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Toxicity to microorganisms

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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
13-01-2015 to 23-01-2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: test according to the guideline under GLP. Slight deviations of the oxygen concentration during the test did not affect the results
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Principles of method if other than guideline:
only the range-finding study was performed
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)
- Method: direct addition to reversed osmosis water, followed by ultrasonication for 15 min. The solution was stirred magnetically for 24 hours at 21-22 °C (shielded from light)
- Differential loading: 0, 10, 100 and 1000 mg/L (at 1000 mg/L 3 replicates)
- Controls: no loading (4 replicates) and positive control 3,5 dichlorophenol ( 1 replicate at 3.2, 10 and 32 mg/L)
- Evidence of undissolved material: at 1000 mg/L visible dispersed test substance throughout and adhered to the side of flask
- pH adjustment: at 100 and 1000 mg/L
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Source of inoculum/activated sludge: Severn Trent Water Plc sewage treatment plant Loughborough, Leicesterhire, UK
- Storage conditions: under aeration at 21 °C
- Storage length: used on the day of collection
- Preparation of inoculum for exposure: washed once by settlemnt and resuspended
- Concentration of sludge: suspended solids 3.0 g/L on day of the test
- pH: 7.2
Test type:
static
Water media type:
freshwater
Total exposure duration:
3 h
Post exposure observation period:
NA
Test temperature:
20-21 °C
pH:
at 30 min 7.6-7.8
at 3 hours 7.5-8.0
Dissolved oxygen:
measured at 30 minutes
in controls: 62-94% of saturation
in test solutions: 55-58% of saturation
Details on test conditions:
TEST SYSTEM
- Test vessel: 500 mL
- Type: conical (open)
- Aeration: 0.1-0.5 L/min
- No. of vessels per concentration (replicates): 1 for 10 and 100 mg/L, 3 at 1000 mg/L
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Test water: deionized reverse osmosis water
- Concentration of sludge: suspended solids 3.0 g/L on day of the test
- pH: 7.6-7.8 (initial after adjustment)
- respiratory substrate (dissolved in 1 liter of water with the aid of ultrasonication):
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
- Adjustment of pH: yes at 100 and 1000 mg/L
- Test temperature: 20-21 ºC

EFFECT PARAMETERS MEASURED : oxygen consumption at t=30 min and t=3 h (over linear part of the respiration curve)

TEST CONCENTRATIONS: 10, 100 and 1000 mg/L
Reference substance (positive control):
yes
Remarks:
3,5 dichlorophenol
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
oxygen respiration rate controls: 26.65. mg O2/g dry weight sludge/h
coefficient of variation replicates at 1000 mg/L : 4.0%
Results with reference substance (positive control):
EC50 for 3,5 dichlorophenol 7.1 mg/L (oxygen concentration at initial reading at 10 and 32 mg/l was > 7.0 mg/L)
Reported statistics and error estimates:
Bartlett test for homogeneity of variance
Dunnett test for statistically differences between test groups and controls
Validity criteria fulfilled:
yes
Remarks:
it was concluded that the deviations of the oxigen concentrations did not affect the study outcome
Conclusions:
In a respiration inhibition test the NOEC of the test substance was 1000 mg/L
Executive summary:

The test substance was tested in a respiration inhibition test at concentrations of 10, 100 and 1000 mg/L. No effects on the respiratory rate of the inoculum were reported at any of the concentrations tested. The NOEC is therefore 1000 mg/L.

Description of key information

The EC50 and NOEC were determined to be above the nominal tested concentration of 1000 mg/L (Harlan 2015)

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
1 000 mg/L

Additional information