100 mg/l	7	*	*	*	*

* All fish died, no further observations.

Nominal concentration (mg/l)	Number of dead fish observed at different points in time after start of exposure
	2 hrs	24 hrs	48 hrs	72 hrs	96 hrs
pH conditions
Control	8.4	8.6	8.6	8.6	8.5
Test item 1 mg/l	8.1	8.6	8.5	8.5	8.5
Test item 10 mg/l	8	8.5	8.5	8.5	8.5
Test item 100 mg/l	8.4	*	*	*	*
Oxygen concentrations (mg/l)
Control	7.8	7.1	7.1	6.9	6.7
Test item 1 mg/l	7	7.3	7	6.8	6.8
Test item 10 mg/l	7.1	7.2	7	7	7
Test item 100 mg/l	7.3	*	*	*	*
Temperature (°C)
Control	21	21.5	21.5	22	22
Test item 1 mg/l	21	21.5	21.5	22	22
Test item 10 mg/l	21	21.5	21.5	22	22
Test item 100 mg/l	21	*	*	*	*

* All fish died, no further observations.

Conclusions:

LC50 (96h): 10 -100 mg/l (nominal)

Executive summary:

The acute lethal toxicity of test item to Poecilia reticulata (Guppy) was investigated under static exposure conditions over a period of 96 h. The nominal applied test concentrations were 1, 10 and 100 mg/l. Therefore, the study represents a screening test. The individual test concentrations were prepared by dilution of a 200 mg/l stock solution. No chemical analyses of the test concentrations were conducted.

All fish were dead at the nominal concentration of 100 mg/l after 2 h of exposure. No mortality was observed at 10 and 1 mg/l and in the control after 96 h of exposure. Other toxic effects than mortality, e.g. loss of coordination, hypoactivity and swimming on the back, were not observed.

Based on the recorded data the nominal lethal concentration (LC50) of test item to Poecilia reticulata was estimated to be 10-100 mg/l. The no-observed-effect concentration (LC0) was 10 mg/l; while, the 100 % mortality (LC100) was observed at the nominal concentration of 100 mg/l after 2 h of exposure.

Conclusion

LC50 (96h): 10 -100 mg/l (nominal)

Description of key information

LC50 (96h): 10 -100 mg/l (nominal)

Additional information

The acute lethal toxicity of Acid Blue 225 to Poecilia reticulata (Guppy) was investigated under static exposure conditions, over a period of 96 hours. The experiment was conducted in accordance with the OECD guideline 203. The nominal applied test concentrations were 1, 10 and 100 mg/l. No chemical analyses of the test concentrations were conducted.

All fish were dead at the nominal concentration of 100 mg/l after 2 hours of exposure. No mortality was observed at 10 and 1 mg/l and in the control after 96 hours of exposure. Other toxic effects than mortality, e.g. loss of coordination, hypoactivity and swimming on the back, were not observed.