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Ecotoxicological information

Toxicity to microorganisms

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Reference
Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Data based on the inhibition control of a ready biodegradability study. This approach is in accordance with the Guidance on information requirements and chemical safety assessment (Chapter R.7b: Endpoint specific guidance, ECHA 2017).
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Version / remarks:
adopted July 1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)
Version / remarks:
No 440/2008, 2008
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
Ministerium für Arbeit, Integration und Soziales des Landes Nordrhein-Westfalen, Germany
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- 25 mg of the test item and 25 mg of the reference compound were weighed out on aluminium foil and both were added to the test flasks, filled with 200 mL of mineral medium. Afterwards the flask volume was made up to 250 mL with mineral medium containing the inoculum to give a test concentration of 100 mg test item and reference compound/L.
Test organisms (species):
other: activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum: mixed population of aquatic microorganisms (activated sludge), aeration tank of a wastewater plant treating predominantly domestic sewage (Wupper area water authority, WWTP Odenthal), date of collection: 27 Aug 2018
- Storage conditions: Before use, the inoculum was stored for one day at room temperature under continuous stirring with aeration
- Preparation of inoculum for exposure: The sludge was washed twice by adding mineral medium and centrifuging for 10 min at 2000 rpm and 20 °C and decanting off the supernatant.
- Pretreatment: An aliquot of the wet sludge was dried in order to determine the wet weight / dry weight ratio of the sludge and to prepare a stock suspension (activated sludge) of 3 g dw/L. The calculated amount of sludge, needed to achieve 300 mL of this stock suspension, was dissolved in mineral medium and then filled up to a defined end volume.
- Concentration of sludge: 30 mg/L suspended solids
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
14 d
Test temperature:
22 ± 1 °C
Details on test conditions:
- Composition of medium: The mineral medium was prepared from stock solutions of mineral components according to the guideline.
- Test temperature: 22 ± 1 °C
- pH: 7.4 ± 0.2
- pH adjusted: no
- Suspended solids concentration: 30 mg/L suspended solids
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: OxiTopControl System (WTW)
- Number of culture flasks/concentration: 3 for test item concentration 100 mg/L
- Method used to create aerobic conditions: Test vessels were stirred by an inductive stirring system for a maximum test period of 56 days.
- Measuring equipment: During the test the O2 uptake was continuously measured with a manometrie BOD measuring device (OxiTop®-C measuring head (WTW Wissenschaftlich-Technische Werkstätten GmbH & Co. KG, Weilheim, Germany).

SAMPLING
- Sampling frequency: continously

CONTROL AND BLANK SYSTEM
- Inoculum blank: 3 replicates
- Toxicity control: 1 replicates
- Reference control: 2 replicates

STATISTICAL METHODS: not applicable
Reference substance (positive control):
yes
Remarks:
sodium benzoate
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Result of toxicity control from ready biodegradability test
Results with reference substance (positive control):
The reference compound sodium benzoate showed 88% degradation after 14 days.

The toxicity control attained 34% degradation after 14 days of incubation.

“If in a toxicity test, containing both the test and reference substance, less than 35% degradation (based on total DOC) or less than 25% (based on total ThOD or ThCO2) occurred within 14 days, the test substance can be assumed to be inhibitory.” (OECD Guideline 301)

Since more than 25% degradation occurred in the toxicity control, the substance is with high probability not toxic to aquatic microorganisms. The test item concentration in the toxicity control of 100 mg/L can be used as NOEC value for the toxicity to aquatic microorganisms.

Validity criteria fulfilled:
yes

Description of key information

NOEC (14 d) ≥ 100 mg/L (toxicity control, OECD 301F)

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
100 mg/L

Additional information

One study investigating the toxicity to activated sludge microorganisms is available. The endpoint is based on the inhibition control of a ready biodegradability study. In this study according to OECD 301F a 14 d-NOEC of ≥ 100 mg/L could be determined, based on the toxicity control.

One study investigating the biodegradation of the test item was investigated in an O2-consumption test according to OECD guideline 301F using domestic activated sludge as inoculum (CURRENTA GmbH & Co., 2019). This test also consisted of a toxicity control. The toxicity control was tested with a test concentration of the test item of 100 mg/L (+ reference item at a concentration of 100 mg/L). The toxicity control attained 34% degradation after 14 days of incubation. If in a toxicity control, containing both the test substance and a reference compound, more than 25% degradation (based on total ThOD) occurred within 14 days, the test substance can be assumed to be non-inhibitory to microbial organisms. This approach is in line with the OECD Guideline 301 as well as the ECHA Guidance R.7b (ECHA, 2017). Thus, the test item concentration in the toxicity control of 100 mg/L can be used as NOEC value for the toxicity to aquatic microorganisms (NOEC (14 d) ≥ 100 mg/L).