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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 Jan 2018 - 17 Jan 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
2004
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23
Version / remarks:
2000
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Solubility in water: 43593 mg/L at 24°C for the main constituents (benzoic acid)
Stability in water: somewhat stable
Vapour pressure: 1.4 Pa
Analytical monitoring:
yes
Remarks:
TOC analysis
Details on sampling:
Test concentrations were verified by chemical analysis. Water samples (40 mL, in duplicate) were taken from the test vessels of the control and each exposure level at the start and end of both 24-h (renewal) periods. At the end of each renewal period, replicates were not pooled at each concentration before sampling. Instead, samples were taken from replicates A and B of each test group. Samples were stored in a refrigerator (2 - 8°C) until analysis.


Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Water Acc ommodated Fractions individuallly prepared at 1.0, 10, and 100 mg/L. Before weighing, the test item was heated in the oven at 75ºC; 24 hours for test item used to prepare the test solutions for the first renewal and 48 hours for test solutions made for the second renewal. The container remained closed during heating. Approximately one hour of ultrasonic waves followed by two days of magnetic stirring was applied to ensure maximum dissolution of the test item in medium. After stirring, the aqueous WAFs were collected by filtration through a 0.45 µm membrane filter (RC55, Whatman) and used as test concentrations. All test solutions were clear and colorless at the end of the preparation procedure.
- Controls: Test medium without test item or other additives.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna (Straus 1820)
- Strain: not stated
- Source:In-house laboratory culture with a known history
- Age of parental stock: : >2 weeks (originating from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and without delay in the production of the first brood)
- Age of test organisms at test initiation: young daphnids <24 h
- Method of breeding: start of each batch with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium in an all-glass culture vessel.
- Renewal of the cultures: after 7 days of cultivation half of the medium twice a week.
- Feeding during test: no

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
180 mg/L expressed as CaCO3
Test temperature:
20-21°C
pH:
Fresh solutions: 7.6-7.9
Old solutions: 7.6-8.0
Dissolved oxygen:
Fresh solutions: 8.5-9.7 mg/L
Old solutions: 8.0-9.2 mg/L
Nominal and measured concentrations:
Nominal loading rates: 1.0, 10, 100 mg/L

Bezoin resoid loading rate ( mg/L): control and 100
Measured concentration (t=0): n.q, 4.1 mg TOC/L
Measured concentration (t=24, old): n.q, 1.2 (30) mg TOC/L
Measured concentration (t=24, fresh): n.q, 0.94 mg TOC/L
Measured concentration (t=48): n.q, 0.27(29) mg TOC/L

( ) between brackets, percentage of initially measured concentration are given. n.q. not quantifiable.
Details on test conditions:
TEST SYSTEM
- Test vessel: 60 mL, all-glass, air-tight closed without headspace
- Aeration: no
- Renewal rate of test solution: yes, after 24 h
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4 for 100 mg/L, 2 for 1.0 and 10 mg/L
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Test medium: Adjusted ISO medium (tap water purified by Reverse Osmosis and macro salts, according to OECD202).
- Total organic carbon in Control: not quantifiable
- Culture medium different from test medium: yes, culture medium: Elendt M7 medium.
- Intervals of water quality measurement: pH and dissolved oxygen: at the beginning and at the end of both renewal periods, in the old and freshly prepared test solutions for the control and WAF prepared at 100 mg/L. In addition, after 24 hours in fresh and old solutions of all groups.
Temperature of medium: continuously in a temperature control vessel, beginning at the start of the test. In addition, temperature was measured in old and freshly prepared test solutions of all groups after 24 hours.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h photoperiod daily

EFFECT PARAMETERS MEASURED:
- Immobility (including mortality): at 24 h and at 48 h. Daphnia were considered to be immobilised if they were unable to swim for approximately 15 seconds after gentle agitation.


RANGE-FINDING STUDY:
- A combined limit/range-finding test was performed.


Reference substance (positive control):
yes
Remarks:
Potassium dichromate (test performed Jan 2018)
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Behavioural abnormalities: none observed in test concentrations. One daphnid in Control was trapped at the surface at 48h.
- Mortality of control: no
- Effect concentrations exceeding solubility of substance in test medium: yes
- Samples taken of the control and WAF prepared at a loading rate of 100 mg/L were analysed for TOC concentration. The maximum soluble concentration in the present study was measured to be 4.1 mg TOC/L and was measured in the freshly prepared WAF of 100 mg/L at the start of the test. The concentration measured in the freshly prepared test solution for the second renewal was lower, i.e. was 0.94 mg/L. In both renewals, the measured concentrations were not stable, i.e. decreased to ~30% of initially measured, despite the closed vessels. It should be noted that values below 1.0 mg/L are considered as indicative.

- The reason for the difference between the initially measured concentrations is unknown. The difference in the applied heating period could play a role. However possibility that the measured concentrations were oversaturated cannot be excluded as a possible explanation. Although solubility of one of the components was indicated, the solubility in water of the substance was unknown.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Dose-response test, concentrations: 0.10, 0.18, 0.32, 0.56, 1.0 and 1.8 mg/L
- 48h-EC50: 0.54 mg/L(95% confidence interval 0.49 - 0.61 mg/L).

        
Number of Introduced Daphnids and Incidence of Immobility

Time (h)

Replicate

Benzoin Resoid; Loading rate (mg/L)

Control

1.0

10

100

0

A

5

5

5

5

B

5

5

5

5

C

5

 

 

5

D

5

 

 

5

Total introduced

20

10

10

20

24

A

0

0

0

0

B

0

0

0

0

C

0

 

 

0

D

0

 

 

0

Total immobilised

0

0

0

0

Effect %

0

0

0

0

 

 

 

 

48

A

0(1)

0

0

0

B

0

0

0

0

C

0

 

 

0

D

0

 

 

0

Total immobilised

0

0

0

0

Effect %

0

0

0

0

( ) 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.

Validity criteria fulfilled:
yes
Remarks:
<10% immobilization in control group, O2 conc. at end is >=3 mg/L in control and test vessels
Conclusions:
The 48h-EL50 for Daphnia magna exposed to Benzoin Resoid exceeded the nominal loading rate of 100 mg/L, the maximum loading tested.
Executive summary:

A study was performed to assess the acute toxicity of Benzoin Resoid toDaphnia magnaunder semi-static conditions (renewal period: 24 hours). The study was conducted in accordance with OECD Guideline for Testing of Chemicals No. 202 and GLP. A combined limit/range-finding test was performed with untreated control, and Water Accommodated Fractions prepared at loading rates of 1.0, 10, and 100 mg/L. The incidence of immobilisation was recorded for each treatment group and controls at 24 hours and at 48 hours and TOC analysis was performed. Effect parameters were reported in terms of nominal loading rates initially prepared.

Samples taken from the control and WAF at 100 mg/L at the start and the end of both 24-hour renewal periods were analysed. The maximum soluble concentration was measured to be 4.1 mg TOC/L. In both renewals, the measured concentrations were not stable.

The 48h-EL50 for Daphnia magna exposed to Benzoin resoid exceeded the nominal loading rate of 100 mg/L, the maximum loading tested.

Description of key information

A study was performed to assess the acute toxicity of Benzoin Resoid toDaphnia magnaunder semi-static conditions (renewal period: 24 hours). The study was conducted in accordance with OECD Guideline for Testing of Chemicals No. 202 and GLP. A combined limit/range-finding test was performed with untreated control, and Water Accommodated Fractions prepared at loading rates of 1.0, 10, and 100 mg/L. The incidence of immobilisation was recorded for each treatment group and controls at 24 hours and at 48 hours and TOC analysis was performed. Effect parameters were reported in terms of nominal loading rates initially prepared.

Samples taken from the control and WAF at 100 mg/L at the start and the end of both 24-hour renewal periods were analysed. The maximum soluble concentration was measured to be 4.1 mg TOC/L. In both renewals, the measured concentrations were not stable.

The 48h-EL50 for Daphnia magna exposed to Benzoin resoid exceeded the nominal loading rate of 100 mg/L, the maximum loading tested.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
100 mg/L

Additional information

48h-EL50: >100 mg/L.