reaction mass of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-2H -isothiazol-3-one

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1995

Report date:

1995

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Test material form:

other: orange-yellow liquid

Details on test material:

- Name of test material (as cited in study report): Kathon™ WT

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Rohm and Haas, Batch No. J 97030

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Stability under test conditions: Stable at room temperature
- Solubility and stability of the test substance in the solvent/vehicle: Soluble and stable
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Dilution in water
- Final dilution of a dissolved solid, stock liquid or gel: 0.0010, 0.010, 0.10, 1.0 and 10 mg whole material/L

OTHER SPECIFICS: Purity of test material was 14%

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Dispersion
- Controls: Negative control and positive control (3,5-dichlorophenol)
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No

Test organisms

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Name and location of sewage treatment plant where inoculum was collected: Municipal wastewater treatment plant in Newburyport, Massachusetts, USA which treats predominantly domestic waste.
- Method of cultivation: Not applicable
- Preparation of inoculum for exposure: Aeration
- Pretreatment: Incubated at room temperature
- Initial biomass concentration: Total suspended solids = 9.7 g/L on day of collection and 4.2 g/L on the day of the test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

10 min

Test conditions

Hardness:

Not described

Test temperature:

20 +/- 2°C

pH:

7.9 at test initiation

Dissolved oxygen:

7.9-9.6 mg/L

Salinity:

Not applicable

Conductivity:

Not described

Nominal and measured concentrations:

Nominal: 0 (control), 6.5, 13, 25, 50, and 100 mg/L test substance

Details on test conditions:

TEST SYSTEM
- Test vessel: Static
- Material, size, headspace, fill volume: 1 L glass beaker, 500 mL headspace
- Aeration: 0.5 to 1.0 L air/min
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- Sludge concentration (weight of dry solids per volume): 4.2 g/L
- Nutrients provided for bacteria: 16 mL synthetic sewage per vessel
- Nitrification inhibitor used (delete if not applicable): none

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Synthetic sludge

OTHER TEST CONDITIONS
- Adjustment of pH: adjusted to 7.5 with 0.1 N HCl
- Photoperiod: all light
- Light intensity: ambient lighting, 5 µEin/m^2s

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Dissolved oxygen measurement for first 10 minutes that sludge was in the BOD bottle.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0.0010, 0.010, 0.10, 1.0, and 10 mg/L
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

4.5 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Effect concentrations 2

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

0.91 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

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: No
- Adsorption (e.g. of test material to the walls of the test container): Not described
- Blank controls oxygen uptake rate: 60 mg/L/h
- Coefficient of variation of oxygen uptake rate in control replicates: Not described

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: EC50 = 9.6 mg/L

Reported statistics and error estimates:

The respiration rate was calculated as the mg oxygen/liter/hour consumed between 6.5 and 2.5 mg/L or over a 10 minute respiration period if the respiration rate was low. The percent inhibition, which was used to calculate EC50 values, was calculated as 1 minus ([2 times the respiration rate of the TS concentration] divided by the sum of the 2 control respiration rates) times 100.
The probit method was used to calculate the reference toxicant 3-h EC50 using nominal concentrations (Stephan, 1983, Computer Program for Calculation of LC50 values, personal communication).

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Based on the results, the 3h EC50 for CMIT/MIT based on inhibition of respiration was determined to be 4.5 mg/L. The 3h NOEC was 0.91 mg/L.

Executive summary:

Exposure of activated sludge to Kathon(TM) WT 14% resulted in a 3 hour EC50 of 4.5 mg/L with a 95% confidence interval of 3.9 to 5.2 mg/L, on an active ingredient basis. The 3 hour NOEC is 0.91 mg/L on an active ingredient basis. Both parameters are based on inhibition of respiration after a 3 h exposure to the test material.