Quaternary ammonium compounds, C12-14-alkyl(hydroxyethyl)dimethyl, chlorides

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1993-09-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted following GLP according to OECD guideline 209 "Activated sludge: Respiration inhibition test" and Directive 87/302/ECC Part C "Methods for the Determination of Ecotoxicity: Biotic degradation-Activated Sludge: Respiration Inhibition test."

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)

Principles of method if other than guideline:

N/A

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

[see concordance under 1.1. of the CSR]

IUPAC Name:

[see concordance under 1.1. of the CSR]

Details on test material:

- Name of test material (as cited in study report): C12/14 Dimethyl hydroxyethyl ammonium chloride (KDB-base), E-4408.01
- Molecular formula (if other than submission substance): N/A
- Molecular weight (if other than submission substance): N/A
- Smiles notation (if other than submission substance): N/A
- InChl (if other than submission substance): N/A
- Structural formula attached as image file (if other than submission substance): N/A
- Substance type: N/A
- Physical state: colorless liquid
- Expiration date of the lot/batch: N/A
- Radiochemical purity (if radiolabelling): N/A
- Specific activity (if radiolabelling): N/A
- Locations of the label (if radiolabelling): N/A
- Expiration date of radiochemical substance (if radiolabelling): N/A
- Stability under test conditions: N/A
- Storage condition of test material: N/A
- Other: Non volatile, density ~1, highly water soluble

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
- Melting point: N/A
- Boiling point: N/A
- Vapour pressure: N/A
- Water solubility (under test conditions): N/A
- Henry's law constant: N/A
- log Pow: N/A
- pKa: N/A
- Stability in water: N/A
- Stability in light: N/A
- pH dependance on stability: N/A


OTHER PROPERTIES (if relevant for this endpoint)
- Results of test for ready biodegradability: N/A
- Other:N/A

Sampling and analysis

Analytical monitoring:

no

Details on sampling:

- Concentrations: N/A
- Sampling method: N/A
- Sample storage conditions before analysis: N/A

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock of 2 g active/L was prepared by dissolving 5 g test substance in 1 L demineralized water.
- Eluate: N/A
- Differential loading: N/A
- Controls: The respiration rate of activated sludge as measured in two solutions containing no test substance is used as a control value.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N/A
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): N/A
- Evidence of undissolved material (e.g. precipitate, surface film, etc): N/A

Test organisms

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: No
- Method of cultivation: Activated sludge obtained from the waste water treatment plant of Bocholt.
- Preparation of inoculum for exposure: Upon return to the laboratory the sludge is sieved through a 0.5 mm sieve in order to remove large particles
- Pretreatment: N/A
- Initial biomass concentration: suspended solids of 4.6 g/L corresponding to a concentration of 1.84 g/L in final test solution.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

N/A

Test conditions

Hardness:

N/A

Test temperature:

22.3 degrees C

pH:

7.87-8.23

Dissolved oxygen:

N/A

Salinity:

N/A

Nominal and measured concentrations:

5.8, 10, 18 and 58 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 1000 ml borosilicate glass beakers.
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: The test system consisted of:
16 mL synthetic sewage feed
appropriate amount of stock solution of test substance
tap water to bring volume up to 300 mL
200 mL activated sludge mixed liquor (inoculums)
500 mL total volume/beaker
- Aeration: Aeration provided by Pasteur pipettes
- Type of flow-through (e.g. peristaltic or proportional diluter): N/A
- Renewal rate of test solution (frequency/flow rate): N/A
- No. of organisms per vessel: N/A
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): N/A
- Biomass loading rate: N/A


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: aerated tap water
- Total organic carbon: N/A
- Particulate matter: N/A
- Metals: N/A
- Pesticides: N/A
- Chlorine: N/A
- Alkalinity: N/A
- Ca/mg ratio: N/A
- Conductivity: N/A
- Culture medium different from test medium: N/A
- Intervals of water quality measurement: N/A


OTHER TEST CONDITIONS
- Adjustment of pH: N/A
-A synthetic sewage feed was prepared by dissolving the following nutrients in 1 L deionized water:
16 g peptone
11 g meat extract
3 g urea
0.7 g NaCl
0.4 g CaCl2*2H2O
0.2 g MgSO4*7H2O
2.8 g K2HPO4
- Photoperiod: N/A
- Light intensity: N/A


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Repiration rate is measured over a period of 10 minutes


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.7-1.8
- Justification for using less concentrations than requested by guideline: N/A
- Range finding study
- Test concentrations: 10, 100, 1000 mg/L
- Results used to determine the conditions for the definitive study: 10mg/L=32.69% inhibition, 100mg/L=94.23% inhibition, 1000mg/L=98.08% inhibition

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

19 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Effect concentrations 2

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

4 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Effect concentrations 3

Duration:

3 h

Dose descriptor:

other: EC90

Effect conc.:

88 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: N/A
- Effect concentrations exceeding solubility of substance in test medium:N/A

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: The 3 hours EC50 values was well within the acceptable range of 5 to 30 mg/L (11.4 mg 3,5-dichlorophenol/L)
- Other:N/A

Reported statistics and error estimates:

N/A

Any other information on results incl. tables

Table 1. Activated Sludge Respiration Inhibition by KDB

Nominal Concentration of KDB (mg/L)	Respiration Rate (mg O2/Lh)	Inhibition (%)
5.8	15.8	17.0
10	13.7	28.3
18	10.4	45.3
32	6.48	66.0
58	2.88	84.9

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

The respiration rates of the blanks did not differ more than 15% and the 3-h EC50 value of reference 3.5-dichlorophenol was well within the acceptable range.

Conclusions:

The concentration of KDB that reduced the respiration rate of activated sludge by 50% within 3 hours was 19 mg/L.

Executive summary:

The inhibitory effect of C12/14 dimethyl hydroxyethyl ammonium chloride (KDB base) on activated sludge microorganisms was determined in an OECD 209 guideline test. The nominal test concentrations were 5.8, 10, 18, 32, and 58 mg a.i/L. The activated sludge microorganisms were from a predominantly domestic source.

The 3 hour EC50 was 19 mg a.i./L (% inhibition of respiration of activated sludge microorganisms). The EC10 was 4 mg/L, and the EC90 was 88 mg/L. This toxicity study satisfies the guideline requirements for the OECD 209 activated sludge respiration inhibition test.