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

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

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: Product no. B4429, Lot no. SLBK5235V
- Expiration date of the lot/batch: 30 April 2018
- Purity test date: na

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature

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

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 100 mg/L
- Sampling method: - no data
- Sample storage conditions before analysis: no information

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: mechanical dispersion
- Eluate: not applicable
- Differential loading: not applicable
- Controls: untreated and reference (tested in September 2016)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): not applicable
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): not applicable
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): none

PRELIMINARY TESTS: none

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: 61.81 SAG
- Source (laboratory, culture collection): SAG: Collection of Algal Cultures, 37073 Göttingen, Germany
- Age of inoculum (at test initiation): -
- Method of cultivation: algae were kept as stock culture on solid agar

ACCLIMATION
- Acclimation period: -
- Culturing media and conditions (same as test or not): -
- Any deformed or abnormal cells observed: - none

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Post exposure observation period:

none

Hardness:

no data

Test temperature:

22.2 - 22.4°C (measured in the flasks)
22.1 - 23.8°C (in the climate chamber)

pH:

Start: 7.85 - 7.76 in the five test concentrations(adjusted); 8.18 in the control
End: 8.38 - 8.64 (test concentrations and control)

Dissolved oxygen:

no data

Salinity:

no data

Nominal and measured concentrations:

The measured concentrations showed very good correlation with the nominal concentrations. The recovery rate after 72 hours was 102%.
Therefore the nominal concentration was used for the determination of the results.

Details on test conditions:

TEST SYSTEM
- Test vessel: glass flasks 250 ml
- Type (delete if not applicable): air-permeable stoppers
- Material, size, headspace, fill volume: -
- Aeration:-
- Type of flow-through (e.g. peristaltic or proportional diluter): none
- Renewal rate of test solution (frequency/flow rate): none
- Initial cells density: 10000 cells/ml
- Control end cells density: approx. 10000000 cells/ml
- No. of organisms per vessel:-
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): -

GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used: -

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted algal growth medium (acc. to OECD TG 201)

OTHER TEST CONDITIONS
- Sterile test conditions: no data
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: 7649 Lux
- Salinity (for marine algae): -

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: manual counting at 24, 48 and 72 h
- Chlorophyll measurement: -
- Other: -

TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable
- Justification for using less concentrations than requested by guideline: limit test based on concentration-range finding test
- Range finding study: yes
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: no effects up to 100 mg/L in preliminary tests

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): none reported
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

- Results with reference substance valid? YES
- EC50: 0.71 mg/L (ErC50) (tested from September 20 -23, 2016)
- Other: -

Reported statistics and error estimates:

not applicable as no effects were observed

The acute toxicity of the substance was investigated in a GLP-study according to OECD test guideline 201 using unicellular freshwater green algae Raphidocelis subcapitata under static conditions.

The study was performed using one concentration containing 100 mg/L with a treatment period of 72 hours. The cell number of each replicate was determined by manual counting. Then the growth rate and the yield were determined.

At the start and at the end of the test, the content of the test item in the test solutions was determined using LC/MS. The recovery after 72 hours was 102 % of the start concentration. Therefore, the determination of the results was based on the nominal concentrations.

The EC50s of the positive reference control potassium dichromate were tested in a non-current reference test. The values lay within the normal range of the laboratory.

As no toxicity of the substancewas observed, a NOEC (growth rate and biomass) of 100 mg/L and an EC50 (growth rate and biomass) of > 100 mg/L were determined.

Detailed results:

Growth rates (µ) and %-inhibition of µ:

	growth rates (µ) and %-inhibition of µ
Test Group	0 -24h		0 -48 h		0 -72h
	µ	%	µ	%	µ	%
Control	0.0345	0.0	0.0471	0.0	0.0532	0.0
100 mg/L	0.0373	-8.2	0.0465	1.3	0.0524	1.4

Yield (y) and %-inhibition of y:

	yield (y) and %-inhibition of y
Test Group	0 -72h
	y			%
Control	45.3			0.0
100 mg/L	42.8			5.5

Validity criteria fulfilled:

yes

Executive summary:

The acute toxicity of the substance was investigated in a GLP-study according to OECD test guideline 201 using

unicellular freshwater green alga Raphidocelis subcapitata.

The study was performed as a static limit test with 100 mg/L with a treatment period of 72 hours. The cell number of each replicate was determined by manual counting. Then the growth rate and the yield were determined.

At the start and at the end of the test, the content of the test item in the test solutions was determined by HPLC. The recovery after 72 hours was 102% of the start concentration. Therefore, the determination of the results was based on the nominal concentrations.

The EC50s of the positive reference control potassium dichromate were tested in a non-current reference test. The values lay within the normal range of the laboratory.

As no toxicity of the substance was observed, the NOEC (growth rate and biomass) of 100 mg/L and the EC50 (growth rate and biomass) of > 100 mg/L were determined.

Description of key information

In a GLP-study according to OECD test guideline 201 (toxicity to algae, static conditions), the substance did show no toxicity at the test concentration of 100 mg/L. Therefore, it has a NOEC (growth rate and biomass) of 100 mg/L and an EC50 (growth rate and biomass) of >100 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:

100 mg/L

EC10 or NOEC for freshwater algae:

100 mg/L

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