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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:
1994-09-13 to 1995-04-21
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
The study follows the 301B OECD guideline, GLP procedures and all validity criteria are fulfilled. The test solution is prepared via a volatil solvent but no solvent control was performed The test solution is slightly heated (at 40°C) before the beginning of the test but there is no information of the impact of this warming on the stability of esterol A and hence on its biodegradability in the test conditions.
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
yes
Remarks:
for details see material and methods freetext
GLP compliance:
yes
Remarks:
acreditation number 1-0210 (COFRAC)
Oxygen conditions:
aerobic
Inoculum or test system:
other: Secondary effluent from a biologic treatment plant
Details on inoculum:
- Type of sludge: other: Secondary effluent from a biologic treatment plant (domestic sewage)
- Sampling site: Versailles (France, 78)
- Preparation of inoculum: The effluent sample was collected the day before the test. Then it was centrifugated at 20°C during 20 min at 4000g and the pellet resuspended in test medium to reach a concentration factor of 40 in comparison to initial effluent sample.
- Initial cell concentration (after concentration treatment): 2.7 10E4 bact/mL
- cell concentration after preparation: 2.7 10E2 bact/mL
- Pretreatment: aeration until inoculation
- adaptation: no data is given about an eventual adaptation of he inoculum to the test solution, therefore it is considered by default that the inoculum had not been pre-adapted.

Duration of test (contact time):
29 d
Initial conc.:
20 mg/L
Based on:
other: TOC
Initial conc.:
25.7 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST SYSTEM
- Culturing apparatus: 3-litre Erlenmeyer flasks.
- Number of culture flasks per concentration: 2.
- Aeration device: Bubbling CO2-free air in darkness.
The CO2 elimination process is materialized by the aeration of flasks with CO2-free air after addition of the substance and before introducing
inoculum.
- Measuring equipment: Jenway 3410, Maïhak Tocor 100.
- Closed vessels used: Yes.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 2 flasks (inoculum)
- Abiotic sterile control: yes (test sub., inoculum and sterilising agent)
- Surface active agent control: 1 flask (emulsion of surface active agent, and inoculum)
- Toxicity control: 1 flask (test sub. reference sub., and inoculum)
- Procedure control: 1 flask (ref. sub. and inoculum)

METHOD OF PREPARATION OF TEST SOLUTION:
The substance was dissolved in a water-miscible and very volatile solvent:the Dichloro-1,1-fluoro-1-ethane.
The substance emulsion was realised by the addition of Synperonic P94 (surface active agent not biodegradable nor toxic).
This mixture was heated up to 40°C in order to facilitate the evaporation of the solvent.

ANALYTICAL PARAMETER:
Quantitative analysis of CO2 released.
The CO2 produced was trapped in barium hydroxide (0.0125M) and measured by titration of the residual hydroxide.

SAMPLING: 0, 1, 4, 5, 8, 11, 13, 18, 21, 28, 29 days.

TEST CONDITIONS
- Composition of medium: (same as requested in the guideline)
For 1 litre:
1) 10 ml of solution a:
8.5g KH2PO4
21.75g K2HPO4
33.4 g Na2HPO4,H2O
0.5g NH4Cl
ultrapure water up to 1000ml .
2) 1 ml of solution b: 27.5g CaCl2 or 36.40g CaCl2,2H2O in 1000 ml of ultrapure water.
3) 1 ml of solution c: 22.5g MgSO4,7H2O in 1000 ml of ultrapure water.
4 ) 1 ml of solution d: 0.25g FeCl3,6H2O in 1000 ml of ultrapure water (this solution was prepared at the beginning of the test).
5 ) 800 ml of pure water.
6 ) ultrapure water up to 1000 ml.

- Additional substrate: No.

- Test temperature: 22+/-2°C.

Reference substance:
benzoic acid, sodium salt
Key result
Parameter:
% degradation (CO2 evolution)
Value:
75
Sampling time:
29 d
Details on results:
- 65% biodegradation in the 10-d window (see figure 1 in document attached: EsterolA_5.1.2_Thiebaud1995_fig1 kinetic of degradation.doc)
- procedure control: 90% biodegraded after 29 days
- surface active agent control similar to blank
- No inhibition of inoculum observed.
- The degradation of the substance is only attributed to micro-organism. No physico-chemical degradation can be attributed to the degradation of the Esterol A.
Results with reference substance:
see table 1 kinetic of degradation

Degradation products: not measured. No effect of the surface active agent added in the test suspension Table 1: Kinetics of degradation (in %) of Esterol A (test substance), benzoic acid (ref substance) and toxicity control (EsterolA + benzoic acid)

Day

0

1

4

5

8

11

13

18

21

28

29

Test suspension 1

0

0

36

48

60

66

69

74

76

79

79

Test suspension 2

0

0

35

45

56

61

63

68

70

71

71

Test suspension Average

0

0

36

46

58

63

66

71

73

75

75

Inoculum blank

0

20

68

72

78

83

84

86

87

88

90

Toxicity control

0

2

39

49

61

66

68

72

74

76

76

Validity criteria fulfilled:
yes
Remarks:
for details see freetext overall remarks
Interpretation of results:
readily biodegradable
Conclusions:
The maximum biodegradation rate of Esterol A is 75% in 28d. Esterol A is readily biodegradable.
Executive summary:

The study of the ready biodegradability of Esterol A according to OECD 301B guideline, show a maximum level of biodegradation of 75% in 28days.

The percentage of biodegradation at the end of the 10-d window is 65%.

In the test conditions, no inhibitory effect of Esterol A on the micro-organism of the inoculum nor abiotic degradation were observed and all validity criteria were fulfilled.

Esterol A is considered as readily biodegradable.

Description of key information

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable
Type of water:
freshwater

Additional information

Biodegradability in water has been assessed on the registering substance: 75% in 28 days with 60% in the 10 -day windows, half life of 6/7 days