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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
07 October 2017 - 09 November 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
April 13 2004
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 1704HH0700/0
- Expiration date of the lot/batch: 29th April 2018

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All test concentrations and control
- Sampling method: 2.0 mL from the approximate centre of the test vessels was sampled at 0 and 48 hours. At the end of the exposure period, the replicates were pooled at each concentration before sampling.
- Sample storage conditions before analysis: Not applicable, samples were analysed on the day of sampling
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Loading rates were individually prepared. A three-day period of magnetic stirring was applied to ensure maximum dissolution of the test item in test medium. The solutions were allowed to settle for a period of one hour. Thereafter, the aqueous Water Accommodated Fractions (WAFs) were collected by means of siphoning through glass wool and used as test concentrations.
- Controls: Test medium only (adjusted ISO medium)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): No vehicle used
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): In the main test, all test solutions were clear and colorless at the end of the preparation procedure.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna (Straus, 1820)
- Source: In-house laboratory culture with a known history
- Age of parental stock (mean and range, SD): The test used young daphnids <24 hours old from parental daphnids of more than two weeks old.
- Feeding during test: None during test.

ACCLIMATION
- Acclimation period: None

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
180 mg/L as CaCo3
Test temperature:
20°C (measured in a temperature control vessel)
pH:
Test start: 7.9
Test end: 8.0
Dissolved oxygen:
Test start: 8.7 - 9.4 mg/L
Test end: 9.2 - 9.4 mg/L
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal loading rates: 1.0, 1.8, 3.2, 5.6 and 10 mg/L
Measured concentrations at test start: 0.100, 0.115, 0.117, 0.325, 1.05 mg/L
Measured concentrations at test end: 0.0415, 0.0656, 0.0750, 0.214, 0.764 mg/L
Mean measured concentrations: 0.064, 0.087, 0.093, 0.26, 0.89 mg/L
Mean measured concentrations were used for reporting of results.
Details on test conditions:
TEST SYSTEM
- Test vessel: 100 mL glass vessels
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 80 mL
- Aeration: None
- Renewal rate of test solution (frequency/flow rate): No renewal
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 5 Daphnids per 80 mL test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Adjusted ISO medium (Elendt M7)
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH and dissolved oxygen measured at the start and end of the test in all concentrations and the control

OTHER TEST CONDITIONS
- Adjustment of pH: Not reported
- Photoperiod: 16 hour photoperiod
- Light intensity: Not reported

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility assessed at 24 and 48 hours

VEHICLE CONTROL PERFORMED: No

RANGE-FINDING STUDY
- Test concentrations: 1.0, 10 and 100 mg/L loading rates
- Results used to determine the conditions for the definitive study: Yes, loading rates between 1.0 and 10 mg/L used in the main study based on the rangefinder results
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
0.2 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% confidence intervals: 0.16 - 0.24 mg/L
Details on results:
No immobility was observed in the control or the three lowest concentrations. Effects increased with increasing concentration at the two highest test concentrations. After 24 hours of exposure, 30 and 85% of exposed daphnids were immobilised, respectively, while 80 and 100% of exposed daphnids, respectively, were immobilised at the end of the test.
Results with reference substance (positive control):
- Results with reference substance valid? Yes (tested 25 - 27 September 2017)
- Relevant effect levels: 0.10, 0.18, 0.32, 0.56, 1.0, 1.8 mg/L
- Limit test: No
- ECx:
24h-EC50: 0.59 mg/L (95% confidence interval 0.52 - 0.68 mg/L) 48h-EC50: 0.41 mg/L (95% confidence interval 0.34 - 0.46 mg/L)

Reported statistics and error estimates:
The 48h-EC50-value was calculated by applying the Trimmed Spearman-Karber procedure on the percentages of affected daphnids and the logarithms of the corresponding average test item concentrations.

ToxRat Professional v 3.2.1 (ToxRat Solutions® GmbH, Germany) was used to perform the analyses.

Table 1 Average Exposure Concentration Versus Nominal Concentration

Test item
WAF Loading rate (mg/L)

Measured concentrations (mg/L)

Average exposure (mg/L)

t=0h

t=48h

1.0

0.100

0.0415

0.064

1.8

0.115

0.0656

0.087

3.2

0.117

0.0750

0.093

5.6

0.325

0.214

0.26

10

1.05

0.764

0.89

Table 2 Number of Introduced Daphnids and Incidence of Immobility in the Final Test

 

Time (h)

Replicate

Test item; Averagre conc. (mg/L)

Control

0.064

0.087

0.093

0.26

0.89

0

A

5

5

5

5

5

5

B

5

5

5

5

5

5

C

5

5

5

5

5

5

D

5

5

5

5

5

5

Total introduced

20

20

20

20

20

20

24

A

0

0

0

0

1 (1)

5

B

0

0

0

0 (1)

1 (1)

5

C

0

0

0

0

2#(1)

4 (1)

D

0

0

0

0 (1)

2

3

Total immobilised

0

0

0

0

6

17

Effect %

0

0

0

0

30

85

 

 

 

 

 

 

48

A

0

0 (1)

0

0

2

5

B

0

0

0

0

4

5

C

0

0

0

0

5

5

D

0

0

0

0

5

5

Total immobilised

0

0

0

0

16

20

Effect %

0

0

0

0

80

100

( ) between brackets: number of daphnia observed trapped at the surface of the test solutions. These organisms were reimmersed into the respective solutions before recording of mobility.

#Microscopic observation revealed no test item stuck to the daphnids

Table 3 Effect Parameters

Parameter

Test item

Average conc. (mg/L)

95%-confidence interval (mg/L)

24h-EC50

0.42

0.32-0.58

48h-EC50

0.20

0.16-0.24

Validity criteria fulfilled:
yes
Remarks:
In the control, no Daphnids were immobilised or showed other signs of disease, The oxygen concentration in the control and test vessels was ≥3 mg/L at the end of the test.
Conclusions:
The 48-hour EC50 for the test item is 0.20 mg/L (95% confidence interval 0.16 - 0.24 mg/L), based on measured concentrations.
Executive summary:

Augusiak (2018) is a GLP-compliant study following OECD guideline 202. A static test system was employed, with 48-hour exposure. Water Accommodated fractions were prepared at loading rates of 1.0, 1.8, 3.2, 5.6 and 10 mg/L (mean measured concentrations 0.064, 0.087, 0.093, 0.26 and 0.89 mg/L). Loading rates were selected based on a range-finding study and mean measured concentrations were used for reporting of results. 4 replicates of 5 Daphnids were exposed to each test concentration and the control. All validity criteria for the study were met. The 48-hour EC50 for the test item is 0.20 mg/L (95% confidence interval 0.16 - 0.24 mg/L), based on measured concentrations.

Description of key information

The 48-hour EC50 for the test item is 0.20 mg/L (95% confidence interval 0.16 - 0.24 mg/L), based on measured concentrations.

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
0.2 mg/L

Additional information

Augusiak (2018) is a GLP-compliant study following OECD guideline 202. A static test system was employed, with 48-hour exposure. Water Accommodated fractions were prepared at loading rates of 1.0, 1.8, 3.2, 5.6 and 10 mg/L (mean measured concentrations 0.064, 0.087, 0.093, 0.26 and 0.89 mg/L). Loading rates were selected based on a range-finding study and mean measured concentrations were used for reporting of results. 4 replicates of 5 daphnids were exposed to each test concentration and the control. All validity criteria for the study were met. The 48-hour EC50 for the test item is 0.20 mg/L (95% confidence interval 0.16 - 0.24 mg/L), based on measured concentrations.