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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
14 Feb - 20 April 2012
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
no detailed analysis of the WAF
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Principles of method if other than guideline:
closed system
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The Water Accommodated Fractions (WAFs) for the Range-Finding- and Definite Daphnia-Test with Bis-(isopropyl)naphthalene were prepared by
stirring various amounts of the test item in Daphnia-medium for 24 h with magnetic stirrers.
The WAFs were prepared in closed 2 L decanters. The test loading rates of Bis-(isopropyl)naphthalene were weighed in weighing scoops that
afterwards were placed into the decanters. The decanters were then filled with Daphnia medium and closed immediately.
The mixing was carried out at a speed that was slow enough not to cause dispersion or emulsification of the undissolved fraction of the test item.
To ensure this, the vortex developed at the surface by stirring was set at ~ 10 % of the water depth. After stirring for 24 h the WAFs were allowed
to stand for 1 h before use to facilitate phase separation and were then transferred directly into the test vessels, i.e. 50 mL glass vessels.
The test vessels were closed with ground-in glass stoppers directly after the transfer.

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Clone 5
- Source: German Envirinmental Ministry
- Age at study initiation: <= 24 h
- Cultivation in a climate controlled room (light/dark = 14 h/10 h; illumination: Fluorescent tubes)
- Type and amount of food: Algae (Desmodesmus subspiccatus)
- Feeding during test: none

ACCLIMATION
- Acclimation period:
- Acclimation conditions (same as test or not):
- Type and amount of food: Algae (Desmodesmus subspiccatus)
- Feeding frequency:
- Health during acclimation (any mortality observed):

Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
no data
Test temperature:
20°C +/- 2°C
pH:
8.0 -8.3 (measured at 0, 24, and 48 h)
Dissolved oxygen:
7.9 - 8.4 mg/ml (measured at 0, 24, and 48 h)
Salinity:
no data
Nominal and measured concentrations:
Range finding test: WAF (Water accommodated fraction) in test medium, 1.0, 10.0, 100 mg/L nominal
Definitive test: WAF (Water accomodated fraction) in test medium, 1.0, 2.5, 5.0, 10, and 25 mg/L nominal
Details on test conditions:
TEST SYSTEM
- Test vessel: 50 ml glass vessels, closed with ground-in glass stoppers, test volume 50 mL
- Type: closed
- Aeration: none
- Renewal rate of test solution: - static (range finding test),
- semi static, renewal of test solution after 24 h (definite test)
- No. of organisms per vessel: 5
- No. of vessels per concentration and control (replicates): 2 (range finding test), 4 (definitive test)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionised water
- Total organic carbon: no data
- Chlorine: no data
- Alkalinity: 0.8 mmol/l
- Na/K ratio: 10:1
- Ca/Mg ratio: 4:1
- Conductivity: no data
- Culture medium different from test medium: no
- Intervals of water quality measurement: no data

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light/ 8 h dark (fluorescent tubes)
- Light intensity: no data

EFFECT PARAMETERS MEASURED: Immobilised daphnia, immobilised daphnia that were trapped at the surface of the water, daphnia that were not
immobilised but trapped at the surface of the water, daphnia that were not immobilised but weak and were observed mostly at the bottom of the test vessel, number of carapace moults.
The observations were made at 0 h, 24 h and 48 h.

TEST CONCENTRATIONS
- Preparation of test material: The test item Bis-(isopropyl)naphthalene is a liquid poorly soluble in water comprising volatile components. To avoid
adverse physical effects of undissolved or emulsified test material on the animals, the test was carried out with aqueous extracts
(WAF = Water Accommodated Fraction) at various loading rates in Daphnia-medium.
- Range finding study: with concentrations 1, 10, 100 mg/L
- Test concentrations (definitive test): 1, 2.5, 5, 10, 25 mg/L (due to the results of the range finding test)
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
1.7 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
NOELR
Effect conc.:
< 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: derived from the result of immobilisation (20 %) at the loading rate of 1.0 mg/L
Key result
Duration:
48 h
Dose descriptor:
EL10
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: from dose-response curve at 1 mg/L minus 10% mobility in control.
Duration:
48 h
Dose descriptor:
other: EL90
Effect conc.:
ca. 3.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Mortality of control: 1 of 20 animals after 48 h
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no data
Reported statistics and error estimates:
Determination of the EL50 (EL = Effective Loading) of the immobilisation rate of the Definite Test was accomplished using Probit-analysis
(Finney-method, lognormal distribution; confidence limit 95 %, significance level: 0.05).

Immobilisation rate in dependence of time and loading rate, number of carapace moults, and number of living animals trapped at the surface

[number per total animals, %], (excerpt from original report).

Control

1

mg/L

2.5

mg/L

5

mg/L

10

mg/L

25

mg/L

Num-ber

[%]

Num-ber

[%]

Num-ber

[%]

Num-ber

[%]

Num-ber

[%]

Num-ber

[%]

Immobilisation
0 – 24 h start

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

Immobilisation
0 – 24 h end

0/20

0

0/20

0

2/20

10

19/20

95

13/20

65

15/20

75

Immobilisation
24 – 48 h start

0/20

0

0/20

0

2/20

10

19/20

95

13/20

65

15/20

75

Immobilisation
24 – 48 h end

1/20

5

4/20

20

15/20

75

19/20

95

20/20

100

20/20

100

Moults 48 h

15/20

75

16/20

80

5/20

25

3/20

15

5/20

25

6/20

30

Living animals at
the surface 48 h

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

Immobilisation: 20 % at 1 mg/L, 75 % at 2.5 mg/L, 95 % at 5 mg/L and 100 % at 10 mg/L, and 25 mg/L after 48 h.

Validity criteria fulfilled:
yes
Remarks:
No more than 10 % immobilisation of the Control daphnia. The dissolved oxygen concentration during the test should be >= 3 mg/L and must not be < 2 mg/L.
Conclusions:
The test item Bis-(isopropyl)naphthalene, mixture of isomers, is considered to be acutely toxic to daphnia.
[Test items with an EL50 (48h) of < 100 mg/L are considered to be acutely toxic to daphnia]
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: DIN 38412, Teil 11 - Bestimmung der Wirkung von Wasserhaushaltsstoffen auf Kleinkrebse (Daphnien-Kurzzeittest)
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
yes
Remarks:
: only 10 test organisms instead of 20 / Chemical analysis only in stock solution
Principles of method if other than guideline:
National test guideline
GLP compliance:
no
Analytical monitoring:
yes
Details on sampling:
Only the stock solution was analysed for the test substance concentration.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 0.5 mg/L test substance in dilution water was stirred for 48 h and then filtrated over glass fiber filters.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain: IRCHA
- Source: no data
- Age at study initiation (mean and range, SD): < 24 h
- Feeding during test: no
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 1°C
pH:
7.8 -8.1
Dissolved oxygen:
98.5 - 100% O2 saturation
Nominal and measured concentrations:
Test solutions
nominal concentrations: 0.05, 0.066, 0.1, and 0.16 mg/L
Stock solution
nominal concentration: 0.5 mg/L
measured concentration: 0.2 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: no data
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: artificial freshwater according to DIN 38412, part 11

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: test performed in the dark (no lighting period)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : mobility

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.3 - 1.6
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
0.16 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 0.16 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility

No immobilization was noted after 24 h.

 

 

Results after 48 h

 

Test concentration
[mg/L]

Animals alive

Animals dead
(immobilized)

pH
(final)

O2
(final) [%]

0 (control)

10

0

8.09

98.9

0.16

10

0

7.85

98.5

0.10

8

2

7.89

100

0.066

9

1

7.98

98.9

0.005

10

0

7.82

98.9

 

 

No effect was observed in the range of water-solubility.

Validity criteria fulfilled:
yes
Conclusions:
In an acute 48 h daphnia immobilization test, diisopropylnaphthtalene did not exhibit an immobilizing effect on the test organisms within the range of water-solubility. The EC50 was determined to be > 0.16 mg/L (nominal concentration).

Description of key information

Under closed test conditions, diisopropylnaphthalene produced significant acute intoxication of the daphnia at low loadings: The EL50 (as loading) was 1.7 mg/L. In early studies, only slight effects on mobility of daphnia were observed in short-term toxicity tests within the range of water solubility of the test substance (0.1 to 0.2 mg/L).

Key value for chemical safety assessment

Additional information

For assessment of the short-term toxicity to aquatic invertebrates, two valid studies are available. In a third study, a vehicle was used to solubilise test substance and the exposure period was only 24 h. The recent study (Aniol et al 2012) was conducted with water accommodated fractions (WAF) of different loadings under closed conditions.

 

Study

Test concentration

Result

Experimental conditions

Reliability

AniolS, Blum Th, Honnen W 2012 (Daphnia magna)

Nominal/loading:

1.0, 2.5, 5.0, 10.0, and 25 mg/L

EL50: 1.7 mg/L
NOELR 1 mg/L

no vehicle, 48 h, semi-static, closed conditions,
testing of WAFs,
20 test organisms

1

WaBoLu/BGA 1990

nominal: 0.05, 0.066, 0.1, and 0.16 mg/L;
stock solution:
nominal 0.5 mg/L
measured: 0.2 mg/L

EC0: 0.066 mg/L
EC50: > 0.16 mg/L

no vehicle, 48 h, static, 
only 10 test organisms

2

Shigeoka/MITES 1982

Nominal: 0.4, 0.72, 1.3, 2.3, 4.2, 7.6, and 14 mg/L

EC0: 0.4 mg/L
EC50: 2.3 mg/L

vehicle: Tween 80 (10 mg/mL) in DMSO,
24 h, static,
20 test organisms

3