Amidinourea phosphate

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Remarks:

Daphnia magna

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 August 2017 to 19 Sept 2017

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

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 16VL8189
- Expiration date of the lot/batch: 04 Feb 2018
- Purity test date: 05 Aug 2016

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: ambient (=< 30 deg C), dark, dry

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: No

Analytical monitoring:

yes

Details on sampling:

Analytical data are required by the guidelines for verification of test item concentrations as well as the stability of the test item over the entire test period. Analytical samples were taken from all test item concentrations and control at test start and after 24 hours from fresh and aged solutions and after 48 hours from aged solutions. For each sampling also a retain sample was taken.
All samples were stored deep frozen until they were transferred to the analytical laboratory.

Vehicle:

no

Details on test solutions:

Based on the results of a non GLP range-finding test, the following nominal concentrations were tested in the main test: 100, 50.0, 25.0, 12.5, 6.25 mg/L and control. The necessary amount of test item for preparing the stock solution S1 was weighed on a weighing scoop and transferred to a volumetric flask. Test medium (see Appendix B) was added up to the bench mark and the solution was treated with 10 minutes of ultrasonication. Afterwards the solution was clear and transparent. Lower test solutions were prepared by dilution of the appropriate solutions with test medium. The preparation procedure was repeated after 24 hours. ~ 50 mL of the prepared solutions were transferred to each test vessel

Test organisms (species):

Daphnia magna

Details on test organisms:

Daphnia magna Straus, Clone V, was used as the test organism. The animals are continuously bred in the laboratory and were originally purchased in a healthy condition from the Federal Environment Agency in Berlin/Germany. Daphnia magna was bred as single culture (1 daphnid per 100 mL) in Elendt M4 medium. The pHvalue of the aerated water was within a range of 6.0 – 9.0. The dissolved oxygen was above 60 % saturation and the total hardness 140 - 250 mg/L (as CaCO3), corresponding to 7.8 - 14°dH. The animals were fed with single cell green algae (Desmodesmus subspicatus, formerly Scenedesmus subspicatus) at least three times a week.
The daphnids were reared at a temperature of 20 ± 2 °C in a climatic chamber with 16 hours of illumination and 8 hours of darkness. The medium was changed three times per week. A pipette was used to separate the young daphnids from the adults.
Freshly hatched daphnids less than 24 hours old were used for the test. The first brood progeny was not used

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Remarks on exposure duration:

None

Post exposure observation period:

None

Hardness:

13°dH (232 mg/L as CaCO3)

Test temperature:

19.0 – 19.8 °C

pH:

6.85 – 7.97

Dissolved oxygen:

>= 8.9 mg/L

Salinity:

Not applicable

Conductivity:

Not measured

Nominal and measured concentrations:

100, 50.0, 25.0, 12.5, 6.25 mg/L and control.

Details on test conditions:

Test procedure: semi-static
Duration: 48 hours
Temperature: 19.0 – 19.8 °C
Oxygen concentration: >= 8.9 mg/L
pH-value: 6.85 – 7.97
Exposure to light: 16 hours photoperiod /8 hours darkness daily
Feeding: none
Test vessels: four 100 mL glass beakers per concentration, each filled with ~ 50 mL, one additional replicate for pysico - chemical measurements without test organisms
Loading: ~ 10 mL of test solution for each animal
Aeration: none
Number of animals: 20 per concentration in 4 replicates of 5

Biological Assessment
After 24 h and 48 h the immobilised daphnids were counted. All daphnids not able to swim within 15 seconds after gentle agitation of the test vessel were considered to be immobilised. If present, behavioural changes of daphnids were recorded at 24 and 48 hours after starting the test.
Physico- Chemical Assessment
The test temperature and the pH-value as well as the oxygen concentration of the test solutions were measured at all concentrations at t = 0 h fresh and t = 24 h from fresh test solutions and after t = 24 h and t = 48 h from aged test solutions in one separate replicate per test item concentration without test organisms

Reference substance (positive control):

yes

Key result

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

>= 50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Validity Criteria of the Study
Control immobilisation
The percentage of immobilisation should be 10 %. In this study the control immobilisation was 0 %.
Oxygen concentration
The dissolved oxygen concentration at the end of the test should be 3 mg/L in all test units. In this test, the dissolved oxygen concentration at the end of the test was 8.9 mg/L.
Biological Results
After 24 hours of exposure no immobilisation was observed in the control and up to and including 100 mg/L. After 48 hours of exposure no immobilisation was observed in the control and all test item concentrations up to and including 50.0 mg/L. 30 % immobilisation was observed at 100 mg/L.
No behavioural changes could be observed after 24 hours. After 48 hours the daphnids were sluggish in the highest test item concentration of 100 mg/L.
Analytical Results
The measured content of A.I./test item was between 94 % and 98 % of nominal in the fresh samples with a mean initial concentration of 96 % of nominal. In the aged samples the measured content was between 92 % and 97 % of nominal with a mean measured concentration of 95 % of nominal. Therefore toxicological endpoints were evaluated using the nominal of the test item.

Results with reference substance (positive control):

In order to check the validity of the results, the toxicity of the reference item potassium dichromate was tested at 1.00 and 2.00 mg/L with 20 test organisms per test concentration.
The results indicate an EC50 (24 h) of the reference item potassium dichromate around 1.00 mg/L. Since the results are in accordance with the requirements of the OECD guideline 202 and fall within the historical data generated with the reference item at the testing facility, the daphnids were suitable for the determination of the toxicological effects of the test item.

Reported statistics and error estimates:

All toxicological endpoints were evaluated using nominal concentrations.

Results of test: 48 hours – immobilised Daphnia

mg/L	Control	6.25	12.5	25.0	50.0	100
Group 1	0	0	0	0	0	0
Group 2	0	0	0	0	0	2
Group 3	0	0	0	0	0	3
Group 4	0	0	0	0	0	1
Ʃ	0	0	0	0	0	6
%	0	0	0	0	0	30

Validity criteria fulfilled:

yes

Conclusions:

According to the results of the test, the EC50 (48 h) for immobilisation was supposed to be > 100 mg/L (nominal). The corresponding NOEC (48 h) was 50.0 mg/L (nominal).

Executive summary:

The acute toxicity of the test substance to aquatic invertebrates (Daphnia magna) was performed in accordance with OECD Guideline 202.

A semi-static main test with nominal concentrations of 100, 50.0, 25.0, 12.5 6.25 mg/L and control was performed using twenty organisms per test concentration (4 replicates of 5 animals each). The duration of the test was 48 hours.

Temperature, pH-value and oxygen concentration of the test solutions measured after 0, 24 hours fresh and 48 hours aged are reported. Hardness of the test water was measured on the day of application. Analytical samples taken at 0 hours (initial value) and 24 hours from fresh and 48 hours aged test solutions were analysed from control and 50.0 and 100 mg/L.

According to the results of the test, the EC50 (48 h) for immobilisation was supposed to be > 100 mg/L (nominal). The corresponding NOEC (48 h) was 50.0 mg/L (nominal).

Description of key information

The acute toxicity of the test substance to aquatic invertebrates (Daphnia magna) was performed in accordance with OECD Guideline 202.

A semi-static main test with nominal concentrations of 100, 50.0, 25.0, 12.5 6.25 mg/L and control was performed using twenty organisms per test concentration (4 replicates of 5 animals each). The duration of the test was 48 hours.

Temperature, pH-value and oxygen concentration of the test solutions measured after 0, 24 hours fresh and 48 hours aged are reported. Hardness of the test water was measured on the day of application. Analytical samples taken at 0 hours (initial value) and 24 hours from fresh and 48 hours aged test solutions were analysed from control and 50.0 and 100 mg/L.

According to the results of the test, the EC50 (48 h) for immobilisation was supposed to be > 100 mg/L (nominal). The corresponding NOEC (48 h) was 50.0 mg/L (nominal).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L