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Toxicity to microorganisms

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Endpoint:
activated sludge nitrification inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Principles of method if other than guideline:
Inhibition of nitrification process
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
no
Test organisms (species):
activated sludge of a predominantly domestic sewage
Test type:
static
Water media type:
freshwater
Limit test:
no
Remarks on exposure duration:
2-4 h incubation period
Test temperature:
25 °C
pH:
7.6 - 7.8
Dissolved oxygen:
> 1mg/L
Salinity:
BOD 250 mg/L
50-80 mg/L ammonia
0.003 mol/L NaHCO3
0.002 mol/L (NH4)2SO4
Details on test conditions:
pre-cleaned activated sludge in particle-free communal waste water (BOD5: 250 mg/l; NH4-N/l: 50-80 mg)
Duration:
4 h
Dose descriptor:
other: EC75
Effect conc.:
11.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of nitrification rate

2 -4 h incubation

mayor scope of study was to develop method

Executive summary:

Nitrification inhibition by m-cresol was measured by a method similar to ISO/DIS 9509 yielding an EC75 of 11.4 mg/L during a 2 -4 h incubation period.

Endpoint:
toxicity to microorganisms, other
Remarks:
cell multiplication inhibition test
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Principles of method if other than guideline:
cell multiplication inhibition test
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
a stock solution was prepared; to achive the test concentrations an defined amount of the stock solution was dissolved
Test organisms (species):
Entosiphon sulcatum
Details on inoculum:
holozoic bacteriovorous flagellate
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
25°C
pH:
initial pH 6.9
Duration:
72 h
Dose descriptor:
other: TT
Effect conc.:
31 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition

TT = Toxicity threshold; determined at 5 % effect compared to control

Executive summary:

In a cell multiplication inhibition test conducted with Entosiphon sulcatum a 72h-NOEC of 31 mg/l is determined.

Endpoint:
toxicity to microorganisms, other
Remarks:
cell multiplication test
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Principles of method if other than guideline:
Method: other: Cell multiplication inhibition test
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
a stock solution was prepared; to achive the test concentrations a defined amount of the stock solution was dissolved
Test organisms (species):
Pseudomonas putida
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
16 h
Test temperature:
25°C
Duration:
16 h
Dose descriptor:
other: TT
Effect conc.:
53 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition

TT = Toxicity threshold; determined at 3 % effect compared to control

Executive summary:

For Pseudomonas putida a NOEC of 53 mg/l was obtained in a cell multiplication inhibition test during 16 hours of exposure.

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
yes
Remarks:
Incresed buffer capacity (K2HPO4)
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
no
Test organisms (species):
activated sludge, domestic
Details on inoculum:
Activated sludge obtained from local municipal wastewater treatment plant, settled and liquor dicarded. Solids detremined by gravimetry. Sediment wahsed three times. Resuspended and sludge adjusted to 4000 +/- 400 mg dw/L. Aerated at 0.5 L/min per 9 L of suspension. Daily resupplied with synthetic sewage stock solution according to OECD guideline with the exception of K2HPO4 which was incresed to 28 g/L in the stock solution
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3.08 h
Post exposure observation period:
5 min to determine respiration rate
Test temperature:
21 °C
pH:
7.0 adjusted with H3PO4 in sludge, and 7.5 during incubations
Dissolved oxygen:
not known, but aerated at 0.1 to 1 L/min to ensure mixing
Details on test conditions:
Type: aquatic
Duration:
3 h
Dose descriptor:
IC50
Effect conc.:
461.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Results with reference substance (positive control):
3,5-Dichorophenol

synthetic sewage stock solution slightly different from OECD
guideline
Validity criteria fulfilled:
yes
Executive summary:

The sensitivity of activated sludge to m-cresol was determined by a respiration inhibition test according to OECD Guideline 209. The 3h IC50 for respiration inhibion of microorganisms is 462 mg/L.

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Principles of method if other than guideline:
methanogens: CH4 release from acetate in the presence of chemical additives
aerobic heterotrophs: inhibition of respiration according to Owen WF (1979) Bioassay for monitoring biochemical methane potential and anaerobic toxicity. Water Res. 13, 485 (prolonged incubation compared with ISO 8192)
GLP compliance:
no
Analytical monitoring:
no
Test organisms (species):
other: mixed liquor of activated sludge treatment plant
Details on inoculum:
Nitrosomonas: seed bacteria were obtained from mixed liquor of activated sludge treatment plant
methanogens: anaerobic toxicity test cultue reared in the laboratory for 10 years
aerobic heterotrophs: seed bacteria were obtained from mixed liquor of activated sludge treatment plant
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
24 h
Remarks on exposure duration:
Mehtnaogens 1,2,3, or 4 d, aerobic heterotrphs 15, 27,38, and 49 h
Test temperature:
25 °C (methanogens 35°C; aerobic heterotrophs 25 and 35°C)
pH:
6.5-8 (methanogens and aerobic heterotrophs 7)
Dissolved oxygen:
not measured but atmospheric composition (by volumes):
Nitrosomonas: N2:O2 = 1.6
methanogens: N2:CO2 = 2
aerobic heterotrophs: N2:O2 = 1
Details on test conditions:
Type: aquatic
Duration:
96 h
Dose descriptor:
IC50
Effect conc.:
890 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: methane production
Duration:
49 h
Dose descriptor:
IC50
Effect conc.:
440 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate

Test with Nitrosomas sp.: In principal the test is comparable to standard methods, but the authors state that the compounds with log IC50 <1.5 µmol/l had questionable results. The result is considered invalid by the authors.

Executive summary:

The effect of m-cresol on activated sludge was determined by a respiration inhibition test. An 49 h-IC50 of 440 mg/L was obtained.

Description of key information

Regarding all available test on toxicity to microorganisms, the most relevant test resulting in the lowest PNEC value is used for assessment.


The sensitivity of activated sludge to m-cresol was determined by a respiration inhibition test according to OECD Guideline 209 yielding an 3h- IC50 of 462 mg/L. In a non-guideline study the effect of m-cresol on activated sludge was determined by a respiration inhibition test resulting in an 49 h-IC50 of 440 mg/L. Nitrification inhibition by m-cresol was measured by a method similar to ISO/DIS 9509 yielding an EC75 of 11.4 mg/L during a 2-4 h incubation period. In a cell multiplication inhibition test conducted with Entosiphon sulcatum a 72h-NOEC of 31 mg/l is determined. For Pseudomonas putida a NOEC of 53 mg/l was obtained in a cell multiplication inhibition test during 16 hours of exposure. In order to protect the efficiency of sewage treatment plants, inhibition of nitrification is identified to be the most sensitive test system.

Key value for chemical safety assessment

EC50 for microorganisms:
11.4 mg/L

Additional information

Several tests are available to evaluate the toxicity of m-cresol to microorganisms.

The influence of m-cresol on the nitrification process in domestic wastewater is described in an experiment. A purified activated sludge was incubated for 2 to 4 hours to the wastewater (BSB5 = 250 mg/l, 50 to 80 mg NH4-N/l) fortified with different concentrations of m-cresol. The effect on the inhibition of the nitrification was determined by photometric analysis of nitrite and nitrate. An EC75-value of 11.4 mg/l is given for the inhibition of the reaction step of ammonia to nitrite.

Concerning toxicity towards single species, the toxicity of m-cresol to Pseudomonas putida was assessed in a cell multiplication inhibition test according to Bringmann. Bacteria were exposed for 16 hours at a temperature of 25°C to the test substance. A toxicity threshold concentration of 53 mg/l was determined after 16 hours exposure. The result is equivalent to an EC3 or a NOEC. The endpoint biomass is measured by turbidity determination with a photometric detector.

The protozoa Entosiphon sulcatum was exposed to m-cresol for 72 hours using the cell multiplication inhibition test method. A toxicity threshold of 31 mg/l was obtained which is equivalent to a NOEC or an EC3.

The sensitivity of activated sludge to m-cresol was determined by a respiration inhibition test according to OECD Guideline 209. The 3h IC50 for respiration inhibion of microorganisms is 462 mg/L. In another experiment, the effect of m-cresol on activated sludge was determined by a respiration inhibition test. An 49 h-IC50 of 440 mg/L was obtained.