N-carbamoylmethyliminodi(acetic 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:

2017-01-10 to 2017-12-20

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:

adopted 13th April 2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

May 30, 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1996

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 100 mg/L

- Sampling method: For determination of the test item concentration, four replicate samples were taken from the test item treatment group and one from the control at the start and four replicate samples were taken from the test item treatment group and one from the control at the end of the experiment. The samples were directly injected and analysed by the above detailed HPLC method.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The test solution used in the test was prepared by mechanical dispersion. For preparation of test solutions a stock solution was first prepared by dissolving an amount of 0.03 g test item in 300 mL ISO medium (pH: 7.83) to obtain the 100 mg/L nominal concentration. This solution was placed into an ultrasonic bath for 5 minutes. The pH of the stock solution (100 mg/L) was acidic (pH: 5.02). Therefore, the pH was adjusted to the pH level of the ISO medium with NaOH (according to OECD 23). (The pH level of the stock solution was 7.80 after the pH adjustment). This did not cause any effect (sedimentation, degradation) on the test item and the test solution. The test solution was freshly prepared in the testing laboratory just before introduction of the test animals (start of the experiment).

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus
- Source: Laboratory of Hydrobiology, 2100 Gödöllö, Kotlan S. u. 3, Hungary

ACCLIMATION
Test animals were bred under similar (or the same) conditions as that used during the exposure period (holding water, temperature, background colour etc.), therefore additional acclimatization before the test was not necessary. Brood daphnids were maintained in dilution water at the test temperature for at least 48 hours prior to the start of the test.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Remarks on exposure duration:

none

Post exposure observation period:

no

Hardness:

249 mg/L (as CaCO3)

Test temperature:

19.8 – 20.2 °C

pH:

7.38 – 7.81 (in the treatment group); 7.65 – 8.10 (in the control group)

Dissolved oxygen:

7.24 - 8.12 mg/L

Salinity:

not applicable

Conductivity:

not specified

Nominal and measured concentrations:

Nominal concentration: 100 mg/L;
Measured concentrations reported as nominal as recovery was 106 % (start) and 103 % (end).

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beaker; volume app. 50 mL
- Type: open
- Fill volume: 40 mL
- 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 acc. to OECD TG 202)

OTHER TEST CONDITIONS
- Adjustment of pH: none during test, only for test item solution before application
- Photoperiod: 16 hours light and 8 hours darkness

EFFECT PARAMETERS MEASURED: immobilisation (24 and 48h)

RANGE-FINDING STUDY
- Test concentrations: 0.01, 0.1, 1, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

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:

LOEC

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

Details on results:

There was no immobilisation observed in the control and in the 100 mg/L concentration group. In both groups no abnormal behaviour or appearance of test animals was noted.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: 24h EC50: 1.57 mg/L, (95 % confidence limits: 1.31 – 1.79 mg/L)
- Limit test: no
- Dose-response test: yes

Validity of the Study

Immobilisation was not observed 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 in all cases. All validity criteria were within acceptable limits and therefore the study was considered as valid.

 

Analytical Results

A concentration of 100 mg/L (limit concentration) and a concurrent control was tested in the main test. The concentration of the test item was analytically determined at the start and at the end of the experiment. Four replicate samples were taken from the test item treatment group and one from the control at the start and four replicate samples were taken from the test item treatment group and one from the control at the end of the study.

The test item was not detected in the untreated control group. In the treated group mean measured concentrations of the test item were 106 % of the nominal at the start and 103 % at the end of the test. Since the measured test item concentration remained within ± 20 % of the nominal during the test period of 48 hours, the biological results are given at nominal concentration.

Table 1: Test item concentrations measured in the test solutions

Nominal Concentration (mg/L)	Measured concentrations (mg/L)	In percentage of the nominal	Measured concentrations (mg/L)	In percentage of the nominal
100	106.0	106%	100.7	101%
	105.6	106%	105.0	105%
	105.4	105%	104.2	104%
	105.5	105%	101.4	101%
Mean with 95% confidence interval, (mg/L)	105.6 ± 0.4	106%	102.8 ± 3.4	103%

 

Biological Results

There was no immobilisation observed in the control and in the 100 mg/L concentration group. In both groups no abnormal behaviour or appearance of test animals was noted. The 48-h NOEC was determined to be 100 mg/L. The 48-h LOEC and the EC₅₀ values were determined to be > 100 mg/L. Results are based on the nominal concentration.

Table 2: Summary of the Biological Endpoints

Parameter	Concentration [mg/L] based on nominal concentration
48 h-EC50 95 % conf. limits	> 100 not calculated
48 h-NOEC	100
48 h-LOEC	> 100

 

Table 3: Results of the preliminary range-finding tests

1. Preliminary range-finding test*
Nominal concentrations [mg/L]	Untreated control	0.01	0.1	1	10	100
Number of treated animals	10	10	10	10	10	10
Number of immobilised animals (at 48h)	0	0	0	0	0	0
2. Preliminary range-finding test**
Nominal concentrations [mg/L]	Untreated control			100
Number of treated animals	10			10
Number of immobilised animals (at 48h)	0			10

*: the pH level of the stock solution was adjusted to the pH level of the ISO medium

**: the pH level was not adjusted

 

Table 4: Immobilization of the test animals

Test Group	Replicate	Number of Treated animals	Number of immobilised animals
			24h	48h
Control	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0
100 mg/L	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0

Validity criteria fulfilled:

yes

Conclusions:

In a GLP-study according to OECD test guideline 202 the substance did show no toxicity to daphnia magna at the limit test concentration of 100 mg/L. Therefore, it has a NOEC of 100 mg/L and an EC50 of >100 mg/L.

Executive summary:

The aquatic toxicity of the test item was assessed using Daphnia magna in a 48-hour Acute Immobilisation Test according to OECD TG 202. For this purpose, young Daphnia were exposed to aqueous test media containing the test item for 48 hours. Based on results obtained in the non-GLP preliminary experiments Daphnia were exposed to a single concentration of 100 mg/L (limit concentration). The test included a concurrent negative control. A static test was performed as the test item was previously shown to be stable in the test medium for 48 hours. The purpose of the analytical part of this study was to verify the concentration of the test item in the test medium. The analytically measured test item concentration remained within ± 20 % of the nominal during the test period of 48 hours; therefore, biological results are based on the nominal concentration. Healthy, young female daphnids with known origin, history and acclimatization conditions were used. The test item and the control were tested using 20 Daphnia (divided into concurrent subgroups) each. Each test vessel contained approximately 40 mL test medium. Fully characterized (content, physicochemical characteristics) ISO Medium was used as test medium. The immobilisation of the test animals was recorded 24 and 48 hours after treatment. Environmental conditions were recorded. All measured values remained within acceptable ranges. All validity criteria were met and therefore the study was considered as valid. The 48-h NOEC was determined to be 100 mg/L and the LOEC and the EC₅₀ values were determined to be > 100 mg/L.

Description of key information

In a GLP-study according to OECD test guideline 202 the substance did show no toxicity to daphnia magna at the limit test concentration of 100 mg/L. Therefore, it has a NOEC of 100 mg/L and an EC₅₀ of > 100 mg/L (reference 6.1.3-1).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

The aquatic toxicity of the test item was assessed using Daphnia magna in a 48-hour Acute Immobilisation Test according to OECD TG 202. For this purpose, young Daphnia were exposed to aqueous test media containing the test item for 48 hours. Based on results obtained in the non-GLP preliminary experiments Daphnia were exposed to a single concentration of 100 mg/L (limit concentration). The test included a concurrent negative control. A static test was performed as the test item was previously shown to be stable in the test medium for 48 hours. The purpose of the analytical part of this study was to verify the concentration of the test item in the test medium. The analytically measured test item concentration remained within ± 20 % of the nominal during the test period of 48 hours; therefore, biological results are based on the nominal concentration. Healthy, young female daphnids with known origin, history and acclimatization conditions were used. The test item and the control were tested using 20 Daphnia (divided into concurrent subgroups) each. Each test vessel contained approximately 40 mL test medium. Fully characterized (content, physicochemical characteristics) ISO Medium was used as test medium. The immobilisation of the test animals was recorded 24 and 48 hours after treatment. Environmental conditions were recorded. All measured values remained within acceptable ranges. All validity criteria were met and therefore the study was considered as valid. The 48-h NOEC was determined to be 100 mg/L and the LOEC and the EC₅₀ values were determined to be > 100 mg/L.