[2S-[2α,5α,6β(S*)]]-6-[[[[(4-ethyl-2,3-dioxopiperazin-1-yl)carbonyl]amino]phenylacetyl]amino]-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17/07/2018 - 19/07/2018

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0(control) and 100 mg/L
- Samples were taken from the test solution and from the control solution at the beginning and at the end of the renewal periods (24h). The samples were analyzed by a HPLC - UV method.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
A stock solution with a nominal concentration of 100 mg/L was prepared with direct addition of the test item, mixed into the test medium (ISO Medium) using ultrasonic bath (5 minutes). As a Limit test was carried out, further dilution of stock solution was not performed.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Strain: Daphnia magna
- Source: István Szent University, 2100 Gödöllő, Páter Károly u. 1, Hungary
- Age of the animals: less than 24 h old at the beginning of the test.
- Feeding during test: No

ACCLIMATION: There was no acclimatization because the water used was similar to the culture water.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

245 mg/L (as CaCO3)

Test temperature:

19.7 – 21.0ºC

pH:

7.14 – 7.75

Dissolved oxygen:

8.1 – 8.6 mg/L.

Nominal and measured concentrations:

Nominal concentrations: 0 (control), 100 mg/L.
Measured concentrations: 0 (control), 98.7 mg/L (geometric mean, measured)

Details on test conditions:

TEST SYSTEM
- Test vessel: Beaker
- Material, size, headspace, fill volume: glass beaker with a volume of 50 mL
- Aeration: No
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water (ISO medium, according to OECD 202)

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16-hour light and 8-hour dark cycle

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
The immobility or mortality of the Daphnia was determined by visual observation 24 and 48 hours after the start of the test. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile.

RANGE-FINDING STUDY
A concentration range-finding test was conducted to determine the approximate toxicity of the test item so that appropriate test concentrations could be selected for use in the definitive test.
- Test concentrations: 0, 0.1, 1, 10 and 100 mg/L (10 daphnids per dose, semi-static, 48 hours)
- Results used to determine the conditions for the definitive study: Because toxic response was not observed during the preliminary concentration range-finding test, a limit test was carried out using only one test concentration (100 mg/L) and a control group in a semi-static system.

Reference substance (positive control):

yes

Remarks:

(Potassium dichromate is tested at least twice a year, last 20/06/2018)

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: No abnormal behaviour, appearance or any other effect on test animals were detected.
- Mortality of control: 0/20

Results with reference substance (positive control):

- Results with reference substance valid? Yes
Potassium dichromate is tested at least twice a year to demonstrate satisfactory test conditions.
The date of the last study (Study Code: 18/178-023DA) with reference item Potassium dichromate (batch no.: A0345704) is: 19- 20 June 2018.
The 24h EC50: 0.71 mg/L, (95 % confidence limits: 0.66 – 0.76 mg/L)

Reported statistics and error estimates:

No statistical analysis was performed because of the lack of toxic effects. The EC50, NOEC, LOEC and EC100 values were determined directly from the raw data.

Table 2. Number and percentage of immobilised animals (definitive test).

Test Group	Number of treated animals	Number of immobilised animals
		24 hours		48 hours
		number	percent	number	percent
Control	20	0	0	0	0
100 mg/L	20	0	0	0	0

Validity criteria fulfilled:

yes

Remarks:

There was no immobilization in the control group and the dissolved oxygen concentration at the end of the test in control and test vessels was more than 3 mg/L.

Conclusions:

The 48h-EC50 in Daphnia magna was found to be higher tan 100 mg/L and the NOEC was equal to 100 mg/L.

Executive summary:

An acute immobilisation test on Daphnia magna was performed in accordance with OECD Guideline 202 and EU method C.2. Based on the results obtained in the preliminary range-finding test where no toxic response was observed, a Limit Test was carried out using only one test concentration (100 mg/L) and one control group in a semi-static system (24h renewal periods). Twenty animals, divided into four groups (glass beaker) of five animals each were exposed to the test item or the control for 48 h. Test concentrations were analytically determined at the beginning and at the end of the test and did not deviate more than 20% from the nominal. All validity criteria were met. The 48h-EC50 was determined to be higher than 100 mg/L and the 48h-NOEC was determined to be 100 mg/L, both based on mobility.

Description of key information

Key study. Method according to OECD 202 (limit test), GLP study. The 48h-EC50 was determined to be higher than 100 mg/L and the 48h-NOEC was determined to be 100 mg/L, both based on mobility.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

Key study. An acute immobilisation test on Daphnia magna was performed in accordance with OECD Guideline 202 / EU method C.2. Based on the results obtained in the preliminary range-finding test, a Limit Test was carried out using only one test concentration (100 mg/L) and one control group in a semi-static system (24h renewal periods). Twenty animals, divided into four groups of five animals each, were exposed to the test item or the control for 48 h. Test concentrations were analytically determined at the beginning and at the end of the test and did not deviate more than 20% from the nominal. All validity criteria were met. The 48h-EC50 was determined to be higher than 100 mg/L and the 48h-NOEC was determined to be 100 mg/L, both based on mobility.