2-[(3-aminopropyl)methylamino]ethanol

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-08-16 to 2017-08-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test (Carbon and Ammonium Oxidation))

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: PFW110370
- Expiration date of the lot/batch: 2018-12-31
- Purity: 99.10%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature in the dark
- Stability under test conditions: not specified

OTHER SPECIFICS:
Physical state/Appearance: clear colorless liquid

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 2500 mg (nominal) of test item was dissolved in water and the volume adjusted to 1L to obtain a stock solution from which dilutions of 250 and 25 mg/L were made. Subsequently, a solution of 10 mg/L was made with an 200 mL aliquot from the 25 mg/L stock solution which was dispersed with 16mL of synthetic sewage, 250 mL of activated sewage sludge and water, to a final volume of 500 mL. Test concentrations of 100 and 1000 mg/L (in triplicates) were also prepared similarly. The volumetric flasks containing the stock solutions were inverted several times to ensure homogeneity.
- Eluate: not specified
- Differential loading: 10, 100 and 1000 mg/L (nominal concentrations)
- Controls: 4 vessels of negative control (not exposed to test material) and 3 vessels of positive control (with reference item) in concentrations of 3.2, 10 and 32 mg/L
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): not relevant
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): not relevant
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): not specified

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: A mixed population of activated sewage sludge micro-organisms from the aeration stage
- Name and location of sewage treatment plant where inoculum was collected: Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK
- Preparation of inoculum for exposure: The activated sewage sludge sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21 ºC overnight prior to use in the test. On the day of collection the activated sewage sludge (10 liters) was fed synthetic sewage (500 mL). The pH of the sample on the day of the test was 7.4 measured using a Hach HQ40d Flexi handheld meter. Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 mL) of the activated sewage sludge by suction through a pre-weighed GF/A filter paper* using a Buchner funnel which was then rinsed 3 times with 10 mL of deionized reverse osmosis water and filtration continued for 3 minutes. The filter paper was then dried in an oven at approximately 105 ºC for at least one hour and allowed to cool before weighing. This process was repeated until a constant weight was attained.
- Method of cultivation: activated sewage sludge (10 liters) was fed synthetic sewage (500 mL)
- Initial biomass concentration: 3 g SS/L

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Test temperature:

Controlled room temperature of approximately 21°C

pH:

- Negative control (not exposed to the test item): 0 hr: 7.2-7.6; 3 hrs: 7.4 - 7.6
- Test item: 0 hr: 7.3 - 7.8; 3 hrs: 7.5 - 7.8
- Positive control (3,5- dichlorophenol): 0 hr: 7.3 - 7.5; 3 hrs: 7.6 - 8.0

Dissolved oxygen:

- Negative control (not exposed to the test item): 5.41 - 6.85 mg O2/L
- Test item: 5.45 - 6.30 mg O2/L
- Positive control (3,5- dichlorophenol): 7.17 - 8.24 mg O2/L

In some instances, the initial and final DO concentrations were outside those recommended in the test guidelines (7 mg O2/L and 2 mg O2/L respectively). This was considered to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was determined over the linear portion of the oxygen consumption trace.

Salinity:

not applicable

Nominal and measured concentrations:

loading rates for the test substance (mg/L): 10, 100 and 1000

Details on test conditions:

TEST SYSTEM
- Test vessel: conical flask
- Material, size, headspace, fill volume: 500 mL
- Aeration: yes, with clean, oil-free compressed air
- No. of vessels per concentration (replicates): 1 per concentration (except 3 replicates of the 1000 mg/L test concentration)
- No. of vessels per control (replicates): 2
- Sludge concentration (weight of dry solids per volume): 3 g SS/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: A synthetic sewage feed was made by dissolving the following amounts of substances in 1 litre of deionized water:
16.0 g peptone,
11.0 g meat extrakt,
3.0 g urea,
0.7 g NaCl,
0.4 g CaCl2 x 2 H2O,
0.2 g MgSO4 x 7 H2O,
2.8 g K2HPO4
OTHER TEST CONDITIONS
- Adjustment of pH: The pH of the test item stock solutions were measured using a Hach HQ40d Flexi handheld meter and adjusted to between pH 7.0 to pH 8.0.
- Light intensity: laboratory lighting

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): The respiration rate of an activated sludge fed with a standard amount of synthetic sewage feed was measured after a contact time of 3 hours.
TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Range finding study
- Test concentrations: 10, 100, 1000
- Results used to determine the conditions for the definitive study: No statistically significant toxic effects were shown at all of the test concentrations employed. It was therefore considered justifiable not to perform a definitive test.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Observations were made on the test preparations throughout the test period. Observations of the test item vessels at 0 hours were made prior to addition of activated sewage sludge.
- Observations on Test Preparations:
* Negative control (not exposed to the test item): after 3hrs: Dark brown dispersion (all replicates)
* Test item: Dark brown dispersion, no undissolved test item visible (all test concentrations)
* Positive control (3,5- dichlorophenol): Dark brown dispersion, no undissolved reference item visible (all test concentrations)

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: 3-hr EC50 value of 6.2 mg/L, 95% C.L. 4.7 to 8.2 mg/L.

Reported statistics and error estimates:

The percentage inhibition values were plotted against concentration, a line fitted using the Xlfit software package (IDBS) and the EC10, EC20, EC50 and EC80 values for the reference item and the EC10 and EC20 values for the test item determined from the equation for the fitted line.
The EC50 and EC80 values for the test item were determined by inspection of the inhibition of respiration rate data.
95% confidence limits were calculated for the reference item EC50 value using the method of Litchfield and Wilcoxon (Litchfield and Wilcoxon, 1949).
In order to determine the NOEC, one way analysis of variance incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf, 1981) and Dunnett's multiple comparison procedure for comparing several treatments with a control (Dunnett, 1955) was carried out on the oxygen consumption data after 3 hours for the control and all test concentrations to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package (SAS, 1999 - 2001).

Validity criteria fulfilled:

yes

Conclusions:

An activated sludge respiration inhibition test (according to OECD guideline 209) with activated sludge from a predominantly domestic sewage treatment plant resulted in a 3-h EC50 > 1000 mg/L. The No Observed Effect Concentration (NOEC) after 3 hours exposure was 100 mg/L. The results of the study are reliable without restriction.

Description of key information

Based on the guideline study by Best (2017), the effect of the test item on the respiration of activated sewage sludge micro-organisms gave a 3-Hour EC50 value of greater than 1000 mg/L.  The No Observed Effect Concentration (NOEC) after 3 hours exposure was 100 mg/L. Results of this study are reliable without restrictions.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

100 mg/L