3,5,8-Trioxa-4-phosphaundec-10-en-1-aminium, 4-hydroxy-N,N,N,10-tetramethyl-9-oxo-, inner salt, 4-oxide

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:

1 October to 8 November 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Commission Directive 92/69/EEC

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(Date of Inspection: 30 August 2005 Date of Signature: 21 November 2005)

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

417-560-0

EC Name:

-

Cas Number:

67881-98-5

Molecular formula:

C11H22NO6P

IUPAC Name:

trimethyl[2-({2-[(2-methylprop-2-enoyl)oxy]ethyl phosphonato}oxy)ethyl]azanium

Details on test material:

- Name of test material (as cited in study report): MPC
- Substance type: White powder
- Physical state: Solid
- Lot/batch No.: 270103
- Storage condition of test material: Approximately 4ºC in the dark.

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): A mixed population of activated sewage sludge micro-organisms was obtained on 8 October 2007 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, Unied Kingdom, which treats predominantly domestic sewage.

- Preparation of inoculum for exposure: The activated sewage sludge sample was washed three times by settlement and resuspension in culture medium to remove any excessive amounts of dissolved organic carbon (DOC) that may have been present. The washed sample was then maintained on continuous aeration in the laboratory at a temperature of approximately 21°C and used on the day of collection.
Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 ml) of the washed activated sewage sludge by suction through pre-weighed GF/A filter paper using a Buchner funnel. Filtration was then continued for a further 3 minutes after rinsing the funnel three successive times with 10 ml of deionised reverse osmosis water. The filter paper was then dried in an oven at approximately 105°C for at least 1 hour and allowed to cool before weighing. This process was repeated until a constant weight was attained.

- Concentration of sludge: The suspended solids concentration was equal to 3.7 g/l prior to use.

- Initial cell/biomass concentration: 30 mg suspended solids (ss)/l.

- Type and size of filter used, if any: Pre-weighed GF/A filter paper using a Buchner funnel.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

10 other: mg carbon/l

Based on:

test mat.

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium:
Solution a
KH2PO4 8.50 g/l
K2HPO4 21.75 g/l
Na2HPO4.2H2O 33.40 g/l
NH4Cl 0.50 g/l

pH = 7.4

Solution b
CaCl2 27.50 g/l

Solution c
MgSO4.7H2O 22.50 g/l

Solution d
FeCl3.6H2O 0.25 g/l

To 1 litre (final volume) of purified water (reverse osmosis purified and deionised water (Elga Optima 15+ for Elga Purelab Option R-15 BP) was added the following volumes of solutions a – d.

10 ml of Solution a
1 ml of Solution b
1 ml of Solution c
1 ml of Solution d

- Test temperature: Approximately 21ºC.

- pH: 7.0 - 7.2.

- pH adjusted: no

TEST SYSTEM
- Culturing apparatus: Five litre glass culture vessels each containing 3 litres of solution. The culture vessels were sealed and CO2-free air bubbled through the solution at a rate of approximately 40 ml/minute and stirred continuously by magnetic stirrer.
The CO2-free air was produced by passing compressed air through a glass column containing self-indicating soda lime (Carbosorb®) granules.

- Number of culture flasks/concentration: 2.

- Number of standard control: 2

- Method used to untreated conditions: 2.

- Measuring equipment: The CO2 produced by degradation was collected in two 500 ml Dreschel bottles containing 350 ml of 0.05 M NaOH. The CO2 absorbing solutions were prepared using purified de-gassed water.

- Test performed in : closed systems.

CONTROL AND BLANK SYSTEM
- Untreated control: Consisting of inoculated culture medium.

- Toxicity control: For the purposes of the test a toxicity control, containing the test material and sodium benzoate, was prepared in order to assess any toxic effect of the test material on the sewage sludge microorganisms used in the test.
An aliquot (224 ml) of the test material stock solution was dispersed in inoculated culture medium along with an aliquot (51.4 ml) of the sodium benzoate stock solution. The volume was adjusted to 3 litres to give a final concentration of 22.4 mg test material/l plus 17.1 mg sodium benzoate/l, equivalent to a total of 20 mg carbon/l.

- Other: Standard Material
Sodium benzoate (Sigma Lot No 095K0681), a final concentration of 17.1 mg/l, equivalent to 10 mg carbon/l.

Reference substance

Reference substance:

other: sodium benzoate

Results and discussion

Preliminary study:

The results obtained from the samples taken for DOC analysis from the preliminary investigational work indicated that the test material did not absorb to filter matrices or to activated sewage sludge. For the purpose of the study, the samples taken for DOC analysis were filtered to remove the suspended solids present without the loss of any test material.

Test performance:

The total CO2 evolution in the control vessesl on Day 28 was 26.13 mg/l.
The IC/TC ratio of the test material suspension in the mineral medium at the start of the test was below 5%.
The difference between the values for CO2 production at the end of the test for the replicate vessels was < 20%.
Above satisfied the validation criterion give in the OECD Test Guidelines.

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

> 100

Sampling time:

28 d

Details on results:

Points of degradation plot (test substance): 6 % degradation after 1 d 33 % degradation after 8 d 69 % degradation after 16 d 96 % degradation after 24 d 100 % degradation after 28 d

BOD5 / COD results

Results with reference substance:

Points of degradation plot (reference substance): 7 % degradation after 1 d 63 % degradation after 8 d 77 % degradation after 16 d 75 % degradation after 24 d 71 % degradation after 28 d

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

The test material attained 105% degradation after 28 days and satisfied the 10-Day window validation criterion, whereby 60% degradation must be attained within 10 days of the degradation exceeding 10%. The test material can therefore be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301B.

Executive summary:

Introduction.A study was performed to assess the ready biodegradability of the test material in an aerobic aqueous medium. The method followed that described in the OECD Guidelines for Testing of Chemicals (1992) No 301B, "Ready Biodegradability; CO2 Evolution Test" referenced as Method C.4-C of Commission Directive 92/69/EEC (which constitutes Annex V of Council Directive 67/548/EEC), and US EPA Fate, Transport, and Transformation Test Guidelines OPPTS 835.3110 Paragraph (m).

 

Methods.The test material, at a concentration of 10 mg Carbon/I, was exposed to activated sewage sludge micro-organisms with culture medium in sealed culture vessels in the dark at approximately 21°C for 28 days.

The degradation of the test material was assessed by the determination of carbon dioxide

produced. Control solutions with inoculum and the standard material, sodium benzoate, together with a toxicity control were used for validation purposes.

 

Results.The test material attained 105% degradation after 28 days and satisfied the 10-Day window validation criterion, whereby 60% degradation must be attained within 10 days of the degradation exceeding 10%. The test material can therefore be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No 301B.

Degradation values in excess of 100% were considered to be due to an increase in the numbers of viable micro-organisms in the test material vessels as a result of the readily biodegradable nature of the test material. This effect occurs due to the micro-organisms utilizing the test material as a carbon source for cellular growth resulting in a greater number of viable micro-organisms in these vessels when compared to the control vessels. This increased number of micro-organisms in these vessels gave rise to increased respiration rates and hence background CO2 evolution was greater than in the control vessels. This increase in background CO2 evolution resulted in biodegradation

rates in excess of 100%.