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Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
31.08. - 18.09.2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Version / remarks:
2008
Deviations:
yes
Remarks:
The study was performed without the analytical determination of the test substance concentrations. For all evaluations and results nominal concentrations of the test substance were used.
GLP compliance:
yes
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
Since the test substance is poorly soluble in dilution water at concentrations needed for the test, a stock saturated solution (nominal concentration 100 mg/L) of the test substance in dilution water was prepared for preliminary and limit test.
The stock saturated solution was prepared as follows: the suspension of 100 mg of the test substance in 1000 mL of dilution water was stirred for suitable period on a shaft stirrer and consequently filtered through 0.45 μm filter.
An experiment was performed for the determination of suitable period of stirring in procedure of saturated solution preparation. During this experiment the concentrations of metals (Al, Ca, Fe, K, Mg, Na, P and Ti) in resulted solutions were determined. The saturated solutions were prepared in dilution water, which contents metals as Ca, Mg, Na and K. For this reason the analytical determination of the concentrations of metals was performed also in pure dilution water.
Test organisms (species):
Poecilia reticulata
Details on test organisms:
TEST ORGANISM
Poecilia reticulata
- Source: certified supplier EMPLA s.r.o., Hradec Králové (Czech Republic)
- Age at study initiation (mean and range, SD): 3-4 months
- Length at study initiation (length definition, mean, range and SD): 2-3 cm
- Housing: The fish were kept in all-glass aquaria in aerated drinking water.
- Diet: “Tropica” - feed for aquarium fish, ad libitum and sometimes live daphnia.
- Medical treatment: None
- Weight at study initiation (mean and range, SD):
- Feeding during test: without feeding


ACCLIMATION
- Acclimation period: 7 days in dilution water; they were not fed during 72 hours before the test.
- Acclimation conditions (same as test or not): same as the test
- Type and amount of food: Tropica” - feed for aquarium fish, ad libitum and sometimes live daphnia.
- Health during acclimation (any mortality observed): healthy animals

STUDY TIME SCHEDULE
Experimental part of study: 31.08. – 18.09.2009
Evaluation of results and final report elaboration: 18.09. – 23.10.2009
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Hardness:
Preliminary test: content of (Ca + Mg) ions 2.44 mmol/L
Limit test: content of (Ca + Mg) ions 2.50 mmol/L
Test temperature:
22 ± 2°C, with a variation of ± 1°C at each individual test
pH:
Preliminary test: pH of dilution water 7.7
Limit test: pH of dilution water 7.7
Dissolved oxygen:
Preliminary test: 8.8 mg/L at temperature of 21.8 C
Limit test: 7.7 mg/L at temperature 21.5 °C
Conductivity:
Preliminary test: 0.107 mS·m-1
Limit test: 0.112 mS·m-1
Nominal and measured concentrations:
Preliminary test: 100, 50, 10, 5, 1 mg/L
Limit test: 100, 91, 83, 75, 68, 62 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type: open
- Material, size, headspace, fill volume: glass aquariums, fill volume 2 l
- Aeration: without aretation
- No. of organisms per vessel: 7 for preliminary test, 10 for limit test
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 1g per liter

TEST MEDIUM / WATER PARAMETERS
Standard dilution water was prepared just before the test by admixing 2.5 mL of stored solutions A, B, C and D in 1000 mL of deionized water.
A. 117.6 g CaCl2.2H20 in 1L deionized water
B. 49.3 g MgSO4.7H2O in 1L deionized water
C. 25.9 g NaHCO3 in 1L deionized water
D. 2.3 g KCl in 1L deionized water
The dilution water should be of pH 7.8 ± 0.2. It is saturated by oxygen by the aeration at temperature of 20 – 24 °C. The content of sum Ca + Mg ions should have the value of 2.5 mmol/L.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: natural variation of daily light and dark
- Light intensity: day light

EFFECT PARAMETERS MEASURED
The fish mortality was observed each 24 hours. The pH, temperature and dissolved oxygen were measured at the same intervals.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.1
- Justification for using less concentrations than requested by guideline: Since the test substance is poorly soluble in dilution water at concentrations needed for the test, a stock saturated solution (nominal concentration 100 mg/L) of the test substance in dilution water was prepared for preliminary and limit test.
- Range finding study: 100, 50, 10, 5, 1 mg/L
- Test concentrations: 100, 91, 83, 75, 68, 62 mg/L (nominal concentration)
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
24 h
Dose descriptor:
LC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
72 h
Dose descriptor:
LC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
83 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
83 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
83 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
83 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
24 h
Dose descriptor:
LC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
LC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
72 h
Dose descriptor:
LC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
96 h
Dose descriptor:
LC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Behavioural abnormalities: not observed
- Observations on body length and weight:not observed
- Other biological observations: not observed
- Mortality of control: none
- Other adverse effects control: not observed
- Abnormal responses:not observed
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Mortality: yes
The sensitivity of the test species and correctness of test performance is periodically verified in three-month period by testing the reference substance, potassium dichromate.
The results of verification test with K2Cr2O7, carried out in period from 08.06. to 12.06.2009, are the following:
48h – LC50 = 147 mg/L (95 % confidence limit: 118 – 176 mg/L)
96h – LC50 = 120 mg/L (95 % confidence limit: 103 – 137 mg/L)

The results of the last interlaboratory test arranged by Centre for Assessment of Laboratories of T.G.Masaryk Water Research Institute Prague in 2008 for potassium dichromate were:
96 h - LC50 = 81.2 – 190 mg/L
The mortality of test organisms in the reference substance solution meets criteria determined by the interlaboratory test.

Table No. 1 - Fish mortality – preliminary test

Nominal concentration

mg·L-1

100

50

10

5

1

C

Saturated solution dosing

mL·L-1

1000

500

100

50

10

C

Number of live fish

2 hours

7

7

7

 7

7

7

Mortality %

2 hours

0

0

0

0

0

0

Number of live fish

24 hours

7

7

7

 7

7

7

Mortality %

24 hours

0

0

0

0

0

0

Number of live fish

48 hours

7

7

7

 7

7

7

Mortality %

48 hours

0

0

0

0

0

0

Number of live fish

72 hours

7

7

7

 7

7

7

Mortality %

72 hours

0

0

0

0

0

0

Number of live fish

96 hours

7

7

7

 7

7

7

Mortality %

96 hours

0

0

0

0

0

0

Table No. 2 - Fish mortality – limit test

Nominal concentration

mg·L-1

100

91

83

75

68

62

C

Saturated solution dosing

mL·L-1

1000

910

830

750

680

620

C

Number of live fish

2 hours

10

10

10

10

10

10

10

Mortality %

2 hours

0

0

0

0

0

0

0

Number of live fish

24 hours

10

9

10

10

10

10

10

Mortality %

24 hours

0

10

0

0

0

0

0

Number of live fish

48 hours

10

9

10

10

10

10

10

Mortality %

48 hours

0

10

0

0

0

0

0

Number of live fish

72 hours

10

9

10

10

10

10

10

Mortality %

72 hours

0

10

0

0

0

0

0

Number of live fish

96 hours

10

9

10

10

10

10

10

Mortality %

96 hours

0

10

0

0

0

0

0

Validity criteria fulfilled:
yes
Conclusions:
The highest observed mortality was 10 % in the limit test. Therefore exact values of LC50 could not be calculated and the values of LC are given in the form of a range.
The NOEC values are identified by direct observation. The nominal concentrations of the test substance were used for all evaluations and results.
Test results
24h – LC50 > 100 mg/L (nominal concentration)
48h – LC50 > 100 mg/L (nominal concentration)
72h – LC50 > 100 mg/L (nominal concentration)
96h – LC50 > 100 mg/L (nominal concentration)

24h – NOEC = 83 mg/L (nominal concentration)
48h – NOEC = 83 mg/L (nominal concentration)
72h – NOEC = 83 mg/L (nominal concentration)
96h – NOEC = 83 mg/L (nominal concentration)

24h – LC100 > 100 mg/L (nominal concentration)
48h – LC100 > 100 mg/L (nominal concentration)
72h – LC100 > 100 mg/L (nominal concentration)
96h – LC100 > 100 mg/L (nominal concentration)
Executive summary:

The test substance, Semi Dry Absorption (SDA) Product, was tested for acute toxicity on fish Poecilia reticulata.

The test was performed according to method C.1 – Acute toxicity for fish, Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008.

Since the test substance is poorly soluble in dilution water at concentrations needed for the test, a stock saturated solution (nominal concentration 100 mg·L-1) of the test substance in dilution water was prepared for preliminary and limit test.

The preliminary test was performed in range of test substance nominal concentrations 1 – 100 mg·L-1. Because no mortality of fish was observed in the preliminary test, the limit test was performed. The limit test was performed in range of test substance nominal concentrations 62 – 100 mg·L-1. The test was performed as static.

The study was performed without the analytical determination of the test substance concentrations. The nominal concentrations of the test substance were used for all evaluations and results. Only concentrations of metals in stock saturated solution were analysed in the limit test.

Test results:

24h – LC50 > 100 mg·L-1 (nominal concentration)

48h – LC50 > 100 mg·L-1 (nominal concentration)

72h – LC50 > 100 mg·L-1 (nominal concentration)

96h – LC50 > 100 mg·L-1 (nominal concentration)

 

24h – NOEC = 83 mg·L-1 (nominal concentration)

48h – NOEC = 83 mg·L-1 (nominal concentration)

72h – NOEC = 83 mg·L-1 (nominal concentration)

96h – NOEC = 83mg·L-1 (nominal concentration)

 

24h – LC100 > 100 mg·L-1 (nominal concentration)

48h – LC100 > 100 mg·L-1 (nominal concentration)

72h – LC100 > 100 mg·L-1 (nominal concentration)

96h – LC100 > 100 mg·L-1 (nominal concentration)

 

Classification of the test substance on the basis of hazard to aquatic environment was performed according to the Directive 93/21/EEC, Annex IV (article 5.2.1.).

According to the classification criteria given in quoted Directive the test substance, Semi Dry Absorption (SDA) Product, is not assigned to any of quoted toxicity categories.

Description of key information

EU Method C.1 – Acute toxicity for fish, Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008.

GLP study.

Key value for chemical safety assessment

LC50 for freshwater fish:
100 mg/L

Additional information

The highest observed mortality was 10 % in the limit test. Therefore exact values of LC50 could not be calculated and the values of LC are given in the form of a range.

The NOEC values are identified by direct observation. The nominal concentrations of the test substance were used for all evaluations and results.

Test results

24h – LC50 > 100 mg/L (nominal concentration)

48h – LC50 > 100 mg/L (nominal concentration)

72h – LC50 > 100 mg/L (nominal concentration)

96h – LC50 > 100 mg/L (nominal concentration)

 

24h – NOEC = 83 mg/L (nominal concentration)

48h – NOEC = 83 mg/L (nominal concentration)

72h – NOEC = 83 mg/L (nominal concentration)

96h – NOEC = 83 mg/L (nominal concentration)

 

24h – LC100 > 100 mg/L (nominal concentration)

48h – LC100 > 100 mg/L (nominal concentration)

72h – LC100 > 100 mg/L (nominal concentration)

96h – LC100 > 100 mg/L (nominal concentration)