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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:
16 Mar - 13 Apr 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 310 (Ready Biodegradability - CO2 in Sealed Vessels (Headspace Test)
Version / remarks:
March 2006
Qualifier:
according to
Guideline:
ISO 14593:1999 (Water quality - Evaluation of ultimate aerobic biodegradability of organic compounds in aqueous medium - Method by analysis of inorganic carbon in sealed vessels (CO2 headspace test))
Version / remarks:
1999
GLP compliance:
yes (incl. certificate)
Remarks:
Staatliches Gewerbeaufsichtsamt, Hildesheim, Germany
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: aqueous phase of non adapted activated sludge, municipal sewage treatment plant, Hildesheim, Germany
- Pretreatment: sludge washed twice with autoclaved tap water; settled sludge was resuspended in mineral salts medium, aerated (4 h) and homogenized (blender); supernatant was decanted and maintained at aerobic conditions (aerated for 2 days with CO2-free air)
- Initial cell concentration: 1.3 x 10E+7 colony forming units (CFU)/L in the test vessel at test start
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
test mat.
Initial conc.:
14.5 mg/L
Based on:
other: TOC
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: according to guideline
- Test temperature: 21 - 23 °C (down to 20 °C for 18 h)
- Continuous darkness: no, low light conditions
- Agitation: 150 - 200 rpm

TEST SYSTEM
- Number of culture flasks/concentration: triplicates (5 replicates at the last sampling)
- Measuring equipment: inorganic carbon measurement (according to DIN EN 1484) by a carbon analyser Multi N/C 3000, Analytic Jena
- Test performed in closed vessels: yes
- Details of trap for CO2 and volatile organics if used: sodium hydroxide solution used to convert CO2 to carbonate

SAMPLING
- Sampling frequency: on days 0, 2, 5, 7, 12, 14, 21, 28
- Sample storage before analysis: at 6 ± 2 °C if necessary

CONTROL AND BLANK SYSTEM
- Functional control: yes, triplicates (5 replicates at the last sampling)
- Inoculum control: yes, triplicates (5 replicates at the last sampling)
- Toxicity control: yes, triplicates (5 replicates at the last sampling)
Reference substance:
benzoic acid, sodium salt
Key result
Parameter:
% degradation (CO2 evolution)
Value:
79
Sampling time:
28 d
Details on results:
10% of the biodegradation occured after 4 days, 60% was reached after 12 days and 79% of the test item was biodegraded after 28 days. The 95% confidence interval on day 28 was 74 - 84%.
Results with reference substance:
Transition of adaptation to degradation phase occured after 1 day (>=10% biodegradation). 60% of the substance was biodegraded after 3 days, which fulfills the validity criterion of 60% degradation after 14 days. 92% of the substance was biodegrated after 28 days. The 95% confidence interval on day 28 was 91 - 93%.

The biodegradation of the toxicity control reached 73% within 14 days and it came to 84% after 28 days. The biodegradation of the reference item was not inhibited by the test item in the toxicity control. The 95% confidence interval on day 28 was 83 – 85%.

Table 1: Biodegradation in the test item, toxicity control and functional control samples in % as well as the corresponding 95% confidential interval on day 28

 

Degradation [%]

Confidence Interval on Day 28

Day

0

2

5

7

12

14

21

28

P=95%

Test Item, 20 mg/L

-

0

20

49

62

65

72

79

74 – 84

Functional Control, 30 mg/L

-

54

76

80

86

90

87

92

91 – 93

Toxicity Control, 20mg/L Test Item + 30 mg/L Reference Item

-

-

-

58

-

73

78

84

83 - 85

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The substance was found to be readily biodegradable under the test conditions, with a mean percentage biodegradation of 79% at the end of the 28-day exposure period. The pass level for ready biodegradability was reached on exposure day 12.

Description of key information

Readily biodegradable (79% within 28 d, OECD 310)

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

One GLP guideline study according to OECD 310 is available investigating the ready biodegradability of the substance. A mixed microbial population of activated sludge of a local municipal STP was inoculated with a nominal concentration of 20.0 mg/L test substance (corresponding to a carbon content of 14.5 mg C/L) and incubated for 28 d under controlled conditions at low light conditions. Degradation was followed by the determination of dissolved oxygen content after 0 h, 7, 14, 21 and 28 d. An inoculum blank, a functional control (with 30 mg reference substance/L) and a toxicity control (20 mg/L test item and 30 mg/L Reference item) were run in parallel. The 10% level (beginning of biodegradation) was reached after 4 days. The pass level of 60% was reached after 12 days and the biodegradation came to 79% after 28 days. The percentage degradation of the functional control reached the pass level of 60% after 3 days and came to 90% after 28 days. In the toxicity control containing both test and reference item a biodegradation rate of 73% occurred within 14 days and it came to 84% after 28 days. Based on these facts the test item is classified as readily biodegradable. All validity criteria of the study were met.