Registration Dossier

Ecotoxicological information

Toxicity to microorganisms

Currently viewing:

Administrative data

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:
from 1993-11-10 to 1994-02-24
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: OECD Guideline study, sludge density lower than guideline recommendations.
Justification for type of information:
The justification for read across is provided as an attachment in IUCLID Section 13.
Reason / purpose:
read-across: supporting information
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
yes
Remarks:
sludge density lower than guideline recommendations
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
No test surrogate or analogue material
Analytical monitoring:
no
Details on sampling:
Not relevant (no analytical monitoring)
Vehicle:
no
Details on test solutions:
The test susbtance was known to be water soluble up to 10 mg/L.
To prepare the 1 mg/L test medium, a stock solution (10 of test substance per liter) was made approimately 1 hour 30 minutes before the start of the study, dissolving the test substance into dilution water (distilled water); pH of this stock solution was not adjusted.
To prepare the 10, 50 and 100 mg/L test media, no stock solution was prepared (the test substance was directly added to the test media).
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Source:activated sludge from a sewage treatment plant treating predominantly domestic sewage (Pierre-Bénite - F - 69310 Lyon)
- Method of cultivation: the following stock solution was made for activated sludge alimentation (synthetic seawage), and used at the rate of 16 mL for 500 mL fo final medium:
- Composition (grammes per litre of distilled water):
Peptone 16.0
Meat extract 11.0
Urea 3.0
NaCl 0.7
CaCl2-2H2O 0.4
MgSO4-7H2O 0.2
K2HPO4 2.8
- pH: 7
- Preparation of inoculum for exposure:
- The sludge was not used on the day of collection: it was maintained aerated at thermoregulated temperature (20°C), and added with synthetic sewage.
- Coarse particles were removed after settling for 15 minutes.
- The concentration of suspended solids was evaluated from a small amount of sludge.
- The sludge was centrifuged and the suspended solids obtained were washed before to be resuspended for the preparation of the inoculum (centrifuged 3 times with synthetic sewage for 5 minutes at 2,500 r.p.m., with discarding of the supernatants).
- The suspended solids were then re-suspended into synthetic sewage so as to obtain a concentration of suspended solids of approximately 2 grammes per litre in the inoculum.
- The inoculum was then kept aerated and at thermoregulated temperature (20°C) until its using for the preparation of the media.
- Pretreatment: no pretreatment
- Initial biomass concentration: 800 mg of suspended solids per litre of control, reference and test media.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Remarks on exposure duration:
No remarks
Post exposure observation period:
No post-exposure
Hardness:
No data
Test temperature:
17.5 - 21.9 °C (see table 2 §Remarks on results including tables and figures)
pH:
7.77 - 8.03 (see table 2 § Remarks on results including tabkes and figures)
Dissolved oxygen:
Measurements of the dissolved oxygen concentration over a period of 10 minutes (or until the concentration reaches 1 mg/L) after 3 hours exposure, in 500 mL samples from each container, performed in 500mL BOD flasks, with O2-electrode.
Salinity:
Not relevant (frehswater study)
Nominal and measured concentrations:
Nominal concenrations (range-finding test) : 0, 1, 10, 50 and 100 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): closed with covers
- Material, size, headspace, fill volume: 1000 mL beakers containing 500ml test medium
- Aeration: continuous aeration (air-flow: 0.5 to 1 litre per minute) and continuous magnetic stirring
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- Biomass loading rate: 800 mg/L


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: distilled water
- Adjustment of pH: no


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Oxygen consumption

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0, 1, 10, 50 and 100 mg/L
- Results used to determine the conditions for the definitive study: No definitive study (EC50 not reached in the range-finding test)
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Remarks on result:
other: no remarks
Details on results:
See table 1 §Remarks on results including tables anf figures
Results with reference substance (positive control):
See table 1 §Remarks on results including tables anf figures
Reported statistics and error estimates:
No statistics (range-finding study): it was imposible to reach the EC50 with the concenrations tested

Table 1. Respiration rates and percent inhibitions (rounded values)

 

 

Flask number

Respiration rates (mg/L/h)

Inhibition of respiration (%)

Control assay

FB1

40

/

FB2

41

/

Test substance assay

FT1                    1 mg/L

88

-116

FT10                 10 mg/L

58

-42

FT50                 50 mg/L

50

-23

FT100              100 mg/L

60

-47

Reference assay

FR5                    5 mg/L

57

-41

FR15                 15 mg/L

28

31

FR30                 30 mg/L

15

63

Physico-chemical assay

FPC

0

100

 

 

Table 2. pH and temperatures at the start and the end of the study

 

 

Flask number

Start of the study

End of the study

 

pH value

Temperature °C

pH value

Temperature °C

Control assay

FB1

7.99

20.7

8.25

20.3

FB2

8.01

19.8

8.69

17.5

Test substance assay

FT1         1 mg/L

7.77

21.5

7.95

21.9

FT10      10 mg/L

7.87

20.7

8.12

19.8

FT50      50 mg/L

8.00

20.1

8.47

18.4

FT100   100 mg/L

8.03

20.4

8.15

21.1

Reference substance assay

FR5         5 mg/L

7.99

20.8

8.17

20.7

FR15      15 mg/L

7.84

20.9

8.32

20.7

FR30      30 mg/L

7.86

21.2

8.27

20.3

Physico-chemical assay

FPC

5.95

20.0

6.47

20.0

Validity criteria fulfilled:
yes
Conclusions:
The 3h-EC50 of HDF 200 for respiration inhibition in activated sludge is >100 mg/L
Executive summary:

Microorganisms from activated sludge (predominantly domestic) were exposed to a range of concentrations of test substance for 3 hours: 1, 10, 50 and 100 mg test subtance per litre of test medium according to OECD guideline 209. Two control media (without test substance) were tested. A "physico-chemical assay" was also performed in the presence of mercuric chloride as biological oxygen respiration inhibitor, in order to determine the total consumption of oxygen as a result of non-biological processes.

The dissolved oxygen concentration was measured in 500 mL from each medium after approximately 3 hours of exposure.

A reference substance, 3,5 - dichlorophenol, was tested under the same conditions in order to control the sensitivity of the activated sludge (3 concentrations).

The 3h-EC50 value was higher than 100 mg/L in the range-finding study (phase I), then the study was considered as complete and no further evaluation of this value was performed.

The criteria quality for the study, as notified in the OECD and EEC standards and relative levels of oxygen consumption in the two control media and to the toxicity of 3,5 -dichlorophenol were respected.

No inhibition of respiration was observed under the experimental conditions employed, until the concentration of 100 mg of test substance per liter of test medium; the 3h-EC50 (effective concentration which inhibits respiration in activated sludge by 50%) may be expressed as follows:

EC50 - 3 hours = higher than 100 mg/L

Description of key information

C14-20 Aliphatics (≤2% aromatic) are not toxic to microorganisms: the EC50 is greater than 100 mg/L.

Key value for chemical safety assessment

Additional information

One study dealing with toxicity to microorganisms was performed with Hydrocarbons, C14-C18, n-alkanes, isoalkanes, cyclics, <2% aromatics (TOTAL, 1994). The EC50was reported to be greater than 100 mg/L.

Moreover, the biodegradation studies showed that C14-20 Aliphatics (≤2% aromatic) are readily biodegradable and therefore they are not toxic to microorganisms.