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Environmental fate & pathways

Biodegradation in water: screening tests

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Administrative data

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-01-12 to 2012-03-13
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: EU, OECD method, GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
Species/Origin: Aerobic activated sludge, micro organisms from a domestic waste water treatment plant was supplied by the sewage plant Rossdorf, Germany.Conditioning:The aerobic activated sludge used for this study was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in tap water and centrifuged again. This procedure was done three times. The sediment was re-suspended in test water and aerated overnight. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight was determined. Based on this ratio, calculated aliquots of washed sludge suspension, corresponding to 1.5 g dry material per litre were mixed with test water. This suspension was used for the experiment.
Duration of test (contact time):
60 d
Initial test substance concentration
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
The purpose of this study was to determine the ready biodegradability of the test item bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate. The test item was exposed to aerobic activated sludge from the aeration tank of a domestic waste water treatment plant for 60 days. The biodegradation was followed by the oxygen uptake of the micro organisms during exposure. As a reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control.This type of study is recognized by the international test guidelines and should provide a rational basis to assess the ready biodegradation properties of the test item when incubated with aerobic activated sludge.The test flasks prepared according paragraph 6.6 were incubated at 22 * 1 °C. The pressure decrease in the reaction vessels was measured over complete experimental phase of 60 days using the BSB Sensomat system, Aqualytic Dortmund. The test flasks were closed gas-tight by a measuring head. Potassium hydroxide solution (45%) was used for trapping the produced carbon dioxide. The amount of O2 consumed by the activated sludge was calculated from the decrease of pressure in the reaction vessel.Preparation of Test Flasks:The amounts of test item were dissolved in acetone and dropped onto 100 mg silica gel. After evaporation of the solvent (for about 24 hours) the treated silica gel (230 – 400 mesh) was directly dosed into the flasks (ISO 10634; Handley et al., 2002). The inoculum control contained untreated silica gel with 100 µl acetone, which was evaporated overnight.The reference item was directly weighed into the test flasks. Incubation:The closed test flasks were incubated in a climatised room under continuous stirring. The consumption of oxygen was determined by measuring the change of pressure in the flasks. Evolved carbon dioxide was absorbed in an aqueous solution (45%) of potassium hydroxide.Test Duration:60 daysMeasurement of Oxygen:The change of pressure in the test flasks was measured by means of a manometric method (BSB/BOD-Sensor-System, Aqualytic Dortmund, Germany) each day.Temperature:Temperature was measured each working day in the climatised room.pH-Value:pH-values were measured in control, procedure control and a separately prepared test flask with test item at test start (to prevent loss of test item in the test flasks) and in all flasks at the end of the test using a pH-electrode WTW pH 340i.
Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

% Degradation
Parameter:
% degradation (O2 consumption)
Value:
94
Sampling time:
60 d
Details on results:
Biodegradation of Test ItemPercentage Biodegradation:ThODNH4The mean percentage biodegradation at the end of the 28-day exposure period was 53% (ThODNH4). The 10-day window was not passed. The incubation period was prolonged up to 60 days. Conclusion:The biodegradation of bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate was 94% after 60 days of incubation.Whilst the test item cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F, the evidence of aerobic degradation (94%) that did not plateau before the end of the 60-day indicate the potential for complete mineralisation in the environment. Therefore, bis(2,3-epoxypropyl)cyclohexane-1,2-dicarboxylate can be considered to be inherently biodegradable.

BOD5 / COD results

BOD5 / COD
Parameter:
COD
Value:
1.3 mg O2/g test mat.
Results with reference substance:
Biodegradation of Reference Item Sodium BenzoatePercentage Biodegradation:The reference item sodium benzoate was sufficiently degraded to 84% after 14 days and to 93% after 28 days of incubation. The results are represented in Tables 2 and 3 and Figure 1.Conclusion:The percentage biodegradation of the reference item con-firms the suitability of the used aerobic activated sludge inoculum.

Any other information on results incl. tables

Table.Percentage Biodegradation of Test Item, of Sodium Benzoate and of the Toxicity Control based on ThODNH4

Time

Percentage Biodegradation1

(Days)

bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate1

Sodium Benzoate2

Toxicity
control1, 2

 

Flask 1 [%]

Flask 2 [%]

Flask 5 [%]

Flask 7 [%]

1

0

0

35

20

2

0

-4

44

26

3

-2

-2

63

39

4

-4

-4

64

41

5

-4

-4

70

43

6

0

-4

73

46

7

-2

-2

77

45

8

2

2

77

48

9

4

8

78

51

10

8

8

81

53

11

15

11

81

54

12

17

17

83

57

13

15

19

81

57

14

19

23

84

59

15

23

23

84

59

16

21

24

83

60

17

23

23

84

61

18

26

26

84

62

19

30

26

84

64

20

30

30

84

64

21

34

34

87

66

22

38

34

87

67

23

38

38

90

69

24

41

41

90

69

25

43

43

92

68

26

49

45

90

67

27

53

49

93

69

28

56

49

93

69

29

56

53

93

69

30

62

54

92

70

31

64

56

93

71

32

68

56

93

71

33

66

58

94

70

34

68

60

93

69

35

71

64

93

69

36

71

68

93

69

37

75

71

93

69

38

75

71

93

69

39

75

75

93

69

40

75

75

93

69

41

75

75

93

69

42

75

79

93

69

43

79

79

93

69

44

79

79

93

69

45

79

83

93

69

46

83

83

93

69

47

83

83

93

69

48

86

86

93

69

49

86

86

93

69

50

86

86

93

69

51

86

86

93

69

52

86

90

93

69

53

86

90

93

69

54

90

90

93

69

55

90

94

93

69

56

90

94

93

69

57

90

94

93

69

58

94

94

93

69

59

94

94

93

69

60

94

94

93

69

            1ThODNH4of bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate: 1.30 mg O2/mg test item
            2ThODNH4of sodium benzoate: 1.666 mg O2/mg reference item

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
inherently biodegradable
Conclusions:
The biodegradation of bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate was 94% after 60 days of incubation.Whilst the test item cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F, the evidence of aerobic degradation (94%) that did not plateau before the end of the 60-day indicate the potential for complete mineralisation in the environment. Therefore, bis(2,3-epoxypropyl)cyclohexane-1,2-dicarboxylate can be considered to be inherently biodegradable.
Executive summary:

The ready biodegradability of the test item in a manometric respirometry test was determided based on the proceduresindicated by the following internationally accepted methods:

- EU method C.4 -D

- OECD method 301 F

The test item was investigated for its ready biodegradability in a manometric respirometry test over a period of 60 days. The biodegradation was followed by the oxygen uptake of the micro-organisms during exposure. As a reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control.

For the test item the evaluation of biodegradation has to be based on COD determination.

The criterion for ready biodegradability under the conditions of a manometric respirometry test is the 10-day window, describing the period between reaching at least 10% degradation and 60% degradation. This period should not exceed 10 days. The 10-day window criterion was not passed, because bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate was degraded to 53% after 28 days.

The mean biodegradation after 60 days of bis(2,3-epoxypropyl) cyclohexane-1,2-dicarboxylate was 94% (ThODNH4).

Whilst the test item cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F, the evidence of aerobic degradation (94%) that did not plateau before the end of the 60-day indicate the potential for complete mineralisation in the environment. Therefore, bis(2,3-epoxypropyl)cyclohexane-1,2-dicarboxylate can be considered to be inherently biodegradable.

The reference item sodium benzoate was sufficiently degraded to 84% after 14 days and to 93% after 28 days of incubation, thus confirming the suitability of theaerobic activatedsludge inoculum used.

In the toxicity control containing both, the test item and the reference item sodium benzoate, 59% biodegradation was noted within 14 days and 69% biodegradation after 28 days of incubation. Thus, the test item can be assumed to be not inhibitory to theaerobic activatedsludge micro organisms.