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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 2008).
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: OECD Guideline 301 A (Ready Biodegradability: DOC Die Away Test)
GLP compliance:
yes (incl. certificate)
Remarks:
Sächsisches Ministerium für Umwelt und Landwirtschaft, Germany
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The weighed amount of the test item or reference item was mixed with mineral medium (prepared in accordance with OECD guideline 301 A) to prepare a test item stock solution (400 mg test item/L) and a reference item stock solution (1000 mg reference item/L).

Preparation of the toxicity control solution (2 replicates):
1420 mL mineral medium (prepared in accordance with OECD guideline 301 A), 200 mL test item stock solution (400 mg test item/L), 200 mL reference item stock solution (1000 mg reference item/L) and 180 mL Inoculum were pipetted into a conical glass flask (2 L glass flask).

Afterwards, the prepared solutions (2000 mL/flask) were mixed and a sample was taken from each flask to determine the initial concentration of DOC. The openings of the flasks were covered with aluminium foil in such a way as to allow free exchange of air between the flask and the atmosphere. The test solutions
were shaken using a shaking machine.
Test organisms (species):
other: secondary effluent of a wastewater treatment plant
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): The sample was taken on 01.06.2012 at the wastewater treatment plant “Parthe” (Am Klärwerk 49, 04451 Borsdorf, Saxony, Germany)
- Preparation of inoculum for exposure: pre-conditioned by aerating for 5 days (from 01. - 06.06.2012) under test conditions
- Initial cell/biomass concentration: 22.37 mg suspended solids/L in the test solutions
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
28 d
Details on test conditions:
TEST CONDITIONS
- Composition of medium: The mineral medium was prepared in accordance with OECD guideline 301 A (1992).
- Test temperature: 23.1 - 23.9 °C in a controlled-environment test room
- pH: 7.4
- Suspended solids concentration: 22.37 mg suspended solids/L
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 2 L conical glass flask
- Number of culture flasks/concentration: 2
- Measuring equipment: DOC analyser
- Test performed in open system: The openings of the flasks were covered with aluminium foil in such a way as to allow free exchange of air between the flask and the atmosphere.

SAMPLING
- Sampling frequency: 0, 1, 2, 10, 22, 27 and 28 days after test start
- Sampling method: Before sampling water loss was compensated if necessary by adding deionised water.
- Sample storage before analysis: If possible the analysis was performed immediately after sampling. Otherwise the samples were stored at 2-4 °C for a maximum of 48 hours or below -18 °C.

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes, 2 bottles
- Abiotic sterile control: no
- Toxicity control: yes, 1 bottle
- Reference item: yes, 1 bottle
Reference substance (positive control):
yes
Remarks:
Sodium acetate
Duration:
10 d
Dose descriptor:
NOEC
Effect conc.:
>= 40 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):
23.4% biodegradation after 1 days
89.2% biodegradation after 2 days
89.0% biodegradation after 10 days
79.5% biodegradation after 22 days
79.9% biodegradation after 27 days
81.8% biodegradation after 28 days

The toxicity control attained 51.4% degradation after 10 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 35% degradation occurred in the toxicity control, the substance is expected to have low toxicity towards aquatic microorganisms. The test item concentration in the toxicity control of 40 mg/L can be used as NOEC value for the toxicity to aquatic microorganisms.

Description of key information

NOEC (10 d) ≥ 40 mg/L (OECD 301 A, toxicity control)

Key value for chemical safety assessment

Additional information

Since no studies on the toxicity of Reaction mass of 2-(3,4-dimethyl-1H-pyrazol-1-yl)succinic acid and 2-(4,5-dimethyl-1H-pyrazol-1-yl)succinic acid to aquatic microorganisms are available a ready biodegradability study is used to derive a NOEC for the toxicity to aquatic microorganisms. If a compound does not inhibit the degradation of a positive control at a certain concentration, this concentration can be used as a NOEC value. This approach is in accordance with the Guidance on information requirements and chemical safety assessment (Chapter R.7b: Endpoint specific guidance, ECHA 2017).

A substance can be assumed to be not inhibitory to aquatic microorganisms, if in the toxicity control of a ready biodegradation test, more than 35% degradation based on DOC removal occurred within 14 days (OECD guidance 301). For the present test substance a biodegradation test according to OECD guideline 301 A is available (Schulz, 2012). The test includes a toxicity control, which contains 40 mg/L of the test substance and 100 mg/L of the reference material sodium acetate. The toxicity control attained 51.4% degradation after 10 days of incubation. Hence, the substance is not expected to be toxic to aquatic microorganisms and the test item concentration of 40 mg/L can be used as NOEC.