2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol

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:

2016-10-14 to 2016-10-27

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:

April 13, 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:

April 1996

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

For determination of the test item concentration, five replicate samples were taken from the test item treatment group and two from the control at the start and four replicate samples were taken from the test item treatment group and two from the control at the end of the experiment. The analysis was performed using HPLC method with UV detection.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The test solution used in the test was prepared by mechanical dispersion. An amount of 0.03 g test item was dissolved in 300 mL dilution water (ISO medium) in order to obtain the concentration of 100 mg/L. The test solution was freshly prepared in the testing laboratory just before introduction of test animals (start of the experiment).

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna (Straus)
- Source: Laboratory of Hydrobiology (Central Agricultural Office, Directorate of Plant-, and Soil Protection), 2100 Gödöllő, Kotlán S. u. 3. Hungary
- Breeding: The Daphnia are bred in the Laboratory of TOXI-COOP ZRT.
- Age of animals: Less than 24 h old at the beginning of the test.
- Sex: Female
- Feeding during test: no

ACCLIMATION
- Acclimation Period: 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

Hardness:

249 mg/L (as CaCO3)

Test temperature:

20.1 - 20.2 °C (Test vessel)
19.7 - 20.6 °C (Climate chamber)

pH:

7.86 - 7.98

Dissolved oxygen:

7.95 - 8.07 mg/L

Nominal and measured concentrations:

Nominal: 100 mg/L
Measured: within ± 20 % of nominal concentration

Details on test conditions:

TEST SYSTEM
- Test vessel: beaker
- Material, size, fill volume: glass, 50 mL, 40 mL.
- Aeration: None
- Type: open
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 5

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
- Photoperiod: 16 hours light (artificial illumination) and 8 hours darkness.

EFFECT PARAMETERS MEASURED: Daphnia were observed visually for immobility or mortality after 24 and 48 hours of exposure. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile. In addition to immobility any abnormal behaviour or appearance is reported.

RANGE-FINDING STUDY
In order to select appropriate test concentrations for use in the definitive test, a non-GLP preliminary range-finding test was conducted to determine the approximate toxicity of the test item. For preparation of the test solutions a stock solution (100 mg/L nominal concentration) was first prepared by dispersing/dissolving an appropriate amount of test item into the test medium (ISO medium). Subsequent lower concentrations were prepared by appropriate dilution of this stock solution. Per test concentration and control ten daphnids (divided into 2 replicates) were exposed for 48 hours.
The concentration levels used and results (48-h) of the preliminary range-finding test are summarised in table 1, in the section “Any other information on materials and methods incl. tables”.
As no toxic effects were observed during the preliminary test, a single concentration of 100 mg/L (limit concentration) and a concurrent control were tested in the main test.
The analytically measured test item concentration remained within ± 20 % of the nominal concentration during the test period of 48 hours. Therefore, biological results were based on the nominal concentration.

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

Details on results:

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:
The quantification of the test item in the test solutions was performed by a previously validated method. 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.
The test item was not detected in the untreated control group. In the treated group mean measured concentrations of the test item were 99 % of the nominal at the start and 102 % 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.

Biological Results:
No immobilization or any abnormal behaviour and appearances were observed in the control group and in the 100 mg/L concentration group after 48 hours of exposure.

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)

All validity criteria were met. Therefore, the study was considered as valid.

Results are based on the nominal concentration.

Biological endpoints of the study are summarised below:

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

Validity criteria fulfilled:

yes

Conclusions:

In the 48-h acute toxicity of the test item on Daphnia magna according to OECD T 202, 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. All validity criteria were met. Therefore, the study was considered as valid. Results are based on the nominal concentration.

Executive summary:

The influence of the test item on Daphnia magna was assessed in a 48-h Acute Immobilisation Test according to OECD TG 202. Young Daphnia were exposed to aqueous test media containing the test item for 48 hours. Based on results obtained from a non-GLP preliminary experiment 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 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 concentration of 100 mg/L and the control (ISO Medium) were tested using 20 Daphnia (divided into 4 concurrent subgroups) each. 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.

No immobilisation or any abnormal behaviour was observed in the control and the 100 mg/L concentration group after 48 hours of exposure. All validity criteria were within acceptable limits and therefore the study was considered valid.

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.

Description of key information

In the 48-h acute toxicity study of the test item on Daphnia magna according to OECD T 202, 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 (reference 6.1.3-1).

Additional information

The influence of the test item on Daphnia magna was assessed in a 48-h Acute Immobilisation Test according to OECD TG 202. Young Daphnia were exposed to aqueous test media containing the test item for 48 hours. Based on results obtained from a non-GLP preliminary experiment 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 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 concentration of 100 mg/L and the control (ISO Medium) were tested using 20 Daphnia (divided into 4 concurrent subgroups) each. 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.

No immobilisation or any abnormal behaviour was observed in the control and the 100 mg/L concentration group after 48 hours of exposure. All validity criteria were within acceptable limits and therefore the study was considered valid.

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.