N-nitro-N-(3-methyl-3,6-dihydro-2H-1,3,5-oxadiazin-4-yl)amine

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1996-01-17 until 1996-04-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline-conform study under GLP without deviations.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

431-060-1

EC Name:

-

Cas Number:

153719-38-1

Molecular formula:

Hill formula: C4H8N4O3 CAS formula: C4H8N4O3

IUPAC Name:

3-methyl-N-nitro-3,6-dihydro-2H-1,3,5-oxadiazin-4-amine

Details on test material:

- Name of test material (as cited in study report): CA 2343 A (Intermediate of CGA 293343)
- Substance type: mono-constituent organic substance
- Physical state: solid, fine, white powder
- Analytical purity: 96.7 %
- Impurities (identity and concentrations): no data
- Purity test date: no data
- Lot/batch No.: P.503005
- Expiration date of the lot/batch: Date of reanalysis: December 1996
- Stability under test conditions: not reported
- Storage condition of test material: 0 - 5 °C

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
not applicable

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge (adaptation not specified)

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Activated sludge collected from the sewage treatment plant of CH-4153 Reinach on 22/01/96. No details on sampling procedure and contamination history of the facility reported.
- Laboratory culture: Before application, the inoculum was pre-acclimated to the test medium over night. The aeration was performed inside 1.5 l water, containing the amount of mineral salts for 1.5 l.
- Method of cultivation: Before application, the inoculum was pre-acclimated to the test medium over night. The aeration was performed inside 1.5 l water, containing the amount of mineral salts for 1.5 l.
- Storage conditions: Before application, the inoculum was pre-acclimated to the test medium over night. The aeration was performed inside 1.5 l water, containing the amount of mineral salts for 1.5 l.
- Storage length: Before application, the inoculum was pre-acclimated to the test medium over night. The aeration was performed inside 1.5 l water, containing the amount of mineral salts for 1.5 l.
- Preparation of inoculum for exposure: Before application, the inoculum was pre-acclimated to the test medium over night. The aeration was performed inside 1.5 l water, containing the amount of mineral salts for 1.5 l.
- Pretreatment: Acc. to the study reported "The preparation (of the sludge) was carried out according to the method described in the guideline."
- Concentration of sludge: The concentration of the inoculum in the test was 25.2 mg sludge/l.
- Initial cell/biomass concentration: not reported
- Water filtered: no
- Type and size of filter used, if any: not applicable

Duration of test (contact time):

29 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

16.2 mg/L

Based on:

DOC

Initial test substance concentration 2

Initial conc.:

16.1 mg/L

Based on:

DOC

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: The reconstituted water was prepared according to the method described in the guideline.
- Additional substrate: not applicable
- Solubilising agent (type and concentration if used): not applicable
- Test temperature: 21 ± 2°C
- pH: 7.8
- pH adjusted: no
- CEC (meq/100 g): no data
- Aeration of dilution water: The reconstituted water was prepared according to the method described in the guideline.
- Suspended solids concentration: 2.5 g/l
- Continuous darkness: yes
- Other:


TEST SYSTEM
- Culturing apparatus:
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: stirrer
- Method used to create anaerobic conditions: not applicable
- Measuring equipment: CO2-Scrubbers
- Test performed in closed vessels due to significant volatility of test substance: not applicable
- Test performed in open system: not applicable
- Details of trap for CO2 and volatile organics if used: : Approx 250 ml scrubbers with gas inlet tubes equipped with sintered glass tips containing 200 ml 0.05 N sodium hydroxide. Only one C02 scrubber was used per test bottle. Theoretically, a solution of 0.05 M NaOH is sufficient to trap at least two times more C02 than the maximum ThC02 which can evolve in each test bottle.


SAMPLING
- Sampling frequency: C02 trapped by the 0.05 N NaOH solution of the blank and test substance was measured at 0,3, 6, 8, 10, 13, 15, 17, 20,23,28 and 29 days.
- Sampling method: not reported
- Sterility check if applicable: not applicable
- Sample storage before analysis: not reported
- Other:


CONTROL AND BLANK SYSTEM
- Inoculum blank: test medium and inoculum
- Abiotic sterile control: not applicable
- Toxicity control: reference substance. test substance and inoculum
- Other: procedure control: reference substance and inoculum


STATISTICAL METHODS:

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

Preliminary study:

not applicable

Test performance:

- test substance and inoculum: very limited biodegradation of the test substance after 29 days
- reference substance and inoculum: 69% biodegradation after 29 days
- test substance and reference substance and inoculum: no inhibition observed

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

3

Sampling time:

29 d

Details on results:

Points of degradation plot (test substance):
0 % degradation after 3 d
0 % degradation after 6 d
0 % degradation after 8 d
0 % degradation after 10 d
0 % degradation after 13 d
0 % degradation after 15 d
0 % degradation after 17 d
0 % degradation after 20 d
0 % degradation after 23 d
0 % degradation after 28 d
3 % degradation after 29 d

BOD5 / COD results

Results with reference substance:

Points of degradation plot (reference substance):
45 % degradation after 3 d
57 % degradation after 6 d
60 % degradation after 8 d
60 % degradation after 10 d
65 % degradation after 13 d
64 % degradation after 15 d
65 % degradation after 17 d
68 % degradation after 20 d
67 % degradation after 23 d
69 % degradation after 28 d
69 % degradation after 29 d

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

The biodegradation of CA 2343 A was 3 % in 29 days. Based on the results of this test and using the classification range given in OECD-GHS and directive 67/548EEC, the ecotoxicological classification is " not readily biodegradable"

Executive summary:

The purpose of the study was the determination of the ready biodegradability of test substance oxadiazinamine (CA 2343 A) by measurement of the carbon dioxide formation in per cent of ThCO2 (Theoretical carbon dioxide) calculated from the ThOC (Theoretical organic carbon). The method acc. to OECD guideline 301B was applied. As inoculum activated sludge from a communal wastewater treatment facility in Switzerland was being used.

The biodegradation of CA 2343 A was 3 % in 29 days. Based on the results of this test and using the classification range given in OECD-GHS and directive 67/548EEC, the ecotoxicological classification is " not readily biodegradable"