Registration Dossier

Administrative data

Endpoint:
activated sludge respiration inhibition testing
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
on day 1998-08-27
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report date:
1998

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes (incl. QA statement)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Tri (hexyl, octyl, decyl) citrate
- Substance type: pure active substance
- Physical state: liquid
- Stability under test conditions: in water for at least 2 hours
- Storage condition of test material: in original container, at room temperature, in the dark
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
- Melting point: -29°C (freezing point: -35.5°C)
- Boiling point: test substance showed decomposition above 260°C
- Vapour pressure: 8.5 x 10E-3 p at 25°C
- Water solubility (under test conditions): < 1 mg/L
- Solubility in organic solvents: soluble
- log Pow: > 6
- pKa: not applicable
- Base or acid catalysis of test material: no data
- UV absorption: no data
- Stability of test material at room temperature: stable
- pH dependance on stability: no data


OTHER PROPERTIES (if relevant for this endpoint)
- Adsorption characteristics: no data

Sampling and analysis

Analytical monitoring:
no

Test solutions

Vehicle:
no
Details on test solutions:
The test substance was directly weighed into the test flasks. Then, the test flasks were filled each with 284 mL tap water.

Test organisms

Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Laboratory culture: no
- Method of cultivation: not applicable
- Preparation of inoculum for exposure: The sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was resuspended in tap water and again centrifuged. The latter procedure was repeated twice. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight determined.
- Pretreatment: Calculated aliquots of washed sludge suspension, corresponding to 3 g dry material/L were made up with tap water. To this mixture, 50 mL synthetic sewage feed/L was added daily, starting 3 days prior to use, and the sludge was kept at room temperaure under continuous aeration until use. Just before the start of the test the dry weight was again measured and adjusted to 2 g dry weight/L.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Post exposure observation period:
none

Test conditions

Hardness:
not mentioned
Test temperature:
19 °C
pH:
7.6 - 8.3
Dissolved oxygen:
5.4 - 9.2 mg/L
Nominal and measured concentrations:
Nominal: 10, 32, 100, 320, 1000 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 1 L glass flask with a fill volume of 500 mL
- Aeration: compressed air at an air-flow rate of approx. 0.6 L/min
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water, synthetic sewage feed according to guideline


OTHER TEST CONDITIONS
- Adjustment of pH: no


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Respiration rate of each test medium after 3 hours was measured with an oxygen electrode (ECM-multi, Dr. Lange, Germany), and was recorded for 10 minutes. During measurement, the samples were continuously stirred on a magnetic stirrer. The pH amd the oxygen concentrations were determined before the start of the test and at the end of the incubation period in all test concentrations and controls. The water temperature was measured in one control medium at the start and the end of the incubation period.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: not mentioned
- Range finding study. Not performed
Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol (final concentrations in the test: 10, 32 and 100 mg/L)

Results and discussion

Effect concentrations
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
Remarks:
respiration rate
Details on results:
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: yes
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Relevant effect levels: 3-hour EC50 = 15 mg/L
Reported statistics and error estimates:
no statistics performed

Any other information on results incl. tables

Table: Influence of test substance and reference substance on oxygen consumption of activated sludge

Treatment   Concentration [mg/L]  Oxygen consumption[mg O2/L min]  Inhibition[%]
Control   -  0.411 / 0.379*  -
 Test substance  10  0.337  14.8
 Test substance  32  0.362  8.3
 Test substance  100  0.378  4.4
 Test substance  320  0.316  20.1
 Test substance  1000  0.349  11.7
 3,5 -DCP**  10  0.288  27.1
 3,5 -DCP**  32  0.036  90.9
 3,5 -DCP**  100  0.039  90.1

* repeated test ** reference substance 3,5 -Dichlorophenol

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
The respiration rates of the two control did not differ by more than 15%. The 3-hour EC50 fo the reference substance was in the range of 5 to 30 mg/L. The concentration of dissolved ocygen did not drop below 2.5 mg/L during the incubation period.
Conclusions:
In comparison to the inoculum controls the respiration rate of the activated sludge was inhibited between 4.4 % and 20.1 % up to the highest tested concentration of nominal 1000 mg test substance/L. However, no concentration-effect relationship could be determined. Thus, the respiration rate was not significantly inhibited up to the highest tested concentration. Concentrations exceeding 1000 mg/L nominal were not tested.
From these inhibition rates, the 3-hour EC20 and EC50 could not be quantified. Nevertheless, the 3-hour EC50 was clearly higher than 1000 mg test substance/L under the present test conditions.