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Toxicity to microorganisms

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Reference
Endpoint:
toxicity to microorganisms, other
Remarks:
Biodegradation in water: screening test, toxicity control is used to derive effect concentration
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: EU method C.4-D
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
other: OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Version / remarks:
1992
Deviations:
no
Principles of method if other than guideline:
From biodegradation in water: screening test, toxicity control is used to derive effect concentration
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The test item was a 5% aquatic dispersion.The concentration in the test assays were 2000 mg dispersion per litre mineral test medium and 400 mg dispersion per litre mineral test medium, respectively. 2000 mg test item correspond to 290 mg ThOD, 400 mg test item correspond to 58 mg ThOD.
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
Fresh samples of activated sludge are withdrawn on June 10th, 2014 from the sewage treatment plant Ruhrverband Kläranlage, Sunthelle 6, 57392 Schmallenberg, Germany, which is mainly fed with municipal wastewater. Since it was not necessary, the samples were not washed with mineral medium after the arrival at the laboratory but kept aerobic until use.The concentration used in the test was 29.6 mg dry mass/litre (7.39 mg dry mass/250 mL).
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
28 d
Test temperature:
22 °C ± 1°C
pH:
7.4 ± 0.2 at test start
Nominal and measured concentrations:
test item at 2000 mg per litre + reference item at 100 mg per litre
Details on test conditions:
TEST CONDITIONS
- Composition of medium:
( a )
KH2PO4: 8.50 g/L
K2HPO4: 21.75 g/L
Na2HPO4 x 12 H2O: 67.13 g/L
NH4Cl: 0.50 g/L
adjusted to 7.4 ± 0.2 with H2SO4 (50 g/L)
( b )
CaCl2 x 2 H2O: 36.40 g/L
( c ) MgSO4 x 7 H2O: 22.50 g/L
( d ) FeCl3 x 6 H2O: 0.25 g/L.
The mineral medium applied in the test contained 10 mL/L of mineral stock solution a and 1 mL/L of the mineral stock solution b–d, respectively.
- Test temperature: 22 °C
- pH: 7.4 +/- 0.2
- pH adjusted: no
- Continuous darkness: yes

TEST SYSTEM
- Test vessel: 500 mL glass vessels, fill volume 250 mL
- Aeration: yes
- Number of culture flasks/concentration: 2 vessels containing test item (2000 mg/L) and inoculum; 2 vessels containing test item (400 mg/L) and inoculum; 2 vessels containing only inoculum; 2 vessels containing test item (2000 mg/L) and a sterilising agent
- Method used to create aerobic conditions: The suspension was aerated during the whole test.
- Measuring equipment: SAPROMAT respirometer (VOITH Inc.).

SAMPLING
- Sampling frequency: The measurement and recording of the oxygen demand was carried out continuously

CONTROL AND BLANK SYSTEM
- Inoculum blank: The blank control consists of inoculated mineral medium only
- Abiotic sterile control: An abiotic control containing test item at 2000 mg per litre sterilized mineral test medium was applied. The assay was sterilized by adding HgCl2.
- Toxicity control: A toxicity control containing test item at 2000 mg per litre and reference item at 100 mg per litre mineral test medium was applied.

STATISTICAL METHODS:
Theoretical Oxygen Demand (ThOD):
The Theoretical Oxygen Demand (ThOD) was calculated on the basis of the sum formula of the test and reference item by:
ThOD [g/g] = 16 * (2 C + 1/2 H + 5/2 N + 1/2 Na – O) / Molecular weight.
The ThOD values for Na-benzoate, the test item and the toxicity control were determined as follows:
ThODPC-2014-536: 0.145 mg O2/mg test item
ThODNa-Benzoate: 1.665 mg O2/mg reference item
ThODtoxicity control: 0.218 mg O2/mg substance mixture
Biochemical Oxygen Demand (BOD):
The Biochemical Oxygen Demand (BOD) was calculated on the basis of the test raw data by BOD [mg/mg] = mg O2 uptake corrected by blank per mg test item.
The percent degradation was calculated according to the following formula:
Dt = [(Ct –Cb) / ThOD] x 100Dt: degradation (%) at time t;
Ct: mean oxygen consumption (mg/L) in the test suspension at time t;
Cb: mean oxygen consumption (mg/L) in the blanks at time t;
ThOD: Theoretical oxygen demand of the test suspension (mg/L).
Reference substance (positive control):
no
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
2 000 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
inhibition of total respiration
Remarks on result:
other: comparison of procedural and toxicity control
Details on results:
The biodegradation of the item mixture in the toxicity control was found to be 31.1% after 14 days of incubation. Thus, according to the test guidelines, the test item had no inhibitory effect on activated sludge microorganisms at the tested concentration of 2000 mg/L, as the biodegradation in the toxicity control was higher than 25% within 14 days of incubation.
Validity criteria fulfilled:
yes
Conclusions:
The toxicity of C16 Alkylamidopropyltrimethylammonium Chloride to microorganisms was investigated during a ready biodegradation study. The test item had no inhibitory effect on activated sludge microorganisms at the tested concentration of 2000 mg/L.
Executive summary:

The toxicity of C16 Alkylamidopropyltrimethylammonium Chloride to microorganisms was investigated during a ready biodegradation study according to EU Method C.4-D (2008) and OECD guideline 301 F (1992), Manometric Respirometry Test over a period of 28 days and using an inoculum obtained from activated sludge from a predominantly domestic sewage treatment plant.

Inoculum blank, procedural/functional control with the reference substance Sodium benzoate and a toxicity control with 2000 mg/L test item and 100 mg/L reference item Sodium benzoate were performed.

The reference item degraded normally (91.6% by within 14 days). By the end of the test, the reference item was degraded to an average of 96.4%.

The biodegradation of the item mixture in the toxicity control was found to be 31.1% after 14 days of incubation. Thus, according to the test guidelines, the test item had no inhibitory effect on activated sludge microorganisms at the tested concentration of 2000 mg/L, as the biodegradation in the toxicity control was higher than 25% within 14 days of incubation.

Description of key information

14 d NOEC = 2000 mg/L; toxicity control from OECD Guideline 301 F (1992); RL1; GLP

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
2 000 mg/L

Additional information

The toxicity of C16 Alkylamidopropyltrimethylammonium Chloride (95% a.i.) to microorganisms was investigated during a ready biodegradation study according to EU Method C.4-D (2008) and OECD guideline 301 F (1992), Manometric Respirometry Test over a period of 28 days and using an inoculum obtained from activated sludge from a predominantly domestic sewage treatment plant.

Inoculum blank, procedural/functional control with the reference substance Sodium benzoate and a toxicity control with 2000 mg/L test item and 100 mg/L reference item Sodium benzoate were performed.

The reference item degraded normally (91.6% by within 14 days). By the end of the test, the reference item was degraded to an average of 96.4%.

The biodegradation of the item mixture in the toxicity control was found to be 31.1% after 14 days of incubation. Thus, according to the test guidelines, the test item had no inhibitory effect on activated sludge microorganisms at the tested concentration of 2000 mg/L, as the biodegradation in the toxicity control was higher than 25% within 14 days of incubation.