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Toxicity to microorganisms

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Reference
Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication which meets basic scientific principles
Qualifier:
no guideline followed
Principles of method if other than guideline:
TETRATOX assay: Tetrahymena pyriformis population growth impairment test
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
not specified
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- stock solutions were prepared by mixing test substance with sterile distilled water or with reagent-grade dimethyl sulfoxide (DMSO) just prior to use (within 1 hour). Concentration of DMSO in the test medium did not exceed 0.75% (max. 375 µL stock solution per 50 mL of medium). This concentration had no effect on the growth of the test organisms.
Test organisms (species):
Tetrahymena pyriformis
Details on inoculum:
- Laboratory culture: yes
- Test organism: Tetrahymena pyriformis (strain GL-c) in log-growth-phase
- Initial biomass concentration: 2500 cells/mL (i.e. 0.20 mL of a 48-h culture)
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
40 h
Remarks on exposure duration:
test duration allowes for 8 to 9 cell cycles in controls.
Test temperature:
27 ± 1°C
Nominal and measured concentrations:
nominal concentrations; at least five graduate concentration levels, individual concentrations not reported
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): Erlenmeyer flasks, foam-stoppered
- Material, size, headspace, fill volume: glas, 250 mL, 150 mL, 50 mL
- Aeration: no data
- No. of organisms per vessel: 1.25 E5 (2500 cell/mL) in log-growth phase
- No. of vessels per concentration (replicates): 2 + 1 blank
- No. of tests per substance: 3 replicate independent tests

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: sterile distilled water
- Culture medium different from test medium: no
- Test medium: Proteose peptone (5 g), D-glucose (5 g), Yeast extract (1 g,) Tris_HCl (1.2114 g) in 1000 mL distilled Water supplemented with 100 mL each of two salt solutions (Chlorides and Sulfates). Medium is sterilized.

OTHER TEST CONDITIONS
- Adjustment of pH: test solution not neutralized

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
- Growth impairment: population density is measured spectrophotometrically (absorbance at 540 nm)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: no data
- Range finding study: preliminary to the definite test (3 independent replicate assays) a range finding assay was performed (no further data reported)

STATISTICAL EVALUATION
- results are given as 50 % inhibitory growth concentration in mg/L (IGC50)
- IGC50 values and the 95% fiducial intervals are determined by Probit analysis using the percent control-normalized absorbance as the dependent variable and the toxicant concentration in mg/L as the independent variable
Reference substance (positive control):
not specified
Duration:
40 h
Dose descriptor:
IC50
Effect conc.:
190 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Remarks on result:
other: reported value: Log (ICG50)E-1 = -0.3334
Reported statistics and error estimates:
no statistics reported
Validity criteria fulfilled:
not applicable
Remarks:
no guideline study
Conclusions:
The concentration of isobutyric acid to inhibit the growth of Tetrahymena pyriformis by 50 % (spectrophotometric measurement of cell density) was determined to be 2.15 mg/L.
Executive summary:

In a valid 40 hour growth inhibition study, test cultures of the cilliate Tetrahymena pyriformis were exposed to isobutyric acid at five graduate concentrations under static conditions (individual concentrations not reported). The assay was performed in three independent replicate runs on the basis of a preceding range finding test. The IC50 value based on cell density was determined to be 2.15 mg/L indicating low to moderate toxicity of isobutyric acid to aqueous microorganisms (cilliates) (Schultz, 2006).

Description of key information

Isobutyric acid inhibited the cell growth of Tetrahyma pyriformis test cultures by 50 %  at a concentration of 2.15 mg/mL.

Key value for chemical safety assessment

EC50 for microorganisms:
2.15 mg/L

Additional information

From three tests identified, only one test is valid (Schulz, 1997). The documentation of the other two (BASF AG, 1989; Hoechst AG 1979) is insufficient for assessment.

Schulz 1997

In a valid 40 hour growth inhibition study, test cultures of the cilliate Tetrahymena pyriformis were exposed to isobutyric acid at five graduate concentrations under static conditions (individual concentrations not reported). The assay was performed in three independent replicate runs on the basis of a preceding range finding test.

The IC50 value based on cell density was determined to be 2.15 mg/L indicating low to moderate toxicity of isobutyric acid to aqueous microorganisms (cilliates) (Schultz, 2006).