2,2'-[azobis(1-methylethylidene)]bis[4,5-dihydro-1H-imidazole dihydrochloride

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:

24/3-25/3/14

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))

GLP compliance:

yes (incl. QA statement)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: direct addition
- Eluate:
- Differential loading:
- Controls:
- Chemical name of vehicle (organic solvent, emulsifier or dispersant):
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)):
- Evidence of undissolved material (e.g. precipitate, surface film, etc.):

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture:
- Name and location of sewage treatment plant where inoculum was collected: WWTP Mannheim, Germany (collection date: 24.03.14)
- Method of cultivation:
- Preparation of inoculum for exposure: sieving (1 mm mesh)
- Pretreatment: pre-aeration overnight, washing once with tap water
- Initial biomass concentration: 3 g/L dw

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Test temperature:

20.9-21.2 °C

pH:

pre incubation: 7.4-7.6
post incubation: 7.9 - 8.1

Nominal and measured concentrations:

1000, 500, 250, 125, 62.5 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beakers
- Type (delete if not applicable): open / closed
- Material, size, headspace, fill volume: glass, 1 L, 500 mL, 500 mL
- Aeration:
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates):
- No. of vessels per abiotic control (replicates):
- Sludge concentration (weight of dry solids per volume): 1.5 g/L dw
- Weight of dry solids per volume of reaction mixture per unit of time:
- Nutrients provided for bacteria:
- Nitrification inhibitor used (delete if not applicable): none / N-allylthiourea
- Biomass loading rate:
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: local tap water
- Particulate matter:
OTHER TEST CONDITIONS
- Adjustment of pH:
- Photoperiod:
- Light intensity:
- Details on termination of incubation:
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using fewer concentrations than requested by guideline:
- Range finding study
- Test concentrations:
- Results used to determine the conditions for the definitive study:

Reference substance (positive control):

yes

Remarks:

3,5-DCP

Key result

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

410 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Duration:

3 h

Dose descriptor:

other: EC20

Effect conc.:

680 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

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

Results with reference substance (positive control):

The EC50 of the reference substance 3,5-dichlorophenol was in the range of 2-25 mg/L in 3 hours.

Reported statistics and error estimates:

The degree of inhibition was evaluated by Probit analysis according to Finney.

Validity criteria fulfilled:

yes

Executive summary:

In conclusion the 3 hours effect concentrations for Azo Initiator VA044 in the Activated Sludge Respiration Inhibition Test are:

The following effect concentrations (mg/L) were obtained:

Toxicity Response Variable	Based on nominal effect concentration [mg/L]	Confidence Limits (95%) lower/upper
EC10	410	302.2/555.7
EC20	680	572.3/810.5
EC50	> 1000	-
EC80	> 1000	-

The value of effect concentrations of EC10 EC20 were given with an accuracy of 2 significant digits. The degree of inhibition was evaluated by Probit analysis according to Finney.

The results in this study are consistent with all validity criteria and the test is valid according to the guidelines used for this study.

No deviations from the test guidelines or other incidents occurred during the course of the reported test which may have influenced the results.

Description of key information

Based on the findings in an OECD 209 study exposing activated sludge with the test substance an EC10 value of 410 mg/L and an EC50 value > 1000 mg/L after an exposure period of 3 hours were determined.

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L

EC10 or NOEC for microorganisms:

410 mg/L