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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:
29 Oct - 26 Nov 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Version / remarks:
17 Jul 1992
Deviations:
no
Qualifier:
according to
Guideline:
other: MEP, P.R. China, the Guideline for the Testing of Chemicals, Degradation and Accumulation (2nd edition), No. 301F "Manometric Respirometry Test"
Version / remarks:
2nd edition, 2013.9
Deviations:
no
Qualifier:
according to
Guideline:
other: SEPA, P.R. China, the Guidelines for the Hazard Evaluation of New Chemical Substances (HJ/T 154-2004)
Version / remarks:
2004
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Health Care Inspectorate, Ministry of Health, Welfare and Sport, Den Haag, The Netherlands (30 Apr 2014)
Test material information:
Composition 1
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Aeration tank of Shenyang Northern Sewage Treatment Plant (sampled 1 d prior test)
- Storage conditions: Aerobic conditions, 20.7 - 22.9 °C
- Storage length: 1 d
- Preparation of inoculum: Coarse particles were removed by filtration through a fine sieve and the sample was settled. The supernatant was discarded and the sludge was washed twice with mineral medium. The concentrated sludge was suspended in mineral medium. 10 mL suspended sludge (3 replicates) was weighed and dried at 105 °C for 2 h and 10 min and reweighed to calculate initial sludge concentration. The measured concentration of suspended sludge was 4.28 g/L as suspended solids (SS). On the day of the test start, 5 replicates of 10 mL sludge were weighed and dried at 105 °C for about 1.5 h and reweighed. The mean concentration of suspended sludge measured was 4030 mg/L as SS.
- Initial suspended solids concentration in test vessels: 30 mg/L
Duration of test (contact time):
28 d
Initial conc.:
48.7 mg/L
Based on:
test mat.
Initial conc.:
95.9 mg/L
Based on:
ThOD
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Composition of medium: According to guideline
- Test temperature: The BOD meter requires a test temperature of 20 ± 1 °C. The OECD testing guideline 301F requires a test temperature of 22 ± 2 °C.
- pH: The pH of the mineral medium was adjusted to 7.46 with HCl. The pH of the various test vessels was measured at the end of the incubation period on Day 28.
- Suspended solids concentration: 30 mg/L. Mineral medium and 14.885 mL pretreated activated sludge were mixed in a 2 L flask to give a concentration of about 30 mg/L microbial inoculum (suspended solids).
- Continuous darkness: Yes
- Other: The inoculated mineral medium was brought into an overflow measuring flask (244 mL) to precisely measure the volume of the sample (244 mL).

TEST SYSTEM
- Culturing apparatus: 500 mL Biochemical Oxygen Demand (BOD) bottles, filled with 244 mL test solution with sealed BOD sensor
- Number of culture flasks/concentration: 2
- Measuring equipment: Biochemical Oxygen Demand (BOD) bottles, equipped with magnetic stirring rod and BOD sensors.
- Details of trap for CO2: 5 drops of KOH (45% w/w) in a seal gasket inserted in the neck of the BOD bottle.
- Other: Incubation temperature was measured and recorded at 1 h intervals by automated temperature monitoring system and the measured range was 19.24 - 20.67 °C.

SAMPLING
- Sampling frequency: Oxygen consumption was continuously measured over the 28 d test period.

CONTROL AND BLANK SYSTEM
- Inoculum blank (2 replicates): Inoculated medium only
- Procedure control (1 replicate): 56.9 mg/L reference substance sodium benzoate (equivalent to 95.0 mg/L as ThOD)
- Toxicity control (1 replicate): 48.5 mg/L test item (equivalent to 95.5 mg/L as ThOD) + 56.5 mg/L reference item sodium benzoate (equivalent to 94.4 mg/L ThOD)
Reference substance:
benzoic acid, sodium salt
Remarks:
56.9 mg/L
Parameter:
% degradation (O2 consumption)
Value:
54.8
Sampling time:
28 d
Details on results:
Degradation did not reach the pass level of 60% within the 10 d window, at the plateau and at the end of the test.
The procedure control confirmed the suitability of the inoculum (81.5% after 28 d).

VALIDITY CRITERIA

The study fulfilled the validity criteria (Table 1). The pH values ranged from 7.38 – 7.71 and were within the required range of 6.0 – 8.5.

 

Table 1: Validity criteria OECD 301F

Criterion from the guideline

Outcome

Validity criterion fulfilled

Difference of extremes of replicate values of the removal of the test chemical at the plateau, at the end of the test or at the end of the 10-d window, as appropriate, is less than 20%.

After 28 d, degradation reached 54.6% in replicate n° 1 and 55.0% in replicate n° 2.

Yes

Percentage degradation of the reference compound has reached the pass levels by day 14.

The percentage degradation of the reference item reached 78.4% by Day 14.

Yes

The oxygen uptake of the inoculum blank is normally 20-30 mg O2/L and should not be greater than 60 mg/L in 28 days.

Oxygen uptake by the inoculum was 15.5 mg O2/L in 28 d.

Yes

 

BIODEGRADATION OF THE TEST ITEM

After 7 d, the average biodegradation of the test item reached 30.3%, which was the first day on which the 10% level was exceeded, marking the beginning of the 10-d window (degradation phase). Biodegradation reached 51.2% (Day 17) at the end of the 10-d window on Day 17 and 54.8% after 28 d. Therefore, the 60% pass level for biodegradation was not reached within the 10-d window.

Table 2. Results biodegradation of the test item (28 d)

Day

O2 uptake

[mg O2/L]

BOD

[mg O2/mg test item]

Biodegradation

[%]

 

IC-1

IC-2

TS-1

TS-2

IC average

TS-1

TS-2

TS average

TS-1

TS-2

TS average

1

UFL

UFL

UFL

0

0

0

0

0

0

0

0

2

0

0

UFL

1

0

0

0.021

0.011

0

1.1

0.6

3

1

1

2

2

1.0

0.021

0.021

0.021

1.1

1.1

1.1

4

2

2

4

3

2.0

0.041

0.021

0.031

2.1

1.1

1.6

5

4

3

5

6

3.5

0.031

0.052

0.042

1.6

2.6

2.1

6

5

4

7

8

4.5

0.051

0.072

0.062

2.6

3.7

3.2

7

6

4

35

33

5.0

0.615

0.578

0.597

31.2

29.3

30.3

8

7

5

48

46

6.0

0.861

0.825

0.843

43.7

41.9

42.8

9

8

7

52

51

7.5

0.912

0.897

0.905

46.3

45.5

45.9

10

8

8

54

54

8.0

0.943

0.949

0.946

47.9

48.2

48.1

11

10

8

56

55

9.0

0.964

0.949

0.957

47.9

48.2

48.1

12

11

9

57

56

10.0

0.964

0.949

0.957

48.9

48.2

48.6

13

11

10

57

57

10.5

0.953

0.959

0.956

48.4

48.7

48.6

14

11

10

58

57

10.5

0.974

0.959

0.967

49.4

48.7

49.1

15

11

11

60

59

11

1.005

0.990

0.998

51

50.3

50.7

16

12

11

61

60

11.5

1.015

1

1.008

51.5

50.8

51.2

17

12

11

61

60

11.5

1.015

1

1.008

51.5

50.8

51.2

18

12

12

63

61

12

1.046

1.011

1.029

53.1

51.3

52.2

19

13

12

64

62

12.5

1.056

1.021

1.039

53.6

51.8

52.7

20

13

12

65

63

12.5

1.076

1.042

1.059

54.6

52.9

53.8

21

13

13

65

64

13

1.066

1.052

1.059

54.1

53.4

53.8

22

14

14

66

65

14

1.066

1.052

1.059

54.1

53.4

53.8

23

13

14

66

65

13.5

1.076

1.062

1.069

54.6

53.9

54.3

24

14

15

66

65

14.5

1.056

1.042

1.049

53.6

52.9

53.3

25

14

14

67

66

14

1.087

1.073

1.080

55.2

54.5

54.9

26

15

15

68

67

15

1.087

1.073

1.080

55.2

54.5

54.9

27

15

16

69

67

15.5

1.097

1.062

1.080

55.7

53.9

54.8

28

15

16

68

68

15.5

1.076

1.083

1.080

54.6

55

54.8

IC          = Inoculum control

TS          = Test suspension

UFL       = measured value below 0

 

REFERENCE STUBSTANCE

Biodegradation of the reference substance (sodium benzoate) reached 62.0% on Day 4, 78.4% on Day 14 and 81.5% on Day 28.

TOXICITY CONTROL

In the toxicity control, biodegradation reached 54.0% within 14 d and the value exceeded 25% based on total ThOD, which indicates that the test item is not inhibitory to microorganisms of activated sludge under the tested conditions.

Validity criteria fulfilled:
yes
Remarks:
For further details please refer to “Any other information on results incl. tables”.
Interpretation of results:
not readily biodegradable
Conclusions:
The test item is not readily biodegradable according to the guideline criteria (54.8% degradation after 28 d, OECD 301 F) and is not inhibitory to microorganisms of activated sludge (54.0% degradation after 14 d).

Description of key information

Not readily biodegradable (54.8% after 28 d, OECD 301 F)

Key value for chemical safety assessment

Biodegradation in water:
inherently biodegradable

Additional information

There is one study available, in which the biodegradability of 2-Hydroxycyclohepta-2,4,6-trienone (CAS 533-75-5) was assessed according to OECD guideline 301 F and GLP.

In the Manometric Respirometry Test, 48.7 mg/L test item was inoculated with unacclimatised sewage microorganisms at a concentration of 30 mg/L (suspended solids) in mineral medium for 28 d in the dark. Biodegradation was quantified by continuously measuring oxygen consumption (uptake) throughout the test period. An inoculum control , procedure control and toxicity control were run in parallel.

The degradation phase, demarked by the 10% pass level, began on Day 7 when the average biodegradation of the test item reached 30.3% (2 replicates). After 28 d, biodegradation reached 54.8%. Therefore, the pass level of 60% for ready biodegradability was not reached after 28 d. The procedure control with the reference substance sodium benzoate reached 78.4% degradation after 14 d, confirming the suitability of the inoculum. The toxicity control attained 54.0% degradation within 14 d, indicating that the test item is not inhibitory to microorganisms of activated sludge under the tested conditions.

In conclusion, the test item is not readily biodegradable according to OECD guideline criteria.