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

Biodegradation in soil

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Administrative data

Endpoint:

biodegradation in soil: simulation 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:

2006

Report date:

2006

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)

Version / remarks:

OECD Draft Document, April 2000

Deviations:

no

GLP compliance:

yes

Test type:

laboratory

Test material

Test material information

Constituent 1

Test material form:

not specified

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. 1063.0008

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: 96.9%
- Specific activity: 48.5 mCi/g
- Locations of the label: 14C label was at the 4 and 5 position

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Not reported
- Stability under test conditions: Half-life in aerobic water:sediment simulation studies range from 0.38 – 1.4 days
- Solubility and stability of the test substance in the solvent/vehicle: Water solubility (deionized water) > 1000 ppm
- 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: Dissolution in water
- Final dilution of a dissolved solid, stock liquid or gel: 0.966 ppm 14C-MIT

FORM AS APPLIED IN THE TEST (if different from that of starting material)
Liquid

Radiolabelling:

yes

Study design

Oxygen conditions:

aerobic

Soil classification:

USDA (US Department of Agriculture)

Year:

2006

Soil properties

Soil no.:

#1

Soil type:

silt loam

% Clay:

26

% Silt:

54

% Sand:

20

pH:

6.3

CEC:

9.6 meq/100 g soil d.w.

Bulk density (g/cm³):

0.99

Details on soil characteristics:

SOIL COLLECTION AND STORAGE
- Geographic location: Spring House, Pennsylvania, USA
- Sampling depth (cm): 20
- Soil preparation (e.g., 2 mm sieved; air dried etc.): The soil was sieved through a 2 mm sieve and preconditioned at 20 ± 2°C prior to use

PROPERTIES OF THE SOILS (in addition to defined fields)
- Moisture at 1/3 atm (%): 25.6
- Bulk density (g/cm3): 0.99

Duration of test (contact time)

Soil No.:

#1

Duration:

<= 100 d

Initial test substance concentration

Soil No.:

#1

Initial conc.:

0.966 ppm

Based on:

test mat.

Parameter followed for biodegradation estimation:

radiochem. meas.

Experimental conditions

Soil No.:

#1

Temp.:

20.1 ± 0.4°C

Humidity:

25.6% at 1/1 Bar

Microbial biomass:

6 x 10e6 cfu/g

Details on experimental conditions:

1. PRELIMINARY EXPERIMENTS:
A total of 28 glass bottles at 250ml were prepared by adding 50 g (dwb) of soil to each. Twelve bottles were dosed at 1 ppm 14C MIT, 12 at 5 ppm, and 4 at 10 ppm. The bottles were sealed with 2-hole stoppers containing glass tubes. The glass tubes of the bottles in series were connected by plastic tubing allowing a vacuum flow through the system which was maintained in a dark incubator at 20 ± 2°C. The flow through system consisted of a bottle with water at the vacuum inlet followed by the bottles containing soil and terminating with volatile traps containing ethylene glycol 1.0N KOH closest to the vacuum source. Samples were taken at 0, 3, 24, 48, and 120 hours for the 1 ppm and 5 ppm dosed samples and at 24 and 120 hours for the 10 ppm dosed samples.

2. EXPERIMENTAL DESIGN
- Soil preincubation conditions (duration, temperature if applicable): 20 ± 2°C
- Soil condition: fresh
- Soil (g/replicate): 50
- Control conditions, if used (present differences from other treatments, i.e., sterile/non-sterile, experimental conditions): Sterilized soil
- No. of replication controls, if used: 2
- No. of replication treatments: 2
- Test apparatus (Type/material/volume): 250ml glass bottles
- Details of traps for CO2 and organic volatile, if any: 50 ml of ethylene glycol (trap for volatile organic compounds) and 50 ml of 1.0N KOH (trap for CO2).
- Identity and concentration of co-solvent: Water

Test material application
- Volume of test solution used/treatment: 102 μl
- Application method (e.g. applied on surface, homogeneous mixing etc.): Applied on surface
- Is the co-solvent evaporated: Not applicable

Any indication of the test material adsorbing to the walls of the test apparatus: No

Experimental conditions (in addition to defined fields)
- Moisture maintenance method: Air flow
- Continuous darkness: Yes/No

3. OXYGEN CONDITIONS (delete elements as appropriate)
- Methods used to create the an/aerobic conditions: Air flow

4. SUPPLEMENTARY EXPERIMENTS:
A second set of soils were dosed at 1, 10, and 25 ppm MIT containing a 1:1 mixture of 14C:13C-MIT and incubated 3 to 120 hours to assist in metabolite identification.

5. SAMPLING DETAILS
- Sampling intervals: Hours 0, 2, 5, 22, 48, and 120 and Days 10, 30, and 100
- Sampling method for soil samples: Whole soil sample was extracted using solvents (methanol:acetonitrile and methanol:KOH )
- Method of collection of CO2 and volatile organic compounds: The presence of 14CO2 in the KOH traps was confirmed by adding BaCl2 to aliquots of the trap solution and the resulting BaCO3 radioassayed
- Sampling intervals/times for:
> Sterility check, if sterile controls are used: Not reported
> Moisture content: At each sample interval
> Redox potential/other: Not reported
> Sample storage before analysis: Not reported

Results and discussion

Material (mass) balance

Soil No.:

#1

% Recovery:

100.6

St. dev.:

2.2

% Degradation

% Degradation 1

Soil No.:

#1

% Degr.:

16.8

Parameter:

radiochem. meas.

Sampling time:

0 h

% Degradation 2

Soil No.:

#1

% Degr.:

46.5

Parameter:

radiochem. meas.

Sampling time:

2 h

% Degradation 3

Soil No.:

#1

% Degr.:

92.7

Parameter:

radiochem. meas.

Sampling time:

22 h

% Degradation 4

Soil No.:

#1

% Degr.:

95.1

Parameter:

radiochem. meas.

Sampling time:

48 h

% Degradation 5

Soil No.:

#1

% Degr.:

96.1

Parameter:

radiochem. meas.

Sampling time:

120 h

% Degradation 6

Soil No.:

#1

% Degr.:

93.2

Parameter:

radiochem. meas.

Sampling time:

10 d

% Degradation 7

Soil No.:

#1

% Degr.:

98.1

Parameter:

radiochem. meas.

Sampling time:

30 d

% Degradation 8

Soil No.:

#1

% Degr.:

99.3

Parameter:

radiochem. meas.

Sampling time:

100 d

Half-life / dissipation time of parent compound

Half-life / dissipation time of parent compound 1

Key result

Soil No.:

#1

DT50:

21.7 h

Type:

(pseudo-)first order (= half-life)

Temp.:

20 °C

Remarks on result:

other: 0-22 h

Half-life / dissipation time of parent compound 2

Soil No.:

#1

DT50:

1 645 h

Type:

second order

Temp.:

20 °C

Remarks on result:

other: 0-100 d

Transformation products:

yes

Identity of transformation products

Identity of transformation products 1

No.:

#1

Identity of transformation products 2

No.:

#2

Details on transformation products:

- Formation and decline of each transformation product during test: The two main metabolites were present at concentrations greater than 21% at Hour 22 but were transient decreasing to less than 1.3% by Day 100
- Pathways for transformation: Cleavage of the isothiazolone ring with the ultimate metabolite being CO2

Evaporation of parent compound:

no

Volatile metabolites:

yes

Remarks:

Only CO2

Residues:

yes

Details on results:

TEST CONDITIONS
- Aerobicity (or anaerobicity), moisture, temperature and other experimental conditions maintained throughout the study: Yes

MAJOR TRANSFORMATION PRODUCTS
- Range of maximum concentrations in % of the applied amount and day(s) of incubation when observed:
- Range of maximum concentrations in % of the applied amount at end of study period:
on the - the and -th day of incubation, respectively. At the end of the study period, the corresponding concentrations were - and -- % of the applied amount, respectively.

EXTRACTABLE RESIDUES
- % of applied amount at day 0: 93.7
- % of applied amount at end of study period: 13.3

NON-EXTRACTABLE RESIDUES
- % of applied amount at day 0: 0 6.2
- % of applied amount at end of study period: 38.8

MINERALISATION
- % of applied radioactivity present as CO2 at end of study: 46.6

VOLATILIZATION
- % of the applied radioactivity present as volatile organics at end of study: 0

STERILE TREATMENTS (if used)
- Transformation of the parent compound: Not described
- Formation of transformation products: No
- Formation of extractable and non-extractable residues: No
- Volatilization: 0.2% CO2

Applicant's summary and conclusion

Conclusions:

MIT rapidly biodegrades in soil with a half-life of 6.5 hours in a silt loam soil incubated at 20.1 ± 0.4°C. Metabolism involves cleavage of the isothiazolone ring with the ultimate metabolite being CO2.

Executive summary:

The test guideline was OECD Draft Document for a New Guideline 307: Aerobic and Anaerobic Transformation in Soil, April 2000. The half-life based on the data from sampling intervals 0- 22 hours is 6.5 hours. Thus, MIT is rapidly biodegraded in soil. The total extractable activity decreased rapidly from 93.7% of the applied activity at Hour 0 to 66.6% at Hour 48 and 13.3% at Day 100. Simultaneous evolved 14CO2 increased from 1.8% of applied activity at Hour 2 to 46.6% on Day 100. Bound residue (post extraction solids) increased from 6.2% of the applied activity at Hour 0 to 39.7% on Day 30. Recovery averaged 100.6 ± 2.2% The decrease in extractable activity and increase in evolved14CO2observed correlated with the decreasing amount of MIT with time. MIT decreased from 83.2% of applied activity at Hour 0 to 37.9% at Hour 5 and 0.7% on Day 100. Five metabolites plus CO2 were detected. Besides CO2, two metabolites, M3 and M4 were predominant; 2-(methylcarbamoyl)-ethene sulfonic acid and 2-(methylcarbamoyl)-1-oxo-ethane sulfinic acid (current data suggests that these are actually the cis and trans isomers of 2-(methylcarbamoyl)-ethene sulfonic acid). M3 and M4 were present at greater than 21% at Hour 22 but were transient decreasing to less than 1.3% by Day 100. M1, M2, and M5 comprised multiple compounds none exceeding 5%. Evolved14CO2consistently increased throughout the study so that it becomes the predominate metabolites (46.6% on Day 100). This indicates that MIT, the metabolites, and the bound residue were biodegrading to CO2.