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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
March - May 2008
Reliability:
1 (reliable without restriction)
Justification for type of information:
test item is unstable in water, therefore the hydrolysis product PMA was tested
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
- Description: white powder
- Batch number: 8M30N
- Storage conditions: room temperature in the dark upto 04 April 2008, thereafter at approximately 4 °C, over silica gel, in the dark
Oxygen conditions:
aerobic
Inoculum or test system:
sewage, predominantly domestic (adaptation not specified)
Details on inoculum:
- Source of inoculum/activated sludge: Severn Trent Water PLC sewage treatment plant at Loughborough, Leicestershire, UK (delivery: 07 April 2008)
- Treatment: sample was washed three times by settlement and resuspension in culture medium
- Storage conditions: washed sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21°C
- Storage length: used on the day of collection
- Concentration of sludge: suspended solids concentration was equal to 2.3 g/1
- Water filtered: yes
- Type and size of filter used: GF/A filter paper
Duration of test (contact time):
28 d
Initial conc.:
10 mg/L
Based on:
DOC
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: as recommended in OECD-guidelines
- Solubilising agent (type and concentration if used): no
- Test temperature: 21°C
- pH: 7.6
- pH adjusted: no
- Suspended solids concentration: 30 mg suspended solids ss/l
- Continuous darkness: yes

TEST SYSTEM
The following test preparations were prepared and inoculated in 5 litre glass culture vessels each containing 3 litres of solution:
a) A control, in duplicate, consisting of inoculated culture medium.
b) The standard material (sodium benzoate), in duplicate, in inoculated culture medium to give a final concentration of 10 mg carbon/l
c) The test material, in duplicate, in inoculated culture medium to give a final concentration of 10 mg carbon/l
d) The test material plus the standard material in inoculated culture medium to give a final concentration of 20 mg carbon/l to act as a toxicity control (one vessel only).

SAMPLING
Samples (2 ml) were taken from the first C02 absorber vessel on Days 0, I, 2, 3, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24,28 and 29. The second absorber vessel was sampled on Days 0 and 29. The samples taken on Days 0, 1, 2, 3, 6, 8, 10, 14, 16, 20, 22, 24, 28 and 29 were analysed for CO2 immediately. The samples taken on Days 12 and 18 were stored at approximately -20°C. However, these samples were not analysed for CO2 as the results obtained from previous and subsequent analyses showed that degradation of the test material had met the 10-day window validation criterion given in the OECD Guidelines and therefore additional analyses were considered to be unnecessary.
On Day 28, 1 ml of concentrated hydrochloric acid was added to each vessel to drive off any inorganic carbonates formed. The vessels were resealed, aerated overnight and the final samples taken from both absorber vessels on Day 29. The samples were analysed for CO2 using a Tekmar-Dohrmann Apollo 9000 TOC analyser and a Shimadzu TOC-V csH TOC analyser. Samples (300 or 50 ul) were injected into the IC (Inorganic Carbon) channel of the TOC analyser. Inorganic carbon analysis occurs by means of the conversion of an aqueous sample to CO2 by orthophosphoric acid using zero grade air as the carrier gas. Calibration was by standard solutions of sodium carbonate (Na2C03). Each analysis was carried out in triplicate.
Reference substance:
benzoic acid, sodium salt
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. Therefore, 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:
Each test vessel was inoculated with the prepared inoculum at a final concentration of 30 mg suspended solids (ss)/l. The test was carried out in a temperature controlled room at approximately 21 °C, in darkness. Approximately 24 hours prior to addition of the test and standard materials the vessels were filled with 2400 ml of culture medium and 39.1 ml of inoculum and aerated overnight. On Day 0 the test and standard materials were added and the volume in all the vessels adjusted to 3 litres by the addition of culture medium.
The culture vessels were sealed and CO-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). 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.
Parameter:
% degradation (CO2 evolution)
Value:
0
Sampling time:
0 d
Parameter:
% degradation (CO2 evolution)
Value:
19
Sampling time:
1 d
Parameter:
% degradation (CO2 evolution)
Value:
20
Sampling time:
3 d
Parameter:
% degradation (CO2 evolution)
Value:
23
Sampling time:
6 d
Parameter:
% degradation (CO2 evolution)
Value:
88
Sampling time:
10 d
Parameter:
% degradation (CO2 evolution)
Value:
100
Sampling time:
14 d
Parameter:
% degradation (CO2 evolution)
Value:
100
Sampling time:
22 d
Key result
Parameter:
% degradation (CO2 evolution)
Value:
100
Sampling time:
28 d
Details on results:
see table below
Remarks on result:
not measured/tested
Results with reference substance:
see table below

Day

% Degradation

 

Sodium benzoate

% Degradation

 

Test Material

% Degradation

 

Test Material & Sodium

Benzoate Toxicity

Control

0

0

0

0

1

24

19

18

2

40

12

31

3

48

20

36

6

84

23

44

8

96

67

60

10

111

88

67

14

111

105

77

16

110

100

73

20

112

108

73

22

114

95

66

24

113

107

68

28

112

111

92

29

111

113

94

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test material attained 100% 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:

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.

The test material, at a concentration of 10 mg Carbon/l, 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.

The test material attained 100% 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.

Description of key information

The test material attained 100% 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.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

Source: GLP-report