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Toxicity to microorganisms

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Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
other information
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
This study is the only reliable experiment on bacterial population. It is well described in terms of experimental conditions. The only restriction concerns the description of the test item which is not fully detailed.
Qualifier:
no guideline followed
Principles of method if other than guideline:
Methanogenic activity inhibition test: evaluation of the toxicity to unacclimated methanogenic sludge.
GLP compliance:
not specified
Analytical monitoring:
yes
Details on sampling:
No data
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Granular sludge (1 g of VSS) was transferred to the serum vials containing 100 mL of the basal medium and acetate was added from concentrated stock solution to obtain a final concentration of 2 g COD/L (readjusted by addition of acetate). The liquid was flushed with nitrogen gas and the flasks were sealed with a rubber septum and a screw cap and placed in reciprocating shaker at 30°C. Catechol was added to each flask to provide toxic concentrations to be investigated.
- Controls: yes (without toxicant).
Test organisms (species):
anaerobic bacteria
Details on inoculum:
- Preparation of inoculum for exposure: Elutriated sludge from a full scale UASB reactor treating distillery wastewater was used as inoculum.
The granular sludge was stored at 4°C and pre-acclimated by incubation at 30°C, in the presence of VFA. The sludge used was not previously acclimated to aromatic compounds.
No more data
Test type:
not specified
Water media type:
freshwater
Total exposure duration:
3 d
Nominal and measured concentrations:
No data
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 0.315 L glass serum flasks
No more data

TEST MEDIUM / WATER PARAMETERS
- source preparation of dilution water: Basal medium consisting (in mg/L): NaHCO3 (400), NH4Cl (280), CaCl2 2H2O (10), K2HPO4 (250), MgSO4 7H2O (100), yeast extract (100). Per liter of medium, 1 ml of a trace element solution according to Zenhder et al (1980) was added.
- Total organic carbon: 2 g COD/L
No more data

EFFECT PARAMETERS MEASURED:
Specific methanogenic activity measurement was performed. After flushing the headspace with nitrogen gas, the flasks were again incubated for 1 hour prior to the determination of the methane production rate. The methane composition in the headspace content of each serum flasks was determined periodically during the 4 to 5 hour period that followed by gas chromatography. The compound concentration that caused 50% and 80% inhibition of the methanogenic activity was determined by graphic method.

TEST CONCENTRATIONS: the toxicant concentrations were chosen as to cause an inhibition of the acetoclastic activity ranging from 0 to 100%. The required amount of inhibitory compound was added to each flask to provide the toxic concentration to be investigated. No toxicant was added to the controls.
Duration:
3 d
Dose descriptor:
IC50
Effect conc.:
1 814 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: methanogenic activity
Duration:
3 d
Dose descriptor:
other: IC80
Effect conc.:
2 715 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: methanogenic activity
Reported statistics and error estimates:
The specific acetoclastic activity was calculated from the slope of the methane production versus time curve and the quantity of VSS (volatile suspended solid).
Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
The study is well described. The only restriction is that it is not known whether the purity of catechol was maximal (>= 95%).
Qualifier:
no guideline followed
Principles of method if other than guideline:
Toxicity to Tetrahymena pyriformis according to the Tetratox protocol.
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
yes
Details on test solutions:
- Method: dissolution of the substance in the test medium with vehicle
- Controls: Test medium + Inoculum
- Chemical name of vehicle: Solvent dimethyl sulfoxide (DMSO)
- Concentration of vehicle in test medium: Stock solution : < 0.75%
Test organisms (species):
Tetrahymena pyriformis
Details on inoculum:
- Laboratory culture: yes, strain GL-C
- Method of cultivation: growth medium mainly composed with proteose peptone (20 g/L), D-glucose (5 g/L), yeast extract (1 g/L) and Fe EDTA (1 ml of a 3% solution (w/v) solution) in 1000 ml distilled water. The experiments were conducted with log phase cultures.
- Preparation of inoculum for exposure: toxicity tests were conducted in 250-ml Erlenmeyer flasks containing 50 ml of medium (described above) and inoculated with 0.25 ml of two-day-old log phase culture.
- Initial biomass concentration: approximately 36000 cells per ml
Test type:
static
Water media type:
freshwater
Total exposure duration:
48 h
Hardness:
No data
Test temperature:
27°C +/- 1°C
pH:
7.35
Dissolved oxygen:
No data
Salinity:
Not applicable
Nominal and measured concentrations:
5 different concentrations. No more data.
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 ml glass Erlenmeyer flasks, sealed with
- Three replicate tests were performed with catechol. A minimum of 5 different concentrations of catechol was used with duplicate flasks of each concentration. 
Duplicate controls, which have no test material but were inoculated with T. pyriformis strain GL-C, and a blank were used.

EFFECT PARAMETERS MEASURED
The effect parameter measured was the impairment of population growth, using spectrophotometric analyses at 540 nm.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: No data
- Range finding study: Yes, at least 2 tests
Key result
Duration:
48 h
Dose descriptor:
IC50
Effect conc.:
19.58 mg/L
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Details on results:
In the publication, the result corresponds to the log of the inverse of the IC50:
log (1/IC50) = 0.75 mmol/L

Below is the reasoning used to calculate IC50 from this result:
log (1/IC50) = log (1) - log (IC50)
log (1) - log (IC50) = 0.75
0-log (IC50) = 0.75
log (IC50) = -0.75
IC50 = exp (ln(10) x (-0.75))
IC50 = 0.1778 mmol/L

To obtain an IC50 value expressed in g/L, the IC50 value expressed in mol/L has to be multiplied by the molecular weight of catechol:
IC50 (g/L) = IC50 (mol/L) x Molecular Weight (g/mol)
IC50 = 0.0001778 x 110.112 = 0.01958 g/L
IC50 = 19.58 mg/L
Reported statistics and error estimates:
The 50% inhibitory growth concentration was calculated with the probit procedure.
Validity criteria fulfilled:
not specified
Conclusions:
Catechol is harmful to Tetrahymena pyriformis under the tested conditions.
Executive summary:

In a 48-hour toxicity study (Schultz et al. 1990), a two-day-old log-phase culture of Tetrahymena pyriformis was exposed to five different concentrations of catechol. The effect parameter measured was the impairment of population growth, using spectrophotometric analyses at 540 nm.

On this basis, the 48h-EC50 was determined to be 19.58 mg/L.

As a consequence, catechol is harmful to Tetrahymena pyriformis under the tested conditions.

Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
The study is well described, and the results are in the same range (i.e. 10 to 100 mg/L) than the previous reliable study. The only restriction is that it is not known whether the purity of catechol was maximal (>= 95%).
Principles of method if other than guideline:
Toxicity toTetrahymena pyriformis according to Tetratox protocol.
GLP compliance:
not specified
Analytical monitoring:
no
Vehicle:
yes
Details on test solutions:
Stock solutions were prepared in dimethylsulfoxide (DMSO) at concentrations of 5, 10, 25 or 50 mg/L. In all cases, the volume of stock solution added to each flask was not in excess of 0.75% DMSO. This concentration does not alter Tetrahymena population growth.
Test organisms (species):
Tetrahymena pyriformis
Details on inoculum:
Each flask was inoculated with log-phase culture of T. pyriformis to attain a starting concentration of 2500 cells/mL.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
No data
Test temperature:
27 +/- 1°C
pH:
7.35
Dissolved oxygen:
No data
Salinity:
Not applicable
Nominal and measured concentrations:
5, 10, 25 or 50 mg/L
Details on test conditions:
- Population growth impairment assays:
This assay uses population density measured spectrophotometrically at 540 nm as its endpoint. Tests were conducted in 250-mL Erlenmeyer flasks containing 50 mL of sterile medium.
Each chemical was tested in a range-finder, followed by testing in duplicate with three or more replicates. Each replicate was a 5 to 10-step concentration series using freshly prepared stock solutions.
Only replicates with control values in late log-growth-phase (absorbance from 0.6 to 0.9) were used in the analyses.

Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
57.14 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Remarks on result:
other: 95% CL: 39.81 - 88.37
Validity criteria fulfilled:
not specified
Conclusions:
Catechol is harmful to Tetrahymena pyriformis under the tested concentrations.
Executive summary:

In a 48-hour toxicity study (Schultz et al. 1996), log-phase culture of Tetrahymena pyriformis was exposed to catechol at nominal concentrations of 5, 10, 25 or 50 mg/L under static conditions. This assay used population density, measured spectrophotometrically at 540 nm, as its endpoint.

On this basis, the 48h-EC50 was determined to be 57.14 mg/L.

As a consequence, catechol is harmful for Tetrahymena pyriformis under the tested concentrations.

Description of key information

Several studied  were available. After assessment, the results retained were the following:
48h-EC50 values (Tetrahymena pyriformis) were 57.14 mg/L and 19.58 mg/L.
Based on these studies, Catechol could be considered as harmful to some tested microorganisms.

Key value for chemical safety assessment

EC50 for microorganisms:
19.58 mg/L

Additional information

Most data were available on isolated species. Two of these studies were scored as reliability 2 according to Klimisch and selected as key studies. Based on growth inhibition of Tetrahymena pyriformis, 48h-EC50 values of 57.14 mg/L (Schultz, 1996) and 19.58 mg/L (Schultz, 1990) were reported. In view of the chemical safety assessment, the lowest value was retained.

The only reliable data available on mixed inoculum concerned anaerobic sludge, and was of reliability 2. A 3d-EC50 = 1814 mg/L, based on methane production inhibition, was reported in this study (Sierra-Alvarez, 1989).

The other available data contain few details. They were performed using species which are either not among those recommended by the guidelines, or considered of a low relevance when assessing the risk for micro-organisms from STP. Furthermore, some of the methods applied are not enough standardised to conclude on the toxicity of catechol to micro-organisms.

 

Based on these results, Catechol could be harmful to some of the tested microorganisms.