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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:
2017-04-28 to 2017-10-30
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test material information:
Composition 1
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
Test Inoculum
Activated sludge was collected from the Easton Wastewater Treatment Facility, Easton, Maryland on April 28, 2017. The Easton facility treats predominantly residential wastes. The sludge was sieved using a 2-mm screen, adjusted to approximately 1,200 mg total suspended solids/L with mineral media and then aerated at test temperature until use. A total suspended solids measurement and standard plate count were performed on the inoculum on the day of use in the test. Plates were incubated at 20 ± 3ºC for approximately 48 hours.
Duration of test (contact time):
28 d
Initial conc.:
10 mg/L
Based on:
DOC
Parameter followed for biodegradation estimation:
CO2 evolution
Reference substance:
benzoic acid, sodium salt
Key result
Parameter:
% degradation (CO2 evolution)
Value:
97.3
Sampling time:
28 d

RESULTS AND DISCUSSION

Carbon Analysis

The measured total organic carbon (TOC) concentration of the reference substance stock solution was 400.0 mg C/L. The volumes of stock solution used to dose the reference and toxicity control chambers was adjusted based on the measured TOC value of the appropriate solution so that approximately 10 mg C/L was delivered.

Observations and Measurements

The temperature range recorded during the test was 19.99-22.24ºC and was within the protocol specified range throughout the test. The results of the standard plate count and TSS measurement performed on the inoculum were 86 x 103 CFU/mL and 1140 mg/L,respectively.

The measured dissolved organic carbon (DOC) on Day 29 and pH values of the test chamber contents on Day 28 are presented in Table 1. The measured concentrations of inorganic carbon in the carbon dioxide trapping solutions are presented in Table 2. The cumulative amounts of CO2 produced over the test period are presented in Table 3. The cumulative percent of theoretical carbon dioxide

(% TCO2) evolved is presented in Table 4.

Table 1 Dissolved Organic Carbon (DOC) and pH of Test Solutions at Test Termination

 

 

DOC1

 

 

Test Chamber (Nominal Concentration)

(mg C/L)

pH

 

 

 

 

 

Control Rep. 1

1.495

7.42

 

Control Rep. 2

1.314

7.42

 

 

 

 

 

 

 

Sodium Benzoate Rep. 1 (10 mg C/L)

1.458

7.50

 

Sodium Benzoate Rep. 2 (10 mg C/L)

1.414

7.50

 

 

 

 

 

 

 

1, 12-Dodecanediol Bismethacrylate  Rep. 1 (10 mg C/L)

1.709

7.42

 

1, 12-Dodecanediol Bismethacrylate Rep. 2 (10 mg C/L)

1.499

7.44

 

 

 

 

Toxicity Control (10/10 mg C/L)

1.416

7.50

 

 

 

 

 

1Samples were filtered (0.45 µm) and acidified prior to analysis.

 

Table 2 Measured Inorganic Carbon Concentration of Trapping Solutions (mg C/L)**

 

 

Date

 

Day

Blank

Control

Rep. 1

Blank

Control

Rep. 2

Sodium

Benzoate

Rep. 1

Sodium

Benzoate

Rep. 2

1, 12-Dodecanediol BismethacrylateRep. 1

1, 12-Dodecanediol BismethacrylateRep. 2

 

Toxicity Control

 

KOH

Blank

5-May-17

8-May-17

11-May-17

15-May-17

18-May-17

23-May-17

25-May-17

30-May-17

1-Jun-17

2

5

8

12

15

20

22

27

29

25.50

39.12

32.94

34.80

25.99

24.96

20.29

25.20

18.24

24.92

36.18

34.83

32.99

25.26

25.61

16.82

22.79

18.27

86.84

153.90

72.35

62.34

37.69

38.02

19.82

27.80

20.82

102.70

157.40

83.44

61.24

34.85

31.83

17.67

25.80

18.62

32.44

125.5

109.90

89.93

52.88

51.66

22.53

39.38

21.15

35.72

132.1

100.30

84.49

49.64

44.51

23.43

31.07

22.10

110.40

277.30

136.80

94.67

54.20

50.14

23.16

35.13

22.65

10.29

11.08

8.24

9.59

11.42

5.21

6.69

4.13

7.57

                                                                *Concentrated HCl was added on Day 28. Samples were analyzed on Day 29.

                                                **Values in Table 2 were rounded for presentation purposes.

Table 3 Cumulative Milligrams of Carbon Dioxide Evolved1,2

 

 

Date

 

Day

Blank

Control

Rep. 1

Blank

Control

Rep. 2

 

Sodium3

Benzoate

Rep. 1

Sodium3

Benzoate

Rep. 2

 

1, 12-Dodecanediol Bismethacrylate3

Rep. 1

1, 12-Dodecanediol Bismethacrylate3

Rep. 2

 

 

Toxicity Control3

 

KOH

Blank

5-May-17

8-May-17

11-May-17

15-May-17

18-May-17

23-May-17

25-May-17

30-May-17

1-Jun-17

2

5

8

12

15

20

22

27

284

9.4

23.7

35.8

48.6

58.1

67.3

74.7

84.0

90.7

9.1

22.4

35.2

47.3

56.6

66.0

72.2

80.5

87.2

 

22.6

65.3

79.4

89.8

94.3

98.9

99.4

100.8

101.7

28.4

72.4

90.6

100.6

104.0

106.4

106.1

106.7

106.9

 

2.7

34.9

62.8

83.4

93.4

103.0

104.5

110.1

111.2

3.9

38.5

62.9

81.5

90.3

97.3

99.1

101.7

103.1

 

31.3

119.2

157.0

179.3

189.8

198.9

200.6

204.7

206.3

3.8

7.8

10.9

14.4

18.6

20.5

22.9

24.5

27.2

1The results of the inorganic carbon analyses of the CO2traps were converted to mg CO2produced using the following equation:

 mg CO2= cumulative result (mg C/L) X vol. of KOH (L) X 3.67 mg CO2/mg C

2Calculations performed in Excel 2010 full precision mode. Manual calculations may differ.

3Corrected for the CO2attributed to the inoculum and the KOH by subtracting the average amount of CO2evolved by the controls.

4Concentrated HCL was added on Day 28. Samples were analyzed on Day 29.

Table 4 Cumulative Percent of Theoretical Carbon Dioxide Evolved1,2

 

 

Date

 

Day

Blank

Control

Rep. 1

Blank

Control

 Rep. 2

 

Sodium

Benzoate

Rep.1

Sodium

Benzoate

Rep.2

 

1, 12-Dodecanediol BismethacrylateRep. 1

1, 12-Dodecanediol BismethacrylateRep. 2

 

 

 

Toxicity Control

 

5-May-17

8-May-17

11-May-17

15-May-17

18-May-17

23-May-17

25-May-17

30-May-17

1-Jun-17

2

5

8

12

15

20

22

27

283

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

NA

 

20.5

59.3

72.1

81.6

85.6

89.9

90.3

91.6

92.4

25.8

65.7

82.3

91.4

94.5

96.6

96.3

96.9

97.1

 

2.4

31.7

57.0

75.7

84.8

93.6

94.9

100.0

101.0

3.5

35.0

57.1

74.0

82.0

88.4

90.0

92.4

93.7

 

14.2

54.1

71.3

81.4

86.2

90.3

91.1

93.0

93.7

 

 

Cumulative Average Day 28 (N=2)

Standard Deviation

 

94.7

3.3

 

                                        97.3

                                          5.2

 

 

1The percentage of TCO2was calculated using the following equation:

 

% TCO2=X 100

 

2Calculations performed in Excel 2010 full precision mode. Manual calculations may differ.

3Concentrated HCl was added on Day 28. Samples were analyzed on Day 29.

NA – Not Applicable.

 

 

The blank control chambers evolved an average of 61.7 mg CO2 over the test period. This value has been corrected for the amount of CO2 in the trapping solution since potassium hydroxide solution, even when freshly prepared, contains carbonates. The amount of CO2 evolved by the blank control chambers did not exceed the 40 mg/L (120 mg total) value considered the acceptable limit for CO2 evolution tests. The viability of the inoculum and validity of the test were supported by the results of the reference substance, sodium benzoate, from which an average of 94.7% of theoretical CO2 was evolved. An average percent biodegradation of greater than 60% was achieved by Day 5, thereby fulfilling the criteria for a valid test by reaching the pass level by Day 14. The final mean percent biodegradation for 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) was 97.3% by Day 28. 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) may be considered readily biodegradable since the pass level of 60% TCO2 was achieved within 10 days of achieving 10% TCO2. The toxicity control achieved > 25% degradation by Day 14 and may be considered non-inhibitory at the concentration tested in this study.


Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
CONCLUSIONS
Evidence of ready biodegradability in a Carbon Dioxide Evolution Test is 60% TCO2 within the 28-day test period (1,2). In addition, the pass level must be reached within 10 days of achieving 10% TCO2 (1,2). The final mean percent biodegradation for 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) was 97.3% by Day 28. 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5), may be considered readily biodegradable since the pass level of 60% TCO2 was achieved within 10 days of
achieving 10% TCO2. The toxicity control achieved > 25% degradation by Day 14 and may be considered non-inhibitory at the concentration tested in this study.

Executive summary:

The ready biodegradability of 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) was determined by the Carbon Dioxide Evolution Test Method (OECD Guideline 301B). Tests of ready biodegradability are stringent tests that provide limited opportunity for acclimation and biodegradation to occur. In the CO2 test, inoculated mineral medium was dosed with a known amount of test substance as the nominal sole source of organic carbon and aerated with CO2-free air. The CO2 produced from the mineralization of organic carbon within the test chambers was displaced by the flow of CO2-free air and trapped as K2CO3 in KOH trapping solution. The amount of CO2 produced by the test substance (corrected for that evolved by the blank inoculum) is expressed as a percentage of the theoretical amount of CO2 (TCO2) that could have been produced if complete biodegradation of the test substance occurred. The test contained a blank control group, a reference group, a treatment group and a single toxicity control. Each group contained two replicate test chambers. The blank control was used to measure the background CO2 production of the inoculum and was not dosed with a carbon source. The reference chambers were dosed with sodium benzoate, a substance known to be biodegradable, at a nominal concentration of 10 mg C/L. The treatment group test chambers were used to evaluate 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) at a nominal concentration of 10 mg C/L. The toxicity control was used to evaluate the toxicity of the test substance to the inoculum and was dosed with both the reference (10 mg C/L) and test substances (10 mg C/L). The results indicated that the activated sludge inoculum was active, degrading the reference substance an average of 94.7% and that the test substance was not inhibitory to the inoculum at the concentration tested, as the toxicity control exceeded 25% degradation by Day 14 of the study. The average cumulative percent biodegradation for 1,12-Dodecanediol Bismethacrylate (CAS#72829-09-5) was 97.3% by Day 28.

 

 

Test Substance

Average

Cumulative Percent

Biodegradation1

 

Average

Final

pH1

 

 

Readily Biodegradable2

 

1, 12-Dodecanediol Bismethacrylate (CAS#72829-09-5)

 

 

97.3

 

 

7.43

 

 

Yes

 

1     Average of two replicates.

2     Meets or exceeds the OECD criteria for ready biodegradability (60% of TCO2within a 10-day window of reaching 10% TCO2).

Description of key information

A CO2 Evolution study assessing the ready biodegradability of the test item was conducted in accordance with the OECD 301B Test Guideline and GLP requirements.

The average cumulative percent biodegradation for the test substance was 97.3% by Day 28 and the 10-day window criterion was fulfilled; the test substance was thus considered to be readily biodegradable under the conditions of this test.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information