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Biodegradation in water: screening tests

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Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
13 August 2003 to 10 September 2003
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
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
Qualifier:
according to
Guideline:
EPA OPPTS 835.3110 (Ready Biodegradability)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic (adaptation not specified)
Details on inoculum:
- Source of inoculum/activated sludge: final effluent stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK
- Laboratory culture: the sample of effluent was filtered through coarse filter paper (first approximate 200 mL discarded) and maintained, on aeration, in a temperature controlled room at 21 ± 1 °C
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Composition of medium: According to the OECD and EU guidelines:
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:
MgSO4.7H20:22.50 g/l

Solution d:
FeCl3.6H2O: 0.25 g/l

To 1 litre (final volume) of purified water 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
The pH of the final culture medium was 7.4.

- Test temperature: 21.1 - 21.4 °C
- pH: 7.6 - 8.1
- pH adjusted: no
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 500 mL bottles
- Number of culture flasks/concentration: four replicates (inoculated culture medium); three replicates (inoculated culture medium and 100 mg/L aniline); three replicates (inoculated culture medium and 100 mg/L test material); 2 replicates (inoculated culture medium, 100 mg/L aniline and 100 mg/L test material). All vessels were inoculated with the prepared inoculum at a rate of 1% v/v.
- Measuring equipment: Manometric biochemical oxygen demand (BOD) apparatus
- Details of trap for CO2: ethanolamine solution placed in a sealed cup in the neck of each bottle

OBSERVATIONS
- Frequency: daily
- Method: Manometer readings

CONTROL AND BLANK SYSTEM
- Inoculum blank: a control set up with no test material or aniline
- Toxicity control: test material and aniline at nominal concentrations of 100 mg/L each
Reference substance:
aniline
Parameter:
% degradation (O2 consumption)
Value:
27.5
Sampling time:
7 d
Remarks on result:
other: mean value
Parameter:
% degradation (O2 consumption)
Value:
37.5
Sampling time:
14 d
Remarks on result:
other: mean value
Parameter:
% degradation (O2 consumption)
Value:
47.5
Sampling time:
28 d
Remarks on result:
other: mean value
Details on results:
The test material attained 47.5 % degradation after 28 days and therefore cannot be considered to be readily biodegradable. The toxicity control attained 56 % degradation after 14 days and 64 % degradation after 28 days thereby confirming that the test material was not toxic to the sewage treatment micro-organisms used in the study.
Results with reference substance:
Aniline attained 76 % degradation after 14 days and 89 % degradation after 28 days thereby confirming the suitability of the inoculum and test conditions. Aniline attained degradation after 28 days, calculated from the results of the DOC analyses performed on days 0 and 28.

Table 2: Daily BOD Values

Day

BOD (mg O2/L)

Control

Aniline

Test material

Toxicity control

R1

R2

R1

R2

0

0.00

0.00

0.00

0.00

0.00

0.00

1

2.54

2.58

2.80

18.50

18.12

13.12

2

4.84

4.84

4.74

38.44

35.02

36.90

3

6.16

5.92

6.12

47.74

43.28

84.80

4

7.20

6.96

7.54

55.22

50.28

161.76

5

8.58

8.04

9.54

64.40

65.10

229.04

6

9.70

9.08

12.66

80.80

76.26

268.10

7

10.88

10.38

19.50

91.38

87.22

287.68

8

12.00

11.62

29.98

99.54

94.58

304.04

9

13.08

12.82

41.24

106.70

101.30

316.74

10

14.00

13.92

54.40

113.12

107.34

325.58

11

15.12

15.12

69.22

118.58

111.46

334.00

12

16.16

16.24

121.32

123.12

114.96

341.04

13

17.08

17.24

162.14

126.94

118.04

346.78

14

18.40

18.66

252.10

131.04

121.90

 352.02

15

19.58

19.78

256.38

134.82

125.74

356.64

16

20.08

20.28

263.96

138.24

128.58

360.60

17

20.86

21.16

273.60

142.44

131.66

366.28

18

21.78

22.12

279.18

146.40

135.02

372.18

19

22.74

23.16

282.50

150.18

138.40

378.30

20

23.36

23.86

286.38

153.48

140.98

383.92

21

24.04

24.74

289.22

156.48

143.36

388.52

22

24.78

25.62

291.96

159.22

145.66

392.64

23

25.54

26.48

294.48

161.86

147.90

396.30

24

26.08

27.24

296.54

164.22

149.90

399.46

25

27.04

28.32

298.38

166.84

152.24

402.72

26

27.86

29.32

300.48

169.40

154.44

405.68

27

28.48

30.24

303.96

171.64

156.52

408.46

28

29.48

31.18

306.02

173.68

160.52

411.38

R1, R2 = replicates 1 and 2

Table 3: Daily Percentage Degradation Values

Day

Aniline

Test material

Toxicity control

R1

R2

0

0

1

0

0

1

0

6

5

2

2

0

12

10

5

3

0

14

13

13

4

0

17

15

26

5

0

19

20

37

6

1

25

23

43

7

3

28

27

46

8

6

30

29

49

9

9

32

31

51

10

13

34

32

52

11

18

36

33

53

12

34

37

34

54

13

47

38

35

55

14

76

39

36

56

15

77

40

37

56

16

79

41

38

57

17

82

42

38

58

18

83

43

39

59

19

84

44

40

59

20

85

45

41

60

21

86

46

41

61

22

86

46

42

61

23

87

47

42

62

24

87

48

43

62

25

88

48

43

63

26

88

49

44

63

27

89

49

44

63

28

89

50

45

64

R1, R2 = replicates 1 and 2

Table 4: BOD and Degradation Values

Sample

ThOD  (mg O2/L)

Day 7

Day 14

Day 28

Mean degradation (%)

BOD (mg O2/L)

Degradation (%)

BOD (mg O2/L)

Degradation (%)

BOD (mg O2/L)

Degradation (%)

Control

R1

-

10.88

-

18.40

-

29.48

-

-

R2

-

10.38

-

18.66

-

31.18

-

Aniline

309

19.50

3

252.10

76

306.02

86

-

Test material

R1

289

91.38

28

131.04

39

173.68

50

48

R2

289

87.22

27

121.90

36

160.52

45

Toxicity control

598

287.68

46

352.02

56

411.38

64

-

R1, R2 = replicates 1 and 2

Degradation (%) = [BODtest — BOD controlmean]/[ThOD] x 100

ThOD of test material = 2.89 mg O2/mg

Validity criteria fulfilled:
yes
Interpretation of results:
other: not readily biodegradable
Conclusions:
Under the conditions of the study, the test material attained 47.5 % degradation after 28 days and therefore cannot be considered as readily biodegradable. The study is considered to be reliable, relevant and adequate for risk assessment and classification and labelling purposes.
Executive summary:

The ready biodegradability of the test material was determined in accordance with the standardised guidelines OCED 301 F, EU Method C.4-D and EPA OPPTS 835.3110 in a manometric respirometric test. The test material at a concentration of 100 mg/L was exposed to 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 measurement of daily oxygen consumption values on days 0 to 28. Control solutions with inoculum and the standard reference material, aniline, together with a toxicity control were used for validation purposes. Under the conditions of the study, the test material attained 47.5 % degradation after 28 days and therefore cannot be considered as readily biodegradable.

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Justification for type of information:
See read-across justification attached in Section 13.
Reason / purpose:
read-across source
Parameter:
% degradation (O2 consumption)
Value:
27.5
Sampling time:
7 d
Remarks on result:
other: mean value
Parameter:
% degradation (O2 consumption)
Value:
37.5
Sampling time:
14 d
Remarks on result:
other: mean value
Parameter:
% degradation (O2 consumption)
Value:
47.5
Sampling time:
28 d
Remarks on result:
other: mean value

Description of key information

The analogue test material (EC 271-638-7) was determined to be not readily biodegradable (OECD 301F, EU Method C.4-D, EPA OPPTS 850.3110).

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed

Additional information

The ready biodegradability of an analogue test material (EC 271-638-7) was determined in accordance with the standardised guidelines OCED 301 F, EU Method C.4-D and EPA OPPTS 835.3110 in a manometric respirometric test. The test material at a concentration of 100 mg/L was exposed to 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 measurement of daily oxygen consumption values on days 0 to 28. Control solutions with inoculum and the standard reference material, aniline, together with a toxicity control were used for validation purposes. Under the conditions of the study, the test material attained 47.5 % degradation after 28 days and therefore cannot be considered as readily biodegradable.

The study was performed in line with GLP and accepted standardised guidelines with a high standard of reporting. The study was assigned a reliability score of 1 and considered suitable for assessment as an accurate reflection of the test material.