N,N-bis(2-hydroxyethyl)glycine

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 March 2017 - 31 March 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

13 April 2004

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N,N-bis(2-hydroxyethyl)glycine

EC Number:

205-755-1

EC Name:

N,N-bis(2-hydroxyethyl)glycine

Cas Number:

150-25-4

Molecular formula:

C6H13NO4

IUPAC Name:

N,N-bis(2-hydroxyethyl)glycine

impurity 1

Reference substance name:

N-(2-hydroxyethyl)ethylenediaminetriacetic acid

EC Number:

205-759-3

EC Name:

N-(2-hydroxyethyl)ethylenediaminetriacetic acid

Cas Number:

150-39-0

Molecular formula:

C10H18N2O7

IUPAC Name:

N-{2-[bis(carboxymethyl)amino]ethyl}-N-(2-hydroxyethyl)glycine

impurity 2

Reference substance name:

Not specified

IUPAC Name:

Not specified

impurity 3

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

Dihydrogen oxide

Test material form:

solid: crystalline

Details on test material:

Appearance: white crystalline powder
Storage conditions: at room temperature protected from light

Specific details on test material used for the study:

Solubility in water: 347 g/L (20°C) (see section 4.8)
Stability in water: not indicated

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Single samples for possible analysis were taken from all test concentrations and the control according to the schedule below.
Frequency at t=0 h and t=48 h
Volume 3.6 mL from the approximate centre of the test vessels
Storage Samples were stored in a freezer (=-15°C) until analysis at the analytical laboratory of the test facility
At the end of the exposure period, the replicates were pooled at each concentration before sampling.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION:
- Correction for purity: no correction for purity was made.
- Method: Preparation of test solutions started with the highest concentration of 100 mg/L. No other treatment than vigorous shaking was needed to completely dissolve the test item in medium. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure.
- Controls: test medium without test item or other additives.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Straus, 1820
- Source: in-house laboratory culture with a known history. At least third generation, obtained by a cyclical parthenogenesis under specified breeding conditions.
- Age of parental stock: Daphnids originated 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 there was no delay in the production of the first brood.
- Age at study initiation: < 24 hours, from parental daphnids of more than two weeks old.
- Feeding during test: no

ACCLIMATION
- Acclimation period: no

BREEDING:
- Method: each batch was started with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium (M7) in an all-glass culture vessel. Cultures were max. 4 weeks old. After 7 days of cultivation half of the medium was renewed twice a week.
- Feeding: daily, a suspension of fresh water algae.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

180 mg/L

Test temperature:

20°C

pH:

7.3-7.9

Dissolved oxygen:

9.0-9.2 mg/L

Nominal and measured concentrations:

Nominal concentrations: 0.10, 1.0, 10 and 100 mg/L
Measured concentrations: 100 mg/L: 140 mg/L at t=0; 109 mg/L at t=48

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml, all-glass
- Fill volume: 80 mL
- Aeration: no
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4, for the highest test concentration, 2 for the other test concentrations
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Standard medium used: yes, adjusted ISO medium
- Source/preparation of dilution water: tap water purified by reverse osmosis
- Culture medium different from test medium: yes, M7 was used as culture medium

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: the test was performed in the dark

EFFECT PARAMETERS MEASURED: mobility at 24 and 48 hours.
- Additional measurements: pH and dissolved oxygen at the beginning and at the end of the test, in the highest test concentration and the control. Temperature of the medium: continuously in a temperature control vessel.

COMBINED LIMIT/RANGE-FINDING STUDY: yes, reported as the final test

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (Feb 2017)

Results and discussion

Effect concentrations

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 124 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

No immobility was observed at any of the test concentrations after 24 hours. In the nominal concentration of 100 mg/L, 5% of the daphnids were immobilized after 48 hours of exposure.
- No biological, behavioural or other abnormalities were observed in the control group or any of the test concentrations.
- Analyses of the samples taken from the nominal concentration of 100 mg/L showed a concentration of 140 mg/L at the start of the test. The measured concentration decreased to 78% of initial at the end of the test (109 mg/L). Based on this result, a geometric average exposure concentration of 124 mg/L was used to determine effect parameters.
- Individual pH, temperature and dissolved oxygen values remained within acceptable limits throughout the duration of the study.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- 48h-EC50: 0.48 mg/L, 95%-confidence interval: 0.43-0.53 mg/L
- Other: results fell within the historical range.

Any other information on results incl. tables

Table 1 Measured concentrations in test samples

Time of sampling [hours]	Concentration [mg/L]		Relative to nominal [%]	Relative to initial [%]
	Nominal	Analyzed
0	0	0.0572^2	n.a.
	100	140^3	140
48	0	n.d.	n.a.	n.a.
	100	109	109	78

1 Samples were stored in the freezer (= -15°C) until the day of analysis.

2 The maximum contribution to the sample at the lowest concentration level was 0.043%.

3 Estimated value, calculated by extrapolation of the calibration curve.

n.d. Not detected.

n.a. Not applicable.

Table 2 Number of introduced Daphnids and incidence of immobility

Time (h)	Replicate	Nominal concentration Bicine (mg/L)
		Control	0.1	1.0	10	100
0	A	5	5	5	5	5
	B	5	5	5	5	5
	C	5				5
	D	5				5
	Total introduced	20	10	10	10	20
24	A	0	0	0	0	0
	B	0	0	0	0	0
	C	0				0
	D	0				0
	Total immobilised	0	0	0	0	0
	Effect %
48	A	0	0	0	0	0
	B	0	0	0	0	0
	C	0	0			0
	D	0	0			1
	Total immobilised	0	0	0	0	1
	Effect %	0	0	0	0	5

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

1). In the control, no daphnids became immobilised or showed other signs of disease or stress. 2). The oxygen concentration at the end of the test was = 3 mg/L in control and test vessels (e.g., =9.1 mg/L).

Conclusions:

The 48h-EC50 of the substance towards Daphnia magna, exceeded a mean measured exposure concentration of 124 mg/L.